The vital organs of both the mouse that died and those that were euthanized were individually observed for overt pathology by necropsy, and the 50% lethal dose (LD50) was calculated from the Bliss method on day 14. this study of nearly 390,000 compounds screened by virtual database screening, the top 29 compounds were identified as candidate small-molecule EGFR/Eps8 complex inhibitors and evaluated by using cell-based assays. The compound EE02 was identified as the best match to our selection criteria. Further investigation shown that EE02 directly bound to the JXM website of EGFR and disrupted EGFR/Eps8 complex formation. EE02 selectively suppressed growth and induced apoptosis in EGFR-positive and Eps8-positive breast tumor and NSCLC cells. More importantly, the PI3K/Akt/mTOR and MAPK/Erk pathways downstream of the EGFR/Eps8 complex were suppressed by EE02. In addition, the suppressive effect of EE02 on tumor growth in vivo was comparable to that of erlotinib at the same dose. Conclusions We recognized EE02 as an EGFR/Eps8 complex inhibitor that shown encouraging antitumor effects in breast tumor and NSCLC. Our data suggest that the EGFR/Eps8 complex gives a novel tumor drug target. the peptide E1 derived from the JXM website can bind to the EGFR JXM website and effectively reduce EGFR dimerization, which therefore affects EGFR activity and reduces cell viability . Boran once shown the JXM website of EGFR is essential to the activation of EGFR, and the JXM website activates and regulates EGFR activation and is a potential target for the development of fresh EGFR inhibitors . Epidermal growth element receptor pathway substrate 8 (Eps8) is an important active kinase substrate of EGFR [2, 20]. EPS8 is definitely efficiently phosphorylated by numerous tyrosine kinases, both receptor (RTK) and nonreceptor types, and is UK 5099 a typical signaling protein, having a molecular excess weight of 97?kDa and containing a phosphotyrosine binding protein (PTB) website, an Src homology 3 (SH3) website and a sterile alpha-pointed (SAM-PNT) website [21, 22]. Eps8 is frequently overexpressed in breast, lung and additional malignancies but hardly ever in normal cells [23C26]. Further studies of UK 5099 EPS8 have revealed that a website that encompasses amino acids 298 to 362provides a binding surface for the JXM website of EGFR . Studies by Fazioli et al. and Castagnino et al. have shown that Eps8 directly binds to the JXM website of EGFR and is phosphorylated, which activates a series of downstream signaling pathways [20, 27], and thus promotes tumor progression. Furthermore, the aberrant manifestation of Eps8 often suggests anunfavorable prognosis for malignancy individuals [20, 27, 28]. Consequently, Eps8 is considered a novel CYSLTR2 UK 5099 potential target for specific tumor therapy. Eps8 directly binds to the JXM website of EGFR and forms an EGFR/Eps8 complex. Studies within the EGFR/Eps8 complex in malignancies are limited. In the present study, we focused on the EGFR/Eps8 complex as a encouraging tumor target for malignancy therapy. The Eps8-derived 9-amino acid peptide 327, which UK 5099 partly mimics the EGFR binding region of Eps8, functions like a protein-protein connection module that can disrupt the EGFR/Eps8 complex, prevent the activity of the downstream EGFR pathway, and exert antitumor effects . In our opinion, there is another approach to disrupt the EGFR/Eps8 complex. Direct inhibition of the EGFR/Eps8 complex by using drug-like, nonpeptide small molecules have several advantages, including obstructing the activity mediated by EGFR/Eps8 complex activation, improved cell permeability and better in vivo stability and bioavailability. Based on the high-resolution X-ray 3D crystal structure of JXM and UK 5099 the kinase website of EGFR, the JXM website is critical for EGFR activation and functions as a binding site for Eps8 [13, 27]. Consequently, we.