COX-independent Mechanisms Research Articles

Neural circuits mediating circulating interleukin-1β-evoked fever in the absence of prostaglandin E2 production

Infectious diseases and inflammatory conditions recruit the immune system to mount an appropriate acute response that includes the production of cytokines. Cytokines evoke neurally-mediated responses to fight pathogens, such as the recruitment of thermoeffectors, thereby increasing body temperature and leading to fever. Studies suggest that the cytokine interleukin-1β (IL-1β) depends upon cyclooxygenase (COX)-mediated prostaglandin E2 production for the induction of neural mechanisms to elicit fever. However, COX inhibitors do not eliminate IL-1β-induced fever, thus suggesting that COX-dependent and COX-independent mechanisms are recruited for increasing body temperature after peripheral administration of IL-1β. In the present study, we aimed to build a foundation for the neural circuit(s) controlling COX-independent, inflammatory fever by determining the involvement of brain areas that are critical for controlling the sympathetic outflow to brown adipose tissue (BAT) and the cutaneous vasculature. In anesthetized rats, pretreatment with indomethacin, a non-selective COX inhibitor, did not prevent BAT thermogenesis or cutaneous vasoconstriction (CVC) induced by intravenous IL-1β (2 µg/kg). BAT and cutaneous vasculature sympathetic premotor neurons in the rostral raphe pallidus area (rRPa) are required for IL-1β-evoked BAT thermogenesis and CVC, with or without pretreatment with indomethacin. Additionally, activation of glutamate receptors in the dorsomedial hypothalamus (DMH) is required for COX-independent, IL-1β-induced BAT thermogenesis. Therefore, our data suggests that COX-independent mechanisms elicit activation of neurons within the DMH and rRPa, which is sufficient to trigger and mount inflammatory fever. These data provide a foundation for elucidating the brain circuits responsible for COX-independent, IL-1β-elicited fevers.

Interactions of Analgesics with Cisplatin: Modulation of Anticancer Efficacy and Potential Organ Toxicity.

Cisplatin (CDDP), one of the most eminent cancer chemotherapeutic agents, has been successfully used to treat more than half of all known cancers worldwide. Despite its effectiveness, CDDP might cause severe toxic adverse effects on multiple body organs during cancer chemotherapy, including the kidneys, heart, liver, gastrointestinal tract, and auditory system, as well as peripheral nerves causing severely painful neuropathy. The latter, among other pains patients feel during chemotherapy, is an indication for the use of analgesics during treatment with CDDP. Different types of analgesics, such as acetaminophen, non-steroidal anti-inflammatory drugs (NSAIDS), and narcotic analgesics, could be used according to the severity of pain. Administered analgesics might modulate CDDP’s efficacy as an anticancer drug. NSAIDS, on one hand, might have cytotoxic effects on their own and few of them can potentiate CDDP’s anticancer effects via inhibiting the CDDP-induced cyclooxygenase (COX) enzyme, or through COX-independent mechanisms. On the other hand, some narcotic analgesics might ameliorate CDDP’s anti-neoplastic effects, causing chemotherapy to fail. Concerning safety, some analgesics share the same adverse effects on normal tissues as CDDP, augmenting its potentially hazardous effects on organ impairment. This article offers an overview of the reported literature on the interactions between analgesics and CDDP, paying special attention to possible mechanisms that modulate CDDP’s cytotoxic efficacy and potential adverse reactions.

Anti-proliferative effects of indomethacin, acemetacin and their tromethamine salts in HCT116 human colon cancer cells

