Toxicology Assay Kits

Rapidly assess therapeutic candidates’ potential for drug-drug interactions and accurately predict xenobiotic-induced liver toxicity. INDIGO’s toxicology assay kits provide rapid, reliable results for:

Prospective screening

Retrospective screening

Safety pharmacology

upcycte Hepatocytes in INDIGO Toxicology Kits

Xenobiotic-induced liver injury is a major cause of human morbidity and mortality. A key reason for this problem is our inability to predict hepatotoxicity at the preclinical stage using currently available model systems. Such models include in vivo animal models and in vitro models based on human-derived liver cells or transformed cell systems. Species differences in xenobiotic disposition and mechanisms of cytotoxicity can make whole animal studies unreliable for extrapolation to humans. In addition, whole animal models are costly and of low throughput. Therefore, it is essential to develop in vitro models that are more predictive of hepatotoxicity, particularly those that are based on human or "humanized" component cells.

There are two major limitations to the use of human liver cells or their derivatives. First, there are currently limited sources of fresh human hepatocytes worldwide and, when available, they often suffer from low viability and high batch-to-batch variability. Second, the freezing process used to preserve primary hepatocytes and the transformation process needed to make stable and proliferating cell lines results in changes in cell differentiation, proliferation, and metabolic processes.

INDIGO's in vitro toxicology platform was developed to meet the increasing demand for more predictive models for liver toxicity due to the fact that hepatotoxicity remains a major reason for drug withdrawal from pharmaceutical development and clinical use. Consequently, the use of hepatocytes for the early identification of drug candidates that induce acute hepatotoxicity provides a powerful predictive tool that can inform drug development decisions.

Application of this platform is to screen chemicals for induction and activity of hepatic drug metabolizing enzymes (DME), which have been linked to Non-Alcoholic Fatty Liver Disease, Steatosis, Cholestasis and other conditions.

The platform now utilizes upcyte^® human liver cells to assess chemical and drug-induced toxicity.

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Expression Profiling of Clinically Relevant CYPs

Cytochrome p450 (CYP) enzymes are responsible for the Phase I metabolism of most drugs. And, it is noteworthy that the expression of cytochrome p450 (CYP) genes are predominantly regulated by ligand-activated receptors/transcription factors such as pregnane X receptor (PXR, NR1I2), constitutive androstane receptor (CAR, NR1I3), aryl hydrocarbon receptor (AhR), farnesoid x receptor (FXR, NR1H4), glucocorticoid receptor (GR, NR3C1), and to lesser degrees liver X receptors (LXR, NR1C3), vitamin D receptor (VDR, NR1I1), and peroxisome proliferator-activated receptor alpha (PPARα, NR1C1). Consequently, drugs that activate any of these xenobiotic sensing receptors can dramatically change the endogenous levels of CYP expression in the liver, potentially impacting the rate of their own metabolism, as well as the metabolism of all other co-administered drugs. Of particular concern are metabolic outcomes that transform a drug to greater potency, or to an altered bioactivity.

Assessing drug-induced changes in the expression of CYP genes provides a reliable predictive indicator of altered (either heightened or inhibited) metabolic activities leading to drug-drug interactions in vivo. Cytochrome p450 enzymes with the greatest clinical relevance belong to CYP1, 2, and 3 families. It is estimated that they are involved in the metabolism of 70-80% of drugs currently on the market. For example, CYP3A4 is estimated to be involved in ~30% of all prescription drugs.

INDIGO's assay kit for the Expression Profiling of Clinically Relevant CYPs contains optimized reagents for the culturing and treatment of upcyte^® hepatocytes to assess drug-induced changes in seven clinically-relevant CYPs: CYP3A4. CYP1A1, CYP2B6, CYP2C8, CYP2C9, CYP2C19, and CYP2E1.

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What is MDR1?

Human P-glycoprotein (P-gp) is also known as MDR1 or ABCB1, the human multidrug resistance protein 1. It is a 170kDa transmembrane glycoprotein that functions as an ATP-dependent efflux transporter present in the human body’s epithelial tissues.

P-gp consists of twelve membrane-spanning domains and two cytosolic nucleotide-binding domains. The transmembrane domains comprise several substrate binding pockets capable of interacting with a broad range of both endogenous and foreign small molecule chemotypes. Xenobiotic substrates of P-gp range from pollutants, such as those encountered through unintended exposure to industrial and agricultural chemicals, to small molecule drugs that are intentionally administered for therapeutic benefit.

P-gp is highly expressed in gastrointestinal epithelium, liver, pancreatic and kidney cells, and capillary endothelial cells that establish the blood-brain barrier. Multidrug resistance research has found MDR1 expressed at high levels on the lines of transformed and tumor cells. In combination with the activities of Cytochrome P450 oxidases, the robust efflux activity of P-gp plays a critical role in limiting the absorption and systemic physiological distribution of xenobiotics and facilitating their ultimate elimination from the body. The MDR1 transporter role is important for drug-drug interaction (DDI) and drug safety assessments.

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