Targeting epidermal growth factor receptor (EGFR) with tyrosine kinase inhibitors (TKI) has been widely exploited to disrupt aberrant phosphorylation flux in cancer. However, a bottleneck of potent TKIs is the acquisition of drug resistance mutations, secondary effects, and low ability to attenuate tumor progression. We have developed an alternative means of targeting EGFR that relies on protein degradation through two consecutive routes, ultimately leading to cancer cell detachment-related death. We describe furfuryl derivatives of 4-allyl-5-[2-(4-alkoxyphenyl)-quinolin-4-yl]-4H-1,2,4-triazole-3-thiol that bind to and weakly inhibit EGFR tyrosine phosphorylation and induce strong endocytic degradation of the receptor in cancer cells. The compound-promoted depletion of EGFR resulted in the sequestration of non-phosphorylated Bim, which no longer ensured the integrity of the cytoskeleton machinery, as shown by the detachment of cancer cells from the extracellular matrix (ECM). Of particular note, the longer CH3(CH2)n chains in the terminal moiety of the anti-EGFR molecules confer higher hydrophobicity in the allosteric site located in the immediate vicinity of the catalytic pocket. Small compounds accelerated and enhanced EGFR and associated proteins degradation during EGF and/or glutamine starvation of cultures, thereby demonstrating high potency in killing cancer cells by simultaneously modulating signaling and metabolic pathways. We propose a plausible mechanism of anti-cancer action by small degraders through the allosteric site of EGFR. Our data represent a rational and promising perspective in the treatment of aggressive tumors.

Chemotherapeutic interruption of signaling pathways at receptor tyrosine kinases is an important strategy for attenuating cancer progression. We have synthesized a new generation of polyfunctionalized heterocyclic compounds that bind to an allosteric site in the tyrosine kinase domain of the epidermal growth factor receptor (EGFR). The bound compounds induce degradation of the receptor by endocytosis in cancer cells. The Hsp90α chaperone binds to EGFR and is significantly lost during endocytosis, thereby contributing to the reduction of client proteins. Induced EGFR depletion leads to inactivation of downstream signaling due to sequestration of the Bim sensor protein for cytoskeletal proteins, provoking cell detachment from the extracellular matrix and ultimately cancer cell death. The role of glutamine in maintaining the phosphorylation status of EGFR-mediated signaling pathways can be postulated as «no glutamine, no EGFR signaling». Targeted degradation of EGFR is attractive for aiming to attenuate metastatic progression and to override the drug resistance of malignant tumors.

Targeted protein degradation is an attractive technology for cancer treatment due to its ability to overcome the unpredictability of small molecule inhibitors that cause resistance mutations. In recent years, various targeted protein degradation strategies have been developed based on the ubiquitin-proteasome system in the cytoplasm or the autophagy-lysosomal system during endocytosis. In this review, we describe and compare technologies for targeted inhibition and targeted degradation of the epidermal growth factor receptor (EGFR), one of the major proteins responsible for the onset and progression of many types of cancer. In addition, we have developed an alternative strategy, called alloAUTO, based on the binding of new heterocyclic compounds to an allosteric site located in close proximity to the EGFR catalytic site. These compounds cause targeted degradation of the transmembrane receptor, simultaneously activating both systems of protein degradation in cells. Damage to EGFR signaling pathways promotes inactivation of Bim sensor protein phosphorylation, which leads to disintegration of the cytoskeleton, followed by detachment of cancer cells from the extracellular matrix and, ultimately, to cancer cell death. This hallmark of targeted cancer cell death suggests an advantage over other targeted protein degradation strategies, namely that the fewer cancer cells survive, the fewer chemotherapy-resistant mutants appear.