stearoyl-CoA Desaturase Inhibitors Research Articles

Stearoyl-CoA desaturase activity in bovine cumulus cells protects the oocyte against saturated fatty acid stress.

Metabolic rich and poor conditions are both characterized by elevated free fatty acid levels and have been associated with impaired female fertility. In particular, saturated free fatty acids have a dose-dependent negative impact on oocyte developmental competence, while monounsaturated free fatty acids appear less harmful. Cumulus cells seem to protect the oocyte against free fatty acids, and the aim of this study was to determine the mechanism behind this protection In particular, the role of the enzyme stearoyl-CoA desaturase (SCD) that converts saturated into monounsaturated fatty acids was investigated. SCD gene and protein were abundantly expressed in cumulus cells, but expression was low in oocytes. The level of SCD protein expression in cumulus cells did not change when COCs were exposed to saturated stearic acid during maturation. SCD inhibition in the presence of stearic acid significantly reduced the developmental competence of oocytes and increased the incidence of apoptosis in cumulus cells. The esterified oleic/stearic acid ratio of the neutral lipid fraction in cumulus cells decreased in the presence of SCD inhibitors when COCs were exposed to saturated free fatty acids during maturation, indicating the SCD-specific conversion of saturated fatty acids under noninhibiting conditions. The observation that cumulus cells can desaturate the potentially toxic stearic acid into oleic acid via SCD activity provides a mechanistic insight into how the cumulus cells protect the oocyte against toxicity by saturated fatty acid.

Inhibitory effects of fenretinide metabolites N-[4-methoxyphenyl]retinamide (MPR) and 4-oxo-N-(4-hydroxyphenyl)retinamide (3-keto-HPR) on fenretinide molecular targets β-carotene oxygenase 1, stearoyl-CoA desaturase 1 and dihydroceramide Δ4-desaturase 1.

The therapeutic capacity of fenretinide (N-[4-hydroxyphenyl] retinamide; 4-HPR) has been demonstrated for several conditions, including cancer, obesity, diabetes, and ocular disease. Yet, the mechanisms of action for its pleiotropic effects are still undefined. We hypothesized that investigation of two of the major physiological metabolites of fenretinide, N-[4-methoxyphenyl]retinamide (MPR) and 4-oxo-N-(4-hydroxyphenyl)retinamide (3-keto-HPR), might begin to resolve the multifaceted effects of this synthetic retinoid. We analyzed the effects of fenretinide, MPR, 3-keto-HPR, and the non-retinoid RBP4 ligand A1120, on the activity of known targets of fenretinide, stearoyl-CoA desaturase 1 (SCD1) and dihydroceramide Δ4-desaturase 1 (DES1) in ARPE-19 cells, and purified recombinant mouse beta-carotene oxygenase 1 (BCO1) in vitro. Lipids and retinoids were extracted and quantified by liquid chromatography-mass spectrometry and reversed phase HPLC, respectively. The data demonstrate that while fenretinide is an inhibitor of the activities of these three enzymes, that 3-keto-HPR is a more potent inhibitor of all three enzymes, potentially mediating most of the in vivo beneficial effects of fenretinide. However, while MPR does not affect SCD1 and DES1 activity, it is a potent specific inhibitor of BCO1. We conclude that a deeper understanding of the mechanisms of action of fenretinide and its metabolites provides new avenues for therapeutic specificity. For example, administration of 3-keto-HPR instead of fenretinide may be preferential if inhibition of SCD1 or DES1 activity is the goal (cancer), while MPR may be better for BCO1 modulation (carotenoid metabolism). Continued investigation of fenretinide metabolites in the context of fenretinide’s various therapeutic uses will begin to resolve the pleotropic nature of this compound.

In vitro and in vivo antitumor activities of T-3764518, a novel and orally available small molecule stearoyl-CoA desaturase 1 inhibitor

