Current Clinical Therapies for HIV
Remission
David Margolis MDUNC HIV Cure Center
Cohen J. Science 2014
Aiming for sustained remission off ART
Luzuriaga et al. NEJM 372;8: 786
Effect of vorinostat, hydroxychloroquine and maraviroc combination therapy on viremia
following treatment interruption in individuals treated
during acute HIV infection (Fiebig III-IV)
Eugène Kroon, Jintanat Ananworanich, Keith Eubanks,
Jintana Intasan, Suteeraporn Pinyakorn, Nicolas Chomont,
Sharon R Lewin, Sarah Palmer, Lydie Trautmann, Hua Yang,
Nitiya Chomchey, Nittaya Phanuphak, Ken Cooper,
Praphan Phanuphak, Mark de Souza
on behalf of the SEARCH 019 study group
HIV Rebound after ART Interruption
Median time to first VL detection: 22 days (range 14 to 77 days)
ART resume at VL > 1000 c/ml
Stochastic
Reversal
of Latency
Chronically producing
cell
Host cell modification
KO: CCR5 (Sangamo)KI: sh5/C46 (Calimmune)
• Challenges moving forward:– Is cytoreductive therapy needed?
Acceptable?
– Is there X4 escape?
– Scalability? Cost?
• CCR5-modified CD4 T cells at 1 week post infusion constituted 13.9% of
circulating CD4 T cells
• Modified cells had an estimated mean half-life of 48 weeks
• After ART interruption, decline in circulating CCR5-modified cells (−1.81 cells
per day) was significantly less than the decline in unmodified cells (−7.25 cells
per day) (P = 0.02)
• HIV RNA became undetectable in one of four patients who could be evaluated
A first step to eliminate
latent HIV infection
Latency
Reversal
A second step to eliminate
latent HIV infection
Latency Reversal and Clearance Candidates
• Latency Reversing Agents (LRAs)
– HDAC inhibitors• “lack potency and killing as single agents” (Mellors)
• “HDAC inhibitors do not kill non-transformed cells at clinical exposures” and “Appropriate serial dosing regimens of HDAC inhibitors have not yet been performed” (Margolis)
Apologies, too many references to cite!
Pt. 1 Pt. 2 Pt. 3 Pt. 4 Pt. 5 Pt. 6 Pt. 7 Pt. 80
20
40
60
200
400
600
800
100
Baseline ART
VOR 400 mg
Re
lativ
e H
IV-1
ga
g R
NA
co
pie
s
2012!
Rasmussen Lancet ID 2014
Thrice weekly cycles of Panobinostat
**!**!
**!**!*!
Days!
Lewin CROI 2013!14 daily doses of vorinostat! Elliot&PLoS&Path&2014&
Sogaard IAS 2014
Total CD4 cell-associated unspliced HIV-1 RNA!
Weekly Romidepsin !
Sogaard&PLoS&Path&2015&
Latency Reversal and Clearance Candidates
• Latency Reversing Agents (LRAs)
– HDAC inhibitors• “lack potency and killing as single agents” (Mellors)
• “HDAC inhibitors do not kill non-transformed cells at clinical exposures” and “Appropriate serial dosing regimens of HDAC inhibitors have not yet been performed” (Margolis)
Apologies, too many references to cite!
– PKC agonists• most potent activators but toxicity of concern
• Bryostatin clinical trial did not achieve effective drug exposures
– TLR agonists: • activate HIV expression and immune control in SIV/macaques
TLR7 Agonists Induce Transient Plasma Viremia
Time after TLR7 agonist dose (Hours)
102
103
Pla
sm
a S
IV R
NA
co
pie
s/m
l
Pla
sm
a S
IV
(lo
g1
0R
NA
co
pie
s/m
L)
V1
V2 0
24
48
72
16
8 02
44
87
2
16
8 02
44
87
2
16
8 02
44
87
2
16
8 02
44
87
2
16
8 02
44
87
2
16
8 02
44
87
2
16
8 02
44
87
2
16
8 02
44
87
2
16
8 02
44
87
2
16
8
102
103
Vehicle
V1
V2 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168 0
24
48
72
168
Vehicle
Placebo GS-986 (0.1
mg/kg)
GS-9620 (0.05 mg/kg) GS-9620 (0.15
mg/kg)
• Reduced frequency of blips 38-75% (doses 3-10)
• No more blips from animals dosed after 3 month
pause (doses # 11-19)
Whitney et al., CROI 2016
Phase I/II trial of GS-9620 in HIV-
infectionDesign:
• 3 escalating dose cohorts– 1 mg, 2 mg, 4 mg every 2 weeks for 6 doses
• Placebo-controlled, randomized, double-blinded (6 active, 2 placebo per cohort)
Study Population:
• HIV-infected adults (n=24)
• Virologically suppressed ≥12 months on ART
Study Monitoring:
• Close follow-up – VL 2-3x/w
• Repeat dosing only if VL <50 copies/mL
• Safety review prior to initiation of each cohort
Current status: Enrolling Cohort 3
Latency Reversal and Clearance Candidates
• Latency Reversing Agents (LRAs)
– HDAC inhibitors• “lack potency and killing as single agents” (Mellors)
• “HDAC inhibitors do not kill non-transformed cells at clinical exposures” (Margolis)
– PKC agonists• most potent activators but toxicity of concern
• Bryostatin clinical trial did not achieve effective drug exposures
– TLR agonists: • activate HIV expression and immune control in SIV/macaques
Apologies, too many references to cite!
