KI-CDK9d-32
Based on 1 Customer Validation
KI-CDK9d-32 is a selective CDK9 PROTAC degrader (IC50 = 3 nM; DC50 = 0.89 nM). KI-CDK9d-32 induces CDK9 degradation via the ubiquitin-proteasome pathway to abrogates CDK9 enzymatic and scaffolding functions, downregulates MYC expression and MYC-dependent signaling, represses TNF-alpha signaling pathways activity and pre-rRNA levels. KI-CDK9d-32 disrupts nucleolar homeostasis, blocks cell cycle progression and cellular translation by reducing 4EBP1 phosphorylation, and elicits cancer cell cytotoxicity in a CRBN-dependent manner, while high ABCB1 activity attenuates the efficacy. KI-CDK9d-32 can be used for the research of acute lymphoblastic leukemia, rhabdomyosarcoma, pancreatic adenocarcinoma.
(Pink: CDK9 ligand (HY-153718); Blue: Cereblon ligand (HY-163233); Black: linker (HY-W011657)).
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研究用途以外に使用した場合、当社は一切の責任を負いかねます。
- 純度 : 99.03%
- CAS 番号: 3054009-82-1
- 分子式: C39H45N9O4
- 分子量:703.83
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保管条件:Powder -20°C, 3 years , 4°C, 2 years ; In solvent -80°C, 6 months , -20°C, 1 month
To place orders, for customer services and technical support, please contact: MedChemExpress USA
Tel: 609-228-6898 E-mail: [email protected] [email protected]
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生物活性
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化学情報
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溶剤 & 溶解度
- プロトコル
- 純度とドキュメンテーション
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生物活性
製品説明
IC50 & Target
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CDK9 0.89 nM (DC50) |
CDK9 3 nM (IC50) |
体外実験
KI-CDK9d-32 (0.004-5000 nM; 1-12 h) potently degrades CDK9 in MOLT-4 cells with a DC50 of 0.89 nM after 4 h, achieves near-complete degradation sustained for at least 12 h, and exhibits a maximal degradation rate of 1.04 h-1 at 126 nM[1].
KI-CDK9d-32 (50 nM; 4 h) reduces CDK9 protein levels 5-fold and MYC protein levels 3-fold in MOLT-4 cells, along with downregulating MYC-regulated kinases and disrupting cell cycle and ribosome biogenesis pathways[1].
KI-CDK9d-32 (50 nM; 4 h) selectively inhibits CDK9-mediated phosphorylation in MOLT-4 cells, reduces 4EBP1 phosphorylation to inhibit translation, and disrupts phosphosites linked to mRNA and ribosome-related processes[1].
KI-CDK9d-32 (15-125 nM; 2-8 h) dose- and time-dependently suppresses MYC mRNA levels in MOLT-4 cells, bypassing the compensatory MYC increase seen with CDK9 inhibition, and reduces CDK9 mRNA levels by 5-fold at 8 h[1].
KI-CDK9d-32 (15 nM; 2-8 h) induces rapid, sustained downregulation of MYC mRNA (over 6-fold relative to inhibition) in MOLT-4 cells, downregulates TNF-α signaling via NF-κB across MOLT-4, PSN-1, and RH-4 cells, and disrupts MYC target pathways linked to ribosome biogenesis[1].
KI-CDK9d-32 (up to 1.5 μM; 120 h) has a mean IC50 of 82.6 nM across ~800 human cancer cell lines, with sensitivity driven by CRBN expression and resistance driven by ABCB1 expression[1].
KI-CDK9d-32 (2 h) induces rapid (within 2 h) disruption of nucleolar homeostasis in HeLa cells by compromising the NPM1-marked nucleolar rim, without affecting the DDX21-marked core granular compartment[1].
KI-CDK9d-32 (0.1-10 μM) potently inhibits CDK9/cyclin T1 with a biochemical IC50 of 3 nM at 10 mM ATP, exhibits >1403-fold selectivity over most tested kinases, and inhibits ~90% of CDK9 activity at 100 nM with no off-target kinase inhibition[1].
KI-CDK9d-32 (10-100 nM; 4 h) destabilizes nucleolar homeostasis in HeLa cells[1].
KI-CDK9d-32 (100 nM; 4 h) induces significant disruption and dissolution of nucleolar structure in HeLa cells[1].
KI-CDK9d-32 (100 nM; 4 h) significantly reduces pre-rRNA (5'-ETS) levels in HeLa cells[1].
KI-CDK9d-32 (15 nM; 4-8 h) significantly downregulates multiple core regulatory circuitry gene transcripts in MOLT-4, PSN-1, and RH-4 cells, with stronger effects observed at 8 h[1].
