Polyvalent mpox mRNA vaccines elicit robust immune responses and confer potent protection against vaccinia virus.

The 2022 mpox outbreak led the World Health Organization (WHO) to declare it a public health emergency of international concern (PHEIC). There is a need to develop more effective and safer mpox virus (MPXV)-specific vaccines in response to the mpox epidemic. The mRNA vaccine is a promising platform to protect against MPXV infection. In this study, we construct two bivalent MPXV mRNA vaccines, designated LBA (B6R-A29L) and LAM (A35R-M1R), and a quadrivalent mRNA vaccine, LBAAM (B6R-A35R-A29L-M1R). The immunogenicity and protective efficacy of these vaccines alone or in combination were evaluated in a lethal mouse model. All mRNA vaccine candidates could elicit potential antigen-specific humoral and cellular immune responses and provide protection against vaccinia virus (VACV) infection. The protective effect of the combination of two bivalent mRNA vaccines and the quadrivalent vaccine was superior to that of the individual bivalent mRNA vaccine. Our study provides valuable insights for the development of more efficient and safer mRNA vaccines against mpox.

Enhancing nicotinamide N-methyltransferase bisubstrate inhibitor activity through 7-deazaadenosine and linker modifications.

Nicotinamide N-methyltransferase (NNMT) catalyzes the transfer of a methyl group from S-adenosylmethionine (SAM) to nicotinamide (NAM) and other pyridine-related compounds and is involved in various metabolic processes in the human body. In addition, abnormal expression of NNMT occurs under various pathological conditions such as cancer, diabetes, metabolic disorders, and neurodegenerative diseases, making it a promising drug target worthy of in-depth research. Small-molecule NNMT inhibitors with high potency and selectivity are necessary chemical tools to test biological hypotheses and potential therapies. In this study, we developed a series of highly active NNMT inhibitors by modifying N7 position of adenine. Among them, compound 3-12 (IC50 = 47.9 ± 0.6 nM) exhibited potent inhibitory activity and also had an excellent selectivity profile over a panel of human methyltransferases. We showed that the N7 position of adenine in the NNMT bisubstrate inhibitor was a modifiable site, thus offering insights into the development of NNMT inhibitors.

A small-molecule Skp1 inhibitor elicits cell death by p53-dependent mechanism.

Skp1 overexpression promotes tumor growth, whereas reduced Skp1 activity is also linked with genomic instability and neoplastic transformation. This highlights the need to gain better understanding of Skp1 biology in cancer settings. To this context, potent and cellularly active small-molecule Skp1 inhibitors may be of great value. Using a hypothesis-driven, structure-guided approach, we herein identify Z0933M as a potent Skp1 inhibitor with KD ∼0.054 µM. Z0933M occupies a hydrophobic hotspot (P1) - encompassing an aromatic cage of two phenylalanines (F101 and F139) - alongside C-terminal extension of Skp1 and, thus, hampers its ability to interact with F-box proteins, a prerequisite step to constitute intact and active SCF E3 ligase(s) complexes. In cellulo, Z0933M disrupted SCF E3 ligase(s) functioning, recapitulated previously reported effects of Skp1-reduced activity, and elicited cell death by a p53-dependent mechanism. We propose Z0933M as valuable tool for future efforts toward probing Skp1 cancer biology, with implications for cancer therapy.

Design, synthesis, and biological evaluation of novel 2'-methyl-2'-fluoro-6-methyl-7-alkynyl-7-deazapurine nucleoside analogs as anti-Zika virus agents.

Zika virus (ZIKV) is a mosquito-borne flavivirus and outbreaks of ZIKV have been reported in Africa, Americas and other parts of the world lately. The ZIKV epidemic has received extensive attention due to its ability to cause serious medical consequences and complications such as microcephaly and Guillain-Barre syndrome in recent years. Up to now, there are no specific treatments or vaccines available for ZIKV infection, which highlights the urgent need for developing new therapies. In this work, we designed and synthesized a series of novel 6-methyl-7-acetylenenyl-7-deazapurine nucleoside analogs as potential inhibitors of ZIKV replication. The biological activities against ZIKV replication were evaluated and the structure-activity relationship (SAR) was also studied. Among the compounds evaluated, nucleoside analog 38 (EC50 = 2.8 ± 0.8 µM, EC90 = 6.8 ± 2.3 µM) showed the most potent anti-ZIKV activity with low cytotoxicity (CC50 = 54.1 ± 6.9 µM) in an A549 based cellular model. The inhibitory activity of 38 was about 5 times more potent than the positive control NITD008. Notably, 38 showed similar inhibition potency against different ZIKV strains (ZG-01 and MR766) in a variety of host cell types including SNB19, A549, Huh7, Vero. In addition, 38 (Kd = 1.87 µM) has a stronger affinity to ZIKV RNA-dependent RNA polymerase (RdRp) protein than NITD008 (Kd = 3.43 µM) in the non-phosphorylation assay. These results indicated that compound 38 may serve as a promising candidate in future anti-ZIKV drug discovery.

