Resveratrol belongs to a class of polyphenolic compounds called stilbenes. This natural polyphenol can be detected in >70 plant species (36). Resveratrol decreases CYP1A1 and ABCG2 expression, and impairs gefitinib elimination from cells, increasing intracellular gefitinib concentration, which helps to overcome gefitinib resistance (37). The accumulated gefitinib triggers apoptosis, autophagy and senescence in gefitinib-resistant NSCLC cells (37). Therefore, co-treatment with gefitinib and resveratrol potentiates the anti-NSCLC effects of gefitinib.

Gallic acid is a member of the hydroxybenzoic acids commonly found in a number of herbs and fruits. Gallic acid accelerates EGFR turnover (38), inhibits the Src-signal transducer and activator of transcription-3 (STAT3) signaling pathway, and induces apoptosis and cell cycle arrest in EGFR-TKI-resistant NSCLC, but not in EGFR-TKI-sensitive NSCLC, suggesting that gallic acid overcomes the acquired resistance to EGRF-TKI resistance in NSCLC (39).

Curcumin, also called diferuloylmethane, is isolated from the rhizome of Curcuma longa. Curcumin has anti-NSCLC effects; it inhibits NSCLC cell proliferation and induces apoptosis by increasing caspase-3 activity and the expression of miR-192-5p, which suppresses the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway in NSCLC (40). miR-192-5p mimics enhance the effects of curcumin on cell viability and apoptosis, and suppress the PI3K/Akt signaling pathway in NSCLC cells, while anti-miR-192-5p mimics reverse the effect of curcumin on NSCLC cells (40). Curcumin also suppresses the proliferation and invasion of NSCLC by inhibiting the metastasis-associated protein-1 (MTA-1)-activated Wnt/β-catenin pathway (41). MTA-1 promotes NSCLC cell invasion and metastasis (42). The decrease in MTA-1 by curcumin treatment inhibits NSCLC growth. Furthermore, curcumin induces autophagy in NSCLC and the autophagy inhibitor 3-methyladenine (3-MA) partly blocks the inhibitory growth effect of curcumin on cancer cells (43).

Curcumin also potentiates the therapeutic effects of gefitinib. Curcumin induces apoptosis in TKI-resistant NSCLC cells by decreasing EGFR phosphorylation and increasing EGFR degradation, thus inhibiting cancer growth (44). More importantly, the combination treatment of curcumin and EGFR-TKI markedly inhibits NSCLC growth by decreasing the expression of EGFR, c-MET and cyclin D1. The combination treatment confers a better survival rate and decreases intestinal mucosal damage in intestinal epithelial cells by regulating mitogen-activated protein kinase activity (45). Furthermore, the combination of curcumin and gefitinib causes marked autophagy induction, autophagic cell death and autophagy-mediated apoptosis when compared with curcumin or gefitinib treatment alone. Pharmacological autophagy inhibitors bafilomycin A1 or 3-MA, or knockdown of beclin-1 or autophagy-related 7, ameliorate treatment-induced autophagic cell death (45). Erlotinib and afatinib are other EGFR-TKIs used for frontline treatment. Co-administration of erlotinib and curcumin significantly decreases NSCLC cell viability by inducing apoptosis and increasing the expression of IκB (46), which restricts nuclear factor-κB (NF-κB) to the cytoplasm and inhibit its DNA binding activity (47). The combination treatment also significantly increases apoptosis by decreasing the expression of EGFR and survivin, and inhibiting NF-κB activity in erlotinib-resistant NSCLC cells (48). The application of curcumin and afatinib as a combination treatment for gefitinib- and erlotinib-resistant NSCLC has already been patented (CN105476996A).

Curcumin also overcomes chemotherapeutic resistance in NSCLC. A recent study implicated HIF-1α in the development of chemotherapeutic resistance in cancer; therefore, targeting HIF-1α either by RNA-interference or small interfering RNA may overcome the resistance to cisplatin (49). It has been reported that combined curcumin and cisplatin treatment markedly inhibits cisplatin-resistant NSCLC cell proliferation and triggers apoptotic death, by promoting HIF-1α degradation and activating caspase-3, respectively (49). Curcumin also reverses cisplatin resistance by increasing cisplatin-induced apoptosis by inducing generation of intracellular reactive oxygen species (ROS) and proteosomal degradation of Bcl-2 in NSCLC (50).

