Ceramide (Cer) and sphingosine (Sph) interfere with critical cellular functions relevant for cancer progression and cell survival. While Cer has already been investigated as a potential drug target for lymphoma treatment, information about the potency of sphingosine is scarce. The aim of this study therefore was to evaluate Sph and its synthetic stereoisomer L-threo- sphingosine (Lt-Sph) as potential treatment options for aggressive lymphomas. Methods: Diffuse large B cell lymphoma (DLBCL) cell lines were incubated with Sph and Lt-Sph and consequently analysed by flow cytometry (FACS), enzyme- linked immunosorbent assay (ELISA), liquid chromatography coupled to triple- quadrupole mass spectrometry (LC/MS/MS), electron microscopy, and Western blot. Results: Sph induced cell death and blocked cell growth independently of S1P receptors in different DLBCL cell lines. Three different modes of Sph- mediated cell death were observed: Apoptosis, autophagy, and protein kinase C (PKC) inhibition. Generation of pro-apoptotic Cer accounted only for a minor portion of the apoptotic rate. Conclusion: Sph and its analogues could evolve as alternative treatment options for aggressive lymphomas via PKC inhibition, apoptosis, and autophagy. These physiological responses induced by different intracellular signalling cascades (phosphorylation of JNK, PARP cleavage, LC3-II accumulation) identify Sph and analogues as potent cell death inducing agents.