Microarray analysis of leukocyte gene expression in hepatitis C patients treated with interferon-a

Xuhuai Ji¹, Xiaosong He¹, Ramsey Cheung¹, Stewart Cooper², Pat Brown¹,

Harry B. Greenberg^1,3

¹Stanford University School of Medicine, ²UCSF School of medicine, ³Aviron Inc.

Background. Interferon-a (IFN) is the only FDA-approved treatment for chronic hepatitis C. The treatment has a sustained response rate of 20-50%, depending on genotype of HCV. The mechanisms for the antiviral effects of IFN are believed to be: (1) the IFN-induced changes in hepatocyte gene expression that may directly suppress HCV replication; and (2) the modulation of functions of immune cells, especially lymphocytes and antigen presenting cells that play important roles in antiviral immunity. The most direct method of monitoring the effects of IFN on these host cells is to examine the gene expression changes in cells obtained from patients under IFN treatment. The hypothesis to be tested in this study is the outcome of IFN therapy is associated with different response patterns to IFN in the global gene expression of the treated patient.

Methods. We are carrying out microarray studies with two strategies: (1) in vitro strategy. Blood samples from chronic hepatitis C patients to be treated with IFN are incubated with or without IFN in vitro for several hours; (2) In vivo strategy. Blood samples are drawn from patients right before they receive the first dose of IFN injection, as well as several hours after the injection. RNAs are prepared from the blood samples, followed by reverse transcription to synthesize cDNA and in vitro transcription to amplify mRNA. The amplified antisense RNA samples are labeled with Cy3 and Cy5 and hybridized to cDNA microarrays. The outcome of treatment in each patient will be associated with the microarray analysis when the treatment is finished in 6 or 12 months.

Results. We have performed in vitro and in vivo studies in 8 patients starting IFN therapy and more patients are being enrolled. In all patients tested, a series of previously identified IFN-inducable genes were significantly upregulated in both in vitro and in vivo samples after IFN treatment. Many other genes, including genes involving in antigen presentation, lymphocyte activation and proliferation, T cell function regulation, apoptosis and signal transduction, were also upregulated after IFN treatment. Detailed analysis of the data is underway and will be presented.

Conclusion. Microarray analysis can be used to study the in vivo effect of IFN on the global gene expression in leucocytes of people. This study may lead to insights into the mechanisms of the antiviral effect of IFN, especially those mediated by the immune system, as well as the underlining factors affecting the outcome of treatment in patients with chronic HCV infection..