CerS2 Overexpression Enhances Palmitoyl‐ CoA –Induced Cytotoxicity and Alters Ceramide Species Composition in Breast Cancer Cells

Sphingolipid metabolism, particularly the balance between pro‐apoptotic ceramide and pro‐survival sphingosine‐1‐phosphate (S1P), has been associated with breast cancer progression. The aim of the present study was to examine whether ceramide synthase 2 (CerS2) overexpression is associated with changes in increased palmitoyl‐CoA (PCA) based ceramide buildup and cell fate changes in MCF‐7 breast cancer cells, a hormone receptor–positive model. CERS2 plasmid transfected cells were treated with PCA and/or the CerS inhibitor, fumonisin B1 (FB1). Cell viability, the proliferation marker proliferating cell nuclear antigen (PCNA), apoptosis (TUNEL and cleaved caspase‐3), and sphingolipid profiles were evaluated. High‐dose PCA (100 μM) significantly reduced MTT signal and PCNA expression and increased apoptotic markers, with stronger effects in CerS2‐overexpressing cells. Across short‐ and longer‐term exposures, the response pattern was concentration‐ and time‐responsive but not uniformly monotonic. Lipidomic analysis revealed that PCA and CerS2 overexpression increased C16 ceramide and very‐long‐chain ceramides (C22–C24), respectively. FB1 decreased the level of ceramide, elevated S1P and partly counteracted PCA‐induced cytotoxicity. FB1 pre‐treatment which was applied sequentially restricted but did not eliminate apoptosis. These findings indicate that CerS2 overexpression reshapes the cellular response to substrate‐driven sphingolipid stress by altering acyl‐chain–specific ceramide composition and the balance between ceramide‐ and S1P‐associated signaling. Rather than reflecting isolated pathway effects, the results support a context‐dependent role for CerS2 in modulating apoptotic sensitivity in MCF‐7 breast cancer cells.