Drug Discovery and Regulatory Considerations for Improving In Silico and In Vitro Predictions that Use Caco-2 as a Surrogate for Human Intestinal Permeability Measurements

Abstract

ABSTRACT. There is a growing need for highly accurate in silico and in vitro predictive models to facilitate drug discovery and development. Results from in vitro permeation studies across the Caco-2 cell monolayer are commonly used for drug permeability screening in industry and are also accepted as a surrogate for human intestinal permeability measurements by the US FDA to support new drug applications. Countless studies carried out in this cell line with published permeability measurements have enabled the development of many in silico prediction models. We identify several common cases that illustrate how using Caco-2 permeability measurements in these in silico and in vitro predictive models will not correlate with human intestinal permeability and will further lead to inaccuracies in these models. We provide guidelines and recommendations for improving these models to more accurately predict clinically relevant information, thereby enhancing the drug discovery, development, and regulatory approval processes.

Predicting when Biliary Excretion of Parent Drug is a Major Route of Elimination in Humans

“…Finally, low permeability compounds were highly represented as substrates of hepatic basolateral uptake transporters (32). Importantly, their data indicate that in vitro permeability rate can be used as a surrogate for extent of metabolism for new molecular entities (NMEs) when clinical data is unavailable, as has been proposed by our laboratory (31,33,34). Specifically, compounds with permeability rates equal to or exceeding a standard, e.g., metoprolol, are likely extensively metabolized in vivo in humans, while those with permeability rates lower than the standard are likely eliminated primarily as unchanged drug in either the bile or the urine.…”

“…Prior to any studies in animals or humans for an NME, in vitro permeability data, as demonstrated by Varma et al (32) and initially proposed by our laboratory (4,33,34), can identify which drugs are primarily eliminated by metabolic or non-metabolic (biliary, renal) routes. Highly permeable drugs are likely extensively metabolized in vivo.…”

Non-animal models of epithelial barriers (skin, intestine and lung) in research, industrial applications and regulatory toxicology

“…However, in spite of this achievement, as well as the various developments detailed above, it also needs to be noted that we are a long way from precise prediction of human bioavailability. Several examples are available of instances where in silico and in vitro predictive models based on Caco-2 permeability measurements do not correlate with human intestinal permeability (Larregieu and Benet, 2013).…”

Non-animal approaches for toxicokinetics in risk evaluations of food chemicals

“…Though Caco-2 cells are proficient in the main transporters, including P-glycoprotein (PgP), multidrug resistance protein 2 (MRP2), and breast cancer resistance proteins (BCRP), expression levels of these transporters are generally quite variable (Larregieu and Benet, 2013;Harwood et al, 2013Harwood et al, , 2016. In addition, the under-expression of transporters such as peptide transporter 1 (PEPT1), organic cation transporters (OCTs), and organic anion transporters (OATs), makes the model less suitable for compounds that use these transporters (Larregieu and Benet, 2013).…”

“…Though Caco-2 cells are proficient in the main transporters, including P-glycoprotein (PgP), multidrug resistance protein 2 (MRP2), and breast cancer resistance proteins (BCRP), expression levels of these transporters are generally quite variable (Larregieu and Benet, 2013;Harwood et al, 2013Harwood et al, , 2016. In addition, the under-expression of transporters such as peptide transporter 1 (PEPT1), organic cation transporters (OCTs), and organic anion transporters (OATs), makes the model less suitable for compounds that use these transporters (Larregieu and Benet, 2013). Under-expression of metabolic enzymes (e.g., CYP3A4) and different sulfotransferase and uridine diphospho-glucuronosyltransferase (UGT) enzymes, as compared to the human small intestine, also makes Caco-2 cells a poor model for studying intestinal metabolism (Gregory et al, 2004;Meinl et al, 2008;Peters et al, 2016;Schmiedlin-Ren et al, 1997).…”