Jim Corbett, CEO of Emulate, Inc., released testimony today in support of the FDA Modernization Act of 2021 (H.R 2565 and S. 2952). This submission represents Emulate’s enthusiastic endorsement of the act and recognition that this step is essential for increasing the safety and efficacy of pharmaceutical development. Standards for drug development have remained essentially unchanged since the passage of the Federal Food, Drug, and Cosmetic Act (FFDCA) in 1938, which required all drugs in development to be tested in animals before reaching humans. While this did lead to a safer drug-development process at the time, species differences between animals and humans often mean the results from preclinical research cannot accurately apply to humans. This means that a drug’s toxicity could remain hidden until it reaches human trial or the market.
It is for these reasons that Corbett proudly submits testimony in support of the FDA Modernization Act, which will allow sponsor organizations to submit data from studies using microphysiological systems and other non-animal alternatives, such as Emulate’s Organs-on-Chips.
Jim Corbett, CEO Emulate, Inc.
Before the Subcommittee on Health of the Committee on Energy and Commerce Thursday, March 17, 2022
The Future of Medicine: Legislation to Encourage Innovation and Improve Oversight
On behalf of Emulate, Inc., the leading provider of organ-on-a-chip technology, I offer this testimony in support of the FDA Modernization Act of 2021 (H.R 2565 and S. 2952).
Since the Federal Food, Drug, and Cosmetics Act (FFDCA) of 1938 mandated that all new drugs be tested for toxicity in animals prior to human studies, scientific advancements have been abundant. However, predicting human toxicity through animals models still leaves us with a public health issue.
There is no doubt that animal models have contributed to major advances in medicine and have contributed to safe and effective drugs making it to market. However, these models have the difficult job of approximating the human body, and sometimes they get it wrong.
A growing body of evidence suggests that animal models are lacking in both sensitivity and specificity when it comes to predicting drug toxicity in humans.1-3 A 2014 study analyzing the effects of 2,366 drugs in both animals and humans found that “tests on animals (specifically rat, mouse and rabbit models) are highly inconsistent predictors of toxic responses in humans and are little better than what would result merely by chance.”4 A 2008 review found similar results, concluding that animal models predicting drug toxicity in humans may have sensitivity and specificity values below 70%.2
The cost of poor specificity and selectivity is too often passed onto the patient. A review of 578 discontinued and withdrawn drugs in Europe and the United States showed that nearly half halted distribution due to post-approval toxicity.5 Similarly, a 2012 analysis of 43 post-approval drugs with serious toxicity effects found that only 19% of them showed indications of toxicity in animal studies.6
In a recent study published to bioRxiv, researchers found the human Liver-Chip to have an 87% sensitivity and 100% specificity when differentiating hepatotoxic from non-hepatotoxic small molecules.7 Importantly, all 22 hepatotoxic drugs included in the study had previously been classified as safe due to a lack of toxicity in animal models. Collectively, these compounds resulted in 208 patient fatalities and 10 liver transplants. Had the human Liver-Chip been used during preclinical screening of these compounds, it’s likely that many of these fatalities could have been avoided.
Animal models have played an undeniably significant role in the evolution of medicine, and will continue to do so, but to make the drug development process safer, more efficient, and more humane, we must take a hard look at how we can leverage scientific advancements to continuously improve patient safety.
Van Norman GA. Limitations of animal studies for predicting toxicity in clinical trials: Is it time to rethink our current approach? JACC Basic Transl Sci. 2019;4(7):845-854. 2019. doi: 10.1016/j.jacbts.2019.10.008
Matthews RA. Medical progress depends on animal models – doesn’t it? J R Soc Med. 2008;101(2):95-98. doi: 10.1258/jrsm.2007.070164
Bailey J, Thew M, Balls M. An analysis of the use of animal models in predicting human toxicology and Drug Safety. Altern Lab Anim. 2014;42(3):181-199. doi: 10.1177/026119291404200306
Siramshetty VB, Nickel J, Omieczynski C, Gohlke BO, Drwal MN, Preissner R. WITHDRAWN–a resource for withdrawn and discontinued drugs. Nucleic Acids Res. 2016;44(D1):D1080-D1086. doi: 10.1093/nar/gkv1192
van Meer PJK, Kooijman M, Gispen-de Wied CC, Moors EHM, Schellekens H. The ability of animal studies to detect serious post marketing adverse events is limited. Regul Toxicol Pharmacol. 2012;64(3):345-349. doi: 10.1016/j.yrtph.2012.09.002
Ewart, Lorna, et al. Qualifying a Human Liver-Chip for Predictive Toxicology: Performance Assessment and Economic Implications. 16 Dec. 2021, 10.1101/2021.12.14.472674. Accessed 8 Mar. 2022.
