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A KNOCKOUT FOR CURE? BENIGNO RODRÍGUEZ, MD, MSC, FIDSA
DISCLAIMERS
¡ Consulting fees: Gilead
¡ Funding: NIH, Merck, Gilead
OUTLINE
¡ Challenges of HIV eradication and/or durable control
¡ Survey of approaches in clinical phases of study
¡ Use of CCR5 disruption as a reservoir-targeting strategy
nAbs
CD4 Abs
CKRAs FIs
NRTIs NNRTIs
INSTIs
PIs
MIs
0.00001 0.0001 0.001 0.01 0.1
1 10
100 1000
10000
0 1 2 3 4 5 6 7 8 Time on HAART (years)
Late
ntly
infe
cted
cel
ls (I
UP
M)
-
t 1/2 = 44 months Eradication time = 73 years
WHY NOT JUST “WAIT OUT” THE CELLS WITH INTEGRATED VIRUS?
SILICIANO ET AL., NATURE MEDICINE 9, 727-8, 2003
LIMITATIONS OF LONG-TERM ART
¡ Toxicity
¡ Stigma
¡ Emergence of resistance
¡ Fatigue and non-adherence
¡ Cost
CHALLENGES OF ELIMINATING THE HIV LATENT RESERVOIR
No productive replication and no viral protein expression
Sanctuary sites, host genome integration, immune silence/exhaustion
Prolonged survival with continual reseeding
EXPOSE CLEAR PROTECT
• Latency-reversing agents
• CTL enhancement • ADCC (inc. passive Ab transfer) • Therapeutic vaccination • Checkpoint inhibitors • Focused immune modulation • Viral genome excision? • Immune system replacement?
• Gene therapies • Extended half-life
nAbs • Early ART
After Mellors
• IL-2 • TLR7 agonists • Combinations
HIV genome
Latency reversal
‘Shock’ ‘Kill’
Immune system
Dying infected
cell
ART
HIV proteins
1. Deeks SG. Nature 2012;487:439–40. 2. Walker-Sperling VE, et al. J Virol 2015;89:9631–8.
HIV CURE APPROACH “KICK AND KILL”
HITTING ALL THREE: THE RIVER STUDY DESIGN
Individual with defined PHI
Primary outcome: total proviral DNA in CD4+ T cells
Secondary outcomes
Undetectable viral load
Randomisation
ART only ART + V + V
Vaccines
HDACi
Immediate standard ART (irrespective of CD4) + integrase inhibitor
Approx. 24 w
eeks 18 w
eeks
Vaccination: Prime ChAdV63.HIVconsv at randomisation Boost MVA.HIVconsv week 8 post-randomisation Vorinostat 400mg od every 72 hours* total 10 doses
*Archin et al J Clin Invest 2017 Aug 1;127(8):3126-3135.
FIDDLER S, ET AL. IAS 2018 A-899-0059-12977
RIVER STUDY: ART + V + V DOES WHAT IT IS SUPPOSED TO DO…
10
Overall difference in mean change between the arms: p = 0.038
Ahmed et al Vaccine.
