Interleukin-6
(IL-6)

ß-2 interferon
B-cell differentiation factor (BCDF)
B-cell stimulatory factor-2 (BSF-2)
IL-1-inducible 26 kD protein
Cytotoxic T-cell differentiation factor
Uvomorulin
See also: IL-6 at COPE

Gene: maps to 7p21-p14. It has a length of about 5 kb and contains five exons. Repression of the Il-6 gene promoter by p53 and the retinoblastoma susceptibility gene product has been demonstrated (Santhanam U. et al., 1991).
mRNA: size: ~1.2 kb in breast cancer cells (Lacroix M. et al., 1998).
Protein: glycoprotein; 22-27 kD; variably glycosylated; the level of glycosylation has no effect on bioactivity; translated as a 212 aminoacid (aa) molecule (28-aa signal sequence, 184-aa mature portion).
IL-6 is the prototype of a family of cytokines including interleukin-11 (IL-11), leukemia inhibitory factor (LIF), oncostatin M (OSM), ciliary neurotrophic factor (CNTF), and cardiotrophin-1 (CT-1). All these cytokines induce expression of acute phase proteins in the liver.

Cell lines:
- It was shown that breast fibroblasts-derived IL-6 stimulated the ability of MCF-7 breast cancer cells (BCC) to convert oestrone (E₁) to the biologically more active 17 beta-oestradiol (E₂). This effect was mediated by an increase in reductive E₂ oxidoreductase (EOR) activity (Adams E.F. et al., 1991).

- Subcloning of the breast ductal carcinoma T47-D cells yielded clones with high and low sensitivity to the growth inhibitory effect of IL-6. The subclones varied 40 fold in their sensitivity for inhibition of colony formation in sparse cultures and 200 fold for inhibition of thymidine incorporation in subconfluent cultures. Binding studies with 125I-rIL6 showed that T47-D cells and their subclones, as well as SK-BR-3 and MCF-7 cells, expressed high-affinity receptors for IL-6. The number and affinity constant of these receptors were comparable to those on lymphocytic and myeloid cells, and show no correlation with the high or low sensitivity phenotype. Proliferation of the breast cancer cells was inhibited by IL-6 in a cell density dependent manner, and was not due to a cytotoxic effect. In addition, IL-6 induced a morphological change with loss of epithelial characteristics and of cell-cell adhesion. Sensitivity to growth inhibition by IL-6 was independent from that of IFN-beta 1, IFN-gamma or TNF (Chen L. et al., 1991).

- IL-6 was found to inhibit DNA synthesis in ZR-75-1 and T-47D BCC. The partial escape of ZR-75-1 BCC from this inhibition suggests that IL-6 could prolong the G₁ phase of the cell cycle without necessarily preventing the ultimate entry of cells into S phase. IL-6 also caused considerable cell-cell separation of ZR-75-1 BCC, with some cells remaining connected through long processes. Time-lapse cinemicrography demonstrated that the individual cells were locally motile and occasionally traversed distances many times greater than the cell diameter (Tamm I. et al., 1991).

- Synergistic interaction between IL-6 and TNF alpha, on the growth inhibition (50% reduction in the percentage of S-phase cells) and the upregulation of ICAM-1 expression (4 to 11-fold increase) was shown in MCF-7 breast cancer cells (BCC) using flow cytometric methods. IL-6 and TNF alpha alone had negligible effect on the cell cycle. The individual effect of IL-6 resulted in down-regulation of ICAM-1 expression (30-35%), while TNF alpha always upregulated ICAM-1 (1.5 to 4-fold increase). The combined effect of IL-6 and TNF alpha consistently caused an increased expression of ICAM-1, which was greater than the sum of each one alone and also sustained for 72 h following cytokine withdrawal (Bajaj P. et al., 1993).

- Four human BCC lines (T47-D, ZR-75-1, MCF-7 and HS578T) were examined for the effects of cytokines on expression of ICAM-1. Interferon-gamma, tumour necrosis factor-alpha, interleukin (IL)-1 alpha, IL-1 beta and IL-6 up-regulated the expression of ICAM-1 in all the BCC lines. Conditioned medium (CM) generated from human fibroblasts, and in particular from foetal cells, was highly effective in up-regulating expression of ICAM-1 in the breast cancer cell lines. ICAM-1 induction correlated with IL-6 bioactivity in these CMs. Combinations of IL-6 with other cytokines, such as IL-1, resulted in further increases in ICAM-1 expression (Hutchins D. and Steel C.M., 1994).

- The effects of IL-6 on normal mammary epithelial cells (NMEC) and on 4 BCC lines were studied. In NMEC, IL-6 was mainly growth inhibitory, whereas in MCF-7 BCC, the cytokine had growth-inhibitory and anti-adhesive effects. In T-47D and ZR-75-1 BCC, the anti-adhesive effects were prominent although the growth-inhibitory effects were not. These anti-adhesive effects were associated with epithelioid to fibroblastoid morphological changes and a local decrease in E-cadherin expression. In the highly invasive BCC line MDA-MB-231, which does not express E-cadherin, no effects of IL-6 were seen (Ásgeirsson K.S. et al., 1998).

- ER-negative breast cancer cell-derived interleukin 1 alpha was shown to induce NF-kappaB transcription factor activation in breast fibroblasts. This was accompanied by induction of IL-6 and urokinase plasminogen activator (uPA), both of which promote angiogenesis and metastasis (Bhat-Nakshatri P. et al., 1998).

- IL-6 is produced by some BCC lines (MDA-MB-231, Hs578T), where it is co-expressed with Il-11. The cytokine is also present in MCF/Adr (IL-11 production not tested), but absent in a series of other BCC lines (MCF-7, T-47D, BT-20, SK-BR-3, CAMA-1, EVSA-T, ZR-75-1, IBEP-1, IBEP-2, IBEP-3, MDA-MB-435,...)(Lacroix M. et al., 1998). IL-6 expression in MDA-MB-231 and Hs578T is inhibited by the aspirin metabolite salicylate (Sotiriou C. et al., 1999). IL-6 inhibits the growth of MCF-7, T-47D and ZR-75-1 BCC, but not that of MDA-MB-231, Hs578T and MCF-7/Adr cells. It has been suggested that Il-6 produced by oestrogen receptor (ER) negative BCC could have paracrine anti-proliferative effects on local ER positive BCC, thereby inducing a selective growth advantage (Chiu J.J. et al., 1996; Lacroix M. et al., 1998).

Tumors:
- Il-6 immunoreactivity was found in 40 of 66 paraffin embedded breast tumour specimens. This immunoreactivity did not correlate with the clinical evaluation of oestrogen receptor positivity. However, there was a correlation between Il-6 and Il-1-alpha immunoreactivity (Robinson E.K. et al., 1998).

- The risk of developing breast cancer is higher in women presenting gross cystic disease (cysts > 3 mm in diameter) of the breast with intracystic K⁺/Na⁺ > 3 as compared with K⁺/Na⁺ < 3. The concentrations of tumour necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), and IL-6 were determined in the breast cyst fluid of 54 women with gross cystic disease. No significant differences were found in the cystic levels of IL-1 between cysts with intracystic K⁺/Na⁺ < 3 and > 3. However, in cysts with intracystic K⁺/Na⁺ > 3 we found a lower concentration of IL-6 and TNF-alpha than in those with intracystic K⁺/Na⁺ < 3. The concentration of IL-6 in breast cyst fluid was predicted statistically by a positive regression coefficient for the concentration of TNF-alpha. The concentration of TNF-alpha in breast cyst fluid was predicted statistically by a positive regression coefficient for the concentration of IL-6 (Herruzo A. et al., 1999).

