Quick Foreword: The Palace, (and before that, The Terrace)
remains the best board to get into serious discussions regarding Peregrine. The
Terrace is run by long time Peregrine champion, TerryGD. Due to the “Members
Only” format, this board is top notch without any of the BS that can permeate
other forums. This compilation is not 100% complete, I have left some of
Golfdad’s less science-oriented messages at the Palace.
Thank you for visiting.
VISIT Terry's
Peregrine Palace
Generally,
BLUE TEXT = DR. PHILIP THORPE
GREEN TEXT = DR. ALAN EPSTEIN: COTARA (TNT)
and VEA
BLACK TEXT = ONCOLYM
THE GOLFDAD MONOLOGUES
11/17/1998
Rutuxan:
Idec-C2B8 (Rituxan): This is a chimeric
antibody directed against a B-cell specific antigen expressed on NHL cells. The
target binding site of this antibody appears to allow several favorable
biological effects, including mediation of complement-dependent lysis of
malignant cells (a natural killing method mediated by serum proteins found in
the course of many infectious states), the ability to provoke
antibody-dependent cell killing (another natural consequence of binding
antibodies to foreign cells) and the Rituxan antibody can directly inhibit the
proliferation of NHL cells. Hence, these properties may be sufficient to not
seek any radiolabel (which adds direct killing power to the vicinity of cells
that bind the radiolabeled antibody), and allows its use as a "naked"
antibody, although high concentrations (perhaps on the order of gram amounts)
need to be infused.
Initial Phase I trials (and subsequent
studies) were conducted in patients who eventually failed further chemotherapy
and had few alternative therapies. The use of this antibody is attractive
because of its structure (chimerized, therefore little impact on patients
developing antibodies to Rituxan), the fact that it is not coupled to any toxic
material (such as a radioactive isotope or drug) and can be administered
repeatedly with few toxic side effects, other than eventual depletion of normal
B-cells (which are needed for proper immune protection of the patient). The
effects on immune cell depletion and function can be closely monitored. To
summarize the early results in brief fashion (how can golfdad ever do
that?)...a total response rate of about 45-50% was observed in patients with
refractory, low-grade lymphoma. It must be emphasized that complete and partial
responses are not entirely the same quantities between different clinical
groups, but in general, in the lymphoma studies, a complete response requires
no evidence for any disease for at least 1 month, whereas a partial response in
normally considered to be a reduction of anywhere between 50-70% of tumor
volumes measured by various parameters (dimensions off of radiograph films, CT
scans, etc). However, many descriptions by academic physicians hedge different
terms using phraseology like a "marked reduction", "although not
significant, about 25% less disease", and remarks like that...but us
simple folk know what we want to see: (1) percent of complete response, (2)
percent of partial responses, (3) duration of the remission (time to treatment
failure or time to relapse), and (4) overall survival. These early studies,
like most Phase I set the dose, schedule and probable patient effects to
monitor and watch for as the trials progress. They settled on about 750
mg/patient (depending on body size) once a week for 4 weeks, observed a fairly
rapid depletion of B-cells and malignant cells (normal B-cell numbers would
return in about 3-6 months) and the clinical responses lasted about 8-12
months, depending on the study reported. Low doses of the antibody resulted in
some clinical responses that were encouraging but not impressive. Only a few
patients with intermediate or high-grade lymphoma were attempted in the early
trials. Responses tend to favor patients with follicular disease (a kind of
localized network of webbed malignant cells, strung together in lymph node
structures). The half life of the antibody is rather long...and increases to
nearly 20-30 days as the number of infusions of the antibody increase. They
also conducted a small Japanese study in Phase I at the Japan NCI and observed
about a 40% response rate in relapsed patients.
The company quickly moved to Phase II trials
in a multi-center study (31 centers, about 170 patients) and used the 750
mg/dose (approx) and 4 weekly doses as the gold standard. They observed a 48%
total response rate (note that this was 6% complete response and 42% partial
response, where a complete response was defined as complete clearing of the
bone marrow and nodal sites and partial was a reduction of 50% or more) with a
duration of response of about 13 months. Toxicity was mild and the effects on
B-cell depletion were expected and monitored.
The company also performed studies in Europe and Australia and tested the
product in more aggressive forms of lymphoma, including intermediate and high-grade
lymphomas (diffuse large B-cell lymphoma, mantle cell lymphoma). They performed
a prospective randomized Phase II study including patients if they were in
first or second relapse, refractory to initial therapy, or elderly (>60 yo)
and not previously treated. The patients received 8 weekly infusions of Rituxan
at 750 mg/dose in one arm of the study and the patients in the other arm got
one dose of 750 mg/dose followed by 7 weekly infusions of 1000 mg/dose
(approx). 54 patients were evaluated, with 5 complete responses (10%) and a
partial response of 22%, with an average duration of response of about 8
months, suggesting that this modest clinical activity (in my opinion) needed to
be combined with chemotherapy...or this form of therapy limited mainly to low-grade
lymphoma. I did not find any parameters that may "predict" for
patients with high-grade lymphoma who would respond to this form of therapy
(that is, how do you determine those 17 patients out of the 54 before starting
therapy; is there any biological parameter you could measure, like extent of
antibody binding to lymphoma cells, or??).
The company has recently reported pilot
studies where Rituxan was combined with CHOP chemotherapy in patients with
previously untreated intermediate and high-grade lymphoma (I will refrain from
marketing remarks like..."in an attempt to break into that cohort of
patient numbers"...). Rituxan was given on day 1 (750 mg/dose) followed a
couple of days later by the usual CHOP chemo cycle and these cycles were
repeated 6 times. Of the 30 evaluable patients, they reported a 63% complete
response rate and 33% partial response. The authors do not clearly state
whether this is better than what they would have observed using CHOP alone
(with no Rituxan) on patients receiving their first chemotherapy, but I believe
CHOP therapy alone in this grade of lymphoma would have resulted in about a 65%
response rate...suggesting that this combination therapy may be of benefit as
front-line therapy. But since they don't say it...I won't say it. Clearly these
studies are continuing.
A multicenter study has also been performed with Rituxan combined with
alpha-interferon (a protein produced by lymphocytes that modulate a variety of
immune functions, including upregulation of cell antigens, direct
antiproliferative effects on tumor cells, inhibition of cellular products that
assist tumor growth and on and on). This study included 26 low-grade lymphoma
patients that were refractory to therapy and treatment consisted of modest
amounts of interferon given several times a week followed by 4 weeks of the
gold standard Rituxan therapy. 8% complete response and 50% partial response
was noted...and I suspect these studies will fade away...
Bexxar:
Anti-B1 antibody (Coulter). This is a
different antibody targeted to the CD20 antigen of B-lymphocytes and is used as
a radiolabel conjugate and is the direct correlate of radioimmunotherapy
currently in use by TCLN and its Lym-1 antibody (to be described later). This
form of therapy is championed by M. Kaminski and the group at Ann Arbor,
Michigan and in my opinion, these people strongly believe in the clinical
efficacy and approaches they use in treatment with this reagent. As you will
see...they are willing to push the envelope in attempting to make this a
front-line therapy for NHL.
Initial studies of Phase I focused upon the
use of the I-131 (will confine remarks to iodine-based radiotherapy) conjugated
to the B1 antibody and like all Phase I studies, they were limited to patients
in dire need of further therapy with few alternatives. Studies defined the
parameters of safety, dose and with this form of leukemia, one could look for
therapeutic results. There are caveats to this form of therapy (radiolabel) not
found with the use of naked antibodies...since we are delivering radiation to
target sites in the body, and the toxicity will be limited by effects on normal
tissue no different than observed for whole body irradiation, targeted beam
radiotherapy or chemotherapy with toxic agents. Initial studies have to
determine that the radiation uptake by areas of tumor is greater than that of
normal organs and close monitoring of vital blood counts, effects on bone
marrow, GI tract, central nervous system, etc is necessary. The logistics and
expenses of this form of therapy can be extensive.
Pilot studies in 34 patients with NHL were performed several years ago and the
dosimetry was established (conditions under which one could accurately estimate
the amount of radiation being delivered to critical areas containing lymphoma).
Hematologic toxicity was limiting and doses were escalated until about 750 rads
(a measure of radiation being actually deposited in tissue) was found to be the
upper limit, and represented about 50-75 mCi of radioactive material. A single dose
of B1 (when I say B1, it is always radiolabeled with I131) was given as
therapy, and of the 28 patients that could be evaluated, about 50% achieved a
complete response (see earlier definitions) and a partial response (Bexxar
studies usually use 50% reduction as partial) in 8 patients. The median
duration of remission was about 16.5 months. These results could be broken down
further in that of the 28 patients, 21 patients were low-grade or transformed
NHL and of these patients, 19/21 responded and of the 7 intermediate-grade
lymphoma patients, 3 responded. Hence, again, as with most therapies, better
responses were observed in the low-grade lymphomas.
A comment on these initial studies is warranted.
Radiotherapy with antibodies tends to unlease his hematologic effects due to
resident time in the bone marrow...and if bulky disease or extensive bone
marrow involvement is present, then non-specific toxicity will increase, just
because of the targeting of the antibody. This is a common event in higher
stage disease (say III and IV) because the bulk of the disease is not confined
to discrete areas, such as lymph nodes. Hence, not only are there different
risk factors than observed for the use of naked antibody, the whole system
becomes complex in considering specific sites of disease and effects of the
radiolabel...something to keep in mind.
These initial studies were reported a couple of years later (in a recent paper
in Journal of Nuclear Medicine) and described results of 34 patients. The
maximum tolerated dose was established as about 750 rads and in line with
intuition, the patients who received higher doses of the therapy and responses
were as described above. Some patients developed antimouse antibodies (since
this antibody is mouse), with additional data that complete responses averaged
about 13 months and the median duration of response for all patients who did
respond about 11.5 months.
This group quickly took this form of therapy to the front line in several
respects. First, the final Phase I/II results were reported this spring and
involved 59 patients (28 low grade, 14 transformed low-grade, and 16
intermediate and high grade lymphomas). The median number of prior
chemotherapies was 4 and remember that these studies involve patients who are
non-responsive to further chemotherapy. Without intervening for bone marrow
loss (for example, will discuss autologous bone marrow transplantation below),
the therapeutic dose remained at about 750 rads. 24/28 of the low-grade
patients responded and a complete response was observed in 46% of these
patients. The mean tumor dose was about 14.5 times the whole body dose, meaning
that localization to the tumor was good and much higher than that received by
normal tissues, on the average. 15% of the patients developed antimouse
responses which can limit further therapy. To be honest, I could not calculate
how the intermediate grade patients fared on this therapy, since the given
numbers (ASCO) were confusing to my trained eye.
However, the power of clinical funding for
this therapy is apparent. Also reported this spring are several other
aggressive moves with B1 antibody. For example, the Michigan group showed that
NHL patients who relapse from the initial B1 therapy (from the Phase I/II
described above) can be retreated. 13 patients were retreated and of these 13,
12 responded to the initial therapy....described above. 11 of these patients
were from the low-grade NHL group (suggesting that the intermediate grade
patients above did not do well?) and of these 13, 8 responded to re-treatment
with a median duration of about 8 months. 4 of the 13 had a complete remission
(second remission) of about 10 months. Again, it was noted that the higher the
dose received (for whatever reason), the likelihood of response was
greater...hence, this strategy was modestly successful, in that re-treatment of
relapse patients can be attempted with some degree of confidence. But
wait...there's more.
In collaboration with the O. Press group at University of Washington, the
Coulter people have examined the effects of Bexxar therapy with
chemotherapy....which we all know we are going to have to do. They described at
ASCO, a Phase I/II trial of combining high-dose I131-B1 with chemotherapy
(etoposide, cyclophosphamide) and autologous bone marrow transplantation in
relapsed patients. Now what this means is as follows....they are going to
administer whopping doses of the radioactive material couple to the antibody,
and they know it is going to wipe out the bone marrow...and they are going to
give very potent and toxic chemotherapy, which they know is also
myelosuppressive and they are going to "rescue" these patients from
the myeloablative effects of these therapies by reinfusing their bone marrow
stem cells that will give rise to new blood cells...and recovery from this
massive attack on the lymphoma that will not go away. Very aggressive therapy,
but cancer is an aggressive disease. They report treatment ranging up to 850
mCi (contrast that with perhaps 50-75 mCi of single dose administration)
calculated to deliver up to nearly 3000 rads to critical organs. Of the 37
evaluable patients (26 with indolent, low-grade lymphoma and 12 with aggressive
NHL originally enrolled in study), 33 are alive and 29 are progression free
(not necessarily disease free) after 1.5 years. 4 patients have died. This
study will continue.
In the meantime, the Coulter people have
evaluated the with an independent review...the response to B1 antibody therapy
in low-grade (or transformed low-grade) NHL...and this is the source of the
artistic statistics that Berblady commented on a few days ago, with her astute
and discerning fashion. Be that has it may...using the patients that were
refractory to further chemotherapy, they examined whether treatment with B1
gave a remission duration or response that was "at least as good" as
the last chemotherapy they received...which is an academic question of how are
you going to approach treating the refractory patient...and can you do as good
as chemotherapy. And wouldn't we rather have anything than chemotherapy,
perhaps? But I will not digress at this late stage of information. To make a
long analysis short...of the 45 patients evaluated, the question was asked how
did they do compared to their last chemotherapy. The facts are: 11 patients did
as good with either therapy...19 did better with B1 (at a design cutoff, I
believe of at least 1 month median difference) and 11 did better with their
last chemotherapy. I will leave it to all the statistic buffs to play with
these results...but wait,...
They have at least got the ambition to take on first line therapy, since they
now describe two other approaches involving the use of B1. First, there is a
recent paper (J. Clin. Oncol. 16: 3270, 1998) in which they report the use
again of massive doses of B1 therapy with bone marrow rescue (they purge the
bone marrow of potential malignant cells as best they can by a variety of
biological techniques) in patients who have relapsed from prior chemotherapy.
Hence, this study is like that described above except they don't add the new
chemotherapy. The therapy was administered to 29 patients (19 low grade and 10
high grade), most with stage III and IV disease which we would expect. Dosing
ranged up to nearly 800 mCi. The results are not easy to interpret (always
encounter this with the Press group, but no offense intended)...but as best as
I can decipher...25/29 patients responded, and 23 of these were complete
responses and the best responses were in the low-grade group (that is as good
as I can see for the moment). 11 of 19 patients with indolent lymphoma remain
in remission up to about 3-6 years and only 3 of the patients with aggressive
disease remain in remission (27, 37 and 87 months). The overall survival rate
for low-grade is 78% at 4 years...and the median time to treatment failure is
about 3 years for low-grade and about 2 years for the high-grade group.
But it ain't over...they have described a study back at Michigan where this
form of therapy has moved to first line with NO prior treatment of any kind in
follicular lymphoma, a type that is amenable to this form of therapy (when you
analyze all the groups). They received therapy up to the 750 rad limit and the
intent is to enroll 60 patients...21 are now evaluable. Complete response in
71% and partial response in 29% (100% total) and of the complete responses, 2
patients have relapsed (median duration not yet reached at about 17 months) and
5 out of the 6 patients who had the partial response have relapsed...and so the
intent is to back this treatment philosophy up until limits of success are
reached.
Oncolym:
Oncolym, interestingly, is the marketed trade name for the
radiolabeled Lym-1 antibody when it is ready for infusion...piece of trivia...in
contrast to Lym-1 which is the antibody (mouse class IgG2a, originally
described by Epstein in 1987 concerning his studies with a monoclonal antibody
against an antigen defined on the membrane of a cultured Burkitt lymphoma cell
line. This antigen was found to be a mutated form of a Class II recognition
antigen (in human systems, called HLA, for Human Leukocyte Antigen)...and
specifically, a recognition antigen found on B-lymphocytes (and other cell
types) of a subcategory called HLA-DR. These molecules are involved in cell to
cell communication and exchange of molecular information regarding the
structure of foreign antigens; hence we make immune responses using these
recognition molecules. Now, in this lymphoma, this molecule was mutated and Epstein
was able to isolate a mouse lymphocyte that makes monoclonal antibodies to it.
You can scale up and make a ton of it if necessary and try to treat lymphoma
patients if they carried the same molecule on the surface of their B-cell
lymphomas (and the rest is history). The best I could track in the literature
and I assume there are people familiar with the TCLN message board who go back
more than a decade into the beginning of TCLN and may be able to offer more
insight into any early studies regarding the use of Lym-1 in therapy of
lymphoma, but I could only really track the scientific literature.
Most of the published work on Lym-1 comes from the
prolific laboratory of the academic group of Sally and Gerald DeNardo at the
University of California at Davis. However, there are two major differences
that I observed in my reading of the literature on the clinical use of Lym-1
and the literature associated with Bexxar and Rituxan...namely, (1) the lack of
apparent funding for the expanded clinical trials needed for Lym-1 as
determined by the limited number of trials being conducted and (2) the
marvelous scientific methodologies used by the DeNardos in attempts to clearly
define the parameters of successful therapy of lymphoma using radiolabeled
antibodies. They have set the standard for defining conditions of antibody
localization, dosimetry and predictive parameters that help to define exactly
who and under what conditions patients will respond to this form of therapy.
Will touch on this matter later, just for personal interest. Although published
perhaps in only abstract form years ago, some initial therapy was attempted
with the “naked” Lym-1 antibody in lymphoma patients, which would be the
natural thing to do, since the antibody is of a subtype (IgG2a) that could
cause killing of the lymphoma cells under favorable circumstances after
binding...but this was not observed. Interestingly, one of the first
manuscripts was a case study involving a patient with Ricter’s syndrome, a
somewhat bizarre form of chronic lymphocytic leukemia and the patient was
nearly moribund with rapidly growing tumor. The patient was successfully
treated with a series of infusions with I131-Lym-1 with rapid shrinkage of the
tumor sites and the patient was described to be followed over the next 10
months (an average survival for this syndrome was 4 months)...but I have not
located (nor searched very hard) a follow-up for this particular patient since
the study was an isolated incident published over a decade ago. But it was
worthy of historical note, and the initial enthusiasm must have been exciting
for the medical staff as well as the family of this patient.
Pilot studies in Phase I were taking place before 1987
since one of the first studies was published in 1988 describing treatment of 10
patients with progressive, refractory B-cell malignancies with radiolabeled
Lym-1 (everything I write about here involves the use of radiolabeled Lym-1). I
will tend to skip through these early studies (1988-1990) rather quickly since
the follow-up described in later papers are repetitive and the Phase I studies
were examining distribution of antibody, relative uptake at site of tumor
compared to normal tissues, etc. Early therapy consisted of fractionated,
relatively low doses (25-60 mCi) in intermediate and high-grade patients,
almost all with stage III and IV disease, hence the toughest populations of
lymphoma patients were being treated...late stage disease, refractory to
further chemotherapy and progressive. Of an initial 18 patients, 10 responded (56%,
complete or partial) by parameters described in my earlier posts about the
Rituxan and Bexxar therapies. Toxicity with these low-doses was modest.
However, a brief study published in 1994 described the use of Lym-1 in 5
refractory CLL patients and the use of high-end fractionated doses (up to 65
mCi at 2-6 week intervals) were leading to thrombocytopenia and the group
realized that strategies were needed to support this cytopenia before high-dose
radiotherapy would be a commonplace modality.
Several studies up to the present time from this first
definitive study of Lym-1 therapy have focused upon a reevaluation of data
already obtained and available while doing Phase I/II studies, such as
determining that there is no real reason to exclude patients with splenomegaly
(enlarged spleen, usually with tumor burden) from therapy using Lym-1. A paper
describing this evaluation of a small subset of patients with splenomegaly was
given at the 6th Monoclonal Antibody Conference and also published in Cancer.
At the Eleventh International Conference for Use of Monoclonal Antibodies in
Cancer, in February of this year, DeNardo presented a paper in the milestones
of Lym-1 development, describing in detail their analysis of importance of
LDH/KS in determining the response to this therapy (and probably similar
therapies under these conditions). In addition, at this meeting, Jamie Oliver
described the Phase I/II studies using multidose therapy with Lym-1 in patients
with intermediate or high-grade lymphoma. This was co-authored by the DeNardos
and carried the byline of Techniclone Corp and USC-Davis. 33 patients with
recurrent or refractory intermediate/high grade disease. Doses ranging from
2—200 mCi. Overall, 52% of the patients responded to multidose therapy (CR or
PR) with a mean durability of 13 months and those receiving the higher doses
had a higher rate of response. Thrombocytopenia is dose-limiting (at about
160-200 mCi). An early report from the multicenter Phase II?III (?) study was
also given at this meeting, representing physicians from M.D. Anderson Cancer
Center, Cornell Univ., George Washington Med Center, Iowa City VA, and Univ of
Miami. Patients enrolled received at least 1 prior chemotherapy and were
eligible for up to 4 doses at 160 mCi. 15 patients enrolled, and 9 received at
least one therapeutic dose...3 patients with response (1CR and 2PR) and clearly
this study is too early to talk about or successfully evaluate...this was
earlier this year.
This same story was repeated by both sets of authors at
ASCO in the spring. For those interested in detailed analysis, the DeNardos
published a detailed paper about a year ago in Clinical Cancer Research
outlining the analysis of predictive parameters in patients upon which they had
a very lot of clinical and laboratory data. Suffice it to say that the
development of antimouse antibodies turns out to be a good prognostic factor
for ultimate survival time after therapy...i.e., those patients who develop
HAMA actually do better...representing some facet of the immune response of
these patients not readily apparent in its tie to lymphoma. Two additional
items. One, I posted a summary of the recent paper published in Oct describing
the value of high-dose, fractionated radiotherapy with Lym-1 and I hesitate to
repeat that post here.... (See Post # 181 and 182 to Investor CG). Finally,
much of the summary of the therapy involving 88 patients and a description of
Phase III studies are given from a description of prior news releases and can
be found at:
http://www.wizard.com/NHL/research/Oncolym.htm
Relevant References : Sang-Moo, L., et al, Prediction of
myelotoxicity using radiation doses to marrow from body, blood and marrow
sources. Journal Nuc Med 38:, 1374-, 1997; DeNardo, D.A., et al, Imaging for
improved prediction of myelotoxicity after radioimmunotherapy. Sixth Conf on
Radioimmunodec and Radioimmunother of Cancer, Suppl to Cancer, ACS, 2558, 1997;
Shen, S., et al,, Impact of splenomagaly on therapeutic response and I-131
LYM-1 dosimetry in patients with B-lymphocytic malignancies., ibid, 2553, 1997;
DeNardo, et al, Increased survivial associated with radiolabeled Lym-1 therapy
for non-Hodgkin's lymphoma and chronic lymphocytic leukemia., ibid, 2706, 1997;
DeNardo, G.L., et al., Overview of radiation myelotoxicity secondary to
radioimmunotherapy using I131-Lym-1 as a model Cancer, 73:, 1038-, 1994.;
DeNardo, S.J., et al. A direct apporach for determining marrow radiation from
MoAb therapy, in Biology of Radionuclide Therapy, Washington, DC, Amer Coll Nuc
Phys, , 110, 1989; DeNardo, G.L., et al, Body and blood clearance and marrow
radiation dose of I131-Lym-1 in patients with B-cell malignancies. Nucl Med
Commun, 14: 587-, 1993.
