Influence of Macrophages on Vascular Invasion of Inflammatory Breast Cancer Emboli Measured Using an In Vitro Microfluidic Multi-Cellular Platform.

Inflammatory breast cancer (IBC) is an aggressive disease with a poor prognosis and a lack of effective treatments. It is widely established that understanding the interactions between tumor-associated macrophages (TAMs) and the tumor microenvironment is essential for identifying distinct targeting markers that help with prognosis and subsequent development of effective treatments. In this study, we present a 3D in vitro microfluidic IBC platform consisting of THP1 M0, M1, or M2 macrophages, IBC cells, and endothelial cells. The platform comprises a collagen matrix that includes an endothelialized vessel, creating a physiologically relevant environment for cellular interactions. Through the utilization of this platform, it was discovered that the inclusion of tumor-associated macrophages (TAMs) led to an increase in the formation of new blood vessel sprouts and enhanced permeability of the endothelium, regardless of the macrophage phenotype. Interestingly, the platforms containing THP-1 M1 or M2 macrophages exhibited significantly greater porosity in the collagen extracellular matrix (ECM) compared to the platforms containing THP-1 M0 and the MDA-IBC3 cells alone. Cytokine analysis revealed that IL-8 and MMP9 showed selective increases when macrophages were cultured in the platforms. Notably, intravasation of tumor cells into the vessels was observed exclusively in the platform containing MDA-IBC3 and M0 macrophages.

Towards integration of time-resolved confocal microscopy of a 3D in vitro microfluidic platform with a hybrid multiscale model of tumor angiogenesis.

The goal of this study is to calibrate a multiscale model of tumor angiogenesis with time-resolved data to allow for systematic testing of mathematical predictions of vascular sprouting. The multi-scale model consists of an agent-based description of tumor and endothelial cell dynamics coupled to a continuum model of vascular endothelial growth factor concentration. First, we calibrate ordinary differential equation models to time-resolved protein concentration data to estimate the rates of secretion and consumption of vascular endothelial growth factor by endothelial and tumor cells, respectively. These parameters are then input into the multiscale tumor angiogenesis model, and the remaining model parameters are then calibrated to time resolved confocal microscopy images obtained within a 3D vascularized microfluidic platform. The microfluidic platform mimics a functional blood vessel with a surrounding collagen matrix seeded with inflammatory breast cancer cells, which induce tumor angiogenesis. Once the multi-scale model is fully parameterized, we forecast the spatiotemporal distribution of vascular sprouts at future time points and directly compare the predictions to experimentally measured data. We assess the ability of our model to globally recapitulate angiogenic vasculature density, resulting in an average relative calibration error of 17.7% ± 6.3% and an average prediction error of 20.2% ± 4% and 21.7% ± 3.6% using one and four calibrated parameters, respectively. We then assess the model's ability to predict local vessel morphology (individualized vessel structure as opposed to global vascular density), initialized with the first time point and calibrated with two intermediate time points. In this study, we have rigorously calibrated a mechanism-based, multiscale, mathematical model of angiogenic sprouting to multimodal experimental data to make specific, testable predictions.

Collagen/kerateine multi-protein hydrogels as a thermally stable extracellular matrix for 3D in vitro models.

Objective: To determine whether the addition of kerateine (reduced keratin) in rat tail collagen type I hydrogels increases thermal stability and changes material properties and supports cell growth for use in cellular hyperthermia studies for tumor treatment.Methods: Collagen type I extracted from rat tail tendon was combined with kerateine extracted from human hair fibers. Thermal, mechanical, and biocompatibility properties and cell behavior was assessed and compared to 100% collagen type I hydrogels to demonstrate their utility as a tissue model for 3D in vitro testing.Results: A combination (i.e., containing both collagen 'C/KNT') hydrogel was more thermally stable than pure collagen hydrogels and resisted thermal degradation when incubated at a hyperthermic temperature of 47°C for heating durations up to 60 min with a higher melting temperature measured by DSC. An increase in the storage modulus was only observed with an increased collagen concentration rather than an increased KTN concentration; however, a change in ECM structure was observed with greater fiber alignment and width with an increase in KTN concentration. The C/KTN hydrogels, specifically 50/50 C/KTN hydrogels, also supported the growth and of fibroblasts and MDA-MB-231 breast cancer cells similar to those seeded in 100% collagen hydrogels.Conclusion: This multi-protein C/KTN hydrogel shows promise for future studies involving thermal stress studies without compromising the 3D ECM environment or cell growth.

