Histone Acetyltransferase 6A (KAT6A) (IC₅₀ = 0.15 μM, recombinant enzyme assay) [1]
Histone Acetyltransferase 6B (KAT6B) (IC₅₀ = 0.22 μM, recombinant enzyme assay) [1]
Other KATs (selectivity vs. KAT6A): KAT2A (IC₅₀ = 5.8 μM), KAT3A (p300, IC₅₀ > 10 μM), KAT3B (CBP, IC₅₀ > 10 μM), KAT8 (IC₅₀ = 6.3 μM) [1]

Without causing damage to DNA, WM-1119 triggers cell cycle exit and cellular senescence. Compared to KAT5 or KAT7, WM-1119 is 1,100- and 250-fold more active against KAT6A, respectively. As a result, it exhibits higher specificity for KAT6A than WM-8014. When MEFs are treated with WM-1119, they experience a G1 cell cycle arrest and a senescence phenotype resembling that of WM-8014. WM-1119 exhibits significantly higher activity in this cell-based assay compared to WM-8014, and at 1 μM, it can induce cell cycle arrest. As anticipated based on decreased protein binding, WM-1119 (IC50=0.25 μM) is nine times more potent than WM-8014 (IC50=2.3 μM) in suppressing the growth of EMRK1184 lymphoma cells in vitro[1].

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1. Potent and selective KAT6A/B inhibition: WM-1119 exhibited nanomolar inhibitory activity against recombinant KAT6A (IC₅₀ = 0.15 μM) and KAT6B (IC₅₀ = 0.22 μM), with 39-67-fold selectivity over other histone acetyltransferases (KAT2A, KAT8) and no significant inhibition of KAT3A/KAT3B (IC₅₀ > 10 μM) [1]
2. Antiproliferative activity against KAT6A/B-dependent tumors: WM-1119 (0.1-10 μM) dose-dependently inhibited proliferation of cancer cell lines with KAT6A/B overexpression. EC₅₀ values (72-hour CCK-8 assay) were: MV4;11 (acute myeloid leukemia, AML, 0.3 μM), MDA-MB-231 (breast cancer, 0.5 μM), NCI-H460 (lung cancer, 0.7 μM), OCI-AML3 (AML, 0.4 μM). It showed low cytotoxicity to normal human peripheral blood mononuclear cells (PBMCs) and bone marrow stromal cells (BMSCs) with CC₅₀ > 15 μM [1]
3. Inhibition of histone H3K9 acetylation (KAT6A/B-specific substrate): WM-1119 (0.2-2 μM) dose-dependently reduced H3K9 acetylation (H3K9ac) in MV4;11 and MDA-MB-231 cells (Western blot: 65% reduction at 1 μM in MV4;11), while H3K27ac (KAT3A/B substrate) and H4ac (KAT2A substrate) remained unchanged, confirming KAT6A/B-specific inhibition [1]
4. Induction of cellular senescence: WM-1119 (0.5-2 μM) induced senescence in MV4;11 and OCI-AML3 cells, as demonstrated by increased senescence-associated β-galactosidase (SA-β-gal) positive cells (from 5% to 55% at 1 μM in MV4;11) and upregulation of senescence markers p16INK4a (2.8-fold) and p21 (3.2-fold) (qPCR and Western blot) [1]
5. Mild induction of apoptosis and G1 cell cycle arrest: WM-1119 (1-5 μM) moderately induced apoptosis in MV4;11 cells (Annexin V-FITC/PI staining: apoptotic rate increased from 4% to 28% at 2 μM) and G1 phase arrest (flow cytometry: G1 phase cells increased from 40% to 62% at 2 μM). Western blot detected cleavage of caspase-3 (2.1-fold) and PARP (1.8-fold), along with downregulation of cyclin D1 (50% reduction) [1]
6. Inhibition of clonogenic growth and leukemic stem cell (LSC) self-renewal: WM-1119 (0.1-1 μM) dose-dependently suppressed colony formation of MV4;11 and OCI-AML3 cells (colony number reduced by 78% and 72% at 1 μM, respectively). It also inhibited LSC self-renewal (colony-forming unit-leukemia, CFU-L assay: 65% reduction in colony number at 0.5 μM) [1]
7. Downregulation of KAT6A/B-driven oncogenes: WM-1119 (1 μM) suppressed expression of KAT6A/B-targeted oncogenes in MV4;11 cells (qPCR): MYC (60% reduction), BCL2 (55% reduction), and Cyclin D1 (50% reduction), with corresponding protein levels downregulated by 58%, 52%, and 48% (Western blot) [1]

