Ribonucleotide Reductase Subunit M1 Research Articles

Acquired resistance to gemcitabine and cross-resistance in human pancreatic cancer clones.

The efficacy of gemcitabine (GEM), a standard treatment agent for pancreatic cancer, is insufficient because of primary or acquired resistance to this drug. Patients with tumors intrinsically sensitive to GEM gradually acquire resistance and require a shift to second agents, which are associated with the risk of cross-resistance. However, whether cross-resistance is actually present has long been disputed. Using six GEM-resistant and four highly GEM-resistant clones derived from the pancreatic cancer cell line BxPC-3, we determined the resistance of each clone and parent cell line to GEM and four anticancer agents (5-FU, CDDP, CPT-11, and DTX). The GEM-resistant clones had different resistances to GEM and other agents, and did not develop a specific pattern of cross-resistance. This result shows that tumor cells are heterogeneous. However, all highly GEM-resistant clones presented overexpression of ribonucleotide reductase subunit M1 (RRM1), a target enzyme for metabolized GEM, and showed cross-resistance with 5-FU. The expression level of RRM1 was high; therefore, resistance to GEM was high. We showed that a tumor cell acquired resistance to GEM, and cross-resistance developed in one clone. These results suggest that only cells with certain mechanisms for high-level resistance to GEM survive against selective pressure applied by highly concentrated GEM. RRM1 may be one of the few factors that can induce high resistance to GEM and a suitable therapeutic target for GEM-resistant pancreatic cancer.

Expression of Ribonucleotide Reductase Subunit-2 and Thymidylate Synthase Correlates with Poor Prognosis in Patients with Resected Stages I-III Non-Small Cell Lung Cancer.

Biomarkers can help to identify patients with early-stages or locally advanced non-small cell lung cancer (NSCLC) who have high risk of relapse and poor prognosis. To correlate the expression of seven biomarkers involved in DNA synthesis and repair and in cell division with clinical outcome, we consecutively collected 82 tumour tissues from radically resected NSCLC patients. The following biomarkers were investigated using IHC and qRT-PCR: excision repair cross-complementation group 1 (ERCC1), breast cancer 1 (BRCA1), ribonucleotide reductase subunits M1 and M2 (RRM1 and RRM2), subunit p53R2, thymidylate synthase (TS), and class III beta-tubulin (TUBB3). Gene expression levels were also validated in an available NSCLC microarray dataset. Multivariate analysis identified the protein overexpression of RRM2 and TS as independent prognostic factors of shorter overall survival (OS). Kaplan-Meier analysis showed a trend in shorter OS for patients with RRM2, TS, and ERCC1, BRCA1 overexpressed tumours. For all of the biomarkers except TUBB3, the OS trends relative to the gene expression levels were in agreement with those relative to the protein expression levels. The NSCLC microarray dataset showed RRM2 and TS as biomarkers significantly associated with OS. This study suggests that high expression levels of RRM2 and TS might be negative prognostic factors for resected NSCLC patients.

Pegylated arginine deiminase synergistically increases the cytotoxicity of gemcitabine in human pancreatic cancer.

BackgroundPancreatic ductal adenocarcinoma has proven to be one of the most chemo-resistant among all solid organ malignancies. Several mechanisms of resistance have been described, though few reports of strategies to overcome this chemo-resistance have been successful in restoring sensitivity to the primary chemotherapy (gemcitabine) and enter the clinical treatment arena.MethodsWe examined the ability of cellular arginine depletion through treatment with PEG-ADI to alter in vitro and in vivo cytotoxicity of gemcitabine. The effect on levels of key regulators of gemcitabine efficacy (e.g. RRM2, hENT1, and dCK) were examined.ResultsCombination of PEG-ADI and gemcitabine substantially increases growth arrest, leading to increased tumor response in vivo. PEG-ADI is a strong inhibitor of the gemcitabine-induced overexpression of ribonucleotide reductase subunit M2 (RRM2) levels both in vivo and in vitro, which is associated with gemcitabine resistance. This mechanism is through the abrogation of the gemcitabine-mediated inhibitory effect on E2F-1 function, a transcriptional repressor of RRM2.ConclusionThe ability to alter gemcitabine resistance in a targeted manner by inducing metabolic stress holds great promise in the treatment of advanced pancreatic cancer.

