how does the reservoir influence the clinician's...
TRANSCRIPT
How does the reservoir influence the clinician's decisions
Prof. Carlo Federico Perno
Acknowledgements
University of Milan, Milan Italy: C. Alteri.
Policlinic of Rome Tor Vergata, Rome, Italy: M. Andreoni, L. Sarmati, M. Viscione, S. Gini, C. Cerva, V. Malagnino, K. Stingone,
T. Guenci, F. Stazi, S. Giannella, V. Serafini, M. Ciotti, P. Paba. S. Grelli.
INMI L Spallanzani, Rome, Italy: A. Antinori, R. Bellagamba, C. Pinnetti, S. Cicalini, R. Gagliardini, A. Mondi, A. Vergori, A.
Sanpaoloesi, G. De Carli, F.M. Fusco, L. Lo Iacono, M.L. Giancola, G. Liuzzi, R. Acinapura, P. Scognamiglio, N. Orchi, E. Girardi,
M.R. Capobianchi, C. Gori, F. Forbici, G. Berno, D. Pizzi, A. Giannetti, P. Lorenzini, A. Navarra, R. Libertone, G. Ippolito.
San Gallicano Hospital, Rome, Italy: A. Latini, M. Colafigli, M. Giuliani, A. Pacifici, A. Cristaudo. General Hospital Umberto I:
V. Vullo, G. D’Ettorre, F. Falasca, O. Turriziani, G. Antonelli. San Giovanni Addolorata Hospital, Rome, Italy: F. Montella, F. Di
Sora, W. Leti, F. Iebba. Sant’Andrea Hospital, Sapienza University, Rome, Italy: A. Pennica. Rebibbia, Rome, Italy: S. Marcellini.
Bambin Gesù Hospital, Rome Italy: S. Bernardi, H Tchidjou Kuekou. Polo Pontino, Sapienza University, Rome, Italy: C.
Mastroianni, M. Lichtner, V.S. Mercurio, C. Del Borgo, R. Marrocco. Frosinone Hospital, Frosinone, Italy: G. Farinelli, E.
Anzalone, M. Limodio, L. Sarracino. Rieti Hospital, Italy: G. Natalini Raponi, M.E. Bonaventura. Viterbo Hospital, Viterbo, Italy:
G. Maffongelli, G. Bernardini, A. Caterini, F. Ferri, A. Ialungo, E. Liguori, D. Migliorini, R. Monarca, R. Preziosi, E. Rastrelli, G.
Starnini, G. Sebastiani.
University of Turin, Turin, Italy: G. Di Perri, S. Bonora, A. Calcagno, V. Ghisetti, G. Vandemmiati, T. Allice.
Modena Hospital, Modena, Italy: C. Mussini, V. Borghi, W. Gennari, A. Cossarizza, M. Nasi, M. Di Gaetano.
Pescara General Hospital, Pescara, Italy: G. Parruti, F. Vadini, F. Sozio, E. Mazzott, T. Ursini, E. Polilli, P. Di Stefano, M.
Tontodonati, G. Calella. San Salvatore, L’Aquila, Italy: A. Grimaldi, A. Cellini. Ancona Hospital, Ancona, Italy: A. Mataloni
Paggi. Giuseppe Mazzini Hospital, Teramo, Italy: Di Giammartino, L. Falconi, P. Tarquini. San Salvatore – Muraglia- Hospital,
Pesaro, Italy: E. Petrelli, G. Corbelli, P. Tarquini. Avezzano Hospital, Avezzano, Italy: M. Paoloni, R. Mariani. AO Papa Giovanni
XXIII, Bergamo, Italy: F. Maggiolo, AP Callegaro. AO Careggi, Florence, Italy: K. Sterrantino.
Cotugno Hospital, Naples, Italy: A. Chirianni, M. Gargiulo. University of Campania Vanvitelli, Italy: S. Marini, N. Coppola.
Bisceglie-Trani Hospital, Bisceglie, Italy: R. Losappio. Catania Hospital, Catania, Italy: R. La Rosa. Enna Hospital, Enna, Italy:
L. Guarneri. Palermo Hospital, Palermo, Italy: F. Di Lorenzo T. Prestileo, A. Cascio.
The Patients
University of Rome “Tor Vergata”, Rome Italy: F. Ceccherini Silberstein, V. Svicher, A. Bertoli, M.C. Bellocchi, L. Fabeni, B. Yagai
Romeo, R. Salpini, R. Scutari, S. Barbaliscia, M. Brugneti, A. Biddittu, M. Bruni, L. Carioti, P. Saccomandi.
Unicamillus, Rome Italy: D. Armenia.
Thanks to the modern therapies, today around
95% of HIV infected individuals achieve
virological suppression
The success of antiretroviral therapy
allowed to a consequent dropping of
resistance development (at least in high income countries)
56%60% 60% 57%
51% 54%45%
41%46%
33%23%
27%21%
17% 16% 19% 17% 18%
0%
10%
20%
30%
40%
50%
60%
70%
80%
90%
100%
% o
f is
ola
tes
P<0.001 P=0.705
Among 9014 isolates from cART failing patients the prevalence of M184Vdramatically decreased from 56% in 1999 to 21% in 2013. In the last 5 years theM184V prevalence is stably settled at 17%.
Analysis performed on 9014 isolates from cART failing patients (Update August 2018)
Prevalence of M184V among isolates from cART failing patients over time
Armenia & Santoro, Unpublished data 2019
59%
0%0%
10%
20%
30%
40%
50%
60%
70%
80%
90%
100%
% o
f re
gim
en
sat
M1
84
V d
ete
ctio
n
Suboptimal
Prevalence of regimens used at the moment of M184V detection over time
In the past M184V was mainly detected in isolates under suboptimal regimensbased on NRTIs or unboosted PIs
P<0.001
Analysis performed on 3475 isolates for whom GRT revealed M184V from 2388 cART failing patients (Update August 2018)
Armenia & Santoro, Unpublished data 2019
• A new era for antiretroviral therapy is approaching
• New strategies are needed to maintain virus under control for decades, and preserve immune functions• Long-acting therapies• Simpler drug regimens
• while….
• Cure options are under study and clinical assessment
• The lower is the total viral burden, the higher is the chance that the patient could be eligible for a cure approach
The HIV hiding places
2016
S.L. Lamers 2016; 20:8968-83
229 varied autopsy tissues from 20 ART-treated patients with low or undetectableplasma viremia and cerebral fluid (CSF) VL prior to death, were analysed. HIV-DNA (>200 cp/106 cell) was identified in 48/87 brain tissues and 82/142 non-brain tissues. Abnormal histological findingswere identified in all partecipants (brain, spleen, lung, lymph node, liver, aorta and kidney).
Tissues assayed with the number of HIV+ (red) and HIV- (green) tissues identified
2019
HIV DNA was detected in most body tissues despite long-term ART and despite confirmed undetectable
HIV RNA in plasma at the time of death. The majority of full-length (FL) HIV-env sequences appeared to
be intact.
In the quest for a functional cure or eradication of HIV infection, we need to know howlarge the reservoirs are from which infection rebounds when treatment is interrupted.To that end, we quantified SIV and HIV tissue burdens in tissues of infected non-humanprimates and lymphoid tissue (LT) biopsies from infected humans. Before antiretroviraltherapy (ART), LTs harbor more than 98 percent of the SIV RNA+ and DNA+ cells.While ART substantially reduced their numbers, vRNA+ cells were still detectable andtheir persistence was associated with relatively low drug concentrations in LT comparedto peripheral blood. Prolonged ART also reduced the level of SIV and HIV-DNA+ cells, butthe estimated size of the residual tissue burden of 108 vDNA+ cells that potentiallyharbor replication competent proviruses, along with the evidence for continuing virusproduction in LT despite ART, identify two important sources for rebound followingtreatment interruption.The large sizes of these tissue reservoirs underscore the challenges in developing “HIVcure” strategies that target multiple sources of virus production
Jacob D Estes Nature Medicine 2017
Defining total-body AIDS-virus burden with implications for curative strategies
Graphical representation of the proportion of vRNA+ cells in eachorgan system before and during suppressive ART.
