Satish K. Pillai, Ph.D.

MODULE LEADER

Module R:Recording the HIV Reservoir

Satish Pillai

TimothyHenrich

MichaelSamuels

ChristosPetropoulos

BenjaminFranc

MichaelBusch

HenryVanBrocklin

SheilaKeating

SulggiLee

AhmedTawakol

PriscillaHsue

amfAR Institutefor HIV Cure Research 

Module R Team

Fundamentals of HIV persistence

Untreated HIV infection

HIV-infectedcell

Virus

Uninfected cellsNewly-infected cells

Fundamentals of HIV persistence

Treated HIV infection

HIV-infectedcell

Virus

Uninfected cellsNewly-infected cells

• Long-lived, “latently-infected” cells are invisible to the immune system and may survive for years during ART

When antiretroviral therapy is stopped, the virus returns

Time

Vira

l loa

d

Antiretroviral therapy

We need tools to predict IF and WHEN the virus will rebound

?Time

Vira

l loa

d

Antiretroviral therapy

HIV reservoir size determines timing of viral rebound

Time

Vira

l loa

d

Antiretroviral therapy

Time

Vira

l loa

d

Antiretroviral therapy

HIV reservoir size determines timing of viral rebound

We need a revolution in reservoir recording

We need more sensitive recording tools

Challenge #1:HIV persists at low levels during ART

HIV-infected cells are extremely rare in treated individuals

Fewer than 10 out of a million CD4+ T cells may harbor HIV during antiretroviral therapy

(Abdel-Mohsen et al, AIDS 2015)

We will find more needles by sampling more of the haystack.

We are using digital droplet technology that enables analysis of 10-fold more cells and DNA than standard approaches

We can now detect a single copy of HIV DNAin a million CD4+ T cells using digital PCR

Uninfected primary cells(background)

Cells containing

a single copy of HIV

+

HIV DNA

HIV + Human DNA

HumanDNA

No DNA

We are using digital PCR to understand how HIV cure strategies affect both virus and host

(Adapted from Abdel-Mohsen et al, PLoS Pathogens 2016)

Unstimulated CD4+ cells Cells treated with Galectin-9

• Galectin-9 potently reactivates latent HIVand induces host immunity against HIV

MohamedAbdel‐Mohsen

Challenge #2:Most viruses are defective and cannot grow

HIV has a high mutation rate that enables immune escape and evolution of drug resistance

HIV genomes can be classified into three types

• Infectious, replication competent virus

We are developing a microfluidic chip that will allow us to efficiently measure infectious virus in a clinical sample

Traditional quantitative viral outgrowth assays (“QVOA”) are extremely expensive and laborious

HIV genomes can be classified into three types

• Benign, defective viral genomes

HIV genomes can be classified into three types

• Inflammatory, defective viral genomes

?

Circulating antibodies in blood may inform us about HIV persistence in tissues

Challenge #3:HIV persistence in tissues is likely critical

• The host immune response against HIV may give us critical info about the size of the latent HIV reservoir

Antibodies against HIV may “see” virus in tissues and predict time until viral rebound

We are using in vivo imaging to visualize the latent HIV reservoir in infected individuals

Antibody

Radiolabel

HIV

HIV antibody radiolabeling approach

Dr. VanBrocklin and team have labeled VRC01 (broadly neutralizing antibody against HIV) with zirconium-89 and are gathering the data for an FDA

Investigational New Drug (IND) submission in Q1 2017

89Zr‐VRC0189Zr‐Pembroluzimab

We have performed FDG-PET/CT imaging of lymph nodes to measure the HIV reservoir in humans

(Tawakol and Hsue, in press 2016)

Metabolic activity in lymph nodes is associated with HIV reservoir size

HIV Imaging Roadmap

Biomarker Development

18FDGTSPO

18F‐Raltegravir18F‐GE‐18018F‐AraG

89Zr‐VRC‐01PD‐1/PD‐L1TLR

Direct Disease Marker

Today      

5 years

The work we have done to identify a reservoir “biomarker”

The work we have done to identify a reservoir “biomarker”

The work we have done to identify a reservoir “biomarker”

The work we have done to identify a reservoir “biomarker”

The HIV reservoir may be more “active” than we thought

Analytical treatment interruption will help us to build accurate reservoir recording tools

Time

Vira

l loa

d

Antiretroviral therapy

Collect specimens and record measurements

Analytical treatment interruption will help us to build accurate reservoir recording tools

Time

Vira

l loa

d

Antiretroviral therapy

Our recording tells us the reservoir is depleted