Publications by authors named "Eva Riveira-Munoz"

51 Publications

SARS-CoV-2 Infection Modulates ACE2 Function and Subsequent Inflammatory Responses in Swabs and Plasma of COVID-19 Patients.

Viruses 2021 08 28;13(9). Epub 2021 Aug 28.

IrsiCaixa-AIDS Research Institute, Health Research Institute Germans Trias i Pujol (IGTP), Universitat Autònoma de Barcelona, 08916 Badalona, Spain.

Angiotensin converting enzyme 2 (ACE2) is a host ectopeptidase and the receptor for the SARS-CoV-2 virus, albeit virus-ACE2 interaction goes far beyond viral entry into target cells. Controversial data exists linking viral infection to changes in ACE2 expression and function, which might influence the subsequent induction of an inflammatory response. Here, we tested the significance of soluble ACE2 enzymatic activity longitudinally in nasopharyngeal swabs and plasma samples of SARS-CoV-2 infected patients, along with the induction of inflammatory cytokines. Release of soluble functional ACE2 increases upon SARS-CoV-2 infection in swabs and plasma of infected patients, albeit rapidly decreasing during infection course in parallel with gene expression. Similarly, SARS-CoV-2 infection also induced the expression of inflammatory cytokines. These changes positively correlated with the viral load. Overall, our results demonstrate the existence of mechanisms by which SARS-CoV-2 modulates ACE2 expression and function, intracellular viral sensing and subsequent inflammatory response, offering new insights into ACE2 dynamics in the human upper respiratory tract and pointing towards soluble ACE2 levels as a putative early biomarker of infection severity.
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http://dx.doi.org/10.3390/v13091715DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8471465PMC
August 2021

Critical Presentation of a Severe Acute Respiratory Syndrome Coronavirus 2 Reinfection: A Case Report.

Open Forum Infect Dis 2021 Jul 23;8(7):ofab329. Epub 2021 Jun 23.

IrsiCaixa-AIDS Research Institute and Germans Trias i Pujol Health Research Institute, Badalona, Catalonia, Spain.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reinfections have been reported; however, most cases are milder than the primary infection. We report the first case of a life-threatening critical presentation of a SARS-CoV-2 reinfection.

Methods: A 62-year-old man from Palamós (Spain) suffered a first mild coronavirus disease 2019 (COVID-19) episode in March 2020, confirmed by 2 independent SARS-CoV-2 nasopharyngeal polymerase chain reaction (PCR) assays and a normal radiograph. He recovered completely and tested negative on 2 consecutive PCRs. In August 2020, the patient developed a second SARS-CoV-2 infection with life-threatening bilateral pneumonia and Acute respiratory distress syndrome criteria, requiring COVID-19-specific treatment (remdesivir + dexamethasone) plus high-flow oxygen therapy. Nasopharyngeal swabs from the second episode were obtained for virus quantification by real-time PCR, for virus outgrowth and sequencing. In addition, plasma and peripheral blood mononuclear cells during the hospitalization period were used to determine SARS-CoV-2-specific humoral and T-cell responses.

Results: Genomic analysis of SARS-CoV-2 showed that the virus had probably originated shortly before symptom onset. When the reinfection occurred, the subject showed a weak immune response, with marginal humoral and specific T-cell responses against SARS-CoV-2. All antibody isotypes tested as well as SARS-CoV-2 neutralizing antibodies increased sharply after day 8 postsymptoms. A slight increase of T-cell responses was observed at day 19 after symptom onset.

Conclusions: The reinfection was firmly documented and occurred in the absence of robust preexisting humoral and cellular immunity. SARS-CoV-2 immunity in some subjects is unprotective and/or short-lived; therefore, SARS-CoV-2 vaccine schedules inducing long-term immunity will be required to bring the pandemic under control.
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http://dx.doi.org/10.1093/ofid/ofab329DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8320276PMC
July 2021

A Cluster-Randomized Trial of Hydroxychloroquine for Prevention of Covid-19.

N Engl J Med 2021 02 24;384(5):417-427. Epub 2020 Nov 24.

From the Fight AIDS and Infectious Diseases Foundation (O.M., M.C.-M., M.U., A.A., C.S., E.B., C.A.P., P.A., N.R.-M., P.L., J.M., M.C., L.B., M.S., S.G., A.N., J. Puig, F.R.-V., A. Sierra, M.V.-M., C.G.-B., B.C.), Hospital Universitari Germans Trias i Pujol, Universitat Autònoma de Barcelona (O.M., J.A., M.F., C.Q., M.V.-M., B.C.), IrsiCaixa AIDS Research Institute, Germans Trias i Pujol Research Institute (E.B., A.E.-T., E.R.-M., L.R.), and the Center of Epidemiologic Studies of HIV/AIDS and STI of Catalonia, Catalan Institute of Oncology-Departament de Salut, Generalitat de Catalunya (E.M., J.R.-U., A. Sentis), Badalona, Facultat de Medicina-Universitat de Barcelona (M.C.-M., M.U.), Institute of Environmental Assessment and Water Research, Spanish Council for Scientific Research (A.T.), Direcció-Gerència, Institut Català de la Salut (J.M.A., J.C.), Equip d'Atenció Primària de Sarrià (G.C.), Synlab Diagnósticos (P.C.), Direcció General de Recerca i Innovació en Salut, Generalitat de Catalunya (R.F.), TFS Clinical Contract Research Organization (C.L., J.Z.), Gerència Territorial de Barcelona, Institut Català de la Salut (N.N.), ISGlobal, Hospital Clínic-Universitat de Barcelona (S.S.), and Agència de Qualitat i Avaluació Sanitàries de Catalunya (C.V., R.M.V.-H.), Barcelona, Bellvitge Biomedical Research Institute, L'Hospitalet de Llobregat (C.T., J. Peñafiel), Gerència Territorial de Catalunya Central, Institut Català de la Salut, Sant Fruitós de Bages (A.F.), Xarxa Santa Tecla Sanitària i Social, Tarragona (G.F.-M.), Entitat de Base Asociativa Centelles-Atenció Primària, Centelles (S.N.), Gerència Territorial de Àmbit Metropolità Nord, Institut Català de la Salut, Sabadell (N.P.), Centro de Investigación Biomédica en Red de Epidemiología y Salud Pública, Madrid (J.C.), and Universitat de Vic-Universitat Central de Catalunya, Vic (O.M., B.C.) - all in Spain; and Lihir Medical Center-International SOS, Lihir Island, Papua New Guinea (O.M.).