Most cancer cells are characterized by elevated lipid biosynthesis. The rapid proliferation of cancer cells requires de novo synthesis of fatty acids. Stearoyl-CoA desaturase-1 (SCD1), a key enzyme for lipogenesis, is overexpressed in various types of cancer and plays an important role in cancer cell proliferation. Therefore, it has been studied as a candidate target for cancer therapy. In this study, we demonstrate the pharmacological properties of T-3764518, a novel and orally available small molecule inhibitor of SCD1. T-3764518 inhibited stearoyl-CoA desaturase-catalyzed conversion of stearoyl-CoA to oleoyl-CoA in colorectal cancer HCT-116 cells and their growth. Further, it slowed tumor growth in an HCT-116 and a mesothelioma MSTO-211H mouse xenograft model. Comprehensive lipidomic analyses revealed that T-3764518 increases the membrane ratio of saturated: unsaturated fatty acids in various lipid species such as phosphatidylcholines and diacylglycerols in both cultured cells and HCT-116 xenografts. Treatment-associated lipidomic changes were followed by activated endoplasmic reticulum (ER) stress responses such as increased immunoglobulin heavy chain-binding protein expression in HCT-116 cells. These T-3764518-induced changes led to an increase in cleaved poly (ADP-ribose) polymerase 1 (PARP1), a marker of apoptosis. Additionally, bovine serum albumin conjugated with oleic acid, an SCD1 product, prevented cell growth inhibition and ER stress responses by T-3764518, indicating that these outcomes were not attributable to off-target effects. These results indicate that T-3764518 is a promising new anticancer drug candidate.

Exogenous lipids promote the growth of breast cancer cells via CD36.

Cancer cells present sustained de novo fatty acid (FA) synthesis with increased production of saturated fatty acids (SFAs) and monounsaturated fatty acids (MUFAs). This change in FA metabolism is associated with overexpression of stearoyl-CoA desaturase 1 (SCD1), which catalyses the transformation of SFAs into MUFAs (e.g., oleic acid). In this study, we provide new evidence that SCD1 inhibition leads to the anti-proliferation effect of breast cancer cells through induction of apoptosis, cell cycle arrest and migration prevention. However, the antitumor effect of the SCD1 inhibitor can be reversed by exogenous oleic acid. We hypothesize that, in addition to de novo synthesis, cancer cells may uptake exogenous FAs actively. CD36, also known as FA translocase (FAT), that functions as a transmembrane protein and mediates the uptake of FAs, is observed to be highly expressed in breast cancer tissues. Furthermore, the anti-proliferation effect caused by the SCD1 inhibitor can not be reversed by exogenous oleic acid supplementation in CD36 knockdown breast cancer cells. Our study revealed that the lipid metabolism of breast cancer is regulated not only by de novo lipogenesis but also by the availability of lipids outside cancer cells. Consistent with FA synthesis, FA uptake and transport will be another important target pathway for anticancer therapy, and the FA channel protein CD36 may provide a promising therapeutic target. Lipogenesis combined with FA transport will be a new orientation for antitumor therapy.

Role of Stearoyl-CoA Desaturase 1 and 5 in breast cancer cell migration and survival

We previously reported that a major component of breast tumor stroma, the “cancer-associated fibroblasts” (CAFs), induced epithelial-mesenchymal transition and an increase in cell membrane fluidity as well as in migration speed and directness in poorly (MCF-7) and highly invasive (MDA-MB-231) breast cancer cells. We next investigated the mechanisms responsible for the CAF-promoted tumor cell migration demonstrating the crucial role of Stearoyl-CoA desaturase 1 (SCD1), one of the main enzyme regulating membrane fluidity. We found SCD1 to be upregulated in tumor cells co-cultured with CAFs and that its inhibition (pharmacological or siRNA-based) impaired both intrinsic and CAF-driven tumor cell migration. In the present study, we deepen the understanding of the mechanisms involved in the SCD1-based modulation of tumor cell migration, as well as the possible role of the other human SCD isoform, SCD5. Thus, in the two above mentioned cell lines we studied whether the inhibitory effect produced on cell migration by SCD1 depletion was due to the deficiency of oleic acid (OA), the main SCD1 enzymatic product. By a wound healing assay, we found that the addition of OA nullified the inhibitory effects produced on tumor cell migration by the SCD1 inhibition in both the cell lines while SCD5 appeared not to be involved in the regulation of their motility but it was upregulated in MCF-7 cells co-cultured with CAFs. Because of the high number of detached MCF-7 cells silenced for SCD5, we investigated the role of the desaturase on tumor cell survival and an induction of necrosis was found. Consistently with the promotion of tumor cell migration, CAFs have also been found to induce the activated form of the hepatocyte growth factor receptor, p-MET, in the two cell lines. These results provide further insights in understanding the role of SCD1 in both intrinsic and CAF-stimulated mammary tumor cell migration. Moreover, our data seem to suggest the ability of CAFs to promote the maintenance of tumor cell survival by the induction of SCD5 levels.

Loss of Stearoyl-CoA Desaturase-1 Activity Induced Leptin Resistance in Neuronal Cells.

