Human Monocytes and Monocyte-Like Cells Are Highly Sensitive to Plitidepsin-Induced Cell Death In In Vitro Assays

Abstract

AbstractAbstract 4942Plitidepsin (Aplidin®) is a marine-derived cyclic depsipeptide with strong cytotoxic effects against a variety of cancer cell types. It is currently in Phase II/III clinical trials for solid and hematologic malignancies. We evaluated the cytotoxic effects of Plitidepsin on lymphoid and monocyte cells from healthy donors and analysed the mechanisms involved. We found that monocyte cells showed a markedly higher sensitivity to Plitidepsin compared to lymphoid subsets as determined by incorporation of 7-AAD and flow cytometry analysis. Plitidepsin induced a dose- and time-dependent cell death on freshly isolated monocytes (7-AAD+ cells at 24 h: control: 3±1% versus Plitidepsin 10 nM: 47±5%, mean±SEM, n=6) and on macrophages (5±2 vs 49±3, n=5) and dendritic cells (4±1 vs 21±4, n=5) differentiated in vitro. By contrast, resting or mitogen-activated lymphocytes were not affected by Plitidepsin at 10 nM. The mechanisms of induced cell death in monocytes involved the early exposure of phosphatidylserine to the outer leaflet of plasma membrane, activation of caspase-3 and subsequent PARP fragmentation, indicative of death via apoptosis. Incubation of monocytes with Plitidepsin (10 or 100 nM) for 30 minuntes induced ROS production. On the other hand, the cytotoxic effect of Plitidepsin was significantly diminished when monocytes were pre-incubated with the antioxidant reagent Ebselen, which acts as an scavenger of peroxynitrite. We also determined Plitidepsin activity on malignant cells from chronic myelomonocytic leukemia (CMML) and chronic lymphocytic leukemia (CLL) patients. Monocyte-like cells obtained from 3 CMML samples resulted sensitive to Plitidepsin, though to a different extent (viable CD14+ cells at 24 h: control 100%, incubated with Plitidepsin 10 nM: sample 1: 79%, sample 2: 82%, sample 3: 29%). Finally we evaluated the effects of Plitidepsin on nurse-like cells (NLC) from CLL patients. These cells represent a subset differentiated from monocytes that favours leukemic cell progression through pro-survival signals. NLC were very sensitive to Plitidepsin and, more importantly, their death indirectly decreased neoplasic clone viability (7-AAD+ leukemic B cells co-cultured with control NLC: 36±10 versus 7-AAD+ leukemic B cells co-cultured with NLC exposed to Plitidepsin 100 nM for 24 h: 47±6, n=5, p<0.05). Myeloid cells are present in tumor microenvironment and actively affect the malignant cells, both directly and indirectly via the suppression of host immunity. Given their relevance in supporting tumor growth, the sensitivity of monocyte-like cells to Plitidepsin may contribute to its antitumoral effects. Disclosures:Galmarini: PharmaMar: Employment, Equity Ownership. Giordano: PharmaMar: Research Funding.