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Cancer cells rely on lysosome-dependent degradation to recycle nutrients that serve their energetic and biosynthetic needs. Despite great interest in repurposing the antimalarial hydroxychloroquine as a lysosomal inhibitor in clinical oncology trials, the mechanisms by which hydroxychloroquine and other lysosomal inhibitors induce tumor-cell cytotoxicity remain unclear. In this issue of the JCI, Bhardwaj et al. demonstrate that DC661, a dimeric form of chloroquine that inhibits palmitoyl-protein thioesterase 1 (PPT1), promoted lysosomal lipid peroxidation, resulting in lysosomal membrane permeabilization and tumor cell death. Remarkably, this lysosomal cell death pathway elicited cell-intrinsic immunogenicity and promoted T lymphocyte-mediated tumor cell clearance. The findings provide the mechanistic foundation for the potential combined use of immunotherapy and lysosomal inhibition in clinical trials.

Background: The spread of Plasmodium vivax strains resistant to chloroquine (CQ) has posed a challenge to control strategies aimed at eliminating malaria. Molecular analysis of candidate resistance markers is very important for monitoring the P. vivax resistance to CQ in different endemic regions. In the present study, the multidrug resistance 1 (pvmdr1) gene, a possible marker for CQ resistance in P. vivax, was evaluated by molecular methods., Methods: A simple PCR-RFLP method was developed for mutation analysis in pvmdr1 gene. A number of 120 blood spots were obtained from patients with P. vivax mono-infection in 2021. All of the samples were collected from Pakistani patients who travelled to Iran., Results: None of the samples had any mutation at codon 976 of pvmdr1, while the 1076 mutation was detected in 96.2% of the examined isolates. Only two pvmdr1 haplotypes were identified, including the single mutant (Y976/1076L) as the most prevalent haplotype (with 96.2% frequency) and the wild type (Y976/F1076; with 3.8% frequency)., Conclusions: In this study, the major CQ resistance-mediating mutation and multiple mutant haplotypes of the pvmdr1 gene was not detected. However, continuous monitoring of drug resistance markers and close supervision of the efficacy of CQ is essential to detect the potential emergence of CQ-resistant P. vivax isolates in Iran. This data is important for performing future epidemiological surveillance to monitor CQ resistance in this endemic area and the bordering regions., (© 2023. The Author(s).)

We report the structural functionalization of the terminal amino group of N 1 -(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., (© 2023 Wiley-VCH GmbH.)

In the present work, a library of 120 compounds was prepared using various aliphatic and aromatic amines. Finally, 10 compounds were selected through in silico screening carrying 4-aminobenzoyl-l-glutamic acid and 1,3,5-triazine moiety. The docking results of compounds 4d16 and 4d38 revealed higher binding interaction with amino acids Asp54 (-537.96 kcal/mol) and Asp54, Phe116 (-618.22 kcal/mol) against wild (1J3I) and quadruple mutant (1J3K) type of Pf-DHFR inhibitors and were comparable to standard WR99210. These compounds were developed by facile and microwave-assisted synthesis via nucleophilic substitution reaction and characterized by different spectroscopic methods. In vitro antimalarial assay results also suggested that these two compounds were having higher antimalarial activity against chloroquine-sensitive (3D7) and chloroquine-resistant (Dd2) strain out of the ten synthesized compounds with IC 50 13.25 μM and 14.72 μM, respectively. These hybrid scaffolds might be useful in the lead discovery of a new class of Pf-DHFR inhibitors., (© 2022 Wiley Periodicals LLC.)

The emergence of resistance to conventional antimalarial treatments remains a major cause for concern. New drugs that target the distinct development stages of Plasmodium parasites are required to address this risk. Herein, water-soluble aggregation-induced emission active cyclometalated iridium(III) polypyridyl complexes (Ir1-Ir12) are developed for the elimination of malaria parasites. Remarkably, these complexes show potent antimalarial activity in low nanomolar range against 3D7 (chloroquine and artemisinin sensitive strain), RKL9 (chloroquine resistant strain), and R539T (artemisinin resistant strains) strains of Plasmodium falciparum with faster killing rate of malaria parasites. Concomitantly, these complexes exhibit efficient in vivo antimalarial activity against both the asexual and gametocyte stages of Plasmodium berghei malaria parasite, suggesting promising transmission-blocking potential. The complexes tend to localize into mitochondria of P. falciparum determined by image and cell-based assay. The mechanistic studies reveal that these complexes exert their antimalarial activity by increasing reactive oxygen species levels and disrupting its mitochondrial membrane potential. Furthermore, the mitochondrial-dependent antimalarial activity of these complexes is confirmed in yeast model. Thus, this study for the first time highlights the potential role of targeting P. falciparum mitochondria by iridium complexes in discovering and developing the next-generation antimalarial agents for treating multidrug resistant malaria parasites., (© 2022 Wiley-VCH GmbH.)

Background: Outer membrane protein A (OmpA) is the major virulence factor of Acinetobacter baumannii and plays a wide role in the pathogenesis and antimicrobial resistance of A. baumannii. Dendritic cells (DCs) are the most effective antigen-presenting cells and play a crucial role in regulating the immune response to multiple antigens and immune sentries. We aimed to study the role and molecular mechanisms of OmpA-induced mouse bone marrow-derived dendritic cells (BMDCs) autophagy in the immune response of A. baumannii., Methods: First, purified A. baumannii OmpA was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blot. OmpA effect on BMDCs viability was evaluated by MTT assay. BMDCs were pretreated with autophagy inhibitor chloroquine or transfected with overexpression plasmids (oe-NC or oe-PI3K). Then BMDCs apoptosis, inflammatory cytokines, protein kinase B (PI3K)/mammalian target of rapamycin (mTOR) pathway, and autophagy-related factors levels were evaluated., Results: SDS-PAGE and western blot verified the successful purification of OmpA. BMDCs viability repressed gradually with the increase of OmpA concentration. OmpA treatment of BMDCs led to apoptosis and inflammation in BMDCs. OmpA caused incomplete autophagy in BMDCs, and light chain 3 (LC3), Beclin1, P62, and LC3II/I levels were significantly elevated with the increase of the time and concentration of OmpA treatment. Chloroquine reversed OmpA effects on autophagy in BMDCs, that was, LC3, Beclin1, and LC3II/I levels were reduced, while P62 level was elevated. Furthermore, chloroquine reversed OmpA effects on apoptosis and inflammation in BMDCs. PI3K/mTOR pathway-related factor expression was affected by OmpA treatment of BMDCs. After overexpression of PI3K, these effects were reversed., Conclusions: A. baumannii OmpA induced autophagy in BMDCs involving the PI3K/mTOR pathway. Our study may provide a novel therapeutic target and theoretical basis for treating infections caused by A. baumannii., (© 2023 The Authors. Immunity, Inflammation and Disease published by John Wiley & Sons Ltd.)

Host immunity has been suggested to clear drug-resistant parasites in malaria-endemic settings. However, the immunogenetic mechanisms involved in parasite clearance are poorly understood. Characterizing the host's immunity and genes involved in controlling the parasitic infection can inform the development of blood-stage malaria vaccines. This study investigates host regulatory cytokines and immunogenomic factors associated with the clearance of Plasmodium falciparum carrying a chloroquine resistance genotype. Biological samples from participants of previous drug efficacy trials conducted in two Malian localities were retrieved. The P. falciparum chloroquine resistance transporter (Pfcrt) gene was genotyped using parasite DNA. Children carrying parasites with the mutant allele (Pfcrt-76T) were classified based on their ability to clear their parasites. The levels of the different cytokines were measured in serum. The polymorphisms of specific human genes involved in malaria susceptibility were genotyped using human DNA. The prevalence of the Pfcrt-76T was significantly higher in Kolle than in Bandiagara (81.6 % vs 38.6 %, p < 10 -6 ). The prevalence of children who cleared their mutant parasites was significantly higher in Bandiagara than in Kolle (82.2 % vs 67.4 %, p < 0.05). The genotyping of host genes revealed that IFN-γ -874 T and TNF-α -308A alleles were positively associated with parasite clearance. Cytokine profiling revealed that IFN-γ level was positively associated with parasite clearance (p = 0.04). This study highlights the role of host's immunity and immunogenetic factors to clear resistant parasites, suggesting further characterization of these polymorphisms may help to develop novel approaches to antiparasitic treatment strategies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Crown Copyright © 2023. Published by Elsevier Ltd. All rights reserved.)

