Inhibition Of acetyl-CoA Carboxylase Research Articles

Combinations of an acetyl CoA carboxylase inhibitor with hepatic lipid modulating agents do not augment antifibrotic efficacy in preclinical models of NASH and fibrosis.

Dysregulated hepatocyte lipid metabolism is a hallmark of hepatic lipotoxicity and contributes to the pathogenesis of nonalcoholic steatohepatitis (NASH). Acetyl CoA carboxylase (ACC) inhibitors decrease hepatocyte lipotoxicity by inhibiting de novo lipogenesis and concomitantly increasing fatty acid oxidation (FAO), and firsocostat, a liver‐targeted inhibitor of ACC1/2, is under evaluation clinically in patients with NASH. ACC inhibition is associated with improvements in indices of NASH and reduced liver triglyceride (TG) content, but also increased circulating TG in subjects with NASH and preclinical rodent models. Here we evaluated whether enhancing hepatocyte FAO by combining ACC inhibitors with peroxisomal proliferator‐activated receptor (PPAR) or thyroid hormone receptor beta (THRβ) agonists could drive greater liver TG reduction and NASH/antifibrotic efficacy, while ameliorating ACC inhibitor–induced hypertriglyceridemia. In high‐fat diet–fed dyslipidemic rats, the addition of PPAR agonists fenofibrate (Feno), elafibranor (Ela), lanifibranor (Lani), seladelpar (Sela) or saroglitazar (Saro), or the THRb agonist resmetirom (Res), to an analogue of firsocostat (ACCi) prevented ACCi‐induced hypertriglyceridemia. However, only PPARα agonists (Feno and Ela) and Res provided additional liver TG reduction. In the choline‐deficient high‐fat diet rat model of advanced liver fibrosis, neither PPARα (Feno) nor THRβ (Res) agonism augmented the antifibrotic efficacy of ACCi. Conclusion: These data suggest that combination therapies targeting hepatocyte lipid metabolism may have beneficial effects on liver TG reduction; however, they may not be sufficient to drive fibrosis regression.

Abstract 3018: Targeting lipid metabolism and diet in advanced prostate cancer

Abstract Lipid metabolism reprogramming provides the energetic needs of prostate cancer cells (PCa) to support growth and cell progress through the cell cycle, sustaining a highly proliferative phenotype by increasing the expression levels of several enzymes involved in fatty acid (FAs) synthesis. Fatty acid synthase (FASN), a key enzyme in de novo fatty acid lipogenesis (DNL), is upregulated in metastatic, castration-resistant prostate cancer (CRPC), and its overexpression is related to carcinogenesis, growth, and metastasis. We previously demonstrated that FASN inhibitors show antitumor activity suppressing PCa cell proliferation and slowing tumor growth, both in vitro and in vivo, via rewiring of lipid cancer metabolism and inducing endoplasmic reticulum stress.We observed that impairment of DNL increases polyunsaturated fatty acids (PUFA) uptake as a compensatory mechanism of survival. Exogenously acquired PUFAs cause the remodeling of the prostate cancer cell lipidome, manifesting an enhanced acyl chain unsaturation across several lipid species. Lipid peroxidation and ROS accumulation are also observed, suggesting cell susceptible to oxidative damage following FASN blockade. Alterations in metabolic pathways cause a higher uptake of acetate and glutamine in PCa cells, as assessed by 14C-nutrient incorporation following FASN inhibition. Mitochondrial dysfunction is induced by reduction in several respiratory parameters, including ATP production and basal respiration. This may be, at least in part, the result of a reduction in phosphatidylglycerol, a cardiolipin precursor, altering mitochondrial membrane composition. Supplementation with the PUFA docosahexaenoic acid (DHA) increases phosphatidylcholine levels and total phospholipid acyl chain unsaturation, which can lead to membrane disorganization in prostate cancer cells. Superoxide anion production is enhanced by DHA treatment, as expected, and increased levels of glucose and palmitate oxidation by mitochondria are observed. Interestingly, AR and AR-V7 protein expression, as well as c-MYC and PSA, is reduced with DHA treatment. We then evaluated how de novo lipid synthesis inhibition combined with manipulation of the exogenous supply of fatty acids could impair tumor cell progression. Due to the obligate cell dependence on PUFA following DNL suppression, we combined FASN and Acetyl-CoA carboxylase (ACC) inhibitors with DHA supplementation and observed a significant cell growth inhibition in CRPC cell lines and prostate cancer organoids.Altogether, our results suggest a novel mechanism to inhibit the growth of castration-resistant prostate cancer, combining lipogenesis inhibition with PUFA-enriched diet to overcome advanced disease. Citation Format: Silvia D. Rodrigues, Caroline Fidalgo Ribeiro, Guilherme H. Tamarindo, Hubert Pakula, Massimo Loda. Targeting lipid metabolism and diet in advanced prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3018.

