Objective
Malignant cells can adapt their lipid metabolism to attain higher rates of proliferation, and resist chemotherapy-induced apoptosis. The aim of our study was to first identify the changes in lipid metabolism in multiple myeloma (MM) patients, and to further investigate the role of this altered metabolism in vitro. 

Methods
We performed a lipidomics analysis to compare plasma from MM patients to healthy controls. Based on these results, lipid spike-in assays enabled us to evaluate effects on MM cell  proliferation. Sphingomyelinase (SMase) content of MM cells was measured, before and after treatment with standard-of-care drugs, using an AmplexRed assay. We inhibited both neutral and acid SMase in MM cells to evaluate effects on drug efficacy. To eliminate possible off-target effects, our findings were confirmed by silencing SMPD1 expression. 

Results
Lipidomics analysis revealed that several ceramide species were significantly more present in MM patient plasma, while sphingomyelin seemed to be decreased. When adding ceramide in vitro to MM cells, we observed higher viability and proliferation. The increased ceramide/sphingomyelin ratio in plasma lead us to believe that the sphingomyelinase enzyme, which converts sphingomyelin into ceramide, was upregulated in MM cells. This was confirmed in BM samples, comparing CD138+ MM cells to the CD138 negative fraction.  In the MM cell lines JJN3, OPM2, LP1 and U266, we observed an increased expression of acid and total SMase when treating these cells with melphalan or bortezomib for 24hInterestingly, amitryptiline, an inhibitor of acid SMase, often prescribed in MM patients for neuropathic pain, increased the efficacy of both drugs in vitro, and also induced apoptosis in primary MM cells as a single agent. 

Conclusion
SMase is increased in primary MM cells and cell lines upon treatment with melphalan and bortezomib, while inhibiting this expression using amitryptilin leads to a higher efficacy of drug therapy.

Amitryptilin strengthens effects of bortezomib and melphalan treatment in Multiple Myeloma by inhibiting acid sphingomyelinase.