Fatty Acid Synthase Inhibitor C75 Research Articles

Fatty Acid Synthase Promotes Hepatocellular Carcinoma Growth via S-Phase Kinase-Associated Protein 2/p27KIP1 Regulation.

Background and Objectives: Aberrant upregulation of fatty acid synthase (FASN), catalyzing de novo synthesis of fatty acids, occurs in various tumor types, including human hepatocellular carcinoma (HCC). Although FASN oncogenic activity seems to reside in its pro-lipogenic function, cumulating evidence suggests that FASN's tumor-supporting role might also be metabolic-independent. Materials and Methods: In the present study, we show that FASN inactivation by specific small interfering RNA (siRNA) promoted the downregulation of the S-phase kinase associated-protein kinase 2 (SKP2) and the consequent induction of p27KIP1 in HCC cell lines. Results: Expression levels of FASN and SKP2 directly correlated in human HCC specimens and predicted a dismal outcome. In addition, forced overexpression of SKP2 rendered HCC cells resistant to the treatment with the FASN inhibitor C75. Furthermore, FASN deletion was paralleled by SKP2 downregulation and p27KIP1 induction in the AKT-driven HCC preclinical mouse model. Moreover, forced overexpression of an SKP2 dominant negative form or a p27KIP1 non-phosphorylatable (p27KIP1-T187A) construct completely abolished AKT-dependent hepatocarcinogenesis in vitro and in vivo. Conclusions: In conclusion, the present data indicate that SKP2 is a critical downstream effector of FASN and AKT-dependent hepatocarcinogenesis in liver cancer, envisaging the possibility of effectively targeting FASN-positive liver tumors with SKP2 inhibitors or p27KIP1 activators.

Inhibition of palmitoyltransferase ZDHHC12 sensitizes ovarian cancer cells to cisplatin through ROS-mediated mechanisms.

Platinum-based therapies have revolutionized the treatment of high-grade serous ovarian cancer (HGSOC). However, high rates of disease recurrence and progression remain a major clinical concern. Impaired mitochondrial function and dysregulated reactive oxygen species (ROS), hallmarks of cancer, hold potential as therapeutic targets for selectively sensitizing cisplatin treatment. Here, we uncover an oncogenic role of the palmitoyltransferase ZDHHC12 in regulating mitochondrial function and ROS homeostasis in HGSOC cells. Analysis of The Cancer Genome Atlas (TCGA) ovarian cancer data revealed significantly elevated ZDHHC12 expression, demonstrating the strongest positive association with ROS pathways among all ZDHHC enzymes. Transcriptomic analysis of independent ovarian cancer datasets and the SNU119 cell model corroborated this association, highlighting a strong link between ZDHHC12 expression and signature pathways involving mitochondrial oxidative metabolism and ROS regulation. Knockdown of ZDHHC12 disrupted this association, leading to increased cellular complexity, ATP levels, mitochondrial activity, and both mitochondrial and cellular ROS. This dysregulation, achieved by the siRNA knockdown of ZDHHC12 or treatment with the general palmitoylation inhibitor 2BP or the fatty acid synthase inhibitor C75, significantly enhanced cisplatin cytotoxicity in 2D and 3D spheroid models of HGSOC through ROS-mediated mechanisms. Markedly, ZDHHC12 inhibition significantly augmented the anti-tumor activity of cisplatin in an ovarian cancer xenograft tumor model, as well as in an ascites-derived organoid line of platinum-resistant ovarian cancer. Our data suggest the potential of ZDHHC12 as a promising target to improve the outcome of HGSOCs in response to platinum-based chemotherapy.