Label-Free Imaging of Lipid Droplets in Prostate Cells Using Stimulated Raman Scattering Microscopy and Multivariate Analysis.

Hyperspectral stimulated Raman scattering (SRS) microscopy is a powerful imaging modality for the analysis of biological systems. Here, we report the application of k-means cluster analysis (KMCA) of multi-wavelength SRS images in the high-wavenumber region of the Raman spectrum as a robust and reliable method for the segmentation of cellular organelles based on the intrinsic SRS spectrum. KMCA has been applied to the study of the endogenous lipid biochemistry of prostate cancer and prostate healthy cell models, while the corresponding SRS spectrum of the lipid droplet (LD) cluster enabled direct comparison of their composition. The application of KMCA in visualizing the LD content of prostate cell models following the inhibition of de novo lipid synthesis (DNL) using the acetyl-coA carboxylase inhibitor, 5-(tetradecyloxy)-2-furoic acid (TOFA), is demonstrated. This method identified a reliance of prostate cancer cell models upon DNL for metabolic requirements, with a significant reduction in the cellular LD content after treatment with TOFA, which was not observed in normal prostate cell models. SRS imaging combined with KMCA is a robust method for investigating drug–cell interactions in a label-free manner.

275-OR: Inhibition of Hepatic ACC Decreases Ketogenesis during Fasting due to Elevated Amino Acid Availability

Activation of fatty acid oxidation is a promising therapy for NAFLD. Lowering acetyl-CoA carboxylase (ACC) activity depletes malonyl-CoA, activates CPT1, and increases mitochondrial fatty acid delivery. ACC1/2 liver double KO mice (LDKO) are protected from hepatic steatosis and have elevated ketogenesis in the prandial state. However, fasted LDKO mice exhibit an unexpected inhibition of ketogenesis, indicated by lower blood ketones and reduced in vivo ketone turnover of [3,4-13C2]acetoacetate and [U-13C4]β-hydroxybutyrate. Despite impaired fasting ketogenesis in vivo, perfused livers isolated from fasted mice and starved primary hepatocytes had normal or elevated ketone production, suggesting that a substrate-mediated mechanism inhibits ketosis in vivo. There were no differences in plasma NEFAs, but hepatic lactate was elevated during fasting in LDKO mice. The increase in lactate was sufficient to suppress ketone production in isolated perfused liver. An in vivo [U-13C3]lactate/pyruvate challenge was more rapidly diluted in fasted LKDO mice, indicating that alternative endogenous sources of pyruvate were available. Indeed, essential amino acids were elevated in fasted LDKO liver, but not plasma, suggestive of hepatic proteolysis as a source of amino acids. Liver size was 20% larger in LDKO mice, independent of glycogen storage or triglyceride content, but was consistent with increased insulin action and activation of Nrf2 which have been linked to hepatomegaly. Autophagy during fasting was normal or increased, suggesting that hepatomegaly supplies excess amino acids in the fasted state. Injection of exogenous alanine or glutamine in fasted WT mice recapitulated the fasted LDKO phenotype by increasing plasma glucose and lactate and decreasing ketone concentrations. Thus, hepatic growth is increased in the absence of malonyl-CoA synthesis, which provides a surplus of amino acids, anaplerotic maintenance of the TCA cycle and lower ketogenesis during fasting. Disclosure S.Deja: None. S.C.Burgess: n/a. J.A.Fletcher: None. B.Kucejova: None. M.N.Mizerska: None. X.Fu: None. C.Kim: None. J.Browning: None. J.Young: Consultant; Pfizer Inc., Research Support; Merck & Co., Inc., Stock/Shareholder; Metalytics, Inc. J.D.Horton: Board Member; Draupnir Bio, Consultant; Eli Lilly and Company, ESPERION Therapeutics, Inc., Gilead Sciences, Inc., Janssen Pharmaceuticals, Inc., Pfizer Inc., Regeneron Pharmaceuticals Inc. Funding National Institutes of Health (R01DK078184, P41EB015908)

Inhibition of ATP-citrate lyase improves NASH, liver fibrosis, and dyslipidemia

Elevated liver de novo lipogenesis contributes to non-alcoholic steatohepatitis (NASH) and can be inhibited by targeting acetyl-CoA carboxylase (ACC). However, hypertriglyceridemia limits the use of pharmacological ACC inhibitors as a monotherapy. ATP-citrate lyase (ACLY) generates acetyl-CoA and oxaloacetate from citrate, but whether inhibition is effective for treating NASH is unknown. Here, we characterize a new mouse model that replicates many of the pathological and molecular drivers of NASH and find that genetically inhibiting ACLY in hepatocytes reduces liver malonyl-CoA, oxaloacetate, steatosis, and ballooning as well as blood glucose, triglycerides, and cholesterol. Pharmacological inhibition of ACLY mirrors genetic inhibition but has additional positive effects on hepatic stellate cells, liver inflammation, and fibrosis. Mendelian randomization of human variants that mimic reductions in ACLY also associate with lower circulating triglycerides and biomarkers of NASH. These data indicate that inhibiting liver ACLY may be an effective approach for treatment of NASH and dyslipidemia.

