Metastasis is a dynamic process in which cancer cells navigate the tumor microenvironment, largely guided by external chemical and mechanical cues. Our current understanding of metastatic cell migration has relied primarily on studies of single cell migration, most of which have been performed using two-dimensional (2D) cell culture techniques and, more recently, using three-dimensional (3D) scaffolds. However, the current paradigm focused on single cell movements is shifting toward the idea that collective migration is likely one of the primary modes of migration during metastasis of many solid tumors. Not surprisingly, the mechanics of collective migration differ significantly from single cell movements. As such, techniques must be developed that enable in-depth analysis of collective migration, and those for examining single cell migration should be adopted and modified to study collective migration to allow for accurate comparison of the two. In this review, we will describe engineering approaches for studying metastatic migration, both single cell and collective, and how these approaches have yielded significant insight into the mechanics governing each process.

Aligned collagen architecture is a characteristic feature of the tumor extracellular matrix (ECM) and has been shown to facilitate cancer metastasis using 3D in vitro models. Additional features of the ECM, such as pore size and stiffness, have also been shown to influence cellular behavior and are implicated in cancer progression. While there are several methods to produce aligned matrices to study the effect on cell behavior in vitro, it is unclear how the alignment itself may alter these other important features of the matrix. In this study, we have generated aligned collagen matrices and characterized their pore sizes and mechanical properties at the micro- and macro-scale. Our results indicate that collagen alignment can alter pore-size of matrices depending on the polymerization temperature of the collagen. Furthermore, alignment does not affect the macro-scale stiffness but alters the micro-scale stiffness in a temperature independent manner. Overall, these results describe the manifestation of confounding variables that arise due to alignment and the importance of fully characterizing biomaterials at both micro- and macro-scales.

Hepatocyte growth factor (HGF) has been shown to have anti-fibrotic, pro-angiogenic, and cardioprotective effects; however, it is highly unstable and expensive to manufacture, hindering its clinical translation. Recently, a HGF fragment (HGF-f), an alternative c-MET agonist, was engineered to possess increased stability and recombinant expression yields. In this study, we assessed the potential of HGF-f, delivered in an extracellular matrix (ECM)-derived hydrogel, as a potential treatment for myocardial infarction (MI). HGF-f protected cardiomyocytes from serum-starvation and induced down-regulation of fibrotic markers in whole cardiac cell isolate compared to the untreated control. The ECM hydrogel prolonged release of HGF-f compared to collagen gels, and in vivo delivery of HGF-f from ECM hydrogels mitigated negative remodeling, improved fractional area change (FAC), and increased arteriole density in rat myocardial infarction model. These results indicate that HGF-f may be a viable alternative to using recombinant HGF, and that an ECM hydrogel can be employed to increase growth factor retention and efficacy.

Injectable biomaterials have been evaluated as potential new therapies for myocardial infarction (MI) and heart failure. These materials have improved left ventricular (LV) geometry and ejection fraction, yet there remain concerns that biomaterial injection may create a substrate for arrhythmia. Since studies of this risk are lacking, we utilized optical mapping to assess the effects of biomaterial injection and interstitial spread on cardiac electrophysiology. Healthy and infarcted rat hearts were injected with a model poly(ethylene glycol) hydrogel with varying degrees of interstitial spread. Activation maps demonstrated delayed propagation of action potentials across the LV epicardium in the hydrogel-injected group when compared to saline and no-injection groups. However, the degree of the electrophysiological changes depended on the spread characteristics of the hydrogel, such that hearts injected with highly spread hydrogels showed no conduction abnormalities. Conversely, the results of this study indicate that injection of a hydrogel exhibiting minimal interstitial spread may create a substrate for arrhythmia shortly after injection by causing LV activation delays and reducing gap junction density at the site of injection. Thus, this work establishes site of delivery and interstitial spread characteristics as important factors in the future design and use of biomaterial therapies for MI treatment.