Sortilin-Driven Cancer Secretome Enhances Tumorigenic Properties of Hepatocellular Carcinoma via de Novo Lipogenesis

A growing body of evidence suggests de novo lipogenesis as a key metabolic pathway adopted by cancers to fuel tumorigenic processes. While increased de novo lipogenesis has also been reported in hepatocellular carcinoma (HCC), understanding on molecular mechanisms driving de novo lipogenesis remains limited. In the present study, the functional role of sortilin, a member of the vacuolar protein sorting 10 protein receptor family, in HCC was investigated. Sortilin was overexpressed in HCC and was associated with poorer survival outcome. In functional studies, sortilin-overexpressing cells conferred tumorigenic phenotypes, namely, self-renewal and metastatic potential, of HCC cells via the cancer secretome. Proteomic profiling highlighted fatty acid metabolism as a potential molecular pathway associated with sortilin-driven cancer secretome. This finding was validated by the increased lipid content and expression of fatty acid synthase (FASN) in HCC cells treated with conditioned medium collected from sortilin-overexpressing cells. The enhanced tumorigenic properties endowed by sortilin-driven cancer secretome were partly abrogated by co-administration of FASN inhibitor C75. Further mechanistic dissection suggested protein stabilization by post-translational modification with O-GlcNAcylation as a major mechanism leading to augmented FASN expression. In conclusion, the present study uncovered the role of sortilin in hepatocarcinogenesis via modulation of the cancer secretome and deregulated lipid metabolism.

The fatty acid synthase inhibitor C75 differentially affects the adipogenic differentiation of multipotent cells and preadipocytes.

Previously, we revealed the dual enhancing effect of netoglitazone, an agonist of the peroxisome proliferator-activated receptor γ, on adipogenesis and osteoblastogenesis, and reported that fatty acid synthase (FASN) knockdown selectively repressed its pro-adipogenic effect. Here, we examined if a FASN inhibitor, C75, could selectively repress the pro-adipogenic effect of netoglitazone. Surprisingly, C75 promoted the adipogenic differentiation of multipotent C3H10T1/2 cells but inhibited 3T3-L1 preadipocytes. By identifying glycogen synthase kinase-3β and intracellular cAMP levels as regulatory targets of C75, we ultimately found the differential expression of adenosine receptor 3 (AR3) and AR2a on these cells. Inhibition of AR3 on C3H10T1/2 and AR2a on 3T3-L1 inhibited the effects of C75 on the differentiation of these cells. Our findings imply that cell-type-specific AR expression might account for the differential adipogenic effects of C75.

Apolipoprotein E enhance survival of effector memory regulatory T lymphocytes by regulating caspase-dependent apoptosis and lipid metabolism.

Abstract Mechanisms underlying the pleiotropic anti-inflammatory effect of HDL remain incompletely understood. Regulatory T cells (Tregs) are essential for maintaining immune homeostasis in cardiovascular diseases (CVD) and metabolic disorders. We recently show Tregs, particularly effector memory Tregs (emTregs), bind/internalize HDL, which promotes their survival by limiting caspase-dependent apoptosis. As HDL contain a complex mix of proteins and lipids, we investigated which components of HDL mediate this process and found HDL proteins were the most critical. Using reconstituted HDL mimic vesicles containing recombinant HDL proteins and POPC, we found that APOE, but none of the APOA proteins, significantly decreased caspase expression in emTregs similarly to total HDL. Neither APOE nor HDL had any effect on naïve Tregs. APOE-depleted HDL did not rescue emTregs, confirming the critical role of APOE. Because Tregs have distinct metabolic requirements, we next investigated the effect of HDL, APOE3-POPC and APOE-depleted HDL on lipid metabolism in emTregs. APOE3-POPC and HDL significantly increased the concentration of long chain (14:0 20:0) and polyunsaturated fatty acids (FA, 18:3 22:6) in emTregs. Our data also suggested the involvement of de novo FA synthesis, as APOE3-POPC and HDL increased the concentration of palmitate (16:0), the earlier precursor of this pathway. Further, treatment of Tregs with the fatty acid synthase inhibitor C75 abolished HDL survival effect. Our results show that HDL, specifically APOE, promote emTregs survival by regulating de novo FA synthesis and caspase-dependent apoptosis. HDL-APOE levels seem to be dysregulated in CVD and obesity, suggesting this mechanism may contribute to HDL protective role

Palmitoylation of SARS-CoV-2 S protein is critical for S-mediated syncytia formation and virus entry.

Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is the cause of the ongoing coronavirus disease 2019 (COVID‐19) pandemic. The S protein is the key viral protein for associating with ACE2, the receptor for SARS‐CoV‐2. There are many kinds of posttranslational modifications in S protein. However, the detailed mechanism of palmitoylation of SARS‐CoV‐2 S remains to be elucidated. In our current study, we characterized the palmitoylation of SARS‐CoV‐2 S. Both the C15 and cytoplasmic tail of SARS‐CoV‐2 S were palmitoylated. Fatty acid synthase inhibitor C75 and zinc finger DHHC domain‐containing palmitoyltransferase (ZDHHC) inhibitor 2‐BP reduced the palmitoylation of S. Interestingly, palmitoylation of SARS‐CoV‐2 S was not required for plasma membrane targeting of S but was critical for S‐mediated syncytia formation and SARS‐CoV‐2 pseudovirus particle entry. Overexpression of ZDHHC2, ZDHHC3, ZDHHC4, ZDHHC5, ZDHHC8, ZDHHC9, ZDHHC11, ZDHHC14, ZDHHC16, ZDHHC19, and ZDHHC20 promoted the palmitoylation of S. Furthermore, those ZDHHCs were identified to associate with SARS‐CoV‐2 S. Our study not only reveals the mechanism of S palmitoylation but also will shed important light into the role of S palmitoylation in syncytia formation and virus entry.

Progesterone receptor isoform-dependent cross-talk between prolactin and fatty acid synthase in breast cancer.

Progesterone receptor (PR) isoforms can drive unique phenotypes in luminal breast cancer (BC). Here, we hypothesized that PR-B and PR-A isoforms differentially modify the cross-talk between prolactin and fatty acid synthase (FASN) in BC. We profiled the responsiveness of the FASN gene promoter to prolactin in T47Dco BC cells constitutively expressing PR-A and PR-B, in the PR-null variant T47D-Y cell line, and in PR-null T47D-Y cells engineered to stably re-express PR-A (T47D-YA) or PR-B (T47D-YB). The capacity of prolactin to up-regulate FASN gene promoter activity in T47Dco cells was lost in T47D-Y and TD47-YA cells. Constitutively up-regulated FASN gene expression in T47-YB cells and its further stimulation by prolactin were both suppressed by the prolactin receptor antagonist hPRL-G129R. The ability of the FASN inhibitor C75 to decrease prolactin secretion was more conspicuous in T47-YB cells. In T47D-Y cells, which secreted notably less prolactin and downregulated prolactin receptor expression relative to T47Dco cells, FASN blockade resulted in an augmented secretion of prolactin and up-regulation of prolactin receptor expression. Our data reveal unforeseen PR-B isoform-specific regulatory actions in the cross-talk between prolactin and FASN signaling in BC. These findings might provide new PR-B/FASN-centered predictive and therapeutic modalities in luminal intrinsic BC subtypes.

Cytotoxicity and Radiosensitizing Activity of the Fatty Acid Synthase Inhibitor C75 Is Enhanced by Blocking Fatty Acid Uptake in Prostate Cancer Cells

Prostate cancers, like many other types of cancer, express elevated levels of fatty acid synthase (FASN) to make more fatty acids, which are required for energy, signaling, and proliferation. Because inhibition of FASN has been shown to sensitize tumors to chemotherapy and radiation, we studied the effect of C75, a radiosensitizing FASN inhibitor, and compared its single agent and radiosensitizing activities in 2 prostate cancer cell lines, PC3 and LNCaP, with alternative FASN inhibitors that have progressed into clinical trials. We also investigated the effect of serum and fatty acid supplementation on responses to FASN inhibitors, probing expression of key proteins related to fatty acid uptake in response to FASN inhibition, irradiation, and serum lipid concentration and how this may be modulated to increase the potency of C75. We demonstrated that C75 was the only FASN inhibitor to sensitize cells to ionizing radiation; no sensitization was apparent with FASN inhibitors TVB-3166 or Orlistat. The prostate cancer cell lines were able to take up fatty acids from the culture medium, and the availability of fatty acids affected sensitivity of these cells to C75 but not the other FASN inhibitors tested. C75 also increased expression of fatty acid transporter proteins FATP1 and CD36. Furthermore, blocking CD36 with antibody increased the sensitivity of cells to C75. We suggest that the potency of C75 is affected by fatty acid availability and that the effectiveness of FASN inhibitors in combination with ionizing radiation can be further enhanced by regulating fatty acid uptake.

