44 results on '"Buzon MJ"'
Search Results
2. Reactivation of latent HIV-1 in central memory CD4⁺ T cells through TLR-1/2 stimulation.
- Author
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Verdin, Eric, Novis, CL, Archin, NM, Buzon, MJ, Round, JL, Lichterfeld, M, Margolis, DM, Planelles, V, and Bosque, A
- Abstract
Toll-like receptors (TLRs) are crucial for recognition of pathogen-associated molecular patterns by cells of the innate immune system. TLRs are present and functional in CD4⁺ T cells. Memory CD4⁺ T cells, predominantly central memory cells (TCM), constitut
- Published
- 2013
3. Executive summary of the GeSIDA consensus document on control and monitoring of HIV-infected patients
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Crespo, M, Lozano, F, Buzon, MJ, Curran, A, Estrada, V, Garcia, F, Imaz, A, Cortes, LL, Losa, JE, Masia, M, Merchante, N, Marino, A, Ocampo, A, Perez-Molina, JA, Poveda, E, Riera, M, Santin, M, Santos, J, Valencia, E, Arazo, P, de la Torre, J, Aldeguer, JL, Palacios, R, Rivero, A, Rubio, R, Sanz, J, and Tellez, MJ
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Human immunodeficiency virus ,Acquired immune deficiency syndrome ,Consensus document ,Guidelines ,Recommendations ,GeSIDA - Abstract
The continuous increase in our knowledge of HIV medicine and antiretroviral treatment has led us to draft specific consensus documents focused on topics other than antiretroviral therapy, such as treatment of opportunistic diseases, pre- and post-exposure prophylaxis, metabolic abnormalities, treatment of HBV or HCV coinfection, treatment of patients coinfected with tuberculosis, osteoporosis, kidney disorders, and cardiovascular risk. Accordingly, the AIDS Study Group (GeSIDA) of the Spanish Society of Infectious Diseases and Clinical Microbiology has promoted the drafting of this consensus document on the control and monitoring of adult patients infected with HIV. The document provides recommendations on the initial evaluation and subsequent monitoring of HIV-infected patients that will prove useful for all professionals involved in the management of this infection. (C) 2018 Elsevier Espana, S.L.U. and Sociedad Espanola de Enfermedades lnfecciosas y Microbiologia Clinica. All rights reserved.
- Published
- 2019
4. Episomal HIV-1 DNA and its relationship to other markers of HIV-1 persistence
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Martinez-Picado, J, Zurakowski, R, Buzon, MJ, and Stevenson, M
- Abstract
Reverse transcription of HIV-1 results in the generation of a linear cDNA that serves as the precursor to the integrated provirus. Other classes of extrachromosomal viral cDNA molecules can be found in acutely infected cells including the 1-LTR and 2-LTR circles of viral DNA, also referred as episomal HIV-1 DNA. Circulating CD4(+) T-cells of treatmentnaive individuals contain significant levels of unintegrated forms of HIV-1 DNA. However, the importance of episomal HIV-1 DNA in the study of viral persistence during antiviral therapy (ART) is debatable. 2-LTR circles are preferentially observed in the effector memory CD4(+) T cell subset of long-term treated subjects. Treatment intensification of standard regimens has been used to determine if more potent ART can impact viral reservoir activity. Adding a potent antiretroviral drug to a stable triple-drug regimen has no measurable impact on plasma HIV-1 RNA levels, suggesting that ongoing cycles of HIV-1 replication are not a major mechanism driving persistent plasma viremia during triple-drug ART. However, in randomized clinical trials of HIV-1-infected adults on apparently effective ART, the addition of an integrase inhibitor (raltegravir) to stable regimens resulted in a transient increase in 2-LTR circles in some patients, suggesting a pre-intensification steady-state in which the processes of virion generation and de novo infection were occurring. Mathematical modeling of 2-LTR production during integrase inhibitor intensification suggests the coexistence, at different levels, of ongoing de novo infection and de novo replication mechanisms, specifically in inflamed lymphoid drug sanctuaries. Most reports looking into potential changes in 2-LTR circles in interventional clinical studies have simultaneously assessed other potential surrogate markers of viral persistence. Transient increases in 2-LTR circles have been correlated to decreases in CD8(+) T-cell activation, transient CD45RA(-)CD4(+) T-cell redistribution, and decreases in the hypercoagulation biomarker D-dimer in ART-intensified individuals. It is difficult, however, to establish a systematic association because the level of correlation with different types of markers differs significantly among studies. In conclusion, despite suppressive ART, a steady-state of de novo infection may persist in some infected individuals and that this may drive immune activation and inflammation changes reflecting residual viral reservoir activity during otherwise apparently suppressive ART.
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- 2018
5. Transient therapy with quadruple NRTI provides immune stability in patients with multidrug resistant HIV-1 and no options for suppressive regimens
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Bonjoch, A, primary, Llibre, JM, additional, Negredo, E, additional, Puig, J, additional, Pérez-Álvarez, N, additional, Buzon, MJ, additional, Martinez-Picado, J, additional, and Clotet, B, additional
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- 2008
- Full Text
- View/download PDF
6. Zinc pyrithione is a potent inhibitor of PLPro and cathepsin L enzymes with ex vivo inhibition of SARS-CoV-2 entry and replication
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Jerneja Kladnik, Ana Dolinar, Jakob Kljun, David Perea, Judith Grau-Expósito, Meritxell Genescà, Marko Novinec, Maria J. Buzon, Iztok Turel, Institut Català de la Salut, [Kladnik J, Dolinar A, Kljun J, Novinec M, Turel I] Faculty of Chemistry and Chemical Technology, University of Ljubljana, Ljubljana, Slovenia. [Perea D, Grau-Expósito J, Genescà M, Buzon MJ] Vall d’Hebron Institut de Recerca (VHIR), Barcelona, Spain. Servei de Malalties Infeccioses, Vall d’Hebron Hospital Universitari, Barcelona, Spain. Universitat Autònoma de Barcelona, Bellaterra, Spain, and Vall d'Hebron Barcelona Hospital Campus
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Pharmacology ,compuestos orgánicos::compuestos organometálicos [COMPUESTOS QUÍMICOS Y DROGAS] ,SARS-CoV-2 ,pirition ,zinc ,Otros calificadores::Otros calificadores::/farmacoterapia [Otros calificadores] ,Virus Diseases::RNA Virus Infections::Nidovirales Infections::Coronaviridae Infections::Coronavirus Infections [DISEASES] ,General Medicine ,Other subheadings::Other subheadings::/drug therapy [Other subheadings] ,udc:546.47:61:577 ,cink ,inhibition ,Chemical Actions and Uses::Pharmacologic Actions::Therapeutic Uses::Anti-Infective Agents::Antiviral Agents [CHEMICALS AND DRUGS] ,inhibicija ,COVID-19 (Malaltia) - Tractament ,acciones y usos químicos::acciones farmacológicas::usos terapéuticos::antiinfecciosos::antivíricos [COMPUESTOS QUÍMICOS Y DROGAS] ,antiviral agents ,Drug Discovery ,virosis::infecciones por virus ARN::infecciones por Nidovirales::infecciones por Coronaviridae::infecciones por Coronavirus [ENFERMEDADES] ,Medicaments antivírics - Ús terapèutic ,protivirusna zdravila ,Compostos organometàl·lics ,Organic Chemicals::Organometallic Compounds [CHEMICALS AND DRUGS] ,pyrithione - Abstract
As SARS-CoV-2 triggered a global health crisis, there is an urgent need to provide patients with safe, effective, accessible, and preferably oral therapeutics for COVID-19 that complement mRNA vaccines. Zinc compounds are widely known for their antiviral properties. Therefore, we have prepared a library of zinc complexes with pyrithione (1-hydroxy-2(1H)-pyridinethione) and its analogues, all of which showed promising in vitro inhibition of cathepsin L, an enzyme involved in SARS-CoV-2 entry, and PLPro, an enzyme involved in SARS-CoV-2 replication both in (sub)micromolar range. Zinc pyrithione 1a is a well-established, commercially available antimicrobial agent and was therefore selected for further evaluation of its SARS-CoV-2 entry and replication inhibition in an ex vivo system derived from primary human lung tissue. Our results suggest that zinc pyrithione complex 1a provides a multitarget approach to combat SARS-CoV-2 and should be considered for repurposing as a potential therapeutic against the insidious COVID-19 disease.Featured imageIn our study, we show that zinc pyrithione holds immense potential for the development of a possible out-patient treatment for SARS-CoV-2 due to its inhibition of viral entry and replication.
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- 2022
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7. Resident memory T cells are a cellular reservoir for HIV in the cervical mucosa
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[Cantero-Pérez J, Grau-Expósito J, Serra-Peinado C, Rosero DA, Luque-Ballesteros L, Astorga-Gamaza A, Falcó V, Buzon MJ, Genescà M] Servei de Malalties Infeccioses, Vall d’Hebron Hospital Universitari, Barcelona, Spain. Grup de Recerca en Malalties Infeccioses, Vall d’Hebron Institut de Recerca (VHIR), Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain. [Castellví J] Servei d’Anatomia Patològica, Vall d’Hebron Hospital Universitari, Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain. [Mañalich-Barrachina L, Centeno-Mediavilla C] Servei de Ginecologia i Obstetrícia, Vall d’Hebron Hospital Universitari, Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain and Hospital Universitari Vall d'Hebron
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Hemic and Immune Systems::Hemic and Immune Systems::Immune System::Leukocytes::Leukocytes, Mononuclear::Lymphocytes::T-Lymphocytes::CD4-Positive T-Lymphocytes [ANATOMY] ,Cèl·lules T ,Coll uterí ,Sistemas Sanguíneo e Inmunológico::Sistemas Sanguíneo e Inmunológico::Sistema Inmunológico::Leucocitos::Leucocitos Mononucleares::Linfocitos::Linfocitos T::Linfocitos T CD4-Positivos [ANATOMÍA] ,Urogenital System::Genitalia::Genitalia, Female::Uterus::Cervix Uteri [ANATOMY] ,Infeccions per VIH ,sistema urogenital::genitales::genitales femeninos::útero::cuello del útero [ANATOMÍA] ,Enfermedades del Sistema Inmune::Síndromes de Inmunodeficiencia::Infecciones por VIH [ENFERMEDADES] ,Immune System Diseases::Immunologic Deficiency Syndromes::HIV Infections [DISEASES] - Published
- 2021
8. Expression of CD20 after viral reactivation renders HIV-reservoir cells susceptible to Rituximab
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[Serra-Peinado C, Grau-Expósito J, Luque-Ballesteros L, Astorga-Gamaza A, Navarro J, Gallego-Rodriguez J, Martin M, Curran A, Burgos J, Ribera E, Raventós B, Willekens R, Torrella A, Planas B, Badía R, Genescà M, Falcó V, Buzon MJ] Servei de Malalties Infeccioses, Hospital Universitari Vall d’Hebron, Barcelona, Spain. Vall d’Hebron Institut de Recerca (VHIR). Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain. [Castellví J] Servei de Patologia, Hospital Universitari Vall d’Hebron. Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain and Hospital Universitari Vall d'Hebron
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Hemic and Immune Systems::Hemic and Immune Systems::Immune System::Leukocytes::Leukocytes, Mononuclear::Lymphocytes::T-Lymphocytes::CD4-Positive T-Lymphocytes [ANATOMY] ,Cèl·lules T ,Amino Acids, Peptides, and Proteins::Proteins::Blood Proteins::Immunoproteins::Immunoglobulins::Antibodies::Antibodies, Monoclonal::Antibodies, Monoclonal, Murine-Derived::Rituximab [CHEMICALS AND DRUGS] ,Aminoácidos, Péptidos y Proteínas::Proteínas::Proteínas Sanguíneas::Inmunoproteínas::Inmunoglobulinas::Anticuerpos::Anticuerpos Monoclonales::Anticuerpos Monoclonales de Origen Murino::Rituximab [COMPUESTOS QUÍMICOS Y DROGAS] ,Sistemas Sanguíneo e Inmunológico::Sangre::Células Sanguíneas::Leucocitos::Leucocitos Mononucleares::Linfocitos::Linfocitos T::Sistemas Sanguíneo e Inmunológico::Linfocitos T CD4-Positivos [ANATOMÍA] ,Infeccions per VIH ,Rituximab ,Enfermedades del Sistema Inmune::Síndromes de Inmunodeficiencia::Infecciones por VIH [ENFERMEDADES] ,Immune System Diseases::Immunologic Deficiency Syndromes::HIV Infections [DISEASES] - Published
- 2021
9. Resident memory T cells are a cellular reservoir for HIV in the cervical mucosa
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Cantero, Jon, Grau-Expósito, Judith, Serra-Peinado, Carla, Rosero, D. A., Luque-Ballesteros, Laura, Astorga-Gamaza, Antonio, Castellvi, Josep., Sanhueza, T., Tapia Melendo, Gustavo, Lloveras, B., Fernández, Marco A., Prado, Julia G., Solé-Sedeno, J. M., Tarrats, A., Lecumberri, Carla, Mañalich-Barrachina, L., Centeno-Mediavilla, C., Falcó, Vicenç, Buzón, Maria José, Genescà Ferrer, Meritxell., Universitat Autònoma de Barcelona, [Cantero-Pérez J, Grau-Expósito J, Serra-Peinado C, Rosero DA, Luque-Ballesteros L, Astorga-Gamaza A, Falcó V, Buzon MJ, Genescà M] Servei de Malalties Infeccioses, Vall d’Hebron Hospital Universitari, Barcelona, Spain. Grup de Recerca en Malalties Infeccioses, Vall d’Hebron Institut de Recerca (VHIR), Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain. [Castellví J] Servei d’Anatomia Patològica, Vall d’Hebron Hospital Universitari, Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain. [Mañalich-Barrachina L, Centeno-Mediavilla C] Servei de Ginecologia i Obstetrícia, Vall d’Hebron Hospital Universitari, Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain, and Vall d'Hebron Barcelona Hospital Campus
- Subjects
CD4-Positive T-Lymphocytes ,0301 basic medicine ,CD32 ,Human immunodeficiency virus (HIV) ,Urogenital System::Genitalia::Genitalia, Female::Uterus::Cervix Uteri [ANATOMY] ,General Physics and Astronomy ,HIV Infections ,Cervix Uteri ,medicine.disease_cause ,Pathogenesis ,0302 clinical medicine ,Coll uterí ,sistema urogenital::genitales::genitales femeninos::útero::cuello del útero [ANATOMÍA] ,Dna viral ,lcsh:Science ,Multidisciplinary ,biology ,virus diseases ,Middle Aged ,Viral Load ,Phenotype ,3. Good health ,Cèl·lules T ,Anti-Retroviral Agents ,Female ,HIV infections ,Adult ,Female circumcision ,Science ,Article ,General Biochemistry, Genetics and Molecular Biology ,Viral reservoirs ,03 medical and health sciences ,enfermedades del sistema inmune::síndromes de inmunodeficiencia::infecciones por VIH [ENFERMEDADES] ,VIH (Virus) ,medicine ,Humans ,Mucosa cervical ,Aged ,Disease Reservoirs ,Hemic and Immune Systems::Hemic and Immune Systems::Immune System::Leukocytes::Leukocytes, Mononuclear::Lymphocytes::T-Lymphocytes::CD4-Positive T-Lymphocytes [ANATOMY] ,Mucous Membrane ,Cluster of differentiation ,RNA ,General Chemistry ,Virology ,Immune System Diseases::Immunologic Deficiency Syndromes::HIV Infections [DISEASES] ,sistemas sanguíneo e inmunológico::sistemas sanguíneo e inmunológico::sistema inmunológico::leucocitos::leucocitos mononucleares::linfocitos::linfocitos T::linfocitos T CD4-positivos [ANATOMÍA] ,030104 developmental biology ,HIV-1 ,biology.protein ,Infeccions per VIH ,lcsh:Q ,Immunologic Memory ,030217 neurology & neurosurgery - Abstract
HIV viral reservoirs are established very early during infection. Resident memory T cells (TRM) are present in tissues such as the lower female genital tract, but the contribution of this subset of cells to the pathogenesis and persistence of HIV remains unclear. Here, we show that cervical CD4+TRM display a unique repertoire of clusters of differentiation, with enrichment of several molecules associated with HIV infection susceptibility, longevity and self-renewing capacities. These protein profiles are enriched in a fraction of CD4+TRM expressing CD32. Cervical explant models show that CD4+TRM preferentially support HIV infection and harbor more viral DNA and protein than non-TRM. Importantly, cervical tissue from ART-suppressed HIV+ women contain high levels of viral DNA and RNA, being the TRM fraction the principal contributor. These results recognize the lower female genital tract as an HIV sanctuary and identify CD4+TRM as primary targets of HIV infection and viral persistence. Thus, strategies towards an HIV cure will need to consider TRM phenotypes, which are widely distributed in tissues., Using cervical explant models and cervical tissue from ART-suppressed HIV+ women, the authors here show that resident memory T cells (TRM) in the cervical mucosa are preferentially infected and harbor more viral DNA, RNA and protein than non-TRM, suggesting that TRM may serve as viral reservoir in the cervical mucosa.
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- 2019
10. Latency reversal agents affect differently the latent reservoir present in distinct CD4+ t subpopulations
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Grau-Expósito, Judith, Luque-Ballesteros, Laura, Navarro Mercadé, Jordi, Curran, Adrian, Burgos, Joaquín, Ribera, Esteban, Torrella Domingo, Adriana, Planas, Bibiana, Badía, Rosa, Martin-Castillo, Mario, Fernández-Sojo, Jesús, Genescà Ferrer, Meritxell., Falcó, Vicenç, Buzon, Maria J., Universitat Autònoma de Barcelona, [Grau-Expósito J, Luque-Ballesteros L, Navarro J, Curran A, Burgos J, Ribera E, Torrella A, Planas B, Badía R, Martin-Castillo M, Genescà M, Falcó V, Buzon MJ] Servei de Malalties Infeccioses, Hospital Universitari Vall d’Hebron, Barcelona, Spain. Vall d’Hebron Institut de Recerca (VHIR). Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain. [Fernández-Sojo J] Banc de Sang i Teixits, Hospital Universitari Vall d'Hebron, Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain, and Vall d'Hebron Barcelona Hospital Campus
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RNA viruses ,CD4-Positive T-Lymphocytes ,Hemic and Immune Systems::Blood::Blood Cells::Leukocytes::Leukocytes, Mononuclear::Lymphocytes::T-Lymphocytes::Hemic and Immune Systems::CD4-Positive T-Lymphocytes [ANATOMY] ,Apoptosis ,HIV Infections ,Pathology and Laboratory Medicine ,Toxicology ,medicine.disease_cause ,Microbiological Phenomena::Virus Physiological Phenomena::Virus Latency [PHENOMENA AND PROCESSES] ,Memory T cells ,Romidepsin ,White Blood Cells ,Immunodeficiency Viruses ,Animal Cells ,Depsipeptides ,Medicine and Health Sciences ,Biology (General) ,acciones y usos químicos::acciones farmacológicas::usos terapéuticos::antiinfecciosos::antivíricos::antirretrovirales::fármacos anti-VIH [COMPUESTOS QUÍMICOS Y DROGAS] ,0303 health sciences ,Cell Death ,T Cells ,030302 biochemistry & molecular biology ,Antiretrovirals ,Chemical Actions and Uses::Pharmacologic Actions::Therapeutic Uses::Anti-Infective Agents::Antiviral Agents::Anti-Retroviral Agents::Anti-HIV Agents [CHEMICALS AND DRUGS] ,Viral Load ,Viral Persistence and Latency ,3. Good health ,Virus Latency ,medicine.anatomical_structure ,Cèl·lules T ,Medical Microbiology ,Cell Processes ,Viral Pathogens ,Viruses ,Pathogens ,Cellular Types ,Stem cell ,Diterpenes ,Viral load ,fenómenos microbiológicos::fenómenos fisiológicos de los virus::latencia viral [FENÓMENOS Y PROCESOS] ,Research Article ,medicine.drug ,QH301-705.5 ,Viral protein ,Anti-HIV Agents ,Immune Cells ,T cell ,Immunology ,Biology ,Microbiology ,03 medical and health sciences ,Virology ,Retroviruses ,Genetics ,medicine ,Humans ,Microbial Pathogens ,Molecular Biology ,030304 developmental biology ,Blood Cells ,Toxicity ,Lentivirus ,Virus - Reproducció ,Organisms ,Biology and Life Sciences ,HIV ,RNA ,Cell Biology ,RC581-607 ,Viral Replication ,sistemas sanguíneo e inmunológico::sistemas sanguíneo e inmunológico::sistema inmunológico::leucocitos::leucocitos mononucleares::linfocitos::linfocitos T::linfocitos T CD4-positivos [ANATOMÍA] ,Viral replication ,HIV-1 ,Parasitology ,Virus Activation ,Immunologic diseases. Allergy ,Ex vivo - Abstract
Latency reversal agents (LRAs) have proven to induce HIV-1 transcription in vivo but are ineffective at decreasing the size of the latent reservoir in antiretroviral treated patients. The capacity of the LRAs to perturb the viral reservoir present in distinct subpopulations of cells is currently unknown. Here, using a new RNA FISH/flow ex vivo viral reactivation assay, we performed a comprehensive assessment of the viral reactivation capacity of different families of LRAs, and their combinations, in different CD4+ T cell subsets. We observed that a median of 16.28% of the whole HIV-reservoir induced HIV-1 transcripts after viral reactivation, but only 10.10% of these HIV-1 RNA+ cells produced the viral protein p24. Moreover, none of the LRAs were powerful enough to reactivate HIV-1 transcription in all CD4+ T cell subpopulations. For instance, the combination of Romidepsin and Ingenol was identified as the best combination of drugs at increasing the proportion of HIV-1 RNA+ cells, in most, but not all, CD4+ T cell subsets. Importantly, memory stem cells were identified as highly resistant to HIV-1 reactivation, and only the combination of Panobinostat and Bryostatin-1 significantly increased the number of cells transcribing HIV within this subset. Overall, our results validate the use of the RNA FISH/flow technique to assess the potency of LRAs among different CD4+ T cell subsets, manifest the intrinsic differences between cells that encompass the latent HIV reservoir, and highlight the difficulty to significantly impact the latent infection with the currently available drugs. Thus, our results have important implications for the rational design of therapies aimed at reversing HIV latency from diverse cellular reservoirs., Author summary HIV infection is an incurable disease. Despite antiretroviral therapy, a pool of cells with HIV in a latent state persists and precludes fully eradication of the viral infection. The cells that contain this latent viral reservoir are very diverse, and therefore different therapeutic strategies would be necessary to target and eliminate all infected cells. Latency Reversal Agents (LRAs) are compounds able to awake the latent virus from its dormant state with the purpose of making infected cells visible to the immune system. But the ability of the LRAs to target different cell types containing HIV is currently unknown. Here, using a novel methodology that interrogates individual cells, we found that current LRAs do not impact equally all infected cells. In fact, certain types of memory lymphocytes, recognized to harbor latent HIV for decades, are not fully impacted by most of the LRAs tested. Our study highlights the difficulty to cure HIV with the currently available LRAs. Different therapeutic approaches aimed at reversing HIV latency from diverse cellular reservoirs are needed to reduce HIV persistence.
