Oxaliplatin-resistant Colorectal Cancer Research Articles

LINC00460-miR-149-5p/miR-150-5p-Mutant p53 Feedback Loop Promotes Oxaliplatin Resistance in Colorectal Cancer

Oxaliplatin resistance is a major challenge in the clinical treatment for advanced colorectal cancer (CRC). Long non-coding RNAs (lncRNAs) are involved in tumorigenesis and progression as critical regulators, while their potential roles in chemoresistance are poorly understood. In this study, we report that the LINC00460-miR-149-5p/miR-150-5p-mutant p53 feedback loop is responsible for oxaliplatin resistance in CRC. First, LINC00460 was found to exhibit higher expression in oxaliplatin-resistant CRC (CRC/OxR) cells compared with parental oxaliplatin-sensitive ones, and this expression pattern depends on mutant p53 (SW480/OxR), not wild-type p53 (HCT116/OxR). Oxaliplatin-induced LINC00460 in SW480/OxR cells was mainly located in the cytoplasm and was associated with AGO2 protein. LINC00460 functions as a competing endogenous RNA (ceRNA) to promote oxaliplatin resistance through sequestering miR-149-5p/miR-150-5p and upregulating the expression of the microRNA (miRNA) target p53. Knockdown of LINC00460 sensitized SW480/OxR cells to oxaliplatin by modulating p53 in vitro and in vivo. In turn, mutant p53 positively regulated the expression of LINC00460, thus forming a feedback loop. Clinical data showed that LINC00460 was upregulated in CRC tissues compared with paired normal tissues and was significantly correlated with clinical stage and node (N) status. Our findings uncover a mechanism for the LINC00460-miR-149-5p/miR-150-5p-mutant p53 feedback loop in oxaliplatin resistance of CRC, and they provide potential therapeutic targets for tumor chemoresistance.

Genome-wide single-nucleotide resolution of oxaliplatin–DNA adduct repair in drug-sensitive and -resistant colorectal cancer cell lines

Platinum-based chemotherapies, including oxaliplatin, are a mainstay in the management of solid tumors and induce cell death by forming intrastrand dinucleotide DNA adducts. Despite their common use, they are highly toxic, and approximately half of cancer patients have tumors that are either intrinsically resistant or develop resistance. Previous studies suggest that this resistance is mediated by variations in DNA repair levels or net drug influx. Here, we aimed to better define the roles of nucleotide excision repair and DNA damage in platinum chemotherapy resistance by profiling DNA damage and repair efficiency in seven oxaliplatin-sensitive and three oxaliplatin-resistant colorectal cancer cell lines. We assayed DNA repair indirectly as toxicity and directly measured bulky adduct formation and removal from the genome by slot blot and repair capacity in an excision assay, and used excision repair sequencing (XR-seq) to map repair events genome-wide at single-nucleotide resolution. Using this combinatorial approach and proxies for oxaliplatin-DNA damage, we observed no significant differences in repair efficiency that could explain the relative sensitivities and chemotherapy resistances of these cell lines. In contrast, the levels of oxaliplatin-induced DNA damage were significantly lower in the resistant cells, indicating that decreased damage formation, rather than increased damage repair, is a major determinant of oxaliplatin resistance in these cell lines. XR-seq-based analysis of gene expression revealed up-regulation of membrane transport pathways in the resistant cells, and these pathways may contribute to resistance. In conclusion, additional research is needed to characterize the factors mitigating cellular DNA damage formation by platinum compounds.

Inhibition of miR-19a partially reversed the resistance of colorectal cancer to oxaliplatin via PTEN/PI3K/AKT pathway.

