Abstract

Atiprimod (N-N-diethl-8, 8-dipropyl-2-azaspiro [4.5] decane-2-propanamine) is an orally bio-available cationic amphiphilic agent previously studied for its in vivo anti-inflammatory activity. We previously demonstrated that Atiprimod, in a time- and dose-dependent manner, inhibits myeloma cell growth and survival in IL-6 independent (OPM1) as well as dependent (INA-6) MM cell-lines with an IC50 in the range of 0.5–2 μM. We have now further characterized the molecular changes in MM cells induced by Atiprimod, evaluating the gene expression profile of INA-6 and OPM-1 cells exposed to this drug. Following 24 h treatment, Atiprimod induced significant down-regulation of genes involved in growth and cell-cycle control (CDC14, CDC2-like 5, Cyclin-E binding protein 1 and MDM2), adhesion (ITGA-6, ITGA-X, ADAM-17, CDH-3, CDH-6, CTNND-1, PECAM-1 and MADCAM-1) cell-signalling pathways (PRKAB-1, Mapk-7, GPR-125 and GPR-32), up-regulation of genes implicated in apoptotic cascades (TNFAIP-3, TNFSF-10, TNFRSF-10c, CDKN2A and CDKN1A) and in negative regulation of signal transduction (RGS-4 and IGFBP-5). We next evaluated the in vivo activity of Atiprimod using three SCID mouse models of human MM: 1) To evaluate effects of Atiprimod directly on MM cells, we used SCID mice bearing subcutaneous OPM1 tumors and treated i.p. with Atiprimod or vehicle alone (PBS) on alternate days for 7 days. In this model 31%, 48% and 55% inhibition of tumor growth were observed in mice treated with Atiprimod at 20, 30 and 50 mg/kg, respectively, compared to control group. 2) To evaluate effects of Atiprimod on MM cells in the context of a human BM microenvironment, we used SCID mice implanted with a human fetal bone chip (SCID-hu) engrafted with IL-6-dependant INA-6 cells and treated i.p. with Atiprimod (40 mg/kg) or vehicle alone (PBS) for two weeks. The response was evaluated by detection of serum soluble human IL-6 receptor (shuIL-6R) released by MM cells in murine serum. We observed a 60% reduction in shuIL-6R level in mice treated with Atiprimod versus control group. 3) Finally, to evaluate effects of Atiprimod on primary patient cells in a human microenvironment, we used SCID-hu mice engrafted with patient MM cells (IgG λ) and treated i.p. with Atiprimod (50 mg/kg) or vehicle alone (PBS) for 4 weeks. Treatment with Atiprimod induced a reduction in both human IgG and λ light chain levels in mouse sera, whereas levels of both proteins continued to rise in mice treated with vehicle alone. Taken together these data demonstrate the in vivo anti-tumor activity of Atiprimod and provide a rational for its clinical evaluation in MM. Based on these data the drug is presently in a multi-site Phase I/IIa clinical trial in patients with relapsed or refractory multiple myeloma (MM).

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