Introduction

Epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase (RTK) superfamily that consists of exon boundaries and associated extracellular, transmembrane, and intracellular protein domains. EGFR is involved in multiple signaling pathways and regulates numerous cell functions (Fig. 1A). This transmembrane glycoprotein is composed of a cysteine-rich extracellular ligand binding domain, hydrophobic transmembrane domain, cytoplasmic RTK domain, and C-terminal domain. The RTK domain contains an N-lobe consisting of five β-sheet strands and one αC helix and a C-lobe containing the main helices of a highly flexible activation loop (A-loop) [1]. The deep cleft at the junction of these two lobes forms the binding pocket for the adenine ring of ATP. The conformation of three conserved structural elements, namely the Asp-Phe-Gly (DFG) motif, αC helix, and A-loop, critically regulates the activation or inactivation of the catalytic domain. When EGFR is in the active state, the important catalytic residue D855 is located in the ATP binding site, stabilizing the ATP-loaded complex (DFG-in) and αC helix (αC-in). In the inactive state, EGFR forms a Src-like structure, including a closed A-loop, αC-out, and DFG-in [2]. (Fig. 1B). EGFR can dimerize upon binding by ligands, such as amphiregulin, β-cytokines, epidermal growth factor (EGF), heparin-binding EGF-like growth factor (HB-EGF), and transforming growth factor (TGF). The activation of the intracellular tyrosine kinase domain and autophosphorylation, which initiates the Ras/RAF/MEK, signal transducer and activator of transcription (STAT), PI3K/AKT/mTOR and other downstream signaling pathways, are closely related to embryonic development and stem cell division [2,3,4]. Overexpression of wild-type (WT) EGFR protein with or without EGFR gene amplification or a kinase-activating mutation further enhances cell proliferation, migration, survival, and antiapoptotic responses through signaling cascades, and these processes are closely related to the occurrence and development of many types of epithelial-derived cancer, such as non-small cell lung cancer (NSCLC), breast cancer, glioma, head and neck cancer, cervical cancer, and bladder cancer. Among these cancers, lung cancer appears to be the most common and has the characteristics of aberrant proliferation, metastasis, and drug resistance [5,6,7,8]. Thus, EGFR has become a promising target for anticancer drug design and development. EGFR tyrosine kinase inhibitors (EGFR-TKIs) have achieved remarkable results in the clinic [9]. However, most patients develop acquired drug resistance to first- and second-generation EGFR-TKIs after 1–2 years. The mechanism of drug resistance for nearly half of cases relates to the T790M mutation. Third-generation EGFR-TKIs that target EGFR-TKI-sensitive mutations and the T790M mutation have been developed [10].

Fig. 1
figure 1

Structure and functions of EGFR. A EGFR exon boundaries and associated extracellular, transmembrane, and intracellular protein domains. EGFR is involved in multiple signaling pathways and regulates numerous cell functions. B The tyrosine kinase domain of EGFR and the activation or inactivation of the catalytic domain. C EGFR domains and the molecular mechanisms of acquired resistance. The intracellular domain contains a juxtamembrane domain, tyrosine kinase domain, and multiple C-terminal tyrosine residues. Multiple mutations within the tyrosine kinase domain are associated with resistance and sensitivity to EGFR-TKIs

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Unfortunately, drug resistance caused by less-common mutations in the EGFR gene and components of signal transduction pathways continues to emerge. In addition to common secondary (T790M) and tertiary (C797S) mutations, other EGFR mutations (such as the L718Q, L796S, and L792H mutations and the exon 20 insertion), MET amplification, phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) mutations, HER2 amplification, oncogene fusions, and alterations in cell cycle-related genes have been observed [11] (Fig. 1C). There is an urgent need for better strategies to combat the inevitable molecular-targeted drug resistance associated with third-generation inhibitors. This review aims to provide a comprehensive overview of the mechanisms of resistance to third-generation EGFR-TKIs and to explore new insights and strategies for overcoming acquired resistance.