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- 2019
11. Expression of CD20 after viral reactivation renders HIV-reservoir cells susceptible to Rituximab
- Author
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Serra-Peinado, Carla, Grau-Expósito, Judith, Luque-Ballesteros, Laura, Astorga-Gamaza, Antonio, Navarro, Jordi, Gallego-Rodriguez, Jenny, Martín Castillo, Mario, Curran, Adrian, Burgos, Joaquín, Ribera, Esteban, Raventós, Berta, Willekens, Rein, Torrella Domingo, Adriana, Planas, Bibiana, Badía, Rosa, García, Felipe, Castellvi, Josep, Genescà Ferrer, Meritxell, Falcó, Vicenç, Buzón, Maria José, Universitat Autònoma de Barcelona, [Serra-Peinado C, Grau-Expósito J, Luque-Ballesteros L, Astorga-Gamaza A, Navarro J, Gallego-Rodriguez J, Martin M, Curran A, Burgos J, Ribera E, Raventós B, Willekens R, Torrella A, Planas B, Badía R, Genescà M, Falcó V, Buzon MJ] Servei de Malalties Infeccioses, Hospital Universitari Vall d’Hebron, Barcelona, Spain. Vall d’Hebron Institut de Recerca (VHIR). Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain. [Castellví J] Servei de Patologia, Hospital Universitari Vall d’Hebron. Barcelona, Spain. Universitat Autònoma de Barcelona, Barcelona, Spain, and Vall d'Hebron Barcelona Hospital Campus
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0301 basic medicine ,CD4-Positive T-Lymphocytes ,General Physics and Astronomy ,HIV Infections ,02 engineering and technology ,Lymphocyte Activation ,hemic and lymphatic diseases ,Virus latency ,lcsh:Science ,Lymph node ,CD20 ,Multidisciplinary ,biology ,Amino Acids, Peptides, and Proteins::Proteins::Blood Proteins::Immunoproteins::Immunoglobulins::Antibodies::Antibodies, Monoclonal::Antibodies, Monoclonal, Murine-Derived::Rituximab [CHEMICALS AND DRUGS] ,021001 nanoscience & nanotechnology ,Flow Cytometry ,3. Good health ,Virus Latency ,medicine.anatomical_structure ,Cell killing ,Cèl·lules T ,RNA, Viral ,Rituximab ,0210 nano-technology ,Cell activation ,Infection ,medicine.drug ,Anti-HIV Agents ,Science ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,Viral reservoirs ,Antigen ,enfermedades del sistema inmune::síndromes de inmunodeficiencia::infecciones por VIH [ENFERMEDADES] ,medicine ,Humans ,Immunologic Factors ,Hemic and Immune Systems::Hemic and Immune Systems::Immune System::Leukocytes::Leukocytes, Mononuclear::Lymphocytes::T-Lymphocytes::CD4-Positive T-Lymphocytes [ANATOMY] ,General Chemistry ,Translational research ,medicine.disease ,Antigens, CD20 ,Immune System Diseases::Immunologic Deficiency Syndromes::HIV Infections [DISEASES] ,sistemas sanguíneo e inmunológico::sistemas sanguíneo e inmunológico::sistema inmunológico::leucocitos::leucocitos mononucleares::linfocitos::linfocitos T::linfocitos T CD4-positivos [ANATOMÍA] ,030104 developmental biology ,aminoácidos, péptidos y proteínas::proteínas::proteínas sanguíneas::inmunoproteínas::inmunoglobulinas::anticuerpos::anticuerpos monoclonales::anticuerpos monoclonales de origen murino::rituximab [COMPUESTOS QUÍMICOS Y DROGAS] ,Immunology ,biology.protein ,HIV-1 ,Leukocytes, Mononuclear ,lcsh:Q ,Infeccions per VIH ,Virus Activation ,Lymph Nodes ,Immunologic Memory ,Ex vivo - Abstract
The identification of exclusive markers to target HIV-reservoir cells will represent a significant advance in the search for therapies to cure HIV. Here, we identify the B lymphocyte antigen CD20 as a marker for HIV-infected cells in vitro and in vivo. The CD20 molecule is dimly expressed in a subpopulation of CD4-positive (CD4+) T lymphocytes from blood, with high levels of cell activation and heterogeneous memory phenotypes. In lymph node samples from infected patients, CD20 is present in productively HIV-infected cells, and ex vivo viral infection selectively upregulates the expression of CD20 during early infection. In samples from patients on antiretroviral therapy (ART) this subpopulation is significantly enriched in HIV transcripts, and the anti-CD20 monoclonal antibody Rituximab induces cell killing, which reduces the pool of HIV-expressing cells when combined with latency reversal agents. We provide a tool for targeting this active HIV-reservoir after viral reactivation in patients while on ART., Here, the authors identify B lymphocyte antigen CD20 as a marker for HIV-infected T cells and provide evidence for the potential use of anti-CD20 antibodies in combination with latency reversing agents for depletion of viral reactivated CD4 T cells in patients on antiretroviral therapy.
- Published
- 2018
12. NKG2C and NKG2A Co-expression Defines a Highly Functional Antiviral NK Population in Spontaneous HIV Control.
- Author
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Sanchez-Gaona N, Gallego-Cortés A, Astorga-Gamaza A, Rallón N, Benito J, Ruiz-Mateos E, Curran A, Burgos J, Navarro J, Suanzes P, Falco V, Genescà M, and Buzon MJ
- Abstract
Elite controllers (EC), a unique group of people with HIV (PWH), exhibit remarkable control of viral replication in the absence of antiretroviral therapy. In this study, we comprehensively characterized the NK cell repertoire in EC after long-term viral control. Phenotypic profiling of NK cells revealed profound differences compared with other PWH, but marked similarities to uninfected individuals, with a distinctive prevalence of NKG2C+CD57+memory-like NK cells. Functional analyses indicated that EC had limited production of functional molecules upon NK stimulation and consequently reduced natural cytotoxicity against non-HIV target cells. Importantly, EC showed an exceptional ability to kill primary HIV-infected cells by the antibody-dependent cell cytotoxicity (ADCC) adaptive mechanism, which was achieved by a specific memory-like NK population expressing CD16, NKG2A, NKG2C, CD57 and CXCR3. In-depth single-cell RNA sequencing unveiled a unique transcriptional signature in these NK cells linked to increased cell metabolism, migration, chemotaxis, effector functions, cytokine secretion, and antiviral response. Our findings underscore a pivotal role of NK cells in the immune control of HIV and identify specific NK cells as emerging targets for immunotherapies.
- Published
- 2024
- Full Text
- View/download PDF
13. KLRG1 expression on natural killer cells is associated with HIV persistence, and its targeting promotes the reduction of the viral reservoir.
- Author
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Astorga-Gamaza A, Perea D, Sanchez-Gaona N, Calvet-Mirabent M, Gallego-Cortés A, Grau-Expósito J, Sanchez-Cerrillo I, Rey J, Castellví J, Curran A, Burgos J, Navarro J, Suanzes P, Falcó V, Genescà M, Martín-Gayo E, and Buzon MJ
- Subjects
- Humans, Killer Cells, Natural, Lectins, C-Type genetics, Virus Latency, HIV Infections drug therapy, HIV Infections genetics, HIV-1, Receptors, Immunologic genetics
- Abstract
Human immunodeficiency virus (HIV) infection induces immunological dysfunction, which limits the elimination of HIV-infected cells during treated infection. Identifying and targeting dysfunctional immune cells might help accelerate the purging of the persistent viral reservoir. Here, we show that chronic HIV infection increases natural killer (NK) cell populations expressing the negative immune regulator KLRG1, both in peripheral blood and lymph nodes. Antiretroviral treatment (ART) does not reestablish these functionally impaired NK populations, and the expression of KLRG1 correlates with active HIV transcription. Targeting KLRG1 with specific antibodies significantly restores the capacity of NK cells to kill HIV-infected cells, reactivates latent HIV present in CD4
+ T cells co-expressing KLRG1, and reduces the intact HIV genomes in samples from ART-treated individuals. Our data support the potential use of immunotherapy against the KLRG1 receptor to impact the viral reservoir during HIV persistence., Competing Interests: Declaration of interests The authors declare no competing interest., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)- Published
- 2023
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14. Schlafen 12 restricts HIV-1 latency reversal by a codon-usage dependent post-transcriptional block in CD4+ T cells.
- Author
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Kobayashi-Ishihara M, Frazão Smutná K, Alonso FE, Argilaguet J, Esteve-Codina A, Geiger K, Genescà M, Grau-Expósito J, Duran-Castells C, Rogenmoser S, Böttcher R, Jungfleisch J, Oliva B, Martinez JP, Li M, David M, Yamagishi M, Ruiz-Riol M, Brander C, Tsunetsugu-Yokota Y, Buzon MJ, Díez J, and Meyerhans A
- Subjects
- Codon Usage, RNA, Viral genetics, Virus Latency genetics, CD4-Positive T-Lymphocytes, HIV-1 physiology
- Abstract
Latency is a major barrier towards virus elimination in HIV-1-infected individuals. Yet, the mechanisms that contribute to the maintenance of HIV-1 latency are incompletely understood. Here we describe the Schlafen 12 protein (SLFN12) as an HIV-1 restriction factor that establishes a post-transcriptional block in HIV-1-infected cells and thereby inhibits HIV-1 replication and virus reactivation from latently infected cells. The inhibitory activity is dependent on the HIV-1 codon usage and on the SLFN12 RNase active sites. Within HIV-1-infected individuals, SLFN12 expression in PBMCs correlated with HIV-1 plasma viral loads and proviral loads suggesting a link with the general activation of the immune system. Using an RNA FISH-Flow HIV-1 reactivation assay, we demonstrate that SLFN12 expression is enriched in infected cells positive for HIV-1 transcripts but negative for HIV-1 proteins. Thus, codon-usage dependent translation inhibition of HIV-1 proteins participates in HIV-1 latency and can restrict the amount of virus release after latency reversal., (© 2023. The Author(s).)
- Published
- 2023
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15. Zinc pyrithione is a potent inhibitor of PL Pro and cathepsin L enzymes with ex vivo inhibition of SARS-CoV-2 entry and replication.
- Author
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Kladnik J, Dolinar A, Kljun J, Perea D, Grau-Expósito J, Genescà M, Novinec M, Buzon MJ, and Turel I
- Subjects
- Antiviral Agents pharmacology, Cathepsin L, Humans, Organometallic Compounds, Pyridines, SARS-CoV-2, Ruthenium, COVID-19 Drug Treatment
- Abstract
Zinc pyrithione ( 1a ), together with its analogues 1b - h and ruthenium pyrithione complex 2a , were synthesised and evaluated for the stability in biologically relevant media and anti-SARS-CoV-2 activity. Zinc pyrithione revealed potent in vitro inhibition of cathepsin L (IC
50 =1.88 ± 0.49 µM) and PLPro (IC50 =0.50 ± 0.07 µM), enzymes involved in SARS-CoV-2 entry and replication, respectively, as well as antiviral entry and replication properties in an ex vivo system derived from primary human lung tissue. Zinc complexes 1b - h expressed comparable in vitro inhibition. On the contrary, ruthenium complex 2a and the ligand pyrithione a itself expressed poor inhibition in mentioned assays, indicating the importance of the selection of metal core and structure of metal complex for antiviral activity. Safe, effective, and preferably oral at-home therapeutics for COVID-19 are needed and as such zinc pyrithione, which is also commercially available, could be considered as a potential therapeutic agent against SARS-CoV-2.- Published
- 2022
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16. Identification of HIV-reservoir cells with reduced susceptibility to antibody-dependent immune response.
