How does the reservoir influence the clinician's decisions Prof. Carlo Federico Perno Acknowledgements The Patients University of Milan, Milan Italy: C. Alteri. 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. 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. 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) Among 9014 isolates from cART failing patients the prevalence of M184V dramatically decreased from 56% in 1999 to 21% in 2013. In the last 5 years the M184V prevalence is stably settled at 17%. Prevalence of M184V among isolates from cART failing patients over time 100% 90% 80% 70% 60% 60% 56% 57% 60% 51% 54% 45% 46% 50% 41% 40% 33% 27% % of isolates 30% 23% 21% 17% 16% 19% 17% 18% 20% P<0.001 P=0.705 10% 0% Analysis performed on 9014 isolates from cART failing patients (Update August 2018) Armenia & Santoro, Unpublished data 2019 In the past M184V was mainly detected in isolates under suboptimal regimens based on NRTIs or unboosted PIs Prevalence of regimens used at the moment of M184V detection over time Suboptimal 100% 90% 80% detection 70% 60% 59% 50% M184V at 40% 30% 20% P<0.001 10% regimens 0% 0% % of 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 229 varied autopsy tissues from 20 ART- treated patients with low or undetectable plasma 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 findings were identified in all partecipants (brain, spleen, lung, lymph node, liver, aorta and kidney). Tissues assayed with the number of HIV+ (red) S.L. Lamers 2016; 20:8968-83 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. Defining total-body AIDS-virus burden with implications for curative strategies In the quest for a functional cure or eradication of HIV infection, we need to know how large 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-human primates and lymphoid tissue (LT) biopsies from infected humans. Before antiretroviral therapy (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 and their persistence was associated with relatively low drug concentrations in LT compared to peripheral blood. Prolonged ART also reduced the level of SIV and HIV-DNA+ cells, but the estimated size of the residual tissue burden of 108 vDNA+ cells that potentially harbor replication competent proviruses, along with the evidence for continuing virus production in LT despite ART, identify two important sources for rebound following treatment interruption. The large sizes of these tissue reservoirs underscore the challenges in developing “HIV cure” 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 During ART the numbers of virus (v) RNA+ cells substantially decreased but remained detectable. Graphical representation of the proportion of vRNA+ cells in each organ 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 decreased but remained detectable. Graphical representation of the proportion of vRNA+ cells in each organ system before and during suppressive ART. Jacob D Estes Nature Medicine 2017 « Size » of the HIV reservoir The « real reservoir » ? • The vast majority of proviruses that persist on ART are defective. Ho et al. Cell 2013 • 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. 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. Bruner et al. Nature 2019 2018 The Remarkable Frequency of Human Immunodeficiency Virus Type 1 Genetic Recombination A Onafuwa-Nuga and A Telesnitsky Microbiology and Molecular Biology Reviews - 2009 The vast majority of acute transforming retroviruses are replication defective, with the oncogene-containing genome being transmissible 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 recombining with its replication-competent “helper.” In fact, there is some evidence that Rous sarcoma virus, possibly the only naturally arising replication-competent retrovirus containing a host oncogene, was replication defective initially Infectivity of recombinant viruses generated following transfection of 8E5 cells with defective molecular clones of HIV. As we observed for 8E5 in this study, genetic recombination could generate replication-competent