Please use this identifier to cite or link to this item: http://prr.hec.gov.pk/jspui/handle/123456789/2514
Title: Studies on the use of HBV epitope Chimeras
Authors: Malik, Imran Riaz
Keywords: Natural Sciences
Biology
Genetics & evolution
Biotechnology
Issue Date: 2014
Publisher: Quaid-i-Azam University, Islamabad, Pakistan
Abstract: Hepatitis B virus (HBV) is a common cause of viral hepatitis with significant health complications including cirrhosis and hepatocellular carcinoma. Major advances have been made in the treatment of Hepatitis B recent years. However, much remains to be accomplished because current antiviral drugs do not eradicate infection. The currently licensed hepatitis B vaccines, consisting of recombinant hepatitis B surface antigen (HBsAg) and alum, are highly effective and induce protective antibody titers in 95% of vaccinated individuals after 3 immunizations before onset of HBV infection. Present studies were aimed to develop chimeric vaccine (proteins) that may stimulate B- and T- cell responses and generate enhance immune response to treat the HBV patients after infection. In this regard, five HBV chimeric vaccine constructs were designed and developed using the HBcAg gene encoding amino acids 1-78 and amino acids 80-144. Amino acid- position 79 where deleted and replaced with preS epitopes sequences. All five chimeric constructs (pIJMcsc-1, pIJMcsc-2, pIJMcsc-3. pIJMcsc-4 and pIJMcsc-5) contained HBcAg as a carrier molecule for the epitopes of preS regions primed HBcAg- specific antibodies response. These constructs (chimeric plasmids) were transformed into E.coli to see the protein product on SDS-PAGE. The expressed proteins (csc-1, csc-2, csc-3. csc-4 and csc-5) were purified based on affinity chromatography using His tag at the C terminal of expressed proteins. The purified chimeric proteins were further subjected to Enzyme linked Immunosorbent assay (ELISA) and western blotting assays using secondary anti- rabbit AP conjugated antibody and secondary anti-mouse AP-conjugated antibody respectively. All five chimeric proteins showed positive antibody response. In vivo study was performed for chimeric protein (csc-5) only. This chimeric protein (csc-5) was evaluated in C57BL/6J mice for activation of humoral and cellular immune responses, primed both HBcAg- specific T cells and antibodies to preS1. IXResult obtained indicated that the csc-5 protein can induce HBV-specific antibodies and T cells, two functions that well complement the activity of antiviral compounds. Thus, the csc-5 protein may be a future component in therapy for chronic HBV infections where the host is gaining a sustained control of the HBV infection and also a viable approach to develop an effective bi-functional therapeutic vaccine as an add-on for treatment of chronic HBV.
URI:  http://prr.hec.gov.pk/jspui/handle/123456789//2514
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