Percorrer por autor "Borrego, Pedro"
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- An ancestral HIV-2/simian immunodeficiency virus peptide with potent HIV-1 and HIV-2 fusion inhibitor activityPublication . Borrego, Pedro; Calado, Rita; Marcelino, José M.; Pereira, Patrícia; Quintas, Alexandre; Barroso, Helena; Taveira, Nuno"Objectives: To produce new fusion inhibitor peptides for HIV-1 and HIV-2 based on ancestral envelope sequences. Methods: HIV-2/simian immunodeficiency virus (SIV) ancestral transmembrane protein sequences were reconstructed and ancestral peptides were derived from the helical region 2 (HR2). The activity of one ancestral peptide (named P3) was examined against a panel of HIV-1 and HIV-2 primary isolates in TZM-bl cells and peripheral blood mononuclear cells and compared to T-20. Peptide secondary structure was analyzed by circular dichroism. Resistant viruses were selected and resistance mutations were identified by sequencing the env gene. Results: P3 has 34 residues and overlaps the N-terminal pocket-binding region and heptad repeat core of HR2. In contrast to T-20, P3 forms a typical a-helical structure in solution, binds strongly to the transmembrane protein, and potently inhibits both HIV-2 (mean IC50, 63.8 nmol/l) and HIV-1 (11 nmol/l) infection, including T-20-resistant isolates. The N43K mutation in the HR1 region of HIV-1 leads to 120-fold resistance to P3 indicating that the HR1 region in transmembrane glycoprotein is the target of P3. No HIV-2-resistant mutations could be selected by P3 suggesting that the genetic barrier to resistance is higher in HIV-2 than in HIV-1. HIV-1-infected patients presented significantly lower P3-specific antibody reactivity compared to T-20. Conclusion: P3 is an HIV-2/SIV ancestral peptide with low antigenicity, high stability, and potent activity against both HIV-1, including variants resistant to T-20, and HIV-2. Similar evolutionary biology strategies should be explored to enhance the production of antiviral peptide drugs, microbicides, and vaccines."
- Antagonism of BST-2/Tetherin is a conserved function of the Env glycoprotein of primary HIV-2 isolatesPublication . Chen, Chia-Yen; Shingai, Masashi; Welbourn, Sarah; Martin, Malcolm A.; Borrego, Pedro; Taveira, Nuno; Strebel, KlausAlthough HIV-2 does not encode a vpu gene, the ability to antagonize bone marrow stromal antigen 2 (BST-2) is conserved in some HIV-2 isolates, where it is controlled by the Env glycoprotein. We previously reported that a single-amino-acid difference between the laboratory-adapted ROD10 and ROD14 Envs controlled the enhancement of virus release (referred to here as Vpu-like) activity. Here, we investigated how conserved the Vpu-like activity is in primary HIV-2 isolates. We found that half of the 34 tested primary HIV-2 Env isolates obtained from 7 different patients enhanced virus release. Interestingly, most HIV-2 patients harbored a mixed population of viruses containing or lacking Vpu-like activity. Vpu-like activity and Envelope functionality varied significantly among Env isolates; however, there was no direct correlation between these two functions, suggesting they evolved independently. In comparing the Env sequences from one HIV-2 patient, we found that similar to the ROD10/ROD14 Envs, a single-amino-acid change (T568I) in the ectodomain of the TM subunit was sufficient to confer Vpu-like activity to an inactive Env variant. Surprisingly, however, absence of Vpu-like activity was not correlated with absence of BST-2 interaction. Taken together, our data suggest that maintaining the ability to antagonize BST-2 is of functional relevance not only to HIV-1 but also to HIV-2 as well. Our data show that as with Vpu, binding of HIV-2 Env to BST-2 is important but not sufficient for antagonism. Finally, as observed previously, the Vpu-like activity in HIV-2 Env can be controlled by single-residue changes in the TM subunit.
- Assessment of the Cavidi ExaVir load assay for monitoring plasma viral load in HIV-2-infected patientsPublication . Borrego, Pedro; Gonçalves, Maria Fátima; Gomes, Perpétua; Araújo, Lavínia; Moranguinho, Inês; Figueiredo, Inês Brito; Barahona, Isabel; Rocha, José; Mendonça, Claudino; Cruz, Maria Cesarina; Barreto, Jorge; Taveira, NunoHIV plasma viral load is an established marker of disease progression and of response to antiretroviral therapy, but currently there is no commercial assay validated for the quantification of viral load in HIV-2-infected individuals. We sought to make the first clinical evaluation of Cavidi ExaVir Load (version 3) in HIV-2- infected patients. Samples were collected from a total of 102 individuals living in Cape Verde, and the HIV-2 viral load was quantified by both ExaVir Load and a reference in-house real-time quantitative PCR (qPCR) used in Portugal in 91 samples. The associations between viral load and clinical prognostic variables (CD4 T cell counts and antiretroviral therapy status) were similar for measurements obtained using ExaVir Load and qPCR. There was no difference between the two methods in the capacity to discriminate between nonquantifiable and quantifiable HIV-2 in the plasma. In samples with an HIV-2 viral load quantifiable by both methods (n 27), the measurements were highly correlated (Pearson r 0.908), but the ExaVir Load values were systematically higher relative to those determined by qPCR (median difference, 0.942 log10 copies/ml). A regression model was derived that enables the conversion of ExaVir Load results to those that would have been obtained by the reference qPCR. In conclusion, ExaVir Load version 3 is a reliable commercial assay to measure viral load in HIV-2-infected patients and therefore a valuable alternative to the inhouse assays in current use.
- Baseline susceptibility of primary HIV-2 to entry inhibitorsPublication . Borrego, Pedro; Calado, Rita; Marcelino, José M; Bártolo, Inês; Rocha, Cheila; Cavaco-Silva, Patrícia; Doroana, Manuela; Antunes, Francisco; Maltez, Fernando; Caixas, Umbelina; Barroso, Helena; Taveira, NunoBackground: The baseline susceptibility of primary HIV-2 to maraviroc (MVC) and other entry inhibitors is currently unknown.
- Development of synthetic light-chain antibodies as novel and potent HIV fusion inhibitorsPublication . Cunha-Santos, Catarina; Figueira, Tiago N.; Borrego, Pedro; Oliveira, Soraia S.; Rocha, Cheila; Couto, Andreia; Cantante, Cátia; Santos- Costa, Quirina; Azevedo-Pereira, José M.; Fontes, Carlos M. G. A.; Taveira, Nuno; Aires-Da-Silva, Frederico; Castanho, Miguel A. R. B.; Veiga, Ana Salomé; Gonçalves, João"Objective: To develop a novel and potent fusion inhibitor of HIV infection based on a rational strategy for synthetic antibody library construction. Design: The reduced molecular weight of single-domain antibodies (sdAbs) allows targeting of cryptic epitopes, the most conserved and critical ones in the context of HIV entry. Heavy-chain sdAbs from camelids are particularly suited for this type of epitope recognition because of the presence of long and flexible antigen-binding regions [complementary-determining regions (CDRs)]."
