Target-based drug design strategies to overcome resistance to antiviral agents: opportunities and challenges.

Viral infections have a major impact in human health. Ongoing viral transmission and escalating selective pressure have the potential to favor the emergence of vaccine- and antiviral drug-resistant viruses. Target-based approaches for the design of antiviral drugs can play a pivotal role in combating drug-resistant challenges. Drug design computational tools facilitate the discovery of novel drugs. This review provides a comprehensive overview of current drug design strategies employed in the field of antiviral drug resistance, illustrated through the description of a series of successful applications. These strategies include technologies that enhance compound-target affinity while minimizing interactions with mutated binding pockets. Furthermore, emerging approaches such as virtual screening, targeted protein/RNA degradation, and resistance analysis during drug design have been harnessed to curtail the emergence of drug resistance. Additionally, host targeting antiviral drugs offer a promising avenue for circumventing viral mutation. The widespread adoption of these refined drug design strategies will effectively address the prevailing challenge posed by antiviral drug resistance.

May I Help You with Your Coat? HIV-1 Capsid Uncoating and Reverse Transcription.

The human immunodeficiency virus type 1 (HIV-1) capsid is a protein core formed by multiple copies of the viral capsid (CA) protein. Inside the capsid, HIV-1 harbours all the viral components required for replication, including the genomic RNA and viral enzymes reverse transcriptase (RT) and integrase (IN). Upon infection, the RT transforms the genomic RNA into a double-stranded DNA molecule that is subsequently integrated into the host chromosome by IN. For this to happen, the viral capsid must open and release the viral DNA, in a process known as uncoating. Capsid plays a key role during the initial stages of HIV-1 replication; therefore, its stability is intimately related to infection efficiency, and untimely uncoating results in reverse transcription defects. How and where uncoating takes place and its relationship with reverse transcription is not fully understood, but the recent development of novel biochemical and cellular approaches has provided unprecedented detail on these processes. In this review, we present the latest findings on the intricate link between capsid stability, reverse transcription and uncoating, the different models proposed over the years for capsid uncoating, and the role played by other cellular factors on these processes.

Predicting kidney outcomes among Latin American patients with lupus nephritis: The prognostic value of interstitial fibrosis and tubular atrophy and tubulointerstitial inflammation.

OBJECTIVE: To assess the effect of tubulointerstitial inflammation (TII) and interstitial fibrosis and tubular atrophy (IFTA) on kidney survival in lupus nephritis (LN). METHODS: Two hundred eighty five patients with biopsy-proven LN were retrospectively studied. Kidney survival was defined as the time from initial biopsy to end-stage kidney disease (ESKD), dialysis, or transplant. Kidney survival analysis was performed by the Kaplan-Meier method and the statistical difference between survival curves compared by the log-rank test. Cumulative incidence functions with competing risk of death for kidney survival were also graphed. Multivariable Cox proportional hazards regression and competing-risk analyses were performed to identify independent predictors of ESKD. RESULTS: Fifty-seven patients (20%) progressed to ESKD during a median time of 4.2 (2.0-55.2) months after biopsy. TII was present in 206 (72.3%) biopsies, while IFTA in 99 (34.7%) biopsies. Patients with moderate-to-severe IFTA had worse kidney survival than those with none or mild IFTA in both the Kaplan-Meier (p = 0.018) and the competing-risk analyses (p = 0.017). Patients with class IV ± V LN had worse kidney survival than those with non-class IV LN by the Kaplan-Meier method (p = 0.050), but not in the competing-risk analysis (p = 0.154). Worse kidney survival was also found among those with fibrous crescents than those without, in both the Kaplan-Meier (p = 0.010) and the competing-risk (p = 0.011) analyses. By multivariable Cox regression analysis, older age (HR 1.04, 95% CI 1.01-1.07) and class IV ± V LN (HR 5.06, 95% CI 1.82-14.09) were associated with higher risk of ESKD after adjusting for sex, ethnicity, TII, and IFTA. By competing-risk analyses, class IV ± V LN (SHR 3.32, 95% CI 1.25-8.83) and no response to immunosuppressive therapy (SHR 4.55, 95% CI 1.54-13.41) were associated with a higher risk of ESKD, while eGFR >90 mL/min/1.73 m2 (SHR 0.98 for each ml/min/1.73 m2, 95% 0.97-0.99) with a lower risk. CONCLUSIONS: Patients with moderate-to-severe IFTA had worse kidney survival than those with none or mild IFTA. Worse kidney survival was also found among those with class IV LN and fibrous crescents versus those without IV LN and fibrous crescents, respectively.

