Abstract 3375: Activation of the acid sphingomyelinase pathway as a potential mechanism of chemotherapy-induced normal tissue vasculature damage

Abstract

Abstract Background: Several classes of chemotherapy confer substantial risk for late-term vascular morbidity. We aimed to investigate the mechanisms of chemotherapy-induced vascular toxicity in normal tissues. Methods: The effect of Doxorubicin and Cisplatin has been studied in-vitro using bovine aortic endothelial cells (BAEC) as a model of normal tissue vascular endothelium. Specifically, we investigated activation of the acid sphingomyelinase (ASMase) pathway, which may lead to endothelial dysfunction (vasoconstriction) and apoptosis following incubation with different concentrations of Doxorubicin and Cisplatin. ASMase activity was measured using radioenzymatic assay with [N-methyl-14C] sphingomyelin. Cellular Reactive Oxygen Species (ROS) production was assessed using the 2’,7’-dichlorofluorescin diacetate (DCFDA) detection assay kit (Abcam®). Endothelial apoptosis was quantified using the 3,5-bis-benzamide assay (Sigma-Aldrich®). For the in-vivo studies, mouse femoral arterial blood flow was measured in real-time during and after Doxorubicin (8 mg/kg intravenously) or Cisplatin (8 mg/kg) administration. Visualization of ovarian and femoral microvasculature was obtained by fluorescence optical imaging system, equipped with a confocal fiber microscope (Cell-viZio). Results: Incubation of BAEC with Doxorubicin (0.25 μM or 0.50 μM) resulted in increased ASMase activity, production of ROS and induction of apoptosis in a dose-dependent fashion. Cisplatin (25 μM or 50 μM) generated significantly reduced effects in BAEC yet apoptosis and ROS were notable. In-vivo endoscopic fibered confocal microscopy following chemotherapy administration revealed a differential pattern between the two agents. While post Doxorubicin injection, there was a marked reduction in fluorescent signal in small vessels (<15 μM) indicating vessel constriction and disintegration of vessel's wall in larger vessels, Cisplatin treatment resulted in very minor vascular changes. Conclusions: These findings suggest that chemotherapy-induced vascular damage in normal tissues may represent different modes of toxicity in vivo and in vitro, while is mediated via activation of the ASMase pathway in response to Doxorubicin and to a lesser extent in response to Cisplatin. Citation Format: Aviram Mizrachi. Activation of the acid sphingomyelinase pathway as a potential mechanism of chemotherapy-induced normal tissue vasculature damage. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3375.