Antimalarial activities of benzothiazole analogs: A systematic review.

BACKGROUND: Benzothiazole derivatives have been reported to possess a wide range of biological activities, including antimalarial activity. This systematic review aims to summarize and evaluate the antimalarial activities of benzothiazole analogs. METHODS: We conducted an electronic search using nine databases in October 2017 and subsequently updated in September 2022. We included all original in vitro and in vivo studies that documented the antimalarial activities of compounds containing benzothiazole analogs with no restriction. The risk of bias of each included study was assessed by ToxRTool. RESULTS: Twenty-eight articles were included in our study, which are in vitro, in vivo, or both. Of these, 232 substances were identified to have potent antiplasmodial activity against various strains of the malaria parasite. Benzothiazole analogs show different antimalarial mechanisms, including inhibition of Plasmodium falciparum enzymes in in vitro studies and inhibition of blood parasites in in vivo studies. CONCLUSIONS: Benzothiazole derivatives are promising substances for treating malaria. The structure-activity relationship studies suggest that the substitution pattern of the benzothiazole scaffold plays a crucial role in determining the antimalarial activity of the analog.

Screening and Synthesis of Tetrazole Derivatives that Inhibit the Growth of Cryptococcus Species.

Cryptococcosis has become a major health problem worldwide and caused morbidity and mortality in immunocompromised patients, especially those infected with human immunodeficiency virus (HIV). Despite the global distribution of cryptococcosis, the number and types of the available antifungals are limited, and the treatment outcomes in HIV patients are generally poor. In this study, we screened a compound library and identified one tetrazole derivative as an efficient inhibitor of Cryptococcus neoformans and Cryptococcus gattii. We further designed and synthesized a series of tetrazole derivatives and determined their structure-activity relationship, demonstrating that tetrazole backbone-containing compounds could be developed as novel antifungal drugs with distinct mechanisms against Cryptococcus spp. Our findings provide a starting point for novel target identification and structural optimization to develop a distinct class of therapeutics for patients with cryptococcosis.

Neutrophil S100A9 supports M2 macrophage niche formation in granulomas.

Mycobacterium infection gives rise to granulomas predominantly composed of inflammatory M1-like macrophages, with bacteria-permissive M2 macrophages also detected in deep granulomas. Our histological analysis of Mycobacterium bovis bacillus Calmette-Guerin-elicited granulomas in guinea pigs revealed that S100A9-expressing neutrophils bordered a unique M2 niche within the inner circle of concentrically multilayered granulomas. We evaluated the effect of S100A9 on macrophage M2 polarization based on guinea pig studies. S100A9-deficient mouse neutrophils abrogated M2 polarization, which was critically dependent on COX-2 signaling in neutrophils. Mechanistic evidence suggested that nuclear S100A9 interacts with C/EBPß, which cooperatively activates the Cox-2 promoter and amplifies prostaglandin E2 production, followed by M2 polarization in proximal macrophages. Because the M2 populations in guinea pig granulomas were abolished via treatment with celecoxib, a selective COX-2 inhibitor, we propose the S100A9/Cox-2 axis as a major pathway driving M2 niche formation in granulomas.

Challenges Based on Antiplasmodial and Antiviral Activities of 7-Chloro-4-aminoquinoline Derivatives.

We report the structural functionalization of the terminal amino group of N1 -(7-chloroquinolin-4-yl) butane-1,4-diamine, leading to a series of 7-chloro-4-aminoquinoline derivatives, and their evaluation as potent anti-malarial and anti-viral agents. Some compounds exhibited promising anti-malarial effects against the Plasmodium falciparum 3D7 (chloroquine-sensitive) and Dd2 (chloroquine-resistant) strains. In addition, these compounds were assayed in vitro against influenza A virus (IAV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Compound 5 h, bearing an N-mesityl thiourea group, displayed pronounced anti-infectious effects against malaria, IAV, and SARS-CoV-2. These results provide new insights into drug discovery for the prevention or treatment of malaria and virus co-infection.

Identification of novel chemical compounds targeting filovirus VP40-mediated particle production.