Effect of interferon gamma on neutral lipid levels, lipid droplet formation, and antiviral responses in pancreatic islets and INS‐1 β cells

Interferon (IFN)‐regulated pathways play a critical role in the development of type 1 diabetes (T1D) following exposure to environmental insults including viral infection. IFNs have been shown to modulate lipid metabolism in both immune and non‐hematopoietic cells in response to pathogens. Little is known, however, about the impact of IFN signaling on lipid metabolism in pancreatic β cells and whether IFN‐induced changes in β cell lipid metabolism are associated with antiviral responses and β cell dysfunction. To characterize islet lipid changes potentially mediated by IFNs during initiation of β cell autoimmunity in vivo, juvenile LEW.1WR1 rats (an IFN‐dependent, inducible‐T1D model) were treated with the viral analog polyinosinic:polycytidylic (PIC) every other day for 6 days then islets were isolated 16h and 48h post last treatment for gene expression and non‐targeted lipidomics, respectively. Islets from PIC‐treated animals displayed a robust induction of IFN gamma (IFNγ) target genes, e.g. IFNγ‐inducible protein and ubiquitin D, and significantly increased islet triacylglyceride (TAG) and non‐esterified fatty acids (NEFA). Similar increases in TAG and NEFA were observed when isolated human islets were treated ex vivo for 24h with mixtures of IFNγ and IL1β, or IFNγ and TNFα, suggesting that IFNγ drives TAG synthesis or accumulation. To investigate the direct effect of IFNγ, INS‐1 β cells were treated with IFNγ for 24h. Consistent with the in vivo and ex vivo islet results, IFNγ‐treated cells had significantly higher TAG levels. IFNγ also increased mRNA levels of genes involved in lipid droplet (LD) formation and clustering (e.g. perilipins and fat‐induced storage membrane protein 1) and led to the appearance of clusters of large cytoplasmic LDs. Treatment of INS‐1 cells with IFNγ also increased NEFA levels, most notably arachidonic acid. Although the source for increased TAG and NEFA is uncertain, IFNγ induced fatty acid synthase (FASN) gene expression suggesting increased de novo FA synthesis. IFNγ‐mediated changes in lipid metabolism were associated with impaired insulin secretion. IFNγ, however, did not increase the expression of endoplasmic reticulum (ER) stress markers, but sensitized INS‐1 cells to TNFα‐ or IL1β‐induced ER stress and apoptosis. Importantly, pretreatment of INS‐1 β cells with IFNγ for 12 h markedly increased PIC‐induced expression of antiviral genes (e.g. Ifnb, Mx1) and this synergistic response was abrogated by the FASN inhibitor C75. Inhibition of FASN with C75 also led to a redistribution of IFNγ‐mediated LD clusters into more discrete LDs. Overall, these data suggest that IFNγ promotes LD formation and clustering in β cells, and these LD clusters serve as scaffolds to enhance expression of antiviral response genes. Unfortunately, these lipid metabolism changes are also associated with impaired secretory function and increased susceptibility to cytokine‐induced ER stress and β cell death.Support or Funding InformationJDRF grant 3‐SRE‐2014‐43‐Q‐R to LKO.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

EGCG-Derivative G28 Shows High Efficacy Inhibiting the Mammosphere-Forming Capacity of Sensitive and Resistant TNBC Models.