Learn how these two complementary technologies can be combined for improved physiological relevance
In recent years, organoids—tiny, self-organized, three-dimensional cell models—have emerged as a promising technology for researching human physiology and disease. A major advantage of organoids is that they can be developed from induced pluripotent stem cells (iPSCs) or stem cells from primary human biopsies. As a result, they are able to differentiate into a variety of cell types to contain a greater range of cellular diversity than conventional models such as immortalized Caco-2 cell lines.
However, organoids lack some critical elements of the in vivo intestinal microenvironment, which limits their physiological relevance, such as the presence of vasculature and the mechanical forces caused by fluid flow and peristalsis. Additionally, their spherical structure results in several experimental challenges, including inconsistencies in size and shape, poor experimental control of key variables, and access to only one side of the epithelium.
Fortunately, researchers can unlock the full potential of organoids by using them as a robust cell source for Organ-Chips, enabling the creation of more accurate human biological models such as the Colon Intestine-Chip and Duodenum Intestine-Chip. Combining these technologies improves the organoids’ cellular morphology and functionality, results in more in vivo-like gene expression, and opens the door for new experimental designs—from simple drug permeability assays to more complex studies of colon inflammation, immune cell recruitment, and colorectal cancer tumor cell invasion.
Combining Organ-Chips and Organoids
Each Emulate Organ-Chip contains a top (epithelial) channel and a bottom (endothelial) channel separated by a thin, flexible, porous membrane that enables cell-cell interaction. This membrane is coated with tissue-specific extracellular matrix (ECM) on top of which human cells can be cultured. To create the Intestine-Chip, intestinal epithelial organoids are first established from endoscopic biopsies of healthy adults.
They are then dissociated into fragments and seeded onto the ECM-coated porous membrane in the top channel. Meanwhile, primary microvascular endothelial cells are seeded on the other side of the ECM-coated porous membrane in the endothelial channel. Importantly, both the organoids and microvascular endothelial cells are intestine-specific, meaning researchers can model particular sections of the intestine, such as the duodenum, jejunum, ileum, or colon.
Distinct media flows through each channel to promote cellular differentiation, and stretch can be applied at different amplitudes and frequencies to create intestinal peristalsis-like motions. Over several days, the epithelial cells form a confluent monolayer in the top channel, and the endothelial cells form a complete blood vessel in the bottom channel.
The advantages of Organ-on-a-Chip technology
Using organoids as a cell source for Organ-Chips enables researchers to create more physiologically relevant models and allows for a greater range of study possibilities. Some of the advantages include:
Endothelial co-culture and tissue-tissue interactions
Organ-Chips allow for the inclusion of tissue-specific endothelial cells to recreate the epithelial-endothelial interface of the intestinal barrier and support tissue-tissue interactions—critical drivers of cellular function that organoids lack. This endothelial co-culture was shown to result in several distinct advantages in a study using the Colon Intestine-Chip, including enhanced epithelial polarity, correct localization of tight junction markers, a tight epithelial barrier with low permeability, and the formation of a mature brush border with densely packed and elongated microvilli. In addition, researchers can administer their drug candidate of interest through the vascular channel of Organ-Chips, recapitulating how many therapeutics reach the intended tissue in vivo.