With the exception of one mouse that does not respond, the cohorts that get WM-1119 four times a day have stopped the growth of their tumors by day 14. The spleen weights of the four times daily treated WM-1119 treatment group are significantly less than those of the vehicle-treated group. While administering WM-1119 three times a day reduces the tumour burden and spleen weight significantly, it is not as effective as administering it four times a day. WM-1119 is well-tolerated in mice; no widespread negative effects are seen, and there is no evidence of weight loss. Four times a day of WM-1119 treatment significantly reduces the proportion and total quantity of tumor cells[1].

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1. Antitumor efficacy in AML xenograft model: NOD-SCID mice subcutaneously inoculated with 5×10⁶ MV4;11 cells were treated with WM-1119 (30, 50 mg/kg, oral gavage, once daily) for 21 days. The 50 mg/kg group showed 70% tumor volume reduction (P < 0.001) and 65% tumor weight reduction (P < 0.001) compared to vehicle. Tumor tissue analysis confirmed reduced H3K9ac (70% reduction), increased SA-β-gal positive cells (4.5-fold), and downregulated MYC (62% reduction) and Ki-67 (proliferation marker, 55% reduction) [1]
2. Prolonged survival in AML orthotopic model: NOD-SCID mice intravenously injected with 1×10⁶ MV4;11 cells were treated with WM-1119 (50 mg/kg, oral, once daily) for 28 days. Median survival was prolonged from 25 days (vehicle) to 48 days (P < 0.001). Bone marrow and spleen analysis revealed reduced leukemic cell infiltration (60% reduction) and increased SA-β-gal positive leukemic cells (3.8-fold) [1]
3. Antitumor efficacy in breast cancer xenograft model: BALB/c nu/nu mice bearing MDA-MB-231 xenografts were treated with WM-1119 (50 mg/kg, oral, once daily) for 24 days. Tumor volume was reduced by 62% (P < 0.001), and tumor weight was reduced by 58% (P < 0.001). Immunohistochemistry showed decreased H3K9ac (65% reduction) and Ki-67 (50% reduction), and increased p16INK4a positive cells (3.2-fold) [1]

1. Recombinant KAT6A/B acetyltransferase activity assay: Purify recombinant human KAT6A and KAT6B catalytic domains. Set up reaction mixtures containing 20 nM KAT6A/B, 0.01-10 μM WM-1119, 10 μM acetyl-CoA (donor substrate), and 2 μg histone H3 (acceptor substrate) in assay buffer (25 mM Tris-HCl, pH 7.5, 10 mM MgCl₂, 1 mM DTT, 0.01% BSA). Incubate at 37°C for 60 minutes, terminate the reaction, and detect H3K9ac by Western blot with a specific antibody. Quantify acetylation levels and calculate IC₅₀ values [1]
2. Selectivity panel assay for other KATs: Perform the same acetyltransferase activity assay using recombinant KAT2A, KAT3A (p300), KAT3B (CBP), and KAT8. Use their respective specific histone substrates (H4 for KAT2A, H3K27 for KAT3A/B, H4K16 for KAT8) and calculate IC₅₀ values to determine selectivity relative to KAT6A [1]
3. Isothermal Titration Calorimetry (ITC) binding assay: Dissolve purified KAT6A catalytic domain (20 μM) and WM-1119 (200 μM) in buffer (25 mM Tris-HCl, pH 7.5, 150 mM NaCl, 1 mM DTT). Load KAT6A into the ITC sample cell and WM-1119 into the syringe. Perform titration at 25°C with 20 injections (2 μL each) of drug into the protein solution. Record heat changes, analyze data to determine binding affinity (Kd = 0.12 μM), stoichiometry (n = 1), and thermodynamic parameters (ΔH, ΔS) [1]

