Inhibiting ACSL1-Related Ferroptosis Restrains Murine Coronavirus Infection

Murine hepatitis virus strain A59 (MHV-A59) was shown to induce pyroptosis, apoptosis, and necroptosis of infected cells, especially in the murine macrophages. However, whether ferroptosis, a recently identified form of lytic cell death, was involved in the pathogenicity of MHV-A59 is unknown. We utilized murine macrophages and a C57BL/6 mice intranasal infection model to address this. In primary macrophages, the ferroptosis inhibitor inhibited viral propagation, inflammatory cytokines released, and cell syncytia formed after MHV-A59 infection. In the mouse model, we found that in vivo administration of liproxstatin-1 ameliorated lung inflammation and tissue injuries caused by MHV-A59 infection. To find how MHV-A59 infection influenced the expression of ferroptosis-related genes, we performed RNA-seq in primary macrophages and found that MHV-A59 infection upregulates the expression of the acyl-CoA synthetase long-chain family member 1 (ACSL1), a novel ferroptosis inducer. Using ferroptosis inhibitors and a TLR4 inhibitor, we showed that MHV-A59 resulted in the NF-kB-dependent, TLR4-independent ACSL1 upregulation. Accordingly, ACSL1 inhibitor Triacsin C suppressed MHV-A59-infection-induced syncytia formation and viral propagation in primary macrophages. Collectively, our study indicates that ferroptosis inhibition protects hosts from MHV-A59 infection. Targeting ferroptosis may serve as a potential treatment approach for dealing with hyper-inflammation induced by coronavirus infection.

Inhibiting ACSL1 related ferroptosis restrains MHV-A59 infection

Murine hepatitis virus strain A59 (MHV-A59) belongs to the β-coronavirus and is considered as a representative model for studying coronavirus infection. MHV-A59 was shown to induce pyroptosis, apoptosis and necroptosis of infected cells, especially the murine macrophages. However, whether ferroptosis, a recently identified form of lytic cell death, was involved in the pathogenicity of MHV-A59, is unknown. Here, we demonstrate inhibiting ferroptosis suppresses MHV-A59 infection. MHV-A59 infection upregulates the expression of Acsl1, a novel ferroptosis inducer. MHV-A59 upregulates Acsl1 expression depending on the NF-kB activation, which is TLR4-independent. Ferroptosis inhibitor inhibits viral propagation, inflammatory cytokines release and MHV-A59 infection induced cell syncytia formation. ACSL1 inhibitor Triacsin C suppresses MHV-A59 infection induced syncytia formation and viral propagation. In vivo administration of liproxstatin-1 ameliorates lung inflammation and tissue injuries caused by MHV-A59 infection. Collectively, these results indicate that ferroptosis inhibition protects hosts from MHV-A59 infection. Targeting ferroptosis may serves as a potential treatment approach for dealing with hyper-inflammation induced by coronavirus infection.

A metabolic modeling approach reveals promising therapeutic targets and antiviral drugs to combat COVID-19

In this study we have developed a method based on Flux Balance Analysis to identify human metabolic enzymes which can be targeted for therapeutic intervention against COVID-19. A literature search was carried out in order to identify suitable inhibitors of these enzymes, which were confirmed by docking calculations. In total, 10 targets and 12 bioactive molecules have been predicted. Among the most promising molecules we identified Triacsin C, which inhibits ACSL3, and which has been shown to be very effective against different viruses, including positive-sense single-stranded RNA viruses. Similarly, we also identified the drug Celgosivir, which has been successfully tested in cells infected with different types of viruses such as Dengue, Zika, Hepatitis C and Influenza. Finally, other drugs targeting enzymes of lipid metabolism, carbohydrate metabolism or protein palmitoylation (such as Propylthiouracil, 2-Bromopalmitate, Lipofermata, Tunicamycin, Benzyl Isothiocyanate, Tipifarnib and Lonafarnib) are also proposed.