Organocatalytic Asymmetric Arylation of p-Quinone Phosphonates: A Green Access to Biaryl Monophosphorus Ligands.

Herein, we report a highly efficient organocatalytic asymmetric synthesis of axially chiral biaryl phosphonates with p-quinone phosphonates and 2-naphthols via CPA-catalyzed asymmetric arylations. A series of chiral biaryl monophosphonates were obtained in excellent yields and enantioselectivities (up to 99% yield and 95% ee). This reaction could be operated at a gram scale with a low catalyst loading (0.5 mol %). Remarkably, our approach provides a green and ready access to chiral biaryl monophosphorus ligands. Compound 4ca was successfully converted to novel chiral biaryl monophosphorus ligands 7a, 7b, and 8 with high enantioselectivities in three steps.

The development of HEC-866 and its analogues for the treatment of idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease with a typical survival time between three to five years. Two drugs, pirfenidone and nintedanib have been approved for the treatment of IPF, but they have limited efficacy. Thus, the development of new drugs to treat IPF is an urgent medical need. In this paper we report the discovery of a series of orally active pyrimidin-4(3H)-one analogs which exhibit potent activity in in vitro assays. Among them, HEC-866 showed promising efficacy in rat IPF models. Since HEC-866 also had good oral bioavailability, a long half-life and favorable long-term safety profiles, it was selected for further clinical evaluation.

Design, Synthesis, and Resolution of Spirocyclic Bisoxindole-Based C2-Symmetric Diols.

A new type of spirocyclic bisoxindole-based C2-symmetric diols (SBIDOLs) was designed and synthesized. A series of racemic SBIDOL derivatives (6a-6g) were readily synthesized from commercially available 2-halo-5-methoxyanilines 1 (X = Cl or Br) through N-mono alkylation, acylation, oxidation, double intramolecular Friedel-Crafts reaction, and demethylation reactions. The optical resolution of racemic 6b was achieved via fractional crystallization of their bis-l-menthoxycarboxylates. Further modifications of SBIDOLs were investigated, leading to 5,5'-diaryl SBIDOL derivatives (11a and 11b) through Pd-catalyzed Suzuki coupling and DM-SBIDOL 12 by Pd/C-catalyzed hydrogenative dechlorination reactions.

Synthesis and computational studies of highly selective inhibitors of human recombinant tissue non-specific alkaline phosphatase (h-TNAP): A therapeutic target against vascular calcification.

In this study, we have discovered small druglike molecules as selective inhibitors of human tissue-nonspecific alkaline phosphatase (h-TNAP), an enzyme critical for the regulation of extracellular matrix calcification. The upregulation of h-TNAP is associated with various pathologies particularly the vascular calcification (VC). Selective inhibition of h-TNAP over h-NPP1 may serve as a useful therapeutic strategy against vascular calcification. A series of novel triazolyl pyrazole derivatives (10a-y) in which thiol bearing triazole moiety as the zinc binding functional group was introduced to a pyrazole based pharmacophore was synthesized and evaluated as potent and selective inhibitors of h-TNAP over h-NPP1. The biological screening against h-TNAP, h-IAP, h-NPP1 and h-NPP3 showed that many of the synthesized compounds are selective inhibitors of TNAP. Particularly, the compounds 10a-h, 10j, 10m-q, 10u, 10w and 10x displayed high potency and complete selectivity towards h-TNAP over h-NPP1. Compound 10q emerged as a highly potent inhibitor (IC50 = 0.16 µM or 160 nM) against h-TNAP with 127-fold increased inhibition compared to levamisole. On the other hand, compound 10e was found to be most selective inhibitor against the tested APs and NPPs (IC50 = 1.59 ± 0.36 µM). Binding sites architecture analysis, molecular-docking and molecular dynamics simulations (MDS), revealed the basis for h-TNAP and h-IAP ligand selectivity as well as selectivity towards h-TNAP over h-NPP1. These newly discovered inhibitors are believed to represent valuable lead structures to further streamline the generation of candidate compounds to target VC.

