Vetrei Cinzia, Passariello Margherita, Froechlich Guendalina, Rapuano Lembo Rosa, Zambrano Nicola, De Lorenzo Claudia
Department of Molecular Medicine and Medical Biotechnology, University of Naples "Federico II", 80131 Naples, Italy.
Ceinge-Biotecnologie Avanzate s.c. a.r.l., via Gaetano Salvatore 486, 80145 Naples, Italy.
Cancers (Basel). 2021 Jun 8;13(12):2858. doi: 10.3390/cancers13122858.
Antibodies targeting Immune Checkpoints (IC) on tumor infiltrating lymphocytes improve immune responses against cancer. Recently, the expression of some ICs has also been reported on cancer cells. We used the clinically validated Ipilimumab and Nivolumab and other novel human antibodies targeting Cytotoxic T- lymphocyte-antigen 4 (CTLA-4), Programmed Death receptor-1 (PD-1) and Programmed Death Ligand 1 (PD-L1) to shed light on the functions of these ICs in cancer cells. We show here for the first time that all these antagonistic mAbs are able to reduce Erk phosphorylation and, unexpectedly, to induce a significant increase of ICs expression on tumor cells, involving a hyperphosphorylation of NF-kB. On the contrary, agonistic PD-L1 and PD-1 recombinant proteins showed opposite effects by leading to a significant reduction of PD-1 and PD-L1, thus also suggesting the existence of a crosstalk in tumor cells between multiple ICs. Since the immunomodulatory mAbs show their higher anti-tumor efficacy by activating lymphocytes against cancer cells, we also investigated whether it was possible to identify the most efficient combinations of immunomodulatory mAbs for achieving potent anti-tumor efficacy associated with the lowest adverse side effects by setting up novel simple and predictive in vitro models based on co-cultures of tumor cells or human fetal cardiomyocytes with lymphocytes. We demonstrate here that novel combinations of immunomodulatory mAbs with more potent anti-cancer activity than Ipilimumab and Nivolumab combination can be identified with no or lower cardiotoxic side effects. Thus, we propose these co-cultures-based assays as useful tools to test also other combinatorial treatments of emerging immunomodulatory mAbs against different ICs for the early screening of most potent and safe combinatorial therapeutic regimens.
靶向肿瘤浸润淋巴细胞上免疫检查点（IC）的抗体可改善针对癌症的免疫反应。最近，也有报道称某些IC在癌细胞上有表达。我们使用了临床验证有效的伊匹木单抗和纳武单抗以及其他靶向细胞毒性T淋巴细胞相关抗原4（CTLA-4）、程序性死亡受体-1（PD-1）和程序性死亡配体1（PD-L1）的新型人源抗体，以阐明这些IC在癌细胞中的功能。我们首次在此表明，所有这些拮抗单克隆抗体都能够降低细胞外信号调节激酶（Erk）的磷酸化，并且出乎意料的是，会诱导肿瘤细胞上IC的表达显著增加，这涉及核因子κB（NF-κB）的过度磷酸化。相反，激动性PD-L1和PD-1重组蛋白则产生相反的效果，导致PD-1和PD-L1显著减少，因此也表明多种IC在肿瘤细胞中存在相互作用。由于免疫调节单克隆抗体通过激活淋巴细胞对抗癌细胞而显示出更高的抗肿瘤疗效，我们还研究了是否有可能通过建立基于肿瘤细胞或人胎儿心肌细胞与淋巴细胞共培养的新型简单且具有预测性的体外模型，来确定免疫调节单克隆抗体的最有效组合，以实现与最低副作用相关的强效抗肿瘤疗效。我们在此证明，可以鉴定出比伊匹木单抗和纳武单抗组合具有更强抗癌活性且无心脏毒性副作用或心脏毒性副作用更低的免疫调节单克隆抗体的新型组合。因此，我们提出这些基于共培养的检测方法作为有用的工具，也可用于测试针对不同IC的其他新兴免疫调节单克隆抗体的联合治疗，以早期筛选出最有效和安全的联合治疗方案。