Jacob D Estes Nature Medicine 2017
Defining total-body AIDS-virus burden with implications for curative strategies
During ART the numbers of virus (v) RNA+ cells substantially decreasedbut remained detectable.
Graphical representation of the proportion of vRNA+ cells in eachorgan system before and during suppressive ART.
Jacob D Estes Nature Medicine 2017
Defining total-body AIDS-virus burden with implications for curative strategies
During ART the numbers of virus (v) RNA+ cells substantially decreasedbut remained detectable.
« Size » of the HIV reservoir
Ho et al. Cell 2013
The « real reservoir » ?
• The vast majority of
proviruses that persist on
ART are defective.
• These “ZOMBIE” proviruses (Imamichi, H. et al., International AIDS
Conference, 2014) lack the ability to produce intact viruses but can
inflict harm by producing foreign nucleic acids and proteins.
Persistence of these proviruses may explain the persistent
seropositivity to HIV-1 and persistent immune activation seen in
patients with "undetectable" virus.
• Of the minority proviruses that are intact (~2%), the
fractions that are latent or replicative competent are not
known.
Bruner et al. Nature 2019
DNA PCR assays predominantly measure defective proviruses. Proviruses persisting in CD4+ T cells of individuals
on suppressive ART as detected by nFGS (near full genome sequencing). The near full genome sequencing (nFGS) are
methods used identify defects throughout the genome except the 5′ long terminal repeat (LTR). Defects include internal
stop codons, deletions not attributable to normal length polymorphisms, and APOBEC3G/F mediated hypermutation
(G→A). Most deletions were large except for those in the packaging signal (ψ) or major splice donor site. Analysis is
based on 211 sequences from individuals initiating ART during chronic infection.
2018
The vast majority of acute transforming retroviruses are replicationdefective, with the oncogene-containing genome beingtransmissible only during mixed infection with a replication-competent virus. A defective retrovirus that relies on complementing functions can, in some instances, become replication competent by recombiningwith its replication-competent “helper.” In fact, there is some evidence that Rous sarcoma virus, possiblythe only naturally arising replication-competent retrovirus containing a host oncogene, was replication defective initially
The Remarkable Frequency of Human ImmunodeficiencyVirus Type 1 Genetic RecombinationA Onafuwa-Nuga and A Telesnitsky
Microbiology and Molecular Biology Reviews - 2009
As we observed for 8E5 in this study, genetic recombination
could generate replication-competent viruses from such a
collection of defective proviral sequences.
Infectivity of recombinant viruses
generated following transfection
of 8E5 cells with defective
molecular clones of HIV.
• The HIV-1 sanctuary: the meaning of
compartmentalization
Lee, et al Abstr 407, CROI 2017
• This is the first study to evaluate GALTand LN tissue concentrations in patientsreceiving RAL and 800 mg daily DRV.
• Tissue:plasma ratios were higher inileum>rectum as shown previously, andlowest in lymph node.
• In a limited number of participants,concentrations of RAL were significantlylower in lymph nodes vs. GALT, supportingprior observations.
• These results support the current limiteddata on tissue ART drug concentrations andhave potential implications on HIV curestrategies.
UntreatedTreated
RALT: rectum
COMPARATIVE
LYMPHOID
TISSUE
PHARMACOKINE
TICS OF
INTEGRASE
INHIBITORS.
C. V. Fletcher
CROI 2018
In the contest of long-term strategies, a deepen
evaluation of viral reservoir with biomarkers easy to be
measured in clinical practice is today fundamental!
In this frame, an evaluation of quantity of HIV-DNA (as a
measure of HIV reservoir) and the archived resistance
needs to be considered.
The quantification of total HIV-DNA in PBMCs provides
a reliable and easy way of measuring the size of the
cellular reservoirs of HIV.