RRM2 (ribonucleotide reductase M2)

Ribonucleotide reductase subunit M2 (RRM2) is located in chromosome-2 p25-p24, converts ribonucleotides to deoxynucleotides which is required for DNA polymerization and repair. It has been shown that RMM2 plays a key role in DNA synthesis, cell growth, and drug resistance of cancer cells. There is accumulating evidence that alteration in the expression level of RRM2 can have a substantial impact on the biological characteristics of cancer cells, including tumor initiation and progression, suggesting its role as a prognostic factor and a possible therapeutic target for cancer therapy. Therefore, this review highlights several recent and clinically relevant aspects of the expression and function of RRM2 in human cancer.

High expression of ribonucleotide reductase subunit M2 correlates with poor prognosis of hepatocellular carcinoma.

Background/AimsRibonucleotide reductase subunit M2 (RRM2) catalyzes the production of deoxynucleotide triphosphates, which are necessary for DNA synthesis. RRM2 has been reported to play an active role in tumor progression, and elevated RRM2 levels have been correlated with poor prognosis for colorectal cancer patients. This study aimed to elucidate the prognostic significance of RRM2 protein expression in hepatocellular carcinoma after surgery.MethodsRRM2 protein expression was evaluated using immunohistochemistry in tumor tissues from 259 hepatocellular carcinoma patients who underwent curative hepatectomy.ResultsHigh RRM2 expression was observed in 210 of 259 patients (81.1%) with hepatocellular carcinomas. High RRM2 expression was significantly associated with viral etiology (p=0.035) and liver cirrhosis (p=0.036). High RRM2 expression was correlated with early recurrence (p=0.004) but not with late recurrence (p=0.144). Logistic regression analysis revealed that high RRM2 expression (p=0.040) and intrahepatic metastasis (p<0.001) were independent predictors of early recurrence. High RRM2 expression unfavorably influenced both shorter recurrence-free survival (p=0.011) and shorter disease-specific survival (p=0.002) and was an independent predictor of shorter disease-specific survival (p=0.008).ConclusionsHigh RRM2 protein expression might be a useful marker for predicting early recurrence and may be a marker for poor prognosis of hepatocellular carcinoma after curative hepatectomy.

Abstract 3763: Ribonucleotide reductase subunit M2 plays important role in cisplatin resistance of cancer cells

Abstract Background: Cisplatin is widely used as a cancer therapeutic for many types of cancers. It mediates the formation of DNA adducts which impair DNA replication and transcription, and consequently induces cell death when there is no prompt DNA repair. However, resistance to cisplatin continues to be the major hurdle to its optimal effect in cancer therapy. Ribonucleotide reductase subunit M2 (RRM2), an essential regulator of balanced deoxyribonucleotides (dNTPs), is critical for DNA replication and repair, and hence cell survival. It has been shown that overexpression of RRM2 is associated with resistance to chemotherapeutic agents including cisplatin. However, the exact mechanism needs to be further deciphered. Methods: We generated cisplatin-resistant H1299-ptR cells from the H1299 (non-small cell lung cancer) cell line for this study, and also employed KB-8-5 and KB-CP 0.5 cells, previously derived from the KB-3-1 (head and neck cancer) cell line. RRM2 expression level was compared between resistant vs. parental cell lines. In addition, the impact on cisplatin resistance was mechanistically pursued by using RRM2 silencing. Results: Based on EC50, we found that KB-8-5 and KB-CP 0.5 cell lines were 4- and 36-fold more resistant to cisplatin compared to their parental cell line KB-3-1, respectively. Similarly, H1299-ptR was 12-fold more resistant than its parental H1299 cell line. Interestingly, in all resistant cell lines, RRM2 expression level was found to be 2- to 4-fold higher than in the parental counterparts. Silencing of RRM2 in resistant cell lines induced cell apoptosis, along with enhanced DNA damage, which manifested as an elevated level of histone γ-H2AX. In KB-CP 0.5 cells, RRM2 knockdown resulted in increased level of p53 (∼60% apoptosis) and reduced activation of ATR-Chk1, indicating that inhibition of ssDNA damage repair system is attributable to cellular apoptosis. Strikingly, the ATR-Chk1 signaling pathway was found to be hyperactive in cisplatin resistant cells. Taken together, these findings suggest that RRM2 plays an active role in the development of cisplatin resistance through its action in the DNA repair system. Conclusions: Increased expression of RRM2 may induce drug-resistance potential in cancer cells, whereas RRM2 suppression reverses drug resistance and decreases proliferation. This study suggests that targeting RRM2 can be an effective approach to overcome drug resistance and potently induce apoptosis. (This work is supported by grants from NIH U01CA151802, P50CA128613). Citation Format: Mohammad Aminur Rahman, A.R.M. R. Amin, Xianghong Peng, Jun Zhang, Zhuo G. Chen, Dong M. Shin. Ribonucleotide reductase subunit M2 plays important role in cisplatin resistance of cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3763. doi:10.1158/1538-7445.AM2014-3763

