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Fatty acid metabolism can provide various products essential for viral infections. How vaccinia virus (VACV), the prototype of poxviruses, modulates fatty acid metabolism is not well understood. Here, we show that VACV infection results in increased neutral lipid droplet synthesis, the organelles that play a crucial role in storing and mobilizing fatty acids for energy production via β-oxidation. Citrate is the first tricarboxylic acid (TCA) cycle intermediate that can be transported to the cytosol to be converted to acetyl-CoA for de novo fatty acid biosynthesis. We found that VACV infection stimulates the S455 phosphorylation of ATP citrate lyase (ACLY), a pivotal enzyme that links citrate metabolism with lipid metabolism. We demonstrate that the inhibition of neutral lipid droplet synthesis and ACLY severely suppresses VACV replication. Remarkably, we found that virus growth factor (VGF)-induced signaling is essential for the VACV-mediated upregulation of ACLY phosphorylation and neutral lipid droplets. Finally, we report that VGF-induced EGFR-Akt pathway and ACLY phosphorylation are important for VACV stimulation of neutral lipid synthesis. These findings identified a new way of rewiring cell metabolism by a virus and a novel function for VGF in the governance of virus-host interactions through the induction of a key enzyme at the crossroads of the TCA cycle and fatty acid metabolism. Our study also provides a mechanism for the role played by VGF and its downstream signaling cascades in the modulation of lipid metabolism in VACV-infected cells.IMPORTANCENeutral lipid droplets are vital players in cellular metabolism. Here, we showed that VACV induces neutral lipid droplet synthesis in infected primary human foreskin fibroblasts and identified the cellular and viral factors needed. We identified VACV encoded growth factor (VGF) as an essential viral factor that induces cellular EGFR-Akt signaling to increase lipid droplets. Interestingly, VACV increases the S455 phosphorylation of ACLY, a key metabolic enzyme that sits at the crossroads of carbohydrate and lipid metabolism in a VGF-EGFR-Akt-dependent manner. We also found that ACLY is vital for VACV-induced lipid droplet synthesis. Our findings identified the modulation of ACLY by a virus and identified it as a potential target for antiviral development against pathogenic poxviruses. Our study also expands the role of growth factor signaling in boosting VACV replication by targeting fatty acid metabolism., Competing Interests: The authors declare no conflict of interest.

Purpose: Chemotherapy is one of the most commonly used treatments for oral squamous cell carcinoma (OSCC), but its use is limited by drug resistance and severe systemic toxicity. To eliminate these side effects and improve anti-tumor efficacy, several therapeutic approaches have been developed for use with chemotherapy. In this study, we developed a graphene-based lipid modulation nanoplatform (NSD) that carries SB-204990, a small molecule inhibitor specific for ATP citrate lyase (ACLY), and doxorubicin (DOX), a chemotherapeutic agent, and the trio enables synergistic treatment of OSCC with lipid starvation, chemotherapy, and photothermal therapy., Methods: We first determined whether ACLY expression was upregulated in OSCC, and then assessed the growth inhibitory effects of SB-204990 on SCC-15 cells and changes in lipid (acetyl coenzyme A, free fatty acids, and cholesterol) levels. We characterized NSD and then evaluated the stability, photothermal properties, drug loading, and release ability of NSD. Finally, the therapeutic effects of NSD on OSCC were investigated by in vitro and in vivo experiments, and the changes in lipid levels in OSCC tissues after ACLY inhibition were further evaluated., Results: The results showed that ACLY was highly expressed in OSCC, and ACLY inhibition produced reproductive suppression and decreased lipid levels in SCC-15 cells. The NSD nanoplatform possessed good stability, photothermal properties, high drug loading capacity and controlled release. In addition, the triple therapy achieved satisfactory anticancer effects in both in vivo and in vitro assays, and the inhibition rate of tumors was as high as 99.4% in the NSD+Laser treatment group., Conclusion: The changes in tumor cell lipid levels and cell proliferation arrest induced by ACLY inhibition suggest that ACLY may be a promising target for lipid starvation therapy and resistance to chemoresistance, and its inhibitors are expected to become new anticancer drugs. The NSD nanocarrier system enables synergistic treatment with lipid starvation, chemotherapy, and photothermal therapy, which represents an innovative approach to combating tumors., Competing Interests: 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., (© 2024 Li et al.)

Spliceosome dysfunction and aberrant RNA splicing underline unresolved inflammation and immunopathogenesis. Here, we revealed the misregulation of mRNA splicing via the spliceosome in the pathogenesis of rheumatoid arthritis (RA). Among them, decreased expression of RNA binding motif protein 25 (RBM25) was identified as a major pathogenic factor in RA patients and experimental arthritis mice through increased proinflammatory mediator production and increased hyperinflammation in macrophages. Multiomics analyses of macrophages from RBM25-deficient mice revealed that the transcriptional enhancement of proinflammatory genes (including Il1b, Il6, and Cxcl10) was coupled with histone 3 lysine 9 acetylation (H3K9ac) and H3K27ac modifications as well as hypoxia inducible factor-1α (HIF-1α) activity. Furthermore, RBM25 directly bound to and mediated the 14 th exon skipping of ATP citrate lyase (Acly) pre-mRNA, resulting in two distinct Acly isoforms, Acly Long (Acly L) and Acly Short (Acly S). In proinflammatory macrophages, Acly L was subjected to protein lactylation on lysine 918/995, whereas Acly S did not, which influenced its affinity for metabolic substrates and subsequent metabolic activity. RBM25 deficiency overwhelmingly increased the expression of the Acly S isoform, enhancing glycolysis and acetyl-CoA production for epigenetic remodeling, macrophage overactivation and tissue inflammatory injury. Finally, macrophage-specific deletion of RBM25 led to inflammaging, including spontaneous arthritis in various joints of mice and inflammation in multiple organs, which could be relieved by pharmacological inhibition of Acly. Overall, targeting the RBM25-Acly splicing axis represents a potential strategy for modulating macrophage responses in autoimmune arthritis and aging-associated inflammation., (© 2024. The Author(s), under exclusive licence to CSI and USTC.)

Background and Aims: Hepatic ischemia-reperfusion (IR) injury is the most common complication that occurs in liver surgery and hemorrhagic shock. ATP citrate lyase (Acly) plays a pivotal role in chromatin modification via generating acetyl-CoA for histone acetylation to influence biological processes. We aim to examine the roles of Acly, which is highly expressed in hepatocytes, in liver IR injury., Approach and Results: The functions of Acly in hepatic IR injury were examined in the mouse model with a hepatocyte-specific knockout of Acly . The Acly target genes were analyzed by CUT&RUN assay and RNA sequencing. The relationship between the susceptibility of the steatotic liver to IR and Acly was determined by the gain of function studies in mice. Hepatic deficiency of Acly exacerbated liver IR injury. IR induced Acly nuclear translocation in hepatocytes, which spatially fueled nuclear acetyl-CoA. This alteration was associated with enhanced acetylation of H3K9 and subsequent activation of the Foxa2 signaling pathway. Nuclear localization of Acly enabled Foxa2-mediated protective effects after hypoxia-reperfusion in cultured hepatocytes, while cytosolic Acly demonstrated no effect. The presence of steatosis disrupted Acly nuclear translocation. In the steatotic liver, restoration of Acly nuclear localization through overexpression of Rspondin-1 or Rspondin-3 ameliorated the IR-induced injury., Conclusions: Our results indicate that Acly regulates histone modification by means of nuclear AcCoA production in hepatic IR. Disruption of Acly nuclear translocation increases the vulnerability of the steatotic liver to IR. Nuclear Acly thus may serve as a potential therapeutic target for future interventions in hepatic IR injury, particularly in the context of steatosis., (Copyright © 2023 American Association for the Study of Liver Diseases.)

Lipid metabolism in various adipose tissue depots can differ vastly. This also applies to lipogenesis, the process of synthesizing fatty acids from acetyl-CoA. This study compared the expression of some lipogenic enzymes: fatty acid synthase (FASN), ATP-citrate lyase (ACLY), and malic enzyme 1 (ME1) in different regions of the posterior subcutaneous adipose tissue in rats. Methods and Results: Posterior subcutaneous adipose tissue collected from twelve-month-old Wistar rats was divided into six parts (A-F). The expression of genes encoding lipogenic enzymes was assessed by measuring their activity and mRNA levels using real-time PCR. In the gluteal region of the fat pad, there were much higher levels of activity and mRNA for these lipogenic enzymes compared to the dorsolumbar region. The mRNA level of FASN increased by more than twentyfold, whereas the level of ME1 and ACLY increased eight- and fivefold respectively. This phenomenon was observed in both old and young animals. Furthermore, the lack of uncoupling protein one (Ucp1) expression suggests that neither the presence of brown adipocytes in the gluteal part nor the transformation of white adipocytes into beige contributed to the observed differences. Conclusion: These results indicate that the gluteal white adipose tissue appears to be a unique and separate subcutaneous fat depot.

