Abstract 2027: Tamoxifen amplifies antitumor impact of nanoformulated ceramide- pan efficacy in a myriad of tumor cell types.
Authors: Morad Samy A.F. , Levin Jonathan C. , Abdelmageed Noha , Kester Mark , Shanmugavdlandy Sriram S. , Paulson Robert F. , Rosenberg Daniel W. , Madigan James P. , Cabot Myles C. 
Published: August 2014

Abstract
Although ceramide has potent tumor suppressor properties, its use in cancer therapeutics has been limited; this is mainly due to rapid metabolism of ceramide by cancer cells, the inability to attain therapeutic levels when ceramide-generating agents are employed, and the inherent insolubility of ceramide as a therapeutic. Our approach circumvents these disadvantages and provides advantages over existing methodologies. For example, rather than promoting intracellular ceramide generation by application of exogenous drugs that activate de novo and sphingomyelinase pathways, we have chosen to administer ceramide exogenously in the form of short-chain C6-ceramide nanoliposomes; this overcomes solubility issues and improves uptake. In addition, nanoliposomes are efficient platforms for delivering other hydrophobic chemotherapeutics. The antiestrogen tamoxifen was selected for our studies because of its unusual “off-target” actions, which include inhibition of ceramide metabolism at glycosylation and hydrolysis. Blocking these junctures increases C6-ceramide intracellular residence time and diminishes production of sphingosine 1-phosphate, a mitogenic lipid derived via the action of ceramidase and sphingosine kinase. Hence, C6-ceramide and tamoxifen make up a versatile drug duo. Using in vitro models of human colon and breast cancer, acute mylogenous leukemia (AML), and melanoma, we show that simultaneous administration of nanoliposomal C6-ceramide and tamoxifen potently and synergistically decreased cell viability, compared to the single agents. The mix induced caspase-dependent apoptosis, cell cycle arrest at G1 and G2, upregulation of JNK, p38, and p53, and downregulation of Akt and survivin. In triple negative, hormone-insensitive breast cancer cells, C6-ceramide-tamoxifen induced cell cycle arrest, caspase-dependent apoptosis, and lysosomal and mitochondrial membrane permeability. Evaluation of single agents showed that tamoxifen elicited lysosomal membrane permeability and inhibited acid ceramidase. We have obtained similar results in in vitro models of human melanoma and AML. In an in vivo retroviral transduced murine AML model, the combination lowered leukemia burden in spleen and marrow. The combination also perpetuated synergistic increases in long-chain ceramides, likely compounding the overall ceramide effect. Of interest regarding in vivo utility, the major metabolites of tamoxifen, N-desmethyltamoxifen and 4-hydroxytamoxifen, were effective in combination with C6-ceramide. Our findings show that tamoxifen magnifies the antiproliferative effects of C6-ceramide via aggregate mechanisms. This proficient, ceramide-based nanoliposomal drug regimen may have potential as an effective anticancer therapeutic.
Supported by NIGMS 77391; ABC's, Los Angeles; Fashion Footwear Association of New York Charitable Foundation.
Citation Format: Samy A.F. Morad, Jonathan C. Levin, Noha Abdelmageed, Mark Kester, Sriram S. Shanmugavdlandy, Robert F. Paulson, Daniel W. Rosenberg, James P. Madigan, Myles C. Cabot. Tamoxifen amplifies antitumor impact of nanoformulated ceramide- pan efficacy in a myriad of tumor cell types. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2027. doi:10.1158/1538-7445.AM2013-2027