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No Evidence of HIV-1 Evolution in Circulating Reservoirs in Children during Long-Term Antiretroviral Therapy

This study investigates the dynamics of HIV-1 reservoirs in HIV-infected children receiving long-term antiretroviral therapy (ART). The results show no evidence of viral evolution in circulating reservoirs despite detectable levels of cell-associated HIV-RNA, suggesting the absence of ongoing viral replication.

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No Evidence of HIV-1 Evolution in Circulating Reservoirs in Children during Long-Term Antiretroviral Therapy

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  1. Abstract: WEAA0104 No evidence of HIV-1 evolution in circulating reservoirs in children during more than six years of early suppressive antiretroviral therapy despite detectable levels of cell associated-HIV-RNA • M. Moragas, MD. Golemba, M. Distefano, R. Bologna, A. Mangano • Hospital de Pediatría “Prof. Dr. Juan P. Garrahan” - ARGENTINA

  2. Conflicts of Interest • No conflicts of interest to declare

  3. Whyisitimportant to understandthedynamics of HIV-1 reservoirs? • Since 1997 it has been possible to achieve virological suppression due to cART. • Today, HIV cure is still a challenge. • The long-term persistence of replication competent HIV-1 forms is the principal obstacle for a cure. Deeks SG. et al. Nat Medicine 2016

  4. Possiblemechanisms of HIV-1 reservoirmaintenance Molecular markers tHIV-DNA 2-LTR circles HIV-1 usRNA HIV-1 msRNA Virus evolution Eisele E. et al. Immunity 2012

  5. Questions addressed in the study • Isthere viral evolution in childrenundercART and long-termvirologicalsuppression? • Howistheinterplaybetweenreservoirsize and thedynamics of viral variants?

  6. Study design Study population selection Dynamics of circulating HIV-1 reservoirs HIV variantsdynamics Phylogenetic analysis Phylogenetic analysis HIV variantsdynamics Dynamics of circulating HIV-1 reservoirs

  7. Study population selection HIV infected children (n=1,181) Incomplete virological response (n=1,155) Achieved viral suppression <12 mo of life (n=26) Rapid virological failure (n=16) Sustained virological suppression (n=10) Less than 6 yrs of virological suppression (n=4) Maintain virological suppression >6 yrs (n=6) Non-available samples: at ART and VS initiation and during VS (n=1) Patients with available samples (n=5)

  8. Measurement of circulating HIV-1 reservoirs Total HIV-1 DNA • ART initiation • VS initiation • During VS Semi-nested Real Time PCR (ltr/gag) • Lastvisit (≥6 yrs of VS) 2-LTRs circles HIV-1 usRNA HIV-1 antibody

  9. HIV-1 variant pipeline BIOINFORMATIC WORKFLOW LABORATORY WORKFLOW Indexing Mergeintohaplotypes HIV-1 env Library (783 bp) Assembly HIV-1 gag Library (423 bp) Defectivehaplotypeexclusion Ampliconsequencingmethod NGS sequencing MiSeqIllumina In process • PBMC • ART initiation • VS initiation • During VS Tagmentation & Indexing Seq. with freq. < ER were mixed with most common seq. if they differ in 1 nt • Stop codon • Hypermutation

  10. Patient characteristics aClassification criteria based on ACTG 076: Complete, zidovudine administration to HIV-1 infected women during pregnancy and to the child during the first six weeks of life; Partial, zidovudine administration only to the pregnant women infected with HIV-1 or to the child. Abbreviations: ART, antiretroviral therapy; AZT, zidovudine; LPV/r, lopinavir/ritonavir; NFV, nelfinavir; NVP, nevirapine; pVL, plasma viral load; 3TC, lamivudine.

  11. IS THERE VIRAL EVOLUTION UNDER LONG-TERM VIROLOGICAL SUPPRESSION

  12. Pt. 5 Pt. 4 Pt. 3 Phylogeneticcharacteristics Pt. 2 Pt. 1 ML tree for HIV-1 gag region All haplotypes of each patient have a common ancestor There was no cross-contamination between patients

  13. Phylogeneticanalysis VS initiation 17-35 mo. of VS 69-108 mo. of VS 127-142 mo. of VS 69-108 mo. of VS 17-35 mo. of VS VS initiation pre-ART pre-ART 138-179 mo. of VS Pt. 2 Pt. 2 Pt. 3 Pt. 4 Pt. 5 Pt. 1 Pt. 1 r= 0.409 r=-0.445 r=-0.274 r=-0.115 r= 0.017

  14. HOW IS THE DYNAMIC OF HIV-1 HAPLOTYPES

  15. Dynamics of HIV-1 gag haplotypes Pt. 1 Haplotypefrequency -3 0 17 Time to VS (mo) 69 6 1,053 ND 18 24,853 153 14 6,213 3 9 NA NA Time to VS (mo) Nº haplotypes tHIV-1 DNA (cp/106 PBMC) 2-LTRs circles (cp/106 PBMC) NA: notavailable; ND: notdetected

  16. Dynamics of HIV-1 gag haplotypes Pt. 2 Haplotypefrequency -6 0 20 Time to VS (mo) 97 10 3,847 ND 28 61,183 232 24 19,757 83 4 NA NA Time to VS (mo) Nº haplotypes tHIV-1 DNA (cp/106 PBMC) 2-LTRs circles (cp/106 PBMC) NA: notavailable; ND: notdetected

  17. Dynamics of HIV-1 gag haplotypes Pt. 4 Haplotypefrequency -5 3 30 106 140 179 Time to VS (mo) 19 422 ND 4 ND ND 4 559 ND 16 NA NA 3 618 6 3 NA NA Time to VS (mo) Nº haplotypes tHIV-1 DNA (cp/106 PBMC) 2-LTRs circles (cp/106 PBMC) NA: notavailable; ND: notdetected

  18. Dynamics of HIV-1 gag haplotypes Pt. 5 Haplotypefrequency -5 7 35 108 127 138 Time to VS (mo) 31 2,705 ND 6 NA NA 16 NA 5 11 1,883 ND 6 NA NA 25 4,936 ND Time to VS (mo) Nº haplotypes tHIV-1 DNA (cp/106 PBMC) 2-LTRs circles (cp/106 PBMC) NA: notavailable; ND: notdetected

  19. Dynamics of HIV-1 gag haplotypes Pt. 3 Haplotypefrequency -6 0 17 107 142 160 Time to VS (mo) 5 ND ND 20 8 ND 14 29,572 128 14 NA NA 22 NA NA 30 NA NA Time to VS (mo) Nº haplotypes tHIV-1 DNA (cp/106 PBMC) 2-LTRs circles (cp/106 PBMC) NA: notavailable; ND: notdetected

  20. Dynamics of HIV-1 gag haplotypes Pt. 2 Pt. 4 Pt. 1 Pt. 5 Pt. 3

  21. Conclusions Therewas no evidence of ongoing viral replicationbyreservoirsizemarkers. CLONALEXPANSION HIV RESERVOIR Circulating DNA reservoir showed no signal of statistically significant evolution during suppresive ART Themostabundanthaplotypesfoundduring 6 to 14 yrs. of VS wereidentical to those at pre-cART .

  22. THANK YOU MUCHAS GRACIAS Scholarship

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