2016 Feb 24;34(9):1215-24.
FIDDLER S, ET AL. IAS 2018 A-899-0059-12977
…BUT NOT WHAT WAS HOPED IT WOULD DO
11
ART only ART +V+V
Primary endpoint: Difference (ART+V+V minus ART only) in mean log10 HIV DNA copies/million CD4+T cells averaged across PR weeks 16 and 18: 0.04 (95% CI: -0.03 to 0.11); p=0.26
FIDDLER S, ET AL. IAS 2018 A-899-0059-12977
JUST TREAT EARLY
¡ Features of early HIV Infection:
¡ Low viral genetic diversity
¡ Preserved (?) cellular immune response capacity
¡ Small established reservoir
¡ The Mississippi baby Persaud D, et al. N Engl J Med 2013
¡ VISCONTI cohort Sáez-Cirión A et al. PLoS Pathog 2013
¡ Not reproduced in the San Diego Primary Infection Cohort Gianella S et al ADIS 2015
¡ Induction of elite controller phenotype in macaques by combination bNABS Nishimura et al. Nature 2017
BLOCKADE OF A4B7: INITIALLY PROMISING
MACAQUES HUMANS (n=18)
DI MASCIO M ET AL. IAS 2018 TUAA0206LB & FAUCI A, IAS 2018 PLENARY
¡ Other immunomodulatory strategies: ¡ TLR7 agonists ¡ PD-1, S1P1 blockade ¡ Interferons
BNABS DELAY VIRAL REBOUND
¡ Combination human bNAbs administered during acute SHIV Infection in macaques led to elite controller phenotype in some monkeys Nishihima Y et al. Nature 2017
¡ Effect was mediated by CTL responses (viremia rebounded after CD8 depletion)
BAR KJ, ET AL. N ENGL J MED 2016
INTERVAL SUMMARY
¡ The presence of a long-lived latent HIV reservoir containing integrated replication-competent HIV genomes is the major obstacle to HIV eradication
¡ Strategies targeting latency, the immune response, the virus itself, or combinations thereof, have been tested in humans
¡ But MOSTLY have not led to demonstrable reservoir reduction or durable control of HIV replication
IL-2: FORSAKEN, BUT NOT FORGOTTEN
¡ IL-2: A potent lymphocyte growth factor that increases:
¡ Proliferation of T- and B-cells
¡ Cytolytic activity
¡ NK cell function
¡ Total CD4+ T cell counts in HIV infection and other settings…
¡ But does not protect against clinical HIV disease progression
INSIGHT–ESPRIT STUDY GROUP & SILCAAT SCIENTIFIC COMMITTEE. N ENGL J MED 2009
IL2: SOME INTERESTING OBSERVATIONS
¡ In pooled analyses of clinical trials, recipients of IL-2+ART had more profound VL decline than ART-alone recipients
¡ In a subset of clinical trials participants, replication-competent HIV was detected more often in ART-alone recipients
¡ Despite this, two participants who stopped ART rebounded
CHUN TW ET AL. PROC NATL ACAD SCI USA 1997; EMERY S ET AL. J INFECT DIS 2000
ADCC: WHAT IT IS, WHY IT MATTERS
Target cell
Punch 1: Activate
cell
Punch 2: Target
response
Punch 3: Turbocharge
response
IL-2 GETS BY WITH A LITTLE HELP FROM ITS FRIENDS
¡ A controlled clinical trial of rIL-2 +/- VRC07 for suppressed HIV infection (n=20)
¡ General criteria:
¡ Age 18-65
¡ CD4+ T cell count >350
¡ Plasma HIV RNA BLD x 1 year
¡ No active Hep B or Hep C
¡ Generally healthy
¡ Regimen:
¡ Eight 5-day cycles of subcutaneous rIl-2, twice a day, 8 weeks apart
¡ (+/-)VRC07 infused once IV at the beginning of each cycle
¡ Primary endpoint: Reservoir size
INTERVAL SUMMARY
¡ A strategy that delivers EXPOSE + CLEAR + PROTECT may be feasible
¡ No clinical data to predict whether or not such an approach will have a measurable effect
¡ Clinical trial expected to open to accrual in Fall 2018 in Cleveland
EARLY EVENTS IN HIV CELL ENTRY: ROLE OF THE CCR5 CORECEPTOR
CCR5 AND ITS “KNOCKOUT”
¡ 32 bp deletion in the CCR5 gene results in defective protein
¡ But few detectable adverse clinical consequences
¡ Persons homozygous for this knockout are almost completely resistant to HIV infection
¡ And those who are heterozygous might have slower disease progression
¡ About 1% of Caucasians are born with two defective alleles
¡ If it could be produced on demand, it would be the ultimate “PROTECT” weapon
¡ But still would need the “CLEAR”
ENTER TIMOTHY BROWN, THE MAN WHO ONCE HAD HIV
¡ HIV+ patient (CCR5 wt/Δ32) suppressed on HAART developed AML and failed chemotherapy
¡ During first induction, HAART held and viremia rebounded.
¡ After recurrence, conditioned with: TBI; amsacrine, fludarabine, cytarabine, cyclophosphamide, ATG)
¡ Allo stem cell transplant (CCR5 Δ32/Δ32 donor); HAART held.
