Dr. Jeffrey Dorfman Cellular Immunology ICGEB Cape Town Autoreactivity of anti-HIV-1 neutralizing antibodies does not prevent broad antibody responses

Problems to solve in HIV vaccine research No vaccine has induced broadly neutralising antibodies. Epitope modelling approaches have never before been used to generate any licensed vaccine. Autoreactivity and polyreactivity – common among anti-HIV neutralising Abs – though to pose barriers to the development of neutralisation breadth. – Verkoczy, L. & Diaz, M. (2014). Autoreactivity in HIV-1 broadly neutralizing antibodies: implications for their function and induction by vaccination. Curr Opin HIV AIDS 9,

Targets for broadly neutralizing antibodies on HIV envelope proteins, gp120 and gp41 Burton DR et al, Cell Host and Microbe 2012 Broadly neut Narrowly neut Broadest antibodies What about the rest of the continuum?

What determines if an antibody is broad? 1.Antibody response to an epitope must be made. 2.Antibody response to an epitope must mature. One component of this is based upon immunodominance and overall dynamics of the antibody response. Antibody genes must mutate, and these mutations must lead it on a path to greater neutralisation breadth. Vast differences in rates & likelihood noted. (Doria-Rose, Nature 2014 vs Liao HX, Nature 2013) Greater neutralisation breadth in a cross sectional cohort should reflect: higher likelihood of reaching breadth higher average breadth achieved faster rate of reaching breadth This will mostly reflect the difficulty in achieving neutralisation breadth for antibodies directed against that site in a human population.

Targets for broadly neutralizing antibodies on HIV envelope proteins, gp120 and gp41 Burton DR et al, Cell Host and Microbe 2012

Autoreactivity and polyreactivity Liu M et al J Virol 89(1): Reactivity of indicated anti-HIV-1 mAb Reactivity of control mAb anti-MPER bnmAb anti-V2 g-site bnmAb anti-V3/glycan bnmAb Antibody equality controls

Our study –which epitopes contribute to breadth? Measure neutralisation breadth and potency of a set of sera from chronically HIV-infected ART-naive individuals. Map sera for anti-MPER and dominant anti-V2 glycan site and dominant anti-V3/glycan antibodies. Assess likely contribution of antibodies to each site to neutralisation breadth and potency.

Cohort and virus panel 177 chronically HIV-1-infected individuals – Infected >1 yr – Antiretroviral naive (except PMTCT) 2 locations in Cape Town, South Africa – Groote Schuur Hospital – Khayelitsha Site B clinic (primary) 24 pseudovirus panel – Hard to neutralize (Tier 2–3) Five: Tier 3 (Seaman et al 2010) Six: more resistant than Tier 2 but not consistently grouping w/ Tier 3 – Geographically & clade distributed (A, B, C, G, CRF02_AG)

Mapping methods For MPER: – Swap in an HIV-1 MPER into an HIV-2 envelope – Test 3 HIV-1 MPER sequences (control=HIV-2) For V2 glycan site and V3/glycans: – Mutate key residues in epitope, measure drop in neut – 160, 169 for V2 glycan site; 301, 332 for V3/glycans – Test 3 parent viruses HIV-2 HIV-1 *

Sera from 177 chronically HIV-infected individuals Jacob RA, Moyo T et al J Virol 89(10): Geometric mean ID Neutralization potency Number of panel viruses neutralized (n=24) % of samples 25 Neutralization breadth 18% (32/177)16% (29/177)

Sera with anti-MPER or dominant anti-V3/glycan antibodies are broader and more potent neutralizers Difference in neutralisation breadth Ratio of neutralisation potencies

Significance Previous HIV-vaccine candidates have failed to: – Induce broadly neutralising antibodies – Substantially protect vaccinees New approaches are being used to design candidate HIV vaccines – Never successfully used to design any licensed vaccine Candidate immunogens defined by targets of broadly neutralising antibodies Antibodies to some targets are autoreactive and/or polyreactive – Thought to pose direct and indirect barriers to inducing neutralisation breadth Yet it is these targets whose antibodies are associated with breadth Autoreactivity/polyreactivity effects are not decisive and these targets should remain high priority vaccine candidates

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Detecting anti-MPER antibodies 3 HIV-2/HIV-1 chimeric viruses C1C, C1, MPER Chimera Has an exposed, easy-to-neutralise HIV-1 MPER Compare neutralisation between parent HIV-2 virus and HIV-2/HIV-1 chimeric viruses HIV-2 HIV-1 (1/3 sequences or HIV-2 control)

Detecting anti-V3/glycan antibodies Key mutations for V3/glycan &2G12-like antibody binding: N301A, N332A (Walker et al., 2011, Nat.) Du156 N301A.N332A QH343.A10 N301A.N332A Cap45 N301A

Detecting anti-V2/glycan site antibodies Key mutations for V3/glycan antibody binding: N160A, K169E (Moore et al. 2011; Tomaras et al. 2011) Du156 N160K, K169E QH343.A10 N160A, I169E Cap45 N160A, K169E

Sera from 177 chronically HIV-infected individuals Jacob RA, Moyo T et al J Virol 89(10): Geometric mean ID Neutralization potency Number of panel viruses neutralized (n=24) % of samples 25 Neutralization breadth 18% (32/177)16% (29/177)

Distribution of specificities I Fold drop in ID % of samples mapped N160A/K mapping for V2/glycan site K/I169E mapping for V2/glycan site % of samples mapped Fold drop in ID 50 29% (34/118)

Distribution of specificities II % of samples mapped Fold drop in ID 50 N301A/N332A mapping For anti-V3/glycans < % of samples ID 50 of MPER chimaera constructs, maximum of 3 constructs 64 Anti-MPER antibody levels 19% (33/177)19% (21/113)

Presence of anti-MPER and dominant anti-V3/glycan abs (& not anti- V2-g-site abs) are associated with neutralisation breadth and potency Category Less potent a Potently neutralizing b Relative Risk (95% CI) p value (  2 ) Less broad c Broadly neutralizing d Relative Risk (95% CI) p value (  2 ) Anti –MPER neg (reference) (reference) Anti-MPER pos (0.99, 3.91) (1.04, 3.78)0.043 Anti-V2/glycan site neg (reference) (reference) Anti-V2/glycan site pos (0.24, 1.43) (0.37, 1.67)0.522 Anti-V3/glycan neg (reference) (reference) Anti-V3/glycan pos (1.21, 4.46) (1.10, 3.92)0.033 a Geometric mean ID 50 <220 b Geometric mean ID 50 >220 c <18/24 panel viruses neutralized d >18/24 panel viruses neutralized