IGF2 Mediates Resistance to Isoform-Selective-Inhibitors of the PI3K in HPV Positive Head and Neck Cancer

In the current study, we delineate the molecular mechanisms of acquisition of resistance to two isoform-selective inhibitors of PI3K (isiPI3K), alpelisib and taselisib, in human papillomavirus positive head and neck cell lines. By comparing RNA sequencing of isiPI3K-sensitive tumor cells and their corresponding isiPI3K-acquired-resistant tumor cells, we found that overexpression of insulin growth factor 2 (IGF2) is associated with the resistance phenotype. We further demonstrated by gain and loss of function studies that IGF2 plays a causative role in limiting the sensitivity of human papillomavirus-positive head and neck cell lines. Moreover, we show that blocking IGF2 stimulation activity, using an inhibitor of the IGF1 receptor (IGF1R), enhances isiPI3K efficacy and displays a synergistic anti-tumor effect in vitro and superior anti-tumor activity ex vivo and in vivo. we generated two acquired-resistant cell line models from UM-SCC47 and UT-SCC60A cells.UM-SCC47 and UT-SCC60A tumor cell lines were exposed to increasing concentrations of either GDC0032 or BYL719, respectively, for four to eight months, until resistance developed. We mapped the transcriptional landscape of isiPI3K-sensitive and isiPI3K-resistant cell lines by RNA sequencing (RNAseq). Specifically, we used the parental UM- SCC47 and UT-SCC60A and their corresponding acquired resistance models, UM- SCC47-Res2 and UT-SCC60A-Res2. Importantly, RNA extraction from the resistant cells was performed following 48 h of drug-free culture.

本研究阐明了人类乳头瘤病毒（human papillomavirus, HPV）阳性头颈部细胞系对两种磷脂酰肌醇3-激酶（phosphatidylinositol 3-kinase, PI3K）亚型选择性抑制剂（isiPI3K）——阿培利司（alpelisib）与他赛利昔布（taselisib）产生获得性耐药的分子机制。通过对比isiPI3K敏感肿瘤细胞及其对应获得性耐药肿瘤细胞的RNA测序（RNA sequencing）数据，我们发现胰岛素样生长因子2（insulin growth factor 2, IGF2）的过表达与耐药表型密切相关。进一步通过功能获得与功能缺失实验证实，IGF2在降低HPV阳性头颈部细胞系的药物敏感性中发挥了直接致病作用。此外，本研究表明，采用胰岛素样生长因子1受体（insulin-like growth factor 1 receptor, IGF1R）抑制剂阻断IGF2的刺激活性，可增强isiPI3K的抗肿瘤效力，并在体外（in vitro）展现出协同抗肿瘤效应，在离体（ex vivo）及体内（in vivo）模型中则表现出更优的抗肿瘤活性。我们从UM-SCC47与UT-SCC60A细胞系中构建了两株获得性耐药细胞模型：将UM-SCC47和UT-SCC60A肿瘤细胞系分别置于浓度逐步递增的GDC0032与BYL719环境中培养4至8个月，直至其产生耐药性。我们通过RNA测序（RNAseq）绘制了isiPI3K敏感与耐药细胞系的转录组全景。具体而言，本研究采用亲本株UM-SCC47、UT-SCC60A及其对应的耐药模型UM-SCC47-Res2与UT-SCC60A-Res2。值得注意的是，耐药细胞的RNA提取于无药培养48小时后的样本。