Cell interactions and cytoarchitecture
Although the cells in organoids are in a 3D structure, their organization does not resemble what it would be in vivo. The lack of directional cues results in somewhat random tissue organization, and the spherical shape results in reduced oxygen exposure in their center, often resulting in necrotic cores. In contrast, Organ-Chips provide the appropriate microenvironmental conditions for epithelial cells to spontaneously organize into physiological cytoarchitecture, including correct polarity and the formation of microvilli.
Dynamic media flow
Unlike organoids in static culture, Emulate Organ-Chips are designed to allow for continuous, unidirectional media flow, enabling steady-state nutrient levels and recreating the dynamic shear forces cells experience in the body. In the Duodenum Intestine-Chip, this media flow was shown to positively affect tissue architecture, resulting in increased cell height, cobblestone-like morphology, well-defined cell-cell junction formation, and dense microvilli.
With the Zoë Culture Module, researchers can fine-tune the frequency and strain of the chip’s flexible membrane to create peristalsis-like mechanical forces, enabling studies not possible with animals or alternative in vitro models. Recently, researchers at the Ellison Institute of USC applied this unique functionality to study the role of peristalsis in colorectal cancer tumor cell invasion. The Pasteur Institute has also leveraged this capability to study the impact of mechanical forces on Shigella infection and found that peristalsis is critical for specific stages of the infection process.
Improved gene expression
Multiple RNA-Seq analyses have shown that organoid-derived epithelial cells cultured in Emulate Organ-Chips have a transcriptome profile significantly closer to in vivo tissue than those same organoids in suspension. Analysis of specific pathways revealed differences in epithelial differentiation and key metabolic enzymes and pathways, indicating enhanced cell differentiation. This reinforces the advanced functionality of endothelial co-culture and the dynamic chip microenvironment. Given the closer in vivo gene expression, Organ-Chip models with organoids are more likely to express drug targets than organoids alone.
Immune cell incorporation
The fluidic nature of Emulate Organ-on-a-Chip technology allows users to introduce circulating immune cells through the chip channels, which is critical for modeling some aspects of disease. Additionally, research published in eLife used this approach to evaluate the safety of T-cell bispecific antibodies, a cancer immunotherapy difficult to study in animals due to fundamental species differences in immunological response.
Increase the physiological complexity of your organoid studies
Creating the next generation of effective therapeutics requires more human-relevant models of health and disease. While organoids offer several advantages over traditional monolayers, it is only when they are combined with Organ-on-a-Chip technology that their full potential can be realized, with improvements to cell morphology, functionality, and gene expression. By leveraging these advanced in vitro models, researchers can model more complex human biological mechanisms—including peristalsis, tumor cell migration, and immune cell interaction—enabling studies not possible with conventional models.
Contact us to learn more about how Organ-on-a-Chip technology can help you improve the physiological relevance of your research.
See how the Colon Intestine-Chip has been used to model cytokine-mediated intestine inflammation and barrier disruption.
Bacteria, viruses, and potential toxins all transit through the human intestines. In healthy conditions, the intestinal barrier serves as a protective wall, helping to prevent the engagement of these would-be biological agents. However, when this protective barrier breaks down, problems arise. Intestinal barrier dysfunction is often associated with chronic inflammation and is increasingly linked topathological conditions ranging from inflammatory bowel disease (IBD) to Parkinson’s Disease. These observations suggest that intestinal barrier deterioration may influence pathogenesis of some diseasesand have value as a therapeutic target.
understanding of the processes that lead to intestinal barrier deterioration is limited, due in part to a lack of human-relevant models. The intestineis a dynamic organ consisting of many diverse cell types whose behaviors are influenced by the complex milieu of cell-cell interactions, peristaltic contractions, and various environmental factors. Normally, it is exceedingly difficult for single-model systems to capture this complexity. However, recent evidence suggests Organ–a-Chip technology can provide a strong approximation of in vivo conditions, making it an invaluable tool for studying intestinal biology.