Oxaliplatin is a platinum-based chemotherapeutic drug that is effective and commonly used in the treatment of colorectal cancer (CRC). However, long-term use of oxaliplatin usually induces significant drug resistance. It is urgent to develop strategies to reverse the oxaliplatin resistance to CRC cells. In the present study, we established the model of oxaliplatin-resistant CRC cell lines (SW480/R and HT29/R) through continuous treatment of SW480 and HT29 cells with oxaliplatin. Results of qRT-PCR analysis showed that expression of miR-19a was significantly increased in SW480/R and HT29/R compared to their parental SW480 and HT29. However, combination treatment with anti-miR-19a, an antisense oligonucleotide of miR-19a, was found to resensitize SW480/R and HT29/R cells to oxaliplatin treatment. In the mechanism research, we found that anti-miR-19a increased the expression of PTEN and thus inhibited the phosphorylation of PI3K and AKT in SW480/R and HT29/R cells. As a result, mitochondrial apoptosis induced by oxaliplatin was expanded. We demonstrated that PTEN was the target of miR-19a and inhibition of miR-19a partially reversed the resistance of colorectal cancer to oxaliplatin via PTEN/PI3K/AKT pathway.

Thioredoxin domain-containing protein 9 (TXNDC9) contributes to oxaliplatin resistance through regulation of autophagy-apoptosis in colorectal adenocarcinoma

Colorectal cancer (CRC) is the third most commonly malignancy worldwide. The incidence of CRC is on the rise and leads to indisputable society burden due to the high cost of cancer treatments. Resistance to oxaliplatin-chemotherapy is the major cause for treatment failure and CRC-related death. In this study, we anticipated that TXNDC9 might demonstrate a protective role in oxaliplatin-resistant CRC cells. TXNDC9 was found significantly upregulated when treated with oxaliplatin. Manipulation of TXNDC9 expression largely affected the oxaliplatin-induced cell death. Moreover, TXNDC9 regulates autophagy and apoptosis in response to oxaliplatin treatment in HT29 cells via the Nrf2 pathway. Taken together, our findings explore the biological role of TXNDC9 in oxaliplatin resistance in CRC cells and may identify a novel therapeutic target to counteract drug resistance to oxaliplatin in CRC.

A role for ceramide glycosylation in resistance to oxaliplatin in colorectal cancer

There is growing evidence to support a role for the ceramide-metabolizing enzyme, glucosylceramide synthase (GCS), in resistance to a variety of chemotherapeutic agents. Whether GCS contributes to oxaliplatin resistance in colorectal cancer (CRC) has not yet been determined. We have addressed this potentially important clinical issue by examining GCS function in two panels of oxaliplatin-resistant, isogenic CRC cell lines. Compared to parental cell lines, oxaliplatin-resistant cells have increased expression of GCS protein associated with increased levels of the pro-survival ceramide metabolite, glucosylceramide (GlcCer). Inhibition of GCS expression by RNAi-mediated gene knockdown resulted in a reduction in cellular GlcCer levels, with restored sensitivity to oxaliplatin. Furthermore, oxaliplatin-resistant CRC cells displayed lower ceramide levels both basally and after treatment with oxaliplatin, compared to parental cells. GlcCer, formed by GCS-mediated ceramide glycosylation, is the precursor to a complex array of glycosphingolipids. Differences in cellular levels and species of gangliosides, a family of glycosphingolipids, were also seen between parental and oxaliplatin-resistant CRC cells. Increased Akt activation was also observed in oxaliplatin-resistant CRC cell lines, together with increased expression of the anti-apoptotic protein survivin. Finally, this study shows that GCS protein levels are greatly increased in human CRC specimens, compared to matched, normal colonic mucosa, and that high levels of UGCG gene expression are significantly associated with decreased disease-free survival in colorectal cancer patients. These findings uncover an important cellular role for GCS in oxaliplatin chemosensitivity and may provide a novel cellular target for augmenting chemotherapeutic drug effectiveness in CRC.

Abstract C014: Development of an injectable water-soluble anticancer drug CMG for oxaliplatin-resistant colorectal cancer with KRAS mutation