Total HIV DNA is associated with other easier-
to-measure parameters in patients under
successful therapy
• Pre-therapy plasma HIV RNA
– Hocqueloux, JAC 2013; Lambert-Niclot, PLoS ONE 2012
• Residual viremia, even when simply classified as detectable vs.
undetectable
– Chun, JID 2011; Lambert-Niclot, PLoS ONE 2012; Mexas, AIDS
; Parisi et al., JCM 2012; Falasca et al., JAIDS 2015; Parisi, CMI
2015.
• Nadir CD4 counts
– Watanabe, BMCID 2011; Lambert-Niclot, PLoS ONE 2012
• Duration of suppression of plasma HIV RNA
– Watanabe, BMCID 2011
• Earlier treatment start
– Hocqueloux, JAC 2013
Ceccherini-Silberstein et al. , JAC Dec 2018
Pre-ART HIV-DNA correlates with pre-ARTHIV-RNA, CD4+ T-cells, CD4/CD8 ratio.
By considering the 397 patients achieving virological suppression, theprobability of experiencing virological rebound, defined by 2 confirmedplasma HIV-RNA >50 copies/mL, was 12% (95% CI: 8.6-15.5). By stratifyingpatients for the 3 different pre-ART HIV-DNA levels, increasing rates ofvirological rebound were found by increasing pre-ART HIV-DNA.
17.2%
7.4%4.3%
15.2%
4.8%
0%
Ceccherini-Silberstein et al. JAC 2018
HIV-1 DNA may be a strong predictor of :• Immunological progression (CD4 <350) in absence of
cART.
• Immunological progression (CD4 <350) following
treatment interruption.
• Time to viral rebound.
Williams, eLife 2014
Various studies have shown that the level
of baseline HIV-DNA can influence the maintenance
of virological success under simplification therapy
2013
Virological Factors Associated With Outcome of Dual ETR/RAL Therapy (ANRS-163 Trial)
Cathia Soulie, Lambert Assoumou, Sophie Sayon, Thuy Nguyen, Marc-Antoine Valantin, Virginie Ferre, Chakib Alloui, Brigitte Montes,
Véronique Avettand-Fenoel, Constance Delaugerre, Diane Descamps, Esteban Martinez, Jacques Reynes, Gilles Peytavin, Dominique
Costagliola, Christine Katlama, Vincent Calvez, Anne-Geneviève Marcelin.
PS6/4
FACTORS PREDICTING VIROLOGICAL FAILURE DURING DOLUTEGRAVIR MAINTENANCE MONOTHERAPYIngeborg Wijting, Sofie L. Rutsaert, Casper Rokx, David M. Burger, Elrozy Andrinopoulou1, Linos Vandekerckhove, Bart Rijnders
Defining a Total HIV DNA threshold as
guidance for therapy simplification strategies
S. Rutsaert1, I. Wijting2, W. De Spiegelaere3, L. De Clercq1, B. Rijnders2, L. Vandekerckhove1
HIV Cure and Research Symposium, Ghent
Therapy
simplification studies
PROTEA DOMONO
PROTEA▪ Substudy of the PROTEA (NCT01448707)
▪ Randomized clinical trial
HIV-1 infected patients
• First-line ART• VL undetectable• CD4 nadir >100
cells/mm3
• CD4 at baseline ≥200 cells/mm3
Triple therapy
DRV/r 800/100mg +2 NRTIs
Monotherapy
DRV/r 800/100mg N= 77
W48 W96
▪ Substudy of the DOMONO (NCT02401828)
▪ Randomized clinical trial
HIV-1 infected patients
• VL undetectable• CD4 nadir ≥200
cells/mm3
• HIV RNA zenith < 105
copies/ml
Monotherapy
DTG 50mg N= 77
W48
DOMONO
Undetected Detected Failers Undetected Detected Failers
PROTEA DOMONO
RESULTS
*
*
*
*
: p<0,05
Still……. a relevant clinical cut-off of total HIV DNA
needs to be defined in both drug-naïve and
virologically suppressed patients.
