Understanding the potential mechanisms of HIV cure by stem ...€¦ · Hospital Düsseldorf), Rolf...
Transcript of Understanding the potential mechanisms of HIV cure by stem ...€¦ · Hospital Düsseldorf), Rolf...
Understanding the potential
mechanisms of HIV cure by stem cell
transplantation
Annemarie Wensing, MD, PhD
University Medical Center Utrecht
WITS RHI University of the Witwatersrand Johannesburg
• HIV-DNA persists as viral reservoir
• If ART is interrupted HIV rebounds from this reservoir
Besson, G. J. et al. Clin. Infect. Dis. 2014
HIV-DNA remains despite ART
https://www.semanticscholar.org/paper/Are-T-cells-the-only-HIV-1-reservoir-Kandathil-Sugawara
HIV-RNA = viral load after start ART
HIV-DNA after start ART
Time (days) Time (years)
Proviral DNA in CD4 T-cells
CD4 T-cells
Where is this reservoir?
The reservoir is not only present in blood
but throughout the body
gp41
gp120
V3 loop
CD4 Binding
CD4
Co-receptor Binding
CD4 positive human cell
• HIV uses two human receptors to invade cells: CD4 and a co-receptor
Different co-receptors are used by HIV for
entry
CCR5
Hartley et al. Aids Res Hum Retroviruses 2005;21(2):171-89
CXCR4
R5
X4
Dual
Early in infection uses HIV the CCR5 receptor for entry = R5 troop
Late in infection HIV may start to use the CXCR4 receptor for entry = X4 troop
Cure Strategies
To limit the
establishment of the
reservoir
To reduce the size of
the reservoir
To control the
reservoir
Early
ART
Render uninfected
cells resistant to
HIV
Deplete infected
cells
Flush out the latent reservoir
Vaccine Immuno-therapy
Measuring the reservoir
Inspired by Nicholos Chomont. HIV reservoirs and strategies towards an HIV cure. HIV-NAT 2017
Markers of persistence Single Copy Assay: an extremely sensitive method that can even measures one copy of HIV-RNA at a time. Input: Plasma
https://www.projectinform.org/
PCR 2-LTR circles: ongoing replication qPCR/ddPCR can measure cell-
associated HIV RNA (caRNA) or total HIV DNA but does not discriminate between replication competent and defective HIV.
The Tat/Rev Inducible Limiting Dilution Assay (TILDA) is a polymerase-chain-reaction-based (PCR-based) assay that measures intracellular HIV RNA, no full distinction between replication competent variants or not
Bruner et al. Nature 2016
>95% of the measured HIV DNA is defective and
does not reflect replication competent virus
Markers of persistence
https://www.projectinform.org/
PCR 2-LTR circles: ongoing replication
Quantitative viral outgrowth assay (QVOA) is meant to measure replication-competent latent provirus. Input: CD4+ T cells retrieved by leukapheresis.
The Tat/Rev Inducible Limiting Dilution Assay (TILDA) is a polymerase-chain-reaction-based (PCR-based) assay that measures intracellular HIV RNA, no full distinction between replication competent variants or not
Applied techniques may overestimate or
underestimate the HIV reservoir
Challenges in qualifying the reservoir
• Only a small amount of CD4 T cells in PLWH on ART harbor HIV
• Intact or defect provirus
• Compartimentalization
• Which successes do we know?
Patient no more: The Berlin Patient
Stem cell transplantation procedure
https://www.miltenyibiotec.com/BE-en/products/cell-therapy/clinimacs-cd34-reagent-system-fda-approved/Patients/what-is-allogeneic-stem-cell-transplantation.html
= 100% chimera
Berlin Patient follow up
Gero Hütter et al. NEJM 2009
Stop HAART
18
wt/wt wt/32 32/32
“Resistant” to HIV infection
Normal R5 receptor
Heterozygote: less CCR5
Homozygote gene defect: no CCR5
Liu R, et al. Cell. 1996;86:367-367. Samson M, et al. Nature. 1996;382:722-725. Dean M, et al. Science. 1996;273:1856-1862. Huang Y, et al. Nat Med.1996;2:1240-1243. Michael NL, et al. Nat Med. 1997;3:1160-1162. Eugen-Olsen J, et al. AIDS. 1997;11:305-310.
1-2% of white people (Caucasians) do have a gene defect (Δ32 DNA) IN these people the CCR5 receptor is not present at the cell surface
Timothy Brown received stem cells with a
gene defect
How was Timothy Brown cured?
