Herein, we report the identification of the HIV protease inhibitor amprenavir as a selective cannabinoid receptor 2 (CB2) agonist and describe structure-activity relationship (SAR) studies toward repurposing this peripherally restricted scaffold for high CB2 potency and CNS exposure. This exercise yielded compounds with exceptional CB2 potency (EC50s <10 nM), no appreciable activity at the CB1 receptor, and high predicted permeability/low P-gp efflux activity. Selected compounds were profiled in rat i.v. dosing cassettes; several novel amprenavir analogues displayed good t1/2 (>2 h), moderate plasma clearance, and appreciable brain exposure. Additionally, fully flexible protein-ligand docking studies with molecular dynamics (MD) simulations were used to predict the most likely mode of interaction of highly potent analogue VU6077967 with CB2 and to provide a rationale for the observed selectivity of this series relative to CB1.

Organic anion-transporting polypeptides (OATPs) are key transporters of hepatic uptake for endogenous compounds and xenobiotics. Human OATP1B1 and OATP1B3 are well-studied due to their role in drug-drug interactions. In contrast, data on murine OATP1B2, the rodent orthologue of these transporters, are limited, despite its importance in early drug development. Here, we systematically compared the transport characteristics of mouse and rat OATP1B2 under identical experimental conditions. The Km values for estrone-3-sulfate (E1S) and taurocholate (TCA) were 242 and 73 µM for mOATP1B2 and 90 and 16 µM for rOATP1B2. Nine clinically relevant drugs were evaluated for inhibitory effects, showing strong correlation between species. Cyclosporine A, ritonavir, odevixibat, rosuvastatin, and rifampicin markedly inhibited uptake. Rifampicin demonstrated species-specific differences, with higher IC50 values for mOATP1B2 (E1S: 9.6 µM; TCA: 7.7 µM) than rOATP1B2 (E1S: 1.1 µM; TCA: 2.4 µM). A comparison of the rodent data with the human orthologues revealed similar inhibition patterns but distinct substrate selectivity: hOATP1B1 showed high affinity for E1S but negligible TCA uptake, while hOATP1B3 transported TCA weakly but not E1S. This study provides insights into species-specific differences in OATP-mediated hepatic uptake and is therefore valuable for the interpretation of preclinical studies and their transfer to human pharmacology.

Background/Objectives: Predicting whether a compound is subject to active transport is crucial in drug development. We propose a simple threshold for passive membrane permeability (Pm), derived from the cell's energy limitation, to identify compounds unlikely to be actively effluxed. Results: By considering fundamental cellular energy constraints, our approach provides a mechanistic rationale for why compounds with very high passive permeability in combination with low applied concentration will not undergo active efflux. This moves beyond the empirical observation (such as in previous systems that associate fast-permeating, poorly soluble compounds with low transporter activity) by grounding the prediction in the cell's energetic limitations. For MDCK (Madin-Darby canine kidney) cells, this threshold-normalized to the applied compound concentration (Cext)-was determined to be Pm×Cext = 10[-1.7] cm/s×µM. Methods: To derive this threshold, we conducted an extensive analysis of literature-reported efflux ratios (ERs) in MDCKII cells overexpressing efflux transporters (MDR1, BCRP, MRP2; 294 datapoints across 136 unique compounds). Concentration-dependent measurements for Amprenavir, Eletriptan, Loperamide, and Quinidine-chosen because these borderline compounds exhibited the highest Pm×Cext while still being actively effluxed-enabled the most accurate determination of the threshold. Literature ER values were re-evaluated through the experimental determination of reliable Pm values, as well as newly measured ER values with MDCK efflux assays. Conclusions: The results of these assays and the re-evaluation allowed us to reclassify all but three outliers (compounds with ER > 2.5 and log(Pm×Cext) > -1.7). In contrast, more than 60% of the compounds analyzed without significant ER values (123 compounds) fell above the threshold, in strong agreement with our theory of an energy limitation to active transport. This permeability threshold thus provides a simple and broadly applicable criterion to identify compounds for which active efflux is energetically not feasible and may serve as a practical tool for early drug discovery and optimization, pending further validation in practical applications.

