HLA and Drug Resistance Thomas Harrer Dept. of Medicine 3 University Hospital Erlangen Sandra Müller Birgit Schätz Bernd Spriewald Kathrin Eismann Hauke Walter Silke Bergmann Klaus Korn Ellen Harrer Barbara Schmidt Michael Bäuerle Heinrich Sticht SFB 466
HLA A 2 HLA CW HLA A3 8 CTL TCR HIV HLA B27 HLA CW 4 HLA B7
Tat vif Rev vpu Gag LTR LTR vpr Pol Env Nef HLA A2,A3 HLA A24,A32 B27,B8 B14,B51 Cw3,4 Cw5,8
Influence of M184V 3TC-Resistance mutation on CTL-recognition CTL CTL CTL TCR TCR TCR Q Y D V Y M Y D V D I I IVY I Q Y D D L L I I Q V V Y D L + - + Schmitt et al, AIDS 2000 Harrer E et al, JID 1996
CTL I TCR E Y I QY M Q Y D YM D D D I I L L V - +
AZT-Resistance mutation M41L Gnerates an HLA-A2 Neo-Epitope: Wilde type: ALVEICTEM does not bind to HLA A2 CTL mutated ALVEICTEL binds to HLA A2 Samri et al, JVi 2000 TCR M T E E C I I C T E A V E L A V L L - +
Correlations between AA substitutions in the PR and HLA class I alleles in 94 patients Published and A74 B44 B44 A2 A2 Cw4 Cw6 A2 A3/A11 A3/A11 potential HLA class I A2 Cw3 B13 Cw3 restricted epitopes A19 A68 Cw3 A3 A2 B51 A68 A2 A2/Cw6 B44 B62 B62 Negative HLA class I B62 A23 associations A3 B51 Cw5 A26 Cw6 Positive HLA class I B13 A2 A2 B44 associations A11 Cw7 A2 B18 B62 B44 Cw6 Cw3 Cw2 A23 B44 B44 Cw3 B62 B62 60 � � � PR variability in 94 patients (%) N37 L63 V3 89% 88% 83% 50 I93 ♦ E35 ♦ 40 I62 ♦ A71 Δ L10 V77 Δ M36 30 I15 M41 I64 V82 I13 ♦ H69 ♦ M46 L90 20 K20 L19 I54 T12 D60 ♦ K14 K43 ♦ L33 L89 G16 ♦ 10 E65 G73 I84 L24 T74 ♦ P39 V32 I47 F53 Q58 ♦ R70 D30 G48 N83 ♦ N88 I50 0 1 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95 amino acids described epitopes potential epitopes mutations are classified according to Müller et al, JVI 2007 Johnson et al. 2005 ♦ mutations contributing to TPV or ATV major mutations relevant correlations resistance minor mutations
Relevant correlations between HLA class I alleles and amino acid substitutions in the PR known potential univariate statistical analyses position HLA P -value OR n* epitopes epitopes** (Fisher’s exact test) major mutations L33 ♦Δ A2 0.005 12.6 9 A2 M46 ♦Δ A2 0.049 2.9 14 +/A2 minor mutations L10 ♦Δ Cw2 0.032 4.0 9 A71 Δ B44 0.013 3.5 14 B44 Bonferroni correction: p ≤ 0.00003 V77 B62 0.037 3.5 9 polymorphisms biologically relevant I13 ♦ A11 0.012 5.6 6 +/A3 supertype I13 ♦ B62 0.018 0.0027 0 correlations may be missed I13 ♦ B13 0.020 8.7 4 B13 K14 B51 0.001 13.0 6 B51 I15 A23 0.012 7.5 5 I15 Cw7 0.019 0.27 6 Cw7 E35 ♦ B44 1.2x10E-9 34.4 21 +/B44 E35 ♦ Cw5 0.000014 2543.6 10 to confirm correlations P39 B44 0.048 6.1 4 +/B44 P39 B18 0.046 10.6 2 B18 biological assays were R41 A26 0.032 9.1 3 A26 K43 ♦ B62 0.012 6.7 5 B62 conducted K43 ♦ Cw3 0.000073 21.5 8 Cw3 I54 ♦ B44 0.017 4.6 7 B44 I62 Δ A2 0.027 3.0 20 A2 I62 Δ A23 0.028 0.0017 0 I64 A3 0.030 0.2 2 A3 E65 Cw6 0.045 6.3 3 Cw6 T74 ♦ Cw6 0.028 8.4 3 Cw6 I93 Δ B62 2.0x10E-6 31.5 15 B62 I93 Δ Cw3 0.00038 8.9 17 Cw3
Recognition of EEMNLPGRW and variant peptides EEMNLPGRW EEMNLPGRW -D------- -D------- -DI------ -DI------ SFU/100 000 cells SFU/100 000 cells ---D----- ---D----- F S s 0 l e c 0 1 / U 0 U / 1 0 F c e l s S F S 0 0 s e c l 1 / U --I------ --I------ -----S--- -----S--- -------K- -------K- no peptide no peptide 0 200 400 600 800 0 200 400 600 800 EW9 E/D EW9 N/D EW9 M/I EW9 EW9 DI EEMNLPGRW -D------- -DI------ SFU/100 000 cells F 0 S l U c 1 0 s e / ---D----- --I------ -----S--- -------K- no peptide 0 50 100 150 200 250
E35D: polymorphism E35D negative effect on substrate binding affinity: Meiselbach et al. 2006 Reduced in vitro activity of ritonavir/amprenavir Associated with early treatment failure to PIs (Alexander et al. AIDS 2001)
Recognition of newly predicted epitopes containing HLA class I linked mutations aa position HLA m otif frequency of recognition K14 B51 Q RPLVTVKIG 11/16 L33 A2 LLDTG ADDTVL 29/76 K43 Cw 3 LPG RW KPKM I 4/15 K43 Cw 3 W KPKM IG G I 6/12 I62 A2 KVRQ YDQ IL 25/76 I64 A3 ILIEICG HK * 17/33 I62 B13 RQ YDQ IPIEI* 12/18 E65 B13 RQ YDQ IPIEI* 12/18 K70 A3 ILIEICG HK * 17/33 A71 B44 IEICG HK AIG 12/23 I93 B62 TQ IG CTLNF 4/6 I93 Cw 3 TQ IG CTLNF 4/9 I93 B62/Cw 3 TQ IG CTLNF 16/17 Mutations correlating to HLA class I alleles are shown in red
Einfluss von Mutationen auf die CTL Erkennung im pairm ent of im provem ent w ild-type m utant H L A C D 8+ T-cell of C D 8+ T-cell sequence peptides activity* activity * Q R P LV TV K IG Q R P LV TV R IG B 51 5/6 0/6 LLD TG AD D TV L LLD TG AD D TV F A2 6/12 2/12 LLD TG AD D TV L LID TG AD D TV F A2 4/12 7/12 K V R Q YD Q IL K V R Q YD Q V L A2 0/2 1/2 R Q YD Q IP IE I R Q YD Q V P IE I B 13 6/8 2/8 ILIE IC G H K ILIE IC G H R A3 9/12 1/12 IE IC G H K AIG IE IC G H K V IG B 44 3/11 7/11 TQ IG C TLN F TQ LG C TLN F B 62 3/6 3/6 B 62/ TQ IG C TLN F TQ LG C TLN F 5/12 4/12 C w 3 Mutations correlating to HLA class I alleles are shown in red
Recognition of KI10 and M46I / M54V mutants KMIGGIGGFI KMIGGIGGFI SFU/100 000 cells SFU/100 000 cells ---------V ---------V l e c s 0 0 1 / U F S . -I-------- -I-------- 0 / 1 0 . e c l s F U S no peptide no peptide 0 500 1000 1500 0 200 400 600 800 KI10-I KI10 KMIGGIGGFI KMIGGIGGFI F s l e c U / 1 0 . 0 S SFU/100 000 cells SFU/100 000 cells ---------V -I-------- l e s c . 0 U 0 / F S 1 -I-------- no peptide no peptide 0 200 400 600 800 0 200 400 600 800 KMIGGIGGFI (KI10) is highly conserved Contains the resistance mutations M46I/L and I54V M46I can abrogate CTL recognition = CTL escape mutation M46I in HLA A2+ Patienten more frequently (OR 2,9)
Summary • Significant Significant associati ssociation of frequent f frequent HLA alleles HLA alleles with occurrence ccurrence of defined f defined resis resistance ance mutations mutations and pol nd polymorphism morphisms • Definiti Definition of ten new on of ten new CTL epi TL epitopes topes • Dru Drug escap escape variants variants may may inhibit nhibit or or au augment gment the the CTL response, dependent CTL response, dependent on t the T e T-cell r l repertoire o e of t the p e patients. • Some Some patients patients mount mount an oligoclonal n oligoclonal immune response mmune response targeting targeting dru rug resis resistant ant viruses iruses • CTL induce CTL induce muta mutati tions ons presumably resumably have ave an important n important influence nfluence on the on the development development of drug of drug mutati mutation ons and mutati nd mutational onal pathways pathways
Selection by ART and CTL HIV-Genome ART ART + CTL CTL
HIV- -1 1 Protease Protease: : Structure Structure and and Resistance Resistance HIV ~ ¼ of all amino acids can mutate: Drug resistance and Polymorphisms : Los Alamos. http://hiv-web.lanl.gov/
Protease Inhibitor Resistance L K L V M M I A G V V I L 71 82 IDV 10 20 24 32 36 46 54 73 77 90 84 99 1 IRV MR I I I IL V VT SA I AFTS V M L 32 82 RTV 20 33 36 46 54 71 77 90 10 84 F IL VL FIRV MR I VT I AFTSV M G 71 82 SQV 10 48 54 73 77 90 84 IRV V VL I M VT S A V L D N 82 88 NFV 10 30 36 46 71 77 84 90 FI N I IL VT I AFTS V DS M V I I 46 APV 10 32 47 50 54 84 V V LMV FIRV I IL V L L 82 LPV/RTV 20 24 53 54 71 90 10 46 84 I FIRV MR IL F VL VT AFTS V M Antiretroviral Resistance Mutations, A collaborative effort of the International AIDS Society-USA Resistance Testing Panel and HIV InSite. HIV Clinical Trials 2001;2(4):346-355
Influence of resistance associated mutations on CD8+ T-cell recognition IEICGHKAIG TQIGCTLNF SFU/100 000 cells SFU/100 000 cells -------V-- --L------ 1 U / 0 0 c e l s S F S s U / 1 0 0 c e F l no peptide no peptide 0 100 200 300 400 500 600 0 50 100 150 200 IG10-V TF9-L IG10 TF9 VRQYDQIPIEI VRQYDQIPIEI SFU/100 000 cells SFU/100 000 cells ------V---- ------V---- no peptide. no peptide 0 100 200 300 400 500 600 0 200 400 600 800 VI11-V VI11-V VI11 VI11
Priming of M46I-Viruses by CTL strong CTL-response against KI10: Selection of M46I because of strong fitness disadvantage wild type M46 more prevalent in plasma on therapy wild type decreases and M46I prevails hypothesis: HLA A2+ u. B62 + patients develop M46I – Mutation more rapidly planned: Prospective study in patients failing PIs with M46I
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