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Optimization of compound 11L led to the identification of novel HIV capsid modulators, quinazolin-4-one-bearing phenylalanine derivatives, displaying potent antiviral activities against both HIV-1 and HIV-2. Notably, derivatives 12a2and 21a2showed significant improvements, with 2.5-fold over 11L and 7.3-fold over PF74 for HIV-1, and approximately 40-fold over PF74 for HIV-2. The X-ray co-crystal structures confirmed the multiple pocket occupation of 12a2and 21a2in the binding site. Mechanistic studies revealed a dual-stage inhibition profile, where the compounds disrupted capsid-host factor interactions at the early stage and promoted capsid misassembly at the late stage. Remarkably, 12a2and 21a2significantly promoted capsid misassembly, outperforming 11L, PF74, and LEN. The substitution of easily metabolized amide bond with quinolin-4-one marginally enhanced the stability of 12a2in human liver microsomes compared to controls. Overall, 12a2and 21a2highlight their potential as potent HIV capsid modulators, paving the way for future advancements in anti-HIV drug design.

In the current landscape of antiretroviral options, there remains an urgent need for novel non-nucleoside reverse transcriptase inhibitors (NNRTIs) with improved resistance profiles and safety properties. Herein, a series of novel tetrahydropyrido[4,3-d]pyrimidine derivatives were discovered utilizing the “escape from flatland” strategy. The most potent inhibitor 10cwas endowed with broad-spectrum antiviral activity and improved resistance profiles against NNRTI-resistant variants compared to efavirenz and etravirine. Molecular simulations were investigated to furnish insights into the biological results. Drug-likeness assessment showed that 10cexhibited desirable physicochemical properties and in vitro metabolic stability. Notably, lower cytochrome P450 inhibition and human ether-à-go-go-related gene blockade liability were observed for 10cthan those for etravirine and rilpivirine. Besides, 10cwas characterized by excellent in vivo safety properties without acute/subacute toxicity and organ pathological damage. Overall, our multiparameter optimization campaign led to the identification of 10cwith excellent antiviral activities and favorable drug-like profiles that could serve as an ideal drug candidate for further development.

Although non-nucleoside reverse transcriptase inhibitors (NNRTIs) exhibit potent anti-HIV-1 activity and play an important role in the active antiretroviral therapy of AIDS, the emergence of drug-resistant strains has seriously reduced their clinical efficacy. Here, we report a series of 2,4,5-trisubstituted pyrimidines as potent HIV-1 NNRTIs by exploiting the tolerant regions of the NNRTI binding pocket. Compounds 16band 16cwere demonstrated to have excellent activity (EC50= 3.14–22.1 nM) against wild-type and a panel of mutant HIV-1 strains, being much superior to that of etravirine (EC50= 3.53–52.2 nM). Molecular modeling studies were performed to illustrate the detailed interactions between RT and 16b, which shed light on the improvement of the drug resistance profiles. Moreover, 16bpossessed favorable pharmacokinetic (T1/2= 1.33 h, F= 31.8%) and safety profiles (LD50> 2000 mg/kg), making it a promising anti-HIV-1 drug candidate for further development.

HIV-1 capsid (CA) is an attractive target for its indispensable roles in the viral life cycle. We report the design, synthesis, and mechanistic study of a novel series of 2-piperazineone peptidomimetics as HIV capsid modulators by mimicking the structure of host factors binding to CA. F-Id-3owas the most potent compound from the synthesized series, with an anti-HIV-1 EC50value of 6.0 μM. However, this series of compounds showed a preference for HIV-2 inhibitory activity, in which Id-3orevealed an EC50value of 2.5 μM (anti-HIV-2 potency), an improvement over PF74. Interestingly, F-Id-3odid bind HIV-1 CA monomers and hexamers with comparable affinity, unlike PF74, consequently showing antiviral activity in the early and late stages of the HIV-1 lifecycle. Molecular dynamics simulations shed light on F-Id-3oand Id-3obinding modes within the HIV-1/2 CA protein and provide a possible explanation for the increased anti-HIV-2 potency. Metabolic stability assays in human plasma and human liver microsomes indicated that although F-Id-3ohas enhanced metabolic stability over PF74, further optimization is necessary. Moreover, we utilized computational prediction of drug-like properties and metabolic stability of F-Id-3oand PF74, which correlated well with experimentally derived metabolic stability, providing an efficient computational pipeline for future preselection based on metabolic stability prediction. Overall, the 2-piperazineone-bearing peptidomimetics are a promising new chemotype in the CA modulators class with considerable optimization potential.