Elevated lipid synthesis is one of the best-characterized metabolic alterations in cancer and crucial for membrane expansion. As a key rate-limiting enzyme in de novo fatty acid synthesis, ATP-citrate lyase (ACLY) is frequently up-regulated in tumors and regulated by posttranslational modifications (PTMs). Despite emerging evidence showing O -GlcNAcylation on ACLY, its biological function still remains unknown. Here, we observed a significant upregulation of ACLY O -GlcNAcylation in various types of human tumor cells and tissues and identified S979 as a major O -GlcNAcylation site. Importantly, S979 O -GlcNAcylation is required for substrate CoA binding and crucial for ACLY enzymatic activity. Moreover, it is sensitive to glucose fluctuation and decisive for fatty acid synthesis as well as tumor cell proliferation. In response to EGF stimulation, both S979 O -GlcNAcylation and previously characterized S455 phosphorylation played indispensable role in the regulation of ACLY activity and cell proliferation; however, they functioned independently from each other. In vivo, streptozocin treatment- and EGFR overexpression-induced growth of xenograft tumors was mitigated once S979 was mutated. Collectively, this work helps comprehend how cells interrogate the nutrient enrichment for proliferation and suggests that although mammalian cell proliferation is controlled by mitogen signaling, the ancient nutrition-sensing mechanism is conserved and still efficacious in the cells of multicellular organisms., Competing Interests: Competing interests statement:The authors declare no competing interest.

The starch metabolic network was investigated in relation to other metabolic processes by examining a mutant with altered single-gene expression of ATP citrate lyase (ACL), an enzyme responsible for generating cytosolic acetyl-CoA pool from citrate. Previous research has shown that transgenic antisense plants with reduced ACL activity accumulate abnormally enlarged starch granules. In this study, we explored the underlying molecular mechanisms linking cytosolic acetyl-CoA generation and starch metabolism under short-day photoperiods. We performed transcriptome and quantification of starch accumulation in the leaves of wild-type and antisense seedlings with reduced ACL activity. The antisense- ACLA mutant accumulated more starch than the wild type under short-day conditions. Zymogram analyses were conducted to compare the activities of starch-metabolizing enzymes with transcriptomic changes in the seedling. Differential expression between wild-type and antisense- ACLA plants was detected in genes implicated in starch and acetyl-CoA metabolism, and cell wall metabolism. These analyses revealed a strong correlation between the transcript levels of genes responsible for starch synthesis and degradation, reflecting coordinated regulation at the transcriptomic level. Furthermore, our data provide novel insights into the regulatory links between cytosolic acetyl-CoA metabolism and starch metabolic pathways.

Cardiac hypertrophy is a classical forerunner of heart failure and myocardial structural and metabolic remodeling are closely associated with cardiac hypertrophy. We aim to investigate the characteristics of myocardial structure and central carbon metabolism of cardiac hypertrophy at different stages. Using echocardiography and pathological staining, early and compensatory cardiac hypertrophy were respectively defined as within 7 days and from 7 to 14 days after transverse aortic constriction (TAC) in mice. Among mass-spectrometry-based metabolomics, we identified 45 central carbon metabolites. Differential metabolite analysis showed that six metabolites, including citrate, cis -aconitate and so on, decreased significantly on day 1 after TAC. Ten metabolites, including l-lactate, (S)-2-hydroxyglutarate and so on, were obviously changed on days 10 and 14. Pathway analysis showed that these metabolites were involved in seven metabolic pathways, including carbohydrates, amino acids and so on. Western blot showed the expression of ATP-citrate lyase, malate dehydrogenase 1 and lactate dehydrogenase A in myocardium changed markedly on day 3, while the phosphorylation level of AMP-activated protein kinase did not show significantly difference. We hope our research will promote deeper understanding and early diagnosis of cardiac hypertrophy in clinical practice. All raw data were deposited in MetaboLights (MTBLS10555).

An extensive phytochemical investigation on the EtOAc-soluble fraction of the 90% MeOH extract from the twigs and needles of the 'vulnerable' Chinese endemic conifer Tsuga forrestii (Forrest's hemlock) led to the isolation and characterization of 50 structurally diverse diterpenoids, including 15 unreported C-18 carboxylated ones (tsugaforrestiacids A-O, 1-15, resp.). Among them, compounds 1-7 are abieten-18-oic acids, compound 8 is an abieten-18-succinate, and compounds 10-12 are podocarpen-18-oic acids, whereas compounds 13-15 are pimarane-type, isopimarane-type, and totarane-type diterpenoid acids, respectively. Their structures and absolute configurations were determined by a combination of spectroscopic methods, GIAO NMR calculations and DP4+ probability analyses, electronic circular dichroism (ECD) data, and single crystal X-ray diffraction analyses. All the isolates were evaluated for their inhibitory activities against the ATP-citrate lyase (ACL), a key enzyme in cellular metabolism. Tsugaforrestiacids E (5) and H (8) were found to have significant inhibitory effects against ACL, with IC 50 values of 5.3 and 6.2 μM, respectively. The interactions of the bioactive molecules with the ACL enzyme were examined by molecular docking studies. The isolated diterpenoids also provide chemotaxonomic evidence to support the delimitation of Tsuga from its closest sister group (Nothotsuga). The above findings highlight the importance of protecting plant species with unique and diverse secondary metabolites, which may be potential sources of new therapeutic agents for the treating ACL-associated diseases., 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 © 2024 Elsevier Ltd. All rights reserved.)

The search for novel tumor biomarkers and targets is of significant importance for the early clinical diagnosis and treatment of Hepatocellular Carcinoma (HCC). The mechanisms by which ATP citrate lyase (ACLY) promotes HCC progression remain unclear, and the connection between ACLY and REGγ has not been reported in the literature. In vitro, we will perform overexpression/knockdown of ACLY or overexpression/knockdown of REGγ to investigate the impact of ACLY on HCC cells and its underlying mechanisms. In vivo, we will establish mouse tumor models with overexpression/knockdown of ACLY or overexpression/knockdown of REGγ to study the effect of ACLY on mouse tumors and its mechanisms. Firstly, ACLY overexpression upregulated REGγ expression and activated the REGγ-proteasome pathway, leading to changes in the expression of downstream signaling pathway proteins. This promoted HCC cell proliferation, invasion, and migration in vitro, as well as tumor growth and metastasis in vivo. Secondly, ACLY overexpression increased acetyl-CoA production, upregulated the acetylation level of the REGγ promoter region histone H3K27ac, and subsequently induced REGγ expression. Lastly, enhanced acetylation of the REGγ promoter region histone H3K27ac resulted in upregulated REGγ expression, activation of the REGγ-proteasome pathway, changes in downstream signaling pathway protein expression, and promotion of HCC cell proliferation, invasion, and migration in vitro, as well as tumor growth and metastasis in vivo. Conversely, REGγ knockdown reversed these effects. ACLY and REGγ may serve as potential biomarkers and clinical therapeutic targets for HCC., (© 2024 Wiley Periodicals LLC.)

Background and Aim: Cholangiocarcinoma (CCA) is a rare cancer, yet its incidence and mortality rates have been steadily increasing globally over the past few decades. Currently, there are no effective targeted treatment strategies available for patients. ACLY (ATP Citrate Lyase), a key enzyme in de novo lipogenesis, is aberrantly expressed in several tumors and is associated with malignant progression. However, its role and mechanisms in CCA have not yet been elucidated., Methods: The expression of ACLY in CCA was assessed using transcriptomic profiles and tissue microarrays. Kaplan-Meier curves were employed to evaluate the prognostic significance of ACLY in CCA. Functional enrichment analysis was used to explore the potential mechanisms of ACLY in CCA. A series of assays were conducted to examine the effects of ACLY on the proliferation and migration of CCA cells. Ferroptosis inducers and inhibitors, along with lipid peroxide probes and MDA assay kits, were utilized to explore the role of ACLY in ferroptosis within CCA. Additionally, lipid-depleted fetal bovine serum and several fatty acids were used to evaluate the impact of fatty acids on ferroptosis induced by ACLY inhibition. Correlation analyses were performed to elucidate the relationship between ACLY and tumor stemness as well as tumor microenvironment., Results: The expression of ACLY was found to be higher in CCA tissues compared to adjacent normal tissues. Patients with elevated ACLY expression demonstrated poorer overall survival outcomes. ACLY were closed associated with fatty acid metabolism and tumor-initiating cells. Knockdown of ACLY did not significantly impact the proliferation and migration of CCA cells. However, ACLY inhibition led to increased accumulation of lipid peroxides and enhanced sensitivity of CCA cells to ferroptosis inducers. Polyunsaturated fatty acids were observed to inhibit the proliferation of ACLY-knockdown cells; nonetheless, this inhibitory effect was diminished when the cells were cultured in medium supplemented with lipid-depleted fetal bovine serum. Additionally, ACLY expression was negatively correlated with immune cell infiltration and immune scores in CCA., Conclusion: ACLY promotes ferroptosis by disrupting the balance of saturated and unsaturated fatty acids. ACLY may therefore serve as a potential diagnostic and therapeutic target for CCA., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Sun, Zhao, Wang, Liu, Wei, Xu, Wang and Yang.)

