Disulfiram: Mechanisms, Applications, and Challenges

Rennar G.A., Gallinger T.L., Mäder P., Lange-Grünweller K., Haeberlein S., Grünweller A., Grevelding C.G., Schlitzer M.

Schistosomiasis is a neglected tropical disease with more than 200 million new infections per year. It is caused by parasites of the genus Schistosoma and can lead to death if left untreated. Currently, only two drugs are available to combat schistosomiasis: praziquantel and, to a limited extent, oxamniquine. However, the intensive use of these two drugs leads to an increased probability of the emergence of resistance. Thus, the search for new active substances and their targeted development are mandatory. In this study the substance class of "dithiocarbamates" and their potential as antischistosomal agents is highlighted. These compounds are derived from the basic structure of the human aldehyde dehydrogenase inhibitor disulfiram (tetraethylthiuram disulfide, DSF) and its metabolites. Our compounds revealed promising activity (in vitro) against adults of Schistosoma mansoni, such as the reduction of egg production, pairing stability, vitality, and motility. Moreover, tegument damage as well as gut dilatations or even the death of the parasite were observed. We performed detailed structure-activity relationship studies on both sides of the dithiocarbamate core leading to a library of approximately 300 derivatives (116 derivatives shown here). Starting with 100 μm we improved antischistosomal activity down to 25 μm by substitution of the single bonded sulfur atom for example with different benzyl moieties and integration of the two residues on the nitrogen atom into a cyclic structure like piperazine. Its derivatization at the 4-nitrogen with a sulfonyl group or an acyl group led to the most active derivatives of this study which were active at 10 μm. In light of this SAR study, we identified 17 derivatives that significantly reduced motility and induced several other phenotypes at 25 μm, and importantly five of them have antischistosomal activity also at 10 μm. These derivatives were found to be non-cytotoxic in two human cell lines at 100 μm. Therefore, dithiocarbamates seem to be interesting new candidates for further antischistosomal drug development.

Lajarin-Reinares M., Martinez-Esteve E., Pena-Rodríguez E., Cañellas-Santos M., Bulut S., Karabelas K., Clauss A., Nieto C., Mallandrich M., Fernandez-Campos F.

Scabies and hair lice are parasitic diseases that affect human skin and hair, respectively. The incidence and resistances of these infections are increasing. Tenutex® (disulfiram and benzyl benzoate emulsion) is an alternative to standard insecticides to avoid resistances. The aim of the work is to evaluate the transdermal absorption and the in vitro efficacy against scabies and hair lice after different exposition times. Dermatomed human skin was used to assess the dermal absorption using a validated High Performance Liquid Chromatography (HPLC) method. HEK001 keratinocytes were used to evaluate the cytotoxicity of benzyl benzoate. Only benzyl benzoate was able to cross the skin, but it did not show cytotoxicity at any of the tested concentrations. The product efficacy was tested on Psoroptes ovis after direct contact and after administration on sheep skin explants at different contact times. Permethrin/malathion-resistant strains of Pediculus humanis capitis adults and eggs were directly exposed to Tenutex, and the vitality and hatchability, respectively, were evaluated. The anti-scabies study demonstrated that exposure for 6 or 24 h completely eradicated the parasite. The pediculicidal activity of Tenutex exhibited superior efficacy than standard treatment on resistant lice. The positive results obtained suggest that Tenutex® is a good treatment option, especially in drug resistance situations.

Almeida-Silva J., Menezes D.S., Fernandes J.M., Almeida M.C., Vasco-dos-Santos D.R., Saraiva R.M., Viçosa A.L., Perez S.A., Andrade S.G., Suarez-Fontes A.M., Vannier-Santos M.A.

Chagas disease (CD) affects at least 6 million people in 21 South American countries besides several thousand in other nations all over the world. It is estimated that at least 14,000 people die every year of CD. Since vaccines are not available, chemotherapy remains of pivotal relevance. About 30% of the treated patients cannot complete the therapy because of severe adverse reactions. Thus, the search for novel drugs is required. Here we tested the benznidazole (BZ) combination with the repositioned drug disulfiram (DSF) and its derivative diethyldithiocarbamate (DETC) upon Trypanosoma cruzi in vitro and in vivo. DETC-BZ combination was synergistic diminishing epimastigote proliferation and enhancing selective indexes up to over 10-fold. DETC was effective upon amastigotes of the BZ- partially resistant Y and the BZ-resistant Colombiana strains. The combination reduced proliferation even using low concentrations (e.g., 2.5 µM). Scanning electron microscopy revealed membrane discontinuities and cell body volume reduction. Transmission electron microscopy revealed remarkable enlargement of endoplasmic reticulum cisternae besides, dilated mitochondria with decreased electron density and disorganized kinetoplast DNA. At advanced stages, the cytoplasm vacuolation apparently impaired compartmentation. The fluorescent probe H2-DCFDA indicates the increased production of reactive oxygen species associated with enhanced lipid peroxidation in parasites incubated with DETC. The biochemical measurement indicates the downmodulation of thiol expression. DETC inhibited superoxide dismutase activity on parasites was more pronounced than in infected mice. In order to approach the DETC effects on intracellular infection, peritoneal macrophages were infected with Colombiana trypomastigotes. DETC addition diminished parasite numbers and the DETC-BZ combination was effective, despite the low concentrations used. In the murine infection, the combination significantly enhanced animal survival, decreasing parasitemia over BZ. Histopathology revealed that low doses of BZ-treated animals presented myocardial amastigote, not observed in combination-treated animals. The picrosirius collagen staining showed reduced myocardial fibrosis. Aminotransferase de aspartate, Aminotransferase de alanine, Creatine kinase, and urea plasma levels demonstrated that the combination was non-toxic. As DSF and DETC can reduce the toxicity of other drugs and resistance phenotypes, such a combination may be safe and effective.

Yazdi A.K., Pakarian P., Perveen S., Hajian T., Santhakumar V., Bolotokova A., Li F., Vedadi M.

SARS-CoV-2 non-structural protein 13 (nsp13) is a highly conserved helicase and RNA 5′-triphosphatase. It uses the energy derived from the hydrolysis of nucleoside triphosphates for directional movement along the nucleic acids and promotes the unwinding of double-stranded nucleic acids. Nsp13 is essential for replication and propagation of all human and non-human coronaviruses. Combined with its defined nucleotide binding site and druggability, nsp13 is one of the most promising candidates for the development of pan-coronavirus therapeutics. Here, we report the development and optimization of bioluminescence assays for kinetic characterization of nsp13 ATPase activity in the presence and absence of single-stranded DNA. Screening of a library of 5000 small molecules in the presence of single-stranded DNA resulted in the discovery of six nsp13 small-molecule inhibitors with IC50 values ranging from 6 ± 0.5 to 50 ± 6 μM. In addition to providing validated methods for high-throughput screening of nsp13 in drug discovery campaigns, the reproducible screening hits we present here could potentially be chemistry starting points toward the development of more potent and selective nsp13 inhibitors, enabling the discovery of antiviral therapeutics.

Mego M., Svetlovska D., Angelis V D., Kalavska K., Lesko P., Makovník M., Obertova J., Orszaghova Z., Palacka P., Rečková M., Rejlekova K., Z S., Mardiak J., Chovanec M.

Multiple relapsed/refractory germ cell tumor (GCT) patients have extremely poor prognosis. Cisplatin resistant testicular GCTs overexpress aldehyde-dehydrogenase (ALDH) isoforms and inhibition of ALDH activity by disulfiram is associated with reconstitution of cisplatin sensitivity in vitro as well as in animal model. This study aimed to determine the efficacy and toxicity of ALDH inhibitor disulfiram in combination with cisplatin in patients with multiple relapsed/refractory GCTs. Disulfiram was administered at a dose of 400 mg daily until progression or unacceptable toxicity, cisplatin was administered at dose 50 mg/m2 day 1 and 2, every 3 weeks. Twelve evaluable patients had to be enrolled into the first cohort, and if 0 of 12 patients had treatment response, the study was to be terminated. The results of the first stage of the trial are presented in this report. Twelve patients with multiple relapsed/refractory GCTs were enrolled in the phase II study from May 2019 to September 2021. Median number of treatment cycles was 2 (range 1–6). None of patients achieved objective response to treatment, therefore the study was terminated in first stage. Median progression-free survival was 1.4 months, 95% CI (0.7–1.5 months), and median overall survival was 2.9 months 95% CI (1.5–4.7 months). Disease stabilization for at least 3 months was observed in 2 (16.7%) patients. Treatment was well tolerated, however, 5 (41.7%) of patients experienced grade 3/4 fatigue, 4 (33.3%) thrombocytopenia, 3 (25.0%) anemia, while 2 (16.7%) experienced neutropenia, nausea and infection. This study failed to achieve its primary endpoint and our data suggest limited efficacy of disulfiram in restoring sensitivity to cisplatin in multiple relapsed/refractory GCTs.

Cabello F.C., Embers M.E., Newman S.A., Godfrey H.P.

The annual incidence of Lyme disease, caused by tick-transmitted Borreliella burgdorferi , is estimated to be at least 476,000 cases in the United States and many more worldwide. Ten to 20% of antimicrobial-treated Lyme disease patients display posttreatment Lyme disease syndrome (PTLDS), a clinical complication whose etiology and pathogenesis remain uncertain.

Zhang C., Niu H., Wan C., Yu X., Xin G., Zhu Y., Wei Z., Li F., Wang Y., Zhang K., Li S., Dong Y., Li Y., Huang W.