• Use of donor cells with genetic defect: CCR5Δ3232
• Absence of viruses able to use other co-receptors (i.e. X4 tropic-virus)
But
• Heavy Immune suppressive treatment (i.e. ATG levels)
• Total body irradiation
• Graft Versus Host Disease
• Two Stam Cell Transplantations
• Patient’s own CCR5 heterozygous state (smaller HIV-DNA reservoir?)
-
Gero Hütter et al. NEJM 2009
• Does this work for all patients who receive a SCT?
The Essen patient
Kordelas et al. N Engl J Med 2014 , Verheyen CID 2018.
• ART was stopped before SCT
• Treatment interruption led to rapid rebound of X4 tropic virus
Verheyen et al. CID, 2018
• clearly two distinct viral populations • 100% of the viral sequences detected after SCT were predicted to be X4-tropic • dominant X4-tropic viral sequence observed after SCT -present prior to SCT (4.4% of proviral DNA population) -present prior to treatment interuption
-287d/RNA/V03 -103d/DNA/V02
-103d/DNA/V11 -103d/RNA/V03 -103d/RNA/V04 -103d/RNA/V01 -103d/RNA/V05 -103d/RNA/V02
-18d/DNA/V05 -103d/DNA/V04
-103d/DNA/V10 -287d/RNA/V01 -103d/DNA/V01
-103d/DNA/V08 -18d/DNA/V06
-18d/DNA/V04 -18d/DNA/V02
-18d/DNA/V03 -287d/RNA/V02 -18d/DNA/V01
-103d/DNA/V03 -103d/DNA/V09
-18d/DNA/V07 -103d/DNA/V07
-103d/DNA/V06 -287d/RNA/V04
+373d/RNA/V06 +20d/RNA/V01 +373d/RNA/V02 -103d/DNA/V05 +373d/DNA/V01
+373d/RNA/V03 +373d/RNA/V08 +20d/RNA/V02 +373d/RNA/V01 +373d/RNA/V05 +373d/DNA/V02 +373d/DNA/V03 +373d/RNA/V04 +373d/RNA/V07
0.01
The Essen patient
Henrich, et al. Ann. Intern. Med. 2014
• Boston patients were transplanted with wildtype CCR5 donor cells
• Despite drastic reduction of the reservoir, HIV was able to rebound after ATI at 12 cq 32 weeks
The Boston patients
International collaboration to guide and investigate the potential for HIV cure in HIV-infected patients requiring allogeneic stem cell transplantation for hematological disorders
AIM 1 To guide clinicians involved in allogeneic SCT procedures in HIV infected individuals
AIM 2 To better understand the underlying biological processes leading to viral reservoir reduction and potential cases of HIV-1 eradication/remission.
www.icistem.org
Principal Investigators:
Javier Martinez Picado
Annemarie Wensing
Registration overview
Analyses of the dynamics of the viral reservoir
These unpublished data are not available at the website. For more information: See Salgado et al. https://www.annals.org/article.aspx?doi=10.7326/M18-0759
• How do we find out whether these patients has been
cured?
IciStem #19 The “Duesseldorf patient”
Stem cell transplantation with CCR5Δ32 Homozygous Allo-HSCT
ATI Since November 2018 without viral rebound
120
100
80
60
40
20
0
900 700 500 300 100 0
Jensen et al. CROI 2019.
• No proviral DNA detectable in blood, bone marrow, lymph nodes, liqour and rectum
• DNA and RNA scope positive (Jake Estes) • Western blot fading (antibodies)
IciStem #19 The “Duesseldorf patient”
Jensen et al. CROI 2019.
• DNA and RNA scope positive (Jake Estes)
IciStem #19 The “Duesseldorf patient”
IciStem #36 The “London patient”
Days post allo-HSCT
Gupta et al. Nature 2019
Stem cell transplantation with CCR5Δ32 Homozygous Allo-HSCT
Gupta et al. Nature 2019
IciStem #36 The “London patient”
CCR5 genotyping Pre-SCT: wildtype
Post-SCT: CCR5∆32
Immune responses post-SCT
Time (days) Gupta et al. Nature 2019
IciStem #36 The “London patient”
Overview CCR5∆32 homozygous HSCT
transplants
“Berlin patient” “London patient” “Duesseldorf patient”
CCR5∆ 32 heterozygous WT CCR5 homozygous WT CCR5 homozygous
R5 tropic HIV infection R5 tropic HIV infection R5 tropic HIV infection
Acute myeloid leukemia Hodgekin lymphoma Acute myeloid leukemia
2 HSCTs Single HSCT Single HSCT
Total body irradiation No irradiation No irradiation
Full intensity conditioning Reduced intensity
conditioning
Reduced intensity
conditioning
Mild GvHD (skin) Mild GvHD (skin/gut) Moderate GvHD (liver)
100% T-cell donor
chimerism
100% T-cell donor
chimerism
100% T-cell donor
chimerism
• Allogeneic Stem cell transplantation has a high mortality risk
• Only suitable for patients with a severe haematological malignancy
• Chances to find an adult matching donor for non Caucasian patients are extremely low
Is this approach safe and scalable?