– Other Epigenetic targets• Bromodomain inhibitors
• Histone methyltransferase inhibitors (eg. EZH2 inhibitors)
• Others in development
– High-throughput library screening efforts
Merck HIV Latency HDAC Inhibitor
Synergy Screen
HIV LTR Induction
With 250nM SAHA2,900,000 Compounds
Confirmatory Assays, n=3HIV LTR Induction
With 250nM SAHA
Dose Response Assays, n=2
HIV LTR Induction
Without SAHA
HIV LTR Induction
With 250nM SAHA
NFkB BLA Reporter
Counter Screen
Toxicity @ 48 Hours
CTG
Data Analysis/
Hit Selection
Follow-up
Analysis~4,500 Compounds
Initial Screen, n=3
Objective: To identify compounds that potentially act synergistically
with HDACis to induce HIV transcription
Compounds with unknown mechanism
Known HDAC InhibitorsKnown Farnesyl-Transferase Inhibitors
66.5%16.1%
17.4%
Testing interventions in vivo
ART *After
intervention
• Leukapheresis for QVOA and ca-HIV RNA
• SCA• Immune assays• Host cell assays & biomarkers• Novel assays, eg. Quanterix
Simoa
• Leukapheresis for QVOA and ca-HIV RNA
• SCA• Immune assays• Host cell assays & biomarkers• Novel assays, eg. Quanterix
Simoa
Baseline
LRAsImmunodulators
HIV vaccinesNovel approaches
Reduction in:• Resting CD4
cell infection• Low-level
viremia
HIV RNAin cells or culture
HIV DNA, RNA, antigen & viruses
Ho, Cell 2013
Ericksson, PLoS Path 2013
HIV Antigen
(protein)
detector
The “Real”
Reservoir
HIV DNA
QVOAReplication-
competent
virus
1 2 3 4 5 6 7Time on ART (years)
-
0.0001
0.001
0.01
0.1
1
10
100
0
Fre
quency
(per
10
6cells
)
0.00001
Margolis, Garcia, Hazuda, Haynes Science 2016
Time to eradication > 73.4 years
Persistent HIV infection despite ART
Residual
Replication
or Cell
Proliferation
Viral
Activation &
Clearance
or Cell Death
Crooks et al. JID 2015
• Given assay variance, a more than 6-fold RCI decrease would have likelihood 0.023 (2.3%)
• Therefore a measurable goal is therapy that can reduce the latent reservoir by half a log
Challenges to clearing persistent infection after latency reversal
• Recent absence of antigen – low frequency of HIV-specific antiviral responses
• Immune dysfunction, deletion, or exhaustion• Archived viral diversity, including immune escape• Viral antigen is rare, dispersed,
compartmentalized, and may be transient• Latency Reversing Agents (LRAs) are host-
targeted, and alone or in combination may alter antiviral immune response
Latency Reversal Agent Discovery
• New metric of viral antigen production or
presentation
- Antigen required to allow clearance
• Assessment of LRA effect on immune function as
part of development path
- Immune function required for clearance
Quanterix Simoa™ Technology
Quanterix Simoa Platform• Ultrasensitive platform (sub-pg/mL sensitivity)
• Full automation (samples in – data out)
– Rapid readout (~1 hr), >500 data points per day
• Broad dynamic range (>4 logs)
• Commercial p24 assay applied to plasma and serum (Chang L, et al. J Virol Methods. 2013;188(1-2):153-160.)
