Milciclib

Catalog No.S2751 Batch:S275102

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Technical Data

Formula

C25H32N8O
Molecular Weight 460.57 CAS No. 802539-81-7
Solubility (25°C)* In vitro DMSO 17 mg/mL (36.91 mM)
Water Insoluble
Ethanol Insoluble
In vivo (Add solvents to the product individually and in order)
Clear solution
30%propylene glycol 5%Tween80 65%D5W
30.0mg/ml Taking the 1 mL working solution as an example, add 300 μL of 100 mg/ml clarified propylene glycol stock solution to 50 μL of Tween 80, mix evenly to clarify it; then continue to add 650 μL of D5W to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results. 
* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

Preparing Stock Solutions

Biological Activity

Description Milciclib is a potent, ATP-competitive CDK inhibitor for CDK2 with IC50 of 45 nM. It is >3-fold more selective for CDK2 than CDK1, 2, 4, 5, and 7, it's also an inhibitor of TRKA with IC50 of 53nM. Milciclib (PHA-848125) induces cell death through autophagy. Phase 2.
Targets
CDK2/CyclinA [1]
(Cell-free assay)		 TrkA [1]
(Cell-free assay)		 CDK7/CyclinH [1]
(Cell-free assay)		 CDK4/CyclinD1 [1]
(Cell-free assay)		 CDK5/p35 [1]
(Cell-free assay)		 View More
45 nM 53 nM 150 nM 160 nM 265 nM
In vitro PHA-848125 inhibits, although with lower potency, the activities of cyclin H/CDK7, cyclin D1/CDK4, p35/CDK5, as well as cyclin E/CDK2 and cyclin B/CDK1 with IC50 values of 0.15, 0.16, 0.265, 0.363, 0.398 μM, respectively. [1] Thropomyosin receptor kinase A is also inhibited by PHA-848125 in the same nanomolar range as CDKs. In the most PHA-848125-sensitive cell line, PHA-848125 induces a concentration-dependent G(1) arrest.[2] PHA-848125 also impairs phosphorylation of the retinoblastoma protein at CDK2 and CDK4 specific sites, reduces retinoblastoma protein and cyclin A levels, and increases p21(Cip1), p27(Kip1) and p53 expression. PHA-848125 is added to the cells 48 h after TMZ and cell growth is evaluated after 3 additional days of culture. [2] A drug combination of TMZ, BG and PHA-848125 induces an additive or synergistic effect on cell growth, depending on the cell line. [2] In the absence of BG, the combination is still more active than the single agents in cell lines moderately sensitive to TMZ, but comparable to PHA-848125 alone in the two TMZ-resistant cell lines. When TMZ plus BG are used in combination with PHA-848125 against cultured normal melanocytes, neither synergistic nor additive antiproliferative effects are observed.[2]
In vivo In the preclinical xenograft A2780 human ovarian carcinoma model, PHA-848125 reveals good efficacy and is well tolerated upon repeated daily treatments. Treatment of K-Ras(G12D)LA2 mice with PHA-848125 (40 mg/kg twice daily for 10 days) results in significant tumor growth inhibition at the end of the treatment and is accompanied by a reduction in the cell membrane turnover.[3] On the other hand, following oral administration, PHA-848125 shows significant antitumor activity in various human xenografts, carcinogen-induced tumors and in disseminated primary leukemia models; the plasma concentrations in rodents being in the same range as those found inhibiting cancer cell proliferation.[4]

Protocol (from reference)

Kinase Assay:[4]
  • Biochemical kinase inhibition assays

    Inhibition of kinase activity by PHA-848125 is assessed using a strong anion exchanger (Dowex 1X8 resin)–based assay in robotized format run on 384-well plates. In this assay, specific peptides or protein substrates are transphosphorylated by their specific kinase in the presence of ATP traced with [γ-33P]ATP using optimal buffers and cofactors. The potency of PHA-848125 toward CDKs and 38 additional kinases belonging to an in-house Kinase Selectivity Screening panel is evaluated, and the relevant IC50s are determined. For each enzyme, the absolute KM values for ATP and the specific substrate are calculated and each assay is then run at optimized ATP (2KM) and substrate (5KM) concentrations. This setting enables direct comparison of IC50 values of PHA-848125 across the panel for evaluation of its biochemical profile.
Cell Assay:[1]
  • Cell lines