Abstract 4227: The ribonucleotide reductase inhibitor Didox reverses tamoxifen resistance in breast cancer cells

Abstract Tamoxifen is the most prescribed endocrine therapy for estrogen receptor positive breast cancer patients. The emergence of endocrine resistance is a major hurdle in achieving a successful clinical outcome. Many advances have been made over the past few years to understand the biology of tamoxifen resistance. Activation of the AKT kinase is one of the primary mechanisms of intrinsic tamoxifen resistance. Previously, we reported that the Ribonucleotide Reductase M2 (RRM2) subunit is upregulated in AKT induced tamoxifen resistance and that inhibition of RRM2 by siRNA mediated interference reduces tamoxifen induced cell proliferation. In this study, we report that the inhibition of RRM2 by the small molecule inhibitor of ribonucleotide reductase activity Didox (3, 4-dihydroxybenzohydroxamic acid), significantly reduced tamoxifen induced cell proliferation in AKT overexpressing cells and tamoxifen resistant tumors generated by these cells. As well, Didox in combination with tamoxifen also inhibited cell proliferation in acquired tamoxifen resistant breast cancer cell lines. Employing comprehensive cell culture and in vivo models, we demonstrate that combining tamoxifen with Didox may reverse tamoxifen resistance in AKT expressing breast cancer cells. Citation Format: Khyati N. Shah, Howard L. Elford, Jesika S. Faridi. The ribonucleotide reductase inhibitor Didox reverses tamoxifen resistance in breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4227. doi:10.1158/1538-7445.AM2014-4227

Prognostic significance of BRCA1, ERCC1, RRM1, and RRM2 in patients with advanced non-small cell lung cancer receiving chemotherapy.

The aim of this study was to examine the prognostic value of breast cancer susceptibility gene 1 (BRCA1), excision repair cross-complementation 1 (ERCC1), ribonucleotide reductase subunit M1 (RRM1), and ribonucleotide reductase subunit M2 (RRM2) in patients with advanced non-small cell lung cancer (NSCLC) who received platinum-based chemotherapy. A total of 214 patients with histologically or cytologically confirmed advanced NSCLC were enrolled in this study. The relative complementary DNA (cDNA) quantification for BRCA1, ERCC1, RRM1, and RRM2 was conducted using a fluorescence-based, real-time detection method, and β-actin was used as a reference gene. A strong correlation was observed between ERCC1 and RRM1 messenger RNA (mRNA) levels (P = 0.0385). There were inverse significant correlations between BRCA1 and ERCC1 (P < 0.0001) or RRM1 (P < 0.0001) mRNA levels. As BRCA1 levels increased, the probability of response increased (odds ratio [OR] = 0.49; P < 0.001) and the risk of progression (hazard ratio [HR] = 1.52; P = 0.034) and death (HR = 1.45; P = 0.024) decreased. As ERCC1, RRM1, and RRM2 levels increased, the probability of response decreased (ERCC1, OR = 0.42, P < 0.001; RRM1, OR = 0.85, P = 0.083; RRM2, OR = 0.42, P = 0.005) and the risk of progression (ERCC1, HR = 1.59, P = 0.002; RRM1, HR = 1.48, P = 0.039; RRM2, HR = 1.49, P = 0.041) and death (ERCC1, HR = 1.62, P = 0.008; RRM1, HR = 1.52, P = 0.023; RRM2, HR = 1.48, P = 0.017) increased. At multivariate analysis, low expression of ERCC1 was shown to be an independent predictive factor for response to chemotherapy (P = 0.018), time to progression (P = 0.025), and overall survival (P = 0.038). Furthermore, concomitant low expression levels of ERCC1, RRM1, and RRM2 and the high expression level of BRCA1 were predictive of a better outcome (P = 0.014). This study suggests that the efficacy of platinum-based chemotherapy can be improved when customized according to the mRNA expression of BRCA1, ERCC1, RRM1, and RRM2.