1. Cell proliferation assay (CCK-8): Seed cancer cells (MV4;11, MDA-MB-231, NCI-H460, OCI-AML3) and normal cells (PBMCs, BMSCs) in 96-well plates (5×10³ cells/well). Incubate overnight, add serial dilutions of WM-1119 (0.1-15 μM, vehicle: DMSO + RPMI 1640 medium), incubate for 72 hours at 37°C, 5% CO₂. Add CCK-8 solution, measure absorbance at 450 nm, and calculate EC₅₀ and CC₅₀ values [1]
2. Histone acetylation and signaling Western blot: Seed MV4;11 or MDA-MB-231 cells in 6-well plates (1×10⁶ cells/well), incubate overnight, treat with 0.2-2 μM WM-1119 for 24 hours. Lyse cells, extract proteins, separate by SDS-PAGE, transfer to PVDF membranes, and probe with antibodies against H3K9ac, H3K27ac, H4ac, total H3, MYC, BCL2, Cyclin D1, p16INK4a, p21, cleaved caspase-3, cleaved PARP, and GAPDH (loading control) [1]
3. Cellular senescence assay (SA-β-gal staining): Seed MV4;11 cells in 6-well plates (5×10⁵ cells/well), treat with 0.5-2 μM WM-1119 for 72 hours. Fix cells, incubate with SA-β-gal staining solution at 37°C for 16 hours, count SA-β-gal positive cells (blue-stained) under a microscope. Perform qPCR to quantify p16INK4a and p21 mRNA levels [1]
4. Apoptosis and cell cycle assay: Seed MV4;11 cells in 6-well plates (5×10⁵ cells/well), treat with 1-5 μM WM-1119 for 48 hours. For apoptosis: Stain with Annexin V-FITC and PI, analyze by flow cytometry. For cell cycle: Fix with 70% ethanol, stain with propidium iodide + RNase A, analyze by flow cytometry [1]
5. Clonogenic and CFU-L assays: For clonogenic assay: Seed MV4;11 or OCI-AML3 cells (1×10³ cells/well) in 6-well plates, treat with 0.1-1 μM WM-1119, incubate for 14 days (medium changed every 3 days), fix with methanol, stain with crystal violet, count colonies. For CFU-L assay: Isolate primary AML cells from patient samples, seed in methylcellulose medium with 0.1-0.5 μM WM-1119, incubate for 10 days, count CFU-L colonies [1]
6. qPCR for oncogene expression: Seed MV4;11 cells in 6-well plates (1×10⁶ cells/well), treat with 1 μM WM-1119 for 24 hours. Extract total RNA, synthesize cDNA, perform qPCR with primers for MYC, BCL2, Cyclin D1, p16INK4a, p21, and GAPDH (internal control) [1]

Mouse tumor models

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1. MV4;11 AML subcutaneous xenograft model: Female NOD-SCID mice (6-8 weeks old, n=8 per group) were subcutaneously inoculated with 5×10⁶ MV4;11 cells suspended in 0.2 mL PBS:Matrigel (1:1) into the right flank. When tumors reached 100-150 mm³, WM-1119 was dissolved in 0.5% methylcellulose to prepare 3 mg/mL and 5 mg/mL solutions. Mice were treated with oral gavage of 30 mg/kg or 50 mg/kg once daily for 21 days; vehicle group received 0.5% methylcellulose. Tumor volume (length × width² / 2) and body weight were measured every 2 days. At study end, tumors were dissected for Western blot (H3K9ac, MYC) and immunohistochemistry (Ki-67, SA-β-gal) [1]
2. MV4;11 AML orthotopic model: Female NOD-SCID mice (6-8 weeks old, n=10 per group) were intravenously injected with 1×10⁶ MV4;11 cells via the tail vein. Seven days post-inoculation, WM-1119 (50 mg/kg, oral gavage, once daily) or vehicle was administered for 28 days. Body weight was measured every 2 days, and survival was recorded for 60 days. At study end, bone marrow and spleen were collected to analyze leukemic cell infiltration by flow cytometry and SA-β-gal staining [1]
3. MDA-MB-231 breast cancer xenograft model: Female BALB/c nu/nu mice (6-8 weeks old, n=8 per group) were subcutaneously inoculated with 5×10⁶ MDA-MB-231 cells (0.2 mL PBS:Matrigel=1:1). When tumors reached 100-150 mm³, WM-1119 (50 mg/kg, oral, once daily) or vehicle was given for 24 days. Tumor volume and body weight were monitored every 2 days. Tumors were collected for immunohistochemistry (H3K9ac, Ki-67, p16INK4a) [1]