The central role of Ebola virus (EBOV) VP40 in nascent virion assembly and budding from infected host cells makes it an important therapeutic target. The mechanism of dimerization, following oligomerization of VP40 leading to the production of virus-like particles (VLP) has never been investigated for the development of therapeutic candidates against Ebola disease. Molecular dynamics-based computational screening targeted VP40 dimer with 40,000,000 compounds selected 374 compounds. A novel in vitro screening assay selected two compounds, NUSU#1 and NUSU#2. Conventional VLP assays consistently showed that both compounds inhibited EBOV VP40-mediated VLP production. Intriguingly, NUSU#1 inhibited the VP40-mediated VLP production in other ebolavirus species and the Marburg virus, but did not inhibit Lassa virus Z-mediated VLP production. These results strongly suggested that the selected compounds are potential lead drug candidates against Filovirus disease via disruption of VP40-mediated particle production.

Lead Optimization of Influenza Virus RNA Polymerase Inhibitors Targeting PA-PB1 Interaction.

Influenza viruses are responsible for contagious respiratory illnesses in humans and cause seasonal epidemics and occasional pandemics worldwide. Previously, we identified a quinolinone derivative PA-49, which inhibited the influenza virus RNA-dependent RNA polymerase (RdRp) by targeting PA-PB1 interaction. This paper reports the structure optimization of PA-49, which resulted in the identification of 3-((dibenzylamino)methyl)quinolinone derivatives with more potent anti-influenza virus activity. During the optimization, the hit compound 89, which was more active than PA-49, was identified. Further optimization and scaffold hopping of 89 led to the most potent compounds 100 and a 1,8-naphthyridinone derivative 118, respectively. We conclusively determined that compounds 100 and 118 suppressed the replication of influenza virus and exhibited anti-influenza virus activity against both influenza virus types A and B in the range of 50% effective concentration (EC50) = 0.061-0.226 µM with low toxicity (50% cytotoxic concentration (CC50) >10 µM).

Trifluoromethylthiolation of Hindered α-Bromoamides with Nucleophilic Trifluoromethylthiolating Reagents.

General methods have not been previously developed for the synthesis of sterically hindered α-SCF3-substituted carbonyl compounds using nucleophilic trifluoromethylthiolating reagents. Thus, we herein report sp3C-SCF3 bond formation in hindered α-bromoamides containing 3-bromo-oxindoles and linear α-bromoamides using CuSCF3 or AgSCF3 under mild conditions to access sterically hindered α-SCF3-substituted amides. This transformation is applicable to not only 3-SCF3-substituted oxindoles but also primary and secondary amides and reveals a broad functional group tolerance. This method will benefit the fields of medicinal and agricultural chemistry.

2D-quantitative structure-activity relationships model using PLS method for anti-malarial activities of anti-haemozoin compounds.

BACKGROUND: Emergence of cross-resistance to current anti-malarial drugs has led to an urgent need for identification of potential compounds with novel modes of action and anti-malarial activity against the resistant strains. One of the most promising therapeutic targets of anti-malarial agents related to food vacuole of malaria parasite is haemozoin, a product formed by the parasite through haemoglobin degradation. METHODS: With this in mind, this study developed two-dimensional-quantitative structure-activity relationships (QSAR) models of a series of 21 haemozoin inhibitors to explore the useful physicochemical parameters of the active compounds for estimation of anti-malarial activities. The 2D-QSAR model with good statistical quality using partial least square method was generated after removing the outliers. RESULTS: Five two-dimensional descriptors of the training set were selected: atom count (a_ICM); adjacency and distance matrix descriptor (GCUT_SLOGP_2: the third GCUT descriptor using atomic contribution to logP); average total charge sum (h_pavgQ) in pKa prediction (pH = 7); a very low negative partial charge, including aromatic carbons which have a heteroatom-substitution in "ortho" position (PEOE_VSA-0) and molecular descriptor (rsynth: estimating the synthesizability of molecules as the fraction of heavy atoms that can be traced back to starting material fragments resulting from retrosynthetic rules), respectively. The model suggests that the anti-malarial activity of haemozoin inhibitors increases with molecules that have higher average total charge sum in pKa prediction (pH = 7). QSAR model also highlights that the descriptor using atomic contribution to logP or the distance matrix descriptor (GCUT_SLOGP_2), and structural component of the molecules, including topological descriptors does make for better anti-malarial activity. CONCLUSIONS: The model is capable of predicting the anti-malarial activities of anti-haemozoin compounds. In addition, the selected molecular descriptors in this QSAR model are helpful in designing more efficient compounds against the P. falciparum 3D7A strain.

