Small molecule targeting the Rac1-NOX2 interaction prevents collagen-related peptide and thrombin-induced reactive oxygen species generation and platelet activation

Autor: Xin Duan, S. Saleem, Wolfgang Bergmeier, Ashley Kuenzi Davis, Yi Zheng, Huzoor Akbar, N. N. Tandon, Raymond Piatt
Rok vydání: 2018
Předmět:
Zdroj: Journal of thrombosis and haemostasis : JTH. 16(10)
ISSN: 1538-7836
Popis: Superoxide anion (O2−) and its derivatives, collectively called reactive oxygen species (ROS), are generated in platelets and contribute to signaling events involved in platelet activation [1–4]. The exact mechanisms of ROS generation and signaling mechanisms involved in ROS-mediated platelet activation are still being elucidated. Although diverse biochemical reactions contribute to ROS generation in platelets, nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) oxidases (NOX) have emerged as critical sources of agonist-induced ROS generation. Six homologs of the cytochrome subunit of phagocyte NADPH oxidase (also known as NOX2 or gp91phox), namely NOX1, NOX3, NOX4, NOX5, DUOX1 and DUOX2, have been characterized in various cells [5]. Human platelets appear to possess NOX1 [6], NOX2 [6,7], NOX4 [8] and NOX5 [9]. Platelets also contain cytosolic cofactors, namely p22phox, p67phox [10] and p47phox, and Rac GTPase [11,12]. NOX2 is constitutively associated with p22phox and its activation requires translocation of p47phox, p67phox and p40phox as well as RacGTP to the NOX2/p22phox complex. The assembled NOX2 complex generates superoxide by transferring an electron from NADPH in the cytosol to oxygen in the extracellular space. With the emergent role of NOX in platelet activation [13–15], it has been suggested that NOX2 may be a novel target for antithrombotic treatment [3,8]. A direct link between NOX2 activity, ROS generation and platelet activation has been shown in patients with X-linked chronic granulomatous disease (X-CGD). These patients are genetically deficient in NOX2 and exhibit diminished ROS generation and CD40 ligand expression in response to collagen or thrombin [12]. Moreover, plasma levels of soluble CD40 ligand and soluble P-selectin, two markers of in vivo platelet activation, are significantly diminished in X-CGD patients [16]. Recently, the role of NOX2 in platelet activation and thrombosis has also been reported using mice genetically deficient in NOX2 [8]. Rac1 GTPase plays a critical role in regulation of both NOX2 [17–19] and NOX1 [20,21]. The binding of Rac1-GTP to p67phox facilitates its binding to NOX2 and its activation [18], whereas binding of Rac1-GTP to NOXA1 enhances its binding to NOX1 and its activation. We have shown earlier that a rationally designed small molecule inhibitor of Rac1–p67phox interaction (Phox-I) inhibits ROS generation [22]. In this study, we tested our hypothesis that if Rac1–p67phox complex formation is critical in ROS generation, then interrupting this complex formation by Phox-I should inhibit ROS generation and consequently platelet activation. Here we report that Phox-I, by inhibiting Rac1–p67phox interaction in platelets, prevents GPVI- and non-GPVI-dependent NOX2 activation and the consequent ROS generation, as well as the in vivo and in vitro platelet activation without affecting the hemostatic response, presenting small molecule targeting of the Rac1-NOX2 interaction as a useful antithrombosis regimen.
Databáze: OpenAIRE
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