Viral Tissue Reservoirs Are Determined Early and Little Viral

RNA Is Detected during Suppression in the

Macaque ModelZandrea Ambrose, Ph.D.

Division of Infectious Diseases, University of Pittsburgh School of

Medicine, Pittsburgh, PA, USA

“Towards an HIV Cure” Pre-Conference Symposium20 & 21 July 2012

• infection, pathogenesis and immunity can be similar to that of HIV-infected humans

• procedures can be performed more easily and/or ethically than in humans• removal of vital organs at necropsy• acquisition of longitudinal samples • changing/testing novel treatment regimens

• model can be controlled in a way that humans cannot• known infecting virus, timing, and route of infection• known treatment adherence, past regimens, and timing of

treatments• scheduled blood and tissue sampling

Macaque Models of HIV/AIDS

HIV-1 Does Not Replicate Well in Monkeys

HIV-1

SIVLTRLTR gag

PRenv

vif

vpx vpr

revtat nefRT IN

LTRLTR gagPR

env

vif

vpr vpu

revtat nefRT IN

RT-SHIVLTRLTR gag

PRenv

vif

vpx vpr

revtat nefINRT

Study Objectives

• Determine which tissues harbor viral DNA that could contribute to the reservoir• How early are these reservoirs established?• Are the levels different after suppressive therapy?

• Determine which tissues contain viral RNA• Are multiple sites contributing to viremia in untreated

animals?• Is there detectable viral RNA in the tissues of ART-

suppressed animals?

RT-SHIV Plasma Viremia

Therapy

1

2

3

4

5

6

7

0 5 10 15 20 25 30 35Weeks post-challenge

Vira

l RN

A co

pies

/ml p

lasm

a (lo

g)

PT8272PT8030GN19GG45GV08GV40

TNV/FTC/EFV

TNV/FTC/EFV/L-870812

Therapy

No Therapy

RT-SHIV gag DNA vs. Host Cell DNA (CCR5)

Viral DNA is Mainly in Lymphoid and Gut Tissues

gag DNA copies/106 CCR5 DNA copies in each tissue

1

10

100

1000

10000

10000 100000 1000000 10000000

Week 1 Plasma Viral Load

Lym

phoi

d Ti

ssue

vD

NA

(nor

mal

ized

to 1

06 CC

R5)

0.996p < 0.0001

Week 1 Plasma Viremia Correlates with Lymphoid Viral DNA Levels

Untreated ART-Treated

RT-SHIV gag DNA copies in PBMC

Little Viral RNA Detected in Tissues of Suppressed Animals

gag RNA copies/106 CD4 RNA copies in each tissue

PBMC Viral RNA Level is not Reflective of Plasma Viremia or Most Tissues

6760

8433

8232

GT29GR65

8272

8030

GN19GG45

GV08GV40

1

10

100

1000

10000

100000

1000000

10000000PBMCPlasma

vRN

A (p

er 1

0^5

cells

or m

l pla

sma)

6760

8433

8232

GT29GR65

8272

8030

GN19GG45

GV08GV40

1

10

100

1000

10000

100000

1000000PBMCPlasma

vRN

A (p

er 1

0^5

cells

or m

l pla

sma)

Untreated ART-Treated Untreated ART-Treated

Week 1/2 Week 28/30

• Viral DNA is detected in many tissues and PBMC of untreated and ART-treated macaques- < 4% of viral DNA from 2-LTR circles

• Levels of viral DNA in lymphoid tissues correlated with week 1 plasma viremia- reservoir size is established early after infection

• Tissue viral RNA levels were lower and less frequent in ART-treated macaques- No replication-competent virus was detected in LN resting

CD4+ T cells of ART-treated animals treated with 4 (< 0.5 IUPM)

• Viral RNA levels in PBMC were not consistent with treatment or plasma viremia

Conclusions

AcknowledgementsUniversity of Pittsburgh

Chris KlineJean NdjomouTamera Franks

John Mellors

AIDS & Cancer Virus Program, NCIJeff LifsonJake Estes

Mike Piatak, Jr.Jeremy Smedley

Vicky CoalterRebecca Kiser

MerckDaria HazudaJulio Quintero

Gilead SciencesNorbert Bischofberger

Detection of Replication-Competent Virus from Lymph Node CD4+

Lymphocytes

R2 = 0.997

CCR5 Standard Curve

R2 = 0.987

gag Standard Curve

RT-SHIV gag DNA vs. Host Cell DNA (CCR5)