An estrogen analogue and promising anticancer agent refrains from inducing morphological damage and reactive oxygen species generation in erythrocytes, fibrin and platelets: a pilot study

Cancer Cell International, Jun 2014

Background 2-Methoxyestradiol is known to have antitumour and antiproliferative action in vitro and in vivo. However, when 2-methoxyestradiol is orally administered, it is rapidly oxidized by the enzyme 17β-hydroxysteriod dehydrogenase in the gastrointestinal tract. Therefore, 2-methoxyestradiol never reaches high enough concentrations in the tissue to be able to exert these antitumour properties. This resulted in the in silico-design of 2-methoxyestradiol analogues in collaboration with the Bioinformatics and Computational Biology Unit (UP) and subsequent synthesis by iThemba Pharmaceuticals (Pty) Ltd (Modderfontein, Midrand, South Africa). One such a novelty-designed analogue is 2-ethyl-3-O-sulphamoyl-estra-1, 3, 5(10)16-tetraene (ESE-16). Methods This pilot study aimed to determine the morphological effect and possible generation of reactive oxygen species by ESE-16 on erythrocytes and platelet samples (with and without added thrombin) by means of scanning electron microscopy, transmission electron microscopy and flow cytometry. Results Erythrocytes and platelets were exposed to ESE-16 at a concentration of 180nM for 24 hours. Scanning- and transmission electron microscopy indicated that ESE-16 did not cause changes to erythrocytes, platelets or fibrin networks. Flow cytometry measurements of hydrogen peroxide and superoxide indicated that ESE-16 does not cause an increase in the generation of reactive oxygen species in these blood samples. Conclusion Further in vivo research is warranted to determine whether this novel in silico-designed analogue may impact on development of future chemotherapeutic agents and whether it could be considered as an antitumour agent.

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An estrogen analogue and promising anticancer agent refrains from inducing morphological damage and reactive oxygen species generation in erythrocytes, fibrin and platelets: a pilot study

Cancer Cell International An estrogen analogue and promising anticancer agent refrains from inducing morphological damage and reactive oxygen species generation in erythrocytes, fibrin and platelets: a pilot study Lisa Repsold 0 Etheresia Pretorius 0 Annie Margaretha Joubert 0 0 Department of Physiology, School of Medicine, Faculty of Health Sciences, University of Pretoria , Pretoria , South Africa Background: 2-Methoxyestradiol is known to have antitumour and antiproliferative action in vitro and in vivo. However, when 2-methoxyestradiol is orally administered, it is rapidly oxidized by the enzyme 17β-hydroxysteriod dehydrogenase in the gastrointestinal tract. Therefore, 2-methoxyestradiol never reaches high enough concentrations in the tissue to be able to exert these antitumour properties. This resulted in the in silico-design of 2-methoxyestradiol analogues in collaboration with the Bioinformatics and Computational Biology Unit (UP) and subsequent synthesis by iThemba Pharmaceuticals (Pty) Ltd (Modderfontein, Midrand, South Africa). One such a novelty-designed analogue is 2-ethyl-3-O-sulphamoyl-estra-1, 3, 5(10)16-tetraene (ESE-16). Methods: This pilot study aimed to determine the morphological effect and possible generation of reactive oxygen species by ESE-16 on erythrocytes and platelet samples (with and without added thrombin) by means of scanning electron microscopy, transmission electron microscopy and flow cytometry. Results: Erythrocytes and platelets were exposed to ESE-16 at a concentration of 180nM for 24 hours. Scanning- and transmission electron microscopy indicated that ESE-16 did not cause changes to erythrocytes, platelets or fibrin networks. Flow cytometry measurements of hydrogen peroxide and superoxide indicated that ESE-16 does not cause an increase in the generation of reactive oxygen species in these blood samples. Conclusion: Further in vivo research is warranted to determine whether this novel in silico-designed analogue may impact on development of future chemotherapeutic agents and whether it could be considered as an antitumour agent. 2-Methoxyestradiol analogues; 2-Ethyl-3-O-sulphamoyl-estra-1; 3; 5(10)16-tetraene; Reactive oxygen species; Cancer Introduction Cancer is currently one of the leading causes of mortality across the world’s population [ 1 ]. Research aimed at finding possible compounds to treat or cure cancer is therefore of extreme importance. 2-Methoxyestradiol (2ME) (Figure 1) is a metabolite of 17β-estradiol which has been tested for its antitumour and antiangiogenic properties in vitro and in vivo [ 2,3 ]. 2ME however, has poor bioavailability limiting the action in vivo as indicated in earlier phase I studies where 2ME was orally administered to cancer patients in the form of a capsule [ 5 ]. The rate at which 2ME becomes available to the site of drug action is low since the rate of oxidation of 2ME is higher than absorption [ 5 ]. 2ME is rapidly oxidized in the gastrointestinal tract when orally administered by the enzyme 17β-hydroxysteriod dehydrogenase [ 6,7 ]. This leads to decreased levels of 2ME, thus being too low to exert significant antitumour effects [8]. The development of a 2ME NanoCrystal® colloidal dispersion (NCD) was subsequently formulated. The latter increased bioavailability, however, the antitumour effects were not optimal in patients [ 9 ]. This led to the development and synthesis of analogues of 2ME to test for a compound with increased antitumour and antiangiogenic properties with increased bioavailability [ 10,11 ]. Such analogues of 2ME have been in silicodesigned in our laboratory (Figure 2). One novel in silicodesigned analogue is 2-ethyl-3-O-sulphamoyl-estra-1, 3, 5 (10)16-tetraene (ESE-16) (Figure 2). 2ME exerts its anticancer signaling by disrupting mitochondrial function, generating reactive oxygen species (ROS) and by targeting microtubule dynamics in vitro and in vivo [ 13 ]. 2ME results in apoptosis involving both the extrinsic and intrinsic pathways [ 14 ]. 2ME activates apoptosis via the extrinsic pathway by means of death receptors (DRs) such as death receptor 5 (DR5) and activates caspases known to promote the extrinsic pathway [ 14 ]. The extrinsic pathway of apoptosis comprises of the instigation of caspase 8, caspase 3, and upregulation of DR5 with activation of caspase 9 present in the intrinsic pathway through crosstalk with caspase 8 [ 14,15 ]. Production of ROS in 2ME- and analogue-treated cells is likely to be a result of the inhibition of the mitochondrial electron transport complex I [ 13 ] leading to autophagy by means of beclin-1 upregulation and autophagy protein 4 (Atg4) inactivation [ 16 ]. Generation of ROS upregulates beclin-1 which results in the increased incidence of autophagy, while hydrogen peroxide causes autophagy and the inactivation of Atg4 responsible for the induction of autophagy [ 17 ]. 2ME also induces apoptosis in cancer cells as a result of the upreg (...truncated)


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Lisa Repsold, Etheresia Pretorius, Annie Joubert. An estrogen analogue and promising anticancer agent refrains from inducing morphological damage and reactive oxygen species generation in erythrocytes, fibrin and platelets: a pilot study, Cancer Cell International, 2014, pp. 48, 14, DOI: 10.1186/1475-2867-14-48