Functional evolution of HIV-1 subtype C Envelope: Implications for disease progression and immune escape

Aktivitet: Tale eller præsentation - typerForedrag og mundtlige bidrag

Se relationer på Aarhus Universitet

Martin Roelsgaard Jakobsen - Foredragsholder

  • Infektionsmedicinsk Afdeling Q, SKS
Introduction: The HIV-1 envelope glycoproteins (Env) mediate viral entry into cells via the sequential interaction with CD4 and then a coreceptor, which in vivo is either CCR5 or CXCR4. Furthermore, Env determines cell tropism, causes significant cytotoxicity in infected cells, and is the primary target of HIV-1 neutralising antibodies. In HIV-1 subtype B (B-HIV), the evolution of Env to use both CCR5 and CXCR4 for entry in vivo (R5X4), and sometimes minor alternative coreceptors in vitro is associated with rapid disease progression. Comparatively little is known about Env determinants contributing to HIV-1 subtype C (C-HIV) disease progression, despite C-HIV being the predominant HIV-1 subtype worldwide. This study aims to elucidate changes in C-HIV Env function that contribute to pathogenicity, by establishing and characterising an extensive longitudinal panel of Envs derived from a well-characterised cohort of untreated subjects who experienced progressive C-HIV infection.
Methods: Plasma samples were obtained from the Mupfure Schistosomiasis and HIV cohort (MUSH), which is a longitudinal Zimbabwean cohort of 196 subjects followed for 3 years (1, 2). We selected 60 plasma samples from 20 patients who experienced disease progression toward AIDS. Specifically, we selected 2 samples from each patient spanning a chronically phase of infection with stable CD4+ T-cell count (>250 cells/l; t = 0 and 2 years) and a later sample after sustained CD4+ T-cell decline (<170 cells/l; t = 3 years). HIV-1 RNA was extracted from plasma, and the gp160 Env gene PCR-amplified and cloned into the pSVIII-Env expression plasmid. Functional Envs capable of mediating HIV-1 entry were identified using single round entry assays in the CD4/CCR5/CXCR4-expressing JC53 cell line with Env-pseudotyped GFP reporter viruses. Functional Envs were sequenced and subjected to phylogenetic analysis. The ability of cloned Envs to use CCR5, CXCR4 or the alternative coreceptors CCR2b, CCR3, CCR8, Gpr1, Gpr15, Strl33 or Apj for HIV-1 entry was determined using single round entry assays in Cf2th cells expressing CD4 and either coreceptor and Env-pseudotyped luciferase reporter viruses.
Results: The GFP reporter virus entry assays identified 5 functional Envs cloned from each plasma sample. Thus, the Env bank created is extensive, and consists of 300 fully-functional Envs sampled from subjects spanning chronic to late stages of infection. Preliminary sequence analysis on a subset of Envs confirmed subtype C infection with no evidence of inter-subtype recombinants. At present, 127 C-HIV Envs cloned from 12 subjects have been tested for the ability to use CCR5, CXCR4 or alternative coreceptors for HIV-1 entry. All Envs used CCR5 for HIV-1 entry. CXCR4-usage was evident in 4 subjects at baseline and in 2 additional subjects at follow-up. There was a relatively high frequency of alternative coreceptor usage, with CCR2b, CCR3, CCR8, Gpr1, Gpr15, Strl33 and/or Ajp able to mediate the entry of up to 18% of C-HIV Envs. The R5 Envs showed relatively limited use of alternative coreceptors, whereas R5X4 Envs had comparatively broader usage of alternative coreceptors.
Discussion: In contrast to previous studies on C-HIV, our interim data illustrates prominent CXCR4-usage, occurring at a rate which is similar to progressive B-HIV infection. The overall frequency of alternative coreceptor usage appears higher in C-HIV infection compared to B-HIV infection, but the significance of alternative coreceptors in mediated C-HIV infection in vivo is presently unclear. Our results suggest pathogenicity of C-HIV infection may involve adaptive changes in Env, but further studies are required to determine the nature of these changes. The analysis of the remaining ~170 Env clones is underway and we expect these results to be available at the time of presentation.
4 jun. 2009

Begivenhed (Konference)

Titel5<sup>th</sup> National Scientific Workshop
Dato04/06/200904/06/2009
ByTerrigal
LandAustralien

ID: 18430456