Third-generation EGFR-TKIs and drug resistance mechanisms

The development of third-generation EGFR-TKIs

The first-generation EGFR-TKIs form hydrogen bonds with Met793 in the ATP binding pocket of EGFR and reversibly compete with ATP for binding. Drug resistance occurs due to the EGFR T790M mutation (Thr790 in the hydrophobic ATP binding site encoded on exon 20 is replaced by methionine), subclonal selection (of a genetically resistant clone), and rare EGFR mutations (such as G719X, S768I, and L861Q). Thereafter, the development of second-generation EGFR-TKIs was reported; these inhibitors have the same quinazoline scaffold as first-generation EGFR-TKIs, but the side chain can irreversibly bind to Cys797 to inhibit the tyrosine kinase activity of EGFR. For example, the anilinoquinazoline derivative forms hydrogen bonds with the backbone of Met793 in the hinge region and interacts with the hydrophobic region. The acrylamide group binds covalently to Cys797 in the active conformation of EGFR, the furanyl group is exposed to solvent, and the 3-chloro-4-fluorophenyl group is situated next to the gatekeeper residue [12,13,14]. However, mutations such as T790M still emerge upon treatment with second-generation EGFR-TKIs, which have limited selectivity against WT-EGFR, resulting in serious side effects [15]. Fortunately, third-generation covalent inhibitors that bind irreversibly to the target and are mutation-selective have been developed. These compounds were designed based on a new aminopyrimidine scaffold and show preferable biological activities [16]. Early clinical trials have proven that these third-generation EGFR-TKIs are effective in patients with double-mutated tumors (EGFR L858R/T790M or ex19del/T790M) and have high selectivity for mutant EGFR, thereby eliminating the side effects in the skin and gastrointestinal system associated with the nonselective inhibition of WT-EGFR [17]. For example, the crystal structures of rociletinib (CO-1686) in complex with EGFR T790M and EGFR L858R have been published; in EGFR T790M, the anilinopyrimidine group of rociletinib forms hydrogen bonds with the Met793 amide and the carbonyl backbone, whereas in EGFR L858R, hydrophobic interactions between rociletinib and the protein were due to hydrogen bonds between nitrogens in the pyrimidine group and between the fluoromethyl and Thr790. In addition, the acrylamide group in rociletinib covalently binds to Cys797 in the DFG-in/αC-in active conformations [18]. The specificity for EGFR T790M may stem from hydrophobic interactions between the large methionine in mutant EGFR and pyrimidines. Drugs that have been approved for marketing include osimertinib (US), almonertinib (China), lazertinib (South Korea), and alflutinib (China) (Fig. 2).

Fig. 2
figure 2

Development of third-generation EGFR-TKIs

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Mechanisms of resistance to third-generation EGFR-TKIs

Due to the covalent bond between the acrylamide (Michael acceptor) of third-generation EGFR-TKIs and the active thiol in the EGFR kinase domain, highly selective inhibitory activity has been achieved by targeting Cys797 and irreversible binding EGFR; thus, these compounds show excellent antitumor activity. Targeted therapy for patients with EGFR T790M and EGFR-activating mutations showed good efficacy in both first- and second-line settings. In patients who developed resistance to third-generation EGFR-TKIs as first-line therapy, genetic changes such as MET amplification, EGFR C797X mutation, PIK3CA amplification and mutation, HER2 amplification and mutation, K-RAS mutation, and BRAF mutation, as well as changes in cell cycle-related genes and oncogene fusions, have been reported, but no T790M mutations have been detected. The mechanism of resistance to second-line therapy is more complicated. Acquisition or deletion of the T790M mutation has been detected in patients [19], and other EGFR mutations (such as L718Q, L796S, L792H, and exon 20 insertion) have also been observed (Fig. 1B). In addition, the mechanisms of acquired resistance to third-generation EGFR-TKIs include alternative pathway activation and histologic and phenotypic transformation (Fig. 3); the details will be discussed in the following sections.

