Surveillance of IL-2 inducing CD4+ T cell epitopes in
This presentation is the property of its rightful owner.
Sponsored Links
1 / 16

Surveillance of IL-2 inducing CD4+ T cell epitopes in PowerPoint PPT Presentation


  • 70 Views
  • Uploaded on
  • Presentation posted in: General

Surveillance of IL-2 inducing CD4+ T cell epitopes in acute HIV-1 infection for the emergence of escape mutants. R. Brad Jones 1 , Feng Yun Yue 2 , Colin Kovacs 3 , Ruqaya Mohamed 2 , Kelly Macdonald 1,2 , Mario Ostrowski 1,2. 1 Department of Immunology, University of Toronto

Download Presentation

Surveillance of IL-2 inducing CD4+ T cell epitopes in

An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -

Presentation Transcript


Surveillance of il 2 inducing cd4 t cell epitopes in

Surveillance of IL-2 inducing CD4+ T cell epitopes in

acute HIV-1 infection for the emergence of escape mutants

R. Brad Jones1, Feng Yun Yue2, Colin Kovacs3,

Ruqaya Mohamed2, Kelly Macdonald1,2,

Mario Ostrowski1,2

1 Department of Immunology, University of Toronto

2 Clinical Sciences Division, University of Toronto

3 Canadian Immunodeficiency Collaborative


Surveillance of il 2 inducing cd4 t cell epitopes in

Introduction

  • CD4+ T cell responses are critical in control of other chronic viral

  • infections including: gamma herpesvirus (Cardin, 1996),

  • LCMV and vaccinia (Leist, 1989)

  • Strong HIV-specific CD4+ T cell proliferation is maintained only

  • in long-term nonprogressors (LTNP), (Pontesilli, 1999)

  • Vigorous HIV-1-specific IL-2 producing CD4+ T cell responses

  • are associated with control of viremia (Rosenberg, 1997)

  • Is this cause or effect? High level HIV-1 viremia suppresses viral

  • antigen specific CD4+ T cell proliferation (McNeil, 2001)

  • Prospective study suggests that IL-2 producing CD4+ T cell

  • response to gag does not have prognostic value for rate

  • of progression to AIDS (Miedema, 2006)


Surveillance of il 2 inducing cd4 t cell epitopes in

Delineating Cause and Effect

  • HIV-1 is capable of rapidly acquiring mutations which confer

  • escape from selective pressure

  • We see this with gp120 mutations which escape antibody

  • responses, drug-resistance mutations, and certain CD8+

  • T cell responses

  • If CD4+ T cells are capable of exerting immunological

  • pressure on HIV-1, we should see the emergence of CD4

  • epitope escape mutations


Surveillance of il 2 inducing cd4 t cell epitopes in

Subject: OM214

  • Acute seroconverter - symptomatic: fever, rash

HAART


Surveillance of il 2 inducing cd4 t cell epitopes in

Methods

Cloning:

  • Sample obtained from leukopheresis

  • After CD8+ depletion, cells were stimulated overnight with p55

  • Enrichment for HIV-p55 specific CD4+ T cells achieved with IL-2

  • secretion assay (Miltenyi) and MACS

  • Plated at limiting dilution with irradiated feeder cells

  • p55 specificity was screened by ELISPOT and confirmed by FACS

Determining Eptiope Specificity of Clones:

  • Gross specificity determined by overlapping gag peptide pool

  • ELISPOT and confirmed by FACS

  • Minimal epitope determined using truncated peptides


Surveillance of il 2 inducing cd4 t cell epitopes in

Results

Gag p17:

Gag p24:

“HIVWASRELER”

“FRDYVDRFYK”

Gag p24:

“REPRGSDIAGT”


Surveillance of il 2 inducing cd4 t cell epitopes in

HLA Restriction

  • ELISPOTs repeated with core peptides in presence of either anti-DQ,

  • anti-DR, or isotype controls

Peptide + clone +

B cell line

Clone +

BCL

Clone +

BCL + anti-DR

Clone +

BCL + anti-DQ

  • Two clones from OM214 ‘MREPRGSDIAGT’ and ‘FRDYVDRFYK’ are DQ

  • restricted

  • Specifically ‘FRDYVDRFYK’ is restricted by DQB1*05011/DQA1*010101


Surveillance of il 2 inducing cd4 t cell epitopes in

Epitope Responses in ex vivo PBMCs

Month 2:

Control

“FRDYVDRFYK”

Autologous p55

“REPRGSDIAGT”

“HIVWASRELER”

0.025

0.021

0

0.053

0.015

IL-2

CD69

Month 12:

Control

“FRDYVDRFYK”

Autologous p55

“REPRGSDIAGT”

“HIVWASRELER”

0.005

0

0

0

0

IL-2

CD69


Surveillance of il 2 inducing cd4 t cell epitopes in

Sequencing

  • Performed on circulating plasma viruses

  • Limiting dilution methodology with direct sequencing

  • from PCR product

  • Sequences screened for hypermutation

  • Phylogenetic trees constructed to ensure that patient’s

  • sequences cluster together


Surveillance of il 2 inducing cd4 t cell epitopes in

Types ofMutations Observed

Mutations in Core Epitope

Extended Epitope/Processing Mutations


Surveillance of il 2 inducing cd4 t cell epitopes in

Frequencies of Mutations Observed


Surveillance of il 2 inducing cd4 t cell epitopes in

Frequencies of Mutations Observed


Surveillance of il 2 inducing cd4 t cell epitopes in

Epitope Mutation

Clone A2:

FRDYVDRFYK

FRDYVDQFYK

55.9

0

PBMCs:

Control

FRDYVDQFYK

FRDYVDRFYK

0.006

0.025

0


Surveillance of il 2 inducing cd4 t cell epitopes in

Flanking Mutations

  • Clone and express autologous p55 with and without

  • flanking mutations

  • Test ability to stimulate clones

Flanking mutations(FM)

wt

2ug/ml

FM p55

2ug/ml

SUMO-CAT

2.5ug/ml

wt p55

10ug/ml

p55

1ug/ml

p55

10ug/ml

p55

1ug/ml

p55

Med

Med

SEB

SEB

Clone A2

‘FRDYVDRFYKT’

Clone B2

‘REPRGSDIAGT’

  • Flanking mutations do not confer escape to A2 or B2


Surveillance of il 2 inducing cd4 t cell epitopes in

Conclusions

  • Rapid progression occurred in OM214 despite early

  • induction of an IL-2 producing CD4+ T cell response -

  • including a potent MHR-directed response

  • Loss of IL-2 secreting CD4+ T-cell response accompanied

  • disease progression

  • An escape mutation in an IL-2 inducing CD4+ T cell

  • epitope within the MHR was confirmed

  • This mutation arose within 6 months of infection and was

  • maintained at a frequency of 10%, for at least 1 year

  • IL-2 producing CD4+ T cell responses are capable of

  • exerting immune pressure on HIV-1, resulting in escape

  • mutations

  • Generalized loss of IL-2 responses with time suggests that

  • immune dysfunction due to viremia is an important

  • mechanism for viral escape from immune pressure


Surveillance of il 2 inducing cd4 t cell epitopes in

Acknowledgments

Elizabeth Yue

Mario Ostrowski

Maple Leaf Clinic:

Colin Kovacs

Roberta Halpenny

Macdonald Lab:

Ruqaya Mohamed

David Willer

Kaul Lab

Sample Donors


  • Login