The lack of response to leptin's actions in the brain, "leptin resistance," is one of the main causes of the pathogenesis of obesity. However, although high-fat diets affect sensitivity to leptin, the underlying mechanisms of leptin resistance are still an enigma. Here we examined the effect of excess saturated fatty acids (SFAs) on leptin signaling in human neuronal cells. Palmitate, the principle source of SFAs in diet, induced leptin resistance in a human neuroblastoma cell line stably transfected with the Ob-Rb leptin receptor (SH-SY5Y-ObRb). We next investigated the function of stearoyl-CoA desaturase-1 (SCD1), an enzyme which converts SFAs into monounsaturated fatty acids (MUFAs), on leptin-induced signaling. We found that reduction of SCD1 activity, through SCD1 inhibition and knockdown, impairs leptin-induced signal transducer and activator of transcription 3 (STAT3) phosphorylation in human neuronal cells. Our findings suggested that SCD1 plays a key role in the pathophysiology of leptin resistance in neuronal cells associated with obesity.

Stearoyl-CoA desaturase 1 deficiency reduces lipid accumulation in the heart by activating lipolysis independently of peroxisome proliferator-activated receptor α

Stearoyl-CoA desaturase 1 (SCD1) has recently been shown to be a critical control point in the regulation of cardiac metabolism and function. Peroxisome proliferator-activated receptor α (PPARα) is an important regulator of myocardial fatty acid uptake and utilization. The present study used SCD1 and PPARα double knockout (SCD1−/−/PPARα−/−) mice to test the hypothesis that PPARα is involved in metabolic changes in the heart that are caused by SCD1 downregulation/inhibition. SCD1 deficiency decreased the intracellular content of free fatty acids, triglycerides, and ceramide in the heart of SCD1−/− and SCD1−/−/PPARα−/− mice. SCD1 ablation in PPARα−/− mice decreased diacylglycerol content in cardiomyocytes. These results indicate that the reduction of fat accumulation in the heart associated with SCD1 deficiency occurs independently of the PPARα pathway. To elucidate the mechanism of the observed changes, we treated HL-1 cardiomyocytes with the SCD1 inhibitor A939572 and/or PPARα inhibitor GW6471. SCD1 inhibition decreased the level of lipogenic proteins and increased lipolysis, reflected by a decrease in the content of adipose triglyceride lipase inhibitor G0S2 and a decrease in the ratio of phosphorylated hormone-sensitive lipase (HSL) at Ser565 to HSL (pHSL[Ser565]/HSL). PPARα inhibition alone did not affect the aforementioned protein levels. Finally, PPARα inhibition decreased the phosphorylation level of 5′-adenosine monophosphate-activated protein kinase, indicating lower mitochondrial fatty acid oxidation. In summary, SCD1 ablation/inhibition decreased cardiac lipid content independently of the action of PPARα by reducing lipogenesis and activating lipolysis. The present data suggest that SCD1 is an important component in maintaining proper cardiac lipid metabolism.

SCD1 deficiency protects mice against ethanol-induced liver injury

Stearoyl-CoA desaturase 1 (SCD1) is a delta-9 fatty acid desaturase that catalyzes the synthesis of mono-unsaturated fatty acids (MUFA). SCD1 is a critical control point regulating hepatic lipid synthesis and β-oxidation. Scd1 KO mice are resistant to the development of diet-induced non-alcoholic fatty liver disease (NAFLD). Using a chronic-binge protocol of ethanol-mediated liver injury, we aimed to determine if these KO mice are also resistant to the development of alcoholic fatty liver disease (AFLD).Mice fed a low-fat diet (especially low in MUFA) containing 5% ethanol for 10days, followed by a single ethanol (5g/kg) gavage, developed severe liver injury manifesting as hepatic steatosis. This was associated with an increase in de novo lipogenesis and inflammation. Using this model, we show that Scd1 KO mice are resistant to the development of AFLD. Scd1 KO mice do not show accumulation of hepatic triglycerides, activation of de novo lipogenesis nor elevation of cytokines or other pro-inflammatory markers. Incubating HepG2 cells with a SCD1 inhibitor induced a similar resistance to the effect of ethanol, confirming a role for SCD1 activity in mediating ethanol-induced hepatic injury.Taken together, our study shows that SCD1 is a key player in the development of AFLD and associated deleterious effects, and suggests SCD1 inhibition as a therapeutic option for the treatment of this hepatic disease.