This review meticulously examines the development, design, and pharmacological assessment of both well known antiviral and antihypertensive medications all time employing new chemical techniques and structure-based drug design to design and synthesize vital therapeutic entities such as aliskiren (renin inhibitor), captopril (a2-ACE-Inhibitor), dorzolamide (inhibitor of carbonic anhydrase) the review demonstrates initial steps regarding the significance of stereoselective synthesis, metal chelating pharmacophores, and rational molecular properties. More importantly, protease inhibitors (i.e., saquinavir, ritonavir, indinavir, amprenavir, etc.) and more contemporary agents (i.e., oseltamivir, nirmatrelvir/ritonavir (Paxlovid), etc.) were identified in the review and form a basis of advancement in antiviral therapy. Using the above-mentioned computational applications (i.e., molecular dockings, structure-activity relationships (SAR)), quantitative structure-activity relationships (QSAR modeling), and ADMET profile information, selectivity, safety, and therapeutic reliability of compounds toward antiviral activity and antihypertensive activity have improved further as outcomes of drug discovery research. Through the combinatorial application of computational drug design and experimental chemistry, researchers have significantly optimized the drug discovery process, minimized off-target effects, and expedited the pathway to develop therapeutically useful medications. In conclusion, this review highlights the transformative power of interdisciplinary approaches, including structure-based design, computational modeling, and the emerging approaches of drug repurposing and virtual screening. This hybrid approach provides a greater opportunity to improve drug therapy for cardiovascular disease and viral infections, and represents the rewards of collaboration.

Gastroesophageal reflux disease (GERD) is associated with inflammatory and neoplastic changes in the esophageal epithelium. Despite widespread PPI use, esophageal adenocarcinoma (EAC) incidence continues to rise, implicating non-acidic reflux components such as pepsin in disease progression. We performed transcriptomic profiling to assess pepsin-induced changes and the protective effect of amprenavir in vitro. Het-1A (normal) and BAR-T (Barrett's) cells (n = 3) were treated at pH 7.0 with pepsin and/or 10 μM amprenavir for 1 h. RNA-seq identified DEGs (FDR ≤ 0.05, |log2FC| ≥ 0.375), and Ingenuity Pathway Analysis revealed enriched pathways. Pepsin exposure altered mitochondrial function, oxidative phosphorylation, epithelial integrity, signaling, and inflammatory pathways in both cell lines. Amprenavir attenuated these transcriptomic perturbations, preserving mitochondrial and stress-response pathways. Notably, BAR-T cells exhibited heightened activation of wound-healing and epithelial repair pathways, whereas Het-1A cells showed greater mitochondrial and systemic stress pathway alterations. Pepsin drives transcriptomic dysregulation in esophageal epithelial cells under non-acidic conditions, and amprenavir shows potential to counteract peptic injury. Further studies are needed to validate these findings and explore amprenavir's therapeutic utility in GERD management and EAC prevention.

The current analytical work focuses on the development of a bioanalytical method for anti-HIV drugs, nevirapine, fosamprenavir calcium, and its metabolite amprenavir. Fosamprenavir is a prodrug of amprenavir, digested by cellular phosphatases in vivo. Rat plasma was isolated from the collected rat blood by centrifugation and used for preparing stock solutions. A reverse-phase column (ODS C-18) was used for the development of the method and validation. The bioanalytical method was developed using the isocratic mode. The solvent system used is Methanol: Water: ACN (800 mL: 200 mL: 50 mL). The method was run at a flow rate of 0.5 mL/min, and 258 nm was the detection wavelength. The developed method showed retention at 8.705 min for nevirapine, 11.923 min for fosamprenavir calcium, and 14.391 min for amprenavir. The calibration curve was linear with a correlation coefficient (r[2]) of 0.9916 for nevirapine, 0.9909 for fosamprenavir calcium, and 0.9879 min for amprenavir. The RSD of the accuracy, precision, and stability study of the method was found to be less than 2 % and was found to be acceptable. The method is reliable and does not show any kind of interference due to the plasma sample. Thus, the results support that a reliable, reproducible, and efficient method was developed, and validation was carried out for the estimation of the drug nevirapine, fosamprenavir, and its metabolite amprenavir in rat plasma samples.