Decoding molnupiravir-induced mutagenesis in SARS-CoV-2.

Molnupiravir, a prodrug of the nucleoside derivative ß-D-N4-hydroxycytidine (NHC), is currently in clinical trials for COVID-19 therapy. However, the biochemical mechanisms involved in molnupiravir-induced mutagenesis had not been explored. In a recent study, Gordon et al. demonstrated that NHC can be incorporated into viral RNA and subsequently extended and used as template for RNA-dependent RNA synthesis, proposing a mutagenesis model consistent with available virological evidence. Their study uncovers molecular mechanisms by which molnupiravir drives SARS-CoV-2 into error catastrophe.

Efficacy and Safety of Intravitreal Aflibercept Treat and Extend for Polypoidal Choroidal Vasculopathy in the ATLANTIC Study: A Randomized Clinical Trial.

IMPORTANCE: Polypoidal choroidal vasculopathy (PCV) is far less common and studied in a Caucasian population than in an Asian population, and the optimal treatment approach remains to be confirmed. METHODS: A 52-week, double-masked, sham-controlled, phase 4, investigator-initiated randomized clinical trial (RCT) in naive symptomatic Caucasian patients with PCV treated with aflibercept in a treat-and-extend regimen (T&E) (intravitreal aflibercept injection [IVAI] T&E). Patients were randomized at week 16 to receive IVAI T&E plus either sham photodynamic therapy (PDT) or standard fluence PDT with verteporfin. The main outcome measures were changes in best-corrected visual acuity (BCVA) from baseline to 52 weeks and polyp occlusion at week 52. Data are presented as median (interquartile range [IQR]) for BCVA, number of IVAI, and change in central retinal thickness (CRT). RESULTS: Of the 50 patients included in the study, 48 patients completed the 52 weeks of follow-up. During this period, a significant median (IQR) BCVA gain of 6 [2-12] Early Treatment Diabetic Retinopathy Study letters was observed for all patients (p < 0.001), after 8 (7-9) injections, with a significant reduction of -93.0 [-154.0, -44.0] µm in central macular thickness (p < 0.001). Using indocyanine green angiography, a complete occlusion of polypoidal lesions was documented in 72% of the cases. Still, no significant difference was detected between the sham PDT and the aflibercept PDT arms, at week 52, for BCVA change (6.5 [2-11] vs. 5 [2-13] letters (p = 0.98)), number of IVAIs (8.5 [7-9] vs. 8 [7-9] (p = 0.21)), change in CRT (-143 [-184; -47] vs. -89 [-123; -41.5] µm [p = 0.23]), and rates of complete polyp occlusion: 77 versus 68% (p = 0.53) or presence of fluid: 68 versus 57% (p = 0.56). No serious ocular adverse events were registered in the 2 arms. CONCLUSIONS AND RELEVANCE: To our knowledge, this is the first RCT to compare aflibercept T&E monotherapy with aflibercept T&E plus verteporfin PDT in a Caucasian population with PCV. Aflibercept monotherapy in a T&E showed to be effective and safe with a significant median BCVA improvement of 6 letters and a complete occlusion of polypoidal lesions in near 3 quarters of the eyes, at 1 year. As only 22% of the eyes underwent PDT treatment, the benefit of combined treatment for PCV in Caucasian patients could not be definitively elucidated from this study. TRIAL REGISTRATION: The clinical trial was registered in ClinicalTrials.gov Identifier NCT02495181 and the European Union Drug Regulating Authorities Clinical Trials Database EudraCT No. 2015-001368-20.

Medicinal chemistry strategies for discovering antivirals effective against drug-resistant viruses.

