fk-866 and Multiple-Myeloma

Antibodies targeting CD38, a NAD

Topics: Acrylamides; Adenosine; Adenosine Diphosphate Ribose; ADP-ribosyl Cyclase 1; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Antineoplastic Agents, Immunological; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Cytokines; Drug Synergism; Humans; Membrane Glycoproteins; Multiple Myeloma; NAD; Niacinamide; Nicotinamide Phosphoribosyltransferase; Piperidines; T-Lymphocytes, Cytotoxic; T-Lymphocytes, Regulatory; Tumor Escape; Warburg Effect, Oncologic

Multiple myeloma (MM) cells typically grow in focal lesions, stimulating osteoclasts that destroy bone and support MM. Osteoclasts and MM cells are hypermetabolic. The coenzyme nicotinamide adenine dinucleotide (NAD(+)) is not only essential for cellular metabolism; it also affects activity of NAD-dependent enzymes, such as PARP-1 and SIRT-1. Nicotinamide phosphoribosyltransferase (NAMPT/PBEF/visfatin, encoded by PBEF1) is a rate-limiting enzyme in NAD(+) biosynthesis from nicotinamide. Coculture of primary MM cells with osteoclasts induced PBEF1 upregulation in both cell types. PBEF1 expression was higher in experimental myelomatous bones than in nonmyelomatous bone and higher in MM patients' plasma cells than in healthy donors' counterparts. APO866 is a specific PBEF1 inhibitor known to deplete cellular NAD(+). APO866 at low nanomolar concentrations inhibited growth of primary MM cells or MM cell lines cultured alone or cocultured with osteoclasts and induced apoptosis in these cells. PBEF1 activity and NAD(+) content were reduced in MM cells by APO866, resulting in lower activity of PARP-1 and SIRT-1. The inhibitory effect of APO866 on MM cell growth was abrogated by supplementation of extracellular NAD(+) or NAM. APO866 inhibited NF-κB activity in osteoclast precursors and suppressed osteoclast formation and activity. PBEF1 knockdown similarly inhibited MM cell growth and osteoclast formation. In the SCID-rab model, APO866 inhibited growth of primary MM and H929 cells and prevented bone disease. These findings indicate that MM cells and osteoclasts are highly sensitive to NAD(+) depletion and that PBEF1 inhibition represents a novel approach to target cellular metabolism and inhibit PARP-1 and bone disease in MM.

Topics: Acrylamides; Animals; Bone and Bones; Cell Differentiation; Coculture Techniques; Cytokines; Enzyme Induction; Enzyme Inhibitors; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Humans; Mice; Mice, SCID; Multiple Myeloma; NAD; Neoplasm Proteins; NF-kappa B; Niacinamide; Nicotinamide Mononucleotide; Nicotinamide Phosphoribosyltransferase; Osteoclasts; Osteolysis; Piperidines; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Rabbits; Sirtuin 1; Tumor Cells, Cultured; Up-Regulation

We recently demonstrated that Nicotinamide phosphoribosyltransferase (Nampt) inhibition depletes intracellular NAD⁺ content leading, to autophagic multiple myeloma (MM) cell death. Bortezomib has remarkably improved MM patient outcome, but dose-limiting toxicities and development of resistance limit its long-term utility. Here we observed higher Nampt messenger RNA levels in bortezomib-resistant patient MM cells, which correlated with decreased overall survival. We demonstrated that combining the NAD⁺ depleting agent FK866 with bortezomib induces synergistic anti-MM cell death and overcomes bortezomib resistance. This effect is associated with (1) activation of caspase-8, caspase-9, caspase-3, poly (ADP-ribose) polymerase, and downregulation of Mcl-1; (2) enhanced intracellular NAD⁺ depletion; (3) inhibition of chymotrypsin-like, caspase-like, and trypsin-like proteasome activities; (4) inhibition of nuclear factor κB signaling; and (5) inhibition of angiogenesis. Furthermore, Nampt knockdown significantly enhances the anti-MM effect of bortezomib, which can be rescued by ectopically overexpressing Nampt. In a murine xenograft MM model, low-dose combination FK866 and Bortezomib is well tolerated, significantly inhibits tumor growth, and prolongs host survival. Taken together, these findings indicate that intracellular NAD⁺ level represents a major determinant in the ability of bortezomib to induce apoptosis in MM cells and provide proof of concept for the combination with FK866 as a new strategy to enhance sensitivity or overcome resistance to bortezomib.