In drug-naïve patients starting a first-line regimen, the risk of virological
rebound was significantly higher in patients with a pre-cART total HIV-1 DNA
>10,000 copies/106 CD4+ T cells than in those with a total HIV-1 DNA ranging from
1,000 to 10,000 copies/106 CD4+ T cells and <1,000copies/106 CD4+ T cells.
Ceccherini-Silberstein et al., JAC 2018
In virologically suppressed patients who switched to a PI-sparing regimen,
higher HIV DNA levels (>226 copies/106 PBMCs) at baseline were independently
associated to an increased risk of virological failure or viral blip.
Sarmati et al., J Med Virol 2007
More data are required to set up specific cut offs relevant for the virological
outcome and significant progress in this area is awaited depending on
availability and widened use of standardized HIV DNA assays.
What about the impact of archived resistance in virologically
suppressed patients that need a therapy switch?
2016
RT resistance mutations detected by population sequencing (PS) and deep sequencing (DS), with indication of the number of patients in whom the mutations were present.
Of the 20 patients selected according to the Sanger sequencing results, 17 of them had valid RNA and DNAdeep sequencing results. The DS results shown are the results after read data processing and quality filteringbut before hypermutation cleanup. Mutations that result from a G-to-A transition are underlined.
Dauwe et al, J Clin Microbiol 2017
A
Patients with pre-existent NRTI+NNRTI resistance had a higher probability of
experiencing VR compared to those harboring pre-existent NRTI or NNRTI
resistance and to those without pre-existent RTI resistance.
Armenia et al., JAC Jan 2017
0 12 24 36 48 60 72
0.0
0.2
0.4
0.6
0.8
1.0
224 197 175 140252
12 24 36 480
Time (Weeks)
No. at risk
0.0
0.2
0.4
0.6
0.8
1.0
Pro
ba
bilit
y o
f e
xp
eri
en
cin
g
vir
olo
gic
al
reb
ou
nd
34 32 30 2839
12 9 7 718
9.4%11.5%
39.2%
p<0.0001
Pre-existent RTI-resistance before switching:
No resistance
NRTI or NNRTI
NRTI + NNRTI
102
60
18
7
75
72
13
6
The presence of historical M184V was associated with the risk ofexperiencing viral blips
Gagliardini et al Open Forum Infectious Diseases2018
Estimated probability of remaining free from VFin dual therapy for different time of viralsuppression
45Gagliardini et al Open Forum Infectious Diseases 2018
In an additional analysis selecting patients with equal to or less than3 years of viral suppression, the respective 1- and 3-year probabilities of remaining free from virological failure were 100.0% and 67.7% in the M184V+ group and 97.3% an 96.2% in theM184V- group (P = .002)
67.7%
96.2%
JAC 2018
Switch Strategies for Virologically Suppressed
Persons
A complete ARV history with HIV-VL,
tolerability issues and cumulative genotypic
resistance history should be analysed prior to
any drug switch……
October 2017
…..Together with duration of viral
suppression and the type of mutations (and
their fitness)!
