Novel 6, 7-disubstituted 7H-purine analogues as potential EGFR/HER2 dual kinase inhibitors overcome lapatinib resistance: Design, synthesis, in-vitro and in-vivo evaluation

We synthesized and evaluated a new series of 6, 7 disubstituted-7H-purine derivatives (10a-u) that inhibit cancer growth via dual inhibition of EGFR/HER2. A total of twenty-one compounds were synthesized employing effective synthetic strategies and characterized. Initially, all compounds were screened in-vitro for kinase inhibition against a panel of seven kinases, including four members of the EGFR family, VEGFR, PDGFR alpha, and PDFGR beta. Five compounds from three synthetic series, 10d, 10e, 10i, 10 m, and 10r, were identified as potent EGFR family kinase inhibitors. Compound 10r effectively inhibited EGFR/HER2 dual kinases with an IC50 of 0.017 +/- 0.003/0.014 +/- 0.002 mu M. In addition, in-vitro cytotoxic data indicated that compounds 10i and 10r significantly altered the growth, resulting in potential anti-proliferation in a panel of breast cancer cells except for MCF-7. Following the findings of kinase inhibition, 10r demonstrated the lowest growth inhibition (IC50) against all breast cancer cells, ranging from 1.537 to 3.196 mu M, except MCF-7 cells. Interestingly, 10i and 10r showed promising IC50 ranging from 1.682 to 2.236 mu M against the Lapatinib resistant lines BT-474/L and SKBR3/L. Furthermore, apoptotic analysis of 10r revealed that the most significant percentage of cells, including Lapatinib-sensitive and resistant BT-474 and SKBR-3 cells, were forced into the accelerated death phase over 10r treatment, indicating effective apoptotic behaviour. The selective downregulation of dual EGFR/HER2 and their downstream signaling regulators ERK1/2 and AKT were observed over the treatment of 10r in both BT-474/L SKBR-3/L cells. Inconsistent with the in-vitro effects, 10r exhibited the same result in BT-474 and BT-474/L xenograft mice models. Finally, molecular docking studies revealed that 10r strongly binds to EGFR and HER2 via 'H' and 'pi' bond interactions, resulting in irreversible dual kinase inhibition. Thus our findings show that substituting 6, 7 positions in the purine core might result in potential anti-cancer agents, including combating the ability of Lapatinib resistance.