Background: Current strategies for preventing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection are limited to nonpharmacologic interventions. Hydroxychloroquine has been proposed as a postexposure therapy to prevent coronavirus disease 2019 (Covid-19), but definitive evidence is lacking.

Methods: We conducted an open-label, cluster-randomized trial involving asymptomatic contacts of patients with polymerase-chain-reaction (PCR)-confirmed Covid-19 in Catalonia, Spain. We randomly assigned clusters of contacts to the hydroxychloroquine group (which received the drug at a dose of 800 mg once, followed by 400 mg daily for 6 days) or to the usual-care group (which received no specific therapy). The primary outcome was PCR-confirmed, symptomatic Covid-19 within 14 days. The secondary outcome was SARS-CoV-2 infection, defined by symptoms compatible with Covid-19 or a positive PCR test regardless of symptoms. Adverse events were assessed for up to 28 days.

Results: The analysis included 2314 healthy contacts of 672 index case patients with Covid-19 who were identified between March 17 and April 28, 2020. A total of 1116 contacts were randomly assigned to receive hydroxychloroquine and 1198 to receive usual care. Results were similar in the hydroxychloroquine and usual-care groups with respect to the incidence of PCR-confirmed, symptomatic Covid-19 (5.7% and 6.2%, respectively; risk ratio, 0.86 [95% confidence interval, 0.52 to 1.42]). In addition, hydroxychloroquine was not associated with a lower incidence of SARS-CoV-2 transmission than usual care (18.7% and 17.8%, respectively). The incidence of adverse events was higher in the hydroxychloroquine group than in the usual-care group (56.1% vs. 5.9%), but no treatment-related serious adverse events were reported.

Conclusions: Postexposure therapy with hydroxychloroquine did not prevent SARS-CoV-2 infection or symptomatic Covid-19 in healthy persons exposed to a PCR-positive case patient. (Funded by the crowdfunding campaign YoMeCorono and others; BCN-PEP-CoV2 ClinicalTrials.gov number, NCT04304053.).
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http://dx.doi.org/10.1056/NEJMoa2021801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7722693PMC
February 2021

New indolylarylsulfone non-nucleoside reverse transcriptase inhibitors show low nanomolar inhibition of single and double HIV-1 mutant strains.

Eur J Med Chem 2020 Dec 11;208:112696. Epub 2020 Aug 11.

Laboratory Affiliated to Istituto Pasteur Italia - Fondazione Cenci Bolognetti, Dipartimento di Chimica e Tecnologie Del Farmaco, Sapienza Università di Roma, Piazzale Aldo Moro 5, 00185, Rome, Italy. Electronic address:

We designed and synthesized 21 new indolylarylsulfones (IASs) as new HIV-1 NNRTIs. Among these, IAS 12 exhibited a remarkable antiviral activity against single and double mutants (K103N EC = <0.7 nM; Y181C EC = <0.7 nM; Y188L EC = 21.3 nM; K103N-Y181C EC = 6.2 nM), resulting equally or more active than previuosly reported IAS 6 and some approved anti-HIV-1 drugs. Docking and molecular dynamics simulations of compound 12 in complex with WT, Y181C, Y188L, K103N and K103N-Y181C RTs clarified a general binding mode that was consistent with biological results. Kinetic experiments disclosed that derivative 12 preferentially binds WT and K103N-Y181C RTs to binary and ternary complexes, respectively.
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http://dx.doi.org/10.1016/j.ejmech.2020.112696DOI Listing
December 2020

Hydroxychloroquine for Early Treatment of Adults with Mild Covid-19: A Randomized-Controlled Trial.

Clin Infect Dis 2020 Jul 16. Epub 2020 Jul 16.

Gerència territorial de Barcelona, Institut Català de la Salut, Barcelona, Spain.

Background: No therapeutics have yet been proven effective for the treatment of mild-illness caused by SARS-CoV-2. We aimed to determine whether early treatment with hydroxychloroquine (HCQ) would be more efficacious than no-treatment for outpatients with mild Covid-19.

Methods: We conducted a multicenter, open label, randomized controlled trial in Catalonia (Spain) between March 17, and May 26, 2020. Eligible Covid-19 cases were non-hospitalized adult patients with recently confirmed SARS-CoV-2 infection and less than five days of symptoms. Patients were assigned to receive HCQ (800 mg on day 1, followed by 400 mg once daily for 6 days) or no antiviral treatment (not-placebo controlled). Study outcomes were the reduction of viral RNA load in nasopharyngeal swabs up to 7 days after treatment start, patient disease progression using the WHO scale up to 28 days, and time to complete resolution of symptoms. Adverse events were assessed up to 28 days.

Results: A total of 293 patients were eligible for intention-to-treat analysis: 157 in the control arm and 136 in the intervention arm. The mean age was 41.6 years (SD 12.6), mean viral load at baseline was 7.90 (SD 1.82) Log10 copies/mL, and median time from symptom onset to randomization was 3 days. No significant differences were found in the mean reduction of viral load at day 3 (-1.41 vs. -1.41 Log10 copies/mL in the control and intervention arm, respectively; difference 0.01 [95% CI -0.28;0.29]) or at day 7 (-3.37 vs. -3.44; d -0.07 [-0.44;0.29]). This treatment regimen did not reduce risk of hospitalization (7.1%, control vs. 5.9%, intervention; RR 0.75 [0.32;1.77]) nor shortened the time to complete resolution of symptoms (12 days, control vs. 10 days, intervention; p = 0.38). No relevant treatment-related AEs were reported.

Conclusions: In patients with mild Covid-19, no benefit was observed with HCQ beyond the usual care.
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http://dx.doi.org/10.1093/cid/ciaa1009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7454406PMC
July 2020

Targeting HIV-1 RNase H: -(2-Hydroxy-benzylidene)-3,4,5-Trihydroxybenzoylhydrazone as Selective Inhibitor Active against NNRTIs-Resistant Variants.

Viruses 2020 07 6;12(7). Epub 2020 Jul 6.

Department of Life and Environmental Sciences University of Cagliari, Cittadella Universitaria di Monserrato, 09042 Monserrato, Italy.

HIV-1 infection requires life-long treatment and with 2.1 million new infections/year, faces the challenge of an increased rate of transmitted drug-resistant mutations. Therefore, a constant and timely effort is needed to identify new HIV-1 inhibitors active against drug-resistant variants. The ribonuclease H (RNase H) activity of HIV-1 reverse transcriptase (RT) is a very promising target, but to date, still lacks an efficient inhibitor. Here, we characterize the mode of action of -(2-hydroxy-benzylidene)-3,4,5-trihydroxybenzoylhydrazone (compound ), an N-acylhydrazone derivative that inhibited viral replication (EC = 10 µM), while retaining full potency against the NNRTI-resistant double mutant K103N-Y181C virus. Time-of-addition and biochemical assays showed that compound targeted the reverse-transcription step in cell-based assays and inhibited the RT-associated RNase H function, being >20-fold less potent against the RT polymerase activity. Docking calculations revealed that compound binds within the RNase H domain in a position different from other selective RNase H inhibitors; site-directed mutagenesis studies revealed interactions with conserved amino acid within the RNase H domain, suggesting that compound can be taken as starting point to generate a new series of more potent RNase H selective inhibitors active against circulating drug-resistant variants.
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http://dx.doi.org/10.3390/v12070729DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7412550PMC
July 2020

Pharmacological Modulation of SAMHD1 Activity by CDK4/6 Inhibitors Improves Anticancer Therapy.

Cancers (Basel) 2020 Mar 18;12(3). Epub 2020 Mar 18.

IrsiCaixa AIDS Research Institute, Badalona, 08916 Catalonia, Spain.

Sterile alpha motif and histidine-aspartic acid domain-containing protein 1 (SAMHD1) is a dNTP triphosphohydrolase involved in the regulation of the intracellular dNTP pool, linked to viral restriction, cancer development and autoimmune disorders. SAMHD1 function is regulated by phosphorylation through a mechanism controlled by cyclin-dependent kinases and tightly linked to cell cycle progression. Recently, SAMHD1 has been shown to decrease the efficacy of nucleotide analogs used as chemotherapeutic drugs. Here, we demonstrate that SAMHD1 can enhance or decrease the efficacy of various classes of anticancer drug, including nucleotide analogues, but also anti-folate drugs and CDK inhibitors. Importantly, we show that selective CDK4/6 inhibitors are pharmacological activators of SAMHD1 that act by inhibiting its inactivation by phosphorylation. Combinations of a CDK4/6 inhibitor with nucleoside or folate antimetabolites potently enhanced drug efficacy, resulting in highly synergic drug combinations (CI < 0.04). Mechanistic analyses reveal that cell cycle-controlled modulation of SAMHD1 function is the central process explaining changes in anticancer drug efficacy, therefore providing functional proof of the potential of CDK4/6 inhibitors as a new class of adjuvants to boost chemotherapeutic regimens. The evaluation of SAMHD1 expression in cancer tissues allowed for the identification of cancer types that would benefit from the pharmacological modulation of SAMHD1 function. In conclusion, these results indicate that the modulation of SAMHD1 function may represent a promising strategy for the improvement of current antimetabolite-based treatments.
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http://dx.doi.org/10.3390/cancers12030713DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7140116PMC
March 2020

ADAR1 function affects HPV replication and is associated to recurrent human papillomavirus-induced dysplasia in HIV coinfected individuals.

Sci Rep 2019 12 27;9(1):19848. Epub 2019 Dec 27.

AIDS Research Institute-IrsiCaixa, Badalona, Spain.

Infection by human papillomavirus (HPV) alters the microenvironment of keratinocytes as a mechanism to evade the immune system. A-to-I editing by ADAR1 has been reported to regulate innate immunity in response to viral infections. Here, we evaluated the role of ADAR1 in HPV infection in vitro and in vivo. Innate immune activation was characterized in human keratinocyte cell lines constitutively infected or not with HPV. ADAR1 knockdown induced an innate immune response through enhanced expression of RIG-I-like receptors (RLR) signaling cascade, over-production of type-I IFNs and pro-inflammatory cytokines. ADAR1 knockdown enhanced expression of HPV proteins, a process dependent on innate immune function as no A-to-I editing could be identified in HPV transcripts. A genetic association study was performed in a cohort of HPV/HIV infected individuals followed for a median of 6 years (range 0.1-24). We identified the low frequency haplotype AACCAT significantly associated with recurrent HPV dysplasia, suggesting a role of ADAR1 in the outcome of HPV infection in HIV+ individuals. In summary, our results suggest that ADAR1-mediated innate immune activation may influence HPV disease outcome, therefore indicating that modification of innate immune effectors regulated by ADAR1 could be a therapeutic strategy against HPV infection.
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http://dx.doi.org/10.1038/s41598-019-56422-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6934649PMC
December 2019

Dual effect of the broad spectrum kinase inhibitor midostaurin in acute and latent HIV-1 infection.

Antiviral Res 2019 08 8;168:18-27. Epub 2019 May 8.

AIDS Research Institute - IrsiCaixa, AIDS Unit and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain. Electronic address:

Midostaurin is a multi-kinase inhibitor with antineoplastic activity. We assessed the capacity of midostaurin to affect early and late steps of HIV-1 infection and to reactivate HIV-1 latently infected cells, alone or in combination with histone deacetylase inhibitors (HDACi) known to act as latency-reversing agents (LRA). Acute HIV-1 infection was assessed by flow cytometry in three cell types treated with midostaurin in the presence or absence of SAMHD1. Non-infected cells were treated with midostaurin and harvested for Western blot analysis. Macrophage infections were also measured by quantitative RT-PCR. HIV-1 latency reactivation was assessed in several latency models. Midostaurin induced G2/M arrest and inhibited CDK2, preventing the phosphorylation of SAMHD1 associated to inhibition of its dNTPase activity. In the presence of SAMHD1, midostaurin blocked HIV-1 DNA formation and viral replication. However, following Vpx-mediated SAMHD1 degradation, midostaurin increased viral transcripts and virus replication. In three out of four HIV-1 latency models, including primary CD4 T cells, midostaurin effectively reversed HIV-1 latency and was synergistic in combination with LRA vorinostat and panobinostat. Our study describes a dual effect for midostaurin in HIV-1 infection, antiviral or proviral depending on SAMHD1 activation, and highlights a role for active SAMHD1 in regulating the activity of potential HIV-1 latency reversal agents.
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http://dx.doi.org/10.1016/j.antiviral.2019.05.003DOI Listing
August 2019

Effect of α-Methoxy Substitution on the Anti-HIV Activity of Dihydropyrimidin-4(3 H)-ones.

J Med Chem 2019 01 20;62(2):604-621. Epub 2018 Dec 20.

Dipartimento di Chimica e Tecnologie del Farmaco , Università degli Studi di Roma "La Sapienza" , P. le A. Moro 5 , 00185 Roma , Italy.