• In-house Merck assay optimized to reduce nonspecific binding and enhance sensitivity, thus enabling p24 quantitation in cell lysates
1. Single-protein molecules are captured and labeled on individual beads using standard ELISA reagents
2. Beads + substrate are loaded in individual femtoliter wells. Oil added to seal well
3. Digital or analog fluorescence readout of individual beads
http://www.quanterix.com/Howell et al. submitted
CD8+
LatencyReversing
Agent
PBMCs
No Effectors
or or
AddEffectors
LimitingDilution
Co-culture
Measure HIV
Productionat 2 weeks
CD8+ by negative selection
CultureResting CD4+cells
Resting CD4+ cells bynegativeselection
RemoveEffectors
CD8+,HXTCs
,or
DARTs
Latency Clearance Assay
Patient
425Patient
532Patient
250
0
20
40
60
80
100
120
% V
ira
l re
cover
y
Patient
423
Patient
250
Patient
231
Patient
492
Patient
532
Patient
425
0
1
2
3
4
5
6
7
8N
um
ber
of
posi
tive w
ells
(ou
t of
12
tota
l)
No Effectors
CD8HXTC
††
††
†
Patient 425
PHA
VOR
†p<0.05 compared to No Effector
††p<0.05 compared to BOTH
HXTCs Reduce Recovery of Virus from autologous
resting CD4+ T cells stimulated with:
††
††
†
†
Sung et al. JID 2015
Latency Reversal and Clearance Candidates• Natural and Engineered Antibodies
– Broadly-neutralizing monoclonal antibodies (bnMAbs)• Can delay rebound and promote cell clearance in humans (3BNC117)
• Resistance can rapidly develop (VRC01, 3BNC117)
• Effect in individuals on ART? (VRC01)
– Engineered bnMAb• Can prolong half-life and enhance Fc effector functions (e.g. PGT-151)
– Bispecific Ab (anti-HIV/anti-host, e.g. CD3 or CD16)• Enhance effector function ex vivo and in animal models
Apologies, too many references to cite!
A5342/VRC01 Study• Double-blind, randomized, placebo-controlled, Phase I study
• 40 participants (20 per arm)
• VRC01 40 mg/kg IV at Day 0 & 21 (Arm A) or Day 42 & 63 (Arm B)
Trial Completed and Analyses Underway!
• SIV-infected monkeys were treated with a 90-day course of ART initiated 5 weeks post infection
• 9 weeks post infection infused with primatizedmonoclonal antibody against the α4β7 integrin every 3 weeks until week 32
• All animals subsequently maintained low to undetectable viral loads and normal CD4+ T cell counts for more than 9 months, after all treatment was withdrawn.
• Human trial underway at NIH
Dual Affinity ReTargeting (DARTs) Molecules for HIV
• Do not require pre-existing HIV specificity• Not impacted by archived CTL escape variants
• Anti-Env arm based on well characterized mAbs that have: • Are broad neutralizing antibodies -- bind virions and infected cells: DH542 (V3
glycan bnAb), CH557 (CD4bs bnAb), DH511-K3 (gp41 MPER bnAb)• Are ADCC mediating antibodies -- bind only infected cells: (7B2, gp41
immunodominant), A32 (C1)
Dual Affinity Re-Targeting proteins direct T cell –mediated cytolysis of latently HIV-infected cells.Sung, JA, Pickeral, J, Liu, L, Stanfield-Oakley, SA, Lam, CY, Garrido, C, Pollara, J, LaBranche C. Bonsignori, M. Moody, MA, …..Haynes, BF, Nordstrom JL, Margolis, DM, Ferrari, G. JCI 2015
Sung et al.:• Screened ADCC, non-neutralizing
Abs that bound HIV-infected CD4 T cells for optimal ADCC
• Constructed DARTs with non-Neutralizing mAbs A32XCD3 and 7B2XCD3
• Showed DARTs + CD8 CTL eliminated HIV-infected CD4 T cells in vitro by CD8 T cell-mediated cytolysis.
Sloan et al.:• Constructed bnAb PGT145 (V1V2),
PGT121 (V3 glycan), VRC01 (CD4 bs), and 10E8 (distal MPER)---compared to A32 and 7B2 non-neutralizing DARTs
• Best were PGT121, A32 and 7B2 DARTs
Targeting HIV Reservoir in Infected CD4 T cells by DARTs that bind Envelope and Recruit CTLs. Sloan DD et al. PLoS Pathogens, 2015
HIVxCD3 DART-mediated virus clearance in 4 of 4 patients (longer time needed for Pt 795)
V O R -E x p o s e d L a te n tly In fe c te d C e lls
6 7 4 4 0 8 4 0 7 7 9 5 7 9 5
0
2 0
4 0
6 0
8 0
1 0 0
1 2 0
P a tie n t
2 4 h r c u ltu re w ith D A R T s 9 6 h r
%v
ira
l re
co
ve
ry
(no
rm
ali
ze
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ffe
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rs
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N o E ffe c to rs
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C D 3 x4 4 2 0
C o m b o D A R T s
00
Sung, et al. JCI 2015
HIVxCD3 DART Mediated Clearance of Resting
Patient CD4 Cells Exposed to Vorinostat
• Immune Checkpoint Blocking Antibodies– Major advance in cancer immunotherapy
– Reverse immune exhaustion
– Examples: Anti-PD-1/PD-L1, LAG-3, 2B4, CD160, TIM-3, others
• Cellular therapies– CD8+T-cells with chimeric antigen receptors
– Ex-vivo Effector cell expansion/re-infusion
– Activated NK cells
• Therapeutic Vaccines– Multiple approaches
– Chimp Adeno vector, CMV vector, VSV vector, Ad26/MVA vectors, Dendritic cells
– Can induce broad CTL responses
Covering immune escape variants is critical!