- Development of water-soluble polyanionic carbosilane dendrimers as novel and highly potent topical anti-HIV-2 microbicidesPublication . Briz, Verónica; Sepúlveda-Crespo, Daniel; Diniz, Ana Rita; Borrego, Pedro; Rodes, Berta; Javier de la Mata, Francisco; Gómez, Rafael; Taveira, Nuno; Muñoz-Fernández, Mª Ángeles"The development of topical microbicide formulations for vaginal delivery to prevent HIV-2 sexual transmission is urgently needed. Second- and third-generation polyanionic carbosilane dendrimers with a silicon atom core and 16 sulfonate (G2-S16), napthylsulfonate (G2-NS16) and sulphate (G3-Sh16) end-groups have shown potent and broad-spectrum anti-HIV-1 activity. However, their antiviral activity against HIV-2 and mode of action have not been probed. Cytotoxicity, anti-HIV-2, anti-sperm and antimicrobial activities of dendrimers were determined. Analysis of combined effects of triple combinations with tenofovir and raltegravir was performed by using CalcuSyn software. We also assessed the mode of antiviral action on the inhibition of HIV-2 infection through a panel of different in vitro antiviral assays: attachment, internalization in PBMCs, inactivation and cell-based fusion. Vaginal irritation and histological analysis in female BALB/c mice were evaluated. Our results suggest that G2-S16, G2-NS16 and G3-Sh16 exert anti-HIV-2 activity at an early stage of viral replication inactivating the virus, inhibiting cell-to-cell HIV-2 transmission, and blocking the binding of gp120 to CD4, and the HIV-2 entry. Triple combinations with tenofovir and raltegravir increased the anti-HIV-2 activity, consistent with synergistic interactions (CIwt: 0.33–0.66). No vaginal irritation was detected in BALB/c mice after two consecutive applications for 2 days with 3% G2-S16. Our results have clearly shown that G2-S16, G2-NS16 and G3-Sh16 have high potency against HIV-2 infection. The modes of action confirm their multifactorial and non-specific ability, suggesting that these dendrimers deserve further studies as potential candidate microbicides to prevent vaginal/rectal HIV-1/HIV-2 transmission in humans."
- Epidemic history and baseline resistance to NS5A-specific direct acting drugs of hepatitis C virus in SpainPublication . Palladino, Claudia; Ezeonwumelu, Ifeanyi Jude; Mate-Cano, Irene; Borrego, Pedro; Martínez-Román, Paula; Arca-Lafuente, Sonia; Resino, Salvador; Taveira, Nuno; Briz, VerónicaHepatitis C virus (HCV) infection remains a global health problem. Previously, the prevalence of NS5A resistance-associated substitutions (RASs) to elbasvir, a new direct-acting antiviral (DAA) against the NS5A viral protein was assessed by our group before its introduction into clinical use in Spain. However, the origin, epidemic history, transmission dynamics, diversity and baseline RASs to NS5A direct-acting agents of HCV-GT1a in Spain remain unknown. A nationwide cross-sectional survey of individuals chronically-infected with HCV-G1a and DAAs-naïve was performed. HCV population sequencing, phylogenetic analysis and Bayesian methods were used. GT1a clade II was more prevalent than clade I (82.3% vs. 17.7%; P < 0.001) and older (estimated origin in 1912 vs. 1952). Clade II epidemic is currently declining whereas clade I epidemic has reached equilibrium. A total of 58 single RASs were identified, which account for the moderate level (10%) of baseline resistance observed. When considering the regional data, marked differences were observed, with thirteen regions showing an intermediate level (5–15%) and one a high level (20%) of resistance. Current HCV-GT1a epidemic in Spain is driven by clade I which seem to have different dissemination routes relative to clade II. A moderate level of baseline RASs to NS5A-DAAs with marked differences among regions was observed. Close surveillance of response to treatment with DAAs will be important.