Silver-Promoted Fluorination Reactions of α-Bromoamides.

Silver-promoted C-F bond formation in α-bromoamides by using AgF under mild conditions is reported. This simple method enables access to tertiary, secondary, and primary alkyl fluorides involving biomolecular scaffolds. This transformation is applicable to primary and secondary amides and shows broad functional-group tolerance. Kinetics experiments revealed that the reaction rate increased in the order of 3°>2°>1° α-carbon atom. In addition, it was found that the acidic amide proton plays an important role in accelerating the reaction. Mechanistic studies suggested generation of an aziridinone intermediate that undergoes subsequent nucleophilic addition to form the C-F bond with stereospecificity (i.e., retention of configuration). The synthesis of sterically hindered alcohols and ethers by using AgI is also demonstrated. Examples of reactions of α-bromoamides with O nucleophiles are presented.

An Antiviral Drug Screening Platform with a FRET Biosensor for Measurement of Arenavirus Z Assembly.

The smallest arenavirus gene product, Z protein, plays critical roles in the virus life cycle. Z is the major driving force of budding and particle production because of a unique property that defines self-assembly. In addition to the roles in budding, Z also participates in the suppression of type I interferon production to evade host antiviral immunity. Therefore, Z and its assembled form are an attractive drug target for arenaviral hemorrhagic fever, such as Lassa fever. Here, we developed a biosensor that enabled the evaluation of the prototype arenavirus, lymphocytic choriomeningitis virus (LCMV), Z assembly using the principle of Förster resonance energy transfer (FRET). This FRET biosensor consisted of three tandem Z that were sandwiched between super-enhanced cyan-emitting fluorescent protein and variant of a yellow-emitting mutant of green fluorescent protein so that Z-Z intermolecular binding via the really interesting new gene finger domain increased the emission ratio. To identify novel anti-arenavirus compounds, the FRET biosensor was employed to screen the PathogenBox400 for inhibitors of Z assembly in a 96-well plate format. The assay performed well, with a Z'-factor of 0.89, and identified two compounds that decreased the emission ratio of the FRET biosensor in a dose-dependent manner. Of them, the compound, 5,6,7,8-tetrahydro-7-(benzyl)-pyrido[4',3':4,5]thieno[2,3-d]pyrimidin-2,4-diamine, was found to significantly inhibit LCMV propagation in infected cells. Thereby, the present study demonstrated that a novel FRET biosensor incorporating Z assembly built on FRET and named Zabton, was a valuable screening tool to identify anti-arenavirus compounds in the context of inhibition of Z assembly.Key words: Arenavirus, Förster resonance energy transfer, anti-viral drugs, Z protein.

Novel and potent antimicrobial effects of caspofungin on drug-resistant Candida and bacteria.

Echinocandins, including caspofungin, micafungin, and anidulafungin, are first-line antifungal agents for the treatment of invasive candidiasis. They exhibit fungicidal activity by inhibiting the synthesis of ß-1,3-D-glucan, an essential component of the fungal cell wall. However, they are active only against proliferating fungal cells and unable to completely eradicate fungal cells even after a 24 h drug exposure in standard time-kill assays. Surprisingly, we found that caspofungin, when dissolved in low ionic solutions, had rapid and potent antimicrobial activities against multidrug-resistant (MDR) Candida and bacteria cells even in non-growth conditions. This effect was not observed in 0.9% NaCl or other ion-containing solutions and was not exerted by other echinocandins. Furthermore, caspofungin dissolved in low ionic solutions drastically reduced mature biofilm cells of MDR Candida auris in only 5 min, as well as Candida-bacterial polymicrobial biofilms in a catheter-lock therapy model. Caspofungin displayed ion concentration-dependent conformational changes and intracellular accumulation with increased reactive oxygen species production, indicating a novel mechanism of action in low ionic conditions. Importantly, caspofungin dissolved in 5% glucose water did not exhibit increased toxicity to human cells. This study facilitates the development of new therapeutic strategies in the management of catheter-related biofilm infections.

