Ribonucleotide Reductase Subunit M1 Expression Research Articles

Double-staining Immunohistochemistry Reveals in Malignant Pleural Mesothelioma the Coexpression of ERCC1 and RRM1 as a Frequent Biological Event Related to Poorer Survival.

Malignant pleural mesothelioma (MPM) is a rare cancer with a poor prognosis. To date, standard MPM therapy is still limited to surgery, radiotherapy, and chemotherapy, including pemetrexed and platinum compounds. The main mechanisms of platinum resistance are associated with DNA repair pathways. Excision repair cross-complementing group 1 (ERCC1) and ribonucleotide reductase subunit M1 (RRM1) are important components of the DNA repair, considered as prognostic and predictive biomarkers in various cancer types. The main goal of the present study was to investigate the ERCC1 and RRM1 expression and their potential impact on outcome in this tumor. A series of 73 MPM, mainly treated with a platin-based regimen, was collected and the immunohistochemistry tests were performed to assess ERCC1 and RRM1 expression. In addition, a multiplex immunohistochemistry has been validated to detect simultaneously the 2 proteins on the same slide. In our series, 36 of 73 cases showed ERCC1 expression and 55 of 73 showed RRM1 expression. The double immunohistochemical staining showed the coexpression of ERCC1/RRM1 in 34 of 73 cases. A significant association between ERCC1 and RRM1 expression was observed in our series (P<0.05). Patients with ERCC1/RRM1 coexpression experienced shorter median overall survival (6.6 vs. 13.8 mo, log-rank=7688; P=0.006). Our results suggest that the coexpression of ERCC1/RRM1 could define a group of MPM patients with the worst prognosis who should need likely alternative treatment. In conclusion, we propose the putative usefulness of ERCC1/RRM1 coexpression as prognostic biomarkers for overall survival in MPM.

RRM1 predicts clinical outcome of high-and\xa0intermediate-risk non-muscle-invasive bladder cancer patients treated with intravesical gemcitabine monotherapy

BackgroundThe expression level of ribonucleotide reductase subunit M1 (RRM1) is closely related to the effect of gemcitabine-based therapy in advanced bladder cancer. However, the value of RRM1 expression in predicting progression-free survival in non-muscle-invasive bladder cancer (NMIBC) patients treated with intravesical gemcitabine chemotherapy has not been elucidated.MethodsThis study randomly assigned 162 patients to either the RRM1-known group or the unknown group. We collected cancer tissues from 81 patients to evaluate the mRNA expression of RRM1 by using liquid chip technology. All patients were diagnosed and then treated with intravesical gemcitabine monotherapy immediately after transurethral resection of the bladder tumour (TURBT).ResultsRRM1 expression was high in 21% (17/81) of patients. The RRM1 mRNA level was not correlated with sex, age, weight, performance status, or CUA/EAU risk (p > 0.05). Progression-free survival (PFS) was significantly longer for patients with low RRM1 expression than for patients with high and unknown RRM1 expression (p = 0.009). Additionally, the 1- and 2-year relapse rates also differed according to RRM1 expression level. The 1-year relapse rates for RRM1-low, RRM1-high and RRM1-unknown patients were 0, 17.7 and 6.2% (p = 0.009), while the 2-year relapse rates for these groups were 3.1, 29.4, and 11.1% (p = 0.005), respectively.ConclusionsThis preliminary study showed that low RRM1 expression was associated with longer progression-free survival and lower 1-year/2-year relapse rates in NMIBC patients treated with intravesical gemcitabine monotherapy, despite the need for further verification with large sample sizes and considering more mixed factors and biases.

Comparison of the expression of human equilibrative nucleotide transporter 1 (hENT1) and ribonucleotide reductase subunit M1 (RRM1) genes in seven non-Hodgkin lymphoma cell lines.