Summary
• IciStem has identified > 40.000 CCR5Δ32 donors that can be used for allo-SCT also IciStem has compiled the largest registry of allo-transplants in people living with HIV
• After allo-SCT, a sharp decline in HIV DNA in the blood and
tissues is observed to below the level of detection in most transplanted patients
• Important determinants for cure after allo-SCT are: single
transplant CCR5Δ32 donor cells, no TBI, no full intensity chemo,
• The case of the Berlin Patient was not a miracle
• “Everything that happens once can never happen again. But everything that happens twice will surely happen a third time.” (Paulo Coelho)
IciStem research workgroup: Julià Blanco (Immunologist, AIDS Research Institute IrsiCaixa, Barcelona), Jorge Carrillo (Immunologist, AIDS Research Institute IrsiCaixa, Barcelona), Johanna Eberhard (Immunologist, University Medical Center Hamburg-Eppendorf), Mi Kwon (Hematologist, Hospital Gregorio Marañón), María Salgado (Virologist, AIDS Research Institute IrsiCaixa, Barcelona) Management team: Tineke Johnston and Josien Straver (Financial management/project assistance University Medical Center Utrecht), Laura Huyveneers (Medical coordination/IRB University Medical Center Utrecht), Pascual Balsalobre (Gregorio Marañón, Madrid), Judith Dalmau (AIDS Research Institute IrsiCaixa, Barcelona) Scientific Advisors: Koen van Besien (New York-Presbyterian hospital) IciStem participants Belgium: Linos Vandekerckhove, Marie-Angélique de Scheerder, Eva Steel (University of Ghent) Canada: Lisa Barrett, Sharon Oldford, Jill Moore, Clarissa Brisseau (NSHA/Dalhousie University, Halifax) Germany: Dieter Häussinger, Guido Kobbe, Björn Jensen (University Hospital Düsseldorf), Rolf Kaiser, Elena Knops (University of Cologne) Maximilian Christopeit (University Medical Center Hamburg-Eppendorf)
Italy: Alessandra Bandera, Antonio Muscatello and Alessandro Soria (San Gerardo Hospital, Monza, Italy), Gabriella Scarlatti, Simona Piemontese (Istituto Scientifico San Raffaele) Netherlands: Pauline Ellerbroek, Lodewijk Brosens, Anke Bruns, Erik van Maarseveen (UMC Utrecht) Jan van der Meer, Sacha Zeerleder (AMC), Jaap Jan Boelens (University Medical Center Utrecht/Memorial Sloan Kettering Cancer Center, New York) Spain: Jon Badiola, Manuel Jurado Chacón (Complejo Hospitalario Universitario de Granada), Raquel Saldana (Hospital General de Jerez de la Frontera), Luz Martín Carbonero (La Paz Hospital), Ildefonso Espigado (University Hospital Virgen del Rocío, Seville) Sweden Piotr Nowak (Karolinska Institutet) Switzerland Mitja Nabergoj, Alexandra Calmy (Hôpitaux Universitaires de Genève) United Kingdom: Kavita Raj, Fabio Cruciani, Varun Mehra, Carmel Rice (Kings College Hospital, London) Angela Bailey (Imperial College Healthcare NHS Trust, London), Waseem Qasim (Institute of Child Health & Great Ormond Street Hospital, London), Ravindra Gupta (University College London Hospital)
Acknowledgements The IciStem consortium
Javier Martinez-Picado (Co-PI,
Virologist, AIDS Research Institute IrsiCaixa, Barcelona)
Annemarie Wensing (Co-PI, Clinical
Virologist, University Medical Center Utrecht)
Jose L. Díez Martin (Hematologist,
Hospital Gregorio Marañón, Madrid)
Gero Hütter (Hematologist, Cellex
Dresden) Jürgen Kuball (Hematologist, University
Medical Center Utrecht)
Monique Nijhuis (Virologist,
University Medical Center Utrecht)
Vanderson Rocha (Hematologist
Cord Blood Bank Specialist Oxford University)
Asier Sáez-Cirión (Immunologist,Pasteur Institute, Paris) Julian Schulze zur Wiesch (Infectious disease specialist, UMC Hamburg-Eppendorf)
Translational Virology, UMC Utrecht