- Author
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Astorga-Gamaza A, Grau-Expósito J, Burgos J, Navarro J, Curran A, Planas B, Suanzes P, Falcó V, Genescà M, and Buzon MJ
- Subjects
- Anti-Retroviral Agents therapeutic use, CD4-Positive T-Lymphocytes, HIV Antibodies, Humans, Immunity, HIV Infections, HIV-1 physiology
- Abstract
Human immunodeficiency virus (HIV) establishes a persistent infection in heterogeneous cell reservoirs, which can be maintained by different mechanisms including cellular proliferation, and represent the main obstacle to curing the infection. The expression of the Fcγ receptor CD32 has been identified as a marker of the active cell reservoirs in people on antiretroviral therapy (ART), but if its expression has any role in conferring advantage for viral persistence is unknown. Here, we report that HIV-infected cells expressing CD32 have reduced susceptibility to natural killer (NK) antibody-dependent cell cytotoxicity (ADCC) by a mechanism compatible with the suboptimal binding of HIV-specific antibodies. Infected CD32 cells have increased proliferative capacity in the presence of immune complexes, and are more resistant to strategies directed to potentiate NK function. Remarkably, reactivation of the latent reservoir from antiretroviral-treated people living with HIV increases the pool of infected CD32 cells, which are largely resistant to the ADCC immune mechanism. Thus, we report the existence of reservoir cells that evade part of the NK immune response through the expression of CD32., Competing Interests: AA, JG, JB, JN, AC, BP, PS, VF, MG, MB No competing interests declared, (© 2022, Astorga-Gamaza et al.)
- Published
- 2022
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17. Evaluation of SARS-CoV-2 entry, inflammation and new therapeutics in human lung tissue cells.
- Author
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Grau-Expósito J, Perea D, Suppi M, Massana N, Vergara A, Soler MJ, Trinite B, Blanco J, García-Pérez J, Alcamí J, Serrano-Mollar A, Rosado J, Falcó V, Genescà M, and Buzon MJ
- Subjects
- Adult, Animals, Antiviral Agents pharmacology, COVID-19 immunology, COVID-19 pathology, Cells, Cultured, Chlorocebus aethiops, Drug Evaluation, Preclinical, Drugs, Investigational pharmacology, Drugs, Investigational therapeutic use, HEK293 Cells, Host-Pathogen Interactions drug effects, Humans, Inflammation pathology, Inflammation therapy, Inflammation virology, Lung pathology, SARS-CoV-2 drug effects, Vero Cells, Antiviral Agents therapeutic use, Lung virology, SARS-CoV-2 physiology, Virus Internalization drug effects, COVID-19 Drug Treatment
- Abstract
The development of physiological models that reproduce SARS-CoV-2 infection in primary human cells will be instrumental to identify host-pathogen interactions and potential therapeutics. Here, using cell suspensions directly from primary human lung tissues (HLT), we have developed a rapid platform for the identification of viral targets and the expression of viral entry factors, as well as for the screening of viral entry inhibitors and anti-inflammatory compounds. The direct use of HLT cells, without long-term cell culture and in vitro differentiation approaches, preserves main immune and structural cell populations, including the most susceptible cell targets for SARS-CoV-2; alveolar type II (AT-II) cells, while maintaining the expression of proteins involved in viral infection, such as ACE2, TMPRSS2, CD147 and AXL. Further, antiviral testing of 39 drug candidates reveals a highly reproducible method, suitable for different SARS-CoV-2 variants, and provides the identification of new compounds missed by conventional systems, such as VeroE6. Using this method, we also show that interferons do not modulate ACE2 expression, and that stimulation of local inflammatory responses can be modulated by different compounds with antiviral activity. Overall, we present a relevant and rapid method for the study of SARS-CoV-2., Competing Interests: The authors have declared that no competing interests exist.
- Published
- 2022
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18. The active human immunodeficiency virus reservoir during antiretroviral therapy: emerging players in viral persistence.
- Author
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Astorga-Gamaza A and Buzon MJ
- Subjects
- CD4-Positive T-Lymphocytes, Genome, Viral, HIV genetics, Humans, Viral Load, Virus Latency, Virus Replication, HIV Infections drug therapy
- Abstract
Purpose of Review: To discuss the role of CD4+ T cells with active Human immunodeficiency virus (HIV), meaning infected cells with transcriptional and/or translational viral activity during antiretroviral therapy (ART), focusing on new technologies for its detection, potential cell markers for its characterization, and evidences on the contribution of the active HIV reservoir to long-term viral persistence., Recent Findings: HIV-infected cells expressing viral ribonucleic acid are systematically detected in subjects on long-term ART. In recent years, powerful new tools have provided significant insights into the nature, quantification, and identification of cells with active HIV, including the identification of new cell markers, and the presence of viral activity in specific cell populations located in different cellular and anatomical compartments. Moreover, studies on viral sequence integrity have identified cell clones with intact viral genomes and active viral transcription that could potentially persist for years. Together, new investigations support the notion that the active reservoir could represent a relevant fraction of long-term infected cells, and therefore, the study of its cell sources and mechanisms of maintenance could represent a significant advance in our understanding of viral persistence and the development of new curative strategies., Summary: The presence of HIV-infected cells with viral expression during ART has been traditionally overlooked for years. Based on recent investigations, this active viral reservoir could play an important role in HIV persistence., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc.)
- Published
- 2021
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19. Antibody cooperative adsorption onto AuNPs and its exploitation to force natural killer cells to kill HIV-infected T cells.
- Author
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Astorga-Gamaza A, Vitali M, Borrajo ML, Suárez-López R, Jaime C, Bastus N, Serra-Peinado C, Luque-Ballesteros L, Blanch-Lombarte O, Prado JG, Lorente J, Pumarola F, Pellicer M, Falcó V, Genescà M, Puntes V, and Buzon MJ
- Abstract
HIV represents a persistent infection which negatively alters the immune system. New tools to reinvigorate different immune cell populations to impact HIV are needed. Herein, a novel nanotool for the specific enhancement of the natural killer (NK) immune response towards HIV-infected T-cells has been developed. Bispecific Au nanoparticles (BiAb-AuNPs), dually conjugated with IgG anti-HIVgp120 and IgG anti-human CD16 antibodies, were generated by a new controlled, linker-free and cooperative conjugation method promoting the ordered distribution and segregation of antibodies in domains. The cooperatively-adsorbed antibodies fully retained the capabilities to recognize their cognate antigen and were able to significantly enhance cell-to-cell contact between HIV-expressing cells and NK cells. As a consequence, the BiAb-AuNPs triggered a potent cytotoxic response against HIV-infected cells in blood and human tonsil explants. Remarkably, the BiAb-AuNPs were able to significantly reduce latent HIV infection after viral reactivation in a primary cell model of HIV latency. This novel molecularly-targeted strategy using a bispecific nanotool to enhance the immune system represents a new approximation with potential applications beyond HIV.
- Published
- 2021
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20. Recommendations for measuring HIV reservoir size in cure-directed clinical trials.
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Abdel-Mohsen M, Richman D, Siliciano RF, Nussenzweig MC, Howell BJ, Martinez-Picado J, Chomont N, Bar KJ, Yu XG, Lichterfeld M, Alcami J, Hazuda D, Bushman F, Siliciano JD, Betts MR, Spivak AM, Planelles V, Hahn BH, Smith DM, Ho YC, Buzon MJ, Gaebler C, Paiardini M, Li Q, Estes JD, Hope TJ, Kostman J, Mounzer K, Caskey M, Fox L, Frank I, Riley JL, Tebas P, and Montaner LJ
- Subjects
- CD4-Positive T-Lymphocytes virology, Clinical Trials as Topic, Humans, Mass Screening methods, Viral Load drug effects, Virus Latency drug effects, Anti-Retroviral Agents therapeutic use, Disease Reservoirs virology, HIV Infections diagnosis, HIV Infections drug therapy, HIV-1 drug effects
- Abstract
Therapeutic strategies are being clinically tested either to eradicate latent HIV reservoirs or to achieve virologic control in the absence of antiretroviral therapy. Attaining this goal will require a consensus on how best to measure the numbers of persistently infected cells with the potential to cause viral rebound after antiretroviral-therapy cessation in assessing the results of cure-directed strategies in vivo. Current measurements assess various aspects of the HIV provirus and its functionality and produce divergent results. Here, we provide recommendations from the BEAT-HIV Martin Delaney Collaboratory on which viral measurements should be prioritized in HIV-cure-directed clinical trials.
- Published
- 2020
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21. Latency reversal agents affect differently the latent reservoir present in distinct CD4+ T subpopulations.
- Author
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Grau-Expósito J, Luque-Ballesteros L, Navarro J, Curran A, Burgos J, Ribera E, Torrella A, Planas B, Badía R, Martin-Castillo M, Fernández-Sojo J, Genescà M, Falcó V, and Buzon MJ
- Subjects
- CD4-Positive T-Lymphocytes drug effects, CD4-Positive T-Lymphocytes virology, Depsipeptides pharmacology, Diterpenes pharmacology, HIV Infections drug therapy, HIV Infections virology, HIV-1 drug effects, Humans, Viral Load, Virus Activation drug effects, Virus Latency drug effects, Anti-HIV Agents pharmacology, CD4-Positive T-Lymphocytes immunology, HIV Infections immunology, HIV-1 immunology, Virus Activation immunology, Virus Latency immunology
- Abstract
Latency reversal agents (LRAs) have proven to induce HIV-1 transcription in vivo but are ineffective at decreasing the size of the latent reservoir in antiretroviral treated patients. The capacity of the LRAs to perturb the viral reservoir present in distinct subpopulations of cells is currently unknown. Here, using a new RNA FISH/flow ex vivo viral reactivation assay, we performed a comprehensive assessment of the viral reactivation capacity of different families of LRAs, and their combinations, in different CD4+ T cell subsets. We observed that a median of 16.28% of the whole HIV-reservoir induced HIV-1 transcripts after viral reactivation, but only 10.10% of these HIV-1 RNA+ cells produced the viral protein p24. Moreover, none of the LRAs were powerful enough to reactivate HIV-1 transcription in all CD4+ T cell subpopulations. For instance, the combination of Romidepsin and Ingenol was identified as the best combination of drugs at increasing the proportion of HIV-1 RNA+ cells, in most, but not all, CD4+ T cell subsets. Importantly, memory stem cells were identified as highly resistant to HIV-1 reactivation, and only the combination of Panobinostat and Bryostatin-1 significantly increased the number of cells transcribing HIV within this subset. Overall, our results validate the use of the RNA FISH/flow technique to assess the potency of LRAs among different CD4+ T cell subsets, manifest the intrinsic differences between cells that encompass the latent HIV reservoir, and highlight the difficulty to significantly impact the latent infection with the currently available drugs. Thus, our results have important implications for the rational design of therapies aimed at reversing HIV latency from diverse cellular reservoirs., Competing Interests: The authors have declared that no competing interests exist.
- Published
- 2019
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22. Expression of CD20 after viral reactivation renders HIV-reservoir cells susceptible to Rituximab.
- Author
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Serra-Peinado C, Grau-Expósito J, Luque-Ballesteros L, Astorga-Gamaza A, Navarro J, Gallego-Rodriguez J, Martin M, Curran A, Burgos J, Ribera E, Raventós B, Willekens R, Torrella A, Planas B, Badía R, Garcia F, Castellví J, Genescà M, Falcó V, and Buzon MJ
- Subjects
- Anti-HIV Agents therapeutic use, CD4-Positive T-Lymphocytes immunology, CD4-Positive T-Lymphocytes metabolism, Flow Cytometry, HIV Infections drug therapy, HIV Infections immunology, HIV-1, Humans, Immunologic Memory, Leukocytes, Mononuclear, Lymph Nodes cytology, Lymphocyte Activation immunology, RNA, Viral, Rituximab therapeutic use, Antigens, CD20 metabolism, CD4-Positive T-Lymphocytes drug effects, HIV Infections metabolism, Immunologic Factors pharmacology, Rituximab pharmacology, Virus Activation, Virus Latency
- Abstract
The identification of exclusive markers to target HIV-reservoir cells will represent a significant advance in the search for therapies to cure HIV. Here, we identify the B lymphocyte antigen CD20 as a marker for HIV-infected cells in vitro and in vivo. The CD20 molecule is dimly expressed in a subpopulation of CD4-positive (CD4
+ ) T lymphocytes from blood, with high levels of cell activation and heterogeneous memory phenotypes. In lymph node samples from infected patients, CD20 is present in productively HIV-infected cells, and ex vivo viral infection selectively upregulates the expression of CD20 during early infection. In samples from patients on antiretroviral therapy (ART) this subpopulation is significantly enriched in HIV transcripts, and the anti-CD20 monoclonal antibody Rituximab induces cell killing, which reduces the pool of HIV-expressing cells when combined with latency reversal agents. We provide a tool for targeting this active HIV-reservoir after viral reactivation in patients while on ART.- Published
- 2019
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23. CD32 is expressed on cells with transcriptionally active HIV but does not enrich for HIV DNA in resting T cells.