Multiple resistance to ACCase- and ALS-inhibiting herbicides in black-grass (Alopecurus myosuroides Huds.) in China

Two black-grass (Alopecurus myosuroides Huds.) populations (R2105 and R1027) that were suspected to be resistant to clodinafop-propargyl, an acetyl-CoAcarboxylase (ACCase) inhibitor, were found in winter wheat fields in China. Research was carried out to investigate whether resistance to clodinafop-propargyl was present and the molecular mechanism of herbicide resistance in these two populations. Dose–response assays confirmed high level resistance to clodinafop-propargyl in both R2105 and R1027 populations, with resistance indexes 25.1 and 22.1. ACCase gene sequence comparison revealed three amino acid mutations (Trp-1999-Leu, Ile-2041-Asn, or Asp-2078-Gly) in R2105 population and Ile-2041-Asn mutation in R1027 population. Sensitivity to other herbicides assays indicated that R2105 and R1027 populations were cross resistant to fenoxaprop-P-ethyl and multiple resistant to pyroxsulam and mesosulfuron-methyl. The ALS gene sequence analysis revealed that all resistant individuals in R2105 and R1027 populations had the Trp-574-Leu mutation. Applying malathion, significantly decreased the rate of metabolism of clodinafop-propargyl in both R2105 and R1027 populations. This is the first report of multiple resistance to ACCase- and ALS-inhibiting herbicides conferred by target-site mutations and enhanced metabolism in black-grass in China.

Comparison between the mechanisms of Clearfield ® wheat and Lolium rigidum multiple resistant to acetyl CoA carboxylase and acetolactate synthase inhibitors

Clearfield® wheat (Triticum aestivum) have helped eliminate the toughest grasses and broadleaf weeds in Spain since 2005. This crop production system includes other tolerant cultivars to the application of imidazolinone (IMI) herbicides. However, the continuous use and off-label rates of IMI herbicides can contribute to the development of resistance in Lolium rigidum and other weed species. In this research, the main objectives were to study the resistance mechanisms to acetolactate synthase (ALS) and acetyl coenzyme A carboxylase (ACCase) inhibitors in a L. rigidum accession (LrR) from a Clearfield® wheat field, with a long history rotating these IMI-tolerant crops and compare them with those present in the IMI-tolerant wheat. The resistance to ACCase inhibitors in LrR was due to point mutations (Ile1781Leu plus Asp2078Gly) of the target site gene plus an enhanced herbicide metabolism (EHM), on the other hand, in wheat accessions was due only by EHM. Mechanisms involved in the resistance to ALS inhibitors were both point mutations of the target gene and EHM in the IMI-tolerant wheat, while only evidence of mutation (Trp574Leu) was found in the multiple herbicide resistant L. rigidum accession. This research demonstrates that if crop rotation is not accompanied by the use of alternative sites of action in herbicide-tolerant crops, resistant weeds to herbicide to which crops are tolerant, can easily be selected. Moreover, repeated and inappropriate use of Clearfield® crops and herbicide rotations can lead to the evolution of multiple resistant weeds, as shown in this study, and have also inestimable environmental impacts.

Dissecting weed adaptation: Fitness and trait correlations in herbicide-resistant Alopecurus myosuroides.

BACKGROUNDUnravelling the genetic architecture of non‐target‐site resistance (NTSR) traits in weed populations can inform questions about the inheritance, trade‐offs and fitness costs associated with these traits. Classical quantitative genetics approaches allow study of the genetic architecture of polygenic traits even where the genetic basis of adaptation remains unknown. These approaches have the potential to overcome some of the limitations of previous studies into the genetics and fitness of NTSR.RESULTSUsing a quantitative genetic analysis of 400 pedigreed Alopecurus myosuroides seed families from nine field‐collected populations, we found strong heritability for resistance to the acetolactate synthase and acetyl CoA carboxylase inhibitors (h 2 = 0.731 and 0.938, respectively), and evidence for shared additive genetic variance for resistance to these two different herbicide modes of action, r g = 0.34 (survival), 0.38 (biomass). We find no evidence for genetic correlations between life‐history traits and herbicide resistance, indicating that resistance to these two modes of action is not associated with large fitness costs in blackgrass. We do, however, demonstrate that phenotypic variation in plant flowering characteristics is heritable, h 2 = 0.213 (flower height), 0.529 (flower head number), 0.449 (time to flowering) and 0.372 (time to seed shed), demonstrating the potential for adaptation to other nonchemical management practices (e.g. mowing of flowering heads) now being adopted for blackgrass control.CONCLUSIONThese results highlight that quantitative genetics can provide important insight into the inheritance and genetic architecture of NTSR, and can be used alongside emerging molecular techniques to better understand the evolutionary and fitness landscape of herbicide resistance. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Pinoxaden resistance in Lolium perenne L. is due to both target-site and non-target-site mechanisms