Acute pancreatitis (AP) is one of the most common causes of hospitalization for gastrointestinal diseases, with high morbidity and mortality. Endoplasmic reticulum stress (ERS) and Gasdermin D (GSDMD) mediate AP, but little is known about their mutual influence on AP. Diosgenin has excellent anti-inflammatory and antioxidant effects. This study investigated whether Diosgenin derivative D (Drug D) inhibits L-arginine-induced acute pancreatitis through meditating GSDMD in the endoplasmic reticulum (ER). Our studies were conducted in a mouse model of L-arginine-induced AP as well as in an in vitro model on mouse pancreatic acinar cells. The GSDMD accumulation in ER was found in this study, which caused ERS of acinar cells. GSDMD inhibitor Disulfiram (DSF) notably decreased the expression of GSDMD in ER and TXNIP/HIF-1α signaling. The molecular docking study indicated that there was a potential interaction between Drug D and GSDMD. Our results showed that Drug D significantly inhibited necrosis of acinar cells dose-dependently, and we also found that Drug D alleviated pancreatic necrosis and systemic inflammation by inhibiting the GSDMD accumulation in the ER of acinar cells via the TXNIP/HIF-1α pathway. Furthermore, the level of p-IRE1α (a marker of ERS) was also down-regulated by Drug D in a dose-dependent manner in AP. We also found that Drug D alleviated TXNIP up-regulation and oxidative stress in AP. Moreover, our results revealed that GSDMD-/- mitigated AP by inhibiting TXNIP/HIF-1α. Therefore, Drug D, which is extracted from Dioscorea zingiberensis, may inhibit L-arginine-induced AP by meditating GSDMD in the ER by the TXNIP /HIF-1α pathway.

Meewan I., Kattoula J., Kattoula J.Y., Skinner D., Fajtová P., Giardini M.A., Woodworth B., McKerrow J.H., Lage de Siqueira-Neto J., O’Donoghue A.J., Abagyan R.

One inhibitor of the main SARS-CoV-2 protease has been approved recently by the FDA, yet it targets only SARS-CoV-2 main protease (Mpro). Here, we discovered inhibitors containing thiuram disulfide or dithiobis-(thioformate) tested against three key proteases involved in SARS-CoV-2 replication, including Mpro, SARS-CoV-2 papain-like protease (PLpro), and human cathepsin L. The use of thiuram disulfide and dithiobis-(thioformate) covalent inhibitor warheads was inspired by an idea to find a better alternative than disulfiram, an approved treatment for chronic alcoholism that is currently in phase 2 clinical trials against SARS-CoV-2. Our goal was to find more potent inhibitors that target both viral proteases and one essential human protease to reduce the dosage, improve the efficacy, and minimize the adverse effects associated with these agents. We found that compounds coded as RI175, RI173, and RI172 were the most potent inhibitors in an enzymatic assay against SARS-CoV-2 Mpro, SARS-CoV-2 PLpro, and human cathepsin L, with IC50s of 300, 200, and 200 nM, which is about 5-, 19-, and 11-fold more potent than disulfiram, respectively. In addition, RI173 was tested against SARS-CoV-2 in a cell-based and toxicity assay and was shown to have a greater antiviral effect than disulfiram. The identified compounds demonstrated the promising potential of thiuram disulfide or dithiobis-(thioformate) as a reactive functional group in small molecules that could be further developed for treatment of the COVID-19 virus or related variants.

Bahji A., Bach P., Danilewitz M., Crockford D., Devoe D.J., el-Guebaly N., Saitz R.

We aimed to determine medications' comparative efficacy and safety for adults with alcohol use disorders.We searched eleven electronic data sources for randomized clinical trials with at least 4 weeks of treatment reporting on alcohol consumption (total abstinence and reduced heavy drinking), dropouts, and dropouts due to adverse events. We conducted network meta-analyses using random-effects, frequentist models, and calculated summary rate ratios (RRs) with 95% confidence intervals (CIs).We included 156 trials (N = 27,334). Nefazodone (RR = 2.11; 95% CI, 1.42-3.13), aripiprazole (RR = 1.97; 95% CI, 1.36-2.88), carbamazepine (RR = 1.85; 95% CI, 1.03-3.32), and nalmefene (RR = 1.17; 95% CI, 1.01-1.35) were associated with the most dropouts. Baclofen (RR = 0.83; 95% CI, 0.70-0.97) and pregabalin (RR = 0.63; 95% CI, 0.43-0.94) caused fewer dropouts than placebo. Nalmefene (RR = 3.26; 95% CI, 2.34-4.55), fluvoxamine (RR = 3.08; 95% CI, 1.59-5.94), and topiramate (RR=2.18; 95% CI, 1.36-3.51) caused more dropouts from adverse events over placebo. Gamma-hydroxy-butyrate (RR = 1.90; 95% CI, 1.03-3.53), baclofen (RR = 1.80; 95% CI, 1.39-2.34), disulfiram (RR = 1.71; 95% CI, 1.39-2.10), gabapentin (RR = 1.66; 95% CI, 1.04-2.67), acamprosate (RR = 1.33; 95% CI, 1.15-1.54), and oral naltrexone (RR = 1.15; 95% CI, 1.01-1.32) improved total abstinence over placebo (Fig. 3C). For reduced heavy drinking, disulfiram (RR = 0.19; 95% CI, 0.10-0.35), baclofen (RR = 0.72; 95% CI, 0.57-0.91), acamprosate (RR = 0.78; 95% CI, 0.70-0.86), and oral naltrexone (RR = 0.81; 95% CI, 0.73-0.90) were efficacious against placebo.The current meta-analyses provide evidence that several medications for AUDs are effective and safe and encourage the expanded use of these medications in the clinical setting. Our review found that acamprosate (2-3 g/d), disulfiram (250-500 mg/d), baclofen (30 mg/d), and oral naltrexone (50 mg/d) had the best evidence for improving abstinence and heavy drinking for patients with AUD.CRD42020208946.