¡ Mild GVHD (treated with immune suppressants)
¡ Plasma levels of virus remained undetectable off HAART. No viral RNA or DNA in blood, BM, rectal biopsy, meningeal biopsy.
¡ CD4 T cell counts rose, serum Abs to gag/pol proteins decreased and T cell responses to HIV fell to background levels
¡ He remains well without evidence of virus recurrence more than 9 years after transplant and off antiretrovirals HÜTTER G ET AL. N ENGL J MED 2019
¡ 2 HIV+ pts with lymphoma
¡ Allogeneic stem cell transplantation (donors CCR5 wt/wt)
¡ On HAART 2.6 and 4.3 yrs with ¡ No HIV detected in plasma
¡ No HIV DNA found in multiple assays of 5 million CD4 cells
¡ No HIV DNA in rectal bx of one patient
¡ ART withdrawn
¡ Initially, no virus or viral sequences in blood or tissues. HIV Antibody levels fell
¡ But viral rebound after 12 and 32 weeks off ART
¡ Virus persisted somewhere at low frequency
SCT AS AN ERADICATION STRATEGY: SUBSEQUENT EXPERIENCE
HÜTTER G. N ENGL J MED 2014
SCT AS AN ERADICATION STRATEGY: SCALING IT UP - ICISTEM
WENSING AMJ, ET AL. AIDS 2018
• 37 patients registered from 9 different countries • 30 patients transplanted • Mean follow-up: 887 days
12 patients deceased after SCT (Δ32/Δ32) 12 patients beyond 2nd year post-SCT
ICISTEM: EFFECTS ON THE RESERVOIR
1
10
100
1000
10000
-100 0 100 200 300 400 500 600 700
numbe
rofH
IVDNAcopies(log(cop
ies/millioncells))
timebefore/afterSCT(days)
Ultra-sensitiveHIVDNAquantificationintotalPBMCDNA
WENSING AMJ, ET AL. AIDS 2018
INTERVAL SUMMARY
¡ Homozygosity for the d32 mutation confers almost absolute protection against HIV Infection
¡ And successful complete replacement of wt cells with d32 may result in sterilizing cure of HIV Infection
¡ Which components of the intervention are responsible is unclear: induction? transplantation? GVHD?
ZFP
ZFP
§ Comprises two domains:
- Nuclease domain of FokI restriction enzyme - Engineered zinc finger protein (ZFP) provides DNA binding specificity - Targets 12 nucleotides each for a total of 24-bps of DNA
§ ZFN cleaves genomic DNA as a heterodimer within a 5-6 bp gap between the two binding domains
§ Repair typically renders a deletion in the target gene § Introduction of this vector into host target cells or host stem cells can render the target
or stem cell progeny resistant to HIV infection (if both gene copies are knocked out)
DNA
ZINC FINGER NUCLEASES CAN TARGET HOST ELEMENTS AND DELETE THEM
ADOPTIVE TRANSFER OF CCR5 GENE MODIFIED AUTOLOGOUS CD4+ T-CELLS
• This immunotherapy is minimally invasive, with no severe adverse events
• It is more
accessible than HSCT; it removes the need of searching for compatible donors and the risks associated with GVHD
Enrich CD4+
Infusion
SB-728 CCR5 ZFNs
CCR5 gene disruption
Apheresis
Expand, formulate and test
MedianCCR5modification
~25%Single
InfusionofSB-728-T
ZFN
ZFN
DNA
HIV+subjects
INFUSION OF AUTOLOGOUS CD4 CELLS WITH MODIFIED CCR5
Dramatic CD4 rises – mechanism? Modified cells persisted durably (were they protected?)
