The use of modern genomic, proteomic and functional analytical techniques has resulted in a substantial increase in our ability to identify novel genes and signaling networks that are involved in determining the responsiveness of tumors to a particular drug treatment. The mechanisms of resistance to ‘classical’ cytotoxic chemotherapeutics and to therapies that are designed to be selective for specific molecular targets share many features, such as alterations in the drug target, activation of pro-survival pathways and ineffective induction of cell death [
9]. Pre-clinical and clinical studies have already shown that combining information from more than one molecular biomarker increases our ability to predict tumour drug response [
19]. Hence, understanding how tumors evolve on a molecular level to overcome the cytotoxic effects of chemotherapy is a critical step in developing therapeutic approaches that will prevent or overcome chemoresistance [
20].
Alterations in gene expression profiles and epigenetic changes are known to be early events in the multi-steps of carcinogenesis [
21]. On an epigenome-wide basis, multiple DNA methylation changes in the cancer methylome have been reported to occur during the acquisition of drug resistance [
22]. Here, we explored the contributory role of cell cycle dependent protein GTSE1 in gastric cancer chemoresistance. GTSE1 is highly expressed in cancers and was shown to be clinically associated with drug resistance [
18]. Analysis of public gene array databases showed high GTSE1 expression in gastric cancer [
23‐
25]. Since aberrant DNA methylation as a marker of platinum-resistance was shown in lung and ovarian cancer cell lines [
26,
27]; we examined the methylation status of GTSE1 in gastric cancer patients who underwent neoadjuvant DCX-combinational chemo therapy. Although GTSE1 appeared hypomethylated in both responders and non responders of DCX treatment, a clear distinction could be seen between the two groups wherein responders showed a higher methylation index. However, the current study was unable to show any significant survival benefit of GTSE1 methylation in gastric cancer patients. This could be due to the small sample size and thus hinders evaluation of GTSE1 methylation as a predictive biomarker for DCX treatment response. Consistently, GTSE1 was shown to be hypomethylated in gastric cancer cell lines. All hypomethytlated cell lines showed significant expression levels of GTSE1. Intriguingly, cell lines that are sensitive to cisplatin treatment showed a significantly different methylation index for GTSE1. Since, the predominant model of regulation of gene expression underscores an inverse correlation between DNA methylation and expression [
28], the higher methylation index of GTSE1 in DCX responders and in cisplatin sensitive cell lines could be suggestive of its lower expression levels. However, this study did not investigate this correlation in the DCX patient cohort. Hence, clinical implications of GTSE1 methylation status needs further investigation regarding the possible rationales raised in this study
Gene expression profiling in cancer have provided substantial information on the oncogenic potential of genes controlling essential pathways and other cellular events. Cell proliferation and growth inhibition are tightly regulated processes which are key in the maintenance of normal cell growth homeostasis and viability [
29]. However deregulation of these processes occur during tumor development and is often contributory to drug resistance. The present study demonstrated a major role of GTSE1 in conferring cisplatin resistance as knock down of GTSE1 expression in gastric cancer cells enhanced cisplatin sensitivity. Moreover, cisplatin treatment induced a dose dependent upregulation as well as nuclear translocation of GTSE1 in gastric cancer cells. The increase in expression and change in membrane localization of GTSE1 falls in line with a previous study that showed up regulation and nuclear import of this protein in response to various DNA- damaging agents [
14]. The nuclear translocation of GTSE1 is also suggestive of the cellular response to cisplatin induced DNA damage stress, where its expression counteracts with drug sensitivity. Enhanced proliferation and colony formation ability of cells are well associated with malignant transformation in many cancers [
30]. Although anti-cancer therapy targets apoptosis of hyper-proliferative cells, its efficacy may vary according to cancers and its molecular sub-types. In our study, loss of GTSE1 expression enhanced the anti-proliferative and growth-inhibitory effects of cisplatin in gastric cancer cells. Of note, the decrease in cell proliferation and colony formation observed in GTSE1 knocked-down cells is also suggestive of its tumorogenic potential that could be utilized in targeted therapies in gastric cancer. Consistent with these observations, expression of GTSE1 was found inhibitory to cisplatin cytotoxicity as its knock down lead to a two-fold increase in apoptosis of gastric cancer cells. This increase in apoptosis could well be explained by the observed increase of p53 expression in GTSE1 knocked-down cells. P53 is a potent inhibitor of cell growth and its function is tightly controlled to allow normal growth development [
31]. In response to DNA damage insults, p53 induces cell cycle arrest and activates the intrinsic apoptotic signaling pathway [
32]. P53 up regulation also resulted in activation of its downstream effectors- bax. This could attribute to chromatin condensation, DNA fragmentation and finally apoptosis [
33]. Hence, up regulation of p53 and its downstream effectors in cisplatin treated cells can be inferred as a cellular response to DNA damage. Therefore, by identifying a reduced expression profile of p53 in normal cells against GTSE1 knocked-down cells, our study demonstrates a role of GTSE1 in attenuating p53 mediated apoptotic response in cisplatin treated gastric cancer cells. However, we did not see any significant difference in caspase 3 activation in GTSE1 knock down cells upon cisplatin treatment, as compared to its parental counterparts (Additional file
1: Figure S4). Our findings suggest that GTSE1 mediates a caspase 3 independent cascade of apoptotic repression in cisplatin treated gastric cancer cells. This is consistent with previous studies by and Cui
et al. and Hu
et al. that have shown a p53 dependent but caspase 3 independent mechanism of apoptosis [
34,
35]. In addition, GTSE1 expression was also seen to be associated with high expression of p21 as knockdown of GTSE1 caused a reduction in p21 turn over. This falls in agreement with studies by Bublik
et al. that showed GTSE1 as a regulator of p21 stability by protecting it from proteosome-dependent degradationEarlier, the association of GTSE1 with p21 was shown to modulate cellular response to paclitaxel induced apoptosis in cervical cancer cells [
36]. Moreover, high levels of p21 in cancers have been linked to poor prognosis [
37]. Thus, by oppositely regulating p53 and p21, GTSE1 protein may display a combined role in promoting cell survival by shifting the equilibrium of p53 response from apoptosis to survival [
23]. Hence, targeting GTSE1 in cancer therapy could enhance a p53 mediated pro-apoptotic response with a reflective decrease in p21 induced cell cycle arrest.