Conformational restriction applied to dihydrobenzylpyrimidin-4-(3 H)-ones (DABOs) by the intoduction of a methyl group at the α-benzylic position is known to massively improve the anti-HIV-1 activity of these compounds. Here, we report the effects of methoxy substitution at the α-benzylic position in S-, NH-, and N, N-DABOs carrying 2,6-difluoro, 2-chloro-6-fluoro, or 2,6-dichloro substituted benzyl moieties. The various α-methoxy DABO series (12-14) present different SAR at the dihalo benzyl substitution, with the most potent compounds (12d,e and 13c) showing similar (picomolar/nanomolar) anti-HIV-1 potency as the corresponding α-methyl analogues against wt HIV-1, and 10-100-fold increased potency (up to low nanomolar) against clinically relevant K103N, Y181C, Y188L, IRLL98, and K103N+Y181C HIV-1 mutant strains, highlighting the importance of the α-methoxy substitution to provide highly efficient DABOs as "second generation" NNRTIs. HPLC enantioseparation of three of the most potent derivatives (12d, 13c, and 14c) provided single enantiomers with significant enantioselectivity in HIV-1 inhibition. Computational studies allowed to correlate the best antiviral activity with the ( R) absolute configuration at the α-methoxy stereogenic center.
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http://dx.doi.org/10.1021/acs.jmedchem.8b01238DOI Listing
January 2019

Meeting report: 31 International Conference on Antiviral Research.

Antiviral Res 2018 10 4;158:88-102. Epub 2018 Aug 4.

AIDS Research Institute - Irsicaixa, Hospital Germans Trias i Pujol, Universitat Autónoma de Barcelona, Badalona, Spain. Electronic address:

The 31 International Conference on Antiviral Research (ICAR) was held in Porto, Portugal from June 11-15, 2018. In this report, volunteer rapporteurs provide their summaries of scientific presentations, hoping to effectively convey the speakers' goals and the results and conclusions of their talks. This report provides an overview of the invited keynote and award lectures and highlights of short oral presentations, from the perspective of experts in antiviral research. Of note, a session on human cytomegalovirus included an update on the introduction to the clinic of letermovir for the prevention of CMV infection and disease. The 31 ICAR successfully promoted new discoveries in antiviral research and drug development. The 32 ICAR will be held in Baltimore, Maryland, USA, May 6-10, 2019.
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http://dx.doi.org/10.1016/j.antiviral.2018.08.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7113893PMC
October 2018

CD32 expression is associated to T-cell activation and is not a marker of the HIV-1 reservoir.

Nat Commun 2018 07 16;9(1):2739. Epub 2018 Jul 16.

AIDS Research Institute-IrsiCaixa and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916, Badalona, Spain.

CD32 has been shown to be preferentially expressed in latently HIV-1-infected cells in an in vitro model of quiescent CD4 T cells. Here we show that stimulation of CD4+ T cells with IL-2, IL-7, PHA, and anti-CD3/CD28 antibodies induces T-cell proliferation, co-expression of CD32 and the activation of the markers HLA-DR and CD69. HIV-1 infection increases CD32 expression. 79.2% of the CD32+/CD4+ T cells from HIV+ individuals under antiretroviral treatment were HLA-DR+. Resting CD4+ T cells infected in vitro generally results in higher integration of provirus. We observe no difference in provirus integration or replication-competent inducible latent HIV-1 in CD32+ or CD32- CD4+ T cells from HIV+ individuals. Our results demonstrate that CD32 expression is a marker of CD4+ T cell activation in HIV+ individuals and raises questions regarding the immune resting status of CD32+ cells harboring HIV-1 proviruses.
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http://dx.doi.org/10.1038/s41467-018-05157-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6048139PMC
July 2018

ADAR1 affects HCV infection by modulating innate immune response.

Antiviral Res 2018 08 12;156:116-127. Epub 2018 Jun 12.

AIDS Research Institute-IrsiCaixa and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain. Electronic address:

The hepatitis C virus (HCV) is a globally prevalent infectious pathogen. As many as 80% of people infected with HCV do not control the virus and develop a chronic infection. Response to interferon (IFN) therapy is widely variable in chronic HCV infected patients, suggesting that HCV has evolved mechanisms to suppress and evade innate immunity responsible for its control and elimination. Adenosine deaminase acting on RNA 1 (ADAR1) is a relevant factor in the regulation of the innate immune response. The loss of ADAR1 RNA-editing activity and the resulting loss of inosine bases in RNA are critical in producing aberrant RLR-mediated innate immune response, mediated by RNA sensors MDA5 and RIG-I. Here, we describe ADAR1 role as a regulator of innate and antiviral immune function in HCV infection, both in vitro and in patients. Polymorphisms within ADAR1 gene were found significantly associated to poor clinical outcome to HCV therapy and advanced liver fibrosis in a cohort of HCV and HIV-1 coinfected patients. Moreover, ADAR1 knockdown in primary macrophages and Huh7 hepatoma cells enhanced IFN and IFN stimulated gene expression and increased HCV replication in vitro. Overall, our results demonstrate that ADAR1 regulates innate immune signaling and is an important contributor to the outcome of the HCV virus-host interaction. ADAR1 is a potential target to boost antiviral immune response in HCV infection.
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http://dx.doi.org/10.1016/j.antiviral.2018.05.012DOI Listing
August 2018

RNA editing by ADAR1 regulates innate and antiviral immune functions in primary macrophages.

Sci Rep 2017 10 17;7(1):13339. Epub 2017 Oct 17.

AIDS Research Institute-IrsiCaixa and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain.

ADAR1-dependent A-to-I editing has recently been recognized as a key process for marking dsRNA as self, therefore, preventing innate immune activation and affecting the development and resolution of immune-mediated diseases and infections. Here, we have determined the role of ADAR1 as a regulator of innate immune activation and modifier of viral susceptibility in primary myeloid and lymphoid cells. We show that ADAR1 knockdown significantly enhanced interferon, cytokine and chemokine production in primary macrophages that function as antiviral paracrine factors, rendering them resistant to HIV-1 infection. ADAR1 knockdown induced deregulation of the RLRs-MAVS signaling pathway, by increasing MDA5, RIG-I, IRF7 and phospho-STAT1 expression, an effect that was partially rescued by pharmacological blockade of the pathway. In summary, our results demonstrate a role of ADAR1 in regulating innate immune function in primary macrophages, suggesting that macrophages may play an essential role in disease associated to ADAR1 dysfunction. We also show that viral inhibition is exclusively dependent on innate immune activation consequence of ADAR1 knockdown, pointing towards ADAR1 as a potential target to boost antiviral immune response.
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http://dx.doi.org/10.1038/s41598-017-13580-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5645456PMC
October 2017

Evaluation of the Innate Immune Modulator Acitretin as a Strategy To Clear the HIV Reservoir.

Antimicrob Agents Chemother 2017 11 24;61(11). Epub 2017 Oct 24.

AIDS Research Institute IrsiCaixa, AIDS Unit and Health Research Institute Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain

The persistence of HIV despite suppressive antiretroviral therapy is a major roadblock to HIV eradication. Current strategies focused on inducing the expression of latent HIV fail to clear the persistent reservoir, prompting the development of new approaches for killing HIV-positive cells. Recently, acitretin was proposed as a pharmacological enhancer of the innate cellular defense network that led to virus reactivation and preferential death of infected cells. We evaluated the capacity of acitretin to reactivate and/or to facilitate immune-mediated clearance of HIV-positive cells. Acitretin did not induce HIV reactivation in latently infected cell lines (J-Lat and ACH-2). We could observe only modest induction of HIV reactivation by acitretin in latently green fluorescent protein-HIV-infected Jurkat cells, comparable to suboptimal concentrations of vorinostat, a known latency-reversing agent (LRA). Acitretin induction was insignificant, however, compared to optimal concentrations of LRAs. Acitretin failed to reactivate HIV in a model of latently infected primary CD4 T cells but induced retinoic acid-inducible gene I (RIG-I) and mitochondrial antiviral signaling (MAVS) expression in infected and uninfected cells, confirming the role of acitretin as an innate immune modulator. However, this effect was not associated with selective killing of HIV-positive cells. In conclusion, acitretin-mediated stimulation of the RIG-I pathway for HIV reactivation is modest and thus may not meaningfully affect the HIV reservoir. Stimulation of the RIG-I-dependent interferon (IFN) cascade by acitretin may not significantly affect the selective destruction of latently infected HIV-positive cells.
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http://dx.doi.org/10.1128/AAC.01368-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5655051PMC
November 2017

Chiral Indolylarylsulfone Non-Nucleoside Reverse Transcriptase Inhibitors as New Potent and Broad Spectrum Anti-HIV-1 Agents.

J Med Chem 2017 08 5;60(15):6528-6547. Epub 2017 Jul 5.

Istituto Pasteur Italia-Fondazione Cenci Bolognetti, Dipartimento di Chimica e Tecnologie del Farmaco, Sapienza Università di Roma , Piazzale Aldo Moro 5, I-00185 Roma, Italy.

We designed and synthesized a series of chiral indolyarylsulfones (IASs) as new HIV-1 NNRTIs. The new IASs 8-37 showed potent inhibition of the HIV-1 WT NL4-3 strain and of the mutant K103N, Y181C, Y188L, and K103N-Y181C HIV-1 strains. Six racemic mixtures, 8, 23-25, 31, and 33, were separated at semipreparative level into their pure enantiomers. The (R)-8 enantiomer bearing the chiral (α-methylbenzyl) was superior to the (S)-counterpart. IAS derivatives bearing the (S) alanine unit, (S)-23, (S,R)-25, (S)-31, and (S)-33, were remarkably more potent than the corresponding (R)-enantiomers. Compound 23 protected hippocampal neuronal cells from the excitotoxic insult, while efavirenz (EFV) did not contrast the neurotoxic effect of glutamate. The present results highlight the chiral IASs as new NNRTIs with improved resistance profile against the mutant HIV-1 strains and reduced neurotoxic effects.
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http://dx.doi.org/10.1021/acs.jmedchem.6b01906DOI Listing
August 2017

Antiviral treatment strategies based on gene silencing and genome editing.

Curr Opin Virol 2017 06 21;24:46-54. Epub 2017 Apr 21.

AIDS Research Institute-IrsiCaixa and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain. Electronic address:

The ability of some viruses to establish latently infected chronic reservoirs that escape to immune control becomes a major roadblock that impedes the cure of these infections. Therefore, new alternatives are needed to pursuit the eradication of viral persistent infections. Gene silencing technologies are in constant evolution and provide an outstanding sequence specificity that allows targeting any coding sequence of interest. Here we provide an overview of the development of gene silencing technologies ranging from initially RNA interference to the recently developed CRISPR/Cas9 and their potential as new antiviral strategies focusing on the eradication of HIV.
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http://dx.doi.org/10.1016/j.coviro.2017.04.001DOI Listing
June 2017

SAMHD1 is active in cycling cells permissive to HIV-1 infection.

Antiviral Res 2017 06 28;142:123-135. Epub 2017 Mar 28.

AIDS Research Institute - IrsiCaixa and Health Research, Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain. Electronic address:

SAMHD1 is a triphosphohydrolase that restricts HIV-1 by limiting the intracellular dNTP pool required for reverse transcription. Although SAMHD1 is expressed and active/unphosphorylated in most cell lines, its restriction activity is thought to be relevant only in non-cycling cells. However, an in depth evaluation of SAMHD1 function and relevance in cycling cells is required. Here, we show that SAMHD1-induced degradation by HIV-2 Vpx affects the dNTP pool and HIV-1 replication capacity in the presence of the 3'-azido-3'-deoxythymidine (AZT) in cycling cells. Similarly, in SAMHD1 knockout cells, HIV-1 showed increased replicative capacity in the presence of nucleoside inhibitors, especially AZT, that was reverted by re-expression of wild type SAMHD1. Sensitivity to non-nucleoside inhibitors (nevirapine and efavirenz) or the integrase inhibitor raltegravir was not affected by SAMHD1. Combination of three mutations (S18A, T21A, T25A) significantly prevented SAMHD1 phosphorylation but did not significantly affect HIV-1 replication in the presence of AZT. Our results demonstrate that SAMHD1 is active in HIV-1 permissive cells, does not modify susceptibility to HIV-1 infection but strongly affects sensitivity to nucleoside inhibitors.
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http://dx.doi.org/10.1016/j.antiviral.2017.03.019DOI Listing
June 2017

The G1/S Specific Cyclin D2 Is a Regulator of HIV-1 Restriction in Non-proliferating Cells.

PLoS Pathog 2016 08 19;12(8):e1005829. Epub 2016 Aug 19.

AIDS Research Institute-IrsiCaixa, Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain.

Macrophages are a heterogeneous cell population strongly influenced by differentiation stimuli that become susceptible to HIV-1 infection after inactivation of the restriction factor SAMHD1 by cyclin-dependent kinases (CDK). Here, we have used primary human monocyte-derived macrophages differentiated through different stimuli to evaluate macrophage heterogeneity on cell activation and proliferation and susceptibility to HIV-1 infection. Stimulation of monocytes with GM-CSF induces a non-proliferating macrophage population highly restrictive to HIV-1 infection, characterized by the upregulation of the G1/S-specific cyclin D2, known to control early steps of cell cycle progression. Knockdown of cyclin D2, enhances HIV-1 replication in GM-CSF macrophages through inactivation of SAMHD1 restriction factor by phosphorylation. Co-immunoprecipitation experiments show that cyclin D2 forms a complex with CDK4 and p21, a factor known to restrict HIV-1 replication by affecting the function of the downstream cascade that leads to SAMHD1 deactivation. Thus, we demonstrate that cyclin D2 acts as regulator of cell cycle proteins affecting SAMHD1-mediated HIV-1 restriction in non-proliferating macrophages.
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http://dx.doi.org/10.1371/journal.ppat.1005829DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4991798PMC
August 2016

Long-term HIV-1 infection induces an antiviral state in primary macrophages.

Antiviral Res 2016 09 7;133:145-55. Epub 2016 Aug 7.

AIDS Research Institute - IrsiCaixa and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain.

HIV-1 infection is thought to impair type I interferon (IFN-I) production in macrophages, a cell type that is also relatively resistant to HIV-1 cytotoxic effects. Here, we show that monocyte differentiation into macrophages by M-CSF led to cell proliferation and susceptibility to HIV-1 infection that induced cell cycle arrest and increased cell death. Established HIV-1 infection of monocyte-derived macrophages induced the upregulation of the pattern recognition receptors MDA5 and Rig-I that serve as virus sensors; production of interferon-β, and transcription of interferon-stimulated genes including CXCL10. Infected macrophages showed increased expression of p21 and subsequent inactivation of cyclin-CDK2 activity leading to a hypo-phosphorylated active retinoblastoma protein (pRb) and deactivation of E2F1-dependent transcription and CDK1 downregulation. Additionally, HIV-1 infection limited deoxynucleotide pool by downregulation of the ribonucleotide reductase subunit R2 (RNR2) and reactivation of the HIV-1 restriction factor SAMHD1 together with increased cell death. In conclusion, HIV-1 induced an innate antiviral mechanism associated to IFN-I production, interferon stimulated gene activation, and p21-mediated G2/M arrest leading to elevated levels of cell death in monocyte derived macrophages. Upregulation of MDA5 and Rig-I may serve as targets for the development of antiviral strategies leading to the elimination of HIV-1 infected cells.
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http://dx.doi.org/10.1016/j.antiviral.2016.08.004DOI Listing
September 2016

Inhibition of herpes simplex virus type 1 by the CDK6 inhibitor PD-0332991 (palbociclib) through the control of SAMHD1.

J Antimicrob Chemother 2016 Feb 4;71(2):387-94. Epub 2015 Nov 4.

AIDS Research Institute - IrsiCaixa, and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain

Objectives: Sterile α motif and histidine-aspartate domain-containing protein 1 (SAMHD1) has been shown to restrict retroviruses and DNA viruses by decreasing the pool of intracellular deoxynucleotides. In turn, SAMHD1 is controlled by cyclin-dependent kinases (CDK) that regulate the cell cycle and cell proliferation. Here, we explore the effect of CDK6 inhibitors on the replication of herpes simplex virus type 1 (HSV-1) in primary monocyte-derived macrophages (MDM).

Methods: MDM were treated with palbociclib, a selective CDK4/6 inhibitor, and then infected with a GFP-expressing HSV-1. Intracellular deoxynucleotide triphosphate (dNTP) content was determined using a polymerase-based method.

Results: CDK6 inhibitor palbociclib blocked SAMHD1 phosphorylation, intracellular dNTP levels and HSV-1 replication in MDM at subtoxic concentrations. Treatment of MDM with palbociclib reduced CDK2 activation, measured as the phosphorylation of the T-loop at Thr160. The antiviral activity of palbociclib was lost when SAMHD1 was degraded by viral protein X. Similarly, palbociclib did not block HSV-1 replication in SAMHD1-negative Vero cells at subtoxic concentrations, providing further evidence for a role of SAMHD1 in mediating the antiviral effect.

Conclusions: SAMHD1-mediated HSV-1 restriction is controlled by CDK and points to a preferential role for CDK6 and CDK2 as mediators of SAMHD1 activation. Similarly, the restricting activity of SAMHD1 against DNA viruses suggests that control of dNTP availability is the major determinant of its antiviral activity. This is the first study describing the anti-HSV-1 activity of palbociclib.
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http://dx.doi.org/10.1093/jac/dkv363DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4710216PMC
February 2016

The thioacetate-ω(γ-lactam carboxamide) HDAC inhibitor ST7612AA1 as HIV-1 latency reactivation agent.

Antiviral Res 2015 Nov 5;123:62-9. Epub 2015 Sep 5.

AIDS Research Institute - IrsiCaixa and Health Research Institute Germans Trias i Pujol (IGTP), Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain. Electronic address:

Antiretroviral therapy (ART) is unable to cure HIV infection. The ability of HIV to establish a subset of latent infected CD4(+) T cells, which remain undetectable to the immune system, becomes a major roadblock to achieve viral eradication. Histone deacetylase inhibitors (HDACi) have been shown to potently induce the reactivation of latent HIV. Here, we show that a new thiol-based HDACi, the thioacetate-ω(γ-lactam carboxamide) derivative ST7612AA1, is a potent inducer of HIV reactivation. We evaluated HIV reactivation activity of ST7612AA1 compared to panobinostat (PNB), romidepsin (RMD) and vorinostat (VOR) in cell culture models of HIV-1 latency, in latently infected primary CD4(+) T lymphocytes and in PBMCs from HIV(+) patients. ST7612AA1 potently induced HIV-1 reactivation at submicromolar concentrations with comparable potency to panobinostat or superior to vorinostat. The presence of known antiretrovirals did not affect ST7612AA1-induced reactivation and their activity was not affected by ST7612AA1. Cell proliferation and cell activation were not affected by ST7612AA1, or any other HDACi used. In conclusion, our results indicate that ST7612AA1 is a potent activator of latent HIV and that reactivation activity of ST7612AA1 is exerted without activation or proliferation of CD4(+) T cells. ST7612AA1 is a suitable candidate for further studies of HIV reactivation strategies and potential new therapies to eradicate the viral reservoirs.
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http://dx.doi.org/10.1016/j.antiviral.2015.09.004DOI Listing
November 2015

Cyclin D3-dependent control of the dNTP pool and HIV-1 replication in human macrophages.