Recent studies showed that Fatty Acid Synthase (FASN), a lipogenic enzyme overexpressed in several carcinomas, plays an important role in drug resistance. Furthermore, the enrichment of Breast Cancer Stem Cell (BCSC) features has been found in breast tumors that progressed after chemotherapy. Hence, we used the triple negative breast cancer (TNBC) cell line MDA-MB-231 (231) to evaluate the FASN and BCSC population role in resistance acquisition to chemotherapy. For this reason, parental cell line (231) and its derivatives resistant to doxorubicin (231DXR) and paclitaxel (231PTR) were used. The Mammosphere-Forming Assay and aldehyde dehydrogenase (ALDH) enzyme activity assay showed an increase in BCSCs in the doxorubicin-resistant model. Moreover, the expression of some transcription factors involved in epithelial-mesenchymal transition (EMT), a process that confers BCSC characteristics, was upregulated after chemotherapy treatment. FASN inhibitors C75, (−)-Epigallocatechin 3-gallate (EGCG), and its synthetic derivatives G28, G56 and G37 were used to evaluate the effect of FASN inhibition on the BCSC-enriched population in our cell lines. G28 showed a noticeable antiproliferative effect in adherent conditions and, interestingly, a high mammosphere-forming inhibition capacity in all cell models. Our preliminary results highlight the importance of studying FASN inhibitors for the treatment of TNBC patients, especially those who progress after chemotherapy.

An unexpected link between fatty acid synthase and cholesterol synthesis in proinflammatory macrophage activation

Different immune activation states require distinct metabolic features and activities in immune cells. For instance, inhibition of fatty acid synthase (FASN), which catalyzes the synthesis of long-chain fatty acids, prevents the proinflammatory response in macrophages; however, the precise role of this enzyme in this response remains poorly defined. Consistent with previous studies, we found here that FASN is essential for lipopolysaccharide-induced, Toll-like receptor (TLR)-mediated macrophage activation. Interestingly, only agents that block FASN upstream of acetoacetyl-CoA synthesis, including the well-characterized FASN inhibitor C75, inhibited TLR4 signaling, while those acting downstream had no effect. We found that acetoacetyl-CoA could overcome C75's inhibitory effect, whereas other FASN metabolites, including palmitate, did not prevent C75-mediated inhibition. This suggested an unexpected role for acetoacetyl-CoA in inflammation that is independent of its role in palmitate synthesis. Our evidence further suggested that acetoacetyl-CoA arising from FASN activity promotes cholesterol production, indicating a surprising link between fatty acid synthesis and cholesterol synthesis. We further demonstrate that this process is required for TLR4 to enter lipid rafts and facilitate TLR4 signaling. In conclusion, we have uncovered an unexpected link between FASN and cholesterol synthesis that appears to be required for TLR signal transduction and proinflammatory macrophage activation.

Lipid Bodies as Sites of Prostaglandin E2 Synthesis During Chagas Disease: Impact in the Parasite Escape Mechanism.

During Chagas disease, the Trypanosoma cruzi can induce some changes in the host cells in order to escape or manipulate the host immune response. The modulation of the lipid metabolism in the host phagocytes or in the parasite itself is one feature that has been observed. The goal of this mini review is to discuss the mechanisms that regulate intracellular lipid body (LB) biogenesis in the course of this parasite infection and their meaning to the pathophysiology of the disease. The interaction host–parasite induces LB (or lipid droplet) formation in a Toll-like receptor 2-dependent mechanism in macrophages and is enhanced by apoptotic cell uptake. Simultaneously, there is a lipid accumulation in the parasite due to the incorporation of host fatty acids. The increase in the LB accumulation during infection is correlated with an increase in the synthesis of PGE2 within the host cells and the parasite LBs. Moreover, the treatment with fatty acid synthase inhibitor C75 or non-steroidal anti-inflammatory drugs such as NS-398 and aspirin inhibited the LB biogenesis and also induced the down modulation of the eicosanoid production and the parasite replication. These findings show that LBs are organelles up modulated during the course of infection. Furthermore, the biogenesis of the LB is involved in the lipid mediator generation by both the macrophages and the parasite triggering escape mechanisms.

