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Immunogenicity of a PCR-based humanized HIV envelope DNA vaccine in mice

หน่วยงาน จุฬาลงกรณ์มหาวิทยาลัย

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ชื่อเรื่อง : Immunogenicity of a PCR-based humanized HIV envelope DNA vaccine in mice
นักวิจัย : Sunee Sirivichayakul
คำค้น : AIDS (Disease) , Vaccines
หน่วยงาน : จุฬาลงกรณ์มหาวิทยาลัย
ผู้ร่วมงาน : Praphan Phanuphak , Thaweesak Tirawatnapong , Chulalongkorn University. Graduate School
ปีพิมพ์ : 2546
อ้างอิง : 9741734301 , http://cuir.car.chula.ac.th/handle/123456789/5462
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Thesis (Ph.D.)--Chulalongkorn University, 2003

Human immunodeficiency virus type 1 (HIV-1) infection is a major public health problem causing epidemic worldwide. It infects mainly CD4+T lymphocyte and destroys the cells resulting in progressive immunodeficiency. Although many potent antiretroviral drugs acting at different stages of HIV life cycle are currently available, the majority of patients worldwide will not have easy access to these expensive and complicated therapeutic regimens. Protection including prophylactic vaccination represents one of the best and sustainable solutions to curb the worldwide epidemic. Many attempts to produce an effective HIV vaccine have been made without much success up to now. To explore more about HIV vaccine, three HIV-1 DNA constructs, namely 297-bp humanized DNA encompassing the immunodominant epitopes of the V3 and its adjacent regions of multi-clade HIV-1 representing the last common ancestor consensus sequence of 8 HIV-1 subtypes circulating wordwide (297-bp hu V3 DNA), 297-bp non-humanized DNA and 2.5 kb full-length envelope DNA were studied. The 297-bp hu DNA was produced by a PCR-based method while the rests were directly amplified from DNA isolated from peripheral blood mononuclear cells (PBMC) of HIV-1 subtype A/E-infected individual. All 3 PCR products were successfully cloned into appropriate plasmid vectors to be used as HIV DNA vaccines. The 297-bp multi-clade hu V3 DNA and the 2.5 kb full-length envelope DNA were well expressed in vitro by transfection experiments as 13 kd and as 160, 120 and 38 Kd respectively but not the 297-bp non-humanized DNA. However, all 3 constructs were shown to be immunogenic in mice by in vivo DTH skin testing (footpad swelling) with specific V3 peptide. The in vitro correlates of the immunogenicity study was best demonstrate by Enzyme Linked Immunospot Assay (ELISPOT). Intracellular cytokine staining (ICCS) response could be well demonstrated only in 2.5 kb full-length envelope DNA primed/recombinant gp120 (rgp120) boosted group. Both the ELISPOT and ICCS responses were relatively weak but positive. No antibodies of antigen-stimulated lymphoproliferative response could be shown in this study. The attempt to demonstrate cross-reactivity of the hu V3 DNA by boosting with recombinant vaccinia gp160(E) yielded negative results which may be due to the low immunogenicity of the constructs or the techniques used. In conclusion, we were able to produce DNA constructs including the humanized DNA of the envelope region to be tested as HIV-1 vaccine. Humanized DNA was better expressed than the non-humanized counterpart. The 3 DNA constructs were shown to have some immunogenicity in mice. Further improvements are needed to enhance their immunogenicity in order to allow for further pre-clinical and clinical testings.

บรรณานุกรม :
Sunee Sirivichayakul . (2546). Immunogenicity of a PCR-based humanized HIV envelope DNA vaccine in mice.
    กรุงเทพมหานคร : จุฬาลงกรณ์มหาวิทยาลัย.
Sunee Sirivichayakul . 2546. "Immunogenicity of a PCR-based humanized HIV envelope DNA vaccine in mice".
    กรุงเทพมหานคร : จุฬาลงกรณ์มหาวิทยาลัย.
Sunee Sirivichayakul . "Immunogenicity of a PCR-based humanized HIV envelope DNA vaccine in mice."
    กรุงเทพมหานคร : จุฬาลงกรณ์มหาวิทยาลัย, 2546. Print.
Sunee Sirivichayakul . Immunogenicity of a PCR-based humanized HIV envelope DNA vaccine in mice. กรุงเทพมหานคร : จุฬาลงกรณ์มหาวิทยาลัย; 2546.