We investigated the variability in the expression of human equilibrative nucleoside transporter 1 (hENT1) and ribonucleotide reductase subunit M1 (RRM1) in non-Hodgkin lymphoma cell lines. hENT1 and RRM1 mRNA expression levels in natural killer (NK) cells and seven non-Hodgkin lymphoma cell lines (YTS, SNK-6, Jeko-1, ly-1, Raji, Karpas, and Jurket) were studied using reverse-transcription quantitative real-time polymerase chain reaction (RT-qPCR) and the results were compared using the Student t-test. mRNA expression of hENT1 was detectable in YTS, SNK-6, Jeko-1, ly-1, Raji, Karpas, Jurket, and NK cells, which revealed variability in gene expression. There were significant differences in the mRNA expression values of hENT1 (P = 0.021) and RRM1 (P = 0.002) compared to those in NK cells. mRNA expression of both hENT1 and RRM1 was closely associated with non-Hodgkin lymphoma cell proliferation. Differential expression analysis of hENT1 and RRM1 in non-Hodgkin lymphoma cell lines may provide novel drug leads for precision medicine.

Prognostic significance of the mRNA expression of ERCC1, RRM1, TUBB3 and TYMS genes in patients with non-small cell lung cancer.

The present study aimed to investigate the prognostic value of excision repair cross-complementing 1 (ERCC1), ribonucleotide reductase subunit M1 (RRM1), class III β-tubulin (TUBB3) and thymidylate synthase (TYMS) in patients with non-small cell lung cancer (NSCLC) receiving platinum-based adjuvant chemotherapy. The mRNA expression of these genes was assessed in 72 tumor tissue samples obtained following surgery, using multiplex branched-DNA technology. Subsequent to surgery, all 72 patients with NSCLC were treated with platinum-based chemotherapy. The expression of these five genes was analyzed and the correlation with clinical characteristics and patient survival was investigated. Among the 72 samples, the incidence rate of mRNA expression of ERCC1 was 38.9% (28/72), RRM1 was 55.6% (40/72), TUBB3 was 47.2% (34/72) and TYMS was 62.5% (45/72). The incidence rate of ERCC1 expression in adenocarcinoma (34.2%) was significantly lower than that in non-adenocarcinoma (44.1%; P<0.05). Furthermore, the incidence rates of TYMS and TUBB3 expression in the high-median differentiation tissue samples were significantly lower than those in the low differentiation tissue samples (P<0.05). When the correlation of gene expression and patient survival was analyzed, high expression of ERCC1, RRM1, TUBB3 or TYMS was found to be associated with poor prognosis (P<0.001, P=0.001, P=0.001 and P=0.001, respectively). ERCC1, RRM1, TUBB3 and TYMS are key factors involved in survival following surgical treatment in patients with NSCLC. The mRNA expression of these genes may have prognostic value for patients with NSCLC treated with platinum-based chemotherapy.

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.

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.

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.

Abstract 3545: ATM, TS and RRM1 protein expression in nasopharyngeal carcinomas (NPC) treated with curative intent.

Abstract Background: Although NPC is a radiation and chemotherapy sensitive tumour, some patients still relapse with distant metastatic disease. Biomarkers are needed to help identify which patients are at higher risk of relapse and/or who will respond to therapy. Ataxia telangiectasia mutated (ATM) is a putative marker for radiation sensitivity while, thymidylate synthetase (TS), and ribonucleotide reductase subunit M1 (RRM1) levels have been associated with response to 5FU and gemcitabine, two chemotherapy agents often used in NPC. We characterized the protein expression of ATM, TS, and RRM1 in tumours from 146 patients with non-metastatic NPC treated at two Canadian institutions between Jan 2000-Dec 2007 and explored the relationship of each marker with clinical outcomes. Methods: Pre-treatment, formalin-fixed, paraffin-embedded NPC tumour specimens were assembled in a tissue microarray. ATM, TS and RRM1 protein expression were evaluated by quantitative fluorescence immunohistochemistry and with automated quantitative digital image analysis (AQUA) using the Y170, TS106 and 60073-2-Ig monoclonal antibodies respectively. Protein expression levels were assessed in tumour, tumour cytoplasm, tumour nuclear, and non-malignant tumour stroma compartments. The effect of protein expression levels on overall survival (OS) and disease-free survival (DFS) was examined using Cox regression. For all biomarkers, AQUA scores were analyzed in quartiles. Results: Patient characteristics were: mean age=52 years (SD=12.2; range 18 to 85), 67% male, 73% KPS ≥ 90%, WHO type 1/2/3=11%/28%/61%, stage III/IV=65%. Fifty-eight (40%) patients received platinum-based CRT; the remainder was treated with RT alone. With a median follow-up of 50 months (range 3 to 120), the 5 year survival rates were 71% (95% CI=62%-78%) for OS and 48% (95% CI=39%-57%) for DFS. After adjusting for KPS, stage, and WHO type, OS was worse for the group with tumour:stromal (T/S) ATM ratios in the top quartile versus those in the lowest quartile (p=0.049). OS was also worse for tumours with TS levels in the lowest quartile versus the top quartile (p=0.033). RRM1 was not associated with outcomes (p=0.748). There was no significant effect of any of the biomarkers on disease-free survival (ATM T/S p=0.708; TS p=0.978; RRM1 p=0.918). Conclusions: In our cohort of non-metastatic NPC patients, relative overexpression of ATM and low TS protein levels were associated with worse outcomes suggesting further investigation of these potential biomarkers is warranted. Citation Format: Alexander C. Klimowicz, Amanda Jagdis, Tien Phan, Janessa J. Laskin, Harold Y. Lau, Jodi E. Siever, Stephanie K. Petrillo, Thomas A. Thomson, M. Sarah Rose, Anthony M. Magliocco, Desirée Hao. ATM, TS and RRM1 protein expression in nasopharyngeal carcinomas (NPC) treated with curative intent. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3545. doi:10.1158/1538-7445.AM2013-3545