1. Sarmati L, Nicastri E, Uccella I, et al. 2003. J Clin Microbiol 41:1760-2.
2. Parisi SG, Boldrin C, Cruciani M, et al. 2007. J Clin Microbiol 45:1783-1788.
3. Turriziani O, Bucci M, Stano A, et al. 2007. J Acquir Immune Defic Syndr 44:518-524.
4. Palmisano L, Galluzzo CM, Giuliano M. 2009. J Acquir Immune Defic Syndr 51:233-234.
5. Banks L, Gholamin S, White E, et al. 2012. J AIDS Clin Res 3:141-147.
6. Delaugerre C, Braun J, Charreau I, et al. 2012. HIV Med 13:517-525.
7. Bon I, Turriziani O, Musumeci G, et al. J Med Virol 2015 87:315-322.
8. Fabeni L, Berno G, Svicher V, et al. 2015. J Clin Microbiol 53:2935-41.
9. Gallien S, Charreau I, Nere ML, et al. 2015. J Antimicrob Chemother 70:562-565.
10. Lubke N, Di Cristanziano V, Sierra S, et al. 2015. Intervirology 58:184-189.
11. Gantner P, Morand-Joubert L, Sueur C, et al. 2016. J Antimicrob Chemother 71:751-61.
12. Michelini Z, Galluzzo CM, Pirillo MF, et al. 2016. J Med Virol. doi: 10.1002/jmv.24581.
13. Fernández-Caballero JÁ, Chueca N, Álvarez M, et al. 2016. BMC Infect Dis. 16:197.
14. Zaccarelli M, Santoro MM, Armenia D, et al. 2016. J Clin Virol 82:94-100.
15. Lambert-Niclot S, Allavena C, Grude M, et al. 2016. J Antimicrob Chemother. 71:2248-51.
16. Rodallec A, Le Guen L, Leplat A, et al. 2017. IAS. Abstract MOPEB0270.
17. Allavena C, Rodallec A, Leplat A, et al. J Virol Methods. 2018 Jan;251:106-110.
18. Armenia D, Zaccarelli M, Borghi V et al. J Clin Virol. 2018 Jul;104:61-64.
19. Boukli N, Boyd A, Collot M, et al. J Antimicrob Chemother. 2018 Aug 17.
20. Rodriguez C, Nere ML, et al. J Antimicrob Chemother. 2018 Aug 20.
21. Sotillo A, Sierra O, Martínez-Prats L, et al. J Virol Methods. 2018 Oct;260:1-5.
HIV DNA Genotypic Resistance Test is a good tool for therapy optimization
in both drug-naïve and drug-experienced patients
Journal of Clinical Virology 2016
Proportion of Patients with MRM in PBMCs and Cumulative Plasma(149 Patients with DNA GRT and ≥2 Plasma GRTs, 9 Patients for INSTI)
50.343
30.2
11.1
61.1
38.3
28.2
16.811.1
51
11.4 10.1
1.3 0.3
20.1
0
10
20
30
40
50
60
70
80
90
100
NRTI NNRTI PI INSTI Overall(PI/NNRTI/NRTI)
Re
sist
ance
pre
vale
nce
(%
) Mutations detected only in cumulative plasma
Mutations detected in PBMCs and cumulative plasma
Mutations detected only in PBMCs
1.29 (±1.67) 0.74 (±1.00) 0.77 (±1.39) 0.11 (±0.33) 2.79 (±3.26)
1.03 (±1.66) 0.38 (±0.72) 0.34 (±8.84) 0.44 (±1.33) 1.75 (±2.57)
0.13 (±0.37) 0.14 (±0.47) 0.02 (±0.18) 0.00 (±0.00) 0.29 (±0.71)
Mean (±SD) number of MRM,
Zaccarelli et al., JCV2016
By exploring plasma cumulative resistance for any class andresistance detected in PBMC, 20.1% of patients harboured majorresistance mutations (MRMs) not detected in any of previous GRTsperformed in plasma.
Resistance detected in PBMCs predicts virological rebound in HIV-1
suppressed patients switching treatment
Armenia S et al. J Clin Virol 2018;104:61-64
Probability of viral rebound at month 24
Conclusions (I)
• The construction and management of antiretroviral therapy
is designed to take into account a long-term strategy
finalized to decrease to the lowest possible level of HIV
replication and disease /comorbidity progression.
• In this contest, the evaluation of clinically relevant
virological biomarkers not considered in the past (such as
HIV-DNA) is today crucial to ensure a long term control
replication.
• Moreover, they can also be useful in better understanding
factors assessing off-therapy virological remission, and
thus could be relevant for therapeutic strategies aimed at
achieving HIV cure.
• The improvement of resistance testing, finalized to detect
resistance even at undetectable viremia, may allow
clinicians to optimize therapy in the case of switch for
treatment simplification.
• Further investigation, through ultra-sensitive technology,
is needed to clarify the clinical impact of resistance
present in PBMCs.
• Joint efforts among virologists, immunologists, and
clinicians are necessary in order to properly position these
virological parameters into clinical practice and current
guidelines.
Conclusions (II)
Thank you for
the attention!