CDK2 (IC50 = 3.27 nM); CDK9 (IC50 = 5.78 nM); CDK1 (IC50 = 8.17 nM); CDK4 (IC50 = 8.18 nM); CDK6 (IC50 = 37.68 nM); CDK7 (IC50 = 134.26 nM); CDK1 (Ki = 4 nM); CDK2 (Ki = 3 nM); CDK9 (Ki = 4 nM); CDK7 (Ki = 91 nM)
CDK9 (IC50 = 5.78 ± 0.05 nM) [1]
CDK2 (IC50 = 3.27 ± 0.15 nM) [1]
CDK1 (IC50 = 8.17 ± 0.07 nM) [1]
CDK4 (IC50 = 8.18 ± 0.58 nM) [1]
CDK6 (IC50 = 37.68 ± 0.81 nM) [1]
CDK7 (IC50 = 134.26 ± 1.29 nM) [1]

CDKI-73 primarily targets CDK9 and downregulates the expression of anti-apoptotic proteins Bcl-2, Mcl-1, and XIAP, causing cancer cells to undergo apoptosis. The bone marrow cells of healthy donors, however, are comparatively less toxic to it.[2]

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CDKI-73 inhibited the proliferation of six acute leukemia cell lines (CCRF-CEM, U937, Molt-4, HL-60, THP-1, Jurkat) with IC50 values ranging from 100 to 200 nM (mean IC50 = 163 ± 48 nM), comparable to flavopiridol (mean IC50 = 147 ± 25 nM). [1]
CDKI-73 decreased the survival of primary lymphocytes derived from 21 ALL pediatric patients with a mean LD50 of 67 nM, and from 15 AML pediatric patients with a mean LD50 of 102 nM. [1]
In a series of 41 additional kinase counter screens, CDKI-73 was selective for the CDK family. Only GSK3β activity was inhibited by 55% at 100 nM. [1]
In HL-60 and CCRF-CEM cells, CDKI-73 (0.1-0.4 μM) dose-dependently induced apoptosis, with approximately 20% of HL-60 cells undergoing apoptosis at 0.1 μM and 80% at 0.4 μM after 24 hours. [1]
CDKI-73 (0.2 μM) significantly increased apoptotic cells in HL-60 cells within 12 hours, with the apoptotic population continuing to increase over 48 hours. [1]
CDKI-73 (0.2 and 0.4 μM) induced cell apoptosis in patient-derived primary lymphocytes. [1]
In HL-60, CCRF-CEM, and Molt-4 cells, combined application of CDKI-73 with the Bcl-2 inhibitor ABT-199 (1 or 2 μM) markedly increased cell apoptosis, with combination index (CI) values of 0.44 ± 0.03, 0.61 ± 0.17, and 0.54 ± 0.13, respectively, indicating synergism (CI < 0.8). Maximal decrease in XIAP levels was observed with the combination. [1]

CDKI-73 targets CDK9 to downregulate anti-apoptotic proteins. It is highly effective against MLL-AML MV4-11 xenografts and is orally bioavailable in vivo.[2]

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In vitro kinase assay for CDKs: The inhibition of CDKs (CDK1, CDK2, CDK4, CDK6, CDK7, CDK9) by CDKI-73 was measured using mobility shift assays. [1]
In vitro kinase assay for non-CDK kinases: The inhibition of 41 non-CDK kinases (including ABL, AKT1, AKT2, Aurora A, GSK3β, etc.) by CDKI-73 was measured at concentrations of 10 μM and 100 nM using enzyme immunosorbent assays or by a commercial service provider. [1]

MV4-11 (1 × 10⁵/well) cells are seeded and incubated for an entire night at 37ºC with 5% CO2. Cells are gathered and centrifuged (300 g, 5 min) following a 24- or 48-hour incubation period with the indicated concentrations of CDKI-73. The pellets are resuspended in cold 70% ethanol for the cell cycle assay, fixed on ice for 15 minutes, and then centrifuged (300 g, 5 min). Following a 1-hour incubation period at 37ºC with 200 μL staining solution, the collected pellets are examined using a Gallios flow cytometer. Cells are taken and processed in accordance with the directions provided in the commercial annexin V-FITC/PI kit to measure apoptosis.

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Cell proliferation assay (Resazurin): Cells were seeded in 96-well plates and exposed to increasing doses of CDKI-73 or flavopiridol for 72 hours. 10% resazurin solution (1.5 mg/mL) was then added, and cells were incubated for another 2-4 hours. Fluorescence intensity was detected at excitation and emission wavelengths of 540±35 nm and 590±35 nm, respectively. IC50 values were calculated using non-linear 4-parameter regression analysis. [1]
Apoptosis assay (Annexin V/PI double staining): Cells (5 × 10⁵ per well in 6-well plates) were incubated with various concentrations of CDKI-73 for 24 hours or with 0.2 μM of the test compounds for 12, 24, or 48 hours. Samples were collected, stained with Annexin V-FITC and propidium iodide (PI), and detected using flow cytometry. [1]
Western blotting: Cells were cultured with various concentrations of CDKI-73 for 2 or 24 hours, harvested, washed, and lysed. Protein lysates were resolved by SDS-PAGE, transferred to nitrocellulose membranes, and probed with primary antibodies against RNA pol II, p-RNA pol II (ser2), p-RNA pol II (ser5), RB, p-RB, PP1α, p-PP1α, β-tubulin, PARP, XIAP, Mcl-1, Bcl-2, and cleaved-caspase3. Proteins were detected using enhanced chemiluminescence. [1]
Real-time PCR: Cells were left untreated or exposed to CDKI-73 for 6 hours. Total RNA was extracted, reverse transcribed into cDNA, and real-time PCR was performed using SYBR-Green Master mix with specific primers for Mcl-1, XIAP, and GAPDH. [1]
Primary lymphocyte isolation: Bone marrow samples from ALL and AML pediatric patients were collected. Mononuclear cells were separated using density gradient centrifugation. Enriched cells were maintained in RPMI-1640 medium with 10% FBS and seeded for viability tests (72 h treatment), apoptosis detection (48 h treatment), or protein expression analysis (12 h treatment) with CDKI-73. [1]

female nude (nu/nu) Balb/C mice aged 6–8 weeks
25 mg/kg
Oral gavage

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[1]. Acta Pharmacol Sin . 2016 Nov;37(11):1481-1489.

[2]. Invest New Drugs . 2019 Aug;37(4):625-635.

CDKI-73 (also known as LS-007) is a potent CDK9 inhibitor that exhibits potent antitumor activity against chronic lymphocytic leukemia and ovarian cancer cells. [1]
CDKI-73 induces cell apoptosis predominantly through CDK9 inhibition-related dephosphorylation at the serine 2 residue of RNA polymerase II and the corresponding depletion of short-lived anti-apoptotic proteins, especially Mcl-1 and XIAP. [1]
The combination of CDKI-73 with the Bcl-2 inhibitor ABT-199 (venetoclax) shows remarkable synergy in acute leukemia cells. [1]

C15H15FN6O2S2

394.44

394.068

C, 45.68; H, 3.83; F, 4.82; N, 21.31; O, 8.11; S, 16.26

1421693-22-2

1421693-22-2

71561915

Light yellow to yellow solid powder

4.407

3

10

5

26

577

0

S(C1=C([H])C([H])=C([H])C(=C1[H])N([H])C1=NC([H])=C(C(C2=C(C([H])([H])[H])N=C(N([H])C([H])([H])[H])S2)=N1)F)(N([H])[H])(=O)=O

GAIOPWBQKZMUNO-UHFFFAOYSA-N

InChI=1S/C15H15FN6O2S2/c1-8-13(25-15(18-2)20-8)12-11(16)7-19-14(22-12)21-9-4-3-5-10(6-9)26(17,23)24/h3-7H,1-2H3,(H,18,20)(H2,17,23,24)(H,19,21,22)

3-[[5-fluoro-4-[4-methyl-2-(methylamino)-1,3-thiazol-5-yl]pyrimidin-2-yl]amino]benzenesulfonamide

CDKI73; CDKI-73; CDKI 73; asnuciclib [INN]; asnuciclib

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture.

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

DMSO: ~79 mg/mL (~200.3 mM)
Ethanol: ~1.5 mg/mL (~3.8 mM)

Solubility in Formulation 1: ≥ 2.67 mg/mL (6.77 mM) (saturation unknown) in 10% DMSO + 40% PEG300 +5% Tween-80 + 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 26.7 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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 (Please use freshly prepared in vivo formulations for optimal results.)