Quinoline Derivatives Kill Mycobacterium tuberculosis by Activating Glutamate Kinase.

There is a great need for identification and development of new anti-tuberculosis drugs with novel targets. Recent drug-discovery efforts typically focus on identifying inhibitors but not activators that perturb metabolic enzymes' functions as a means to kill Mycobacterium tuberculosis (Mtb). Here, we describe a class of quinoline compounds, Z0933/Z0930, which kill Mtb by acting as activators of glutamate kinase (GK), a previously untargeted enzyme catalyzing the first step of proline biosynthesis. We further show that Z0933/Z0930 augment proline production and induce Mtb killing via proline-derived redox imbalance and production of reactive oxygen species. This work highlights the effectiveness of gain-of-function probes against Mtb and provides a framework for the discovery of next-generation allosteric activators of GK.

BubR1 phosphorylates CENP-E as a switch enabling the transition from lateral association to end-on capture of spindle microtubules.

Error-free mitosis depends on accurate chromosome attachment to spindle microtubules, powered congression of those chromosomes, their segregation in anaphase, and assembly of a spindle midzone at mitotic exit. The centromere-associated kinesin motor CENP-E, whose binding partner is BubR1, has been implicated in congression of misaligned chromosomes and the transition from lateral kinetochore-microtubule association to end-on capture. Although previously proposed to be a pseudokinase, here we report the structure of the kinase domain of Drosophila melanogaster BubR1, revealing its folding into a conformation predicted to be catalytically active. BubR1 is shown to be a bona fide kinase whose phosphorylation of CENP-E switches it from a laterally attached microtubule motor to a plus-end microtubule tip tracker. Computational modeling is used to identify bubristatin as a selective BubR1 kinase antagonist that targets the αN1 helix of N-terminal extension and αC helix of the BubR1 kinase domain. Inhibition of CENP-E phosphorylation is shown to prevent proper microtubule capture at kinetochores and, surprisingly, proper assembly of the central spindle at mitotic exit. Thus, BubR1-mediated CENP-E phosphorylation produces a temporal switch that enables transition from lateral to end-on microtubule capture and organization of microtubules into stable midzone arrays.

Rational design, synthesis and biological evaluation of ubiquinone derivatives as IDO1 inhibitors.

Indoleamine 2,3-dioxygenase 1 (IDO1) is an attractive therapeutic target for the treatment of cancer, chronic viral infections and neurological disorders characterized by pathological immune stimulation. Herein, a series of known metal-chelating ubiquinone derivatives were designed, synthesized and evaluated for the IDO1 inhibiting activities. The docking studies showed that the compounds 11, 16, 18 and coenzyme-Q1 exhibited different binding modes to IDO1 protein. Among these compounds, the most active compound is 16d with an IC50 of 0.13 µM in enzymatic assay. The results reveal that a possible halogen bonding interaction between the bromine atom (3-Br) and Cys129 significantly enhances the inhibition activity against IDO1. This study provides structural insights of the interactions between ubiquinone analogues and IDO1 protein for the further modification and optimization.

CXCL13/CXCR5 signaling axis in cancer.

The tumor microenvironment comprises stromal and tumor cells which interact with each other through complex cross-talks that are mediated by a variety of growth factors, cytokines, and chemokines. The chemokine ligand 13 (CXCL13) and its chemokine receptor 5 (CXCR5) are among the key chemotactic factors which play crucial roles in deriving cancer cell biology. CXCL13/CXCR5 signaling axis makes pivotal contributions to the development and progression of several human cancers. In this review, we discuss how CXCL13/CXCR5 signaling modulates cancer cell ability to grow, proliferate, invade, and metastasize. Furthermore, we also discuss the preliminary evidence on context-dependent functioning of this axis within the tumor-immune microenvironment, thus, highlighting its potential dichotomy with respect to anticancer immunity and cancer immune-evasion mechanisms. At the end, we briefly shed light on the therapeutic potential or implications of targeting CXCL13/CXCR5 axis within the tumor microenvironment.

Ligand binding and heterodimerization with retinoid X receptor α (RXRα) induce farnesoid X receptor (FXR) conformational changes affecting coactivator binding.

