SMO基因突变与牙源性角化囊肿发病关系及分子靶向干预研究

牙源性角化囊肿(OKC)是常见的颌骨囊性病损。既往研究表明PTCH1基因突变与OKC发生密切相关。本课题组前期研究发现，除了PTCH1突变，OKC亦存在SMO基因突变。已检测到的3例SMO突变OKC中2例不含PTCH1突变，polyphen2软件预测显示：该2例SMO突变很可能导致其蛋白功能受到损害，提示SMO基因突变亦可能与OKC发病有关，据此提出“SMO基因突变与OKC发病关系及分子靶向干预研究”的科学问题。围绕此科学问题，本研究拟构建突变型SMO真核表达质粒，转染HEK293细胞，探讨SMO突变对SHH通路状态的影响；同时体外培养OKC病变成纤维细胞，筛查SMO突变，探讨SMO突变对OKC成纤维细胞SHH通路活化状态、增殖能力、侵袭能力的影响，探索应用靶向SHH通路不同水平的抑制剂治疗SMO突变OKC是否可行、有效，以进一步完善对OKC发病机制的认识，为OKC临床治疗提供新思路。

Odontogenic Keratocysts (OKCs) are common cystic lesions that can occur sporadically or in association with nevoid basal cell carcinoma syndrome (NBCCS). OKCs are locally aggressive, can cause marked jaw bone destruction, and often recur after surgery. PTCH1 mutations are frequently detected in both NBCCS-related and sporadic OKCs, suggesting that PTCH1 inactivation might constitutively activate sonic hedgehog (SHH) signaling and play a major role in disease pathogenesis. In addition to loss-of-function mutations in PTCH1, gain-of-function mutations in SMO have also been found in NBCCS as well as in its related sporadic tumors. However, few reports have analyzed SMO mutations in OKCs. In our previous study, we identified 3 SMO mutations in OKCs, among which, 2 SMO mutations were not paired with PTCH1 mutations. According to the Polyphen 2 software, the 2 mutations result in a damaging effect on their SMO proteins, suggesting SMO mutations can drive initial OKC tumorigenesis. Therefore, based on the newly detected SMO mutations, we will construct three mutant SMO eukaryotic expression vectors by site-directed mutagenesis and transfect them into HEK293 cells，and the status of SHH signaling will be investigated in vitro. Small molecule SHH inhibitors will be tested in these SMO-mutant transfected HEK293 cells. Furthermore，small molecule SHH inhibitors will then be tested for their effects on the cell proliferation as well as invasion in the SMO-mutant OKC fibroblasts. Thus this study will provide further insight into the molecular mechanism of OKC and new strategies for OKC treatment of molecular approaches.