Melatonin Modulates Lipid Metabolism in Porcine Cumulus–Oocyte Complex via Its Receptors

Lipid is a crucial energy resource for mammalian oocyte. Melatonin could benefit the maturation of porcine oocyte in vitro, but the related mechanism is not elucidated yet. In the current study, methods to monitor lipid metabolism in single live oocytes were firstly established using probes (Lipi-Blue and Lipi-Green). It was observed that both lipid biogenesis and lipolysis occurred in maturing oocyte, but the general level of lipids dropped. Then maturing oocytes stained with probes were treated with melatonin or lipid metabolic-related inhibitors (triacsin C, rotenone, or etomoxir). The results showed that the lipid metabolism and maturation of porcine oocytes were all disrupted and that melatonin rescued the oocytes treated with triacsin C or rotenone, but not those treated with etomoxir. Further investigation demonstrated that cumulus cells are able to transfer lipids to oocytes via gap junctions. It was also observed that melatonin receptors exist in cumulus cells and are required for oocytes to maintain lipid metabolism. Meanwhile, the global gene expressing in cumulus cells was also modulated by melatonin, especially the genes related to antioxidants (SOD1, GPX1, GPX3, GPX4, PRDX2, and PRDX5), lipid metabolism (FABP3, FABP5, ACACB, TECR, etc.), and mitochondrial respiration (GPD1, ETFB, CYC1, and the genes of ATP synthase). Altogether the current research demonstrates that melatonin modulates lipid metabolism in maturing oocytes through its receptors in cumulus cells and benefits the developmental competence of oocytes.

Enhanced lipid metabolism induces the sensitivity of dormant cancer cells to 5-aminolevulinic acid-based photodynamic therapy

Cancer can develop into a recurrent metastatic disease with latency periods of years to decades. Dormant cancer cells, which represent a major cause of recurrent cancer, are relatively insensitive to most chemotherapeutic drugs and radiation. We previously demonstrated that cancer cells exhibited dormancy in a cell density-dependent manner. Dormant cancer cells exhibited increased porphyrin metabolism and sensitivity to 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT). However, the metabolic changes in dormant cancer cells or the factors that enhance porphyrin metabolism have not been fully clarified. In this study, we revealed that lipid metabolism was increased in dormant cancer cells, leading to ALA-PDT sensitivity. We performed microarray analysis in non-dormant and dormant cancer cells and revealed that lipid metabolism was remarkably enhanced in dormant cancer cells. In addition, triacsin C, a potent inhibitor of acyl-CoA synthetases (ACSs), reduced protoporphyrin IX (PpIX) accumulation and decreased ALA-PDT sensitivity. We demonstrated that lipid metabolism including ACS expression was positively associated with PpIX accumulation. This research suggested that the enhancement of lipid metabolism in cancer cells induces PpIX accumulation and ALA-PDT sensitivity.

Induction of apoptosis in RL95-2 human endometrial cancer cells by combination treatment with docosahexaenoic acid and triacsin C

IntroductionDocosahexaenoic acid (DHA) supplementation has been reported to negatively correlate with cancer cell proliferation and tumour development in many cancer types. Although cumulative evidence has demonstrated the apoptotic effect and cytotoxicity of DHA against tumour development in many cell types, the precise cellular and biochemical mechanisms of DHA-induced apoptosis in human endometrial cancer cells have not been investigated.Material and methodsMTT assay was performed to confirm the degree of apoptosis by combining treatment with DHA and triacsin C in endometrial cancer cell line. The synergistic effects of triacsin C and DHA were identified by performing flowcytometry and immunoblotting analysis.ResultsCombined treatment with DHA and triacsin C significantly induced apoptosis in RL95-2 endometrial carcinoma cells. Combined treatment with 125 μM DHA and 5 μM triacsin C significantly increased the sub-G1 population and apoptotic fragments in endometrial carcinoma cells. It was also demonstrated that DHA and triacsin C induced apoptosis through mitochondrial pathways via caspases-9, -3, and -7 as well as through the extrinsic pathway by activation of caspase-8/BID.ConclusionsFurther elucidation of the apoptotic mechanisms involving DHA treatment with ACS ablation could shed light on possible new treatment strategies for endometrial cancer. In addition, further research into the mechanisms of DHA and triacsin C-induced apoptotic mechanisms may lead to the development of therapeutic strategies for endometrial cancer.