Application of herbicides inhibiting acetyl CoA carboxylase (ACCase) has been one of the main strategies for selectively controlling grass weed species such as perennial ryegrass (Lolium perenne L.) in wheat and barley crops in New Zealand. In this study, we have confirmed and characterized resistance to pinoxaden, an ACCase-inhibiting herbicide, in a population of L. perenne. Dose-response experiments were conducted to assess the level of pinoxaden resistance, and based on the LD50 values, the studied population was 41.4-times more resistant to pinoxaden than a susceptible population. Application of malathion, an inhibitor of the cytochrome P450s, preceding pinoxaden treatment reduced the level of resistance to 9.7-fold. However, pre-treatment with the glutathione S-transferase (GST) inhibitor 4-chloro7- nitrobenzoxadiazole prior to pinoxaden treatment did not affect pinoxaden resistance. Partial sequencing of the ACCase gene revealed that the resistant population had an isoleucine to valine replacement at position 2041. These results suggest that both cytochrome P450-based and target-site mechanisms are jointly associated with this instance of pinoxaden resistance in L. perenne. The pinoxaden-resistant L. perenne individuals were also resistant to quizalofop-p-ethyl (108.6-fold), but they were susceptible to clethodim, which can, therefore, be used to manage this pinoxaden-resistant L. perenne. This is the first report of a L. perenne population in which a rare target-site mutation works in concert with enhanced cytochrome P-450 activity to confer pinoxaden resistance. Evolution of resistance to ACCase-inhibiting herbicides in this L. perenne population indicates that integrated weed management practices are required to prevent widespread resistance developing in New Zealand cereal crop systems.

Obesity promotes lipid accumulation in mouse cartilage-A potential role of acetyl-CoA carboxylase (ACC) mediated chondrocyte de novo lipogenesis.

Obesity promotes the development of osteoarthritis (OA). It is also well-established that obesity leads to excessive lipid deposition in nonadipose tissues, which often induces lipotoxicity. The objective of this study was to investigate changes in the levels of various lipids in mouse cartilage in the context of obesity and determine if chondrocyte de novo lipogenesis is altered.We used Oil Red O to determine the accumulation of lipid droplets in cartilage from mice fed high-fat diet (HFD) or low-fat diet (LFD). We further used mass spectrometry-based lipidomic analyses to quantify levels of different lipid species. Expression of genes involving in fatty acid (FA) uptake, synthesis, elongation, and desaturation were examined using quantitative polymerase chain reaction. To further study the potential mechanisms, we cultured primary mouse chondrocytes under high-glucose and high-insulin conditions to mimic the local microenvironment associated with obesity and subsequently examined the abundance of cellular lipid droplets. The acetyl-CoA carboxylase (ACC) inhibitor, ND-630, was added to the culture medium to examine the effect of inhibiting de novo lipogenesis on lipid accumulation in chondrocytes.When compared to the mice receiving LFD, the HFD group displayed more chondrocytes with visible intracellular lipid droplets. Significantly higher amounts of total FAs were also detected in the HFD group. Five out of six significantly upregulated FAs were ω-6 FAs, while the two significantly downregulated FAs were ω-3 FAs. Consequently, the HFD group displayed a significantly higher ω-6/ω-3 FA ratio. Ether linked phosphatidylcholine was also found to be higher in the HFD group. Fatty acid desaturase (Fad1-3), fatty acid-binding protein 4 (Fabp4), and fatty acid synthase (Fasn) transcripts were not found to be different between the treatment groups and fatty acid elongase (Elovl1-7) transcripts were undetectable in cartilage. Ceramide synthase 2 (Cers-2), the only transcript found to be changed in these studies, was significantly upregulated in the HFD group. In vitro, chondrocytes upregulated de novo lipogenesis when cultured under high-glucose, high-insulin conditions, and this observation was associated with the activation of ACC, which was attenuated by the addition of ND-630.This study provides the first evidence that lipid deposition is increased in cartilage with obesity and that this is associated with the upregulation of ACC-mediated de novo lipogenesis. This was supported by our observation that ACC inhibition ameliorated lipid accumulation in chondrocytes, thereby suggesting that ACC could potentially be targeted to treat obesity-associated OA.