The root extract of Nauclea xanthoxylon (A.Chev.) Aubrév. displayed significant 50 % inhibition concentration (IC 50 s) of 0.57 and 1.26 μg/mL against chloroquine resistant and sensitive Plasmodium falciparum (Pf) Dd2 and 3D7 strains, respectively. Bio-guided fractionation led to an ethyl acetate fraction with IC 50 s of 2.68 and 1.85 μg/mL and subsequently, to the new quinovic acid saponin named xanthoxyloside (1) with IC 50 s of 0.33 and 1.30 μM, respectively against the tested strains. Further compounds obtained from ethyl acetate and hexane fractions were the known clethric acid (2), ursolic acid (3), quafrinoic acid (4), quinovic acid (5), quinovic acid 3-O-β-D-fucopyranoside (6), oleanolic acid (7), oleanolic acid 3-acetate (8), friedelin (9), β-sitosterol (10a), stigmasterol (10b) and stigmasterol 3-O-β-D-glucopyranoside (11). Their structures were characterised with the aid of comprehensive spectroscopic methods (1 and 2D NMR, Mass). Bio-assays were performed using nucleic acid gel stain (SYBR green I)-based fluorescence assay with chloroquine as reference. Extracts and compounds exhibited good selectivity indices (SIs) of >10. Significant antiplasmodial activities measured for the crude extract, the ethyl acetate fraction and xanthoxyloside (1) from that fraction can justify the use of the root of N. xanthoxylon in ethnomedicine to treat malaria., (© 2023 Wiley-VHCA AG, Zurich, Switzerland.)

Parkinson's disease (PD) is a neuro-motor ailment that strikes adults in their older life and results in both motor and non-motor impairments. In neuronal and glial cells, PD has recently been linked to a dysregulated autophagic system and cerebral inflammation. Chloroquine (CQ), an anti-malarial drug, has been demonstrated to suppress autophagy in a variety of diseases, including cerebral ischemia, Alzheimer's disease (AD), and Traumatic brain injury (TBI), while its involvement in PD is still unclear. BALB/c mice were randomly allocated to one of four groups: 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP), CQ treatment with or without MPTP, or control. The CQ treatment group received CQ (intraperitoneally, 8 mg/kg body weight) after 1 h of MPTP induction on day 1, and it lasted for 7 days. CQ therapy preserves dopamine levels stable, inhibits tyrosine hydroxylase (TH) positive dopaminergic cell death, and lowers oxidative stress. CQ reduces the behavioural, motor, and cognitive deficits caused by MPTP after injury. Furthermore, CQ therapy slowed aberrant neuronal autophagy (microtubule-associated protein-1 light chain 3B; LC3B & Beclin1) and lowered expression levels of the inflammatory cytokines interleukin 1 (IL-1β) and tumour necrosis factor (TNF-α) in the mice brain. In addition, CQ's antioxidant and anti-inflammatory effects were also tested in MPTP-mediated cell death in PC12 cells, demonstrating that CQ has a neurorestorative impact by successfully rescuing MPTP-induced ROS generation and cell loss. Our findings show that CQ's can help to prevent dopaminergic degeneration and improve neurological function after MPTP intoxication by lowering the harmful effects of neuronal autophagy and cerebral inflammation., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Aims: Non-small cell lung cancer (NSCLC) is one of the aggressive tumors mostly diagnosed in the advanced stage. Therapeutic failure and drug resistance pose a major problem in NSCLC treatment primarily due to alterations in autophagy and loss of apoptosis. Therefore, the present study aimed to investigate the importance of the second mitochondria-derived activator of caspase mimetic BV6 and autophagy inhibitor chloroquine (CQ) on the regulation of apoptosis and autophagy, respectively., Subjects and Methods: Study was conducted on NCI-H23 and NCI-H522 cell lines to evaluate the effect of BV6 and CQ on the transcription and translation level of LC3-II, caspase-3, and caspase-9 genes by quantitative real-time-polymerase chain reaction and western blotting techniques., Results: In NCI-H23 cell line, BV6 and CQ treatments showed increased mRNA and protein expression of caspase-3, and caspase-9 compared to its untreated counterpart. BV6 and CQ treatments also caused downregulation of LC3-II protein expression compared to its counterpart. In NCI-H522 cell line, BV6 treatment showed a significantly increased expression of caspase-3 and caspase-9 mRNA and protein expression levels whereas BV6 treatment downregulated the expression level of LC3-II protein. A similar pattern was also observed in CQ treatment when compared with the respective controls. Both BV6 and CQ modulated in vitro expression of caspases and LC3-II which have critical regulatory roles in apoptosis and autophagy, respectively., Conclusions: Our findings suggest that BV6 and CQ could be promising candidates in NSCLC treatment and there is a need to explore them in vivo and in clinical applications., Competing Interests: None

Hepatic stellate cell (HSC) activation via the autophagy pathway is a critical factor in liver fibrogenesis. This study tests the hypothesis that chloroquine (CQ) treatment can prevent autophagy and HSC activation in vitro and in vivo in bile-duct-ligated (BDL) mice. Sham-operated and BDL mice were treated with either PBS or CQ in two 60 mg/kg doses the day (D) before and after surgery. On day 2 (2D), HSCs were isolated, and their biological activities were evaluated by measuring intracellular lipid content, α-sma/collagen, and expression of autophagy lc3, sqstm1/p62 markers. The treatment efficacy on liver function was evaluated with serum albumin, transaminases (AST/ALT), and hepatic histology. Primary HSCs were treated in vitro for 24 h with CQ at 0, 2.5, 5, 10, 30, and 50 µM. Autophagy and HSC activation were assessed after 2D of treatment. CQ treatment improved serum AST/ALT, albumin, and bile duct proliferation in 2D BDL mice. This is associated with a suppression of HSC activation, shown by higher HSC lipid content and collagen I staining, along with the blockage of HSC autophagy indicated by an increase in p62 level and reduction in lc3 staining. CQ 5 µM inhibited autophagy in primary HSCs in vitro by increasing p62 and lc3 accumulation, thereby suppressing their in vitro activation. The autophagy inhibitor CQ reduced HSC activation in vitro and in vivo. CQ improved liver function and reduced liver injury in BDL mice.

Neutrophil extracellular traps (NETs) occur when chromatin is decondensed and extruded from the cell, generating a web-like structure. NETs have been implicated in the pathogenesis of several sterile disease states and thus are a potential therapeutic target. Various pathways have been shown to induce NETs, including autophagy, with several key enzymes being activated like peptidyl arginine deiminase 4 (PAD4), an enzyme responsible for citrullination of histones, allowing for DNA unwinding and subsequent release from the cell. Pre-clinical studies have already demonstrated that chloroquine (CQ) and hydroxychloroquine (HCQ) are able to reduce NETs and slow disease progression. The exact mechanism as to how these drugs reduce NETs has yet to be elucidated. CQ and HCQ decrease NET formation from various NET activators, independent of their autophagy inhibitory function. CQ and HCQ were found to inhibit PAD4 exclusively, in a dose-dependent manner, confirmed with reduced CitH3+ NETs after CQ or HCQ treatment. Circulating CitH3 levels were reduced in pancreatic cancer patients after HCQ treatment. In silico screening of PAD4 protein structure identified a likely binding site interaction at Arg639 for CQ and Trp347, Ser468, and Glu580 for HCQ. SPR analysis confirmed the binding of HCQ and CQ with PAD4 with KD values of 54.1 µM (CQ) and 88.1 µM (HCQ). This data provide evidence of direct PAD4 inhibition as a mechanism for CQ/HCQ inhibition of NETs. We propose that these drugs likely reduce NET formation through multiple mechanisms; the previously established TLR9 and autophagy inhibitory mechanism and the novel PAD4 inhibitory mechanism., (© The Author(s) 2023. Published by Oxford University Press on behalf of the British Society for Immunology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Structural modification of existing chemical scaffolds to afford new molecules able to circumvent drug resistance constitutes one of the rational approaches to antimalarial drug discovery. Previously synthesized compounds based on the 4-aminoquinoline core hybridized with a chemosensitizing dibenzylmethylamine side group showed in vivo efficacy in Plasmodium berghei -infected mice despite low microsomal metabolic stability, suggesting a contribution from their pharmacologically active metabolites. Here, we report on a series of these dibemequine (DBQ) metabolites with low resistance indices against chloroquine-resistant parasites and improved metabolic stability in liver microsomes. The metabolites also exhibit improved pharmacological properties including lower lipophilicity, cytotoxicity, and hERG channel inhibition. Using cellular heme fractionation experiments, we also demonstrate that these derivatives inhibit hemozoin formation by causing a buildup of toxic "free" heme in a similar manner to chloroquine. Finally, assessment of drug interactions also revealed synergy between these derivatives and several clinically relevant antimalarials, thus highlighting their potential interest for further development.

