C22:0- and C24:0-dihydroceramides Confer Mixed Cytotoxicity in T-Cell Acute Lymphoblastic Leukemia Cell Lines

Abstract

We previously reported that fenretinide (4-HPR) was cytotoxic to acute lymphoblastic leukemia (ALL) cell lines in vitro in association with increased levels of de novo synthesized dihydroceramides, the immediate precursors of ceramides. However, the cytotoxic potentials of native dihydroceramides have not been defined. Therefore, we determined the cytotoxic effects of increasing dihydroceramide levels via de novo synthesis in T-cell ALL cell lines and whether such cytotoxicity was dependent on an absolute increase in total dihydroceramide mass versus an increase of certain specific dihydroceramides. A novel method employing supplementation of individual fatty acids, sphinganine, and the dihydroceramide desaturase-1 (DES) inhibitor, GT-11, was used to increase de novo dihydroceramide synthesis and absolute levels of specific dihydroceramides and ceramides. Sphingolipidomic analyses of four T-cell ALL cell lines revealed strong positive correlations between cytotoxicity and levels of C22:0-dihydroceramide (ρ = 0.74–0.81, P ≤ 0.04) and C24:0-dihydroceramide (ρ = 0.84–0.90, P ≤ 0.004), but not between total or other individual dihydroceramides, ceramides, or sphingoid bases or phosphorylated derivatives. Selective increase of C22:0- and C24:0-dihydroceramide increased level and flux of autophagy marker, LC3B-II, and increased DNA fragmentation (TUNEL assay) in the absence of an increase of reactive oxygen species; pan-caspase inhibition blocked DNA fragmentation but not cell death. C22:0-fatty acid supplemented to 4-HPR treated cells further increased C22:0-dihydroceramide levels (P ≤ 0.001) and cytotoxicity (P ≤ 0.001). These data demonstrate that increases of specific dihydroceramides are cytotoxic to T-cell ALL cells by a caspase-independent, mixed cell death mechanism associated with increased autophagy and suggest that dihydroceramides may contribute to 4-HPR-induced cytotoxicity. The targeted increase of specific acyl chain dihydroceramides may constitute a novel anticancer approach.

A prospective analysis of circulating saturated and monounsaturated fatty acids and risk of non‐Hodgkin lymphoma

“…We also found an inverse association between VLCSFA and T‐NHL. An in vitro study reported that certain acyl fatty acids, such as 22:0 and 24:0, but not shorter chain acyl fatty acids, are cytotoxic to T cell acute lymphoblastic leukemia cell lines . However, it remains unknown whether a similar effect exists in vivo or would confer protection against developing T‐NHL.…”

Ceramide induced mitophagy and tumor suppression

“…On the other hand, C 16 -ceramide promotes head and neck cancer cell proliferation and its increased serum levels associate with a positive lymph node status in breast cancer patients [38, 39]. On the other hand, there are studies showing that C 16 -ceramide is pro-apoptotic, whereas C 24 -ceramide protects from cell death [40, 41]. Thus, overall, distinct roles of ceramides appear to be context dependent, especially that knockout mice for multiple CerS enzymes exhibit different phenotypes: mice that express a mutant CerS1 (toppler mice) exhibit neurological disorders associated mainly with alterations in Purkinjee cells; mice with genetic loss of CerS2 results in liver damage; mice with CerS6 knockout exhibit neurological/behavioral alterations; and CerS4 knockout mice exhibit severe alopecia with alterations in sebaceous glands and sebum contents [42-43,204].…”

Inhibition of dihydroceramide desaturase activity by the sphingosine kinase inhibitor SKI II

“…Blockade of Des1 produces an increase in dihydroceramides (dhCers), which have emerged as bioactive lipids and the target of several drugs ( 34 ), including fenretinide. Several studies have shown that cells respond to fenretinide treatment with a robust production of dhCers (35)(36)(37)(38)(39)(40)(41)(42)(43)(44), and that this increase induces autophagy ( 45,46 ). Furthermore, experimental evidence exists on the connection between resistance to fenretinide and increased S1P production ( 38,44 ) due to an increased SK activity and SK1 mRNA and protein levels ( 38 ).…”