During the last forty years we have witnessed impressive advances in the field of antiviral drug discovery culminating with the introduction of therapies able to stop human immunodeficiency virus (HIV) replication, or cure hepatitis C virus infections in people suffering from liver disease. However, there are important viral diseases without effective treatments, and the emergence of drug resistance threatens the efficacy of successful therapies used today. In this review, we discuss strategies to discover antiviral compounds specifically designed to combat drug resistance. Currently, efforts in this field are focused on targeted proteins (e.g. multi-target drug design strategies), but also on drug conformation (either improving drug positioning in the binding pocket or introducing conformational constraints), in the introduction or exploitation of new binding sites, or in strengthening interaction forces through the introduction of multiple hydrogen bonds, covalent binding, halogen bonds, additional van der Waals forces or multivalent binding. Among the new developments, proteolysis targeting chimeras (PROTACs) have emerged as a valid approach taking advantage of intracellular mechanisms involving protein degradation by the ubiquitin-proteasome system. Finally, several molecules targeting host factors (e.g. human dihydroorotate dehydrogenase and DEAD-box polypeptide 3) have been identified as broad-spectrum antiviral compounds. Implementation of herein described medicinal chemistry strategies are expected to contribute to the discovery of new drugs effective against current and future threats due to emerging and re-emerging viral pandemics.

Kidney transplantation outcomes in lupus nephritis: A 37-year single-center experience from Latin America.

OBJECTIVE: We assessed patient and graft outcomes and prognostic factors in kidney transplantation in patients with end-stage kidney disease (ESKD) secondary to lupus nephritis (LN) undergoing kidney transplantation from August 1977 to December 2014 in a Latin American single center. METHODS: The primary endpoint was patient survival, and the secondary endpoints were death-censored graft survival for the first renal transplant and the rate of recurrent LN (RLN). Kaplan-Meier method was used for survival analysis. Factors predicting patient and death-censored graft survivals were examined by Cox proportional-hazards regression analyses. RESULTS: 185 patients were retrospectively evaluated. Patient survival rates were 88% at one year, 82% at three years, 78% at five years, and 67% at ten years. Death-censored graft survival for the first renal transplant was 93% at one year, 89% at three years, 87% at five years, and 80% at ten years. RLN was diagnosed in 2 patients (1.08%), but no graft was lost because of RLN. Thirty-nine (21.1%) patients died, and 65 (35.1%) patients experienced graft loss during the follow-up. By multivariable analyses, older recipient age and 1-month posttransplantation eGFR <45 ml/min/1.73m2 were associated with lower patient survival and an increased risk of graft loss, while induction immunosuppressive therapy exerted a protective effect on patients' survival. In the subgroup of patients in whom disease activity was measured at the time of transplantation, a higher SLEDAI score was also associated with lower patient survival and an increased risk of graft loss. CONCLUSION: In a mostly Mestizo population, kidney transplantation is an excellent therapeutic alternative in LN patients with ESKD. Older recipient age, an eGFR <45 ml/min/1.73m2 at one month posttransplantation, and disease activity at the time of transplantation are predictive of a lower patient and death-censored graft survival, while induction immunosuppressive therapy has a protective effect on patient survival. RLN is rare and does not influence the risk of graft loss.

Novel indolylarylsulfone derivatives as covalent HIV-1 reverse transcriptase inhibitors specifically targeting the drug-resistant mutant Y181C.

Non-nucleoside reverse transcriptase inhibitors (NNRTIs) are widely used in combination therapies against HIV-1. However, emergent and transmitted drug resistance compromise their efficacy in the clinical setting. Y181C is selected in patients receiving nevirapine, etravirine and rilpivirine, and together with K103N is the most prevalent NNRTI-associated mutation in HIV-infected patients. Herein, we report on the design, synthesis and biological evaluation of a novel series of indolylarylsulfones bearing acrylamide or ethylene sulfonamide reactive groups as warheads to inactivate Cys181-containing HIV-1 RT via a Michael addition reaction. Compounds I-7 and I-9 demonstrated higher selectivity towards the Y181C mutant than against the wild-type RT, in nucleotide incorporation inhibition assays. The larger size of the NNRTI binding pocket in the mutant enzyme facilitates a better fit for the active compounds, while stacking interactions with Phe227 and Pro236 contribute to inhibitor binding. Mass spectrometry data were consistent with the covalent modification of the RT, although off-target reactivity constitutes a major limitation for further development of the described inhibitors.