- Author
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Abdel-Mohsen M, Kuri-Cervantes L, Grau-Exposito J, Spivak AM, Nell RA, Tomescu C, Vadrevu SK, Giron LB, Serra-Peinado C, Genescà M, Castellví J, Wu G, Del Rio Estrada PM, González-Navarro M, Lynn K, King CT, Vemula S, Cox K, Wan Y, Li Q, Mounzer K, Kostman J, Frank I, Paiardini M, Hazuda D, Reyes-Terán G, Richman D, Howell B, Tebas P, Martinez-Picado J, Planelles V, Buzon MJ, Betts MR, and Montaner LJ
- Subjects
- Anti-Retroviral Agents therapeutic use, CD4-Positive T-Lymphocytes metabolism, HIV Infections drug therapy, Humans, In Vitro Techniques, Lymphocytes metabolism, Receptors, CCR4 metabolism, Receptors, CCR6 metabolism, Receptors, CXCR3 metabolism, HIV Infections metabolism, Receptors, IgG metabolism
- Abstract
The persistence of HIV reservoirs, including latently infected, resting CD4
+ T cells, is the major obstacle to cure HIV infection. CD32a expression was recently reported to mark CD4+ T cells harboring a replication-competent HIV reservoir during antiretroviral therapy (ART) suppression. We aimed to determine whether CD32 expression marks HIV latently or transcriptionally active infected CD4+ T cells. Using peripheral blood and lymphoid tissue of ART-treated HIV+ or SIV+ subjects, we found that most of the circulating memory CD32+ CD4+ T cells expressed markers of activation, including CD69, HLA-DR, CD25, CD38, and Ki67, and bore a TH 2 phenotype as defined by CXCR3, CCR4, and CCR6. CD32 expression did not selectively enrich for HIV- or SIV-infected CD4+ T cells in peripheral blood or lymphoid tissue; isolated CD32+ resting CD4+ T cells accounted for less than 3% of the total HIV DNA in CD4+ T cells. Cell-associated HIV DNA and RNA loads in CD4+ T cells positively correlated with the frequency of CD32+ CD69+ CD4+ T cells but not with CD32 expression on resting CD4+ T cells. Using RNA fluorescence in situ hybridization, CD32 coexpression with HIV RNA or p24 was detected after in vitro HIV infection (peripheral blood mononuclear cell and tissue) and in vivo within lymph node tissue from HIV-infected individuals. Together, these results indicate that CD32 is not a marker of resting CD4+ T cells or of enriched HIV DNA-positive cells after ART; rather, CD32 is predominately expressed on a subset of activated CD4+ T cells enriched for transcriptionally active HIV after long-term ART., (Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)- Published
- 2018
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24. A Novel Single-Cell FISH-Flow Assay Identifies Effector Memory CD4 + T cells as a Major Niche for HIV-1 Transcription in HIV-Infected Patients.
- Author
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Grau-Expósito J, Serra-Peinado C, Miguel L, Navarro J, Curran A, Burgos J, Ocaña I, Ribera E, Torrella A, Planas B, Badía R, Castellví J, Falcó V, Crespo M, and Buzon MJ
- Subjects
- Adult, CD4-Positive T-Lymphocytes ultrastructure, HIV Infections immunology, HIV-1 immunology, HIV-1 physiology, Humans, Immunologic Memory, Leukocytes, Mononuclear immunology, Leukocytes, Mononuclear virology, Middle Aged, RNA, Viral genetics, Receptors, IgG genetics, Single-Cell Analysis, Viral Load, Virus Latency, CD4-Positive T-Lymphocytes immunology, CD4-Positive T-Lymphocytes virology, Flow Cytometry methods, HIV Infections virology, HIV-1 genetics, In Situ Hybridization, Fluorescence methods, Transcription, Genetic
- Abstract
Cells that actively transcribe HIV-1 have been defined as the "active viral reservoir" in HIV-infected individuals. However, important technical limitations have precluded the characterization of this specific viral reservoir during both treated and untreated HIV-1 infections. Here, we used a novel single-cell RNA fluorescence in situ hybridization-flow cytometry (FISH-flow) assay that requires only 15 million unfractionated peripheral blood mononuclear cells (PBMCs) to characterize the specific cell subpopulations that transcribe HIV RNA in different subsets of CD4
+ T cells. In samples from treated and untreated HIV-infected patients, effector memory CD4+ T cells were the main cell population supporting HIV RNA transcription. The number of cells expressing HIV correlated with the plasma viral load, intracellular HIV RNA, and proviral DNA quantified by conventional methods and inversely correlated with the CD4+ T cell count and the CD4/CD8 ratio. We also found that after ex vivo infection of unstimulated PBMCs, HIV-infected T cells upregulated the expression of CD32. In addition, this new methodology detected increased numbers of primary cells expressing viral transcripts and proteins after ex vivo viral reactivation with latency reversal agents. This RNA FISH-flow technique allows the identification of the specific cell subpopulations that support viral transcription in HIV-1-infected individuals and has the potential to provide important information on the mechanisms of viral pathogenesis, HIV persistence, and viral reactivation. IMPORTANCE Persons infected with HIV-1 contain several cellular viral reservoirs that preclude the complete eradication of the viral infection. Using a novel methodology, we identified effector memory CD4+ T cells, immune cells preferentially located in inflamed tissues with potent activity against pathogens, as the main cells encompassing the transcriptionally active HIV-1 reservoir in patients on antiretroviral therapy. Importantly, the identification of such cells provides us with an important target for new therapies designed to target the hidden virus and thus to eliminate the virus from the human body. In addition, because of its ability to identify cells forming part of the viral reservoir, the assay used in this study represents an important new tool in the field of HIV pathogenesis and viral persistence., (Copyright © 2017 Grau-Expósito et al.)- Published
- 2017
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25. Lack of concordance between residual viremia and viral variants driving de novo infection of CD4(+) T cells on ART.
- Author
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Puertas MC, Noguera-Julian M, Massanella M, Pou C, Buzon MJ, Clotet B, Stevenson M, Paredes R, Blanco J, and Martinez-Picado J
- Subjects
- Antiretroviral Therapy, Highly Active, DNA, Viral genetics, HIV Infections drug therapy, HIV-1 drug effects, HIV-1 physiology, Humans, Male, Phylogeny, Proviruses genetics, RNA, Viral genetics, T-Lymphocyte Subsets virology, Viral Load, Viral Tropism, Anti-HIV Agents therapeutic use, CD4-Positive T-Lymphocytes drug effects, CD4-Positive T-Lymphocytes virology, Genetic Variation, HIV Infections virology, HIV-1 genetics, Viremia virology
- Abstract
Background: In most patients, current antiretroviral therapy (ART) regimens can rapidly reduce plasma viral load. However, even after years of effective treatment, a significant proportion of patients show residual plasma viremia below the clinical detection limit. Although residual viremia might be associated with increased chronic immune activation and morbidity, its origin and its potential role in the replenishment of the viral reservoir during suppressive ART is not completely understood. We performed an in-depth genetic analysis of the total and episomal cell-associated viral DNA (vDNA) repertoire in purified CD4(+) T cell subsets of three HIV-infected individuals, and used phylogenetic analysis to explore its relationship with plasma viruses., Results: The predominant proviral reservoir was established in naïve or memory (central and transitional) CD4(+) T cell subsets in patients harboring X4- or R5-tropic viruses, respectively. Regardless of the viral tropism, most plasma viruses detected under suppressive ART resembled the proviral reservoir identified in effector and transitional memory CD4(+) T-cell subsets in blood, suggesting that residual viremia originates from these cells in either blood or lymphoid tissue. Most importantly, sequences in episomal vDNA in CD4(+) T-cells were not well represented in residual viremia., Conclusions: Viral tropism determines the differential distribution of viral reservoir among CD4(+) T-cell subsets. In spite of viral tropism, the effector and transitional memory CD4(+) T-cells subsets are the main source of residual viremia during suppressive ART, even though their contribution to the total proviral pool is small. However, the lack of concordance between residual viremia and viral variants driving de novo infection of CD4(+) T cells on ART may reflect the predominance of defective plasma HIV RNA genomes. These findings highlight the need for monitoring the multiple viral RNA/DNA persistence markers, based on their differential contribution to viral persistence.
- Published
- 2016
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26. A Subset of Latency-Reversing Agents Expose HIV-Infected Resting CD4+ T-Cells to Recognition by Cytotoxic T-Lymphocytes.
- Author
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Jones RB, Mueller S, O'Connor R, Rimpel K, Sloan DD, Karel D, Wong HC, Jeng EK, Thomas AS, Whitney JB, Lim SY, Kovacs C, Benko E, Karandish S, Huang SH, Buzon MJ, Lichterfeld M, Irrinki A, Murry JP, Tsai A, Yu H, Geleziunas R, Trocha A, Ostrowski MA, Irvine DJ, and Walker BD
- Subjects
- Enzyme-Linked Immunosorbent Assay, Enzyme-Linked Immunospot Assay, Flow Cytometry, Humans, Polymerase Chain Reaction, Recombinant Fusion Proteins, Virus Activation drug effects, Antiviral Agents pharmacology, CD4-Positive T-Lymphocytes virology, HIV Infections immunology, Proteins pharmacology, T-Lymphocytes, Cytotoxic immunology, Virus Latency drug effects
- Abstract
Resting CD4+ T-cells harboring inducible HIV proviruses are a critical reservoir in antiretroviral therapy (ART)-treated subjects. These cells express little to no viral protein, and thus neither die by viral cytopathic effects, nor are efficiently cleared by immune effectors. Elimination of this reservoir is theoretically possible by combining latency-reversing agents (LRAs) with immune effectors, such as CD8+ T-cells. However, the relative efficacy of different LRAs in sensitizing latently-infected cells for recognition by HIV-specific CD8+ T-cells has not been determined. To address this, we developed an assay that utilizes HIV-specific CD8+ T-cell clones as biosensors for HIV antigen expression. By testing multiple CD8+ T-cell clones against a primary cell model of HIV latency, we identified several single agents that primed latently-infected cells for CD8+ T-cell recognition, including IL-2, IL-15, two IL-15 superagonists (IL-15SA and ALT-803), prostratin, and the TLR-2 ligand Pam3CSK4. In contrast, we did not observe CD8+ T-cell recognition of target cells following treatment with histone deacetylase inhibitors or with hexamethylene bisacetamide (HMBA). In further experiments we demonstrate that a clinically achievable concentration of the IL-15 superagonist 'ALT-803', an agent presently in clinical trials for solid and hematological tumors, primes the natural ex vivo reservoir for CD8+ T-cell recognition. Thus, our results establish a novel experimental approach for comparative evaluation of LRAs, and highlight ALT-803 as an LRA with the potential to synergize with CD8+ T-cells in HIV eradication strategies.
- Published
- 2016
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27. Correction: Integrated and Total HIV-1 DNA Predict Ex Vivo Viral Outgrowth.
- Author
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Kiselinova M, De Spiegelaere W, Buzon MJ, Malatinkova E, Lichterfeld M, and Vandekerckhove L
- Published
- 2016
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28. Integrated and Total HIV-1 DNA Predict Ex Vivo Viral Outgrowth.