Using a recently elucidated atomic-resolution cryogenic electron microscopy (cryo-EM) structure for the Plasmodium falciparum chloroquine resistance transporter (PfCRT) protein 7G8 isoform as template [Kim, J.; Nature 2019, 576, 315-320], we use Monte Carlo molecular dynamics (MC/MD) simulations of PfCRT embedded in a 1-palmitoyl-2-oleoyl- sn -glycero-3-phosphocholine (POPC) membrane to solve energy-minimized structures for 7G8 PfCRT and two additional PfCRT isoforms that harbor 5 or 7 amino acid substitutions relative to 7G8 PfCRT. Guided by drug binding previously defined using chloroquine (CQ) photoaffinity probe labeling, we also use MC/MD energy minimization to elucidate likely CQ binding geometries for the three membrane-embedded isoforms. We inventory salt bridges and hydrogen bonds in these structures and summarize how the limited changes in primary sequence subtly perturb local PfCRT isoform structure. In addition, we use the "AlphaFold" artificial intelligence AlphaFold2 (AF2) algorithm to solve for domain structure that was not resolved in the previously reported 7G8 PfCRT cryo-EM structure, and perform MC/MD energy minimization for the membrane-embedded AF2 structures of all three PfCRT isoforms. We compare energy-minimized structures generated using cryo-EM vs AF2 templates. The results suggest how amino acid substitutions in drug resistance-associated isoforms of PfCRT influence PfCRT structure and CQ transport.

Pharmacological inhibition of KRAS>RAF>MEK1/2>ERK1/2 signaling has provided no clinical benefit to patients with pancreatic ductal adenocarcinoma (PDAC). Interestingly, combined inhibition of MEK1/2 (with trametinib [T]) plus autophagy (with chloroquine [CQ] or hydroxychloroquine [HCQ]) demonstrated striking anti-tumor effects in preclinical models and in a patient (Patient 1). However, not all patients respond to the T/HCQ regimen, and Patient 1 eventually developed resistant disease. Here we report that primary or acquired resistance is associated with focal DNA copy number gains encompassing c-MYC. Furthermore, ectopic expression of c-MYC in PDAC cell lines rendered them T/HCQ resistant. Interestingly, a CDK4/6 inhibitor, palbociclib (P), also induced autophagy and overrode c-MYC-mediated T/HCQ resistance, such that P/HCQ promoted regression of T/HCQ-resistant PDAC tumors with elevated c-MYC expression. Finally, P/HCQ treatment of Patient 1 resulted in a biochemical disease response. These data suggest that elevated c-MYC expression is both a marker and a mediator of T/HCQ resistance, which may be overcome by the use of P/HCQ., (This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.)

Acidic lysosomes are indispensable for cancer development and linked to chemotherapy resistance. Chloroquine (CQ) and functional analogues have been considered as a potential solution to overcome the cancer progression and chemoresistance by inhibiting the lysosome-mediated autophagy and multidrug exocytosis. However, their anti-cancer efficacy in most clinical trials demonstrated modest improvement. In this study, we investigated the detailed mechanisms underlying the acquired resistance of K562 leukemic cells to CQ treatment. In response to 5-80 μM CQ, the lumen pH of endosomal-lysosomal system immediately increased and gradually reached dynamic equilibrium within 24 h. Leukemic cells produced more acidic organelles to tolerate 5-10 μM CQ. CQ (20-80 μM) concentration-dependently triggered cytosolic pH (pHi) rise, G0/G1 arrest, mitochondrial depolarization/fragmentation, and necrotic/apoptotic cell death. Oxidant induction by CQ was responsible for the mitochondria-dependent cytotoxicity and partial pHi elevation. Cells that survived the CQ cytotoxicity were accompanied with increased mitochondria. Under the 80 μM CQ challenge, co-treatment with the inhibitor of F 0 part of mitochondrial H + -ATP synthase, oligomycin (40 nM), prevented the elevation of oxidants as well as pHi, and attenuated stresses on mitochondria, cell survival, and cell proliferation. Besides, oligomycin-treated cells obviously displayed the lysosomal peripheralization and plasma membrane blebbing, suggesting that these cells were in process of lysosomal exocytosis and microvesicle release. Enhanced motion of these secretory processes allowed the cells to exclude CQ and repair necrotic injury. Together, the oxidant production and the proton dynamic interconnection among lysosomes, mitochondria, and cytosol are crucial for leukemic susceptibility to lysosomotropic chemotherapeutics., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to disclose., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Continued emerging resistance of pathogens against the clinically approved candidates and their associated limitations continuously demand newer agents having better potency with a more suited safety profile. Quinoline nuclei containing scaffolds of natural and synthetic origin have been documented for diverse types of pharmacological activities, and a number of drugs are clinically approved. In the present review, we unprecedentedly covered the biological potential of 4-substituted quinoline and elaborated a rationale for its special privilege to afford the significant number of approved clinical drugs, particularly against infectious pathogens. Compounds with 4-substituted quinoline are well documented for antimalarial activity, but in the last two decades, they have been extensively explored for activity against cancer, tuberculosis, and several other pathogens including viruses, bacteria, fungi, and other infectious pathogens. In the present study, the anti-infective spectrum of this scaffold is discussed against viruses, mycobacteria, malarial parasites, and fungal and bacterial strains, along with recent updates in this area, with special emphasis on the structure-activity relationship., (© 2022 Deutsche Pharmazeutische Gesellschaft.)

Chloroquine (CQ) and hydroxychloroquine (HCQ) show good efficacy in the treatment of SARS-CoV-2 in the early stage, while they are no longer recommended due to their side effects. As an important drug delivery carrier, serum albumin (SA) is closely related to the efficacy of drugs. Here, the affinity behaviour of chloroquine and hydroxychloroquine with two SA were investigated through the multispectral method of biochemistry and computer simulation. The results showed that the intrinsic emission of both SA was quenched by CQ and HCQ in a spontaneous exothermic entropy reduction static process, which relied mainly on hydrogen bonding and van der Waals forces. The lower binding constants suggested weak binding between the two drugs and SA, which might lead to differences in efficacy and possibly even to varying side effects. Binding site recognition demonstrated that CQ preferred to bind to the two sites of both SA, while HCQ tended to bind to site I of SA. The results of conformational studies demonstrated that CQ and HCQ could affect the structure of both SA by slightly increasing the α-helix content of SA. Finally, we combine the results from experimental start with molecular simulations to suggest drug modifications to guide the design of drugs. This work has important implications for guiding drug design improvements to select CQ derivatives with fewer side effects for the treatment of COVID-19., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Glioma is difficult-to-treat because of its infiltrative nature and the presence of the blood-brain barrier. Temozolomide is the only FDA-approved drug for its management. Therefore, finding a novel chemotherapeutic agent for glioma is of utmost importance. Magnolol, a neolignan, has been known for its apoptotic role in glioma. In this work, we have explored a novel anti-glioma mechanism of Magnolol associated with its role in autophagy modulation. We found increased expression levels of Beclin-1, Atg5-Atg12, and LC3-II and lower p62 expression in Magnolol-treated glioma cells. PI3K/AKT/mTOR pathway proteins were also downregulated in Magnolol-treated glioma cells. Next, we treated the glioma cells with Insulin, a stimulator of PI3K/AKT/mTOR signaling, to confirm that Magnolol induced autophagy by inhibiting this pathway. Insulin reversed the effect on Magnolol-mediated autophagy induction. We also established the same in in vivo glioma model where Magnolol showed an anti-glioma effect by inducing autophagy. To confirm the cytotoxic effect of Magnolol-induced autophagy, we used Chloroquine, a late-stage autophagy inhibitor. Chloroquine efficiently reversed the anti-glioma effects of Magnolol both in vitro and in vivo. Our study revealed the cytotoxic effect of Magnolol-induced autophagy in glioma, which was not previously reported. Additionally, Magnolol showed no toxicity in non-cancerous cell lines as well as rat organs. Thus, we concluded that Magnolol is an excellent candidate for developing new therapeutic strategies for glioma management., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Toxoplasma gondii, the etiologic agent of toxoplasmosis, infects about 30 - 50% of the world population. The currently available anti-Toxoplasma agents have serious limitations. The present study aimed to investigate the effects of two antimalarials; buparvaquone (BPQ) and chloroquine (CQ), on immunocompromised mice with chronic cerebral toxoplasmosis, using spiramycin as a reference drug. The assessed parameters included the estimation of mortality rates (MR) among mice of the different study groups, in addition to the examination of the ultrastructural changes in the brain tissues by transmission electron microscopy. The results showed that only CQ treatment could decrease the MR significantly with zero deaths, while both spiramycin and BPQ caused an insignificant reduction of MR compared to the infected non-treated group. All the used drugs decreased the number of mature ruptured cysts significantly compared to the infected non-treated group, while only CQ increased the number of atrophic and necrotic cysts significantly. Furthermore, both spiramycin and BPQ improved the microvasculopathy and neurodegeneration accompanying the infection with different degrees of reactive astrocytosis and neuronal damage with the best results regarding the repair of the microvascular damage with less active glial cells, and normal neurons in the CQ-treated group. In conclusion, this study sheds light on CQ and its excellent impact on treating chronic cerebral toxoplasmosis in an immunocompromised mouse model.