Elevated production of extracellular matrix (ECM) in tumor stroma is a critical obstacle for drug penetration. Here we demonstrate that ATP-citrate lyase (ACLY) is significantly upregulated in cancer-associated fibroblasts (CAFs) to produce tumor ECM. Using a self-assembling nanoparticle-design approach, a carrier-free nanoagent (CFNA) is fabricated by simply assembling NDI-091143, a specific ACLY inhibitor, and doxorubicin (DOX) or paclitaxel (PTX), the first-line chemotherapeutic drug, via multiple noncovalent interactions. After arriving at the CAFs-rich tumor site, NDI-091143-mediated ACLY inhibition in CAFs can block the de novo synthesis of fatty acid, thereby dampening the fatty acid-involved energy metabolic process. As the lack of enough energy, the energetic CAFs will be in a dispirited state that is unable to produce abundant ECM, thereby significantly improving drug perfusion in tumors and enhancing the efficacy of chemotherapy. Such a simple drug assembling strategy aimed at CAFs' ACLY-mediated metabolism pathway presents the feasibility of stromal matrix reduction to potentiate chemotherapy.

Coordination of cellular metabolism is essential for optimal T cell responses. Here, we identify cytosolic acetyl-CoA production as an essential metabolic node for CD8 T cell function in vivo. We show that CD8 T cell responses to infection depend on acetyl-CoA derived from citrate via the enzyme ATP citrate lyase (ACLY). However, ablation of ACLY triggers an alternative, acetate-dependent pathway for acetyl-CoA production mediated by acyl-CoA synthetase short-chain family member 2 (ACSS2). Mechanistically, acetate fuels both the TCA cycle and cytosolic acetyl-CoA production, impacting T cell effector responses, acetate-dependent histone acetylation, and chromatin accessibility at effector gene loci. When ACLY is functional, ACSS2 is not required, suggesting acetate is not an obligate metabolic substrate for CD8 T cell function. However, loss of ACLY renders CD8 T cells dependent on acetate (via ACSS2) to maintain acetyl-CoA production and effector function. Together, ACLY and ACSS2 coordinate cytosolic acetyl-CoA production in CD8 T cells to maintain chromatin accessibility and T cell effector function., (© 2024 Kaymak et al.)

Insulin-like growth factor-1-induced activation of ATP citrate lyase (ACLY) improves muscle mitochondrial function through an Akt-dependent mechanism. In this study, we examined whether Akt1 deficiency alters skeletal muscle fiber type and mitochondrial function by regulating ACLY-dependent signaling in male Akt1 knockout (KO) mice (12-16 weeks old). Akt1 KO mice exhibited decreased body weight and muscle wet weight, with reduced cross-sectional areas of slow- and fast-type muscle fibers. Loss of Akt1 did not affect the phosphorylation status of ACLY in skeletal muscle. The skeletal muscle fiber type and expression of mitochondrial oxidative phosphorylation complex proteins were unchanged in Akt1 KO mice compared with the wild-type control. These observations indicate that Akt1 is important for the regulation of skeletal muscle fiber size, whereas the regulation of muscle fiber type and muscle mitochondrial content occurs independently of Akt1 activity., (© 2024 The Author(s). Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

ATP citrate lyase (ACLY) inhibitors have the potential of modulating central processes in protein, carbohydrate, and lipid metabolism, which can have relevant physiological consequences in aging and age-related diseases. Here, we show that hepatic phospho-active ACLY correlates with overweight and Model for End-stage Liver Disease score in humans. Wild-type mice treated chronically with the ACLY inhibitor potassium hydroxycitrate exhibited delayed early mortality. In AML12 hepatocyte cultures, the ACLY inhibitors potassium hydroxycitrate, SB-204990, and bempedoic acid fostered lipid accumulation, which was also observed in the liver of healthy-fed mice treated with potassium hydroxycitrate. Analysis of soleus tissue indicated that potassium hydroxycitrate produced the modulation of wound healing processes. In vivo, potassium hydroxycitrate modulated locomotor function toward increased wire hang performance and reduced rotarod performance in healthy-fed mice, and improved locomotion in mice exposed to cardiotoxin-induced muscle atrophy. Our findings implicate ACLY and ACLY inhibitors in different aspects of aging and muscle regeneration., (© 2024 The Authors. Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

In pancreatic ductal adenocarcinoma (PDAC), metabolic rewiring and resistance to standard therapy are closely associated. PDAC cells show enormous requirements for glucose-derived citrate, the first rate-limiting metabolite in the synthesis of new lipids. Both the expression and activity of citrate synthase (CS) are extraordinarily upregulated in PDAC. However, no previous relationship between gemcitabine response and citrate metabolism has been documented in pancreatic cancer. Here, we report for the first time that pharmacological doses of vitamin C are capable of exerting an inhibitory action on the activity of CS, reducing glucose-derived citrate levels. Moreover, ascorbate targets citrate metabolism towards the de novo lipogenesis pathway, impairing fatty acid synthase (FASN) and ATP citrate lyase (ACLY) expression. Lowered citrate availability was found to be directly associated with diminished proliferation and, remarkably, enhanced gemcitabine response. Moreover, the deregulated citrate-derived lipogenic pathway correlated with a remarkable decrease in extracellular pH through inhibition of lactate dehydrogenase (LDH) and overall reduced glycolytic metabolism. Modulation of citric acid metabolism in highly chemoresistant pancreatic adenocarcinoma, through molecules such as vitamin C, could be considered as a future clinical option to improve patient response to standard chemotherapy regimens., (© 2024 The Authors. Molecular Oncology published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Parrotia subaequalis, an endangered Tertiary relict tree native to China and a member of the Hamamelidaceae family, is one of several host plant species in this family that exhibit unique ecological habits, such as gall formation. Tree galls are the results of complex interactions between gall-inducing insects and their host plant organs. The formation of galls may serve to protect other regions of the plant from potential damage, often through the production of phytoalexins. In this study, a preliminary investigation was carried out on the metabolites of the 90% MeOH extract derived from the closed spherical galls on the twigs of P. subaequalis. Consequently, nine previously undescribed benzofuran-type and dibenzofuran-type phytoalexins (parrotiagallols A-I, 1-9, respectively) were isolated and characterized, along with several known miscellaneous metabolites (10-17). Their chemical structures and absolute configurations were elucidated using spectroscopic methods, a combination of calculated and experimental electronic circular dichroism data, and single crystal X-ray diffraction analyses. Among these compounds, 1 and 2 are identified as neolignan derivatives, while compounds 3-5 are classified as 9,10-dinorneolignans. Compound 6 represents a rare 2,3-seco-neolignan, and compounds 7-9 are dihydroxy-dimethyl-dibenzofuran derivatives. Parrotiagallol A (1) showed considerable antibacterial activity against Staphylococcus aureus, with an MIC value of 14 μM. Additionally, parrotiagallol E (5) and methyl gallate (17) exhibited inhibitory effects against ATP-citrate lyase (ACL), a potential therapeutic target for hyperlipidemia, with IC 50 values of 5.1 and 9.8 μM, respectively. The findings underscore that galls not only serve as physical defense barriers but also benefit from the chemical defense system of the host plants. These insights provide avenues for exploring potential new therapeutic agents for S. aureus infections and ACL-related diseases, while also promoting scientific conservation strategies for P. subaequalis., 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 © 2024 Elsevier Ltd. All rights reserved.)

The senescent microenvironment and aged cells per se contribute to tissue remodeling, chronic inflammation, and age-associated dysfunction. However, the metabolic and epigenomic bases of the senescence-associated secretory phenotype (SASP) remain largely unknown. Here, we show that ATP-citrate lyase (ACLY), a key enzyme in acetyl-coenzyme A (CoA) synthesis, is essential for the pro-inflammatory SASP, independent of persistent growth arrest in senescent cells. Citrate-derived acetyl-CoA facilitates the action of SASP gene enhancers. ACLY-dependent de novo enhancers augment the recruitment of the chromatin reader BRD4, which causes SASP activation. Consistently, specific inhibitions of the ACLY-BRD4 axis suppress the STAT1-mediated interferon response, creating the pro-inflammatory microenvironment in senescent cells and tissues. Our results demonstrate that ACLY-dependent citrate metabolism represents a selective target for controlling SASP designed to promote healthy aging., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Differentiation of cardiac fibroblasts to myofibroblasts is necessary for matrix remodeling and fibrosis in heart failure. We previously reported that mitochondrial calcium signaling drives α-ketoglutarate-dependent histone demethylation, promoting myofibroblast formation. Here we investigate the role of ATP-citrate lyase (ACLY), a key enzyme for acetyl-CoA biosynthesis, in histone acetylation regulating myofibroblast fate and persistence in cardiac fibrosis. We show that inactivation of ACLY prevents myofibroblast differentiation and reverses myofibroblasts towards quiescence. Genetic deletion of Acly in post-activated myofibroblasts prevents fibrosis and preserves cardiac function in pressure-overload heart failure. TGFβ stimulation enhances ACLY nuclear localization and ACLY-SMAD2/3 interaction, and increases H3K27ac at fibrotic gene loci. Pharmacological inhibition of ACLY or forced nuclear expression of a dominant-negative ACLY mutant prevents myofibroblast formation and H3K27ac. Our data indicate that nuclear ACLY activity is necessary for myofibroblast differentiation and persistence by maintaining histone acetylation at TGFβ-induced myofibroblast genes. These findings provide targets to prevent and reverse pathological fibrosis., (© 2024. The Author(s).)

