N-stearoylethanolamine (NSE), a lipid mediator that belongs to the N-acylethanolamine (NAE) family, has anti-inflammatory, antioxidant and membranoprotective actions. In contrast to other NAEs, NSE does not interact with cannabinoid receptors. The exact mechanism of its action remains unclear. The aim of this study was to evaluate the action of NSE on activation, aggregation and adhesion of platelets that were chosen as a model of cellular response.Aggregation of platelets was measured to analyze the action of NSE (10 -6 -10 -10 M) on platelet reactivity. Changes in granularity and shape of resting platelets and platelets stimulated with ADP in the presence of NSE were monitored by flow-cytometry, platelet deganulation was monitored by spectrofluorimetry. In vivo studies were performed using obese insulin-resistant rats. Binding of fibrinogen to the GPIIb/IIIa receptor was estimated using indirect ELISA and scanning electron microscopy (SEM). It was found that NSE inhibits the activation and aggregation of human platelets. Our results suggest that NSE may decrease the activation and subsequent aggregation of platelets induced by ristocetin, epinephrine and low doses of ADP.NSE also reduced the binding of fibrinogen to GPIIb/IIIa on activated platelets. These effects could be explained by the inhibition of platelet activation mediated by integrin receptors: the GPIb-IX-V complex for ristocetin-induced activation and GPIIb/IIIa when epinephrine and low doses of ADP were applied. The anti-platelet effect of NSE complements its anti-inflammatory effect and allows us to prioritize studies of NSE as a potent anti-thrombotic agent. Significance StatementN-stearoylethanolamine (NSE) was shown to possess inhibitory action on platelet activation, adhesion and aggregation. The mechanism of inhibition possibly involves integrin receptors. This finding complements the known anti-inflammatory effects of NSE.

This study reports a dose-dependent pro-apoptotic action of synthetic cannabimimetic N-stearoylethanolamine (NSE) on diverse cancer cell lines, including multidrug-resistant models. No antioxidant or cytoprotective effects of NSE were found when it was applied together with doxorubicin. A complex of NSE with the polymeric carrier poly(5-(tert-butylperoxy)-5-methyl-1-hexen-3-yn-co-glycidyl methacrylate)-graft-PEG was synthesized. Co-immobilization of NSE and doxorubicin on this carrier led to a 2-10-fold enhancement of the anticancer activity, particularly, against drug-resistant cells overexpressing ABCC1 and ABCB1. This effect might be caused by accelerated nuclear accumulation of doxorubicin in cancer cells, which led to the activation of the caspase cascade, revealed by Western blot analysis. The NSE-containing polymeric carrier was also able to significantly enhance the therapeutic activity of doxorubicin in mice with implanted NK/Ly lymphoma or L1210 leukemia, leading to the complete eradication of these malignancies. Simultaneously, loading to the carrier prevented doxorubicin-induced elevation of AST and ALT as well as leukopenia in healthy Balb/c mice. Thus, a unique bi-functionality of the novel pharmaceutical formulation of NSE was revealed. It enhanced doxorubicin-induced apoptosis in cancer cells in vitro and promoted its anticancer activity against lymphoma and leukemia models in vivo. Simultaneously, it was very well tolerated preventing frequently observed doxorubicin-associated adverse effects.