Engineered Hepatocellular Models for Drug Development

Abstract

Primary hepatocytes of adult human and rodent origin are essential components for developing drugs against infectious pathogens and for studying drug mediated liver toxicity. One of the key drawbacks limiting the use of these primary hepatocytes in vitro is their rapid loss of differentiated function, polarity, inability to recapitulate drug responses accurately and failure to capture the life cycle of pathogens. Although multiple platforms have been developed to improve functional maintenance of hepatocytes in culture, there is little understanding on the utility of these models for applications like toxicology and infections by various liver specific pathogens. In this thesis we have studied the utility of spheroid cultures of human hepatocytes to support hepatitis C infection and replication and sandwich culture of rat hepatocytes and co-culture of rat hepatocytes with fibroblasts for drug testing applications. Spheroid culture models of human hepatocytes and human hepatoma cells maintain and enhance liver specific functions, while localizing various liver specific proteins at domains similar to that found in vivo. These spheroid models maintain polarity over prolonged cultures and support glycoprotein mediated HCV entry. Huh 7.5 also support higher levels of replication of HCV virus in vitro. This makes it a suitable model to screen for drugs inhibiting HCV entry and replication. Rat hepatocyte culture with fibroblasts (co-culture) enhances hepatocyte specific synthetic and metabolic functions. However co-culture of hepatocytes with fibroblasts inhibits drug-induced CYP 450 responses. We found that TGF?1 is an important cytokine in co-culture responsible for repression of drug-induced responses. Soluble factor mediated repression of drug-induced CYP 450 responses makes co-culture an unsuitable model to study drug induction/inhibition and drug-drug interactions. We have analyzed the strengths of different hepatocyte culture models and demonstrated the strengths of different models for applications pertaining to drug development.