Novel Compounds Identified by Structure-Based Prion Disease Drug Discovery Using In Silico Screening Delay the Progression of an Illness in Prion-Infected Mice.

The accumulation of abnormal prion protein (PrPSc) produced by the structure conversion of PrP (PrPC) in the brain induces prion disease. Although the conversion process of the protein is still not fully elucidated, it has been known that the intramolecular chemical bridging in the most fragile pocket of PrP, known as the "hot spot," stabilizes the structure of PrPC and inhibits the conversion process. Using our original structure-based drug discovery algorithm, we identified the low molecular weight compounds that predicted binding to the hot spot. NPR-130 and NPR-162 strongly bound to recombinant PrP in vitro, and fragment molecular orbital (FMO) analysis indicated that the high affinity of those candidates to the PrP is largely dependent on nonpolar interactions, such as van der Waals interactions. Those NPRs showed not only significant reduction of the PrPSc levels but also remarkable decrease of the number of aggresomes in persistently prion-infected cells. Intriguingly, treatment with those candidate compounds significantly prolonged the survival period of prion-infected mice and suppressed prion disease-specific pathological damage, such as vacuole degeneration, PrPSc accumulation, microgliosis, and astrogliosis in the brain, suggesting their possible clinical use. Our results indicate that in silico drug discovery using NUDE/DEGIMA may be widely useful to identify candidate compounds that effectively stabilize the protein.

Design and Development of an HBT-Based Ratiometric Fluorescent Probe to Monitor Stress-Induced Premature Senescence.

Stress-induced premature senescence (SIPS) can be induced in tumor cells by reactive oxygen species (ROS) or oncogenes. The antineoplastic drugs cause apoptosis and senescence by damaging the DNA. Although the detection of cellular senescence is important to monitor drug response during anticancer therapy, only a few probes have been studied for imaging SIPS. In this study, we developed 2-(2'-hydroxyphenyl)benzothiazole (HBT)-based fluorescent probes to determine SIPS by monitoring the oxidative stress and ß-galactosidase activity. HBT is a commonly used fluorophore because of its luminescence mechanism via excited-state intramolecular proton transfer, and it has attractive properties, such as a four-level photochemical process and large Stokes shift (151 nm). A novel fluorescent probe, (2-(benzo[d]thiazol-2-yl)phenyl)boronic acid, was prepared for the detection of ROS, including H2O2, via the oxidation reaction of arylboronic acids to form the fluorescent phenol, HBT. In addition, to determine the enzymatic activity of ß-galactosidase, a 2-(4'-chloro-2'-hydroxyphenyl)benzothiazole (CBT)-based enzymatic turn-on probe (CBT-ß-Gal) was designed and synthesized. ß-Galactosidase catalyzed the hydrolysis of ß-galactopyranoside from CBT-ß-Gal to release the fluorescent CBT. These probes were capable of ratiometric imaging the accumulation of H2O2 and the degree of ß-galatosidase activity in contrast to H2O2-untreated and H2O2-treated HeLa cells. Furthermore, these probes were successfully employed for imaging the increased levels of ROS and ß-galactosidase activity in the doxorubicin-treated HeLa cells.

A Quinolinone Compound Inhibiting the Oligomerization of Nucleoprotein of Influenza A Virus Prevents the Selection of Escape Mutants.

The emergence of resistance to currently available anti-influenza drugs has heightened the need for antivirals with novel mechanisms of action. The influenza A virus (IAV) nucleoprotein (NP) is highly conserved and essential for the formation of viral ribonucleoprotein (vRNP), which serves as the template for replication and transcription. Recently, using in silico screening, we identified an antiviral compound designated NUD-1 (a 4-hydroxyquinolinone derivative) as a potential inhibitor of NP. In this study, we further analyzed the interaction between NUD-1 and NP and found that the compound interferes with the oligomerization of NP, which is required for vRNP formation, leading to the suppression of viral transcription, protein synthesis, and nuclear export of NP. We further assessed the selection of resistant variants by serially passaging a clinical isolate of the 2009 H1N1 pandemic influenza virus in the presence of NUD-1 or oseltamivir. NUD-1 did not select for resistant variants after nine passages, whereas oseltamivir selected for resistant variants after five passages. Our data demonstrate that NUD-1 interferes with the oligomerization of NP and less likely induces drug-resistant variants than oseltamivir; hence, it is a potential lead compound for the development of novel anti-influenza drugs.

Synthesis and Immunomodulatory Activity of Fluorine-Containing Bisphosphonates.

Immune checkpoint blockade using anti-PD-1/PD-L1 or anti-CTLA-4 monoclonal antibodies (mAbs) has revolutionized cancer treatment. However, many types of cancer do not respond and for those that do, only a minority of patients achieve durable remissions. Therefore, oncoimmunologists are working to develop adoptive cell therapies for non-hematopoietic tumors by harnessing immune effector cells such as αß T cells and γδ T cells. In contrast to conventional αß T cells that recognize peptides in the context of MHC class I or II molecules, γδ T cells expressing Vγ2Vδ2 T cell receptors (also termed Vγ9Vδ2) are stimulated by isoprenoid metabolites (phosphoantigens) such as isopentenyl diphosphate in a butyrophilin-3A1-dependent manner. Vγ2Vδ2 T cells kill almost all types of tumor cells that have been treated with bisphosphonates. In this study, we synthesized a series of fluorine-containing bisphosphonates based on current drugs and found that they stimulated Vγ2Vδ2 T cell killing of tumor cells. A fluorine-containing prodrug analogue of zoledronate where phosphonate moieties were masked with pivaloyloxymethyl groups markedly enhanced Vγ2Vδ2 T-cell-mediated cytotoxicity, and also promoted the expansion of peripheral blood Vγ2Vδ2 T cells. These results demonstrate that a prodrug of a fluorine-containing zoledronate analogue can sensitize tumor cells for killing as well as expand Vγ2Vδ2 T cells for adoptive cell therapy.

Determination of human γδ T cell-mediated cytotoxicity using a non-radioactive assay system.

The adoptive transfer of immune effector cells, such as CD8+ killer αß T cells, γδ T cells, NK (natural killer) cells, and genetically-modified T cells, has been receiving increasing attention. It is essential to determine cellular cytotoxicity so as to monitor the function and quality of ex vivo-expanded immune effector cells before infusion. The most common method is the [51Cr]-sodium chromate release assay. It is, however, preferable to avoid the use of radioactive materials in clinical laboratories. In order to establish a non-radioactive alternative to the standard radioactive assay, we previously synthesized a chelate-forming prodrug (BM-HT) and demonstrated that a combination of BM-HT and europium (Eu3+) was useful to determine NK cell-mediated cytotoxicity. In the present study, we examined whether or not this improved assay system could be used to determine the cellular cytotoxicity exhibited by Vγ2Vδ2+ γδ T cells. In addition, we compared Eu3+ and terbium (Tb3+) in the measurement of cellular cytotoxicity. Our assay system using BM-HT could be used successfully for the analysis of both γδ T cell receptor (TCR)- and CD16-mediated cytotoxicity. When the intensity of fluorescence was compared between Eu3+ and Tb3+, Tb3+ chelate was more sensitive than Eu3+ chelate, suggesting that the detection system using Tb3+ is superior to Eu3+ when tumor cells are not efficiently labeled with BM-HT. The method established herein is expected to promote the development of novel adoptive cell therapies for cancer.

Synthesis of Trifluoromethyl-α,ß-unsaturated Lactones and Pyrazolinones and Discovery of Influenza Virus Polymerase Inhibitors.

