Synthesis and evaluation of 3-phenylpyrazolo[3,4-d]pyrimidine-peptide conjugates as Src kinase inhibitors

3-Phenylpyrazolo[3,4-d]pyrimidine (PhPP) derivatives substituted with an alkyl or aryl carboxylic acid at the N1-endocyclic amine, such as PhPP-CH ₂COOH (IC₅₀=250 μm), and peptides Ac-CIYKYY (IC ₅₀=400 μm) and Ac-YIYGSFK (IC₅₀=570 μm) were weak inhibitors of polyE₄Y phosphorylation by active c-Src. A series of PhPP-peptide conjugates were synthesized using PhPP as an ATP mimic and CIYKYY or YIYGSFK as a peptide substrate to improve the inhibitory potency against active c-Src kinase. PhPP derivatives were attached to the Nterminus or the side chain of amino acids in the peptide template. Two N-terminal substituted conjugates, PhPP-CH₂CO-CIYKYY (IC₅₀=0.38 μm) and PhPP-CH₂CO-YIYGSFK (IC₅₀=2.7 μm), inhibited the polyE₄Y phosphorylation by active c-Src significantly higher than that of the parent compounds. The conjugation of PhPP with the peptides produced a synergistic inhibition effect possibly through creation of favorable interactions between the conjugate and the kinase domain as shown by molecular modeling studies.