ATP-citrate lyase (ACLY) links carbohydrate and lipid metabolism and provides nucleocytosolic acetyl-CoA for protein acetylation. ACLY has two major splice isoforms: the full-length canonical "long" isoform and an uncharacterized "short" isoform in which exon 14 is spliced out. Exon 14 encodes 10 amino acids within an intrinsically disordered region and includes at least one dynamically phosphorylated residue. Both isoforms are expressed in healthy tissues to varying degrees. Analysis of human transcriptomic data revealed that the percent spliced in (PSI) of exon 14 is increased in several cancers and correlated with poorer overall survival in a pan-cancer analysis, though not in individual tumor types. This prompted us to explore potential biochemical and functional differences between ACLY isoforms. Here, we show that there are no discernible differences in enzymatic activity or stability between isoforms or phosphomutants of ACLY in vitro. Similarly, both isoforms and phosphomutants were able to rescue ACLY functions, including fatty acid synthesis and bulk histone acetylation, when re-expressed in Acly knockout cells. Deletion of Acly exon 14 in mice did not overtly impact development or metabolic physiology nor did it attenuate tumor burden in a genetic model of intestinal cancer. Notably, expression of epithelial splicing regulatory protein 1 (ESRP1) is highly correlated with ACLY PSI. We report that ACLY splicing is regulated by ESRP1. In turn, both ESRP1 expression and ACLY PSI are correlated with specific immune signatures in tumors. Despite these intriguing patterns of ACLY splicing in healthy and cancer tissues, functional differences between the isoforms remain elusive., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Berberine was used as the lead compound in the present study to design and synthesize novel berberine derivatives by splicing bromine bridges of different berberine carbon chain lengths coupled nitric oxide donors, and their lipid lowering activities were assessed in a variety of ways. This experiment synthesized 17 new berberine nitric oxide donor derivatives. Compared with berberine hydrochloride, most of the compounds exhibited certain glycerate inhibitory activity, and compounds 6a, 6b, 6d, 12b and 12d showed higher inhibitory activity than berberine, with 6a, 6b and 6d having significant inhibitory activity. In addition, compound 6a linked to furazolidone nitric oxide donor showed better NO release in experiments; In further mechanistic studies, we screened and got two proteins, PCSK9 and ACLY, and docked two proteins with 17 compounds, and found that most of the compounds bound better with ATP citrate lyase (ACLY), among which there may be a strong interaction between compound 6a and ACLY, and the interaction force was better than the target drug Bempedoic Acid, which meaning that 6a may exert hypolipidemic effects by inhibiting ACLY; moreover, we also found that 6a may had the better performance in gastrointestinal absorption, blood-brain barrier permeability, Egan, Muegge class drug principle model calculation and bioavailability., Competing Interests: Declaration of competing interest This Project Supported by Natural Science Foundation of Heilongjiang Province(LH2022H013)., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background & Aims: Hypertrophic scar (HS) is a skin fibroproliferative disorder occurring after burns, surgeries or traumatic injuries, and it has caused a tremendous economic and medical burden. Its molecular mechanism is associated with the abnormal proliferation and transition of fibroblasts and excessive deposition of extracellular matrix. Cartilage intermediate layer protein 2 (CILP2), highly homologous to cartilage intermediate layer protein 1 (CILP1), is mainly secreted predominantly from chondrocytes in the middle/deeper layers of articular cartilage. Recent reports indicate that CILP2 is involved in the development of fibrotic diseases. We investigated the role of CILP2 in the progression of HS., Methods and Results: It was found in this study that CILP2 expression was significantly higher in HS than in normal skin, especially in myofibroblasts. In a clinical cohort, we discovered that CILP2 was more abundant in the serum of patients with HS, especially in the early stage of HS. In vitro studies indicated that knockdown of CILP2 suppressed proliferation, migration, myofibroblast activation and collagen synthesis of hypertrophic scar fibroblasts (HSFs). Further, we revealed that CILP2 interacts with ATP citrate lyase (ACLY), in which CILP2 stabilizes the expression of ACLY by reducing the ubiquitination of ACLY, therefore prompting Snail acetylation and avoiding reduced expression of Snail. In vivo studies indicated that knockdown of CILP2 or ACLY inhibitor, SB-204990, significantly alleviated HS formation., Conclusion: CILP2 exerts a vital role in hypertrophic scar formation and might be a detectable biomarker reflecting the progression of hypertrophic scar and a therapeutic target for hypertrophic scar., 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 © 2024 Elsevier B.V. All rights reserved.)

The impaired osteogenic capability of bone marrow mesenchymal stem cells (BMSCs) caused by persistent inflammation is the main pathogenesis of inflammatory bone diseases. Recent studies show that metabolism is disturbed in osteogenically differentiated BMSCs in response to Lipopolysaccharide (LPS) treatment, while the mechanism involved remains incompletely revealed. Herein, we demonstrated that BMSCs adapted their metabolism to regulate acetyl-coenzyme A (acetyl-CoA) availability and RNA acetylation level, ultimately affecting osteogenic differentiation. The mitochondrial dysfunction and impaired osteogenic potential upon inflammatory conditions accompanied by the reduced acetyl-CoA content, which in turn suppressed N4-acetylation (ac 4 C) level. Supplying acetyl-CoA by sodium citrate (SC) addition rescued ac 4 C level and promoted the osteogenic capacity of LPS-treated cells through the ATP citrate lyase (ACLY) pathway. N-acetyltransferase 10 (NAT10) inhibitor remodelin reduced ac 4 C level and consequently impeded osteogenic capacity. Meanwhile, the osteo-promotive effect of acetyl-CoA-dependent ac 4 C might be attributed to fatty acid oxidation (FAO), as evidenced by activating FAO by L-carnitine supplementation counteracted remodelin-induced inhibition of osteogenesis. Further in vivo experiments confirmed the promotive role of acetyl-CoA in the endogenous bone regeneration in rat inflammatory mandibular defects. Our study uncovered a metabolic-epigenetic axis comprising acetyl-CoA and ac 4 C modification in the process of inflammatory osteogenesis of BMSCs and suggested a new target for bone tissue repair in the context of inflammatory bone diseases., (Copyright © 2024. Published by Elsevier B.V.)

Lipid metabolic reprogramming of tumor cells has been proven to play a critical role in tumor initiation and development. However, lipid metabolism in cancer-associated fibroblasts (CAFs) has rarely been studied, particularly in CAFs of oral squamous cell carcinoma (OSCC). Additionally, the molecular mechanism by which tumor cells regulate lipid metabolism in fibroblasts is unclear. In this study, we found that phosphorylated ATP citrate lyase (p-ACLY), a key lipid metabolic enzyme, was upregulated in OSCC CAFs. Compared to paracancerous normal fibroblasts, CAFs showed enhanced lipid synthesis, such as elevated cytosolic acetyl-CoA level and accumulation of lipid droplets. Conversely, reduction of p-ACLY level blocked this biological process. In addition, blocking lipid synthesis in CAFs or inhibiting fatty acid uptake by OSCC cells reduced the promotive effects of CAFs on OSCC cell proliferation, invasion, and migration. These findings suggested that CAFs are one of lipid sources required for OSCC progression. Mechanistically, AKT signaling activation was involved in the upregulation of p-ACLY level and lipid synthesis in CAFs. Interleukin-8 (IL8), an exocrine cytokine of OSCC cells, could activate AKT and then phosphorylate ACLY in fibroblasts. This study suggested that the IL8/AKT/p-ACLY axis could be considered as a potential target for OSCC treatment., (© 2024 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

ATP citrate lyase, the first rate-limiting enzyme in de novo lipogenesis, plays a crucial role in tumour progression. This study explores ATP citrate lyase's potential as a tumour biomarker and its role in cutaneous squamous cell carcinoma. ATP citrate lyase expression patterns were analysed using TCGA and TIMER databases, and patient skin specimens were collected for immunohistochemistry to determine ATP citrate lyase levels. Cell proliferation, cell cycle, apoptosis, and c-Myc expression were assessed in A431 and SCL-1 cells. Stable cell lines with reduced ATP citrate lyase expression were obtained and subcutaneously implanted into nude mice to evaluate in vivo tumour growth. Ki67, c-Myc expression and TUNEL staining were analysed in subcutaneous tumours. ATP citrate lyase exhibited upregulation in various tumours, and showed significant associations with prognosis and immune infiltrate. Moreover, ATP citrate lyase was highly expressed in cutaneous squamous cell carcinoma. After ATP citrate lyase silencing, cutaneous squamous cell carcinoma cell growth decelerated, the cell cycle halted, cell apoptosis increased, and c-Myc expression decreased. Animal experiments revealed that, following ATP citrate lyase knockdown, tumour tissue growth slowed down, and there was a reduction in Ki-67 and c-Myc expression, accompanied by enhanced TUNEL staining. In conclusion, ATP citrate lyase may serve as a tumour biomarker. It is highly expressed in cutaneous squamous cell carcinoma and may serve as a therapeutic target.