- Author
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Kiselinova M, De Spiegelaere W, Buzon MJ, Malatinkova E, Lichterfeld M, and Vandekerckhove L
- Subjects
- Adult, Cohort Studies, DNA, Viral analysis, DNA, Viral isolation & purification, Disease Reservoirs virology, Female, HIV-1 genetics, Humans, Linear Models, Male, Middle Aged, RNA, Viral analysis, RNA, Viral isolation & purification, Viral Load, HIV Infections virology, HIV-1 growth & development, Virus Integration
- Abstract
The persistence of a reservoir of latently infected CD4 T cells remains one of the major obstacles to cure HIV. Numerous strategies are being explored to eliminate this reservoir. To translate these efforts into clinical trials, there is a strong need for validated biomarkers that can monitor the reservoir over time in vivo. A comprehensive study was designed to evaluate and compare potential HIV-1 reservoir biomarkers. A cohort of 25 patients, treated with suppressive antiretroviral therapy was sampled at three time points, with median of 2.5 years (IQR: 2.4-2.6) between time point 1 and 2; and median of 31 days (IQR: 28-36) between time point 2 and 3. Patients were median of 6 years (IQR: 3-12) on ART, and plasma viral load (<50 copies/ml) was suppressed for median of 4 years (IQR: 2-8). Total HIV-1 DNA, unspliced (us) and multiply spliced HIV-1 RNA, and 2LTR circles were quantified by digital PCR in peripheral blood, at 3 time points. At the second time point, a viral outgrowth assay (VOA) was performed, and integrated HIV-1 DNA and relative mRNA expression levels of HIV-1 restriction factors were quantified. No significant change was found for long- and short-term dynamics of all HIV-1 markers tested in peripheral blood. Integrated HIV-1 DNA was associated with total HIV-1 DNA (p<0.001, R² = 0.85), us HIV-1 RNA (p = 0.029, R² = 0.40), and VOA (p = 0.041, R2 = 0.44). Replication-competent virus was detected in 80% of patients by the VOA and it correlated with total HIV-1 DNA (p = 0.039, R² = 0.54). The mean quantification difference between Alu-PCR and VOA was 2.88 log10, and 2.23 log10 between total HIV-1 DNA and VOA. The levels of usHIV-1 RNA were inversely correlated with mRNA levels of several HIV-1 restriction factors (TRIM5α, SAMHD1, MX2, SLFN11, pSIP1). Our study reveals important correlations between the viral outgrowth and total and integrated HIV-1 DNA measures, suggesting that the total pool of HIV-1 DNA may predict the size of the replication-competent virus in ART suppressed patients.
- Published
- 2016
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29. Potent Cell-Intrinsic Immune Responses in Dendritic Cells Facilitate HIV-1-Specific T Cell Immunity in HIV-1 Elite Controllers.
- Author
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Martin-Gayo E, Buzon MJ, Ouyang Z, Hickman T, Cronin J, Pimenova D, Walker BD, Lichterfeld M, and Yu XG
- Subjects
- Blotting, Western, Flow Cytometry, Gene Knockdown Techniques, Humans, Lymphocyte Culture Test, Mixed, Polymerase Chain Reaction, RNA, Small Interfering, CD8-Positive T-Lymphocytes immunology, Dendritic Cells immunology, HIV Infections immunology, HIV-1 immunology, Immunity, Cellular immunology
- Abstract
The majority of HIV-1 elite controllers (EC) restrict HIV-1 replication through highly functional HIV-1-specific T cell responses, but mechanisms supporting the evolution of effective HIV-1-specific T cell immunity in these patients remain undefined. Cytosolic immune recognition of HIV-1 in conventional dendritic cells (cDC) can facilitate priming and expansion of HIV-1-specific T cells; however, HIV-1 seems to be able to avoid intracellular immune recognition in cDCs in most infected individuals. Here, we show that exposure of cDCs from EC to HIV-1 leads to a rapid and sustained production of type I interferons and upregulation of several interferon-stimulated effector genes. Emergence of these cell-intrinsic immune responses was associated with a reduced induction of SAMHD1 and LEDGF/p75, and an accumulation of viral reverse transcripts, but inhibited by pharmacological blockade of viral reverse transcription or siRNA-mediated silencing of the cytosolic DNA sensor cGAS. Importantly, improved cell-intrinsic immune recognition of HIV-1 in cDCs from elite controllers translated into stronger abilities to stimulate and expand HIV-1-specific CD8 T cell responses. These data suggest an important role of cell-intrinsic type I interferon secretion in dendritic cells for the induction of effective HIV-1-specific CD8 T cells, and may be helpful for eliciting functional T cell immunity against HIV-1 for preventative or therapeutic clinical purposes.
- Published
- 2015
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30. Transcriptional Changes in CD8(+) T Cells During Antiretroviral Therapy Intensified With Raltegravir.
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Ouyang Z, Buzon MJ, Zheng L, Sun H, Yu XG, Bosch RJ, Mellors JW, Eron JJ, Gandhi RT, and Lichterfeld M
- Abstract
Background. Intensification of antiretroviral therapy with raltegravir does not affect levels of residual human immunodeficiency virus (HIV)-1 viremia, but it has led to increased levels of episomal HIV-1 DNA in some patients, suggesting antiviral activity against otherwise unresponsive components of the viral reservoir. Effects of raltegravir on host cells remain less well understood. Methods. We used comprehensive and unbiased microarray-based transcriptional profiling to analyze gene expression changes in CD8(+) T cells from participants in a randomized clinical trial (AIDS Clinical Trials Group [ACTG] A5244) comparing raltegravir-intensified to nonintensified antiretroviral therapy. Results. Although raltegravir intensification failed to induce statistically significant changes in HIV-1 DNA or residual plasma viremia, we observed significant increases in the expression intensity of 121 host gene transcripts. In functional annotations of these transcripts, we found that they were mainly involved in glucose and carbohydrate metabolism, immune regulation, control of cell proliferation, and tumor suppression. Two of the raltegravir-responsive gene transcripts were statistically correlated with levels of residual HIV-1 RNA, but none of the remaining 119 transcripts were associated with immunologic or virologic characteristics of the study patients. Conclusions. Together, these findings demonstrate that raltegravir intensification can induce previously unrecognized, statistically significant gene expression changes in host CD8(+) T lymphocytes.
- Published
- 2015
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31. Long-term antiretroviral treatment initiated at primary HIV-1 infection affects the size, composition, and decay kinetics of the reservoir of HIV-1-infected CD4 T cells.
- Author
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Buzon MJ, Martin-Gayo E, Pereyra F, Ouyang Z, Sun H, Li JZ, Piovoso M, Shaw A, Dalmau J, Zangger N, Martinez-Picado J, Zurakowski R, Yu XG, Telenti A, Walker BD, Rosenberg ES, and Lichterfeld M
- Subjects
- Adult, Cohort Studies, DNA, Viral analysis, DNA, Viral genetics, Female, HIV Infections immunology, Humans, Male, Middle Aged, Time Factors, Treatment Outcome, Anti-Retroviral Agents administration & dosage, CD4-Positive T-Lymphocytes immunology, CD4-Positive T-Lymphocytes virology, HIV Infections drug therapy, HIV Infections virology, HIV-1 isolation & purification
- Abstract
Unlabelled: Initiation of antiretroviral therapy during the earliest stages of HIV-1 infection may limit the seeding of a long-lasting viral reservoir, but long-term effects of early antiretroviral treatment initiation remain unknown. Here, we analyzed immunological and virological characteristics of nine patients who started antiretroviral therapy at primary HIV-1 infection and remained on suppressive treatment for >10 years; patients with similar treatment duration but initiation of suppressive therapy during chronic HIV-1 infection served as controls. We observed that independently of the timing of treatment initiation, HIV-1 DNA in CD4 T cells decayed primarily during the initial 3 to 4 years of treatment. However, in patients who started antiretroviral therapy in early infection, this decay occurred faster and was more pronounced, leading to substantially lower levels of cell-associated HIV-1 DNA after long-term treatment. Despite this smaller size, the viral CD4 T cell reservoir in persons with early treatment initiation consisted more dominantly of the long-lasting central-memory and T memory stem cells. HIV-1-specific T cell responses remained continuously detectable during antiretroviral therapy, independently of the timing of treatment initiation. Together, these data suggest that early HIV-1 treatment initiation, even when continued for >10 years, is unlikely to lead to viral eradication, but the presence of low viral reservoirs and durable HIV-1 T cell responses may make such patients good candidates for future interventional studies aiming at HIV-1 eradication and cure., Importance: Antiretroviral therapy can effectively suppress HIV-1 replication to undetectable levels; however, HIV-1 can persist despite treatment, and viral replication rapidly rebounds when treatment is discontinued. This is mainly due to the presence of latently infected CD4 T cells, which are not susceptible to antiretroviral drugs. Starting treatment in the earliest stages of HIV-1 infection can limit the number of these latently infected cells, raising the possibility that these viral reservoirs are naturally eliminated if suppressive antiretroviral treatment is continued for extremely long periods of time. Here, we analyzed nine patients who started on antiretroviral therapy within the earliest weeks of the disease and continued treatment for more than 10 years. Our data show that early treatment accelerated the decay of infected CD4 T cells and led to very low residual levels of detectable HIV-1 after long-term therapy, levels that were otherwise detectable in patients who are able to maintain a spontaneous, drug-free control of HIV-1 replication. Thus, long-term antiretroviral treatment started during early infection cannot eliminate HIV-1, but the reduced reservoirs of HIV-1 infected cells in such patients may increase their chances to respond to clinical interventions aiming at inducing a drug-free remission of HIV-1 infection., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)
- Published
- 2014
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32. A cell-intrinsic inhibitor of HIV-1 reverse transcription in CD4(+) T cells from elite controllers.
- Author
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Leng J, Ho HP, Buzon MJ, Pereyra F, Walker BD, Yu XG, Chang EJ, and Lichterfeld M
- Subjects
- Amino Acid Sequence, Case-Control Studies, Cyclin-Dependent Kinase 2 genetics, Cyclin-Dependent Kinase Inhibitor p21 genetics, Enzyme Stability, HIV Reverse Transcriptase genetics, HIV-1 physiology, Host-Pathogen Interactions, Humans, Molecular Sequence Data, Phosphorylation, Reverse Transcription, Up-Regulation, Virus Replication, CD4-Positive T-Lymphocytes virology, Cyclin-Dependent Kinase 2 metabolism, Cyclin-Dependent Kinase Inhibitor p21 metabolism, HIV Reverse Transcriptase metabolism
- Abstract
HIV-1 reverse transcription represents the predominant target for pharmacological inhibition of viral replication, but cell-intrinsic mechanisms that can block HIV-1 reverse transcription in a clinically significant way are poorly defined. We find that effective HIV-1 reverse transcription depends on the phosphorylation of viral reverse transcriptase by host cyclin-dependent kinase (CDK) 2 at a highly conserved Threonine residue. CDK2-dependent phosphorylation increased the efficacy and stability of viral reverse transcriptase and enhanced viral fitness. Interestingly, p21, a cell-intrinsic CDK inhibitor that is upregulated in CD4(+) T cells from "elite controllers," potently inhibited CDK2-dependent phosphorylation of HIV-1 reverse transcriptase and significantly reduced the efficacy of viral reverse transcription. These data suggest that p21 can indirectly block HIV-1 reverse transcription by inhibiting host cofactors supporting HIV-1 replication and identify sites of viral vulnerability that are effectively targeted in persons with natural control of HIV-1 replication., (Copyright © 2014 Elsevier Inc. All rights reserved.)
- Published
- 2014
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33. Hepatitis C therapy with interferon-α and ribavirin reduces CD4 T-cell-associated HIV-1 DNA in HIV-1/hepatitis C virus-coinfected patients.