11/22/1998
With respect to the article published this past month by
the DeNardo's (J Clin Oncol 16: 3246, 1998), it was asked whether this is
"old" or "new" data, with inflection, I assume on whether
these terms represent progress in the company's trials for this product. All
published data is rather "old" due to the lag in publication time
found for academia and clinical results always represent a follow-up period of
substantial length due to the nature of the disease in its time of remission or
ultimate progression. I prefer to look at the data in this article as a natural
extension of PhaseI/II studies designed to optimize the parameters by which
they can administer the highest possible radiation dose to the site of lymphoma
and maintain acceptable toxicity. This study defined the ability to
"fractionate" the total radiation delivered into separate infusions
of labeled antibody (about 4 weeks apart, up to 4 doses, but most received two
therapeutic doses). All patients, as usual, were refractory to further chemotherapy
and all grades (low, intermediate and high) of NHL were included. Several
objective results were found and can be summarized in my ususal simplistic
manner: (1) if you do NOT respond to radiolabeled antibody therapy, you are in
big trouble and will succumb quickly to progressive disease, (2) of the 21
"entries" (1 patient entered the trial twice) into this analysis,
about 50% of the patients were responders (33% complete) and the best overall
response was observed with the low-grade NHL (in terms of survival and duration
of remission) although half of the intermediate and high-grade patients also
responded to this therapy, and (3) if you are able to achieve high-dose
delivery (about 200mCi per patient), the response rate approaches 100%.
This publication does define what can be accomplished in
terms of maximum dose delivery and value of fractionating the radiation. This
information, together with the knowledge of toxicities carefully outline the
nature of how clinical treatment with this agent is packaged for Phase III
trials. However, what remains to be determined with Lym-1 is how quickly the
studies can be pushed to front-line status, in the following sense: (1) DeNardo
mentions that it is likely that patients can respond to repetitive rounds of
Lym-1 therapy since relapsed patients retain expression of the Lym-1 antigen,
which is encouraging. This is being done by the Bexxar group (using the CD20
antigen) and there has been considerable discussion regarding this strategy in
light of the short responses (or none) to continued chemotherapy (or different
chemotherapy) by M. Winn, BobLLL and Berblady that has been interesting and
quite informative. (2) How does one start needed studies on combination therapy
(chemo + Lym-1) and what is the optimal schedule, dose and timing of that form
of therapy? Finally, how does one put together an informative, knowledgeable
and meaningful discussion of comparing Bexxar, Rituxan and Lym-1 together? It
is extremely tedious and laborious to sort out the nuances of the clinical studies
of different groups.
Interesting clinical case study that outlines in detail
how therapy with Oncolym goes in the clinic. As brief as I can make
it...patient diagnosed in 1985 with NHL, underwent 6 cycles of CHOP chemo to
complete remission...relapse in 1986...another year or so on chlorambucil and
prednisone...poor reponse then progression...9 cm scalp mass, cervical node
involvement...occipital mass, etc. May, 1988, underwent three courses of Lym-1
with progressive shrinkage of masses...(quickly developed human anti-mouse
antibodies, HAMA, final comment on this later)...Jan, 1989 had two rounds of
localized radiation therapy to resolve sclertotic abnormality on skull
x-ray...went 7 years with no evidence of disease...recurred in abdomen in Dec
96. Tough disease, aggressive therapy name of the game). Couple of interesting
clinical points. Within one month of initial Lym-1 treatment, patient had high
titer of HAMA (which may preclude further therapy with mouse antibodies in some
instances...but "humanized" antibody hybrid molecules are overcoming
that...)...but they manipulated the plasma levels of HAMA sufficiently to have
three cycles of Lym-1 treatment. What was of interest...besides the patient
making an immune response against the mouse antibody (Lym-1)...the patient also
made antibodies against the targeting sequence of the Lym-1...in other
words...an additonal family of antibodies that may have actually helped in
targeting lymphoma cells. This may have also been a factor in the 7-year
disease free interval. For those who want to appreciate the clinical course of
NHL and therapy...this article reads easily, and is found in Cancer Biotherapy
& Radiopharmaceuticals 13: 1-12, 1998, entitled, "Prolonged survival
associated with immune response in a patient treated with Lym-1 mouse
monoclonal antibody", by SJ Denardo, et al.
12/28/1998
Maximum-Tolerated Dose, Toxicity, and Efficacy of
131I-Lym-1 Antibody for Fractionated Radioimmunotherapy of Non-Hodgkin's
Lymphoma
By Gerald L. DeNardo, Sally J. DeNardo, Desiree S. Goldstein, Linda A. Kroger,
Kathleen R.
Lamborn, Norman B. Levy, John P. McGahan, Qansy Salako, Sui Shen, and Jerry P.
Lewis
Purpose: Lym-1, a monoclonal antibody that preferentially targets malignant
lymphocytes, has induced remissions in patients with non-Hodgkin's lymphoma
(NHL) when labeled with iodine 131(131I). Based on the strategy of
fractionating the total dose, this study was designed to define the
maximum-tolerated dose (MTD) and efficacy of the first two, of a maximum of
four, doses of 131I-Lym-1 given 4 weeks apart. Additionally, toxicity and
radiation dosimetry were assessed.
Materials and Methods: Twenty patients with advanced NHL entered the study a
total of 21 times. Thirteen (62%) of the 21 entries had diffuse large-cell
histologies. All patients had disease resistant to standard therapy and had
received a mean of four chemotherapy regimens. 131I-Lym-1 was given after Lym-1
and 131I was escalated in cohorts of patients from 40 to 100 mCi (1.5 to 3.7
GBq)/m2 body surface area.
Results: Mean radiation dose to the bone marrow from body and blood 131I was
0.34 (range, 0.16 to 0.63) rad/mCi (0.09 mGy/MBq; range, 0.04 to 0.17 mGy/MBq).
Dose-limiting toxicity was grade 3 to 4 thrombocytopenia with an MTD of 100
mCi/m2 (3.7 GBq/m2) for each of the first two doses of 131I-Lym-1 given 4 weeks
apart. Nonhematologic toxicities did not exceed grade 2 except for one instance
of grade 3 hypotension. Ten (71%) of 14 entries who received at least two doses
of 131I-Lym-1 therapy and 11 (52%) of 21 total entries responded. Seven of the
responses were complete, with a mean duration of 14 months. All three entries
in the 100 mCi/m2 (3.7 MBq/m2) cohort had complete remissions (CRs). All
responders had at least a partial remission (PR) after the first therapy dose
of 131I-Lym-1.
Conclusion: 131I-Lym-1 induced durable remissions in patients with NHL
resistant to chemotherapy and was associated with acceptable toxicity. The
nonmyeloablative MTD for each of the first two doses of 131I-Lym-1 was 100
mCi/m2 (total, 200 mCi/m2) (3.7 GBq/m2; total, 7.4 GBq/m2).
J Clin Oncol 16:3246-3256.
Address reprint requests to Gerald L. DeNardo, MD, Molecular Cancer Institute,
1508 Alhambra
Blvd, Room 3100, Sacramento, CA 95816; Email [email protected].
Interesting correspondence (J. Clin. Oncol. 16: 3916,
1998): We have discussed before the theoretical possibility that sustained
treatment with antiCD20 (and presumably other antigen directed antibody approaches)
could eventually select for the emergence of a subpopulation of lymphoma cells
that would be negative for the expressed intact CD20 antigen target molecule.
Such a case has been reported (cited above). Japanese group report that 5
months after Rituximab antibody therapy, relapse with lymphoma-positive bone
marrow revealed variant lymphoma population that was CD20, presumably
precluding further therapy with Rituximab. This has been reported before (J.
Clin. Oncol. 16: 1635, 1998). Interestingly, when one speaks of retreatment of
patients and an overall 40% response rate...about half of the patients do not
respond (due to unknown mechanisms). Now, this report leads to a clue, i.e., a
reduction, mutation, or loss of the target antigen on the relapse lymphoma
cells. Certainly, this could happen with Lym-1 also, but I do not know of
retreatment data at the present time. Theory in biology usually means it will
happen, given enough observations...
01/05/1999
Provided by a colleague: (edited), filed on UPI
under Health Notes.NEW BRAIN CANCER THERAPY: Each year, 15,000 Americans are
diagnosed with terminal brain cancer. Now, there's a glimmer of hope for some
of them, thanks to a new targeting agent called Tumor Necrosis Therapy.
Developed by Techniclone Corp. and undergoing Phase II human clinical trials,
the therapy allows doctors to deliver high concentrations of radiation directly
to the tumor's core through the use of a low-pressure intra-tumoral catheter,
essentially killing the tumor from the inside out while potentially causing no
damage to healthy surrounding tissue and with minimal adverse side effects. Of
the six malignant glioma brain tumor patients who underwent Phase I trials, all
showed stabilization of the tumor and three actually had tumors that shrank
despite receiving doses that were 1/3 to 1/2 of what is considered to be an
effective dose to treat this disease. In the past, victims diagnosed with this
form of brain cancer were faced with two treatment options, both of which have
proven ineffective over the long term. If the tumor is operable, doctors are
only able to remove the bulk of the tumor, leaving behind microscopic
``tentacles'' which ultimately results in re-growth. External treatments such
as radiation and chemotherapy have always been limited in their effectiveness,
since doses are regulated in order to minimize damage to the surrounding brain
tissue. TNT, however, may prove in clinical trials to severally reduce the
amount of toxic agents necessary to kill the tumor by directly targeting its
core, says Larry Bymaster, Techniclone's CEO. ___ By LIDIA WASOWICZ - UPI
Science Writer=
01/08/1999
This is a remarkable document. Thorpe is a
first-class scientist. While I understand the science contained within the
granted patent, I am unable to entirely grasp the protection it may afford TCLN
regarding this type of methodology. It is important to discern between what
might be argued as comprehensive in a scientific discussion and what may be
argued in a legal battle over patent rights regarding the delivery of reagents
to a targeted vascular bed of blood vessels within a tumor. Hence, I am not
able to speak of what could be challenged or undone. However, in my opinion, if
one contemplates the delivery of nearly any reagent used in the diagnosis,
imaging or therapy of a vascularized tumor by an antibody-based technology, no
matter the form that antibody may be constructed (genetically engineered or
produced monoclonal, or even induced polyclonal); then Thorpe's patent has it
covered. No matter the target antigen on the endothelial cell that at least I
can envision, if one wants to target that molecule (assuming that it is
preferentially displayed in tumor endothelium), then the patent has it covered.
It is comprehensive and complete in that respect. In lay terms, the delivery of
any agent via an antibody construct to tumor blood vessels belongs to TCLN
according to how I read the granted claims of this patent. It seems
straight-forward. TCLN knows how to write a patent.
Now, what does this mean in the world of
anti-angiogenesis (the ability to prevent or eradicate the blood supply of a
growing tumor)? The inhibition of endothelial cell division, or migration of
those cells, or their ability to form proper tubules within a tumor would
inhibit tumor growth. Hence, regarding these properties, there may be several
expressed properties and receptors of endothelial cells that could be exploited
as therapeutic targets. The most obvious and well-known molecules, angiostatin
and endostatin (ENMD) inhibit the growth of endothelium by a manner not
entirely characterized, but these molecules are not delivered to the blood
vessels by antibody or similar vehicles, hence they remain outside the scope of
this patent in my opinion. Not addressed in this patent is the targeting of the
well-known VEGF-receptor being addressed by companies such as Genentech or
Chiron, but small molecular-weight compounds that target the enzymatic
properties of this receptor would seem not to fall inside this patent. In other
words, the world of anti-angiogenesis is filled with companies attempting to
evaluate new kinds of molecules that have specific molecular targets and these
molecules are NOT delivered by immunologically-based devices, such as
antibodies or antibody-constructs. Examples include angiostatin-endostain
fusion protein (Genetix); combretastatinA4 (OxiGene), Flt-1 soluble receptor
(Merck); Kringle5 (Abbot); 2-methoxy-estradiol (ENMD), carbohydrate-RG8803
(Repligen); PD-173074 kinase inhibitor (Parke-Davis); S-836, a integrin
antagonist(Monsanto); small molecular antagonists, TBC series (Tex Biotech
Cor); Angiozyme, a catalytic destroyer of VEGF-receptor RNA (Ribozyme Pharma);
NX1838 antagonist of VEGF (Nexstar); SU5416, an inhibitor of the VEGF-receptor
(Sugen); ZD101, an endotoxin that may stimulate an immune response against
blood vessels (Zeneca); AG3340, a matrix enzyme inhibitor (Agouron, Phase III);
Marimastat, similar inhibitor (Brit Biotech, Phase III);...and on and on. Many
of these molecules have not yet made it into clinical trials. But the list of
compounds being synthesized and evaluted in preclinical animal studies is
growing. However, the antibodies against the VEGF-receptor and cadherins being
pursued by ImClone could be sticky with respect to this patent. But that's for
lawyers.
Finally, outside of the scope of this patent
are approaches championed by Ruoslahti who demonstrated in a series of elegant
studies the ability to select phage (bacterial viruses) that preferentially
bind to endothelium in tumor sites and has used these phage with attached drugs
to kill tumors in mice. There are efforts to construct peptides with particular
amino acid sequences (kind of a combination biochemistry synthetic
antibody-type molecule) that bind to target molecules on endothelial cells and
these peptides could be argued not to be of an immunological delivery system,
i.e., not fashioned using variable regions of immunoglobulin genes. In
addition, there are a number of studies attempting viral-mediated delivery of
genes and reagents to endothelium that could be argued to fall outside the
scope of this patent, in my opinion. Certainly Thorpe has done a magnificent
job with his studies, and the scope of this patent with respect to protection
of the VTA technology as envisioned by TCLN. Remember, everything I have said
is pure academic opinion which is a lot different than business-based
competition or the currency of lawyers.
regards.
01/09/1999
Cancer Treatments
Gee, doccg, you are a brave person to ask me
about expressing my views on an approach to cancer treatment. Let me extend
that question to an excuse to give the following views:
Malignant disease: Cancer is a collection of heterogeneous diseases that
follows an unchecked genetic, and heritable change in the DNA of a single cell
that results in unregulated proliferation of that cell and its daughters until
the mass/metabolism of the primary tumor or its metastasis shuts down normal
function of vital organs resulting in death. The cancerous changes that occur
(e.g., leukemias; immune cells; sarcomas, of connective tissue and bone;
carcinomas, the common solid tumors of gastric, colon, kidney, liver, breast,
prostate) may be unique to the tissue type and perhaps to the genetic
susceptibility of the individual. For example, loss of the p53 suppressor gene
(the normal function of this gene is to regulate just how many times a cell can
undergo cell division without being called into question about its motives for
continually dividing) is known to occur in about 40-50% of carcinomas that are
aggressive, highly metastatic, autonomous from tissue regulation and deadly.
Other examples of “suppressor” genes include for instance, the retinoblastoma
gene, RB, the breast cancer set, BRCA1 and 2; and other less common or
mentioned genes. Of course, on the other side of the coin is the activation of
tumor-inducing genes (which functionally inactivate suppressor genes), which
can occur by genetic alteration of environmental insults (tobacco carcinogens,
hydrocarbon-damage, UV induced damage, etc). Suffice it to say, once the
malignant transformation of a single cell occurs, forces are set into motion
that determine the destiny of the patient...does that cell survive which such
… (?message truncated?)
Therapies: Everything is based upon the
premise that cancer cells have metabolic properties that are unique or abundant
compared to normal tissue and therefore are more sensitive to toxic agents
designed to kill dividing cells. Name of the game. Kill dividing cells. Kill
relatively more cancer cells than normal cells. Over the past 3 decades, most
chemotherapeutic regimens have been based upon exploiting natural biochemical
pathways known to be essential to dividing cells and are somewhat sensitive in
tumor cells. That’s why chemotherapy is eventually limited by normal cell
toxicity (GI tract, bone marrow; neuronal); our natural need to replenish vital
cell populations involved in the everyday housekeeping functions of our body
and maintain normal cell function (platelets, leukocytes, mucosal cells of the
GI tract, functions of neurons) ultimately limit our ability to poison our
bodies with drugs designed to kill anybody that’s moving. Hence, for purposes
of this discussion, chemotherapy will always fail. Always. Why?
1. Because drug-limiting toxicity is of normal tissue, not cancer.
2. Because the pressure of drug administration always selects drug-resistant
cells out, even though this phenomenon is highly sensitive to variables of dose
and schedule in the administration of drug.
3. There are sites that harbor tumor cells that may not allow penetration or
proper metabolism of the chemotherapeutic drug, thereby sparing dormant tumor.
Therefore, the continued search for toxic agents is starting to require very
stringent conditions in order to improve upon our toxic therapy available
today. Let me explain. If you discovered a compound that interfered with DNA
synthesis in a very simple way (say, inhibition of a required, though boring
enzyme), chances are your therapy of cancer with that compound would quickly
reach unmanageable doses because even the normal cells that divide in our body
require that everyday, boring enzyme, therefore, your new drug is not very
effective; because it did not have a “therapeutic index” of being more
important to the cancer cell than the normal cell. That’s why. Trust me. I am
eventually getting to your question, doc.
Modern “chemotherapy” is attempting to exploit specific metabolic pathways that
may be very, very sensitive and required and relied upon by cancer cells; yes,
these pathways are found in normal cells, but not to the same sensitivity. The
tumor may have left itself vulnerable, because it needed to synthesize more of
a particular enzyme, or express more of a certain receptor than its
corresponding normal cell in order to metastasize to a particular organ and
survive (remember, the cancer cell is always “out of place”; a feature to
mention later). Therefore, new medicinal chemistry of cancer is very much
centered upon inhibition of over-expressed molecules (figuring that if the
cancer cell didn’t need so many of these proteins, they wouldn’t express so
many). Turns out these overexpressed proteins are very important to continued
proliferation by the cancer cells, the so-called growth factor receptors, the
kinases, the enzyme machinery of rapidly dividing cells, they are now the
targets of the new “molecular therapy”. These inhibitors are in clinical
trials; they do not have quite the classical toxicity of ragged chemotherapy,
but we need to see if they can eradicate the tumor. What new molecular tricks
will tumor cells play to always produce a progeny capable of circumventing the
ability of us to turn off their vital pathways. There will be alternate
pathways the cells will invoke (select for daughter cells that have this
property); or inactivation pathways of our targeting agent...etc. We just don’t
really know yet.
Eradication of cancer (theory): The stuff of
ordinary, but very bright research people at the bench that have little
interest in the stock market, but just do good and imaginative work everyday.
Well, you can carry the comments of chemotherapy into radiation therapy very
quickly and come to similar conclusions. Radiation therapy (external beam of
photons or particles) has deleterious side effects that eventually render this
modality of suspect effectiveness. I have also mentioned several times over the
past months about the possibility that toxic drugs and radiation delivered by
targeted vehicles (such as antibody; Rituxan, Bexxar, lym-1) can be eventually
limited by the expression of the target antigen by malignant cells, or the
penetration of these agents into privileged areas of the body that may harbor
cancer cells but not allow entrance of large molecular weight antibodies. I am
particularly enamored by the delivery of molecules to the vicinity of the tumor
by the TNT-generation of antibodies and fusion peptides for reasons thoroughly
discussed before; because of the exploitation of a natural necrosis that occurs
in the vicinity of a rapidly growing tumor mass by the TNT targeting and the
ability of such targeting to induce inflammatory changes that, in my opinion,
are favorable to subsequent host defense responses against the tumor.
In my opinion, based upon my experience in preclinical studies of cancer,
curative procedures will only be obtained by ultimate recognition of the
malignant cell by a competent immune system capable of mounting an effective
immune response against the neoplasm, or by systemic initiation of a cessation
of cell division and induction of a terminal differentiation of cancer cells,
who have suddenly been induced to realize that they are part of a programmed
tissue that no longer needs to divide. I see no other potential therapeutic
interventions that can achieve permanent regression of malignant tissue. The
first modality, immunotherapy, has been attempted in cancer for over 30 years
with very limited success and we are coming to realize that the differences we
have been looking for (the BIG foreign antigen that makes the cancer cell an
immunological monster easily recognized and eradicated) are not there...the
changes are very subtle, but perhaps noticeable. As our knowledge of classical
immunology expands, (especially transplantation immunology, our creative juices
in the area of cancer immunology improves). There are several viable approaches
today that are very promising. The second property, that of programmed normal
behavior is rapidly evolving...and in an interesting fashion. Gene therapy is
actually based upon the premise that one can somehow restore normal cell
function to the cancer cell by replacing the known defective genetic elements
that have gone astray (say, that p53 gene mentioned earlier; put in a new
one!). Yet, it is tough to imagine how we are gonna deliver genes to big tumor
masses or scattered small metastases throughout the body. There are thoughts,
but no concrete or convincing approaches to that yet. We can envision agents
that induce new pathways, kind of an embryo-approach...that says to the cancer cell,
picture yourself as a liver cell, or colon cell, or well-behaved lung cell,
please. The cells stop dividing, become quiescent and as mannered as a
boarding-school freshman.
02/03/1999
Fusion peptides: Need to discuss some
properties of two cytokines; interleukin-2 (IL-2) and interferon-gamma (FN-g),
which are molecules released by lymphocytes that have a myriad of
immune-stimulating properties and are very important in antitumor responses.
IL-2 has been used (with modest success) in a variety of cancers, but has a
deleterious side effect that limits its use, namely, vascular-leak syndrome,
where leakage of the vascular compartment takes place into tissues. This is
because IL-2 causes an increase in the permeability of endothelial cells to
water (either by direct or indirect mechanisms); and this syndrome is
life-threatening. IFN-g activates a variety of immune cells which aids in
antitumor activity and also causes tumor cells (and other immune cells) to
express new levels of needed cell-surface molecules that assist in destruction
of tumor. Now, one other facet of this strategy must be mentioned, the TNT
antibody molecule; which is an antibody directed to nuclear antigens found in
necrotic regions of growing tumor and is the basis of TNT-therapy with which we
are familiar.
Epstein has taken advantage of the exquisite
localization of TNT antibody to the region of tumor by considering also the
local delivery of IL-2 or IFN-g and at the same time, exploited that side
effect of IL-2, i.e., its effect on local permeability of endothelial cells.
Now, antibodies are formed by the linking of several protein molecules due to
gene rearrangement prior to their synthesis. The antibodies are secreted by
lymphocytes and this synthesis of the TNT antibody (to which the isotope I-131
can be added is toxic to cells in the vicinity of its binding, like in
treatment of glioma) can be duplicated in the laboratory in expression systems
that artificially make tons of antibody of against a specific target. Hence,
the TNT therapy machine. However, by genetic engineering, we can take the
molecular targeting of the TNT antibody (to vicinity of tumor) and hook up
genes that will also attach the IL-2 (or IFN-g) molecule directly to the
antibody. Now we have one antibody-interleukin protein, a so-called fusion
peptide, since it is due to the “fusion” of two distinct gene-products normally
not related.
Epstein now proposes (and he has good funding
for these projects) to synthesize these fusion-peptides to cause localized
changes in the tumor microenvironment that will aid in other types of
therapies. The I-131 is no longer attached. For example, the TNT/IL-2
constructed peptide will cause localized permeability changes in the vicinity
of the tumor and allow easier delivery of other agents that might have a tough
time penetrating the blood vessels (endothelial cells) to get inside the tumor
mass. These agents might include chemotherapeutic drugs normally used in cancer
therapy or larger molecules, such as other types of antibodies or related
agents. On the other hand, the fusion-peptide containing IFN-g may allow
localized upregulation of antigens on cells that aid in immune responses. Only
the creative imagination of the scientist in this setting limits the
possibilities of new types of therapeutic approaches using fusion-peptides of
different constructs. This is the essence of this technology and I view it as
imaginative (Epstein is a very gifted scientist) and filled with possible new
treatment regimens. Of course, the test is in the clinic. But he has published
about preclinical models. When I have time I can post some of those references.
