TG2抑制剂KCC009增强喉癌细胞放射敏感性的作用机制研究

TGase 2 is a calcium-dependent enzyme that catalyzes the formation of covalent bonds between free amine groups in one protein and protein-bound glutamines of another, creating highly cross-linked protein complexes. TG2 is ubiquitous and most diverse member of the transglutaminase family of enzymes. TG2 are implicated in cell differentiation, receptor-mediated endocytosis, cell adhesion, and induction of apoptosis. TGase 2 has also been associated with various types of cancer. In our previous study, immunostaining with antibodies specific to TG2 was performed in formalin-fixed, paraffinembedded specimens from 148 laryngeal SCC patients. we found that TG2 was highly positive in 47.3% (70/148) of the tumors and Patients with low TG2 expression had a significantly better overall survival compared to patients with high TG2 expression (P < 0.001). TG2 may therefore offer a novel therapeutic target to intervene in laryngeal cancer. As a result, the present study aims to observe and explore the radiosensitization effects of KCC009, the recently developed irreversible active-site inhibitor of TG2, on laryngeal cancer and its underlying mechanism. The Hep-2 laryngeal cancer cells were cultured to evaluate the antitumor activity of KCC009 combined with radiation (KCC009 + radiation) treatment compared with monotherapy (KCC009 or radiation). The rate of apoptotic cancer cells and cancer cells' Colony forming ability were measured to evaluate the antitumor effect. The result of our present study will verify that TG2 can offer a novel therapeutic target for laryngeal cancer and the expression level of TG2 is novel predictive factor of radiosensibility of laryngeal cancer.

放射治疗诱导肿瘤细胞凋亡是放疗引起肿瘤细胞死亡的重要机制。寻求各种手段来促进细胞凋亡以提高肿瘤治疗的效率是目前放射增敏研究的热点内容。既往研究已经证实组织型谷氨酰胺转氨酶（TG2）基因的过量表达与多种肿瘤细胞系的凋亡抵抗及化疗药物耐药相关，但与喉癌放疗敏感性的研究还未见报道。我们前期研究发现TG2在喉癌中高表达率达42.3%，接受术后放疗喉鳞癌患者中，高表达TG2者预后差。TG2通过何种机制影响喉癌放射治疗的疗效以及TG2能否成为放射治疗增敏的靶点成为我们进一步研究的方向。本课题通过TG2抑制剂KCC009与放射治疗联合作用于喉癌Hep-2细胞，研究其对肿瘤细胞克隆形成能力和凋亡的影响，通过体外实验探讨KCC009对放疗的增敏作用。本研究将明确TG2基因能否作为一种新型的喉癌放射增敏靶基因以及KCC009对喉癌的具体放射增敏疗效。

我们前期研究发现TG2在喉癌中高表达率达42.3%，接受术后放疗喉鳞癌患者中，高表达TG2者预后差。TG2通过何种机制影响喉癌放射治疗的疗效以及TG2能否成为放射治疗增敏的靶点成为我们进一步研究的方向。本课题通过TG2抑制剂KCC009与放射治疗联合作用于喉癌Hep-2细胞，研究其对肿瘤细胞的存活分数、克隆存活率、细胞周期、细胞周期调控蛋白的影响。结果表明：（1）KCC009对肿瘤细胞的抑制率呈浓度和时间依赖性增加；（2）KCC009能明显增加喉癌Hep-2细胞的辐射敏感性；（3）KCC009主要通过诱导喉癌Hep-2细胞凋亡，发挥辐射增敏作用；（4）KCC009对喉癌Hep-2细胞周期的影响主要是诱导肿瘤细胞产生S期阻滞，而对G2/M期没有阻滞效应；（5）KCC009对喉癌Hep-2细胞的放疗增敏作用呈浓度依赖性增加。