The current work aims to develop a reliable and robust RP-HPLC method for analyzing Fosamprenavir and its potential impurities, including isomer, amino, propyl, nitro, and Amprenavir. The method used a Zobrax C18 column with a mobile phase of 0.1% V/V orthophosphoric acid in water and acetonitrile in gradient elution at a flow rate of 1 mL/min to accomplish efficient separation with detection at 264 nm and column temperature of 30 ± 2[0]C. A diluent with a 1:1 water-to-acetonitrile ratio was used to prepare standard and sample solutions. The developed approach was validated as per ICH Q2(R1) guidelines. Fosamprenavir, Amino, Propyl, Isomer, Nitro impurities, and Amprenavir impurities were eluted at retention time (RT) of 5.3 min, 2.3 min, 4.3 min, 4.7 min, 8.1 min and 8.6 min correspondingly with good resolution within a 10-minute run time. Method validation confirmed system suitability, linearity (R[2] = 0.999), good sensitivity (LOD/LOQ), specificity, precision (% RSD: 0.5-1.7), accuracy (% recovery: 90.9-104.3%), and robustness. The optimized approach excelled existing methods in lower retention time, run time, sensitivity, and linearity for all potential impurities, making it a novel and trustworthy method for monitoring Fosamprenavir drug quality and assessing stated impurities. The established method allows precise measurement of Fosamprenavir and related substances, supporting drug safety and regulatory compliance.

The combined effort of medical, scientific and pharmaceutical research has transformed the HIV infection from an inevitably fatal disease into a manageable chronic ailment. Especially after the introduction of protease (PR) inhibitors the efficacy of the treatment regimen has been increased. However, the development of mutant PR enzymes is a major concern. The present study focuses on to understand the alterations in wild-type and mutants (D30N and L76V) PR structure by interaction of first-generation PR inhibitors amprenavir, indinavir, nelfinavir, ritonavir and saquinavir. This information is extracted using molecular docking and MD simulation. Further, post-MD analysis such as RMSD, RMSF, PCA, DCCM, intermolecular interactions and binding free energy values calculation has been done. In comparison to L76V, D30N mutation affected the stable binding of the most of the studied PR inhibitors. The positively charged NH2 and thiazole group of amprenavir and nelfinavir is repelled by NH2 of Asn30 in D30N mutant. This repulsion pushes the drugs towards flap domain and causes flap opening. The MD simulation reveals that amprenavir, nelfinavir and saquinavir drugs have good compatibility with L76V mutant PR enzyme whereas indinavir was affected by the same. L76V mutant does not form any direct interaction with the first line drugs; hence the drugs forming strong interactions with the active site, flap domain and substrate binding region residues are quite enough to manage the resistance provided by L76V mutant. The results provided the insight into the drug-resistant or drug-susceptibility mechanism of L76V and D30N mutation against first-generation PR inhibitors.

Visceral leishmaniasis (VL) is an opportunistic infection in HIV patients with higher relapse and mortality rate. The number of HIV-VL patients is comparatively higher in areas where both infections are endemic. However, the conventional chemotherapeutic agents have limited success due to drug toxicity, efficacy variance and overall cost of treatment. Therefore, it is crucial to discover and develop newer potent antileishmanial agents for successful eradication of the disease. Our previous report, for the first time showed the leishminicidal effect of amprenavir (APV) mediated by inhibition of L.donovani Topoisomerase I (LdTopILS). So, we intended to demonstrate the effect of APV in combination with ritonavir (RTV). The present study revealed that the complete catalytic inhibition of LdTopILS by APV (10 μM) in combination with RTV (5 μM), compare to APV (20 μM) as previously reported (Roy et al., 2021). Moreover, APV (5 μM) in combination with RTV (4 μM) exhibited promastigote inhibition with IC50 values of 2.4 ± 0.6 μM at 12 h and 1.6 ± 0.7 μM at 24 h, respectively. The study was extended in animal model where the in vivo antileishmanial efficacy of APV-RTV in BALB/c mice demonstrated that treatment of APV in combination with RTV led to significant splenic and hepatic protection as compared to single dose of APV. Moreover, the antileishmanial activity of APV in combination with RTV was exerted via inhibition of LdTopILS at much lower concentration of APV and this inhibition of the enzyme induced programmed cell death in Leishmania parasites by generating oxidative stress within the cells. From the in vitro, ex vivo and in vivo studies, it was indicated that lower dose of APV in combination with RTV elevated the effective killing of the parasites as compared to the single higher dose of APV. Thus, the current study highlights repurposing of available protease inhibitors in combination, which might be exploited further for the therapeutic development against VL as well as HIV-VL co-infection.

