Extracellular vesicle isolation methods identify distinct HIV-1 particles released from chronically infected T-cells.

The current study analyzed the intersecting biophysical, biochemical, and functional properties of extracellular particles (EPs) with the human immunodeficiency virus type-1 (HIV-1) beyond the currently accepted size range for HIV-1. We isolated five fractions (Frac-A through Frac-E) from HIV-infected cells by sequential differential ultracentrifugation (DUC). All fractions showed a heterogeneous size distribution with median particle sizes greater than 100 nm for Frac-A through Frac-D but not for Frac-E, which contained small EPs with an average size well below 50 nm. Synchronized and released cultures contained large infectious EPs in Frac-A, with markers of amphisomes and viral components. Additionally, Frac-E uniquely contained EPs positive for CD63, HSP70, and HIV-1 proteins. Despite its small average size, Frac-E contained membrane-protected viral integrase, detectable only after SDS treatment, indicating that it is enclosed in vesicles. Single particle analysis with dSTORM further supported these findings as CD63, HIV-1 integrase, and the viral surface envelope (Env) glycoprotein (gp) colocalized on the same Frac-E particles. Surprisingly, Frac-E EPs were infectious, and infectivity was significantly reduced by immunodepleting Frac-E with anti-CD63, indicating the presence of this protein on the surface of infectious small EPs in Frac-E. To our knowledge, this is the first time that extracellular vesicle (EV) isolation methods have identified infectious small HIV-1 particles (smHIV-1) that are under 50 nm. Collectively, our data indicate that the crossroads between EPs and HIV-1 potentially extend beyond the currently accepted biophysical properties of HIV-1, which may have further implications for viral pathogenesis.

A scientific career from the early 1960s till 2023: A tale of the various protagonists.

In this era spanning more than 60 years (from the early 1960s till today (2023), a broad variety of actors played a decisive role: Piet De Somer, Tom C. Merigan, Paul A. Janssen, Maurice Hilleman, and Georges Smets. Two protagonists (Antonín Holý and John C. Martin) formed with me a unique triangle (the Holý Trinity). Walter Fiers' group (with the help of Jean Content) contributed to the cloning of human ß-interferon, and Piet Herdewijn accomplished the chemical synthesis of an array of anti-HIV 2',3'-dideoxynucleoside analogues. Rudi Pauwels, Masanori Baba, Dominique Schols, Johan Neyts, Lieve Naesens, Anita Van Lierde, Graciela Andrei, Robert Snoeck and Dirk Daelemans, as members of my team, helped me in achieving the intended goal, the development of a selective therapy for virus infections. The collaboration with "Lowie" (Guangdi Li) generated a new dimension for the future.

HIV Infection, Antiretroviral Drugs, and the Vascular Endothelium.

Endothelial cell activation, injury, and dysfunction underlies the pathophysiology of vascular diseases and infections associated with vascular dysfunction, including human immunodeficiency virus (HIV) and acquired immunodeficiency syndrome. Despite viral suppression with combination antiretroviral therapy (ART), people living with HIV (PLWH) are prone to many comorbidities, including neurological and neuropsychiatric complications, cardiovascular and metabolic diseases, premature aging, and malignancies. HIV and viral proteins can directly contribute to the development of these comorbidities. However, with the continued high prevalence of these comorbidities despite viral suppression, it is likely that ART or some antiretroviral (ARVs) drugs contribute to the development and persistence of comorbid diseases in PLWH. These comorbid diseases often involve vascular activation, injury, and dysfunction. The purpose of this manuscript is to review the current literature on ARVs and the vascular endothelium in PLWH, animal models, and in vitro studies. I also summarize evidence of an association or lack thereof between ARV drugs or drug classes and the protection or injury/dysfunction of the vascular endothelium and vascular diseases.

Selected Milestones in Antiviral Drug Development.

This review article will describe the (wide) variety of approaches that I envisaged to develop a specific therapy for viral infections: (i) interferon and its inducers, (ii) HSV, VZV and CMV inhibitors, (iii) NRTIs (nucleoside reverse transcriptase inhibitors), NtRTIs (nucleotide reverse transcriptase inhibitors) and NNRTIs (non-nucleoside reverse transcriptase inhibitors) as HIV inhibitors, (iv) NtRTIs as HBV inhibitors, and finally, (v) the transition of an HIV inhibitor to a stem cell mobilizer, as exemplified by AMD-3100 (Mozobil®).