Enhanced Lipid Metabolism Induces the Sensitivity of Dormant Cancer Cells to 5-aminolevulinic acid-based Photodynamic Therapy

Cancer can develop into a recurrent metastatic disease with latency periods of years to decades. Dormant cancer cells, which represent a major cause of recurrent cancer, are relatively insensitive to most chemotherapeutic drugs and radiation. We previously demonstrated that cancer cells exhibited dormancy in a cell density-dependent manner. Dormant cancer cells exhibited increased porphyrin metabolism and sensitivity to 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT). However, the metabolic changes in dormant cancer cells or the factors that enhance porphyrin metabolism have not been fully clarified. In this study, we revealed that lipid metabolism was increased in dormant cancer cells, leading to ALA-PDT sensitivity. We performed microarray analysis in non-dormant and dormant cancer cells and revealed that lipid metabolism was remarkably enhanced in dormant cancer cells. In addition, triacsin C, a potent inhibitor of acyl-CoA synthetases (ACSs), reduced protoporphyrin IX (PpIX) accumulation and decreased ALA-PDT sensitivity. We demonstrated that lipid metabolism including ACS expression was positively associated with PpIX accumulation. This research suggested that the enhancement of lipid metabolism in cancer cells induces PpIX accumulation and ALA-PDT sensitivity.

Acyl-CoA Synthetase Long Chain Family Inhibition with Triacsin C Inhibits Multiple Myeloma Cell Proliferation and Survival

Multiple myeloma (MM) is defined by the clonal expansion of malignant plasma cells in the bone marrow (BM) and has a 5-year survival rate of 50% (Siegel el al. 2018, Cancer J. Clin.). MM remains incurable due to the development of resistance to current chemotherapies; therefore, it is paramount to investigate novel treatments and the mechanisms of drug resistance in MM cells. Interestingly, obesity correlates with increased incidence of MM and high body mass index correlates with a poor treatment response (Marinac et al. 2019, JNCI Cancer Spectr, Groß et al. 2017, Oncotarget). Obesity is a major risk factor for many cancers, however, given the complexity of obesity, there are an array of mechanisms by which obesity may support tumor cells. Studies of obesity and MM are mainly at the epidemiological level and have not extensively explored the mechanism of this relationship. Therefore, there is a critical need to understand how obesity contributes to support cancers such as MM. Possible mechanisms may be through the increased availability of free fatty acids or through other factors that are found in obese patients. We hypothesize that lipid metabolism contributes to obesity-linked cancers such as MM. Recently, changes in lipid metabolism have been shown to support the proliferation, migration and the development of drug resistance in other blood cancers such as acute myeloid leukemia (Tabe et al. 2017, Cancer Res, Tabe et al. 2018, Sci. Reports) and solid tumors such as breast (Wang et al. 2017, JCI Insights) and prostate (Mitra et al. 2017, BMC Cancer) cancer. However, the role of lipid metabolism in MM cells has been understudied. Therefore, we hypothesized that genes within the Hallmark Fatty Acid Metabolism gene set (https://www.gsea-msigdb.org) would be differentially expressed between healthy patients and those with MM. We mined the clinical data (GSE6477, Chng et al. 2007, Cancer Res.) and found that transcripts of an enzyme critical for lipid metabolism, acyl-CoA synthetase long chain member 1 (ACSL1), was significantly downregulated (Figure 1A, Log2(Fold Change)=-2.33, adjusted p value=1.64*10-5, false discovery rate) in patients with newly diagnosed MM relative to normal plasma cells. Therefore, we hypothesized that ACSL1 may act as a tumor suppressor in MM. In order to test the role of the ACSL family as tumor suppressors, we treated human (MM1.S, OPM2 and RPMI-8226) and mouse myeloma (5TGM1) cell lines with an inhibitor (Triacsin C, TriC) of four of the five human acyl-CoA synthetase long chain family members (ACSL1,3,4 and 5). Contrary to our hypothesis, TriC treatment significantly decreased MM cell proliferation (Figure 1B, p<0.0001, One-way ANOVA Tukey's multiple comparisons test is used throughout unless otherwise noted), increased apoptosis (Figure 1C, p<0.001) and caused G0 arrest (Figure1D, p<0.0001) in a dose-dependent manner. Motivated to understand if TriC's toxicity was due to changes in metabolic dynamics, MM1.S cells were treated with 1 μM TriC for 30 minutes and subjected to a metabolic flux assay (Seahorse XF, Agilent). TriC treatment significantly reduced ATP-dependent respiration from fatty acid oxidation (FAO) (Figure 1F, p<0.0001 Student's t-test) and increased proton leak (p<0.0001). Taken together, our data demonstrate that TriC-mediated ACSL inhibition in MM cells decreases proliferation, induces G0 arrest, apoptosis and decreases FAO-dependent respiration and mitochondrial function. It is unclear what ACSL family member is responsible for the phenotype we report here. To address these questions, future studies will focus on genetically targeting individual ACSL family members and characterizing the lipidomic profile of MM ACSL mutants. Our data also suggests that fatty acids are used as an energy source, therefore we will explore how FAO contributes to MM cell proliferation and survival. Disclosures No relevant conflicts of interest to declare.