Tan H., Ma C., Wang J.

The COVID-19 pandemic spurred a broad interest in antiviral drug discovery. The SARS-CoV-2 main protease (Mpro) and papain-like protease (PLpro) are attractive antiviral drug targets given their vital roles in viral replication and modulation of host immune response. Structurally disparate compounds were reported as Mpro and PLpro inhibitors from either drug repurposing or rational design. Two polyphenols dieckol and 1,2,3,4,6-pentagalloylglucose (PGG) were recently reported as SARS-CoV-2 Mpro inhibitors. With our continuous interest in studying the mechanism of inhibition and resistance of Mpro inhibitors, we report herein our independent validation/invalidation of these two natural products. Our FRET-based enzymatic assay showed that neither dieckol nor PGG inhibited SARS-CoV-2 Mpro (IC50 > 20 µM), which is in contrary to previous reports. Serendipitously, PGG was found to inhibit the SARS-CoV-2 PLpro with an IC50 of 3.90 µM. The binding of PGG to PLpro was further confirmed in the thermal shift assay. However, PGG was cytotoxic in 293T-ACE2 cells (CC50 = 7.7 µM), so its intracellular PLpro inhibitory activity could not be quantified by the cell-based Flip-GFP PLpro assay. In addition, we also invalidated ebselen, disulfiram, carmofur, PX12, and tideglusib as SARS-CoV-2 PLpro inhibitors using the Flip-GFP assay. Overall, our results call for stringent hit validation, and the serendipitous discovery of PGG as a putative PLpro inhibitor might worth further pursuing.

Wang X., Ye H., Yang S., Sha X., Wang X., Zhang T., Chen R., Xiao W., Yang H.

Fibrosis of extraocular muscles (EOMs) is a marker of end-stage in Graves’ orbitopathy (GO). To determine the antifibrotic and anti-inflammatory therapeutic effects and the underlying molecular mechanisms of disulfiram (DSF) on perimysial orbital fibroblasts (pOFs) in a GO model in vitro, primary cultures of pOFs from eight patients with GO and six subjects without GO (NG) were established. CCK-8 and EdU assays, IF, qPCR, WB, three-dimensional collagen gel contraction assays, cell scratch experiments, and ELISAs were performed. After TGF-β1 stimulation of pOFs, the proliferation rate of the GO group but not the NG group increased significantly. DSF dose-dependently inhibited the proliferation, contraction, and migration of pOFs in the GO group. Additionally, DSF dose-dependently inhibited fibrosis and extracellular matrix production markers (FN1, COL1A1, α-SMA, CTGF) at the mRNA and protein levels. Furthermore, DSF mediates antifibrotic effects on GO pOFs partially through the ERK-Snail signaling pathway. In addition, DSF attenuated HA production and suppressed inflammatory chemokine molecule expression induced by TGF-β1 in GO pOFs. In this in vitro study, we demonstrate the inhibitory effect of DSF on pOFs fibrosis in GO, HA production, and inflammation. DSF may be a potential drug candidate for preventing and treating tissue fibrosis in GO.

Meneguello J.E., Murase L.S., de Souza J.V., de Oliveira C.G., Ghiraldi-Lopes L.D., Teixeira J.J., Scodro R.B., Ferracioli K.R., Siqueira V.L., Campanerut-Sá P.A., Cardoso R.F.

The objective of this systematic review was to retrieve and examine published studies related to in vitro and in vivo evaluation of disulfiram for the treatment of bacterial infections. Five scientific databases (PubMed, Embase, Scopus, Web of Science, and Latin American and Caribbean Health Sciences Literature) were searched to retrieve the maximum literature regarding the study's aim. The search strategy retrieved a total of 870 studies, of which 31 were included and 19 approached disulfiram as the primary aim and 12 included it as a secondary finding from other investigational objectives. The evidence pointed out five main aspects of pre-clinical testing regarding disulfiram antibacterial activity, namely spectrum of antimicrobial action, drug combinations, intracellular studies, animal studies and bacterial targets. Findings to emerge from this study are the observed potential of disulfiram as a non-antibiotic drug being proposed as a potential drug to contribute to the treatment of bacterial diseases usually with few treatment alternatives in the context of drug resistance. We evaluated the potency and selectivity of disulfiram, which indeed until now shows potential to be explored for use as an adjunctive chemical to antimicrobial ones. Even with the level of evidence being reserved, the potential of combining disulfiram with other drugs, already used or new to be used for the treatment of mycobacterial diseases, as well as its likely immunomodulatory effect, deserve to be further investigated. Furthermore, the copper-dependent mode of action in Gram-positive bacteria is an alternative to be explored in drug design or repurposing of chemicals.