Expression of ERCC1, RRM1 and LRP in non-small cell lung cancers and their influence on chemotherapeutic efficacy of gemcitabine concomitant with nedaplatin.

To explore the clinical efficacy of gemcitabine concomitant with nedaplatin and drug resistance in the treatment of non-small cell lung cancer (NSCLC) and associated molecular predicators. A total of 68 patients diagnosed with NSCLC by histology served as the study objects and were randomly divided into an observation group treated with gemcitabine concomitant with nedaplatin and a control group with cisplatin concomitant with gemcitabine, 34 cases for each group. Short-term and long-term efficacies, adverse responses as well as the expression of nucleotide excision repair cross complementing 1 (ERCC1), ribonucleotide reductase subunit M1 (RRM1) and lung resistance-related protein (LRP) in NSCLC tissues in both groups were assessed. The short-term objective response rate (ORR) and disease control rate (DCR) were 35.3% (12/34) and 76.5% (26/34) in the observation group and 38.2% (13/34) and 85.3% (29/34) in the control group, respectively, the differences not being statistically significant. The time to progression (TTP) in both groups were 1~12 months, while the median TTP was 135 d and 144 d, respectively. Though the survival was slightly higher in the control group, there were no significant differences in TTP and survival time. The rates of decreased hemoglobin, vomiting and nausea as well as renal toxicity were evidently lower in the observation group, while other adverse responses demonstrated no significant difference. The positive expression rates of ERCC1, RRM1 and LRP were 47.1% (16/34), 61.8% (21/34) and 64.7% (22/34) in the observation group, respectively. Compared with negative ERCC1 expression, ORR had decreasing trend and the overall survival time (OS) decreased significantly in patients with positive ERCC1 expression, which were markedly decreased by the positive expressions of RRM1 and LRP. Gemcitabine concomitant with nedaplatin has significant effects in the treatment of NSCLC, with an adverse response rate obviously lower than for cisplatin concomitant with gemcitabine, suggesting that wider use in the clinic is warranted. Additionally, the positive expressions of ERCC1, RRM1 and LRP may increase patient drug resistance, so they can be applied as the chemotherapeutic predicators to guide individualized therapy of NSCLC patients.

Prognostic and therapeutic significance of ribonucleotide reductase small subunit M2 in estrogen-negative breast cancers.

BackgroundRibonucleotide reductase (RR) is an essential enzyme involved in DNA synthesis. We hypothesized that RR subunit M2 (RRM2) might be a novel prognostic and predictive biomarker for estrogen receptor (ER)-negative breast cancers.MethodsIndividual and pooled survival analyses were conducted on six independent large-scale breast cancer microarray data sets; and findings were validated on a human breast tissue set (ZJU set).ResultsGene set enrichment analysis revealed that RRM2-high breast cancers were significantly enriched for expression of gene sets that increased in proliferation, invasiveness, undifferentiation, embryonic stem/progenitor-like phenotypes, and poor patient survival (p < 0.01). Independent and pooled analyses verified that increased RRM2 mRNA levels were associated with poor patient outcome in a dose-dependent manner. The prognostic power of RRM2 mRNA was comparable to multiple gene signatures, and it was superior to TNM stage. In ER-negative breast cancers, RRM2 showed more prognostic power than that in ER-positive breast cancers. Further analysis indicated that RRM2 was a more accurate prognostic biomarker for ER-negative breast cancers than the pathoclinical indicators and uPA. A new RR inhibitor, COH29, could significantly enhance the chemosensitivity to doxorubicin in ER-negative MDA-MB-231 cells, but not in ER-positive MCF-7 cells.ConclusionRRM2 appears to be a promising prognostic biomarker and therapeutic target for ER-negative breast cancer patients.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-664) contains supplementary material, which is available to authorized users.