02/03/1999
The TNT antibody currently in use for glioma
(and I suppose the Mexico trials in other solid tumors) uses an attached radioactive
isotope, I-131 as the source of toxicity to the tumor cells. The antibody
attaches to the displayed antigens offered by the tumor region, and the
accompanying radiation kills tumor cells within that region. Now, you may be
confusing the use of Oncolym (which is an antibody also containing the
radioactive isotope used against a lymphoma antigen in NHL), and Rituxan, the
IDEC antibody against lymphoma cells which does not carry a radioactive
molecules attached to it, because it can mediate tumor cell destruction by a
different mechanism. TNT antibody and Oncolym antibody are two different
antibodies against two different target antigens found on different tumors.
chTNT-3 (next generation) can be used to deliver a variety of attache
goodies...kind of in a payload fashion, with the docking end the antibody that
recognizes the good ole TNT antigen. Not related to patent, since it is not
directed to endothelial cell antigens...(without getting into complex legal
interpretations).
Epstein just used the radioactive thymidine molecule as a tracking drug to show
that IF he had used real chemotherapeutic drug (the thymidine was just used in
a radioactive fashion so he could show how it was taken up in tissue)then the
use of the chTNT enhanced uptake of the drug by about 3-fold.
Hope this was helpful...
02/03/1999
Pretreatment with a monoclonal antibody/interleukin-2
fusion protein directed against DNA enhances the delivery of therapeutic
molecules to solid tumors. Clin Can Res 1999
Hornick JL, Khawli LA, Hu P, Sharifi J, Khanna C, Epstein AL
Department of Pathology, University of Southern California School of Medicine,
Los Angeles 90033, USA.
[Medline record in process]
The efficacy of molecular therapies for human malignancies is limited by
inadequate accumulation within solid tumors. Our laboratory has developed a
novel approach that uses monoclonal antibodies (MAbs) to direct vasoactive
proteins to tumor sites to increase local vascular permeability and, in turn,
improve the delivery of therapeutic reagents. Previously, we demonstrated that
pretreatment with immunoconjugates containing interleukin-2 (IL-2) enhances
specific tumor uptake of radiolabeled MAbs without affecting normal tissues. In
the present study, we describe a fusion protein consisting of a chimeric
antinuclear antibody and IL-2 (chTNT-3/IL-2) and illustrate its potential for
improving the delivery of both MAbs and drugs. The ability of pretreatment with
chTNT-3/IL-2 to increase specific tumor uptake of the MAb B72.3 was
demonstrated in LS174T colon tumor-bearing mice. Tumor accretion of B72.3
increased nearly 3-fold, with no changes in normal tissues. Abrogation of this
effect with N(G)-methyl-1-arginine, a chemical inhibitor of nitric oxide
synthase, suggests that rapid generation of nitric oxide in the tumor is
responsible for the enhanced uptake. To demonstrate that pretreatment with
chTNT-3/IL-2 can improve the uptake of other clinically relevant MAbs in
different tumor models, additional studies were performed in both lung and
prostate xenograft models. Pretreatment with the fusion protein increased
specific tumor uptake of the MAb NR-LU-10 in A427 lung tumor-bearing mice and
enhanced tumor uptake of the MAb CYT-351 in LNCaP prostate tumor-bearing mice,
2.1-fold and 1.7-fold, respectively. Finally, tumor uptake of the radiolabeled
thymidine analogue 125IUdR also increased approximately 3-fold after
pretreatment, indicating that this approach can be extended to small molecules
such as chemotherapeutic drugs. Because TNT-3 recognizes a universal nuclear
antigen accessible in degenerating and necrotic cells within all solid tumors,
this strategy may be applicable to the majority of human cancers.
02/05/1999
I have mentioned that one of the strengths of the
DeNardo's is their incessant penchant for refining methodologies associated
with radioimmunotherapy with monoclonal antibodies. This recent paper
(Dec)demonstrates their interest and potential use of yittrium-labeled Lym-1 by
use of new methodology. Productive and cost-effective...good biotech terms.
This paper is of limited clinical interest to readers here, but I offer it as
evidence that DeNardos and the lym-business goes on...
J Nucl Med 1998 Dec;39(12):2105-10
Optimized conditions for chelation of yttrium-90-DOTA immunoconjugates.
Kukis DL, DeNardo SJ, DeNardo GL, O'Donnell RT, Meares CF
Department of Internal Medicine, University of California Davis Medical Center,
Sacramento, USA.
Radioimmunotherapy (RIT) with 90Y-labeled immunoconjugates has shown promise in
clinical trials. The macrocyclic chelating agent
1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid (DOTA)
binds 90Y with extraordinary stability, minimizing the toxicity of 90Y-DOTA
immunoconjugates arising from loss of 90Y to bone. However, reported 90Y-DOTA
immunoconjugate product yields have been typically only < or =50%. Improved
yields are needed for RIT with 90Y-DOTA immunoconjugates to be practical.
METHODS: (S) 2-[p-(bromoacetamido)benzyl]-DOTA (BAD) was conjugated to the
monoclonal antibody Lym-1 via 2-iminothiolane (2IT). The immunoconjugate
product, 2IT-BAD-Lym-1, was labeled in excess yttrium in various buffers over a
range of concentrations and pH. Kinetic studies were performed in selected
buffers to estimate radiolabeling reaction times under prospective
radiopharmacy labeling conditions. The effect of temperature on reaction
kinetics was examined. Optimal radiolabeling conditions were identified and
used in eight radiolabeling experiments with 2IT-BAD-Lym-1 and a second
immunoconjugate, DOTA-peptide-chimeric L6, with 248-492 MBq (6.7-13.3 mCi) of
90Y. RESULTS: Ammonium acetate buffer (0.5 M) was associated with the highest
uptake of yttrium. On the basis of kinetic data, the time required to chelate
94% of 90Y (four half-times) under prospective radiopharmacy labeling
conditions in 0.5 M ammonium acetate was 17-148 min at pH 6.5, but it was only
1-10 min at pH 7.5. Raising the reaction temperature from 25 degrees C to 37
degrees C markedly increased the chelation rate. Optimal radiolabeling
conditions were identified as: 30-min reaction time, 0.5 M ammonium acetate
buffer, pH 7-7.5 and 37 degrees C. In eight labeling experiments under optimal
conditions, a mean product yield (+/- s.d.) of 91%+/-8% was achieved,
comparable to iodination yields. The specific activity of final products was
74-130 MBq (2.0-3.5 mCi) of 90Y per mg of monoclonal antibody. The
immunoreactivity of 90Y-labeled immunoconjugates was 100%+/-11%. CONCLUSION:
The optimization of 90Y-DOTA chelation conditions represents an important advance
in 90Y RIT because it facilitates the dependable and cost-effective preparation
of 90Y-DOTA pharmaceuticals.
TNT
Interesting case study presented 4 years ago at ASCO.
Response of hormonally refractory prostate cancer (HRPC) to TNT-1 monoclonal
antibody (Meeting abstract).
Proc Annu Meet Am Soc Clin Oncol. 13:A742, 1994.
Abstract
Treatment options for HRPC are severely limited with responses to current
chemotherapy regimens usually being of short duration. TNT-1 is a murine
monoclonal antibody which has been shown to bind specifically to degenerating
and dead cells of human cancers (Epstein et al, Cancer Res 48:5842-8, 1988).
The antibody recognizes an antigenic domain of nuclear histones which is
expressed in all cancers and normal tissues studied to date. Inadequate blood
supply and impaired phagocytic responses within the tumor favor the
accumulation of degenerating cells adjacent to viable areas of the tumor. In
contrast, normal tissues have a relatively low rate of cell death and are
characterized by a rapid and orderly removal of necrotic elements. We report
successful treatment with TNT-1 MoAb in a 47 year old man with HRPC with prior
treatment that included orchiectomy, flutamide, suramin, radiation therapy and
doxorubicin. Within 36 days after the administration of 50 mg of biotinylated
TNT-1 followed by 20 millicuries of 131I-streptavidin given weekly times 2, the
PSA declined from 210 to 4.9 and the PAP from 6.5 to 1.2. The patient was
clinically improved and remained stable for 10 months having received only
TNT-1 for 2 such treatment cycles. Moderate thrombocytopenia and leukopenia
with nadirs at day 36 after treatment were noted. Further clinical evaluation
of TNT-1 against a broad spectrum of tumor types is now being considered. (C)
American Society of Clinical Oncology 1997
Granted, we are not out of the first generation TNT
clinical studies, but looking ahead is always a good distraction. Epstein
published a paper last year (Radiochemica Acta 79: 83-86, 1997) on initial
preclinical studies using TNT-IL2, the often mentioned fusion peptide
construct. As we know, TNT is an antibody that targets specific DNA-associated
proteins within necrotic regions of tumor, and this target rarely, if ever,
mutates as do other molecules of cancer cells, thus making these proteins
valuable for docking onto and delivering toxic molecules to the
microenvironment of cancer. The fusion construct is the antibody genetically
combined with an active cytokine, called IL2, which has two major effects (1)
promotes activation and receptor expression of immune cells against tumor and
(2) causes tremendous increase in local permeability of blood vessels allowing
blood-borne molecues to infiltrate the tumor. Hence, targeting the TNT-IL2
molecule to tumor causes the uptake of drugs to be increased about 4-fold over
the use of a placebo antibody...and significantly improves the therapeutic
index while causing no increase in toxicity or uptake in normal tissues over
that observed as normal. More delivery to tumor. This approach is where the
next ideas will be carried out...and represents further potential value in the
product platform of TCLN,
TNT mAb is directed against a nucleohistone antigen
exposed in degenerating cells; and the past and ongoing literature is contained
in (a) Hornick, et al, A chimeric TNT/IL-2 antibody fusion protein as a
universal pretreatment to enhance the delivery of therapeutic molecules to
solid tumors, ProcAACR, 1997; (b) Khawli, et al, Effect of seven new vasoactive
immunoconjugates on the enhancement of monoclonal antibody uptake in tumors,
Cancer 73:, 1994; (c) Khawli, et al, Chemical modification of monoclonal
antibodies; ProcAACR, 1994; (d) Khawli, et al, Enhancement of chemotherapetuic
index by a monoclonal antibody vasoconjugate, ProcAACR, 1994; (e) Strum, et al,
Response of hormonally refractory prostate cancer (HRPC) to TNT-1 monoclonal
antibody; ProcASCO, 1994; (f) Miller, et al, Immunolgoci and biochemical
analysis of TNT-1 and TNT-2 monoclonal antibody binding to histones; Hybridoma
12:, 1993, (f) Epstein, et al, Tumor necrosis treatment of human cancers;
ProcAACR, 1992; (g) Chen, et al, Effects of 131-I-labeled TNT-1
radioimmunotherapy on HT-29 human colon adenocarcinoma spheroids, Can Immunol
Immunother, 33:, 1991; (h) Gaffar, et al, Cell based radioimmunoassays to
quantitate the immunoreactivity of TNT monoclonal antibodies directed against
intracellular antigens, J. Immuno 12:, 1991; (h) Cheng, et al, Diffusion and
binding of monoclonal antibody TNT-1 in multicellular tumor spheroids J. Natl
Cancer Inst 83:, 1991, (i) Chen, et al, Localization of monoclonal antibody
TNT-1 in experimental kidney infarction of the mouse, FASEB J 4:, 1990; (j)
Chen, et al, A comparative autoradiographic study demonstrating differential
intratumor localization of monoclonal antibodies to cell surface (Lym -1) and
intracellular (TNT-1) antigens, J Nucl Med 31:, 1990; (k) Chen, et al, Tumor
necrosis treatment of ME-180 human cervical carcinoma model with 131-I-labeled
TNT-1 monoclonal antibody, Cancer Res 49:, 1989
An encouraging aspect of the TNT Phase I studies being
done at South Carolina is that these studies will be shifted to Phase II
involving 4 medical centers (probably including M. Berger's department at
UCSF). Recalling the news release of Oct 1 (which tend to get lost), TCLN will
recruit 1/2 of the patients that have newly diagnosed glioblastoma or
anaplastic astrocytoma and the other half from patients with recurrent disease
(it almost always recurs). Dosage will be individualized to each patient, (two
doses, 8 weeks apart). Phase II endpoints are efficacy (response) as determined
by stabilization of disease and time to disease progression as well as
continued study on safety and tolerability. Remember, they noted antitumor
responses in all of the initial Phase I patients, even at low doses...but I
have not heard any follow up on those patients. Whether Patel found a way to
make it to the program of the Neuro-Oncology meeting is unknown at this
time...but the secretary of the meeting is here at the Tex Med Center, I will
try to contact her and found out more.
VEA
December 9, 1996 -- Techniclone Corporation
(NASDAQ: TCLN), a biotechnology company engaged in the research and development
of drug delivery systems based on monoclonal antibodies announced today that a
team of scientists from the University of Southern California and the Mayo
Clinic have published an article discussing the combined use of Techniclone’s
patented LYM-1 and Vasopermeation Enhancement in preclinical (animal) studies.
The Company’s Director of Scientific Affairs, Alan L. Epstein, M.D. Ph.D. and
Peter M. Anderson, a member of Techniclone’s Scientific Advisory Board, were
part of the group who published the article.
The article, published in Cancer Research, November 1, 1996, indicated that
Vasopermeation Enhancement technology and administered as a pretreatment,
enhanced the uptake of radiolabeled LYM-1 antibody specifically in the tumor by
an average of 300%. They concluded that Vasopermeation Enhancement increased
the tumor’s permeability, suggesting that this combination may be a new form of
therapy which could be used in humans.
Authors of the article include Peishing Hu, Jason L. Hornick, Michelle S.
Glasky, Aoyun Yun, Mary N. Milkie, Leslie A. Khawli, and Alan L. Epstein, all of
the University of Southern California Department of Pathology, and Peter M.
Anderson of the Mayo Clinic Section of Pediatric Hematology/Oncology.
"This new fusion molecule utilizing our Vasopermeation Enhancement
technology appears to be several times more effective than LYM-1 alone
(presently in Phase III clinical trials), and reflects the efforts of our
scientists to continuously test and improve the Company’s product
pipeline" said CEO Lon H. Stone.
02/05/1999
At the risk of turning this forum into a library, we have
discussed the next generation TNT molecues, i.e., the fusion peptides, where
cytokines or advantageous molecules are genetically linked into the formation
of the TNT antibody. Here is an older paper that describes early work by Epstein
a fusion-peptide with the LYM-1 antibody, the basis of current Oncolym therapy
for NHL.
It does show one that if sufficient funding were available, and clinical trials
are extremely expensive; it would be very interesting to pursue these kind of
"hidden" pipeline methodologies, that probably won't see the light of
day for awhile. TCLN is supporting very bright people, IMO, and the ideas now
available from genetic modifications of the tumor microenvironment are moving
very fast in the drug-discovery business; both academically and within biotech
industry.
Cancer Res 1996 Nov 1;56(21):4998-5004
A chimeric Lym-1/interleukin 2 fusion protein for increasing tumor vascular
permeability and enhancing antibody uptake.
Hu P, Hornick JL, Glasky MS, Yun A, Milkie MN, Khawli LA, Anderson PM, Epstein
AL
Department of Pathology, University of Southern California School of Medicine,
Los Angeles 90033, USA.
A murine antihuman B-cell monoclonal antibody, Lym-1, has shown considerable
promise for the treatment of human malignant lymphomas. To enhance its clinical
potential, a genetically engineered fusion protein consisting of a chimeric
Lym-1 (chLym-1) and interleukin 2 (IL-2) was tested for mediating cytotoxicity,
increasing vasopermeability, and enhancing antibody uptake in human malignant
lymphomas. The chLym-1/IL-2 fusion protein, which was expressed initially in a
baculovirus system and more recently in the glutamine synthetase gene
amplification system, was shown to be processed and assembled into a normal
immunoglobulin monomer with two IL-2 molecules per antibody. It was found to be
equivalent to the chLym-1 antibody in antigen-binding specificity and relative
affinity. In addition, it maintains IL-2 cytokine activity as demonstrated by
support of T-cell proliferation. Moreover, in antibody-dependent cellular
cytotoxicity assays against Raji target cells, chLym-1/IL-2 had approximately
2-fold and 4-fold higher cytotoxicity than chLym-1 and murine Lym-1,
respectively. Used as a pretreatment, chLym-1/IL-2 enhances the uptake of
chLym-1 at the tumor site by altering the permeability of tumor vessels
producing tumor:normal organ ratios of 420:1 for blood and 1708:1 for muscle at
3 days. The in vitro and in vivo activities of chLym-1/IL-2, therefore, suggest
that this genetically engineered antibody fusion protein may represent a new
immunotherapeutic reagent for the treatment of human malignant lymphomas.
Q. Regarding
the chLym-1/IL2 fusion protein that you refer to in the 1996 paper...do you
think that this sort of approach will someday make it to clinical trials and
possibly eventually improve upon the current Oncolym? If so, roughly how close
to clinical trials might it be?
Regarding fusion peptides in general, does anyone hold patents? are many
scientists working on this approach? and how would the patents work? would
separate patents be issued for specific peptide combinations, or is the entire
"fusion of proteins" idea patentable?
Similarly, re the latest Epstein paper on chTNT-3/IL-2,if it is eventually
determined that this approach improves upon current TNT (already in trials), do
you still have to start trials from the beginning, or can you make
modifications to a trial already in progress?
A. Your questions are thoughtful and
insightful, unfortunately for me, they are also difficult to answer. Giving
opinions during seminars or classes or at lunchroom discussions (which can be
heated) is not quite as intimidating as talking out loud on an internet chat
board. Hence, I believe I need to qualify my remarks as much as possible.
Regarding your first question about possible use of
chLym/IL-2 fusion peptides. Certainly I believe the approach will be used in
the clinic; whether it will be that particular combination of targeting/attached
cytokine is entirely unknown. One parameter involves the therapeutic use of
Lym-1 which is iodinated with a radioactive isotope and that is its source of
cellular toxicity (the emission from the isotope). The chLym/IL-2 was used to
target regions of tumor with subsequent changes in permeability of the vascular
bed and is a different form of trying to enhance the therapeutic efficacy of
other agents, perhaps drugs or delivery of antibodies.
Short answer, no, I do not believe that particular combination is close to
clinical trials, especially with the ongoing studies in progress. But I do
stress the idea bank is there.
Patents on fusion-peptides. Wow, what a lawyers
delight! That’s my qualification on this one...I would have no idea of the
patent messes one could get into, but I’ll bet they ain’t pretty. I will say
that with regards to fusion-peptides that target vascular cells by means of a
constructed antibody attached to some neat protein that would alter the
properties of the endothelium in that region...that is covered (IMO) by
Thorpe’s patent. Put a little more scientifically...the construction of
fusion-peptides by means of the utilization of immunoglobulin genes with
specificity toward endothelial antigens (no matter there presence, induction or
ultimate expression) would be covered under the Thorpe patent. Other
combinations of protein constructs would not be covered, IMO.
Finally, your last question regarding chTNT/IL-2
seems as complex as the first one. Tell you why. An analogy is the approved
agent, Rituxan from IDEC. Since it is FDA-approved, it can be prescribed under
conditions or combinations that may not have been ideal or optimal for its
original intended use. IL-2 (free-form, as a protein in solution) is approved.
It can be used in a variety of ways. Hence, their combination in different
clinical settings is easy to do. But it is not easy to combine agents where one
remains an experimental reagent. Regulations may require going back and looking
at parameters that you don’t believe are necessary, but may have to do.
Radiolabeled chTNT is a therapy molecule, but chTNT/IL-2 is a permeation
enhancer using the target of TNT as a local place to dock the molecule. I
realize my answers are not very enlightening, but escalating clinical trials
and using combinations that make perfect sense can always be bogged down by
layers of committees that exist all the way from the hospital to Washington.
02/07/1999
Previous post demonstrated earlier paper on the construction
of a Lym-1/IL-2 fusion peptide; the targeting antibody sequence against NHL
cells. The paper cited below involves the Lym-2 antibody, produced against
chronic lymphocytic leukemia cells, and two cytokines; GM-CSF, a product of
bone marrow macrophages and other cell types which aids in the maturation of
immune cells and causes infiltration of cells into lesions (and by the way,
activates macrophages to produce an enzyme which cleaves a serum protein to
yield the famous angiostatin. A good cytokine. Also studied is the fusion of
Lym-2/Il-2.
Blood 1997 Jun 15;89(12):4437-47
Chimeric CLL-1 antibody fusion proteins containing granulocyte-macrophage
colony-stimulating factor or interleukin-2 with specificity for B-cell
malignancies exhibit enhanced effector functions while retaining tumor
targeting properties.
Hornick JL, Khawli LA, Hu P, Lynch M, Anderson PM, Epstein AL
Department of Pathology, University of Southern California School of Medicine,
Los Angeles 90033, USA.
Although monoclonal antibody (MoAb) therapy of the human malignant lymphomas
has shown success in clinical trials, its full potential for the treatment of
hematologic malignancies has yet to be realized. To expand the clinical
potential of a promising human-mouse chimeric antihuman B-cell MoAb (chCLL-1)
constructed using the variable domains cloned from the murine Lym-2 (muLym-2)
hybridoma, fusion proteins containing granulocyte-macrophage colony-stimulating
factor (GM-CSF) (chCLL-1/GM-CSF) or interleukin (IL)-2 (chCLL-1/IL-2) were generated
and evaluated for in vitro cytotoxicity and in vivo tumor targeting. The
glutamine synthetase gene amplification system was employed for high level
expression of the recombinant fusion proteins. Antigenic specificity was
confirmed by a competition radioimmunoassay against ARH-77 human myeloma cells.
The activity of chCLL-1/GM-CSF was established by a colony formation assay, and
the bioactivity of chCLL-1/IL-2 was confirmed by supporting the growth of an
IL-2-dependent T-cell line. Antibody-dependent cellular cytotoxicity against
ARH-77 target cells demonstrated that both fusion proteins mediate enhanced
tumor cell lysis by human mononuclear cells. Finally, biodistribution and
imaging studies in nude mice bearing ARH-77 xenografts indicated that the
fusion proteins specifically target the tumors. These in vitro and in vivo data
suggest that chCLL-1/GM-CSF and chCLL-1/IL-2 have potential as
immunotherapeutic reagents for the treatment of B-cell malignancies.
01/27/2000
Cancer Res 1998 May 1;58(9):1952-9
Vascular endothelial growth factor as a marker of tumor endothelium.
Brekken RA, Huang X, King SW, Thorpe PE
Cell Regulation Program, Department of Pharmacology, University of Texas
Southwestern Medical Center, Dallas 75235-9041, USA.
Vascular endothelial growth factor (VEGF) is an angiogenic growth factor that
is a primary stimulant of the vascularization of solid tumors. VEGF production
is induced by oncogenic gene mutations in the tumor cells and by hypoxic
conditions inside the tumor mass. Hypoxia and the locally increased
concentration of VEGF lead to an up-regulation of VEGF receptor expression on
tumor endothelial cells. Therefore, in the tumor microenvironment, there is an
up-regulation of both VEGF and its receptor, leading to a high concentration of
occupied receptor on tumor vascular endothelium. The VEGF:receptor complex
presents an attractive target for the specific delivery of drugs or other
effectors to tumor endothelium. In the present study, several hybridomas that
secrete monoclonal antibodies against the VEGF:receptor (Flk-1) complex or
against VEGF itself have been raised. Three of the antibodies (3E7, GV39M, and
11B5) bind with high affinity to the VEGF:Flk-1 complex in ELISA and to tumor
endothelium in frozen sections of human tumors, rodent tumors, and human tumor
xenografts. 3E7 and GV39M localize selectively to tumor endothelium after i.v.
injection into mice bearing human tumor xenografts. Additionally, one antibody
(2C3) was raised that blocks the interaction between VEGF and KDR/Flk-1. 2C3
inhibits VEGF-mediated growth of endothelial cells in vitro and localizes
strongly to connective tissue in tumors after injection into mice bearing human
tumor xenografts. These findings suggest that 3E7, GV39M, and 2C3 are
candidates for targeting and imaging the vasculature or connective tissue of
tumors.
