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Cell Death Discovery Oct 2023Lung ischemia/reperfusion injury (LIRI) is a complex pathophysiological process, with the histopathological hallmark of neutrophils migrating into the lungs. Neutrophil...
Lung ischemia/reperfusion injury (LIRI) is a complex pathophysiological process, with the histopathological hallmark of neutrophils migrating into the lungs. Neutrophil extracellular traps (NETs) have been suggested to exert a critical role in the pathogenesis of inflammation and infection in humans and animals, while the exact functions and underlying mechanisms of NETs in LIRI remain insufficiently elucidated. In this study, we investigated the role of pore-forming protein gasdermin D (GSDMD) on NETs release in LIRI induced by lung ischemia/reperfusion (I/R). We found that disulfiram, a GSDMD inhibitor, dramatically reduced NETs release and pathological injury in lung I/R in vivo and in vitro. Additionally, GSDMD caused mitochondrial DNA (mtDNA) leaking into the neutrophil cytosol, and then the cytoplasmic mtDNA activated the cGAS-STING signaling pathway and stimulated NETs formation in lung I/R. Furthermore, inhibition of cGAS/STING pathway could inhibit cytosol mtDNA mediated NETs formation.
PubMed: 37794018
DOI: 10.1038/s41420-023-01663-z -
Scientific Reports Sep 2023Neuroblastoma, the most common type of pediatric extracranial solid tumor, causes 10% of childhood cancer deaths. Despite intensive multimodal treatment, the outcomes of...
Neuroblastoma, the most common type of pediatric extracranial solid tumor, causes 10% of childhood cancer deaths. Despite intensive multimodal treatment, the outcomes of high-risk neuroblastoma remain poor. We urgently need to develop new therapies with safe long-term toxicity profiles for rapid testing in clinical trials. Drug repurposing is a promising approach to meet these needs. Here, we investigated disulfiram, a safe and successful chronic alcoholism treatment with known anticancer and epigenetic effects. Disulfiram efficiently induced cell cycle arrest and decreased the viability of six human neuroblastoma cell lines at half-maximal inhibitory concentrations up to 20 times lower than its peak clinical plasma level in patients treated for chronic alcoholism. Disulfiram shifted neuroblastoma transcriptome, decreasing MYCN levels and activating neuronal differentiation. Consistently, disulfiram significantly reduced the protein level of lysine acetyltransferase 2A (KAT2A), drastically reducing acetylation of its target residues on histone H3. To investigate disulfiram's anticancer effects in an in vivo model of high-risk neuroblastoma, we developed a disulfiram-loaded emulsion to deliver the highly liposoluble drug. Treatment with the emulsion significantly delayed neuroblastoma progression in mice. These results identify KAT2A as a novel target of disulfiram, which directly impacts neuroblastoma epigenetics and is a promising candidate for repurposing to treat pediatric neuroblastoma.
Topics: Animals; Child; Humans; Mice; Alcohol Deterrents; Cell Line, Tumor; Disulfiram; Down-Regulation; Drug Repositioning; Emulsions; Histone Acetyltransferases; Neuroblastoma
PubMed: 37777587
DOI: 10.1038/s41598-023-43219-2 -
PloS One 2023Disulfiram and hydralazine have recently been reported to have anti-cancer action, and repositioned to be used as adjuvant in cancer therapy. Chemotherapy combined with...
Disulfiram and hydralazine have recently been reported to have anti-cancer action, and repositioned to be used as adjuvant in cancer therapy. Chemotherapy combined with other medications, such as those that affect the immune system or epigenetic cell profile, can overcome resistance with fewer adverse effects compared to chemotherapy alone. In the present study, a combination of doxorubicin (DOX) with hydrazine (Hyd) and disulfiram (Dis), as a triple treatment, was evaluated against wild-type and DOX-resistant MCF-7 breast cancer cell line. Both wild-type MCF-7 cell line (MCF-7_WT) and DOX-resistant MCF-7 cell line (MCF-7_DoxR) were treated with different combination ratios of DOX, Dis, and Hyd followed by measuring the cell viability using the MTT assay. Synergism was determined using a combination index, isobologram analysis, and dose-reducing index. The anti-proliferation activity and mechanism of the triple combination were investigated by apoptosis analysis. The results showed a reduction in the IC50 values of DOX in MCF-7_WT cells (from 0.24 μM to 0.012 μM) and MCF-7_DoxR cells (from 1.13 μM to 0.44 μM) when treated with Dis (0.03μM), and Hyd (20μM) combination. Moreover, The triple combination DOX/Hyd/Dis induced significant apoptosis in both MCF-7_WT and MCF-7_DoxR cells compared to DOX alone. The triple combination of DOX, Dis, and Hyd showed a synergistic drugs combination to decrease the DOX dose needed to kill both MCF-7_WT and MCF-7_DoxR cancer cells and enhanced chemosensitivity to DOX.
