Cell lines:
- S100A4 was found to be expressed in aggressive breast cancer cell lines (MDA-MB-231, Hs578T), not in MCF-7 or ZR-75 cells
in vitro (Pedrocchi M. et al., 1994).
- A human sense S100A4 expression vector was transferred in MCF-7 BCC which do not express endogenous S100A4. In vitro, S100A4 expression did not modify proliferative or invasive properties of transfected MCF-7 cells. In vivo, MCF-7 cells expressing S100A4 were associated with tumors exhibiting necrosis, and abundant fibrous and poorly cellular stroma. Immunohistochemical staining of endothelial cells showed that, in the presence of S100A4, the number and the size of tumoral microvessels were decreased and some of them were collapsed. No metastases were observed in mice with either S100A4-expressing or nonexpressing tumors (Onischenko A. et al., 1996).
- MCF-7 cells were transfected with the S100A4 gene under the control of a strong constitutive promoter. All of the 3 tested clones (MCF-7/S100A4) producing S100A4 protein acquired an ability for hormone-independent growth in nude mice. Tumors derived from S100A4 transfectants revealed local invasiveness into surrounding muscle and adipose tissues and metastasized to regional lymph nodes and lungs, characteristics which are rarely observed with parental MCF-7 cells. Electron-microscopic analysis of MCF-7/S100A4 cells demonstrated structural changes in anchoring junctions, particularly in intermediate filament attachment site (desmosomes). The S100A4-transfected clones expressed
estrogen receptor, and their growth in tissue culture was both estrogen- and anti-estrogen responsive. Changes in regulation of the estrogen-dependent proteins
progesterone receptor and
cathepsin D were observed in some of the transfected clones (Grigorian M. et al., 1996).
- Transfection into the benign rat mammary cell line Rama 37, S100A4 showed metastasis-inducing capabilities (Lloyd B.H. et al., 1998).
- Treatment of MDA-MB-231 BCC by thapsigargin, the ionophore A23187, or cyclic ADP-ribose, to increase [Ca2+]i via different pathways, led to relocation of S100A4, as demonstrated by confocal laser scanning microscopy. These findings support the hypothesis that S100A4 could play a crucial role in the regulation of Ca
2+ homeostasis in cancer cells (Mueller A. et al., 1999).
- Recombinant S100A4 has been reported to interact
in vitro with cytoskeletal components and to form oligomers, particularly homodimers
in vitro. Using the yeast two-hybrid system, a strong interaction between S100A4 and another S100 protein, S100A1, was detected. Site-directed mutagenesis of conserved amino acid residues involved in the dimerization of S100 proteins abolished the interactions. The interaction between S100A4 and S100A1 was also observed in vitro using affinity column chromatography and gel overlay techniques. Both S100A1 and S100A4 can occur in the same cultured mammary cells, suggesting that in cells containing both proteins, S100A1 might modulate the metastasis-inducing capability of S100A4 (Wang G. et al., 2000).
Tumors:
- S100A4 expression in tumors not correlated with
ER expression, positively correlated with
urokinase (Pedrocchi M. et al., 1994).
- In a proportion of human intraductal carcinomas of the breast, a shorter variant S100A4 transcript of approximately 450 nucleotides was found. RT-PCR study showed that the noncoding exon 1a/1b was lost in the variant cDNA. Exons 2 and 3, which code for the protein, seemed to be present in the variant isoform. The RT-PCR products obtained using exons 2- and 3-specific oligonucleotides showed a high degree of sequence homology with exons 2 and 3 of the h-mts1 gene. The expression of the variant transcript could be influencing disease progression, albeit not as effectively as the normal unspliced h-mts1 transcript (Albertazzi E. et al., 1998a).
- In carcinoma of the breast, the level of expression of S100A4 has been found to be closely related to metastatic spread of the cancer to regional lymph nodes (Sherbet G.V. and Lakshmi M.S., 1998).
- In infiltrating ductal carcinomas of the breast, high S100A4 expression was associated with metastatic spread to the regional lymph nodes. Breast cancers with no detectable expression of h-mts1 were found to be estrogen and progesterone receptor positive. Expression of h-mts1 was not related to tumor differentiation. Overall, S100A4 expression appeared to be associated with and indicative of more aggressive disease. Moreover, it was suggested that, complemented with nm23, S100A4 could provide a powerful marker of breast cancer prognosis (Albertazzi E. et al., 1998b).
- The occurrence and prognostic significance over 14-20 years of immunocytochemically detected S100A4 and other tumour variables in primary tumours from 349 patients with operable breast cancer were compared. For a cut-off of 1% staining of the malignant cells, the antibody to S100A4 stains positively 56% of the carcinomas. There was a significant association of staining for S100A4 with tumours fixed to the chest wall, staining for
c-erbB-2, c-erbB-3,
pS2,
cathepsin D and, inversely, at borderline levels with staining for
estrogen receptor. Using Wilcoxon statistics in univariate analyses, staining for S100A4, nodal status, tumour class, histological grade and staining for
c-erbB-2, p53 were associated negatively and staining for
estrogen receptor,
progesterone receptor were associated positively with patient survival times. The survival times of patients with S100A4-negative carcinomas with or without one of the other tumour variables showed no significant differences, whilst those of patients with S100A4-positive carcinomas showed significant differences in a negative or a positive way. Multivariate regression analysis for 137 patients showed that staining for S100A4 is most highly correlated with patient deaths, but involved lymph nodes, fixed tumours, high histological grade and staining for
progesterone receptor were also significant independent prognostic variables. These results suggest that in this set of patients, the tumour variable most tightly correlated with patient death is S100A4 (Platt-Higgins A.M. et al., 2000).
- Rabbit antibodies to recombinant rat S100A4 were shown to recognize specifically human S100A4 using Western blotting techniques and were used to assess the prognostic significance of S100A4 in primary tumors from a group of 349 patients treated between 1976 and 1982 for stage I and stage II breast cancer. The antibody stained normal breast tissue heterogeneously, but stained positively 41% of the carcinomas, leaving the remaining 59% as negatively stained. In addition to the carcinoma cells, some host stromal cells and lymphocytes were also stained, but were discounted in subsequent analyses. There was an association of staining of carcinomas for S100A4 with some tumor variables considered to be associated with poor prognosis for patients: tumor present in axillary lymph nodes, staining for c-erbB-3,
cathepsin D, and
c-erbB-2. The association of staining for S100A4 with patient survival was evaluated using life tables and analyzed using generalized Wilcoxon statistics. Eighty percent of the S100A4-negative patients but only 11% of the S100A4-positive patients were alive after 19 years of follow-up, and this association was highly significant; the former had a median survival of >228 months and the latter 47 months. The other tumor variables that showed significant association with survival time were nodal status, tumor size, histological grade, staining for
c-erbB-2,
estrogen receptor, and p53. Analysis of the association of patients with carcinomas staining for S100A4 and their survival in subgroups defined by these other tumor variables shows that in each subgroup, staining for S100A4 is associated with poorer survival. Patients whose tumors stain for S100A4 and possess involved lymph nodes, which are fixed to the chest wall or which stain for
c-erbB-2, showed a significant reduction in survival times over those with only S100A4-staining tumors. Patients with involved lymph nodes, or staining for
c-erbB-2 in the S100A4-negative group failed to show any significant reduction in survival times. Multivariate regression analysis for 137 patients showed that staining for S100A4 was most highly correlated with patient deaths, but involved lymph nodes, fixed tumors, and high histological grade were also significant independent prognostic variables. These results suggest that in this group of patients, the metastasis-inducing protein S100A4 could be most tightly correlated with patient demise (Rudland P.S. et al., 2000).
