We investigated whether platelets prime colon cancer cells for metastasis and whether pharmacological inhibition of platelet function may prevent it. Coculturing HT29 human colon carcinoma cells with human platelets led to the induction of mesenchymal-like cancer cells characterized by downregulation of E-cadherin and upregulation of Twist1, enhanced cell mobility and a proaggregatory action on platelets. These changes were prevented by different antiplatelet agents, aspirin[an inhibitor of cyclooxygenase(COX)-1], DG-041[an antagonist of prostaglandin(PG)E2 EP3 receptor] and ticagrelor (a P2Y12 receptor antagonist). The injection of HT29 cells, exposed to platelets in vitro, into the tail vein of humanized immunodeficient mice led to higher incidence of lung metastasis compared to the injection of untreated HT29 cells. This effect was associated with enhanced systemic biosynthesis of thromboxane(TX)A2 and PGE2 in vivo. Platelet COX-1 inhibition by aspirin administration to mice prevented the increased rate of metastasis as well as the enhanced production of TXA2 and PGE2 induced by the in vitro priming of HT29 cells by platelets. In conclusion, targeting platelet COX-1 with low-dose aspirin exerts an antimetastatic action by averting the stem cell mimicry of cancer cells associated with enhanced proaggregatory effects induced by platelet-tumor cell interactions. These effects may be shared by other antiplatelet drugs.

Aspirin taken for several years at doses of at least 75 mg daily reduced long-term incidence and mortality due to colorectal cancer. The finding of aspirin benefit at low-doses given once daily, used for cardioprevention, locates the antiplatelet effect of aspirin at the center of its antitumor efficacy. In fact, at low-doses, aspirin acts mainly by an irreversible inactivation of platelet cyclooxygenase (COX)-1 in the presystemic circulation, which translates into a long-lasting inhibition of platelet function. Given the short half-life of aspirin in the human circulation(approximately 20 min) and the capacity of nucleated cells to resynthesize the acetylated COX-isozyme(s), it seems unlikely that a nucleated cell could be the target of aspirin chemoprevention. These findings convincingly suggest that colorectal cancer and atherothrombosis may share a common mechanism of disease, i.e. platelet activation in response to epithelial(in tumorigenesis) and endothelial(in tumorigenesis and atherothrombosis) injury. Activated platelets may also enhance the metastatic potential of cancer cells (through a direct interaction and/or the release of soluble mediators or exosomes) at least in part by inducing the overexpression of COX-2. COX-independent mechanisms of aspirin, such as the inhibition of NF-kB signaling and Wnt/β-catenin signaling and the acetylation of extra-COX proteins, have been suggested to play a role in its chemopreventive effects. However, their relevance remains to be demonstrated in vivo at clinical doses.

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Conventional medical treatment for rheumatoid arthritis and osteoarthritis includes the use of NSAIDs (traditional and selective inhibitors of cyclooxygenase [COX]-2), because they provide unmistakable and significant health benefits in the treatment of pain and inflammation. However, they are associated with an increased risk of serious gastrointestinal (GI) and cardiovascular (CV) adverse events. Both beneficial and adverse effects are due to the same mechanism of action, which is inhibition of COX-dependent prostanoids. Since CV and GI risk are related to drug exposure, a reduction in the administered dose is recommended. However, this strategy will not eliminate the hazard owing to a possible contribution of individual genetic background. Further studies will be necessary to develop genetic and/or biochemical markers predictive of the CV and GI risk of NSAIDs.

The cyclooxygenase-2 (COX-2) enzyme catalyzes the rate-limiting step of prostaglandin formation in pathogenic states and a large amount of evidence has demonstrated constitutive COX-2 expression to be a contributing factor promoting colorectal cancer (CRC). Various genetic, epigenetic, and inflammatory pathways have been identified to be involved in the etiology and development of CRC. Alteration in these pathways can influence COX-2 expression at multiple stages of colon carcinogenesis allowing for elevated prostanoid biosynthesis to occur in the tumor microenvironment. In normal cells, COX-2 expression levels are potently regulated at the post-transcriptional level through various RNA sequence elements present within the mRNA 3′-untranslated region(3′UTR). A conserved AU-rich element(ARE) functions to target COX-2 mRNA for rapid decay and translational inhibition through association with various RNA-binding proteins to influence the fate of COX-2 mRNA. Specific microRNAs bind regions within the COX-2 3′UTR and control COX-2 expression. In this chapter, we discuss novel insights in the mechanisms of altered posttranscriptional regulation of COX-2 in CRC and how this knowledge may be used to develop novel strategies for cancer prevention and treatment.

