HIV broadly neutralizing antibodies: learning lessons from infections Penny Moore National Institute for Communicable Diseases, a division of the National Health Laboratory Service of South Africa, University of the Witwatersrand, Johannesburg, South Africa and the Centre for the AIDS Program of Research in South Africa (CAPRISA) IAS, 2014, Melbourne

Broadly neutralizing antibodies (BNAbs) develop in a fifth of HIV infected people Elite neutralizers (1%) Screening of chronically infected people shows BCN antibodies develop fairly often... Broad neutralizers (20%) No/limited breadth

Isolation of monoclonal anti-HIV antibodies We know (most of) the targets V2/glycan >12 mAbs V3/glycan supersite >25 mAbs gp120-gp41 interface >3 mAbs CD4bs >25 mAbs MPER >5 mAbs Modified from Burton et al., Science 2012

Ontogeny of BNAbs: How do BNAbs develop? Breadth years of infection UCA (unmutated common ancestor)

Ontogeny of BNAbs: How do BNAbs develop? Breadth years of infection UCA

Ontogeny of BNAbs: How do BNAbs develop? Breadth years of infection UCA

Developmental pathways are likely to differ by epitope V2/glycan >12 mAbs Long CDRH3 (>25 aa) to penetrate glycan shield V3/glycan >25 mAbs Heavily mutated (30%) CD4bs >25 mAbs MPER >5 mAbs Modified from Burton et al., Science 2012

Developmental pathways are likely to differ by epitope V2/glycan >12 mAbs V3/glycan >25 mAbs Heavily mutated (30%) CD4bs >25 mAbs MPER >5 mAbs Modified from Burton et al., Science 2012

CD4bs BNAbs – highly affinity matured Unmutated common ancestor Mature BNAb Highly mutated away from their ancestor years of infection

Do long CDR H3s similarly develop gradually? V2/glycan >12 mAbs Long CDRH3 (>25 aa) to penetrate glycan shield V3/glycan >25 mAbs MPER >5 mAbs Modified from Burton et al., Science 2012

CAP256 targets V2 CAP256 Gray et al 2011, Moore et al 2011

Twelve related mAbs isolated by B cell culture V1V2-directed NAbs Neutralization breadth starts Weeks post-infection 15 30 38 48 59 119 176* 206 Week Antibody Somatic mutations (nt) CDR H3 length (aa) Neutralization (46 strains) Vl1-51*02 VH3-30*18 Breadth Potency (IC50, μg/ml) 59 CAP256-VRC26.01 3.9% 8.3% 35 15% 2.93 119 CAP256-VRC26.02 4.9% 8.7% 17% 0.40 CAP256-VRC26.03 7.4% 30% 0.06 CAP256-VRC26.04 8.1% 9.0% 28% 0.25 CAP256-VRC26.05 5.4% 10.1% 22% 0.10 CAP256-VRC26.06 10.8% 36 0.16 CAP256-VRC26.07 7.7% 11.8% 13% 2.02 CAP256-VRC26.08 9.8% 37 46% 0.14 CAP256-VRC26.09 14.2% 0.08 206 CAP256-VRC26.10 24% 0.60 CAP256-VRC26.11 13.7% 11.9% 0.82 CAP256-VRC26.12 8.4% 15.3% 7% 0.49 Doria-Rose, Schramm, Gorman, Moore et al, Nature, 2014

CAP256-VRC26 has typical features of anti-V2 broad neutralizing antibodies PG9 CAP256-VRC26 Antibody BG505 SOSIP HIV Env Jason Gorman and Peter Kwong HL Kim, A Cupo, R Sanders, IA Wilson, JP Moore, AB Ward Ideal for defining the developmental pathways of V2 antibodies with long CDR H3

CAP256-VRC26 heavy chain and light chain antibody sequences VH3-30*18 VL1-52*02 UCA_H UCA_L CAP256-VRC26.01 CAP256-VRC26.10 CAP256-VRC26.11 CAP256-VRC26.06 CAP256-VRC26.05 CAP256-VRC26.02 CAP256-VRC26.04 CAP256-VRC26.03 CAP256-VRC26.07 CAP256-VRC26.12 CAP256-VRC26.09 CAP256-VRC26.08 The UCA had a 35 amino acid CDR H3 fully formed through VDJ recombination. Heavy Chain longitudinal phylogenetic tree Light Chain longitudinal phylogenetic tree Week 38 48 59 119 176 206 Evolutionary distance 0.02 Chaim Schramm and Larry Shapiro

Unmutated common ancestor The CAP256-VRC26 unmutated common ancestor had a fully formed long CDR H3 Long CDRH3s Mature BNAb Unmutated common ancestor

Unmutated common ancestor The CAP256-VRC26 unmutated common ancestor had a fully formed long CDR H3 Long CDRH3s Was this enough for breadth? Mature BNAb Unmutated common ancestor