Two new families of enantiomerically pure carbocyclic nucleoside analogues based on a cyclohexane moiety with five chiral centers and a fused cyclopropyl ring have been synthesized. A highly regio- and stereoselective synthetic approach for the modular construction of the functionalized bicyclo[4.1.0]heptyl azide intermediate 6has been established. Key steps to achieve this asymmetric synthesis involved highly diastereoselective allylic oxidation and hydroboration reactions. The first family of compounds, 1a,band 2, presents different natural nucleobases, whereas the second one 3a–ebears functionalized 1,2,3-triazoles. These derivatives have been tested as antiviral agents, and compound 3dhas shown to display moderate activity against coxsackie B4 virus.

Inspired by our previous studies to discover novel human immunodeficiency virus-1 (HIV-1) non-nucleoside reverse transcriptase inhibitors (NNRTIs) by targeting the tolerant region II of the NNRTIs binding pocket (NNIBP), a series of novel benzo[4,5]thieno[2,3-d]pyrimidine derivatives were designed through structure-based drug design as novel potent HIV-1 NNRTIs. The results showed that compound 16bwas the most active inhibitor, exhibiting 50% effective concentration (EC50) values from 0.021 µmol/L to 0.298 µmol/L against wild-type (WT) and a panel of NNRTIs-resistant HIV-1 strains. Moreover, 16bwas demonstrated with a significantly low 50% cytotoxicity concentration (CC50) value (>200 µmol/L) and high selectivity index (SI) values. In addition, 16byielded moderate reverse transcriptase (RT) enzyme inhibition with a 50% inhibition concentration (IC50) value of 0.183 µmol/L, which demonstrated that it acted as HIV-1 NNRTIs. The binding mode of 16bwith RT was also illustrated viamolecular docking. Overall, this work provided a novel lead compound for developing potent HIV-1 NNRTIs.

Results from recently completed studies suggested that the NH2-naphthyl-diarylpyrimidine JX-7displayed remarkable inhibitory activity against wild-type HIV-1 (EC50= 5 nM) and numerous clinically observed variants in MT-4 cells; however, its high cytotoxicity (CC50= 19 μM) precluded its further development as a clinical candidate. One approach we took to improve the safety involved replacing the naphthyl of JX-7with biphenyl to provide a series of novel NH2-biphenyl-DAPYs. Investigation of the structure–activity relationships (SARs) led to the identification of 4ab, a potent NNRTI with significantly reduced cytotoxicity (CC50= 120 μM), approximately 6-fold lower than JX-7, which maintained remarkable anti-HIV-1 activity against wild-type HIV-1 (EC50= 1.9 nM) and multiple mutant strains simultaneously. Also, 4abdisplayed weak CYP sensitivity, little inhibition of hERG, and no apparent in vivoacute toxicity. These promising results demonstrate that 4abcan be used as a drug candidate for HIV-1 therapy.

Here, we report the design, synthesis, structure–activity relationship studies, antiviral activity, enzyme inhibition, and druggability evaluation of dihydrofuro[3,4-d]pyrimidine derivatives as a potent class of HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs). Compounds 14b(EC50= 5.79–28.3 nM) and 16c(EC50= 2.85–18.0 nM) exhibited superior potency against a panel of HIV-1-resistant strains. Especially, for the changeling mutations F227L/V106A and K103N/Y181C, both compounds exhibited remarkably improved activity compared to those of etravirine and rilpivirine. Moreover, 14band 16cshowed moderate RT enzyme inhibition (IC50= 0.14–0.15 μM), which demonstrated that they acted as HIV-1 NNRTIs. Furthermore, 14band 16cexhibited favorable pharmacokinetic and safety properties, making them excellent leads for further development.

A series of novel heteroaromatic biphenyl-methyl-pyrimidine analogues were designed via hybridization of privileged structures of two HIV-1 inhibitors. Among them, compound 7acontaining 4-pyridinyl-phenyl and methyl-pyrimidine fragments revealed excellent wild-type HIV-1 inhibitory activity with low cytotoxicity. 7ahad favorable solubility and liver microsome stability; moreover, no apparent CYP enzymatic inhibitory activity or acute toxicity was observed. However, its inhibitory activity toward mutant strains and the pharmacokinetic (PK) profiles were still unsatisfactory. Further optimizations resulted in a highly potent compound 9dwithout methyl on the pyrimidine but a heteroaromatic dimethyl-biphenyl on the left rings of difluoro-pyridinyl-diarylpyrimidines (DAPYs). A broad-spectrum activity (EC50= 2.0–57 nM) of 9dagainst resistant strains was revealed. This compound also exhibited good solubility and safety profiles and a good PK profile with an oral bioavailability of 59% in rats. Collectively, these novel heteroaromatic dimethyl-biphenyl-DAPYs represent promising drug candidates for HIV clinical therapy.

To address the drug resistance, a series of novel boronic acid-containing diarylpyrimidine (DAPY) derivatives were designed viabioisosterism and scaffold-hopping strategies, and compound 10jyielded the most potent activity and turned out to be a single-digit nanomolar inhibitor towards the HIV-1 IIIB, L100I and K103N strains.

Enlightened by the available structural biology information, a novel series of dihydrothiopyrano[4,3-d]pyrimidine derivatives were rationally designed viascaffold hopping and molecular hybridization strategies. Notably, compound 20ayielded exceptionally potent antiviral activities (EC50= 4.44–54.5 nM) against various HIV-1 strains and improved resistance profiles (RF = 0.5–5.6) compared to etravirine and rilpivirine. Meanwhile, 20aexhibited reduced cytotoxicity (CC50= 284 μM) and higher SI values (SI = 5210–63992). Molecular dynamics simulations were performed to rationalize the distinct resistance profiles. Besides, 20adisplayed better solubility (sol. = 12.8 μg/mL) and no significant inhibition of the main CYP enzymes. Furthermore, 20awas characterized for prominent metabolic stability and in vivosafety properties. Most importantly, the hERG inhibition profile of 20a(IC50= 19.84 μM) was a remarkable improvement. Overall, 20apossesses huge potential to serve as a promising drug candidate due to its excellent potency, low toxicity, and favorable drug-like properties.

A series of novel heteroaromatic-difluoro-biphenyl-diarylpyrimidines were designed as non-nucleoside anti-HIV inhibitors targeting reverse transcriptase by a fragment-based replacement strategy with the purpose of improving the druggability. Hopping five- or six-membered heterocycle groups on the biphenyl moiety as bioisosterism for intrinsically cyanophenyl gave 23 derivatives. All of these compounds possessed excellent HIV-1 inhibitory activity in the nanomolar range. Among them, 12gwith a 4-pyridine group displayed excellent inhibitory activity toward WT and mutant HIV virus possessing significant selectivity. Moreover, this compound exhibited a decent improvement in druggability than etravirine and rilpivirine: (1) The hydrochloric acid salt of 12gexhibited significantly improved water solubility in different pH conditions. (2) 12gdid not show apparent CYP enzymatic inhibitory activity or acute toxicity. (3) Excellent oral bioavailability was also revealed (F= 126%, rats) in 12g. Collectively, these novel heteroaromatic-biphenyl-DAPYs represent promising drug candidates for HIV clinical therapy.

A series of novel S-DACO derivatives were designed by structure-based linker optimization strategy. Most compounds showed potential activities against HIV-1 with IC50values ranging from 7.55 μmol/L to 0.018 μmol/L. Preliminary SAR and molecular modeling of these novel congeners were investigated.

