Engineering a Vascularized Hypoxic Tumor Model for Therapeutic Assessment

Abstract

Solid tumors in advanced cancer often feature a structurally and functionally abnormal vasculature through tumor angiogenesis, which contributes to cancer progression, metastasis, and therapeutic resistances. Hypoxia is considered a major driver of angiogenesis in tumor microenvironments. However, there remains a lack of in vitro models that recapitulate both the vasculature and hypoxia in the same model with physiological resemblance to the tumor microenvironment, while allowing for high-content spatiotemporal analyses for mechanistic studies and therapeutic evaluations. We have previously constructed a hypoxia microdevice that utilizes the metabolism of cancer cells to generate an oxygen gradient in the cancer cell layer as seen in solid tumor sections. Here, we have engineered a new composite microdevice-microfluidics platform that recapitulates a vascularized hypoxic tumor. Endothelial cells were seeded in a collagen channel formed by viscous fingering, to generate a rounded vascular lumen surrounding a hypoxic tumor section composed of cancer cells embedded in a 3-D hydrogel extracellular matrix. We demonstrated that the new device can be used with microscopy-based high-content analyses to track the vascular phenotypes, morphology, and sprouting into the hypoxic tumor section over a 7-day culture, as well as the response to different cancer/stromal cells. We further evaluated the integrity/leakiness of the vascular lumen in molecular delivery, and the potential of the platform to study the movement/trafficking of therapeutic immune cells. Therefore, our new platform can be used as a model for understanding tumor angiogenesis and therapeutic delivery/efficacy in vascularized hypoxic tumors.

Recapitulating Tumor Hypoxia in a Cleanroom-Free, Liquid-Pinning-Based Microfluidic Tumor Model

“…The phenomenon is mainly caused by a combined effect of oxygen consumption (reaction) and diffusion in the tumor. While this is often simulated mathematically by analyzing a tumor section between two insulated boundaries without oxygen flux, we can experimentally recapitulate it in vitro by culturing a layer of cancer cells between two oxygen diffusion barriers (referred to as the “tumor section”), both in 2-D monolayers and 3-D ECM-embedded formats − (Figure B). Here, we designed a new generation of hypoxia models by integrating a polycarbonate (PC) film directly into a microfluidic device as the oxygen barrier (Figure C).…”

“…We next examined whether the micromilling and liquid pinning approaches can achieve the desired sizes of tumor sections − in the new microfluidic hypoxic tumor model design. We chose the computer numerical control (CNC) micromilling to fabricate the microfluidic mold for its lower cost and shorter turnaround compared to the cleanroom-based photolithographic approaches. , We milled the microfluidic mold in bulk PC and vapor-polished it for casting PDMS microfluidic devices through replica molding , (Figure A).…”

“…We can further integrate tumor vasculature in the microfluidic channels to evaluate tumorvascular crosstalk and angiogenesis. 42 We have shown our platform as a scalable tool for the preclinical evaluation of chemotherapeutics. Additionally, the platform can be used to assess the efficacy of cell-based therapeutics (e.g., conventional and chimeric antigen receptor T cells, natural killer cells).…”

“…To address these limitations, we have previously introduced micromilling-based hypoxia microdevices that naturally induce hypoxia in 2-D and 3-D tumor models, while eliminating the need for photolithography and external gas control. − These microdevices have a “cap-and-base” design composed of cancer cells cultured between two oxygen diffusion barriers, which enables high-content imaging and spatially resolved analyses of cancer cell responses to hypoxic gradients, anticancer drugs, and cell-based therapy. However, the “cap-and-base” design adds an additional handling step for the user, where the cap must be assembled to the base during cell culture and thus serves as a potential source of error in the tumor model fabrication.…”

“…However, the “cap-and-base” design adds an additional handling step for the user, where the cap must be assembled to the base during cell culture and thus serves as a potential source of error in the tumor model fabrication. Additionally, while a microfluidic channel was introduced in the 3-D microdevices, , the single-channel design limits its ability and throughput in assessing combinatory therapies or comparing multiple treatment conditions in the same model in a highly controlled manner.…”