Effect of hypoxia-inducible factor-2α/stearoyl-CoA desaturase-1 pathway on biological behaviors of hepatoma cells induced by hypoxia

Objective: To investigate whether hypoxia-inducible factor-2α (HIF-2α) affects the biological behaviors of hepatoma cells through regulating stearoyl-CoA desaturase-1 (SCD1) in a hypoxic environment. Methods: HepG2 and SMMC-7721 cells were exposed to 1% O2 to establish the hypoxic models. After hepatoma cells were stimulated by hypoxia for 0, 3, 6, 12, and 24 hours, fluorescent quantitative PCR and Western blot were used to measure the mRNA and protein expression of HIF-2α and SCD1 over time. HIF-2α interfering plasmids and SCD1 inhibitor CAY10566 were used to divide the cells into blank group (nomoxia), hypoxic group (1% O2 for 12 h), hypoxic negative control group (negative HIF-2α plasmid+1% O2 for 12 h), hypoxic interference group (HIF-2α interfering plasmid+1% O2 for 12 h), hypoxic CAY group (CAY10566 10 μmol+1% O2 for 12 h), and hypoxic interference+CAY group (HIF-2α interfering plasmid+CAY10566 10 μmol+1% O2 for 12 h). Western blot was used to measure the protein expression of HIF-2α and SCD1 in hepatoma cells, CCK8 assay was used to measure the proliferative capacity of hepatoma cells, Annexin-V/PE flow cytometry was used to measure the apoptosis of hepatoma cells, and transwell invasion assay was used to measure the invasion of hepatoma cells. A one-way analysis of variance was used to compare the means of multiple samples. Results: Both HepG2 and SMMC-7721 cells showed increasing mRNA and protein expression of HIF-2α and SCD1 over the time of hypoxic induction. After the expression of HIF-2α was downregulated in a hypoxic environment, hepatoma cells showed a significant reduction in the protein expression of SCD1; inhibition of SCD1 expression had no significant effect on the protein expression of HIF-2α in hepatoma cells. After HIF-2α was interfered with and SCD1 expression was inhibited, HepG2 and SMMC-7721 cells showed significantly greater reduction in the protein expression of SCD1 than those with HIF-2α or SCD1 inhibition alone (0.53±0.04 vs 1.12±0.04 or 1.12±0.04; 0.44±0.10 vs 0.90±0.10 or 0.99±0.13) (HIF-2α: FhepG2 = 1026.89, PhepG2 = 0.00, FSMMC-7721 = 2186.22, PSMMC-7721 = 0.00; SCD1: FhepG2 = 1347.93, PhepG2 = 0.00, FSMMC-7721 = 46.43, PSMMC-7721 = 0.00). Inhibition of the expression of HIF-2α or SCD1 reduced the proliferation and invasion of HepG2 and SMMC-7721 cells and promoted apoptosis (P < 0.05); interference and downregulation of HIF-2α combined with inhibition of SCD1 expression by CAY10566 achieved significantly greater reductions in proliferation and invasion and a significantly greater increase in apoptosis rate of hepatoma cells, compared with inhibition of HIF-2α or SCD1 alone (P <0.05). Conclusion: HIF-2α/SCD1 pathway may be one of the important mechanisms for hypoxia to regulate the energy metabolism of hepatoma cells and affect their biological behaviors.

Abstract 1003: Targeting liver-tumor initiating cells via hampering the lipogenesis pathways through stearoyl - CoA desaturase

Abstract Hepatocellular carcinoma is one of the most lethal cancers in the world. Most of the patients are diagnosed at advanced stage, and they receive sorafenib as their treatment modality. However, sorafenib can only extend patients’ survival to a median of 3-4 months. Hence, there is an urgent need to identify a new therapeutic target for treatment of this disease. Increasing evidence showed that tumor-initiating cells (T-ICs) are intrinsically resistant to conventional treatments. Identification of the signalling pathways that maintain the functions of T-ICs provides potential targets for treatment of HCC. For this purpose, we cultured serial passages of hepatospheres combined with chemotherapeutic regimens as a strategy to enrich liver T-IC population. Using cDNA microarray, we found upregulation of several key enzymes in lipogenesis in enriched T-IC population, among which Stearoyl CoA desaturase-1 (SCD1), the enzyme involved in the conversion of saturated into monounsaturated fatty acids, was the most significant. In HCC clinical samples, 62% (36/58) was found to be overexpressed (&amp;gt; 2-fold) when compared with non-tumor counterparts and patients with SCD1 overexpression had shorter disease free survival. Using overexpression and knockdown approaches, SCD1 was found to regulate the traits of T-ICs, including tumorigenicity, self-renewal, drug resistance and expression of liver T-IC markers. SCD1 was found critical in survival of T-ICs, as enriched T-IC population are more sensitive to the inhibition of SCD1 compared to adherent lineages. Interestingly, SCD1 was found to be upregulated in sorafenib-resistant HCC cells. Pharmacological inhibition of SCD1 by A939572 not only suppressed self-renewal ability but also consistently enhanced the sensitivity towards sorafenib. Using a patient-derived xenograft model, we found that a new inhibitor against SCD1 demonstrated significant growth suppressive effect. By comparing the genetic profiles between SCD1 knockdown cells and control cells by RNA sequencing analysis, we found that upregulation of ER stress signature genes may be the potential downstream targets of SCD1 to maintain the properties of T-ICs. In conclusion, we demonstrated the crucial role of SCD1 in maintenance of liver T-ICs, and targeting SCD1 in combination with sorafenib might be a novel therapeutic regimen against HCC. Citation Format: Kin Fai Ma, Jessica Lo, Eunice Yuen-Ting Lau, John A. Copland, Irene Oi-Lin Ng, Terence Kin-Wah Lee. Targeting liver-tumor initiating cells via hampering the lipogenesis pathways through stearoyl - CoA desaturase. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1003.

