Gene Expression Profiling in Multiple Myeloma Cells Treated with the Novel Anti-Myeloma Agents Zoledronate and Fluvastatin.

We have shown in vitro that multiple myeloma (MM) cells can be destroyed by treating them with the mevalonate pathway inhibitors zoledronate and fluvastatin. While the efficacy of these compounds singly and combination have been demonstrated, their exact modes of action remain largely unknown. The present study aimed to use microarray and quantitative real-time PCR (QRT-PCR) techniques to analyse gene expression in treated myeloma cells to identify novel genes and pathways involved in the anti-myeloma action of these compounds. The human MM cell line NCI-H929 was treated with zoledronate and fluvastatin singly and in combination, and RNA was extracted and used to interrogate oligonucleotide microarrays consisting of 19,000 features representing known and unknown genes. Quantitative real-time PCR was subsequently used to confirm the expression of several genes of interest. Flow cytometry with Annexin V FITC staining was used to detect apoptosis. It was observed that genes related to apoptosis (caspases and p53-related genes), cell cycle control (cyclins), GTPase signalling (Rabs), and growth and proliferation (growth factors) were particularly affected by zoledronate and fluvastatin, and some of these genetic effects were synergistic when a combination of zoledronate and fluvastatin was used. QRT-PCR confirmed the effects on the caspase- and p53-related apoptotic pathways, and these effects were correlated with increased apoptosis in the myeloma cells. The mevalonate pathway inhibitors fluvastatin and zoledronate are highly efficient at killing MM cells, and their effects appear to be synergistic. Our microarray and QT-PCR analyses demonstrated that the expression of specific groups of genes important to the survival and proliferation of myeloma cells are affected by these compounds. p53 and caspase-dependent pathways appear to be the key apoptotic cascades stimulated. Insights into the mechanisms of these novel therapeutics are important as they might help to define their roles in the treatment of multiple myeloma.