The Anti-tumor Effect Of Anti-ErbB2 Fullv Human Monoclonal Antibody H2-18 On Breast Cancer And Its Mechanism Of Action

Breast cancer is the most lethal malignancy in women world-wide in recent years, and its incidence is still climbing high in both developed and developing countries. The later one, which including china is worse.The mortality rate of human breast cance is staying at a high level, especially the patients with amplification of the Erb B2 gene. Approximately 25%-30% of human breast cancers find amplification of the ErbB2 oncogene. Women whose malignancies contain this alteration have an aggressive form of the disease with significantly shortened disease-free and overall survival.Trastuzumab,(produced by Roche group and its trade name is Herceptin) is a humanized monoclonal antibody(mAb) directed against the extracellular domain IV of Erb B2. And it is the ?rst anti-ErbB2 treatment approved for clinical use for patients with ErbB2-overexpressing metastatic breast cancer by FDA in 1998.However, clinical data shows about 70% of the patients with high-ErbB2-expressing breast cancer still do not respond to trastuzumab treatment, and the majority of trastuzumab-responsive patients develop resistance within 1 year of treatment initiation.Pertuzumab,(also produced by Roche group and its trade name is Perjeta) is another humanized monoclonal antibody(mAb) directed against the extracellular domain II of ErbB2.As Trastuzumab inhibited ErbB2 homodimer and ligand-independent heterodime,r, Pertuzumab ef?ciently inhibited ErbB2–ErbB3 complex formation when cells were stimulated with ErbB3 ligand. The combination of these two anti-ErbB2 antibodies that have complementary mechanisms of action synergistically inhibits the growth of Erb B2-overexpressing breast cancer.Despite the effectiveness of trastuzumab and pertuzumab combination therapy in patients with ErbB2-positive breast cancer whose disease progressed after prior trastuzumab-based therapy, the objective response rate is only 24.2%, and less than 8% of patients experience a complete response. Thus, there is still an urgent need to improve Erb B2-directed therapy. In our previous study, we obtained a fully anti-ErbB2 human monoclonal antibody, denoted as H2-18, through screening a phage display antibody library. Surprisingly, we found that it could strongly induce programmed cell death(PCD) in ErbB2-overexpressing breast cancer cells.In this study, we attempted to investigate the antitumor activity of H2-18 in ErbB2-overexpressing breast cancer cell lines, especially Trastuzumab-resistant cell lines, and elucidated its mechanism of action.The results of cell viability assay indicated that H2-18 was significantly more effective in inhibited the in vitro proliferation of Trastuzumab-resistant ErbB2-overexpressing cell line HCC1954 than Trastuzumab plus Pertuzumab. In another Trastuzumab-resistant ErbB2-overexpressing cell line,HCC1419, H2-18 displayed a similar inhibitory activity as Trastuzumab in combination with Pertuzumab. In Trastuzumab-sensitive ErbB2-overexpressing BT474 and SK-BR-3 cell lines, The growth-inhibitory effect of H2-18 was significantly weaker than that of Trastuzumab and Pertuzumab.The PCD-inducing activity of H2-18 was evaluated by Annexin V/PI assays in BT474, SK-BR-3, HCC1954 and HCC1419 cell lines. The results showed that H2-18 exhibited much more potent PCD-inducing activity compared with Trastuzuamb and Pertuzumab. Next, we investigated the in vivo antitumor effect of H2-18 in nude mice bearing HCC1954 and HCC1419 tumors. We found that H2-18 was significant more effective in inhibiting the growth of HCC1954 and HCC1419 tumors than Trastuzuamb and Pertuzuamb.Furthermore, we confirmed that H2-18-induced PCD was caspase-independent, and BCL-2 family also played no part in it. Based on morphological characteristics of TEM, we concluded that H2-18 could induce programmed necrosis in ErbB2-overexpressing breast cancer. Further data indicated that H2-18 treatment lead to the activation of RIP1,which is upstream of ROS production and JNK activation. We found that H2-18 regulates programmed necrosis through transcriptin factor c-Jun.In summary, H2-18 has potent PCD-inducing activity and efficiently inhibits the growth of Trastuzumab-resistant breast cancer, suggesting that it might be a promising therapeutic agent for Trastuzumab-resistant breast cancer. Our study also may provide new strategies to develop more effective anti-ErbB2 therapeutic antibodies.