The Influence of Nucleoside Reverse Transcriptase Inhibitors on Mitochondrial Activity, Lipid Content, and Fatty-Acid-Binding Protein Levels in Microglial HMC3 Cells.

Despite the availability of a wide range of preventive measures and comprehensive treatment options following infection, the development of acquired immunodeficiency syndrome (AIDS) remains a persistent challenge. Nucleoside reverse transcriptase inhibitors (NRTIs) represent the most commonly utilized therapeutic approach, despite being on the pharmaceutical market for nearly four decades. During this time, a spectrum of side effects ranging from mild discomfort and hypersensitivity reactions to the more prevalent nephrotoxicity and hepatotoxicity has been documented. In light of these considerations, our study aimed to investigate the impacts of two NRTIs, lamivudine and zidovudine, on lipid metabolism in HMC3 microglial cells. Our findings revealed statistically significant reductions in the ATP levels (nearly 8%) and increased mitochondrial superoxide levels (around 10%) after 24 h of treatment with the maximum therapeutic concentration of zidovudine compared to the untreated microglial cells. Furthermore, the concentrations of fatty-acid-binding proteins 4 and 5 were significantly lower (approximately 40%) in the microglial cells that were exposed to NRTIs than in the untreated cells. Notably, the total lipid concentration within the microglial cells markedly increased following NRTI administration with a 13% rise after treatment with 10 µM lamivudine and a remarkable 70% surge following the administration of 6 µM zidovudine. These results suggest that the prolonged administration of NRTIs may potentially lead to lipid accumulation, posing a significant risk to the delicate homeostasis of the neuronal system and potentially triggering a pro-inflammatory response in microglial cells.

The Application of Prodrugs as a Tool to Enhance the Properties of Nucleoside Reverse Transcriptase Inhibitors.

Antiretroviral Therapy (ART) is an effective treatment for human immunodeficiency virus (HIV) which has transformed the highly lethal disease, acquired immunodeficiency syndrome (AIDS), into a chronic and manageable condition. However, better methods need to be developed for enhancing patient access and adherence to therapy and for improving treatment in the long term to reduce adverse effects. From the perspective of drug discovery, one promising strategy is the development of anti-HIV prodrugs. This approach aims to enhance the efficacy and safety of treatment, promoting the development of more appropriate and convenient systems for patients. In this review, we discussed the use of the prodrug approach for HIV antiviral agents and emphasized nucleoside reverse transcriptase inhibitors. We comprehensively described various strategies that are used to enhance factors such as water solubility, bioavailability, pharmacokinetic parameters, permeability across biological membranes, chemical stability, drug delivery to specific sites/organs, and tolerability. These strategies might help researchers conduct better studies in this field. We also reported successful examples from the primary therapeutic classes while discussing the advantages and limitations. In this review, we highlighted the key trends in the application of the prodrug approach for treating HIV/AIDS.

Repurposing nucleoside reverse transcriptase inhibitors (NRTIs) to slow aging.

Repurposing drugs already approved in the clinic to be used off-label as geroprotectors, compounds that combat mechanisms of aging, are a promising way to rapidly reduce age-related disease incidence in society. Several recent studies have found that a class of drugs-nucleoside reverse transcriptase inhibitors (NRTIs)-originally developed as treatments for cancers and human immunodeficiency virus (HIV) infection, could be repurposed to slow the aging process. Interestingly, these studies propose complementary mechanisms that target multiple hallmarks of aging. At the molecular level, NRTIs repress LINE-1 elements, reducing DNA damage, benefiting the hallmark of aging of 'Genomic Instability'. At the organellar level, NRTIs inhibit mitochondrial translation, activate ATF-4, suppress cytosolic translation, and extend lifespan in worms in a manner related to the 'Loss of Proteostasis' hallmark of aging. Meanwhile, at the cellular level, NRTIs inhibit the P2X7-mediated activation of the inflammasome, reducing inflammation and improving the hallmark of aging of 'Altered Intercellular Communication'. Future development of NRTIs for human aging health will need to balance out toxic side effects with the beneficial effects, which may occur in part through hormesis.

HIV and Drug-Resistant Subtypes.

Acquired Immunodeficiency Syndrome (AIDS) is a human viral infectious disease caused by the positive-sense single-stranded (ss) RNA Human Immunodeficiency Virus (HIV) (Retroviridae family, Ortervirales order). HIV-1 can be distinguished into various worldwide spread groups and subtypes. HIV-2 also causes human immunodeficiency, which develops slowly and tends to be less aggressive. HIV-2 only partially homologates to HIV-1 despite the similar derivation. Antiretroviral therapy (ART) is the treatment approved to control HIV infection, based on multiple antiretroviral drugs that belong to different classes: (i) NNRTIs, (ii) NRTIs, (iii) PIs, (iv) INSTIs, and (v) entry inhibitors. These drugs, acting on different stages of the HIV life cycle, decrease the patient's total burden of HIV, maintain the function of the immune system, and prevent opportunistic infections. The appearance of several strains resistant to these drugs, however, represents a problem today that needs to be addressed as best as we can. New outbreaks of strains show a widespread geographic distribution and a highly variable mortality rate, even affecting treated patients significantly. Therefore, novel treatment approaches should be explored. The present review discusses updated information on HIV-1- and HIV-2-resistant strains, including details on different mutations responsible for drug resistance.

Impact of age and gender disparity on CD4+ cell counts to control disease progression using specific HAART in HIV-1 positive patients: A case-control study.

BACKGROUND: Highly Active Antiretroviral Therapy (HAART) is composed of several drugs in the antiretroviral class to better treat human immunodeficiency virus type 1 (HIV-1) patients. The estimation of CD4+ T cell counts and HIV-1 viral load in plasma is required to evaluate the treatment success of a specific HAART. METHODOLOGY: The study included the effects of NRTIs (nucleoside reverse transcriptase inhibitors) and novel protease inhibitors (HAART) on normal control subjects and HIV-1 positive subjects from SGPGIMS, Lucknow, with different age groups and genders. Furthermore, the study was conducted by the estimation of HIV through ELISA, measurement of absolute CD4+ cell count, and the measurement of viral load through qRT-PCR. Furthermore, NRTIs (Retrovir and Epivir) were administered orally one tablet daily in the morning followed by newly FDA-approved protease inhibitors (fosamprenavir and darunavir) orally in the evening at the same dose. Furthermore, CD4+T cell counts and HIV-1 viral load were investigated and correlated in patients with different genders and age groups. RESULT: Administration of NRTIs and novel protease inhibitors (HAART) in HIV patients had a significant effect on the CD4+ cell count in various age intervals among males and females. The mean comparison of viral load distribution based on gender in CD4 +ve patients in the case group exhibited a viral load higher in females compared to males, indicating a statistically significant difference between males and females (p<0.05). A notable association between virological and immunological parameters was observed with a reciprocal relationship between viral load and CD4 cell count in CD4 +ve patients, demonstrating multiple correlation coefficients with an R-value of 0.853. CONCLUSION: The administration of specific HAART (NRTIs and novel protease inhibitors) in HIV patients had a notable improvement in the CD4+ cell count and viral load with significant age and gender disparity.

Can plant-derived anti-HIV compounds be used in COVID-19 cases?

People living with HIV are more exposed to the adverse health effects of the worldwide COVID-19 pandemic. The pandemic's health and social repercussions may promote drug abuse and inadequate HIV management among this demographic. The coronavirus pandemic of 2019 (COVID-19) has caused unprecedented disruption worldwide in people's lives and health care. When the COVID-19 epidemic was identified, people with HIV faced significant obstacles and hurdles to achieving optimal care results. The viral spike protein (S-Protein) and the cognate host cell receptor angiotensin-converting enzyme 2 (ACE2) are both realistic and appropriate intervention targets. Calanolides A, Holy Basil, Kuwanon-L, and Patentiflorin have anti-HIV effects. Our computational biology study investigated that these compounds all had interaction binding scores related to S protein of coronavirus of -9.0 kcal /mol, -7.1 kcal /mol, -9.1 kcal /mol, and -10.3 kcal/mol/mol, respectively. A combination of plant-derived anti-HIV compounds like protease inhibitors and nucleoside analogs, which are commonly used to treat HIV infection, might be explored in clinical trials for the treatment of COVID-19.

Analytical Assessment of the Vela Diagnostics NGS Assay for HIV Genotyping and Resistance Testing: The Apulian Experience.

