Lung cancer is the leading killer of cancer-related cases and has an extremely high incidence worldwide [1], [2]. Currently, there are two main types of lung cancer: NSCLC (Non-Small Cell Lung Cancer) and SCLC (small cell lung cancer), NSCLC accounts for about 85 % of all lung cancer cases, which is a serious threat to human health [3], [4], [5]. Traditional cancer treatments include surgery, radiotherapy, and chemotherapy, which not only lack specificity but also seriously harm the physical and mental health of patients and are often difficult for most patients to bear [6], [7], [8]. In contrast, molecular-targeted therapy has the advantages of good efficacy, low toxicity, and high specificity, and many molecular-targeted drugs have been successfully marketed and applied, becoming an important treatment method for NSCLC [9], [10].

Among the molecularly targeted therapeutic agents for NSCLC, RTKIs (Receptor Tyrosine Kinase Inhibitors) have had a brilliant performance [11]. Among them, the successful development of EGFR-TKIs (Epidermal growth factor receptor- Tyrosine Kinase Inhibitors) is considered a milestone event in the targeted therapy of NSCLC [12]. EGFR belongs to the tyrosine kinase family, and the aberrant expression of EGFR is closely related to the growth and proliferation of tumor cells [13], [14]. Currently, four generations of EGFR inhibitors have been developed according to different mechanisms of action [15].

First-generation EGFR-TKIs are quinazoline-based structures, and they have significant efficacy in patients who develop EGFRdel19 deletion mutations and EGFRL858R mutations, of which Gefitinib was the first FDA-approved EGFR-TKI [16]. However, relevant studies have demonstrated that in approximately 68 % of patients with NSCLC who take first-generation EGFR- TKIs, the amino acid residue at exon 20 was mutated from THR790 to MET790 (T790M mutation), leading to the development of drug resistance [17]. The second generation of EGFR-TKIs introduced acrylamide as the Michael receptor and successfully overcame the T790M mutation problem [18]. Afatinib and Neratinib are representative drugs, but they also inhibit the expression of EGFRWT and do not selectively target the EGFRT790M mutation, which leads to toxic side effects such as diarrhea and dermatitis [19], [20], [21]. The structures of the above EGFR-TKIs are shown in Fig. 1.

The third-generation EGFR-TKIs are based on pyrimidines and retain the structure of acrylamide, effectively solving the problems of poor selectivity and high toxicity of the first two generations of inhibitors [22]. Such inhibitors include Osimertinib [23], WZ-4002 [24], Olmutinib [25], etc., and their structures are shown in Fig. 2. Among them, Osimertinib has an IC50 value of 15 nM against EGFRL858R/T790M in H1975 cells, respectively, and has been approved by the FDA for the treatment of patients with EGFRT790M-positive mutated NSCLC. However, after taking the drug for some time, the EGFRC797S mutation in patients resulted in the disruption of the cysteine 797 binding site, causing the development of Osimertinib resistance problems [26], [27]. Although a small number of fourth-generation EGFR-TKIs have been reported based on the EGFRC797S mutation resistance mechanism, they have not entered clinical application, so third-generation EGFR-TKIs are still a hot spot of research for the treatment of non-small cell lung cancer.

Based on the excellent efficacy of Osimertinib, researchers developed a new EGFR inhibitor, Mobocertinib (TAK-788), with a strategy of structural improvement [28], [29]. Mobocertinib can interact with the C790 residue of EGFR by introducing carboxylated isopropyl ester at the C5 position and selectively targets the EGFRex20 mutation in NSCLC [30], [31], [32]. Mobocertinib has excellent anti-tumor effects in vitro and in vivo and has been approved by the FDA for the treatment of patients with advanced NSCLC [33], [34]. However, Mobocertinib has a large adverse effect on the stomach, intestines, and skin, which is physically hazardous to patients [35], [36]. The structure of Mobocertinib is shown in Fig. 3.

To investigate the inhibitory effect of Mobocertinib derivatives on EGFRT790M mutation and to minimize drug toxicity, we introduced the morpholine structure, which has good biocompatibility in Gefitinib, to promote hepatic metabolism [37], [38]. Therefore, we envisioned retaining the Mobocertinib parental structure and structurally modifying the side chain to investigate the antitumor efficacy of Mobocertinib derivatives against EGFRT790M mutation. We chose Mobocertinib as the lead compound for structural modification and designed and synthesized 63 Mobocertinib derivatives by using molecular docking mimicry and bioelectronic isoarrangement principles. After the biological activity evaluation and anti-tumor activity study of the compounds, it was finally verified that compound H-13 has low toxicity and excellent potency with an anti-tumor effect and can be used as a potential third-generation EGFR inhibitor.

The problem of the high toxicity of Mobocertinib has similarities with the second-generation EGFR inhibitors, and from the structural point of view, the tails of Mobocertinib and the structures of the second-generation EGFR inhibitors, Afatinib and Neratinib, all contain a similar hydrophobic structure. The tails of WZ-4002 and Olmutinib in the third-generation EGFR-TKIs all have a six-membered cyclic piperazine structure, so based on the structural similarity improvement strategy, we considered modifying the structure of the hydrophobic region of the tail end of Mobocertinib to the six-membered cyclic morpholino structure at the tail end in the structure of the first-generation EGFR inhibitor Gefitinib. The morpholino deconjugation in Gefitinib promotes hepatic metabolism and reduces toxicity. In addition, we designed and synthesized Z-series compounds by trying to replace the acrylamide structure with different types of amide structures; meanwhile, based on the principle of bioelectronic isoarrangement, we designed and synthesized H-series compounds by replacing the amides of Z-series with different types of urea structures. The schematic diagram of the atomic design is shown in Fig. 4.

Firstly, we performed molecular docking simulations of Mobocertinib, compounds Z-1 and H-1 with EGFRT790M protein (PDB: 3IKA [24]), respectively. The results are shown in Fig. 5. Fig. 5A observes that the indole pyrimidine structure in the structure of Mobocertinib extends well into the protein cavity, forming a strong affinity interaction; Fig. 5B shows that it is the benzene ring and the acyl chloride in the structure of Z-1 that extends into the protein cavity, and the indole pyrimidine structure is exposed in the solvent region; whereas the indole pyrimidine structure of H-1 enters into the same position of the protein cavity as that of Mobocertinib in Fig. 5C and the urea structure extends into the cavity on the other side of the protein, demonstrating that the three bind to the protein in different ways. Then we carried out the synthesis and bioactivity evaluation of the Z series and H series, trying to investigate the effect of different side chain structure types on the antitumor efficacy.