Pharmacokinetics, Mass Balance, Metabolism, and Excretion of the Liver-Targeted Acetyl-CoA Carboxylase Inhibitor PF-05221304 (Clesacostat) in Humans

The disposition of the hepatoselective ACC inhibitor PF-05221304 (Clesacostat) was studied after a single 50-mg oral dose of [14C]-PF-05221304 to healthy human subjects. Mass balance was achieved with 89.9% of the administered dose recovered in urine and faeces, over the 11-day study period. The total administered radioactivity excreted in faeces and urine was 81.7% and 8.2%, respectively. Unchanged PF-05221304 accounted for 35.6% of the radioactive dose in faeces, suggesting ∼64% of the administered dose was absorbed. PF-05221304 was principally metabolized via oxidative and reductive pathways involving: (a) N-dealkylation, (b) isopropyl group monohydroxylation to yield enantiomeric metabolites (M2a and M2b), (c) hydroxylation on the 3-azaspiro[5.5]undecan-8-one moiety to metabolites M5 and 519c, and (d) carbonyl group reduction to enantiomeric alcohol metabolites M3, and M4. Secondary metabolites (521a, 521b, and 533), derived from a combination of oxidation and reduction of the primary metabolites accounted for ∼14.8% of the dose. In plasma, unchanged PF-05221304 represented 96.1% circulating radioactivity. Metabolites M1, M2b, and M2a represented 1.94, 1.76, and 0.18% of circulating radioactivity, respectively. Overall, these data suggest that PF-05221304 is well absorbed in humans and eliminated largely via phase I metabolism.

New avenues for NASH therapy by targeting ACC

Acetyl-CoA carboxylase (ACC) is one of the more promising therapeutic targets for non-alcoholic steatohepatitis (NASH), but current ACC inhibitors already tested in clinical trials also exert the unwanted adverse side effect of hypertriglyceridemia. In two recent studies by Calle etal. in Nature Medicine and Zhang etal. in Science Translational Medicine, new strategies for ACC targeting were explored for NASH therapy that successfully resolved the adverse effect of hyperlipidemia while maintaining potent anti-NASH efficacy. These findings bring encouraging new momentum to the clinical application of ACC inhibition for NASH therapy.

Thiobencarb resistance mechanism is distinct from CYP81A-based cross-resistance in late watergrass (Echinochloa phyllopogon)

AbstractThe genetic and molecular basis of resistance evolution in weeds to multiple herbicides remains unclear despite being a great threat to agriculture. A population of late watergrass [Echinochloa phyllopogon(Stapf.) Koso-Pol.] was reported to exhibit resistance to ≥15 herbicides from six sites of action, including thiobencarb (TB). While previous studies disclosed that the resistance to a majority of herbicides such as acetolactate synthase (ALS) and acetyl-CoA carboxylase inhibitors is caused by the overexpression of herbicide-metabolizing cytochrome P450s (CYP81A12andCYP81A21), the resistance mechanisms to some herbicides remain unknown. Here, we analyzed the resistance segregation in the progenies between resistant and sensitive populations and performed a transgenic plant sensitivity assay to resolve whether TB resistance is endowed by the sameCYP81A12/21-based cross-resistance mechanism or other unknown multiple-resistance mechanisms. In the F6progenies, resistance to the ALS inhibitor bensulfuron-methyl cosegregated with the resistances to many other herbicides under theCYP81A12/21-based cross-resistance mechanism; however, TB resistance segregated independently. Furthermore,CYP81A12/21failed to confer TB resistance in transgenicArabidopsis thalianaL. Heynh, thus confirming that TB resistance in resistantE. phyllopogonis not endowed by the two P450s that are responsible for the metabolism-based cross-resistance. This study provides evidence that resistance inE. phyllopogonto herbicides with multiple sites of action is endowed by both P450-based and other uncharacterized non–target site based mechanisms. Our findings add another layer in the understanding of resistance evolution to multiple herbicides inE. phyllopogon. Identification of the key genes endowing TB resistance will be the future direction of this research.

Structurally diverse glycosides of secoiridoid, bisiridoid, and triterpene-bisiridoid conjugates from the flower buds of two Caprifoliaceae plants and their ATP-citrate lyase inhibitory activities

The ethanolic extracts of the dried flower buds of two Caprifoliaceae plants, Lonicera japonica and Abelia × grandiflora, showed considerable inhibitory activities against adenosine triphosphate (ATP)-citrate lyase (ACL), a new promising drug target for the treatment of metabolic disorders. Bioassay-guided purification in conjunction with HPLC-PDA profiling led to the isolation and characterization of thirty-five (1–35) and fourteen (1′−14′) structurally diverse compounds from the above two plant extracts, respectively. Compounds 1–9 and 1′−6′ are previously undescribed glycosides. Their structures were elucidated on the basis of spectroscopic data, electronic circular dichroism (ECD), and single crystal X-ray diffraction analyses. In particular, lonicejaposide A (1) has an unprecedented skeleton generated through the coupling of C-7 in secologanin with C-2′′ in phenylacetaldehyde via an aldol condensation. Abeliflorosides A (1′) and B (2′) are hitherto unknown glycosides of triterpene and bisiridoid conjugates constructed through the formation of a 1,3-dioxane moiety. All the isolates were evaluated for their inhibitory activities against ACL. Compounds 9, 25–28, 31, 1′, 2′, and 14′ displayed significant inhibitory effects, with IC50 values ranging from 0.1 to 14.2 μM. The interactions of selected compounds possessing different structure features (e.g., 9, 25, 31, and 2′) with ACL were thereafter performed by employing molecular docking studies. In addition, compound 2′, the most complex triterpene-bisiridoid conjugate glycoside reported herein, also inhibited acetyl-CoA carboxylase 1 (ACC1), with an IC50 value of 7.9 μM. The dried material of the flower buds of L. japonica (honeysuckle) is a well-known traditional oriental medicine (i.e., Flos Lonicerae Japonicae, FLJ) and has long been used in large quantities. The above findings not only provide new insights for the development of multipurpose utilization of FLJ in healthcare community, but also provide profitable clues indicating that the flower buds of A. × grandiflora might be a potential alternative to FLJ in the traditional Chinese medicine market.

Total Synthesis of the Acetyl CoA Carboxylase Inhibitor Soraphen A: Asymmetric Tsuji Reduction Enables Successive Olefin Metathesis.

The total synthesis of soraphen A, a myxobacterial metabolite and inhibitor of acetyl CoA carboxylase, was completed in 11 steps (longest linear sequence), less than half the steps previously required. Seven metal-catalyzed processes were deployed to unlock step-economy (comprising five asymmetric processes and four C-C bond formations). The present route does not utilize chiral auxiliaries, and four of five C-C bond formations exploit non-premetalated partners. To maximize convergency, an asymmetric Tsuji reduction was developed using a Pd-AntPhos catalyst that allows a metathesis-inactive allylic carbonate to serve as a masked terminal olefin, thereby enabling successive olefin metathesis events.

Fenofibrate Mitigates Hypertriglyceridemia in Nonalcoholic Steatohepatitis Patients Treated With Cilofexor/Firsocostat

Patients with advanced fibrosis due to nonalcoholic steatohepatitis (NASH) are at high risk of morbidity and mortality. We previously found that a combination of the farnesoid X receptor agonist cilofexor (CILO) and the acetyl-CoA carboxylase inhibitor firsocostat (FIR) improved liver histology and biomarkers in NASH with advanced fibrosis but was associated with hypertriglyceridemia. We evaluated the safety and efficacy of icosapent ethyl (Vascepa) and fenofibrate to mitigate triglyceride elevations in patients with NASH treated with CILO and FIR. Patients with NASH with elevated triglycerides (≥150 and <500 mg/dL) were randomized to Vascepa 2 g twice daily (n= 33) or fenofibrate 145 mg daily (n= 33) for 2 weeks, followed by the addition of CILO 30 mg and FIR 20 mg daily for 6 weeks. Safety, lipids, and liver biochemistry were monitored. All treatments were well-tolerated; most treatment-emergent adverse events were Grade 1 to 2 severity, and there were no discontinuations due to adverse events. At baseline, median (interquartile range [IQR]) triglycerides were similar in the Vascepa and fenofibrate groups (median, 177 [IQR, 154-205] vs 190 [IQR, 144-258] mg/dL, respectively). Median changes from baseline in triglycerides for Vascepa vs fenofibrate after 2 weeks of pretreatment were-12 mg/dL (IQR,-33 to 7 mg/dL; P= .09) vs-32 mg/dL (IQR,-76 to 6 mg/dL; P= .012) and at 6 weeks were+41 mg/dL (IQR, 16-103 mg/dL; P < .001) vs-2 mg/dL (IQR,-42 to 54 mg/dL; P= .92). In patients with baseline triglycerides <250 mg/dL, fenofibrate was more effective vs Vascepa in mitigating triglyceride increases after 6 weeks of combination treatment (+6 vs+39 mg/dL); similar trends were observed in patients with baseline triglycerides ≥250 mg/d (-61 vs+99 mg/dL). In patients with NASH with hypertriglyceridemia treated with CILO and FIR, fenofibrate was safe and effectively mitigated increases in triglycerides associated with acetyl-CoA carboxylase inhibition. gov, Number: NCT02781584.

