Ruxolitinib

JAK3, JAK2, JAK1

306, 37

6- to 8-week-old female BALB/c mice with Ba/F3-JAK2V617F tumors

INCB018424 has entered in a Phase II clinical trial for the treatment of breast cancer.

180 mg/kg

2.7 nM, 2.7 nM, 2.7 nM, 2.7 nM, 2.7 nM, 2.7 nM

INCB018424 treatment improves viability and splenomegaly in a JAK2V617F-driven model of malignant disease. Spleens from INCB018424-treated mice demonstrates significantly fewer of these neoplastic features and retained normal lymphoid components. The mass of white pulp in INCB018424-treated mice is similar to that of naive and vehicle-treated mice. In agreement with the biochemical selectivity of INCB018424 against JAK3, whereas cyclophosphamide significantly decreases the absolute lymphocyte counts compared with vehicle-treated animals after 2 weeks of treatment, daily administration of INCB018424 for a period of 4 weeks has no significant effect on circulating lymphocyte numbers. Similarly, INCB018424 has minimal effects on thymus weights. [2]

[2], Biochemical assays, The kinase domains of human JAK1 (837-1142), JAK2 (828-1132), JAK3 (781-1124), and Tyk2 (873-1187) are cloned by PCR with N-terminal epitope tags. Recombinant proteins are expressed using Sf21 cells and baculovirus vectors and purified with affinity chromatography. JAK kinase assays used a homogeneous time-resolved fluorescence assay with the peptide substrate (-EQEDEPEGDYFEWLE). Each enzyme reaction is carried out with INCB018424 or control, JAK enzyme, 500 nM peptide, adenosine triphosphate (ATP, 1 mM), and 2.0% dimethyl sulfoxide (DMSO) for 1 hour. The 50% inhibitory concentration (IC50) is calculated as the compound concentration required for inhibition of 50% of the fluorescent signal. Biochemical assays for CHK2 and c-MET enzymes are performed using standard conditions (Michaelis constant [Km] ATP) with recombinantly expressed catalytic domains from each protein and synthetic peptide substrates., An additional panel of kinase assays (Abl, Akt1, AurA, AurB, CDC2, CDK2, CDK4, CHK2, c-kit, c-Met, GFR, EphB4, ERK1, ERK2, FLT-1, HER2, IGF1R, IKKalpha, IKKbeta, JAK2, JAK3, JNK1, Lck, MEK1, p38alpha, p70S6K, PKA, PKCalpha, Src, and ZAP70) is performed using standard conditions using 200 nM INCB018424. Significant inhibition is defined as more than or equal to 30% (average of duplicate assays) compared with control values.

DMSO 61 mg/mL, Water <1 mg/mL, Ethanol 61 mg/mL

(3R)-3-(4-(7H-pyrrolo[2, 3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentylpropanenitrile