Ignite Creation Date:
2024-05-05 @ 11:53 AM
Last Modification Date:
2024-10-26 @ 9:15 AM
Study NCT ID:
NCT00164073
Status:
UNKNOWN
Last Update Posted:
2007-08-01
First Post:
2005-09-13
Brief Title:
Pharmacogenomics of Interferon and Ribavirin Treatment in Patients With Chronic Hepatitis C Virus Infection
Sponsor:
Bayside Health
Organization:
Bayside Health
Study Overview
Official Title:
Pharmacogenomics of Interferon and Ribavirin Treatment in Patients With Chronic Hepatitis C Virus Infection
Status:
UNKNOWN
Status Verified Date:
2005-09
Last Known Status:
RECRUITING
Delayed Posting:
No
If Stopped, Why?:
Not Stopped
Has Expanded Access:
False
If Expanded Access, NCT#:
N/A
Has Expanded Access, NCT# Status:
N/A
Acronym:
None
Brief Summary:
The purpose of this study is to examine gene expression profiles by DNA microarray in patients who are responders and non-responders to interferon and ribavirin treatment for hepatitis C virus HCV Genes involved in inflammation and fibrosis and mediators of the Th-1 lymphocyte response will be looked for It is hoped that genetic targets for future more effective and less toxic treatments will be identified
Detailed Description:
Background and Research Plan
Chronic HCV infection often follows a progressive course over many years and can ultimately result in cirrhosis and the need for liver transplantation or hepatocellular carcinoma The decision to treat patients with chronic hepatitis C infection is based upon several factors including the natural history and stage of the disease and the efficacy and adverse effects related to therapy As a general rule patients who are considered for treatment should have histologic and virologic evidence of chronic infection ie HCV RNA detectable in serum and an elevated serum ALT 1 Combination treatment with Interferon and Ribavirin is currently the best available treatment for chronic HCV infection However response to therapy is suboptimal and there are several side effects
Side effects seen during therapy of patients with Interferon and Ribavirin include flu-like symptoms mostly due to interferon cytopenias and haemolysis induced by Ribavirin Psychiatric changes due to interferon occur in up to 50 and thyroid abnormalities requiring therapy occur in about 1 to 5 treated with interferon Autoimmune disease and retinal haemorrhages have also been reported In addition combination therapy is associated with significant risks in patients with renal dysfunction cardiac disease and haemolytic anaemias as treatment may be associated with anaemia Patients who are depressed or suicidal should also be excluded 2
To this point little DNA microarray work done has been done with HCV However what work has been done is interesting and shows promise In studies comparing Hepatitis B HBV with Hepatitis C HBV infection was found to express genes predominantly involved in inflammation In contrast with HCV infection anti-inflammatory genes were found to be expressed 3 In another study HCV associated cirrhosis has been shown to be characterized by proinflammatory pro-fibrotic and proapoptotic gene expression profiles 4
The progression of HCV infection by gene expression analysis of liver biopsies in acutely infected chimpanzees that develop persistent infection transient viral clearance or sustained clearance has been examined Transient and sustained viral clearance were uniquely associated with induction of IFN-gamma-induced genes and other genes involved in antigen processing and presentation and the adaptive immune response The study revealed genome-wide transcriptional changes that reflect the establishment spread and control of infection and they reveal potentially unique antiviral programs associated with clearance of HCV infection 5
In-vitro studies in the human cell line Huh7 has shown that on exposure of the cells to Interferon 50 genes were up-regulated by at least twofold in control clones whereas induction of 9 of the 50 genes was significantly reduced in those Huh7 clones expressing NS5A Interferon-alpha activity has been found to be inhibited by the HCV protein NS5A The strongest effect of NS5A on Interferon response was observed on the OAS-p69 gene with reduced expression In addition Huh7 cells expressing NS5A showed an up-regulation of interleukin-8 a proinflammatory cytokine As a result this study postulated a mechanism for NS5A mediated interferon resistance 6
In a recent study microarray gene profiling of peripheral blood mononuclear cells from hepatitis C patients treated with Interferon-alpha was performed 88 genes directly relating to functions of immune cells were up-regulated including genes involved in antigen processing and presentation T-cell activation lymphocyte trafficking and effector functions suggesting that Interferon-alpha up-regulates multiple genes involving different aspects of immune responses to enhance immunity against hepatitis C virus 7
Currently treatment for HCV still has a significant non-responder rate and is associated with side effects which limits many individuals from receiving treatment Already molecular profiling by DNA microarrays has identified potential genetic targets which may have a role in the pathogenesis of HCV-associated hepatitis and more importantly possible targets which have a role in response to Interferon treatment The work proposed here hopes to build on this body of knowledge and may lead to improved treatment modalities for HCV
MethodsExperimental Design
Approval from the Alfred Hospital Ethics Committee will be obtained and subjects will be recruited via the Alfred Hospitals Hepatitis Clinics Informed consent for the study will be obtained after its purpose is explained verbally and via a plain English statement Informed consent will also include consent for liver biopsies Initially a pilot study will be undertaken in which only one subject will be recruited to both the experimental and control arm of the study Once the results from this pilot study have been analysed the study will be expanded to encompass a greater number of individuals
Liver biopsies will be performed on the following groups
1 HCV positive patients prior to commencement and at the end of treatment with either standard or pegylated Interferon plus Ribavirin This group will be subdivided into responders and non-responders to treatment Core liver biopsies under ultra-sound guidance are already routinely performed on this group prior to the commencement of treatment For the purpose of this study non-responders to combination therapy are those individuals who remain HCV-RNA positive at the end of treatment
2 Control population consisting of patients undergoing laparotomy for diseases not involving the liver These biopsies would be obtained at the time of surgery
Enough liver tissue approximately 1-2 mg will be obtained to isolate messenger RNA mRNA by oligo dT chromatography Once isolated the mRNA will be reverse transcribed to complementary DNA cDNA This cDNA will then be labeled with a fluorescent reporter molecule Hybridization of the labeled liver cDNA will then occur with the DNA microarray It is envisaged that the microarray will be commercially obtained and consist of human cDNA There is enough DNA on the microarray that two lots of labeled liver cDNA with different fluorescent markers ie red and green can be hybridized to the array at the same time without any interference The following hybridizations would then be set up
Reporter molecule 1 HCV ve group cDNA Reporter molecule 2 HCV -ve group cDNA
Hybridization 1 Responder to Rx Before Rx Control cDNA
Hybridization 2 Responder to Rx After Rx Control cDNA
Hybridization 3 Non responder to Rx Before Rx Control cDNA
Hybridization 4 Non responder to Rx After Rx Control cDNA
Rx treatment
The hybridized array will then be scanned and commercially available software packages will be used to help in the interpretation of the results 8 Genes or groups of genes of particular interest include
1 Pro-inflammatory response genes
Th-1 cytokines such as Gamma Interferon IFN Interleukin-2 IL-2 and Tumour Necrosis Factor-beta TNF-ß Th1 cytokines promote production of opsonizing antibodies eg IgG1 and induction of cellular cytotoxicity and macrophage activation Th1 responses are prominent in the defence against pathogens which replicate intracellularly
Tumour Necrosis Factor TNF and TNF receptor family of molecules which are strongly pro-inflammatory and involved in immune regulation
Interleukin-12 IL-12 which is a key cytokine that protects the host from viral and microbial infection It links the innate and acquired arms of the immune system
Interleukin-18 IL-18 which has been shown to act as a chemoattractant
2 Genes involved in a pro-fibrotic response
Transforming growth factor beta TGFbeta which causes fibroblast proliferation
Procollagen I and II
Platelet-derived growth factor PDGF and PDGF beta receptor over-expression have been linked to development of fibrotic disease as well as cancer and atherosclerosis
Matrix metalloproteinases play a pivotal role in angiogenesis and have a role in tumorigenesis
Fibroblast Growth Factor
Vascular Endothelial Growth Factor VEGF which is known for its angiogenic properties
Connective tissue growth factor which stimulates collagen production
In addition novel genes previously unidentified with differential expression will be looked for and will be characterized
References relevant to this project from literature search
1 Chopra S Treatment of chronic hepatitis C infection Recommendations In Up To Date Rose BREd Up To Date Wellesley MA 2002
2 Bonis PLA Chopra S Administration of combined interferon alfa-2b and ribavirin in the treatment of hepatitis C infection In Up To Date Rose BREd Up To Date Wellesley MA 2002
3 Honda M Kaneko S Kawai H et al Differential gene expression between chronic hepatitis B and C hepatic lesions Gastroenterology 2001 120 955-66
4 Shackel NA McGuinness PH Abbott CA et al Insights into the pathobiology of hepatitis C virus-associated cirrhosis analysis of intrahepatic differential gene expression Am J Pathol 2002 160 641-54
5 Su AI Pezacki JP Wodicka L et al Genomic analysis of the host response to hepatitis C virus infection Proc Natl Acad Sci 20029915669-74