The landscape of clinical management for metastatic melanoma (MM) and other solid tumors has been modernized by the advent of immune checkpoint inhibitors (ICI), including programmed cell death-1 (PD-1), programmed cell death-ligand 1 (PD-L1), and cytotoxic T lymphocyte antigen 4 (CTLA-4) inhibitors. While these agents demonstrate efficacy in suppressing tumor growth, they also lead to immune-related adverse events (irAEs), resulting in the exacerbation of autoimmune diseases such as rheumatoid arthritis (RA), ulcerative colitis (UC), and Crohn's disease (CD). The immune checkpoint inhibitors offer promising advancements in the treatment of melanoma and other cancers, but they also present significant challenges related to irAEs and autoimmune diseases. Ongoing research is crucial to better understand these challenges and develop strategies for mitigating adverse effects while maximizing therapeutic benefits. In this manuscript, we addressed this challenge using network-based approaches by constructing and analyzing the molecular and signaling networks associated with tumor-immune crosstalk. Our analysis revealed that IL6 is the key regulator responsible for irAEs during ICI therapies. Furthermore, we conducted an integrative network and molecular-level analysis, including virtual screening, of drug libraries, such as the Collection of Open Natural Products (COCONUT) and the Zinc15 FDA-approved library, to identify potential IL6 inhibitors. Subsequently, the compound amprenavir was identified as the best molecule that may disrupt essential interactions between IL6 and IL6R, which are responsible for initiating the signaling cascades underlying irAEs in ICI therapies.

Background & objectives Despite advancements in antiretroviral therapy, drug-resistant strains of HIV (human immunodeficiency virus) remain a global health concern. Natural compounds from medicinal plants offer a promising avenue for developing new HIV-1 PR (protease) inhibitors. This study aimed to explore the potential of compounds derived from Calotropis procera, a medicinal plant, as inhibitors of HIV-1 PR. Methods This in silico study utilized natural compound information and the crystal structure of HIV-1 PR. Molecular docking of 17 steroidal cardenolides from Calotropis procera against HIV-1 PR was performed using AutoDock 4.2 to identify compounds with higher antiviral potential. A dynamic simulation study was performed to provide insights into the stability, binding dynamics, and potential efficacy of the top potential antiviral compound as an HIV-1 therapeutic. Results We found that all tested cardenolides had higher binding affinities than Amprenavir, indicating their potential as potent HIV-1 PR inhibitors. Voruscharin and uscharidin displayed the strongest interactions, forming hydrogen bonds and hydrophobic interactions with HIV-1 PR. Voruscharin showed improved stability with lower RMSD (Root Mean Square Deviation) values and reduced fluctuations in binding site residues but increased flexibility in certain regions. The radius of gyration analysis confirmed a stable binding pose between HIV-1 PR and Voruscharin. Interpretation & conclusions These findings suggest that Calotropis procera could potentially be a source of compounds for developing novel HIV-1 PR inhibitors, contributing to the efforts to combat HIV. Further studies and clinical trials are needed to evaluate the safety and efficacy of these compounds as potential drug candidates for the treatment of HIV-1 infection.

Multidrug resistance is a paramount impediment to successful treatment of most hospital acquired bacterial infections. A plethora of bacterial genera exhibit differential levels of resistance to the existing antibiotics. Prevalent Uropathogenic Escherichia coli or UPEC conduce high mortality among them. Multi-Drug Resistant bacterial strains utilize precise mechanisms to bypass effects of antibiotics. This is probably due to their familiar genomic origin. In this article drug repositioning method have been utilised to target 23 enzymes of UPEC strains viz. CFT073, 536 and UTI89. 3-D drug binding motifs have been predicted using SPRITE and ASSAM servers that compare amino acid side chain similarities. From the hit results anti-viral drugs have been considered for their uniqueness and specificity. Out of 14 anti-viral drugs 3 anti-HIV drugs viz. Amprenavir, Darunavir and Saquinavir have selected for maximum binding score or drug targetability. Finally, active sites of the enzymes were analyzed using GASS-WEB for eloquent drug interference. Further analyses with the active sites of all the enzymes showed that the three selected anti-HIV drugs were very much potent to inhibit their active sites. Combination or sole application of Amprenavir, Darunavir and Saquinavir to MDR-UPEC infections may leads to cure and inhibition of mortality.