The proteasome inhibitor carfilzomib functions independently of p53 to induce cytotoxicity and an atypical NF-κB response in chronic lymphocytic leukemia cells.

PURPOSE: The proteasome consists of chymotrypsin-like (CT-L), trypsin-like, and caspase-like subunits that cleave substrates preferentially by amino acid sequence. Proteasomes mediate degradation of regulatory proteins of the p53, Bcl-2, and nuclear factor-κB (NF-κB) families that are aberrantly active in chronic lymphocytic leukemia (CLL). CLL remains an incurable disease, and new treatments are especially needed in the relapsed/refractory setting. We therefore investigated the effects of the proteasome inhibitor carfilzomib (CFZ) in CLL cells. EXPERIMENTAL DESIGN: Tumor cells from CLL patients were assayed in vitro using immunoblotting, real-time polymerase chain reaction, and electrophoretic mobility shift assays. In addition, a p53 dominant-negative construct was generated in a human B-cell line. RESULTS: Unlike bortezomib, CFZ potently induces apoptosis in CLL patient cells in the presence of human serum. CLL cells have significantly lower basal CT-L activity compared to normal B and T cells, although activity is inhibited similarly in T cells versus CLL. Co-culture of CLL cells on stroma protected from CFZ-mediated cytotoxicity; however, PI3K inhibition significantly diminished this stromal protection. CFZ-mediated cytotoxicity in leukemic B cells is caspase-dependent and occurs irrespective of p53 status. In CLL cells, CFZ promotes atypical activation of NF-κB evidenced by loss of cytoplasmic IκBα, phosphorylation of IκBα, and increased p50/p65 DNA binding, without subsequent increases in canonical NF-κB target gene transcription. CONCLUSIONS: Together, these data provide new mechanistic insights into the activity of CFZ in CLL and support phase I investigation of CFZ in this disease.

The novel deacetylase inhibitor AR-42 demonstrates pre-clinical activity in B-cell malignancies in vitro and in vivo.

BACKGROUND: While deacetylase (DAC) inhibitors show promise for the treatment of B-cell malignancies, those introduced to date are weak inhibitors of class I and II DACs or potent inhibitors of class I DAC only, and have shown suboptimal activity or unacceptable toxicities. We therefore investigated the novel DAC inhibitor AR-42 to determine its efficacy in B-cell malignancies. PRINCIPAL FINDINGS: In mantle cell lymphoma (JeKo-1), Burkitt's lymphoma (Raji), and acute lymphoblastic leukemia (697) cell lines, the 48-hr IC(50) (50% growth inhibitory concentration) of AR-42 is 0.61 microM or less. In chronic lymphocytic leukemia (CLL) patient cells, the 48-hr LC(50) (concentration lethal to 50%) of AR-42 is 0.76 microM. AR-42 produces dose- and time-dependent acetylation both of histones and tubulin, and induces caspase-dependent apoptosis that is not reduced in the presence of stromal cells. AR-42 also sensitizes CLL cells to TNF-Related Apoptosis Inducing Ligand (TRAIL), potentially through reduction of c-FLIP. AR-42 significantly reduced leukocyte counts and/or prolonged survival in three separate mouse models of B-cell malignancy without evidence of toxicity. CONCLUSIONS/SIGNIFICANCE: Together, these data demonstrate that AR-42 has in vitro and in vivo efficacy at tolerable doses. These results strongly support upcoming phase I testing of AR-42 in B-cell malignancies.