Hydrochlorothiazide use and risk of non-melanoma skin cancer in Spain: A case/non-case study.

OBJECTIVE: In 2018, the Pharmacological Risk Assessment Committee alerted to a potential relationship between accumulated hydrochlorothiazide dosage and the risk of non-melanoma skin cancer. To study this relationship we used data from the Spanish Pharmacovigilance System for Medicinal Products of Human Use. MATERIALS AND METHODS: Following a case search for every thiazide potentially associated with (SMQ/MedDRA) "Malignant Skin Neoplasms and not Otherwise Specified Skin Neoplasms", a series of disproportionality analyses were conducted by estimating the reporting odds ratio (95% confidence interval). Registered adverse drug reactions and disproportionality through the reported odds ratio were the main outcome measures. RESULTS: For basal cell carcinoma, reporting odds ratio was 4.8 (2.2 - 10.7); squamous cell carcinoma 3.2 (0.9 - 10.5); malignant melanoma, 0.8 (0.2 - 3.5). We found both disproportionality and association between hydrochlorothiazide and basal cell carcinoma, but none of these were found regarding malignant skin melanoma. In the case of squamous cell carcinoma, the lower confidence interval limit was below 1, thus the disproportionality value was not statistically significant. The accumulated hydrochlorothiazide dose was 36,714 mg for basal cell carcinoma; 98,288 mg for squamous cell carcinoma; and 38,444 mg for malignant melanoma. CONCLUSION: The results in Spain, where sun exposure is significant, are consistent with the data in the Pharmacological Risk Assessment Committee's alert, which were obtained in Denmark for both basal cell carcinoma and malignant melanoma. However, the results for squamous cell carcinoma did not reach statistical significance, although the reporting odds ratio value suggested a potential relationship between hydrochlorothiazide and squamous cell carcinoma.

Topical NSAIDs, intravitreal dexamethasone and peribulbar triamcinolone for pseudophakic macular edema.

BACKGROUND: The purpose of this study is to assess the effectiveness of topical nonsteroidal anti-inflammatory drugs (NSAIDs) and corticosteroids (intravitreal dexamethasone and peribulbar triamcinolone) in treating pseudophakic macular edema (PME). METHODS: Retrospective study of 33 eyes. Variables included best corrected visual acuity (BCVA; logMAR scale) and central retinal thickness (CRT) and central choroidal thickness (CCT) assessed with swept-source OCT. All patients were initially prescribed topical NSAIDs and reevaluated after 2 months. If improvement in BCVA or CRT was noted, topical NSAIDs were continued until resolution. If no improvement was observed at 2 months or subsequent visits, intravitreal dexamethasone implant was performed. Patients who refused intravitreal treatment were offered peribulbar triamcinolone. RESULTS: After treatment with topical NSAIDs for a median of 2 months, BCVA increased significantly from 0.5 to 0.3 while CRT decreased significantly from 435 to 316 µm. PME resolved in 19 of the 33 eyes (57.6%). Of the 14 recalcitrant cases, 13 were treated with corticosteroids. Of these 13 cases, 9 (69.2%) resolved. BCVA increased non-significantly from 0.7 to 0.4. CRT and CCT decreased significantly from 492 to 317 µm and from 204 to 182 µm respectively. CONCLUSIONS: The overall success rate of the treatment algorithm was greater than 80%, a remarkable finding considering that no randomized study has yet been conducted to determine the optimal therapeutic protocol for PME. This is the first study to evaluate choroidal thickness in PME using SS-OCT, which could play a key role in its pathophysiology and provide useful information to improve the management of PME.

An Update on Antiretroviral Therapy.