Serafin M.B., Foletto V.S., da Rosa T.F., Bottega A., Viana A.R., Franco L.N., de Lima Marion S., da Mota A.D., de Paula B.R., Krause L.M., Finatto L.J., Hörner M., Hörner R.

The identification of molecules that exhibit potent antibacterial activity and are capable of circumventing resistance mechanisms is an unmet need. The repositioning of approved drugs is considered an advantageous alternative in this case, and has gained prominence. In addition, drug synergism can reduce morbidity and mortality in the treatment of nosocomial infections caused by multi-drug resistant microorganisms (MDR). Whole cell growth inhibition assays were used to define the in vitro antibacterial activity of disulfiram against two standard American Type Culture Collection (ATCC) strains and 35 clinical isolates of vancomycin-resistant enterococci (VRE). The ability of disulfiram to synergize with vancomycin was determined by fractional inhibitory concentration index, preceded by the checkerboard test. The cytotoxicity of drugs alone and in combination was tested against Raw 264.7 cells. Disulfiram exhibited potent antibacterial activity against VRE (MIC 16–64 µg mL−1). Results: Associated with vancomycin, disulfiram it had a reduction in MIC of up to 64 times, with values of 0.5–4 µg mL−1. Vancomycin had a MIC of 128–1024 µg mL−1; combined, reduced this value by up to 124 times (8 µg mL−1), with synergy occurring against all strains. Disulfiram and vancomycin alone and in combination did not show cytotoxicity against the eukaryotic cell line. Based on these results, we suggest that the redirection of disulfiram may be promising in the treatment of infections caused by VRE, since it was able to potentiate the activity of vancomycin against the strains, being able to act as an adjuvant in cases of serious infections caused by Enterococcus.

Zhang T., Kephart J., Bronson E., Anand M., Daly C., Spasojevic I., Bakthavatsalam S., Franz K., Berg H., Karachaliou G.S., James O.G., Howard L., Halabi S., Harrison M.R., Armstrong A.J., et. al.

In preclinical models of prostate cancer (PC), disulfiram (DSF) reduced tumor growth only when co-administered with copper (Cu), and Cu uptake in tumors is partially regulated by androgen-receptor signaling. However, prior trials of DSF in PC used DSF as monotherapy.To assess the safety and efficacy of concurrent administration of DSF with Cu, we conducted a phase 1b clinical trial of patients with metastatic castration-resistant prostate cancer (mCRPC) receiving Cu with DSF.Patients with mCRPC were treated in two cohorts: mCRPC with nonliver/peritoneal metastases (A), and mCRPC with liver and/or peritoneal metastases (B). Baseline Cu avidity was measured by 64 CuCl2 PET scan. Intravenous (IV) CuCl2 was given weekly for three doses with oral daily DSF followed by daily oral Cu gluconate and DSF until disease progression. DSF and metabolite diethyldithiocarbamic acid methyl ester (Me-DDC) levels in plasma were measured. DSF and Me-DDC were then assessed for cytotoxicity in vitro.We treated nine patients with mCRPC (six on cohort A and three on cohort B). Bone and nodal metastases showed differential and heterogeneous Cu uptake on 64 CuCl2 PET scans. No confirmed PSA declines or radiographic responses were observed. Median PFS was 2.8 months and median OS was 8.3 months. Common adverse events included fatigue and psychomotor depression; no Grade 4/5 AEs were observed. Me-DDC was measurable in all samples (LOQ = 0.512 ng/ml), whereas DSF was not (LOQ = 0.032 ng/ml, LOD = 0.01 ng/ml); Me-DDC was not cytotoxic in vitro.Oral DSF is not an effective treatment for mCRPC due to rapid metabolism into an inactive metabolite, Me-DDC. This trial has stopped enrollment and further work is needed to identify a stable DSF formulation for treatment of mCRPC.

Mazmanian K., Chen T., Sargsyan K., Lim C.

