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Constitutive activation of the epidermal growth factor receptor (EGFR) signaling pathway is implicated in the initiation and progression of lung cancer. EGFR tyrosine kinase inhibitor (TKI)-targeted therapy has become the standard treatment for nonsmall cell lung cancer (NSCLC) patients. However, acquired resistance to these agents remains a major obstacle for managing NSCLC. Here, we investigated a novel strategy to overcome EGFR TKI resistance by targeting the stanniocalcin 2 (STC2)-JUN-AXL pathway. We revealed that STC2 was expressed at significantly higher levels in EGFR TKI-resistant cells. Further, clinical analysis showed that STC2 expression was increased after the development of EGFR TKI resistance and that higher levels were correlated with shorter progression-free survival in EGFR TKI-treated lung cancer patients. Moreover, STC2 overexpression in EGFR TKI-sensitive cells resulted in EGFR TKI resistance. Conversely, genetic silencing of STC2 rendered EGFR TKI-resistant cells more sensitive to EGFR TKIs. Mechanically, STC2 enhanced AXL promoter activity by increasing the phosphorylation of c-Jun, which is an indispensable transcription factor that transactivates AXL. STC2 promoted activation of the JUN-AXL-extracellular signal-regulated kinase (ERK) signaling axis in lung cancer cells. Pharmacological or genetic inhibition of AXL-ERK activity inhibited STC2-mediated EGFR TKI resistance. We also demonstrated that PE2988 cells, a C797S-independent osimertinib-resistant primary cancer cell line from a lung cancer patient, responded to combined AXL inhibitor and osimertinib treatment. In conclusion, our research indicates that STC2 overexpression is important for acquired resistance to EGFR TKIs and that STC2-JUN-AXL-ERK signaling might be a potential therapeutic target to overcome resistance to EGFR TKIs.

Four novel complexes based on the ligand piperazine (Pip) in a chair conformation have been synthesized, namely [Pd(5,5′-dcbpy)Pip]·4H2O (1), [Pt(5,5′-dcbpy)Pip]·3H2O (2), [Pd(4,4′-dcbpy)Pip]·4H2O (3), and [Pt(4,4′-dcbpy)Pip]·2H2O (4; 4,4′-dcbpy = 2,2′-bipyridyl-4,4′-dicarboxylate, 5,5′-dcbpy = 2,2′-bipyridyl-5,5′-dicarboxylate). The four complexes were characterized by elemental analysis, FTIR spectroscopy, 1H and 13C NMR spectroscopy, and single-crystal X-ray diffraction analysis. In addition, gel electrophoresis was performed on the four complexes and further insight into these complexes was gained by quantum chemistry calculations. The interaction of these complexes with fish sperm DNA (FS-DNA) was studied by UV/Vis and fluorescence spectroscopy, which revealed that they have the ability to bind to FS-DNA. The IC50 values of the four complexes range from 8.3 to 32.5 μM. Finally, we studied the cytotoxic activities of complexes 1–4 against two different cancer cell lines.

Kadsufolins A–D (1–4, resp.), four new dibenzocyclooctane-type lignans, were isolated from the roots and stems of Kadsura oblongifolia, together with eleven known lignans. Their structures and configurations were elucidated by spectroscopic methods including 2D-NMR techniques. The compounds were also evaluated for cytotoxic activity against human tumor cell lines A549 (lung carcinoma), DU145 (prostate carcinoma), KB (epidermoid carcinoma of the nasopharynx), and HCT-8 (ileocecal carcinoma). Kadsufolin A (1), kadsufolin D (4), angeloylbinankadsurin A, and heteroclitin B were found to show cytotoxic activities against A549, DU145, KB and HCT-8 with GI50 values of 5.1–20.0 μg/ml.