Studying the inhibition mechanism of human acetyl-CoA carboxylase by aromatic ligands in the treatment of fatty acid metabolic syndrome using computational chemistry methods

Background: Metabolic syndrome is a consequence of excess fatty acid accumulation in the human body, causing metabolic disorders related to the role of the two acetyl-CoA carboxylase (ACC) isoforms, ACC1 and ACC2. Inheriting the results from our previous research about "the effective model of human acetyl-CoA carboxylase inhibition by aromatic-structure inhibitors", this research explains the inhibition mechanism of the enzyme by studying the interaction direction, preferential binding sites of aromatic ligands on the isoforms and conformational changes of enzyme-ligand complexes in the body’s environmental conditions (temperature 37 degrees Celsius and water solvent). Methods: This research applies hard docking combined with molecular dynamics and virtual screening of 14 inhibitory ligand groups, which have 50% enzyme inhibitor (IC50) values determined by experiments, including bipiperidylcarboxamide derivatives, polyketides, aryloxyphenoxypropionate, cyclohexanedione, synthetic acyl-CoA fatty acid chains, acylsulfonamide derivatives, Taisho inhibitors, Sanofi-Aventis inhibitors, AstraZeneca inhibitors, Takeda inhibitors, Pfizer inhibitors, Nimbus inhibitors, Amgen inhibitors and Boehringer Ingel-heim inhibitors. The geometric structures of the ligands are modeled and optimized by quantum mechanical methods, including PM3 for ACC1 and DFT at the B3LYP/6-31G(d,p) level for ACC2 based on our previous research about the QSAR enzyme-inhibition models, using Gaussian 09 W software. Results: Binding energy values at the identified preferential binding sites of aromatic ligands are closely correlated with the experimental log(IC50), which has statistical significance (p &lt; 0.05): R2ACC1 = 96.3% and R2ACC2 = 78.9%. Those sites reveal the five reaction niches on ACC1, involving the two preferential binding regions in the order of amino acids 152 to 643 and 1632 to 2100, and the four reaction niches on ACC2, involving the two preferential binding regions in the order of amino acids 270 to 704 and 1831 to 2299. The molecular dynamic simulation of enzyme-ligand complexes in the body’s environmental conditions confirms that if ligands have fewer stereogenic interactions and are ready to form four types of bonds, including hydrogen, pi-alkyl, pi-cation and alkyl-alkyl bonds, their inhibitory activity will be higher. Conclusion: The research clarifies the inhibition mechanism of the enzyme in which ligands prefer to interact with amino acids 152 to 643 and 270 to 704 on the biotin carboxylase domain of ACC and amino acids 1632 to 2100 and 1831 to 2299 on the carboxyl transferase domain of ACC. In addition, the fewer stereogenic interactions of ligands and the easier it is to form four types of bonds, including hydrogen, pi-alkyl, pi-cation and alkyl-alkyl bonds, the higher their inhibitory activity.

First Detection and Characterization of Cross- and Multiple Resistance to Acetyl-CoA Carboxylase (ACCase)- and Acetolactate Synthase (ALS)-Inhibiting Herbicides in Black-Grass (Alopecurus myosuroides) and Italian Ryegrass (Lolium multiflorum) Populations from Ireland

