Smad4-independent TGF-b signaling in p16-negative HNSCC

描述（由申请人提供）：头颈部鳞状细胞癌（HNSCC）是全球第六大常见癌症，其发展的危险因素包括吸烟、饮酒和人乳头瘤病毒（HPV）感染。HPV阳性(+)/p16（+）肿瘤患者预后良好[3年总生存率（OS）： ~80%]，而相似分期的HPV阴性（-）肿瘤患者，通常为p16阴性（-），OS较差（~50%）。因此，有必要改善HPV (-)/p16（-）患者的治疗和临床反应。Smad4（或DPC4）在转化生长因子- β （TGF-ß）信号传导中起核心作用。最近，在小鼠口腔中，条件性Smad4缺失（Smad4-/-）被证明可引起自发肿瘤，其组织学与人类疾病相似。在我们的初步研究中，发现Smad4的表达状态在一组HNSCC细胞系中存在差异表达，并与上皮-间质转化（EMT）相关。此外，Smad4的稳定敲低（KD）诱导了emt样表型和侵袭性增加。TGF-ß1刺激后，SCC25-Smad4 KD和FaDu （smad4缺失的HNSCC细胞系）与SCC25-scramble对照组相比，Smad2的激活增强。我们还确定，从HNSCC患者获得的14/77（18%）肿瘤中Smad4缺失明显，表明与p16（-）肿瘤有显著关联[13/ 53,25 %的p16（-）对1/ 24.4%的p16(+)]。因为p16阳性是一个强有力的替代标记

DESCRIPTION (provided by applicant): Head and neck squamous cell carcinoma (HNSCC) is the sixth most frequent cancer worldwide, and risk factors for development include tobacco use, alcohol consumption, and human papillomavirus (HPV) infection. Patients with HPV positive (+)/p16 (+) tumors have a favorable prognosis [3-year overall survival (OS): ~80%] while patients with HPV-negative (-) tumors of similar stage, typically p16-negative (-), experience a worse OS (~50%). Consequently, there is a great need to improve the treatment and clinical response of HPV (-)/p16 (-) patients. Smad4 (or DPC4) plays a central role in transforming growth factor-beta (TGF-ß) signaling. Recently, conditional Smad4 loss (Smad4-/-) in the mouse oral cavity was shown to cause spontaneous tumors with histology reminiscent of the human disease. In our preliminary investigations, Smad4 expression status was found to be differentially expressed across a panel of HNSCC cell lines and associated with epithelial-to- mesenchymal transition (EMT). Additionally, stable knockdown (KD) of Smad4 induced EMT-like phenotypes and increased invasion. After stimulation with TGF-ß1, SCC25-Smad4 KD and FaDu (Smad4-null HNSCC cell line) demonstrated enhanced activation of Smad2 compared to SCC25-scramble control. We also determined Smad4 loss was evident in 14/77 (18%) tumors obtained from HNSCC patients demonstrating a significant association with p16 (-) tumors [13/53, 25% of p16 (-) vs. 1/24, 4% of p16 (+)]. As p16 positivity is a strong surrogate marker for HPV infection, the vast majority of Smad4 (-) tumors are HPV (-). Patients with Smad4 (-) tumors experienced poor survival and an increased incidence of metastasis. In this application, we propose to; 1) determine the mechanism of Smad4-independent Smad2 activation and its effect on TGF-ß inhibitor sensitivity in HNSCC, and 2) determine the role of Smad4-independent Smad2 activation in the metastatic process using an in vivo orthotopic mouse model which allows for an unbiased evaluation of tumor- microenvironment interactions. Identification of novel therapeutic targets and predictive biomarkers of response will allow for appropriate patient selection in future clinical trials, and potentially impact the survival of HNSCC patients with particularly poor clinical outcome.