Discovery of tumor-specific irreversible inhibitors of stearoyl CoA desaturase.

A hallmark of targeted cancer therapies is selective toxicity among cancer cell lines. We evaluated results from a viability screen of over 200,000 small molecules to identify two chemical series, oxalamides and benzothiazoles, that were selectively toxic to the same four of 12 human lung cancer cell lines at low nanomolar concentrations. Sensitive cell lines expressed cytochrome P450 (CYP) 4F11, which metabolized the compounds into irreversible stearoyl CoA desaturase (SCD) inhibitors. SCD is recognized as a promising biological target in cancer and metabolic disease. However, SCD is essential to sebocytes, and accordingly SCD inhibitors cause skin toxicity. Mouse sebocytes were unable to activate the benzothiazoles or oxalamides into SCD inhibitors, providing a therapeutic window for inhibiting SCD in vivo. We thus offer a strategy to target SCD in cancer by taking advantage of high CYP expression in a subset of tumors.

Stearoyl-CoA desaturase-1 mediated cell apoptosis in colorectal cancer by promoting ceramide synthesis.

Inhibition of stearoyl-CoA desaturase 1 (SCD1) has been found to effectively suppress tumor cell proliferation and induce apoptosis in numerous neoplastic lesions. However, mechanism underlying SCD1-mediated anti-tumor effect has maintained unclear. Herein, we reported endo-lipid messenger ceramides played a critical role in tumor fate modulated by SCD1 inhibition. In vitro study in colorectal cancer cells demonstrated inhibition of SCD1 activity promoted apoptosis attributed to mitochondria dysfunctions, upregulation of reaction oxygen species (ROS), alteration of mitochondrial transmembrane potential and translocation of mitochondrial protein cytochrome C. While these effects were mediated by intracellular ceramide signals through induction of ceramide biosynthesis, rather than exclusive SFA accumulation. In vivo study in xenograft colorectal cancer mice showed pharmacologic administration of SCD1 inhibitor A939 significantly delayed tumor growth, which was reversed by L-cycloserine, an inhibitor of ceramide biosynthesis. These results depicted the cross-talk of SCD1-mediated lipid pathway and endo-ceramide biosynthesis pathway, indicating roles of ceramide signals in SCD1-mediated anti-tumor property.

Design, radiosynthesis, and evaluation of radiotracers for positron emission tomography imaging of stearoyl-CoA desaturase-1.

Design, radiosynthesis, and biological evaluation of two radiotracers (N-(3-[(18)F]fluoropropyl)-6-(4-(trifluoromethyl)benzoyl)-piperazin-1-yl)pyridazine-3-carboxamide ((18)F-FPPPT) and (N-(4-[(18)F]fluoroaniline)-6-(4-(trifluoromethyl)benzoyl)-piperazin-1-yl)pyridazine-3-carboxamide ((18)F-FAPPT)) are described for noninvasive assessment of stearoyl-CoA desaturase-1 (SCD-1). The overexpression of SCD-1 in multiple solid tumors associates with poor survival in cancer patients. The two radiotracers, (18)F-FPPPT and (18)F-FAPPT, were each prepared in three steps in radiochemical yields of 21% and 3%, respectively. The practicality of imaging SCD-1 with (18)F-FPPPT was tested in two mouse models bearing xenograft tumors with different levels of SCD-1 expression, which afforded a 1.8-fold uptake difference correspondingly. Our work indicates that it is possible to develop SCD-1 specific imaging probes from previously reported SCD-1 inhibitors.

Hepatocyte differentiation of human induced pluripotent stem cells is modulated by stearoyl-CoA desaturase 1 activity.

