Catherine K. Koofhethile 21st IAS 2016 18-22 July,2016 Durban, SA CD8+ T cell breadth and ex vivo virus inhibition capacity distinguish between viremic controllers with and without protective HLA class I alleles Catherine K. Koofhethile 21st IAS 2016 18-22 July,2016 Durban, SA

The impact of HLA on HIV disease progression 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 B*57 p < 0.0001 p = 0.0014 p = 0.0006 B*81:01 B*58:01 A*74:01 Mechanism of control? Protective Non protective B*58:02 Kiepiela et al, Nature, 2004 B*18:01 p < 0.0001 p = 0.0008

Gaps in knowledge ? responsible for control and loss of control? baseline non-controllers ? Failing VC (fVC) Viremic controllers (VC) What are the immunological and virological events responsible for control and loss of control?

Study design and methods in brief Protective HLA alleles: HLA- B*57, B*58:01, B*81:01, A*74:01 ARV naïve; at least 2 yr follow up. VL, CD4 count, HLA typing Immune responses – ELISPOT assay; OLP/optimal peptides covering whole HIV proteome. Sequencing of Gag (bulk) – molecular methods Ex vivo Viral inhibition assay SK COHORT n = 453 With protective HLA alleles n = 133 Baseline non-controllers (bNC) VL >100,000 RNA copies/ml (enrollment) n = 15 Baseline viremic controller (bVC+) VL< 2,000 RNA copies/ml n = 20 Without protective HLA alleles n = 320 VL > 100,000 RNA copies/ml n = 25 Baseline viremic controller (bVC) VL< 2,000 RNA copies/ml (enrollment) n = 10 7 VC+ 7 fVC+ 6 VC-

Clinical characteristics of study participants at enrollment Groups Protective HLA alleles +/- n Median years of follow up (IQR) Median viral load (HIV RNA copies/ml) (IQR) Median CD4 count (cells/mm3)(IQR) Baseline VC + 20 5 (4 - 6) 487 (283 – 1,252) 509 (459 – 684) Baseline non-controllers 15 4 (3 - 6) 156,000 (118,000 – 192,000) 329 (257 – 377) - 10 613 (399 – 1,213) 669 (550 – 865) 25 3 (2.5 - 4) 209,000 (132,000 – 330,500) 373 (260 – 467) Protective HLA alleles: HLAs B*57, B*58:01, B*81:01 and A*74:01

CD8+ T cell responses to Gag are associated with lower VL in individuals with protective HLA alleles OLP stim.

There is loss of virologic control amongst baseline viremic controllers with protective HLA alleles VC+ n=7 fVC+ n=7 VC- n=6

Loss of CD8+ T cell breadth is associated with loss of viral control Optimal peptide stim.

Viral escape and non-escape mechanisms of loss of CTL loss fVC+ Baseline Post loss timepoint DIAGTTSTLQEQIAWMTSNP DIAGTTSNLQEQITWMTSNP 200 B*57-TW10 p24 SCENARIO 1 -------------------- Baseline Post loss timepoint TLRAEQATQDVKNWMTDLL 300 B*57-QW9 p24 SCENARIO 2 What remains unknown is the loss of CD8+ T cell responses in the absence of escape

CD8+ T cell ex vivo virus inhibition assay HIV CD4+ T-cells Black Box + Autologous CD8+ T-cells Co-culture infected CD4 T-cells with CD8+ T cells at E:T ratio 1:1 Inhibition of HIV replication measured by p24 in supernatants

Overall inhibition- VC+ display enhanced ex vivo CD8+ T cell inhibitory capacity

Main findings Baseline viremic controllers (bVC) with protective HLA alleles (+) n=14 Broad Gag CTL responses Good virus inhibition capacity Baseline viremic controllers (bVC) without protective HLA alleles (-) n=6 Limited Gag CTL responses, Limited virus inhibition capacity VC (-) n=6 Maintain limited CTL responses CTLs limited ability to inhibit viral replication VC (+) n=7 Maintain broad Gag CTLS CTLs maintained ability to Inhibit viral replication. Failing VC (+) n=7 Lost CTL responses CTLs had limited ability to inhibit viral replication Loss of some CTL responses  escape Mechanism of control- CTL dependent Mechanism of control- Non CTL Loss of some CTL responses NOT due to escape -> ?????? CD4? Virus? Innate immunity? Koofhethile et al. JVI 2016

Acknowledgements Harvard/MGH HPP, UKZN Prof. Bruce Walker Elana Stampouloglou University of Oxford Prof. Philip Goulder AIDS Research Institute IrsiCaixa, Spain Dr Julia Prado Funding Organisation for Women in science for the developing world NIH, Ragon Institute of MGH, MIT and Harvard University. HPP, UKZN Prof. Thumbi Ndung’u Dr Christina Thobakgale Dr Zaza Ndhlovu HPP team Sinikithemba study team Study participants CAPRISA Nonhlanhla Yende