Robust and Scalable Angiogenesis Assay of Perfused 3D Human iPSC-Derived Endothelium for Anti-Angiogenic Drug Screening

Duinen, Vincent van; Stam, Wendy; Mulder, E.; Famili, Farbod; Reijerkerk, Arie; Vulto, Paul; Hankemeier, Thomas; Zonneveld, Anton Jan van

doi:10.3390/ijms21134804

Cited by 29 publications

(33 citation statements)

References 30 publications

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“…This is particularly the case, as more reliable estimates of the therapeutic window for subsequent in vivo experimentation can be deduced from 3D vs. 2D assays through more refined information on dose-dependent drug efficacies and their respective cytotoxicity profiles. Furthermore, in vitro assays generally bear the potential for cost-efficient adaption into a scalable assay platform to screen antiangiogenic compounds in combination with other drugs ( Siller et al, 2020 ; Van Duinen et al, 2020 ). To our knowledge, there has been no in vitro 3D model described to simultaneously validate the antiangiogenic as well as vascular disrupting properties of drugs.…”

Section: Introductionmentioning

confidence: 99%

3D-Cultured Vascular-Like Networks Enable Validation of Vascular Disruption Properties of Drugs In Vitro

Yavvari

Laporte

Elomaa

et al. 2022

Front. Bioeng. Biotechnol.

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Vascular-disrupting agents are an interesting class of anticancer compounds because of their combined mode of action in preventing new blood vessel formation and disruption of already existing vasculature in the immediate microenvironment of solid tumors. The validation of vascular disruption properties of these drugs in vitro is rarely addressed due to the lack of proper in vitro angiogenesis models comprising mature and long-lived vascular-like networks. We herein report an indirect coculture model of human umbilical vein endothelial cells (HUVECs) and human dermal fibroblasts (HDFs) to form three-dimensional profuse vascular-like networks. HUVECs embedded and sandwiched in the collagen scaffold were cocultured with HDFs located outside the scaffold. The indirect coculture approach with the vascular endothelial growth factor (VEGF) producing HDFs triggered the formation of progressively maturing lumenized vascular-like networks of endothelial cells within less than 7 days, which have proven to be viably maintained in culture beyond day 21. Molecular weight-dependent Texas red-dextran permeability studies indicated high vascular barrier function of the generated networks. Their longevity allowed us to study the dose-dependent response upon treatment with the three known antiangiogenic and/or vascular disrupting agents brivanib, combretastatin A4 phosphate (CA4P), and 6´-sialylgalactose (SG) via semi-quantitative brightfield and qualitative confocal laser scanning microscopic (CLSM) image analysis. Compared to the reported data on in vivo efficacy of these drugs in terms of antiangiogenic and vascular disrupting effects, we observed similar trends with our 3D model, which are not reflected in conventional in vitro angiogenesis assays. High-vascular disruption under continuous treatment of the matured vascular-like network was observed at concentrations ≥3.5 ng·ml−1 for CA4P and ≥300 nM for brivanib. In contrast, SG failed to induce any significant vascular disruption in vitro. This advanced model of a 3D vascular-like network allows for testing single and combinational antiangiogenic and vascular disrupting effects with optimized dosing and may thus bridge the gap between the in vitro and in vivo experiments in validating hits from high-throughput screening. Moreover, the physiological 3D environment mimicking in vitro assay is not only highly relevant to in vivo studies linked to cancer but also to the field of tissue regeneration.

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Section: Introductionmentioning

confidence: 99%

3D-Cultured Vascular-Like Networks Enable Validation of Vascular Disruption Properties of Drugs In Vitro

Yavvari

Laporte

Elomaa

et al. 2022

Front. Bioeng. Biotechnol.

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“…In future studies, the coculture model presented here can be used to further investigate tumor cell invasion into lymphatic microvasculature, for instance by studying tumor cell motility in real time or quantifying tumor cell arrival into the lumen of the main lymphatic vessel. Moreover, the coculture model can be used for antilymphangiogenic or other anticancer compounds in high-throughput drug screens, as was reported for human iPSC-derived and HUVEC endothelia in the 3-lane OrganoPlate …”

Section: Discussionmentioning

confidence: 99%

“…In the reported setups aside from the Swartz model, the chips are experimental in nature and do not provide throughput that would be required for routine experimentation. Here, employing a 3-lane OrganoPlate , that allows patterning of an extracellular matrix gel and adjacent growth of a vessel-like structure, we created a microfluidic lymphatic vessel culture, verified its barrier function and showed it capable of sustained lymphangiogenesis. To illustrate our further intentions for using this model for studying lymphatic endothelial cell–cancer cell interaction in the future, we created a coculture model using mouse-derived colon cancer organoids.…”

Section: Introductionmentioning

confidence: 99%

Long-Lived Human Lymphatic Endothelial Cells to Study Lymphatic Biology and Lymphatic Vessel/Tumor Coculture in a 3D Microfluidic Model

Frenkel

Poghosyan

Alarcón

et al. 2021

ACS Biomater. Sci. Eng.

