Tissue-Engineered Prostate Cancer Xenografts: A Biomaterials-Based Approach to Study Tumorigenesis and Dormancy Escape
Long, Thomas Joseph
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Despite massive investments in research and development, it is estimated that 95% of oncology compounds that enter clinical trials ultimately fail to receive FDA approval . This disconnect between pre-clinical testing and clinical success points to a need to develop improved pre-clinical model systems for cancer studies that more accurately reflect human disease states. Toward this goal, biomaterial scaffolds have shown promise as the basis for in vitro and in vivo 3D cancer models. Tumors engineered using biomaterials have shown evidence of being more physiologically relevant than some traditional preclinical model systems, and synthetic biomaterials provide the added potential for enhanced microenvironmental control. In this dissertation, we examine sphere-templated poly(2-hydroxyethyl methacrylate) (pHEMA) scaffolds as the basis for engineering in vivo xenografts from human prostate cancer cell lines. Methods were developed to seed, culture, and measure the proliferation of prostate cancer cells in vitro within these porous hydrogels. A novel capillary force-based seeding method is described that improved cell number and distribution within the scaffolds compared to well-established protocols such as static and centrifugation seeding. Dynamic cell culture improved oxygen diffusion in vitro, and a PicoGreen-based DNA assay was used to evaluate cell proliferation. pHEMA scaffolds seeded and pre-cultured with tumorigenic M12 prostate cancer epithelial cells prior to implantation generated tumors in athymic nude mice, demonstrating the ability of the scaffolds to be used as a synthetic vehicle for xenograft generation. The resulting tumors showed no significant differences in tumor growth kinetics or vascularity compared to standard xenografts derived from Matrigel, which is consistent with observations that highly tumorigenic cells are not affected in vivo by 3D culture within biomaterial scaffolds. Because Matrigel-based xenografts expose cells to exogenous growth factors and ECM proteins, it would be of interest to the cancer research field to develop a controllable, synthetic system as a replacement. We attempted to do this using pHEMA scaffolds seeded with LNCaP C4-2 metastatic prostate cancer cells. LNCaP C4-2 cells ordinarily require Matrigel or stromal cell support to form tumors in vivo, but when implanted within pHEMA, the constructs were poorly tumorigenic. Scaffold surface modification with collagen I did not improve tumorigenicity, but the synthetic nature of the scaffold lends itself to further surface modifications and controlled growth factor release in future studies that may allow tumor development within a controllable microenvironment. Finally, M12mac25 cells, an epithelial prostate cancer cell line that is ordinarily rendered non-tumorigenic through the expression of the tumor suppressor insulin-like growth factor binding protein 7 (IGFBP7), displayed a tumorigenic response when implanted within porous pHEMA scaffolds. These findings show the potential for this biomaterials-based model system to be used in the study of in vivo prostate cancer dormancy and dormancy escape. The M12mac25 tumors showed no significant difference in vascularity compared to their dormant Matrigel counterparts, but did demonstrate a significantly higher macrophage infiltration within the scaffolds mediated by the foreign body response to the materials. Cytokine arrays, DNA oligonucleotide arrays, in vitro macrophage-conditioned media studies, and in vivo studies using clondronate liposomes to eliminate macrophages showed evidence that macrophages could be the key cellular player mediating this dormancy escape.
- Bioengineering