Human immunodeficiency virus (HIV) infection and acquired immune deficiency syndrome (AIDS) still claim many lives across the world. However, research efforts during the last 40 years have led to the approval of over 30 antiretroviral drugs and the introduction of combination therapies that have turned HIV infection into a chronic but manageable disease. In this chapter, we provide an update on current available drugs and treatments, as well as future prospects towards reducing pill burden and developing long-acting drugs and novel antiretroviral therapies. In addition, we summarize efforts to cure HIV, including pharmaceutical strategies focused on the elimination of the virus.

Search, Identification, and Design of Effective Antiviral Drugs Against Pandemic Human Coronaviruses.

Recent coronavirus outbreaks of SARS-CoV-1 (2002-2003), MERS-CoV (since 2012), and SARS-CoV-2 (since the end of 2019) are examples of how viruses can damage health care and generate havoc all over the world. Coronavirus can spread quickly from person to person causing high morbidity and mortality. Unfortunately, the antiviral armamentarium is insufficient to fight these infections. In this chapter, we provide a detailed summary of the current situation in the development of drugs directed against pandemic human coronaviruses. Apart from the recently licensed remdesivir, other antiviral agents discussed in this review include molecules targeting viral components (e.g., RNA polymerase inhibitors, entry inhibitors, or protease inhibitors), compounds interfering with virus-host interactions, and drugs identified in large screening assays, effective against coronavirus replication, but with an uncertain mechanism of action.

Survival in small choroidal melanocytic lesions with risk factors managed by initial observation until detection of tumour growth.

BACKGROUND: The main objective was to describe metastatic and survival rates in patients with small choroidal melanocytic lesions initially managed by observation. METHODS: Retrospective, observational study of consecutive cases recruited from 2001 through 2018, followed for a median (mean, range) of 81.0 (89.3, 10-204) months in a tertiary referral centre for ocular oncology. Seventy-five consecutive patients diagnosed with small choroidal melanocytic lesions with risk factors for growth initially observed and who showed progression during follow-up. Treatment was performed (plaque radiotherapy or enucleation in 96% and 4% of cases, respectively) at detection of tumour growth. RESULTS: Median (mean, range) tumour thickness was 2.2 (2.23, 1.08-3.40) mm, and median maximum basal diameter was 8.5 (8.16, 4-12) mm. At diagnosis, a median (mean, range) of 5 (5.48, 1-8) risk factors for progression were present. Lesions grew at a median (mean, range) rate of 0.42 mm/y (1.12, 0-7.68) in thickness and 1.05 mm/y (3.14, 0-4.8) in maximum diameter. Median (mean, range) time until growth was 17.00 (32.6, 1-161) months post-diagnosis, at which time tumours were treated. Five patients developed local recurrence after brachytherapy requiring enucleation. Four patients developed hepatic metastasis. Melanoma-specific survival was 98% at 5 years (95% CI, 94.2-100%) and 91.6% (95% CI, 82-100%) at 10 and 15 years. CONCLUSION: In small melanocytic lesions with risk factors for growth, initial observation until detection of tumour growth results in a seemingly low risk of metastasis, suggesting that this may be an initial approach to consider in tumours with indeterminate malignant potential.

Economic burden of age-related macular degeneration in routine clinical practice: the RAMDEBURS study.

PURPOSE: To describe and evaluate the main direct health costs, in routine clinical practice, of age-related macular degeneration (AMD) patients, from hospital perspective, in Spain. METHODS: Retrospective, multicenter, and observational study conducted on five third-level Spanish hospitals, between December 2018 and December 2019. The study included patients who were diagnosed of AMD before December 2018. Direct healthcare costs were obtained from a Spanish database. Study variables included demographic and clinical variables, and resources, such as treatment, diagnostic tests, medical examination, and surgery. Among the 1414 screened AMD patients, 1164 patients were included. In the overall study patients, the total cost was €5,386,511.0, with a mean cost per patient of €4627.6 ± 2383.9. The largest cost items were diagnostic examinations (€2.832.902,0) and vascular endothelial growth factor inhibitors (anti-VEGF) treatment (€2.038.257,2). Bevacizumab was administered to 325 (27.9%) patients, ranibizumab to 328 (28.2%), and aflibercept to 626 (53.8%); 115 (10.7%) patients received two anti-VEGF treatments, while 90 (7.7%) did not receive any. Over the course of the study, a total of 6,057 anti-VEGF injections were administered, with a mean (95% confidence interval) of 4.8 (4.4-5.2) injections per patient. Regarding safety, 29 patients experience injection-related adverse events, among them 12 patients had cataract and 11 ones elevated intraocular pressure (IOP). The incidence of endophthalmitis was 0.5% (6/1164). CONCLUSIONS: AMD was associated with considerable healthcare costs for regional healthcare systems. Diagnostic examinations, particularly OCT examinations, and anti-VEGF treatment represented the largest cost items.