Tebas et al NEJM ‘14
SB-728-T INFUSION LEADS TO HIV RESERVOIR DECAY
Ø Decay of the reservoir is a continuous, on-going process
BLyr
2-310
100
1000
10000
Inte
grat
ed H
IV D
NA
(cop
y/10
6 CD
4+ T
cel
ls) 1-01
1-021-032-012-022-033-013-023-03
0.0039
Integrated HIV DNA
BLyr
1yr
210
100
1000
10000
100000
Tota
l HIV
DN
A (c
opy
/106 P
BM
C)
Total HIV DNA
0.0547
0.6523
0.0195
COURTESY OF R. P. SÉKALY
Ø CCR5 mutations that were unique to TSCMs in the infused product were identified using DNA-Seq and tracked in all CD4+ memory subsets at year 3-4
Ø Mutations unique to TSCM were detected in other sorted memory T cell subsets including short lived TEM indicating that TSCM cells could differentiate into other subsets
% C
CR
5 m
ut. a
t yr 3
-4 (
3881
uni
que
mut
. in
R
Ain
t RO
int C
CR
7+C
D27
+ pr
oduc
ts)
TCM TTM TEM
CD45RAintROint TSCM CD45RA+ TSCM
ZFN-INDUCED MUTATION TRACKING CONFIRMS PERSISTENCE AND DIFFERENTIATION CAPACITY OF TSCM
COURTESY OF R. P. SÉKALY
INTERVAL SUMMARY
¡ Reinfusion of autologous ex vivo CCR5-modified CD4+ T cells results in robust CD4+ T cell increase, reduction of the size of the HIV reservoir, and an apparent survival advantage for cells that express a TSCM-like phenotype
¡ Whether this is the result of dilution from infused cells, and whether the replication-competent reservoir is affected, is less clear
T-CELL REINFUSION AFTER INTERFERING WITH LYMPHOCYTE BINDING LOCATION OF AIDS VIRUS THROUGH ZINC-FINGER-NUCLEASE ELIMINATION OF CCR5 RECEPTORS: THE TRAILBLAZER STUDY
THE TRAILBLAZER STUDY
¡ Phase 2 double-blind randomized clinical trial of expanded and CCR5-modified vs. unmodified autologous CD4+ T cells
¡ Will address mechanisms of CD4 gain and reservoir reduction
¡ Extensive tissue sampling
¡ LN
¡ CSF
¡ Rectal mucosa
¡ 24 month primary F/U to assess durability
¡ No treatment interruption
¡ Suppressed patients, 18 to 65, generally healthy
¡ Expected to open to accrual in early December in Cleveland
Screening Obtaininformedconsent,screeninglabs,determineeligibility
Pre-leukapheresis Obtainpre-leukapheresis laboratorytestsasrequiredpersite’sstandards
Blockrandomize
Arm2N=10
UnmodifiedCD4+Tcells
Leukapheresis Perform2.5bloodvolumeleukapheresis,obtainreservoirs izemeasurementsfromleukapheresisproduct
BaselineRoutinelabs.Rectalbiopsy,lymphnodebiopsyorFNAinsubsetofparticipants.NOTE:Tissuespecimencollectionnotnecessarilyonsamedayasotherbaselineevaluations.
Cyclophosphamidedos ing
Pre-doseIVhydrationandpremedication,cyclophosphamideadministration,post-doseIVhydrationandmonitoring.
Intervalfollow-upvis i ts:Day1,day3,week1,week2,week4,month2,months4,6,9
Variousfollow-upassessmentsincludinglumbarpunctureatweek4insubsetofparticipants,refertosection1.3.
Month12Variousfollow-upassessments,2.5bloodvolumeleukapheresis.Rectalbiopsyinsubsetofparticipants.NOTE:Rectaltissuespecimencollectionandleukapheresis notnecessarilyonsamedayasothermonth12evaluations.
Intervalfollow-upvis i ts:Months15,18,21
Variousfollow-upassessments,refertosection1.3.
Month24Finalassessments,2.5bloodvolumeleukapheresis.Rectalbiopsy,lymphnodebiopsyorFNAandlumbarpunctureinsubsetofparticipants.NOTE:Tissue/CSFspecimencol lectionandleukapheresisnotnecessarilyonsamedayasothermonth24evaluations.