Stearoyl-CoA desaturase 1 (SCD1) plays important roles in organ development, glucose tolerance, insulin sensitivity, and cancer. Here, we examined the role of SCD1 for the differentiation of human induced pluripotent stem (hiPS) cells to liver cells by using drug inhibition and biochemical experiments. hiPS cells cultured in a pro-hepatic medium were exposed to an SCD1 inhibitor at various stages throughout differentiation. Liver-specific markers, specifically α-fetoprotein, albumin and urea in conditioned medium, and hepatocyte nuclear factor 4α (HNF4α) and cytochrome P450 7A1 (CYP7A1) gene expressions and triglyceride in cellular extracts were analyzed at various development stages. Measures of hepatocyte-specific function and triglyceride accumulation in later stages were strongly inhibited a minimum of -29% (P < 0.05) by SCD1 inhibitor in the early stage of hepatic differentiation and effectively reversed (>30%, P < 0.01) by the addition of oleate. The results were also reproducible with human primary mononuclear cells (hPMN). SCD1 inhibitor had no significant effect on liver-specific markers when it was added in the hepatic maturation stage. However, it strikingly led to higher albumin (1.6-fold, P = 0.03) and urea (1.9-fold, P = 0.02) production, and HNF4α (1.9-fold, P = 0.02) and CYP7A1 (1.3-fold, P = 0.03) expression upon incubation during the lineage-commitment stage. Hepatic differentiation from cultured hiPS cells is sensitive to SCD1 inhibition and this sensitivity is affected by the stage of cellular differentiation. Notably, findings also indicate that this notion can be extended to hPMN. The requirement for SCD1 activity in functional differentiation of hepatocytes may have relevance for human liver disease and metabolic dysregulation.

Elucidating the roles of stearoyl-CoA desaturase 1 in adipocyte fatty acid metabolism and cellular function

Worldwide obesity rates have risen to epidemic proportions, with over 600 million obese individuals across the globe. These individuals are prone to obesity-related health complications, including type 2 diabetes, hypertension, cancer, and cardiovascular disease. Obesity also coincides with the expansion of adipose tissue, which has an important role storing excess calories in the form of triacylglycerol (TAG) within adipocyte lipid droplets. The predominant fatty acids (FAs), comprising adipocyte TAGs, are monounsaturated FAs, which are produced by stearoyl-CoA desaturase 1 (SCD1). Specifically, SCD1 converts saturated FAs palmitate (PA) and stearate (SA) into palmitoleate and oleate, respectively. Interestingly, whole-body SCD1-deficiency is associated with reduced lipogenesis and adiposity. This places SCD1 as a potential target for obesity therapies; however, our understanding of the mechanisms linking SCD1 with changes in adipocyte function remains unclear. This thesis provides important new insights into how SCD1 impacts adipocyte FA metabolism and cellular function. Using a specific SCD1 inhibitor, changes in lipid metabolism, global gene expression, inflammation, cellular stress, and basal insulin signaling were assessed in 3T3-L1 adipocytes. Results demonstrated that SCD1 inhibition caused a reduction in TAGs and phospholipids, which coincided with the downregulation of genes associated with the biosynthesis of these lipid fractions. Cellular diacylglycerols were increased with SCD1 inhibition and insulin signaling was partially impaired. In contrast, markers of cellular stress were unaltered. Furthermore, the FA composition of each lipid fraction was dramatically modified, with SCD1-inhibited adipocytes specifically upregulating the elongation of PA to SA. Stable isotope tracer experiments revealed that this elongation was occurring via elongase 6. Additionally, reduced SCD1 activity in adipocytes exacerbated the effects of exogenous SA on markers of inflammation. Taken together, SCD1 activity has many indirect influences on adipocyte metabolism in addition to its role in FA desaturation. Importantly, SCD1 facilitates the storage of FAs in TAGs and the ability of adipocytes to handle exogenous FAs. SCD1 also prevents saturated FA and DAG accumulation, and preserves insulin signaling in adipocytes. Ultimately this thesis highlights the importance of SCD1 in the maintenance of adipocyte cellular function, and emphasizes the wide-ranging impact of SCD1 on adipocyte FA metabolism.

Recent progress in the discovery and development of stearoyl CoA desaturase inhibitors

Stearoyl-CoA desaturase 1 (SCD1) is a rate-limiting enzyme in the biosynthesis of monounsaturated fatty acids from their saturated fatty acid precursors. SCD1 introduces a cis-double bond at the Δ9 position (between carbons 9 and 10) of stearoyl (C18:0) and palmitoyl-CoA (C16:0). SCD1 has been shown to be a crucial factor in lipid metabolism and body weight control. In addition, SCD1 inhibitors are claimed to be new treatments for various diseases, such as skin disorders, nonalcoholic steatohepatitis (NASH), hepatitis C virus (HCV), Alzheimer’s disease, or cancer. This review aims to summarize the examples of the recently reported novel SCD1 inhibitors and to highlight the emerging areas of target indications that may hold promise for the development of SCD1 inhibitors.