Interesting twist the anti-VEGF thrust is
taking...from many angles and corporate headings. IMCL has the IMC-1C11, an
antibody targeted against the KDR receptor (I still believe the use of this
antibody is in direct conflict with the Thorpe VTA patent), SUGN has the SU5416
which blocks the activation of the KDR receptor and now, the new Thorpe derived
antibody, 2C3 which blocks the binding of VEGF to its receptor on tumor
endothelium. These strategies, combined with the recent SUPG intent to license
from Techniclone a VEGF-ligand construct to target tumor endothelium
demonstrates the importance of this particular growth factor in the growth of
abnormal, inflammatory-type blood vessel cells.
A question which has yet to be addressed in the science of tumor vasculature,
as well as abnormal behavior of endothelial cells in other areas, such as
macular degeneration and coronary artery inflammation that often follows
angioplasty, as well as plaque formation is exactly how the membrane properties
of these cells, expressing their newly formed VEGF receptors is different than
normal cells that line blood vessels. Exploiting these differences is the grist
for the mill of therapeutic approaches that are innovative with respect to
molecular targeting of today's modern view of anticancer therapy. Certainly
other target antigens, such as the expression of lipid molecules (addressed by
Thorpe in recent studies) will come into play.
Knowledgeable people have told me that the VTA patent may cover distinct
properties of "newly" formed vessels, compared to "old"
vessels found inside the tumor, but I have not seen this addressed in the
patent applications or issued patents. Other knowledgeable people have said that
"just about anything that can be targeted to tumor endothelial cells is
covered by the Thorpe studies".
How this can be partitioned out in licenses remains to be seen. Some may try to
get it all, and at an unreasonable price or conditions with the company. ES and
colleagues may hold their cards close to the vest as the financial conditions
improve. Perhaps we can enter an era where we are have chips on the table in
plain view, and the other players can not just bet pot-limit and take us out of
the hand. We've been there, done that...but no more.
01/28/2000
Q. The use of TNT as an imaging tool has
been discussed many times. This latest addition to our patent coverage brings
up my question.
When used as a imaging technology, I'm assuming that lab trials will be done
and biopsies performed after imaging to determine the precision of the read
out. This process should be quite short as the biopsy can be performed
immediately after the imaging. Hundreds of images and biopsies can be taken
over a period of days/weeks. If successful, would the technique still have to
be performed on humans to obtain FDA approval? Also, would this fall under IND
or Medical Device in seeking approval? The use as an imaging tool seems to me
to be something that could be brought to market in a very short period of time.
A. Well...just my opinion, since I have no
experience in this end of the science. I would think that imaging doses of
labeled TNT would be far less than any therapeutic dose, and would fall into
the categories of agents such as contrast dyes, radionuclides like Tc which is
used to light up bone metastasis, and Prostascint, things like that. Hence, I
would look for quick studies on efficacy of this kind of application with a
rather quick FDA process.
Of course, this topic brings up another quirk that gripes me...see, Terry, this
is another topic that could be discussed publicly, in an exciting fashion by
the company in a news release...
Something like..."New Applications for Vascular Targeting
Discussed..."..etc. We just never seem to do so. Chew (remember him???)
ought to be thinking about these corporate-minded issues...(duh,
HELLLLLOOOOOOOOOO ASCO??????????).
regards.
01/28/2000
Very recent patents awarded to Thorpe.
Certainly relevant to the patent estate of TCLN. Appear to be very specific
claims regarding targeting to endothelial cell antigens.
Issued October 12, 1999. 5,965,132: Methods and compositions for targeting the
vasculature of solid tumors. Describes the delivery of diagnostic or
therapeutic agents via antibodies specific for the antigen formed by the
conjugate of a growth factor AND its receptor (as opposed to targeting either
one alone), including the induction of target antigens on endothelium.
Issued Dec 21, 1999. 6,004,554: Methods for targeting the vasculature of solid
tumors. A clever notion of selectively suppressing certain antigens found on
normal endothelium but inducing these antigens on endothelial cells within the
tumor bed. Then antibody mediated targeting and delivery of diagnostics and
therapeutics to the induced target antigen.
Issued Dec 21, 1999. 6,004,555. Methods for the specific coagulation of
vasculature. Method of delivery clotting inducing ligands to the
microenvironment of the tumor vasculature by a bispecific antibody construct
that binds both a cell antigen and localizes the coagulation factor resulting
in a clotting cascade that infarcts the tumor bed.
At the time of issue, these three patents were assigned to UT Texas. Certainly
a broad-base repertoire of once again, innovative vascular targeting.
03/06/2000
Phil Thorpe gave a talk at the very prestigious
Keystone conference this weekend in Salt Lake City. "Targeting Coagulants
to Tumor Vasculature"..."We have induced major tumor regressions in
mice bearing large solid tumors with immunoconjugates that destroy or occlude
the vasculature of solid tumors. The immunoconjugates employ antibodies that
recognize molecules that are selectively expressed on vascular endothelium in
solid tumors. The molecules either occur naturally, (e.g., VCAM-1 or
E-selectin) or can be induced on tumor endothelium (e.g., MHC Class II). The
antibodies are linked to the extracelluar domain of the human coagulation
initiating protein, tissue factor. Truncated tissue factor is a soluble protein
that is virtually devoid of coagulant activity while free in the blood
ciruclation, but becomes a powerful and specific thrombogen after binding to
the tumor endotheial cell surface. Induction of coagulation by the targeted
tissue factor requires exposure of phosphatidylserine (PS) on the endotheial
cell surface. PS exposure is itself confined to tumor endothelial cells in
mice. The need for coincident expression of the target marker and PS further
restricts the action of the targeted tissue factor to tumor vessels. These new
drugs have the potential for widespread application in the treatment of solid
tumors.
03/31/2001
The annual meeting of the AACR focuses upon
progress of older studies that continue down their natural evolution of
knowledge and occasional wildcards of new, exciting fronts of basic science
that lends itself rapidly to therapy (translational research). The overview I
offer here is a result of my own interest and expertise and other researchers
may have differing and argumentative views. I will start with very general
notions of tumor biology and end with specific thoughts as to the science of
cancer therapy.
The problems of identifying unique characteristics of cancer cells that can be
subsequently exploited for therapy remain somewhat unchanged. A brief review is
probably relevant, in order to assess where current research is headed. The
heterogeneity of tumor cell populations is well established and, for the most
part, the therapy of cancer is the therapy of metastatic disease. What makes therapy
so particularly difficult it two-fold, (a) the more we uncover with regard to
the genes and their products of both normal and cancerous cells the more we
have to unravel intellectually about the constant interplay between metabolic
pathways of cells that ultimately determine sensitive set-points of survival or
cell death; and (b) the inherent genetic instability of tumor cells results in
permutations, amplifications and redundancies that are difficult to attack.
It seems clear that tumor cells are able to subvert homeostasis, capture the
advantageous elements within the tumor microenvironment and through their own
evolution of selection for anonymous subpopulations of cells endowed with
unique characteristics, cancer cells are showing us their ability to mutate and
alter their environment in a manner that assures continued growth. Some
investigators would argue that the focus should be entirely upon tumor blood
vessels and their particular properties since the expanding tumor mass
ultimately depends upon a viable blood supply, but I suspect this will be shown
to be not sufficient. I imagine we will find that the properties of the tumor
vasculature are constantly being modeled and shaped as a result of intimate
communication that takes place between tumor cells and their blood supply.
Current thinking about direct therapy is
aimed squarely at the tumor cells (as always) and the blood vessels of the
tumor. Keep in mind there is a probably a great difference between trying to
"prevent" metastatic growth of a tumor and the direct treatment of
established metastases or primary tumor, even with regard to the properties of
the endothelial cells. Most hard information at the present time is coming from
studies of established disease, even in the animal models. Current efforts seem
to center upon the knowledge about what tumor cells need to do in order to
sustain signaling for continued growth, induce a blood supply and make it all
work in a constantly changing tumor mass. The secretion of growth factors such
as bFGF, VEGF, EGF, TGF-alpha, IL-8, IL-6 and the expression (and secretion) of
a variety of metalloproteases (MMPs), together with the loss of certain
adhesion molecules and the upregulation of a vareity of genes that promote
drug-resistance, avoidance of programmed cell death events (such as the Bcl-2,
bax family of genes), and activation of cell cycle pathways (p53 mutations,
PTEN alterations, CDK genes) all make for a hungry, aggressive population of
cells ready to take on the battles of host response and intervention. In
addition, tumor cells upregulate and express a variety of growth factor
receptors that can be auto-stimulated and the signaling from these receptors
activate survival pathways for the cells. The question becomes, in the
research, how to categorize and make sense of the vulnerabilities of the tumor,
taking into account the needs of normal tissue limits to our assaults.
Progress in identification of unique molecular expressions of tumor cells and
endothelial cells from solid tumors is being made with respect to the
"chip array" technology. DNAchips, RNAchips, CD-expression arrays and
proteomics (still mainly limited to mainstream 2-dimensional gel techniques)
are showing a multitude (too many) of genes and proteins that are somewhat
unique to the tumor with respect to normal tissue around the tumor, or of
endothelial cells isolated from tumor. Unfortunately, the mass of information
is not easily translated to bench experiments about whether the identified
proteins are viable targets for therapy. Yet, the wealth of information (say in
identifying that certain growth factors go up, certain receptors are present,
certain molecules are being secreted) lends itself to putative targeting for
therapy. With respect to direct therapy that results from the knowledge of
reading the 10,000 genes off of a DNAchip, we ain't there yet. Most of the data
are about genes we already are targeting. But the information is coming.
Modern therapy still is focused upon 4 main
areas, (a) direct cytotoxic agents aimed at tumor cells, especially specific
genetic defects of tumor cells, (b) direct inhibition of tumor blood supply,
(c) immunotherapy and (d) gene therapy. I shall write little about gene
therapy, due to the limitations of trying to insert "corrective"
genes into a tumor mass (not convincing) and the clinical troubles resulting
from Pennsylvania studies about a year ago, most gene therapy is at a halt.
Let's work somewhat backwards…
Immunotherapy (my own field of experience) remains a mysterious realm of intrigue
and heart-breaking experience that has persisted for three decades. Modern
techniques of dendritic cell isolation and "loading" with tumor
antigens is very promising, but seems limited at the moment to tumors that we
knew were somewhat recognized by the immune system, such as melanoma and renal
cell carcinoma. Good progress is being made about the requirements of
antigen-presenting cells that result in a demonstrated immune response against
tumor, but these studies are fraught with modulation due to the presence of an
overwhelming amount of immune cytokines within tumors (interleukins 2, 4, 5, 6,
10, 13, IL-1, TGF-beta, TNF-alpha, GMCSF, for example) and various leukocyte
populations that have defects and can be immunosuppressive; we still wait for
some definitive insight and breakthrough experience in this area.
Tumor cell cytotoxics are taking two paths; (a) more and more toxic agents that
just plain kill cells (such as the CPT-11, gemcitabine, etc) and (b) agents
that target the growth factor requirements of tumor cells. We can not ignore
(a) in the context of (b)…and I will explain.
It is well-established that targeting of growth factor receptors (such as the
EGF-receptor, PDGF-receptor, VEGF-receptor) on both tumor cells and endothelium
that make up the blood vessels of tumor cell masses (more on that later), is a
fast-moving, aggressive field of current therapy. The obvious examples are the
Herceptin antibody directed against EGF-r on breast tumor cells, C225 directed
against the same...and in various stages of clinical trials and several
antibodies against the VEGF-receptor. The other strategy is the use of small
molecule tyrosine kinase inhibitors against the growth factor receptors, such
as the marvelous STI571 against the mutated PDGF-r in chronic myelogeneous
leukemia (it seems there are variant subpopulations emerging in some of the
"cured" patients that have further mutated this kinase and is not
targeted by STI571). These tyrosine kinase inhibitors seem to come out of the
woodwork…besides the known SUGN compounds, now Novartis has a couple of very
promising agents, (PKI166, PTK787) that inhibit the growth of tumor cells by
blocking the activation of their growth factor receptors.
Now, here is where the twist on these agents become very interesting. There
must be 25 new tyrosine kinase inhibitors indentified (we only need a few,
excuse me) that target the common growth factor receptors of EGF, VEGF and
PDGF. However, some new ones are against what is known as downstream signals
(into the areas of what is called MAP kinases, NFkappaB, CDKinases) which are
exciting new targets, but I suspect not much better than hitting the initial
receptor. Let me say this another way…when the needed survival and growth
factor, such as EGF (or its analog, TGF-alpha) binds to the receptor on tumor
cells (or some endothelium) it promotes cell division, stabilization and
survival. To target that receptor is excellent…but one can also go after those
events that take place inside the cell after the EGF bound the receptor…in
other words, the "downstream" events. These cascades are complex,
redundant and yet new targets can be found, many involving kinases of a
different variety. A lot of studies have identified these targets, and there
were at least three full sessions devoted to new tyrosine kinase inhibitors.
Let's talk tumor vasculature, before we
combine the cytotoxic agents and the growth factor receptor inhibitors, for
common threads are now being observed. It is obvious to anyone who reads of
modern tumor therapy that inhibition of angiogenesis (the formation of a blood
supply) is the exciting feature of cancer research. Exciting because of the
hope and promise of taking away the blood supply of tumor, resulting in a
collapse of tumor growth and sustained dormancy of remaining tumor cells…and
perhaps the inability of tumors to induce the needed blood vessels for growth
of other tumor deposits not actually detected (occult metastatic disease).
When one considers the physiology of the formation of a new blood supply, one
finds that this event (say, in normal wound healing or menstruation, or
embryogenesis), the migration or differentiation of endothelial cells that form
the new vessels undergo an extemely complex set of moves comprised of cell
division, migration, tubular formation, establishment of cell-cell contacts and
eventual stabilization into blood vessels that have functional viability. It
seems clear that tumor blood vessels differ with respect to other tissue blood
vessels with regards to the expression of cell-surface molecules, such as
integrins, collagen products, matrix proteins, growth factor receptors,
angiopoietins, lipids and a variety of different genes (identified through the
chip arrays mentioned before). This makes them "targetable" and forms
the basis of studies and clinical trials using inhibitors of angiogenesis, such
as interferon-alpha, angiostatin, endostatin, 2ME2, combretastatin,
thalidomide, specific integrin antagonists, antibodies directed against markers
on endothelial cells and perhaps even low-dose chemotherapy. Make no mistake
about it…this is THE area of aggressive preclinical and clinical studies, the
potential of targeting the blood supply of tumors. Pasqualini and her
colleagues have published exciting work in the area of phage-identification of
endothelial cell targets, which are a whole new set of potential proteins found
on exclusively on tumor endothelium that may serve as viable targets for
shutting down the blood supply of tumors.
However, the situation becomes more complex
(sort of a "but wait, there's more"). It turns out that many of the
same growth factor targets found on tumor cells are also found on tumor
endothelium. The implication of this is as follows…suppose a tumor requires the
expression of the EGF-receptor for optimal growth and one observes that an
antibody or tyrosine kinase inhibitor of that receptor inhibits directly the
tumor growth. Well, in addition, it has been observed that the tumor
endothelial cells are likely to upregulate their receptors for EGF also, and
the inhibitor becomes a TWO-pronged attack, both tumor and vasculature.
Moreover, preclinical studies at AACR showed that if one treated tumors (in
animals) with BOTH cytotoxic agents (such as CPT-11, taxol or gemcitabine) PLUS
the growth factor inhibitors, one obtains superb inhibition of tumor growth
(and metastasis) compared to the use of either agent alone. This is not a
trivial observation. It suggests that the optimal attack on tumor, at the
moment, remains the use of cytotoxics combined with antiangiogenic agents. I
doubt that single agent use of pure antiangiogenic compounds will achieve near
the effect as combination therapy with standard cytoreductive therapy.
An additional note on tumor vasculature leads us to the PPHM VTA targeting…that
is, the propensity of tumor beds to have a "coagulytic" environment.
Essentially this means that the vessels found within tumor masses have
properties conducive to coagulation, a process found normally only in the
immediate environment of a wound or repair process. They are "setting on
the edge" with exposed molecular structures that are optimal for one juicy
blood clot if saturated with the right agents, such as Tissue Factor. This is a
unique and unexplored area (except for Thorpe and a couple of groups) that is
ripe for harvest. The transition to this subject will lead me to comments of
the PPPH-related abstracts found this year at the AACR.
Abstract from Epstein group.
Recombinant Truncated Tissue Factor/Rgd Fusion Protein as a Target Anti-Vascular
Therapeutic Agent
Peisheng Hu, Qing Zhou, Thomas Bai, John Shariff, JiaLi Li, Alan Epstein,
University of Southern California, Los Angeles, CA.
Rapidly proliferating tumors require an efficient blood supply to meet their
nutritional needs in both primary and metastatic diseases. Tumor vasculature is
a particular suitable target for cancer therapy because it is composed of
nonmalignant endothelial cells and those cells are genetically stable. In this
study, truncated tissue factor fusion protein chTNT-3/tTF, chTV-1 and RGD/tTF
have been constructed. Truncated tissue factor (tTF) is a 25 KD soluble protein
whose membrane binding domain has been deleted. It has the factor VII
activating capacity of its parent protein. Monoclonal antibodies chTNT-3 and
chTV-1create a binding domain which targets the thrombogenic capacity of the
tTF to the tumor vasculature, therefore initiating coagulation and occlusion of
blood flow within the tumor. The glutamine synthase gene amplification system
was used for statement of these two fusion proteins. A motif contains sequence
Arg-Gly-Asp (RGD) has been identified integrin primarily expressed in tumor
vasculature, RGD/tTF has been inserted into pQE60 and expressed in Top 10. The
statement of chTNT-3/tTF and chTV-1/tTF has been done by standard ELISA assay,
and Coomassie Plus Protein Assay Reagent Kit has been used for RGD/tTF. Animal
studies showed that RGD/tTF had been completely removed 12 hours after
injection, biodistribution study demonstrated that tumor has higher uptake of
fusion protein compared with other organs, and tumor had obviously necrosis due
to the treatment. These results showed that tTF linked to monoclonal antibody
and RGD may be effective immunotherapeutic reagent for the therapy of cancer.
Comment: [This study demonstrates that not only can the truncated tissue factor
protein be targeted to the endothelial cells within a tumor, and results in a
choking of the tumor blood suppy in the tumor microenvironment, but that this
molecule can be hooked to the TNT family of antibodies which are highly
specific for the tumor cells and give rise to the same phenomenon. This finding
expands the concept of targeting the tumor vasculature to include effects from
also targeting antigens found within the tumor cells. Hence, the ARCUS venture
of VTA can include the TNT antibody as well as Thorpe lab-derived targeting
molecules. Advantages include the rapid clearance of this targeting antibody
and tTF construct from normal tissue, and the specific localization within
tumor compared to normal tissue.]
In Vitro and in Vivo Studies of VEGF/RGEL
Fusion Toxin Targeting Tumor Vascular Endothelium.
Hangqing Jin, Sophia Ran, Philip Thorpe, Michael G. Rosenblum, The University
of Texas, MD Anderson Cancer Center, Houston, TX; UT Southwestern, Dallas, TX.
Vascular endothelial growth factor (VEGF) is an angiogenic growth factor which
binds to two
structurally related tyrosine kinase receptors denoted KDR/FLK-1 and FLt-1.
Both KDR/FLK-1 and Flt-1 are overexpressed on the endothelium of solid tumor
vasculature. Therefore, the receptors for VEGF appear to be unique targets for
the development of therapeutic agents that may interfere with tumor growth and
metastatic spread through inhibition of tumor neovascularization. We created a
fusion construct of VEGF121 and the plant toxin gelonin (rGel) and the
resulting VEGF121/rGel fusion protein demonstrated the biological activities of
both VEGF and rGel components. Porcine Aortic Endothelial cells were
transfected with either flt-1 (PAE-flt) or KDR/flk-1 (PAE-KDR) and utilized to
examine whether KDR/FLK-1 or Flt-1 play a role in cytotoxicity of VEGF121/rGel.
Log-phase PAE-KDR cells were ~900 fold more sensitive to VEGF121/rGel compared
to rGel (I.C.50 of 0.5 nM vs 450 nM respectively). The PAE-flt cells
demonstrated no selective sensitivity to either the fusion toxin or free rGel
(I.C.50450 nM). Western analysis demonstrated increased auto-phosphorylation of
the KDR receptor after exposure to either VEGF or VEGF121/rGel. Dose-response and
time courses for this effect were identical for both agents. As expected,
neither VEGF nor VEGF121/rGel were able to induce autophosphorylation of flt-1
expressing cells. These data suggest that the KDR receptor is primarily
responsible for development of VEGF121/rGel cytotoxicity. Studies to examine
the binding, internalization and intracellular routing of VEGF and VEGF121/rGel
after binding to the two receptors are ongoing. In nude mice bearing PC-3
prostate tumors, IV treatment (q 48h X 5) with total doses up to 20 mg/kg
results in impressive suppression of tumor growth compared to controls.
Pharmacokinetic studies of the VEGF121/rGel fusion construct will also be
reported. These data suggest that this construct has significant potential for
clinical use in reduction or prevention of metastatic spread.
Comment: [This is a "trojan" horse approach to vascular targeting, in
that the statement of this particular VEGF-receptor on tumor endothelial cells,
in contrast to "normal endothelium" makes the attachment of the toxic
compound (the gelonin) target only these type of cells in a specific manner.
Since the VEGF-construct will "seek out" the receptor on the tumor
endothelium, it will deliver this toxic payload to these cells and destroy
their ability to support a blood supply to the tumor. Animal data presented
that was not included in this abstract was a >85% reduction in growth of the
A375 human melanoma in nude mice, as well as >90% reduction in growth of the
PC-3 prostate cancer in nude mice. In fact, the prostate cancer seemed to be a
flat line of a very small tumor nodule as a function of time throughout the
study, with exponential growth of the control-treated tumor. This particular
therapy appears to be what SUPG has in mind concerning their interest in the
PPHM collaboration (see press release of 13 Feb 2001 regarding targeting the
VEGF receptor)]
Expression of VCAM-1, E Selectin and PSMA on
Human Normal, Malignant and Inflamed Vessels.
Sophia Ran, Maria Sambade, Philip E. Thorpe,
Southwestern Medical Center, Univ. Dallas at Texas,Dallas, TX; Southwestern
Medical Center, Dallas, TX.
An ideal marker for targeting tumor vasculature would be a luminal protein that
is expressed on a high proportion of tumor endothelium and absent from normal
vessels. Success of the targeting approach will ultimately depend on the extent
of a marker's statement, its consistency and favorable distribution of positive
vessels within the tumor vasculature. It should be also considered whether a
marker is present at sites of inflammation, tissue re-modeling and other
pathological non-malignant conditions that might be found in cancer patients.
We examined immunohistochemically the statement of three vascular markers of
human solid tumors: vascular cell adhesion molecule-1 (VCAM-1), E-selectin and
prostate specific membrane antigen (PSMA). It has been previously shown that
these markers are upregulated on the tumor vessels in various malignancies but
the comparative analysis of the pattern of their statement has not been done.