Cell Cycle 2015 ;14(11):1657-65

a AIDS Research Institute-IrsiCaixa and AIDS Unit; Hospital Germans Trias i Pujol; Universitat Autonoma de Barcelona ; Badalona , Spain.

Cyclins control the activation of cyclin-dependent kinases (CDK), which in turn, control the cell cycle and cell division. Intracellular availability of deoxynucleotides (dNTP) plays a fundamental role in cell cycle progression. SAM domain and HD domain-containing protein 1 (SAMHD1) degrades nucleotide triphosphates and controls the size of the dNTP pool. SAMHD1 activity appears to be controlled by CDK. Here, we show that knockdown of cyclin D3 a partner of CDK6 and E2 a partner of CDK2 had a major impact in SAMHD1 phosphorylation and inactivation and led to decreased dNTP levels and inhibition of HIV-1 at the reverse transcription step in primary human macrophages. The effect of cyclin D3 RNA interference was lost after degradation of SAMHD1 by HIV-2 Vpx, demonstrating the specificity of the mechanism. Cyclin D3 inhibition correlated with decreased activation of CDK2. Our results confirm the fundamental role of the CDK6-cyclin D3 pair in controlling CDK2-dependent SAMHD1 phosphorylation and dNTP pool in primary macrophages.
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http://dx.doi.org/10.1080/15384101.2015.1030558DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4614030PMC
March 2016

Indolylarylsulfones carrying a heterocyclic tail as very potent and broad spectrum HIV-1 non-nucleoside reverse transcriptase inhibitors.

J Med Chem 2014 Dec 24;57(23):9945-57. Epub 2014 Nov 24.

Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Chimica e Tecnologie del Farmaco, Sapienza Università di Roma , Piazzale Aldo Moro 5, I-00185 Roma, Italy.

We synthesized new indolylarylsulfone (IAS) derivatives carrying a heterocyclic tail at the indole-2-carboxamide nitrogen as potential anti-HIV/AIDS agents. Several new IASs yielded EC50 values <1.0 nM against HIV-1 WT and mutant strains in MT-4 cells. The (R)-11 enantiomer proved to be exceptionally potent against the whole viral panel; in the reverse transcriptase (RT) screening assay, it was remarkably superior to NVP and EFV and comparable to ETV. The binding poses were consistent with the one previously described for the IAS non-nucleoside reverse transcriptase inhibitors. Docking studies showed that the methyl group of (R)-11 points toward the cleft created by the K103N mutation, different from the corresponding group of (S)-11. By calculating the solvent-accessible surface, we observed that the exposed area of RT in complex with (S)-11 was larger than the area of the (R)-11 complex. Compounds 6 and 16 and enantiomer (R)-11 represent novel robust lead compounds of the IAS class.
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http://dx.doi.org/10.1021/jm5011622DOI Listing
December 2014

Characterization of the influence of mediator complex in HIV-1 transcription.

J Biol Chem 2014 Oct 6;289(40):27665-76. Epub 2014 Aug 6.

From the AIDS Research Institute-IrsiCaixa and AIDS Unit, Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916 Badalona, Spain

HIV-1 exploits multiple host proteins during infection. siRNA-based screenings have identified new proteins implicated in different pathways of the viral cycle that participate in a broad range of cellular functions. The human Mediator complex (MED) is composed of 28 elements and represents a fundamental component of the transcription machinery, interacting with the RNA polymerase II enzyme and regulating its ability to express genes. Here, we provide an evaluation of the MED activity on HIV replication. Knockdown of 9 out of 28 human MED proteins significantly impaired viral replication without affecting cell viability, including MED6, MED7, MED11, MED14, MED21, MED26, MED27, MED28, and MED30. Impairment of viral replication by MED subunits was at a post-integration step. Inhibition of early HIV transcripts was observed by siRNA-mediated knockdown of MED6, MED7, MED11, MED14, and MED28, specifically affecting the transcription of the nascent viral mRNA transactivation-responsive element. In addition, MED14 and MED30 were shown to have special relevance during the formation of unspliced viral transcripts (p < 0.0005). Knockdown of the selected MED factors compromised HIV transcription induced by Tat, with the strongest inhibitory effect shown by siMED6 and siMED14 cells. Co-immunoprecipitation experiments suggested physical interaction between MED14 and HIV-1 Tat protein. A better understanding of the mechanisms and factors controlling HIV-1 transcription is key to addressing the development of new strategies required to inhibit HIV replication or reactivate HIV-1 from the latent reservoirs.
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http://dx.doi.org/10.1074/jbc.M114.570341DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4183804PMC
October 2014

Increased expression of SAMHD1 in a subset of HIV-1 elite controllers.

J Antimicrob Chemother 2014 Nov 25;69(11):3057-60. Epub 2014 Jul 25.

AIDS Research Institute-IrsiCaixa and AIDS Unit, Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain

Objectives: SAMHD1 and the CDKN1A (p21) cyclin-dependent kinase inhibitor have been postulated to mediate HIV-1 restriction in CD4+ cells. We have shown that p21 affects HIV replication through its effect on SAMHD1. Thus, we aimed at evaluating the expression of SAMHD1 and p21 in different HIV+ phenotypic groups.

Patients And Methods: We evaluated SAMHD1 and CDKN1A mRNA expression in CD4+ T cells from HIV+ individuals including elite controllers (n = 12), individuals who control HIV without the need for antiretroviral treatment, viraemic progressors (n = 10) and HIV-1 seronegative healthy donors (n = 14). Immunological variables were measured by flow cytometry.

Results: We show that a subset of HIV+ elite controllers with lower T cell proliferation levels (Ki67+ cells) expressed higher SAMHD1 compared with healthy donors or viraemic progressors. Conversely, there was no difference in p21 expression before or after T cell activation with a bispecific CD3/CD8 antibody.

Conclusions: Our results suggest that SAMHD1 may play a role in controlling virus replication in HIV+ individuals and slow the rate of disease progression.
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http://dx.doi.org/10.1093/jac/dku276DOI Listing
November 2014

Palbociclib, a selective inhibitor of cyclin-dependent kinase4/6, blocks HIV-1 reverse transcription through the control of sterile α motif and HD domain-containing protein-1 (SAMHD1) activity.