6 Girard S Shalhoub P Lescure P et al An altered cellular response to interferon and up-regulation of interleukin-8 induced by the hepatitis C viral protein NS5A uncovered by microarray analysis Virology 2002 295 272-83
7 Ji X Cheung R Cooper S et alInterferon alfa regulated gene expression in patients initiating interferon treatment for chronic hepatitis C Hepatology 200337610-21
8 Jenkins RE Pennington SR arrays for protein expression profiling towards a viable alternative to two-dimensional gel electrophoresis Proteomics 2001 1 12-29
Chronic HCV infection often follows a progressive course over many years and can ultimately result in cirrhosis and the need for liver transplantation or hepatocellular carcinoma The decision to treat patients with chronic hepatitis C infection is based upon several factors including the natural history and stage of the disease and the efficacy and adverse effects related to therapy As a general rule patients who are considered for treatment should have histologic and virologic evidence of chronic infection ie HCV RNA detectable in serum and an elevated serum ALT 1 Combination treatment with Interferon and Ribavirin is currently the best available treatment for chronic HCV infection However response to therapy is suboptimal and there are several side effects
Side effects seen during therapy of patients with Interferon and Ribavirin include flu-like symptoms mostly due to interferon cytopenias and haemolysis induced by Ribavirin Psychiatric changes due to interferon occur in up to 50 and thyroid abnormalities requiring therapy occur in about 1 to 5 treated with interferon Autoimmune disease and retinal haemorrhages have also been reported In addition combination therapy is associated with significant risks in patients with renal dysfunction cardiac disease and haemolytic anaemias as treatment may be associated with anaemia Patients who are depressed or suicidal should also be excluded 2
To this point little DNA microarray work done has been done with HCV However what work has been done is interesting and shows promise In studies comparing Hepatitis B HBV with Hepatitis C HBV infection was found to express genes predominantly involved in inflammation In contrast with HCV infection anti-inflammatory genes were found to be expressed 3 In another study HCV associated cirrhosis has been shown to be characterized by proinflammatory pro-fibrotic and proapoptotic gene expression profiles 4
The progression of HCV infection by gene expression analysis of liver biopsies in acutely infected chimpanzees that develop persistent infection transient viral clearance or sustained clearance has been examined Transient and sustained viral clearance were uniquely associated with induction of IFN-gamma-induced genes and other genes involved in antigen processing and presentation and the adaptive immune response The study revealed genome-wide transcriptional changes that reflect the establishment spread and control of infection and they reveal potentially unique antiviral programs associated with clearance of HCV infection 5
In-vitro studies in the human cell line Huh7 has shown that on exposure of the cells to Interferon 50 genes were up-regulated by at least twofold in control clones whereas induction of 9 of the 50 genes was significantly reduced in those Huh7 clones expressing NS5A Interferon-alpha activity has been found to be inhibited by the HCV protein NS5A The strongest effect of NS5A on Interferon response was observed on the OAS-p69 gene with reduced expression In addition Huh7 cells expressing NS5A showed an up-regulation of interleukin-8 a proinflammatory cytokine As a result this study postulated a mechanism for NS5A mediated interferon resistance 6
In a recent study microarray gene profiling of peripheral blood mononuclear cells from hepatitis C patients treated with Interferon-alpha was performed 88 genes directly relating to functions of immune cells were up-regulated including genes involved in antigen processing and presentation T-cell activation lymphocyte trafficking and effector functions suggesting that Interferon-alpha up-regulates multiple genes involving different aspects of immune responses to enhance immunity against hepatitis C virus 7
Currently treatment for HCV still has a significant non-responder rate and is associated with side effects which limits many individuals from receiving treatment Already molecular profiling by DNA microarrays has identified potential genetic targets which may have a role in the pathogenesis of HCV-associated hepatitis and more importantly possible targets which have a role in response to Interferon treatment The work proposed here hopes to build on this body of knowledge and may lead to improved treatment modalities for HCV
MethodsExperimental Design
Approval from the Alfred Hospital Ethics Committee will be obtained and subjects will be recruited via the Alfred Hospitals Hepatitis Clinics Informed consent for the study will be obtained after its purpose is explained verbally and via a plain English statement Informed consent will also include consent for liver biopsies Initially a pilot study will be undertaken in which only one subject will be recruited to both the experimental and control arm of the study Once the results from this pilot study have been analysed the study will be expanded to encompass a greater number of individuals