The staining of normal, malignant and inflamed human tissues with anti-VCAM-1,
E-selectin and PSMA antibodies was analyzed according to the following
criteria: a) the percentage of positive vessels compared to total, CD31-stained
vessels; b) the intensity of staining; c) the pattern of distribution of
positive vessels (focal, central, peripheral, patchy or random). The overall
vascular statement in malignancies was compared to that in normal and inflamed
tissues. PSMA was the most abundant and consistent marker, being present in all
12 different types of tumors studied. The percent of PSMA-positive vessels
ranged from 10% in lymphomas to 80% in renal and breast carcinomas. Normal
vessels were devoid of PSMA with the exception of endometrium, ovary and adrenals.
PSMA was also strongly upregulated at inflammatory sites, such as colitis,
cystitis and reactive tonsil. VCAM-1 and E-selectin were found in 9 and 7 out
of 12 tumor types, respectively. The proportion of positive vessels (5 to 50%)
was smaller than for PSMA. VCAM-1 was present on 5-10% of normal vessels in
kidney, thyroid and endometrium, whereas E selectin was completely absent from
normal organs. VCAM-1 also prevailed in the inflammatory sites while E-selectin
did not. This analysis suggests that: a) all three markers are upregulated by
inflammatory or angiogenic conditions; b) PSMA is an attractive marker because
of its widespread and consistent statement in various tumors; c) E-selectin is
an attractive marker because it is the most selective tumor endothelial cell
marker, and is moderately to strongly expressed in Hodgkin's lymphoma,
non-small cell lung carcinoma
and breast tumors.
Comment:[This study falls right into the lap of the Thorpe patent estate on
molecules uniquely targeted on tumor vasculature. This study is important for
in HUMAN tumors, they determined whether or not these molecules they want to
target, (E-selectin, VCAM-1 or PSMA) are widespread in inflammatory conditions
(which patients may have besides their growing tumor) and on normal tissue. The
attractiveness of the two molecules (PSMA and E-selectin) verifies the approach
Thorpe has taken for the VTA. Now, whether PSMA falls under the Cytogen claims
or not is a consideration for the company (?????) but the science is solid. Note
the number of tumor types (lymphoma, lung, breast) that carry these markers,
making it of broad applicability, as desired. This type of targeting can be
hooked to a variety of toxic compounds, or using the tTF (truncated tissue
factor)].
Angiogenesis as a Determinant of Tamoxifen
Response in a MCF-7 Breast Cancer Xenograft Model of Resistance
Rachel Schiff, C Kent Osborne, Philip E. Thorpe, Sophia Ran, Rolf A. Brekken,
Irma Parra, Syed K. Mohsin, Suzanne A. Fuqua, Baylor College of Medicine,
Houston, TX; The University of Texas at Dallas, Dallas, TX; The Hope Heart
Institute, Seattle, WA.
Angiogenesis is known to be important for the prognosis of breast cancer --
cancers with high microvessel counts are associated with increased recurrence
and poorer outcome. Less is known, however, about angiogenesis as a predictive
marker in breast cancer. Pilot clinical studies suggest that high vessel counts
are associated with a reduced response to the antiestrogen tamoxifen. We
therefore utilized a xenograft tamoxifen-resistance (TR) model to examine gene
statement profiles associated with the development of the resistant phenotype.
Using Clontech cDNA arrays, we found that several angiogenic factors and
receptors for these factors were changed in statement as resistance developed.
Among these, total VEGF was reduced in the TR tumors, while a specific potent
isoform, VEGF121, increased. Flt-1, a VEGF receptor, was also found to be
upregulated in the TR tumors. Thus, we hypothesize that VEGF121 and its
receptor may be important determinants of resistance. To test this hypothesis
we are targeting the VEGF receptor complex to reverse or delay the development
of resistance in the xenograft model. Interestingly, when microvessel density
was determined using an antibody to CD31, we did not see a major increase
during the development of resistance. This suggests that these
angiogenesis-related markers may have roles beyond direct stimulation of
microvessel density in the development of tamoxifen resistance in breast cancer.
Comment: [The purpose of this study was to target the VEGF-receptor within
human breast cancer growing in nude mice. This study confirms the possibility
that VEGF itself is a "survival" factor for new blood vessels within
a tumor (some studies have shown that anti-blood vessel effects from some
compounds can be abrogated by the presence of EGF or VEGF). Can we actually
take advantage of the presence of this survival factor? Modern technique of the
DNA array shows that molecular analysis of the tumor can lead to choosing
important molecular targets…nice modern approach]
06/05/2001
There are opportunities for the company to
demonstrate scientific excellence and progress in the preclinical and clinical
evaluation of their approaches to the therapy of cancer. There are periods of
time where there is nothing much that can be said (for instance, during the
treatment and follow-up phase of a clinical trial where one is waiting on
parameters of response that can be measured) and sometimes there are rapid
opportunities such as society meetings, conferences, licensing deals and
significant management decisions that can be made available to the
shareholders. In my opinion, any legitimate opportunity to update the progress
of the technology in the area of research and clinical trials must be seized
and handled with professional savvy.
First, there must be an commitment by management to constantly demonstrate
visibility in the medical center, be it at the lab bench with preclinical
studies or at the bedside with trials of the new products under clinical
evaluation. In my opinion, there are no good strategies that involve silence
about the science or the clinic. Second, there is no place to hide in the
medical center. If a new therapeutic agent or procedure is good, really good,
people will know. Of course, one must have superb experimental design for the
studies, execution of the trials by knowledgeable and excellent physicians and
wisdom and proper insight about clinical parameters
With these introductory remarks, I find it
unacceptable that the company would not follow up their press releases
announcing presentations of preclinical studies at AACR and the Thorpe
presentation in Seattle. Moreover, I was encouraged by the ASCO data on glioma,
but left pondering exactly how the neuro-oncologists view the use of Cotara in
therapy of brain cancer. Let me explain this latter comment in detail.
Regarding the ASCO results from the Phase II brain cancer study: Newly
diagnosed or recurrent brain tumors have to be one of the most complex clinical
issues in oncology to evaluate. When one considers Phase I and Phase II studies
in glioblastoma (or anaplastic astrocytoma) the patient population can quickly
be obscure. The single clinical parameter discussed in the news release (which
is all we have available for interpretation, other than the abstract) was
median time to progression which was about 75% longer for treated patients
compared to "historical controls". Patel was quoted as being
"enthusiastic" regarding the potential of Cotara in treatment of both
newly diagnosed and recurrent malignant gliomas, even compared to
"standard therapy". What was NOT presented was an analysis that
allowed definitive conclusions and comparisons to be drawn in light of many
other Phase I/II studies in glioma. For instance, should one examine the
responses of patients with recurrent disease (many prior therapies?) in light
of only that specific patient population? I assume they did. What about the few
newly diagnosed patients? How did they do (no real mention of two types of
patient groups) compared to newly diagnosed patients that undergo other
"standard" therapies? What were the differences between patients that
received only one dose of Cotara versus two? What about the subsequent quality
of life, regarding neurological defects of brain cancer therapy, behavior and
well being? Is there a subset of glioma patients that benefit more from Cotara
that other patients, perhaps by virtue of their prior therapies, or initial
size of tumor, or features of the histological invasion of surrounding normal
brain? ? The competition at medical centers is usually pretty active among
clinical trials. Is this a treatment where Patel is enthusiastic enough to
attempt to "recruit" patients to Cotara?
There may be limitations to the use of Cotara in
brain cancer. The design of the Phase III trials may be restricted to patient
populations that are well defined in clinical parameters that we know will be
responsive to Cotara therapy in a fashion that offers a new viable option for
their treatment. The results may be encouraging enough to attempt other forms
of brain metastases such as those that occur from primary breast or lung
cancer. Without publication of these data and hard analysis that is expected in
the clinical community, that portion of the potential of this form of therapy
in brain cancer remains elusive to the public.
Now…why press releases announcing presentations
at AACR without any follow up? Perhaps they were about preclinical (animal)
studies that did not constitute "blockbuster" news. So what? It was
an opportunity to write about the progress of the science in the lab. I found
the presentations significant. I assume portions of these studies are supported
by the company; hence, they should have been presented to the shareholder. For
example, Thorpe's paper entitled "Recombinant Truncated Tissue Factor/Rgd
Fusion Protein as a Target Anti-Vascular Therapeutic Agent" involves
cutting-edge research on targeting the blood supply of tumors and should be in
clinical trials as soon as possible. This paper represented an opportunity to
talk about the PPHM/OXGN venture (Arcus), the timelines for potential trials,
the importance of this work and how it represents the modern thinking of
two-compartment targeting of cancer (the tumor cells and their blood supply).
The Thorpe and Rosenblum paper entitled, "In Vitro and In Vivo Studies of
VEGF-RGEL Fusion Toxin Targeting Tumor Vascular Endothelium" was an
excellent opportunity to update the findings of excellent reduction in tumor
growth in the animal studies and speak to the SUPG/PPHM alliance regarding the
use of VEGF targeting. Another opportunity missed. The Thorpe paper entitled,
"Expression of VCAM-1, E Selectin and PSMA on Human Normal, Malignant and
Inflamed Vessels" was an excellent chance to speak about newly identified
vascular targets that may be exploited by the scientists of PPHM and represent
future directions for clinical studies of targeting blood vessels of tumor.
These studies demonstrated forward thinking and expansion of earlier work by
Thorpe. Finally, the presentation by Rachel Schiff (Thorpe as co-author),
"Angiogenesis as a Determinant of Tamoxifen Response in a MCF-7 Breast
Cancer Xenograft Model of Resistance" spoke of potential targeting of the
VEGF-receptor on blood vessels, a potent and modern approach to combination
therapies of several different cancers in the clinic today. Unfortunately, we
chose to remain silent, not out of "strategy" but perhaps out of lack
of an aggressive philosophy to make the science of PPHM visible.
Finally, there was the Seattle presentation by Thorpe
(and Rosenblum) regarding a summation of the modern approaches to vascular
targeting, once again discussing the specific targets mentioned above, such as
VCAM-1, PSMA, VEGF receptors (and their complexes) and the lipid,
phosphatidylserine, which we will hear more about someday. Another wonderful
opportunity for a powerful follow up of a news release that got buried only
within the schedules of the shareholders with little visibility to the biotech
investor community that would be seeing the name of PPHM emerge out of the mass
of new attempts at cancer therapy.
It is disappointing to see theses opportunities go by. Now, later this month,
we have Cotara data out of Mexico City to be presented at the Society for
Nuclear Medicine. Once again we had the news release about the presentation of
this Phase I trial by Cesarman. I wonder what the follow up will be?
06/13/2001
AVN (via Xenerex) has interesting methodology
of generating human monoclonal antibodies to self-antigens. I wonder what
target(s) PPHM has in mind. Several recombinant DNA technologies exist
(probably all under patents)for the creation of antibody-fragments that can
bind to the antigens in question. I wonder if the VTAs will continue to be
created as Thorpe described in this paper from January.
Biotechniques 2001, Jan; 30(1): 190-194
Generation and characterization of recombinant vascular targeting agents from
hybridoma cell lines.
Gottstein C, Wels W, Ober B, Thorpe PE.
University of Texas Southwestern Medical Center, Dallas, TX, USA.
Vascular targeting agents (VTAs) can be produced by linking antibodies or
antibody fragments directed against endothelial cell markers to effector
moieties. So far, it has been necessary to produce the components of VTAs
(antibody, antibody fragment, linker, and effector) separately and,
subsequently, to conjugate them by biochemical reactions. We devised a cloning
and expression system to allow rapid generation of recombinant VTAs from
hybridoma cell lines. The VTAs consist of a single chain Fv antibody fragment
as a targeting moiety and either truncated Pseudomonas exotoxin (resulting in
immunotoxins) or truncated human tissue factor (resulting in coaguligands) as
effectors. The system was applied to generate recombinant immunotoxins and
coaguligands directed against endoglin, vascular endothelial growth factor
(VEGF):VEGF receptor (VEGFR) complex and vascular cell adhesion molecule 1
(VCAM-1). The fusion proteins exhibited similar functional activity to
analogous biochemical constructs. This is the first report to describe the
generation and characterization of recombinant coaguligands.
The Xenerex subsidiary almost seems a
"contract" facility to produce human monoclonal antibodies against
target antigens of the client's choice. Using the PR as a guide, it appears we
have chosen to enter a "collaborative" agreement with AVN regarding
the development of these antibodies. I imagine, from a business point of view,
it is a path that allows future payments against present success. That is…we
want them to produce human monoclonal antibodies against our selected target
antigens, which is an expensive process (and all methods are fraught with
patent infringement problems; including construction of fusion peptides), so we
enter a joint venture with them that may include milestones and rewards.
What are the target antigens? That's the neat question. Well, PPHM is a
vascular targeting company. With the exception of Lym-1 antibody (Oncolym when
radiolabeled with I-131) that targets leukemia cells and TNT which delivers
intracellular radiation (or, in theory, drugs) to the cell interior, the VTAs
of ARCUS and PPHM are seeking targets on the blood vessels of tumors. Many of
these targets are self-antigens.
Example: Prostate Specific Membrane Antigen (PSMA) is found on tumor
endothelium. Thorpe is very busy studying this protein as a target for vascular
targeting. But, PSMA is a self-antigen, meaning that since it is a protein that
we have circulating in our bodies all the time…we can NOT raise antibodies
against it (except using mice). Well, we don't want mouse antibodies for a
variety of reasons, so we need human antibodies to use to target the PSMA on
the cells of the tumor blood vessels, but the Xenerex technology knows how to
"break our tolerance" against this PSMA, and can produce human
antibodies against it. We can then make tons of the stuff and have a targeting
antibody. The same can be said for other antigens that Thorpe has identified on
the surface of endothelial cells that may be "self-antigens". Hence,
the AVN methods are very useful. It will be extremely interesting to see if a
couple of self-antigens, such as the phosphatidylserine complexes described in
his Cancer Research paper of last year can serve as targets. But Thorpe has
also described methods in the Biotechniques paper that I mentioned earlier
today. Only he and the big guns at PPHM know what avenue they are taking on
these studies.
AVN? Yes, a collaborator…but remember, with PPHM in charge and control.
Regards.
07/31/2001
Oakwood and PPHM
The simple view.
This joint venture with Oakwood Labs opens a new application of the TNT
antibodies in targeting. Liposomes (microscopic, spherical globules usually
constructed of synthetic lipids or fats) contain empty spaces that can be
loaded with all kinds of drugs, reagents, genes, toxins, and immunomodulators
to name a few. When injected intravenously, the liposomes are readily absorbed
and taken up by phagocytes in the blood, trapped in the liver and taken out of
the circulation in the lung and spleens. It is necessary to devise tricks in
the composition and targeting of the liposomes to get the specificity of
delivery to site of tumor cells.
Loading (attaching) antibodies specific for tumor cells and a variety of other
means can increase this specificity. Oakwood is working on a therapy of colon
cancer and need to have great specificity for the regions of tumor cells…since
the liposomes will not last long in the liver tissue. They have chosen to
partner with PPHM in targeting by using the TNT antibody, which is highly
specific for tumor tissue. Of course, this approach is not limited to colon
cancer mets in the liver…but is applicable to a wide range of solid tumors.
These preclinical studies should be extremely interesting for Oakwood and PPHM
and the general applicability of this technology will have implications for a
variety of compounds delivered in this manner.
This approach can be a very lucrative and productive form of targeted therapy.
08/31/2001
With respect to the license by PPHM of the
antibody technology that targets the cell surface lipid, phosphatidylserine
(PS), the interest in this particular target is as follows. The stimulation of
new blood vessel formation (which is continual in a growing tumor mass) and
even the inhibition of normal blood vessel formation (as when a wound has
healed) involves elements of the clotting-cascade, a complex set of serum
proteins that ultimately interact in various ways to form clots. Hence, the
blood vessels within a growing tumor are sometimes "thrombogenic",
that is, they are setting right on the edge of "wanting to clot".
About three years ago, Thorpe demonstrated that the targeting of truncated
Tissue Factor (tTF, one of those clotting proteins) to the tumor blood vessels
caused intratumoral clots, necrosis and inhibition of tumor growth. Part of the
excitement of this methodology is based upon the lack of clotting found in
"normal" tissues that could prove to be a clinical problem, such as
clots elsewhere in the body.
Now, why was this phenomenon of clotting limited to the tumor endothelium? The
answer seems to lie in the observations that (a) clotting requires a negatively
charged lipid surface to speed along the chemical reactions of these clotting
factors, and (b) the common lipid available for this function is
phosphatidylserine and (c) many tumor cells and damaged or inflammatory
endothelium express PS on their outer cell membrane.
He stained the tumorsof the mice with an antibody specific for PS and found
this lipid on the surface of tumor blood vessels, but not on the membranes of
normal blood vessels (and some very small tumors). Therefore, the success of
the tTF therapy may have relied upon the expression of PS. However,
conceptually, this means that PS itself becomes a potential target molecule for
directing specific intratumoral therapy, such as attaching tTF to the
antiphosphatidylserine antibody. I assume there must be interesting preclinical
data on this concept to spark the act of PPHM wanting to license this
conceptual targeting moiety.
Whether this approach falls under the VTA of Arcus versus the sole pipeline of
PPHM is a subject, I'm sure, of a much longer essay.
Will write about the 2C3 license at a later time.
09/01/2001
The 2C3 is the designation for an antibody
which targets the receptor for Vascular Endothelial Growth Factor (VEGF) on
tumor blood vessels. VEGF is a protein is released by tumor cells in abundance,
especially in areas of low oxygen or acidic pH, which is turn stimulates
"survival" factors inside of blood vessel cells. VEGF also causes
dramatic changes in the ability of blood vessels to allow nutrients across
their lumens; all of its actions seems to serve to bring oxygen and nutrients
and stimulate new blood vessel formation. The twist of this seemingly important
target is that VEGF is important for normal cell function with different cell
populations in the body. However, the receptors for this protein can be fairly
selective, and the 2C3 antibody targets the one (KDR/flt-1) found on blood
vessel cells within the tumor.
This "vascular targeting" by 2C3, that is, the ability to prevent the
actions of VEGF on the endothelial cells by binding of the antibody to the
receptor, and thus blocking the VEGF has resulted in impressive preclinical
studies published from the Thorpe lab.
The licensing of this antibody from the UT System in Dallas is somewhat
confusing, since TCLN licensed this methodology in 18 months ago (see PR dated
27 Jan 2000) and J. Bonfiglio commented on it about a year ago on the lab
studies (see PR dated 15 Sep 2000). The statement at that time, was "We
are planning to develop it as a stand-alone anti-angiogenesis inhibitor and
with our joint venture partner, OIGENE, we are examining its use as a Vascular
Targeting Agent." This statement seems vague, at best, about how they are
going to develop this therapy. I do not understand what this "new"
licensing is about.
As a side note, therapies that have been directed against growth factor
receptors, such as the VEGF-receptor (which include others, such as the PDGF
receptor, the EGF-receptor [IMCL C225]) invariably are best when used in
combination therapies, such as being combined with chemotherapy and
irradiation. In fact, it appears that inhibition of angiogenesis is best served
by a two-compartment attack upon the tumor, the tumor cells themselves and the
tumor blood supply. Most preclinical data support this concept.
09/02/2001
Arcus and PPHM
VTA, antigangiogenesis, Arcus and PPHM: I have
re-read some statements from the conference call and looked at the recent
science.
Two statements from the conference call imply the
relationship of targeting the vasculature in terms of targeting established
blood vessels within the tumor and targeting newly developing endothelial cells
in attempts to prevent angiogenesis (the formation of new vessels).
Now, the use of truncated Tissue Factor (tTF) in tumor tissue, which causes
thrombosis has relied upon conjugation of the factor to an antibody for
targeting, such as antiVACM-1 (originally published by Thorpe) or like the
Italian study using antiFibronectin to deliver the tTF to the tumor blood
vessel cells. This is vascular targeting with CONJUGATED antibody is the
property of Arcus. This conclusions follows LeGere's statement, " Well,
everything to do with the truncated tissue factor is within Arcus. Any
conjugated vascular targeting agent is within Arcus. So the only thing that is
outside of Arcus is nonconjugated antibodies for vascular targeting. Our lead
compound within Arcus is going to be an antibody that uses the truncated tissue
factor as the coagulation effector. We will move that into the clinic as soon
as we can."
Hence, the use of NONCONJUGATED tTF as an effective agent (which takes
advantage of the expressed phosphatidylserine lipid on endothelial cells; found
to also induce thrombosis in a later study by Thorpe) is the property of PPHM,
inferred by his statement later in the conference call, "I believe the
nonconjugated tTF would be under Peregrine. I have to see some more data on
that before we would move that forward, but Dr. Thorpe has got some tremendous
research going on over there, and there is a lot of stuff he's working on that we
haven't even told you about that we find very exciting, so we look forward to
getting a whole bunch more products out of Dr. Thorpe into the future, and we
are very excited about working with the University of Texas,
Southwestern."
Now, combine this with the statement above, that everything to do with
UNCONJUGATED antibodies is outside of Arcus means that the 2C3 belongs to PPHM
where probably the conjugation of the antiphosphatidylserine antibody to tTF
belongs to Arcus.
Finally, an important statement, "Our lead compound, which would be our
2C3, which I know you have all heard of, is our anti-angiogenesis drug. We want
to move that into clinical studies. I think the important thing to remember
about these other approaches, they are going to be what we call nonconjugated
antibody, or other people maybe call them naked antibodies. Big pharmas love
naked antibodies. They are easy and cheap to manufacture. They can put a 30000
liter reactor on line and pump these things out day after day, and that's
really what Rituxan and Herceptin, both of those drugs are naked antibodies, so
the big pharmas love them, and those are the types of companies we will target
with these programs.
The other one, we'll release more data on the other ones a little bit later. I
don't want to kill a surprise that will be coming up later this year."
This certainly suggests that either licensing or pharma partnership about the
antiPS antibody or related methodology is in the works and will be a
significant announcement.
09/09/2001
Zevalin and Oncolym:
Similarities: radiolabeled antibodies targeted against antigens found on
lymphoma cells, primarily of B-cell origin. Both are murine monoclonal
antibodies.
Differences: Zevalin is targeted against the external, cell-surface antigen,
CD20, whereas Oncolym targets the external, cell-surface antigen, Lym-1, an
altered HLA-Dr transplantation class antigen. Zevalin uses the yittrium-90
isotope, where the standard therapy with Oncolym is with I131 iodine (although
DeNardo has used the 67-Cu isotope with a lym-1 antibody).
Zevalin is the IDEC attempt to answer the problem of probable rituxan
resistance and their attempt to develop radiotherapy of lymphoma using the CD20
antigen. Oncolym remains a very viable candidate for therapy of B-cell
lymphoma.
11/02/2001
Regarding ARCUS. One can only shake their head
at the frustration that Thorpe must feel at the inability of his work to get
into the clinic. The licensing deals, such as SUPG (the VEGF-directed
targeting) are no doubt out of our control.
It may be more fruitful for Thorpe/PPHM to pursue the construct of humanized
antibodies (via AVN, Xenerex) against the putative targets on endothelial cells
of tumors that use unconjugated antibodies, which belong solely to PPHM and go
from there.
11/02/2001
An example of the potential importance of
intratumoral administration of Cotara is the China data on chTNT therapy of
lung cancer. The patient population reported was a mixture of adeno, squamous,
and small cell carcinomas. The partial and complete responses were noted in the
group that received both intratumoral and intravenous administration of the
chTNT, but we do not know if particular types of lung cancer were more
responsive.