Abstract 346: Fatty acid synthase is a risk factor for bleomycin-induced lung fibrosis in mice

Abstract Pulmonary fibrosis is a significant complication in cancer patients treated with radiation or various chemotherapeutic agents. One such reagent, bleomycin, is used to treat a variety of cancers including, but not limited to, lymphoma, ovarian cancer, and melanoma. Its efficacy, however, is impacted by the incidence of significant pulmonary fibrosis, which often becomes dose limiting. In studies investigating the role(s) of metabolism in the profibrotic actions of transforming growth factor beta, we previously determined that increased fatty acid synthase (FASN) expression was observed in the fibrotic areas of lungs from idiopathic pulmonary fibrosis (IPF) patients compared to healthy controls. In that FASN levels are significantly increased in numerous human tumors including breast, prostate, colorectal, and ovarian, we wished to integrate these findings by examining whether inhibiting FASN activity could reduce pulmonary fibrosis. To that end, the cerulenin-derived FASN inhibitor C75 was tested in a murine treatment model of bleomycin-induced lung fibrosis. Following addition of C75, there was significant diminution in a number of profibrotic targets including collagen 1α1, fibronectin, connective tissue growth factor, and total lung collagen as assessed by hydroxyproline content. Not only was there a C75 dose-dependent decrease in the aforementioned targets, but lung function as assessed by peripheral blood oxygenation (on room air) was also stabilized/improved. Thus, FASN inhibition might reflect a possible target to maintain normal lung physiology in the context of cancer chemotherapy. This work was supported by Public Health Service grants GM-055816 and GM-054200 from the National Institutes of General Medical Sciences. Citation Format: Mi-Yeon Jung. Fatty acid synthase is a risk factor for bleomycin-induced lung fibrosis in mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 346. doi:10.1158/1538-7445.AM2017-346

MACC1 decreases the chemosensitivity of gastric cancer cells to oxaliplatin by regulating FASN expression.

The effect of chemotherapeutic agents is limited as a result of drug resistance, which demands prompt solutions provided by clinical studies. To date, the underlying mechanisms of chemotherapy resistance are relatively unknown. Metastasis-associated in colon cancer 1 (MACC1) is an oncogene associated with the progression and prognosis of gastric cancer (GC). Bioinformatic analysis revealed that MACC1 is positively associated with fatty acid synthase (FASN), a major enzyme of lipogenesis, and drives chemoresistance to oxaliplatin in GC. Similar findings were demonstrated in two GC cell lines (BGC-823 and MKN-28) with MACC1 ectopic expression. We next employed FASN inhibitor C75 or siFASN (small interfering RNA targeted to FASN) to block endogenous fatty acid metabolism and it was revealed that cell proliferation and chemoresistance to oxaliplatin induced by MACC1 upregulation were attenuated by FASN blockade to various extents. Conclusively, these outcomes highlight a novel role of MACC1 in GC cell lipogenesis, and suggest that MACC1 may be an attractive target to decrease oxaliplatin resistance in GC.

Differential in radiosensitizing potency of enantiomers of the fatty acid synthase inhibitor C75.

The elevated activity of fatty acid synthase has been reported in a number of cancer types. Inhibition of this enzyme has been demonstrated to induce cancer cell death and reduce tumor growth. In addition, the fatty acid synthase inhibitor drug C75 has been reported to synergistically enhance the cancer‐killing ability of ionizing radiation. However, clinical use of C75 has been limited due to its producing weight loss, believed to be caused by alterations in the activity of carnitine palmitoyltransferase‐1. C75 is administered in the form of a racemic mixture of (−) and (+) enantiomers that may differ in their regulation of fatty acid synthase and carnitine palmitoyltransferase‐1. Therefore, we assessed the relative cancer‐killing potency of different enantiomeric forms of C75 in prostate cancer cells. These results suggest that (−)‐C75 is the more cytotoxic enantiomer and has greater radiosensitizing capacity than (+)‐C75. These observations will stimulate the development of fatty acid synthase inhibitors that are selective for cancer cells and enhance the tumor‐killing activity of ionizing radiation, while minimizing weight loss in cancer patients.

Clinical and therapeutic relevance of the metabolic oncogene fatty acid synthase in HER2+ breast cancer.