Lysosomes are known to influence cholesterol trafficking into endoplasmic reticulum (ER) membranes. Though intracellular cholesterol levels are known to influence the lipid biosynthetic responses in ER, the specific effects of lysosomal modulation on these outcomes is not known. To demonstrate this, C2C12 cells were treated with chloroquine, a lysosomotropic agent, and its effects on cellular biosynthetic capacity, structural and functional status of ER was determined. In addition to its known effects on autophagy reduction, chloroquine treatment induced accumulation of total cellular lipid and ER-specific cholesterol content. It was also observed that chloroquine caused an increase in smooth-ER content with defects in overall protein turnover. Further, since ER and mitochondria function in close association through ER membrane contact sites, it is likely that lysosomal modulation also brings about associated changes in mitochondria. In this regard, we found that chloroquine reduces mitochondrial membrane potential and mitochondrial dynamics. Collectively, the differential biosynthetic response of rise in lipid content, but not protein content, cannot be accounted by merely considering that chloroquine induced suppression of autophagy causes defects in organelle function. In this defective autophagy scenario, both biosynthetic responses such as lipid and protein synthesis are expected to be reduced rather than only the latter, as observed with chloroquine. These findings suggest that cholesterol trafficking/distribution within cellular organelles could act as an intracellular mediator of differential biosynthetic remodelling in interconnected organelles., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Hydroxychloroquine (HCQ) is a lysosomotropic agent that is commonly used for treating Sjögren's disease (SjD). However, its efficacy is controversial because of the divergent response to the drug among patients. In a subgroup of SjD patients, lysosome-associated membrane protein 3 (LAMP3) is elevated in expression in the salivary glands and promotes lysosomal dysregulation and lysosome-dependent apoptotic cell death. In this study, chloroquine (CQ) and its derivative HCQ were tested for their ability to prevent LAMP3-induced apoptosis, in vitro and on a mouse model of SjD. In addition, efficacy of HCQ treatment was retrospectively compared between high LAMP3 mRNA expression in minor salivary glands and those with LAMP3 mRNA levels comparable with healthy controls. Study results show that CQ treatment stabilized the lysosomal membrane in LAMP3-overexpressing cells via deactivation of cathepsin B, resulting in decreased apoptotic cell death. In mice with established SjD-like phenotype, HCQ treatment also significantly decreased apoptotic cell death and ameliorated salivary gland hypofunction. Retrospective analysis of SjD patients found that HCQ tended to be more effective in improving disease activity index, symptom severity and hypergammaglobulinemia in patients with high LAMP3 expression compared those with normal LAMP3 expression. Taken together, these findings suggested that by determining salivary gland LAMP3 mRNA level, a patient's response to HCQ treatment could be predicted. This finding may provide a novel strategy for guiding the development of more personalized medicine for SjD., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Protein ubiquitination is an important posttranslational regulation mechanism that mediates Plasmodium development and modifies parasite responses to antimalarial drugs. Although mutations in several parasite ubiquitination enzymes have been linked to increased drug tolerance, the molecular mechanisms by which ubiquitination pathways mediate these parasite responses remain largely unknown. Here, we investigate the roles of a Plasmodium falciparum ring finger ubiquitin ligase (PfRFUL) in parasite development and in responses to antimalarial drugs. We engineered a transgenic parasite having the Pfrful gene tagged with an HA-2A-NeoR-glmS sequence to knockdown (KD) Pfrful expression using glucosamine (GlcN). A Western blot analysis of the proteins from GlcN-treated pSLI-HA-NeoR-glmS-tagged (PfRFULg) parasites, relative to their wild-type (Dd2) controls, showed changes in the ubiquitination of numerous proteins. PfRFUL KD rendered the parasites more sensitive to multiple antimalarial drugs, including mefloquine, piperaquine, amodiaquine, and dihydroartemisinin. PfRFUL KD also decreased the protein level of the P. falciparum multiple drug resistance 1 protein (PfMDR1) and altered the ratio of two bands of the P. falciparum chloroquine resistance transporter (PfCRT), suggesting contributions to the changed drug responses by the altered ubiquitination of these two molecules. The inhibition of proteasomal protein degradation by epoxomicin increased the PfRFUL level, suggesting the degradation of PfRFUL by the proteasome pathways, whereas the inhibition of E3 ubiquitin ligase activities by JNJ26854165 reduced the PfRFUL level. This study reveals the potential mechanisms of PfRFUL in modifying the expression of drug transporters and their roles in parasite drug responses. PfRFUL could be a potential target for antimalarial drug development.

Background: The incidence of zoonotic Plasmodium knowlesi infections in humans is rising in Southeast Asia, leading to clinical studies to monitor the efficacy of anti-malarial treatments for knowlesi malaria. One of the key outcomes of anti-malarial drug efficacy is parasite clearance. For Plasmodium falciparum, parasite clearance is typically estimated using a two-stage method, that involves estimating parasite clearance for individual patients followed by pooling of individual estimates to derive population estimates. An alternative approach is Bayesian hierarchical modelling which simultaneously analyses all parasite-time patient profiles to determine parasite clearance. This study compared these methods for estimating parasite clearance in P. knowlesi treatment efficacy studies, with typically fewer parasite measurements per patient due to high susceptibility to anti-malarials., Methods: Using parasite clearance data from 714 patients with knowlesi malaria and enrolled in three trials, the Worldwide Antimalarial Resistance Network (WWARN) Parasite Clearance Estimator (PCE) standard two-stage approach and Bayesian hierarchical modelling were compared. Both methods estimate the parasite clearance rate from a model that incorporates a lag phase, slope, and tail phase for the parasitaemia profiles., Results: The standard two-stage approach successfully estimated the parasite clearance rate for 678 patients, with 36 (5%) patients excluded due to an insufficient number of available parasitaemia measurements. The Bayesian hierarchical estimation method was applied to the parasitaemia data of all 714 patients. Overall, the Bayesian method estimated a faster population mean parasite clearance (0.36/h, 95% credible interval [0.18, 0.65]) compared to the standard two-stage method (0.26/h, 95% confidence interval [0.11, 0.46]), with better model fits (compared visually). Artemisinin-based combination therapy (ACT) is more effective in treating P. knowlesi than chloroquine, as confirmed by both methods, with a mean estimated parasite clearance half-life of 2.5 and 3.6 h, respectively using the standard two-stage method, and 1.8 and 2.9 h using the Bayesian method., Conclusion: For clinical studies of P. knowlesi with frequent parasite measurements, the standard two-stage approach (WWARN's PCE) is recommended as this method is straightforward to implement. For studies with fewer parasite measurements per patient, the Bayesian approach should be considered. Regardless of method used, ACT is more efficacious than chloroquine, confirming the findings of the original trials., (© 2023. The Author(s).)

The ongoing COVID-19 pandemic has caused over six million deaths and huge economic burdens worldwide. Antivirals against its causative agent, SARS-CoV-2, are in urgent demand. Previously, we reported that heterocylic compounds, i.e., chloroquine (CQ) and hydroxychloroquine (HCQ), are potent in inhibiting SARS-CoV-2 replication in vitro . In this study, we discussed the syntheses of two novel heterocylic compounds: tert -butyl rel -4-(((3 R ,4 S )-3-(1 H -indol-3-yl)-1-oxo-2-propyl-1,2,3,4-tetrahydroisoquinolin-4-yl)methyl)piperazine-1-carboxylate ( trans - 1 ) and rel -(3 R ,4 S )-3-(1 H -indol-3-yl)-4-(piperazin-1-ylmethyl)-2-propyl-3,4-dihydroisoquinolin-1(2 H )-one ( trans - 2 ), which effectively suppressed authentic SARS-CoV-2 replication in Vero E6 cells. Compound trans - 1 showed higher anti-SARS-CoV-2 activity than trans - 2 , with a half maximal effective concentration (EC 50 ) of 3.15 μM and a selective index (SI) exceeding 63.49, which demonstrated comparable potency to CQ or HCQ. Additional anti-SARS-CoV-2 tests on Calu-3 human lung cells showed that trans - 1 efficiently inhibited viral replication (EC 50 = 2.78 μM; SI: > 71.94) and performed better than CQ (EC 50 = 44.90 μM; SI = 2.94). The time of an addition assay showed that the action mechanism of trans - 1 differed from that of CQ, as it mainly inhibited the post-entry viral replication in both Vero E6 and Calu-3 cells. In addition, the differences between the antiviral mechanisms of these novel compounds and CQ were discussed.