To explore the potential biological activities of trifluoromethyl heterocycles, we recently developed a synthetic approach to access a series of α-trifluoromethyl-α,ß-unsaturated lactones and trifluoromethyl pyrazolinones. The compounds were tested for their antimicrobial activity, and we found that some compounds had anti-influenza viral activity. The ß-aryl-α-trifluoromethyl α,ß-unsaturated lactone derivatives 5 g (5-(4-chlorophenyl)-5-methyl-4-phenyl-3-(trifluoromethyl)furan-2-one), 7 b (4-(4-methoxyphenyl)-3-(trifluoromethyl)spiro[furan-5,1'-indane]-2-one), and the trifluoromethyl pyrazolinone 12 c (1-(6-methoxy-2-naphthyl)-2-(trifluoromethyl)-5,6,7,8-tetrahydropyrazolo[1,2-a]pyridazin-3-one) were found to possess promising inhibitory activity against influenza virus type A, strain A/WSN/33 (H1N1). These three hit compounds were successfully optimized, and we identified that the most potent compound 5 h (5-(4-chlorophenyl)-4-(6-methoxy-2-naphthyl)-5-methyl-3-(trifluoromethyl)furan-2-one) showed inhibitory activity against various types of influenza A and B viruses in the low-micromolar range without showing cytotoxicity. Moreover, 5 h was more effective against the clinical isolate A/California/7/2009 (H1N1pdm) strain than the influenza viral polymerase inhibitor, favipiravir (T-705). We also delineated the structure-activity relationship and obtained mechanistic insight into inhibition of the viral polymerase.

Fragment Molecular Orbital Study of the Interaction between Sarco/Endoplasmic Reticulum Ca2+-ATPase and its Inhibitor Thapsigargin toward Anti-Malarial Development.

Plasmodium falciparum, the causative agent of malignant malaria, is insensitive to thapsigargin (TG), a well-known inhibitor of the human sarco/endoplasmic reticulum Ca2+-ATPase (SERCA). To understand the key factor causing the difference of the sensitivity, the molecular interaction of TG and each SERCA was analyzed by the fragment molecular orbital (FMO) method. While the major component of the interaction energy was the nonpolar interaction, the major difference in the molecular interaction arose from the polar interaction, namely, the hydrogen bonding interaction with a hydroxyl group of TG. Additionally, we successfully confirmed these FMO calculation results by measuring the inhibitory activity of a synthesized TG derivative. Our calculations and experiments indicated that, by replacing the hydroxyl group of TG with another functional group, the sensitivities of TG to human and P. falciparum SERCAs can be reversed. This study provides important information to develop antimalarial compounds targeting P. falciparum SERCA.

Prediction Model for Antimalarial Activities of Hemozoin Inhibitors by Using Physicochemical Properties.

The rapid spread of strains of malaria parasites that are resistant to several drugs has threatened global malaria control. Hence, the aim of this study was to predict the antimalarial activity of chemical compounds that possess anti-hemozoin-formation activity as a new means of antimalarial drug discovery. After the initial in vitro anti-hemozoin-formation high-throughput screening (HTS) of 9,600 compounds, a total of 224 hit compounds were identified as hemozoin inhibitors. These 224 compounds were tested for in vitro erythrocytic antimalarial activity at 10 µM by using chloroquine-mefloquine-sensitive Plasmodium falciparum strain 3D7A. Two independent experiments were conducted. The physicochemical properties of the active compounds were extracted from the ChemSpider and SciFinder databases. We analyzed the extracted data by using Bayesian model averaging (BMA). Our findings revealed that lower numbers of S atoms; lower distribution coefficient (log D) values at pH 3, 4, and 5; and higher predicted distribution coefficient (ACD log D) values at pH 7.4 had significant associations with antimalarial activity among compounds that possess anti-hemozoin-formation activity. The BMA model revealed an accuracy of 91.23%. We report new prediction models containing physicochemical properties that shed light on effective chemical groups for synthetic antimalarial compounds and help with in silico screening for novel antimalarial drugs.