Designing and biological evaluation of single-cycle replicable HIV1 system as a potential vaccine strategy
Abstract number: P1199
Sadat S.M., Zabihollahi R., Vahabpour R., Azadmanesh K., Javadi F., Siadat S.D., Memarnejadian A., Parivar K., Khanahmad Shahreza H., Arabi Mianroodi R., Hekmat S., Aghasadeghi M.R.
Objectives: Infection with human immunodeficiency virus (HIV) is a spreading world health problem, so a vaccine is desperately needed to control the AIDS pandemic. Inactive HIV particles may represent suitable vaccine candidates as they have all the immunogenic structural and surface antigens in their native forms. Accordingly, several preclinical studies have already shown that inactive and attenuated HIV virions can elicit protective humoral and cellular immune responses. Herein, we designed and constructed a novel HIV-1 virion, capable of replicating in a single cycle that provides a more immunogenicity, while prevents any pathologic effects and further evaluated its biological properties.
Methods: The pmzNL43 plasmid, containing the complete HIV genome of NL43 strain with a 2-kb MlsI-digested fragment deletion in reverse transcriptase (RT) and integrase (IN) genes was constructed, confirmed by sequencing reactions and transfected into HEK 293T cell line. By further co-transfection of psPAX2 and pMD2.G plasmids, which encoded HIV Gag-pro-pol and vesicular stomatitis virus surface glycoprotein, into the same pmzNL43-harboring cells, pseudotyped virions were produced, evaluated by electron microscopy and quantified using P24 end-point ELISA assay. Infectivity of mzNL43 virions and their efficiency towards the syncytium formation was evaluated on HIV-sensitive MT-2 cells.
Results: Despite the occurrence of a designed long deletion within the RT and IN genes the production of HIV virions was indicated by the level of P24 protein in culture supernatant of transfected cells and was further confirmed by electron microscopy. Formation of syncytia in MT-2 cells also evidenced for the functionality of the surface glycoproteins in produced pseudotyped virions. Interestingly, infectivity analysis verified that the second generation virions were completely non-replicative.
Conclusion: While the introduced single cycle replicable (SCR) HIV system completely maintained the antigenic structures of HIV-1, by its one cycle replicating properties represented a good implication as a potential vaccine candidate and this guarantees the further investigations towards the assessment of its immunogenicity, which are currently under process. Also, replacement of the deleted fragment in pmzNL43 with various immunostimulatory sequences may present another interesting approach towards the improvement of its application in HIV vaccine researches.
|Session name:||Abstracts 20th European Congress of Clinical Microbiology and Infectious Diseases|
|Location:||Vienna, Austria, 10 - 13 April 2010|
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