PubMed: 37768997
DOI: 10.1371/journal.pone.0291981 -
International Immunopharmacology Nov 2023Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by pruritus, erythema, and skin barrier dysfunction. Gasdermin D (GSDMD) is the key...
Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by pruritus, erythema, and skin barrier dysfunction. Gasdermin D (GSDMD) is the key executioner of an inflammatory cell death mechanism known as pyroptosis. However, the role of GSDMD in the pathogenesis of AD remains unclear. Through the analysis of publicly available Gene Expression Omnibus (GEO) datasets, we observed an upregulation of Gsdmd mRNA in the skin tissue of AD patients. Moreover, we delved into the impact of GSDMD deletion and inhibition on AD-like skin lesions using a mouse model induced by the topical application of oxazolone (Oxa). We found that mice lacking GSDMD exhibited relieved AD signs and symptoms in terms of reduced skin thickness, scarring and scratching behavior compared to wild-type mice after induction of AD-like skin lesions. This was associated with decreased infiltration of inflammatory cells, reduced epidermal thickness, and decreased serum levels of IgE and IL-4. Western blot analysis further revealed decreased GSDMD cleavage in the skin of GSDMD knockout mice, and reduced expression of IL-1β and IL-18. Inhibition of GSDMD using the pharmacological agent disulfiram or the herbal compound matrine significantly attenuated the symptoms of AD-like skin lesions in wild-type mice, GSDMD cleavage and pro-inflammatory cytokines were reduced as well. Our results suggest that GSDMD-mediated pyroptosis plays a critical role in the development of AD-like skin lesions, and targeting GSDMD may be a promising therapeutic strategy for AD.
Topics: Animals; Humans; Mice; Cytokines; Dermatitis, Atopic; Epidermis; Gasdermins; Phosphate-Binding Proteins; Pore Forming Cytotoxic Proteins; Skin
PubMed: 37741129
DOI: 10.1016/j.intimp.2023.110958 -
Nature Communications Sep 2023The poor efficacy of chimeric antigen receptor T-cell therapy (CAR T) for solid tumors is due to insufficient CAR T cell tumor infiltration, in vivo expansion,...
The poor efficacy of chimeric antigen receptor T-cell therapy (CAR T) for solid tumors is due to insufficient CAR T cell tumor infiltration, in vivo expansion, persistence, and effector function, as well as exhaustion, intrinsic target antigen heterogeneity or antigen loss of target cancer cells, and immunosuppressive tumor microenvironment (TME). Here we describe a broadly applicable nongenetic approach that simultaneously addresses the multiple challenges of CAR T as a therapy for solid tumors. The approach reprograms CAR T cells by exposing them to stressed target cancer cells which have been exposed to the cell stress inducer disulfiram (DSF) and copper (Cu)(DSF/Cu) plus ionizing irradiation (IR). The reprogrammed CAR T cells acquire early memory-like characteristics, potent cytotoxicity, enhanced in vivo expansion, persistence, and decreased exhaustion. Tumors stressed by DSF/Cu and IR also reprogram and reverse the immunosuppressive TME in humanized mice. The reprogrammed CAR T cells, derived from peripheral blood mononuclear cells of healthy donors or metastatic female breast cancer patients, induce robust, sustained memory and curative anti-solid tumor responses in multiple xenograft mouse models, establishing proof of concept for empowering CAR T by stressing tumor as a promising therapy for solid tumors.
Topics: Humans; Female; Animals; Mice; Leukocytes, Mononuclear; Receptors, Chimeric Antigen; Tumor Microenvironment; Breast Neoplasms; Disease Models, Animal; Immunosuppressive Agents; T-Lymphocytes
PubMed: 37714830
DOI: 10.1038/s41467-023-41282-x -
International Journal of Pharmaceutics Oct 2023The respiratory tract, as the first and most afflicted target of many viruses such as SARS-CoV-2, seems to be the logical choice for delivering antiviral agents against...