Non-steroidal anti-inflammatory drugs (NSAIDs) are a chemically heterogeneous group of compounds that provide unmistakable and significant health benefits in the treatment of pain and inflammation. They include traditional NSAIDs (tNSAIDs), which act by inhibiting both cyclooxygenase (COX)-1 and COX-2 and selective COX-2 inhibitors (coxibs). The development of biomarkers predictive of the impact of NSAIDs on COX-1 and COX-2 activities in vitro, ex vivo and in vivo has been essential to read out the clinical consequences of selective and non-selective inhibition of COX isozymes in human beings. The analgesic and anti-inflammatory effects of NSAIDs are COX-2-dependent effects, unrelated to COX-2 selectivity. The intensity and duration of these effects are influenced by dose and half-life of the NSAID. However, the inhibition of COX-1 in cells of the gastrointestinal (GI) system and COX-2 in vascular cells translates into increased risk of serious GI adverse events and atherothrombosis and hypertension, respectively. The COX-2 selectivity of NSAIDs can predict, at least in part, the GI toxicity. In contrast, the CV effects are largely COX-2-dependent effects, unrelated to COX-2 selectivity but are dose dependent. The reduction in the dose is recommended and presumably will limit the number of patients exposed to a CV or a GI hazard by NSAIDs and coxibs. It will not, however, eliminate the risk on an individual level because there is a marked variability in how different people react to these drugs, based on their genetic background. The challenge of the next future will be to develop biomarkers useful to identify the individuals who react abnormally to COX inhibition.

3-(5-Chlorofuran-2-yl)-5-methyl-4-phenylisoxazole (P6), a known selective cyclooxygenase-1 (COX-1) inhibitor, was used to design a new series of 3,4-diarylisoxazoles in order to improve its biochemical COX-1 selectivity and antiplatelet efficacy. Structure-activity relationships were studied using human whole blood assays for COX-1 and COX-2 inhibition in vitro, and results showed that the simultaneous presence of 5-methyl (or -CF3), 4-phenyl, and 5-chloro(-bromo or -methyl)furan-2-yl groups on the isoxazole core was essential for their selectivity toward COX-1. 3g, 3s, 3d were potent and selective COX-1 inhibitors that affected platelet aggregation in vitro through the inhibition of COX-1-dependent thromboxane (TX) A2. Moreover, we characterized their kinetics of COX-1 inhibition. 3g, 3s, and 3d were more potent inhibitors of platelet COX-1 and aggregation than P6 (named 6) for their tighter binding to the enzyme. The pharmacological results were supported by docking simulations. The oral administration of 3d to mice translated into preferential inhibition of platelet-derived TXA2 over protective vascular-derived prostacyclin (PGI2).

The design of compounds that are able to inhibit cyclooxygenase (COX) and to release nitric oxide (NO) should give rise to drugs endowed with an overall safer profile for the gastrointestinal and cardiovascular systems. Herein we report a new class of pyrrole-derived nitrooxy esters (11a-j), cyclooxygenase-2 (COX-2) selective inhibitors endowed with NO releasing properties, with the goal of generating new molecules able to both strongly inhibit this isoform and reduce the related adverse side effects. Taking into account the metabolic conversion of nitrooxy esters into corresponding alcohols, we also studied derivatives 12a-j. All compounds proved to be very potent and selective COX-2 inhibitors; nitrooxy derivatives displayed interesting ex vivo NO-dependent vasorelaxing properties. Compounds 11c, 11d, 12c, and 12d were selected for further in vivo studies that highlited good anti-inflammatory and antinociceptive activities. Finally, two selected compounds (11c and 12c) tested in human whole blood (HWB) assay proved to be preferential inhibitors of COX-2.