Nucleocapsid Protein Precursors NCp9 and NCp15 Suppress ATP-Mediated Rescue of AZT-Terminated Primers by HIV-1 Reverse Transcriptase.

In HIV-1, development of resistance to AZT (3'-azido-3'-deoxythymidine) is mediated by the acquisition of thymidine analogue resistance mutations (TAMs) (i.e., M41L, D67N, K70R, L210W, T215F/Y, and K219E/Q) in the viral reverse transcriptase (RT). Clinically relevant combinations of TAMs, such as M41L/T215Y or D67N/K70R/T215F/K219Q, enhance the ATP-mediated excision of AZT monophosphate (AZTMP) from the 3' end of the primer, allowing DNA synthesis to continue. Additionally, during HIV-1 maturation, the Gag polyprotein is cleaved to release a mature nucleocapsid protein (NCp7) and two intermediate precursors (NCp9 and NCp15). NC proteins interact with the viral genome and facilitate the reverse transcription process. Using wild-type and TAM-containing RTs, we showed that both NCp9 and NCp15 inhibited ATP-mediated rescue of AZTMP-terminated primers annealed to RNA templates but not DNA templates, while NCp7 had no effect on rescue activity. RNase H inactivation by introducing the active-site mutation E478Q led to the loss of the inhibitory effect shown by NCp9. NCp15 had a stimulatory effect on the RT's RNase H activity not observed with NCp7 and NCp9. However, analysis of RNase H cleavage patterns revealed that in the presence of NCp9, RNA/DNA complexes containing duplexes of 12 bp had reduced stability in comparison with those obtained in the absence of NC or with NCp7 or NCp15. These effects are expected to have a strong influence on the inhibitory action of NCp9 and NCp15 by affecting the efficiency of RNA-dependent DNA polymerization after unblocking DNA primers terminated with AZTMP and other nucleotide analogues.

EFFICACY AND SAFETY OF A TREAT-AND-EXTEND REGIMEN WITH AFLIBERCEPT IN TREATMENT-NAIVE PATIENTS WITH TYPE 3 NEOVASCULARIZATION: A 52-Week, Single-Arm, Multicenter Trial.

PURPOSE: To evaluate 52-week efficacy and safety of a treat-and-extend regimen of intravitreal aflibercept 2 mg on treatment-naive Type 3 neovascularization lesions. METHODS: Phase IV, prospective, open-label, single-arm, multicenter trial including patients with untreated Stage I/II Type 3 neovascularization lesions and baseline best-corrected visual acuity between 78 and 23 Early Treatment Diabetic Retinopathy Study letters. Primary endpoint: mean change in best-corrected visual acuity from baseline at 52 weeks. RESULTS: Thirty-two eyes from 32 patients were included (mean ± SD age: 78.2 ± 7.7 years, 68.8% females, baseline best-corrected visual acuity: 57.9 ± 15.4 [Snellen fraction 20/70]). Best-corrected visual acuity increased by 10.5 ± 15.9 Early Treatment Diabetic Retinopathy Study letters at Week 52 (P = 0.0001). The mean foveal and choroidal thickness decreased by 129.1 ± 80.1 µm (P < 0.0001) and 64.3 ± 96.5 (P = 0.0001), respectively. The proportion of patients with intraretinal/subretinal fluid decreased from 28 (87.5%) at baseline to 3 (11.5%) at Week 52 (P < 0.0001). Pigment epithelial detachment and lesion area showed nonsignificant changes over 52 weeks. The mean number of injections was 8.0 ± 2.0. Seven (21.9%) patients experienced treatment-related adverse events and two (6.3%) experienced serious adverse events; one (3.1%) ocular serious adverse event requiring treatment withdrawal, endophthalmitis, and one (3.1%) nonocular spontaneously resolved serious adverse event, palpitations. One (3.1%) patient experienced an APTC ATE: nonfatal stroke not related to trial treatment. CONCLUSION: A treat-and-extend regimen of aflibercept improves visual acuity and retinal edema in eyes with Type 3 neovascularization over 52 weeks with good tolerability.