- Author
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Sun H, Buzon MJ, Shaw A, Berg RK, Yu XG, Ferrando-Martinez S, Leal M, Ruiz-Mateos E, and Lichterfeld M
- Subjects
- Adult, Aged, Antiviral Agents therapeutic use, CD4-Positive T-Lymphocytes immunology, Coinfection blood, Coinfection drug therapy, Coinfection immunology, Coinfection virology, Female, HIV Infections immunology, HIV Infections virology, HIV-1 isolation & purification, Hepatitis C immunology, Hepatitis C virology, Humans, Interferon-alpha therapeutic use, Male, Middle Aged, Retrospective Studies, CD4-Positive T-Lymphocytes virology, DNA, Viral blood, HIV Infections drug therapy, HIV-1 genetics, Hepatitis C drug therapy, Ribavirin therapeutic use, Viral Load drug effects
- Abstract
Combined treatment with interferon alpha (IFN-α) and ribavirin (RBV) can effectively cure HCV infection in a significant proportion of patients, but effects of this regimen on cellular reservoirs for human immunodeficiency virus type 1 (HIV-1) are unknown. Here, we show that treatment with IFN-α/RBV led to a moderate but significant and sustained decline of HIV-1 DNA in CD4 T cells from HIV-1/hepatitis C virus-coinfected patients receiving highly active antiretroviral therapy (n = 12). However, in vitro experiments failed to demonstrate an effect of pharmacological doses of IFN-α on HIV-1 reactivation. Together, these data suggest that treatment with IFN-α/RBV can moderately reduce the reservoir of HIV-1-infected CD4 T cells that persists despite suppressive antiretroviral therapy.
- Published
- 2014
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34. HIV-1 persistence in CD4+ T cells with stem cell-like properties.
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Buzon MJ, Sun H, Li C, Shaw A, Seiss K, Ouyang Z, Martin-Gayo E, Leng J, Henrich TJ, Li JZ, Pereyra F, Zurakowski R, Walker BD, Rosenberg ES, Yu XG, and Lichterfeld M
- Subjects
- Antiretroviral Therapy, Highly Active, Base Sequence, Boston, CD4-Positive T-Lymphocytes cytology, Cluster Analysis, Evolution, Molecular, Flow Cytometry, HIV Infections drug therapy, Humans, Molecular Sequence Data, Reverse Transcriptase Polymerase Chain Reaction, Sequence Analysis, DNA, Species Specificity, Statistics, Nonparametric, env Gene Products, Human Immunodeficiency Virus genetics, CD4-Positive T-Lymphocytes virology, Disease Reservoirs virology, HIV Infections virology, HIV-1 genetics, Lymphoid Progenitor Cells cytology, Phylogeny, Virus Latency
- Abstract
Cellular HIV-1 reservoirs that persist despite antiretroviral treatment are incompletely defined. We show that during suppressive antiretroviral therapy, CD4(+) T memory stem cells (TSCM cells) harbor high per-cell levels of HIV-1 DNA and make increasing contributions to the total viral CD4(+) T cell reservoir over time. Moreover, we conducted phylogenetic studies that suggested long-term persistence of viral quasispecies in CD4(+) TSCM cells. Thus, HIV-1 may exploit the stem cell characteristics of cellular immune memory to promote long-term viral persistence.
- Published
- 2014
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35. Intensification of a raltegravir-based regimen with maraviroc in early HIV-1 infection.
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Puertas MC, Massanella M, Llibre JM, Ballestero M, Buzon MJ, Ouchi D, Esteve A, Boix J, Manzardo C, Miró JM, Gatell JM, Clotet B, Blanco J, and Martinez-Picado J
- Subjects
- Adult, DNA, Viral blood, Female, HIV Infections virology, HIV-1 isolation & purification, Humans, Male, Maraviroc, Prospective Studies, RNA, Viral blood, Raltegravir Potassium, Treatment Outcome, Viral Load, Anti-HIV Agents therapeutic use, Antiretroviral Therapy, Highly Active methods, Cyclohexanes therapeutic use, HIV Infections drug therapy, Pyrrolidinones therapeutic use, Triazoles therapeutic use
- Abstract
Background: Latent HIV-1-infected cells generated early in the infection are responsible for viral persistence, and we hypothesized that addition of maraviroc to triple therapy in patients recently infected with HIV-1 could accelerate decay of the viral reservoir., Methods: Patients recently infected (<24 weeks) by chemokine receptor 5 (CCR5)-using HIV-1 were randomized to a raltegravir + tenofovir/emtricitabine regimen (control arm, n = 15) or the same regimen intensified with maraviroc (+MVC arm, n = 15). Plasma viral load, cell-associated HIV-1 DNA (total, integrated, and episomal), and activation/inflammation markers were measured longitudinally., Results: Plasma viral load decayed in both groups, reaching similar residual levels at week 48. Total cell-associated HIV-1 DNA also decreased in both groups during the first month, although subsequently at a slightly faster rate in the +MVC arm. The transient increase in two long terminal repeat (2-LTR) circles observed in both groups early after initiation of treatment decreased earlier in MVC-treated individuals. Early (week 12) increase of CD4 T-cell counts was higher in the +MVC arm. Conversely, CD8 T-cell counts and CD4 T-cell activation decreased slower in the +MVC arm. Absolute CD4 T-cell and CD8 T-cell counts, immune activation, CD4/CD8 T-cell ratio, and soluble inflammation markers were similar in both arms at the end of the study., Conclusion: Addition of maraviroc in early integrase inhibitor-based treatment of HIV-1 infection results in faster reduction of 2-LTR newly infected cells and recovery of CD4 T-cell counts, and a modest reduction in total reservoir size after 48 weeks of treatment. Paradoxically, CCR5 blockade also induced a slower decrease in plasma viremia and immune activation.
- Published
- 2014
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36. Susceptibility to CD8 T-cell-mediated killing influences the reservoir of latently HIV-1-infected CD4 T cells.
- Author
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Buzon MJ, Yang Y, Ouyang Z, Sun H, Seiss K, Rogich J, Le Gall S, Pereyra F, Rosenberg ES, Yu XG, and Lichterfeld M
- Subjects
- Adult, Aged, CD4-Positive T-Lymphocytes physiology, CD8-Positive T-Lymphocytes virology, Cytotoxicity, Immunologic physiology, Female, Flow Cytometry, HIV Infections immunology, HIV Infections virology, Humans, Lymphocyte Subsets physiology, Male, Middle Aged, Young Adult, CD4-Positive T-Lymphocytes virology, CD8-Positive T-Lymphocytes physiology, HIV Long-Term Survivors psychology, HIV-1 physiology
- Abstract
Background: HIV-1 establishes a lifelong infection in the human body, but host factors that influence viral persistence remain poorly understood. Cell-intrinsic characteristics of CD4 T cells, the main target cells for HIV-1, may affect the composition of the latent viral reservoir by altering the susceptibility to CD8 T-cell-mediated killing., Results: We observed that susceptibilities of CD4 T cells to CD8 T-cell-mediated killing, as determined in direct ex vivo assays, were significantly higher in persons with natural control of HIV-1 (elite controllers) than in individuals effectively treated with antiretroviral therapy. These differences were most pronounced in naive and in terminally differentiated CD4 T cells and corresponded to a reduced viral reservoir size in elite controllers. Interestingly, the highest susceptibility to CD8 T-cell-mediated killing and lowest reservoirs of cell-associated HIV-1 DNA was consistently observed in elite controllers expressing the protective HLA class I allele B57., Conclusions: These data suggest that the functional responsiveness of host CD4 T cells to cytotoxic effects of HIV-1-specific CD8 T cells can contribute to shaping the structure and composition of the latently infected CD4 T-cell pool.
- Published
- 2014
- Full Text
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37. Reactivation of latent HIV-1 in central memory CD4⁺ T cells through TLR-1/2 stimulation.
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Novis CL, Archin NM, Buzon MJ, Verdin E, Round JL, Lichterfeld M, Margolis DM, Planelles V, and Bosque A
- Subjects
- Adolescent, Adult, Humans, Lipopeptides immunology, Signal Transduction, Toll-Like Receptor 1 immunology, Toll-Like Receptor 2 immunology, Young Adult, CD4-Positive T-Lymphocytes immunology, CD4-Positive T-Lymphocytes virology, HIV-1 physiology, Lipopeptides metabolism, Toll-Like Receptor 1 agonists, Toll-Like Receptor 2 agonists, Virus Activation
- Abstract
Background: Toll-like receptors (TLRs) are crucial for recognition of pathogen-associated molecular patterns by cells of the innate immune system. TLRs are present and functional in CD4⁺ T cells. Memory CD4⁺ T cells, predominantly central memory cells (TCM), constitute the main reservoir of latent HIV-1. However, how TLR ligands affect the quiescence of latent HIV within central memory CD4⁺ T cells has not been studied., Results: We evaluated the ability of a broad panel of TLR agonists to reactivate latent HIV-1. The TLR-1/2 agonist Pam3CSK4 leads to viral reactivation of quiescent HIV in a model of latency based on cultured TCM and in resting CD4⁺ T cells isolated from aviremic patients. In addition, we investigated the signaling pathway associated with Pam3CSK4 involved in HIV-1 reactivation. We show that the transcription factors NFκB, NFAT and AP-1 cooperate to induce viral reactivation downstream of TLR-1/2 stimulation. Furthermore, increasing levels of cyclin T1 is not required for TLR-mediated viral reactivation, but induction of viral expression requires activated pTEFb. Finally, Pam3CSK4 reactivates latent HIV-1 in the absence of T cell activation or proliferation, in contrast to antigen stimulation., Conclusions: Our findings suggest that the signaling through TLR-1/2 pathway via Pam3CSK4 or other reagents should be explored as an anti-latency strategy either alone or in combination with other anti-latency drugs.
- Published
- 2013
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38. Modelling HIV-1 2-LTR dynamics following raltegravir intensification.
- Author
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Luo R, Cardozo EF, Piovoso MJ, Wu H, Buzon MJ, Martinez-Picado J, and Zurakowski R
- Subjects
- Antiretroviral Therapy, Highly Active methods, Disease Reservoirs virology, Humans, Pyrrolidinones administration & dosage, Raltegravir Potassium, Time Factors, DNA, Circular metabolism, DNA, Viral metabolism, HIV Infections drug therapy, HIV-1 genetics, Models, Biological, Pyrrolidinones pharmacology, Virus Replication drug effects
- Abstract
A model of reservoir activation and viral replication is introduced accounting for the production of 2-LTR HIV-1 DNA circles following antiviral intensification with the HIV integrase inhibitor raltegravir, considering contributions of de novo infection events and exogenous sources of infected cells, including quiescent infected cell activation. The model shows that a monotonic increase in measured 2-LTR concentration post intensification is consistent with limited de novo infection primarily maintained by sources of infected cells unaffected by raltegravir, such as quiescent cell activation, while a transient increase in measured 2-LTR concentration is consistent with significant levels of efficient (R0 > 1) de novo infection. The model is validated against patient data from the INTEGRAL study and is shown to have a statistically significant fit relative to the null hypothesis of random measurement variation about a mean. We obtain estimates and confidence intervals for the model parameters, including 2-LTR half-life. Seven of the 13 patients with detectable 2-LTR concentrations from the INTEGRAL study have measured 2-LTR dynamics consistent with significant levels of efficient replication of the virus prior to treatment intensification.
- Published
- 2013
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39. Increased coronary atherosclerosis and immune activation in HIV-1 elite controllers.
