Multiscale hydrogel biomaterials-enabled 3D modeling of cancer drug resistance

项目总结/摘要 开发新的抗癌药物一直非常缓慢和昂贵。一个主要原因是， 今天用于药物发现的2D癌细胞和动物模型与人类的3D肿瘤非常不同 患者最近，已经通过将肿瘤细胞悬浮在培养基中以形成多细胞的3D肿瘤模型 在一些实施方案中，细胞可以在类球体/类器官中生长，在水凝胶/支架中生长细胞，以及并入血管。不过小 已经完成了建立3D血管化肿瘤模型，用于重现患者肿瘤中的耐药性。 我们最近开发了一种新的多尺度水凝胶生物材料为基础的自下而上的战略，以控制 肿瘤中两个不同组织域的形成--3D血管网络和无血管/血管间 含肿瘤细胞区域，用于研究癌症耐药性。这是通过产生3D无血管 首先是微肿瘤（µtumors），并将其用作与内皮细胞（EC）组装的构建块 在动态灌注培养下，形成具有复杂3D血管网络的3D血管化肿瘤， 模拟体内肿瘤的血管间组织。重要的是，我们的数据定量显示， 这是第一次，3D血管化肿瘤比3D无血管肿瘤更具抗药性， 表明肿瘤血管不仅运输营养/氧气，而且增强癌症耐药性。 然而，没有3D血管化肿瘤是用从癌症干细胞（CSC）分化的细胞在体外建立的。 尽管罕见的CSC被认为具有肿瘤发生的独特能力， 并且由于它们的高耐药性而导致癌症化疗的许多失败。此外，委员会认为， 用现代方法分离的CSC没有显示出分化成不参与细胞分化的细胞类型。 肿瘤的这种跨组织多谱系分化是干细胞的关键特性（例如，脂肪干 细胞可以分化成脂肪中不存在的成骨细胞）。因此，需要一种用于分离真CSC的方法。 我们最近开发了一种新的基于核-壳水凝胶生物材料的方法，用于分离CSCs， 在微胶囊的纳升水凝胶核中培养一个（1）单个癌细胞（来自细胞系）， 孵化前胚胎，其中全能-多能干细胞在人体内培养。重要的是 分离的CSC能够内皮、心脏、神经和成骨分化，并且高度致瘤， 转移性和耐药性，表明用我们的生物启发的1-细胞培养物分离的细胞是真正的CSC。 在这个项目中，我们建议进一步开发新的生物启发的方法，从乳腺中分离CSCs， 人类患者的肿瘤。鉴于用我们的生物启发分离的CSC的高度耐药性质， 1-细胞培养及其在体内分化为EC的能力可能会降低患者的存活率，我们进一步 我建议使用CSC衍生的EC和癌细胞来构建3D血管化肿瘤，以更好地模拟肿瘤细胞。 患者肿瘤中的耐药性优于使用非CSC癌细胞制备的现有3D血管化肿瘤， 非CSC（并且通常是非癌症相关的）EC（例如，人脐静脉内皮细胞，称为HUVEC）。

Project Summary/Abstract Developing new anticancer drugs has been very slow and costly. A major reason is that the commonly used 2D cancer cells and animal models for drug discovery today are very different from the 3D tumors in human patients. Lately, 3D tumor models have been made by suspending tumor cells in medium to form multicellular spheroids/organoids, growing the cells in hydrogels/scaffolds, and incorporating blood vessels. However, little has been done to build 3D vascularized tumor models for recapitulating the drug resistance in patient tumors. We recently developed a novel multiscale hydrogel biomaterials-based bottom-up strategy to control the formation of the two distinct tissue domains in tumors - a 3D vascular network and an avascular/intervascular tumor cell-containing region, for studying cancer drug resistance. This is achieved by producing 3D avascular micro-tumors (µtumors) first and using them as the building blocks for assembling with endothelial cells (ECs) under dynamic perfusion culture, to form 3D vascularized tumors with a complex 3D vascular network that mimics the vascular-intervascular organization of in vivo tumors. Importantly, our data show quantitatively for the first time that, the 3D vascularized tumors are much more drug resistant than 3D avascular µtumors, indicating tumor blood vessels not only transport nutrients/oxygen but also enhance cancer drug resistance. However, no 3D vascularized tumor was built in vitro with cells differentiated from cancer stem cells (CSCs) for drug discovery, although the rare CSCs have been posited to possess the exclusive ability of tumorigenesis and be responsible for the many failures of cancer chemotherapy due to their high drug resistance. Moreover, no CSCs isolated with contemporary methods are shown to differentiate into a type of cells that are not in tumors. This cross-tissue multilineage differentiation is a key property of stem cells (e.g., adipose-derived stem cells can differentiate into osteoblasts that are absent in fat). Thus, a method for isolating true CSCs is in need. We recently developed a novel core-shell hydrogel biomaterials-based approach for isolating CSCs by culturing one (1) single cancer cell (from a cell line) in the nanoliter hydrogel core of microcapsules that mimic pre-hatching embryos, where totipotent-pluripotent stem cells are cultured in human body. Importantly, the isolated CSCs are capable of endothelial, cardiac, neural, and osteogenic differentiations and highly tumorigenic, metastatic, and drug resistant, indicating the cells isolated with our bioinspired 1-cell culture are truly CSCs. In this project, we propose to further develop the novel bioinspired approach for isolating CSCs from breast tumors of human patients. In view of the highly drug resistant nature of the CSCs isolated with our bioinspired 1-cell culture and their ability of differentiating into ECs in vivo that may reduce patient survival, we further propose to use the CSC-derived ECs and cancer cells for building 3D vascularized tumors, to better model the drug resistance in patient tumors than existing 3D vascularized tumors made using non-CSC cancer cells and non-CSC (and often non-cancer-related) ECs (e.g., human umbilical vein endothelial cells known as HUVECs).