Abstract Recent well-designed preclinical studies have demonstrated that KRAS mutations can activate the canonical NF-κB signaling pathway, which is a pivotal role in tumor progression. Therefore, inhibition of this KRAS-NF-κB axis would be a promising target to develop a new molecular targeted anticancer agent. Curcumin, a naturally occurring polyphenol derived from the plant Curcuma longa, has been shown to block NF-κB signaling pathway via at least two mechanisms; inhibition of IkB phosphorylation and inhibition of ubiquitin-dependent degradation of IκB. Recently dual-specificity tyrosine-regulated kinase (DYRK2), a positive regulator of the 26S proteasome, has been reported as a direct molecular target of curcumin, which is consisted with the inhibition of NF-κB signaling pathway by curcumin. Several preclinical studies have clearly demonstrated that curcumin exhibits anticancer effects in colorectal cancer model through NF-κB inhibition. However, since curcumin is highly lipophilic and orally administrated curcumin has very low bioavailability, its clinical application in cancer patients has been hampered. Very recently we developed a synthetic water-soluble prodrug CMG, curcumin mono-β-D-glucuronide, to overcome these problems. In this study, we investigated the anticancer effect of CMG on oxaliplatin (L-OHP) -resistant colorectal cancer cells HCT116 with KRAS mutation and p53 deletion (KRAS+p53- cells). Curcumin, the active substance released from CMG in vivo, showed similar effects regardless of KRAS/p53 mutation status in vitro. In xenograft models using KRAS+p53- HCT116 cells, intraperitoneal administration of CMG significantly superior antitumor effects compared to L-OHP monotherapy. Notably, no adverse events were noted in the CMG group, whereas bodyweight loss, severe myelosuppression, and liver damage were observed in the L-OHP group. Combination of CMG and L-OHP showed additive antitumor effects with moderate suppression of liver damage by L-OHP. These results indicate that CMG could be a novel anticancer drug with minimal toxicities for colon cancer patients with KRAS and p53 mutations. Citation Format: Hideaki Kakeya, Atsuhiro Kishimoto, Hitomi Umeta, Atsushi Imaizumi, Masashi Kanai. Development of an injectable water-soluble anticancer drug CMG for oxaliplatin-resistant colorectal cancer with KRAS mutation [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C014. doi:10.1158/1535-7163.TARG-19-C014

Overexpression of miR-106a enhances oxaliplatin sensitivity of colorectal cancer through regulation of FOXQ1.

Chemotherapy resistance poses a major challenge for the clinical treatment of colorectal cancer, therefore, the aim of the present study was to examine its underlying mechanisms. Reverse transcription-quantitative polymerase chain reaction and western blot analysis were used to determine the microRNA (miRNA)/mRNA and protein expression levels, respectively. A dual luciferase assay was conducted for verification of the interaction between miR-106a and 3′untranslated region (UTR) of Forkhead box Q1 (FOXQ1). Cell viability was assessed using an MTT assay. In the present study, it was demonstrated that miR-106a is involved in regulating oxaliplatin sensitivity of colorectal cancer. Transfection of miR-106a mimics slightly inhibited colorectal cancer cell growth and sensitized colorectal cancer cells to oxaliplatin exposure. In addition, miR-106a overexpression induced a decrease of FOXQ1 at mRNA and protein levels in colorectal cancer cells. The enhanced expression of miR-106a also increased the expression of Wnt target genes, including vascular endothelial growth factor-A and matrix metallopeptidase 2, which were reported to be regulated by FOXQ1. It was predicted and validated that miR-106a could repress FOXQ1 expression via direct binding to 3′UTR. Elevation of miR-106a and a decrease of FOXQ1 expression levels were detected in tumor tissues from patients with oxaliplatin-sensitive colorectal cancer, compared with patients with oxaliplatin-resistant colorectal cancer. Furthermore, there was a significant association between miR-106a and FOXQ1 mRNA levels. In conclusion, the present study demonstrated that miR-106a increased oxaliplatin sensitivity of colorectal cancer cells through direct repression of FOXQ1 expression.

Novel exosomal miR-46146 transfer oxaliplatin chemoresistance in colorectal cancer.

