Xenografted Tumor decellularization provides cell-specific in vitro 3D environment

In vitro cell culture studies are common in cancer research field, and reliable biomimetic 3D models are needed to ensure physiological relevance. In this manuscript, we hypothesized that decellularized xenograft tumors can serve as an optimal 3D substrate to generate a top-down approach for in vitro tumor modeling. Methods: Multiple tumor cell lines were xenografted and the formed solid tumors were recovered for their decellularization by several techniques and further characterization by histology and proteomics techniques. Selected decellularized tumor xenograft samples were seeded with human triple negative breast cancer (TNBC)c ells and cell behavior was compared among them and with other control 2D and 3D cell culture methods. Results: A soft treatment using Freeze-EDTA-DNAse allows proper decellularization of xenografted tumor samples. Interestingly, proteomic data show that samples decellularized from TMBC xenograft models had different extracellular matrix (ECM) composition compared to the rest of xenograft tumors tested. The in vitro re-cellularization of decellularized ECM (dECM) yields tumor-type specific cell-behavior in TNBC context. Conclusions: Data indicate that dECM derived from xenograft tumors is a feasible substrate for re-seeding purposes, thereby promoting tumor-type specific cell behavior. These data serve as a proof-of-concept for further potential generation of patient-specific in vitro research models.

体外细胞培养研究在癌症研究领域应用极为广泛，为保障研究结果的生理学相关性，亟需构建可靠的仿生三维（3D）模型。本研究提出假说：脱细胞异种移植肿瘤可作为最优三维底物，以此构建自上而下的体外肿瘤建模体系。
方法：将多种肿瘤细胞系进行异种移植，回收所形成的实体瘤后采用多种技术开展脱细胞处理，并通过组织学与蛋白质组学技术完成进一步表征。选取部分脱细胞异种移植肿瘤样本，接种人三阴性乳腺癌（triple negative breast cancer, TNBC）细胞，将其细胞行为与其余实验组及二维（2D）、三维（3D）对照细胞培养方法进行对比分析。
结果：采用冷冻-乙二胺四乙酸（EDTA）-脱氧核糖核酸酶（DNAse）的温和处理方案，可实现异种移植肿瘤样本的充分脱细胞。值得注意的是，蛋白质组学数据显示，源自三阴性乳腺癌异种移植模型的脱细胞样本，其细胞外基质（extracellular matrix, ECM）组成与其余受试异种移植肿瘤存在显著差异。对脱细胞细胞外基质（decellularized ECM, dECM）进行体外再细胞化培养，可在三阴性乳腺癌模型中重现肿瘤类型特异性的细胞行为。
结论：研究数据表明，源自异种移植肿瘤的脱细胞细胞外基质可作为可靠的再接种底物，有效诱导肿瘤类型特异性的细胞行为。本研究结果为后续构建患者特异性体外研究模型提供了概念验证依据。