Pyrazole core represents a privilege scaffold in medicinal chemistry; a number of pyrazole compounds are endowed with various pharmacological activities in different therapeutic areas including antimalarial treatment. Supported by this evidence, a series of 5-anilino-3-(hetero)arylpyrazoles were evaluated for their antiplasmodial activity in in vitro assays. The compounds were synthesized according to regioselective and versatile protocols that combine active methylene reagents, aryl isothiocyanates and (substituted)hydrazines. The considered derivatives 2 allowed the definition of consistent structure-activity relationships and compounds 2b , e , k , l were identified as the most interesting derivatives of the series showing micromolar IC 50 values against chloroquine-sensitive and chloroquine-resistant Plasmodium strains. Additionally, the most active anilino-pyrazoles did not show any cytotoxicity against tumor and normal cells and were predicted to have favorable drug-like and pharmacokinetic properties.

A series of seventeen 4-chlorocinnamanilides and seventeen 3,4-dichlorocinnamanilides were characterized for their antiplasmodial activity. In vitro screening on a chloroquine-sensitive strain of Plasmodium falciparum 3D7/MRA-102 highlighted that 23 compounds possessed IC 50 < 30 µM. Typically, 3,4-dichlorocinnamanilides showed a broader range of activity compared to 4-chlorocinnamanilides. (2 E )- N -[3,5-bis(trifluoromethyl)phenyl]-3-(3,4-dichlorophenyl)prop-2-en-amide with IC 50 = 1.6 µM was the most effective agent, while the other eight most active derivatives showed IC 50 in the range from 1.8 to 4.6 µM. A good correlation between the experimental log k and the estimated clogP was recorded for the whole ensemble of the lipophilicity generators. Moreover, the SAR-mediated similarity assessment of the novel (di)chlorinated N -arylcinnamamides was conducted using the collaborative (hybrid) ligand-based and structure-related protocols. In consequence, an 'averaged' selection-driven interaction pattern was produced based in namely 'pseudo-consensus' 3D pharmacophore mapping. The molecular docking approach was engaged for the most potent antiplasmodial agents in order to gain an insight into the arginase-inhibitor binding mode. The docking study revealed that (di)chlorinated aromatic (C-phenyl) rings are oriented towards the binuclear manganese cluster in the energetically favorable poses of the chloroquine and the most potent arginase inhibitors. Additionally, the water-mediated hydrogen bonds were formed via carbonyl function present in the new N -arylcinnamamides and the fluorine substituent (alone or in trifluoromethyl group) of N -phenyl ring seems to play a key role in forming the halogen bonds.

Herein, upgraded chloroquine (CQ) derivatives capable of overcoming Plasmodium resistance and, at the same time, suppressing excessive immune response and risk of concurrent bacteremia were developed. Twelve new ferrocene- CQ hybrids tethered with a small azathia heterocycle (1,3-thiazolidin-4-one, 1,3-thiazinan-4-one, or 5-methyl-1,3-thiazolidin-4-one) were synthesized and fully characterized. All hybrids were evaluated for their in vitro antiplasmodial, antimicrobial, and immunomodulatory activities. Additional assays were performed on selected hybrids to gain insights into their mode of action. Although only hybrid 4a was more potent than the parent drug toward CQ -resistant Dd2 Plasmodium falciparum strain, several other hybrids (such as 6b , 6c , and 6d ) manifested substantially improved antimicrobial and immunomodulatory properties. Interesting structure-activity relationship data were obtained, hinting at future research for the development of new multitarget chemotherapies for malaria and other infectious diseases complicated by drug resistance, bacterial co-infection, and immune-driven pathology issues.

Inhibition of autophagy contributes to the pathophysiology of preeclampsia. Although chloroquine (CHQ) is an autophagy inhibitor, it can reduce the occurrence of preeclampsia in women with systemic lupus erythematosus. To clarify this important clinical question, this study aimed to address the safety of CHQ in trophoblast cells from the viewpoint of homeostasis, in which the anti-oxidative stress (OS) response and autophagy are involved. We used Western blotting to evaluate the protein levels in the trophoblast cells. The expression levels of heme oxygenase-1 (HO-1), an anti-OS enzyme, mediate resistance to OS induced by hydrogen peroxide (H 2 O 2 ) in trophoblast cell lines. Among the autophagy modulators, bafilomycin A1 (BAF), an autophagy inhibitor, but not autophagy activators, suppressed HO-1 expression in BeWo cells; CHQ did not suppress HO-1 expression in BeWo cells. To clarify the role of autophagy in HO-1 induction, we observed no difference in HO-1 induction by H 2 O 2 between autophagy-normal and autophagy-deficient cells. As for the mechanism of HO-1 induction by OS, BAF suppressed HO-1 induction by downregulating the expression of neighbor of BRCA1 gene 1 (NBR1) in the selective p62-NBR1-nuclear factor erythroid 2-related factor 2 (Nrf2) autophagy pathway. CHQ did not inhibit HO-1 expression by sustaining NBR1 expression in human villous tissues compared to BAF treatment. In conclusion, CHQ is a safer medicine than BAF for sustaining NBR1, which resist against OS in trophoblasts by connecting selective autophagy and the anti-OS response., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2022. Published by Elsevier B.V.)

Soft corals distributed across the Red Sea coasts are a rich source of diverse and bioactive natural products. Chemical probing of the Red Sea soft coral Litophyton arboreum led to isolation and structural characterization of an undescribed sesquiterpenoid, litoarbolide A ( 1 ), along with 14 previously reported metabolites ( 2-15 ). The chemical structures of the isolates were assigned based on NMR as well as high resolution electrospray ionization mass spectrometry (HR-ESI-MS) data. Litoarbolide A is supposed to be the biosynthetic precursor to other sesquiterpenoids, which formed via further post-translational modifications. Furthermore, these metabolites were evaluated for anti-malarial activity, where only the acyclic sesquiterpenoid of a sec -germacrane nucleus ( 7 ) showed an activity against chloroquine-sensitive (D6) and chloroquine-resistant (W2) strains of Plasmodium falciparum with IC 50 at 3.7 and 2.2 mg/mL, respectively. Moreover, the isolated metabolites were all non-toxic to the Vero cell line. These findings support the consideration of L. arboreum in further natural anti-malarial studies.

Background: Non-small cell lung carcinoma (NSCLC) is the most common cause of cancer-associated deaths worldwide. Though recent development in targeted therapy has improved NSCLC prognosis, yet there is an unmet need to identify novel causative factors and appropriate therapeutic regimen against NSCLCs., Methods and Results: In this study, we identify key molecular factors de-regulated in NSCLCs. Analyze their expression by real-time PCR and immunoblot; map their localization by immuno-fluorescence microscopy. We further propose an FDA approved drug, chloroquine (CQ) that affects the function of the molecular factors and hence can be repurposed as a therapeutic strategy against NSCLCs. Available NSCLC mutation data reflects a high probabilistic chance of patients harboring a p53 mutation, especially a gain of function (GOF)-R273H mutation. The GOF-P53 mutation enables the P53 protein to potentially interact with non-canonical protein partners facilitating oncogenesis. In this context, analysis of existing transcriptomic data from R273H-P53 expressing cells shows a concomitant up-regulation of Yes-associated protein (YAP) transcriptional targets and its protein partner TEAD1 in NSCLCs, suggesting a possible link between R273H-P53 and YAP. We therefore explored the inter-dependence of R273H-P53 and YAP in NSCLC cells. They were found to co-operatively regulate NSCLC proliferation. Genetic or pharmacological inhibition of YAP and GOF-P53 resulted in sensitization of NSCLC cells. Further analysis of pathways controlled by GOF-P53 and YAP showed that they positively regulate the cellular homeostatic process- autophagy to mediate survival. We hence postulated that a modulation of autophagy might be a potent strategy to curb proliferation. In accordance to above, autophagy inhibition, especially with the FDA-approved drug- chloroquine (CQ) resulted in cytoplasmic accumulation and reduced transcriptional activity of GOF-P53 and YAP, leading to growth arrest of NSCLC cells., Conclusion: Our study highlights the importance of GOF-P53 and YAP in NSCLC proliferation and proposes autophagy inhibition as an efficient strategy to attenuate NSCLC tumorigenesis., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

The use of an effective antimalarial drug is the cornerstone of malaria control. However, the development and spread of resistant Plasmodium falciparum strains have placed the global eradication of malaria in serious jeopardy. Molecular marker analysis constitutes the hallmark of the monitoring of Plasmodium drug-resistance. This study included 96 P. falciparum PCR-positive samples from southern Somalia. The P. falciparum chloroquine resistance transporter gene had high frequencies of K76T, A220S, Q271E, N326S, and R371I point mutations. The N86Y and Y184F mutant alleles of the P. falciparum multidrug resistance 1 gene were present in 84.7 and 62.4% of the isolates, respectively. No mutation was found in the P. falciparum Kelch-13 gene. This study revealed that chloroquine resistance markers are present at high frequencies, while the parasite remains sensitive to artemisinin (ART). The continuous monitoring of ART-resistant markers and in vitro susceptibility testing are strongly recommended to track resistant strains in real time.

Objectives: Chloroquine (CQ) is an antimalarial drug with a growing number of applications as recently demonstrated in attempts to treat Covid-19. For decades, it has been well known that skeletal and cardiac muscle cells might display vulnerability against CQ exposure resulting in the clinical manifestation of a CQ-induced myopathy. In line with the known effect of CQ on inhibition of the lysosomal function and thus cellular protein clearance, the build-up of autophagic vacuoles along with protein aggregates is a histological hallmark of the disease. Given that protein targets of the perturbed proteostasis are still not fully discovered, we applied different proteomic and immunological-based studies to improve the current understanding of the biochemical nature of CQ-myopathy., Methods: To gain a comprehensive understanding of the molecular pathogenesis of this acquired myopathy and to define proteins targets as well as pathophysiological processes beyond impaired proteolysis, utilising CQ-treated C2C12 cells and muscle biopsies derived from CQ-myopathy patients, we performed different proteomic approaches and Coherent Anti-Stokes Raman Scattering (CARS) microscopy, in addition to immunohistochemical studies., Results: Our combined studies confirmed an impact of CQ-exposure on proper protein processing/folding and clearance, highlighted changes in the interactome of p62, a known aggregation marker and hereby identified the Rett syndrome protein MeCP2 as being affected. Moreover, our approach revealed-among others-a vulnerability of the extracellular matrix, cytoskeleton and lipid homeostasis., Conclusion: We demonstrated that CQ exposure (secondarily) impacts biological processes beyond lysosomal function and linked a variety of proteins with known roles in the manifestation of other neuromuscular diseases., (© 2023 British Neuropathological Society.)

Background: Chloroquine (CQ) is an effective and safe antimalarial drug that is also used as a disease-modifying antirheumatic drug. Recent studies have shown that CQ can sensitize cancer cells to anti-cancer therapies., Methods: In this study, we investigated the molecular mechanisms underlying CQ-mediated chemosensitization in human colon carcinoma cells., Results: CQ prevented hypoxia-inducible factor (HIF)-1α protein induction in human colon carcinoma cells. CQ also suppressed HIF-1 activity, as represented by CQ inhibition of HIF-1-dependent luciferase activity and reduced induction of vascular endothelial growth factor. Under hypoxia, CQ restricted HIF-1α synthesis but did not affect HIF-1α transcription and protein stability. The hypoxic state activated ataxia telangiectasia and Rad3-related (ATR) kinase and increased the level of phosphorylated checkpoint kinase 1, a substrate of ATR kinase; however, this was prevented by CQ. An ATR kinase inhibitor suppressed the hypoxic induction of HIF-1α protein and was as effective as CQ. The cytotoxicity of 5-fluorouracil (5-FU), the first choice for the treatment of colorectal cancer, was attenuated under hypoxia. CQ enhanced the cytotoxicity of 5-FU treatment, which was mimicked by the transient transfection with HIF-1α siRNA., Conclusions: Under hypoxia, CQ-mediated sensitization of colon carcinoma HCT116 cells to 5-FU involves HIF-1 inhibition via ATR kinase suppression., (© 2022. The Author(s) under exclusive licence to Maj Institute of Pharmacology Polish Academy of Sciences.)

Recent evidence suggests that autophagy is a governed catabolic framework enabling the recycling of nutrients from injured organelles and other cellular constituents via a lysosomal breakdown. This mechanism has been associated with the development of various pathologic conditions, including cancer and neurological disorders; however, recently updated studies have indicated that autophagy plays a dual role in cancer, acting as a cytoprotective or cytotoxic mechanism. Numerous preclinical and clinical investigations have shown that inhibiting autophagy enhances an anticancer medicine's effectiveness in various malignancies. Autophagy antagonists, including chloroquine and hydroxychloroquine, have previously been authorized in clinical trials, encouraging the development of medication-combination therapies targeting the autophagic processes for cancer. In this review, we provide an update on the recent research examining the anticancer efficacy of combining drugs that activate cytoprotective autophagy with autophagy inhibitors. Additionally, we highlight the difficulties and progress toward using cytoprotective autophagy targeting as a cancer treatment strategy. Importantly, we must enable the use of suitable autophagy inhibitors and coadministration delivery systems in conjunction with anticancer agents. Therefore, this review briefly summarizes the general molecular process behind autophagy and its bifunctional role that is important in cancer suppression and in encouraging tumor growth and resistance to chemotherapy and metastasis regulation. We then emphasize how autophagy and cancer cells interacting with one another is a promising therapeutic target in cancer treatment.

Mitochondria is an important drug target for ailments ranging from neoplastic to neurodegenerative diseases and metabolic diseases. Here, we describe the synthesis of chloroquine analogs and show the results of mitochondrial ATP inhibition testing. The 2,4-dinitrobenzene-based analogs showed concentration-dependent mitochondrial (mito.) ATP inhibition. The most potent mito. ATP inhibitor was found to be N -(4-((2,4-Dinitrophenyl)amino)pentyl)- N -ethylacetamide ( 17 ).

Cold atmospheric plasma (CAP) is an intensively-studied approach for the treatment of malignant neoplasms. Various active oxygen and nitrogen compounds are believed to be the main cytotoxic effectors on biotargets; however, the comprehensive mechanism of CAP interaction with living cells and tissues remains elusive. In this study, we experimentally determined the optimal discharge regime (or semi-selective regime) for the direct CAP jet treatment of cancer cells, under which lung adenocarcinoma A549, A427 and NCI-H23 cells demonstrated substantial suppression of viability, coupled with a weak viability decrease of healthy lung fibroblasts Wi-38 and MRC-5. The death of CAP-exposed cancer and healthy cells under semi-selective conditions was caspase-dependent. We showed that there was an accumulation of lysosomes in the treated cells. The increased activity of lysosomal protease Cathepsin D, the transcriptional upregulation of autophagy-related MAPLC3B gene in cancer cells and the changes in autophagy-related proteins may have indicated the activation of autophagy. The addition of the autophagy inhibitor chloroquine (CQ) after the CAP jet treatment increased the death of A549 cancer cells in a synergistic manner and showed a low effect on the viability of CAP-treated Wi-38 cells. Downregulation of Drp1 mitochondrial protein and upregulation of PINK1 protein in CAP + CQ treated cells indicated that CQ increased the CAP-dependent destabilization of mitochondria. We concluded that CAP weakly activated pro-survival autophagy in irradiated cells, and CQ promoted CAP-dependent cell death due to the destabilization of autophagosomes formation and mitochondria homeostasis. To summarize, the combination of CAP treatment with CQ could be useful for the development of cold plasma-based antitumor approaches for clinical application.

Chloroquine (CQ) is a first-choice drug against malaria and autoimmune diseases. It has been co-administered with zinc against SARS-CoV-2 and soon dismissed because of safety issues. The structural features of Zn-CQ complexes and the effect of CQ on zinc distribution in cells are poorly known. In this study, state-of-the-art computations combined with experiments were leveraged to solve the structural determinants of zinc-CQ interactions in solution and the solid state. NMR, ESI-MS, and X-ray absorption and diffraction methods were combined with ab initio molecular dynamics calculations to address the kinetic lability of this complex. Within the physiological pH range, CQ binds Zn 2+ through the quinoline ring nitrogen, forming [Zn(CQH)Cl x (H 2 O) 3- x ] (3+)- x ( x = 0, 1, 2, and 3) tetrahedral complexes. The Zn(CQH)Cl 3 species is stable at neutral pH and at high chloride concentrations typical of the extracellular medium, but metal coordination is lost at a moderately low pH as in the lysosomal lumen. The pentacoordinate complex [Zn(CQH)(H 2 O) 4 ] 3+ may exist in the absence of chloride. This in vitro / in silico approach can be extended to other metal-targeting drugs and bioinorganic systems.

Background: Drug resistance is a current issue affecting parasites caused by Plasmodium. Therefore, researchers have expanded their studies on nanoparticles to find new and effective drugs that can treat drug-resistant strains. The present study systematically investigates the effect of different nanoparticles, including metal, polymer, and lipid nanoparticles, on Plasmodium berghei., Methods: In this study, English-language online literature was obtained from the databases Science Direct, PubMed, Scopus, Ovid, and Cochrane to conduct a systematic review. In the search, we used the keywords: (Plasmodium Berghei) AND (Malaria) AND (Parasitemia) AND (antimalarial activity) AND (nanoparticles) AND (Solid lipid NPS) AND (Nano lipid carriers) AND (Artemether) AND (Chloroquine) AND (intraperitoneal) AND (in vivo). Initially, a total of 160 studies were retrieved from the search. After removing duplicates, 80 studies remained. After reviewing the title and abstract of each study, 45 unrelated studies were eliminated., Results: The remaining 35 studies were thoroughly reviewed using the full texts. The final result was 21 studies that met the inclusion/exclusion criteria., Conclusion: Using these findings, we can conclude that various nanoparticles possess antiparasitic effects that may be applied to emerging and drug-resistant parasites. Together, these findings suggest that nanostructures may be used to design antiparasitic drugs that are effective against Plasmodium berghei., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The human beta-coronavirus strain, OC43, provides a useful model for testing the antiviral activity of various agents. We compared the activity of several antiviral drugs against OC43, including remdesivir, chloroquine, interferon (IFN)-β, IFN-λ1, and IFN-λ4, in two distinct cell types: human colorectal carcinoma cell line (HCT-8 cells) and normal human bronchial epithelial (NHBE) cells. We also tested whether these agents mediate additive, synergistic, or antagonistic activity against OC43 infection when used in combination. When used as single agents, remdesivir exhibited stronger antiviral activity than chloroquine, and IFN-β exhibited stronger activity than IFN-λ1 or IFN-λ4 against OC43 in both HCT-8 and NHBE cells. Anakinra (IL-1 inhibitor) and tocilizumab (IL-6 inhibitor) did not mediate any antiviral activity. The combination of IFN-β plus chloroquine or remdesivir resulted in higher synergy scores and higher expression of IFN-stimulated genes than did IFN-β alone. In contrast, the combination of remdesivir plus chloroquine resulted in an antagonistic interaction in NHBE cells. Our findings indicate that the combined use of IFN-β plus remdesivir or chloroquine induces maximal antiviral activity against human coronavirus strain OC43 in primary human respiratory epithelial cells. Furthermore, our experimental OC43 virus infection model provides an excellent method for evaluating the biological activity of antiviral drugs.

Tumour relapse, chemotherapy resistance, and metastasis continue to be unsolved issues in cancer therapy. A recent approach has been to scrutinise drugs used in the clinic for other illnesses and modify their structure to increase selectivity to cancer cells. Chloroquine (CQ) and hydroxychloroquine (HCQ), known antimalarials, have successfully treated autoimmune and neoplastic diseases. CQ and HCQ, well-known lysosomotropic agents, induce apoptosis, downregulate autophagy, and modify the tumour microenvironment. Moreover, they affect the Toll 9/NF-κB receptor pathway, activate stress response pathways, enhance p53 activity and CXCR4-CXCL12 expression in cancer cells, which would help explain their effects in cancer treatment. These compounds can normalise the tumourassociated vasculature, promote the activation of the immune system, change the phenotype of tumour-associated macrophages (from M2 to M1), and stimulate cancer-associated fibroblasts. We aim to review the historical aspects of CQ and its derivatives and the most relevant mechanisms that support the therapeutic use of CQ and HCQ for the treatment of cancer., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

As an autophagy inhibitor, chloroquine (CQ) showed anti-tumor effect on several types of cancer and paclitaxel (PTX) is widely used in the treatment of esophageal carcinoma patients, but chemoresistance remains a major hurdle for PTX application due to the cytoprotective autophagy. Therefore, the aim of this study was to investigate whether CQ could elevate the anti-tumor effect of PTX on esophageal carcinoma cell line EC109 and explore the potential molecular mechanisms. We confirmed the suppressive effect of PTX on EC109 by MTT, scratch test, transwell and soft agar assay. And, we detected the key proteins in Akt/mTOR pathway, as well as the autophagy marker LC3 and p62 through Western Blot. In addition, GFP-LC3 plasmid was transfected into EC109 cells to monitor the autophagosome after CQ and PTX treatment. Ultimately, we observed the alterations in the proliferation and colony formation abilities of EC109 after knocking down mTOR by shRNA. We confirmed PTX could suppress the proliferation, migration and colony formation (all P < 0.05) abilities of EC109, and CQ could sensitize the inhibition effect of PTX by inhibiting autophagy through Akt/mTOR pathway. Furthermore, inhibiting Akt/mTOR pathway initiated autophagy and enhanced the sensitivity of EC109 to CQ and PTX. In summary, we suggest CQ could be used as a potential chemosensitizer for PTX in esophageal carcinoma treatment.

Background: Compounds that target tumor epigenetic events are likely to constitute a prominent strategy for anticancer treatment. Histone deacetylase inhibitors (HDACis) have been developed as prospective candidates in anticancer drug development, and currently, many of them are under clinical investigation. We assessed the anticancer efficacy of a now hydroxamate-based HDACi, YF-343, in triple-negative breast cancer development and studied its potential mechanisms., Methods: YF-343 was estimated as a novel HDACi by the HDACi drug screening kit. The biological effects of YF-343 in a panel of breast cancer cell lines were analyzed by Western blot and flow cytometry. YF-343 exhibited notable cytotoxicity, promoted apoptosis, and induced cell cycle arrest. Furthermore, it also induced autophagy, which plays a pro-survival role in breast cancer cells., Results: The combination of YF-343 with an autophagy inhibitor chloroquine (CQ) significantly suppressed breast tumor progression as compared to the YF-343 treatment alone both in vitro and in vivo . Mechanistically, the molecular mechanism of YF-343 on autophagy was elucidated by gene chip expression profiles, qPCR analysis, luciferase reporter gene assay, chromatin immunoprecipitation assays, immunohistochemical analysis, and other methods. E2F7, a transcription factor, promoted the expression of ATG2A via binding to the ATG2A promoter region and then induced autophagy in triple-negative breast cancer cells treated with YF-343., Conclusion: Our studies have illustrated the mechanisms for potential action of YF-343 on tumor growth in breast cancer models with pro-survival autophagy. The combination therapy of YF-343 and CQ maybe a promising strategy for breast cancer therapy., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

In this study, a structurally guided pharmacophore hybridization strategy is used to combine the two key structural scaffolds, para -aminobenzoic acid (PABA), and 1,3,5 triazine in search of new series of antimalarial agents. A combinatorial library of 100 compounds was prepared in five different series as [ 4A ( 1 - 22 ), 4B ( 1 - 21 ), 4 C ( 1 - 20 ), 4D ( 1 - 19 ) and 4E ( 1 - 18 )] using different primary and secondary amines, from where 10 compounds were finally screened out through molecular property filter analysis and molecular docking study as promising PABA substituted 1,3,5-triazine scaffold as an antimalarial agent. The docking results showed that compounds 4A12 and 4A20 exhibited good binding interaction with Phe58, IIe164, Ser111, Arg122, Asp54 (-424.19 to -360.34 kcal/mol) and Arg122, Phe116, Ser111, Phe58 (-506.29 to -431.75 kcal/mol) against wild (1J3I) and quadruple mutant (1J3K) type of Pf -DHFR. These compounds were synthesized by conventional as well as microwave-assisted synthesis and characterized by different spectroscopic methods. In-vitro antimalarial activity results indicated that two compounds 4A12 and 4A20 showed promising antimalarial activity against chloroquine-sensitive (3D7) and chloroquine-resistant (Dd2) strains of Plasmodium falciparum with IC 50 (1.24-4.77 μg mL -1 ) and (2.11-3.60 μg mL -1 ). These hybrid PABA substituted 1,3,5-triazine derivatives might be used in the lead discovery towards a new class of Pf -DHFR inhibitors.Communicated by Ramaswamy H. Sarma.

