Role of PUMA in EGFR targeted therapy in HNSCC

描述(申请人提供)：头颈部鳞状细胞癌(HNSCC)是全球第六大癌症相关死亡原因。在过去的30年里，使用传统疗法并没有显著提高HNSCC患者的存活率。80%以上的HNSCC表达高水平的表皮生长因子受体(EGFR)，这与预后不良有关。用酪氨酸激酶抑制剂(TKI)或抗体靶向EFGR信号转导已成为治疗包括HNSCC在内的癌症的一种有前途的方法。阻断EGFR的抗肿瘤作用被认为涉及多种机制，包括诱导细胞凋亡，其潜在机制尚不清楚。我们和其他人以前发现BH3-Only的BH3-Bcl2家族成员PUMA(P53非调节凋亡调节器)是DNA损伤诱导的细胞凋亡的重要介质。我们最近的初步数据显示，在大多数HNSCC细胞系中，PUMA不能被传统的化疗药物诱导，但可以由临床批准的EGFR靶向药物吉非替尼、厄洛替尼和西妥昔单抗诱导。彪马诱导与EGFR TKI敏感性有关。用小干扰RNA(SiRNA)和小发夹RNA(HsRNA)敲除PUMA可抑制吉非替尼诱导的HNSCC细胞凋亡。此外，EGFR TKI通过作用于P53家族成员P63和P73而诱导PUMA，而不是通过影响P53本身。这些观察结果使我们推测，PUMA诱导是EGFR阻断诱导HNSCC细胞毒性的重要机制。我们建议使用细胞培养、动物肿瘤模型和临床样本来检验这一中心假设。在目标1中，我们将阐明抑制EGFR在HNSCC细胞中诱导PUMA的分子机制。我们将重点介绍p73和p63在PUMA转录调控中的作用。在目的2中，我们将探讨PUMA在EGFR抑制诱导HNSCC细胞凋亡中的作用。我们将重点介绍PUMA在调节线粒体完整性和其他Bcl2家族蛋白中的作用和机制。在目标3中，我们将在细胞培养、异种移植模型和临床样本中确定PUMA水平是否调节EGFR靶向治疗的治疗反应。公共卫生相关性：头颈部鳞状细胞癌(HNSCC)是美国癌症相关死亡的重要原因。本项目旨在了解一类新型抗癌药物--表皮生长因子受体(EGFR)拮抗剂在HNSCC中发挥抗肿瘤作用的机制。这一结果可能有助于开发治疗HNSCC的改进策略和药物，也可能有助于治疗其他EGFR过表达的癌症。

DESCRIPTION (provided by applicant): Abstract Head and neck squamous cell carcinoma (HNSCC) is the sixth leading cause of cancer related death worldwide. The survival rate of HNSCC patients has not been significantly improved over the past 30 years with conventional therapies. More than 80% of HNSCC express high levels of epidermal growth factor receptor (EGFR), which correlate with poor prognosis. Targeting EFGR signaling with tyrosine kinase inhibitors (TKIs) or antibodies has emerged as a promising approach for treating cancers including HNSCC. The antitumor effects of EGFR blockade are thought to involve multiple mechanisms, including induction of apoptosis, whose underlying mechanism is not well understood. We and others previously identified the BH3- only Bcl-2 family member PUMA (p53 unregulated modulator of apoptosis) as an essential mediator of DNA damage-induced apoptosis. Our recent preliminary data showed that PUMA fails to be induced by conventional chemotherapeutic agents, but is induced by clinically approved EGFR targeting agents, gefitinib, erlotinib and cetuximab, in most HNSCC cell lines. PUMA induction is associated with EGFR TKI sensitivity. Knockdown of PUMA by small interference RNA (siRNA) and small hairpin RNA (hsRNA) suppressed gefitinib- induced apoptosis in HNSCC cells. Furthermore, EGFR TKIs induce PUMA through the effects on the p53 family members p63 and p73, but not p53 itself. These observations led us to hypothesize that PUMA induction is an important mechanism of EGFR blockade-induced cytotoxicity in HNSCC cells. We propose to test this central hypothesis using cell culture, animal tumor models and clinical samples. In Aim 1, we will delineate the molecular mechanisms of PUMA induction by EGFR inhibition in HNSCC cells. We will focus on the role of p73 and p63 in the transcriptional regulation of PUMA. In Aim 2, we will investigate the role of PUMA in EGFR inhibition-induced apoptosis in HNSCC cells. We will focus on the role and mechanism of PUMA in the regulation of mitochondrial integrity and other Bcl-2 family proteins. In Aim 3, we will determine whether PUMA levels modulate the therapeutic responses to EGFR targeted therapy in cell culture, xenograft models and clinical samples. PUBLIC HEALTH RELEVANCE: Head and neck squamous cell carcinoma (HNSCC) is a significant cause of cancer related death in the United States. This project seeks to understand the mechanisms by which a novel class of anticancer agent, epidermal growth factor receptor (EGFR) antagonists, exerts their antitumor effects in HNSCC. The results could be useful for developing improved strategies and agents for treatment of HNSCC and possibly other cancers with EGFR overexpression.