Over the past six decades the inflation-adjusted cost to bring a new drug to market has been increasing constantly and doubles every 9 years - now reaching in excess of $2.5 billion. Overall, the ...likelihood of FDA approval for a drug (any disease indication) that has entered phase I clinical trials is a mere 9.6%, with the approval rate for oncology far below average at only 5.1%. Lack of efficacy or toxicity is often not revealed until the later stages of clinical trials, despite promising preclinical data. This indicates that the current in vitro systems for drug screening need to be improved for better predictability of in vivo outcomes. Microphysiological systems (MPS), or bioengineered 3D microfluidic tissue and organ constructs that mimic physiological and pathological processes in vitro, can be leveraged across preclinical research and clinical trial stages to transform drug development and clinical management for a range of diseases. Here we review the current state-of-the-art in 3D tissue-engineering models developed for cancer research, with a focus on tumor-on-a-chip, or tumor chip, models. From our viewpoint, tumor chip systems can advance innovative medicine to ameliorate the high failure rates in anti-cancer drug development and clinical treatment.
This paper reports a method for generating an intact and perfusable microvascular network that connects to microfluidic channels without appreciable leakage. This platform incorporates different ...stages of vascular development including vasculogenesis, endothelial cell (EC) lining, sprouting angiogenesis, and anastomosis in sequential order. After formation of a capillary network inside the tissue chamber via vasculogenesis, the adjacent microfluidic channels are lined with a monolayer of ECs, which then serve as the high-pressure input ("artery") and low pressure output ("vein") conduits. To promote a tight interconnection between the artery/vein and the capillary network, sprouting angiogenesis is induced, which promotes anastomosis of the vasculature inside the tissue chamber with the EC lining along the microfluidic channels. Flow of fluorescent microparticles confirms the perfusability of the lumenized microvascular network, and minimal leakage of 70 kDa FITC-dextran confirms physiologic tightness of the EC junctions and completeness of the interconnections between artery/vein and the capillary network. This versatile device design and its robust construction methodology establish a physiological transport model of interconnected perfused vessels from artery to vascularized tissue to vein. The system has utility in a wide range of organ-on-a-chip applications as it enables the physiological vascular interconnection of multiple on-chip tissue constructs that can serve as disease models for drug screening.
There is a growing awareness that complex 3-dimensional (3D) organs are not well represented by monolayers of a single cell type - the standard format for many drug screens. To address this ...deficiency, and with the goal of improving screens so that drugs with good efficacy and low toxicity can be identified, microphysiological systems (MPS) are being developed that better capture the complexity of in vivo physiology. We have previously described an organ-on-a-chip platform that incorporates perfused microvessels, such that survival of the surrounding tissue is entirely dependent on delivery of nutrients through the vessels. Here we describe an arrayed version of the platform that incorporates multiple vascularized micro-organs (VMOs) on a 96-well plate. Each VMO is independently-addressable and flow through the micro-organ is driven by hydrostatic pressure. The platform is easy to use, requires no external pumps or valves, and is highly reproducible. As a proof-of-concept we have created arrayed vascularized micro tumors (VMTs) and used these in a blinded screen to assay a small library of compounds, including FDA-approved anti-cancer drugs, and successfully identified both anti-angiogenic and anti-tumor drugs. This 3D platform is suitable for efficacy/toxicity screening against multiple tissues in a more physiological environment than previously possible.
Mice are the experimental tool of choice for the majority of immunologists and the study of their immune responses has yielded tremendous insight into the workings of the human immune system. ...However, as 65 million years of evolution might suggest, there are significant differences. Here we outline known discrepancies in both innate and adaptive immunity, including: balance of leukocyte subsets, defensins, Toll receptors, inducible NO synthase, the NK inhibitory receptor families Ly49 and KIR, FcR, Ig subsets, the B cell (BLNK, Btk, and lambda5) and T cell (ZAP70 and common gamma-chain) signaling pathway components, Thy-1, gammadelta T cells, cytokines and cytokine receptors, Th1/Th2 differentiation, costimulatory molecule expression and function, Ag-presenting function of endothelial cells, and chemokine and chemokine receptor expression. We also provide examples, such as multiple sclerosis and delayed-type hypersensitivity, where complex multicomponent processes differ. Such differences should be taken into account when using mice as preclinical models of human disease.