Understanding the resistance spectrum and underlying genetic mechanisms is critical for managing herbicide-resistant populations. In this study, resistance to acetyl CoA carboxylase (ACCase) and acetolactate synthase (ALS) inhibitors was investigated in four suspected resistant populations of Alopecurus myosuroides (ALOMY-001 to ALOMY-004) and Lolium multiflorum (LOLMU-001 to LOLMU-004), collected from cereal production fields in Ireland. Glasshouse assays with three ALOMY-active herbicides [propaquizafop, cycloxydim (ACCase) and mesosulfuron + iodosulfuron (ALS)] or five LOLMU-active herbicides [pinoxaden, propaquizafop, cycloxydim (ACCase) and mesosulfuron + iodosulfuron, pyroxsulam (ALS)], and target-site resistance mechanism studies, based on pyrosequencing, were carried out in each of those populations. For A. myosuroides, Ile-1781-Leu ACCase mutation contributed to propaquizafop and cycloxydim resistance (shoot dry weight GR50 resistance factor (RF) = 7.5–35.5) in all ALOMY populations, and the independent Pro-197-Thr or Pro-197-Ser ALS mutation contributed to mesosulfuron + iodosulfuron resistance (RF = 3.6–6.6), in ALOMY-002 to ALOMY-004. Most of the analyzed plants for these mutations were homo/heterozygous combinations or only heterozygous. For L. multiflorum, phenotypic resistance to mesosulfuron + iodosulfuron (RF = 11.9–14.6) and pyroxsulam (RF = 2.3–3.1) was noted in all LOLMU populations, but the Pro-197-Gln or Pro-197-Leu ALS mutation (mostly in homozygous status) was identified in LOLMU-001, LOLMU-002 and LOLMU-004 only. Additionally, despite no known ACCase mutations in any LOLMU populations, LOLMU-002 survived pinoxaden and propaquizafop application (RF = 3.4 or 1.3), and LOLMU-003 survived pinoxaden (RF = 2.3), suggesting the possibility of non-target-site resistance mechanisms for ACCase and/or ALS resistance in these populations. Different resistance levels, as evidenced by a reduction in growth as dose increased above field rates in ALOMY and LOLMU, were due to variations in mutation rate and the level of heterozygosity, resulting in an overall resistance rating of low to moderate. This is the first study confirming cross- and multiple resistance to ACCase- and ALS-inhibiting herbicides, highlighting that resistance monitoring in A. myosuroides and L. multiflorum in Ireland is critical, and the adoption of integrated weed management strategies (chemical and non-chemical/cultural strategies) is essential.

Acetyl-CoA Carboxylase Inhibitor CP640.186 Increases Tubulin Acetylation and Impairs Thrombin-Induced Platelet Aggregation.

Acetyl-CoA carboxylase (ACC) is the first enzyme regulating de novo lipid synthesis via the carboxylation of acetyl-CoA into malonyl-CoA. The inhibition of its activity decreases lipogenesis and, in parallel, increases the acetyl-CoA content, which serves as a substrate for protein acetylation. Several findings support a role for acetylation signaling in coordinating signaling systems that drive platelet cytoskeletal changes and aggregation. Therefore, we investigated the impact of ACC inhibition on tubulin acetylation and platelet functions. Human platelets were incubated 2 h with CP640.186, a pharmacological ACC inhibitor, prior to thrombin stimulation. We have herein demonstrated that CP640.186 treatment does not affect overall platelet lipid content, yet it is associated with increased tubulin acetylation levels, both at the basal state and after thrombin stimulation. This resulted in impaired platelet aggregation. Similar results were obtained using human platelets that were pretreated with tubacin, an inhibitor of tubulin deacetylase HDAC6. In addition, both ACC and HDAC6 inhibitions block key platelet cytoskeleton signaling events, including Rac1 GTPase activation and the phosphorylation of its downstream effector, p21-activated kinase 2 (PAK2). However, neither CP640.186 nor tubacin affects thrombin-induced actin cytoskeleton remodeling, while ACC inhibition results in decreased thrombin-induced reactive oxygen species (ROS) production and extracellular signal-regulated kinase (ERK) phosphorylation. We conclude that when using washed human platelets, ACC inhibition limits tubulin deacetylation upon thrombin stimulation, which in turn impairs platelet aggregation. The mechanism involves a downregulation of the Rac1/PAK2 pathway, being independent of actin cytoskeleton.

Impact of a Novel W2027L Mutation and Non-Target Site Resistance on Acetyl-CoA Carboxylase-Inhibiting Herbicides in a French Lolium multiflorum Population.

Herbicides that inhibit acetyl-CoA carboxylase (ACCase) are among the few remaining options for the post-emergence control of Lolium species in small grain cereal crops. Here, we determined the mechanism of resistance to ACCase herbicides in a Lolium multiflorum population (HGR) from France. A combined biological and molecular approach detected a novel W2027L ACCase mutation that affects aryloxyphenoxypropionate (FOP) but not cyclohexanedione (DIM) or phenylpyraxoline (DEN) subclasses of ACCase herbicides. Both the wild-type tryptophan and mutant leucine 2027-ACCase alleles could be positively detected in a single DNA-based-derived polymorphic amplified cleaved sequence (dPACS) assay that contained the targeted PCR product and a cocktail of two discriminating restriction enzymes. Additionally, we identified three well-characterised I1781L, I2041T, and D2078G ACCase target site resistance mutations as well as non-target site resistance in HGR. The non-target site component endowed high levels of resistance to FOP herbicides whilst partially impacting on the efficacy of pinoxaden and cycloxydim. This study adequately assessed the contribution of the W2027L mutation and non-target site mechanism in conferring resistance to ACCase herbicides in HGR. It also highlights the versatility and robustness of the dPACS method to simultaneously identify different resistance-causing alleles at a single ACCase codon.