Background and Aims: Since 1980's, several preclinical studies have been published on the anti-colorectal cancer activity of the nonsteroidal anti-inflammatory drug indomethacin. The direct anti-proliferative effect of indomethacin seems to occur via a variety of reported COX-independent mechanisms. Acemetacin is a glycolic acid ester derivative of indomethacin and contrary to indomethacin, there is not much published research on anti-cancer effects of acemetacin. Herein, we compared the in vitro anti-proliferative properties of indomethacin, acemetacin, and their tromethamine salts in HCT116 colon cancer cells. Methods: The tromethamine salts of indomethacin and acemetacin were synthesized and the structures were established by microanalysis, IR, 1 H-NMR, 13C-NMR (APT) and 2D-NMR (HSQC and HMBC) spectrometry. Cell proliferation assays were performed using xCELLigence real-time cell analysis system. Results: Indomethacin exhibited profound inhibitory effects with IC50 values at low micromolar ranges. Acemetacin exhibited far lower cytotoxic activity as compared to that of indomethacin. Surprisingly, indomethacin-tromethamine salt was 2-fold and 4.4-fold more potent than indomethacin at 48 and 72 h, respectively, while maintaining its activity at 24 h. The tromethamine salt of acemetacin was more potent than acemetacin at 24 h and 48 h post-treatment. Conclusion: The anti-proliferative effect of indomethacin in HCT116 cells was found to be at low micro-molar levels. The esterification of indomethacin with glycolic acid caused a strong decrease in anti-proliferative effect. The salt formation caused a positive effect on the anti-proliferative activity of indomethacin and indomethacin-tromethamine salt may be a promising candidate for additional in vivo studies.

Updates on the molecular mechanisms of aspirin in the prevention of colorectal cancer: Review.

Colorectal cancer is one of the commonest malignancies worldwide. The estimated lifetime risk of the disease is about 5% with an incidence of one million new cases and 600,000 deaths worldwide every year. It is estimated that in 2019, approximately 134,490 new cases of colorectal cancer will be diagnosed with 49,190 mortalities. Though the disease is regarded as a disorder of the more developed world, the occurrence is steadily increasing in many developing countries. Since chronic inflammation is a known aggravating risk factor for colorectal cancer, anti-inflammatory agents such as aspirin have been used to prevent the development of colorectal cancer and related mortality. The potential mechanisms for the effect of aspirin in the prevention of colorectal cancer have been proposed and broadly classified as cyclooxygenase (COX) dependent and COX-independent. Some of the primary effectors of COX-dependent mechanisms in carcinogenesis are likely to be prostaglandins. In contrast to the reversible action of other nonsteroidal anti-inflammatory drugs, aspirin is known to irreversibly inactivate COX enzymes to suppress production of prostaglandins. COX-independent mechanisms of anticancer effects of aspirin include down-regulation of nuclear factor kappa B activity and Akt activation, modulation of Bcl-2 and Bax family proteins, suppression of vascular endothelial growth factor, induction of apoptosis, disruption of DNA repair mechanisms, and induction of spermidine/spermine N1-acetyltransferase that modulates polyamine catabolism.

Abstract 656: High resolution metabolomics of aspirin treatment in colon tissue and adenoma risk: Results from a randomized clinical trial

Abstract Although substantial evidence from cell and animal studies, epidemiological studies, and clinical trials supports the chemopreventive effects of aspirin, especially for colorectal cancer, the molecular mechanisms are uncertain. Aspirin incorporates two bioactive components in one molecule, a reactive acetyl moiety and a salicylate group, and is well known for its pleotropic effects. Its best-characterized pharmacologic activity is the irreversible acetylation of the cyclooxygenases (COX-1 and COX-2). However, there is evidence for numerous COX-independent mechanisms that could also modify colorectal carcinogenesis. This study applied an untargeted, discovery-based approach (metabolomics) to elucidate the effects of aspirin on low molecular weight molecules in human colon tissue and assess their impact on colorectal carcinogenesis. We utilized normal mucosal tissue biopsies collected at colonoscopy after about 3 years of treatment from a sub-set of N=325 participants in the Aspirin/Folate Polyp Prevention Study, a randomized, placebo-controlled trial of aspirin (81 or 325 mg/day) for the prevention of colorectal adenomas. The global metabolic effects of aspirin were assessed using a high-resolution Thermo Fusion mass spectrometer coupled with dual chromatography and dual ionization (HILIC positive and C18 negative electrospray ionization). Multivariable linear regression was used to identify metabolic features associated with aspirin treatment adjusting for age, sex and race. Multivariable Poisson regression was used to assess associations with adenoma outcomes of metabolic features associated with aspirin treatment. Products of aspirin metabolism (salicylate, salicyluric acid) were identified and statistically significantly increased in post treatment compared to baseline plasma samples confirming aspirin treatment status. After quality control exclusions, N=4,879 and N=5,390 features were included in analyses from the C18 and HILIC columns, respectively, of which N=244 and N=222 were associated with aspirin treatment at a raw P<0.05. At the pathway level, aspirin treatment was most strongly associated with perturbations in prostaglandin/arachidonic acid metabolism and the carnitine shuttle, which is involved in energy metabolism from fatty acids. At the metabolite level, only one feature (tentatively identified as a pterosin compound) was statistically significantly associated with aspirin treatment after FDR adjustment, but it was not associated with adenoma risk. Top features associated with aspirin treatment that were associated with adenoma outcomes included creatinine but are mostly of unknown identity at this time. In conclusion, a non-targeted high-resolution metabolomics approach has the potential to identify key downstream effects of aspirin involved in colorectal chemoprevention. Citation Format: Elizabeth L. Barry, Karan Uppal, Chunyu Ma, Dean P. Jones, John A. Baron, Veronika Fedirko. High resolution metabolomics of aspirin treatment in colon tissue and adenoma risk: Results from a randomized clinical trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 656.

Inhibition of SIRT1 deacetylase and p53 activation uncouples the anti-inflammatory and chemopreventive actions of NSAIDs

BackgroundNonsteroidal anti-inflammatory drugs (NSAIDs) have been proposed as chemopreventive agents for many tumours; however, the mechanism responsible for their anti-neoplastic activity remains elusive and the side effects due to cyclooxygenase (COX) inhibition prevent this clinical application.MethodsMolecular biology, in silico, cellular and in vivo tools, including innovative in vivo imaging and classical biochemical assays, were applied to identify and characterise the COX-independent anti-cancer mechanism of NSAIDs.ResultsHere, we show that tumour-protective functions of NSAIDs and exisulind (a sulindac metabolite lacking anti-inflammatory activity) occur through a COX-independent mechanism. We demonstrate these NSAIDs counteract carcinogen-induced proliferation by inhibiting the sirtuin 1 (SIRT1) deacetylase activity, augmenting acetylation and activity of the tumour suppressor p53 and increasing the expression of the antiproliferative gene p21. These properties are shared by all NSAIDs except for ketoprofen lacking anti-cancer properties. The clinical interest of the mechanism identified is underlined by our finding that p53 is activated in mastectomy patients undergoing intraoperative ketorolac, a treatment associated with decreased relapse risk and increased survival.ConclusionOur study, for the first-time, links NSAID chemopreventive activity with direct SIRT1 inhibition and activation of the p53/p21 anti-oncogenic pathway, suggesting a novel strategy for the design of tumour-protective drugs.

Aspirin inhibits cancer stem cells properties and growth of glioblastoma multiforme through Rb1 pathway modulation.

Several clinical studies indicated that the daily use of aspirin or acetylsalicylic acid reduces the cancer risk via cyclooxygenases (Cox-1 and Cox-2) inhibition. In addition, aspirin-induced Cox-dependent and -independent antitumor effects have also been described. Here we report, for the first time, that aspirin treatment of human glioblastoma cancer (GBM) stem cells, a small population responsible for tumor progression and recurrence, is associated with reduced cell proliferation and motility. Aspirin did not interfere with cell viability but induced cell-cycle arrest. Exogenous prostaglandin E2 significantly increased cell proliferation but did not abrogate the aspirin-mediated growth inhibition, suggesting a Cox-independent mechanism. These effects appear to be mediated by the increase of p21 waf1 and p27 Kip1 , associated with a reduction of Cyclin D1 and Rb1 protein phosphorylation, and involve the downregulation of key molecules responsible for tumor development, that is, Notch1, Sox2, Stat3, and Survivin. Our results support a possible role of aspirin as adjunctive therapy in the clinical management of GBM patients.