Oxaliplatin (OX) is widely used for patients with advanced colorectal cancer (CRC). However, most of them will turn out to be OX resistant. Therefore, it is necessary to uncover the causes underlying this phenomenon. Emerging works have reported that exosomal miRNAs are linked to chemoresistance in many types of cancer. Hence, we separated exosomes from OX sensitive (Exo-S) and resistant CRC cells (Exo-R) by ultracentrifugation and characterized those exosomes by transmission electron microscope and Nanosight NS300. The differentiated miRNAs between Exo-S and Exo-R were identified by small RNA deep sequencing. The expression of miRNA was examined by quantitative real-time PCR (qRT-PCR). The effect of Exo-R and exosomal miR-46146 was determined by CCK-8 assay and flow cytometry (FCM). The target gene of miR-46146 was predicted by computational algorithms and validated by dual luciferase assay. We found that parental OX sensitive CRC cell acquired increased resistance to the cytotoxicity of OX when they were cocultured with exosomes secreted by OX-resistant CRC. Notably, a novel miRNA miR-46146 was identified and proved to be upregulated in the Exo-R which was internalized by its recipient cells and contributes to the chemoresistance transfer. Furthermore, we demonstrated that PDCD10 was the direct functional target of miR46146 and augmentation of the PDCD10 expression might reverse the effect of Exo-miR-46146-driven chemoresistance. These results indicate that exosomal miR-46146 functions as a vital mediator of OX resistance by targeting PDCD10 and could be a potential target to re-sensitize CRC cell to OX.

TLR 7/8 agonist reverses oxaliplatin resistance in colorectal cancer via directing the myeloid-derived suppressor cells to tumoricidal M1-macrophages.

Resistance to oxaliplatin is a major obstacle hindering the clinical treatment of colorectal cancer, but the underlying immunological mechanism has not yet been well illustrated. As a pivotal immunosuppressive component in tumor microenvironment, myeloid-derived suppressor cells (MDSCs) and their differentiated tumor-associated macrophages (TAMs) have been considered to be associated with resistance to chemotherapy. The aim of current study was to investigate the role of MDSCs in oxaliplatin-resistance and antitumor activity in colorectal cancer. Here, we found that tumor-bearing mice treated with oxaliplatin performed remarkably decreasing M-MDSCs and M1-type TAMs differentiated from MDSCs in tumor site, which inspired us to combine immunotherapy that activates M1-like TAMs to conquer oxaliplatin resistance. In addition, this study further confirmed a dose-dependent improvement of M1-like macrophage supernatants on enhancing pro-apoptotic effect and inhibiting migration and invasion of cancer cells incubated with oxaliplatin. Administration of oxaliplatin combined with Toll-like receptors agonists R848 reversed the functional orientation of MDSCs towards M1-like macrophages and strengthened antitumor effect of oxaliplatin. In this study, we uncovered novel immunological mechanism of oxaliplatin-resistance and showed the great potential of TLR7/8 agonist as a new immunologic adjuvant in chemotherapy for oxaliplatin-resistant colorectal cancer.

Development of new treatment using CMG for oxaliplatin-resistant colorectal cancer with KRAS mutation

Abstract Background KRAS mutations are known as poor prognostic factor for patients with colon cancer who receive chemotherapy. KRAS mutations have been shown to activate NF-kappa B (NF-kB) signaling pathway, that plays a pivotal role in cancer progression or chemoresistance. In this study, we tested whether KRAS mutations could confer chemoresistance to oxaliplatin (L-OHP) in vitro. In addition, we tested the efficacy of the newly developed water-soluble injectable type of curcumin monoglucuronide (CMG) in vivo, that is transformed into an active form of curcumin after the injection and potentially suppresses NF-kB activation. Methods Sensitivity of L-OHP was tested using HCT116/DLD1 colon cancer cell lines with/without KRAS mutations (KRAS+/- cells). Efficacy of CMG was tested using xenograft mouse models. Results IC50 of L-OHP was significantly higher in KRAS+ cells compared to KRAS- cells. Addition of p53 deletion in KRAS+ HCT 116 cells (KRAS+p53- cells) significantly increased chemoresistance to L-OHP compared to KRAS+ cells. Curcumin showed the similar anti-tumor effects regardless of KRAS/p53 mutation status in vitro. Higher NF-kB activity in KRAS+ cells compared to KRAS- cells was detected by using a reporter gene assay and NF-kB activation was suppressed by curcumin in vitro. In xenograft models using KRAS+p53- HCT116 cell line, intraperitoneal administration of CMG demonstrated significantly superior anti-tumor effects compared to L-OHP monotherapy. Notably, no adverse events were noted in the CMG group, whereas bodyweight loss, severe myelosuppression, and liver damage were observed in the L-OHP group. Combination of CMG and L-OHP showed synergic anti-tumor effects. Conclusion These results indicate that CMG could be a novel anti-cancer drug with minimal toxicities for colorectal cancer patients with KRAS mutations.

Exosome-transmitted miR-128-3p increase chemosensitivity of oxaliplatin-resistant colorectal cancer

BackgroundOxaliplatin resistance is a major challenge for treatment of advanced colorectal cancer (CRC). Both acquisition of epithelial-mesenchymal transition (EMT) and suppressed drug accumulation in cancer cells contributes to development of oxaliplatin resistance. Aberrant expression of small noncoding RNA, miR-128-3p, has been shown to be a key regulator in tumorigenesis and cancer development. However, its roles in the progression of CRC and oxaliplatin-resistance are largely unknown.MethodsOxaliplatin-resistant CRC and normal intestinal FHC cells were transfected with a miR-128-3p expression lentivirus. After transfection, FHC-derived exosomes were isolated and co-cultured with CRC cells. miR-128-3p expression in resistant CRC cells, FHC cells, and exosomes was quantified by quantitative real-time PCR (RT-qPCR). The mRNA and protein levels of miR-128-3p target genes in resistant CRC cells were quantified by RT-qPCR and western blot, respectively. The effects of miR-128-3p on CRC cell viability, apoptosis, EMT, motility and drug efflux were evaluated by CCK8, flow cytometry, Transwell and wound healing assays, immunofluorescence, and atomic absorption spectrophotometry. Xenograft models were used to determine whether miR-128-3p loaded exosomes can re-sensitize CRC cells to oxaliplatin in vivo.ResultsIn our established stable oxaliplatin-resistant CRC cell lines, in vitro and vivo studies revealed miR-128-3p suppressed EMT and increased intracellular oxaliplatin accumulation. Importantly, our results indicated that lower miR-128-3p expression was associated with poor oxaliplatin response in advanced human CRC patients. Moreover, data showed that miR-128-3p-transfected FHC cells effectively packaged miR-128-3p into secreted exosomes and mediated miR-128-3p delivery to oxaliplatin-resistant cells, improving oxaliplatin response in CRC cells both in vitro and in vivo. In addition, miR-128-3p overexpression up-regulated E-cadherin levels and inhibited oxaliplatin-induced EMT by suppressing Bmi1 expression in resistant cells. Meanwhile, it also decreased oxaliplatin efflux through suppressed expression of the drug transporter MRP5.ConclusionOur results demonstrate that miR-128-3p delivery via exosomes represents a novel strategy enhancing chemosensitivity in CRC through negative regulation of Bmi1 and MRP5. Moreover, miR-128-3p may be a promising diagnostic and prognostic marker for oxaliplatin-based chemotherapy.

MiR-483-3p regulates oxaliplatin resistance by targeting FAM171B in human colorectal cancer cells

Oxaliplatin resistance limits the efficiency of treatment for colorectal cancer (CRC). Studies have shown that abnormal expression of microRNAs (miRNAs) were associated with tumorigenesis, cancer development and chemoresistance. The purpose of this study was to identify potential miRNAs related to oxaliplatin resistance in CRC cells. In this work, using small RNA sequencing (small RNA-Seq) and transcriptome sequencing (RNA-Seq), we found that down-regulated miR-483-3p was concurrent with up-regulated FAM171B in oxaliplatin-resistant colorectal cancer cell line HCT116/L as compared with its parental cell line HCT116. Transient transfection of miR-483-3p mimics markedly decreased the levels of FAM171B and restored oxaliplatin responsiveness of HCT116/L cells, and this alteration enhanced cell apoptosis and weakened cell migration. Whereas miR-483-3p inhibitor dramatically promoted the expression of FAM171B and enhanced oxaliplatin resistance of HCT116 cells by repressing cell apoptosis. Furthermore, knockdown of FAM171B in HCT116/L cells could also sensitize its reaction of the treatment with oxaliplatin, which was verified by the reduced cell migration. These findings demonstrate that FAM171B is a functional target of miR-483-3p in the regulation of oxaliplatin resistance in human CRC cells.

A Novel Combination Therapy for Human Oxaliplatin-resistant Colorectal Cancer Using Oxaliplatin and Coxsackievirus A11.

