dr penny moore

Characterization of acute infection in South Africa and its relevance to HIV vaccine

discovery - focus on neutralizing antibodies

Dr Penny Moore

CENTRE FOR THE AIDS PROGRAMME OF CENTRE FOR THE AIDS PROGRAMME OF RESEARCH IN SOUTH AFRICARESEARCH IN SOUTH AFRICACCAPRISAAPRISA CAPRISA is a UNAIDS CAPRISA is a UNAIDS

Collaborating Centre for HIV Collaborating Centre for HIV Prevention ResearchPrevention Research

AIDS Virus Research Unit, National Institute for Communicable Diseases, Johannesburg, South Africa

Salim Abdool Karim, PIKoleka MlisanaThumbi Ndung’u

Carolyn Williamson Joanne Passmore

Lynn MorrisClive Gray

Winston Hide

Registration Number: 2002/024027/08 http://www.caprisa.org

CENTRE FOR THE AIDS PROGRAMME OF RESEARCH IN SOUTH AFRICACAPRISA CAPRISA is a UNAIDS

Collaborating Centre for HIV Prevention Research

Western Cape

Eastern Cape

Northern Cape

North West

Free State

Northern Province

Gauteng

KwaZuluNatal

Mpumalanga

Durban

Vulindlela

Introduction – neutralizing antibodies

Neutralizing antibody (nAb) responses develop in most HIV-1 infected individuals within a few months of infection (Moog et al, 1997; Richman et al, 2003; Wei et al, 2004)

Much of the variation that occurs in Env during early infection may be due to nAb pressure (Frost, 2005)

Neutralization escape extensively documented in HIV-1 subtype B and SIV – substitutions, indels, glycan shifts (Richman et al, 2003; Wei et al, 2004)

Autologous nAb response in subtype C develop to higher titres and show greater type-specificity (Li et al, 2006; Gray et al, 2007)





Autologous neutralizing responses(n=14)

0 10 20 30 40 50 60 70 80 90 10010

100

1000

10000

Controllers (n=3)Rapid progressors (n=3)Intermediate progressors (n=8)

weeks post-infection

Ave

rag

e ac

cum

ula

ted

neu

tral

izat

ion

Median initial autologous response is 19 weeks post-infection

Development of the autologous neutralizing antibody response in early subtype C infection

CCAPRISAAPRISAGray ES, Moore PL et al., JV 2007

CCAPRISAAPRISA

CAP 8 CAP 45 CAP 61 CAP 63 CAP 84 CAP 85 CAP 88 CAP 206 CAP 210 CAP 228 CAP 239 CAP 244 CAP 255 CAP 256CAP 8 733 <20 40 <20 29 248 <20 37 <20 <20 <20 29 23 <20CAP 45 <20 6199 <20 <20 <20 <20 <20 <20 <20 <20 <20 <20 <20 <20CAP 61 <20 <20 54 <20 145 84 <20 <20 <20 <20 32 <20 <20 <20CAP 63 <20 <20 <20 5490 <20 <20 <20 <20 <20 <20 <20 <20 <20 <20CAP 84 <20 <20 <20 <20 2585 21 <20 <20 <20 <20 <20 <20 <20 <20CAP 85 29 <20 <20 113 <20 1697 <20 <20 <20 <20 <20 <20 <20 <20CAP 88 <20 <20 <20 <20 <20 33 1097 <20 <20 <20 <20 <20 <20 <20CAP 206 <20 <20 <20 <20 <20 236 <20 2425 <20 <20 <20 <20 <20 <20CAP 210 <20 <20 <20 <20 <20 <20 <20 <20 138 <20 <20 <20 <20 <20CAP 228 <20 36 <20 <20 27 41 <20 51 <20 405 <20 <20 <20 <20CAP 239 <20 <20 37 <20 28 <20 <20 <20 <20 <20 3841 <20 <20 <20CAP 244 <20 <20 <20 <20 <20 <20 <20 <20 <20 <20 <20 416 <20 <20CAP 255 <20 <20 96 <20 <20 78 <20 <20 <20 <20 67 <20 2163 <20CAP 256 <20 195 27 <20 <20 <20 <20 <20 <20 <20 85 <20 <20 188

12th months serum

Enrolment virus

<2020-5050-100100-500500-1000

>1000

Gray ES, Moore PL et al., JV 2007

The Early Neutralizing Antibody Response at 1 Year is Highly Type-specific

V1V2 C3-V4 C3 V4 V5CAP45 ++ ++ - - -CAP84 - +++ - - +/-CAP63 - +++ - +/- -CAP88 ++ +++ +++ - -

Moore PL, Gray ES et al., JV 2008 CCAPRISAAPRISA

The Major Target of the Early Autologous Neutralizing Antibody Response is the C3-V4 Region of the HIV-1

Subtype C Envelope

Numbers and kinetics of autologous nAbs in early infection is not known

0 25 50 75 100 12540

400

4000

40000 88.1m.c1788.6m.c1088.6m.c4488.12m.c288.12m.c1788.12m.c2388.12m.c4588.12m.c46

Weeks p.i.

Tit

er

Neutralization escape in HIV-1 subtype C – CAP88

SGA derived clones from 1 month, 6 months and 12 months p.i. were tested against plasma spanning 2 years p.i. to assess neutralization escape.

12 months p.i.

6 months p.i.

1 month p.i.

Peak 1 Peak 2

Moore et al., PLoS Pathogens, in press

0 25 50 75 100 12540

400

4000

40000 88.1m.c17

63/88/63-C3

CAP6363/88/63-V1V2

Weeks p.i.

Tit

erAnti-C3 Anti-V1V2

Temporal variations in neutralization titers correspond to waves of evolving specificities – CAP88

Use of heterologous chimeric envelopes suggest:

• initial anti-C3 response develops, peaking at 26 weeks p.i., then waning.

• second anti-V1V2 response peaking at 81 weeks p.i.


α2helix __________________ [ V1 V2 ] [ C3 ] ____________________________ _______________________________________ __________________________________________________ 88.1mo.6A CTNVTVVNATHTNKSMNGEIDMKEEMKNCSFKTTTGIRGKKQTEYALFYRPDIVPLSKESSEYILISC KDRWIATLEKVKKKLEEHFPNKTKIKFAPSSGGDLEVTTHSFNCGGEFFY 88.1mo.17 .................................................................... .................................................. 88.1mo.6B ...................T................................................ .................................................. 88.1mo.9 .................................................................... .................................................. 88.1mo.15 .................................................................... .................................................. 88.6mo.29 ................D................................................... ....N............................................. 88.6mo.5 .............T............................................G......... ....N...K......................................... 88.6mo.2 .................E.................................................. ....N...K......................................... 88.6mo.25 .................................................................... ....N...K......................................... 88.6mo.48 .................................................................... ....N...K......................................... 88.6mo.44 ...............R.................................................... ....N..........K.................................. 88.6mo.10 .................E.................................................. ....N..........K.................................. 88.6mo.21 .................E.................................................. ....N..........K.................................. 88.12mo.15 ................T.........................................D--....... ....N..........A.................................. 88.12mo.23 ...............R.........................................NS--....... ....N................N............................ 88.12mo.17 .................V.T......................................D--....... ....N..........K.......-.......................... 88.12mo.35 ................T.........................................D--....... ....N...K..............-.......................... 88.12mo.47 .................V.T......................................D--....... ....N...K..............-.......................... 88.12mo.1 .............R.....T..............P.....E................N.......... ....S..................-.......................... 88.12mo.2 ..................--..............P.....E................N.......... ....N..........K.......-.......................... 88.12mo.46 ..................--....................EK...............N.......... ....N..................-.......................... 88.12mo.16 ..................--..............P.....E................N.......... ....N..................-.......................... 88.12mo.45 ...I..............--.................I..................NN.......... ....N...........K......-.......................... 88.12mo.25 .............R....................-......................N.......... ....N..................-.......................... 88.12mo.21 .............R.....T..............-......................N.......... ....N..................-.......................... 88.12mo.31 ..................--...........Q..P.....................GN.......... ....N..................-..........................

Escape mutations in CAP88





0 25 50 75 100 12540

400

4000

4000088.1m.c17

88.12m.c2

88.12m.c2-V1V2s

88.12m.c2-V1V2s,C3s

88.6m.c10

Weeks p.i.

Tit

er0 25 50 75 100 125

40

400

4000

40000 88.1m.c1788.6m.c1088.6m.c10-C3s

weeks p.i.

Tit

re

Escape at 6 months p.i. is mediated by 2 amino acid changes in C3

Escape at 12 months p.i. is mediated by changes in V1V2 (and C3)

Limited nAb specificities drive sequential escape mutations – CAP88 cont





Limited and sequential nAb specificities drive sequential escape mutations – CAP177

Escape at 6 months p.i. is mediated by changes in C3

Escape at 12 months p.i. is mediated by changes in V1V2

In CAP88 and CAP177, nAbs first target the C3 region, then the V1V2 loop

The V1V2 region is a target of autologous nAbs and changes within V1V2 mediate escape – CAP210

V1V2 is the nAb target in CAP210 (heterologous chimera data)

Escape at 12 months p.i. is mediated by changes in V1V2





A single specificity mediates neutralization of CAP45

0 25 50 75 100 12540

400

4000

4000045.2wk.c945.12m.c7

45.12m.c7 E460K/G462D

Weeks p.i.

Tit

er





K460E,D462G/K460E [ (V5) ] __________________ 45.2wks.9 TRDGGKTDRNDTEIFRPG 45.2wks.8 .................. 45.2wks.A5 .................. 45.2wks.A2 .................. 45.2wks.A4 .................. 45.2wks.B12 .................. 45.2wks.B9 .................. 45.2wks.B1 .................. 45.2wks.B8 .................. 45.2wks.B10 .................. 45.2wks.TA1 .................. 45.2wks.B4 .................. 45.4mo.14 .......G.......... 45.4mo.19 .....---.......... 45.4mo.20 .......G.......... 45.4mo.23 .......G.......... 45.4mo.26 .......G.......... 45.4mo.3 .......G.......... 45.4mo.7 .......G.......... 45.8mo.40 .....E............ 45.8mo.9 .......G.......... 45.8mo.15 .......G.......... 45.8mo.18 .......G...A...... 45.8mo.22 .......G.......... 45.8mo.24 .......G...A...... 45.8mo.27 .......G.......... 45.8mo.7 ...*...G.......... 45.8mo.47 .......G.......... 45.8mo.43 .......G...A...... 45.12mo.7 .....E.G.......... 45.12mo.8 .....E.G.......... 45.12mo.17 .....E..K......... 45.12mo.19 .....E.G..........

K460E,D462G/K460E [ (V5) ] __________________ 45.2wks.9 TRDGGKTDRNDTEIFRPG 45.2wks.8 .................. 45.2wks.A5 .................. 45.2wks.A2 .................. 45.2wks.A4 .................. 45.2wks.B12 .................. 45.2wks.B9 .................. 45.2wks.B1 .................. 45.2wks.B8 .................. 45.2wks.B10 .................. 45.2wks.TA1 .................. 45.2wks.B4 .................. 45.4mo.14 .......G.......... 45.4mo.19 .....---.......... 45.4mo.20 .......G.......... 45.4mo.23 .......G.......... 45.4mo.26 .......G.......... 45.4mo.3 .......G.......... 45.4mo.7 .......G.......... 45.8mo.40 .....E............ 45.8mo.9 .......G.......... 45.8mo.15 .......G.......... 45.8mo.18 .......G...A...... 45.8mo.22 .......G.......... 45.8mo.24 .......G...A...... 45.8mo.27 .......G.......... 45.8mo.7 ...*...G.......... 45.8mo.47 .......G.......... 45.8mo.43 .......G...A...... 45.12mo.7 .....E.G.......... 45.12mo.8 .....E.G.......... 45.12mo.17 .....E..K......... 45.12mo.19 .....E.G..........


Summary of autologous neutralizing antibody targets Summary of autologous neutralizing antibody targets and escape mutationsand escape mutations

CAP88 CAP210CAP177CAP45

C3V4 α2-helix

V1V2 V1V2 V1V2

α2-helix

V1V2V1V2V1V2V5

α2-helix

NeutralizationTargets

Escapemutations

Early targets

Later targets

Early escape mutations

Later escape mutations

Not known

5 10 15 20 25 30

0

25

50

75

100

% Neutralization

100

1000

10000

100000

1000000

VL (overall)WT VL (VL x proportion WT)Mut VL (VL x proportion Mut)

Weeks p.i.