Slug (SNAI2), a member of the well-conserved Snail family of transcription factors, has multiple developmental roles, including in epithelial-to-mesenchymal transition (EMT). Here, we show that Slug ...is critical for the pathological angiogenesis needed to sustain tumor growth, and transiently necessary for normal developmental angiogenesis. We find that Slug upregulation in angiogenic endothelial cells (EC) regulates an EMT-like suite of target genes, and suppresses Dll4-Notch signaling thereby promoting VEGFR2 expression. Both EC-specific Slug re-expression and reduced Notch signaling, either by γ-secretase inhibition or loss of Dll4, rescue retinal angiogenesis in SlugKO mice. Conversely, inhibition of VEGF signaling prevents excessive angiogenic sprouting of Slug overexpressing EC. Finally, endothelial Slug (but not Snail) is activated by the pro-angiogenic factor SDF1α via its canonical receptor CXCR4 and the MAP kinase ERK5. Altogether, our data support a critical role for Slug in determining the angiogenic response during development and disease.
This paper reports a polydimethylsiloxane microfluidic model system that can develop an array of nearly identical human microtissues with interconnected vascular networks. The microfluidic system ...design is based on an analogy with an electric circuit, applying resistive circuit concepts to design pressure dividers in serially-connected microtissue chambers. A long microchannel (550, 620 and 775 mm) creates a resistive circuit with a large hydraulic resistance. Two media reservoirs with a large cross-sectional area and of different heights are connected to the entrance and exit of the long microchannel to serve as a pressure source, and create a near constant pressure drop along the long microchannel. Microtissue chambers (0.12 μl) serve as a two-terminal resistive component with an input impedance >50-fold larger than the long microchannel. Connecting each microtissue chamber to two different positions along the long microchannel creates a series of pressure dividers. Each microtissue chamber enables a controlled pressure drop of a segment of the microchannel without altering the hydrodynamic behaviour of the microchannel. The result is a controlled and predictable microphysiological environment within the microchamber. Interstitial flow, a mechanical cue for stimulating vasculogenesis, was verified by finite element simulation and experiments. The simplicity of this design enabled the development of multiple microtissue arrays (5, 12, and 30 microtissues) by co-culturing endothelial cells, stromal cells, and fibrin within the microchambers over two and three week periods. This methodology enables the culturing of a large array of microtissues with interconnected vascular networks for biological studies and applications such as drug development.
Vascular morphogenesis encompasses a temporally regulated set of morphological changes that endothelial cells undergo to generate a network of interconnected tubules. Such a complex process ...inevitably involves multiple cell signaling pathways that must be tightly coordinated in time and space. The formation of a new capillary involves endothelial cell activation, migration, alignment, proliferation, tube formation, branching, anastomosis, and maturation of intercellular junctions and the surrounding basement membrane. Each of these stages is either known or suspected to fall under the influence of the vascular endothelial growth factor, notch, and transforming growth factor-beta/bone morphogenetic protein signaling pathways. Vascular endothelial growth factor is essential for initiation of angiogenic sprouting, and also regulates migration of capillary tip cells, proliferation of trunk cells, and gene expression in both. Notch has been implicated in the regulation of cell fate decisions in the vasculature, especially the choice between arterial and venular endothelial cells, and between tip and trunk cell phenotype. Transforming growth factor-beta regulates cell migration and proliferation, as well as matrix synthesis. In this review, we emphasize how crosstalk between these pathways is essential for proper patterning of the vasculature and offer a transcriptional oscillator model to explain how these pathways might interact to generate new tip cells during retinal angiogenesis.
There is a growing interest in developing microphysiological systems that can be used to model both normal and pathological human organs in vitro. This "organs-on-chips" approach aims to capture key ...structural and physiological characteristics of the target tissue. Here we describe in vitro vascularized microtumors (VMTs). This "tumor-on-a-chip" platform incorporates human tumor and stromal cells that grow in a 3D extracellular matrix and that depend for survival on nutrient delivery through living, perfused microvessels. Both colorectal and breast cancer cells grow vigorously in the platform and respond to standard-of-care therapies, showing reduced growth and/or regression. Vascular-targeting agents with different mechanisms of action can also be distinguished, and we find that drugs targeting only VEGFRs (Apatinib and Vandetanib) are not effective, whereas drugs that target VEGFRs, PDGFR and Tie2 (Linifanib and Cabozantinib) do regress the vasculature. Tumors in the VMT show strong metabolic heterogeneity when imaged using NADH Fluorescent Lifetime Imaging Microscopy and, compared to their surrounding stroma, many show a higher free/bound NADH ratio consistent with their known preference for aerobic glycolysis. The VMT platform provides a unique model for studying vascularized solid tumors in vitro.
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