The effect of the acid-sensitivity of 4-(N)-stearoyl gemcitabine-loaded micelles on drug resistance caused by RRM1 overexpression

Chemoresistance is a major issue for most gemcitabine-related chemotherapies. The overexpression of ribonucleotide reductase subunit M1 (RRM1) plays a key role in gemcitabine resistance. In this study, we synthesized a new highly acid-sensitive amphiphilic micelle material by conjugating hydrophilic polyethylene glycol with a hydrophobic stearic acid derivative (C18) using a hydrazone bond, which was named as PHC-2. A lipophilic prodrug of gemcitabine, 4-(N)-stearoyl gemcitabine (GemC18), was loaded into micelles prepared with PHC-2, a previously synthesized less acid-sensitive PHC-1, and their acid-insensitive counterpart, PAC. GemC18 loaded in acid-sensitive micelles can overcome gemcitabine resistance, and GemC18 in the highly acid-sensitive PHC-2 micelles was more cytotoxic than in the less acid-sensitive PHC-1 micelles. Mechanistic studies revealed that upon cellular uptake and lysosomal delivery, GemC18 in the acid-sensitive micelles was released and hydrolyzed more efficiently. Furthermore, GemC18 loaded in the highly acid-sensitive PHC-2 micelles inhibited the expression of RRM1 and increased the level of gemcitabine triphosphate (dFdCTP) in gemcitabine resistant tumor cells. The strategy of delivering lipophilized nucleoside analogs using highly acid-sensitive micelles may represent a new platform technology to increase the antitumor activity of nucleoside analogs and to overcome tumor cell resistance to them.

Excision repair cross‐complementing gene‐1, ribonucleotide reductase subunit M1, ribonucleotide reductase subunit M2, and human equilibrative nucleoside transporter‐1 expression and prognostic value in biliary tract malignancy

Tumor expression of excision cross-complementing gene-1 (ERCC1), human equilibrative nucleoside transporter 1 (hENT1), ribonucleotide reductase subunit M1 (RRM1), and ribonucleotide reductase subunit M2 (RRM2), is associated with the efficacy of platinum and gemcitabine chemotherapy. The authors of this report recently demonstrated that high ERCC1 and RRM2 expression levels are independent negative prognostic markers for survival in early stage pancreas cancer. The differential expression and prognostic value of these biomarkers in biliary tract malignancy (BTM) is unknown. In total, 63 patients who had tissue available for analysis were selected from a prospective database of all patients (n = 104) who underwent resection of BTM (intrahepatic, hilar, or distal cholangiocarcinoma; gallbladder carcinoma) between January 2000 and December 2008. Immunohistochemistry for ERCC1, hENT1, RRM1, and RRM2 expression was performed. Staining was scored by a single pathologist who was blinded to patient outcomes. The median patient age was 67 years. The median overall survival (OS) was 16.2 months, and the median follow-up was 32.7 months. Only 3 BTMs (4.8%) had high ERCC1 expression, and 92.1% and 81% of BTMs exhibited high hENT1 and RRM1 expression, respectively. RRM2 expression varied, and 32% of tumors demonstrated high RRM2 expression. ERCC1 and RRM1 were not associated with OS. High RRM2 expression was associated with a trend toward improved OS (30.8 months vs 16.2 months; P = .06), and high hENT1 expression was associated with improved OS (17.7 months vs 9.5 months; P = .04). Most BTMs exhibited low ERCC1 expression and high hENT1 and RRM1 expression, whereas RRM2 expression levels varied. High expression of hENT1 was associated with improved OS. These findings may have implications for the selection of chemotherapy agents (gemcitabine vs platinum) and the stratification of patients in chemotherapy trials that assess outcome.

Expression of RRM1 and its association with resistancy to gemcitabine-based chemotherapy in advanced nasopharyngeal carcinoma

Gemcitabine has high activity against nasopharyngeal carcinoma (NPC). The level of ribonucleotide reductase subunit M1 (RRM1) expression is closely related to the efficacy of gemcitabine on non-small cell lung cancer and pancreatic cancer. However, the expression of RRM1 and its association with sensitivity to gemcitabine-based chemotherapy in advanced NPC is not known. In this study, we retrospectively collected 48 formalin-fixed, paraffin-embedded NPC tissues to evaluate the expression of RRM1 using immunohistochemistry. All patients were diagnosed and treated with gemcitabine-based chemotherapy at Sun Yat-sen University Cancer Center. RRM1 expression was positive in 17 (35%) patients. RRM1 expression was not associated with sex, age, performance status, WHO histological type, number of distant metastases, previous treatment, or cycles of gemcitabine-based chemotherapy (P > 0.05). The progression-free survival of the RRM1-positive group was shorter than that of the RRM1-negative group (5 months vs. 7 months, P = 0.036), and the response rate of the RRM1-positive group was somewhat lower than that of the RRM1-negative group (51.6% vs. 35.3%, P = 0.278). There was no significant difference in median survival between the RRM1-positive and RRM1-negative groups (22 months vs. 19 months, P = 0.540). Our results show that RRM1-negative expression is related with longer progression-free survival in advanced NPC patients treated with gemcitabine-based regimens.

Combined analysis of mRNA expression of ERCC1, BAG-1, BRCA1, RRM1 and TUBB3 to predict prognosis in patients with non-small cell lung cancer who received adjuvant chemotherapy

BackgroundThe aim of this study was to investigate prognostic value of excision repair cross-complementing 1 (ERCC1), BCL2-associated athanogene (BAG-1), the breast and ovarian cancer susceptibility gene 1 (BRCA1), ribonucleotide reductase subunit M1 (RRM1) and class III β-tubulin (TUBB3) in patients with non-small cell lung cancer (NSCLC) who received platinum- based adjuvant chemotherapy.MethodsMessenger RNA expressions of these genes were examined in 85 tumor tissues and 34 adjacent tissue samples using semi-quantitative RT-PCR. The expressions of these five genes were analyzed in relation to chemotherapy and progression-free survival (PFS) and overall survival (OS). Seventy-four patients were enrolled into chemotherapy.ResultsPatients with ERCC1 or BAG-1 negative expression had a significantly longer PFS (P = 0.001 and P = 0.001) and OS (P = 0.001 and P = 0.001) than those with positive expression. Patients with negative ERCC1 and BAG-1 expression benefited more from platinum regimen (P = 0.001 and P = 0.002). Patients with BRCA1 negative expression might have a longer OS (P = 0.052), but not PFS (P = 0.088) than those with BRCA1 positive expression. A significant relationship was observed between the mRNA expression of ERCC1 and BAG-1 (P = 0.042). In multivariate analysis, ERCC1 and BAG-1 were significantly favorable factors for PFS (P = 0.018 and P = 0.017) and OS (P = 0.027 and P = 0.022).ConclusionsERCC1 and BAG-1 are determinants of survival after surgical treatment of NSCLC, and its mRNA expression in tumor tissues could be used to predict the prognosis of NSCLC treated by platinum.

