Data from: An experimental analysis of the molecular effects of trastuzumab (herceptin) and fulvestrant (falsodex), as single agents or in combination, on human HR+/HER2+ breast cancer cell lines and mouse tumor xenografts

Purpose: To investigate the effects of trastuzumab (herceptin) and fulvestrant (falsodex) either in combination or alone, on downstream cell signaling pathways in lab-cultured human HR+/HER2+ breast cancer cell lines ZR-75-1 and BT-474, as well as on protein expression levels in mouse xenograft tissue. Methods: Cells were cultivated in the presence of trastuzumab or fulvestrant or both. Molecular events that resulted in an inhibition of cell proliferation and cell cycle progression or in an increased rate of apoptosis were studied. The distribution and abundance of the proteins p-Akt and p-Erk expressed in these cells in response to single agents or combinatorial treatment were also investigated. In addition, the effects of trastuzumab and fulvestrant, either as single agents or in combination on tumor growth as well as on expression of the protein p-MED1 expressed in in vivo mouse xenograft models was also examined. Results: Cell proliferation was increasingly inhibited by trastuzumab or fulvestrant or both, with a CI<1 and DRI>1 in both human cell lines. The rate of apoptosis increased only in the BT-474 cell line and not in the ZR-75-1 cell line upon treatment with fulvestrant and not trastuzumab as a single agent (P<0.05). Interestingly, fulvestrant, in combination with trastuzumab, did not significantly alter the rate of apoptosis (in comparison with fulvestrant alone), in the BT-474 cell line (P>0.05). Cell accumulation in the G1 phase of cell cycle was investigated in all treatment groups (P<0.05), and the combination of trastuzumab and fulvestrant reversed the effects of fulvestrant alone on p-Akt and p-Erk protein expression levels. Using ZR-75-1 or BT-474 to generate in vivo tumor xenografts in BALB/c athymic mouse models, we showed that a combination of both drugs resulted in a stronger inhibition of tumor growth (P<0.05) and a greater decrease in the levels of activated MED1 (p-MED1) expressed in tumor issues compared with the use of either drug as a single agent. Conclusions: We demonstrate that the administration of trastuzumab and fulvestrant in combination results in positive synergistic effects on both, ZR-75-1 and BT-474 cell lines. This combinatorial approach is likely to reduce physiological side effects of both drugs, thus providing a theoretical basis for the use of such combination treatment in order to resolve HR+/HER2+ triple positive breast cancer that has previously been shown to be resistant to endocrine treatment alone.

研究目的：探究曲妥珠单抗（trastuzumab，商品名赫赛汀Herceptin）与氟维司群（fulvestrant，商品名芙仕得Faslodex）单药或联合给药，对实验室培养的人激素受体阳性（HR+）/人表皮生长因子受体2阳性（HER2+）乳腺癌细胞系ZR-75-1与BT-474的下游细胞信号通路，以及小鼠异种移植瘤组织中蛋白表达水平的影响。
实验方法：将细胞置于含曲妥珠单抗、氟维司群或二者联合的培养基中培养。研究了可抑制细胞增殖、细胞周期进程或促进细胞凋亡的分子事件。同时探究了单药或联合给药后，上述细胞中表达的磷酸化蛋白Akt（p-Akt）与磷酸化蛋白Erk（p-Erk）的分布与丰度。此外，还考察了曲妥珠单抗与氟维司群单药或联合给药对体内小鼠异种移植瘤模型中肿瘤生长，以及肿瘤组织中磷酸化蛋白MED1（p-MED1）表达水平的影响。
实验结果：曲妥珠单抗、氟维司群单药或联合给药均可抑制细胞增殖，且两种人乳腺癌细胞系中均呈现联合指数（CI）<1、剂量减少指数（DRI）>1的协同效应。仅在BT-474细胞系中，氟维司群单药给药可显著提升细胞凋亡率，而曲妥珠单抗单药无此效果（P<0.05）。值得注意的是，在BT-474细胞系中，氟维司群与曲妥珠单抗联合给药与氟维司群单药给药相比，细胞凋亡率无显著变化（P>0.05）。所有给药组均观察到细胞周期G1期阻滞（P<0.05），且曲妥珠单抗与氟维司群联合给药可逆转氟维司群单药对p-Akt与p-Erk蛋白表达水平的影响。通过在BALB/c裸小鼠模型中利用ZR-75-1或BT-474细胞构建体内肿瘤异种移植瘤，我们发现与单药给药相比，联合给药可更强效地抑制肿瘤生长（P<0.05），并更显著地降低肿瘤组织中活化型MED1（p-MED1）的蛋白水平。
研究结论：本研究证实，曲妥珠单抗与氟维司群联合给药对ZR-75-1与BT-474细胞系均具有正向协同效应。该联合疗法有望降低两种药物单独使用时的生理不良反应，为采用此类联合治疗方案以解决既往被证实对单纯内分泌治疗耐药的HR+/HER2+三阳性乳腺癌提供了理论依据。