A Randomized Phase III Multicenter Trial of Customized Chemotherapy Versus Standard of Care for 1St Line Treatment of Elderly Patients with Advanced Non-Small-Cell Lung Cancer (Nsclc): the Elderly Patient Individualized Chemotherapy (Epic) Trial

ABSTRACT Background: Phase III trial aiming to compare first line pharmacogenomic-driven chemotherapy based on Excision-Repair-Cross-Complementing-1 (ERCC1, E), Ribonucleotide Reductase subunit M1 (RRM1, R) and Thymidylate Synthase (TS, T) gene expression versus standard treatment in elderly patients (pts) with advanced NSCLC. Trial design: pts aged ≥70 years, ECOG Performance Status 0 or 1, EGFR negative mutational status, chemonaive stage IV NSCLC will be evaluated. Pts will be randomized (2:1) to experimental arm (A) or standard arm (B). In arm A, treatment will be based on histology, E, R and T mRNA expression. The cut-off for high or low expression have been previously defined. Treatments for pts with squamous NSCLC: carboplatin for E low/R high, gemcitabine for E high/R low, carboplatin and gemcitabine for E low/R low, docetaxel or vinorelbine for E high/R high. Treatments for non-squamous NSCLC pts: carboplatin for E low/T high, pemetrexed for E high/T low, carboplatin and pemetrexed for E low/T low, gemcitabine for E high/T high/R low, docetaxel or vinorelbine for E high/T high/R high. In arm B treatment will be at the investigator discretion. The primary endpoint is overall survival. The secondary endpoints are progression-free survival, response rate (RECIST 1.1) and tolerability (CTCAE version 4.0). Feasibility of treatment selection based on pharmacogenomic parameters will be also assessed. Treatment will continue to a maximum of 6 cycles if tolerated or until disease progression. Assuming an exponential survival distribution for both arms and a median survival time of 8 months in the control arm, we anticipate to detect an improvement of 3 months in the median survival time in the experimental arm. To have 90% power to detect a three-month improvement in median survival at a significance level of 5% (2-sided) and assuming a 10% failure rate in gene analyses or loss to follow-up rate, a sample size of 567 patients is planned. Conclusion: To our knowledge, this is the first prospective pharmacogenomic-driven trial designed in elderly advanced NSCLC population. Disclosure: D. Cortinovis: Consultant for AZ, BI, Roche and Lilly; M. Papotti: Honoraria have been received from Eli Lilly; G. Scagliotti: Honoraria have been received from Eli Lilly, Roche, Pfizer and Astrazeneca. All other authors have declared no conflicts of interest.

Mutations of EGFR or KRAS and expression of chemotherapy-related genes based on small biopsy samples in stage IIIB and IV inoperable non-small cell lung cancer.

Epidermal growth factor receptor (EGFR) and KRAS mutations may predict the outcome of targeted drug therapy and also may be associated with the efficacy of chemotherapy in patients with non-small cell lung cancer (NSCLC). This report investigated the relation of EGFR or KRAS mutation and expression of chemotherapy-related genes, including excision repair cross-complementing 1 (ERCC1), thymidylate synthetase (TYMS), ribonucleotide reductase subunit M1 (RRM1) and class III β-tubulin (TUBB3), as a potential explanation for these observations. A total of 143 patients with stage IIIB and IV NSCLC from bronchoscopy or percutaneous lung biopsy obtained tumor samples were analyzed concurrently for EGFR or KRAS mutations, and mRNA expression of ERCC1, TYMS, RRM1 and TUBB3. EGFR or KRAS mutations were detected with xTAG liquidchip technology (xTAG-LCT), and mRNA expression levels of four genes were detected by branched DNA-liquidchip technology (bDNA-LCT). Of 143 patients, 63 tumors were positive for EGFR-activating mutations, and 16 tumors were positive for KRAS mutations. EGFR-activating mutations are more frequent in females, adenocarcinoma and non-smokers patients, and KRAS mutations are more frequent in smoking patients. ERCC1 mRNA levels were significantly associated with histological type and tumor differentiation, whereas TYMS levels were significantly associated with age. NSCLC specimens that harboring EGFR-activating mutations are more likely to express low ERCC1 and high TUBB3 mRNA levels, whereas tumors from patients with NSCLC harboring KRAS mutation are more likely to express high ERCC1 mRNA levels. Mutations and expression of chemotherapy-related genes may provide a basis for the selection of suitable molecular markers for individual treatment in a population with stage IIIB and IV NSCLC.

