Gandotinib

Concentration notes: Data suggests it could be dosed up to 20 uM in cellular assays.

BID dosing is required to maintain reasonable drug levels in vivo models.  PK parameters are not discussed; dose selection in alternate models would require additional work.

Activation of the JAK-STAT signalling pathway has been associated with cancer progression across multiple tumour types either as a tumour intrinsic driver of cancer cell growth and spread or as a modulator of immunosurveillance (10.3390/cancers12071971).  Constitutive activation of JAK-STAT signalling can arise through a number of different mechanisms, including the elevated expression of cytokines and/or activating mutations in receptors upstream of JAK-STAT signalling. Activating mutations of JAK2 are commonly found in a group of hematologic malignancies known as myeloproliferative neoplasms (MPNs). An acquired point mutation (JAK2 1849G-T) results in a missense substitution of phenylalanine for valine at amino acid position 617 (V617F) within the autoinhibitory pseudokinase domain of JAK2. This substitution results in a gain-of-function activation mutant of JAK2 (JAK2V617F) with subsequent phosphorylation of STAT5 (10.1038/bcj.2013.6).

Biochemical screening assays demonstrated the small molecule compound LY2784544 exhibits 8- and 16-fold selectivity for JAK2 (IC50= 0.0025) over JAK1(IC50=0.0198) and JAK3 (IC50=0.048), respectively. Further assessment of activity against a CEREP kinase panel (total of 99 kinases) demonstrated 20 kinases were inhibited by LY2784544 with IC50 of <0.3 M. with most potent activity upon FLT3, FLT4, FGFR2 with IC50: 0.004, 0.025 and 0.032 respectively (10.1038/bcj.2013.6).  In Ba/F3 cellular assays LY2784544 effectively inhibited JAK2V617F-driven signalling and cell proliferation in Ba/F3 cells (IC50; 0.02 and 0.05 M, respectively). In comparison, LY2784544 was much less potent at inhibiting
interleukin-3-stimulated wild-type JAK2-mediated signalling and cell proliferation (IC50:1.183 and 1.309 M, respectively). In vivo, LY2784544 effectively inhibited STAT5 phosphorylation in Ba/F3-JAK2V617F-GFP (green fluorescent protein) ascitic tumor cells
(TED50.12.7 mg/kg) and significantly reduced Ba/F3-JAK2V617F-GFP tumor burden in the JAK2V617F-induced MPN model (TED50.13.7 mg/kg, twice daily).

LY2784544 is therefore a suitable chemical probe for exploring the activity of the JAK2V617F mutant kinase.  LY2784544 may also represent a useful orthogonal probe for exploring the activity of wild type JAK2, however further validation to rule out an activity against other kinases particularly FLT3 and FLT4 would need to be determined.

Cellular concentration notes: I would recommend this is used as a chemical probe specifically for the JAK2V617F mutant protein at a lower concentration of up to 200 nM to avoid off-target activities

The cpd should also be tested in vitro and in vivo against FLT4 and FLT3 driven tumors and Baf3 cells (may be testing of this cpd against a large panel of Baf3 cell (ACD-Baf3) would be more beneficial than the CEREP biochemical panel.