Development of CAP256-VRC26.01 V1V2-directed NAbs Neutralization breadth starts Weeks post-infection 15 30 38 48 59 119 176* 206 Week Antibody Somatic mutations (nt) CDR H3 length (aa) Neutralization (46 strains) Vl1-51*02 VH3-30*18 Breadth Potency (IC50, μg/ml) 59 CAP256-VRC26.01 3.9% 8.3% 35 15% 2.93 119 CAP256-VRC26.02 4.9% 8.7% 17% 0.40 CAP256-VRC26.03 7.4% 30% 0.06 CAP256-VRC26.04 8.1% 9.0% 28% 0.25 CAP256-VRC26.05 5.4% 10.1% 22% 0.10 CAP256-VRC26.06 10.8% 36 0.16 CAP256-VRC26.07 7.7% 11.8% 13% 2.02 CAP256-VRC26.08 9.8% 37 46% 0.14 CAP256-VRC26.09 14.2% 0.08 206 CAP256-VRC26.10 24% 0.60 CAP256-VRC26.11 13.7% 11.9% 0.82 CAP256-VRC26.12 8.4% 15.3% 7% 0.49 Nicole Doria-Rose, Ryan Staupe, Nancy Longo, Jinal Bhiman

Rapid development of neutralization breadth within the CAP256-VRC26 lineage VRC26-UCA VRC26-I1 VRC26-I2 VRC26.01 Chaim Schramm

Rapid development of neutralization breadth within the CAP256-VRC26 lineage VRC26-UCA VRC26-I1 VRC26-I2 VRC26.01 CAP256 SI The CAP256-VRC26 UCA, despite having a long CDR H3, can ONLY neutralize the virus that infected CAP256 - no neutralization breadth

Rapid development of neutralization breadth within the CAP256-VRC26 lineage VRC26-UCA VRC26-I1 VRC26-I2 VRC26.01 CAP256 SII Chaim Schramm

Only moderate levels of somatic hypermutation needed for breadth Mature BNAb with moderate somatic hypermutation Unmutated common ancestor Weeks…. ✗ years of infection

Ontogeny of BNAbs: Looking backwards from breadth years of infection UCA

Dramatic viral changes immediately precede CAP256 neutralization breadth Maturation of the anti-V2 lineage CAP256-VRC26 was associated with viral replacement in the epitope Broadly neutralizing antibodies emerge in the context of multiple immunotypes created through viral diversification V1V2 mutations 160-171 Duration of infection CAP256-VRC26 emerges in peripheral B cell repertoire

Dramatic viral changes immediately precede CAP256 neutralization breadth Maturation of the anti-V2 lineage CAP256-VRC26 was associated with viral replacement in the epitope Broadly neutralizing antibodies emerge in the context of multiple immunotypes created through viral diversification V1V2 mutations 160-171 Duration of infection CAP256-VRC26 emerges in peripheral B cell repertoire Plasma heterologous neutralization develops Jinal Bhiman, Daniel Sheward, Molati Nonyane, Bronwen Lambson

Emergence of escape mutations drives increased breadth 166 CAP256 infecting virus 94 weeks p.i. R166, a crucial part of the CAP256 epitope, undergoes selection pressure as bNAbs mature Jinal Bhiman, Daniel Sheward

Emergence of new immunotypes drives increased breadth – K166 All subtype C viruses (n=1,005) 166 CAP256 infecting virus 166 94 weeks p.i.

Emergence of new immunotypes drives increased breadth – K166 All subtype C viruses (n=1,005) 166 CAP256 infecting virus 166 94 weeks p.i. CAP256 CAP256 R166K Percent breadth

Summary Isolated PG9-type V1V2 neutralizing mAbs from seroconverting donor CAP256 CAP256 mAbs have a characteristic long anionic CDRH3 formed by initial gene recombination; i.e., present on naïve BCR The UCA was capable of strain-specific neutralization, with modest affinity maturation conferring breadth Viral diversification and co-evolution was associated with breadth, through bNAb tolerance of escape mutations Doria-Rose, Schramm, Gorman, Moore et al, Nature, 2014

Which pathway is more amenable to HIV vaccine design? CD4bs - highly mutated away from their ancestor V1V2 - Long CDRH3s Requires the engagement of a BCR with a long CDR H3 - these B cells are very rare No requirement for long CDR H3, but Ig allele skewing may limit viable BCRs Once stimulated, V1V2 BNAbs can develop within months, not years May need high levels of affinity maturation take years – hard to achieve through vaccination Immunogens may need to recreate antigenic diversity to drive affinity maturation Immunogens may need to recreate antigenic diversity to drive affinity maturation

Acknowledgements NICD Jinal Bhiman IAVI NAC, CHAVI-ID and Scripps Participant CAP256 Slim Abdool Karim, Quarraisha Abdool Karim, Nigel Garrett and the CAPRISA staff NICD Jinal Bhiman Kurt Wibner Molati Nonyane Bronwen Lambson Thandeka Khoza Mashudu Madzivhandila Lynn Morris Mascola lab Nicole Doria-Rose R. Staupe, R. Roark M. Ernandes Rebecca Lynch Rui Kong Nancy Longo Sijy O’Dell Stephen Schmidt Krisha McKee Mark Louder John Mascola Kwong lab Jason Gorman Marie Pancera Tongqing Zhou Baoshan Zhang Ivelin Georgiev Young Do Kwon Yongping Yang Peter Kwong Bioinformatics Chaim Schramm Zhenhai Zhang Cinque Soto Ivelin Georgiev Larry Shapiro IAVI NAC, CHAVI-ID and Scripps Andrew Ward Helen Kim Albert Cupo John Moore Rogier Sanders Wayne Koff Ian Wilson Dennis Burton U. Texas Austin Brandon DeKosky George Georgiou Williamson lab Daniel Sheward Carolyn Williamson Torrey Pines Emma Crooks James Binley NIH Sequencing Core Holly Coleman Brian Schmidt Morgan Park Jim Mullikin NIAID NVITAL Ellen Turk Bob Bailer