Fig. 3
figure 3

Molecular mechanisms of acquired resistance. The mechanisms include target gene modification, parallel alternative pathway activation, downstream pathway activation, and histological/phenotypic transformation. Both amplification and mutation of receptor tyrosine kinases (RTKs) can induce downstream survival signaling pathways. Moreover, direct overexpression and/or mutation of components of downstream pathways can contribute to acquired resistance by promoting cancer cell survival

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Primary/intrinsic resistance

The differential sensitivity of TKIs to different EGFR mutations is a cause of primary drug resistance. In NSCLC patients, the in-frame deletion of exon 19 (ex19del) and the L858R point mutation in exon 21 are the most common somatic mutations, occurring in approximately 80% of cases. During EGFR-TKI treatment, patients with longer median survival have presented with more than 20 unique deletions of exon 19. Intrinsic drug resistance can all be triggered by other nonclassical sensitizing mutations (mainly exon 20 insertion) and inherent secondary genetic changes. Drug-resistant clones (for example, T790M) may already exist within the cancer cell population, leading to drug resistance during treatment [20]. Some studies have found that in nearly 1% of lung cancer patients, 2–3 simultaneous driver mutations can be detected before treatment. Some molecular and genetic changes have been reported to relate to intrinsic drug resistance, such as the lack of K-RAS/phosphatase and tensin homolog (PTEN) expression. These preexisting molecular and genetic alterations can stimulate the Ras/Raf/MEK/ERK and PI3K/AKT downstream pathways to promote cancer progression [21].

BIM deletion polymorphism

BIM is a proapoptotic member of the B-cell lymphoma-2 (Bcl-2) family [22]. Recent studies showed that lung cancer cells with the BIM deletion polymorphism and EGFR mutation are resistant to third-generation EGFR-TKIs, suggesting that the BIM deletion polymorphism has potential as a biomarker to predict the efficacy of third-generation EGFR-TKIs in patients [22].

EGFR exon 20 insertion

The molecular mechanism of drug resistance caused by the exon 20 insertion is not fully understood. Eck et al. [23] hypothesized that this mutation prevents binding to EGFR-TKIs due to the addition of residues to the N-lobe of EGFR. The crystal structure of EGFR exon 20 with the D770_N771insNPG insertion shows an unchanged ATP binding pocket and a rigid active conformation, leading to steric hindrance of the drug binding pocket and resistance to EGFR-TKIs.

Acquired resistance

Acquired drug resistance refers to the process by which tumor cells with prior sensitivity to treatment circumvent the inhibitory effects of drugs by changing their metabolic pathways. The mechanisms of acquired resistance to third-generation EGFR-TKIs can be divided into EGFR-dependent resistance and EGFR-independent resistance [24].

EGFR-dependent drug resistance mechanisms

Reappearance of an EGFR mutation

C797S mutation

One point mutation of EGFR (C797S) involves the replacement of Cys797 within the ATP binding site (exon 20) with serine [25]. Osimertinib binds covalently and irreversibly to EGFR T790M by interacting with Cys797. When the C797S mutation occurs, the osimertinib binding efficiency decreases [10], resulting in tumor resistance to all third-generation EGFR-TKIs.

G796R/D mutation

The G796R mutation has been detected in cancer patients who received treatment with a third-generation EGFR-TKI. Molecular docking predictions revealed that G796R sterically hinders the covalent binding of osimertinib. Because the bulky side chain and hydrophilic group hinder the binding of osimertinib to the hydrophobic region, the change in binding energy renders binding unfavorable. Compared with samples containing the double-mutant EGFR L858R/T790M, those harboring the triple-mutant EGFR L858R/T790M/G796R are 110 times more resistant to osimertinib [26]. G796D was reported for the first time in osimertinib-resistant NSCLC patients. In vitro studies have shown that the G796D mutation causes a 50-fold increase in the growth inhibitory 50% (GI50) value of osimertinib. Structural modeling showed that the side chain of the mutated G796D residue collides with the surface of osimertinib, resulting in steric hindrance and energy repulsion and ultimately the loss of binding affinity [27].

L792 mutation

The mutations at Leu792 include L792F, L792Y, and L792H. Structural prediction revealed that these mutations introduce a benzene ring or imidazole ring to the side chain of the residue at 792, which spatially disrupts the orientation of osimertinib, thereby potentially affecting the binding of osimertinib to the EGFR ATP binding site [28].

M766Q mutation

The homology simulation with the T790M and M766Q double mutant showed that M766Q seems to position T790M in the inhibitor binding site, thereby weakening osimertinib binding [29].

Mutations in exon 18

EGFR L718Q/V

EGFR L718Q was reported for the first time in a cell model of resistance to third-generation EGFR-TKIs. Subsequent studies have shown that NSCLC with EGFR L858R/T790M/L718Q is resistant to all EGFR-TKIs, but that with only L858R/L718Q remains sensitive to afatinib [30]. The crystallographic model revealed that the L718Q mutation reduces the efficiency of the formation of covalent bonds between the acrylamide warhead and the Cys797 thiol group, thus interfering with the irreversible binding of osimertinib [31, 32]. In addition, L718V resistance mutations in the kinase domain of EGFR have been detected, and these may interfere with the binding of osimertinib to the kinase domain [33]. Of note, EGFR L718Q/V is still sensitive to afatinib [32].

EGFR G724S

The G724S mutation in the ATP binding loop enriches this loop in glycine, which can lead to the development of resistance to EGFR-TKIs by changing the protein structure, enhancing ATP affinity, and stabilizing activating mutations [34]. However, this mutation does not lead to resistance to second-generation EGFR inhibitors [34].

Compound mutations

A compound mutation refers to the simultaneous detection of two or more different types of EGFR mutations in patient cancer cells [35]. The impact of compound mutations on EGFR-TKI sensitivity is listed in descending order: double classic mutations, compound mutations involving classic mutations and rare mutations, and compound mutations of only rare mutations [36, 37]. These EGFR mutations caused by treatment with third-generation EGFR-TKIs confer resistance to irreversible pyrimidine TKIs but not to quinazoline EGFR inhibitors [42]. After treatment with third-generation EGFR-TKIs, MET gene amplification can promote drug resistance by activating MAPK/ERK, which is independent of EGFR [43].

HER2 amplification

Hus et al. found that H1975 cells expressing HER2D16 were resistant to osimertinib in vitro. HER2D16 can form a heterodimer with EGFR or a disulfide homodimer, which activates downstream signaling to achieve resistance to osimertinib [44]. HER2D16-driven drug resistance occurs in a manner unrelated to the kinase Src. In addition, other mutations in exon 20 of HER2 have been reported, including point mutations (such as G776C and L755S) and insertions that cause downstream activation [45, 46]. HER2 mutation occurs in approximately 2–4% of NSCLC cases, mostly in lung adenocarcinoma (LUAD) [47]. In NSCLC, HER2 oncogenic amplification occurs in approximately 3% of cases without EGFR-TKI treatment and accounts for approximately 10% of cases with EGFR-TKI resistance [48].

AXL activation

AXL is an RTK that regulates cell survival, proliferation, metastasis, and other cellular functions. Abnormalities in the AXL gene can generate acquired resistance to TKIs by activating relevant downstream signaling pathways. Osimertinib was found to trigger AXL activation by closing the negative feedback loop with SPRY4, thus triggering inherent osimertinib resistance [49].

Overexpression of HGF

Hepatocyte growth factor (HGF) is the ligand of the proto-oncogene c-Met; it can trigger MET activation through EGFR bypass signaling and induce lung cancer resistance to EGFR-TKIs. Yano et al. [50] found that high expression of HGF was related to the acquired and intrinsic drug resistance to EGFR-TKIs in patients with lung cancer. Tumor specimens from patients with acquired drug resistance showed high expression of HGF in the context of MET amplification and the T790M mutation.

Fibroblast growth factor receptor (FGFR) signaling

FGFR is a transmembrane RTK. Studies have shown that FGFR1 is amplified and fibroblast growth factor 2 (FGF2) mRNA levels are increased in patients with osimertinib resistance, suggesting that the FGFR2-FGFR1 autocrine loop may be related to drug resistance [51]. Patients with the T790M mutation have been reported to show disease progression after treatment with osimertinib and nilotinib. The FGFR3-TACC3 fusion was detected in ctDNA [52, 53]. These findings suggest that abnormalities in the FGFR signaling pathway may underlie the mechanism of acquired resistance to third-generation EGFR-TKIs.

Insulin-like growth factor receptor 1 (IGF1R)

IGF1R, a transmembrane heterotetrameric protein encoded by the gene located on chromosome 15q26.3, is involved in promoting the growth of tumor cells. Abnormal activation of IGF1R leads to EGFR-TKI resistance [54].

Aurora kinases (AURKs)

AURKs are an important category of enzymes within the serine/threonine kinase family consisting of three mammalian isoforms: Aurora kinase A (AURK A), AURK B, and AURK C [55, 56]. AURK A and AURK B are highly expressed in dividing cells and play important roles in mitotic progression. Mammalian AURK A and AURK B share approximately 71% similarity in the carboxy-terminal catalytic domain [57]. Aberrant expression of AURK A and AURK B is involved in a broad range of solid cancers and is associated with adverse prognosis and drug resistance [58, 59]. In addition, Tanaka et al. [60] reported that targeting AURK B can prevent and overcome resistance to EGFR inhibitors in lung cancer by enhancing BIM- and PUMA-mediated apoptosis.

Downstream signaling pathway activation

The activation of signaling pathways downstream of oncogenic receptors can regulate cell proliferation, cell cycle progression, and cell survival. Therefore, the direct regulation of downstream signaling pathway-related factors can lead to acquired resistance.

K-RAS mutation

An epidemiological meta-analysis found that K-RAS mutations are present in NSCLC patients, and all patients with K-RAS mutations were resistant to EGFR-TKIs [61]. K-RAS mutation is related to activation of the RAS-MAPK pathway. The common K-RAS mutations include G12S, G12D, G12A, Q61H, and A146T. Studies have found that inhibiting mutant K-RAS can reduce tumor growth and render NSCLC patients sensitive to EGFR inhibitors [62].

BRAF (v-RAF murine sarcoma viral oncogene homologue B1) mutation

BRAF is a serine/threonine protein kinase that plays a key role in the MAPK/ERK pathway, including in EGFR/RAS/RAF signal transduction. BRAF can regulate cell survival, proliferation, differentiation, and apoptosis, as well as tumor induction. Many BRAF mutations (G469A, V600E, and V599E) have been found in cancer, including lung cancer [63]. Ohashi et al. [64] reported that in patients with lung cancer, BRAF mutations can induce acquired resistance to EGFR-TKIs. Preclinical data showed that the BRAF V600E mutation has a strong association with resistance to the third-generation EGFR-TKI osimertinib in patients with T790M-mutated LUAD.

PI3K/AKT/mTOR

PIK3CA is a driver gene of LUAD. Mutation of PIK3CA can promote tumor cell invasion and increase the activity of downstream PI3Ks. Studies have shown that PIK3CA amplification or mutation (including E453K, E545K, and H1047R) may occur in patients with osimertinib resistance [52, 65]. Increased PI3K activity leads to the activation of various downstream kinases, thereby increasing PI3K/AKT/mTOR pathway activity in the absence of coupling to upstream EGFR phosphorylation.

STAT3 activation

STAT proteins, especially STAT3, are key downstream signal sensors of EGFR activation. In studies on NSCLC, Zhao et al. [66] discovered the clinical significance of JAK2/STAT3 in angiogenesis. Chaib et al. [67] found that osimertinib treatment activates not only STAT3 but also SrcYAP1 signaling, which may act downstream of IL-6 to promote disease progression.

Loss of PTEN

PTEN is a tumor suppressor gene that encodes a protein with lipid phosphatase activity and thus regulates cellular protein phosphatase activity. PTEN has dual antitumor effects and is a key component of many signaling pathways in the body. If mutation or deletion of the PTEN gene or downregulation of PTEN expression can reduce or eliminate its antitumor activity [68], loss of PTEN leads to hyperactivation of the PI3K/AKT signaling pathway and resistance to EGFR-TKIs, including osimertinib.

Hyperactivation of activated Cdc42-associated kinase 1 (ACK1)

Hyperphosphorylation of ACK1 and the subsequent activation of antiapoptotic signaling through the AKT pathway are associated with resistance to third-generation EGFR-TKIs [69].

c-Myc gene

The c-Myc gene is an important member of the MYC gene family. The c-Myc gene can induce cells to proliferate indefinitely and can promote cell division; these activities are related to the occurrence and development of various types of cancer. Studies have shown that c-Myc levels are substantially elevated in different EGFR-mutant NSCLC cell lines with acquired resistance to the third-generation EGFR-TKI osimertinib compared with the corresponding parental cell lines; moreover, these increased levels cannot be reduced by osimertinib. Consistently, c-Myc levels are elevated in the majority of EGFR-mutant NSCLC tissues from patients who relapsed on EGFR-TKI treatment compared with the corresponding baseline c-Myc levels prior to treatment [70]. These findings indicate that c-Myc mediates the therapeutic efficacy of third-generation EGFR-TKIs and the development of acquired resistance to these TKIs.

Other mechanisms

Epithelial–mesenchymal transition (EMT)

In EMT, cancer cells lose their epithelial properties through the loss of E-cadherin, leading to increased vimentin expression and transformation into a mesenchymal phenotype. A previous study found that osimertinib-resistant H1975 cells have EMT characteristics in the absence of other EGFR mutations [71]. EMT is a coordinated process involving multiple regulatory factors, such as EMT-induced transcription factors (EMT-TFs), noncoding RNAs (ncRNAs), and various extracellular signals. EMT-TFs play an important role in all stages of EMT; the most well-known EMT-TFs are members of the SNAIL, ZEB, and TWIST families. Many studies have shown that SLUG and SNAIL overexpression can induce drug resistance [72].

miRNAs and EMT

Long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) play important roles in regulating EMT and TKI resistance. Although most miRNAs have been found to inhibit EMT, some have activity that promotes EMT, including miR-21 and miR-155 [73, 74]. Some miRNAs can promote TKI resistance by activating the PI3K/AKT/mTOR signaling pathway; for example, miR-21 and miR-23a can target PTEN and activate AKT, leading to resistance to EGFR-TKIs [75, 76].

Epigenetic alterations

Epigenetic modifications involved in cancer initiation and progression include changes in DNA methylation patterns and histone modifications. Epigenetic changes are common in the development and progression of lung cancer [77]. Studies have shown that epigenetic disorders can make cancer patients susceptible to acquired resistance to EGFR-TKIs [78].

Oncogene fusion

The AURA-3 and FLAURA trials showed that oncogene fusion might be one mechanism of osimertinib resistance; the identified fusions included transforming growth factor receptor (TGFR)-transforming acidic coiled-coil protein 3 (TACC3), neurotrophic receptor tyrosine kinase 1 (NTRK1)-thrombopoietin mimetic peptide 3 (TMP3), ERC1-RET, SPTBN1-ALK, coiled-coil domain-containing protein 6 (CCDC6)-RET, GOPC-ROS1, AGK-BRAF, NCOA4-RET, ESYT2-BRAF, and echinoderm microtubule-associated protein-like 4 (EML4)-ALK. Oncogene fusions can coexist with the EGFR C797S mutation, MET amplification, and BRAF mutation [79].

Cell cycle-related gene abnormalities

Recent studies have shown that changes in cell cycle-related genes, including the CDKN2A E27fs mutation, cyclin D (CCND) amplification, cyclin-dependent kinase 4/6 (CDK4/6) amplification, and cyclin E1 (CCNE1) amplification, can cause resistance to third-generation EGFR-TKIs [65].