Repurposing of Aspirin: Opportunities and Challenges

Repurposing refers to the study of drugs which is already approved to treat one disease for the treatment of other diseases. Aspirin (ASA) is widely used for the treatment of pain, fever or inflammation. It has been repurposed against cardiovascular disorders. In recent years ASA has generated a significant interest for prevention of cancer, central nervous system disorder, diabetes and cryptococcal infection due to its influence on COX dependent and COX independent mechanism. Following oral administration ASA is rapidly hydrolyzed to salicylic acid (SA). Since SA has selective ability to inhibit COX-2, its application for repurposing is limited. Besides, ASA is more potent due to its capacity for irreversible inhibition of COX. Thus for repurposing it is desirable to ensure adequate availability of ASA at site of action. This is a challenge as issues related to solubility, stability and toxicity needs to be simultaneously addressed while repurposing.

The value of aspirin in the prevention of colorectal cancer

Aspirin has demonstrated its efficacy in the prevention of adverse events related to cardiovascular disease, and lots of epidemiological studies suggest that it also can be used to prevent colorectal cancer and to decrease its recurrence after curative resection. The guideline published by USPSTF recently first time affirms the overall health benefit of aspirin in colorectal cancer prevention. Aspirin could prevent the colorectal cancer by the cyclooxygenase-dependent mechanisms and the COX-independent mechanisms. In this review, we will summarize some results of related researches and discuss the possible mechanisms. Key words: Colorectal neoplasms; Aspirin; Prevention

Prostate Cancer and Aspirin Use: Synopsis of the Proposed Molecular Mechanisms.

Background: Prostate cancer (PCa) is a critical health burden, impacting the morbidity and mortality of millions of men around the world. Most of the patients with PCa have their disease at first sensitive to androgen deprivation treatments, but later they develop resistance to therapy and eventually die of metastatic castration-resistant prostate cancer (CRPC). Although the newly developed anti-androgen therapies are effectively alleviating symptoms and prolonging lives of patients, there are still no curable treatments for CRPC. Recently, statistical studies have shown that the chronic use of aspirin might be significantly associated with better outcomes in PCa patients. Through this review, we aim to identify the different proposed molecular mechanisms relating aspirin to the pathobiology of PCa neoplasms, with a major focus on basic research done in this context.Methods: Articles were retrieved via online database searching of PubMed and MEDLINE between 1946 and September 2016. Keywords and combinations related to PCa and aspirin were used to perform the search. s of the articles were studied by two independent reviewers and then data extraction was performed on the relevant articles that met our review objectives.Results: Aspirin, a non-steroidal anti-inflammatory drug (NSAID), affects the proliferation, apoptosis, resistance and metastasis of PCa cell lines, through both COX-dependent and COX-independent mechanisms. It also lowers levels of the PCa diagnostic marker prostate specific antigen (PSA), suggesting that clinicians need to at least be aware if their patients are using Aspirin chronically.Conclusion: This review strongly warrants further consideration of the signaling cascades activated by aspirin, which may lead to new knowledge that might be applied to improve diagnosis, prognosis and treatment of PCa.

The role of aspirin in colorectal cancer chemoprevention.

Considerable interest has emerged over the last decade regarding the role of aspirin in prevention of colorectal cancer. This disease is one of the commonest cancers in the Western World, therefore, the existence of a simple "everyday" agent, which could have the ability to prevent the disease, represents an invaluable opportunity clinicians may be able to exploit. Evidence from case-control and cohort studies, and recent updates of randomised controlled trials have been very encouraging-indicating benefit from long term use of aspirin at low dose. Possible mechanisms of chemoprevention include inhibition of the cyclooxygenase (COX) pathway, or COX-independent mechanisms, for example, the PIK3CA pathway, or therapy-induced senescence of cancer cells. The most serious side effect of prolonged aspirin treatment is haemorrhage, especially from the GI tract. This is likely to be less of a problem with chemoprevention at lower doses. One also needs to consider the impact if aspirin resistance, an increasingly recognised clinical entity.

Small molecule tolfenamic acid and dietary spice curcumin treatment enhances antiproliferative effect in pancreatic cancer cells via suppressing Sp1, disrupting NF-kB translocation to nucleus and cell cycle phase distribution

Combination of dietary/herbal spice curcumin (Cur) and COX inhibitors has been tested for improving therapeutic efficacy in pancreatic cancer (PC). The objective of this study was to identify agent with low toxicity and COX-independent mechanism to induce PC cell growth inhibition when used along with Cur. Anticancer NSAID, tolfenamic acid (TA) and Cur combination were evaluated using PC cell lines. L3.6pl and MIA PaCa-2 cells were treated with Cur (5–25μM) or TA (25–100μM) or combination of Cur (7.5μM) and TA (50μM). Cell viability was measured at 24–72h posttreatment using CellTiter-Glo kit. While both agents showed a steady/consistent effect, Cur+TA caused higher growth inhibition. Antiproliferative effect was compared with COX inhibitors, Ibuprofen and Celebrex. Cardiotoxicity was assessed using cordiomyocytes (H9C2). The expression of Sp proteins, survivin and apoptotic markers (western blot), caspase 3/7 (caspase-Glo kit), Annexin-V staining (flow cytometry), reactive oxygen species (ROS) and cell cycle phase distribution (flow cytometry) was measured. Cells were treated with TNF-α, and NF-kB translocation from cytoplasm to nucleus was evaluated (immunofluorescence). When compared to individual agents, combination of Cur+TA caused significant increase in apoptotic markers, ROS levels and inhibited NF-kB translocation to nucleus. TA caused cell cycle arrest in G0/G1, and the combination treatment showed mostly DNA synthesis phase arrest. These results suggest that combination of Cur+TA is less toxic and effectively enhance the therapeutic efficacy in PC cells via COX-independent mechanisms.

Preserved Microvascular Endothelial Function in Young, Obese Adults with Functional Loss of Nitric Oxide Signaling

Data indicate endothelium-dependent dilation (EDD) may be preserved in the skeletal muscle microcirculation of young, obese adults. Preserved EDD might be mediated by compensatory mechanisms, impeding insight into preclinical vascular dysfunction. We aimed to determine the functional roles of nitric oxide synthase (NOS) and cyclooxygenase (COX) toward EDD in younger obese adults. We first hypothesized EDD would be preserved in young, obese adults. Further, we hypothesized a reduced contribution of NOS in young, obese adults would be replaced by increased COX signaling. Microvascular EDD was assessed with Doppler ultrasound and brachial artery infusion of acetylcholine (ACh) in younger (27 ± 1 year) obese (n = 29) and lean (n = 46) humans. Individual and combined contributions of NOS and COX were examined with intra-arterial infusions of l-NMMA and ketorolac, respectively. Vasodilation was quantified as an increase in forearm vascular conductance (ΔFVC). Arterial endothelial cell biopsies were analyzed for protein expression of endothelial nitric oxide synthase (eNOS). ΔFVC to ACh was similar between groups. After l-NMMA, ΔFVC to ACh was greater in obese adults (p < 0.05). There were no group differences in ΔFVC to ACh with ketorolac. With combined NOS-COX inhibition, ΔFVC was greater in obese adults at the intermediate dose of ACh. Surprisingly, arterial endothelial cell eNOS and phosphorylated eNOS were similar between groups. Younger obese adults exhibit preserved EDD and eNOS expression despite functional dissociation of NOS-mediated vasodilation and similar COX signaling. Compensatory NOS- and COX-independent vasodilatory mechanisms conceal reduced NOS contributions in otherwise healthy obese adults early in life, which may contribute to vascular dysfunction.

Suppression of β-catenin/TCF transcriptional activity and colon tumor cell growth by dual inhibition of PDE5 and 10.

Previous studies suggest the anti-inflammatory drug, sulindac inhibits tumorigenesis by a COX independent mechanism involving cGMP PDE inhibition. Here we report that the cGMP PDE isozymes, PDE5 and 10, are elevated in colon tumor cells compared with normal colonocytes, and that inhibitors and siRNAs can selectively suppress colon tumor cell growth. Combined treatment with inhibitors or dual knockdown suppresses tumor cell growth to a greater extent than inhibition from either isozyme alone. A novel sulindac derivative, ADT-094 was designed to lack COX-1/-2 inhibitory activity but have improved potency to inhibit PDE5 and 10. ADT-094 displayed >500 fold higher potency to inhibit colon tumor cell growth compared with sulindac by activating cGMP/PKG signaling to suppress proliferation and induce apoptosis. Combined inhibition of PDE5 and 10 by treatment with ADT-094, PDE isozyme-selective inhibitors, or by siRNA knockdown also suppresses β-catenin, TCF transcriptional activity, and the levels of downstream targets, cyclin D1 and survivin. These results suggest that dual inhibition of PDE5 and 10 represents novel strategy for developing potent and selective anticancer drugs.