The COVID-19 pandemic poses a challenge in coming up with quick and effective means to counter its cause, the SARS-CoV-2. Here, we show how the key factors governing cysteine reactivity in proteins derived from combined quantum mechanical/continuum calculations led to a novel multi-targeting strategy against SARS-CoV-2, in contrast to developing potent drugs/vaccines against a single viral target such as the spike protein. Specifically, they led to the discovery of reactive cysteines in evolutionary conserved Zn2+-sites in several SARS-CoV-2 proteins that are crucial for viral polypeptide proteolysis as well as viral RNA synthesis, proofreading, and modification. These conserved, reactive cysteines, both free and Zn2+-bound, can be targeted using the same Zn-ejector drug (disulfiram/ebselen), which enables the use of broad-spectrum anti-virals that would otherwise be removed by the virus's proofreading mechanism. Our strategy of targeting multiple, conserved viral proteins that operate at different stages of the virus life cycle using a Zn-ejector drug combined with other broad-spectrum anti-viral drug(s) could enhance the barrier to drug resistance and antiviral effects, as compared to each drug alone. Since these functionally important nonstructural proteins containing reactive cysteines are highly conserved among coronaviruses, our proposed strategy has the potential to tackle future coronaviruses. This article is categorized under:Structure and Mechanism > Reaction Mechanisms and CatalysisStructure and Mechanism > Computational Biochemistry and BiophysicsElectronic Structure Theory > Density Functional Theory.

Harris A.C., Muelken P., Howard S., Wilde S., LeSage M.G.

Xiong W., Li J., Mao X., Qian Z., Tian A., Peng X., Yang Z., Li H., Li Z.

Roy J., Mouawad R., Kyani A., Hanafi M., Xu Y., Wen B., Sun D., Neamati N.

Mackowiak B., Haggerty D.L., Lehner T., Lin Y., Fu Y., Lu H., Pawlosky R.J., Ren T., Seo W., Feng D., Zhang L., Lovinger D.M., Gao B.

AbstractBackgroundEthanol metabolism is intimately linked with the physiological and behavioral aspects of ethanol consumption. Ethanol is mainly oxidized by alcohol dehydrogenase (ADH) to acetaldehyde and further to acetate via aldehyde dehydrogenases (ALDHs). Understanding how ethanol and its metabolites work together to initiate and drive continued ethanol consumption is crucial for identifying interventions for alcohol use disorder (AUD). Therefore, the goal of our study was to determine how ADH1, which is mainly peripherally expressed and metabolizes >90% of ingested ethanol, modulates ethanol metabolite distribution and downstream behaviors.MethodsEthanol consumption in drinking‐in‐the‐dark (DID) and two‐bottle choice (2BC) drinking paradigms, ethanol metabolite concentrations, and lickometry were assessed after ADH1 inhibition and/or in Adh1‐knockout (Adh1 KO) mice.ResultsWe found that Adh1 KO mice of both sexes exhibited decreased ethanol consumption and preference compared with wild‐type (WT) mice in DID and 2BC. ADH1 inhibitor fomepizole (4‐MP) also significantly decreased normal and sweetened ethanol consumption in DID studies. Measurement of ethanol and its metabolites revealed that ethanol was increased at 1 h but not 15 min, peripheral acetaldehyde was slightly decreased at both timepoints, and ethanol‐induced increases in acetate were abolished after ethanol administration in Adh1 KO mice compared with controls. Similarly, ethanol accumulation as a function of consumption was 2‐fold higher in Adh1 KO or 4‐MP‐treated mice compared with controls. We then used lickometry to determine how this perturbation in ethanol metabolism affects drinking microstructure. Adh1 KO mice consume most of their ethanol in the first 30 min, like WT mice, but display altered temporal shifts in drinking behaviors and do not form normal bout structures, resulting in lower ethanol consumption.ConclusionsOur study demonstrates that ADH1‐mediated ethanol metabolism is a key determinant of ethanol consumption, highlighting a fundamental knowledge gap regarding how ethanol and its metabolites drive ethanol consumption.

d’Errico A., Strippoli E., Goldman S.M., Blanc P.D.

Abstract Background Case reports implicate disulfiram treatment in causing parkinsonism, but these observations lack epidemiological confirmation. Aim of the present study was to estimate the risk of incident parkinsonism associated with disulfiram dispensing in a large Italian population. Methods In this observational cohort study, administrative data were used, linking records at the individual level from civic registries, population census, mortality registers, hospital admissions, archives of drug prescriptions, and direct ambulatory drug distribution. Participants included all residents in the Piedmont region of Italy aged ≥ 40 years participating in 2011 census, still resident and alive at the beginning of 2013, followed-up from 2013 to 2019. The outcome was incident parkinsonism identified through multiple prescriptions of levodopa or a hospital admission for Parkinson’s disease or atypical parkinsonism. Exposure to disulfiram and to neuroleptics was assessed through regional drug prescription archives. The association between disulfiram and parkinsonism onset was assessed using Cox proportional hazards models, adjusted for gender, age and neuroleptic use. Results The study population included 2,498,491 individuals (mean age: 62 years). During follow-up, 19,072 parkinsonism cases were identified, 8 of whom had been prescribed disulfiram. Exposure to disulfiram was associated with a three-fold increased risk of parkinsonism (HR = 3.10, 95% CI = 1.55–6.21) that remained significant when adjusted for neuroleptic use (HR = 2.04, 95% CI = 1.01–4.10). The association was stronger among persons unexposed to neuroleptics and among those with more than four disulfiram prescriptions. Conclusions These results support the hypothesis that disulfiram may cause parkinsonism. Clinicians and drug regulatory agencies should consider parkinsonism when assessing the risks and benefits of disulfiram use.

