Vascular endothelial growth factor receptor-2 (VEGFR-2) plays a crucial role in cancer angiogenesis. In this study, a series of novel furo[2,3-d]pyrimidine and thieno[2,3-d]pyrimidine based-derivatives were designed and synthesized as VEGFR-2 inhibitors, in accordance to the structure activity relationship (SAR) studies of known type II VEGFR-2 inhibitors. The synthesized compounds were evaluated for their ability to in vitro inhibit VEGFR-2 kinase enzyme. Seven compounds (15b, 16c, 16e, 21a, 21b, 21c and 21e) demonstrated highly potent dose-related VEGFR-2 inhibition with IC₅₀ values in nanomolar range, of which the thieno[2,3-d]pyrimidine based-derivatives (21b, 21c and 21e) exhibited IC₅₀ values of 33.4, 47.0 and 21 nM respectively. Moreover, furo[2,3-d]pyrimidine-based derivative (15b) showed the strongest inhibition of human umbilical vein endothelial cells (HUVEC) proliferation with 99.5% inhibition at 10 μM concentration. Consistent with our in vitro findings, compounds (21b and 21e) orally administered at 5 and 10 mg/kg/day for 8 consecutive days demonstrated potent anticancer activity in Erhlich ascites carcinoma (EAC) solid tumor murine model. Such compounds blunted angiogenesis in EAC as evidenced by reduced percent microvessel via decreasing VEGFR-2 phosphorylation with subsequent induction of apoptotic machinery. Furthermore, Miles vascular permeability assay confirmed their antiangiogenic effects in vivo. Intriguingly, such compounds showed no obvious toxicity.

In light of the emergence of resistance against the currently available EGFR inhibitors, our study focuses on tackling this problem through the development of dual EGFR/HER2 inhibitors with improved enzymatic affinities. Guided by the binding mode of the marketed dual EGFR/HER2 inhibitor, Lapatinib, we proposed the design of dual EGFR/HER2 inhibitors based on the 6-phenylthieno[2,3-d]pyrimidine as a core scaffold and hinge binder. After two cycles of screening aiming to identify the optimum aniline headgroup and solubilizing group, we eventually identified 27b as a dual EGFR/HER2 inhibitor with IC50 values of 91.7 nM and 1.2 μM, respectively. Notably, 27b dramatically reduced the viability of various patient-derived cancer cells preferentially overexpressing EGFR/HER2 (A431, MDA-MBA-361 and SKBr3 with IC50 values of 1.45, 3.5 and 4.83 μM, respectively). Additionally, 27b efficiently thwarted the proliferation of lapatinib-resistant human non-small lung carcinoma (NCI-H1975) cells, harboring T790 M mutation, with IC50 of 4.2 μM. Consistently, 27b significantly blocked EGF-induced EGFR activation and inactivated its downstream AKT/mTOR/S6 signalling pathway triggering apoptotic cell death in NCI-H1975 cells. The present study presents a promising candidate for further design and development of novel EGFR/HER2 inhibitors capable of overcoming EGFR TKIs resistance.

Epidermal Growth Factor Receptor (EGFR) stands out as a key player in the development of many cancers. Its dysregulation is associated with a vast number of tumors such as non-small-cell lung cancer, colon cancer, head-and-neck cancer, breast and ovarian cancer. Being implicated in the development of a number of the most lethal cancers worldwide, EGFR has long been considered as a focal target for cancer therapies, ever since the FDA approval of "Gefitinib" in 2003 and up to the last FDA approved small molecule EGFR kinase inhibitor "Osimertinib" in 2015. Studies are still going on to find more efficient EGFR inhibitors due to the continuous emergence of resistance to the current inhibitors. Cancerous cells resist EGFR tyrosine kinase inhibitors (TKIs) through various mechanisms, the most commonly reported ones are the T790M mutation and HER2 amplification. Therefore, tackling EGFR TKIs-resistant tumors through a multi-targeting approach comprising a dual EGFR/HER2 inhibitor that is also capable of inhibiting the mutant T790M EGFR is anticipated to overcome drug resistance. In this review, we will survey the structural aspects of EGFR family and the structure-activity relationship of representative dual EGFR/HER2 inhibitors. To follow, we will discuss the structural aspects of the mutation-driven resistance and various design strategies to overcome it. Finally, we will review the SAR of exemplary irreversible dual EGFR/HER2 inhibitors that can overcome the mutation-driven resistance.