COX-2 and EGFR: Partners in Crime Split by Aspirin

COX-2 and EGFR: Partners in Crime Split by Aspirin

Structural analysis of sulindac as an inhibitor of aldose reductase and AKR1B10

Aldose reductase (AR, AKR1B1) and AKR1B10 are enzymes implicated in important pathologies (diabetes and cancer) and therefore they have been proposed as suitable targets for drug development. Sulindac is the metabolic precursor of the potent non-steroidal anti-inflammatory drug (NSAID) sulindac sulfide, which suppresses prostaglandin production by inhibition of cyclooxygenases (COX). In addition, sulindac has been found to be one of the NSAIDs with higher antitumoral activity, presumably through COX inhibition. However, sulindac anticancer activity could be partially mediated through COX-independent mechanisms, including the participation of AR and AKR1B10. Previously, it had been shown that sulindac and sulindac sulfone were good AR inhibitors and the structure of the ternary complex with NADP+ and sulindac was described (PDB ID 3U2C). In this work, we determined the three-dimensional structure of AKR1B10 with sulindac and established structure–activity relationships (SAR) of sulindac and their derivatives with AR and AKR1B10. The difference in the IC50 values for sulindac between AR (0.36μM) and AKR1B10 (2.7μM) might be explained by the different positioning and stacking interaction given by Phe122/Phe123, and by the presence of two buried and ordered water molecules in AKR1B10 but not in AR. Moreover, SAR analysis shows that the substitution of the sulfinyl group is structurally allowed in sulindac derivatives. Hence, sulindac and its derivatives emerge as lead compounds for the design of more potent and selective AR and AKR1B10 inhibitors.

Abstract 814: Novel aspirin based selenium compounds as therapy against pancreatic cancer

Abstract Inflammation plays an important role in tumor growth promotion and has been linked to several human cancers. Evidences have demonstrated that inflammation has a high correlation with pancreatic cancer (PC) growth and promotion. An important mediator of inflammatory process is Cyclooxygenase 2 (COX-2) protein, which has been shown to be over-expressed in PC. Another major pathway which is crucial for inflammatory responses, cell proliferation, and resistance to apoptosis is the nuclear factor κB (NF-κB) pathway. It has been demonstrated that NF-κB pathway is responsible for drug resistance in PC. Aspirin, a well-known non-steroidal anti-inflammatory drug (NSAID), has been shown to have anti-cancer effects against PC. The mechanism of action of aspirin against PC is partially dependent on COX inhibition, as well as COX independent mechanism. Through extensive structure-activity relationship studies on Se-NSAID compounds, we have recently identified two Se-Aspirin compounds, ASD-43 and ASD-49, designed by incorporating selenium (Se) into salicylic acid structure, rendering it lethal to cancer cells. Previously, we had reported that Se-aspirin hybrid compounds were effective in different cancers, including PC. In this study we sought to evaluate the mechanism of action of these compounds in inhibiting the growth of PC. Since aspirin induces its effect partially by inhibiting COX, we hypothesized that these agents (ASD-43 and ASD-49) showed their anti-cancer activity by potently inhibiting COX activity as well as by modulating the inflammatory NF-κB pathway. Our studies, in Panc-1 cells, have shown that both ASD-43 and ASD-49 inhibited the degradation of IκB-alpha, which inhibits NF-κB from going into the nucleus, in presence of tumor necrosis factor (TNF). Upon treatment with ASD-43 and ASD-49, NF-κB regulated proteins like survivin and Bcl-xL expression was down regulated. Furthermore, a dose dependent induction of apoptosis in these cells was detected by activation of caspase-3 and PARP cleavage. Taken together, these results suggest that both ASD-43 and ASD-49 may represent potentially promising therapeutic agents for PC that induce apoptosis via the inhibition of COX and NF-κB pathways. Detailed investigations regarding the efficacy and mechanism of action of these compounds will be presented. Citation Format: Deepkamal N. Karelia, Manoj K. Pandey, Daniel Plano, Shantu Amin, Arun K. Sharma. Novel aspirin based selenium compounds as therapy against pancreatic cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 814. doi:10.1158/1538-7445.AM2014-814

