Constitutive activity of Ras Raf MEK ERK
Constitutive activity of Ras/Raf/MEK/ERK pathway plays a critical role on melanoma growth, thus inhibition of activity of Ras, Raf and MEK can reduce ERK activity, leading to a decrease in melanoma cell proliferation, survival and invasion (Burotto et al., 2014; Paluncic et al., 2016). Our results also reveaed that U0126 alone could inhibit constitutive ERK phosphorylation and reduce survivin expression and cell viability, but had only a slight effect on the constitutive MKP-3 expression in B16 melanoma tyramide (Figs. 7C and 8B and D). These results imply that constitutive MEK activity can stimulate ERK activity to regulate survivin expression while constitutive MKP-3 expression may not be completely mediated by constitutive MEK/ERK signaling in B16 melanoma. Taking together, hinokitiol-induced activation of ERK/MKP-3/proteosome pathway was able to inhibit survivin expression rendering suppression of cell growth and cell migration and induction of apoptosis in B16 melanoma.
Conflicts of interest
Acknowledgements This study was supported by Grants MOST 104-2320-B-320-003 from the Ministry of Science and Technology of Taiwan, and Research Grant, KSB1105-029 and KSC106-014 from Kaohsiung Veterans General Hospital. The authors would like to thank Dr. Js kuo (Department of Pharmacology, Tzu Chi University) for his critical reading of the manuscript.
Introduction Gastric cancer (GC) is the fifth commonest malignancy and the third cause of cancer deaths worldwide [1,2]. An estimated 26,240 new GC cases and 10,800 GC-related deaths occurred in 2018 in the United States alone . GC is often diagnosed at the advanced stage and the prognosis for patients with advanced GC is poor with lower than 20% 5-year overall survival rate after neoadjuvant treatment [4,5]. Chemotherapy is an effective strategy to prevent the development of GC to a certain extent [4,6,7]. However, the existence and development of drug resistance especially multidrug resistance (MDR) is a massive obstacle for successful cancer therapies, strikingly limiting efficacy of chemotherapies . MDR is a phenomenon by which tumor cells develop cross-resistance to multiple structurally and mechanistically unrelated chemotherapeutic agents, limiting the prolonged and effective use of chemotherapeutic drugs . ATP-binding cassette (ABC) efflux transporters including ATP binding cassette subfamily B member 1 (ABCB1; multidrug-resistance gene 1; MDR1) and ATP binding cassette subfamily G member 2 (ABCG2) have been identified as vital mediators of MDR in various cancers including GC [8,10]. Cisplatin (CDDP), a platinum-based anti-cancer drug, has been widely used for the therapy of numerous cancers such as esophageal cancer and ovarian cancer . Moreover, DDP has been identified as the first-line chemotherapeutic drug for GC patients [2,12]. Vincristine (VCR), a vinca alkaloid derived from the plant Catharanthus roseus, has also been used as a chemotherapeutic agent for various cancers including pheochromocytoma and paraganglioma , glioma  and GC . Ribophorin II (RPN2), located on chromosome 20q12-13.1, has been reported to be an oncogene in multiple malignancies such as non-small-cell lung cancer (NSCLC) , osteosarcoma  and colorectal cancer . Also, RPN2 expression was associated with poor prognosis in some cancers including breast cancer , NSCLC  and GC . Previous studies indicated that RPN2 was closely linked with the drug resistance of tumor cells. For instance, RPN2 knockdown reduced the resistance of breast cancer and oesophageal squamous cell cancer cells to docetaxel by inhibiting P‑glycoprotein (P‑gp) glycosylation [21,22]. RPN2 silence relieved DDP resistance in NSCLC cells and NSCLC-bearing mice . Additionally, downregulation of RPN2 alleviated the resistance of GC cells to multiple chemotherapeutic agents [23,24].
Materials and methods
Discussion It has been well documented that ABC transporters-mediated MDR is a vital reason for cancer chemotherapy failure [8,27]. Consequently, it is imperative to seek for potential molecular targets or biomarkers to alleviate MDR. P-gp, encoded by MDR1/ABCB1 gene, can inhibit drug accumulation and cause MDR by ejecting drugs out from tumor cells in an energy-dependent manner . ABCG2 is implicated in the mediating MDR by reducing drug penetration and intestinal absorption . Correspondingly, the inhibition of P-gp and ABCG2 can alleviate MDR phenotypes and improve chemotherapeutic outcome [, , ]. The expressions of MDR1 and ABCG2 were positively associated with advanced pathological stages, higher differentiated level and poor prognosis in GC [, , ]. A prior report pointed out that DDP derived ABCG2 expression in GC . MDR1 mRNA level and P-gp protein level were upregulated in SGC7901/DDP and SGC7901/VCR cells [, , ]. Hence, the expression patterns of MDR1 and ABCG2 were measured at first in SGC7901/DDP and SGC7901/VCR cells. Our results showed that MDR1 mRNA expression, P-gp protein expression along with the mRNA and protein expressions of ABCG2 were upregulated in SGC7901/DDP and SGC7901/VCR cells as compared to those in SGC7901 parental cells, which was in line with previous reports [, , ]. Moreover, consistent with prior findings, SGC7901/DDP and SGC7901/VCR cells presented stronger resistance to DDP and VCR, respectively [35,37,38].