Therapeutic Target For Chemoresistance Research Articles

MicroRNA-34a negatively regulates Netrin1 and mediates MEK/ERK pathway to regulate chemosensitivity of gastric cancer cells.

To explore the mechanism of action of MicroRNAs-34a (miR-34a) and Eurite growth guiding factor 1 (Netrin1) in cisplatin resistance in gastric cancer (GC), providing new clues for overcoming tumor resistance and optimizing anti-tumor therapy for GC. The Cancer Genome Atlas (TCGA), Differentially Expressed MicroRNAs (miRNAs) in human cancers (dbDEMC), and Starbase online databases were used to analyze the correlation between miR-34a and Netrin-1 and prognosis in GC, and to predict and verify the targeted binding of miR-34a to Netrin-1. The experimental methods including Cell transfection, real-time polymerase chain reaction (RT-PCR), Cell-Counting-Kit-8 (CCK8) assay, flow cytometry, wound scratch assay, transwell assay, and western blotting were used to investigate the effects of miR-34a and Netrin1 on chemotherapy resistance and biological characteristics in cisplatin-resistant GC cells (HGC27/DDP), and to analyze the molecular mechanism of cisplatin resistance. miR-34a expression was downregulated in gastric cancer clinical samples and cisplatin-resistant cells, while Netrin1 was upregulated, and was related to overall survival (OS). Upregulation of miR-34a can significantly reduce the IC50 value of cisplatin(0.65 vs 1.6 ng/mL) and Multidrug Resistance 1 (MDR-1) protein level, inhibit the proliferation activity, reduce the expression levels of proliferating cell nuclear antigen (PCNA) and ki-67 protein, and induce the increase of apoptosis rate and the enhancement of cycle arrest. Upregulation of miR-34a can also significantly reduce the expression level of Matrix metalloproteinase 9 (MMP9) protein, promote the expression of E-cadherin protein, reduce the wound healing rate and invasion number to inhibit migration and invasion ability in drug-resistant gastric cancer cells. Moreover, overexpression of Netrin1 on the basis of upregulation of miR-34a can weaken the above changes caused by upregulation of miR-34a. In addition, upregulation of miR-34a can significantly inhibit the Mitogen-activated protein kinase kinase (MEK) / Extracellular regulated protein kinases (ERK) pathway, while overexpression of Netrin1 can activate the MEK/ERK pathway, and inhibition of MEK/ERK pathway can effectively counteract the protein expression of Netrin1, and reverse changes in the expression of cisplatin IC50 and MDR-1 proteins caused by co-upregulation of miR-34a/Netrin1 in HGC27/DDP, as well as changes in proliferation, apoptosis, migration and invasion. In addition, upregulation of miR-34a can significantly inhibit the MEK/ERK pathway, while overexpression of Netrin1 can activate the MEK/ERK pathway. If the MEK/ERK pathway was inhibited, it can effectively counteract the protein overexpression of Netrin1, and reverse the changes in the expression of cisplatin IC50 and MDR-1 proteins in HGC27/DDP induced by co-upregulation of miR-34a / Netrin1, as well as changes in proliferation, apoptosis, migration and invasion. miR-34a targets and negatively regulates Netrin1 to mediate the proliferation, apoptosis, apoptosis, migration, and invasion of drug-resistant gastric cancer cells via the MEK/ERK pathway, and change the chemosensitivity in GC cells. miR-34a/Netrin1/MEK/ERK axis may serve as a novel therapeutic target for chemoresistance in GC, it is of great significance for overcoming drug resistance and developing new therapeutic strategies for GC.

Dichloroacetate Overcomes Oxaliplatin Chemoresistance in Colorectal Cancer through the miR-543/PTEN/Akt/mTOR Pathway.

Chemoresistance is responsible for most colorectal cancer (CRC) related deaths. In this study, we found that dichloroacetate (DCA), a pyruvate dehydrogenase kinase (PDK) inhibitor, can be used as a sensitizer for oxaliplatin (L-OHP) chemoresistant CRC cells. The aim of this study was to explore the ability of DCA to overcome L-OHP resistance in CRC cells and to identify the underlying molecular mechanisms. We found that DCA sensitizes chemoresistant CRC cells to L-OHP-induced cytotoxic effects by inhibiting clone formation capacity and promoting cell apoptosis. A microRNA (miRNA) array was used for screen, and miR-543 was identified and shown to be downregulated after DCA treatment. The expression of miR-543 was higher in chemoresistant CRC cells than in chemosensitive CRC cells. Overexpression of miR-543 increased chemoresistance in CRC cells. The validated target gene, PTEN, was negatively regulated by miR-543 both in vitro and in vivo, and PTEN was upregulated by DCA through miR-543. In addition, overexpression of miR-543 reversed the inhibition of colony formation after DCA treatment. Furthermore, the Akt/mTOR pathway is activated by miR-543 and is involved in the miR-543 induced chemoresistance. There was a significant inverse relationship between miR-543 expression and PTEN level in CRC patients, and high miR-543 expression was associated with worse prognosis. In conclusion, DCA restored chemosensitivity through miR-543/PTEN/Akt/mTOR pathway, and miR-543 may be a potential marker or therapeutic target for chemoresistance in CRC.

Abstract 5840: Inhibition of cystathionine-β-synthase (CBS) sensitizes colon cancer cells to 5-FU via increasing apoptosis and inhibiting cellular bioenergetics

Abstract Increased expression of cystathionine-β-synthase (CBS) has been validated in multiple kinds of tumor tissues, including colorectal cancer. Endogenous hydrogen sulfide (H2S), produced by CBS, was able to promote the proliferation and migration of colon cancer cells by inhibiting apoptosis and injuries caused by noxious environmental factors, including chemotherapeutic agents. Recently, overexpression of CBS has been reported to play a pivotal role in tumorigenesis of colon cancer cells, rendering the normal colon epithelial cell line acquiring the features of malignant cells. However, the potential role of CBS in the development of chemoresistance of colon cancer cells has not be illustrated. In the present study, overexpression of CBS in tumor tissues was further validated by realtime PCR and the effect of inhibition of both the activity and expression of CBS on the sensitivity to 5-fluorouracil (5-FU) was investigated in colon cancer cell lines. Inhibition of the activity of CBS by CBS inhibitor aminooxyacetate (AOAA) significantly increased the sensitivity of colon cancer cell lines to 5-FU, featured by decreased IC50 and colony forming efficiency. However, utilizing a slowly releasing H2S donor GYY4137, the present study suggested that exogenous hydrogen sulfide was able to increase the resistance to 5-FU in both colon cancer cell lines. What's intriguing was that, 5-FU, at clinically relevant concentrations, significantly inhibited the expression of CBS. Co-treatment of AOAA significantly increased the expression of Bax and decreased the expression of Bcl-2, indicating the involvement of increased apoptosis. AOAA also predisposed the colon cancer cells to the inhibition of cellular energetics induced by 5-FU, featured by the further reduced level of cellular ATP. These results suggested that inhibition of CBS promoted the sensitivity of colon cancer cells to 5-FU via increasing apoptosis and inhibiting cellular bioenergetics. Furthermore, inhibition of CBS expression by shRNA sensitizes the 5-FU resistant colon cancer cell line HCT-8/5-FU to treatment with 5-FU. The increased expression of CBS and the consequently increased level of endogenous H2S might be a potential therapeutic target for chemoresistance in colon cancer. Citation Format: Shanwen Chen, Pengyuan Wang, Yisheng Pan, Yucun Liu. Inhibition of cystathionine-β-synthase (CBS) sensitizes colon cancer cells to 5-FU via increasing apoptosis and inhibiting cellular bioenergetics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5840.

Hypoxic tumor microenvironment activates GLI2 via HIF-1α and TGF-β2 to promote chemoresistance in colorectal cancer

Colorectal cancer patients often relapse after chemotherapy, owing to the survival of stem or progenitor cells referred to as cancer stem cells (CSCs). Although tumor stromal factors are known to contribute to chemoresistance, it remains not fully understood how CSCs in the hypoxic tumor microenvironment escape the chemotherapy. Here, we report that hypoxia-inducible factor (HIF-1α) and cancer-associated fibroblasts (CAFs)-secreted TGF-β2 converge to activate the expression of hedgehog transcription factor GLI2 in CSCs, resulting in increased stemness/dedifferentiation and intrinsic resistance to chemotherapy. Genetic or small-molecule inhibitor-based ablation of HIF-1α/TGF-β2-mediated GLI2 signaling effectively reversed the chemoresistance caused by the tumor microenvironment. Importantly, high expression levels of HIF-1α/TGF-β2/GLI2 correlated robustly with the patient relapse following chemotherapy, highlighting a potential biomarker and therapeutic target for chemoresistance in colorectal cancer. Our study thus uncovers a molecular mechanism by which hypoxic colorectal tumor microenvironment promotes cancer cell stemness and resistance to chemotherapy and suggests a potentially targeted treatment approach to mitigating chemoresistance.

MicroRNA-410-3p attenuates gemcitabine resistance in pancreatic ductal adenocarcinoma by inhibiting HMGB1-mediated autophagy.

Gemcitabine-based chemotherapy is the most common treatment option for pancreatic ductal adenocarcinoma (PDAC). However, it offers little therapeutic value in many cases due to the rapid development of chemoresistance. MicroRNAs (miRNAs) have been found to play pivotal roles in the chemotherapeutic resistance of PDAC. We found that miR-410-3p was significantly down-regulated in human pancreatic cancer xenograft (HPCx) tumor tissues from gemcitabine-treated mice. Low miR-410-3p expression correlated with gemcitabine resistance in HPCx tumors and PDAC cells as well as poor prognosis in PDAC patients. We also found that miR-410-3p attenuated the gemcitabine resistance of PDAC by targeting the 3′-UTR of HMGB1. Moreover, our study clearly demonstrated that miR-410-3p enhanced chemosensitivity to gemcitabine via inhibiting HMGB1-induced autophagy during chemotherapy in PDAC cells. Our study suggests that miR-410-3p expression may be a useful indicator of the potential for chemoresistance to gemcitabine and provide a potential new therapeutic target for chemoresistance in PDAC.