Liver biopsies will be performed on the following groups
1 HCV positive patients prior to commencement and at the end of treatment with either standard or pegylated Interferon plus Ribavirin This group will be subdivided into responders and non-responders to treatment Core liver biopsies under ultra-sound guidance are already routinely performed on this group prior to the commencement of treatment For the purpose of this study non-responders to combination therapy are those individuals who remain HCV-RNA positive at the end of treatment
2 Control population consisting of patients undergoing laparotomy for diseases not involving the liver These biopsies would be obtained at the time of surgery
Enough liver tissue approximately 1-2 mg will be obtained to isolate messenger RNA mRNA by oligo dT chromatography Once isolated the mRNA will be reverse transcribed to complementary DNA cDNA This cDNA will then be labeled with a fluorescent reporter molecule Hybridization of the labeled liver cDNA will then occur with the DNA microarray It is envisaged that the microarray will be commercially obtained and consist of human cDNA There is enough DNA on the microarray that two lots of labeled liver cDNA with different fluorescent markers ie red and green can be hybridized to the array at the same time without any interference The following hybridizations would then be set up
Reporter molecule 1 HCV ve group cDNA Reporter molecule 2 HCV -ve group cDNA
Hybridization 1 Responder to Rx Before Rx Control cDNA
Hybridization 2 Responder to Rx After Rx Control cDNA
Hybridization 3 Non responder to Rx Before Rx Control cDNA
Hybridization 4 Non responder to Rx After Rx Control cDNA
Rx treatment
The hybridized array will then be scanned and commercially available software packages will be used to help in the interpretation of the results 8 Genes or groups of genes of particular interest include
1 Pro-inflammatory response genes
Th-1 cytokines such as Gamma Interferon IFN Interleukin-2 IL-2 and Tumour Necrosis Factor-beta TNF-ß Th1 cytokines promote production of opsonizing antibodies eg IgG1 and induction of cellular cytotoxicity and macrophage activation Th1 responses are prominent in the defence against pathogens which replicate intracellularly
Tumour Necrosis Factor TNF and TNF receptor family of molecules which are strongly pro-inflammatory and involved in immune regulation
Interleukin-12 IL-12 which is a key cytokine that protects the host from viral and microbial infection It links the innate and acquired arms of the immune system
Interleukin-18 IL-18 which has been shown to act as a chemoattractant
2 Genes involved in a pro-fibrotic response
Transforming growth factor beta TGFbeta which causes fibroblast proliferation
Procollagen I and II
Platelet-derived growth factor PDGF and PDGF beta receptor over-expression have been linked to development of fibrotic disease as well as cancer and atherosclerosis
Matrix metalloproteinases play a pivotal role in angiogenesis and have a role in tumorigenesis
Fibroblast Growth Factor
Vascular Endothelial Growth Factor VEGF which is known for its angiogenic properties
Connective tissue growth factor which stimulates collagen production
In addition novel genes previously unidentified with differential expression will be looked for and will be characterized
References relevant to this project from literature search
1 Chopra S Treatment of chronic hepatitis C infection Recommendations In Up To Date Rose BREd Up To Date Wellesley MA 2002
2 Bonis PLA Chopra S Administration of combined interferon alfa-2b and ribavirin in the treatment of hepatitis C infection In Up To Date Rose BREd Up To Date Wellesley MA 2002
3 Honda M Kaneko S Kawai H et al Differential gene expression between chronic hepatitis B and C hepatic lesions Gastroenterology 2001 120 955-66
4 Shackel NA McGuinness PH Abbott CA et al Insights into the pathobiology of hepatitis C virus-associated cirrhosis analysis of intrahepatic differential gene expression Am J Pathol 2002 160 641-54
5 Su AI Pezacki JP Wodicka L et al Genomic analysis of the host response to hepatitis C virus infection Proc Natl Acad Sci 20029915669-74
6 Girard S Shalhoub P Lescure P et al An altered cellular response to interferon and up-regulation of interleukin-8 induced by the hepatitis C viral protein NS5A uncovered by microarray analysis Virology 2002 295 272-83
7 Ji X Cheung R Cooper S et alInterferon alfa regulated gene expression in patients initiating interferon treatment for chronic hepatitis C Hepatology 200337610-21
8 Jenkins RE Pennington SR arrays for protein expression profiling towards a viable alternative to two-dimensional gel electrophoresis Proteomics 2001 1 12-29
Study Oversight
Has Oversight DMC:
None
Is a FDA Regulated Drug?:
None
Is a FDA Regulated Device?:
None
Is an Unapproved Device?:
None
Is a PPSD?:
None
Is a US Export?:
None
Is an FDA AA801 Violation?:
None