Apologies, too many references to cite!
Latency Reversal and Clearance Candidates
Tools to Test Latency Reversal and
Clearance
Autologous DC vaccineMinimal effect of VOR on immune function
Clutton
Sci Rep 2016
Baseline Multidose
0
5 0 0
1 0 0 0
1 5 0 0
2 0 0 0
2 5 0 0
p < 0 .0 0 0 1
HIV
-1
ga
g R
NA
co
pie
s p
er
we
llArchin
Keystone Symposium on
HIV Persistence 2016
Persistent LRA activity of multiple VOR doses
Combination Latency Reversal and
Clearance Trial I
Step4
AGSEOS
Term~Week15-
Week24
~Week73-
Week89
Visit8 27
AGSManufacturing EOS
visits-nodosing Leukapheresis&rc-RNA IMandProviral Viralinhibitionandlatencyclearanceassays
visits-Vorinistatdoses Safetylabs SCA Acetylation
visits-AGS-004injections PKSamples ExploratoryImmunology
rc-RNAmeasurementdeterminesprogress
SingleDose
Step7
AGSDosing
Step8
MultipleVORDosing
Step1
Screen
Step2
Enrollment
Step3
Interval
Step5
AGSDosing
Step6
MultipleVORDosing
~Week39-Week54
Visits14-17 Visits18-22
~Week52-Week73
Visits23-26
~Week64-Week81
PairedDoses
Approximately
Weeks0-8
InjectionsX4
IntervaldosingX10doses
InjectionsX4
IntervaldosingX10doses
~Week27-Week46
Visit9-13
BASELINE
~Week6-Week13
Visits1&2 Visit3&4
~Week10-Week
18
Vists5-7
Vaccine Vaccine
LRA x 10
Vaccine = Argos dendritic cell therapy
LRA = vorinostat
LRA x 1 LRA x 2LRA x 10
Combination Latency Reversal and
Clearance Trial II
ART
STEP1PretreatmentPhaseExhibit Ex-vivoresponsetoVOR
STEP2:SingleVOR400mgdose
STEP3:HXTCsproduced
STEP4:FirstCombinationtreatmentVORevery72hoursx10dosesand2HXTCInfusions
RESTPERIOD
STEP5:FirstCombinationtreatmentVORevery72hoursand2HXTCInfusions
STEP 6:ImmuneResponsemonitoring
~4weeks ~8weeks
~6-8weeks
4weeks 2-4weeks
4weeks ~45weeks
Visits12 33.1 45 6789101112 13141516171819 20 2122232425
ImmuneResponseAssays HXTCInfusion LeukapheresisSingleCopyAssayOtherResearchAssays
VORevery72hoursx10doses
VORevery72hoursx10doses
cART
STEP1PretreatmentPhaseExhibit Ex-vivoresponsetoVOR
STEP2:SingleVOR400mgdose
STEP3:HXTCsproduced
STEP4:FirstCombinationtreatmentVORevery72hoursx10dosesand2HXTCInfusions
RESTPERIOD
STEP5:FirstCombinationtreatmentVORevery72hoursand2HXTCInfusions
STEP 6:ImmuneResponsemonitoring
~4weeks ~8weeks
~6-8weeks
4weeks 2-4weeks
4weeks ~45weeks
Visits12 33.1 45 6789101112 13141516171819 20 2122232425
ImmuneResponseAssays HXTCInfusion LeukapheresisSingleCopyAssayOtherResearchAssays
VORevery72hoursx10doses
VORevery72hoursx10doses
cART
HX
TC
HX
TC
HX
TC
HX
TC
HX
TC
Cath Bollard
Clio Rooney
Steps to eliminate
HIV infection
Latency
Reversal
Finally: the addition of durable vaccine protection if rebound or reexposure occurs