Preclinical evaluation of the novel HDAC-inhibitor JNJ-26481585 in multiple myeloma reveals strong anti-tumor activity

738 Considerable efforts are currently being devoted to the development and optimization of pharmacologic histone-deacetylase (HDAC) inhibitors for their use in cancer. Here, we undertook preclinical evaluations of the novel pyrimidyl-hydroxamic-acid analog JNJ-26481585 in cells from patients with multiple myeloma (MM) and in established human MM cell lines.
 Dose-effect curves for JNJ-26481585-mediated cell death were established for MM cell lines and freshly isolated CD138-positive primary myeloma cells. The latter were normally tested in the more rigorous setting of coculture with bone marrow stromal cells. Molecular effects of drug treatment on candidate pharmacodynamic markers were evaluated by Western analysis. Combination experiments for the novel HDAC inhibitor and drugs relevant for the treatment of MM were performed as colorimetric assay (Alamar Blue) and analyzed for synergistic effects using Calcusyn software.
 All MM cell lines tested were profoundly sensitive to treatment with JNJ-26481585, and displayed steep dose-effect curves for drug-induced apoptosis at low nanomolar concentrations (EC50: 3-45nM; EC90: 5-90nM). Primary tumor samples also underwent apoptosis, albeit to various extents, and treatment with 10nM JNJ-26481585 broadly segregated primary MM cells into two groups that were either very sensitive (less than 25% survival relative to DMSO-treated controls, 12/24 samples) or more resilient (40-90% survival, 12/24 samples). Synergistic effects were observed for simultaneous treatment of MM cell lines with the novel HDAC-inhibitor and proteasome inhibitor bortezomib. Treatment with JNJ-26481585 usually induced substantial changes in the amount of Bcl2-family members, which, through decreases of antiapoptotic proteins (Mcl1, Bcl2) or cleavage (and thus activation) of proapoptotic Bid, implied a shift towards apoptosis induction. Considerable variation, as to the specific Bcl2-family members involved, existed between different samples. This observation was also true concerning the level of p21CIP-induction or with regard to acetylation of histones H3 and H4 in response to drug treatment. Ongoing experiments further extend the analysis of combinatorial effects between JNJ-26481584 and other drugs relevant to MM therapy, their verification in primary MM cells and further analysis of potential molecular markers for their usefulness as pharmacodynamic markers in myeloma.
 In summary, preclinical evaluation of JNJ-26481584 showed strong anti-tumor activity against primary MM cells and established human MM cell lines. It is therefore a promising compound for further evaluation as future drug candidate in myeloma therapy.