The HIV-1 CA protein has gained remarkable attention as a promising therapeutic target for the development of new antivirals, due to its pivotal roles in HIV-1 replication (structural and regulatory). Herein, we report the design and synthesis of three series of benzenesulfonamide-containing phenylalanine derivatives obtained by further structural modifications of PF-74to aid in the discovery of more potent and drug-like HIV-1 CA inhibitors. Structure–activity relationship studies of these compounds led to the identification of new phenylalanine derivatives with a piperazinone moiety, represented by compound 11l, which exhibited anti-HIV-1NL4–3activity 5.78-fold better than PF-74. Interestingly, 11lalso showed anti-HIV-2RODactivity (EC50= 31 nM), with almost 120 times increased potency over PF-74. However, due to the higher significance of HIV-1 as compared to HIV-2 for the human population, this manuscript focuses on the mechanism of action of our compounds in the context of HIV-1. SPR studies on representative compounds confirmed CA as the binding target. The action stage determination assay demonstrated that these inhibitors exhibited antiviral activities with a dual-stage inhibition profile. The early-stage inhibitory activity of compound 11lwas 6.25 times more potent as compared to PF-74but appeared to work via the accelerating capsid core assembly rather than stabilization. However, the mechanism by which they exert their antiviral activity in the late stage appears to be the same as PF-74with less infectious HIV-1 virions produced in their presence, as judged p24 content studies. MD simulations provided the key rationale for the promising antiviral potency of 11l. Additionally, 11lexhibited a modest increase in HLM and human plasma metabolic stabilities as compared to PF-74, as well as a moderately improved pharmacokinetic profile, favorable oral bioavailability, and no acute toxicity. These studies provide insights and serve as a starting point for subsequent medicinal chemistry efforts in optimizing these promising HIV inhibitors.

The development of efficacious NNRTIs for AIDS therapy commonly encountered the rapid generation of drug-resistant mutations, which becomes a major impediment to effective anti-HIV treatment. Using a structure-based bioisosterism strategy, a series of piperidine-substituted thiophene[2,3-d]pyrimidine derivatives were designed and synthesized. Compound 9ayielded the greatest potency, exhibiting significantly better anti-HIV-1 activity than ETRagainst all of the tested NNRTI-resistant HIV-1 strains. In addition, the phenotypic (cross)resistance of 9aand other NRTIs to the different selected HIV-1 strains was evaluated. As expected, no phenotypic cross-resistance against the NRTIs (AZT and PMPA) was observed with the mutant 9aresstrain. Furthermore, 9awas identified with improved solubility, lower CYP liability, and hERG inhibition. Remarkably, 9aexhibited optimal pharmacokinetic properties in rats (F= 37.06%) and safety in mice (LD50> 2000 mg/kg), which highlights 9aas a promising anti-HIV-1 drug candidate.

The novel sulfur-containing diarylbenzopyrimidines were designed by fragment-based strategy. The best compound 6emaintained the antiviral activity, low cytotoxicity and cLogP as a novel lead compound for further optimization.

Our previous efforts have led to the development of two potent NNRTIs, K-5a2 and 25a, exhibiting effective anti-HIV-1 potency and resistance profiles compared with etravirine. However, both inhibitors suffered from potent hERG inhibition and short half-life. In this article, with K-5a2 and etravirine as leads, series of novel fluorine-substituted diarylpyrimidine derivatives were designed via molecular hybridization and bioisosterism strategies. The results indicated 24bwas the most active inhibitor, exhibiting broad-spectrum activity (EC50= 3.60–21.5 nM) against resistant strains, significantly lower cytotoxicity (CC50= 155 μM), and reduced hERG inhibition (IC50> 30 μM). Crystallographic studies confirmed the binding of 24band the role of the fluorine atom, as well as optimal contacts of a nitrile group with the main-chain carbonyl group of H235. Furthermore, 24bshowed longer half-life and favorable safety properties. All the results demonstrated that 24bhas significant promise in circumventing drug resistance as an anti-HIV-1 candidate.

A series of nondimethylphenyl-diarylpyrimidines with much lower cytotoxicities than their dimethyl analogues were developed. Compound B13with a difluorobiphenyl moiety showed the highest antiviral activity against WT, mutant strains, and RT. The hydrochloride form of B13exhibited an improved water solubility of 5.6 μg/mL compared with ETR (≪1 μg/mL), better stability in human and rat liver microsomes, and a great oral bioavailability of 44%, making it promising as a drug candidate. In addition, no apparent toxicity was observed in the acute toxicity assay (2 g/kg) and HE staining.

Introducing novel strategies, concepts, and technologies that speed up drug discovery and the drug development cycle is of great importance both in the highly competitive pharmaceutical industry as well as in academia. This Perspective aims to present a “big-picture” overview of recent strategic innovations in medicinal chemistry and drug discovery.

