Downregulation of lactosylceramide synthases B4GALT5 or B4GALT6 alters glycosphingolipid profiles and enhances oxaliplatin sensitivity in colon cancer cells

Abstract

Glycosphingolipids (GSLs) represent an important group of bioactive lipids that serve both as structural components of membranes, and as signaling molecules involved in the regulation of numerous biological processes, such as control of cell growth and cell death. Their significant deregulation in colon cancer cells suggests their potential role in the disease progression. Lactosylceramide (LacCer) is a key intermediate in GSL metabolism and an important precursor of more complex GSLs. Here, we studied the impact of inhibition of LacCer synthesis (using selective targeting of specific LacCer synthases (LCS), B4GALT5 or B4GALT6, via CRISPR/Cas9-mediated gene knockdown) on control of the human DLD-1 colon adenocarcinoma cell proliferation, death and chemosensitivity. The downregulation of the selected enzymes, as verified by a decrease in respective mRNA and protein levels, significantly reduced levels of LacCers and several more complex GSLs (in particular, GM3 and GM1a gangliosides) in LCS-knockdown cells. Importantly, it enhanced the sensitivity of DLD-1 cells to the cytotoxic effects of oxaliplatin, a chemotherapy drug commonly used in colorectal cancer treatment. This was demonstrated by a general decrease in cell viability, an enhanced apoptotic cell death, caspase-8,-9,-3 cleavage/activation and cleavage of caspase substrates. We also observed a modulation of endoplasmic reticulum stress response, in particular decreased levels of ATF6, in LCS-knockdown cells treated with oxaliplatin. The present findings support the functional role of LCS in regulating the chemosensitivity of colon cancer cells towards the action of chemotherapy drugs, such as oxaliplatin, with possible further implications for the mechanisms underlying toxic action of platinum-based drugs in cancer cells.