In a paper published in Cellular and Molecular Gastroenterology and Hepatology, researchers from Emulate characterize a colon intestine model in which patient-derived colonic organoids are cultured in a dynamic Organ-on-a-Chip platform. Unlike conventional models, this “gut-on-a-chip” includes primary human cells that are subject to biomechanical forces and co-cultured with intestine-specific endothelial cells, closely resembling the phenotypic characteristics of in vivo tissue.
Collectively, the data presented in this paper highlights the Colon Intestine-Chip’s ability to provide detailed insights into the human intestine barrier in health and disease settings.
Research Area: Gastroenterology, Disease pathology Organisms: Human Sample Types:Colon Intestine-Chip Research Question: Can this “gut-on-a-chip” be used to model the effects of cytokines, therapeutics, and other agents on intestinal barrier integrity?
Co-culture of colonoids with endothelial cells in the Colon Intestine-Chip results in improved epithelial cell phenotypic and transcriptomic profiles that more accurately represent in vivo observations compared to immortalized epithelial cell monolayers or colonoids cultured in suspension.
Perfusion of the Colon Intestine-Chip vascular chamber with IFN-γ promotes inflammatory phenotypes in epithelial cells, breakdown of tight junctions in the epithelial cell barrier, and subsequent increased barrier permeability.
Treatment of the Colon Intestine-Chip with Interleukin-22 (IL-22) promotes inflammatory signaling and tight junction breakdown, shedding light on the potential role of IL-22 in the pathogenesis of intestinal barrier deterioration.
Conclusion: The Colon Intestine-Chip represents an improved model of the human colon that contains a heterogeneous epithelial cell layer displaying phenotypic and transcriptomic profiles similar to those observed in vivo. Using this model, researchers can effectively investigate the mechanisms behind cytokine-mediated inflammation and the efficacy of therapeutic candidates on human colonic barrier integrity. Because of this, the Colon Intestine-Chip can help shed light on the complex relationship between intestinal barrier integrity and disease pathogenesis.
Modeling a dynamic organ
Researchers aiming to study gastrointestinal physiology and disease primarily rely on three types of models: animals, organoids, and conventional monolayer cultures of immortalized cell-lines. Each of these has been invaluable in advancing our understanding of gut physiology; however, none are able to recreate the critical features of the human intestine that influence cellular response to stressors and—in turn—disease pathogenesis. Because of this, it has been challenging to translate results from these models into effective disease modifying therapies.
Organ-on-a-Chip technology presents a promising alternative. Emulate Organ-Chips are three-dimensional, dynamic systems that co-culture tissue-specific cell types—such as epithelial cells and immune cells—alongside endothelial cells under fluid flow and in the presence of tissue-specific extracellular matrix proteins. Selectively seeding cells into the chip’s two channels enables the formation of a vascular chamber consisting of endothelial cells and a tissue chamber containing the remaining cell types. These two channels are separated by a thin, porous membrane to enable communication between cell chambers while maintaining distinct microenvironments.
To improve on current models of the intestines, researchers from Emulate leveraged Organ-Chip technology to develop a Colon Intestine-Chip.
Colon Intestine-Chip: an improved model of gastrointestinal physiology
To create the Colon Intestine-Chip, Apostolou et al., made use of Emulate’s Organ-on-a-Chip technology, which enables multiple cell types to be co-cultured in a dynamic environment. Colonic organoids (colonoids), which came from healthy patient biopsies and were mechanically dissociated, served as the basis for the chip’s intestinal chamber. In parallel, colonic human intestinal microvascular endothelial cells were seeded in the vascular chamber. When exposed to unidirectional media flow as well as cyclic 10% stretch to emulate peristalsis, the model closely resembles the microenvironment intestinal cells would experience in vivo.
Characterization of the Colon Intestine-Chips revealed a close phenotypic resemblance to healthy colonic barriers, including: The formation of a confluent, highly polarized epithelial cell barrier with low permeability (0.89 x 10-6 cm / s); the localization of tight junction proteins at intercellular junctions; the asymmetric distribution of ion channels; and the formation of a mature brush border with densely packed microvilli.