Ginsenosides are the main saponins in ginseng roots, fruits, stems and leaves. To date, >60 ginsenosides have been isolated and identified. Although the basic structure of ginsenosides are similar, the different arrangements in the four rings of steroid nuclei are different in different sub-types, such as the A-Panaxadiol group (e.g., Rb1, Rb2, Rb3, Rc, Rd, Rg3 and Rh2), the B-Panaxatriol group (e.g. Re, Rg1, Rg2 and Rh1) and the C-Oleanolic acid group (e.g. Ro) (51). One study showed that ginsenoside upregulated vaccinia-related kinase 1 and activated the tumor suppressor p53-binding protein-1, thus inhibiting NSCLC cell proliferation (52).

Ginsenosides also promote the antitumor activity of gefitinib in NSCLC. It has been demonstrated that ginsenoside Rg3 increases the cytotoxic effects of gefitinib in NSCLC cells in a dose- and time-dependent manner (53). The combination treatment of gefitinib and ginsenoside Rg3 significantly increases apoptosis in NSCLC cells by increasing the expression of the pro-apoptotic protein Bax and the activity of caspase-3, and by decreasing the expression of the anti-apoptotic protein Bcl-2 (53). The combination treatment also inhibits NSCLC cell migration by decreasing the protein expression of Snail and Slug (53), which are master regulators that regulate numerous genes that contribute to epithelial-mesenchymal transition (EMT), including E-cadherin (54). Osimertinib is the third-generation EGFR-TKI. A recent study revealed that osimertinib-resistant NSCLC cells displayed stronger stemness than the parental cells, which conferred to them greater metastatic potential (55). Notably, ginsenoside Rg3 attenuates the stemness of NSCLC cells and the resistance to osimertinib, as evidenced by the decreased expression of stemness markers and decreased spheroid formation ability, which is dependent on the Hippo signaling pathway (56). Furthermore, ginsenosides overcome resistance to chemotherapy drugs such as cisplatin (DDP), which is used for the treatment of NSCLC. Clinical research shows that ginsenoside Rg3 enhances the therapeutic effects of chemotherapy in patients with NSCLC. Rg3 treatment prolongs the overall survival time and improves the quality of life for the patients (57). Rg3 restores leucocyte count, decreases vascular endothelial growth factor (VEGF) expression and increases the CD4/CD8 T-cell ratio in these patients. Clinical outcomes show that Rg3 combined with chemotherapy enhances the efficacy and overall survival time of patients with NSCLC. Experimental research also suggests that ginsenoside overcomes the resistance to cisplatin in NSCLC. In cisplatin-resistant NSCLC (A549/DDP) cells, ginsenoside inhibits the NRF2 pathway and significantly sensitizes A549/DDP cells to cisplatin. NRF2 knockdown attenuates the synergistic effects of ginsenoside in cisplatin-resistant cells (58). A previous study also reported that Rg3 alleviated the resistance of A549/DDP cells to cisplatin treatment by decreasing PD-L1 expression via the inhibition of NF-κB p65 and Akt; therefore, restoring the cytotoxicity of T cells to the cancer cells (59).

Astragaloside IV is a bioactive saponin found in the roots of a Chinese medicinal herb Astragalus membranaceus (Fisch.) Bge. Astragaloside IV sensitizes NSCLC to gefitinib by regulating sirtuin 6 (SIRT6), and inhibiting SIRT6 abolishes the sensitization role of astragaloside IV in NSCLC cells including HCI-H1299, HCC827 and A549 cells (60). SIRT6 is an NAD-dependent deacetylase and promotes the metastasis of NSCLC. Depletion of SIRT6 inhibits transforming growth factor-β1-induced EMT in NSCLC cells, which underscores how astragaloside IV potentiates the anti-NSCLC effects of gefitinib.

PPI, PPII and PPVII are the main active saponins isolated from Paris polyphylla. PPI overcomes gefitinib resistance in NSCLC cells by decreasing cell viability and inducing apoptosis via downregulation of MALAT long non-coding RNA and inactivating the STAT3 signaling pathway (61). Both MALAT and STAT3 promote NSCLC growth. Overexpression of MALAT has been associated with a poor prognosis in patients with NSCLC (62), including recurrence and metastasis (63,64). Mechanistic studies show that MALAT activates Akt/mTOR signaling in NSCLC and promotes cancer cell migration and invasion. High expression of STAT3 in patients with NSCLC is also associated with a poor prognosis (65,66), and mediates drug resistance to EGFR-TKIs (67). Therefore, downregulating these two signaling molecules potentiates the anti-NSCLC effects of gefitinib. PPII restores sensitization of resistant NSCLC cells to gefitinib by inhibiting the PI3K/Akt/mTOR signaling pathway, thus potentiating the anti-proliferative effects of gefitinib and gefitinib-induced apoptosis (68). PPVII enhances the anti-proliferative effects of gefitinib and gefitinib-induced G₁ phase arrest by modulating the p21 signaling pathway in gefitinib-resistant NSCLC cells (69). Another study by Lou et al also reported that PPI overcame erlotinib resistance in NSCLC by reversing EMT and modulating the IL-6/STAT3 signaling pathway. Co-treatment of erlotinib and PPI completely abrogated the growth of NSCLC (70).

Both PPI and PPVII have a chemo-sensitizing effect; they overcome cisplatin resistance in NSCLC by enhancing apoptosis, reversing EMT, suppressing the cancerous inhibitor of protein phosphatase 2A (CIP2A)/AKT/mTOR signaling axis and activating autophagy in cancer cells (71,72).

Dioscin is a natural steroidal saponin abundantly found in Dioscorea nipponica Makino. Dioscin overcomes EGFR-TKI resistance in NSCLC by downregulating the expression of SH2 domain-containing phosphatase-2 (SHP2) at the transcriptional level through the induction of ROS production, and therefore decreases the binding of p53 to the SHP2 promoter (73). It is known that SHP2 is required for the growth of KRAS-mutant NSCLC, and the inhibition of SHP2 in KRAS-mutant NSCLC provokes a senescence response (74).

Capilliposide is a saponin in the Chinese medicinal herb Lysimachia capillipes; it inhibits AKT activity, increases the pro-apoptotic effect of gefitinib and decreases EGFR phosphorylation in gefitinib-resistant NSCLC cells (75).

Cucurbitacins, such as cucurbitacin B and D, are tetracyclic triterpenes isolated from the Cucurbitaceae plant family. Cucurbitacin B potently suppresses NSCLC growth by inducing G₂/M cell-cycle arrest and mitochondrial apoptosis. The anti-NSCLC effects of cucurbitacin B are attenuated by the thiol antioxidant N-acetylcysteine, while glutathione synthesis inhibitor butithione-sulfoxime exacerbates the anti-NSCLC effects (76). Further research suggests that NSCLC cells treated with cucurbitacin B have a significant decrease in the ratio of thiols and glutathione to oxidized glutathione, suggesting that disruption of the cellular redox balance mediates the anti-NSCLC effects of cucurbitacin B (77).

Cucurbitacin B and D also overcome gefitinib resistance in NSCLC cells. CIP2A inhibits protein phosphatase-2A (PP2A) (78), which suppresses tumor growth through dephosphorylation of oncogenic kinases and transcription factors (79). Cucurbitacin B suppresses the CIP2A/PP2A/Akt signaling axis in gefitinib-resistant cells and inhibits cancer growth and metastasis (80). Furthermore, the cucurbitacin B-mediated inhibition of the CIP2A/PP2A/Akt axis also induces the lysosomal degradation of EGFR in NSCLC cells (80). In a previous study, cucurbitacin D overcame gefitinib resistance by blocking EGF binding to EGFR, as demonstrated by a solid-phase EGF-EGFR interaction assay. In the analysis, cucurbitacin D directly inhibited the interaction between EGF and EGFR (81). Hence, cucurbitacin D decreases EGFR phosphorylation in gefitinib-resistant NSCLC cells (81).

Oridonin is a diterpenoid compound extracted from Isodon plant Rabdosia rubescens, Isodon japonicus Hara and I. trichocarpus (82). Oridonin inhibits the growth of gefitinib-resistant NSCLC cells. Oridonin decreases the expression of matrix metalloproteinase-12 (MMP-12) and CIP2A, suggesting that oridonin inhibits the growth and metastasis of gefitinib-resistant NSCLC by suppressing the EGFR/ERK/MMP-12 and CIP2A/Akt signaling pathways (83).

Artemisinin is a sesquiterpene lactone in the Chinese medicinal herb Artemisia annua L. In EGFR-TKI-resistant NSCLC cells, artemisinin induces apoptosis and cell cycle arrest at the G₁ phase. Artemisinin decreases Wnt/β-catenin protein levels by stimulating the release of naked cuticle homolog 2 (NKD2) and Axin2 (84). Both NKD2 and Axin2 are negative regulators of the Wnt/β-catenin signaling pathway. Downregulation of NKD2 causes Wnt activation and increases the invasive potential of NSCLC (85). Axin promotes the phosphorylation and the consequent degradation of β-catenin, and thus suppresses the Wnt/β-catenin signaling pathway (86). One study reported that the combination of artemisinin and onconase, a RNase of ribonuclease A, synergistically inhibited NSCLC growth (87). However, whether the combination of artemisinin and onconase can overcome gefitinib resistance in NSCLC is not known.

Dihydroartemisinin is the semi-synthetic derivative of artemisinin. It has been reported that treatment with dihydroartemisinin and gefitinib synergistically inhibits NSCLC cell growth and induces apoptosis via the Akt/mTOR/STAT3 pathway. Furthermore, this combination of treatment induces NSCLC cell cycle arrest in the G₂/M phase, which is associated with decreased expression of G₂/M regulatory proteins, including cyclin B1 and cyclin-dependent kinase 1. Combined treatment with dihydroartemisinin and gefitinib also significantly decreases the metastatic potential of NSCLC compared with treatment with gefitinib alone (88).

Matrine is an alkaloid isolated from Sophora flavescens. Treatment with a combination of matrine and afatinib, an EGFR-TKI, results in increased growth inhibitory effects of NSCLC cells by inhibiting the JAK/STAT3 signaling pathway (89), suggesting that matrine can overcome drug resistance to afatinib in NSCLC.

Furthermore, matrine also overcomes resistance to chemotherapy. In cisplatin-resistant NSCLC cells, matrine inhibits β-catenin activation and survivin expression, and induces apoptotic death, coupled with the loss of mitochondrial membrane potential and activation of caspase-9 and −3 (90).

Oxymatrine is one of the alkaloid components in Sophora flavescens; it inhibits NSCLC growth by suppressing the EGFR signaling pathway by inhibiting the activity of wild-type EGFR and EGFR with exon 19 deletion and L858R/T790M mutation (91). Further studies are needed to verify whether oxymatrine overcomes EGFR-TKI resistance in NSCLC.