A novel COX-independent mechanism of sulindac sulfide involves cleavage of epithelial cell adhesion molecule protein

Non-steroidal anti-inflammatory drugs (NSAIDs) are extensively used over the counter to treat headaches and inflammation as well as clinically to prevent cancer among high-risk groups. The inhibition of cyclooxygenase (COX) activity by NSAIDs plays a role in their anti-tumorigenic properties. NSAIDs also have COX-independent activity which is not fully understood. In this study, we report a novel COX-independent mechanism of sulindac sulfide (SS), which facilitates a previously uncharacterized cleavage of epithelial cell adhesion molecule (EpCAM) protein. EpCAM is a type I transmembrane glycoprotein that has been implemented as an over-expressed oncogene in many cancers including colon, breast, pancreas, and prostate. We found EpCAM to be down-regulated by SS in a manner that is independent of COX activity, transcription regulation, de novo protein synthesis, and proteasomal degradation pathway. Our findings clearly demonstrate that SS drives cleavage of the extracellular portion of EpCAM near the N-terminus. This SS driven cleavage is blocked by a deleting amino acids 55–81 as well as simply mutating arginine residues at positions 80 and 81 to alanine of EpCAM. Proteolysis of EpCAM by SS may provide a novel mechanism by which NSAIDs affect anti-tumorigenesis at the post-translational level.

Novel non-cyclooxygenase inhibitory derivatives of naproxen for colorectal cancer chemoprevention.

A structure-based medicinal chemistry strategy was applied to design new naproxen derivatives that show growth inhibitory activity against human colon tumor cells through a cyclooxygenase (COX)-independent mechanism. In vitro testing of the synthesized compounds against the human HT-29 colon tumor cell line revealed enhanced growth inhibitory activity compared to the parent naproxen with 3a showing IC50 of 11.4 μM (two orders of magnitude more potent than naproxen). Selectivity of 3a was investigated against a panel of three tumor and one normal colon cell lines and showed up to six times less toxicity against normal colonocytes. Compound 3a was shown to induce dose-dependent apoptosis of HT116 colon tumor cells as evidenced by measuring the activity of caspases-3 and 7. None of the synthesized compounds showed activity against COX-1 or COX-2 isozymes, confirming a COX-independent mechanism of action. Compound 3k was found to have no ulcerogenic effect in rats as indicated by electron microscope scanning of the stomach after oral administration. A pharmacophore model was developed for elucidating structure-activity relationships and subsequent chemical optimization for this series of compounds as colorectal cancer chemopreventive drugs.

NSAIDs Inhibit Tumorigenesis, but How?

Numerous epidemiologic studies have reported that the long-term use of nonsteroidal anti-inflammatory drugs (NSAID) is associated with a significant decrease in cancer incidence and delayed progression of malignant disease. The use of NSAIDs has also been linked with reduced risk from cancer-related mortality and distant metastasis. Certain prescription-strength NSAIDs, such as sulindac, have been shown to cause regression of precancerous lesions. Unfortunately, the extended use of NSAIDs for chemoprevention results in potentially fatal side effects related to their COX-inhibitory activity and suppression of prostaglandin synthesis. Although the basis for the tumor growth-inhibitory activity of NSAIDs likely involves multiple effects on tumor cells and their microenvironment, numerous investigators have concluded that the underlying mechanism is not completely explained by COX inhibition. It may therefore be possible to develop safer and more efficacious drugs by targeting such COX-independent mechanisms. NSAID derivatives or metabolites that lack COX-inhibitory activity, but retain or have improved anticancer activity, support this possibility. Experimental studies suggest that apoptosis induction and suppression of β-catenin-dependent transcription are important aspects of their antineoplastic activity. Studies show that the latter involves phosphodiesterase inhibition and the elevation of intracellular cyclic GMP levels. Here, we review the evidence for COX-independent mechanisms and discuss progress toward identifying alternative targets and developing NSAID derivatives that lack COX-inhibitory activity but have improved antineoplastic properties.