Pyrazolo[3,4-d]pyrimidine-based dual EGFR T790M/HER2 inhibitors: Design, synthesis, structure-activity relationship and biological activity as potential antitumor and anticonvulsant agents

Abstract

A new series of pyrazolo[3,4-d]pyrimidine/triazine hybrids 6a-r was designed as antitumor and anticonvulsant agents. All the prepared compounds were evaluated against colon (HCT-116), breast (MCF-7) and normal human fibroblast (WI38) cell lines. The most potent derivatives against HCT-116 and MCF-7 cells were 6o and 6q, with IC₅₀ = 4.80 and 6.50 nM, respectively, when compared to lapatinib, the reference drug (IC₅₀ = 12.00 and 21.00 nM, on HCT-116 and MCF-7, sequentially). All other derivatives exhibited good to moderate cytotoxic activity. Four compounds 6f, 6j, 6o and 6q were evaluated for their EGFR T790M/HER2 inhibitory activity. They revealed 81.81-65.70% and 86.66-54.49% inhibitory activity against EGFR T790M and HER2 in a sequent. The most potent derivatives 6o and 6q were further estimated for cell cycle analysis showing pre G1 apoptotic activity and cell growth arrest at G2/M phase. Apoptotic marker proteins expression levels (caspase-3/7/9, Bax and Bcl-2) were measured for 6o and 6q. They showed pro-apoptotic effect by increasing caspase-3/7/9 protein levels and Bax/Bcl-2 ratio. Moreover, anticonvulsant activity for the prepared compounds 6a-r were evaluated in vivo using lithium-pilocarpine mice model of Status Epilepticus. EEG changes where recorded and MDA, GSH, GABA and glutamate were measured in brain tissue of different groups. All tested compounds revealed variable anti-epileptic effects, the most potent compounds were 6b and 6m. Also 6d, 6e, 6h, 6i, 6k, 6l and 6n compounds exhibited good anti-seizure activity, while compound 6j showed the lower activity. The rest of compounds displayed a neutral activity.

Keywords: Anticonvulsant; Antitumor agents; Pyrazolopyrimidines; Structure-activity relationship; Triazines.