涎腺腺样囊性癌中MYB-NFIB融合基因及亚型与肿瘤细胞特性、侵袭转移的相关性研究

Cribriform architecture, consisting of myoepithelial cell, is the characteristic structure of salivary adenoid cystic carcinoma (SACC). In other salivary benign tumors, they also have cribriform architecture but the biological behaviour is different with SACC. The genetic change may be the main reason for this difference. Translocation of MYB and NFIB gene, with several subtype, is the sole specific molecular feature of tumor tissue of SACC. MYB-NFIB fusion gene can result in high expression of MYB protein. One of the function of MYB protein is activate epithelial-to-mesenchymal transition in human cancer cells of different origin by transactivationg Slug.Previously, we demonstrated that reduced E-cadherin expression in myoepithelial cell in SACC, indicating mesenchymal differentiation of myoepithelial cell, whereas, there was high MYB expression in myoepithelial cell. We therefore raised the question weather specific translocation of MYB and NFIB was responsible for mesenchymal differentiation of myoepithelial cell in SACC and promoted invasion and transvasular metastasis of tumor cell of SACC, furthermore,whether subtype of MYB-NFIB fusion gene was correlated with different biological behaviour of SACC. In this study, using fresh neoplastic tissues and paraffin embedded pathology tissues, we first detected translocation of MYB and NFIB by RT-PCR and sequencing and FISH (fluorescence in situ hybridization) in myoepithelial and glandular cell, also the expression of MYB,Slug and E-cadherin. Then the correlation of translocation of MYB and NFIB, expression of MYB,Slug, E-cadherin, clinicopathological features of tumors and prognosis of patients will be discussed. To further identified the role of MYB-NFIB fusion gene, using with and without translocation of MYB and NFIB cell lines of SACC, we will transduct/inhibit the fusion gene of MYB-NFIB by lentiviral vector, and examine the proliferation, invasion, capability of tumor formation and metastasis in vitro and in vivo,expression of MYB,Slug and E-cadherin before and after transduction/inhibition. By this study, we will reveal the assocation between specific MYB-NFIB fusion gene/subtype and feature of SACC tumor cell,prognosis of patient, also the mechanism of specific MYB-NFIB fusion gene in tumor invasion and metastasis.This study will provide experiment evidence for MYB-NFIB fusion gne as specific molecular diagnosis and therapeutic targets of SACC.

由肿瘤性肌上皮细胞形成的筛状结构是涎腺腺样囊性癌（SACC）特征性组织学形态，但筛状结构也见于某些涎腺良性肿瘤，并呈现不同的生物学行为，其原因可能是肿瘤间"貌似"的肌上皮细胞存在遗传性差异。MYB和NFIB基因融合是近年来发现SACC唯一具特异性的遗传改变，未见于其它涎腺肿瘤。我们前期研究发现MYB-NFIB融合基因存在融合位点差异，且肌上皮细胞MYB蛋白高表达、与SACC侵袭相关蛋白E-cad低表达，而腺上皮细胞表达状况与此相反，上述基因及蛋白表达差异是否是肌上皮细胞"貌似神异"的原因呢？与SACC侵袭转移相关性仍未可知。本研究拟针对SACC临床样本和细胞系，采用FISH、目的基因干扰/载入等方法，多方位阐述MYB和NFIB基因融合及位点、MYB蛋白表达、肌上皮细胞特性相关蛋白Slug、E-cad与肌上皮细胞特性、肿瘤侵袭转移、预后的关系，为MYB融合基因作为分子诊断及化疗靶点提供依据。

SACC(salivary adenoid cystic carcinoma)是唾液腺恶性肿瘤中的常见肿瘤，具有生长缓慢、侵袭性强等特点，分子水平改变MYB和NFIB基因融合在SACC发生发展过程中的作用研究有助于我们进一步深入认识该肿瘤。98例SACC样本利用RT-PCR及测序检测发现，71.4%（70/98）的病例中存在MYB-NFIB的基因融合；利用FISH检测发现，82.6% (81/98) 的病例中存在MYB基因的异常，包括分离、缺失、扩增；IHC方法检测98例样本中MYB蛋白水平的表达，发现77.6% (76/98) 的病例中存在MYB蛋白的高表达。提示在SACC中，MYB-NFIB基因融合是最主要的MYB基因异常方式，除此之外尚存在其他基因异常，这些异常可以导致MYB蛋白的高表达。统计发现，存在MYB-NFIB基因融合的患者预后要略差于不存在MYB-NFIB基因融合者。对4例特殊部位—颌骨中央性的ACC与唾液腺ACC进行了对比分析，发现下颌骨ACC和唾液腺ACC具有相似的临床病理特征、免疫表型和分子改变，提示他们具有相同的来源，MYB蛋白及基因异常可作为与其他颌骨肿瘤鉴别诊断的有用指标。收集了22例与SACC形态极为相似的基底细胞肿瘤，包括18例良性及4例恶性，同时选用20例形态相近的SACC作为对照。用FISH及IHC方法检测其MYB表达及基因异常情况，结果表明，不论是良性还是恶性基底细胞肿瘤中均没有MYB的蛋白表达及基因异常，而对照组中均有较大比例的MYB蛋白表达(11/22)及基因断裂(9/11)。总之，本课题的研究表明，MYB基因的异常为SACC中的常见事件，发生MYB和NFIB基因融合的患者预后稍差，MYB基因异常可以作为诊断SACC的分子指标，颌骨中央型ACC具有与SACC相同的分子学改变。