- Evaluation of the fusion inhibitor P3 peptide as a potential microbicide to prevent HIV transmission in womenPublication . Bártolo, Inês; Diniz, Ana Rita; Borrego, Pedro; Ferreira, João Pedro; Bronze, Maria Rosário; Barroso, Helena; Pinto, Rui; Cardoso, Carlos; Pinto, João F.; Diaz, Rafael Ceña; Broncano, Pilar Garcia; Muñoz-Fernández, Maria Angel; Taveira, NunoMicrobicides are an important strategy for preventing the sexual transmission of HIV but, so far, the most advanced tenofovir-based microbicides have had modest efficacy. This has been related to adherence problems and high prevalence of tenofovir-resistant HIV-1 strains. P3 is a new peptide with potent activity against HIV that may be a good microbicide candidate. In this work P3 was formulated in a gel of hydroxyethyl cellulose and its activity, stability and safety profile in Balb/c mice were evaluated. HIV infection was fully blocked by a 1.5% gel containing P3 at the IC90 (366.4 nM) concentration. The antiviral activity did not change at 4°C during 4 months and at 25, 37 and 65°C for 1 week. P3 was stable and fully functional at acidic pH up to 24h, under different concentrations of hydrogen peroxide and in the presence of genital fluids up to 48h. P3 had no antibacterial activity and did not affect sperm motility and vitality. Finally, P3 didn't cause significant alterations in the vaginal epithelium of Balb/c mice at 0.06 (456.8 μM) and 0.2 mg/day (1522.7 μM) doses. These findings indicate that P3 is an excellent candidate for further development as a microbicide gel for the prevention of HIV transmission in women.
- Evolution of the human immunodeficiency virus type 2 envelope in the first years of infection is associated with the dynamics of the neutralizing antibody responsePublication . Rocha, Cheila; Calado, Rita; Borrego, Pedro; Marcelino, José Maria; Bártolo, Inês; Rosado, Lino; Cavaco-Silva, Patrícia; Perpétua, Gomes; Família, Carlos; Quintas, Alexandre; Skar, Helena; Leitner, Thomas; Barroso, Helena; Taveira, Nuno"Background: Differently from HIV-1, HIV-2 disease progression usually takes decades without antiretroviral therapy and the majority of HIV-2 infected individuals survive as elite controllers with normal CD4+ T cell counts and low or undetectable plasma viral load. Neutralizing antibodies (Nabs) are thought to play a central role in HIV-2 evolution and pathogenesis. However, the dynamic of the Nab response and resulting HIV-2 escape during acute infection and their impact in HIV-2 evolution and disease progression remain largely unknown. Our objective was to characterize the Nab response and the molecular and phenotypic evolution of HIV-2 in association with Nab escape in the first years of infection in two children infected at birth. Results: CD4+ T cells decreased from about 50% to below 30% in both children in the first five years of infection and the infecting R5 viruses were replaced by X4 viruses within the same period. With antiretroviral therapy, viral load in child 1 decreased to undetectable levels and CD4+ T cells recovered to normal levels, which have been sustained at least until the age of 12. In contrast, viral load increased in child 2 and she progressed to AIDS and death at age 9. Beginning in the first year of life, child 1 raised high titers of antibodies that neutralized primary R5 isolates more effectively than X4 isolates, both autologous and heterologous. Child 2 raised a weak X4-specific Nab response that decreased sharply as disease progressed. Rate of evolution, nucleotide and amino acid diversity, and positive selection, were significantly higher in the envelope of child 1 compared to child 2. Rates of R5-to-X4 tropism switch, of V1 and V3 sequence diversification, and of convergence of V3 to a β-hairpin structure were related with rate of escape from the neutralizing antibodies. Conclusion: Our data suggests that the molecular and phenotypic evolution of the human immunodeficiency virus type 2 envelope are related with the dynamics of the neutralizing antibody response providing further support for a model in which Nabs play an important role in HIV-2 pathogenesis."
- Evolutionary and Structural Features of the C2, V3 and C3 Envelope Regions Underlying the Differences in HIV-1 and HIV-2 Biology and InfectionPublication . Barroso, Helena; Borrego, Pedro; Bártolo, Inês; Marcelino, José Maria; Família, Carlos; Quintas, Alexandre; Taveira, NunoBackground: Unlike in HIV-1 infection, the majority of HIV-2 patients produce broadly reactive neutralizing antibodies, control viral replication and survive as elite controllers. The identification of the molecular, structural and evolutionary footprints underlying these very distinct immunological and clinical outcomes may lead to the development of new strategies for the prevention and treatment of HIV infection.
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