使用工程化脑微环境模型研究GBM与血管周围微环境之间的相互作用

Introduction: Glioblastoma (GBM) is the largest histological group among brain tumors, accounting for 47.8% of malignant brain tumors in the adult population; its high infiltrative nature and incidence of tumor recurrence make GBM lethal and untreatable. Vessel co-option occurs when GBM invades the vasculature; tumor cells hijack pre-existing vessels and utilize it to move inside the brain. To understand GBM mechanisms and facilitate therapeutic developments against tumor recurrence, a tumor microenvironment model for vessel co-option is required. Our lab has previously developed a gelatin hydrogel platform using a photopolymerizable methacrylamide-functionalized gelatin (GelMA). This platform was coupled with the synthetic zinc finger gene regulator (SynZiFTR) to overexpress pro- and anti- angiogenic factors to control vascular formation. We aim to use this model to study the interaction between GBM and the perivascular niche. Materials: Sequences of human PDGFR-beta and ANG-2 were obtained from the NCBI database, which were later inserted into the vector backbone using a Gibson assembly cloning kit. The gelatin hydrogel model is generated by encapsulating a mixture of brain microvascular endothelial cells, normal human astrocyte, and pericytes (3:1: 1, 3 x 10 6 BMVECs/mL); PVN cells were resuspended with GBM spheroid in the GelMA-LAP solution and then polymerized by exposure to UV light. Results: SynZiFTR containing human PDGFR-beta was transduced into human brain vascular pericytes (HBVPs). Immunostaining showed a significant increase in PDGFR-beta expression and vessel formation when encapsulated in the GelMA hydrogel with NHAs and BMVECs. Conditioned media collected from this pro-angiogenic microenvironment significantly increased tumor cell migration. We further validated the effect of this vessel network on GBM behavior by encapsulating GBM spheroid into the hydrogel model. We observed a similar increase in cellular outgrowth, and a reduced effect of TMZ on GBM spheroids. Ongoing efforts are expanding the study to consider maturation processes, where we observe overexpression of ANG-2 reduces the overall density of vessel formation, yet conditioned media from more mature vessel networks promotes GBM invasion into the surrounding hydrogel environment. Conclusion: Our data demonstrate that the cytokines and growth factors released from pro- and anti-angiogenic brain microenvironments significantly affect tumor behaviors. Ongoing efforts seek to co-encapsulate GBM spheroids in engineered vascular hydrogels (overexpressed ANG-2) to more deeply investigate reciprocal interactions between GBM and perivascular cell compartments that shape vessel co-option and GBM invasion. We believe this model would greatly facilitate the development of new therapeutics against GBM recurrence and understanding of vessel co-option.