成体腺泡细胞起源的胰腺癌动物模型的建立及分析

Pancreatic cancer is the fourth leading cause of cancer related deaths, with a five-year survival rate of less than 7%, so establishment of better animal models are need urgently to improve pancreatic cancer research. Embryo models are mostly used as existing pancreatic cancer animal models in which alterations of oncogenic genes are induced at embryonic stage and which are inconsistent with the process of human pancreatic cancer that is caused by accumulation of genetic variation in somatic cell during adulthood and these genetic alterations in embryo model could not induce pancreatic cancer in adult animals. Both ductal cells and acinar cells were supposed to be the cell-of-origin of pancreatic cancer. In previous study, we found the driver mutations of pancreatic ductal cells transformation in adult mice and established adult mice model of pancreatic cancer, while recent studies have suggested that acinar cells are the most possible cell-of-origin. In order to explore the driver mutations essential for pancreatic acinar cells transformation and to establish adult pancreatic cancer animal model derived from the transformation. In this proposed study, we will select representative genetic variant combinations in human pancreatic cancer according to the genomic analyses of human pancreatic cancer in TCGA database and the previous study in mouse model. And change the expression of these genes combinations in adult tree shrew pancreatic acinar cells by lentivirus infection to induce malignant transformation of these acinar cells and finally induced adult tree shrew pancreatic cancer. The expected results would be possible to illuminate the driver mutations which are essential for the transformation of adult pancreatic acinar cells and will provide a better platform for the mechanism research and medicinal development of human pancreatic cancer.

胰腺癌在肿瘤死亡原因中排第四位，5年生存率仅7%左右，迫切需要建立更好的动物模型来改善胰腺癌相关研究。现有胰腺癌动物模型大多是从胚胎期就引入基因改变，这与人胰腺癌是成年过程中体细胞遗传变异累积造成的情况不符，且这些遗传变异不能诱导成体动物形成胰腺癌。腺泡和导管细胞被认为是胰腺癌的主要起源，前期我们通过改变成体小鼠导管细胞基因表达，成功建立导管起源的胰腺癌模型，但近年研究表明腺泡细胞更有可能是胰腺癌的起始细胞。为了探究成体腺泡细胞转化所需的遗传变异，并建立其起源的成体动物模型，本项目将根据TCGA数据库对人类胰腺癌基因组的分析及前期小鼠模型的研究，选择人胰腺癌中有代表性的遗传变异组合，利用慢病毒感染技术改变成体树鼩腺泡细胞基因的表达，诱导其发生恶性转化并形成胰腺癌。预期成果将阐明成体腺泡细胞转化所需的遗传变异，并为人类胰腺癌机理研究及药物开发等提供更好的动物模型。

动物模型是探索人类疾病的病因、机制及治疗手段等的重要工具，现有胰腺癌动物模型大多是从胚胎时期就引入基因的改变，研究表明这些基因突变却不足以诱导成体动物形成胰腺癌，而且与人胰腺癌是成年过程中体细胞遗传变异累积造成的情况不符，迫切需要建立更好的动物模型来改善胰腺癌相关研究。腺泡和导管细胞被认为是胰腺癌的主要起源，前期我们通过改变成体小鼠导管细胞基因表达，成功建立导管起源的胰腺癌模型，但近年研究表明腺泡细胞更有可能是胰腺癌的起始细胞。本项目拟探究成体腺泡细胞转化所需的遗传变异，并建立其起源的成体动物模型，. 在本项目中，我们发现胰腺原位注射慢病毒，可以特异性的感染树鼩胰腺腺泡细胞，这为我们探索腺泡细胞转化条件奠定基础。根据前期的研究及TCGA数据库中人类胰腺癌样本中的基因变异情况，选定构建模型所需的基因为KRASG12D、TP53、CDKN2A、CDKN2B，并构建不同的慢病毒：KRASG12D-shTp53、KRASG12D-shTp53-shCdkn2a、KRASG12D-shTp53-shCdkn2b和KRASG12D-shTp53-shCdkn2a/b, 在树鼩胰腺头部原位注射15ul 1x10^7病毒颗粒，只有注射KRAS-shTp53-shCdkn2a/b病毒的树鼩在注射后3-7周陆续发病死亡，并都形成胰腺肿瘤。说明成体腺泡细胞恶性转化需要的遗传改变为过表达KRASG12D，敲降抑癌基因TP53、CDKN2A、CDKN2B。. 本项目中我们利用原位注射慢病毒，改变局部少量细胞基因表达，成功构建了树鼩胰腺癌模型，病理和分子特征分析表明树鼩胰腺癌与人胰腺导管腺癌特征一致。同时通过比较人、小鼠、树鼩胰腺癌基因表达谱，我们发现树鼩胰腺癌基因表达谱与人更相似，同时通过比较胰腺癌通路相关基因，发现树鼩与人同源性更高，修饰位点更相似。说明本项目构建的树鼩胰腺癌模型更能模拟人胰腺癌，将对人类胰腺癌的机理研究及药物开发等提供更好的工具。. 我们的研究成功阐明了成体腺泡细胞恶性转化所需遗传改变，并成功构建了腺泡细胞起源的树鼩胰腺癌动物模型，将为胰腺癌研究提供更好地平台。