Objective: Mucosal T cells play a major role in inflammatory bowel disease (IBD). However, their immunometabolism during intestinal inflammation is poorly understood. Due to its impact on cellular metabolism and proinflammatory immune cell function, we here focus on the enzyme ATP citrate lyase (ACLY) in mucosal T cell immunometabolism and its relevance for IBD., Design: ACLY expression and its immunometabolic impact on colitogenic T cell function were analysed in mucosal T cells from patients with IBD and in two experimental colitis models., Results: ACLY was markedly expressed in colon tissue under steady-state conditions but was significantly downregulated in lamina propria mononuclear cells in experimental dextran sodium sulfate-induced colitis and in CD4 + and to a lesser extent in CD8 + T cells infiltrating the inflamed gut in patients with IBD. ACLY-deficient CD4 + T cells showed an impaired capacity to induce intestinal inflammation in a transfer colitis model as compared with wild-type T cells. Assessment of T cell immunometabolism revealed that ACLY deficiency dampened the production of IBD-relevant cytokines and impaired glycolytic ATP production but enriched metabolites involved in the biosynthesis of phospholipids and phosphatidylcholine. Interestingly, the short-chain fatty acid butyrate was identified as a potent suppressor of ACLY expression in T cells, while IL-36α and resolvin E1 induced ACLY levels. In a translational approach, in vivo administration of the butyrate prodrug tributyrin downregulated mucosal infiltration of ACLY high CD4 + T cells and ameliorated chronic colitis., Conclusion: ACLY controls mucosal T cell immunometabolism and experimental colitis. Therapeutic modulation of ACLY expression in T cells emerges as a novel strategy to promote the resolution of intestinal inflammation., Competing Interests: Competing interests: MFN has served as an advisor for Pentax, Giuliani, MSD, Abbvie, Janssen, Takeda and Boehringer. Moreover, MFN serves as an associated editor of the journal Gut. The remaining authors disclose no conflicts., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Disease-associated astrocyte subsets contribute to the pathology of neurologic diseases, including multiple sclerosis and experimental autoimmune encephalomyelitis 1-8 (EAE), an experimental model for multiple sclerosis. However, little is known about the stability of these astrocyte subsets and their ability to integrate past stimulation events. Here we report the identification of an epigenetically controlled memory astrocyte subset that exhibits exacerbated pro-inflammatory responses upon rechallenge. Specifically, using a combination of single-cell RNA sequencing, assay for transposase-accessible chromatin with sequencing, chromatin immunoprecipitation with sequencing, focused interrogation of cells by nucleic acid detection and sequencing, and cell-specific in vivo CRISPR-Cas9-based genetic perturbation studies we established that astrocyte memory is controlled by the metabolic enzyme ATP-citrate lyase (ACLY), which produces acetyl coenzyme A (acetyl-CoA) that is used by histone acetyltransferase p300 to control chromatin accessibility. The number of ACLY + p300 + memory astrocytes is increased in acute and chronic EAE models, and their genetic inactivation ameliorated EAE. We also detected the pro-inflammatory memory phenotype in human astrocytes in vitro; single-cell RNA sequencing and immunohistochemistry studies detected increased numbers of ACLY + p300 + astrocytes in chronic multiple sclerosis lesions. In summary, these studies define an epigenetically controlled memory astrocyte subset that promotes CNS pathology in EAE and, potentially, multiple sclerosis. These findings may guide novel therapeutic approaches for multiple sclerosis and other neurologic diseases., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

ATP citrate lyase (ACLY), a strategic metabolic enzyme that catalyzes the glycolytic to lipidic metabolism, has gained increasing attention as an attractive therapeutic target for hyperlipidemia, cancers and other human diseases. Despite of continual research efforts, targeting ACLY has been very challenging. In this field, most reported ACLY inhibitors are "substrate-like" analogues, which occupied with the same active pockets. Besides, some ACLY inhibitors have been disclosed through biochemical screening or high throughput virtual screening. In this review, we briefly summarized the cancer-related functions and the recent advance of ACLY inhibitors with a particular focus on the SAR studies and their modes of action. We hope to provide a timely and updated overview of ACLY and the discovery of new ACLY inhibitors., 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 Inc.)

The 2'3'-cyclic GMP-AMP (cGAMP) synthase (cGAS)-stimulator of IFN genes (STING) pathway can sense infection and cellular stress by detecting cytosolic DNA. Upon ligand binding, cGAS produces the cyclic dinucleotide messenger cGAMP, which triggers its receptor STING. Active STING initiates gene transcription through the transcription factors IFN regulatory factor 3 (IRF3) and NF-κB and induces autophagy, but whether STING can cause changes in the metabolism of macrophages is unknown. In this study, we report that STING signaling activates ATP-citrate lyase (ACLY) by phosphorylation in human macrophages. Using genetic and pharmacologic perturbation, we show that STING targets ACLY via its prime downstream signaling effector TANK (TRAF family member-associated NF-κB activator)-binding kinase 1 (TBK1). We further identify that TBK1 alters cellular metabolism upon cGAMP treatment. Our results suggest that STING-mediated metabolic reprogramming adjusts the cellular response to DNA sensing in addition to transcription factor activation and autophagy induction., (Copyright © 2023 by The American Association of Immunologists, Inc.)

The hepatic SLC13A5/SLC25A1-ATP-dependent citrate lyase (ACLY) signaling pathway, responsible for maintaining the citrate homeostasis, plays a crucial role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Bempedoic acid (BA), an ACLY inhibitor commonly used for managing hypercholesterolemia, has shown promising results in addressing hepatic steatosis. This study aimed to elucidate the intricate relationships in processes of hepatic lipogenesis among SLC13A5, SLC25A1, and ACLY and to examine the therapeutic potential of BA in NAFLD, providing insights into its underlying mechanism. In murine primary hepatocytes and HepG2 cells, the silencing or pharmacological inhibition of SLC25A1/ACLY resulted in significant upregulation of SLC13A5 transcription and activity. This increase in SLC13A5 activity subsequently led to enhanced lipogenesis, indicating a compensatory role of SLC13A5 when the SLC25A1/ACLY pathway was inhibited. However, BA effectively counteracted this upregulation, reduced lipid accumulation, and ameliorated various biomarkers of NAFLD. The disease-modifying effects of BA were further confirmed in NAFLD mice. Mechanistic investigations revealed that BA could reverse the elevated transcription levels of SLC13A5 and ACLY, and the subsequent lipogenesis induced by PXR activation in vitro and in vivo. Importantly, this effect was diminished when PXR was knocked down, suggesting the involvement of the hepatic PXR-SLC13A5/ACLY signaling axis in the mechanism of BA action. In conclusion, SLC13A5-mediated extracellular citrate influx emerges as an alternative pathway to SLC25A1/ACLY in the regulation of lipogenesis in hepatocytes, BA exhibits therapeutic potential in NAFLD by suppressing the hepatic PXR-SLC13A5/ACLY signaling axis, while PXR, a key regulator in drug metabolism may be involved in the pathogenesis of NAFLD. SIGNIFICANCE STATEMENT: This work describes that bempedoic acid, an ATP-dependent citrate lyase (ACLY) inhibitor, ameliorates hepatic lipid accumulation and various hallmarks of non-alcoholic fatty liver disease. Suppression of hepatic SLC25A1-ACLY pathway upregulates SLC13A5 transcription, which in turn activates extracellular citrate influx and the subsequent DNL. Whereas in hepatocytes or the liver tissue challenged with high energy intake, bempedoic acid reverses compensatory activation of SLC13A5 via modulating the hepatic PXR-SLC13A5/ACLY axis, thereby simultaneously downregulating SLC13A5 and ACLY., (Copyright © 2023 by The American Society for Pharmacology and Experimental Therapeutics.)

Melanoma is the most lethal skin cancer originating from the malignant transformation of epidermal melanocyte. The dysregulation of cellular metabolism is a hallmark of cancer, including in melanoma. Aberrant branched-chain amino acids (BCAA) metabolism and related enzymes has been greatly implicated in the progression of multiple types of cancer, whereas remains far from understood in melanoma. Herein, we reported that the critical BCAA metabolism enzyme branched-chain amino acid transaminase 2 (BCAT2) is an oncogenic factor in melanoma by activating lipogenesis via the epigenetic regulation of fatty acid synthase (FASN) and ATP-citrate lyase (ACLY) expressions. Firstly, we found that BCAT2 expression was prominently increased in melanoma, and highly associated with clinical stage. Then, it was proved that the deficiency of BCAT2 led to impaired tumor cell proliferation, invasion and migration in vitro, and tumor growth and metastasis in vivo. Further, RNA sequencing technology and a panel of biochemical assays demonstrated that BCAT2 regulated de novo lipogenesis via the regulation of the expressions of both FASN and ACLY. Mechanistically, the inhibition of BCAT2 suppressed the generation of intracellular acetyl-CoA, mitigating P300-dependent histone acetylation at the promoter of FASN and ACLY, and thereby their transcription. Ultimately, zinc finger E-box binding homeobox 1 (ZEB1) was identified as the upstream transcriptional factor responsible for BCAT2 up-regulation in melanoma. Our results demonstrate that BCAT2 promotes melanoma progression by epigenetically regulating FASN and ACLY expressions via P300-dependent histone acetylation. Targeting BCAT2 could be exploited as a promising strategy to restrain tumor progression in melanoma., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

