Gründker C, Völker P, Schulz K D, Emons G
Department of Gynecology and Obstetrics, Georg-August University, Göttingen, D-37075, Germany.
Gynecol Oncol. 2000 Aug;78(2):194-202. doi: 10.1006/gyno.2000.5863.
Spontaneous and epidermal growth-factor-induced proliferation of human gynecological cancer cell lines is dose- and time-dependently reduced by treatment with the luteinizing hormone-releasing hormone (LHRH) agonist triptorelin and antagonist Cetrorelix. This antiproliferative activity is probably directly mediated through the LHRH receptors expressed by the tumor cells interacting with growth-factor-dependent mitogenic signal transduction. We have examined whether epidermal growth-factor (EGF)-induced expression of the early response gene c-fos is reduced by LHRH analogs.
Human endometrial (Ishikawa, Hec-1A), ovarian (EFO-21, EFO-27, SK-OV-3), and breast cancer cell lines (MCF-7) were rendered quiescent by incubation (72 h) in the absence of fetal calf serum and phenol red. This was followed by a 15-min incubation in the absence or presence of the LHRH agonist triptorelin (100 nM) or the antagonist Cetrorelix (100 nM) before the cells were stimulated for 10 min with EGF (100 nM). C-fos mRNA expression was determined by semi-quantitative RT-PCR using a synthetic DNA fragment as internal standard. C-Fos protein synthesis was determined by SDS-PAGE and semi-quantitative Western blotting.
In cells derived from endometrial and ovarian cancer, maximal c-fos mRNA expression (seven- to ninefold over basal level) was obtained 30 min after EGF stimulation. In the breast cancer cell line MCF-7 this effect was obtained 60 min after EGF treatment. In all of the lines expressing LHRH receptor, EGF-induced c-fos mRNA expression as well as c-Fos protein synthesis was dose-dependently reduced by treatment with LHRH agonists and antagonists. At 100 nM concentrations of the LHRH analogs, c-fos expression was reduced to baseline levels. No effect of LHRH analogs on EGF-induced c-fos expression was observed in the ovarian cancer cell line SK-OV-3, which does not express the LHRH receptor.
These results suggest that the binding of LHRH agonists and antagonists to their receptors inhibits the mitogenic signal transduction pathway of the EGF receptor in endometrial, ovarian, and breast cancer cell lines. The coupling of both signal transduction systems mediates the antiproliferative effect of LHRH analogs.
促黄体生成素释放激素（LHRH）激动剂曲普瑞林和拮抗剂西曲瑞克可剂量和时间依赖性地降低人妇科癌细胞系的自发增殖以及表皮生长因子诱导的增殖。这种抗增殖活性可能是通过肿瘤细胞表达的LHRH受体与生长因子依赖性有丝分裂信号转导相互作用直接介导的。我们研究了LHRH类似物是否会降低表皮生长因子（EGF）诱导的早期反应基因c-fos的表达。
将人子宫内膜癌（石川细胞系、Hec-1A细胞系）、卵巢癌细胞系（EFO-21、EFO-27、SK-OV-3）和乳腺癌细胞系（MCF-7）在无胎牛血清和酚红的条件下孵育72小时使其静止。随后，在无或有LHRH激动剂曲普瑞林（100 nM）或拮抗剂西曲瑞克（100 nM）存在的情况下孵育15分钟，然后用EGF（100 nM）刺激细胞10分钟。使用合成DNA片段作为内标，通过半定量逆转录聚合酶链反应（RT-PCR）测定c-fos mRNA表达。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）和半定量蛋白质免疫印迹法测定C-Fos蛋白合成。
在源自子宫内膜癌和卵巢癌的细胞中，EGF刺激30分钟后可获得最大的c-fos mRNA表达（比基础水平高7至9倍）。在乳腺癌细胞系MCF-7中，EGF处理60分钟后可获得此效应。在所有表达LHRH受体的细胞系中，LHRH激动剂和拮抗剂处理可剂量依赖性地降低EGF诱导的c-fos mRNA表达以及C-Fos蛋白合成。在100 nM浓度的LHRH类似物作用下，c-fos表达降至基线水平。在不表达LHRH受体的卵巢癌细胞系SK-OV-3中，未观察到LHRH类似物对EGF诱导的c-fos表达有影响。
这些结果表明，LHRH激动剂和拮抗剂与其受体的结合抑制了子宫内膜癌、卵巢癌和乳腺癌细胞系中EGF受体的有丝分裂信号转导途径。两种信号转导系统的偶联介导了LHRH类似物的抗增殖作用。