This issue of intratumoral administration should not be taken lightly, at least
with the paltry amount of data in the public realm concerning Cotara. The data
presented earlier this summer from Mexico by G. Cesarman was stated by the
company, " The objectives of the study were to evaluate the efficacy,
safety and dosimetry of Cotara for the treatment of primary and unresectable
cancers of the pancreas, liver, prostate or brain. Depending upon the tumor
type, Cotara was injected into the vein, directly into the tumor, into the
spinal canal, or given in combination The drug was well tolerated. Positive
responses were seen, including one prostate cancer patient with stable disease
and one pancreatic patient with significant shrinkage of the tumor. The
pharmacokinetics, biodistribution and dosimetry results showed that Cotara
exhibited the expected patterns for a protein of its molecular weight and size.
The dosimetry profile of Cotara showed that a good therapeutic dose was
administered to the tumor", implying once again, any type of reduction of
tumor was most likely in groups that received intratumoral administration of
Cotara.
Whether this route is necessary (without large dose escalation in using
intravenous administration) in the trials on biliary, pancreatic, soft tissue
sarcoma and liver cancer at Stanford and Mayo is not known. Perhaps the nearest
entry artery of some of these specific organs that contain tumor, such as
pancreas or biliary duct or liver could be cannulated for direct infusion of
Cotara, with a subsequent optimal therapeutic ratio of uptake of Cotara.
11/29/2001
Q. Have we seen or have we checked for any
attachment of Cotara to malignant cells within the blood stream when injected
intravenously?
A. The target antigen(s) of the TNT family of
antibodies are DNA-associated proteins (mainly histones). These are intracellular
antigens that are not normally exposed on the surface of cells, except for
conditions of cell death, or necrosis. Tumors of any size ultimately develop
areas of necrosis, which makes them targets for localizing the radiolabeled TNT
antibody, such as TNT-1 or TNT-3. As cells die...say, during therapy (or even
during irradiation therapy or chemotherapy), the number of target antigens gets
larger for the TNT infusion.
Hence, viable tumor cells that float in the circulation are, in principle, not
"seen" by the TNT antibody since they have no exposed histones. Yes,
TNT would bind to floating dead, junky, crappy tumor cells, but they ain't
gonna do anything anyway. Remember...the really excellent part of the original
Epstein science on developing the TNT antibodies was the premise that, for the
most part, immunotherapy with antibodies depends on the expression of the
target antigen externally (on the cell membrane) by the cancer cells. This is
why, ultimately, tumors develop resistance...such as the case of rituxan,
bexxar, and even presumably oncolym. Because you select for the presence of the
very, very small subpopulation of cells that do not express the target antigen.
Epstein went a step further, and chose antigens that rarely mutate, and are always
present...cause they are "normal" proteins that are only
"recognized" when cells are necrotic.
12/13/2001
Brief view of today's news. The PR has been
anticipated, since the FDA did grant a Fast Track review process for Cotara in
glioma, and EL made a strong point of receiving a go-ahead by the end of the
year.
There is only one source of hard data about the Phase II results with Cotara,
and that is the ASCO meeting abstract of last May. Several items need to be
noted: (a) the data presented at ASCO included 20 patients, 15 of which were
either newly diagnosed or recurrent glioblastoma and 5 with recurrent
anaplastic astrocytoma., (b) the stated results focused upon parameters of
treatment, with a mention of median time to progression as one endpoint, being
about 16 weeks (later revised to 13.5 weeks in the ASCO news release) for the
group, compared to 8 weeks for historical median time of progression of about 8
weeks, and (c) the patient selection was not randomized, meaning a select group
of patients was chosen for this Phase II study.
In the absence of a formal publication by Patel, et al., which describes in
detail the patient populations treated and an analysis of both patient and
treatment parameters that would allow definitive measures of antitumor
response, the duration of the response, median survival and time to
progression, it is very difficult to know exactly what the numbers will be. The
Phase III trial is precisely defined in terms of the patient population, i.e.,
first-time recurrent glioblastoma. This is important in light of the fact that
other brain cancers have different responses to therapies and hence different
outcomes with respect to progression and survival which can skew or confuse
results.
The comparison with temozolomide (Temodar) is appropriate since studies are
starting to indicate that chemotherapy with multiple agents is no better than
the use of Temodar alone (oral administration, usually about 150-200 mg/m2 for
5 days on a monthly cycle). Actually, the data for a Phase II study of Temodar
in patients with first recurrent glioblastoma is published and indicates a
progression-free survival at 6 months of 18%; median progression-free survival
of 9 weeks and median overall survival of 22 weeks. A very low complete
response rate was observed (less than 10%) and stable disease was noted in
about 45% of the patients. The safety profile was similar to that described for
Cotara. There is a good literature on what can be anticipated for the treatment
of these patients with Temodar.
Recurrent glioblastoma is a terrible, fast growing lesion that invades and
recurs in patterns that can be unmanageable. The survival rate and time is
extremely poor. We wish the patients the best of success.
Editorial comment: I would have titled today's PR something like,
"Peregrine Pharmaceuticals Announces the Design of the Fast Track Phase
III Study for Brain Cancer Approved by the FDA".
Of course, we wait on the designation of the trial centers, how fast Europe and
Canada can come on board and the expected accrual rate of patients.
Unfortunately, it will be a quick read-out, since this particular cancer is so
devastating.
01/09/2002
Oncolym remains somewhat of a mystery to me with respect
to status of trials and exactly how it will be used in NHL. Nearly 60 patients
were treated with I-131 labeled Lym-1 antibody between 1985 and 1994 (Clin Can
Res 3: 1253, 1997). The Lym-1 antibody targets a mutated HLA-DR antigen found
on B-cell lymphomas (patients must be positive for the presence of this antigen
to enter this form of therapy). Interestingly, a lot of the dosimetry
measurements (radioactivity found in normal tissue) and biodistribution did not
really predict for response in the patients, but half-life of the radioisotope
was an important predictive parameter. At the same time, the development of
anti-mouse antibodies (the Lym-1 antibody is a mouse hybridoma antibody) did
correlate with improved survival, suggesting that development of these
antibodies by patients, thought to present an obstacle to therapy (and can)
actually may improve the therapeutic benefit of Lym-1 therapy.
The DeNardos championed the use of Lym-1 therapy over the past decade and
published many papers on the use and comparison of radiolabels for this
antibody (more on these comparisons later). Early studies using I-131 Lym-1
indicated that responders had a median survival of about 84 weeks, compared to
22 weeks observed for non-responders (Cancer 80: 2706, 1997).
Early 1998, PPHM announced the start of a Phase II/III
trial of Oncolym in treatment of intermediate- and high-grade refractory or
relapsed NHL. This trial was to evaluate he therapeutic efficacy of two doses
of 60 mCi/m2 (about 120 mCi per patient) given 6 weeks apart.
At ASCO in 1998, a retrospective analysis of this therapy was presented by
Jamie Oliver and the DeNardos. A limited number of patients were used for this
analysis. It was observed that response was dose-dependent (the higher the dose
of radiation, the better response rate and durability of that response) and
repeated dosing was superior. At the same meeting, dosimetry results were given
by the multi-center group (Iowa, M.D. Anderson, Miami, Cornell, George Washing
U). They again spoke of improved responses with subsequent doses of
antibody-delivered I-131. Similar results were presented the summer of 1998 by
the Georgetown U group at a Nuclear Medicine meeting in Toronto. The treatment
protocol was amended that summer by PPHM (with permission of the FDA) to
include patients that actually failed prior therapy with monoclonal antibodies
(presumably patients that had failed Rituxan or Bexxar?), but I have not found
publications on the success or failure of these specific patients treated with
Oncolym. Actually, one can not find a formal publication at all using the name
Oncolym. Strange.
However, the DeNardos remained prolific and productive in
their use of the Lym-1 antibody. They started studies using fractionated doses
of Lym-1 coupled to the 67Cu isotope by a methodology that allowed binding of
the copper atom to the antibody. This method was different than the
chloramine-T method commonly used for iodine (131) coupling to the Lym-1
antibody. Now, radiometal labeled antibodies provide a higher tumor radiation
dose than corresponding I-131 labeled antibodies. This by itself doesn't mean
much, unless you look at how long the radiation stays at the tumor site…then it
becomes important. In other words, one can always up the dose of I-131, but the
question is what is damage to normal tissue and bone marrow and how long is the
isotope in the vicinity of the tumor. They described the use of 67Cu-Lym-1 in
NHL using a minimum of 2, and up to 4, doses (Anticancer Res 18:2779, 1998).
They concluded in this study that the isotope 67Cu gave good imaging, favorable
dosimetry and a high ratio of tumor to marrow radiation (good). Immediately
after that publication, in the summer of 1998, they published a similar study
with I-131 Lym-1 with multiple dosing up to a total of 300 mCi over the course
of several weeks. Tumor regression occurred in 83% of the patients, and was
prolonged enough in 57% of the patients to qualify them as responders (Cancer
Biother Radiopharm 13: 239, 1998). In the Fall of 1998, they published a study
about I-131 Lym-1 therapy describing the maximum tolerated dose & efficacy
for fractionated multiple dose therapy of NHL (J Clin Oncol 16: 3264, 1998).
Ten of 14 patients who received at least two doses of therapy and 11/21 total
entries responded. Seven of the responses were complete with a mean duration of
14 months. All responders had at least a partial remission after the first
therapy dose, suggesting that response after initial dosing was predictive for
subsequent treatment response. A similar report was published in 1999 (J Nucl
Med 40: 302) using 67Cu with superficial lesions of NHL and they achieved about
a 50% regression of measured tumors. The remarkable thing about use of the 67Cu
isotope was the long residence time of the radiation within the tumor and that
even imaging (subtherapeutic) doses showed some regression.
They then published (a study) (Clin Can Res 5: 533, 1999)
where they compared the pharmacokinetics and dosimetry in patients with NHL
using either I-131- or 67Cu-labeled Lym-1 and concluded that due to the better
ratio of tumor/normal tissue delivery of isotope, lower radiation to the bone
marrow by 67Cu and imaging was improved using 67Cu that the use of 67Cu may be
superior to I-131 for Lym-1. Yes, I read the paper in its entirety. They do
mention that 67Cu is not routinely available, is expensive and would require a
ramp-up of accelerator facilities not readily available. Several subsequent
publications review the dosimetry and parameters associated with Lym-1 therapy
(using either 67Cu or I-131; J. Nucl Med 40: 2014, 1999; Clin Can Res 5: 3330s,
1999); J Nucl Med 40: 1317, 1999).
Now the company PR of March 2000 gives a rationale for re-evaluation of Oncolym
in a Phase I setting. Bonfiglio stated that a three treatment dose protocol was
cumbersome, expensive and had difficulty in recruiting patients. And he stated
that an analysis of the data revealed that 80% of the patients treated with a
therapeutic dose of Oncolym responded well to the first dose. I assume he is
referring to those that go on to be responders as found in the earlier studies.
The status of the trials remains somewhat unclear to me. We do list many
centers that are open for enrollment. From a purely scientific point of view, I
would still wonder whether optimal treatment requires multiple doses and the
fact that other radiolabel therapies for NHL, such as Zevalin (yttrium90
isotope) and Bexxar (I-131) use multiple dosing.
01/29/2002
This anti-VEGF subject is a little too complex
for me. Let me explain. VEGF has long been known to be a growth factor that is involved
in myriad of embryonic, adult tissue maintenance and tumor growth and
vascularization events. There are several VEGF receptors (VEGFR) found on
different tissues, some fairly specific, such as the VEGFR2 (KDR/Flt-1) found
on vascular endothelium, although this receptor is important in embryonic
development (Proc Natl Acad Sci U S A 1997 May 13;94(10):5141-6,
"Ligand-dependent development of the endothelial and hemopoietic lineages
from embryonic mesodermal cells expressing vascular endothelial growth factor
receptor 2", Eichmann A, Corbel C, Nataf V, Vaigot P, Breant C, Le Douarin
NM). However, this receptor is pretty specific for tumor vasculature and hence
an attractive target for antiangiogenic therapy.
There are currently several approaches to anti-VEGF therapy, e.g., blocking the
binding of VEGF itself to receptors (a rather risky non-specific issue, due to
need for VEGF in normal tissue function), and the blocking of binding of VEGF
to tumor cell receptors (tumors use and secrete VEGF in large quantities) by
the use of antibodies (Genentech and IMCL). In addition, other strategies have
attempted to specifically block the phosphorylation of these receptors, since
they have function that is known as tyrosine kinases (the SUGEN compounds, 5416
and 6668, and Novartis PTK787). Hence, there is little doubt that the more one
can successfully target the tumor endothelium carrying the KDR receptor, the
more superior the therapy becomes. As an example, the VEGFR-2 has been found in
90% of tumor specimens of head and neck cancer (Neuchrist C, et al.,
Laryngoscope 2001, lll: 1834) and appeared to be localized to vascular
endothelium. This has been observed in many solid tumors and also by Thorpe and
his colleagues (J Control Release 2001 Jul 6;741-3):173-81, "VEGF-VEGF
receptor complexes as markers of tumor vascular endothelium.", Brekken RA,
Thorpe PE) as well as expression of the VEGFR-2 in vasculature of
VEGF-secreting tumors ( Feng, D., Nagy, J.A., Brekken, R.A., Pettersson, A.,
Manseau, E.J., Pyne, K., Mulligan, R., Brekken, R., Thorpe, P.E., Dvorak, H.F.,
& Dvorak, A.M. (2000) Ultrastructural localization of the Vascular
Permeability Factor/Vascular Endothelial Growth Factor (VPF/VEGF) Receptor-2
(FLK-1, KDR) in normal mouse kidney and in hyperpermeable vessels induced by
VPF/VEGF-secreting tumors and adenoviral vectors. J. Histochem. Cytochem.,
48:545-555).
Scientists from Imclone reported several years
ago the value of blocking the VEGFR-2 receptor using the DC101 antibody in mice
(Cancer Metastasis Rev 1998 Jun;17(2):155-61, "Monoclonal antibodies
targeting the VEGF receptor-2 (Flk1/KDR) as an anti-angiogenic therapeutic
strategy", Witte L, Hicklin DJ, Zhu Z, Pytowski B, Kotanides H, Rockwell
P, Bohlen P). These studies evolved into the clinical trials of their
anti-VEGF/KDR receptor antibody, IMC-1C11. In May, 1998, IMCL was awarded US
Patent 5747651 on the use of antibodies against the extracellular portion of
the Flk-1/KDR receptor of VEGF. Early last year, the Imclone scientists
reported the ability to select high affinity antibody construction against
VEGFR-2 by the use of phage-display technology (Int J Cancer 2002 Jan
20;97(3):393-9, "Selection of high affinity human neutralizing antibodies
to VEGFR2 from a large antibody phage display library for antiangiogenesis
therapy; Lu D, Jimenez X, Zhang H, Bohlen P, Witte L, Zhu Z).
The highly specific antibody, described by Thorpe, 2C3, has been shown to have
potent antitumor activity, presumably by targeting the tumor endothelium
(Brekken, R.A., Overholser, J.P., Stasny, V.A., Waltenberger, J., Minna, J.D.,
and Thorpe, P.E. (2000) Selective inhibition of VEGFR2 activity by a monoclonal
anti-VEGF antibody blocks tumor growth in mice. Cancer Res., 60, 5117-24). Now,
the present patents awarded to Thorpe, and I guess, subsequently assigned to
PPHM covers the usual infinite scope of applications that only Thorpe can
envision and articulate, even to the point of delivery of about every
diagnostic and therapeutic agent ever used or being tested (including angiostatin,
endostatin, etc). There seems little doubt that the specificity of such
delivery may exist (what happened to the SUPERGEN studies with VEGF-trojan
horse?) but this enthusiasm needs to be tempered with the reality of PPHM
activities.
First, these studies appear to be preclinical, i.e., promising due to the
observations using good animal tumor models. One of the most exasperating
observations about this company is the inability to get the Thorpe science into
the clinic. We have had VTA in several forms; such as targeting expressed and
induced receptors on tumor endothelium, the delivery of either conjugated or
free truncated tissue factor and the potential antitumor activity associated
with antibodies that target phosphatidylserine on tumor endothelium (Ran, S.,
Rote, N., and Thorpe, P E. Phosphatidylserine is a marker of tumor vasculature
and a potential target for anti-cancer drugs; submitted). However we jockey for
position with ARCUS (and OXGN) on who owns what in what form
(conjugated/non-conjugated), it is time to get some of these molecules into the
clinic. We can't do them all. They all can't be the best. Somebody has to
decide what to develop in terms of endothelial cell targeting, study the
parameters within the confines of good animal models and quickly get into Phase
I trials that can confirm the biodistribution, activity and behavior of the
agent in patients.
An infinite patent package of pipelines that are never
tested doesn't do anybody any good. So, in this regard, I am still focused upon
the activities of the company that give credibility to the products under
development, mainly Cotara and Oncolym.
In that regard, guess I might as well make a few more
comments.
1. Oncolym. What a disaster of such a promising molecule. Fascinating trials
could have been formulated regarding therapy of relapsed lymphoma (every
therapy requiring of course, a strong expression of the HLA-DR10 antigen by
lymphoma cells, but that is easily assayed). I am not sure we ever came away
with a strong indication for optimal dose and scheduling for I131-Lym1 and yet
the company says that strong analysis of the data indicate that a one-dose
scheme should be developed. It really doesn't matter what I think, but the
publications by the DeNardos clearly indicated that the durability and depth of
response was dose-dependent (several treatments). I doubt we can deliver that
accumulated radiation by a single dose, at least using I-131. And of course,
now there is evidence that the Cu67 isotope may have preferable properties. We
have no real word of the status of the Phase I trials using single-dose methods
even though the web site says multi-center trials. I doubt the enrollment is
robust. The front line therapies will be rapidly changing in most cancers and
with the lessons learned by Bexaar, Rituxan and Zevalin (although the FDA is
giving these therapies a rough time; the C225 antibody of IMCL is very good
therapy when combined with conventional cytotoxics), it just appears we make
little progress in the clinic with Oncolym. There could be several indications
for the use of Oncolym, regarding combination therapy or studies to determine
the development of resistance to Oncolym (loss of the target antigen, which is
not unique to antibody-mediated therapies, e.g., Gleevec (STI571, the tryosine
kinase inhibitor of the PDGF-receptor in CML now has resistance based upon
cells that mutate and overexpress that receptor and become resistant to
Gleevec). Except for the published literature by the DeNardos, there is nothing
about "standard" Oncolym therapy in the medical literature. Certainly
no update has been given by the company….that I recall even in enrolling or
treating the first patient in this new Phase I trial. Where the funding comes
from for the Oncolym trials is unknown, since we know the bulk of monies are
directed to the ongoing Phase II and to-start Phase III of Cotara.
2. Cotara. Clinically, the orphan-status of this
agent doesn't mean anything, and the Fast-Trac of Phase III means a lot. The
number of experimental therapies for glioblastoma is enormous, considering the
relative small numbers of patients with this disease compared to mainline
cancers, such as prostate, breast and colon. It is interesting that the FDA
leaned toward a comparative study with temozolomide, which suggests that this
form of chemo is approaching front-line therapy for recurrent glioblastoma.
However, the use of the chemo-wafers, and now small-molecule development
(cytokines, tyrosine kinase inhibitors), as well as other targeting antigens
for antibody-mediated therapy (such as the Bigner group) may be improved with
better delivery methods. And, we know this Phase III trial will be very, very
expensive, especially if we conduct the trial in Canada and Europe as well as
here in the U.S.A. Certainly this clinical use of Cotara is the actual leading
candidate for product-income over the next few years. How the Phase III (or
ongoing Phase II) trials will integrate the collaboration with Image-Guided
Neurologics in their use of the Navigus Array Catheter remains to be presented.
I remain quite disturbed about the lack of
formal publications regarding any case studies involving Cotara in brain tumors
(case studies are interesting, informative and rapid ways to tell how a field
is developing in new therapies). There must be little motivation or incentive
for this Assistant Professor to publish these findings which is extremely
unusual for a young clinician in academic medicine. I assume we will have
something new to say at ASCO, and it is disturbing that Cotara therapy of brain
cancer is NOT well known to the neuro-oncology community and doesn't find it
way to a podium setting at a meeting. As I have mentioned in the past, poster
sessions do not necessarily reflect unenthusiasm for a research presentation, (many
high-profile results of other biotechs are presented in poster sessions), but
ultimately the HOT topics of a field find their way to a podium presentation.
3. Cotara in solid tumors. Multi-site studies at good
medical settings, yes, and again, we have no reports at all concerning the
enrollment or findings of the Phase I trials in solid tumors with Cotara, other
than last years paltry data from Mexico. Yes, I know that Phase I trials are
designed to determine maximum tolerated dose, pharmacokinetics (distribution),
localization ratios of isotope to tumor/normal tissues and other parameters,
but we have really no clue as to the progress of these trials. I guess that is
the disturbing part…progress. We really don't seem to know about progress.
There are so many ways for a company to let the shareholders know that their
compounds and drugs are moving along a track, e.g., seminars, small
conferences, society meetings, and brief publications.
Now, the China SDA approval of chTNT for certain indications (I still emphasize
the probable need for an intratumoral protocol, besides IV injection) will be
overwhelming validation for at least considering the odds of Cotara being a
player in therapy of solid tumors. The transition needed from these data out of
China should be rapid and smooth into the trials of Cotara in this country
(yet, to my knowledge, we have no lung cancer trials ongoing, which is
surprising given the modest amount of data known from China). There seems
little doubt that short-term sentiment about the value of the Cotara therapy
will be established by what China SDA does. As I mentioned before, the link to
the China data (which I can not seem to resurrect stated that approval was on
track for May, 2002).
4. ARCUS. Oh well, that seems a somewhat
distant cause. Unless OXGN really wants to try targeting the tubulin-binding
agents (combretastatin derivatives) using TNT-type antibodies, it seems we are
intent are using un-conjugated VTA agents on our own. The Thorpe anti-PS stuff
is truly frustrating and must be to him also. He has submitted a paper on the
targeting of PS (according to the PPHM website). It seems clear, IMO, that the
anti-PS will be developed via ARCUS, since that is where the patent was
awarded. We really don't know what has happened to the truncated-tissue factor
stuff. ARCUS remains a mystery to me…even more so than the SUPG deal with the
VEGF-gelonin molecule developed by M. Rosenblum.
5. Oakwood Labs: This company develops delivery of
compounds and drugs via liposomes and obviously believe they can improve their
targeting with TNT-guided delivery to the tumor bed by coupling the antibody to
liposomes that contain toxic goodies. This will take some preclinical studies
on animal tumor models, doses, coupling procedures that are optimal and
scheduling parameters. Nothing coming soon here that I can see. But that's
okay. A pipeline development.
6. Merck (Lexigen): This licensing/collaboration with
Lexigen probably centers around the fusion-peptide technology of TNT-IL-2
(Epstein has published on this concept, (Pretreatment with a monoclonal
antibody/interleukin-2 fusion protein directed against DNA enhances the
delivery of therapeutic molecules to solid tumors. Clin Cancer Res. 1999
Jan;5(1):51-60). As I mentioned before, this methodology probably has been
secured by S. Gillis, a recognized world-class researcher in the area of
immunocytokine peptides and Gillis is the head of Lexigen. I note that the
agreement was over a year ago, which means the discussions of this agreement
were probably at least six months before that…and in the year and a half since,
no new data is in the literature that shares a Lexigen/PPHM byline, although
Gillis has published recent papers on the use of the fusion peptide-IL-2
effects ("Improved circulating half-life and efficacy of an
antibody-interleukin 2 immunocytokine based on reduced intracellular
proteolysis." Clin Cancer Res. 2002 8: 210-216; as well as
"Augmentation of antitumor activity of an antibody-interleukin 2
immunocytokine with chemotherapeutic agents." Clin Cancer Res. 2001
Sep;7(9):2862-9.) I assume some sort of preclinical studies are underway in
this area. How this fits into the overall VEA scheme of PPHM is not clear to
me.