KI-CDK9d-32 (50 nM; 4 h) induces potent degradation of CDK9 (fold change -4.81), with no detectable degradation of other tested kinase targets in a quantitative mass spectrometry assay[1].
KI-CDK9d-32 (50 nM; 4 h) induces degradation of CDK9 and c-MYC, and reduces pSer2, POLR2A levels in PSN-1, RH-4, and HeLa cells, with degradation reversed by MG-132 (HY-13259) co-treatment[1].
KI-CDK9d-32 (50 nM; 4 h) does not affect CDK7 protein levels in MOLT-4 cells, demonstrating selectivity for CDK9 over its homologue CDK7[1].
KI-CDK9d-32 (15-125 nM; 4 h) induces dose-dependent degradation of CDK9 and MYC in MOLT-4 cells[1].
KI-CDK9d-32 (15 nM; 4 h) induces significant enrichment of HALLMARK MYC TARGETS V2 and HALLMARK TNFA via NFKB gene sets in MOLT-4 cells, enrichment of these sets in PSN-1 cells, and enrichment of HALLMARK DNA REPAIR in RH-4 cells[1].
KI-CDK9d-32 (0.69 nM-1.5 µM) reduces viability of ~800 tested cancer cell lines in a dose-dependent manner, with cell line sensitivity correlated with CRBN expression and resistance correlated with ABCB1 expression, as measured via the Broad Institute PRISM high-throughput platform[1].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only. Further protocols information, click here.
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Cell Line:MOLT-4 human acute lymphoblastic leukemia cells
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Concentration:15; 50; 125 nM
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Incubation Time:2, 4, 8 h
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Result:Significantly suppressed MYC mRNA transcription in a dose- and time-dependent manner at all tested concentrations, in contrast to the compensatory MYC mRNA increase induced by CDK9 inhibition with KB-0742.
Reduced CDK9 mRNA levels by 5-fold at 8 h, while effects on CDK9 transcript levels were not clearly distinct from inhibition at early time points.
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Cell Line:MOLT-4 human acute lymphoblastic leukemia cells, PSN-1 human pancreatic adenocarcinoma cells, RH-4 human rhabdomyosarcoma cells
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Concentration:Up to 500 nM
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Incubation Time:72, 120 h
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Result:Exhibited potent cytotoxicity with IC50 values of 2.8 nM (72 h) and 1.70 nM (120 h) in MOLT-4 cells; 21.3 nM (72 h) and 73.85 nM (120 h) in PSN-1 cells; 52.9 nM (72 h) and 81.94 nM (120 h) in RH-4 cells.
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Cell Line:PSN-1, RH-4, HeLa cells
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Concentration:50 nM
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Incubation Time:4 h
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Result:Induced degradation of CDK9 and c-MYC, and reduced levels of pSer2, POLR2A in PSN-1, RH-4, and HeLa cells.
Degradation was rescued by co-treatment with MG-132.
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Cell Line:MOLT-4 cells
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Concentration:50 nM
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Incubation Time:4 h
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Result:Did not alter CDK7 protein levels relative to control.
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Cell Line:unspecified cell line
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Concentration:50 nM
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Incubation Time:4 h
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Result:Reduced relative expression of CDK9, MYC, AURKA, and PLK1 compared to DMSO control.
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Cell Line:MOLT-4 cells
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Concentration:15; 50; 125 nM
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Incubation Time:4 h
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Result:Induced degradation of CDK9 and MYC at all tested concentrations.
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Cell Line:HeLa cells
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Concentration:10; 50; 100 nM
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Incubation Time:4 h
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Result:Compromised nucleolar rim structure and altered NPM1 and DDX21 localization at all tested concentrations, with effects visible at the lowest concentration of 10 nM.
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Cell Line:HeLa cells
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Concentration:100 nM
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Incubation Time:4 h
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Result:Induced a significant reduction in pre-rRNA (5'-ETS) levels compared to DMSO control.
化学情報
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CAS 番号 3054009-82-1
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性状 Solid
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分子量 703.83
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分子式 C39H45N9O4
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Color White to off-white
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SMILES
O=C(N[C@H]1C[C@H](NC2=CC(CCC)=NC3=CC=NN23)CC1)C4=CC=C(CN5CCN(C6=CC7=C(C(N(C(CC8)C(NC8=O)=O)C7)=O)C=C6)CC5)C=C4
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輸送条件
Room temperature in continental US; may vary elsewhere.
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保管条件
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month
溶剤 & 溶解度
体外:
DMSO : 100 mg/mL (142.08 mM; Need ultrasonic; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
濃度 (開始) × 体積 (開始) = 濃度 (終了) × 体積 (終了)
体内:
Select the appropriate dissolution method based on your experimental animal and administration route.