Integrating Quantitative Assays with Biologically Based Mathematical Modeling for Predictive Oncology.

We provide an overview on the use of biological assays to calibrate and initialize mechanism-based models of cancer phenomena. Although artificial intelligence methods currently dominate the landscape in computational oncology, mathematical models that seek to explicitly incorporate biological mechanisms into their formalism are of increasing interest. These models can guide experimental design and provide insights into the underlying mechanisms of cancer progression. Historically, these models have included a myriad of parameters that have been difficult to quantify in biologically relevant systems, limiting their practical insights. Recently, however, there has been much interest calibrating biologically based models with the quantitative measurements available from (for example) RNA sequencing, time-resolved microscopy, and in vivo imaging. In this contribution, we summarize how a variety of experimental methods quantify tumor characteristics from the molecular to tissue scales and describe how such data can be directly integrated with mechanism-based models to improve predictions of tumor growth and treatment response.

In vitro vascularized tumor platform for modeling tumor-vasculature interactions of inflammatory breast cancer.

Inflammatory breast cancer (IBC), a rare form of breast cancer associated with increased angiogenesis and metastasis, is largely driven by tumor-stromal interactions with the vasculature and the extracellular matrix (ECM). However, there is currently a lack of understanding of the role these interactions play in initiation and progression of the disease. In this study, we developed the first three-dimensional, in vitro, vascularized, microfluidic IBC platform to quantify the spatial and temporal dynamics of tumor-vasculature and tumor-ECM interactions specific to IBC. Platforms consisting of collagen type 1 ECM with an endothelialized blood vessel were cultured with IBC cells, MDA-IBC3 (HER2+) or SUM149 (triple negative), and for comparison to non-IBC cells, MDA-MB-231 (triple negative). Acellular collagen platforms with endothelialized blood vessels served as controls. SUM149 and MDA-MB-231 platforms exhibited a significantly (p < .05) higher vessel permeability and decreased endothelial coverage of the vessel lumen compared to the control. Both IBC platforms, MDA-IBC3 and SUM149, expressed higher levels of vascular endothelial growth factor (p < .05) and increased collagen ECM porosity compared to non-IBCMDA-MB-231 (p < .05) and control (p < .01) platforms. Additionally, unique to the MDA-IBC3 platform, we observed progressive sprouting of the endothelium over time resulting in viable vessels with lumen. The newly sprouted vessels encircled clusters of MDA-IBC3 cells replicating a key feature of in vivo IBC. The IBC in vitro vascularized platforms introduced in this study model well-described in vivo and clinical IBC phenotypes and provide an adaptable, high throughput tool for systematically and quantitatively investigating tumor-stromal mechanisms and dynamics of tumor progression.

Vascularized microfluidic platforms to mimic the tumor microenvironment.

Microfluidic technology has led to the development of advanced in vitro tumor platforms that overcome the challenges of in vivo animal and in vitro two dimensional models. This paper presents platform designs and methods used to develop complex vascularized in vitro models to mimic the tumor microenvironment. Features of these platforms include a continuous, aligned endothelium that allows for cell-cell interactions between vasculature and tumor cells. A novel platform for fabrication of a single endothelialized microchannel encased within a collagen platform hosting breast cancer cells was developed and utilized to study the influence of cellular interaction on transport phenomenon through vasculature in a hyperpermeable tumor microenvironment. This platform relies on subtractive tissue engineering fabrication techniques. Through confocal imaging we have demonstrated that the platform produces enhanced vessel leakiness recapitulating physiological features of the tumor microenvironment. The influence of tumor endothelial interactions on transport of particles was also demonstrated. Additionally, we designed two more complex and intricate endothelialized microfluidic networks by combining lithographic techniques with additive tissue engineering methods. We created a network platform consisting of interconnected microchannels to model a highly vascularized system and successfully perfused the system with fluorescent particles. Finally, we developed a physiologically representative in vitro microfluidic platform with vasculature patterned from in vivo data showing the versatility of these systems to replicate the complex geometries of tumor microvasculature and dynamically measured particle transport. Overall, we have shown the ability to develop functional microfluidic vascular tumor platforms of varying complexities and demonstrated their utility for studying spatial particle transport within these systems.