Serotonin Receptor Antagonists Have an In Vitro and In Vivo Anti-Myeloma Effect That Is Mainly Mediated by Caspase Dependent Apoptosis.

To date, serotonin (5-hydroxytryptamine: 5-HT) has been associated with the regulation of several central nervous system functions such as mood, temperature, memory, sleep or appetite. Importantly, a proliferative effect of 5-HT in both normal and tumoral tissue has recently been described. Therefore, serotonin-receptor antagonists have been tested for antineoplastic purposes. In this study, we have investigated the in vitro and in vivo anti-multiple myeloma (MM) effect of two analogs of 5-HT-receptor antagonists (ICI-735 and ICI-685). Both compounds induced cytotoxicity (MTT) and a decrease in proliferation (thymidine uptake) in several MM cell lines both sensitive and resistant to conventional therapies, as well as freshly isolated MM cells, with an IC-50 at 24h ranging between 3-10 μM for ICI-735 and 5–20 μM for ICI-685. No significant toxicity was observed at these doses of both drugs in PBMCs obtained from normal donors. In order to analyze the efficacy of the drugs in the presence of the bone marrow (BM) microenvironment, we tested both compounds in MM1S cells co-cultured with IL-6, IGF-1 or long-term patient BM stromal cells (SCs). ICI-735 and ICI-685 overcame the proliferative advantage conferred by the cytokines and BMSCs. Apoptosis was the main mechanism of cytotoxicity, assessed by Annexin-V staining. Treatment with ICI-735 and ICI-685 induced cleavage of PARP and caspase 3, 7, 8 and 9 at early time points, suggesting involvement of both extrinsic and intrinsic pathways of apoptosis. The presence of caspase-induced apoptosis was confirmed since a pan-caspase inhibitor (ZVAD-FMK) partially abrogated apoptosis, as well as the cleavage of PARP and caspases, induced by these compounds. An increase of AIF was also detected in the cytoplasmic fraction of cells treated with ICI-735, indicating a role for caspase-independent apoptosis. The in vivo efficacy was next evaluated in a xenograft murine (CB-17 SCID-mice) model. Five mice were injected intraperitoneally with 20 mg/Kg of ICI-735, while 5 received vehicle alone. No significant toxicity was detected, except for a slight somnolence after the first doses, in the mice that received the drug. A decrease in tumor volume was observed in the treated group and, despite the small number of mice, survival analysis showed a trend toward prolonged survival for mice receiving the drug (p=0.11). Ongoing studies are providing the preclinical rationale for clinical protocols to improve patient outcome in MM.