Fatty acid synthase (FASN) is a key lipogenic enzyme for de novo fatty acid biosynthesis and a druggable metabolic oncoprotein that is activated in most human cancers. We evaluated whether the HER2-driven lipogenic phenotype might represent a biomarker for sensitivity to pharmacological FASN blockade. A majority of clinically HER2-positive tumors were scored as FASN overexpressors in a series of almost 200 patients with invasive breast carcinoma. Re-classification of HER2-positive breast tumors based on FASN gene expression predicted a significantly inferior relapse-free and distant metastasis-free survival in HER2+/FASN+ patients. Notably, non-tumorigenic MCF10A breast epithelial cells engineered to overexpress HER2 upregulated FASN gene expression, and the FASN inhibitor C75 abolished HER2-induced anchorage-independent growth and survival. Furthermore, in the presence of high concentrations of C75, HER2-negative MCF-7 breast cancer cells overexpressing HER2 (MCF-7/HER2) had significantly higher levels of apoptosis than HER2-negative cells. Finally, C75 at non-cytotoxic concentrations significantly reduced the capacity of MCF-7/HER2 cells to form mammospheres, an in vitro indicator of cancer stem-like cells. Collectively, our findings strongly suggest that the HER2-FASN lipogenic axis delineates a group of breast cancer patients that might benefit from treatment with therapeutic regimens containing FASN inhibitors.

Abstract 5103: Crosstalk between osteoprotegerin (OPG), fatty acid synthase (FASN), and cyclooxygenase-2 (COX-2) in breast cancer: implications in carcinogenesis

Abstract The cross-talk between malignant and nonmalignant cells in the tumor microenvironment, as maneuvered by cytokines/chemokines, drives breast cancer progression. We demonstrated that OPG is expressed and secreted at very high levels from the highly invasive breast cancer cell lines SUM149PT and SUM1315MO2 as compared to human normal mammary epithelial HMEC cells (1). OPG was involved in modulating aneuploidy, cell proliferation, and angiogenesis in breast cancer (1). Mass spectrometry analysis performed in this study revealed OPG interacts with fatty acid synthase (FASN), which is a key enzyme of the fatty acid biosynthetic pathway in breast cancer cells. Further, electron microscopy, immunofluorescence, and fluorescence quantitation assay highlighted the presence of a large number of lipid bodies (lipid droplets) in SUM149PT and SUM1315MO2 cells in comparison to HMEC. We recently showed upregulation of COX-2 inflammatory pathway and its metabolite PGE2 secretion in SUM149PT and SUM1315MO2 breast cancer cells (2). Interestingly, human breast cancer tissue samples displayed high expression of OPG, PGE2 and FASN. Immunofluorescence staining exposed the co-existence of COX-2 and FASN in the lipid bodies of breast cancer cells. We reasoned that there might be cross-talk between OPG, FASN, and COX-2 that sustains the inflammatory pathways in breast cancer. Here, we identified cis-acting elements involved in the transcriptional regulation of COX-2 and FASN by recombinant human OPG (rhOPG). Treatment with FASN inhibitor C75 decreased the number of lipid body/cell, downregulated phosphorylation of ERK, GSK3β, and induced apoptosis by caspase-3 and caspapse-9 activation. Collectively, the novel biological cross-talk between OPG, FASN, and COX-2 advocates for combinatorial drug treatment to block these players of carcinogenesis as a promising therapeutic to target highly invasive breast cancer.

Crosstalk between osteoprotegerin (OPG), fatty acid synthase (FASN) and, cycloxygenase-2 (COX-2) in breast cancer: implications in carcinogenesis.