Inhibitors of VPS34 and lipid metabolism suppress SARS-CoV-2 replication

ABSTRACTTherapeutics targeting replication of SARS coronavirus 2 (SARS-CoV-2) are urgently needed. Coronaviruses rely on host membranes for entry, establishment of replication centers and egress. Compounds targeting cellular membrane biology and lipid biosynthetic pathways have previously shown promise as antivirals and are actively being pursued as treatments for other conditions. Here, we tested small molecule inhibitors that target membrane dynamics or lipid metabolism. Included were inhibitors of the PI3 kinase VPS34, which functions in autophagy, endocytosis and other processes; Orlistat, an inhibitor of lipases and fatty acid synthetase, is approved by the FDA as a treatment for obesity; and Triacsin C which inhibits long chain fatty acyl-CoA synthetases. VPS34 inhibitors, Orlistat and Triacsin C inhibited virus growth in Vero E6 cells and in the human airway epithelial cell line Calu-3, acting at a post-entry step in the virus replication cycle. Of these the VPS34 inhibitors exhibit the most potent activity.

A metabolic modeling approach reveals promising therapeutic targets and antiviral drugs to combat COVID-19

In this study we have developed a metabolic modeling approach to identify human metabolic enzymes which can be targeted for therapeutic intervention against COVID-19. A literature search was carried out in order to identify suitable inhibitors of these enzymes, which were confirmed by docking calculations. In total, 10 targets and 12 bioactive molecules have been predicted. Among the most promising molecules we identified Triacsin C, which inhibits ACSL3, and which has been shown to be very effective against different viruses, including positive-sense single-stranded RNA viruses. Similarly, we also identified the drug Celgosivir, which has been successfully tested in cells infected with different types of viruses such as Dengue, Zika, Hepatitis C and Influenza. Finally, other drugs targeting enzymes of lipid metabolism, carbohydrate metabolism or protein palmitoylation (such as propylthiouracil, 2-bromopalmitate, lipofermata, tunicamycin, benzyl isothiocyanate, tipifarnib and lonafarnib) are also proposed.