Topics: Acrylamides; Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Autophagy; Biomarkers, Tumor; Blotting, Western; Boronic Acids; Bortezomib; Case-Control Studies; Caspases; Cell Proliferation; Drug Synergism; Female; Fluorescent Antibody Technique; Gene Expression Profiling; Humans; Male; Mice; Mice, SCID; Multiple Myeloma; NAD; Neoplasm Recurrence, Local; NF-kappa B; Nicotinamide Phosphoribosyltransferase; Oligonucleotide Array Sequence Analysis; Piperidines; Poly(ADP-ribose) Polymerases; Prognosis; Pyrazines; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Survival Rate; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Multiple myeloma (MM) is a clonal B-cell malignancy characterized by the proliferation of plasma cells in the bone marrow. Despite recent therapeutic advances, MM remains an incurable disease. Therefore, research has focused on defining new aspects in MM biology that can be therapeutically targeted. Compelling evidence suggests that malignant cells have a higher nicotinamide adenine dinucleotide (NAD+) turnover rate than normal cells, suggesting that this biosynthetic pathway represents an attractive target for cancer treatment. We recently reported that an intracellular NAD(+)-depleting agent, FK866, exerts its anti-MM effect by triggering autophagic cell death via transcriptional-dependent (transcription factor EB, TFEB) and -independent (PI3K-MTORC1) mechanisms. Our findings link intracellular NAD(+) levels to autophagy in MM cells, providing the rationale for novel targeted therapies in MM.

Topics: Acrylamides; Apoptosis; Autophagy; Cell Death; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Gene Transfer Techniques; Humans; Lentivirus; Multiple Myeloma; NAD; Piperidines; Transcription, Genetic

Malignant cells have a higher nicotinamide adenine dinucleotide (NAD(+)) turnover rate than normal cells, making this biosynthetic pathway an attractive target for cancer treatment. Here we investigated the biologic role of a rate-limiting enzyme involved in NAD(+) synthesis, Nampt, in multiple myeloma (MM). Nampt-specific chemical inhibitor FK866 triggered cytotoxicity in MM cell lines and patient MM cells, but not normal donor as well as MM patients PBMCs. Importantly, FK866 in a dose-dependent fashion triggered cytotoxicity in MM cells resistant to conventional and novel anti-MM therapies and overcomes the protective effects of cytokines (IL-6, IGF-1) and bone marrow stromal cells. Nampt knockdown by RNAi confirmed its pivotal role in maintenance of both MM cell viability and intracellular NAD(+) stores. Interestingly, cytotoxicity of FK866 triggered autophagy, but not apoptosis. A transcriptional-dependent (TFEB) and independent (PI3K/mTORC1) activation of autophagy mediated FK866 MM cytotoxicity. Finally, FK866 demonstrated significant anti-MM activity in a xenograft-murine MM model, associated with down-regulation of ERK1/2 phosphorylation and proteolytic cleavage of LC3 in tumor cells. Our data therefore define a key role of Nampt in MM biology, providing the basis for a novel targeted therapeutic approach.

Topics: Acrylamides; Animals; Antineoplastic Agents; Autophagy; Cell Line, Tumor; Cell Survival; Cytokines; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Enzyme Inhibitors; Gene Expression Regulation, Neoplastic; Humans; Mechanistic Target of Rapamycin Complex 1; Mice; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Molecular Targeted Therapy; Multiple Myeloma; Multiprotein Complexes; NAD; Nicotinamide Phosphoribosyltransferase; Organ Specificity; Piperidines; Proteins; RNA, Small Interfering; Signal Transduction; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays