Fostamatinib (R788) disodium

SYK

624, 42

B6/C3H F1 female mice intraperitoneally injected with TCL1-002, TCL1-551, or TCL1-870 leukemia cells, and Eu-TCL1 transgenic mice

A Phase II study of R935788 in advanced colorectal, non-small cell lung, head and neck, hepatocelluar and renal cell carcinomas and pheochromocytoma and thyroid tumors has been completed.

80 mg/kg/day

41 nM [1], 41 nM [1], 41 nM [1], 41 nM [1], 41 nM [1], 41 nM [1]

Given that plasma half-life of R406 in mice is less than 2 hours, R935788 is administered in 3 divided doses at 3-hour intervals to provide continuous Syk inhibition during each day of treatment, mimicking the longer plasma half-life in humans (15 hours). Despite the relatively modest cytotoxic effect in vitro, R935788 significantly inhibits the proliferation and survival of leukemic cell in vivo, which is associated with the blocking of antigen-dependent B-cell receptor (Bcr) signaling rather than inhibition of constitutive Syk activity. R935788 treatment at 80 mg/kg/day for 18-21 days potently inhibits tumor growth of TCL1-002, TCL1-551 and TCL1-870 in mice with undetectable leukemic CD5+/B220+ cells at the last day of treatment, significantly prolongs the survival of the treated mice with median survival increased from 45/46 days to 170/172 days, and completely eradicates the malignant cells in a substantial proportion of mice after a 6-month follow-up period without affecting the production of normal B lymphocytes. R935788 treatment also induces an early and transient migration of both normal and malignant B cells from spleen and lymph nodes to peripheral blood, which is subsequently followed by selective growth inhibition of the malignant B-cell population. In addition, R935788 is also effective against spontaneously developing TCL1 leukemias in Eu-TCL1 transgenic mice. [3]

In vitro fluorescence polarization kinase assays, R406 (in vitro active form of R935788) is serially diluted in DMSO and then diluted to 1% DMSO in kinase buffer (20 mM HEPES, pH 7.4, 5 mM MgCl2, 2 mM MnCl2, 1 mM DTT, 0.1 mg/mL acetylated BGG). ATP and substrate in kinase buffer are added at room temperature, resulting in a final DMSO concentration on 0.2%. The kinase reactions are performed in a final volume of 20 uL containing 5 uM HS1 peptide substrate and 4 uM ATP and started by addition of 0.125 ng of Syk in kinase buffer. The reaction is allowed to proceed for 40 minutes at room temperature. The reaction is stopped by the addition of 20 uL of PTK quench mix containing EDTA/anti-phosphotyrosine antibody (1 final)/fluorescent phosphopeptide tracer (0.5 final) diluted in FP Dilution Buffer. The plate is incubated for 30 minutes in the dark at room temperature and then read on a Polarion fluorescence polarization plate reader. Data is converted to determine the amount of phosphopeptide present using a calibration curve generated by competition with the phosphopeptide competitor provided in the Tyrosine Kinase Assay Kit. For IC50 determination, R406 is tested at eleven concentrations in duplicate and curve-fitting is performed by non-linear regression analysis using Prism GraphPad Software.

DMSO 0.4 mg/mL, Water <1 mg/mL, Ethanol <1 mg/mL

2H-Pyrido[3, 2-b]-1, 4-oxazin-3(4H)-one, 6-[[5-fluoro-2-[(3, 4, 5-trimethoxyphenyl)amino]-4-pyrimidinyl]amino]-2, 2-dimethyl-4-[(phosphonooxy)methyl]-, sodium salt (1:2)