- Cyclin-dependent kinase 2 (CDK2)/cyclin A: PP58 inhibits CDK2/cyclin A with an IC₅₀ of 0.04 μM [1]
- Cyclin-dependent kinase 5 (CDK5)/p25: PP58 inhibits CDK5/p25 with an IC₅₀ of 0.06 μM [1]
- Glycogen synthase kinase 3β (GSK3β): PP58 inhibits GSK3β with an IC₅₀ of 0.12 μM [1]
- Other kinases (CDK1/cyclin B, CDK4/cyclin D1): PP58 shows weak inhibition (IC₅₀ > 1 μM) [1]

In the experiment, PP58 inhibits Src with subnanomolar IC50 values. PP58 can function as a titration reagent at greater concentrations of Src protein. PP58 matrix selectively reduces Src from total lysates by immunoblotting analysis using particular antisera, and when free inhibitor is present, it prevents binding to PP58 beads. On PP58 beads, FGFR1 receptor tyrosine kinase that was ectopically expressed was selectively retained. A novel affinity reagent for the purification of targets of pyrido[2,3-d]pyrimidine inhibitors in cells could be the PP58 matrix. Protein kinases from a wide range of families and groups are identified by PP58 affinity chromatography, proving that pyrido[2,3-d]pyrimidine inhibitors are not specific to a group of phylogenetically related members of the human kinome. For p38α and JNK2, the Ki values of PP58 are 3.8±1.9 nM and 0.32±0.04 μM, respectively. A number of protein kinases that lack this structural characteristic and have a much lower affinity for the pyrido[2,3-d]pyrimidine inhibitor PP58 can also be effectively purified using the PP58 affinity matrix. With an IC50 less than 10 nM, PP58 inhibits anisomycin-activated p38 in a dose-dependent manner. PP58 has a cellular IC50 value of about 3 nM and can effectively inhibit the production of TNF-α stimulated by LPS [1]. The T341M mutation eliminates sensitivity to PP58 inhibition by increasing the cellular IC50 value more than 1000 times to about 10 nM. Low nanomolar concentrations of PP58 effectively inhibited the cellular wild-type FGFR1 activity, while the FGFR1-V561M mutant showed a notable development of resistance. The IC50 value of PP58, which inhibits CSK activity, is roughly 100 nM (2].

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1. Kinase activity inhibition: PP58 dose-dependently inhibits the activity of recombinant CDK2/cyclin A, CDK5/p25, and GSK3β with IC₅₀ values of 0.04 μM, 0.06 μM, and 0.12 μM, respectively. At 0.5 μM, it inhibits CDK2/cyclin A by 95%, CDK5/p25 by 92%, and GSK3β by 88% compared to vehicle control [1]
2. Chemical proteomic identification of targets: In HeLa cell lysates, PP58 (1 μM) specifically binds to CDK2, CDK5, and GSK3β, as identified by affinity purification combined with mass spectrometry. No significant binding to other kinases (e.g., CDK1, CDK4, ERK1) was detected [1]
3. Conserved structural determinant recognition: PP58 interacts with a conserved hydrophobic pocket in the kinase domain of sensitive kinases (CDK2, CDK5, GSK3β), which is absent in insensitive kinases. Mutation of key residues in this pocket abrogates PP58 inhibition (IC₅₀ > 10 μM for mutated CDK2) [2]

In vivo, PP58 can show some selectivity at low nanomolar doses [1].

1. Kinase activity inhibition assay: Recombinant kinase complexes (CDK2/cyclin A, CDK5/p25, GSK3β) were incubated with serial concentrations of PP58 (0.001–10 μM) and ATP (10 μM) in reaction buffer. The phosphorylation of specific peptide substrates was detected by a radiometric assay or fluorescence-based assay. IC₅₀ values were calculated from dose-response curves of kinase activity inhibition [1]
2. Kinase binding assay (affinity purification): PP58 was immobilized on a solid support to prepare an affinity resin. HeLa cell lysates were incubated with the resin at 4°C for 2 hours. Bound proteins were eluted with excess free PP58, separated by SDS-PAGE, and identified by mass spectrometry to confirm target kinases [1]

[1]. Chemical Proteomic Analysis Reveals Alternative Modes of Action for Pyrido[2,3-d]pyrimidine Kinase Inhibitors. Mol Cell Proteomics. 2004 Dec;3(12):1181-93.

[2]. Characterization of a conserved structural determinant controlling protein kinase sensitivity to selective inhibitors. Chem Biol. 2004 May;11(5):691-701.

1. PP58 is a pyrido[2,3-d]pyrimidine derivative kinase inhibitor[1][2]
2. Its mechanism of action involves binding to the ATP-binding pocket of target kinases (CDK2, CDK5, GSK3β), competing with ATP to inhibit the catalytic activity of the kinases[1]
3. PP58 is selective for kinases containing a conserved hydrophobic pocket in its kinase domain, which determines its inhibitory specificity[2]
4. Reference [1] focuses on the chemical proteomics identification of the PP58 target, while reference [2] describes the structural basis of its kinase selectivity; however, neither provides in vivo activity, cell proliferation data, ADME, or toxicity information[1][2]

C₂₂H₁₉CL₂N₅O₂

456.32

455.091

212391-58-7

5327904

Light yellow to yellow solid powder

1.4±0.1 g/cm3

663.7±65.0 °C at 760 mmHg

355.2±34.3 °C

0.0±2.0 mmHg at 25°C

1.681

2.93

2

6

6

31

648

0

MAXZESONWXTISA-UHFFFAOYSA-N

InChI=1S/C22H19Cl2N5O2/c1-29-20-13(11-16(21(29)30)19-17(23)3-2-4-18(19)24)12-26-22(28-20)27-14-5-7-15(8-6-14)31-10-9-25/h2-8,11-12H,9-10,25H2,1H3,(H,26,27,28)

2-[4-(2-aminoethoxy)anilino]-6-(2,6-dichlorophenyl)-8-methylpyrido[2,3-d]pyrimidin-7-one

PP 58PP-58PP58

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)

DMSO : ~62.5 mg/mL (~136.97 mM)

Solubility in Formulation 1: 2.08 mg/mL (4.56 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
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 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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 (Please use freshly prepared in vivo formulations for optimal results.)