Our Assay-Ready Expanded Hepatocytes have functional CYP activity (
). The basal Phase I enzyme activities are comparable to HepaRG cells. The basal activity of CYP2D6 is low (<10 pmol/mL) in both ARE Hepatocytes and HepaRG cells. Since CYP2D6 is responsible for the metabolism of a significant number of clinically approved drugs, we also offer an isogenic line where CYP2D6 is overexpressed. Our CYP2D6 overexpressing hepatocytes have basal CYP2D6 activity of 1600 pmol/mL. This provides an isogenic hepatocyte cell line model of extensive/ultrarapid metabolizers and poor metabolizers for CYP2D6.
Figure 1. Comparison of the Phase I CYP enzyme activity between ARE Hepatocytes (Donor 4), ARE Hepatocytes (CYP2D6 Overexpressing; Donor 4) and HepaRG cells.
CYP Induction and Inhibition Data
Assay-Ready Expanded (ARE) Hepatocytes can be used in CYP induction and CYP inhibition studies. ARE Hepatocytes have been extensively characterized for CYP enzyme basal activity and fold induction (
Table 1 and Table 2
). Activity of the CYP enzymes was determined by the speed of metabolism of enzyme substrates. We have 4 donor cell lines available from both male and female donors. ARE Hepatocytes can also be used for CYP inhibition studies.
shows the inhibition of CYP enzyme activity in our expanded primary hepatocytes by selective compounds.
Table 1. ARE Hepatocyte CYP characterization data. C: Caucasian, H: Hispanic. CYP1A2 activity determined by phenacetin metabolism. CYP2B6 activity determined by bupropion metabolism. CYP2C9 activity determined by tolbutamide metabolism. CYP3A4 activity determined by testosterone metabolism.
Table 2. Comparison of CYP enzyme activity between ARE Hepatocytes and primary uncultured human hepatocytes
Figure 2. Inhibition of CYP1A2, CYP2B6, CYP2C9 and CYP3A4 enzyme activity of ARE Hepatocytes (Donor 4) by selective compounds.
Hepatic Transporter Data
Our Assay-Ready Expanded Hepatocytes express hepatic transporter genes that are expressed by primary hepatocytes (
). The mRNA expression level of the hepatic transporter genes is significantly higher compared to HepG2 cells. Our expanded hepatocytes can be used for compound uptake and metabolism studies.
Figure 3. Gene expression of the hepatic transporter genes NTCP, BSEP, OCT1, MDR1 and MRP3 in primary hepatocytes, Axol Assay-Ready Expanded Hepatocytes (Donor 3) and HepG2 cells.
ARE Hepatocytes can be used for hepatotoxicity studies and predictive screens of novel compounds.
shows the responsiveness of our expanded primary hepatocytes to known hepatotoxic compounds. Our expanded primary hepatocytes have high tolerance (IC50 > 200 µM) for non-toxic compounds such as phenytoin and ciprofloxacin. ARE Hepatocytes are responsive to moderate hepatic toxins such as methotrexate and are sensitive (IC50 < 30 µM) to hepatotoxins such as ketoconazole and tacrolimus.
Figure 4. Responsiveness of ARE Hepatocytes to hepatotoxic compounds.
Axol Genotoxicity Assay Validated ARE Hepatocytes (ax3703) can be used for micronucleus assays to determine potential genotoxic effects of novel compounds. The hepatocytes can be seeded at subconfluency and will proliferate in ARE Hepatocyte Genotoxicity Assay Medium (ax3715), permitting the effect of compounds that alter cell division and DNA replication to be observed. Genotoxicity Assay Validated ARE Hepatocytes are division-competent hepatocytes unlike uncultured primary hepatocytes and therefore have wider applicability and can be used for assays that require cell division such as micronucleus assays.
shows the effect of cyclophosphamide on ARE Hepatocyte cell viability and micronucleus formation. Cyclophosphamide is a DNA alkylating agent that interferes with DNA replication and is used for cancer chemotherapy.
Figure 5. Increasing cyclophosphamide concentration affects the percentage of cells with micronuclei (% MN) and cell viability of Axol Genotoxicity Assay Validated ARE Hepatocytes.