The crosstalk between malignant and nonmalignant cells in the tumor microenvironment, as maneuvered by cytokines/chemokines, drives breast cancer progression. In our previous study, we discovered Osteoprotegerin (OPG) as one of the cytokines heavily secreted by breast cancer cells. We demonstrated that OPG is expressed and secreted at very high levels from the highly invasive breast cancer cell lines SUM149PT and SUM1315MO2 as compared to normal human mammary epithelial HMEC cells. OPG was involved in modulating aneuploidy, cell proliferation, and angiogenesis in breast cancer. Mass spectrometry analysis performed in this study revealed OPG interacts with fatty acid synthase (FASN), which is a key enzyme of the fatty acid biosynthetic pathway in breast cancer cells. Further, electron microscopy, immunofluorescence, and fluorescence quantitation assays highlighted the presence of a large number of lipid bodies (lipid droplets) in SUM149PT and SUM1315MO2 cells in comparison to HMEC. We recently showed upregulation of the COX-2 inflammatory pathway and its metabolite PGE2 secretion in SUM149PT and SUM1315MO2 breast cancer cells. Interestingly, human breast cancer tissue samples displayed high expression of OPG, PGE2 and fatty acid synthase (FASN). FASN is a multifunctional enzyme involved in lipid biosynthesis. Immunofluorescence staining revealed the co-existence of COX-2 and FASN in the lipid bodies of breast cancer cells. We reasoned that there might be crosstalk between OPG, FASN, and COX-2 that sustains the inflammatory pathways in breast cancer. Interestingly, knocking down OPG by CRISPR/Cas9 gene editing in breast cancer cells decreased FASN expression at the protein level. Here, we identified cis-acting elements involved in the transcriptional regulation of COX-2 and FASN by recombinant human OPG (rhOPG). Treatment with FASN inhibitor C75 and COX-2 inhibitor celecoxib individually decreased the number of lipid bodies/cell, downregulated phosphorylation of ERK, GSK3β, and induced apoptosis by caspase-3/7 and caspase-9 activation. But a more efficient and effective decrease in lipid bodies/cell and survival kinase signaling was observed upon combining the drug treatments for the aggressive cancer cells. Collectively, the novel biological crosstalk between OPG, FASN, and COX-2 advocates for combinatorial drug treatment to block these players of carcinogenesis as a promising therapeutic target to treat highly invasive breast cancer.

2568 Fatty acid synthase inhibitor C75 sensitizes prostate cancer cells to radiation

2568 Fatty acid synthase inhibitor C75 sensitizes prostate cancer cells to radiation

Abstract 819: Fatty acid synthase, cycloxygenase-2 (COX-2) and osteoprotegerin (OPG) expression in invasive breast cancer: Implications in carcinogenesis

Abstract Breast cancer, the leading cause of death among women with an onset frequency of one in eight, is the most common type of cancer among women. However, despite the advancement in therapy, the mortality rate in breast cancer patients still remains high. The role of various different genes, directly or indirectly, regulating these processes have tremendously added to our knowledge of the complex process of proliferation, which has remained the most important prognostic factor so far. In our previous study, we established Osteoprotegerin (OPG)'s role in aneuploidy, cell proliferation and angiogenesis in breast cancer. We demonstrated that OPG is expressed and secreted at very high levels from the highly invasive breast cancer cell lines SUM149PT and SUM1315MO2 as compared to human normal mammary epithelial cells (HMEC). Here, we demonstrated by mass spectrometry that OPG pulls down fatty acid synthase (FASN), a key enzyme of the fatty acid biosynthetic pathway in breast cancer cells. We observed large number of lipid droplets in SUM149PT and SUM1315MO2 cells in comparison to HMEC by electron microscopy. Human breast cancer tissue samples showed high expression of FASN. Treatment with FASN inhibitor C75 decreased the number of lipid droplets/cell, altered expression of genes involved in cell proliferation, cell signaling (Akt/PI3K, ERK, GSK3β) apoptosis (caspase-3 and caspapse-9) and cell cycle. Treatment with C75 lowered the inflammatory pathway enzyme cyclooxygenase-2 (COX-2) expression and reduced secretion of its inflammatory metabolite prosataglandin E2 (PGE2). We reasoned that there might be crosstalk between OPG, FASN, and COX-2, that sustains the inflammatory pathways, and drives the progression of breast cancer. Here, we identified cis-acting elements involved in the transcriptional regulation of COX-2 by recombinant human OPG. Collectively, our study indicates the crosstalk between FASN, COX-2 and OPG. Combinatorial drug treatment of these main players of carcinogenesis can be used as a potential therapeutic target to treat highly invasive breast cancer. Citation Format: Sudeshna Goswami, Neelam Sharma-Walia. Fatty acid synthase, cycloxygenase-2 (COX-2) and osteoprotegerin (OPG) expression in invasive breast cancer: Implications in carcinogenesis. [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 819. doi:10.1158/1538-7445.AM2015-819