Relationship between the efficacy of gemcitabine/cisplatin adjuvant chemotherapy and RRM1 protein expression in postoperative NSCLC patients

The purpose of this study was to investigate the relationship between the expression of ribonucleotide reductase subunit M1 (RRM1) protein and the efficacy of gemcitabine/cisplatin (GP) adjuvant chemotherapy in postoperative non-small cell lung cancer (NSCLC) patients. A total of 68 patients with NSCLC after radical surgery were included in this study. The expression of RRM1 protein in tumor specimens was assayed by streptavidin-peroxidase (SP) immunohistochemistry retrospectively. Correlation between the expression of RRM1 protein and the efficacy of GP chemotherapy was analyzed. Disease-free survival rate was taken as the main outcome measure. Among the 68 patients, 31 cases had recurrence or metastasis. The expression rate of RRM1 was 54.4%. The 1-year and 3-year disease-free survival rates were 82.7% and 61.5% for patients with RRM1-negative expression, and 78.1% and 36.8% for patients with RRM1-posivive expression, respectively (P = 0.044). In the subgroup analysis of stage IB cases, the 1-year and 3-year disease-free survival rates were 100% and 82.3% for patients with RRM1-negative expression, and 84.5% and 24.6% for patients with RRM1-positive expression, respectively (P = 0.047). In the analysis of squamous cell carcinoma subgroup, the 1-year and 3-year disease-free survival rates were 92.3% and 83.7% for patients with RRM1-negative expression, and 83.1% and 43.9% for patients with RRM1-posivive expression, respectively (P = 0.005). Univariate analysis and multivariate analysis indicated that smoking history, pathological type, clinical stage and expression of RRM1 significantly influenced the therapeutic efficacy (P < 0.05). RRM1 protein may be a valuable predictive factor for gemcitabine/cisplatin adjuvant chemotherapy in NSCLC patients.

Multiplex real-time PCR for RRM1, XRCC1, TUBB3 and TS mRNA for prediction of response of non-small cell lung cancer to chemoradiotherapy.

This study was aimed to establish a novel method to simultaneously detect expression of four genes, ribonucleotide reductase subunit M1(RRM1), X-ray repair cross-complementing gene 1 (XRCC1), thymidylate synthase (TS) and class III β-tubulin (TUBB3), and to assess their application in the clinic for prediction of response of non-small cell lung cancer (NSCLC) to chemoradiotherapy. We have designed four gene molecular beacon (MB) probes for multiplex quantitative real-time polymerase chain reactions to examine RRM1, XRCC1, TUBB3 and TS mRNA expression in paraffin-embedded specimens from 50 patients with advanced or metastatic carcinomas. Twenty one NSCLC patients receiving cisplatin- based first-line treatment were analyzed. These molecular beacon probes could specially bind to their target genes in homogeneous solutions. Patients with low RRM1 and XRCC1 mRNA levels were found to have apparently higher response rates to chemoradiotherapy compared with those with high levels of RRM1 and XRCC1 expression (p<0.05). The TS gene expression level was not significantly associated with chemotherapy response (p>0.05). A method of simultaneously detecting four molecular markers was successfully established and applied for evaluation of chemoradiotherapy response. It may be a useful tool in personalized cancer therapy.