The respiratory tract, as the first and most afflicted target of many viruses such as SARS-CoV-2, seems to be the logical choice for delivering antiviral agents against this and other respiratory viruses. A combination of remdesivir and disulfiram, targeting two different steps in the viral replication cycle, has showed synergistic activity against SARS-CoV-2 in-vitro. In this study, we have developed an inhalable dry powder containing a combination of remdesivir and disulfiram utilizing the spray-drying technique, with the final goal of delivering this drug combination to the respiratory tract. The prepared dry powders were spherical, and crystalline. The particle size was between 1 and 5 μm indicating their suitability for inhalation. The spray-dried combinational dry powder containing remdesivir and disulfiram (RD) showed a higher emitted dose (ED) of >88% than single dry powder of remdesivir (R) (∼72%) and disulfiram (D) (∼84%), with a fine particle fraction (FPF) of ∼55%. Addition of L-leucine to RD showed >60% FPF with a similar ED. The in vitro aerosolization was not significantly affected after the stability study conducted at different humidity conditions. Interestingly, the single (R and D) and combined (RD) spray-dried powders showed limited cellular toxicity (CC values from 39.4 to >100 µM), while maintaining their anti-SARS-CoV-2 in vitro (EC values from 4.43 to 6.63 µM). In a summary, a combinational dry powder formulation containing remdesivir and disulfiram suitable for inhalation was developed by spray-drying technique which showed high cell viability in the respiratory cell line (Calu-3 cells) retaining their anti-SARS-CoV-2 property. In the future, in vivo studies will test the ability of these formulations to inhibit SARS-CoV-2 which is essential for clinical translation.
PubMed: 37703955
DOI: 10.1016/j.ijpharm.2023.123411 -
Cell Death & Disease Sep 2023Gasdermin D (GSDMD)-mediated pyroptosis has a significant pro-inflammation characteristic due to dramatic secretion of pro-inflammatory substances. However, its role...
Gasdermin D (GSDMD)-mediated pyroptosis has a significant pro-inflammation characteristic due to dramatic secretion of pro-inflammatory substances. However, its role remains unclear in psoriasis as one chronic inflammatory skin disorder with high prevalence. We found that N-terminal GSDMD (N-GSDMD) was aberrantly expressed in epidermis of skin lesion in psoriasis patients and imiquimod-induced psoriasis-like dermatitis (IIPLD) mice. In epidermis of IIPLD mice and M5 (simulating psoriatic inflammatory challenge)-treated keratinocytes cultured in vitro, cleavage products of caspase-1, GSDMD and IL-1β were increased. M5-stimulated keratinocyte presented typical pyroptosis morphology accompanied with PI-staining. Gsdmd keratinocytes could not present pyroptosis morphology while stimulated with M5. Electroporation of recombinant N-GSDMD could make the pyroptosis morphology reappear. In Gsdmd mice or keratinocyte-specific Gsdmd conditional knockout mice, we observed the alleviation of psoriatic inflammation and epidermal aberrant expression of Ki-67 and differentiation markers (loricrin and keratin 5) after imiquimod stimulation. Transplanting skin tissue from control mice to Gsdmd mice can evoke the response to imiquimod stimulation in the background of Gsdmd mice (not limited in transplanting area). In M5-stimulated keratinocytes, disulfiram or GSDMD siRNA transfection can inhibit pyroptosis and eliminate disproportionate increases of Ki-67 and PI. We further validated that topically application of disulfiram (pyroptosis inhibitor) also alleviated IIPLD in mice. These findings indicate a novel mechanism that GSDMD-mediated keratinocyte pyroptosis facilitates hyperproliferation and aberrant differentiation induced by immune microenvironment in psoriatic skin inflammation, which contributes to pathogenesis of psoriasis. Our study provides an innovative insight that targeting pyroptosis can be considered as a therapeutic strategy against psoriasis.
Topics: Animals; Mice; Gasdermins; Disulfiram; Imiquimod; Ki-67 Antigen; Pyroptosis; Keratinocytes; Psoriasis; Dermatitis; Inflammation
PubMed: 37673869
DOI: 10.1038/s41419-023-06094-3 -
Scientific Reports Sep 2023By utilizing Optum Life Sciences Claims Data, we constructed Real World Data (RWD) cohorts comprising over 3 million patients and simulated a clinical trial... (Observational Study)
Observational Study
By utilizing Optum Life Sciences Claims Data, we constructed Real World Data (RWD) cohorts comprising over 3 million patients and simulated a clinical trial observational study design to evaluate over 200 FDA-approved drugs with COVID-19 repurposing potential, and identified a dozen candidates exhibiting significant reduction in the odds of severe COVID-19 outcomes such as death, intensive care unit (ICU) admission, hospitalization and pneumonia. Notably, certain drug combinations demonstrated effects comparable to those of COVID-19 vaccines. Furthermore, our study revealed a novel finding: a quantitative linear relationship between COVID-19 outcomes and overall patient health risks. This discovery enabled a more precise estimation of drug efficacy using the risk adjustment. The top performing drugs identified include emtricitabine, tenofovir, folic acid, progesterone, estradiol, epinephrine, disulfiram, nitazoxanide and some drug combinations including aspirin-celecoxib.