The dysregulation of branched-chain amino acid (BCAA) metabolism and related enzymes has been greatly implicated in the progression of multiple types of cancer, whereas remains far from understood in melanoma. Here, we explored the role of the BCAA metabolism enzyme BCKDHA in melanoma pathogenesis and elucidated the underlying mechanisms. In vitro cell biology experiments and in vivo pre-clinical mice model experiments were performed to investigate the role of BCKDHA in melanoma progression. RNA sequencing, immunohistochemical/immunofluorescence staining and bioinformatics analysis were used to examine the underlying mechanism. BCKDHA expression was prominently increased in both melanoma tissues and cell lines. The up-regulation of BCKDHA promoted long-term tumour cell proliferation, invasion and migration in vitro and tumour growth in vivo. Through RNA-sequencing technology, it was found that BCKDHA regulated the expressions of lipogenic fatty acid synthase (FASN) and ATP-citrate lyase (ACLY), which was thereafter proved to mediate the oncogenic role of BCKDHA in melanoma. Our results demonstrate that BCKDHA promotes melanoma progression by regulating FASN and ACLY expressions. Targeting BCKDHA could be exploited as a promising strategy to restrain tumour progression in melanoma., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Objective To investigate the role of ATP citrate lyase(ACLY)in the development of hepatocellular carcinoma(HCC)and the impact of this enzyme on the immune microenvironment of HCC.Methods We utilized the University of Alabama at Birmingham Cancer Data Analysis Portal and the Gene Expression Profiling Interactive Analysis to identify the changes in ACLY expression and prognosis across different tumor types from The Cancer Genome Atlas.With HCC as the disease model,we analyzed the ACLY expression in HCC samples from the gene expression database.Furthermore,we collected the clinical specimens from HCC patients to verify the mRNA and protein levels of ACLY.In addition,we conducted transcriptome sequencing after knocking down the expression of ACLY to analyze the differentially expressed genes and investigated the impact of ACLY expression interference on cell proliferation and other functions.Finally,we explored the correlations of ACLY with immune cells and immune infiltration in the tumor microenvironment,new antigens,and immune checkpoint genes.Results ACLY expression was significantly up-regulated in solid tumors including HCC(all P <0.05),and high ACLY expression was associated with overall survival rate in HCC( P =0.005).Furthermore,high ACLY expression affected the presence of immune cells(e.g.,tumor-associated fibroblasts)and the expression of genes involved in lipid metabolism(all P <0.05).Conclusions ACLY is closely related to the occurrence and development of HCC and lipid metabolism abnormalities.Moreover,it has a specific impact on the immune microenvironment of HCC.

Mounting evidence indicates the potential involvement of ATP-citrate lyase (ACLY) in the modulation of various cancer types. Nevertheless, the precise biological significance of ACLY in gastric cancer (GC) remains elusive. This study sought to elucidate the biological function of ACLY and uncover its influence on peritoneal metastasis in GC. The expression of ACLY was assessed using both real-time quantitative PCR and western blot techniques. To investigate the impact of ACLY on the proliferation of gastric cancer (GC) cells, colony formation and 5-ethynyl-2'-deoxyuridine (EdU) assays were performed. The migratory and invasive abilities of GC were evaluated using wound healing and transwell assays. Additionally, a bioinformatics analysis was employed to predict the correlation between ACLY and HIF-1A. This interaction was subsequently confirmed through a chromatin immunoprecipitation (ChIP) assay. ACLY exhibited upregulation in gastric cancer (GC) as well as in peritoneal metastasis. Its overexpression was found to facilitate the proliferation and metastasis of GC cells in both in vitro and in vivo experiments. Moreover, ACLY was observed to play a role in promoting angiogenesis and epithelial-mesenchymal transition (EMT). Notably, under hypoxic conditions, HIF-1A levels were elevated, thereby acting as a transcription factor to upregulate ACLY expression. Under the regulatory influence of HIF-1A, ACLY exerts a significant impact on the progression of gastric cancer, thereby facilitating peritoneal metastasis.

Inflammation-dependent changes in gene expression programs in innate immune cells, such as macrophages, involve extensive reprogramming of metabolism. This reprogramming is essential for the production of metabolites required for chromatin modifications, such as acetyl-CoA, and regulate their usage and availability impacting the macrophage epigenome. One of the most transcriptionally induced proinflammatory mediator is nitric oxide (NO), which has been shown to inhibit key metabolic enzymes involved in the production of these metabolites. Recent evidence indicates that NO inhibits mitochondrial enzymes such as pyruvate dehydrogenase (PDH) in macrophages induced by inflammatory stimulus. PDH is involved in the production of acetyl-CoA, which is essential for chromatin modifications in the nucleus, such as histone acetylation. In addition, acetyl-CoA levels in inflamed macrophages are regulated by ATP citrate lyase (ACLY) and citrate transporter SLC25A1. Interestingly, acetyl-CoA producing enzymes, such as PDH and ACLY, have also been reported to be present in the nucleus and to support the local generation of cofactors such as acetyl-CoA. Here, we will discuss the mechanisms involved in the regulation of acetyl-CoA production by metabolic enzymes, their inhibition by prolonged exposure to inflammation stimuli, their involvement in dynamic inflammatory expression changes and how these emerging findings could have significant implications for the design of novel therapeutic approaches., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Russo, Pileri and Ghisletti.)

Importance: Adenosine triphosphate citrate lyase (ACLY) is a key regulatory enzyme of glucose metabolism, cholesterol and fatty acid synthesis, and the inflammatory cascade. Bempedoic acid, an ACLY inhibitor, significantly reduces atherogenic lipid markers, including low-density lipoprotein cholesterol (LDL-C), non-high-density lipoprotein cholesterol, and apolipoprotein B. Additional effects of ACLY inhibition include antitumor growth; reduction of triglycerides and proinflammatory molecules such as high-sensitivity C-reactive protein; less insulin resistance; reduction of hepatic lipogenesis; and weight loss., Observations: While numerous ACLY inhibitors have been identified, most of the clinical data have focused on bempedoic acid. The Cholesterol Lowering via Bempedoic Acid, an ACL-Inhibiting Regimen (CLEAR) program was a series of phase 3 clinical trials that evaluated its effects on lipid parameters and safety, leading to US Food and Drug Administration approval in 2020. CLEAR Outcomes was a phase 3, double-blind, randomized, placebo-controlled trial in individuals with a history of statin intolerance, serum LDL-C level of 100 mg/dL or higher, and a history of, or at high risk for, cardiovascular disease. Bempedoic acid modestly reduced the primary 4-way cardiovascular composite end point as well as the individual components of myocardial infarction and coronary revascularization but did not reduce stroke, cardiovascular death, or all-cause mortality. Rates of gout and cholelithiasis were higher with bempedoic acid, and small increases in serum creatinine, uric acid, and hepatic-enzyme levels were also observed., Conclusions and Relevance: ACLY inhibition with bempedoic acid has been established as a safe and effective therapy in high-risk patients who require further LDL-C lowering, particularly for those with a history of statin intolerance. The recently published CLEAR Outcomes trial revealed modest reductions in cardiovascular events with bempedoic acid, proportional to its LDL-C lowering, in high-risk individuals with statin intolerance and LDL-C levels of 100 mg/dL or higher. The additional effects of ACLY inhibition have prompted a more thorough search for novel ACLY inhibitors for conditions such as cancer, hypertriglyceridemia, chronic inflammation, type 2 diabetes, fatty liver disease, obesity, and metabolic syndrome. Similarly, therapies that reduce fatty acid synthesis are being explored for their use in cardiometabolic conditions.

Sepsis involves endothelial cell (EC) dysfunction, which contributes to multiple organ failure. To improve therapeutic prospects, elucidating molecular mechanisms of vascular dysfunction is of the essence. ATP-citrate lyase (ACLY) directs glucose metabolic fluxes to de novo lipogenesis by generating acetyl-Co-enzyme A (acetyl-CoA), which facilitates transcriptional priming via protein acetylation. It is well illustrated that ACLY participates in promoting cancer metastasis and fatty liver diseases. Its biological functions in ECs during sepsis remain unclear. We found that plasma levels of ACLY were increased in septic patients and were positively correlated with interleukin (IL)-6, soluble E-selectin (sE-selectin), soluble vascular cell adhesion molecule 1 (sVCAM-1), and lactate levels. ACLY inhibition significantly ameliorated lipopolysaccharide challenge-induced EC proinflammatory response in vitro and organ injury in vivo. The metabolomic analysis revealed that ACLY blockade fostered ECs a quiescent status by reducing the levels of glycolytic and lipogenic metabolites. Mechanistically, ACLY promoted forkhead box O1 (FoxO1) and histone H3 acetylation, thereby increasing the transcription of c-Myc (MYC) to facilitate the expression of proinflammatory and gluco-lipogenic genes. Our findings revealed that ACLY promoted EC gluco-lipogenic metabolism and proinflammatory response through acetylation-mediated MYC transcription, suggesting ACLY as the potential therapeutic target for treating sepsis-associated EC dysfunction and organ injury., (© 2023. The Author(s).)