7. AVID. Well, all of us know what is trying to
happen there. We wait.
I do applaud the very hard work and enthusiasm of Ed L. but I also believe
there are legitimate (and non-shallow) ways for the company to show us that
science and business is progressing in ways that he has not done. He needs to
get his clinicians publishing studies and working their way into podium
sessions, attending more high-profile small conferences (which is the way of
biotech lately, the AACR conferences for example) and hitting all the clinical
meetings.
02/01/02
You ask, "Do any of you longs which know
this company inside out...know what EL's agenda is to promote the China data?
Has EL assembled a crew of top Ph.Ds/Oncologists to decipher these clinical
trials? Is it your opinion Medipharm will let us publish this data to support
our cancer therapy? Does anyone have a grasp on what will happen once we get
the China data?"
I can not speak for Mr. Legere at all, but I would emphasize that the China
data does not belong to PPHM. It is property of Medipharm and their
collaborators (presumably the Epstein company). We are involved for two
important reasons, (a) we have the ability to manufacture the radiolabel
antibody and share royalties, but more importantly, (b) the success of the
China trials have a huge impact on validating the Cotara-based therapy. This is
what is most important to me. I have no idea what China data he has viewed. The
translated abstract of a couple months ago is the only data I am aware of and
it certainly seemed very encouraging with respect to lung cancer and intratumoral
infusion.
You also stated, "When I see EL promoting the mab plant, and not the data
we have from Phase II, it makes me wonder why we are focusing so much attention
on the mab facility. I like the idea of Big Pharma looking at our data and
paying for our trials." In my opinion, Mr. Legere is focusing upon
immediate goals under his control, such as revenue obtained from contracts of
Mab manufacture. I often complain about the lack of peer-review publication and
visibility of the Cotara-glioma results. Of course, having Big Pharma examining
the data and funding the trials does have the cost of "diluting and
sharing" any subsequent revenue.
To me, the biggest benefit of actually knowing the China data is the immediate
decisions about what strategies of Cotara work and don't work, with regard to
tumor type, location and tumor burden. If I knew the China data obtained with
many tumor types, I would focus upon the success and discard the failed
approaches, all techniques, doses and delivery methodology being equal. I do
believe that Mr. Legere is doing his best with all the resources available to
him.
02/01/02
You ask an important question, wondering how
the China data will be utilized for PPHM. I believe there are several features
of the "business" and the "science" of Cotara that are
crucial for the company. First, there is the example that T. Chew successfully
negotiated the regulatory obstacles for Fast Trac evaluation of Cotara in
recurrent glioblastoma and he has successfully brought a product through the FDA
in his prior work. This is encouraging. So, one could feel confident that Chew
can assemble the data for the FDA when it is presented in a format that can be
evaluated and summarized. Will PPHM have the "task force" available
to evaluate the China data? Several avenues are available. First, he has the
principal investigators of the multi-center glioma trials to evaluate the
findings of intratumoral infusion of Cotara in solid tumors. In addition the
company has the resources of the principal investigators of Cotara in other
solid tumors (Stanford and the Mayo Clinic). I may be somewhat simplistic but
the results of China will be, in my opinion, straight-forward, assuming the
methods and measurements of the use of Cotara in China have been controlled.
Remember, the trials of Cotara in solid tumors here in the USA are Phase I,
designed to study toxicity and pharmacokinetics and NOT efficacy. Presumably,
we know quite a bit from the Mexico studies. However, the China data should
outline pure success and failure and in a variety of tumor types. Assuming
their procedures are carefully designed and controlled, the data should be
invaluable in determining the course of studies here in the USA about Cotara.
Through the expertise of Epstein, I would also think that the company has the
resources to review the data and utilize it to its fullest extent. . The China
data should guide our use of Cotara in solid tumors. This requires that the
China data is simple enough in the parameters of the trials (type of tumor, patient
selection, amount and frequency of TNT administered, route of administration,
close follow up of clinical course of disease, influence of prior therapies,
subsequent observations of extent and site of recurrent disease, and observed
toxicities).
In this regard, the track record is not encouraging.
The example using Oncolym is dismal. We went from the clinical use of Oncolym
via the DeNardos and should have entered into pivotal (final) trials in
lymphoma and ended up going backward to Phase I with Schering, based upon
"outside" opinion that one-dose protocols should be developed (in
spite of the published data), and the status of Oncolym appears uncertain. I
have never understood exactly where the wheels came off in the Oncolym
"business".
The personnel to the Mab facility are technical
support and staff associated with manufacture of monoclonal antibodies and the
regulatory requirements of their production and not personnel associated with
research and development of clinical trials. The R & D of PPHM is Epstein
and Thorpe and their labs are supported by funds from PPHM.
02/10/02
#390
The decision to stop the (Sugen) trials
involving the tyrosine kinase inhibitor (RTKi) SU5416 was disappointing in
light of the promise of this form of therapy. As late as last year at ASCO,
this form of therapy was obtaining some meaningful responses in patients when
used with chemotherapy. It is entirely possible that the parameters that
ultimately determine patient responses to two-compartment targeting may be lost
when trials are performed on "all comers" or large groups of patients
that have seemingly homogeneous conditions (metastatic colon carcinoma with
refractory disease) yet very heterogeneous responses that almost ruin any
statistical analysis for that group. The SU5416-chemotherapy trial had some
very dramatic responses that got lost in the final analysis; and I know this is
what Rosen is talking about in "learning" from this type of trial.
Modern concepts of cancer therapy has focused upon two-compartment targeting;
the tumor cell themselves and tumor-associated blood vessels. The strategies
for this concept vary widely. Sugen originally targeted growth factor receptors
thought to be needed for proliferating tumor cells as well as endothelial cells
(such as epidermal growth factor, EGF; vascular endothelial growth factor,
VEGF; basic fibroblast growth factor; bFGF and platelet-derived growth factor,
PDGF). Different companies are attempting to target specific properties of
tumor-associated blood vessels that may be unique to dividing (and forming)
vasculature within an expanding mass of tumor (VTA by Thorpe [via expression of
phosphatidylserine or VCAM-1 using truncated tissue factor], combretastatin by
OXGN, TNP-40, endostatin, angiostatin by ENMD, VEGF-gelonin presumably by SUPG
and a myriad of other molecules that have indirect effects on endothelial
cells.
02/10/02
#391
The optimal result of two-compartment therapy
will be obtained when the tumor has induced the coordinate receptor on the
endothelium for a growth factor they both secrete and express the receptor
(best example is the large secretion of TGF-alpha from tumor cells that
upregulate the EGF-receptor on endothelium in addition to those expressed on
the tumor cells themselves). In theory, this type of tumor will be very
sensitive to targeting the EGF-receptor (such as PKI-166 by Novartis, or the
IMCL C225) and if one could determine prior to therapy that the tumor is
secreting TGF-alpha (the most common ligand for EGF-r) and that the tumor is expressing
the EGF-r as well as the tumor blood vessels, the odds of successful therapy
(at least using that form of therapy) go up tremendously and the patients can
be divided into groups that have certain "markers" that may be
amenable to certain targeting agents (such as RTK-inhibitors). This is being
pursued vigorously in preclinical models (and can be found in the literature).
At the same time, we are recognizing that tumor-associated endothelial cells,
often thought to be the immutable target for direct antiangiogenic therapy has
properties that can be "conferred" by tumor cells that may cause
resistance to apoptotic-inducing agents. This property of having resistance
induced by factors secreted by tumor cells (instead of classical selection as
found in those cells that survive a round of chemotherapy) may cause problems
in formulations of two-compartment targeting. But the SUxxxx compounds have
pioneered a very important concept that has lead to a much better understanding
of the molecular description of what drives the proliferation of tumor cells,
causes the continual formation of new vasculature, confers protection upon
these cells and thwarts direct and indirect targeting of these molecular
pathways.
Date: Wed Mar 13, 2002 12:27 pm
It is completely unacceptable that physicians
running the clinical trials for PPHM Cotara did not submit presentations to
ASCO, the premier clinical oncology meeting in the world. Phase II data was
presented a year ago (May 2001) at ASCO and there has been a year follow up in
that data. In addition, we agreed to examine the use of the Navigus array
catheter (Aug 2001), certainly there is "new" data about this device
i.e., distribution of antibody, localization images, etc that would have made a
nice presentation.
Patel and his multi-center colleagues should
have published a manuscript on interim Phase II Cotara by now. Obviously the
findings are encouraging enough to warrant Fast Track consideration by the FDA
using a comparative study with chemotherapy in glioblastoma. There must have
been interesting findings that would have made a nice case study report (which
is common in medicine).
It has been nearly 15 months since the first
colorectal patient was treated at Stanford with Cotara and almost a year since
the other solid tumor trials at Stanford and Mayo were started. Nothing has
been presented regarding these Phase I trials and ASCO would be the place to
start. The absolute dogma of academic medicine is to publish and present data.
It seems paradoxical that Thorpe and Epstein
have published for years in preclinical models of VTA and VEA, yet nothing is
in the clinic (while) the clinical studies for Cotara have been going on for
years and nothing gets published.
Date: Fri Mar 15, 2002 6:30 am
Subject: Epstein and Thorpe at AACR
LEC/chTNT-3 fusion protein for the immunotherapy of
solid tumors
Jiali Li, Peisheng Hu, Leslie Khawli, Alan Epstein,
University of Southern California, Keck School of Medicine, Los Angeles, CA.
The human chemokine LEC (liver-expression
chemokine), also named HCC-4, NCC-4 and LMC, was originally found in an
expressed sequence tag library and later the gene was located on chromosome 17q
in the CC chemokine gene cluster. LEC has been shown to chemoattract both
monocytes and lymphocytes mediated by interacting with CCR1 and CCR8, but not
neutrophils. In this study, a LEC/chTNT-3 fusion protein has been generated to
target this potent chemokine to tumors in order to generate an effective immune
response. The cDNA of LEC has been cloned by RT-PCR from Hep-2 cells. Because
the N-terminal of chemokines are important for their activity, we fused the
C-terminal of LEC with the N-terminal of chTNT-3 heavy chain variable region
with restriction enzymes Xba1 and Not1 and then the fused LEC/chTNT-3 heavy
chain was inserted into a glutamine synthase gene amplification system
expression vector pEE12 with an antibody leader sequence. The chTNT-3 light
chain was carried by another vector pEE6 and co-transfected into the murine
myeloma cell line NS0. The fusion protein was purified by tandem protein-A
affinity and ion-exchange chromatography. The proper assembling of the fusion
protein was demonstrated by SDS-PAGE which showed a 68KD band compared with a
56KD for chTNT-3 heavy chain under reduced conditions. The bioactivity of the
fusion protein was demonstrated by a chemotasis assay using the human monocyte
cell line, THP-1 and compared with recombinant LEC. The immunoreactivity of the
fusion protein was determined by binding to fixed Raji lymphoma cells. These
binding studies showed that linking the chemokine to the variable region of the
antibody does not interfere with antigen binding under physiologic conditions.
Furthermore, LEC/chTNT-3 retains its chemokine chemotasis functionality. In vivo
immunotherapy studies will focus on the recruitment of immune effector cells to
tumors and on monitoring tumor growth in mice receiving different regimens.
Humanization and mutation of an anti-nuclear
antibody developed for human cancer therapy
Jianghua Yan, Peisheng Hu, Leslie A. Khawli,
Alan L. Epstein,
Department of Pathology, University of Southern
California School of
Medicine, Los Angeles, CA.
Tumor Necrosis Treatment
monoclonal antibody (TNT-3) was developed by our laboratory to target necrotic
regions of solid tumors. As a carrier of radionuclides and immune modulators,
chTNT-3 has been shown to be a promising targeting agent for the radio- and
immunotherapy of solid tumors in animal models and man. To improve its clinical
potential, a humanized TNT-3 was designed and constructed by searching human
anti-DNA antibody data bases and using PCR assemblage. To restore and improve
the affinity of huTNT-3 candidates, CDR3 of huTNT-3 heavy chain genes was
simultaneously mutated by site-specific random mutation. Reconstructed scFv of
huTNT-3 were inserted into the Lambda phage SurfZAP vector and displayed on the
surface of phage. Ten mutants were selected by four rounds of panning against
crude DNA. The light and heavy chains were then reconstructed in pEE6 and pEE12
vectors, respectively, for expression of whole antibody in NSO cells using the
glutamine synthase expression system. Binding studies against crude DNA
demonstrated that eight of the mutants had a 2-8 fold improvement in affinity
compared to chTNT-3 while two mutants had a 1-4 fold weaker affinity. Analysis
of the mutants with improved binding characteristics showed that positively
charged amino acids such as Arg and His replaced existing residues in four of
the mutants while the other three mutants showed replacement with amino acids
containing hydroxyl groups. In one exceptional case, the replacement of Arg
with Leu increased binding activity by four fold. These results suggest that
changes in the electrostatic or hydrogen –bonding potential of residues
contributed to the observed improvement in antibody binding to crude DNA
antigen. The availability of a high affinity humanized TNT-3 reagent will
facilitate the development of clinically relevant
reagents useful for the therapy of solid tumors
in man.
Comparison of three different targeted tissue
factor fusion proteins for inducing tumor vessel thrombosis
Peisheng Hu, Jianghua Yan, Leslie Khawli, Thomas
Bai, Jahangir
Sharifi, Alan Epstein, University of Southern
California, Los Angeles, CA.
Tissue Factor is a cell membrane
receptor protein that is the initiator of the extrinsic pathway of the blood
coagulation cascade and is normally released from damaged tissues. By
substituting the attachment site with a tumor delivery agent, this potent
thrombogenic protein in its truncated form can be targeted to the tumor where
it can initiate clotting thereby occluding the tumor's blood supply to cause
rapid and massive cell death. In this study, we have explored three different
delivery systems, namely, chTNT-3, which targets necrotic regions of tumors,
chTV-1, which binds an exposed constituent of the tumor vessel basement
membrane, and a RGD motif which targets tumor vessel endothelial integrins on
the luminal side of the vessel. The resultant recombinant fusion proteins,
chTNT-3/tTF, chTV-1/tTF, and RGD/tTF were expressed either in NSO cells using
the glutamine synthase expression system or, as in the case of the RGD reagent,
in E. coli. Antigen binding studies were performed to demonstrate retention of
the binding activity of the antibody moiety and clotting studies revealed
potent activity of the Tissue Factor constituent of the fusion proteins. After
intravenous injection in MAD 109 lung carcinoma-bearing BALB/c mice,
histological analysis revealed that all three reagents induced microregional
thrombosis and subsequent necrosis in tumors. Of interest, the chTV-1/tTF and
the RGD/tTF fusion proteins induced thrombosis in small and medium sized tumor
vessels while the chTNT-3/tTF induced clotting in larger vessels. These studies
demonstrate that multiple targets exist which can be used to localize truncated
Tissue Factor to occlude tumor vessels in this mouse tumor model. Further
experiments are underway to explore additional target sites, which may be more
efficient and/or specific for the delivery of truncated Tissue Factor in order
to identify a single reagent, which can be used in experimental therapeutic
studies in man.
Characterization of antigens recognized by chimeric
Tumor Necrosis Treatment anti-nuclear monoclonal antibodies
Meg L. Flanagan, Jianghua Yan, Peisheng Hu, Alan L.
Epstein, University of Southern California Keck School of Medicine, Los
Angeles, CA.
Our laboratory has developed a
panel of anti-nuclear monoclonal antibodies for targeting necrotic regions of
solid tumors. This panel includes three chimeric monoclonal antibodies
(chTNT-1, -2, -3) and a phage display-generated human monoclonal antibody
(NHS76) generated against the same antigen as chTNT-1. To characterize the
antigenic binding sites of these reagents, ELISA studies were conducted using
various nuclear antigens, including crude DNA, single- and double-stranded 25bp
DNA oligonucleotides, total cellular RNA, and histone subunits H1, H2a, H2b,
and H4. In response to reports of charged-based affinity of anti-DNA antibodies
for glycosaminoglycans of the extracellular matrix, we also used chondroitin
sulfate, heparin sulfate, and hyaluronic acid as potential antigens. From these
ELISA we found that chTNT-2 binds histone but none of the nucleic acid or
glycosaminoglycan species tested, whereas chTNT-3 and NHS76 bind both crude DNA
and histone. chTNT-2, which has been previously shown by immunogold electron
microscopy studies performed in our laboratory to bind heterochromatin, may
have additional uses as a targeting vehicle. Interestingly, chTNT-3 and NHS76
both bound crude DNA and histone, demonstrating that these MAbs may bind
similar epitopes on nucleosomal complexes. chTNT-1 also bound crude DNA and
histone as expected, but exhibited binding to several other species as well. It
may be that chTNT-1, which is currently showing promise in a number of clinical
trials for the radioimmunotherapy of solid tumors, exhibits this promiscuity due
to its known high isoelectric point. In the future we plan to examine in
greater detail the localization of these antibodies by confocal microscopy and,
where possible, use surface plasmon resonance technology to examine the
kinetics governing the binding relationships between our TNT MAbs and their
antigens.
In vitro and animal model studies of VEGF121/rGel
fusion toxin effects on breast cancer cells
Khalid A. Mohamedali, Lawrence H. Cheung, Sophia
Ran, Philip Thorpe, Michael G. Rosenblum, M.D. Anderson Cancer Center, Houston,
TX; U.T. Southwestern, Dallas, TX.
The angiogenic factor VEGF121 fused
with the recombinant plant toxin gelonin (rGel) exhibits potent cytotoxicity
towards tumor neovasculature and inhibits tumor growth by virtue of its selective
targeting of tumor vessels. We have used in vitro and animal models to further
understand the mode of action of VEGF/rGel on tumor cells in vivo. Preliminary
studies have suggested that interruption of blood flow to tumors with agents
targeting tumor vasculature generates intratumoral hypoxia and concomitant
changes in the surrounding pH. Vascular endothelial cells over-expressing flk-1
(PAE/flk-1) were treated with an IC50 dose of VEGF/rGel for 24, 48 and 72
hours. At the appropriate timepoints treated and control cells were harvested
and the effect of VEGF/rGel on intracellular events was examined by extraction
of mRNA and microarray analysis of proteins involved in signal transduction,
stress response, cell cycle control, hypoxia and metastasis. To examine the
effect of VEGF/rGel on tumor cells, MDA-MB-231 breast cancer tumor cells were
co-cultured with PAE/flk-1 with the two cell lines separated by a thin
membrane, allowing exchange of nutrients. The endothelial cells were treated
with various doses of VEGF/rGel and the effect on MDA-MB-231 cells was examined
visually after 72 hours. PAE/flk-1 cells in the upper chamber were treated with
an IC50 dose of VEGF-rGel over 72 hours after which both control and treated
PAE/flk-1 and MDA-MB231 cells were harvested, RNA extracted and subjected to
microarray analysis. We repeated the above experiment and incubated the cells
under hypoxic conditions to examine genetic changes associated with hypoxic
stress. These results are under analysis and will be reported. Internalization
of VEGF-rGel into PAE/flk-1 cells was also investigated 1 hr, 2hr, 10 hr, 16 hr
and 24 hrs after treating with VEGF-rGel. VEGF-rGel was detected by both
anti-VEGF and anti-rGel antibodies using immunofluorescence, and suggests
internalization within one hour of incubation. Finally, the plasma clearance
and tissue distribution of VEGF-rGel in tumor-bearing mice was examined. Nude
mice bearing orthotopic MDA-MB231 tumors were injected with 1 ìCi of
125I-labeled VEGF-rGel and tissue distribution and plasma clearance was
assessed 24, 48 and 72 hrs post injection. Research conducted, in part, by the
Clayton Foundation for Research.
Angiopoietin-2 transfection does not enhance the
sensitivity of the NCI-H358 lung tumor to VEGF-neutralizing therapy
Xianming Huang, Sophia Ran, Chelsea Swandal,
Joanna Brochu, Mary Bennett, Philip Thorpe, UT Southwestern Medical Center at
Dallas, Dallas, TX.
Angiopoietin 2 (Ang-2) is an
angiogenic factor that might coordinate with vascular endothelial cell growth
factor (VEGF) in the modulation of angiogenesis. It has been suggested that, in
the presence of VEGF, Ang-2 induces angiogenesis, but, in the absence of VEGF,
it induces vessel regression. In the present study, we examined the
complementary and coordinate action of Ang-2 and VEGF in tumor growth and
angiogenesis. Human non-small cell lung carcinoma NCI-H358 cells were stably
transfected with Ang-2. In vitro, the Ang-2 transfected cells grew at the same
rate as did tumor cells transfected with empty vector. However, they grew
significantly faster as tumors in mice than did mock transfected tumor cells
and had higher proliferation indices, as judged by PCNA staining. Ang-2
transfected tumors had aberrant vessels: they grew as dense nests of tumor
cells that possessed few microvessels surrounded by large sinusoid-like
vessels. Treatment of mice bearing Ang-2-transfected tumors with the
neutralizing anti-VEGF antibody, 2C3, reduced but did not prevent tumor growth,
whereas 2C3 treatment abolished the growth of mock-transfected tumor cells. No
increase in apoptosis was observed in the Ang-2 transfected tumors in mice,
whereas mock transfected tumor showed increased apoptosis. These data suggested
that Ang-2 might play a supplementary role to VEGF in promoting tumor angiogenesis.
Anti-VEGF antibody, 2C3, down-regulates expression
of mouse VEGFR2 receptor Flk-1, inhibits tumor microvasculature and suppresses
growth of orthotopic 231 breast carcinoma xenografts
Sophia Ran, Wei Zhang, Maria Sambade, Philip
Thorpe, Southwestern Medical Center, Dallas, TX; Maryland General Hospital,
Baltomore, MD.
Vascular endothelial growth factor
(VEGF) is a multifunctional angiogenic growth factor that is a primary
stimulant of tumor angiogenesis. We previously raised a neutralizing anti-VEGF
monoclonal antibody 2C3, that blocks the interaction of human VEGF with either
human or mouse VEGF-R2 (KDR/Flk-1) but not with VEGF-R1 (FLT-1). 2C3 had potent
anti-tumor activity, inhibiting the growth of both newly injected and
established human tumor xenografts growing subcutaneously in mice. In the
current study, we tested the therapeutic effects of 2C3 on tumor growth in an
orthotopic model of MDA-MB-231 human breast carcinoma implanted in the mammary
fat pads (MFP) of nude mice. Administration of 2C3 to mice with 100-150 mm3
tumors inhibited tumor growth up to 75%, as compared to recipients of the
isotype-matched irrelevant control IgG, C44. 2C3 treatment also inhibited the
establishment of tumor colonies and reduced tumor burden in the lungs of mice
injected intravenously with 231 cells. No toxicity of the antibody to the mice
was observed. The effect of 2C3 on tumor angiogenesis was evaluated
immunohistochemically. The mean microvascular density (MVD) of tumors in
2C3-treated mice was 55.71± 4.7 per mm2, as compared to 187.7 ±5.1 per mm2 in
the C44-treated control group. The decrease in MVD closely correlated with
degree of inhibition of tumor growth (p<0.001), suggesting that in this
model human VEGF plays a predominant role in driving murine angiogenesis. The
treated tumors mostly contained mid-size and large vessels. Microvessels were
mainly confined to the peripheral layer bordering with normal MFP epithelium.