Pich E.M., Tarnanas I., Brigidi P., Collo G.

Sridharan K.

Shenoy A., Jasty V.S., Uttal S., Nasser S., Winder G.S., Mellinger J., Fontana R.J.

The number of alcohol use disorder patients with mild (n = 101) and severe alcoholic liver disease (n = 112) increased between 2013 and 2023 (P = 0.06). Naltrexone was prescribed in 65%, acamprosate in 26%, and disulfiram in 9%. Internal medicine providers were the most frequent prescribers (51%), followed by psychiatry (30%) and gastroenterology/hepatology (9%) (P = 0.001). Duration of use was longest with naltrexone compared with acamprosate and disulfiram (360 vs 251 vs 190 days, P = 0.032). The rate of alcohol use disorder medication discontinuation for adverse events was similar in both groups (14% vs 12%). Naltrexone, acamprosate, and disulfiram are increasingly being prescribed in American patients with mild and severe alcohol-related liver disease with similar tolerability and efficacy.

Mackowiak B., Haggerty D.L., Lehner T., Lin Y., Fu Y., Lu H., Pawlosky R.J., Ren T., Seo W., Feng D., Zhang L., Lovinger D.M., Gao B.

AbstractBackgroundEthanol metabolism is intimately linked with the physiological and behavioral aspects of ethanol consumption. Ethanol is mainly oxidized by alcohol dehydrogenase (ADH) to acetaldehyde and further to acetate via aldehyde dehydrogenases (ALDHs). Understanding how ethanol and its metabolites work together to initiate and drive continued ethanol consumption is crucial for identifying interventions for alcohol use disorder (AUD). Therefore, the goal of our study was to determine how ADH1, which is mainly peripherally-expressed and metabolizes >90% of ingested ethanol, modulates ethanol metabolite distribution and downstream behaviors.MethodsEthanol consumption in drinking-in-the-dark (DID) and two-bottle choice (2BC) drinking paradigms, ethanol metabolite concentrations, and lickometry were assessed after ADH1 inhibition and/or inAdh1-knockout (Adh1KO) mice.ResultsWe found thatAdh1KO mice of both sexes exhibited decreased ethanol consumption and preference compared to wild-type (WT) mice in DID and 2BC. ADH1 inhibitor fomepizole (4-MP) also significantly decreased normal and sweetened ethanol consumption in DID studies. Measurement of ethanol and its metabolites revealed that ethanol was increased at 1h but not 15 min, peripheral acetaldehyde was slightly decreased at both time points, and ethanol-induced increases in acetate were abolished after ethanol administration inAdh1KO mice compared to controls. Similarly, ethanol accumulation as a function of consumption was 2-fold higher inAdh1KO or 4-MP treated mice compared to controls. We then used lickometry to determine how this perturbation in ethanol metabolism affects drinking microstructure.Adh1KO mice consume most of their ethanol in the first 30 min like WT mice but display altered temporal shifts in drinking behaviors and do not form normal bout structures, resulting in lower ethanol consumption.ConclusionsOur study demonstrates that ADH1-mediated ethanol metabolism is a key determinant of ethanol consumption, highlighting a fundamental knowledge gap around how ethanol and its metabolites drive ethanol consumption.

Kim J.K., Sapkota A., Roh T., Jo E.

Zhou Y., Wen R., Song T., Long B., Deng G.

Disulfide bonds, especially unsymmetric disulfide bonds, have important applications in bioactivity and drug molecules, but the synthesis of unsymmetric disulfide bonds remains challenging due to efficiency and selectivity issues. Herein, this work utilizes anthraquinone (AQ) and cyclictriphosphonononitrile through a nucleophilic substitution reaction to synthesize an organic polymer (ANTH-AMI) that incorporates an ortho-polyquinone (o-polyquinone) redox center. The anthraquinone molecule functions as a redox center, capable of accepting photoinduced electrons and subsequently transferring them to initiate an electron-coupled hydrogenation reaction (AQ to AQH). Moreover, the proximity of the o-polyquinone redox sites facilitates the catalysis of unsymmetric disulfide bond formation. Consequently, the ANTH-AMI photocatalysts demonstrate exceptional yields (up to 82 %), substrate versatility, cycling stability, and scalable preparation in promoting unsymmetric coupling reactions of thiol. This work offers a solution for designing organic polymer photocatalysts with adjacent multiple redox centers for cross-coupling reactions.

Kim K., Chae J.