% n

eutr

aliz

atio

n a

t 1:

45d

ilu

tio

n

VL

(cop

ies / ml)

5 10 15 20 25 300

25

50

75

100 Wildtype (I339)Mutant (I339N)

% Neutralization

0

25

50

75

100

Weeks p.i.

% n

eutr

aliz

atio

n a

t 1:

45d

ilu

tio

n

% o

f po

pu

lation

Do autologous nAbs affect viral load?

7-fold 4-fold





The relationship between autologous nAbs and the development of cross-neutralizing breadth

A vaccine should aim to elicit cross-neutralizing antibodies

The specificities of antibodies in sera with neutralization breadth are largely unknown (Binley et al, 2008; Sather et al, 2009; Gray

et al, in press )

The mechanisms that lead to the development of breadth are not known – likely to be dependent in part on the autologous infecting envelopes (Rademeyer et al, 2007)

The contribution of autologous neutralizing antibodies to the development of breadth has not been explored





The Neutralizing Antibody Response at 3 Years post-infection shows increased breadth

ConC Du156 Du422 CAP206 CAP210 CAP228 ZM53M ZM135M ZM197M ZM214M CAAN AC10 6535 Q23 Q842

12 1 8 E8 51 PB12 PL10a PB7 PL15 5342 0.29 3 17 d12

CAP8 >1245 299 48 <45 <45 <45 <45 <45 51 <45 155 169 439 636 <45 40%

CAP40 <45 <45 <45 <45 <45 <45 <45 46 <45 <45 <45 <45 <45 <45 <45 0%CAP45 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 0%CAP61 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 0%CAP65 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 0%CAP84 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 69 <45 0%CAP85 nd 276 74 <45 <45 <45 <45 <45 201 <45 <45 <45 172 <45 <45 20%CAP88 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 0%CAP129 341 <45 <45 <45 <45 166 <45 <45 51 56 <45 <45 151 <45 <45 20%CAP137 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 0%CAP177 1053 219 106 <45 122 145 <45 162 <45 <45 261 787 386 626 <45 67%

CAP200 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 187 7%CAP206 <45 692 388 >1245 <45 <45 <45 107 751 <45 <45 111 <45 736 <45 43%

CAP217 <45 309 136 624 121 <45 <45 424 <45 <45 <45 <45 <45 <45 <45 33%CAP221 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 0%CAP222 <45 <45 <45 <45 71 <45 <45 53 <45 <45 <45 <45 <45 <45 <45 0%CAP225 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 0%CAP228 nd <45 <45 <45 <45 310 <45 <45 <45 <45 <45 <45 46 69 <45 0%CAP229 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 46 <45 <45 0%CAP239 187 <45 786 <45 52 <45 <45 <45 <45 <45 <45 <45 170 576 <45 27%CAP244 nd <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 96 <45 0%CAP248 >1245 160 147 841 511 535 <45 <45 98 <45 <45 <45 210 331 248 60%

CAP255 392 520 334 <45 <45 <45 <45 <45 <45 <45 81 49 112 624 <45 33%CAP256 >1245 >1245 >1245 >1245 >1245 >1245 >1245 69 216 920 <45 48 581 >1245 658 80%CAP257 >1245 143 147 78 <45 95 99 63 84 386 <45 <45 88 615 373 40%

CAP261 193 62 <45 <45 64 66 <45 <45 131 <45 <45 <45 <45 <45 <45 13%CAP262 104 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 <45 7%CAP264 <45 47 <45 183 <45 <45 57 86 <45 <45 <45 <45 <45 <45 <45 7%

nd: not done

% viruses neutralized

Viruses (ID50 titer)

Serum ID

Subtype C Subtype B Subtype A

See poster by Maphuti Madiga, MOPEA003 - “Development of intra- and inter-subtype cross-neutralizing antibodies in HIV-1 subtype C infection”

CAP256 80%

0 25 50 75 100 125 15010

100

1000

10000

100000 CAP256 (AUTOLOGOUS)

Weeks PI

ID50

Development of the autologous neutralizing response in CAP256

Note: CAP256 was super-infected at about 13 weeks p.i. – Carolyn Williamson et al, unpublished data

0 25 50 75 100 12510

100

1000

10000CAP25684-256-84 V1V2CAP84

super-infection(13 w eeks p.i.)

weeks p.i.