To address drug resistance to HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs), a series of novel diarylpyrimidine (DAPY) derivatives targeting “tolerant region I” and “tolerant region II” of the NNRTIs binding pocket (NNIBP) were designed utilizing a structure-guided scaffold-hopping strategy. The dihydrofuro[3,4-d]pyrimidine derivatives 13c2and 13c4proved to be exceptionally potent against a wide range of HIV-1 strains carrying single NNRTI-resistant mutations (EC50= 0.9–8.4 nM), which were remarkably superior to that of etravirine (ETV). Meanwhile, both compounds exhibited comparable activities with ETV toward the virus with double mutations F227L+V106A and K103N+Y181C. Furthermore, the most active compound 13c2showed favorable pharmacokinetic properties with an oral bioavailability of 30.96% and a half-life of 11.1 h, which suggested that 13c2is worth further investigation as a novel NNRTI to circumvent drug resistance.

Diarylpyrimidine derivatives (DAPYs) exhibit robust anti-HIV-1 potency, although they have been compromised by E138K variant and severe side-effects and been suffering from poor water solubility. In the present work, hydrophilic morpholine or methylsulfonyl and sulfamide-substituted piperazine/piperidines were introduced into the right wing of DAPYs targeting the solvent-exposed tolerant region I. The anti-HIV-1 activities of 11c(EC50(WT)= 0.0035 μM, EC50(E138K)= 0.0075 μM) were the same as and 2-fold better than that of the lead etravirine against the wild-type and E138K mutant HIV-1, respectively, with a relative low cytotoxicity (CC50≥ 173 μM). Further test showed a significant improvement in the water solubility of 11c. Besides, 11cdisplayed no significant inhibition on main cytochrome P450 enzymes and exhibited no acute/subacute toxicities at doses of 2000 mg·kg–1/50 mg·kg–1in mice. Taken together, we consider that 11cis a promising lead for further structural optimization.

Background: Hydrazide-hydrazones constitute an important class of compounds for new drug development. In this study, a series of 39 new acylhydrazones (3-41), derived from (2S)-3-methyl- 2-[[(4-methylphenyl)sulfonyl]amino]butanoic acid hydrazide were synthesized with further aim to achieve biologically active acylhydrazones carrying an amino acid side chain. Methods: Compounds 3-41 were synthesized by microwave-assisted method. All synthesized compounds have been tested for their anti-HIV activity compound 21 was subjected to a new set of 2DNMR analysis for the characterization of the isomers in solution and determination of its 3D structure. Results: The IC50 values for compounds 2-40 were found between >125-10.90 µg/ml against HIV- 1(IIIB) and HIV-2(ROD) strains in MT-4 cells. Compounds 3, 6, 10, 12, 23, 24, 27, 32, and 37 with CC50 values between 10.90-14.50 µg/ml were selected to evaluate for their antileukemia activity. IC50 values for these mentioned compounds were found as >100μM on human chronic myelogenous leukemia, K562 cell line. Conclusion: Some compounds with IC50 values between 10.90-14.50 μg/ml will be of benefit in the development of novel leads.

Graphene quantum dots (GQD) are the next generation of nanomaterials with great potential in drug delivery and target-specific HIV inhibition. In this study we investigated the antiviral activity of graphene based nanomaterials by using water-soluble GQD synthesized from multiwalled carbon nanotubes (MWCNT) through prolonged acidic oxidation and exfoliation and compared their anti-HIV activity with that exerted by reverse transcriptase inhibitors (RTI) conjugated with the same nanomaterial. The antiretroviral agents chosen in this study, CHI499 and CDF119, belong to the class of non-nucleoside reverse transcriptase inhibitors (NNRTI). From this study emerged the RTI-conjugated compound GQD-CHI499 as a good potential candidate for HIV treatment, showing an IC50of 0.09 μg/mL and an EC50value in cell of 0.066 μg/mL. The target of action in the replicative cycle of HIV of the drug conjugated samples GQD-CHI499 and GQD-CDF119 was also investigated by a time of addition (TOA) method, showing for both conjugated samples a mechanism of action similar to that exerted by NNRTI drugs.