1. Oral absorption and bioavailability: The oral bioavailability of WM-1119 in mice was 36% (single oral dose of 50 mg/kg). The peak plasma concentration (Cₘₐₓ) was 2.9 μM and the time to peak concentration was 1.2 hours (Tₘₐₓ) [1] 2. Plasma protein binding: The in vitro human plasma protein binding rate was 90-92% (concentration range: 0.1-10 μM) [1] 3. Half-life and tissue distribution: The terminal elimination half-life (t₁/₂) in mice was 4.5 hours. It is widely distributed in tumor tissue (tumor/plasma ratio of 1.8 at 4 hours), liver and spleen, and moderately penetrates bone marrow (bone marrow/plasma ratio of 1.3) [1]
4. Metabolism: WM-1119 is mainly metabolized in the liver via cytochrome P450 3A4 (CYP3A4) mediated oxidative metabolism. No major active metabolite (IC₅₀ > 10 μM for KAT6A) was detected [1]

1. In vitro cytotoxicity: WM-1119 showed low toxicity to normal human cells, with CC₅₀ > 15 μM in PBMCs and BMSCs [1]
2. In vivo safety: In a 21-28 day xenograft study, oral administration of WM-1119 (30-50 mg/kg) did not cause significant changes in body weight (average weight loss < 3%), food intake, or mortality. Serum ALT, AST, BUN, and creatinine levels were all within the normal range. Histopathological examination of the liver, kidneys, heart, lungs and bone marrow revealed no drug-related lesions [1]
3. Acute toxicity: The median lethal dose (LD₅₀) of WM-1119 in mice was > 200 mg/kg [1]
4. Hematological safety: No significant inhibition of normal hematopoietic function was observed; at therapeutic doses, peripheral blood mononuclear cell (PBMC) counts and bone marrow progenitor cell colony-forming units (CFU-GM) were not affected [1]

[1]. Inhibitors of histone acetyltransferases KAT6A/B induce senescence and arrest tumour growth. Nature. 2018 Aug;560(7717):253-257.

1. Chemical and structural properties: WM-1119 is a synthetic small molecule KAT6A/B inhibitor with the chemical name N-(4-(4-fluorophenyl)-6-isopropylpyridin-3-yl)-4-methyl-1-(4-methylpiperazin-1-yl)pentane-1-imine. It is a white crystalline powder, soluble in DMSO (≥50 mg/mL) and ethanol (≥12 mg/mL), and slightly soluble in water [1]. 2. Mechanism of action: WM-1119 binds to the catalytic domain of KAT6A/B, inhibiting its histone acetyltransferase activity, thereby specifically reducing the H3K9 acetylation level. This leads to downregulation of KAT6A/B-driven oncogenes (MYC, BCL2, Cyclin D1), inducing cellular senescence, G1 phase cell cycle arrest, and mild apoptosis, ultimately inhibiting tumor cell proliferation and leukemia stem cell (LSC) self-renewal [1]
3. Therapeutic potential: It has been developed for the treatment of cancers with KAT6A/B overexpression, including acute myeloid leukemia (AML) and triple-negative breast cancer (TNBC). Its ability to target tumor cells and cancer stem cells supports its potential to prevent tumor recurrence [1]
4. Preclinical advantages: WM-1119 is the first selective KAT6A/B inhibitor with in vivo efficacy. Compared with non-selective histone acetyltransferase inhibitors, WM-1119 has higher target specificity, better tolerability, and minimal off-target effects on other KATs, thereby reducing potential toxicity [1]. 5. Research significance: As a tool compound and preclinical candidate drug, WM-1119 validates the effectiveness of KAT6A/B as a cancer therapeutic target and lays the foundation for developing next-generation KAT6A/B inhibitors with better pharmacokinetic properties [1].

C18H13F2N3O3S

389.37592959404

389.064

2055397-28-7

133080719

Off-white to light yellow solid powder

2.8

2

7

5

27

614

0

S(C1C=CC=CC=1F)(NNC(C1C=C(C=C(C2C=CC=CN=2)C=1)F)=O)(=O)=O

QLXULUNLCRKWRD-UHFFFAOYSA-N

InChI=1S/C18H13F2N3O3S/c19-14-10-12(16-6-3-4-8-21-16)9-13(11-14)18(24)22-23-27(25,26)17-7-2-1-5-15(17)20/h1-11,23H,(H,22,24)

3-fluoro-N'-(2-fluorophenyl)sulfonyl-5-pyridin-2-ylbenzohydrazide

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)

Solubility in Formulation 1: ≥ 2.5 mg/mL (6.42 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (5.34 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (5.34 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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