- Il-6 was expressed in a majority of 149 cases of invasive breast carcinomas. Il-6 expression was inversely associated with histological tumour grade, but not with tumour size and nodal status. A direct association was found between the percentage of Il-6-positive cells and that of oestrogen receptor and progesterone receptor-positive cells (Fontanini G. et al., 1999).

Adams E.F. et al. (1991) Interleukin 6 is secreted by breast fibroblasts and stimulates 17 beta-oestradiol oxidoreductase activity of MCF-7 cells: possible paracrine regulation of breast 17 beta-oestradiol levels. Int. J. Cancer 49, 118-121.
Armenante F. et al. (1999) Repression of the IL-6 gene is associated with hypermethylation. Biochem. Biophys. Res. Commun. 258, 644-647.
Ásgeirsson K.S. et al. (1998) The effects of Il-6 on cell adhesion and E-cadherin expression in breast cancer. Cytokine 10, 720-728.
Bajaj P. et al. (1993) Interleukin-6 and tumour necrosis factor alpha synergistically block S-phase cell cycle and upregulate intercellular adhesion molecule-1 expression on MCF-7 breast carcinoma cells. Cancer Lett. 71, 143-149.
Bhat-Nakshatri P. et al. (1998) NF-kappaB activation and interleukin 6 production in fibroblasts by estrogen receptor-negative breast cancer cell-derived interleukin 1 alpha. Proc. Natl. Acad. Sci. USA 95, 6971-6976.
Chen L. et al. (1991) IL-6 receptors and sensitivity to growth inhibition by IL-6 in clones of human breast carcinoma cells. J. Biol. Regul. Homeost. Agents 5, 125-136.
Chiu J.J. et al. (1996) Interleukin-6 acts as a paracrine growth factor in human mammary carcinoma cell lines. Clin. Cancer Res. 2, 215-221.
Fontanini G. et al. (1999) Expression of interleukin 6 (IL-6) correlates with oestrogen receptor in human breast carcinoma. Br. J. Cancer 80, 579-584.
Herruzo A. et al. (1999) Tumour necrosis factor-alpha and interleukin-1 and -6 in fibrocystic breast disease. Breast Cancer Res. Treat. 54, 159-164.
Hirano T. et al. (1986) Complementary DNA for a novel human interleukin (BSF-2) that induces B lymphocytes to produce immunoglobulin. Nature 324, 73-76.
Hutchins D. and Steel C.M. (1994) Regulation of ICAM-1 (CD54) expression in human breast cancer cell lines by interleukin 6 and fibroblast-derived factors. Int. J. Cancer 58, 80-84.
Lacroix M. et al. (1998) Production and regulation of interleukin-11 by breast cancer cells. Cancer Lett. 127, 29-35 (see abstract).
Robinson E.K. et al. (1998) Correlation of interleukin 6 with interleukin 1 alpha in human mammary tumours, but not with oestrogen receptor expression. Cytokine 10, 970-976.
Santhanam U. et al. (1991) Repression of the interleukin 6 gene promoter by p53 and the retinoblastoma susceptibility gene product. Proc. Natl. Acad. Sci. USA 88: 7605-7609.
Sotiriou C. et al. (1999) The aspirin metabolite salicylate inhibits breast cancer cells growth and their synthesis of the osteolytic cytokines interleukins-6 and -11. Anticancer Res. 19, 2997-3006 (see abstract).
Tamm I et al. (1991) Interleukin-6 and 12-0-tetradecanoyl phorbol-13-acetate act synergistically in inducing cell-cell separation and migration of human breast carcinoma cells. Cytokine 3, 212-223.
Zilberstein A. et al. (1986) Structure and expression of cDNA and genes for human interferon-beta-2, a distinct species inducible by growth-stimulatory cytokines. EMBO J. 5, 2529-2537.

Leukemia inhibitory factor, oncostatin M, interleukin-11, Il-6 at COPE