Targeting the DNA Damage Response in HPV+ Head and Neck Cancer

项目摘要 人乳头瘤病毒(HPV)阳性的头颈部鳞状细胞癌(HNSCC)是一个日益增长的公众 健康负担，已超过宫颈癌成为#年最常见的HPV相关恶性肿瘤 美国。虽然HPV+HNSCC患者总体上生存良好，但他们患有终生 与放化疗相关的疾病。目前的数据不足以为标准的去强化提供信息 化学放射治疗或靶向治疗的发展。我的最终目标是理解 HPV在HNSCC发展过程中干扰DNA损伤反应(DDR)信号的机制， 从而为新的靶向治疗的合理设计提供信息。在考虑新的战略以有效地 对照HPV+HNSCC，我注意到HPV的致癌E6和E7蛋白抑制肿瘤抑制通路 并破坏DDR信号，导致基因组不稳定。门德斯实验室和其他实验室已经建立了DDR 通过特异性抑制剂AZD1775(WEE1i)抑制WEE1作为HNSCC的一种新的治疗策略， 而且HPV+HNSCC肿瘤是过敏性的。Wee1抑制引起S相复制应激(RS) 以及无法修复的DNA损伤。与基因毒性化疗(如顺铂)相结合，WEE1i可消除 G2/M检查点，并导致过早有丝分裂。我最近发现HPV16E6/E7癌蛋白使 HNSCC细胞通过激活驱动有丝分裂基因的FOXM1-CDK1通路进行WEE1i单药治疗 表达和DNA损伤。我还发现基础FOXM1活性升高易患HPV+HNSCC 对WEE1i诱导的毒性的影响。接下来，我使用RNAi遗传屏幕来识别协同作用的RS和DDR靶点 对于WEE1i；根据我到目前为止的发现，我假设E6/E7对RS和DDR通路的干扰 为还包括WEE1i的组合靶向治疗提供可利用的漏洞。我打算澄清一下 HPV使癌细胞对WEE1i诱导的复制失败敏感的机制(目标1)和 在WEE1抑制时妥协DNA修复途径(目标2)。我会用小鼠癌症模型来测试 针对HPV+HNSCC中RS/DDR缺陷的新治疗组合并确定 它们是最有效的。同时，我将使用一种有针对性的定量蛋白质组学方法来确定 E6/E7特异性RS和DDR对WEE1i的反应，以及多板流式细胞术确定WEE1i- 免疫活性小鼠中E6/E7驱动的肿瘤免疫格局的相关变化。本奖项 将帮助我发展我的科学思想，并提高我使用鼠标模型的能力 忠实地概括了人类癌症。我在这次培训期间取得的科学进步将是 对我的最终目标至关重要，即建立一个独立的研究计划，专注于HPV是如何驱动的 HNSCC的发展以及如何更安全有效地治疗HPV+HNSCC。

Project Summary Human papilloma virus (HPV)-positive head and neck squamous cell carcinoma (HNSCC) is a growing public health burden and has already surpassed cervical cancer as the most common HPV-related malignancy in the United States. While HPV+ HNSCC patients have generally good survival, they suffer from life-long chemoradiotherapy-related morbidities. Current data is insufficient to inform de-intensification of standard chemoradiotherapy or the development of targeted therapies. My ultimate goal is to understand the mechanisms by which HPV disrupts DNA damage response (DDR) signaling during HNSCC development, and to thereby inform the rational design of new targeted therapies. In considering new strategies to effectively control HPV+ HNSCC, I noted that HPV’s oncogenic E6 and E7 proteins abrogate tumor suppressor pathways and impair DDR signaling to cause genomic instability. The Mendez lab and others have established DDR kinase WEE1 inhibition via the specific inhibitor AZD1775 (WEE1i) as a new therapeutic strategy in HNSCC, and that HPV+ HNSCC tumors are hypersensitive. WEE1 inhibition causes S-phase replication stress (RS) and irreparable DNA damage. Combined with genotoxic chemotherapy (e.g., cisplatin), WEE1i abrogates the G2/M checkpoint and causes premature mitosis. I recently showed that HPV16 E6/E7 oncoproteins sensitize HNSCC cells to WEE1i monotherapy through activation of a FOXM1-CDK1 circuit that drives mitotic gene expression and DNA damage. I also showed that elevated basal FOXM1 activity predisposes HPV+ HNSCC to WEE1i-induced toxicity. Next, I used an RNAi genetic screen to identify RS and DDR targets that synergize with WEE1i; based on my findings to date, I hypothesize that disruption of RS and DDR pathways by E6/E7 provide exploitable vulnerabilities for a combination targeted therapy that also includes WEE1i. I plan to clarify the mechanisms by which HPV sensitizes cancer cells to WEE1i-induced replication failure (Aim 1) and compromises DNA repair pathways upon WEE1 inhibition (Aim 2). I will use murine cancer models to test novel therapeutic combinations for targeting RS/DDR defects in HPV+ HNSCC and identify the situations in which they are most effective. In parallel, I will use a targeted quantitative proteomics approach to determine the E6/E7-specific RS and DDR responses to WEE1i, and multi-panel flow cytometry to determine the WEE1i- associated changes in the immune landscape of E6/E7-driven tumors in immunocompetent mice. This award will help me develop my scientific ideas and increase my competency in working with the mouse models that faithfully recapitulate human cancer. The scientific advances that I make during this training period will be critical to my ultimate goal of establishing an independent research program that focuses on how HPV drives HNSCC development and how HPV+ HNSCC might be more safely and effectively treated.