Presentation on theme: "Neutralisation with Antisera The neutralisation sensitivity of B-clade primary isolates (3 clones each), isolated at baseline (table on the left) and at."— Presentation transcript:
Neutralisation with Antisera The neutralisation sensitivity of B-clade primary isolates (3 clones each), isolated at baseline (table on the left) and at week 52 (table on the right), by autologous and heterologous antisera is shown. Primary isolates isolated at baseline are readily neutralisable by their week 52 autologous sera (reciprocal dilution of 50 or greater to achieve IC 90 ). However, primary isolates isolated at week 52 are resistant to neutralisation by their week 52 antisera, suggesting that the viruses developed escape mechanism rapidly. The neutralisation sensitivity of a standard panel of C-clade isolates against the C-clade antisera taken from SPARTAC patients at week 52 is shown. The Zambian isolates (5 out of 6) are neutralised poorly by the antisera (minimum reciprocal dilution of 50 to achieve IC 50 ), while the South African isolates are more sensitive to neutralisation by antisera which are taken from South African patients. We would like to thank the Wellcome Trust for funding SPARTAC and its sub-studies. All the NAbs are obtained from the Centre for AIDS Reagents of NIBSC. The reference panel of C-clade virus is obtained from the AIDS Research and Reference Program of the NIH. Reference Kaufmann GR, Zaunders JJ, Cunningham P, Kelleher AD, Grey P, Smith D, Carr A, Cooper DA. 2000. Rapid restoration of CD4 T cell subsets in subjects receiving antiretroviral therapy during primary HIV-1 infection. AIDS 14:2643-51. Fidler S, Oxenius A, Brady M, Clarke J, Cropley I, Babiker A, Zhang HT, Price D, Phillips R, Weber J. 2002. Virological and immunological effects of short-course antiretroviral therapy in primary HIV infection. AIDS 16:2049-54. Acknowledgement Conclusion The B-clade primary isolates from week 52 are equally or more neutralisable by 2F5 and b12, but more resistant to 2G12, than their baseline counterparts, suggesting that NAbs targeting epitopes on the glycans may be more effective in neutralising viruses in vivo than those targeting gp120 or gp41. We have identified a primary isolate (from patient F) with env sequence highly similar to a laboratory-adapted strain, which is neutralised well by heterologous antisera. Potential use as an immunogen? The antisera taken from C-clade patients can only neutralise viruses isolated from the same geographic location. There is a direct correlation between baseline viral load and env variation. Is high initial viral load a driving force behind viral diversity? Patients This study selected 18 patients from the SPARTAC cohort. Six are UK patients with B-clade virus and 12 of the patients are from S.A. with C- clade virus. Patient M from the U.K. has X4-tropic virus and all others are R5-tropic. Abstract Background: The Short Pulse Antiretroviral Therapy at Seroconversion (SPARTAC) clinical trial recruited 371 patients with acute HIV-1 infection from centres world-wide. The patients were randomised to one of three arms: a short course of antiretroviral therapy (ART) for 12 weeks, or a long course of ART for 48 weeks, or no therapy. This investigation aims to examine the development neutralising antibody (NAb) responses in association with viral diversity and evolution. Methods: A panel of 18 patients (6 with clade-B and 12 with clade-C virus) were selected blind for detailed study. Full-length HIV-1 envelope (env) cassettes, constructed from all patients from samples taken within six months of seroconversion and one year later, were co-transfected with a laboratory-strain (HXB2) backbone into 293T cells to produce pseudovirions. Neutralisation of pseudovirions by Nabs: b12, 2F5, 2G12 or heterologous and autologous sera was assayed. Clonal sequencing of the env V1-V3 regions was carried-out and viral diversity and evolution analysed in relation to baseline viral load were analysed. Result: The clade-B viruses taken at week 52 were equally or more readily neutralised by b12 and 2F5, but were more resistant to 2G12 neutralisation compared to their baseline counterparts. While the baseline antisera were poorly neutralising, antisera taken at week 52 neutralised baseline isolates (1/125) and, to a lesser extent (1/50), isolates from week 52. Figures in parenthesis represent reciprocal dilutions of sera effecting 90% neutralisation. Viral load did not correlate with neutralisation by NAbs. However, antisera from patients with low viral load neutralised early viruses. A direct correlation between viral load and viral variation was found. Conclusion: This study confirms that virus isolated at the time of acute infection is resistant to neutralisation and appeared to have developed early escape mechanisms. Moreover, high viral load at seroconversion may enhance viral diversification and contribute to the weak neutralising responses. Evolutionary History of the Primary Isolates Left: Phylogenetic tree of env isolated from B-clade primary isolates. Except for patient H, the env gene for all other patients have evolved little over 52 weeks. Of note, patient F has env sequence highly similar to laboratory- adapted strains HXB2 and NL.671. R 2 = 0.992 Right: A preliminary phylogenetic tree of env isolated from C-clade primary isolates. Left: For B-clade isolates, intra- patient env variation at baseline directly correlates with its corresponding viral load. Right: Except for patient H (green dot), there is direct correlation between baseline viral load and evolutionary distance of env over 52 weeks. Neutralisation of B-clade isolates with NAbs Pseudoviruses containing primary env, isolated from B- clade patients at baseline and at week 52, are subjected to neutralisation by 50μg/ml of 2F5 (top left), b12 (top right) and 2G12 (bottom). Three env clones were assayed per patient and the average % residual infectivity is shown. Evolution of HIV-1 and Concomitant Antibody Responses in HIV-1 Seroconverters Rachel P.J. Lai, David Bonsall, Anna Helander, Jonathan N. Weber, Myra O. McClure and SPARTAC Trial Steering Committee Section of Infectious Diseases, Wright Fleming Institute, Imperial College London, St Mary’s Hospital, London W2 1PG, United Kingdom PatientCountrySubtype Co-receptor Usage Baseline Viral Load (copies/ml) C United Kingdom B R574,321 FR5421,500 HR554,780 JR5315,764 MX444,100 XR5267,362 1 South AfricaC R598,100 2R5315,000 3R527,100 4R5122,000 5R5294,000 6R5494,000 7R5222,000 8R579,200 9R5104,000 10R5168,000 11R5275,000 12R5150,000 Baseline 1° Isolate (3 clones) Control (negative pooled sera) Baseline SeraWeek 52 Sera CFHJMXCFHJMX C6205020 5020<5050 C20205020 <2020125<5050 C2820<2020 <202012550 F420 50125<2050<20<50 F620 50<5050 <50 F8205020 <202050 <2050 <50 H7205020 <20 5020<5050 125 H9205020 5020<20 <50<2050 H2120<20205020502050<20<5050<2050 J33205020 502050<2050125<2050 J4220 502050 12550 J50205020 50<20 50 <12550<20 M2205020 502050<5050 12550 M14205020 502050 12550 M17205020 502050 <505012550 X10205020 5020502050 X2720<2020 502050 X2920<2020 <20 50 Week 52 virus (3 clones) Control (negative pooled sera) Baseline SeraWeek 52 Sera CFHJMXCFHJMX C1120 50205020<2050 C1620 50<2020<2050 C2120 <205020 F1720 <20 2012550 12550125 F2320 50 1252050 F3520 502012550 H1120 50<2020<20 H2420 50 20<20 H2520 50 J220 50 J1620 <2020<20 J4020 502050 M520<2020 <2020502050<2050 M920 <20205020<202050 M4320 5020<2050 X420 502050 <50 X1020 50 <5050 X1820 <2050 Wk 52 Serum Du 156.12 Du 172.17 Du 422.1 ZM197M.PB7 ZM214M.PL15 ZM233M.PB6 ZM249M.PL1 ZM53M. PB12 ZM109F. PB4 CAP45.2. 00.G3 CAP210. 2.00.E8 SA Zambia SA C1312.5781.25125 205020 312.5 C2781.25 125781.2520312.520 12550 C3781.25312.520312.5205020 125 C4781.25312.550781.2520312.5502050 C5312.5781.2550781.2550 20 5012550 C6781.2512550781.2550 312.5 C7781.2512520125<2020 502050 C8312.5 20312.5<2012520 50 C9312.5125201252012520 50 C101255020312.5<2020 <2020 C11312.5 20312.5<2020 12520 C12312.55020312.5205020 50
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