Gemcitabine, Dexamethasone and Cisplatin (GDP regimen) As First Salvage Treatment of Patients with Refractory or Relapsed Diffuse large B Cell Non Hodgkin Lymphoma (DLBCL)

Abstract 2696 Background:Many chemotherapy regimens have been used for patients with refractory or relapsed DLBCL. No regimen has demonstrated superiority to another in this setting. Specific markers could predict the response to certain agents. Aim: to evaluate the response of GDP regimen in relapsed and refractory DLBCL patients and to assess ribonucleotide reductase subunit M1 (RRM1) as a possible predictor marker to Gemcitabine response. Patients and method: Patients with Relapsed or refractory DLBCL after one previous anthracycline-containing chemotherapy regimen were treated with the GDP regimen. RRM1 was assessed by immunohistochemistry in 55 cases and its expression was correlated to treatment outcome. Patients who could not proceed to stem cell transplantation (SCT) were followed for chemotherapy response and their results are presented. Results: The study included 70 patients with a median age of 40 years (range 18–73). At start of GDP, 19 patients (27.1%) were refractory and 51 (72.9%) were relapsed. After 4 cycles of treatment, 42 patients achieved CR, with a CR rate of 60% (95% CI: 53–68%). The median DFS was 6 months (95% CI: 5 to7 months), this DFS didn’t include patients who underwent auto SCT. After a median follow-up of 20 months (range 6 to 30 months), the median OS of the patients who achieved CR was not reached, while those who didn’t achieve CR had a median OS of 27.4 months (p= 0.01). Correlation between response and the pretreatment prognostic factors including IPI score or any of its elements, previous line of chemotherapy, time to relapse and status at time of salvage were studied with only significant difference in response to GDP between patients with refractory and those with relapsed disease (CR= 21.1% versus 60% respectively, p < 0.001). There was significant correlation between RRMI study results and response to GDP, 30/31 cases with low RRM1 expression achieved CR (96.8%), while only 7/24 cases with high expression achieved CR (29.2%), (p=0.001). No significant relation could be found between RRM1 expression and DFS. Conclusion: GDP regimen is active for patients with refractory or relapsed DLBCL however, the duration of response is short and high-dose therapy with SCT support is the reference postremission treatment. RRM1expression can predict response to Gemcitabine-based chemotherapy. Disclosures:No relevant conflicts of interest to declare.

Ribonucleotide reductase subunit M1 expression in resectable, muscle-invasive urothelial cancer correlates with survival in younger patients

To assess whether high ribonucleotide reductase subunit M1 (RRM1) expression in patients with resected, muscle-invasive (T2-4NxM0) urothelial carcinoma (UC) correlated with longer overall survival (OS). RRM1 is the primary cellular target of gemcitabine and previous studies in resected early-stage lung cancer have shown a survival benefit for patients with high expression. In all, 84 radical cystectomy specimens with muscle-invasive UC were identified from existing tissue microarrays. The patients' medical records were retrospectively reviewed to confirm pathology and stage. Specimens were analysed for RRM1 expression using automated quantitative analysis. The median value of RRM1 was established a priori as the threshold for high and low expression. The median age of the patients was 69 years. Stages were nearly equally distributed: 30%, 38%, and 32% for stage II, III, and IV, respectively. Most were high grade (99%) with no nodal involvement (69%). The median (range) OS was 2.0 (0-13.1) years. Tumoral RRM1 levels did not correlate with OS for the entire cohort, but when adjusted for age, high tumoral RRM1 expression in younger patients (aged <70 years) correlated with increased survival. Younger patients with high RRM1 expression had a median OS of 10.6 years compared with 1.6 years in older patients (P= 0.001). There was no difference in OS among low RRM1 expressors: 2.3 vs 1.6 years in younger and older patients, respectively (P= 0.22). Our results suggest that high RRM1 expression may be prognostic for improved survival in patients with muscle-invasive UC aged <70 years.