HB (hepatoblastoma) is most common in children with liver cancer and few options for treating HB. Thus, it is of great significance to investigate the regulatory mechanism of HB and/or identify new therapeutic targets for clinical treatment of HB. Here, we showed that ACLY (ATP citrate lyase), an important lipometabolic enzyme for de novo biosynthesis of fatty acids and steroids, has a higher expression in HB tissues than noncancerous tissues, and is required for HB cell proliferation. Moreover, knocking down ACLY in HB cells caused severe S-phase arrest and apoptosis. Mechanistically, ACLY knockdown significantly silenced the Wnt signaling pathway and reduced β-catenin expression in HB cells. Conversely, the apoptotic alleviation of HB cells by overexpressing ACLY was blocked by silencing β-catenin, suggesting the modulation of HB cells by ACLY-β-catenin axis. Our results uncovered the role of ACLY in HB cells and presented a theoretical approach for HB targeted therapy in the future., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Acetyl-CoA utilized by histone acetyltransferases (HAT) for chromatin modification is mainly generated by ATP-citrate lyase (ACL) from glucose sources. How ACL locally establishes acetyl-CoA production for histone acetylation remains unclear. Here we show that ACL subunit A2 (ACLA2) is present in nuclear condensates, is required for nuclear acetyl-CoA accumulation and acetylation of specific histone lysine residues, and interacts with Histone AcetylTransferase1 (HAT1) in rice. The rice HAT1 acetylates histone H4K5 and H4K16 and its activity on H4K5 requires ACLA2. Mutations of rice ACLA2 and HAT1 (HAG704) genes impair cell division in developing endosperm, result in decreases of H4K5 acetylation at largely the same genomic regions, affect the expression of similar sets of genes, and lead to cell cycle S phase stagnation in the endosperm dividing nuclei. These results indicate that the HAT1-ACLA2 module selectively promotes histone lysine acetylation in specific genomic regions and unravel a mechanism of local acetyl-CoA production which couples energy metabolism with cell division., (© 2023. The Author(s).)

Proliferating cells rely on acetyl-CoA to support membrane biogenesis and acetylation. Several organelle-specific pathways are available for provision of acetyl-CoA as nutrient availability fluctuates, so understanding how cells maintain acetyl-CoA homeostasis under such stresses is critically important. To this end, we applied 13 C isotope tracing cell lines deficient in these mitochondrial [ATP-citrate lyase (ACLY)]-, cytosolic [acetyl-CoA synthetase (ACSS2)]-, and peroxisomal [peroxisomal biogenesis factor 5 (PEX5)]-dependent pathways. ACLY knockout in multiple cell lines reduced fatty acid synthesis and increased reliance on extracellular lipids or acetate. Knockout of both ACLY and ACSS2 (DKO) severely stunted but did not entirely block proliferation, suggesting that alternate pathways can support acetyl-CoA homeostasis. Metabolic tracing and PEX5 knockout studies link peroxisomal oxidation of exogenous lipids as a major source of acetyl-CoA for lipogenesis and histone acetylation in cells lacking ACLY, highlighting a role for inter-organelle cross-talk in supporting cell survival in response to nutrient fluctuations.

Background: New therapeutic approaches are required to improve the outcomes of lung cancer (LC), a leading cause of cancer-related deaths worldwide. Chinese herbal medicine formulae widely used in China provide a unique opportunity for improving LC treatment, and the Shuang-Huang-Sheng-Bai (SHSB) formula is a typical example. However, the underlying mechanisms of action remains unclear., Purpose: This study aimed to confirm the efficacy of SHSB against lung adenocarcinoma (LUAD), which is a major histological type of LC, unveil the downstream targets of this formula, and assess the clinical relevance and biological roles of the newly identified target., Methods: An experimental metastasis mouse model and a subcutaneous xenograft mouse model were used to evaluate the anti-cancer activity of SHSB. Multi-omics profiling of subcutaneous tumors and metabolomic profiling of sera were performed to identify downstream targets, especially the metabolic targets of SHSB. A clinical trial was conducted to verify the newly identified metabolic targets in patients. Next, the metabolites and enzymes engaged in the metabolic pathway targeted by SHSB were measured in clinical samples. Finally, routine molecular experiments were performed to decipher the biological functions of the metabolic pathways targeted by SHSB., Results: Oral SHSB administration showed overt anti-LUAD efficacy as revealed by the extended overall survival of the metastasis model and impaired growth of implanted tumors in the subcutaneous xenograft model. Mechanistically, SHSB administration altered protein expression in the post-transcriptional layer and modified the metabolome of LUAD xenografts. Integrative analysis demonstrated that SHSB markedly inhibited acetyl-CoA synthesis in tumors by post-transcriptionally downregulating ATP-citrate lyase (ACLY). Consistently, our clinical trial showed that oral SHSB administration declined serum acetyl-CoA levels of patients with LC. Moreover, acetyl-CoA synthesis and ACLY expression were both augmented in clinical LUAD tissues of patients, and high intratumoral ACLY expression predicted a detrimental prognosis. Finally, we showed that ACLY-mediated acetyl-CoA synthesis is essential for LUAD cell growth by promoting G1/S transition and DNA replication., Conclusion: Limited downstream targets of SHSB for LC treatment have been reported in previous hypothesis-driven studies. In this study, we conducted a comprehensive multi-omics investigation and demonstrated that SHSB exerted its anti-LUAD efficacy by actively and post-transcriptionally modulating protein expression and particularly restraining ACLY-mediated acetyl-CoA synthesis., Competing Interests: Declaration of Competing Interest The authors indicate no conflicts of interest., (Copyright © 2023 The Authors. Published by Elsevier GmbH.. All rights reserved.)

ATP citrate lyase (ACLY) is the predominant nucleocytosolic source of acetyl-CoA and is aberrantly regulated in many diseases making it an attractive therapeutic target. Structural studies of ACLY reveal a central homotetrameric core citrate synthase homology (CSH) module flanked by acyl-CoA synthetase homology (ASH) domains, with ATP and citrate binding the ASH domain and CoA binding the ASH-CSH interface to produce acetyl-CoA and oxaloacetate products. The specific catalytic role of the CSH module and an essential D1026A residue contained within it has been a matter of debate. Here, we report biochemical and structural analysis of an ACLY-D1026A mutant demonstrating that this mutant traps a (3S)-citryl-CoA intermediate in the ASH domain in a configuration that is incompatible with the formation of acetyl-CoA, is able to convert acetyl-CoA and OAA to (3S)-citryl-CoA in the ASH domain, and can load CoA and unload acetyl-CoA in the CSH module. Together, this data support an allosteric role for the CSH module in ACLY catalysis., (© 2023. The Author(s).)

ATP-citrate lyase is a central integrator of cellular metabolism in the interface of protein, carbohydrate, and lipid metabolism. The physiological consequences as well as the molecular mechanisms orchestrating the response to long-term pharmacologically induced Acly inhibition are unknown. We report here that the Acly inhibitor SB-204990 improves metabolic health and physical strength in wild-type mice when fed with a high-fat diet, while in mice fed with healthy diet results in metabolic imbalance and moderated insulin resistance. By applying a multiomic approach using untargeted metabolomics, transcriptomics, and proteomics, we determined that, in vivo, SB-204990 plays a role in the regulation of molecular mechanisms associated with aging, such as energy metabolism, mitochondrial function, mTOR signaling, and folate cycle, while global alterations on histone acetylation are absent. Our findings indicate a mechanism for regulating molecular pathways of aging that prevents the development of metabolic abnormalities associated with unhealthy dieting. This strategy might be explored for devising therapeutic approaches to prevent metabolic diseases., (© 2023. The Author(s).)

The generation of neurons from quiescent radial-glia-like neural stem cells (RGLs) in adult brain goes hand in hand with the modulation of cellular metabolism. However, it is still unclear how the exact metabolic program governs the balance between quiescent and activated RGLs. Here, we find that loss of mitochondrial D-2-hydroxyglutarate dehydrogenase (D2HGDH) leads to aberrant accumulation of D-2-hydroxyglutarate (D-2-HG) and impaired RGL activation. Mechanistically, accumulated D-2-HG bonds directly to ATP-citrate lyase and competitively inhibits its enzymatic activity, thereby reducing acetyl-CoA production and diminishing histone acetylation. However, administration of acetate restores the acetyl-CoA levels via acetyl-CoA synthetase-mediated catabolism and rescues the deficiencies in histone acetylation and RGL activation caused by loss of D2HGDH. Therefore, our findings define the role of cross talk between mitochondria and the nucleus via a mitochondrial metabolite, D-2-HG, the aberrant accumulation of which hinders the regulation of histone acetylation in RGL activation and attenuates continuous neurogenesis in adult mammalian brain., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Macropinocytosis is an actin-dependent mode of nonselective endocytosis that mediates the uptake of extracellular fluid-phase cargoes. It is now well recognized that tumor cells exploit macropinocytosis to internalize macromolecules that can be catabolized and used to support cell growth and proliferation under nutrient-limiting conditions. Therefore, the identification of molecular mechanisms that control macropinocytosis is fundamental to the understanding of the metabolic adaptive landscape of tumor cells. Here, we report that the acetyl-CoA-producing enzyme, ATP citrate lyase (ACLY), is a key regulator of macropinocytosis and describes a heretofore-unappreciated association of ACLY with the actin cytoskeleton. The cytoskeletal tethering of ACLY is required for the spatially defined acetylation of heterodimeric actin capping protein, which we identify as an essential mediator of the actin remodeling events that drive membrane ruffling and macropinocytosis. Furthermore, we identify a requirement for mitochondrial-derived citrate, an ACLY substrate, for macropinocytosis, and show that mitochondria traffic to cell periphery regions juxtaposed to plasma membrane ruffles. Collectively, these findings establish a mode of metabolite compartmentalization that supports the spatiotemporal modulation of membrane-cytoskeletal interactions required for macropinocytosis by coupling regional acetyl-CoA availability with dynamic protein acetylation.