Colorectal cancer (CRC) is a major cause of morbidity and mortality throughout the world. It is the third most common cancer worldwide and the fourth most common cause of cancer-related death. FOLFOX, a combination of leucovorin calcium, fluorouracil, and oxaliplatin, is the first-line chemotherapy for stage III and stage IV CRC. However, patients with FOLFOX-resistant CRC have a poor prognosis. In recent years, virochemotherapy has been proposed as a potential treatment for chemotherapy-resistant cancer. Through our first screening assay, we found that coxsackievirus A11 (CVA11) displayed potent oncolytic activities. We tested whether coxsackievirus A11 (CVA11) has oncolytic activity in human CRC cells in vitro and in vivo. We also examined whether pretreatment of oxaliplatin-resistant CRC cells with oxaliplatin enhances the oncolytic activity of CVA11. We found that CVA11 was potently oncolytic against the oxaliplatin-sensitive Caco-2 cell line, but had little effect on the oxaliplatin-resistant line WiDr. However, pretreatment of WiDr cells with oxaliplatin enhanced the oncolytic activity of CVA11, and the combination therapy was more cytotoxic than either oxaliplatin treatment or CVA11 infection alone. Furthermore, growth of subcutaneous WiDr tumors in a xenograft model was significantly lower in mice treated with oxaliplatin followed by intratumoral CVA11 injection compared with mice receiving either treatment alone. Oxaliplatin pretreatment sensitized oxaliplatin-resistant CRC cells to lysis by CVA11 infection in vitro and in vivo. Taken together, these findings identify a novel potential chemovirotherapeutic modality for the treatment of oxaliplatin-resistant human CRC.

Overexpression of MEG3 sensitizes colorectal cancer cells to oxaliplatin through regulation of miR-141/PDCD4 axis

The existence of drug resistance is the main reason for chemotherapeutic failure in malignancies. Long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) is implicated with tumorigenesis and chemoresistance. In the present study, we aimed to identify the role of MEG3 in oxaliplatin-resistant colorectal cancer (CRC) and its potential mechanisms. MEG3 was down-regulated in oxaliplatin-resistant CRC tissues and cell lines. Low MEG3 expression was correlated with poor prognosis of CRC patients. Overexpression of MEG3 improved oxaliplatin sensitivity of HT29/OXA and HCT116/OXA cells. MEG3 suppressed miR-141 expression in HCT116/OXA cells. Moreover, MEG3 elevated PDCD4 expression through targeting miR-141, acting as a competing endogenous RNA (ceRNA). miR-141 inhibition or PDCD4 up-regulation could mimic the functional role in oxaliplatin resistance, which was counteracted by overexpression of MEG3. Collectively, MEG3 facilitated the sensitivity of CRC cells to oxaliplatin by regulating miR-141/PDCD4 axis, providing a novel therapeutic strategy for CRC.

Melatonin Promotes Apoptosis of Oxaliplatin-resistant Colorectal Cancer Cells Through Inhibition of Cellular Prion Protein.

Drug resistance restricts the efficacy of chemotherapy in colorectal cancer. However, the detailed molecular mechanism of drug resistance in colorectal cancer cells remains unclear. The level of cellular prion protein (PrPC) in oxaliplatin-resistant colorectal cancer (SNU-C5/Oxal-R) cells was assessed. PrPC level in SNU-C5/Oxal-R cells was significantly increased compared to that in wild-type (SNU-C5) cells. Superoxide dismutase and catalase activities were higher in SNU-C5/Oxal-R cells than in SNU-C5 cells. Treatment of SNU-C5/Oxal-R cells with oxaliplatin and melatonin reduced PrPC expression, while suppressing antioxidant enzyme activity and increasing superoxide anion generation. In SNU-C5/Oxal-R cells, endoplasmic reticulum stress and apoptosis were significantly increased following co-treatment with oxaliplatin and melatonin compared to treatment with oxaliplatin alone. Co-treatment with oxaliplatin and melatonin increased endoplasmic reticulum stress in and apoptosis of SNU-C5/Oxal-R cells through inhibition of PrPC, suggesting that PrPC could be a key molecule in oxaliplatin resistance of colorectal cancer cells.