The interactions of cells and surrounding tissues with biomaterials used in tissue engineering, wound healing, and artificial organs ultimately determine their fate in vivo. We have demonstrated the ability to tune fibroblast responses with the use of varied material chemistries. In particular, we examined cell morphology, cytokine production, and collagen fiber deposition angles in response to a library of arginine-based polymeric materials. The data presented here shows a large range of vascular endothelial growth factor (VEGF) secretion (0.637 ng/10(6) cells/day to 3.25 ng/10(6) cells/day), cell migration (∼15 min < persistence time < 120 min, 0.11 μm/min < speed < 0.23 μm/min), and cell morphology (0.039 < form factor (FF) < 0.107). Collagen orientation, quantified by shape descriptor (D) values that ranges from 0 to 1, representing completely random (D = 0) to aligned (D = 1) fibers, exhibited large variation both in vitro and in vivo (0.167 < D < 0.36 and 0.17 < D < 0.52, respectively). These findings demonstrate the ability to exert a certain level of control over cellular responses with biomaterials and the potential to attain a desired cellular response such as, increased VEGF production or isotropic collagen deposition upon exposure to these materials in wound healing and tissue engineering applications.

Injectable biomaterials are promising as new therapies to treat myocardial infarction (MI). One useful property of biomaterials is the ability to protect and sustain release of therapeutic payloads. In order to create a platform for optimizing the release rate of cardioprotective molecules we utilized the tunable degradation of acetalated dextran (AcDex). We created microparticles with three distinct degradation profiles and showed that the consequent protein release profiles could be modulated within the infarcted heart. This enabled us to determine how delivery rate impacted the efficacy of a model therapeutic, an engineered hepatocyte growth factor fragment (HGF-f). Our results showed that the cardioprotective efficacy of HGF-f was optimal when delivered over three days post-intramyocardial injection, yielding the largest arterioles, fewest apoptotic cardiomyocytes bordering the infarct and the smallest infarcts compared to empty particle treatment four weeks after injection. This work demonstrates the potential of using AcDex particles as a delivery platform to optimize the time frame for delivering therapeutic proteins to the heart.

During metastasis, cells break free from the primary tumor, migrate through the stroma and enter either the vascular or lymph systems to disseminate into secondary sites. To better understand this process, many groups have developed in vitro models of cancer metastasis using microfluidic based platforms to mimic the tumor microenvironment and the intravasation/extravasation of tumor cells. While these models have provided significant new insights into the mechanisms by which metastatic cells move across the endothelium, they are most often focused on the movement of single cells. Notably, there is significant clinical evidence to suggest that tumor cells can travel collectively though the stroma and can circulate as cell aggregates in the vasculature. Recent advances in the understanding of cancer metastasis have shown that cancer may move more aggressively as a collection of cell when traversing the extracellular matrix (ECM) of the tumor microenvironment as opposed to more studied singular cellular motif. To better mimic these collective movements, we have developed a model that incorporates the following essential elements that exist in the tumor microenvironment: a cylindrical channel lined with a confluent monolayer of endothelial cells, a metastatic tumor analog in close proximity to the channel, the ability to incorporate flow with tunable properties to simulate shear stress due to blood flow, and a tunable collagen microenvironment. Briefly, in a 3D printed cube, Type 1 collagen is added at physiological stiffness. As the collagen is polymerizing in the device, a metastatic tumor analog (spheroid) consisting of labeled MDA-MB-231 cells is placed in proximity of the vessel that is lined with endothelial cells. The platform allows for the visualization of metastatic cell intravasation using confocal microscopy. Notably, this model is easy to build and implement with fairly accessible tools and without the use of a clean room or microfabrication facilities. Importantly, going forward this model has the ability to study the cellular behavior post-intravasation due to the incorporation of physiological flows within the channel. Citation Format: Adam Munoz, Joseph Miller, Cynthia A. Reinhart-King. Development of an in vitro 3D vessel-spheroid model for investigating cancer metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr A15.