PO.ET01.02 · 实验与分子治疗
种系BRCA2突变携带者中的胰腺癌建模
Pancreatic cancer modeling in germline BRCA2 mutation carriers
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摘要 Abstract
中文摘要
背景：BRCA2的种系致病性突变（BRCA2 mut）是男性和女性胰腺癌最强的遗传风险因素；该基因的致病性变异约与5%具有该癌症类型遗传风险的男性和女性相关。此外，BRCA2 mut特别与西班牙裔/拉丁裔人群胰腺癌风险增加相关。患者特异性诱导多能干细胞（iPSC）方法为体外建模人类疾病创造了机会。源自已知遗传突变患者的iPSC携带患者独特的遗传背景，为研究特定基因的功能效应提供平台。多个由iPSC构建的遗传性疾病模型已成功再现了高危癌症。本研究旨在利用基于iPSC的建模来研究致病性BRCA2 mut对早期胰腺组织（PT）的功能影响以及促进胰腺癌（PC）进展的基因组改变。
方法：我们从BRCA2 mut阳性和BRCA野生型（BRCA WT）的女性和男性中生成iPSC。从这些BRCA2 mut和BRCA WT iPSC中，我们构建了胰腺导管和腺泡前体的类器官模型。此外，我们使用慢病毒靶向来模拟参与早期胰腺癌发病机制的特定基因的功能效应，具体包括TP53敲低（使用TP53 siRNA），以及TP53和/或KRAS过表达（分别为TP53 R175H和R273H；KRAS G12V）。
结果：分化为PT类器官后，BRCA2 mut杂合的胰腺导管细胞与BRCA WT对照相比表现出特定的细胞异常——肿瘤性转化、导管形成、癌症特异性生物标志物表达——提示BRCA2单倍体不足对所观察到的表型有所贡献。来自BRCA2 mut受试者的PC模型表现出类似早期肿瘤发育的异常。重要的是，具有BRCA2 mut、TP53和KRAS改变组合的PC类器官与单独的BRCA2 mut和BRCA WT类器官相比表现出更具侵袭性的PC表型，并与原发性PC共享基因组特征。
结论与意义：iPSC来源的类器官能够准确再现BRCA2 mut和BRCA WT胰腺前体组织。这使得癌症可以在培养皿中演化，使其成为用于机制研究和筛选方法以鉴定新药物靶点的理想模型。
查看英文原文 English abstract
Background: Germline, pathogenic mutations in BRCA2 ( BRCA2 mut ) are the strongest genetic risk factor for pancreatic cancer in men and women; pathogenic variants in this gene are implicated in about 5% of men and women with heritable risks of this cancer type. Moreover, BRCA2 mut are associated with increased risk of pancreatic cancer inHispanic/Latino populations specifically. Patient-specific induced pluripotent stem cell (iPSC) methods create opportunities to modelhuman diseases in vitro . iPSCs derived from patients with known genetic mutations carry thepatient's unique genetic background, to provide platforms for studying the functional effects ofspecific genes. Several inherited disease models created from iPSCs have successfully replicated high-risk cancers. This study aimed to utilize iPSC-based modeling to investigate thefunctional impact of pathogenic BRCA2 mut on early-stage pancreatic tissues (PT) and genomicalterations that contribute to pancreatic cancer (PC) progression.
Methods: We generated iPSC from BRCA2 mut positive and BRCA widtype ( BRCA WT ) women and men. From these BRCA2 mut and BRCA WT iPSCs we generated organoid models of pancreatic ductal and acinar precursors. In addition, we used lentiviral targeting to mimic thefunctional effects of specific genes involved in early stage pancreatic cancer pathogenesis, specifically TP53 knockdown (using a TP53 siRNA), and TP53 and/or KRAS overexpression ( TP53 R175H and R273H; KRAS G12V) respectively.
Results: Following differentiation into PT organoids heterozygous for BRCA2 mut pancreatic ductal cells show specific cellular abnormalities - neoplastic transformation, ductal formation, expression of cancer-specific biomarkers - compared to BRCA WT controls suggesting that BRCA2 haploinsufficiency contributes to the observed phenotype. PC models from BRCA2 mut subjects exhibit abnormalities reminiscent of early-stage neoplastic development. Importantly, PC organoids with combinations of BRCA2 mut , TP53 and KRAS alterations exhibited a moreaggressive PC phenotypes compared to BRCA2 mut and BRCA WT organoids alone, and sharedgenomic signatures with primary PCs.
Conclusion and Impact: iPSC-derived organoids can accurately replicate BRCA2 mut and BRCA WT pancreatic precursor tissues. This allows cancer to evolve in a dish, making it an ideal model for mechanistic studies and screening approaches to identify novel drug targets.
利益披露 Disclosure
Y. Xu, None..
N. Yucer, None..
A. Okimoto, None..
B. J. Rimel, None..
K. Lawrenson, None..
S. A. Gayther, None.