Histologic and phenotypic transformation

Histopathological transformation to small cell lung cancer (SCLC) from NSCLC has been reported as a mechanism of acquired resistance to EGFR-TKIs in 3–15% of patients [80,81,82,83]. Transformed SCLC mainly occurs in Asian patients with adenocarcinoma harboring EGFR-TKI-sensitive mutations (such as the EGFR ex19del/T790M mutation) who are nonsmokers. The widely accepted hypothesis for this transformation posits that adenocarcinoma and SCLC originate from type II alveolar cells. RB1 and TP53 mutations might be involved in SCLC transformation but are not sufficient for the induction of complete transformation. Additional genomic alterations, including those that activate the PI3K/AKT family and downregulate NOTCH signaling and those affecting the MYC and SOX families, AKT pathway activation and other molecules, also participate in the transformation from EGFR-mutant NSCLC. However, the precise mechanisms in other cases are unclear [84]. In addition, squamous cell transformation was recently identified as a mechanism of acquired EGFR-TKI resistance that occurs in approximately 15% of patients who received osimertinib as both first- and second-line therapy. Similar to the case in SCLC transformation, the primary EGFR mutation is preserved in squamous cell transformation [85].

Immune escape

EGFR is expressed in different hematopoietic cell types, including macrophages, monocytes, and certain T-cell subsets. Therefore, it is likely that EGFR inhibitors can interfere with the function of these leukocytes. Immune checkpoint inhibitors (ICIs) have adverse effects and poor efficacy in patients with an EGFR mutation or a secondary T790M mutation, largely because of low tumor mutational burden and a noninflamed tumor microenvironment [86,87,88]. A previous study showed that secreted phosphoprotein 1 (SPP1) promotes macrophage M2 polarization and PD-L1 expression in LUAD, which may influence the response to immunotherapy. SPP1 levels might be a useful marker of immunosuppression in patients with an EGFR mutation and could provide therapeutic insight [207]. It is a promising target for the design of selective conformationally restricted drugs, with great potential in terms of affinity, efficacy, and selectivity.

DZ-SIM inhibitors

In addition, researchers found that a group of near-infrared heptamethine carbocyanine (DZ) fluorescent dyes, the prototype of which is heptamethylamine carbocyanine dye (IR-783) (104) (Fig. 14), have tumor-targeting activity through differentially expressed organic anion transport peptides on cancer cells [208]. This group of organic dyes can specifically deliver therapeutic payloads to tumor cells in the form of chemical conjugates. DZ-SIM was preliminarily synthesized; SIM specifically targets 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) in the endoplasmic reticulum. After specific uptake by tumor cells, DZ-SIM was enriched in subcellular organelles (including mitochondria and lysosomes). NSCLC cells were killed by mitochondrial damage, which mainly led to cytochrome C release into the cytoplasm, thereby activating the caspase-3-dependent apoptosis cascade. DZ-SIM inhibited the formation of cancer cell colonies resistant to first-generation (H1650 and H1975) and third-generation EGFR-TKIs (PC9AR), and most IC50 values were lower than 10 μmol/L. DZ-SIM represents a promising new therapy to overcome drug resistance in patients with EGFR-mutant disease.

Selection of individualized combination therapy

For patients who experience SCLC transformation, chemotherapy after the development of osimertinib resistance is an option. Research has shown that patients with transformation to SCLC have higher response rates to etoposide, cisplatin, and paclitaxel. For patients with unclear resistance mechanisms, chemotherapy is still a treatment option. If the patient is asymptomatic or has symptomatic local progression, osimertinib can be combined with local treatment according to National Comprehensive Cancer Network (NCCN) guidelines. Carboplatin, paclitaxel, bevacizumab, and atezolizumab (anti-PD-L1 antibody) are also options for patients who experience systemic progression after osimertinib treatment [

Conclusion

As a crucial “controller” that is related to the inhibition of tumor cell proliferation, angiogenesis, invasion, metastasis, and apoptosis, EGFR actively participates in malignant disease progression. However, the intrinsic and acquired resistance in primary and recurrent cancer which is mediated by EGFR mutations after target treatment leads to difficult therapeutic. Understanding the complex resistance mechanisms of EGFR-TKIs and develo** potential strategies to combat it could be of potential interest for improving the individual therapeutic strategies for cancer.