切除修复交叉互补基因1、核苷酸还原酶M1亚基和β-微管蛋白III的表达对I～III期非小细胞肺癌术后辅助化疗疗效的预测意义

To determine the predictive value of excision repair cross complementation group 1 (ERCC1),ribonucleotide reductase subunit M1 (RRM1), and β-tubulin 3 expressions in postoperative patients with stage 1- 3 non-small cell lung cancer (NSCLC) receiving adjuvant chemotherapy. All NSCLC patients received surgery therapy followed by at least one cycle of adjuvant chemotherapy in our hospital from January 2004 to December 2007. The expressions of ERCC1, RRM1, and β-tubulin 3 were detected by immunohistochemical methods. The relationships among clinicopathologic characteristics, chemotherapy regimens,biomarkers' expressions and disease-free survival (DFS) were analyzed. The high-expression rates of ERCC1, RRM1, and β-tubulin 3 were 36.4%,43.7%,and 38.4%,respectively. The expressions of these three biomarkers were not correlated. After a median follow-up of 35.8 months, 80 patients experienced metastatic or recurrent tumors and 40 patients died. The median overall survival was not reached and the median DFS was 24.1 months. Univariate survival analysis showed that sex, clinical stage,and adenocarcinoma or not were related to DFS, while age, smoke history, chemotherapy regimens, and expression levels of ERCC1, RRM1, and β-tubulin 3 has no prognostic significance in these surgically resected NSCLC patients who were receiving adjuvant chemotherapy. Male (P=0.036), earlier clinical stage (P=0.001), and non-adenocarcinoma (P=0.004) predicted better DFS. Stratified analysis indicated that in RRM1 high-expression strata,the regimens with gemcitabine had curtailed DFS compared with other regimens (P=0.054); in β-tubulin 3 high-expression strata,the regimens containing taxane (including paclitaxel and docetaxel subgroups) had curtailed DFS compared with other regimens (P=0.076), although there was no statistical significance. However,there were no similar predictive significance in RRM1 and β-tubulin 3 low-expression strata or in ERCC1 strata with different expression levels. COX proportional regression analysis showed that adenocarcinoma or not and clinical stage were independent risk factors of DFS in this population. In postoperative NSCLC patients who are receiving adjuvant chemotherapy, patients with high expression of RRM1 tends to be resistant to gemcitabine and patients with high expression of β-tubulin 3 tends to be resistant to taxane drugs. ERCC1, RRM1, and β-tubulin 3 detected by immunohistochemistry can be biomarkers to help to choose better chemotherapy regimen and predict the effectiveness of adjuvant chemotherapy.

Predicting gemcitabine transport and toxicity in human pancreatic cancer cell lines with the positron emission tomography tracer 3′-deoxy-3′-fluorothymidine

The abundance of human equilibrative nucleoside transporter 1 (hENT1) has recently been shown to be a predictive marker of benefit from gemcitabine therapy in patients with pancreatic cancer. Since hENT1 is also important for the uptake of positron emission tomography (PET) tracer 3′-deoxy-3′-fluorothymidine (FLT) in various cultured human cell lines, this study was undertaken to determine if FLT uptake predicts gemcitabine uptake and/or toxicity in a panel of human pancreatic cancer cell lines (Capan-2, AsPC-1, BxPC-3, PL45, MIA PaCa-2, and PANC-1). Capan-2 cells displayed the lowest levels of (1) extracellular nitrobenzylmercaptopurine ribonucleoside (NBMPR) binding, which represents cell-surface hENT1, (2) FLT and gemcitabine uptake during short (1–45 s) and prolonged (1 h) periods, and (3) gemcitabine sensitivity. Exposure to NBMPR (inhibits only hENT1) or dilazep (inhibits hENT1 and hENT2) reduced FLT and gemcitabine uptake and gemcitabine sensitivity, with dilazep having greater effects than NBMPR. Gemcitabine permeation was almost completely mediated, primarily by hENT1 and to a lesser extent by hENT2, whereas FLT permeation included a substantial component of passive diffusion. In five of six cell lines, correlations were observed between (1) FLT and gemcitabine initial rates of uptake, (2) gemcitabine uptake and gemcitabine toxicity, (3) FLT uptake and gemcitabine toxicity, and (4) ribonucleotide reductase subunit M1 expression and gemcitabine toxicity. FLT and gemcitabine uptake were comparable for predicting gemcitabine toxicity in the tested pancreatic cancer cell lines suggesting that FLT PET may provide clinically useful information about tumor gemcitabine transport capacity and sensitivity.