Notably, the epithelial cells’ mature phenotype was dependent on the presence of endothelial cells within the chip’s vascular chamber. Exclusion of endothelial cells from the Colon Intestine-Chip led to increased barrier permeability, decreased tight junction formation, and decreased epithelial cell polarization, collectively demonstrating the importance of endothelial co-culture with epithelial cells in modeling the intestinal barrier.
Given these findings, it’s unlikely that colonoids or conventional monolayer culture models could mimic in vivo conditions as well as the Colon Intestine-Chips. This claim is reinforced by transcriptomic data collected from Colon Intestine-Chips and colonoid cells grown in suspension, which showed that the presence of endothelial cells and periodic stretching in Colon Intestine-Chips produced superior gene expression profiles.
Using the Colon Intestine-Chip to model gut barrier breakdown
To study To study the pathophysiology of gut barrier dysfunction, the team perfused the vascular chamber with interferon gamma (IFN-γ), a cytokine known to affect the pathogenesis of inflammatory bowel disease.
Within two days of treatment, the epithelial cell barrier showed clear signs of distress. Epithelial barrier permeability increased in an IFN-γ-concentration-dependent manner, tight junction proteins were sequestered to the cellular cytoplasm, and F-actin staining revealed cell deformations and poorly defined cell borders. Collectively, these findings indicate that the presence of IFN-γ was driving a breakdown in the epithelial cell barrier—a conclusion that was strongly reinforced by an increase in epithelial cell death (as indicated by elevated levels of cleaved caspase).
What’s more, treatment with IFN-γ prompted an increase in the cytokine IL-6 and vascular adhesion molecule-1—both of which have been found in the sera of patients with inflammatory bowel disease—reinforcing that this model is able to accurately recreate aspects of an IBD-like clinical phenotype.
Advancing our understanding of gut barrier physiology and pathophysiology
Interleukin-22 (IL-22) is a cytokine released by various immune cells in response to pathogens. To date, our understanding of IL-22’s role in intestinal health and disease is incomplete. Various ulcerative colitis studies using mice have found conflicting results, with some results suggesting IL-22 has pro-inflammatory effects and others suggesting it has anti-inflammatory effects.
After showing that the Colon Intestine-Chip model can accurately recreate a mature intestinal epithelial cell barrier phenotype and model the effect of well-characterized, barrier-disrupting cytokine IFN-γ, the authors evaluated whether the model could shed light on the true role of IL-22 in intestinal barrier function.
Before administering IL-22, the team first confirmed the expression of the IL-22 receptor and found that expression was higher in the Colon Intestine-Chip compared with organoids in suspension. These results suggest that our incomplete understanding of IL-22’s role in barrier homeostasis may be due in part to limited gene expression in other models.
Perfusing IL-22 through the vascular chamber negatively affected epithelial cell barrier function in the Colon Intestine-Chip model. Barrier permeability increased, cell morphology became aberrant, and transcriptomic profiles and immunofluorescent staining revealed a marked increase in apoptosis. Taken together, these results suggest that IL-22 drives barrier dysfunction.
ln total, this study showed that the Colon Intestine-Chip can be a powerful model for studying intestinal barrier dysfunction. Immunofluorescent staining, scanning electron microscopy, and RNA sequencing data show that the Colon Intestine-Chip model produces a mature epithelial cell phenotype that responds to inflammatory cytokines, such as IFN-γ, in ways that reflect observations from patients with inflammatory bowel disease.
Importantly, this study showed that endothelial co-culture is critical to promote a mature, functional epithelial phenotype, driving positive effects on cell morphology, polarization, and barrier formation. This insight highlights an advantage Organ-Chips have over intestinal models without endothelial co-culture, such as organoids in suspension.
The team’s use of the Colon Intestine-Chip to study IL-22’s effects on intestinal barrier integrity demonstrates the potential to apply this model in studying mechanisms of intestinal barrier dysfunction. Collectively, this study shows how the Colon Intestine-Chip is a more physiologically relevant model of the human colon that researchers can use to study gastrointestinal disease pathogenesis and the efficacy or safety of preclinical drug candidates in preclinical stages.