The Promise of In Vitro 3D Organoid Models: Meeting the ADME-Tox Testing Needs of the Pharmaceutical Industry
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Inadequate prediction of drug metabolism or toxicity is the Achilles heel of the pharmaceutical industry, leading to high drug attrition rates. This can have significant implications for companies, with late-stage failure typically resulting in the loss of substantial financial investments, time and resources. By defining and predicting the Absorption, Distribution, Metabolism, Excretion and Toxicity (ADME-Tox) profile of compounds as early as possible, companies can focus their resources on the most relevant candidates to increase the likelihood of successfully bringing new safe and efficient therapies to the market.
To implement early ADME-Tox testing, it’s crucial to have access to valid in vitro models of the key sites of drug metabolism — the liver, kidneys and intestines. Preclinical models that reliably replicate the in vivo cellular environment of these organs enable scientists to study and accurately predict drug metabolism, transport and toxicity prior to targets entering the clinical trial phase, ultimately helping to lower attrition rates.
For many years, biologists have relied on in vitro 2-dimensional (2D) cell culture models to perform preclinical ADME-Tox assays. While these models are useful for informing aspects of drug discovery, such as cytotoxicity, they are limited in their translatability to drug metabolism and toxicity in humans. Recently, in vitro 3-dimensional (3D) organoid models that more closely mimic human biological systems show promise in supporting ADME-Tox studies in the early stages of the drug development process, compared to their 2D counterparts.
The limitations of conventional in vitro cellular models
Traditionally, in vitro 2D cell cultures derived from human cells provided an in vivo platform for testing responses to drug candidates. Although valuable for preclinical drug testing, the data generated by traditional 2D cell cultures don’t always faithfully recapitulate the in vivo situation. To-date, much work has been done to improve the physiological relevance of in vitro liver model systems, but less progress has been made with regard to intestine and kidney models.
For instance, a monolayer of cultured human colon epithelial cancer cells (Caco-2 cells) on top of a support membrane is often used to model the human intestinal absorption of drugs. Displaying tight junctions and expressing proteins involved in drug transport, Caco-2 cells function as an in vitro model of the permeability, of test substances.1 However, Caco-2 monolayers have demonstrated low permeability compared to the in vivo situation, and also poor and variable enzymatic activity, which limits their translatability.2
For modeling kidney cell function in vitro and to evaluate general nephrotoxicity, researchers use various types of human proximal tubule cell lines. These cells demonstrate specific properties of the kidney epithelium, such as the transport of solutes, which plays an important role in drug excretion,. Limitations of these in vitro models are due to the different cell types not expressing all necessary transporters, metabolizing enzymes or biomarkers at physiological levels.3
Given these challenges, there is a clear need for advanced in vitro 3D models that more accurately emulate drug permeability, metabolism, transport and toxicity in humans.
More physiologically relevant modeling using advanced in vitro 3D platforms
Novel in vitro 3D models have been designed to bridge the gap between conventional in vivo models and 2D cell cultures. In some instances, seeding primary cells that were originally cultured on 2D surfaces into a 3D format can restore some of their physiological morphologies and functions.4 As a result, it becomes possible to develop miniature organs — so-called organoids — to evaluate drug responses.
Organoids are tissue cultures derived from primary cells or progenitor cells that, when grown in appropriate 3D conditions, can spontaneously self-organize into differentiated functional cell types. Organoids can replicate at least some of the functions of human organs, enabling ADME-Tox assays to be performed in vitro.
Due to a greater understanding of the cell microenvironment, organoid technology has advanced over the last few years and has the potential to streamline the drug development process. Using organoids to support in vitro ADME-Tox studies can help to predict metabolic mechanisms and ascertain key safety and efficacy measures before commencing human clinical trials. Evidence suggests that kidney and intestine organoid models could be more valuable to investigate the metabolism, transport and toxicity of drugs.3,4 Indeed, a kidney organoid model has recently been cultured using human induced pluripotent stem cells. Composed of both glomerular tissue, as well as proximal and distal tubule cells, this model offers a more accurate representation of the kidney. As such, it has the potential to inform and refine preclinical toxicity screening studies.3
In another recent study, human primary cells from intestinal epithelium was engineered into a 3D intestinal organoid using a scaffold system . This model showed complex tissue properties and characteristics of mature epithelium, including the four main types of differentiated epithelial cells (enterocytes, goblet cells, paneth cells and enteroendocrine cells). The tight junction formation, microvilli polarization, digestive enzyme secretion and low oxygen tension in the lumen were also representative of mature epithelium. In addition to these physical properties, the organoids also exhibited complex behavior, such as innate antibacterial responses to E. coli similar to those observed in patients with inflammatory bowel disease (IBD). This suggests that the model could be used in in vitro studies investigating host-microbe-pathogen interplay and IBD pathogenesis.
These findings highlight the future potential of in vitro 3D kidney and intestine organoids for drug development. Because these systems closer resemble in vivo tissues, they could help predict drug responses early in development and offer vast possibilities for modeling many diseases in the future.
Optimizing drug development with the latest advances in organ modeling technology
To accelerate the launch of innovative therapeutics, more reliable means of assessing drug permeability, metabolism, transport and toxicity in the early stages of the development process are essential. Technology advances have enabled more physiologically relevant in vitro 3D organoid liver models to be cultured, which are now routinely used to support ADME-Tox studies in preclinical drug development. The natural next step would be to develop similar kidney and intestine models to allow for an accurate monitoring of drug safety and efficacy, such as intestinal disposition and kidney toxicity.
1. Van Breemen R.B and Li Y. Caco-2 cell permeability assays to measure drug absorption. Expert Opin Drug Metab Toxicol. 2005 Aug;1(2):175–85
2. Yamaura Y, et al. Functional Comparison of Human Colonic Carcinoma Cell Lines and Primary Small Intestinal Epithelial Cells for Investigations of Intestinal Drug Permeability and First-Pass Metabolism. Drug Metabolism and Disposition. March 2016;44:329–335
3. Bajaj P, et al. Emerging Kidney Models to Investigate Metabolism, Transport, and Toxicity of Drugs and Xenobiotics. Drug Metabolism and Disposition. November 2018;46(11):1692–1702
4. Chen Y, et al. In vitro enteroid-derived three-dimensional tissue model of human small intestinal epithelium with innate immune responses. PLoS One. 2017;12(11): e0187880