Tit

re

Specificity of the autologous neutralizing response in CAP256

Use of heterologous chimeric envelopes suggest an initial (unknown) specificity, followed by an anti-V1V2 response

0 25 50 75 100 125 15010

100

1000

10000

100000

CAP210

ZM53M

ZM109F

ZM233M

WITO4160

Q23

+6CAP256 (AUTOLOGOUS)

Weeks PI

ID50

Development of the heterologous neutralizing response in CAP256

0 25 50 75 100 125 15010

100

1000

10000

100000

CAP206

CAP210

CAP239

CAP63

CAP255

Du156

Du422

ZM53M

ZM109F

ZM197M

ZM233M

WITO4160

6535

Q23

+6 +9CAP256 (AUTOLOGOUS)

CAP45

Weeks PI

ID50


0 25 50 75 100 125 15010

100

1000

10000

100000

CAP8

CAP61

CAP206

CAP210

CAP239

CAP228

CAP63

CAP255

CAP84

Du156

Du422

ZM53M

ZM109F

ZM197M

ZM233M

WITO4160

6535

Q461

Q23

Q842

Q168

+6 +9 +2 +1 +4CAP256 (AUTOLOGOUS)

CAP45

Weeks PI

ID50


See poster by Maphuti Madiga, MOPEA003 - “Development of intra- and inter-subtype cross-neutralizing antibodies in HIV-1 subtype C infection”

CAP256(autologous)

0 25 50 75 100 12510

100

1000

10000CAP25684-256-84 V1V2CAP84

super-infection(13 weeks p.i.)

weeks p.i.

Tit

re

CAP45

0 25 50 75 100 12510

100

1000

10000

100000

CAP84

CAP45 G384-45-84 V1V2

weeks p.i.

Tit

re

CAP63

0 25 50 75 100 12510

100

1000

CAP88 B5

CAP63 A988-63-88 V1V2

weeks p.i.

Tit

re

CAP210

0 25 50 75 100 12510

100

1000

10000

100000

CAP8484-210-84 V1V2CAP210

weeks p.i.

Tit

re

CAP239

0 25 50 75 100 12510

100

1000

10000

CAP84

CAP23984-239-84 V1V2

weeks p.i.

Tit

re

CAP255

0 25 50 75 100 12510

100

1000

CAP84

CAP25584-255-84 V1V2

weeks p.i.

Tit

re

Specificity of the heterologous neutralizing response in CAP256 mirrors the specificity of the autologous

response

V1V2 involved in epitope V1V2 not involved in epitope

Conclusions

One or two potent but highly-type specific neutralizing antibody specificities develop in the first year of infection

The C3 and V1V2 regions are frequent targets, and changes in these regions directly mediate escape

These antibody specificities arise sequentially with titers waning as escape occurs





Conclusions

Autologous nAbs were temporally associated with decreased viral load in CAP88, which was abrogated by escape mutations

In CAP256, antibodies mediating autologous neutralization contributed to heterologous neutralization and this involved determinants in the V1V2 region.





Acknowledgements

CAPRISA 002 Salim Abdool Karim (PI, UKZN)Koleka Mlisana (co-chair, UKZN)Carolyn Williamson (co-chair, UCT)CAPRISA 002 study team

NICD Lynn Morris Nthabeleng RanchobeBronwen LambsonElin GrayMaphuti MadigaEleanor CaveSarah CohenMary Phoswa

UCTCarolyn WilliamsonMelissa Rose-AbrahamsGama BandaweFlorette Treurnicht

CAPRISA is supported by the National Institute of Allergy and infectious Disease (NIAID), National Institutes of Health (NIH) (grant# AI51794), the National Research Foundation (grant # 67385), the Columbia University-Southern African Fogarty AIDS International Training and Research Programme (AITRP) funded by the Fogarty International Center, NIH (grant # D43TW00231) and a training grant from LifeLab, a biotechnology centre of the South African Government Department of Science and Technology.Columbia University - Southern Africa Fogarty AIDS International Training and Research Program (AITRP)





dr penny moore

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