Drug-resistance monitoring is one of the hardest challenges in HIV management. Next-generation sequencing (NGS) technologies speed up the detection of drug resistance, allowing the adjustment of antiretroviral therapy and enhancing the quality of life of people living with HIV. Recently, the NGS Sentosa® SQ HIV Genotyping Assay (Vela Diagnostics) received approval for in vitro diagnostics use. This work is the first Italian evaluation of the performance of the Vela Diagnostics NGS platform, assessed with 420 HIV-1 clinical samples. A comparison with Sanger sequencing performance is also reported, highlighting the advantages and disadvantages of the Sentosa® NGS assay. The precision of the technology was studied with reference specimens, while intra- and inter-assay reproducibility were evaluated for selected clinical samples. Vela Diagnostics' NGS assay reached an 87% success rate through 30 runs of analysis in a real-world clinical context. The concordance with Sanger sequencing outcomes was equal to 97.2%. Several detected mismatches were due to NGS's superior sensitivity to low-frequency variants. A high accuracy was observed in testing reference samples. Repeatability and reproducibility assays highlighted the good performance of the NGS platform. Beyond a few technical issues that call for further optimization, the key improvement will be a better balance between costs and processing speed. Once these issues have been solved, the Sentosa® SQ HIV Genotyping Assay will be the way forward for HIV resistance testing.

Synthesis of carbon-14 and stable isotope labeled Censavudine.

Censavudine is a nucleoside reverse transcriptase inhibitor (NRTI) explored clinically by Bristol Myers Squibb for the treatment of human immunodeficiency virus-1 (HIV-1). As part of the development process, a carbon-14 labeled analog was synthesized for use in a human absorption, distribution, metabolism, and excretion (ADME) study. A stable isotope labeled analog was also synthesized for use as a mass spectrum internal standard in bioanalytical assays to accurately quantify the concentration of the drug in biological samples. Carbon-14 labeled Censavudine was synthesized in 10 steps in a 9% overall yield from carbon-14 labeled trimethylsilylacetylene. A total of 4.44 mCi of material was prepared with a specific activity of 0.25 µCi/mg. The radiochemical and UV purities were 99% and it met all of the specifications for use in a human clinical study. Deuterium labeled Censavudine was synthesized in two steps in a 68% overall yield from [D4 ]-thymine. A total of 237 mg were prepared with a UV purity of 99%.

Novel Therapies of Hepatitis B and D.

Hepatitis B virus (HBV) infection is a global public health issue and is a major cause of cirrhosis and hepatocellular carcinoma (HCC). Hepatitis D virus (HDV) requires the hepatitis B surface antigen (HBsAg) to replicate. The eradication of HBV, therefore, can also cure HDV. The current therapies for chronic hepatitis B and D are suboptimal and cannot definitely cure the viruses. In order to achieve functional or complete cure of these infections, novel therapeutic agents that target the various sites of the viral replicative cycle are necessary. Furthermore, novel immunomodulatory agents are also essential to achieve viral clearance. Many of these new promising compounds such as entry inhibitors, covalently closed circular DNA (cccDNA) inhibitors, small interfering RNAs (siRNAs), capsid assembly modulators and nucleic acid polymers are in various stages of clinical developments. In this review article, we provided a comprehensive overview of the structure and lifecycle of HBV, the limitations of the current therapies and a summary of the novel therapeutic agents for both HDV and HBV infection.

Human PrimPol Discrimination against Dideoxynucleotides during Primer Synthesis.

PrimPol is required to re-prime DNA replication at both nucleus and mitochondria, thus facilitating fork progression during replicative stress. ddC is a chain-terminating nucleotide that has been widely used to block mitochondrial DNA replication because it is efficiently incorporated by the replicative polymerase Polγ. Here, we show that human PrimPol discriminates against dideoxynucleotides (ddNTP) when elongating a primer across 8oxoG lesions in the template, but also when starting de novo synthesis of DNA primers, and especially when selecting the 3'nucleotide of the initial dimer. PrimPol incorporates ddNTPs with a very low efficiency compared to dNTPs even in the presence of activating manganese ions, and only a 40-fold excess of ddNTP would significantly disturb PrimPol primase activity. This discrimination against ddNTPs prevents premature termination of the primers, warranting their use for elongation. The crystal structure of human PrimPol highlights Arg291 residue as responsible for the strong dNTP/ddNTP selectivity, since it interacts with the 3'-OH group of the incoming deoxynucleotide, absent in ddNTPs. Arg291, shown here to be critical for both primase and polymerase activities of human PrimPol, would contribute to the preferred binding of dNTPs versus ddNTPs at the 3'elongation site, thus avoiding synthesis of abortive primers.

Development of Human Immunodeficiency Virus Type 1 Resistance to 4'-Ethynyl-2-Fluoro-2'-Deoxyadenosine Starting with Wild-Type or Nucleoside Reverse Transcriptase Inhibitor-Resistant Strains.

4'-Ethynyl-2-fluoro-2'-deoxyadenosine (EFdA, MK-8591, islatravir) is a nucleoside reverse transcriptase translocation inhibitor (NRTTI) with exceptional potency against wild-type (WT) and drug-resistant HIV-1 in phase III clinical trials. EFdA resistance is not well characterized. To study EFdA resistance patterns that may emerge in naive or tenofovir (TFV)-, emtricitabine/lamivudine (FTC/3TC)-, or zidovudine (AZT)-treated patients, we performed viral passaging experiments starting with WT, K65R, M184V, or D67N/K70R/T215F/K219Q HIV-1. Regardless of the starting viral sequence, all selected EFdA-resistant variants included the M184V reverse transcriptase (RT) mutation. Using recombinant viruses, we validated the role for M184V as the primary determinant of EFdA resistance; none of the observed connection subdomain (R358K and E399K) or RNase H domain (A502V) mutations significantly contributed to EFdA resistance. A novel EFdA resistance mutational pattern that included A114S was identified in the background of M184V. A114S/M184V exhibited higher EFdA resistance (∼24-fold) than either M184V (∼8-fold) or A114S alone (∼2-fold). Remarkably, A114S/M184V and A114S/M184V/A502V resistance mutations were up to 50-fold more sensitive to tenofovir than was WT HIV-1. These mutants also had significantly lower specific infectivities than did WT. Biochemical experiments confirmed decreases in the enzymatic efficiency (kcat/Km) of WT versus A114S (2.1-fold) and A114S/M184V/A502V (6.5-fold) RTs, with no effect of A502V on enzymatic efficiency or specific infectivity. The rather modest EFdA resistance of M184V or A114S/M184V (8- and 24-fold), their hypersusceptibility to tenofovir, and strong published in vitro and in vivo data suggest that EFdA is an excellent therapeutic candidate for naive, AZT-, FTC/3TC-, and especially tenofovir-treated patients.

Nucleoside Analogues Are Potent Inducers of Pol V-mediated Mutagenesis.

Drugs targeting DNA and RNA in mammalian cells or viruses can also affect bacteria present in the host and thereby induce the bacterial SOS system. This has the potential to increase mutagenesis and the development of antimicrobial resistance (AMR). Here, we have examined nucleoside analogues (NAs) commonly used in anti-viral and anti-cancer therapies for potential effects on mutagenesis in Escherichia coli, using the rifampicin mutagenicity assay. To further explore the mode of action of the NAs, we applied E. coli deletion mutants, a peptide inhibiting Pol V (APIM-peptide) and metabolome and proteome analyses. Five out of the thirteen NAs examined, including three nucleoside reverse transcriptase inhibitors (NRTIs) and two anti-cancer drugs, increased the mutation frequency in E. coli by more than 25-fold at doses that were within reported plasma concentration range (Pl.CR), but that did not affect bacterial growth. We show that the SOS response is induced and that the increase in mutation frequency is mediated by the TLS polymerase Pol V. Quantitative mass spectrometry-based metabolite profiling did not reveal large changes in nucleoside phosphate or other central carbon metabolite pools, which suggests that the SOS induction is an effect of increased replicative stress. Our results suggest that NAs/NRTIs can contribute to the development of AMR and that drugs inhibiting Pol V can reverse this mutagenesis.

Development and Validation of an Up-to-Date Highly Sensitive UHPLC-MS/MS Method for the Simultaneous Quantification of Current Anti-HIV Nucleoside Analogues in Human Plasma.

Therapeutic options to treat HIV infection have widened in the past years, improving both effectiveness and tolerability, but nucleoside reverse transcriptase inhibitors (NRTIs) are still considered the standard backbone of the combination regimens. Therapeutic drug monitoring (TDM) can be useful for these drugs, due to concentration-effect relationship, with risk of ineffectiveness, toxicity or adherence concerns: in this scenario, robust and multiplexed methods are needed for an effective TDM activity. In this work, the first validated ultra-high spectrometry liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) method is described for the high-sensitive simultaneous quantification of all the currently used NRTIs in human plasma, including tenofovir alafenamide (TAF), following FDA and EMA guidelines. The automated sample preparation consisted in the addition of an internal standard (IS) working solution, containing stable-isotope-linked drugs, protein precipitation and drying. Dry extracts were reconstituted with water, then, these underwent reversed phase chromatographic separation: compounds were detected through electrospray ionization and multiple reaction monitoring. Accuracy, precision, recovery and IS-normalized matrix effect fulfilled guidelines' requirements. The application of this method on samples from people living with HIV (PLWH) showed satisfactory performance, being capable of quantifying the very low concentrations of tenofovir (TFV) in patients treated with TAF.

