pazopanib and Leukemia--Lymphocytic--Chronic--B-Cell

Topics: Activin Receptors, Type II; Antibodies, Monoclonal, Humanized; Antineoplastic Agents; beta-Thalassemia; Graft vs Host Disease; Hematologic Diseases; Humans; Immunoglobulin Fc Fragments; Indazoles; Interleukin-1 Receptor-Associated Kinases; Internet; Leukemia, Lymphocytic, Chronic, B-Cell; Leukemia, Myeloid, Acute; Lymphoma, B-Cell; Multiple Myeloma; Protein Kinase Inhibitors; Pyrimidines; Recombinant Fusion Proteins; Sulfonamides; Telangiectasia, Hereditary Hemorrhagic

We have recently shown that the gene encoding the truncated form of protein tyrosine phosphatase receptor-type O (PTPROt) expressed predominantly in hematopoietic cells is epigenetically silenced in human primary chronic lymphocytic leukemia (B-CLL). To determine whether increased phosphorylation of the PTPROt substrates following PTPROt suppression alters signal transduction pathway(s) that impart a growth advantage to the leukemic lymphocytes, it is critical to discern the key substrates of PTPROt. Here, we used substrate-trapping assay to identify two novel substrates of PTPROt, the tyrosine kinases Lyn and ZAP70. Both Lyn and ZAP70 were dephosphorylated by wild-type PTPROt, but not by its catalytic site (CS) mutant. A critical phosphorylation site in Lyn, Y397, essential for its activity was dephosphorylated by PTPROt. Consequently, the activity of Lyn kinase was compromised when co-expressed with PTPROt-WT compared to vector control or upon co-expression with PTPROt-CS. Ectopic expression of PTPROt in Raji cells reduced phosphorylation of Lyn in the absence of any change in its protein levels. These results have revealed the physiological importance of PTPROt in regulating B-cell receptor signaling at Lyn kinase. Further, ectopic expression of PTPROt also sensitized the cells to the VEGF-R inhibitor Pazopanib.

Topics: B-Lymphocytes; Base Sequence; Blotting, Western; Catalytic Domain; Cell Line, Tumor; DNA Primers; Humans; Indazoles; Leukemia, Lymphocytic, Chronic, B-Cell; Phosphorylation; Pyrimidines; Receptor-Like Protein Tyrosine Phosphatases, Class 3; Receptors, Vascular Endothelial Growth Factor; src-Family Kinases; Substrate Specificity; Sulfonamides; ZAP-70 Protein-Tyrosine Kinase

There is evidence that vascular endothelial growth factor (VEGF) is a critical microenvironmental factor that exerts angiogenesis-independent effects on the survival of chronic lymphocytic leukemia (CLL) cells. Vatalanib and pazopanib are potent orally available VEGF receptor tyrosine kinase inhibitors. We investigated the efficacy and selectivity of both compounds in CLL cells, simulated potential combination with conventional cytostatics, and tested the effect of both substances on CLL-like tumor xenografts.. Primary CLL and normal peripheral blood cells were tested for viability after incubation with varying concentrations of both inhibitors. Further, phosphorylation status of VEGF receptor on treatment, caspase activation, and poly(ADP-ribose) polymerase cleavage were assessed. Combinations of each inhibitor with fludarabine, vincristine, and doxorubicin were analyzed for possible synergistic effects in vitro. For in vivo testing, mice grafted with the CLL-like cell line JVM-3 were treated orally with each inhibitor.. Vatalanib and pazopanib decreased phosphorylation of the VEGF receptor, along with induction of apoptosis in CLL cells in clinically achievable concentrations. Healthy B cells were only mildly affected. Immunoblots showed downregulation of the antiapoptotic proteins XIAP and MCL1, whereas poly(ADP-ribose) polymerase cleavage was increased. Combinations with conventional cytostatic agents resulted in synergistic effects. Treatment of xenografted mice with 100 mg/kg of body weight for 21 days resulted in tumor inhibition rates of 76% (vatalanib) and 77% (pazopanib). In two mice, a total tumor eradication could be observed. No gross systemic toxicity occurred.. We conclude that VEGF inhibition is a promising new therapeutic approach in CLL. Vatalanib and pazopanib seem to be effective and safe candidates to be further evaluated for this purpose.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Caspases; Cell Line, Tumor; Dose-Response Relationship, Drug; Humans; Indazoles; Leukemia, Lymphocytic, Chronic, B-Cell; Mice; Phosphorylation; Phthalazines; Protein Kinase Inhibitors; Pyridines; Pyrimidines; Receptors, Vascular Endothelial Growth Factor; Sulfonamides; Xenograft Model Antitumor Assays