MiR-29b reverses oxaliplatin-resistance in colorectal cancer by targeting SIRT1.

Oxaliplatin is a commonly used chemotherapeutic drug for the treatment of advanced colorectal cancer. However, acquired drug resistance against oxaliplatin remains a major obstacle for efficient use of it, and mechanisms underlying oxaliplatin resistance are still required to be explored. In the present study, we exposed colorectal cancer cell line SW480 to oxaliplatin for a long time to obtain oxaliplatin-resistant colorectal cancer cell model (OR-SW480). We found that intracellular expression of miR-29b was decreased when the SW480 cells became oxaliplatin-resistant. More importantly, overexpression of miR-29b resensitized OR-SW480 cells to oxaliplatin treatment. Mechanically, gene of SIRT1 was identified to be the target of miR-29b. Overexpression of miR-29b in oxaliplatin-treated OR-SW480 decreased the expression of SIRT1 to enhance the ROS production and JNK phosphorylation, and thus promoting apoptosis via activation of caspase 9, 7 and 3. On the other hand, expression plasmid of SIRT1, N-acetyl cysteine or SP600125 (JNK specific inhibitor) abolished the effect of miR-29b on oxaliplatin-treated OR-SW480. We therefore demonstrated that miR-29b reverses oxaliplatin-resistance in colorectal cancer by targeting SIRT1/ROS/JNK pathway.

MiR-122 Targets X-Linked Inhibitor of Apoptosis Protein to Sensitize Oxaliplatin-Resistant Colorectal Cancer Cells to Oxaliplatin-Mediated Cytotoxicity

Background/Aims: Although oxaliplatin is one of the most effective chemotherapeutic drugs used to treat colorectal cancer (CRC), long-term administration usually induces acquired drug resistance during the course of treatment. Thus, there is an urgent need to explore novel strategies to improve the efficiency of cancer therapy. The aim of this study was to explore the effect of microRNA-122 (miR-122) on reversing oxaliplatin resistance in CRC. Methods: The expression of miR-122 in CRC cells was examined by quantitative reverse transcriptase real-time PCR. The cytotoxicity of oxaliplatin against CRC cells was evaluated by Cell Counting Kit-8 assays. Mitochondrial membrane potentials and cell apoptotic rates were measured by flow cytometry. Cellular protein expression and interactions were detected by western blot and co-immunoprecipitation. Results: Established oxaliplatin-resistant SW480 and HT29 cells (SW480/OR and HT29/OR) expressed significantly higher levels of X-linked inhibitor of apoptosis protein (XIAP) and lower levels of miR-122 compared with normal SW480 and HT29 cells, respectively. Our results showed that the downregulation of miR-122 was responsible for the overexpression of XIAP in these oxaliplatin-resistant CRC cells. We then found that the recovery of miR-122 expression can sensitize SW480/OR and HT29/OR cells to oxaliplatin-mediated apoptosis through the inhibition of XIAP expression. Conclusion: Upregulation of XIAP in CRC cells is responsible for the acquired resistance to oxaliplatin. Furthermore, miR-122 reversed oxaliplatin resistance in CRC by targeting XIAP.

P52 confers resistance to oxaliplatin in oxaliplatin resistant colon cancer

Objective To assess the role of p52 in resistance to oxaliplation in colon cancer. Methods Colon cancer cell lines SW480, LoVo and resistant cells (SW480/L-OHP, LoVo/L-OHP) were cultured. The inhibiting rates of SW480, LoVo cells and resistant cells by oxaliplatin were determined by cell counting kit-8 (CCK-8) assay. Western blotting was applied to assess induced p52 expression after cultured with oxaliplatin contained medium for 24 h in SW480 cells. We adopted chromatin immunoprecipitation (CHIP) to validate translational regulation of B cell lymphoma/leukemia-2 (bcl-2) by p52. SW480/L-OHP cells were cultured with oxaliplatin contained medium for 24 h after tranfected with siP52, then stained with Annexin V/propidium iodide (PI) in the detection of apoptosis cells by flow cytometry. Total protein was extracted and Western blotting was used to detect the expression of bcl-2, p52 after transfected with small interfering RNA (siRNA) targeting p52. Results Induced p52 expression was seen in SW480 cells. Translational regulation of bcl-2 by p52 was validated by CHIP assays. Down-regulation of bcl-2 was induced by sip52. The proportion of Annexin V/PI positive cells elevated in sip52 silenced SW480/L-OHP cells after treated with oxaliplatin for 24 h. Conclusion p52 confers resistance to oxaliplatin in oxaliplatin resistant colorectal cancer by inhibiting apoptosis. Key words: Colorectal cancer; Oxaliplatin; P52

