Biotherapy Section, Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.
Division of Preclinical Innovation, Early Translation Branch, National Center for Advancing Translational Sciences, National Institutes of Health, Rockville, Maryland, USA.
FASEB J. 2023 Dec;37(12):e23292. doi: 10.1096/fj.202301052R.
Immunotoxins (ITs) target cancer cells via antibody binding to surface antigens followed by internalization and toxin-mediated inhibition of protein synthesis. The fate of cells responding to IT treatment depends on the amount and stability of specific pro-apoptotic and pro-survival proteins. When treated with a pseudomonas exotoxin-based immunotoxin (HB21PE40), the triple-negative breast cancer (TNBC) cell line MDA-MB-468 displayed a notable resistance to toxin-mediated killing compared to the epidermoid carcinoma cell line, A431, despite succumbing to the same level of protein synthesis inhibition. In a combination screen of ~1912 clinically relevant and mechanistically annotated compounds, we identified several agents that greatly enhanced IT-mediated killing of MDA-MB-468 cells while exhibiting only a modest enhancement for A431 cells. Of interest, two Smac mimetics, birinapant and SM164, exhibited this kind of differential enhancement. To investigate the basis for this, we probed cells for the presence of inhibitor of apoptosis (IAP) proteins and monitored their stability after the addition of immunotoxin. We found that high levels of IAPs inhibited immunotoxin-mediated cell death. Further, TNFα levels were not relevant for the combination's efficacy. In tumor xenograft studies, combinations of immunotoxin and birinapant caused complete regressions in MDA-MB-468tumor-bearing mice but not in mice with A431 tumors. We propose that IAPs constitute a barrier to immunotoxin efficacy which can be overcome with combination treatments that include Smac mimetics.
免疫毒素（ITs）通过抗体与表面抗原结合，随后内吞和毒素介导的蛋白质合成抑制来靶向癌细胞。对 IT 治疗有反应的细胞的命运取决于特定的促凋亡和促存活蛋白的数量和稳定性。当用基于绿脓杆菌外毒素的免疫毒素（HB21PE40）处理时，与表皮癌细胞系 A431 相比，三阴性乳腺癌（TNBC）细胞系 MDA-MB-468 对毒素介导的杀伤表现出明显的抗性，尽管其蛋白质合成抑制水平相同。在对约 1912 种具有临床相关性和机制注释的化合物进行的组合筛选中，我们发现了几种能够显著增强 MDA-MB-468 细胞对免疫毒素杀伤作用的药物，而对 A431 细胞的增强作用则较为温和。有趣的是，两种 Smac 模拟物，birinapant 和 SM164，表现出这种差异化增强。为了研究这种现象的基础，我们检测了细胞中凋亡抑制剂（IAP）蛋白的存在，并在添加免疫毒素后监测它们的稳定性。我们发现高水平的 IAPs 抑制了免疫毒素介导的细胞死亡。此外，TNFα 水平与该组合的疗效无关。在肿瘤异种移植研究中，免疫毒素和 birinapant 的组合在 MDA-MB-468 荷瘤小鼠中引起完全消退，但在携带 A431 肿瘤的小鼠中则没有。我们提出，IAPs 构成了免疫毒素疗效的障碍，可以通过包括 Smac 模拟物在内的联合治疗来克服。