- Author
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Pereyra F, Lo J, Triant VA, Wei J, Buzon MJ, Fitch KV, Hwang J, Campbell JH, Burdo TH, Williams KC, Abbara S, and Grinspoon SK
- Subjects
- Angiography, CD4 Lymphocyte Count, Calcinosis physiopathology, Coronary Artery Disease diagnostic imaging, Coronary Artery Disease physiopathology, Disease Progression, Female, HIV Infections diagnostic imaging, HIV Infections physiopathology, Humans, Male, Middle Aged, Prevalence, Viral Load immunology, Calcinosis diagnostic imaging, Coronary Artery Disease immunology, HIV Infections immunology, HIV-1 immunology, Lymphocyte Activation immunology
- Abstract
HIV-1 elite controllers spontaneously maintain suppressed levels of viremia, but exhibit significant immune activation. We investigated coronary atherosclerosis by coronary computed tomography angiography (CTA) in elite controllers, nonelite controller, chronically HIV-1 infected, antiretroviral therapy (ART)-treated patients with undetectable viral load ('chronic HIV'), and HIV-negative controls. Prevalence of atherosclerosis (78 vs. 42%, P < 0.05) and markers of immune activation were increased in elite controllers compared with HIV-negative controls. sCD163, a monocyte activation marker, was increased in elite controllers compared with chronic HIV-1 (P < 0.05) and compared with HIV-negative controls (P < 0.05). These data suggest a significant degree of coronary atherosclerosis and monocyte activation among elite controllers.
- Published
- 2012
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40. CD4 T-cell regeneration in HIV-1 elite controllers.
- Author
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Yang Y, Al-Mozaini M, Buzon MJ, Beamon J, Ferrando-Martinez S, Ruiz-Mateos E, Rosenberg ES, Pereyra F, Yu XG, and Lichterfeld M
- Subjects
- ADP-ribosyl Cyclase 1 metabolism, Antiretroviral Therapy, Highly Active, CD4 Lymphocyte Count, CD4-Positive T-Lymphocytes immunology, CD8-Positive T-Lymphocytes metabolism, Case-Control Studies, Disease Progression, Flow Cytometry, HIV Infections drug therapy, HLA-DR Antigens metabolism, Humans, Membrane Glycoproteins metabolism, Protein-Tyrosine Kinases metabolism, CD4-Positive T-Lymphocytes metabolism, HIV Infections immunology, HIV Infections metabolism, HIV-1 immunology, Virus Replication
- Abstract
Background: Elite controllers spontaneously control HIV-1 replication, which in many cases is associated with preservation of normal CD4 T-cell counts. However, a subset of elite controllers has progressive CD4 T-cell losses despite undetectable viral loads, for reasons that remain undefined. Here, we assessed mechanisms of CD4 T-cell homeostasis in elite controllers with progressive vs. nonprogressive HIV-1 disease courses., Methods: Flow cytometry assays were used to determine the proliferation, activation and apoptosis levels of naive T cells in elite controllers with high or low CD4 T-cell counts and reference cohorts of HIV-1-negative and HAART-treated persons. Thymic output was measured by single-joint T-cell receptor excision circle (sjTREC)/β T-cell receptor excision circle (βTREC) ratios, and the frequency of circulating recent thymic emigrants was flow cytometrically determined by surface expression of protein tyrosine kinase 7., Results: Proportions of naive T cells in elite controllers were severely reduced and closely resemble those of HIV-1 patients with progressive disease. Despite reductions in naive T cells, most elite controllers were able to maintain normal total CD4 T-cell counts by preservation of uncompromised thymic function in conjunction with extrathymic processes that led to elevated levels of circulating recent thymic emigrants. In contrast, elite controllers with low CD4 T-cell counts had reduced thymic output that mirrored thymic dysfunction during untreated progressive HIV-1 infection., Conclusion: These results indicate that both thymic and extrathymic mechanisms contribute to CD4 T-cell maintenance in elite controllers and support the idea that CD4 T-cell homeostasis and control of viral replication are distinct but frequently coinciding processes.
- Published
- 2012
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41. A non-infectious cell-based phenotypic assay for the assessment of HIV-1 susceptibility to protease inhibitors.
- Author
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Buzon MJ, Erkizia I, Pou C, Minuesa G, Puertas MC, Esteve A, Castello A, Santos JR, Prado JG, Izquierdo-Useros N, Pattery T, Van Houtte M, Carrasco L, Clotet B, Ruiz L, and Martinez-Picado J
- Subjects
- Blotting, Western, Cells, Cultured, Cloning, Molecular, Genes, Reporter, Green Fluorescent Proteins genetics, Green Fluorescent Proteins metabolism, HIV Infections virology, HIV Protease genetics, HIV Protease metabolism, HIV-1 isolation & purification, Humans, Microbial Sensitivity Tests methods, Real-Time Polymerase Chain Reaction, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Anti-HIV Agents pharmacology, HIV Protease Inhibitors pharmacology, HIV-1 drug effects
- Abstract
Objectives: HIV-1 genotyping is widely accepted as a diagnostic tool to optimize therapy changes in patients whose antiretroviral regimen is failing. Phenotyping can substantially complement the information obtained from genotyping, especially in the presence of complex mutational patterns. However, drug susceptibility tests are laborious and require biosafety facilities. We describe the molecular mechanism of a non-infectious HIV-1 protease phenotypic assay in eukaryotic cells and validate its applicability as a tool for monitoring drug resistance., Methods: A cloning vector containing the fusion protein green fluorescent protein-HIV-1 protease (GFP-PR) was modified to facilitate the insertion of HIV-1 protease from infected subjects. Real-time quantitative PCR and western blot analysis were used to establish the molecular mechanism of the new phenotypic assay. The method was validated by analysing HIV-1 protease from 46 clinical isolates. Statistical comparisons were made between values obtained using our assay and those reported from alternative standardized phenotypic assays., Results: The capacity of HIV-1 protease to cleave cellular translation factors, such as the eukaryotic translation initiation factor 4 (eIF4GI) and the poly(A)-binding protein (PABP), led to cyclical accumulation of GFP that varied with the dose of protease inhibitors. Validation and comparison revealed a significant correlation with the Virco TYPE HIV-1 test (P < 0.0001, Spearman's ρ = 0.60), the Antivirogram test (P = 0.0001, Spearman's ρ = 0.60) and the Stanford HIVdb (P < 0.0001, Spearman's ρ = 0.69)., Conclusions: This cell-based non-infectious phenotypic method with a well-understood molecular mechanism was highly reliable and comparable to other widely used assays. The method can be used for both phenotyping of HIV-1 viral isolates resistant to protease inhibitors and screening of new protease inhibitors.
- Published
- 2012
- Full Text
- View/download PDF
42. Inhibition of HIV-1 integration in ex vivo-infected CD4 T cells from elite controllers.
- Author
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Buzon MJ, Seiss K, Weiss R, Brass AL, Rosenberg ES, Pereyra F, Yu XG, and Lichterfeld M
- Subjects
- Cells, Cultured, Humans, CD4-Positive T-Lymphocytes immunology, CD4-Positive T-Lymphocytes virology, HIV Infections immunology, HIV Long-Term Survivors, HIV-1 immunology, HIV-1 pathogenicity, Virus Integration
- Abstract
Elite controllers spontaneously maintain undetectable levels of HIV-1 replication for reasons that remain unclear. Here, we show that in elite controllers, direct ex vivo infection of purified CD4 T cells without prior in vitro activation results in disproportionately low levels of integrated HIV-1 DNA relative to the quantity of reverse transcripts, while the levels of two-long terminal repeat (2-LTR) circles were excessively elevated relative to those of integrated HIV-1 DNA. This indicates that chromosomal HIV-1 integration is inhibited in ex vivo-infected CD4 T cells from elite controllers. This defect in HIV-1 integration was unrelated to p21, a host protein that can restrict early HIV-1 replication steps, and was not visible following infection of in vitro-activated CD4 T cells from elite controllers. These data contribute to increasing evidence that intrinsic inhibition of specific HIV-1 replication steps plays an important role in the ability of elite controllers to maintain undetectable viral loads.
- Published
- 2011
- Full Text
- View/download PDF
43. Transcriptional profiling of CD4 T cells identifies distinct subgroups of HIV-1 elite controllers.
- Author
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Vigneault F, Woods M, Buzon MJ, Li C, Pereyra F, Crosby SD, Rychert J, Church G, Martinez-Picado J, Rosenberg ES, Telenti A, Yu XG, and Lichterfeld M
- Subjects
- CD4 Lymphocyte Count, CD8-Positive T-Lymphocytes immunology, Humans, Viral Load, CD4-Positive T-Lymphocytes immunology, CD4-Positive T-Lymphocytes virology, Gene Expression Profiling, HIV Infections immunology, HIV Long-Term Survivors, HIV-1 immunology
- Abstract
Human immunodeficiency virus type 1 (HIV-1) elite controllers maintain undetectable levels of viral replication in the absence of antiretroviral therapy (ART), but their underlying immunological and virological characteristics may vary. Here, we used a whole-genome transcriptional profiling approach to characterize gene expression signatures of CD4 T cells from an unselected cohort of elite controllers. The transcriptional profiles for the majority of elite controllers were similar to those of ART-treated patients but different from those of HIV-1-negative persons. Yet, a smaller proportion of elite controllers showed an alternative gene expression pattern that was indistinguishable from that of HIV-1-negative persons but different from that of highly active antiretroviral therapy (HAART)-treated individuals. Elite controllers with the latter gene expression signature had significantly higher CD4 T cell counts and lower levels of HIV-1-specific CD8(+) T cell responses but did not significantly differ from other elite controllers in terms of HLA class I alleles, HIV-1 viral loads determined by ultrasensitive single-copy PCR assays, or chemokine receptor polymorphisms. Thus, these data identify a specific subgroup of elite controllers whose immunological and gene expression characteristics approximate those of HIV-1-negative persons.
- Published
- 2011
- Full Text
- View/download PDF
44. Transient treatment exclusively containing nucleoside analogue reverse transcriptase inhibitors in highly antiretroviral-experienced patients preserves viral benefit when a fully active therapy was initiated.
- Author
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Bonjoch A, Buzon MJ, Llibre JM, Negredo E, Puig J, Pérez-Alvarez N, Videla S, Martinez-Picado J, and Clotet B
- Subjects
- Adult, Anti-HIV Agents therapeutic use, CD4 Lymphocyte Count, Drug Administration Schedule, Drug Resistance, Viral genetics, Drug Therapy, Combination, Female, HIV Infections immunology, HIV Infections virology, HIV Protease genetics, HIV Reverse Transcriptase genetics, HIV-1 physiology, Humans, Male, Middle Aged, Mutation, Pilot Projects, RNA, Viral blood, Reverse Transcriptase Inhibitors therapeutic use, Treatment Outcome, Viral Load, Anti-HIV Agents administration & dosage, HIV Infections drug therapy, HIV-1 drug effects, Reverse Transcriptase Inhibitors administration & dosage
- Abstract
Background: We determined whether coformulated zidovudine/lamivudine/abacavir plus tenofovir could maintain immune status in comparison with a genotype-guided salvage regimen in highly pretreated patients., Method: This was a randomized pilot control-arm study. The primary endpoint was the proportion of patients who maintained their CD4+ T-cell count at Week 48., Results: Thirteen patients were randomized to the study arm and 10 to the control arm. At 48 weeks, 8 (64%) patients in the study arm and 10 (100%) in the control arm maintained their immune status (p = .09). No new AIDS-defining events occurred. Three patients (27%) in the study arm and 5 (50%) in the control arm achieved an undetectable viral load (p = .39). When a fully suppressive regimen was initiated, 69% of patients in the study arm (9 patients) and 60% (6 patients) in the control arm reached <50 copies at 96 weeks (p = .98)., Conclusion: Although no statistically significant differences in immunological course were observed between the arms, the control group achieved better results after 48 weeks. This transient therapy could be reserved for specific patients in whom the risk of incomplete adherence or toxicity compromises efficacy while they are awaiting a fully active drug, without jeopardizing viral efficacy when a fully suppressive regimen is initiated.
- Published
- 2008
- Full Text
- View/download PDF
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