2-Deoxy-D-glucose suppresses the migration and reverses the drug resistance of colon cancer cells through ADAM expression regulation.

Cancer cell resistance to chemotherapy is associated with a poor prognosis. The compound 2-deoxy-D-glucose (2-DG) enhances the effect of chemotherapy against cancer cells lines in vitro and in vivo. However, its effect on the epithelial to mesenchymal transition (EMT) in drug-resistant cancer cells has not been fully elucidated. In this study, we investigated whether treatment of 5-fluorouracil or oxaliplatin-resistant colorectal cancer (CRC) cells with 2-DG suppressed their migratory activity and enhanced their susceptibility to chemotherapy. Chemoresistant CRC cells stably expressed drug resistance-related proteins (MDR1, MRP1, MRP2, and MRP3) and showed mesenchymal characteristics and a migratory phenotype. 2-DG treatment attenuated glycolysis-related enzyme expression, invasion activity, and EMT-related cytokine secretion in drug-resistant CRC cells. In addition, 2-DG inhibited the activation of a disintegrin and metalloproteinase 10 (ADAM10) and ADAM17. Gene silencing of ADAM10 and ADAM17 with small interfering RNA downregulated mesenchymal properties, reduced EMT-associated cytokine secretion, and rendered chemoresistant cells susceptible to anticancer drug treatment. Collectively, these findings suggest that increased glycolytic metabolism in drug-resistant cells has an effect on both migratory activity and cell viability through the activation of ADAM10 and ADAM17.

MALAT1 Is Associated with Poor Response to Oxaliplatin-Based Chemotherapy in Colorectal Cancer Patients and Promotes Chemoresistance through EZH2.

A major reason for oxaliplatin chemoresistance in colorectal cancer is the acquisition of epithelial-mesenchymal transition (EMT) in cancer cells. The long noncoding RNA (lncRNA), MALAT1, is a highly conserved nuclear ncRNA and a key regulator of metastasis development in several cancers. However, its role in oxaliplatin-induced metastasis and chemoresistance is not well known. In this study, we aim to investigate the prognostic and therapeutic role of lncRNA MALAT1 in colorectal cancer patients receiving oxaliplatin-based therapy and further explore the potential transcriptional regulation through interaction with EZH2 based on the established HT29 oxaliplatin-resistant cells. Our results showed that high MALAT1 expression was associated with reduced patient survival and poor response to oxaliplatin-based chemotherapy in advanced colorectal cancer patients. Oxaliplatin-resistant colorectal cancer cells exhibited high MALAT1 expression and EMT. LncRNA MALAT1 knockdown enhances E-cadherin expression and inhibits oxaliplatin-induced EMT in colorectal cancer cells. EZH2 is highly expressed and associated with the 3' end region of lncRNA MALAT1 in colorectal cancer, and this association suppressed the expression of E-cadherin. Furthermore, targeted inhibition of MALAT1 or EZH2 reversed EMT and chemoresistance induced by oxaliplatin. Finally, the interaction between lncRNA MALAT1 and miR-218 was observed, which further indicated its prognostic value in patients who received standard FOLFOX (oxaliplatin combine with 5-fluorouracil and leucovorin) treatment. In conclusion, this study illuminates the prognostic role of lncRNA MALAT1 in colorectal cancer patients receiving oxaliplatin-based treatment and further demonstrates how lncRNA MALAT1 confers a chemoresistant function in colorectal cancer. Thus, lncRNA MALAT1 may serve as a promising prognostic and therapeutic target for colorectal cancer patients. Mol Cancer Ther; 16(4); 739-51. ©2017 AACR.