- For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
- To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for In Vivo experiments, it is recommended to prepare freshly and use it on the same day.
- The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.
Add each solvent one by one: 10% DMSO 40% PEG300 5% Tween-80 45% Saline
Solubility: ≥ 2.5 mg/mL (3.55 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.
Preparation of Saline: Dissolve 0.9 g sodium chloride in ddH₂O and dilute to 100 mL to obtain a clear Saline solution.
Add each solvent one by one: 10% DMSO 90% (20% SBE-β-CD in Saline)
Solubility: ≥ 2.5 mg/mL (3.55 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 900 μL 20% SBE-β-CD in Saline, and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C, storage for one week): 2 g SBE-β-CD powder is dissolved in 10 mL Saline, completely dissolve until clear.
In Vivo Dissolution Calculator
Please enter the basic information of animal experiments:
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Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.
Please enter your animal formula composition:
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%DMSO +
Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.
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%+
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+%Tween-80 + +
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%Saline +
The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).
Working solution concentration: 0.22 mg/mL
Method for preparing stock solution: mg drug dissolved in μL DMSO. Stock solution concentration: mg/mL.
1. Take μL DMSO stock solution;
2. Add μL .
μL , mix evenly;
3. Then add μL Tween 80, mix evenly;
4. Then add μL
Please ensure that the stock solution in the first step is dissolved to a clear state, and add co-solvents in sequence. You can use ultrasonic heating (ultrasonic cleaner, recommended frequency 20-40 kHz), vortexing, etc. to assist dissolution.
プロトコル
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Kinase activity and phosphorylation assays
Kinase activity assays measure the ability of kinases to transfer phosphate groups from ATP to specific substrates, while phosphorylation assays detect the presence and levels of phosphorylated proteins. Common methods include radiolabeled ATP incorporation (e. g. ,), ADP release detection via bioluminescence (e. g. ,[3]), enzyme-linked immunosorbent assays (ELISA) for phospho-specific epitopes (e. g. ,[6]), and microtiter-based formats for high-throughput screening (e. g. ,[8]). The ADP-Glo assay quantifies kinase activity by measuring ADP produced during phosphorylation using a luciferase-based system. Radiometric assays involve autoradiography or scintillation counting after incorporation of 32P-labeled ATP into substrate proteins. ELISA-based approaches rely on phospho-specific antibodies to detect activated kinases in cell lysates or purified samples.
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RNA extraction experimental
By lysing cells, releasing RNA, and removing impurities such as proteins and DNA, high-purity RNA products are finally obtained. The commonly used traditional method is the guanidine isothiocyanate/phenol/chloroform method (Trizol), which is suitable for a variety of animal materials including animal tissues, microorganisms, cultured cells, etc., and most plant materials.
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Western Blot
Western blotting (WB) is a commonly used experimental method in molecular biology, biochemistry, and immunogenetics for identifying and quantifying target proteins. It combines gel electrophoresis with immunoassay, enabling researchers to analyze protein expression, post-translational modifications, and molecular weight.
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Cell Cytotoxicity Assay
Cytotoxicity assays are usually based on the assessment of cell membrane damage, which can also be indirectly detected by measuring cell viability. Detection methods include MTT assay, CKK-8 assay, LDH assay and ATP assay, etc.
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Flow cytometric DNA-content cell-cycle staining
Flow cytometric DNA-content cell-cycle staining measures the fluorescence intensity of DNA-bound fluorochromes in single cells or nuclei to estimate DNA content distributions, allowing assignment of populations to G0/G1, S, and G2/M phases by DNA histogram deconvolution. Propidium iodide (PI) intercalates into DNA, and PI fluorescence is proportional to cellular DNA content when staining is performed under conditions that make DNA accessible and minimize non-DNA signal. Cells with G2/M DNA content are expected to show approximately twice the fluorescence intensity of G0/G1 cells, while S-phase cells occupy intermediate fluorescence values. PI-based DNA-content analysis can also detect cells with fractional DNA content, often reported as sub-G1, when DNA fragmentation and extraction during staining reduce retained DNA signal in apoptotic cells. DAPI is an alternative DNA fluorochrome for univariate DNA-content analysis, while bivariate approaches combining DNA content with proliferation
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Mammalian live/dead viability and cytotoxicity staining
Live/dead viability and cytotoxicity staining assays are based on the simultaneous detection of intracellular esterase activity in metabolically active (viable) cells and membrane integrity loss in non-viable cells. In commonly used dual-staining approaches, membrane-permeant fluorogenic substrates are converted by intracellular esterases into fluorescent products in live cells, while impermeant DNA-binding dyes selectively enter cells with compromised plasma membranes and label nucleic acids in dead or dying cells, enabling discrimination between viable and non-viable populations by fluorescence microscopy or flow cytometry.