Shikonin is a naphthoquinone compound extracted from the root of the Chinese medicinal herb Lithospermum erythrorhizon. Shikonin exhibits cytotoxicity, increases ROS production and induces apoptosis in NSCLC cells (92). Inhibiting ROS production blocks shikonin-induced apoptosis (92). Furthermore, shikonin also induces necroptosis, which is necrosis in a programmed fashion, mediated by signal transduction from receptor-interacting serine/threonine kinase (RIP)-1 to RIP-3. It has been reported that shikonin increases the expression of RIP-1 and induces necroptosis in NSCLC (93). Notably, the shikonin-induced necroptosis is enhanced by the inhibition of autophagy in NSCLC cell (93). Shikonin also sensitizes NSCLC to EGFR-TKI treatment. Shikonin in combination with erlotinib or gefitinib significantly enhances ROS-mediated apoptosis in NSCLC when compared with EGFR-TKI treatment alone (92,94). Furthermore, the combination of shikonin and gefitinib has synergistic anticancer effects via the inhibition of pyruvate kinase M2 (PKM2) and its downstream signaling molecules, including STAT3 and cyclin D1, in NSCLC cells (95). However, there is lack of consistent evidence suggesting the correlation of PKM2 and tumor grade in NSCLC (96). Another study has suggested that shikonin sensitizes gefitinib-resistant NSCLC cells by enhancing TRAIL-induced cytotoxicity via modulation of the STAT3, c-Jun N-terminal kinase and AKT signaling pathways (97). Preclinical research has shown that overexpression of STAT3 is correlated with chemoresistance and radioresistance in NSCLC cells (65). Further studies may clarify the role of STAT3 in gefitinib resistance in NSCLC. In addition, shikonin suppresses EGFR phosphorylation and increases EGFR proteasomal degradation in NSCLC (92). Decreasing EGFR expression inhibits its downstream signaling. Cbl is a RING E3/protein-ubiquitin ligase recruited to EGFR that mediates EGFR degradation. However, the role of Cbl in shikonin-mediated EGFR degradation has not been studied.

Tanshinone IIA is a diterpenoid naphthoquinone derived from the roots of Salvia miltiorrhiza. A recent study shows that it synergistically enhances the cytotoxic activity of geftinib in gefitinib-resistant NSCLC cells (98). Both in vitro and in vivo studies suggest that tanshinone IIA potentiates the anti-proliferative and anti-metastatic effects of geftinib in TKI-resistant cancer cells, which is attributed to the increased level of cleaved caspase 3 and the inhibition of the VEGF receptor-2/Akt signaling pathway (98).

Gambogenic acid is a polyprenylated xanthone isolated from the traditional Chinese medicinal herb gamboge. Gambogenic acid induces apoptosis in NSCLC cells by inhibiting the JAK/STAT3 signaling pathways (99); it also triggers autophagy by initiating fusion between autophagosomes and lysosomes, and inhibiting acidification in lysosomes, leading to cancer cell death (100). Furthermore, in a previous study, gambogenic acid abrogated the resistance to erlotinib in NSCLC, as demonstrated in a xenograft mouse model and patient-derived xenograft model by inhibiting c-Met activity and decreasing EGFR phosphorylation (99).

Gambogenic acid also overcomes resistance to cisplatin in NSCLC. In cisplatin-resistant NSCLC cells, gambogenic acid arrests the cell cycle at the G₁ phase by decreasing the expression of cyclin D1, cyclin dependent kinase (CDK)4 and CDK6, and upregulating p53 and p21. Moreover, gambogenic acid induces apoptosis by activating caspase-3 and −7 (101).

Cordyceps species such as C. Sinensis are traditional medicinal fungi. The main constituent of the extract derived from this fungus is cordycepin (102,103). Cordycepin, or 3′-deoxyadenosine, is a derivative of the nucleoside adenosine, differing from the nucleoside only by the absence of the hydroxy group in the 3′ position of its ribose moiety. Cordycepin was first found in fermented broth of the medicinal mushroom Cordyceps militaris (102). Cordycepin induces cell cycle arrest at the G₀/G₁ phase and apoptosis in lung cancer cells by inhibiting the phosphorylation of EGFR, Akt and ERK1/2 (104), and by interacting with and activating AMP-activated protein kinase (105,106). Cordycepin is also capable of inhibiting NSCLC cell cycle progression. NSCLC cells containing EGFR mutations are more sensitive to cordycepin treatment than those without EGFR mutations. More importantly, the therapeutic potency of cordycepin is comparable to afatinib and gefitinib, suggesting cordycepin itself, or in combination with gefitinib, can be a novel therapeutic for treating drug-resistant NSCLC (107).