Treatment did not affect MVD of fat and skin surrounding the tumor, indicating
a specific elimination of tumor vasculature. The remaining tumor vessels showed
a significantly decreased expression of VEGF-R2, as was determined by double
labeling using rat anti-Flk-1 and biotinylated anti-CD31 antibodies. This
observation indicates that neutralization of VEGF by 2C3 causes down-regulation
of VEGF-R2 as a secondary event. This, in turn, may limit re-initiation of
angiogenesis triggered by either human or mouse VEGF and render the tumor
dormant for prolonged periods of time. These findings suggest that 2C3 is a
candidate for treating primary cancer and for preventing the outgrowth of tumor
metastases in cancer patients. Research was supported in part by Susan. G.
Komen Breast Cancer Foundation.
Increased exposure of anionic phospholipids on the
surface of activated endothelial cells and tumor blood vessels
Sophia Ran, Amber Downes, Philip E. Thorpe,
Southwestern Medical Center, UT at Dallas, Dallas, TX.
Anionic phospholipids (AP) of the
plasma membrane consist of phosphatidylserine (PS), phosphatidylinositol (PI),
phosphatidic acid (PA) and phosphatidylglycerol (PG). PS comprises 3-7% of the
total phospholipids and is exclusively located on the inner leaflet of the
plasma membrane under normal circumstances. Other AP are present in much lesser
quantities in the plasma membrane. PA is located mainly on the external leaflet
whereas PI is located mainly on the internal leaflet. Annexin V, which
specifically reacts with AP, does not bind to the external surface of normal
quiescent cells, suggesting that the overall distribution of AP is heavily
biased toward the inner leaflet of the plasma membrane. Externalization of PS,
and annexin V binding, occurs upon cell activation, injury, apoptosis and malignant
transformation. We postulated that tumor-derived factors might damage tumor
endothelium and increase exposure of PS and possibly other AP. To test this
hypothesis, we generated a monoclonal antibody, 9D2, that was specifically
reactive with PS, PI and PA but not with other phospholipids. 9D2 is more
specific for AP than is annexin V, which, strongly binds to
phosphatidylethanolamine (PE) in addition to other AP. 9D2 and annexin V did
not bind to cultured endothelial cells under normal circumstances. However,
treatment of the cells with thrombin, inflammatory cytokines, hydrogen
peroxide, hypoxic or acidic conditions induced binding of 9D2 and annexin V.
Treatment with hydrogen peroxide resulted in the highest AP exposure. No
binding was observed when intact endothelial monolayers were treated with a
variety of other factors (VEGF, bFGF, etc) or medium alone. Combined treatment
with inflammatory cytokines and hypoxia had greater than additive effects,
suggesting that factors may interact to give amplified effects on AP exposure
on tumor endothelium in vivo. To determine biodistribution of 9D2 and annexin V
in tumor bearing mice, their localization was determined immunohistochemically
after intravenous injection. 9D2 and annexin V were found on tumor blood vessels,
in necrotic tumor regions and on individual malignant cells. Both reagents did
not localize to normal endothelium. Isotype-matched control antibodies did not
bind detectably to tumor or normal tissues. These results indicate that AP are
present on the external surface of tumor endothelium but not on normal
endothelium, and that AP exposure might be mediated by thrombin, hypoxia,
cytokines and hydrogen peroxide. The externalized AP on the tumor endothelium
could potentially be utilized for tumor vessel targeting and imaging.
Differences between Thorpe and Epstein's work on the
tissue-factor induced tumor clotting. The simplest explanation is as follows:
Tumor capillary beds (the endothelial cells) are notorious for expressing
certain types of lipids that contain negative charges. This expression promotes
a "clotting" situation, since proteins of the clotting cascade (a
series of binding and enzymatic properties of serum proteins that result in a
blood clot) need this negatively-charged surface to start the clotting
sequence. Tissue factor (and truncated Tissue Factor, tTF) easily bind to these
surfaces and start the clotting process.
Thorpe originally exploited this process by delivery
of TF or tTF to the tumor-associated endothelum by coupling TF to antibodies
that would find targets in the endothelium, notably, V-CAM-1, an adhesion
molecule found on endothelium, or prostate-specific membrane antigen (PSMA), or
even the VEGF-receptor complex found on endothelium. This was his original basis
for VTA. It is speculated now…with the talk of the anti-phosphatidylserine
antibodies (the PS antibody) that this is another form of delivery, since PS
itself is one of those negatively-charged lipids (but difficult to raise
antibodies against). So there are several ways to deliver TF or tTF.
Epstein has now taken another step, in creating fusion peptides between tTF and
several delivery vehicles known to get into the tumor microenvironment (via the
blood vessels). He describes the use of TNT (targeting necrotic regions of the
tumor), chTV-1, an antibody that targets a membrane antigen of tumor blood
vessels, and a protein called RGD, which is nothing more than a small protein
that binds to exposed adhesion molecules on endothelium. So, in essence, the
delivery of the TF (or perhaps other targeting molecules, such as toxins) to
the blood vessels is only as good as the specific targeting of the delivery
molecule…in this case, a variety of antibodies or fusion-peptides.
12 April 2002
Overview: Although the AACR every
year shows much progress in exotic techniques of gene regulation involved with
cancer (such as more sophistication using DNA-chip arrays and proteomics), the
meeting was somewhat "stagnant" with respect to novel ideas of
therapy. For example, we seem to be able to identify hundreds subtle (and not
so subtle) changes in gene expression and regulation in tumor cells but we
don't seem so sure which gene products are viable targets for therapy. However,
molecular targeting in the realm of new toxic therapies for cancer cells (in
the spirit of Gleevec, the powerful inhibitor of a needed enzyme in some
leukemia and gastric cancer cells) has emerged as the concept, which is dogma
for investigators trying to find specific approaches to stopping the growth of
cancer.
There is little doubt that the concept and
field of antiangiogenesis is rapidly being incorporated into mainstream
therapeutics. However, the data from the number of papers this year dealing
with "inhibition of tumor growth" using various compounds that
supposedly inhibit the vascularization of tumors clearly indicate that, at the
present time, inhibitors of angiogenesis are not impressive as single-agent
therapy. This year, a great number of papers indicated that the best therapy
with antiangiogenics is combination therapy. This further supports the growing
realization that a two-compartment approach to cancer therapy, i.e., cytotoxic
compounds delivered to the bulk of disease together with AI's is where the
field is headed. This holds true for direct inhibitors of endothelial cell
function, such as angiostatin or endostatin, as well as the indirect inhibitors
such as antibodies against growth factors and their receptors that may be found
on tumor-associated blood vessels. There are now a growing number of studies
that demonstrate solid preclinical evidence that inhibitors of needed growth
factors and survival factors of blood vessels (and tumor cells) combined with
conventional cytotoxic therapies can be very powerful in the rodent models; and
these combinations are making their way to the clinic.
Regarding the direct targeting of
endothelial cells with VTA approaches (which is different in some respects than
"inhibiting the formation of new blood vessels", in my opinion, it is
too early in the preclinical studies to make great judgments as to how these
studies will evolve. As with all new approaches, there are "windows"
of opportunity and interest in these approaches and if they don't rapidly move
from the animal quarters to the clinic, they get lost in the shuffle. It's just
the way it is.
Epstein is a very personable
scientist who grinds out most of the papers himself and has a busy lab of
students and post-docs. The LEC/chTNT-3 fusion peptide study (highlighted in
the AACR press release) was extremely interesting. The nature of the
"humanized" portion of the fusion peptide prevents too many
injections of this peptide in mice (they will build their own antibodies to
it), but with 3-5 injections of this immunomodulatory molecule (stimulating a
local inflammatory reaction with migration of many immune cells into the tumor
area), Epstein was able to demonstrate strong inhibition of tumor growth in
several animal models (renal cell, colon and lung cancer). The criticism
remains that while concepts are being developed (and I suppose, the patent pool
expanded), the reality remains of getting these type of molecules into clinical
trials. The study using targeted tissue factor fusion peptides by Epstein
(presented by Peisheng Hu, who is the scientist involved intimately with the
China trials) was fascinating because one always likes to see tumors blasted in
their wallow of clotted blood and dying from the inside out. Something
clinically pleasant about a collapsing, dying tumor. The one construct using a
simple peptide sequence (the RGD) was not impressive, but the other two, the
chTV-1 and the use of the chTNT-3 as a delivery agent for the truncated tissue
factor was impressive and resulted in strong intra-tumor clotting and collapse
of various animal tumors. It would be very exciting to see this sort of therapy
tried as single agent targeting (together with whatever Thorpe has in mind with
the anti-PS antibody), but we wait the resources, I suppose, to see some
fruition in that area.
I spent a lot of time looking at
immunological approaches to cancer, where the excitement remains in complex
studies of dendritic cells (cells that present tumor antigens to lymphocytes)
and immunotherapy of cancer remains elusive and artsy-craftsy.
There was no real "breakthrough
buzz" at this AACR, which is not that unusual. But the clinical potential
of PPHM remains just that, potential. These areas of Thorpe and Epstein remain
the science of PPHM (also found in the other papers of these two scientists at
AACR), but without absolute solid clinical results from Cotara in glioblastoma,
encouragement from the other Phase I studies using Cotara, or validation of the
approach via China and lung cancer…all of this science remains
potential. I don't put a lot of confidence in patents or potential, just
clinical results.
On this latter point, the abstracts
of Thorpe were
interesting…and P. Thorpe manned his posters
with great interest from the attendees and he pressed the flesh with good
comments to questions that were asked. I was disappointed in the presentation
from Thorpe's associate (S. Ran) on the exposure of phosphatidylserine (PS) on
the surface of tumor blood vessels, in that their presentation did not contain
any therapy data…(but Thorpe did comment on the other side of the room where is
was…that the antibodies are effective against tumors in the mouse models). It
appeared that just preferred not to show that data, which surprised me. The use
of the 2C3 antibody did strongly inhibit the growth of breast carcinoma cells
in mice (and lung metastasis of these cells) but I suspect this form of therapy
(as with all the antibodies to growth factor receptors) would be greatly
enhanced when combined with a chemotherapy. I would like to see that sort of
data. It would probably represent the fastest, most effective clinical proof of
a pipeline product.
Thorpe's collaborative effort with M.
Rosenblum on the use of the VEGF/gelonin toxin (this is where the toxic
molecule, gelonin, is brought to the tumor and blood vessels by attachment to
VEGF, the growth factor and this complex binds the receptor) was interesting
but the concept or data was not entirely new from a clinical point of view. If
it is a viable, good therapy, then clinical trials should be in order. Some say
later this year (via SUPG?).
05/22/2002
Big pharmas have drug-discovery departments that
work closely with people who do the animal models of tumors. Hence, new drugs
are evaluated quickly as to their toxicity and efficacy in standard preclinical
models. These people are very aware of innovative methods of drug delivery and
new trends of drug development. In that sense, there is little doubt that some
will be quite aware of Epstein's work in the area of using TNT-fusion peptides
that alter permeability of blood vessels that allow enhanced local
concentration of toxic drugs within tumors. Epstein has several papers on this
and related topics.
It would be helpful if PPPH had this ASCO poster on it's
website, but I do not see it at the present time. Raw data of uptake and inhibition
of tumor growth would make it easier to form opinions about the study presented
at ASCO.
Several thoughts come to mind about what is known
from the generalized statements found in the Epstein abstract. Regarding the 3
types of responses; there is little doubt that improved local delivery of the
drugs to drug-sensitive tumors is a preferred method of chemotherapy. This
could be utilized by enhanced local concentration of drug given at the normal
(near MTD) dosing, or even using reduced doses with the same effects (I doubt
this would be done clinically, rather treatment options would prefer the MTD
dose to achieve greater tumor cell kill). The last two groups seem to differ in
the following manner. First, "those tumors which normally do not respond
to a given drug (ex. MAD 109 lung carcinoma treated with Taxol) were now found
to have a significant response…" suggests that the basis of their
drug-resistance did not lie in inherent properties of the tumor cells themselves,
but in the lack of drug delivery to the tumor. This is a surprising result.
Evidently, administration of the TNT-fusion peptide altered the local
permeability or homed to distinct regions of the tumor otherwise not accessible
to Taxol in the local circulation This result is distinct to the "type
3" response, described by "those tumors resistant to a given drug…remained
unaffected or had only a minor response by pretreatment [with the TNT-fusion
peptide]". Drug-resistant tumor populations evolve by prior drug therapy
or can be inherently resistant due to the upregulation of molecular pathways
that defeat the mode of action of the toxic drug. Therefore, this finding is
not unexpected for pure drug-resistant tumor cells.
Eventually, these findings will be evaluated by big
pharma, just as the VTA (via Thorpe) needs to be evaluated by companies able to
begin meaningful, large-scale Phase I trials to prove the innovative features
(with subsequent efficacy). As with many of the PPHM techniques, they have been
around for many years and yet have not been translated to the clinic.
6/03/2002
Research and development of new therapeutic agents
for cancer take millions of dollars. And time. I personally know of several innovative
compounds that were not pursued in a clinical trial due to the extremely high
cost of bringing such a compound to market in light of a limited patient
population that might have received limited benefit. The realities of this type
of drug development (are) fearsome. These companies are not developing cough
medicine or healing cold sores one day faster by subjective analysis, but attempting
to alter the natural course of malignant disease by methods that demand
objective evaluation and endpoints of regression, inhibition of tumor growth,
time to progression of disease, survival, and quality of life, with acceptable
toxicity. Several principles must be established in studying innovative methods
of cancer therapy. First, it is important to remember that, (a) THERAPY SELECTS
FOR RESISTANCE; i.e., the act of targeting tumor cells by chemotherapy will
result in a subpopulation of cells that are resistant to the drug, or resistant
to the concentration of drug that can be safely administered to a patient, this
concept holds true for beam-directed and local irradiation, (b) THE MORE UNIQUE
THE TARGET IS TO CANCER CELLS, THE BETTER THE THERAPY, i.e., the more dependence
that a cancer cell has on a metabolic pathway (enzymes, growth factor
receptors, antiapoptotic factors, etc) to survive compared to normal cells, the
better targeting can be accomplished for a compound that targets that unique
pathway (best example Gleevic, STI-571), (c) IT IS NECESSARY TO IDENTIFY
NON-MUTABLE TARGETS; tumors are extremely heterogenous with respect to the emergence
of subpopulations of cells that are endowed with new and unique characteristics
that allow them to modify and change the environment around them (including
their endothelium, the blood vessel cells that supply them nutrients) and the
more that a given therapy depends upon a property of tumor cells that can be
easily mutated (to escape the killing), then a given therapy will be less effective.
Toward this type of therapeutic philosophy, PPHM
has captured the methodology and patents of two very talented scientists, Drs.
Epstein and Thorpe; whose approaches are quite different but based upon similar
premises. Namely, that they have been able to identify vulnerabilities of tumor
cells (or their blood supply) that may be very unique, at least, to the
expansion of the tumor mass (the property of having necrotic regions and the
need for an expanding blood supply). Dr. Epstein has targeted a basic protein
subset that is found exposed in necrotic regions of tumors (the target of the
TNT family of antibodies and the Lym-1 antigen found almost exclusively on
B-cell lymphomas) and Dr. Thorpe has found unique antigens that are found
almost exclusively on tumor-associated blood vessels (the TNT technology has
reached the clinic in various formats; the Vascular Targeting of Thorpe has
not).
The progress of these agents within the clinic was
undertaken by PPHM over the past two decades through times of promise and
despair, mainly in terms of the funding needed to complete such expensive clinical
trials. Simultaneously with this type of innovative science, new formulations
of targeted therapy have arisen in the possession of cash-rich pharmaceutical
companies able to pursue large, multi-center trials with agents that appear to
attempt to achieve that exclusive targeting of cancer cells or their blood vessels.
The competition is extremely tough and requires that clinicians recognize the
value of the therapy being tested and the funds to push the trials to fruition.
Toward this goal, there are recognized steps of
progress that can be objectively measured. Steps of progress that give
encouragement and hope to patients (and investors). First, the publication of
new and encouraging preclinical data that demonstrate the proof of principle and
promise of a given therapy. Toward that end, Epstein and Thorpe have published
for many years the various themes of targeting DNA-histones (TNT) and vascular
antigens (delivery of truncated tissue factor, antiVEGF-complex antibodies, and
now the elusive anti-phosphatidylserine moiety found on tumor-associated blood
vessels). Secondly, there must be well-designed and executed clinical trials of
Phase I and II that promote the therapy to a serious evaluation of showing
improved survival and freedom from disease and enhancement of quality of life
that patients hope and pray for…and this appears to be on-track for TNT in
glioblastoma and nowhere to be found for VTA.
The clinical trials of Oncolym, a promising
radiolabeled antibody that could have easily rivaled Rituxan and Zevalin (and
Bexxar) for therapy of B-cell lymphoma seems lost. In my opinion, this modality
will not be recovered.
Now, in examining the state of the science for TNT and VTA
and VEA, I perceive the following:
(a)
TNT in glioblastoma obviously
lacks funding for the start of Phase III. That's my opinion. Why? The FDA has
approved the Phase II progress, helped in the requirements for evaluation in
Phase III, granted Orphan Status and denoted Fast-Track Phase III. A
multi-center study here and in Europe will cost millions of dollars. The start
has been delayed, (I assume).
(b)
There is no visibility of TNT in
glioblastoma in the mainstream brain cancer centers. Why? Because there has
been no publication of exciting, promising Phase II data by Patel or his
colleagues from the collaborative centers. There has been no academic
publication of Phase I or Phase II studies that were described at ASCO over a
year ago;
(TUSTIN, Calif. -
May 14, 2001--Peregrine Pharmaceuticals Inc. (Nasdaq:PPHM - news) today
announced Dr. Sunil Patel, associate professor of neurosurgery at the Medical
University of South Carolina, presented preliminary efficacy and safety results
from a Phase II brain cancer trial using Cotara(TM), Peregrine's Tumor Necrosis
Therapy drug. Patel presented his findings at the American Society of Clinical
Oncology's (ASCO) 37th annual meeting, being held in San Francisco, where he
was representing the Cotara brain tumor study consortium. The primary objective
of the Phase II Cotara study is to determine the median time to progression of
treated patients compared with a historical control population. Patel presented
data on the first 29 patients in the study. This group possessed a uniformly
poor prognosis, the majority of them having been diagnosed with recurrent
glioblastoma multiforme (GBM), which is a form of malignant brain tumor.
Primary side effects of the Cotara infusion were well tolerated in this patient
population. The overall median time to progression was 13.9 weeks, despite
inclusion of many heavily pretreated patients).
(c)
There has been no abstract or
conference presentation of the sophisticated methods of image-guided catheterization
of glioma treatment as described almost a year ago (Peregrine Pharmaceuticals
and Image-Guided Neurologics Agree to Study Use of Navigus® Array Catheter in
Treatment of Brain Cancer New Catheter May Enhance Delivery of Targeted Cancer
Therapy; 22-Aug-2001) and there has been no publication of the findings of
Phase I trials in any of the solid tumors treated in Mexico or Stanford or the
Mayo Clinic (26-Jun-2001, TUSTIN, Calif.--[BW HealthWire] Peregrine
Pharmaceuticals Inc. (Nasdaq:PPHM - news) today announced preliminary results
from a Phase I/II Malignant Tumor Trial utilizing Cotara, Peregrine's Tumor
Necrosis Therapy (TNT) drug; PEREGRINE ANNOUNCES OPENING OF A NEW STUDY IN
HEPATIC CANCER New trial to explore use of CotaraTM along with ablation of
hepatic tumors , 19-Apr-2001, PEREGRINE OPENS STANFORD UNIVERSITY TRIAL TO
STUDY CANCERS OF THE PANCREAS AND BILIARY SYSTEM Trial to explore new uses of CotaraTM,
12-Apr-2001, PEREGRINE ANNOUNCES OPENING OF A NEW STUDY AT THE STANFORD
UNIVERSITY, STANFORD, CA New trial to explore use of CotaraTM in soft tissue
sarcoma, 9-Apr-2001).
When there are no publications, there are two
interpretations.
(1)
The data are not worth writing
up for the academic oncology community.
(2) There is
no pressure on the investigators to publish their clinical research.
Both of these conclusions are extremely unlikely.
There must be some informative techniques and knowledge gained from these
studies and these investigators must certainly feel an obligation to their
academic careers to publish good clinical research. Why these data are not
published is a very disturbing mystery.
Epstein presented interesting data at ASCO on the
properties of VEA, using the fusion peptide of the permeability portion of the
IL-2 molecule fused to the TNT antibody binding site, but he has published on
this concept for several years. It must now reach the clinic or be lost in the
bin of many good papers and ideas that never reach fruition. Thorpe presented
interesting data at AACR on the exposure of phosphatidylserine on blood
vessels, but gave NO data as to the potential use in the clinic. Very
disappointing. We have heard nothing on the use of truncated tissue factor as a
therapeutic weapon against tumor.
And then there are the other collaborations and
partnerships (I don't know what to call them) that never seem to have a
publication about anything, such as the SUPG deal, which has floated around the
table for a long, long time, (Peregrine Pharmaceuticals and SuperGen Inc. Announce
Finalization of Licensing Deal for VEGF Vascular Targeting Agents 13-Feb-2001),
and the possibility of using TNT antibodies as a
targeting agents coupled to liposomes to deliver antitumor agents to the
interior of tumor cells, a simple concept that has no literature (Tustin, CA -
July 30, 2001- Peregrine Pharmaceuticals (NASDAQ: PPHM) today announced the
signing of a Joint Development Agreement with Oakwood Labs, Oakwood Village,
Ohio. Under the terms of the agreement, the companies will jointly evaluate
therapeutic agents that combine Peregrine’s Tumor Necrosis Therapy (TNT) tumor targeting
technology with Oakwood’s anti-cancer Liposome technology. Under the agreement,
Peregrine will provide TNT antibodies and Oakwood will provide liposomes and
preclinical testing resources. At the completion of preclinical testing,
Peregrine and Oakwood will decide whether to advance the potential products
into human clinical studies under a commercialization agreement).
In addition, the imaging possibilities that the TNT
antibodies offer due to local delivery within the tumor environment, but of
course, no publications on this concept (PEREGRINE ANNOUNCES RESEARCH AND
DEVELOPMENT AGREEMENT WITH PAUL SCHERRER INSTITUTE FOR TNT BASED PET IMAGING AGENTS
Continues Radiolabeling Research and Development 23-Mar-2001),
or the exciting prospect of cytokine-fusion
peptides described by Epstein for several years and awaiting discovery within a
clinical setting ( TECHNICLONE AND MERCK KGAA SIGN AGREEMENT ON USE OF TUMOR NECROSIS
THERAPY (TNT) TECHNOLOGY 23-Oct-2000).
Finally, the confusing aspect of biotech disclosure.
Within the confines of the Society of Nuclear Medicine meeting this month was found
two abstracts from China dealing with the long awaited and greatly elusive data
on therapy of lung cancer and liver cancer. These data were explicitly
embargoed for release until the day of presentation by the SNM meeting
directors, and the abstracts could not be viewed until May 7th, 2002, at which
time, PPHM made a startling announcement of the results and comments by Epstein
(7 May 2002, PEREGRINE PHARMACEUTICALS ANNOUNCES ABSTRACTS OF INTERIM RESULTS
FROM TUMOR NECROSIS THERAPY TREATMENT OF LUNG AND HEPATIC CANCERS Data to be
presented at Society of Nuclear Medicine's 49th Annual Meeting, June 15-19,
2002). The data, in abstract form, seem to contain a small amount of patients,
in light of the common knowledge that this form of therapy had been taken place
for years in China, and very difficult to evaluate. So we await these findings,
the implications of which are nothing more than absolute validation or
refutation of the entire concept of TNT-based therapy, in my opinion. So June
in an important month, yes. Since we have no other clinical barometer by which
to evaluate the validity of TNT nor information as to the start of VTA, license
of VEA, demise (?) of Oncolym, or hope of TNT in Phase III.