Nuclear receptor farnesoid X receptor (FXR) functions as the major bile acid sensor coordinating cholesterol metabolism, lipid homeostasis, and absorption of dietary fats and vitamins. Because of its central role in metabolism, FXR represents an important drug target to manage metabolic and other diseases, such as primary biliary cirrhosis and nonalcoholic steatohepatitis. FXR and nuclear receptor retinoid X receptor α (RXRα) form a heterodimer that controls the expression of numerous downstream genes. To date, the structural basis and functional consequences of the FXR/RXR heterodimer interaction have remained unclear. Herein, we present the crystal structures of the heterodimeric complex formed between the ligand-binding domains of human FXR and RXRα. We show that both FXR and RXR bind to the transcriptional coregulator steroid receptor coactivator 1 with higher affinity when they are part of the heterodimer complex than when they are in their respective monomeric states. Furthermore, structural comparisons of the FXR/RXRα heterodimers and the FXR monomers bound with different ligands indicated that both heterodimerization and ligand binding induce conformational changes in the C terminus of helix 11 in FXR that affect the stability of the coactivator binding surface and the coactivator binding in FXR. In summary, our findings shed light on the allosteric signal transduction in the FXR/RXR heterodimer, which may be utilized for future drug development targeting FXR.

Mechanistic insights into peptide and ligand binding of the ATAD2-bromodomain via atomistic simulations disclosing a role of induced fit and conformational selection.

ATAD2 has emerged as a promising bromodomain (BRD)-containing therapeutic drug target in multiple human cancers. However, recent druggability assessment studies predicted ATAD2's BRD as a target 'difficult to drug' because its binding pocket possesses structural features that are unfeasible for ligand binding. Here, by using all-atom molecular dynamics simulations and an advanced metadynamics method, we demonstrate a dynamic view of the binding pocket features which can hardly be obtained from the "static" crystal data. The most important features disclosed from our simulation data, include: (1) a distinct 'open-to-closed' conformational switch of the ZA loop region in the context of peptide or ligand binding, akin to the induced fit mechanism of molecular recognition, (2) a dynamic equilibrium of the BC loop "in" and "out" conformations, highlighting a role in the conformational selection mechanism for ligand binding, and (3) a new binding region identified distal to the histone-binding pocket that might have implications in bromodomain biology and in inhibitor development. Moreover, based on our simulation results, we propose a model for an "auto-regulatory" mechanism of ATAD2's BRD for histone binding. Overall, the results of this study will not only have implications in bromodomain biology but also provide a theoretical basis for the discovery of new ATAD2's BRD inhibitors.

3-((R)-4-(((R)-6-(2-Bromo-4-fluorophenyl)-5-(ethoxycarbonyl)-2-(thiazol-2-yl)-3,6-dihydropyrimidin-4-yl)methyl)morpholin-2-yl)propanoic Acid (HEC72702), a Novel Hepatitis B Virus Capsid Inhibitor Based on Clinical Candidate GLS4.

The inhibition of hepatitis B virus (HBV) capsid assembly is a novel strategy for the development of chronic hepatitis B (CHB) therapeutics. On the basis of the preclinical properties and clinical results of GLS4, we carried out further investigation to seek a better candidate compound with appropriate anti-HBV potency, reduced hERG activity, decreased CYP enzyme induction, and improved pharmacokinetic (PK) properties. To this end, we have successfully found that morpholine carboxyl analogues with comparable anti-HBV activities to that of GLS4 showed decreased hERG activities, but they displayed strong CYP3A4 induction in a concentration-dependent manner, except for morpholine propionic acid analogues. After several rounds of modification, compound 58 (HEC72702), which had an (R)-morpholine-2-propionic acid at the C6 position of its dihydropyrimidine core ring, was found to display no induction of the CYP1A2, CYP3A4, or CYP2B6 enzyme at the high concentration of 10 µM. In particular, it demonstrated a good systemic exposure and high oral bioavailability and achieved a viral-load reduction greater than 2 log in a hydrodynamic-injected (HDI) HBV mouse model and has now been selected for further development.

ATAD2 in cancer: a pharmacologically challenging but tractable target.

INTRODUCTION: ATAD2 protein is an emerging oncogene that has strongly been linked to the etiology of multiple advanced human cancers. Therapeutically, despite the fact that genetic suppression/knockdown studies have validated it as a compelling drug target for future therapeutic development, recent druggability assessment data suggest that direct targeting of ATAD2's bromodomain (BRD) may be a very challenging task. ATAD2's BRD has been predicted as a 'difficult to drug' or 'least druggable' target due to the concern that its binding pocket, and the areas around it, seem to be unfeasible for ligand binding. Areas covered: In this review, after shedding light on the multifaceted roles of ATAD2 in normal physiology as well as in cancer-etiology, we discuss technical challenges rendered by ATAD2's BRD active site and the recent drug discovery efforts to find small molecule inhibitors against it. Expert opinion: The identification of a novel low-nanomolar semi-permeable chemical probe against ATAD2's BRD by recent drug discovery campaign has demonstrated it to be a pharmacologically tractable target. Nevertheless, the development of high quality bioavailable inhibitors against ATAD2 is still a pending task. Moreover, ATAD2 may also potentially be utilized as a promising target for future development of RNAi-based therapy to treat cancers.