AbstractOrganic disulfides are frequently found in many fields of science, ranging from chemistry to materials science to biology. While symmetrical disulfides, which have the same substitutions on the two sulfurs, are often final products, they are mostly employed as reactants and catalysts in various reactions. In addition to their usage as catalysts, their synthetic applications as reactants can arbitrarily be categorized into three types: (1) surrogates for the corresponding thiols, (2) reactants for unsymmetrical disulfides, and (3) reactants or intermediates for the other compounds. Although the straightforward, traditional way to obtain symmetrical disulfides (RSSR) is the oxidation of their parental thiols (RSH), they can also be obtained from non-thiolic compounds. This short review covers representative synthetic applications of symmetrical disulfides and their preparation methods in which the starting materials are not the corresponding thiols.1 Introduction2 Synthetic Applications of Symmetric Disulfides2.1 Surrogates for Thiols2.2 Synthesis of Unsymmetrical Disulfides2.3 As Reactants and Intermediates3 Synthesis of Symmetrical Disulfides from Non-thiolic Substrates3.1 Synthesis from Organic Halides3.2 Synthesis from Sulfonic Acid Derivatives3.3 Synthesis from Sulfinic Acid Derivatives3.4 Miscellaneous4 Conclusion

Ohno K., Murakami H., Ogo N., Asai A.

The cell painting assay is useful for understanding cellular phenotypic changes and drug effects. To identify other aspects of well-known chemicals, we screened 258 compounds with the cell painting assay and focused on a mitochondrial punctate phenotype seen with disulfiram. To elucidate the reason for this punctate phenotype, we looked for clues by examining staining steps and gene knockdown as well as examining protein solubility and comparing cell lines. From these results, we found that the punctate phenotype was caused by protein insolubility in the mitochondrial matrix. Interestingly, the punctate phenotype of disulfiram was sensitive to the relative expression of LonP1, a protease in the mitochondrial matrix that regulates proteostasis, suggesting that the punctate phenotype manifests when the protein quality control capacity in the mitochondrial matrix is exceeded. Moreover, we discovered that disulfiram and its derivatives, which have all been reported to increase acetaldehyde in the blood after the in vivo intake of alcohol, induced a punctate phenotype as well. The investigated punctate phenotype not only provides a novel clue for elucidating the common mechanism of action among disulfiram derivatives but is also a novel example of chemical perturbation of proteostasis in the mitochondrial matrix.

Evans S.E., Valentine M.E., Gallimore F., Meka Y., Koehler S.I., Yu H.D., Valentovic M.A., Long T.E.

Abstract Aims Disulfiram (Antabuse®) is an oral alcohol sobriety medication that exhibits antimicrobial activity against Gram-positive facultative anaerobes. The aims of this study were to measure the antimicrobial activity against anaerobic bacteria of the gut human microbiome and establish the extent that disulfiram alters the microbial composition of the ileum, cecum, and feces using C57BL/6 mice. Methods and results Antimicrobial susceptibility testing by the microdilution method revealed that disulfiram inhibits the in vitro growth of gut anaerobic species of Bacteroides, Clostridium, Peptostreptococcus, and Porphyromonas. Differential sequencing of 16S rRNA isolated from the ileum, cecum, and feces contents of treated vs. untreated mice showed that disulfiram enriches the Gram-negative enteric population. In female mice, the enrichment was greatest in the ileum, whereas the feces composition in male mice was the most heavily altered. Conclusions Daily administration of oral disulfiram depletes the enteric Gram-positive anaerobe population as predicted by the minimum inhibitory concentration data for isolates from the human gut microbiota.

Luo J., Zeng Y., Chen Z., Luo Y., Shi L., Zhou X.

ObjectiveDisulfiram, an FDA-approved medication for AUD, has shown significant potential as a repurposed drug in therapeutic areas including oncology and infectious diseases. The purpose of study is to analyze adverse events (AEs) associated with disulfiram by examining the FAERS database, with a focus on understanding its safety profile in both traditional and emerging applications.MethodsAE reports concerning disulfiram in the FAERS database from the fourth quarter of 2002 to the third quarter of 2023 were extracted. Various signal detection methods, including ROR, PRR, BCPNN, and MGPS, were used to detect and categorize adverse events.ResultsThe study collected 52,159,321 AE reports, with 508 reports primarily suspecting disulfiram, identifying 104 Preferred Terms (PTs) across 25 System Organ Classes (SOCs). Major categories of AEs included off label use, psychiatric symptom, liver transplant, and polyneuropathy, with off label use being notably the most reported issue. Strong and new potential AEs were identified, including neurological and psychiatric issues like hypomania, delirium, and vocal cord paralysis; cardiac issues such as electrocardiogram st segment depression; and off label use-related issues like Jarisch-Herxheimer reaction.ConclusionDisulfiram poses risks of various adverse reactions while having promise as a “repurposed” agent. In clinical applications, practitioners should closely monitor occurrences of hepatobiliary disorders, psychiatric disorders, and nervous system disorders.