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BrdU Incorporation Assay
Bromodeoxyuridine (BrdU) incorporation assay is based on the principle that BrdU, a thymidine analog, is incorporated into newly synthesized DNA during the S phase of the cell cycle, thereby serving as a marker of DNA replication and cellular proliferation. Incorporated BrdU can be detected using anti-BrdU antibodies following DNA denaturation, enabling visualization or quantification of proliferating cells through immunochemical detection methods such as immunofluorescence or immunohistochemistry.
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Protocol for Cell Cycle
Cell-cycle analysis by flow cytometry measures DNA content in single cells to estimate the fraction of cells in G0/G1, S, and G2/M phases. Propidium iodide intercalates into DNA, and after RNA removal with RNase, fluorescence intensity reflects cellular DNA content: 2N cells are assigned to G0/G1, cells between 2N and 4N to S phase, and 4N cells to G2/M. DNA-content analysis alone cannot reliably separate G0 from G1 or G2 from M. Ki-67 can distinguish quiescent G0 cells from cycling cells, EdU or BrdU incorporation marks active DNA synthesis in S phase, and phospho-histone H3 staining identifies mitotic cells within the 4N population.
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Protocol for Kinase activity and phosphorylation assays
Kinase activity assays measure transfer of phosphate from ATP to a protein or peptide substrate, generating phosphorylated substrate, ADP, or incorporated radiolabeled phosphate as the readout; phosphorylation assays measure site-specific phosphorylation in cells or tissues as a proxy for kinase-pathway activation, inhibition, or substrate regulation. Phosphorylation can be detected by phospho-specific Western blot, immunoprecipitation kinase assay, phospho-immunofluorescence, phospho-flow cytometry, luminescent ADP detection, radiolabeled ATP incorporation, or reporter-based pathway assays, and these readouts can be applied to cancer cells, primary neurons, mouse tumors, organoids, inflammatory macrophages, ferroptosis studies, and mitophagy studies when the kinase target is biologically relevant.
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Cell Viability Determination by MTT Colorimetric Assay
The following protocol uses the MTT colorimetric assay as a classic literature-established method for assessing cell viability/metabolic activity in cultured mammalian cells. MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] is reduced by metabolically active cells to a colored formazan product; the amount of formazan is quantified spectrophotometrically and provides an indirect measure of metabolically active viable cells. Importantly, MTT reduction reflects cellular oxidoreductase/metabolic activity rather than an absolute direct count of living cells, so changes in cellular metabolism can alter the signal independently of cell number.
純度とドキュメンテーション
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データシート (283 KB)
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SDS (254 KB)
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- Français - FR (254 KB)
- Deutsch - DE (254 KB)
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取扱説明書 (2659 KB)
参考文献
Complete Stock Solution Preparation Table
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
| Optional Solvent | Concentration Solvent Mass | 1 mg | 5 mg | 10 mg | 25 mg |
|---|---|---|---|---|---|
| DMSO | 1 mM | 1.4208 mL | 7.1040 mL | 14.2080 mL | 35.5199 mL |
| 5 mM | 0.2842 mL | 1.4208 mL | 2.8416 mL | 7.1040 mL | |
| 10 mM | 0.1421 mL | 0.7104 mL | 1.4208 mL | 3.5520 mL | |
| 15 mM | 0.0947 mL | 0.4736 mL | 0.9472 mL | 2.3680 mL | |
| 20 mM | 0.0710 mL | 0.3552 mL | 0.7104 mL | 1.7760 mL | |
| 25 mM | 0.0568 mL | 0.2842 mL | 0.5683 mL | 1.4208 mL | |
| 30 mM | 0.0474 mL | 0.2368 mL | 0.4736 mL | 1.1840 mL | |
| 40 mM | 0.0355 mL | 0.1776 mL | 0.3552 mL | 0.8880 mL | |
| 50 mM | 0.0284 mL | 0.1421 mL | 0.2842 mL | 0.7104 mL | |
| 60 mM | 0.0237 mL | 0.1184 mL | 0.2368 mL | 0.5920 mL | |
| 80 mM | 0.0178 mL | 0.0888 mL | 0.1776 mL | 0.4440 mL | |
| 100 mM | 0.0142 mL | 0.0710 mL | 0.1421 mL | 0.3552 mL |
KI-CDK9d-32 関連分類
HY-173523 関連分類
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