Relationship between the level of RRM1 expression and the sensitivity to gemcitabine in the esophageal squamous cell carcinoma cell lines

Ribonucleotide reductase subunit M1 (RRM1) is the intracellular target of gemcitabine (GEM). The aim of this study is to explore the relationship between the level of RRM1 expression and the sensitivity to GEM in the esophageal squamous cell carcinoma cell lines. Four esophageal squamous cell carcinoma cell lines (Kyse-150, Kyse-450, 9706 and Eca-109) were cultured in vitro. In the same period, RRM1 expression level was measured by RT-PCR and Western blot, and cell sensitivity to GEM was determined by CCK-8 assay. The relation between cell sensitivity and RRM1 expression was further analyzed. Kyse-450 cells were continuously cultured in the medium containing 50 nM GEM. RRM1 expression was measured at different time points to monitor the dynamic changes in the surviving cells. Inhibition of RRM1 expression by RNAi method was applied and the effect on GEM-sensitivity was further examined. The IC(50) of Eca-109, Kyse-150, Kyse-450 and 9706 cells were (0.92 +/- 0.17), (0.48 +/- 0.11), (0.29 +/- 0.06) and (0.02 +/- 0.01) mmol/L, respectively. The expressions of RRM1 protein and mRNA of Eca-109 cell line were the highest detected by Western blot and RT-PCR, followed by Kyse-150 and Kyse-450, and the lowest one was 9706 cell line. When Kyse-450 cells were continuously treated with 50 nmol/L GEM, the level of RRM1 protein was increasing in the surviving cells. RRM1 siRNA could effectively knock down the expression of RRM1 and significantly increase the cell sensitivity to GEM (P = 0.035). The level of RRM1 expression correlates with the cell sensitivity to gemcitabine. The cells with a lower level of RRM1 expression are more sensitive to gemcitabine.

Positive expression of ERCC1 predicts a poorer platinum-based treatment outcome in Chinese patients with advanced non-small-cell lung cancer

The goal of this study is to determine role of excision repair cross-complementing group 1 gene (ERCC1) and ribonucleotide reductase subunit M1 (RRM1) expression in predicting response and survival in Chinese patients with advanced stage non-small cell lung cancer (NSCLC) treated with platinum-based chemotherapy. Formalin-fixed, paraffin-embedded biopsy tissues were retrospectively obtained from 124 advanced NSCLC patients. Protein expression levels of ERCC1 and RRM1 were determined by immunohistochemistry (IHC). Associations between expression of ERCC1 and RRM1 and clinic-pathologic parameters were analyzed. The study shows that ERCC1 and RRM1 expression was detected in 43 (35%) and 50 (40%) of the 124 tumor samples, respectively. Expression of ERCC1 and RRM1 was negatively associated with tumor response. Fifty-four percent patients whose tumors did not express ERCC1 had partial response (PR) compared to 33% whose tumors expressed the protein (P = 0.022). Similarly, 54% patients whose tumor did not express RRM1 had PR compared to 36% whose tumors expression the protein (P = 0.042). Further, patients whose tumors lacked of ERCC1 but not RRM1 expression had a longer median survival time than those tumors expressed ERCC1 (13.4 months versus 9.1 months; P = 0.006), which is independent of other prognostic factors (P = 0.0066). In conclusion, tumor ERCC1 expression is associated with tumor response and patients' survival in Chinese advanced NSCLC patients treated with platinum-based regimen and may serve as a biomarker in predicting tumor response and clinical outcome in the patient population.

Can We Customize Chemotherapy? Individualizing Cytotoxic Regimens in Advanced Non–Small-Cell Lung Cancer

Metastatic non–small-cell lung cancer remains a fatal disease with a median survival of < 1 year. A critical challenge is to develop predictive markers for customizing platinum-based treatment. The first studies focused on the excision repair cross-complementing 1 (ERCC1) gene in this difficult task. Several layers of evidence indicate that ERCC1 mRNA expression could be a predictive marker for cisplatin alone or in combination with certain drugs such as etoposide, gemcitabine, and 5-fluorouracil but not in combination with antimicrotubule drugs. Several retrospective studies demonstrated an impressive survival advantage for gemcitabine plus cisplatin but not for other combinations in tumors with low ERCC1 expression. A customized phase III ERCC1-based trial met the primary endpoint of improvement in response but not in survival, leading us to hypothesize that docetaxel might not be the most appropriate partner for cisplatin in the presence of low ERCC1 levels or for gemcitabine in the presence of high ERCC1 levels. A phase II study demonstrated the feasibility of combining carboplatin, gemcitabine, docetaxel, and vinorelbine according to ERCC1 and ribonucleotide reductase subunit M1 expression levels. These findings highlight the importance of continual learning, and decision-making strategies for customizing treatment should reflect the limitations of our knowledge.