Selection of CTL escape mutations is determined by both the ability to avoid CTL

recognition and by minimizing the impact on viral replicative capacity

Arne Schneidewind

Partners AIDS Research Center

Boston

HLA-B27HLA-B27

HLA-B27 restricted Epitope KK10

L268M is an early, partial escape mutation

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

R264K is the dominant escape mutation

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

R264K is accompanied by S173A

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

R264K

L268M

S173A

Replicative capacities of KK10-variants

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

Replicative capacities of KK10-variants

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

No reduction in RC for L268M

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

R264K dramatically reduces RC

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

S173A restores RC of R264K

pNL4-3 V I P M F S A L S E G // P P I P V G E I Y K R W I I L G L N K I V R M YL268M . . . . . . . . . . . // . . . . . . . . . . . . . . M . . . . . . . . .R264K . . . . . . . . . . . // . . . . . . . . . . K . . . . . . . . . . . . .S173A . . . . . A . . . . . // . . . . . . . . . . . . . . . . . . . . . . . .

265 270 275170 175 255 260

Replicative capacities of KK10-variants

Impact on replicative capacity and requirement for

compensation delays escape in B27-KK10

Replicative capacities of KK10-variants

Are there alternative ways to escape the CTL pressure against KK10?

Replicative capacities of KK10-variants

Alternative Escape Mutations in KK10

R264T

Alternative Escape Mutations in KK10

Escape Mutations in KK10 abrogate binding to HLA-B27

Alternative Escape Mutations in KK10

Replication capacity of SARKLM > than replication capacity of alternative escape mutants

Simulation of CTL pressure

Simulation of CTL pressure

Simulation of CTL pressure

-------------------------

-------------------------

Conclusions

In VitroReplication Capacity

In Vivo Replication Capacity

Time Time

(R264T/Q/G)KTWIIL/MGLNKKQWIIL/MGLNKKGWIILGLNK

(R264K)KKWIIL/MGLNK

(WT)KRWIIL/MGLNK

(SARKLM)(SARKLM)KKKKWIIMGLNKWIIMGLNK

+ S+ S173173AA

HLA BindingVirus

strong

weak

very weak

weak Rep

licat

ion

Rep

licat

ion

Rep

licat

ion

Rep

licat

ion

Conclusions

- All CTL escape mutations at position P2 of KK10 efficiently abrogate binding to HLA-B27.

- Replicative capacity of the frequent R264K escape mutation is dramatically reduced.

- S173A efficiently compensates for the R264K replicative defect.

Conclusions

-The compensated R264K mutant (including S173A) replicated more efficiently than alternative escape mutations in the presence of KK10-specific CTL pressure.

- Replication capacities in vitro correlate well to the frequency of escape mutations in vivo.

-An in vivo threshold determines the propensity for escape variants to be selected.

AcknowledgementsPartners AIDS Research Center: California State University, San Marcos:

Mark A. Brockman Bianca R. Mothé

Yaoyu E. Wang

Huabiao Chen

Todd J. Suscovich La Jolla Institute for Allergy and Immunity:

Bin Li

Rahma I. Adam John Sidney

Rachel L. Allgaier Alessandro Sette

Thomas Kuntzen

Cesar Oniangue-Ndza

Christian Brander Funding:

Bruce D. Walker

Todd M. Allen R01-AI054178 + R21-AI067078