Macroautophagy/autophagy is a cellular and energy homeostatic mechanism that contributes to maintain the number of primordial follicles, germ cell survival, and anti-ovarian aging. However, it remains unknown whether autophagy in granulosa cells affects oocyte maturation. Here, we show a clear tendency of reduced autophagy level in human granulosa cells from women of advanced maternal age, implying a potential negative correlation between autophagy levels and oocyte quality. We therefore established a co-culture system and show that either pharmacological inhibition or genetic ablation of autophagy in granulosa cells negatively affect oocyte quality and fertilization ability. Moreover, our metabolomics analysis indicates that the adverse impact of autophagy impairment on oocyte quality is mediated by downregulated citrate levels, while exogenous supplementation of citrate can significantly restore the oocyte maturation. Mechanistically, we found that ACLY (ATP citrate lyase), which is a crucial enzyme catalyzing the cleavage of citrate, was preferentially associated with K63-linked ubiquitin chains and recognized by the autophagy receptor protein SQSTM1/p62 for selective autophagic degradation. In human follicles, the autophagy level in granulosa cells was downregulated with maternal aging, accompanied by decreased citrate in the follicular fluid, implying a potential correlation between citrate metabolism and oocyte quality. We also show that elevated citrate levels in porcine follicular fluid promote oocyte maturation. Collectively, our data reveal that autophagy in granulosa cells is a beneficial mechanism to maintain a certain degree of citrate by selectively targeting ACLY during oocyte maturation. Abbreviations: 3-MA: 3-methyladenine; ACLY: ATP citrate lyase; AMA: advanced maternal age; CG: cortical granule; CHX: cycloheximide; CQ: chloroquine; CS: citrate synthase; COCs: cumulus-oocyte-complexes; GCM: granulosa cell monolayer; GV: germinal vesicle; MII: metaphase II stage of meiosis; PB1: first polar body; ROS: reactive oxygen species; shRNA: small hairpin RNA; SQSTM1/p62: sequestosome 1; TCA: tricarboxylic acid; TOMM20/TOM20: translocase of outer mitochondrial membrane 20; UBA: ubiquitin-associated domain; Ub: ubiquitin; WT: wild-type.

Aquatic organisms are exposed to complex mixtures of pesticides in the environment, but traditional risk assessment approaches typically only consider individual compounds. In conjunction with exposure to pesticide mixtures, global climate change is anticipated to alter thermal regimes of waterways, leading to potential co-exposure of biota to elevated temperatures and contaminants. Furthermore, most studies utilize aqueous exposures, whereas the dietary route of exposure may be more important for fish owing to the hydrophobicity of many pesticides. Consequently, the current study aimed to determine the effects of elevated temperatures and dietary pesticide mixtures on swimming performance and lipid metabolism of juvenile Chinook salmon, Oncorhynchus tshawytscha. Fish were fed pesticide-dosed pellets at three concentrations and three temperatures (11, 14 and 17 °C) for 14 days and swimming performance (U max ) and expression of genes involved in lipid metabolism and energetics were assessed (ATP citrate lyase, fatty acid synthase, farnesoid x receptor and liver x receptor). The low-pesticide pellet treatment contained five pesticides, p,p'-DDE, bifenthrin, esfenvalerate, chlorpyrifos and fipronil at concentrations based on prey items collected from the Sacramento River (CA, USA) watershed, with the high-pesticide pellet treatment containing a six times higher dose. Temperature exacerbated effects of pesticide exposure on swimming performance, with significant reductions in U max of 31 and 23% in the low and high-pesticide pellet groups relative to controls at 17 °C, but no significant differences in U max among pesticide concentrations at 11 or 14 °C. At 14 °C there was a significant positive relationship between juvenile Chinook salmon pesticide body residues and expression of ATP citrate lyase and fatty acid synthase, but an inverse relationship and significant downregulation at 17 °C. These findings suggest that temperature may modulate effects of environmentally relevant pesticide exposure on salmon, and that pesticide-induced impairment of swimming performance may be exacerbated under future climate scenarios., 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 © 2022 Elsevier Ltd. All rights reserved.)

ATP-citrate lyase (ACLY) is a key enzyme provoking metabolic and epigenetic gene regulation. Molecularly, these functions are exerted by the provision of acetyl-coenzyme A, which is then used as a substrate for de novo lipogenesis or as an acetyl-group donor in acetylation reactions. It has been demonstrated that ACLY activity can be positively regulated via phosphorylation at serine 455 by Akt and protein kinase A. Nonetheless, the impact of phosphorylation on ACLY function in human myeloid cells is poorly understood. In this study we reconstituted ACLY knockout human monocytic THP-1 cells with a wild type ACLY as well as catalytically inactive H760A, and phosphorylation-deficient S455A mutants. Using these cell lines, we determined the impact of ACLY activity and phosphorylation on histone acetylation and pro-inflammatory gene expression in response to lipopolysaccharide (LPS). Our results show that ACLY serine 455 phosphorylation does not influence the proper enzymatic function of ACLY, since both, wild type ACLY and phosphorylation-deficient mutant, exhibited increased cell growth and histone acetylation as compared to cells with a loss of ACLY activity. Transcriptome analysis revealed enhanced expression of pro-inflammatory and interferon response genes in ACLY knockout and H760A THP-1 cells under unstimulated or LPS-treated conditions. At the same time, S455A ACLY-expressing cells showed a phenotype very similar to wild type cells. Contrary to ACLY knockout, pharmacological inhibition of ACLY in THP-1 cells or in primary human macrophages does not enhance LPS-triggered pro-inflammatory gene expression. Our data thus suggest that ACLY retains functionality in the absence of Akt/PKA-mediated phosphorylation in human myeloid cells. Furthermore, loss of ACLY activity may elicit long-term adaptive mechanisms, increasing inflammatory responses., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Dominguez, Truemper, Mota, Brüne and Namgaladze.)

Aim: The present study aimed mainly to demonstrate the metabolic effects of lycopene (LYC) or atorvastatin (ATOR) in diabetic hyperlipidemic rat model., Main Methods: Rats were randomly classified into four groups; the first was fed normal chow diet (NC) while the other three groups received streptozotocin (STZ) along with CCT-diet. The second group received no treatment (diabetic hyperlipidemic control, DHC), the third one received ATOR (50 mg/kg/day) while the fourth one received LYC (20 mg/kg/day). Serum and tissue samples were collected for biochemical and histological evaluations., Key Findings: DHC rats demonstrated significant hyperglycemia, dyslipidemia, increased hepatic fatty acid synthetase (FAS), malondialdehyde (MDA), tumor necrosis factor- alpha (TNF-α), 3-hydroxy 3-methylglutaryl coenzyme A (HMG-CoA) reductase and ATP citrate lyase (ACLY). However, hepatic reduced glutathione (GSH) and phosphorylated form of AMP-activated protein kinase (AMPK-P) activities showed significant decreases. ATOR or LYC administration induced hypoglycemic and hypolipidemic effects; decreased hepatic levels of MDA, TNF-α, HMG-CoA reductase, ACLY and FAS along with GSH and AMPK-P increases. Histopathological findings showed clear correlation with the biomarkers results., Significance: LYC demonstrated favorable significant effects regarding the biomarkers studied as compared to ATOR and may be expressed as a potent therapeutic agent of natural origin for hyperlipidemia complications either alone or in combination with other hypolipidemic drugs., Competing Interests: Declaration of competing interest The authors declare no conflict of interest to disclose., (Copyright © 2022 Elsevier Inc. All rights reserved.)

ATP citrate lyase (ACLY) is a key enzyme in glucolipid metabolism with therapeutic prospect for treating hyperlipidemia and various cancers. Much effort has been put into discovering ACLY inhibitors. However, current screening approaches have limitations in sensitivity, portability and high-throughput. To develop a general screening assay, we investigated series of conditions affecting the enzymatic reaction based on the ADP-Glo luminescence assay. Bovine serum albumin (0.001%) added triggered strong and stable fluorescence signal. The optimized assay was validated and applied to screen our natural product library. Two novel inhibitors were identified with IC 50 values of 3.86 ± 0.62 μM (2) and 15.48 ± 2.51 μM (4). Their aggregations and target specificities were also examined. 2 was characterized as a noncompetitive inhibitor of ACLY, while 4 was a competitive inhibitor of CoA, which was also elucidated by docking studies. In anticancer activity evaluation, 2 with higher inhibition potency did not exhibit anticancer effect, probably owing to its insufficient cell-permeability. 4 showed moderate inhibition in the proliferation of A549 and PC3 cells. This study not only developed a general approach for ACLY inhibitor discovery, but also identified a new scaffold ACLY inhibitor, which could be served as a hit compound in drug design., 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 © 2022 Elsevier B.V. All rights reserved.)