TAK-659 targets spleen tyrosine kinase (Syk), a non-receptor tyrosine kinase that plays a critical role in B-cell receptor signaling, immune cell activation, and tumor cell survival. It inhibits Syk with an IC50 of 3.2 nM. Importantly, TAK-659 does not inhibit the protein homologue ZAP-70 in T cells, demonstrating its selectivity.

In vitro activity: In a cell proliferation assay, TAK-659 shows inhibition toward a SYK-dependent cell line (OCILY10). TAK-659 is shown to be sensitive toward FLT3-ITD dependent cell lines, MV4-11 and MOLM-13 while the WT FLT3 RS4-11 (ALL cell line) and RA1 (Burkitt’s Lymphoma cell line) are not sensitive toward TAK-659. The sensitivity to TAK-659 is associated with mutations impacting SYK activity in B cell lymphomas, whereas TAK-659 is not cytotoxic for adherent primary or solid tumor cell lines. TAK-659 inhibits the microenvironment-induced activation of Syk and downstream signaling molecules, without inhibiting the protein homologue ZAP-70 in T cells. Importantly, the pro-survival, proliferative, chemoresistant and activation effects promoted by the microenvironment are abrogated by TAK-659, which furthermore blocks CLL cell migration toward BMSC, CXCL12, and CXCL13.
Kinase Assay: TAK-659 hydrochloride is a potent, selective and orally available spleen tyrosine kinase (Syk) inhibitor with an IC50 of 3.2 nM.
Cell Assay: Cells are maintained at 37°C in a humidified atmosphere containing 5-8% CO2. In a panel of hematological and solid tumor cell lines, inhibition of cell viability is determined using the soluble tetrazolium salt, MTS. Cells are seeded in 96-well tissue culture plates and are incubated at 37°C/5% CO2 for 24 hours prior to addition of compounds or DMSO vehicle. After 72 or 96 hours of incubation with compounds, MTS conversion by metabolically active cells is determined by measuring the OD490 nm of the wells using a Thermomax microplate reader. To generate concentration-response curves, cells are treated in duplicate with a range of serial compound dilutions. Prior to addition to cells, compound dilutions are prepared in DMSO. Equal amounts of DMSO are added to cells (final concentration is 0.5%). After background correction and normalization against DMSO-treated cells, EC50 values are calculated by curve-fitting these cell viability results using nonlinear regression analysis.

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In vitro, TAK-659 shows inhibition toward the Syk-dependent cell line OCI-LY10 in cell proliferation assays. It is sensitive toward FLT3-ITD dependent cell lines MV4-11 and MOLM-13, while WT FLT3 cell lines RS4-11 and RA1 are not sensitive. Sensitivity is associated with mutations impacting Syk activity in B-cell lymphomas. It is not cytotoxic for adherent primary or solid tumor cell lines.

TAK-659 is currently undergoing Phase I clinical trials for advanced solid tumor and lymphoma malignancies, a Phase Ib study in advanced solid tumors in combination with nivolumab, and PhIb/II trials for relapsed/refractory AML. TAK-659 blocks anti-IgD (immune-globulin D antibody) stimulated CD86 expression in mouse peripheral B cells in vivo. In the OCI-LY10 xenograft and DLBCL PHTX-95L (primary human tumor graft from DLBCL patient) mouse models, TAK-659 demonstrates potent tumor growth inhibition (TGI) after 20 days of treatment. In the FLT3-dependent MV4-11 xenograft model, TAK-659 shows tumor regression at 60 mg/kg daily after 20 days of dosing

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In vivo, TAK-659 inhibits the microenvironment-induced activation of Syk and downstream signaling molecules. It abrogates pro-survival, proliferative, chemoresistant, and activation effects promoted by the microenvironment. It also blocks CLL cell migration toward BMSC, CXCL12, and CXCL13. Detailed in vivo efficacy data are not provided.

Syk kinase inhibitory activity is determined using cell-free kinase assays with recombinant Syk enzyme, a peptide substrate, and ATP. IC50 values are calculated from dose-response curves. Selectivity is assessed by profiling against a panel of kinases including ZAP-70 to confirm specificity.

Cell viability is determined in a panel of hematological and solid tumor cell lines using the soluble tetrazolium salt MTS. Cells are seeded in 96-well plates and incubated for 24 hours prior to compound addition. After 72 or 96 hours of incubation with compounds, MTS conversion by metabolically active cells is determined by measuring OD490 nm.

0.5% carboxymethylcellulose (CMC); 10, 30, 60 mg/kg QD; by oral gavage
Athymic nude mice

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In vivo animal studies for TAK-659 would be conducted in mouse xenograft models of B-cell lymphomas or leukemia. The compound would be administered orally due to its oral bioavailability, and tumor growth inhibition would be monitored. Detailed protocols are not provided in the available references.

TAK-659 2HCl has the molecular formula C₁₇H₂₁FN₆·2HCl and a molecular weight of 417.31. It is orally bioavailable. The compound is typically handled as a powder with purity ≥98%. Detailed pharmacokinetic parameters such as half-life and bioavailability in preclinical species are not disclosed.

No specific toxicity data are detailed in the available references. As a Syk inhibitor, potential toxicities may include immunosuppression and bleeding risk, which are class effects of Syk inhibitors. The compound's selectivity over ZAP-70 may contribute to a favorable safety profile.

Bioorg Med Chem Lett.2016 Dec 15;26(24):5947-5950;Oncotarget.2017 Jan 3;8(1):742-756.

Mivavotinib is a spleen tyrosine kinase (Syk) inhibitor with potential anti-inflammatory, immunomodulatory, and antitumor activities. After administration, Mivavotinib inhibits Syk activity, thereby blocking downstream B cell receptor (BCR) signaling, leading to suppression of B cell activation, chemotaxis, adhesion, and proliferation. Syk is a BCR-associated non-receptor tyrosine kinase that mediates various cellular responses, including proliferation, differentiation, and phagocytosis. It is expressed in hematopoietic tissues and is often overexpressed in hematopoietic malignancies.
See also: Mivavotinib citrate (note moved to).

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TAK-659 2HCl is a research compound not yet approved for clinical use. It has been studied as a potential anticancer agent, particularly in B-cell malignancies and FLT3-ITD-driven leukemias. Its ability to inhibit microenvironment-induced Syk activation makes it a promising candidate for targeting tumor-stroma interactions.

C17H21FN6.2HCL

417.31

344.176

1312691-41-0

1312691-41-0 (2HCl);1952251-28-3 (HCl);1312691-33-0;

53252276

Typically exists as solid at room temperature

0.5

3

6

3

25

508

2

CN1C=C(C=N1)C2=NC(=C(C3=C2C(=O)NC3)F)N[C@@H]4CCCC[C@@H]4N

MJHOMTRKVMKCNE-NWDGAFQWSA-N

InChI=1S/C17H21FN6O/c1-24-8-9(6-21-24)15-13-10(7-20-17(13)25)14(18)16(23-15)22-12-5-3-2-4-11(12)19/h6,8,11-12H,2-5,7,19H2,1H3,(H,20,25)(H,22,23)/t11-,12+/m0/s1

6-[[(1R,2S)-2-aminocyclohexyl]amino]-7-fluoro-4-(1-methylpyrazol-4-yl)-1,2-dihydropyrrolo[3,4-c]pyridin-3-one

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)