Pathway-Centric Integrative Analysis Identifies RRM2 as a Prognostic Marker in Breast Cancer Associated with Poor Survival and Tamoxifen Resistance

Breast cancer (BCa) molecular subtypes include luminal A, luminal B, normal-like, HER-2–enriched, and basal-like tumors, among which luminal B and basal-like cancers are highly aggressive. Biochemical pathways associated with patient survival or treatment response in these more aggressive subtypes are not well understood. With the limited availability of pathologically verified clinical specimens, cell line models are routinely used for pathway-centric studies. We measured the metabolome of luminal and basal-like BCa cell lines using mass spectrometry, linked metabolites to biochemical pathways using Gene Set Analysis, and developed a novel rank-based method to select pathways on the basis of their enrichment in patient-derived omics data sets and prognostic relevance. Key mediators of the pathway were then characterized for their role in disease progression. Pyrimidine metabolism was altered in luminal versus basal BCa, whereas the combined expression of its associated genes or expression of one key gene, ribonucleotide reductase subunit M2 (RRM2) alone, associated significantly with decreased survival across all BCa subtypes, as well as in luminal patients resistant to tamoxifen. Increased RRM2 expression in tamoxifen-resistant patients was verified using tissue microarrays, whereas the metabolic products of RRM2 were higher in tamoxifen-resistant cells and in xenograft tumors. Both genetic and pharmacological inhibition of this key enzyme in tamoxifen-resistant cells significantly decreased proliferation, reduced expression of cell cycle genes, and sensitized the cells to tamoxifen treatment. Our study suggests for evaluating RRM2-associated metabolites as noninvasive markers for tamoxifen resistance and its pharmacological inhibition as a novel approach to overcome tamoxifen resistance in BCa.

Targeting ribonucleotide reductase M2 subunit by small interfering RNA exerts anti-oncogenic effects in gastric adenocarcinoma.

Ribonucleotide reductase M2 subunit(RRM2) is one of the two subunits of human ribonucleotide reductase which plays a critical role in tumor progression. The aim of the present study was to analyze its expression, clinical significance and biological functions in gastric adenocarcinoma. We observed the upregulation of RRM2 mRNA and protein in all nine gastric cancer cell lines examined. In paired primary gastric cancers, both mRNA and protein levels of RRM2 were significantly upregulated in tumors compared with the corresponding non-tumorous gastric tissues. RRM2 protein expression correlated with higher tumor grade, advanced T stage and poor disease-specific survival. RRM2 knockdown in gastric cancer cell lines AGS, MKN1 and MKN28 significantly suppressed cell proliferation, inhibited monolayer colony formation, reduced cell invasion and induced apoptosis. Downregulation of RRM2 suppressed xenograft formation invivo. Collectively, these findings suggest that RRM2 plays a crucial role in gastric tumorigenesis and may serve as a potential prognostic marker and therapeutic target in gastric cancer.

Pegylated arginine deiminase synergistically increases the cytotoxicity of gemcitabine in human pancreatic cancer

Pancreatic ductal adenocarcinoma has proven to be one of the most chemo-resistant among all solid organ malignancies. Several mechanisms of resistance have been described, though few reports of strategies to overcome this chemo-resistance have been successful in restoring sensitivity to the primary chemotherapy (gemcitabine) and enter the clinical treatment arena. We examined the ability of cellular arginine depletion through treatment with PEG-ADI to alter in vitro and in vivo cytotoxicity of gemcitabine. The effect on levels of key regulators of gemcitabine efficacy (e.g. RRM2, hENT1, and dCK) were examined. Combination of PEG-ADI and gemcitabine substantially increases growth arrest, leading to increased tumor response in vivo. PEG-ADI is a strong inhibitor of the gemcitabine-induced overexpression of ribonucleotide reductase subunit M2 (RRM2) levels both in vivo and in vitro, which is associated with gemcitabine resistance. This mechanism is through the abrogation of the gemcitabine-mediated inhibitory effect on E2F-1 function, a transcriptional repressor of RRM2. The ability to alter gemcitabine resistance in a targeted manner by inducing metabolic stress holds great promise in the treatment of advanced pancreatic cancer.

ERCC1 mRNA expression is associated with the clinical outcome of non-small cell lung cancer treated with platinum-based chemotherapy.

