MAGEA6-p63信号轴抑制细胞焦亡在食管鳞癌化疗抵抗中的作用及机制研究

Chemotherapy resistance is a key factor in poor prognosis esophageal squamous cell carcinoma (ESCC) patients. Pyroptosis is a new type of cell death, and its reduction is one of the causes of chemoresistance. At present, the relationship between pyroptosis and chemoresistance of ESCC is not known. Our previous studies confirmed that MAGEA6 is highly expressed in ESCC and is associated with poor prognosis. Knockdown of MAGEA6 expression not only increased the chemosensitivity of ESCC, but also increased the entry of p63 into the nucleus which is the transcription factor of caspase-1. Overexpression of MAGEA6 reduced the nuclear import of p63. Based on this, we hypothesized that MAGEA6 inhibits the transcriptional activity of caspase-1 by inhibiting the nuclear import of p63, and finally decrease the pyroptosis and promotes chemoresistance in ESCC. To this end, it is proposed to explore (1) the clinical relevance of MAGEA6 and caspase-1 expression levels and chemotherapy response to ESCC. (2) In vitro experiments: The specific mechanism by which MAGEA6 inhibits p63 entry into the nucleus. (3) In vivo experiments: knocking down the expression of MAGEA6 can increase the chemosensitivity of ESCC and improve the efficacy of chemotherapy. This project will provide a theoretical basis for clinical sensitization of ESCC and improvement of patient prognosis.

化疗抵抗是食管鳞癌患者预后差的关键因素。细胞焦亡作为一种新定义的死亡方式，其减少是肿瘤发生化疗抵抗的原因之一。目前，细胞焦亡与食管鳞癌化疗抵抗的关系尚未可知。我们的前期研究证实：MAGEA6在食管鳞癌中高表达，且与不良预后相关。敲低MAGEA6的表达不仅增加食管鳞癌细胞的化疗敏感性，而且增加caspase-1的转录因子p63的入核。过表达MAGEA6则p63的入核减少。据此我们提出假设：MAGEA6通过抑制p63的入核，降低caspase-1的转录活性，抑制细胞焦亡发生，最终促进食管鳞癌细胞产生化疗抵抗。为此，拟探讨⑴MAGEA6和caspase-1表达水平与食管鳞癌化疗反应的临床相关性。⑵体外实验：MAGEA6抑制p63入核的具体机制。⑶体内实验：敲减MAGEA6的表达可以增加食管鳞癌的化疗敏感性，提升化疗疗效。这一项目的完成为临床食管鳞癌化疗增敏，改善患者预后提供理论依据。

化疗抵抗是食管鳞癌患者预后差的关键因素。临床数据表明，MAGEA6表达水平与食管癌患者化疗抵抗和预后显著正相关，通过前期对野生型及MAGEA6-/-食管癌细胞系转录组数据分析，提示细胞焦亡可能是MAGEA6调控食管癌化疗抵抗的下游通路。MAGEA6主要定位于细胞质中，因而其调控转录必然是通过转录因子介导的。通过CUT&Tag-seq和免疫共沉淀实验证实，MAGEA6可与转录因子p63互作，在化疗药物作用后，p63入核可以结合到CASP1的启动子并启动该CASP1的转录从而激活焦亡。MAGEA6蛋白包含一段长的固有无序区序列(IDRs)，并运用光漂白实验及部分体外重建实验初步证实MAGEA6与p63在细胞质中形成相分离。本课题着重探究MAGEA6与p63在细胞质中相互作用形成相分离进而调控焦亡的分子机制及生物学意义，将运用多种模型探究靶向MAGEA6抑制化疗抵抗的可行性，探索本研究内容的转化价值。