    Melanoma cells

  • Concentrations

    5.7 mg/mL

  • Incubation Time

    2 hours

  • Method

    Melanoma cells are suspended in culture media at a concentration of 2 × 104 cells/mL, dispensed in 50 μL aliquots into flat-bottom 96-well plates and allowed to adhere overnight at 37 °C. Graded amounts of PHA-848125 or TMZ are then added to the wells (4 wells per point) in 50 μL of CM and the plates are incubated at 37 °C in a 5% CO2 humidified atmosphere for 5 days. The cytotoxic effects of TMZ are also evaluated in combination with the MGMT inhibitor BG. To this end, 10 μM BG is added to the plates 2 hours before TMZ and left in culture for the entire period of cell exposure to the drug. ContS1017rol groups are represented by untreated cells and cells treated with BG or DMSO alone. The growth of the cells treated with BG or DMSO alone does not differ from that of untreated cells. MGMT activity of BG-treated cells is undetectable 2 hours after the addition of the inhibitor PHA-848125 and remained essentially undetectable up to the end of the assay. Normal melanocytes are suspended in MGM at the concentration of 1.6 × 105 cells/mL, plated (50 μL/well) and exposed to TMZ + BG or to PHA-848125 as described for melanoma cells. At the end of the incubation period, cell growth is evaluated by the MTT assay. Briefly, 0.1 mg of MTT (in 20 μL of PBS) is added to each well and cells are incubated at 37 °C for 4 hours. Cells are then lysed with a buffer (0.1 mL/well) containing 20% SDS and 50% N,N-dimethylformamide, pH 4.7. After overnight incubation, the absorbance is read at 595 nm using a 3550-UV microplate reader. Cell sensitivity to drug treatment is expressed in terms of IC50 (drug concentration producing 50% inhibition of cell growth, calculated on the regression line in which absorbance values at 595 nm are plotted against the logarithm of drug concentration).
Animal Study:[3]
  • Animal Models

    K-Ras(G12D)LA2 mice

  • Dosages

    40 mg/kg twice daily for 10 days

  • Administration

    Oral administration

Customer Product Validation

Data from [Data independently produced by , , PLoS ONE, 2018, 13(2): e0193344]

Selleck's Milciclib has been cited by 18 publications

Spatial proteo-transcriptomic profiling reveals the molecular landscape of borderline ovarian tumors and their invasive progression [ medRxiv, 2023, 2023.11.13.23298409] PubMed: 38014221
Visual barcodes for clonal-multiplexing of live microscopy-based assays [ Nat Commun, 2022, 13(1):2725] PubMed: 35585055
Mechanisms Contributing to Animal Cell Size Homeostasis [ ProQuest, 2022, 29166130] PubMed: none
The role of drug efflux and uptake transporters ABCB1 (P-gp), ABCG2 (BCRP) and OATP1A/1B and of CYP3A4 in the pharmacokinetics of the CDK inhibitor milciclib [ Eur J Pharm Sci, 2021, 159:105740] PubMed: 33524505
Differential reprogramming of breast cancer subtypes in 3D cultures and implications for sensitivity to targeted therapy [ Sci Rep, 2021, 11(1):7259] PubMed: 33790333
The role of drug efflux and uptake transporters ABCB1 (P-gp), ABCG2 (BCRP) and OATP1A/1B and of CYP3A4 in the pharmacokinetics of the CDK inhibitor milciclib [ Eur J Pharm Sci, 2021, S0928-0987(21)00042-7] PubMed: 33524505
Infant high grade gliomas comprise multiple subgroups characterized by novel targetable gene fusions and favorable outcomes. [ Cancer Discov, 2020, 1 pii: CD-19-1030] PubMed: 32238360
A Patient-Derived Cell Atlas Informs Precision Targeting of Glioblastoma [ Cell Rep, 2020, 32(2):107897] PubMed: 32668248
Tethering of Telomeres to the Nuclear Envelope Is Mediated by SUN1-MAJIN and Possibly Promoted by SPDYA-CDK2 During Meiosis [ Front Cell Dev Biol, 2020, 8:845] PubMed: 33015044
Development and validation of an LC-MS/MS method for the quantitative analysis of milciclib in human and mouse plasma, mouse tissue homogenates and tissue culture medium [ J Pharm Biomed Anal, 2020, 190:113516] PubMed: 32784092

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