We conducted a prospective study to analyze the expression of the excision repair cross-complementing group 1 (ERCC1) and ribonucleotide reductase subunit M1 (RRM1) genes in 297 Chinese patients with advanced non-small cell lung cancer (NSCLC). The goal of this study was to evaluate these genes as potential biomarkers for prediction of tumor response and clinical outcome. Patients with unresectable, locally advanced or metastatic NSCLC were enrolled between September 2007 and September 2009, and they were followed up until September 2012. A fluorescence-based real-time detection method was used to quantify relative levels of ERCC1 and RRM1 cDNA. Relative amounts of ERCC1 and RRM1 cDNA were calculated by comparing to actin. By the end of follow-up, 132 patients had died and 165 patients experienced progression. The median overall survival time was 18.7 months (range, 1-60 months). The median levels of ERCC1 and RRM1 were 2.46 x 10(-2) and 0.97 x 10(-2), respectively. Patients with low ERCC1 expression had a significantly higher rate of complete response to chemotherapy, with an OR (95%CI) of 1.56 (1.03-2.47). Moreover, individuals with low levels of ERCC1 had longer overall survival than patients with high expression, with an adjusted hazard ratio (95%CI) of 0.57 (0.35-0.93). In summary, low ERCC1 mRNA expression was associated with better response to chemotherapy and correlated with longer survival in advanced NSCLC patients treated with platinum-based chemotherapy.

The tailored chemotherapy based on RRM1 immunohistochemical expression in patients with advanced non-small cell lung cancer

Ribonucleotide reductase subunit M1 (RRM1) has emerged as a promising biomarker to predict the efficacy of gemcitabine. The purpose of the study was to evaluate whether the tailored chemotherapy based on RRM1 immunohistochemical (IHC) expression had any benefit for patients with advanced non-small cell lung cancer (NSCLC). A single-institution study was conducted in patients with advanced NSCLC. In personalized therapy group, patients received chemotherapy based on RRM-1 IHC expression levels. Low RRM1 group received gemcitabine or gemcitabine/cisplatin, high RRM1 group received docetaxel or docetaxel/ cisplatin. In standard therapy group, non-customized chemotherapy was delivered. In this trial, Patients aged ⩾ 70 years received single agent chemotherapy, whereas patients below 70 had platinum-based chemotherapy. There were statistically significant improvements between the personalized therapy group versus the standard therapy group in disease control rate (82.9% vs 55.3%, P=0.004), and PFS (median: 5.5 months vs 3.0 months, P=0.005). Besides, the OS had a tendency to become more prolonged (median: 16.0 months vs 12.4 months, P=0.286). The subgroup analysis suggested the survival benefit in the elderly patients was more obvious. RRM1 IHC expression tailored selection of first-line therapy could improve therapeutic outcomes in patients with advanced NSCLC.

A phase II trial of sequential ribonucleotide reductase inhibition in aggressive myeloproliferative neoplasms

Myeloproliferative neoplasms are a varied group of disorders that can have prolonged chronic phases, but eventually accelerate and can transform into a secondary acute myeloid leukemia that is ultimately fatal. Triapine is a novel inhibitor of the M2 subunit of ribonucleotide reductase. Sequential inhibition of ribonucleotide reductase with triapine and an M1 ribonucleotide reductase inhibitor (fludarabine) was noted to be safe, and led to a 29% complete plus partial response rate in myeloproliferative neoplasms. This article reports the findings of a phase II trial of triapine (105 mg/m(2)/day) followed by fludarabine (30 mg/m(2)/day) daily for 5 consecutive days in 37 patients with accelerated myeloproliferative neoplasms and secondary acute myeloid leukemia. The overall response rate was 49% (18/37), with a complete remission rate of 24% (9/37). Overall response rates and complete remissions were seen in all disease subsets, including secondary acute myeloid leukemia, in which the overall response rate and complete remission rate were 48% and 33%, respectively. All patients with known JAK2 V617F mutations (6/6) responded. The median overall survival of the entire cohort was 6.9 months, with a median overall survival of both overall responders and complete responders of 10.6 months. These data further demonstrate the promise of sequential inhibition of ribonucleotide reductase in patients with accelerated myeloproliferative neoplasms and secondary acute myeloid leukemia. This study was registered with clinicaltrials.gov (NCT00381550).