INTERNAL LIMITING MEMBRANE PEELING VERSUS NONPEELING TO PREVENT EPIRETINAL MEMBRANE DEVELOPMENT IN PRIMARY RHEGMATOGENOUS RETINAL DETACHMENT: A Swept-Source Optical Coherence Tomography Study With a New Postoperative Classification System.

PURPOSE: To determine whether internal limiting membrane peeling in primary rhegmatogenous retinal detachment prevents epiretinal membrane (ERM) development. Secondarily, we propose a classification system for postoperative ERMs. METHODS: Retrospective, interventional, comparative case series. Consecutive eyes with primary rhegmatogenous retinal detachment (n = 140) treated by a single surgeon. The presence of postoperative ERMs was assessed with swept-source optical coherence tomography. RESULTS: An ERM was detected in 26 eyes (46.4%) in the nonpeeling group and in one eye (1.8%) in the internal limiting membrane peeling group (P ≤ 0.001). The median visual acuity significantly improved in both groups (P ≤ 0.001). Inner retinal dimples were observed in 41.1% of eyes in the internal limiting membrane peeling group versus 0% in the nonpeeling group (P ≤ 0.001), and they were not correlated with visual acuity (r = 0.011; P = 0.941). Based on swept-source optical coherence tomography findings, we identified three different types of ERMs: 7 (26.9%) were classified as Type 1, 12 (46.1%) as Type 2, and 7 (26.9%) as Type 3. Superficial retinal plexus deformations observed on optical coherence tomography angiography and en face images were detected in 100% of Type 3 ERMs, 41.6% of Type 2, and 0% of Type 1 (χ = 14.3; P = 0.001). Interestingly, all of the patients who presented these alterations also had metamorphopsia. CONCLUSION: Internal limiting membrane peeling in primary rhegmatogenous retinal detachment seems to prevent postoperative ERM development. Swept-source optical coherence tomography analysis is helpful to define and classify different types of ERMs and to establish the surgical indication for their removal.

Template-primer binding affinity and RNase H cleavage specificity contribute to the strand transfer efficiency of HIV-1 reverse transcriptase.

During reverse transcription of the HIV-1 genome, two strand-transfer events occur. Both events rely on the RNase H cleavage activity of reverse transcriptases (RTs) and template homology. Using a panel of mutants of HIV-1BH10 (group M/subtype B) and HIV-1ESP49 (group O) RTs and in vitro assays, we demonstrate that there is a strong correlation between RT minus-strand transfer efficiency and template-primer binding affinity. The highest strand transfer efficiencies were obtained with HIV-1ESP49 RT mutants containing the substitutions K358R/A359G/S360A, alone or in combination with V148I or T355A/Q357M. These HIV-1ESP49 RT mutants had been previously engineered to increase their DNA polymerase activity at high temperatures. Now, we found that RTs containing RNase H-inactivating mutations (D443N or E478Q) were devoid of strand transfer activity, whereas enzymes containing F61A or L92P had very low strand transfer activity. The strand transfer defect produced by L92P was attributed to a loss of template-primer binding affinity and, more specifically, to the higher dissociation rate constants (koff) shown by RTs bearing this substitution. Although L92P also deleteriously affected the RT's nontemplated nucleotide addition activity, neither nontemplated nucleotide addition activity nor the RT's clamp activities contributed to increased template switching when all tested mutant and WT RTs were considered. Interestingly, our results also revealed an association between efficient strand transfer and the generation of secondary cleavages in the donor RNA, consistent with the creation of invasion sites. Exposure of the elongated DNA at these sites facilitate acceptor (RNA or DNA) binding and promote template switching.