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The lymphatic system is essential in maintaining tissue fluid homeostasis as well as antigen and immune cell transport to lymph nodes. Moreover, lymphatic vasculature plays an important role in various pathological processes, such as cancer. Fundamental to this research field are representative in vitro models. Here we present a microfluidic lymphatic vessel model to study lymphangiogenesis and its interaction with colon cancer organoids using a newly developed lymphatic endothelial cell (LEC) line. We generated immortalized human LECs by lentiviral transduction of human telomerase (hTERT) and BMI-1 expression cassettes into primary LECs. Immortalized LECs showed an increased growth potential, reduced senescence, and elongated lifespan with maintenance of typical LEC morphology and marker expression for over 12 months while remaining nontransformed. Immortalized LECs were introduced in a microfluidic chip, comprising a free-standing extracellular matrix, where they formed a perfusable vessel-like structure against the extracellular matrix. A gradient of lymphangiogenic factors over the extracellular matrix gel induced the formation of luminated sprouts. Adding mouse colon cancer organoids adjacent to the lymphatic vessel resulted in a stable long-lived coculture model in which cancer cell-induced lymphangiogenesis and cancer cell motility can be investigated. Thus, the development of a stable immortalized lymphatic endothelial cell line in a membrane-free, perfused microfluidic chip yields a highly standardized lymphangiogenesis and lymphatic vessel–tumor cell coculture assay.

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“…The vascularization of whole microfluidic circuits on-chip was shown as early as 2013 (132), followed by fascinating work on the generation of vascularized organ models on-chip (133)(134)(135)(136). As of today, high-throughput platforms generating vascularized single-tissue models on-chip are commercially available (137). The combination of both technologies generating a closed vascularized circuit containing multiple organs onchip is within reach, allowing for the next level of physiological complexity-the perfusion of whole blood or a defined substitute containing all relevant components.…”

Section: Microfluidic Cell Culture Systems-the Key Toward the Integration Of Premature Organoids Into Organismoidsmentioning

confidence: 99%

An Individual Patient's “Body” on Chips—How Organismoid Theory Can Translate Into Your Personal Precision Therapy Approach

Marx

Accastelli²,

David

et al. 2021

Front. Med.

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The first concepts for reproducing human systemic organismal biology in vitro were developed over 12 years ago. Such concepts, then called human- or body-on-a-chip, claimed that microphysiological systems would become the relevant technology platform emulating the physiology and morphology of human organisms at the smallest biologically acceptable scale in vitro and, therefore, would enable the selection of personalized therapies for any patient at unprecedented precision. Meanwhile, the first human organoids—stem cell-derived complex three-dimensional organ models that expand and self-organize in vitro—have proven that in vitro self-assembly of minute premature human organ-like structures is feasible, once the respective stimuli of ontogenesis are provided to human stem cells. Such premature organoids can precisely reflect a number of distinct physiological and pathophysiological features of their respective counterparts in the human body. We now develop the human-on-a-chip concepts of the past into an organismoid theory. We describe the current concept and principles to create a series of organismoids—minute, mindless and emotion-free physiological in vitro equivalents of an individual's mature human body—by an artificially short process of morphogenetic self-assembly mimicking an individual's ontogenesis from egg cell to sexually mature organism. Subsequently, we provide the concept and principles to maintain such an individual's set of organismoids at a self-sustained functional healthy homeostasis over very long time frames in vitro. Principles how to perturb a subset of healthy organismoids by means of the natural or artificial induction of diseases are enrolled to emulate an individual's disease process. Finally, we discuss using such series of healthy and perturbed organismoids in predictively selecting, scheduling and dosing an individual patient's personalized therapy or medicine precisely. The potential impact of the organismoid theory on our healthcare system generally and the rapid adoption of disruptive personalized T-cell therapies particularly is highlighted.

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Robust and Scalable Angiogenesis Assay of Perfused 3D Human iPSC-Derived Endothelium for Anti-Angiogenic Drug Screening

Cited by 29 publications

References 30 publications

3D-Cultured Vascular-Like Networks Enable Validation of Vascular Disruption Properties of Drugs In Vitro

3D-Cultured Vascular-Like Networks Enable Validation of Vascular Disruption Properties of Drugs In Vitro

Long-Lived Human Lymphatic Endothelial Cells to Study Lymphatic Biology and Lymphatic Vessel/Tumor Coculture in a 3D Microfluidic Model

An Individual Patient's “Body” on Chips—How Organismoid Theory Can Translate Into Your Personal Precision Therapy Approach

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