Increased acquired protease inhibitor drug resistance mutations in minor HIV-1 quasispecies from infected patients suspected of failing on national second-line therapy in South Africa.

BACKGROUND: HIV-1C has been shown to have a greater risk of virological failure and reduced susceptibility towards boosted protease inhibitors (bPIs), a component of second-line combination antiretroviral therapy (cART) in South Africa. This study entailed an evaluation of HIV-1 drug resistance-associated mutations (RAMs) among minor viral populations through high-throughput sequencing genotypic resistance testing (HTS-GRT) in patients on the South African national second-line cART regimen receiving bPIs. METHODS: During 2017 and 2018, 67 patient samples were sequenced using high-throughput sequencing (HTS), of which 56 samples were included in the final analysis because the patient's treatment regimen was available at the time of sampling. All patients were receiving bPIs as part of their cART. Viral RNA was extracted, and complete pol genes were amplified and sequenced using Illumina HiSeq2500, followed by bioinformatics analysis to quantify the RAMs according to the Stanford HIV Drug Resistance Database. RESULTS: Statistically significantly higher PI RAMs were observed in minor viral quasispecies (25%; 14/56) compared to non-nucleoside reverse transcriptase inhibitors (9%; 5/56; p = 0.042) and integrase inhibitor RAM (4%; 2/56; p = 0.002). The majority of the drug resistance mutations in the minor viral quasispecies were observed in the V82A mutation (n = 13) in protease and K65R (n = 5), K103N (n = 7) and M184V (n = 5) in reverse transcriptase. CONCLUSIONS: HTS-GRT improved the identification of PI and reverse transcriptase inhibitor (RTI) RAMs in second-line cART patients from South Africa compared to the conventional GRT with ≥20% used in Sanger-based sequencing. Several RTI RAMs, such as K65R, M184V or K103N and PI RAM V82A, were identified in < 20% of the population. Deep sequencing could be of greater value in detecting acquired resistance mutations early.

Trends and strategies to combat viral infections: A review on FDA approved antiviral drugs.

The infectious microscopic viruses invade living cells to reproduce themselves, and causes chronic infections such as HIV/AIDS, hepatitis B and C, flu, etc. in humans which may lead to death if not treated. Different strategies have been utilized to develop new and superior antiviral drugs to counter the viral infections. The FDA approval of HIV nucleoside reverse transcriptase inhibitor, zidovudine in 1987 boosted the development of antiviral agents against different viruses. Currently, there are a number of combination drugs developed against various viral infections to arrest the activity of same or different viral macromolecules at multiple stages of its life cycle; among which majority are targeted to interfere with the replication of viral genome. Besides these, other type of antiviral molecules includes entry inhibitors, integrase inhibitors, protease inhibitors, interferons, immunomodulators, etc. The antiviral drugs can be toxic to human cells, particularly in case of administration of combination drugs, and on the other hand viruses can grow resistant to the antiviral drugs. Furthermore, emergence of new viruses like Ebola, coronaviruses (SARS-CoV, SARS-CoV-2) emphasizes the need for more innovative strategies to develop better antiviral drugs to fight the existing and the emerging viral infections. Hence, we reviewed the strategic enhancements in developing antiviral drugs for the treatment of different viral infections over the years.

Emergence of dual antiretroviral therapy as a viable regimen option for the treatment of patients with HIV infection.

Current antiretroviral therapy is not curative. The need for life-long therapy brings with it concerns regarding long-term toxicity and cost. Thus, investigations into simpler regimens with comparable efficacy and improved safety have been undertaken and continue to be conducted. Various 2-drug combinations have been evaluated with variable results. The combinations of dolutegravir plus lamivudine and dolutegravir plus rilpivirine were found to be comparable in efficacy to conventional 3-drug regimens and have now been approved by the United States Food and Drug Administration (FDA) and have entered into clinical practice. Dolutegravir/rilpivirine was approved for the treatment of adults with HIV-1 infection whose virus has been suppressed on a stable regimen for at least 6 months, with no history of treatment failure and no known substitutions associated with resistance to the individual components of the combination. Dolutegravir/lamivudine was approved for the treatment of HIV infection in adults with no antiretroviral treatment history and with no known or suspected resistance to the individual components of the combination.