In vitro test system to evaluate xenobiotics as immune-modulators of drug transport and metabolism in human hepatocytes

1. An in vitro method of evaluating the effect of a xenobiotic on drug metabolism in hepatocytes, wherein said xenobiotic is a biologic drug, said method comprising:

providing a first culture of hepatocytes;

providing a portion of a xenobiotic-stimulated biological sample by:

providing a first culture of a biological sample;

exposing said first biological sample to said xenobiotic biologic drug for a period of time to yield said xenobiotic-stimulated biological sample, wherein said biologic drug is selected from the group consisting of hormones, growth factors, cytokines, receptors, antibodies, enzymes, oligonucleotides, aptomers, synthetic peptides, and vaccines; and

separating said portion of said xenobiotic-stimulated biological sample, wherein said portion is plasma or supernatant;

transferring said portion of said xenobiotic-stimulated biological sample to said first culture of hepatocytes; and

analyzing the activity, expression, or combination thereof of a biomarker in said hepatocytes to thereby evaluate the effect of said xenobiotic on said drug metabolism in said hepatocytes.

2. The method of claim 1 , wherein said exposing comprises incubating said first biological sample with said xenobiotic for at least about 12 hours prior to said separating.

3. The method of claim 1 , wherein said biological sample has been collected from a subject selected from the group consisting of humans and non-human mammals.

4. The method of claim 1 , wherein said biological sample is selected from the group consisting of whole blood, bone marrow, and isolated peripheral blood mononuclear cells.

5. The method of claim 1 , wherein said biological sample is whole blood and wherein said portion of xenobiotic-stimulated biological sample is xenobiotic-stimulated plasma.

6. The method of 5 , further comprising measuring at least a portion of said xenobiotic-stimulated plasma for cytokines after said separating.

7. The method of 6 , wherein said transferring comprises incubating a portion of said xenobiotic-stimulated plasma with said first culture of hepatocytes.

8. The method of claim 1 , wherein said portion of xenobiotic-stimulated biological sample comprises a plurality of cytokines.

9. The method of claim 1 , further comprising incubating said portion of xenobiotic-stimulated biological sample and said first culture of hepatocytes for at least about 24 hours prior to said analyzing.

10. The method of claim 1 , wherein said analyzing comprises incubating said hepatocytes with an enzyme marker substrate and measuring levels of said substrate or metabolites of the same.

11. The method of claim 1 , wherein said analyzing comprises isolating microsomes from said hepatocytes and measuring the activity of drug-metabolizing enzymes in said microsomes.

12. The method of claim 1 , wherein said analyzing comprises isolating total RNA from said hepatocytes and measuring enzyme mRNA levels of said drug-metabolizing enzymes.

13. The method of claim 1 , wherein said analyzing comprises incubating said hepatocytes with a small molecule drug and measuring metabolism of said drug.

14. The method of claim 1 , further comprising:

providing a second culture of hepatocytes;

exposing said second culture of hepatocytes directly to said xenobiotic; and

analyzing the activity, expression, or combination thereof of a biomarker in said second culture of hepatocytes.

15. The method of claim 14 , further comprising comparing said activity and/or expression of said biomarker in said second culture of hepatocytes to said activity and/or expression of said biomarker in said first culture of hepatocytes.

16. The method of claim 1 , wherein said xenobiotic is dispersed in a carrier prior to said exposing.

17. The method of claim 16 , further comprising:

providing a third culture of hepatocytes;

exposing said third culture of hepatocytes directly to said carrier; and

analyzing the activity, expression, or combination thereof of a biomarker in said third culture of hepatocytes.

18. The method of claim 17 , further comprising comparing said activity and/or expression of said biomarker in said third culture of hepatocytes to said activity and/or expression of said biomarker in said first culture of hepatocytes.

19. The method of claim 16 , further comprising:

providing a second culture of a biological sample and a fourth culture of hepatocytes;

exposing said second biological sample to said carrier for a period of time to yield a carrier-stimulated biological sample;

transferring a portion of said carrier-stimulated biological sample to said fourth culture of hepatocytes; and

analyzing the activity, expression, or combination thereof of a biomarker in said fourth culture of hepatocytes.

20. The method of claim 19 , further comprising comparing said activity and/or expression of said biomarker in said fourth culture of hepatocytes to said activity and/or expression of said biomarker in said first culture of hepatocytes.

21. The method of claim 1 , further comprising:

providing a third culture of a biological sample and a fifth culture of hepatocytes;

exposing said third biological sample to a positive control compound with known cytokine-stimulating effects for a period of time to yield a positive control-stimulated biological sample;

transferring a portion of said positive control-stimulated biological sample to said fifth culture of hepatocytes; and

analyzing the activity, expression, or combination thereof of a biomarker in said fifth culture of hepatocytes.

22. The method of claim 21 , further comprising comparing said activity and/or expression of said biomarker in said fifth culture of hepatocytes to said activity and/or expression of said biomarker in said first culture of hepatocytes.

23. The method of claim 1 , wherein said culture of hepatocytes is a co-culture of hepatocytes and Kupffer cells.

24. The method of claim 1 , wherein said hepatocytes are human hepatocytes.

25. The method of claim 1 , wherein said hepatocytes are pooled hepatocytes.