Abstract 4459: Accelerated drug discovery platform yields synthesis of novel stearoyl-CoA desaturase 1 inhibitors that demonstrate anti-tumor efficacy in several models of aggressive cancer

Abstract Background Intra-tumor heterogeneity is the driving force behind disease progression and metastasis, drug resistance, and relapse in cancer patients. Defining personalized treatment regimens and increasing the arsenal of targeted therapies available will provide the best chance of prolonging survival and finding curative treatments. Recent work by our group and others has shed light on aberrant de novo lipogenesis as a feature of many cancers including kidney, breast, and lung cancer. Of the molecules involved in fatty acid metabolism, stearoyl-CoA desaturase 1 (SCD1) is frequently over-expressed. Targeted inhibition of SCD1 demonstrates a loss of tumor cell viability in a variety of cancer models. While there currently are specific small molecule inhibitors for SCD1 none are under investigation as anti-cancer therapeutics. In order to evaluate the efficacy of SCD1 inhibitors against ccRCC in both a preclinical and eventually a clinical setting we sought to generate unique compounds and test their biological efficacy. Methods Utilizing an innovative in silico approach, we designed new inhibitors for SCD1 via a scaffold hopping approach while searching for different “core” scaffolds. Critical interaction moieties from the chemical R-groups were held fixed, while generated cores were rapidly scanned for best-fit criteria (shape, docking, pharmacophore fit (QSAR)) into our Z-matrix. Following de novo compound generation and synthesis, we implemented iterative analog generation for the top hits. Using this method and diversity of chemical space, we pursued three divergent species of compounds into the nanomolar regime. Biological testing of the novel compounds included high-throughput proliferative-based screening, oleic acid rescue, and evaluation of the endoplasmic stress response. Enzymatic inhibition of SCD1 was evaluated by LC-MS. Preclinical pharmacokinetics was established using a range of doses administered intravenously and orally. Anti-tumor response was evaluated in an ectopic model of renal carcinoma, both as a single agent and in combination with SOC. Results A cohort of potential unique small molecule inhibitors against SCD1 were designed and synthesized. Of these, a select group of compounds demonstrated potent and selective enzyme inhibition and reduced tumor growth in in vitro and in vivo models of ccRCC. These novel inhibitors reliably reproduced activation of ER stress markers, authenticating this biological response as a harbinger for successful abrogation of SCD1 activity. Conclusions We have established an effective, expedited, and economical format for in silico modeling of novel agents based on a scaffold-hopping methodology. Our findings provide compelling evidence supporting the therapeutic benefit of SCD1 inhibitors for clinical use as broad-spectrum anti-tumor agents alone or in combination with SOC. Citation Format: Christina A. Von Roemeling, Thomas R. Caulfield, Derek C. Radisky, Ilah Bok, Laura A. Marlow, James Miller, Mojda Sidiqi, Anthony B. Pinkerton, Winston W. Tan, Amy L. Lane, Han W. Tun, John A. Copland. Accelerated drug discovery platform yields synthesis of novel stearoyl-CoA desaturase 1 inhibitors that demonstrate anti-tumor efficacy in several models of aggressive cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4459. doi:10.1158/1538-7445.AM2015-4459

Obesity Drives Th17 Cell Differentiation by Inducing the Lipid Metabolic Kinase, ACC1

Chronic inflammation due to obesity contributes to the development of metabolic diseases, autoimmune diseases, and cancer. Reciprocal interactions between metabolic systems and immune cells have pivotal roles in the pathogenesis of obesity-associated diseases, although the mechanisms regulating obesity-associated inflammatory diseases are still unclear. In the present study, we performed transcriptional profiling of memory phenotype CD4 T cells in high-fat-fed mice and identified acetyl-CoA carboxylase 1 (ACC1, the gene product of Acaca) as an essential regulator of Th17 cell differentiation in vitro and of the pathogenicity of Th17 cells in vivo. ACC1 modulates the DNA binding of RORγt to target genes in differentiating Th17 cells. In addition, we found a strong correlation between IL-17A-producing CD45RO(+)CD4 T cells and the expression of ACACA in obese subjects. Thus, ACC1 confers the appropriate function of RORγt through fatty acid synthesis and regulates the obesity-related pathology of Th17 cells.