Topics: Humans; COVID-19; COVID-19 Vaccines; Drug Repositioning; COVID-19 Drug Treatment; Aspirin
PubMed: 37666866
DOI: 10.1038/s41598-023-40033-8 -
Frontiers in Bioscience (Landmark... Aug 2023The complex formed by disulfiram (DSF) and copper (Cu) is safe and effective for the prevention and treatment of triple-negative breast cancer (TNBC). Although previous...
BACKGROUND
The complex formed by disulfiram (DSF) and copper (Cu) is safe and effective for the prevention and treatment of triple-negative breast cancer (TNBC). Although previous studies have shown that DSF/Cu induces ferroptosis, the mechanism remains unclear.
METHODS
The mitochondrial morphology of TNBC treated with DSF/Cu was observed by transmission microscopy, and intracellular levels of iron, lipid reactive oxygen species (ROS), malondialdehyde, and glutathione were evaluated to detect the presence of ferroptosis. Target genes for the DSF/Cu-activated ferroptosis signaling pathway were examined by transcriptome sequencing analysis. Expression of the target gene, , was detected by qRT-PCR, immunofluorescence and western blot.
RESULTS
The mitochondria of TNBC cells were significantly atrophied following treatment with DSF/Cu for 24 h. Addition of DSF/Cu supplement resulted in significant up-regulation of intracellular iron, lipid ROS and malondialdehyde levels, and significant down-regulation of glutathione levels, all of which are important markers of ferroptosis. Transcriptome analysis confirmed that DSF/Cu activated the ferroptosis signaling pathway and up-regulated several ferroptosis target genes associated with redox regulation, especially heme oxygenase-1 (HMOX-1). Inhibition of ferroptosis by addition of the ROS scavenger N-acetyl-L-cysteine (NAC) significantly increased the viability of DSF/Cu-treated TNBC cells.
CONCLUSIONS
These results show that DSF/Cu increases lipid peroxidation and causes a sharp increase in HMOX1 activity, thereby inducing TNBC cell death through ferroptosis. DSF/Cu is a promising therapeutic drug for TNBC and could lead to ferroptosis-mediated therapeutic strategies for human cancer.
Topics: Humans; Triple Negative Breast Neoplasms; Copper; Disulfiram; Ferroptosis; Reactive Oxygen Species; Cell Line; Glutathione; Lipids
PubMed: 37664913
DOI: 10.31083/j.fbl2808186 -
Research Square Aug 2023Irradiation (IR) induces immunogenic cell death (ICD) in tumors, but it rarely leads to the abscopal effect (AE). However, combining IR with immune checkpoint inhibitors...
Turning anecdotal irradiation-induced anti-cancer immune responses into reproducible in situ cancer vaccines via disulfiram/copper-mediated enhanced immunogenic cell death of breast cancer cells.
Irradiation (IR) induces immunogenic cell death (ICD) in tumors, but it rarely leads to the abscopal effect (AE). However, combining IR with immune checkpoint inhibitors has shown anecdotal success in inducing AEs. In this study, we aimed to enhance the IR-induced immune response and generate reproducible AEs using the anti-alcoholism drug disulfiram (DSF) and copper complex (DSF/Cu) via induction of tumor ICD. We measured ICD in vitro and in vivo. In mouse tumor models, DSF/Cu was injected intratumorally followed by localized tumor IR, creating an in situ cancer vaccine. We determined the anti-cancer response by primary tumor rejection and assessed systemic immune responses by tumor rechallenge and the occurrence of AEs, i.e., spontaneous lung metastasis. Additionally, we analyzed immune cell subsets and quantified proinflammatory and immunosuppressive chemokines/cytokines in the tumor microenvironment (TME) and blood of the vaccinated mice. Immune cell depletion was investigated for its effects on the vaccine-induced anti-cancer response. The results showed that DSF/Cu and IR induced more potent ICD under hypoxia than normoxia in vitro. Low-dose intratumoral injection of DSF/Cu and IR demonstrated strong anti-primary and -rechallenged tumor effects and robust AEs in mouse models. These vaccinations also increased CD8 + and CD4 + cell numbers while decreasing Tregs and myeloid-derived suppressor cells in the 4T1 model, and increased CD8+, DC, and decreased Treg cell numbers in the MCa-M3C model. Depleting both CD8 + and CD4 + cells abolished the vaccine's anticancer response. Moreover, vaccinated tumor-bearing mice exhibited increased TNFα levels and reduced levels of immunosuppressive chemokines/cytokines. In conclusion, our novel approach generated an anti-cancer immune response, resulting in a lack of or low tumor incidence post-rechallenge and robust AEs, i.e., the absence of or decreased spontaneous lung metastasis in tumor-bearing mice. This approach is readily translatable to clinical settings and may increase IR-induced AEs in cancer patients.
PubMed: 37645899
DOI: 10.21203/rs.3.rs-3195392/v1