AIDS 2014 Sep;28(15):2213-22

aAIDS Research Institute-IrsiCaixa and AIDS Unit, Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain bResearch Group on Neuromuscular and Mitochondrial Disorders, Vall d'Hebron Institut de Recerca, Universitat Autònoma de Barcelona and Biomedical Network Research Centre on Rare Diseases (CIBERER), Instituto de Salud Carlos III, Valencia, Spain. *Eduardo Pauls and Roger Badia contributed equally to this work.

Background: Sterile α motif and HD domain-containing protein-1 (SAMHD1) inhibits HIV-1 reverse transcription by decreasing the pool of intracellular deoxynucleotides. SAMHD1 is controlled by cyclin-dependent kinase (CDK)-mediated phosphorylation. However, the exact mechanism of SAMHD1 regulation in primary cells is unclear. We explore the effect of palbociclib, a CDK6 inhibitor, in HIV-1 replication.

Methods: Human primary monocytes were differentiated into macrophages with monocyte-colony stimulating factor and CD4 T lymphocytes stimulated with phytohaemagglutinin (PHA)/interleukin-2. Cells were treated with palbociclib and then infected with a Green fluorescent protein-expressing HIV-1 or R5 HIV-1 BaL. Viral DNA was measured by quantitative PCR and infection assessed by flow cytometry. Deoxynucleotide triphosphate (dNTP) content was determined using a polymerase-based method.

Results: Pan-CDK inhibitors AT7519, roscovitine and purvalanol A reduced SAMHD1 phosphorylation. HIV-1 replication was blocked by AT7519 (66.4 ± 3.8%; n = 4), roscovitine (47.3 ± 3.9%; n = 4) and purvalanol A (55.7 ± 15.7%; n = 4) at subtoxic concentrations. Palbociclib, a potent and selective CDK6 inhibitor, blocked SAMHD1 phosphorylation, intracellular dNTP levels, HIV-1 reverse transcription and HIV-1 replication in primary macrophages and CD4 T lymphocytes. Notably, treatment of macrophages with palbociclib led to reduced CDK2 activation, measured as the phosphorylation of the T-loop at the Thr160. The antiviral effect was lost when SAMHD1 was degraded by Vpx, providing further evidence for a role of SAMHD1 in mediating the antiretroviral effect.

Conclusions: Our results indicate that SAMHD1-mediated HIV-1 restriction is controlled by CDK as previously suggested but point to a preferential role for CDK2 and CDK6 as mediators of SAMHD1 activation. Our study provides a new signaling pathway susceptible for the development of new therapeutic approaches against HIV-1 infection.
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http://dx.doi.org/10.1097/QAD.0000000000000399DOI Listing
September 2014

Cell cycle control and HIV-1 susceptibility are linked by CDK6-dependent CDK2 phosphorylation of SAMHD1 in myeloid and lymphoid cells.

J Immunol 2014 Aug 11;193(4):1988-97. Epub 2014 Jul 11.

AIDS Research Institute-IrsiCaixa and AIDS Unit, Hospital Germans Trias i Pujol, Autonomous University of Barcelona, 08916 Badalona, Spain;

Proliferating cells are preferentially susceptible to infection by retroviruses. Sterile α motif and HD domain-containing protein-1 (SAMHD1) is a recently described deoxynucleotide phosphohydrolase controlling the size of the intracellular deoxynucleotide triphosphate (dNTP) pool, a limiting factor for retroviral reverse transcription in noncycling cells. Proliferating (Ki67(+)) primary CD4(+) T cells or macrophages express a phosphorylated form of SAMHD1 that corresponds with susceptibility to infection in cell culture. We identified cyclin-dependent kinase (CDK) 6 as an upstream regulator of CDK2 controlling SAMHD1 phosphorylation in primary T cells and macrophages susceptible to infection by HIV-1. In turn, CDK2 was strongly linked to cell cycle progression and coordinated SAMHD1 phosphorylation and inactivation. CDK inhibitors specifically blocked HIV-1 infection at the reverse transcription step in a SAMHD1-dependent manner, reducing the intracellular dNTP pool. Our findings identify a direct relationship between control of the cell cycle by CDK6 and SAMHD1 activity, which is important for replication of lentiviruses, as well as other viruses whose replication may be regulated by intracellular dNTP availability.
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http://dx.doi.org/10.4049/jimmunol.1400873DOI Listing
August 2014

SAMHD1 specifically affects the antiviral potency of thymidine analog HIV reverse transcriptase inhibitors.

Antimicrob Agents Chemother 2014 Aug 9;58(8):4804-13. Epub 2014 Jun 9.

AIDS Research Institute-IrsiCaixa, Hospital Germans Trias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain

Sterile alpha motif and histidine-aspartic domain-containing protein 1 (SAMHD1) is a deoxynucleoside triphosphate (dNTP) triphosphohydrolase recently recognized as an antiviral factor that acts by depleting dNTP availability for viral reverse transcriptase (RT). SAMHD1 restriction is counteracted by the human immunodeficiency virus type 2 (HIV-2) accessory protein Vpx, which targets SAMHD1 for proteosomal degradation, resulting in an increased availability of dNTPs and consequently enhanced viral replication. Nucleoside reverse transcriptase inhibitors (NRTI), one of the most common agents used in antiretroviral therapy, compete with intracellular dNTPs as the substrate for viral RT. Consequently, SAMHD1 activity may be influencing NRTI efficacy in inhibiting viral replication. Here, a panel of different RT inhibitors was analyzed for their different antiviral efficacy depending on SAMHD1. Antiviral potency was measured for all the inhibitors in transformed cell lines and primary monocyte-derived macrophages and CD4(+) T cells infected with HIV-1 with or without Vpx. No changes in sensitivity to non-NRTI or the integrase inhibitor raltegravir were observed, but for NRTI, sensitivity significantly changed only in the case of the thymidine analogs (AZT and d4T). The addition of exogenous thymidine mimicked the change in viral sensitivity observed after Vpx-mediated SAMHD1 degradation, pointing toward a differential effect of SAMHD1 activity on thymidine. Accordingly, sensitivity to AZT was also reduced in CD4(+) T cells infected with HIV-2 compared to infection with the HIV-2ΔVpx strain. In conclusion, reduction of SAMHD1 levels significantly decreases HIV sensitivity to thymidine but not other nucleotide RT analog inhibitors in both macrophages and lymphocytes.
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http://dx.doi.org/10.1128/AAC.03145-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136047PMC
August 2014
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