PPARγ Agonists Attenuate Palmitate-Induced ER Stress through Up-Regulation of SCD-1 in Macrophages.

BackgroundClinical trials have shown that treatment of patients with type 2 diabetes with pioglitazone, a peroxisome proliferator-activated receptor (PPAR)γ agonist, reduces cardiovascular events. However, the effect of PPARγ agonists on endoplasmic reticulum (ER) stress that plays an important role in the progression of atherosclerosis has not been determined. We sought to determine the effect of PPARγ agonists on ER stress induced by palmitate, the most abundant saturated fatty acid in the serum.Methods and ResultsProtein expression of ER stress marker was evaluated by Western blot analysis and stearoyl-CoA desaturase1 (SCD-1) mRNA expression was evaluated by qRT-PCR. Macrophage apoptosis was detected by flowcytometry. Pioglitazone and rosiglitazone reduced palmitate-induced phosphorylation of PERK, a marker of ER stress, in RAW264.7, a murine macrophage cell line. Pioglitazone also suppressed palmitate-induced apoptosis in association with inhibition of CHOP expression, JNK phosphorylation and cleavage of caspase-3. These effects of pioglitazone were reversed by GW9662, a PPARγ antagonist, indicating that PPARγ is involved in this process. PPARγ agonists increased expression of SCD-1 that introduces a double bond on the acyl chain of long-chain fatty acid. 4-(2-Chlorophenoxy)-N-(3-(3-methylcarbamoyl)phenyl)piperidine-1-carboxamide, an inhibitor of SCD-1, abolished the anti-ER stress and anti-apoptotic effects of pioglitazone. These results suggest that PPARγ agonists attenuate palmitate-induced ER stress and apoptosis through SCD-1 induction. Up-regulation of SCD-1 may contribute to the reduction of cardiovascular events by treatment with PPARγ agonists.

17β-estradiol induces stearoyl-CoA desaturase-1 expression in estrogen receptor-positive breast cancer cells.

BackgroundTo sustain cell growth, cancer cells exhibit an altered metabolism characterized by increased lipogenesis. Stearoyl-CoA desaturase-1 (SCD-1) catalyzes the production of monounsaturated fatty acids that are essential for membrane biogenesis, and is required for cell proliferation in many cancer cell types. Although estrogen is required for the proliferation of many estrogen-sensitive breast carcinoma cells, it is also a repressor of SCD-1 expression in liver and adipose. The current study addresses this apparent paradox by investigating the impact of estrogen on SCD-1 expression in estrogen receptor-α-positive breast carcinoma cell lines.MethodsMCF-7 and T47D mammary carcinomas cells and immortalized MCF-10A mammary epithelial cells were hormone-starved then treated or not with 17β-estradiol. SCD-1 activity was assessed by measuring cellular monounsaturated/saturated fatty acid (MUFA/SFA) ratios, and SCD-1 expression was measured by qPCR, immunoblot, and immunofluorescence analyses. The role of SCD-1 in cell proliferation was measured following treatment with the SCD-1 inhibitor A959372 and following SCD-1 silencing using siRNA. The involvement of IGF-1R on SCD-1 expression was measured using the IGF-1R antagonist AG1024. The expression of SREBP-1c, a transcription factor that regulates SCD-1, was measured by qPCR, and by immunoblot analyses.Results17β-estradiol significantly induced cell proliferation and SCD-1 activity in MCF-7 and T47D cells but not MCF-10A cells. Accordingly, 17β-estradiol significantly increased SCD-1 mRNA and protein expression in MCF-7 and T47D cells compared to untreated cells. Treatment of MCF-7 cells with 4-OH tamoxifen or siRNA silencing of estrogen receptor-α largely prevented 17β-estradiol-induced SCD-1 expression. 17β-estradiol increased SREBP-1c expression and induced the mature active 60 kDa form of SREBP-1. The selective SCD-1 inhibitor or siRNA silencing of SCD-1 blocked the 17β-estradiol-induced cell proliferation and increase in cellular MUFA/SFA ratios. IGF-1 also induced SCD-1 expression, but to a lesser extent than 17β-estradiol. The IGF-1R antagonist partially blocked 17β-estradiol-induced cell proliferation and SCD-1 expression, suggesting the impact of 17β-estradiol on SCD-1 expression is partially mediated though IGF-1R signaling.ConclusionsThis study illustrates for the first time that, in contrast to hepatic and adipose tissue, estrogen induces SCD-1 expression and activity in breast carcinoma cells. These results support SCD-1 as a therapeutic target in estrogen-sensitive breast cancer.