SKA1: A therapeutic target for chemo-resistance in human osteosarcoma

SKA1: A therapeutic target for chemo-resistance in human osteosarcoma

Abstract 986: Long noncoding RNA CCAT2 cooperates with miR-145 and miR-21 to regulate colon cancer stem cells

Abstract Colon cancer-associated transcript-2 (CCAT2), a long noncoding RNA (lncRNA), has been reported to be over-expressed in colorectal cancer to promote tumor growth and metastasis and also reduces sensitivity to chemotherapy, a property related to cancer stem cells (CSCs). MicroRNAs, small endogenous noncoding RNAs, negatively control the expression of target genes by cleaving mRNA or through translation repression. We have recently reported cooperation of miR-21 and miR-145 to regulate colon CSCs proliferation and differentiation. Down-regulation of miR-21 enhances CSC enriched chemo-resistant colon cancer cells’ susceptibility to chemo- therapeutic regimens. Although both CCAT2 and miR-21 increase WNT signaling activity and reduce chemosensitivity to 5-FU, a critical evidence on how CCAT2 regulates stemness leading to chemo-resistance is lacking. The primary goal of this investigation was to examine whether CCAT2 associates with miR-145 and miR-21 and whether this association plays a role in regulating stemness of colon cancer cells. We have observed that CCAT2 transcript level is dependent on p53 expression and activity, as evidenced by a marked increase in CCAT2 expression in p53 wild type harboring HCT-116 colon cancer cells compared with p53 null HCT-116 and p53 mutant HT-29 cells. We further revealed that knockdown of CCAT2 increased the expression of miR-145 and negatively regulates miR-21 in HCT-116 cells and impairs proliferation and differentiation of these cells. On the other hand, upregulation of CCAT2 induces proliferation of HCT-116 cells and also increases the expression of CSC markers CD44 and CD166. We have also observed that miR-145 and miR-21 cooperation plays a role in regulating colon cancer stem cells by down-regulating Sox2 and increased activity of Wnt/β-catenin pathway. Our current results suggest that lncRNA CCAT2 cooperates with miR-145 and miR-21 to regulate colon cancer stem cells, which provides a potential therapeutic target for chemoresistance in colorectal cancer. Citation Format: Yingjie Yu, Pratima Nangia-Makker, Lulu Farhana, Adhip P. N. Majumdar. Long noncoding RNA CCAT2 cooperates with miR-145 and miR-21 to regulate colon cancer stem cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 986.

Interaction of translationally controlled tumor protein with Apaf-1 is involved in the development of chemoresistance in HeLa cells

BackgroundTranslationally controlled tumor protein (TCTP), alternatively called fortilin, is believed to be involved in the development of the chemoresistance of tumor cells against anticancer drugs such as etoposide, taxol, and oxaliplatin, the underlying mechanisms of which still remain elusive.MethodsCell death analysis of TCTP-overexpressing HeLa cells was performed following etoposide treatment to assess the mitochondria-dependent apoptosis. Apoptotic pathway was analyzed through measuring the cleavage of epidermal growth factor receptor (EGFR) and phospholipase C-γ (PLC-γ), caspase activation, mitochondrial membrane perturbation, and cytochrome c release by flow cytometry and western blotting. To clarify the role of TCTP in the inhibition of apoptosome, in vitro apoptosome reconstitution and immunoprecipitation was used. Pull-down assay and silver staining using the variants of Apaf-1 protein was applied to identify the domain that is responsible for its interaction with TCTP.ResultsIn the present study, we confirmed that adenoviral overexpression of TCTP protects HeLa cells from cell death induced by cytotoxic drugs such as taxol and etoposide. TCTP antagonized the mitochondria-dependent apoptotic pathway following etoposide treatment, including mitochondrial membrane damage and resultant cytochrome c release, activation of caspase-9, and -3, and eventually, the cleavage of EGFR and PLC-γ. More importantly, TCTP interacts with the caspase recruitment domain (CARD) of Apaf-1 and is incorporated into the heptameric Apaf-1 complex, and that C-terminal cleaved TCTP specifically associates with Apaf-1 of apoptosome in apoptosome-forming condition thereby inhibiting the amplification of caspase cascade.ConclusionsTCTP protects the cancer cells from etoposide-induced cell death by inhibiting the mitochondria-mediated apoptotic pathway. Interaction of TCTP with Apaf-1 in apoptosome is involved in the molecular mechanism of TCTP-induced chemoresistance. These findings suggest that TCTP may serve as a therapeutic target for chemoresistance in cancer treatment.