Design, synthesis, and biological evaluation of novel 7-deazapurine nucleoside derivatives as potential anti-dengue virus agents.

Dengue fever, caused by four distinct serotypes of dengue virus (DENV-1 to -4), has become the fastest spreading human infectious disease in recent years. Despite extensive efforts, there is no specific antiviral treatment approved for dengue until now. Nucleoside inhibitors represent an actively pursued area to develop small-molecule anti-dengue virus agents. In this study, we designed and synthesized a series of 7-deazapurine nucleoside derivatives and evaluated their anti-DENV activity. Our design strategy and structure activity relationship studies revealed 6e as the most potent inhibitor (EC50 = 2.081 ± 1.102 µM) of DENV replication. 6e suppressed RNA levels and DENV E protein expression, without causing any apparent cytotoxicity in A549 and HepG2 cells (CC50 = 150.06 ± 11.42 µM, SI = 72.11 in A549 cells, and CC50 = 146.47 ± 11.05 µM and SI = 63.7 in HepG2 cells). In addition, 6e showed similar inhibition potency against four serotypes of DENV, suggesting that it restrains some evolutionarily conserved targets essential for DENV replication. We conceive that 6e may serve as a promising lead compound for anti-DENV drug development.

Discovery of hepatitis B virus capsid assembly inhibitors leading to a heteroaryldihydropyrimidine based clinical candidate (GLS4).

Inhibition of hepatitis B virus (HBV) capsid assembly is a novel strategy for the development of chronic hepatitis B (CHB) therapeutics. Herein we described our lead optimization studies including the synthesis, molecular docking studies and structure-activity relationship (SAR) studies of a series of novel heteroaryldihydropyrimidine (HAP) inhibitors of HBV capsid assembly inhibitors, and the discovery of a potent inhibitor of HBV capsid assembly of GLS4 (ethyl 4-[2-bromo-4-fluorophenyl]-6-[morpholino-methyl]-2-[2-thiazolyl]-1,4-dihydro-pyrimidine-5-carboxylate) which is now in clinical phase 2. GLS4 demonstrated potent inhibitory activities in HBV HepG2.2.15 cell assay with an EC50 value of 1nM, and it also exhibited high potency against various drug-resistant HBV viral strains with EC50 values in the range of 10-20nM, more potent than the typical HBV polymerase inhibitors such as lamivudine, telbivudine, and entecavir. Pharmacokinetic profiles of GLS4 were favorable and safety evaluation including acute toxicity and repeated toxicity study indicated that GLS4 was safe enough to support clinical experiments in human.

Synthesis and biological evaluation of novel dasatinib analogues as potent DDR1 and DDR2 kinase inhibitors.

Novel dasatinib analogues as DDR1 and DDR2 inhibitors were designed and synthesized. The synthesized compounds were screened for DDR1 and DDR2 kinase inhibitory and cancer cell proliferation inhibitory activities. Some of the compounds showed the potent inhibitory activities against both DDR1 and DDR2, as well as anticancer activity in low nanomolar range against K562 cell line; especially, compound 3j demonstrated significantly better inhibitory potency than the parental dasatinib against both DDRs and also demonstrated the potent inhibitory activity against K562 cell lines (IC50 values of 2.26±0.46 nm for DDR1, 7.04±2.90 nm for DDR2, and 0.125±0.017 nm for K562 cell line).

Synthesis and Evaluation of 2'-Deoxy-2'-Spirodiflurocyclopropyl Nucleoside Analogs.

The preparation of 2'-deoxy-2'-siprodifluorocyclopropany-lnucleoside analogs has been achieved from α-d-glucose in several steps. The key step in the synthesis was the introduction of the difluorocyclopropane through a difluorocarbene type reaction at the 2'-position. Then, a series of novel 2'-deoxy-2'-spirodifluorocyclopropanyl nucleoside analogs were synthesized using the Vorbrüggen method. All the synthesized nucleosides were characterized and subsequently evaluated against hepatitis C and influenza A virus strains in vitro.