Amino acid residues in HIV-2 reverse transcriptase that restrict the development of nucleoside analogue resistance through the excision pathway.

Nucleoside reverse transcriptase (RT) inhibitors (NRTIs) are the backbone of current antiretroviral treatments. However, the emergence of viral resistance against NRTIs is a major threat to their therapeutic effectiveness. In HIV-1, NRTI resistance-associated mutations either reduce RT-mediated incorporation of NRTI triphosphates (discrimination mechanism) or confer an ATP-mediated nucleotide excision activity that removes the inhibitor from the 3' terminus of DNA primers, enabling further primer elongation (excision mechanism). In HIV-2, resistance to zidovudine (3'-azido-3'-deoxythymidine (AZT)) and other NRTIs is conferred by mutations affecting nucleotide discrimination. Mutations of the excision pathway such as M41L, D67N, K70R, or S215Y (known as thymidine-analogue resistance mutations (TAMs)) are rare in the virus from HIV-2-infected individuals. Here, we demonstrate that mutant M41L/D67N/K70R/S215Y HIV-2 RT lacks ATP-dependent excision activity, and recombinant virus containing this RT remains susceptible to AZT inhibition. Mutant HIV-2 RTs were tested for their ability to unblock and extend DNA primers terminated with AZT and other NRTIs, when complexed with RNA or DNA templates. Our results show that Met73 and, to a lesser extent, Ile75 suppress excision activity when TAMs are present in the HIV-2 RT. Interestingly, recombinant HIV-2 carrying a mutant D67N/K70R/M73K RT showed 10-fold decreased AZT susceptibility and increased rescue efficiency on AZT- or tenofovir-terminated primers, as compared with the double-mutant D67N/K70R. Molecular dynamics simulations reveal that Met73influences ß3-ß4 hairpin loop conformation, whereas its substitution affects hydrogen bond interactions at position 70, required for NRTI excision. Our work highlights critical HIV-2 RT residues impeding the development of excision-mediated NRTI resistance.

New Anticoagulant Agents: Incidence of Adverse Drug Reactions and New Signals Thereof.

The aim of this study was to evaluate the adverse drug reaction (ADR) incidence rate and new signals thereof for classic compared with new anticoagulants in real-life ambulatory settings. The authors performed an observational cross-sectional study in two cohorts of surveyed patients treated with vitamin K antagonists (VKAs; acenocoumarol or warfarin) or nonvitamin K antagonist oral anticoagulants (NOACs; apixaban, edoxaban, rivaroxaban, dabigatran etexilate). Descriptive, clinical, and ADRs data were reported and analyzed through a bivariate analysis (odds ratio [OR]) to compare the ADRs incidence rate and an adaptation of Bayesian methodology (false discovery rate [FDR] < 0.05) to detect new signals. A total of 334 patients were surveyed-average international normalized ratio (INR) of 2.6-and 45.4% taking new anticoagulants. Note that 835 ADRs were reported; 2.5 per patient (2.8 in the VKA cohort, 2.1 in the NOAC cohort). The authors obtained higher risk of epistaxis (OR, 2.18; 95% confidence interval [CI], 1.01-4.74) and hematoma (OR, 2.43; 95% CI, 1.39-4.25) with VKAs and lower risk of global bleeding symptoms with NOACs (OR, 0.45; 95% CI, 0.28-0.71). After standardizing the data, a significant risk of diarrhea with VKAs was observed (OR, 3.37; 95% CI, 1.09-10.41). They also detected an intense positive signal regarding the use of VKAs and osteoporosis (FDR < 0.001), specifically acenocoumarol (FDR < 0.002). NOACs presented lower risk of bleeding, especially dabigatran (FDR < 0.031), and of dermatological pathologies with apixaban being the safest (FDR = 0.050). The lower risk of global bleeding and a potential protective effect against osteoporosis in patients treated with NOACs postulate them as safer than VKAs.