Twenty-six unsymmetrical biphenyls were synthesized and evaluated for cytotoxic activity against DU145, A549, KB and KB-Vin tumor cell lines. Three compounds 27 , 35 and 40 showed very potent activity against the HTCL panel with an IC 50 value range of 0.04–3.23 μM. In addition, fourteen active compounds were all more potent against the drug-resistant KB-Vin cell line than the parental KB cell line. Preliminary SAR analysis indicated that two bulky substituents on the 2,2′-positions of unsymmetrical biphenyl skeleton are necessary and crucial for in vitro anticancer activity, thus providing a good starting point to develop unsymmetrical biphenyls as novel anticancer agents.

The Calanthe genus in the Orchidaceae family contains terrestrial perennial herbs that are widely distributed from tropical Africa and Madagascar to tropical and subtropical Asia, China, Japan, southward through Malaysia and Indonesia to the Pacific islands and Australia. This genus includes more than 150 species, but only nineteen are found in Taiwan. Among them Calanthe arisanensis Hayata is endemic to Taiwan and grows in forests from 1000 to 2000 m throughout the island.1 A phytochemical study of this plant has not been reported to date. In cytotoxicity screening of extracts of Formosan plants, an EtOAc extract of C. arisanensis was found to be active against various human cancer cell lines with IC50< 20 µg/mL. Bioassay-directed chromatographic fractionation of this extract produced a new phenanthraquinone calanquinone A (1). Compound 1 showed significant in vitro cytotoxic activicity against seven human cancer cell lines, as described below. The active MeOH extract (225 g) of dry roots of C. arisanensis (5.42 kg) was partitioned between EtOAc and water (1:1, v/v). Further fractionation of the active EtOAc extract (32.7 g) by repeated liquid chromatography on silica gel gave calanquinone A (1). HRESIMS of 1 showed a [M-H]− ion at m/z 313.0705 (C17H14O6 -H), indicating 11 degrees of unsaturation. The IR spectrum showed absorptions for hydroxyl (3348 cm−1), carbonyl (1642 cm−1), and aromatic ring (1626, 1514, 1460, 1411, and 844 cm−1) functional groups. UV absorptions at 242, 308, and 426 nm also indicated an aromatic system. Seventeen carbon signals, including three methoxy, four methine, and ten quaternary carbons, were observed in the NMR spectra of 1 (Table 1). Among the ten quaternary carbons, two were identified as carbonyl carbons on the basis of chemical shifts δC 184.7 and 186.2. Therefore, the data supported the presence of two carbonyls, six olefins, and three ring moieties to fulfill the 11 degrees of unsaturation, and 1 was postulated to be an phenanthrenedione or anthrenedione.2 1D NMR and HSQC data indicated the presence of three methoxy groups at δH 3.96, 4.01, and 4.02 (δC 57.1, 56.2, and 61.0), one pair of ortho-coupled aromatic protons at δH 8.05 (d, J=8.7 Hz, δC 137.1) and 8.10 (d, J=8.7 Hz, δC 122.0), and two olefinic protons at δH 6.15 (s, δC 107.4) and 6.86 (s, δC 101.4). In the HMBC spectra, the olefinic proton at δH 6.86 exhibited 2J interactions with a carbon at δC 155.2 (C-7), as well as 3J interactions with carbons at δC 118.7 (C-4b), 140.4 (C-6), and 137.1 (C-9). The other olefinic proton at δH 6.15 exhibited 2J interactions with a carbon at δC 161.7 (C-3), as well as 3J interactions with carbons at δC 186.2 (C-4) and 133.0 (C-10a). Locations of methoxy groups at C-3, C-6, and C-7 were confirmed by the following NOESY correlations: δH 6.15 (H-2)/3.96 (3-OMe), δH 6.86 (H-8)/4.01 (7-OMe) and 8.05 (H-9), δH 4.01 (7-OMe)/4.02 (6-OMe), and δH 8.05 (H-9)/8.10 (H-10). Thus, compound 1 was identified as 5-hydroxy-3,6,7-trimethoxy-1,4-phenanthrenequinone