Phase1:Prepa
ratory
Phase2:In
fusio
nan
dFollo
w-up
Infusionofstudyproduct,post-infusionmonitoring.Infusion
Arm1N=20
Modi fiedCD4+Tcells
2nd pre-leukapheresis Obtainpre-leukapheresis laboratorytestsasrequiredpersite’sstandards
3rd pre-leukapheresis Obtainpre-leukapheresis laboratorytestsasrequiredpersite’sstandards
CONCLUSIONS
¡ No clinically available intervention has been shown to induce durable control of viral replication or eradication of HIV Infection, with precisely one exception
¡ Strategies targeting just one aspect of the reservoir persistence problem appear to be insufficient
¡ A multi-stage approach to stimulate cells to proliferate, be targeted by bNAbs, and be exposed to enhanced cytotoxicity could prove useful
¡ The phenotype achieved by Timothy Brown can be partially mimicked, without the risk of SCT
¡ Whether there is a specific threshold of CCR5 modification that will replicate the effect seen in Timothy Brown is unknown
¡ Clinical trials evaluating the strategies above will open this year in Cleveland
BACKUP SLIDES
FTY 720 (FINGOLIMOD) ADMINISTRATION RETAINS T CELLS IN LYMPH NODES IN RHESUS
THE HIV RESERVOIR IS NOT ENTIRELY QUIESCENT
COHN ET AL, CROI 2018 #152LB
IN RHESUS MACAQUES TREATED WITH ART EARLY IN INFECTION, COMBINATION THERAPEUTIC VACCINE AND TLR 7 AGONIST HELP TO CONTROL VIREMIA AFTER TREATMENT DISCONTINUATION
Borducchi et al Nature 2016
BCN 02 STUDY
• A rollover from a prior trial that administered two therapeutic vaccines & had started ART within three months of HIV infection
• Participants receive vaccine plus Romipdesin
Better Control of Viremia BCN 02 (Mothe & Brander)
HIV+
8/13 ART
STOP
weeks
4-5/13 ART
STOP
weeks
Mothe et al. presented at CROI (2017)
• Five out of 13 recipients of the interventions have since interrupted ART and displayed control of viral load to low levels for several months (the longest a little over six months)
• The frequency of viral load containment in the cohort (~38%) is higher than has been observed in any studies involving early initiation of ART (where rates have varied from 0-15%).
• The contribution of romidepsin is unclear due to the lack of any control group
BCN 02 - Results
46
Analysis of the primary endpoint
Model N Difference in log10 total HIV-DNA at weeks 16/18: Arm
B vs Arm A
SE 95% CI p-value
a. unadjusted 60 0.107 0.128 (-0.148-0.363) 0.404
b. Adjusted for baseline total HIV-DNA 58 0.039 0.035 (-0.032-0.109) 0.279
c. Adjusted for baseline total HIV-DNA & stratum 58 0.039 0.036 (-0.033-0.110) 0.282
d. as c), with multiple imputation of missing baseline values
60 0.041 0.036 (-0.031- 0.113) 0.256
e. as c), excluding patients with incomplete intervention
55 0.039 0.037 (-0.035-0.113) 0.292
47
------------------------------------------------------------------------------ | Change Std. Err. t P>|t| [95% Conf. Interval] -------------+---------------------------------------------------------------- rx | Arm A | -0.11 0.03 -4.27 0.000 -0.17 -0.06 Arm B | -0.07 0.02 -2.93 0.005 -0.12 -0.02 ------------------------------------------------------------------------------ overall | -0.09 0.02 -5.12 0.000 -0.13 -0.06 ------------------------------------------------------------------------------
Total HIV-DNA: estimated change from baseline
Change from randomisation to PR Week 16/18 : Total HIV-DNA log10 copies/106 CD4 cells
48
Viral outgrowth assay
Model N IUPM at week 16:
Arm B vs Arm A
SE 95% CI p-value
a. Median regression 38 -0.04 0.13 -0.31 - 0.23 0.743
Model N Log10 IUPM at week 16:
Arm B vs Arm A
SE 95% CI p-value
b. Interval regression 56 0.19 0.22 -0.25 - 0.62 0.402
Note: Models adjusted for stratum
DISCUSSION
¡ Did we measure HIV reservoir too soon?
¡ Did we use the best outcome measure of ”cure”
¡ Was this the wrong approach, what is the future for Kick and Kill?