Background: Oxidative stress is an important mechanism in liver ischemia/reperfusion (I/R) injury. Hepatocyte apoptosis and proliferation occur in parallel with liver I/R injury, and the degree of apoptosis and proliferation determines the effects on hepatocytes. Vitamin D receptor (VDR) can lessen liver I/R injury, but previous studies focused mostly on inflammation and immunity., Methods: H 2 O 2 was used to induce hepatocyte injury. Before treatment with H 2 O 2 , Hep-3B cells were pretreated with paricalcitol (PC) and siRNA-VDR. Rapamycin and chloroquine were also applied in the study., Results: The number of apoptotic cells was measured with an annexin V (AV) -fluorescein isothiocyanate apoptosis detection kit. Expression of proteins was measured by western blotting. As compared with the H 2 O 2 +Hep-3B group, levels of AV/PI, cleaved caspase-3, and p62 were lower, and expression levels of Bcl-2, proliferating cell nuclear antigen, and VDR were higher, in the PC+H 2 O 2 +Hep-3B group. When the VDR gene was silenced by siRNA-VDR in the siRNA-VDR+H 2 O 2 +Hep-3B group, expressions of AV/PI, cleaved caspase-3, and p62 were upregulated, and expressions of Bcl-2, proliferating cell nuclear antigen, and VDR were downregulated, as compared with values for the siRNA-NC+H 2 O 2 +Hep-3B group. Treatment with rapamycin or chloroquine partially reversed the effect of PC and siRNA-VDR on apoptosis and proliferation., Conclusions: VDR mediates hepatocyte apoptosis and proliferation through autophagy.

This work deals with the study of the synthesis of new bis-indole analogues with a phenyl linker derived from indole phytoalexins. Synthesis of target bis-indole thiourea linked by a phenyl linker was achieved by the reaction of [1-(tert-butoxycarbonyl)indol-3-yl]methyl isothiocyanate with p-phenylenediamine. By replacing the sulfur of the thiocarbonyl group in bis-indole thiourea with oxygen using mesityl nitrile oxide, a bis-indole homodimer with a urea group was obtained. A cyclization protocol utilizing bis-indole thiourea and methyl bromoacetate was applied to synthesize a bis-indole homodimer with a thiazolidin-4-one moiety. Bis-indole homodimers derived from 1-methoxyspirobrassinol methyl ether were prepared by bromospirocyclization methodology. Among the synthesized analogues, compound 49 was selected for further study. To evaluate the mode of the mechanism of action, we used flow cytometry, Western blot, and spectroscopic analyses. Compound 49 significantly inhibited the proliferation of lung cancer cell line A549 with minimal effects on the non-cancer cells. We also demonstrated that compound 49 induced autophagy through the upregulation of Beclin-1, LC3A/B, Atg7 and AMPK and ULK1. Furthermore, chloroquine (CQ; an autophagy inhibitor) in combination with compound 49 decreased cell proliferation and induced G1 cell cycle arrest and apoptosis. Compound 49 also caused GSH depletion and significantly potentiated the antiproliferative effect of cis-platin.

Background: Despite significant progress in eliminating malaria from the state of Odisha, India, the disease is still considered endemic. Artesunate plus sulfadoxine-pyrimethamine (AS + SP) has been introduced since 2010 as first-line treatment for uncomplicated Plasmodium falciparum malaria. This study aimed to investigate the prevalence of mutations associated with resistance to chloroquine (CQ), sulfadoxine-pyrimethamine (SP), and artesunate (ART) in P. falciparum parasites circulating in the state., Methods: A total of 239 isolates of P. falciparum mono infection were collected during July 2018-November 2020 from the four different geographical regions of the state. Genomic DNA was extracted from 200 µL of venous blood and amplified using nested polymerase chain reaction. Mutations on gene associated with CQ (Pfcrt and Pfmdr1) were assessed by PCR amplification and restriction fragment length polymorphism, artemisinin (Pfk13) gene by DNA sequencing and SP (Pfdhfr and Pfdhps) genes by allele-specific polymerase chain reaction (AsPCR)., Results: The point mutation in Pfcrt (K76T) was detected 2.1%, in Pfmdr1 (N86Y) 3.4%, and no mutations were found in Pfkelch13 propeller domain. Prevalence of Pfdhfr, Pfdhps and Pfhdfr-Pfdhps (two locus) gene mutations were 50.43%, 47.05% and 49.79% respectively. The single, double, triple and quadruple point mutations in Pfdhfr gene was 11.2%, 8.2%, 17.2% and 3.4% while, in Pfdhps gene was 10.9%,19.5%, 9.5% and 2.7% respectively. Of the total 13 haplotypes found in Pfdhfr, 8 were detected for the first time in the state and of the total 26 haplotypes found in Pfdhps, 7 were detected for the fisrt time in the state. The linked quintuple mutation Pfdhfr (N51I-C59R-S108N)-Pfdhps (A437G-K540E) responsible for clinical failure (RIII level of resistance) of SP resistance and A16V-S108T mutation in Pfdhfr responsible for cycloguanil was absent., Conclusion: The study has demonstrated a low prevalence of CQ resistance alleles in the study area. Despite the absence of the Pfkelch13 mutations, high prevalence of Pfdhfr and Pfdhps point mutations undermine the efficacy of SP partner drug, thereby threatening the P. falciparum malaria treatment policy. Therefore, continuous molecular and in vivo monitoring of ACT efficacy is warranted in Odisha., (© 2022. The Author(s).)

Previous studies suggest that 3',5'-dihydro-2'H-spiro[indoline-3,1'-isoquinolin]-2-ones (DSIIQs [spiroquindolones]) are multitarget antiplasmodial agents that combine the actions of spiroindolone and naphthylisoquinoline antimalarial agents. In this study, 12 analogues of compound (±)-5 (moxiquindole), the prototypical spiroquindolone, were synthesized and tested for antiplasmodial activity. Compound (±)-11 (a mixture of compounds 11a and 11b), the most potent analogue, displayed low-nanomolar activity against P. falciparum chloroquine-sensitive 3D7 strain (50% inhibitory concentration [IC 50 ] for 3D7 = 21 ± 02 nM) and was active against all major erythrocytic stages of the parasite life cycle (ring, trophozoite, and schizont); it also inhibited hemoglobin metabolism and caused extensive vacuolation in parasites. In drug-resistant parasites, compound (±)-11 exhibited potent activity (IC 50 for Dd2 = 58.34 ± 2.04 nM) against the P. falciparum multidrug-resistant Dd2 strain, and both compounds (±)-5 and (±)-11 displayed significant cross-resistance against the P. falciparum ATP4 mutant parasite Dd2 SJ733 but not against the Dd2 KAE609 strain. In mice, both compounds (±)-5 and (±)-11 displayed dose-dependent reduction of parasitemia with suppressive 50% effective dose (ED 50 ) values of 0.44 and 0.11 mg/kg of body weight, respectively. The compounds were also found to be curative in vivo and are thus worthy of further investigation.

We generated highly chloroquine (CQ)-resistant (ResCQ) Plasmodium yoelii parasites by stepwise exposure to increasing concentrations of CQ and CQ-sensitive parasites (SenCQ) by parallel mock treatments. No mutations in genes that are associated with drug resistance were detected in ResCQ clones. Autophagy-related genes were highly upregulated in SenCQ compared to ResCQ parasites during CQ treatment. This indicates that CQ resistance can be developed in the malaria parasite by the inhibition of autophagy as an alternative drug resistance mechanism.

The discovery and development of transmission-blocking therapies challenge malaria elimination and necessitate standard and reproducible bioassays to measure the blocking properties of antimalarial drugs and candidate compounds. Most of the current bioassays evaluating the transmission-blocking activity of compounds rely on laboratory-adapted Plasmodium strains. Transmission-blocking data from clinical gametocyte isolates could help select novel transmission-blocking candidates for further development. Using freshly collected Plasmodium falciparum gametocytes from asymptomatic individuals, we first optimized ex vivo culture conditions to improve gametocyte viability and infectiousness by testing several culture parameters. We next pre-exposed ex vivo field-isolated gametocytes to chloroquine, dihydroartemisinin, primaquine, KDU691, GNF179, and oryzalin for 48 h prior to direct membrane feeding. We measured the activity of the drug on the ability of gametocytes to resume the sexual life cycle in Anopheles after drug exposure. Using 57 blood samples collected from Malian volunteers aged 6 to 15 years, we demonstrate that the infectivity of freshly collected field gametocytes can be preserved and improved ex vivo in a culture medium supplemented with 10% horse serum at 4% hematocrit for 48 h. Moreover, our optimized drug assay displays the weak transmission-blocking activity of chloroquine and dihydroartemisinin, while primaquine and oryzalin exhibited a transmission-blocking activity of ~50% at 1 μM. KDU691 and GNF179 both interrupted Plasmodium transmission at 1 μM and 5 nM, respectively. This new approach, if implemented, has the potential to accelerate the screening of compounds with transmission-blocking activity.