and has been named as calanquinone A (1). Table 1 NMR data of calanquinone A (1)a Compound 1 is related in structure to other naturally occurring phenanthrenequinones, including the des-oxy analog sphenone (lacking the C-5 OH group),3 cymbinodin A (lacking the two methoxy groups at C-6 and C-7),4 and annoquinone A (lacking any substituents on ring C).5,6 In prior studies, sphenone and annoquinone A showed cytotoxic activity against the KB cell line (reported EC50 2.73 and 1.65 µg/mL, respectively). Compound 1 exhibited potent cytotoxicity (EC50 0.03–0.45 µg/mL) against human lung (A549), prostate (PC-3 and DU145), colon (HCT-8), breast (MCF7), nasopharyngeal (KB), and vincristine-resistant nasopharyngeal (KB-VIN) cancer cell lines. Paclitaxel was used as a positive control (data shown in Table 2). Interestingly, 1 exhibited comparble potency against both KB and its drug-resistant KB-VIN subline, and thus, showed an improved drug resistance profile compared to paclitaxel. The cytotoxic values demonstrate the strong potential of 1 as a promising lead compound and C. arisanensis as a promising plant source of new agents for cancer chemotherapy. Table 2 Cytotoxicity of calanquinone A (1) isolated from C. arisanensis In order to make sufficient quantities of 1 for extensive biological evaluation, we modified the synthetic procedure of Kraus and co-workers7,8 (Scheme 1) to synthesize 1. As shown in Scheme 2, we prepared 2-methoxy-5-carboxylic acid methyl ester-1,4-quinone (4)9 by AgO oxidation of the methyl ester (3) of commercially available 2,4,5-trimethoxybenzoic acid (2). Compound 4 was coupled with 3,4,5-trimethoxytoluene in the presence of 1 equiv. of trifluoroacetic acid to produce quinone 5, although Kraus reported the production of hydroquinone 10 under these reaction conditions (Scheme 1). The quinone skeleton of 5 was confirmed from 13C-NMR data, which showed two carbonyl groups at 180.6 and 183.6 ppm. In addition, reaction of 5 with Me2SO4 did not give a methoxylated product (i.e., 11 in Scheme 1). We attempted to reduce 5 with Na2S2O4 to obtain the corresponding hydroquinone, which could be transformed to the Kraus type of intermediate 11 (R = OMe) after methylation. Despite extending the reaction time and adding more reducing agent, only starting material was recovered. Scheme 1 Synthetic procedure of Kraus. Scheme 2 Our total synthesis of calanquinone A (1). Therefore, 5 was reduced with LAH (THF, reflux, 1 h) to alcohol, which was oxidized selectively to aldehyde 6 with activated MnO2 (toluene, 110 °C, overnight) (Scheme 2). After an unsuccessful attempt to obtain the phenanthraquinone 9 directly from 6 using P4-tBu, quinone 6 was first reduced to the hydroquinone using aq. Na2S2O4 (CH2Cl2, rt., overnight),10 and then methylated with Me2SO4 in the presence of K2CO3(acetone, 60 °C, 1.5 h) to give the desired compound 7 (Scheme 2). Cyclization of 7 with P4-t Bu (benzene, 100 °C, 63 h) gave 8, which was oxidized with AgO (6N HNO3, acetone, 50 °C, 2–3 min) to phenanthraquinone 9. Compound 9 was converted to calanquinone A (1) by selective demethylation with TMSI in CH2Cl2 at r.t. (Scheme 2). Synthesized 1 and intermediates 5–9 were screened in an in vitro cytotoxicity assay (data shown in Table 3). Compound 1 exhibited the highest potency (EC50 0.15–0.75 µg/mL) against all seven tested cancer cell lines. The remaining compounds showed no (6–8) or only weak (5 and 9) activity. Clearly, the potency of 1 merits further study and our synthetic route can efficiently produce sufficient quantities of 1 for future extensive biological evaluation and SAR investigation. Table 3 Cytotoxicity of synthesized 1 and related intermediates