Excision Repair Cross-complementation Group 1 Protein Expression Research Articles

Multi-index comprehensive evaluation of the efficacy and response mechanism of immunotherapy in non-small cell lung cancer.

This research conducted multi-index comprehensive evaluations of the immunotherapeutic efficacy and response in non-small cell lung cancer (NSCLC). Forty-five patients with epidermal growth factor receptor (EGFR)/anaplastic lymphoma kinase (ALK) wild-type advanced NSCLC who received immunotherapy were included. Immunohistochemistry was adopted to detect the expression levels of programmed death ligand 1 (PD-L1) with X-ray cross-complementing protein 1 (XRCC1) and excision repair cross-complementing group 1 (ERCC1) proteins in tumor tissues. Flow cytometry was utilized to measure the levels of T-cell subsets in peripheral blood before and after treatment. PCR-RELP method was employed to evaluate XRCC1 and ERCC1 gene polymorphisms in peripheral blood. According to the treatment effect, patients evaluated as complete response (CR), partial response (PR), and stable disease (SD) were categorized into the immune response group, and patients evaluated as progressive disease (PD) were categorized into the immune unresponsive group. The correlation between PD-L1 protein expression, XRCC1 and ERCC1 protein expression, gene polymorphisms, T-cell subpopulation levels, and treatment efficacy was analyzed. The therapeutic efficacy of patients with positive PD-L1 expression was better than that of patients with negative PD-L1 expression (P < 0.05). After treatment, peripheral blood CD3+ and CD4+ cell levels and Thl/Th2 cell levels were higher and CD8+ T cells were lower in the immune response group than in the immune unresponsive group (P < 0.05). Among the patients in the immune response group, peripheral blood CD3+ and CD4+ cell levels were higher and CD8+ T cells were lower in patients with positive PD-L1 expression than in patients with negative PD-L1 expression (P < 0.05). In the XRCC1 gene, the proportion of patients in the immune response group carrying the Arg/Trp + Trp/Trp genotype was higher than that of patients in the immune unresponsive group (P < 0.05). In the ERCC1 gene, the proportion of patients in the immune response group carrying the C/T + T/T genotype was higher than that of patients in the immune unresponsive group (P < 0.05). The positive expression rates of XRCC1 and ERCC1 in patients in the immune unresponsive group were higher than those in the immune response group (P < 0.05). PD-L1 protein expression, XRCC1 and ERCC1 protein expression, and gene polymorphisms are associated with immunotherapy outcome in EGFR/ALK wild-type advanced NSCLC patients, and may be biological indicators for predicting immunotherapy outcome in EGFR/ALK wild-type advanced NSCLC patients.

Osteopontin enhances cisplatin resistance of human A549 lung cancer cells via stimulating the PI3K signaling pathway and upregulating ERCC1 expression.

BackgroundTo investigate the regulatory role of osteopontin (OPN) in cisplatin (DDP) resistance in non-small cell lung cancer (NSCLC).MethodsOPN over-expression and interference vectors were constructed. Human NSCLC A549 cells were divided into normal control, shRNA-NC, shRNA-OPN, OPN, negative control (NC), control+DDP, shRNA-NC+DDP, shRNA-OPN+DDP, OPN+DDP, and NC+DDP groups. Cell proliferation was measured by MTT assay. Cell apoptosis was detected by flow cytometry. PI3K (phosphoinositide 3-kinase), p-ERK1/2 (phospho-extracellular signal-regulated kinases) and ERCC1 (excision repair cross-complementation group 1) mRNA and protein expression was determined by real-time PCR and Western blot.ResultsOPN overexpression and interference vectors were successfully constructed. When compared with control group, OPN group had significantly stimulated cell viability, reduced cell apoptosis, and increased expression of PI3K, p-ERK1/2 and ERCC1 (all P<0.05), whereas all other groups had lower cell viability, higher cell apoptosis, and inhibited protein expression. Moreover, OPN+DDP group had significantly higher proliferation, lower cell apoptosis, and enhanced PI3K, p-ERK1/2 and ERCC1 mRNA and protein expression compared with shRNA-OPN+DDP (all P<0.05).ConclusionsOPN can promote tumor cell growth and reduce cell apoptosis by activating PI3K pathway, which may induce the resistance of NSCLC to cisplatin. Moreover, the stimulation of ERCC1 expression may also increase the cisplatin resistance of NSCLCs.

Prognostic and Predictive Role of Excision Repair Cross-complementation Group 1 and Thymidylate Synthase in Colorectal Carcinoma Patients Received FOLFOX Chemotherapy: An Immunohistochemical Study.

We aim to determine the frequency of thymidylate synthase (TS) and excision repair cross-complementation group 1 (ERCC-1) immunohistochemical (IHC) expression and its relationship with clinicopathologic variables in colorectal carcinoma (CRC) patients. In addition, we aim to assess the correlation between TS and ERCC-1 expression and the response of these cases to oxaliplatin and 5-fluorouracil chemotherapy (FOLFOX). Fifty-one CRC patients were prepared for IHC analysis of ERCC-1 and TS protein expression. All patients received oxaliplatin and 5-fluorouracil combined chemotherapy (FOLFOX) and were followed up for 24 months. The data analysis showed that high ERCC-1 and TS expression was significantly associated with early treatment failure (P=0.020 and 0.000). In contrast, TS immunoexpression affects the disease-free survival rate (P=0.010). The presence of deep tumor invasion, distant metastasis, lymph node metastasis, and high Dukes' classification were significantly statistically associated with early treatment failure (P=0.001, 0.000, 0.041, and 0.015, respectively). Our results showed that both ERCC-1 and TS are predictive factors for early treatment failure in CRC patients. TS protein is a prognostic factor for disease-free survival rates. This supports the theory that both ERCC-1 and TS can be used to improve chemotherapeutic outcomes in CRC patients. High expression of TS and ERCC-1 helps in the identification of cases that will get fewer benefits from FOLFOX chemotherapy. As an innovative strategy, in these cases, we can use alternative chemotherapeutic regimens or add an extra agent. In addition, Dukes' classification and lymph node metastasis are predictive factors for early treatment failure. Thus, all those values can be used to predict CRC patients with bad prognosis and those who will get fewer benefits from FOLFOX.

Excision repair cross-complementing group 1 (ERCC1) overexpression inhibits cell apoptosis and is associated with unfavorable prognosis of esophageal squamous cell carcinoma.

Excision repair cross-complementing group 1 (ERCC1) functions as a nucleotide excision repair (NER) enzyme. Altered ERCC1 expression or function is closely associated with cancer development and progression. This study determined the association of ERCC1 expression with survivin expression, clinicopathological characteristics, and survival of esophageal squamous cell carcinoma (ESCC) patients after postoperative concurrent chemoradiotherapy.Tissue specimens from 102 resected ESCC patients were acquired for immunohistochemical analysis of ERCC1 and survivin protein expression.ERCC1 expression was detected in 62.7% of ESCC tissues and in 9.8% of normal squamous epithelium tissues (P < .01), while survivin expression was detected in 60.8% of ESCC tissues and in 19.6% of normal squamous epithelia (P < .01). ERCC1 overexpression associated with advanced tumor clinical stage and lymph node metastasis (P < .05), but not with tumor size, depth of invasion, or differentiation (P > .05). ERCC1 overexpression was also associated with survivin levels (r = 0.42, P < .01) and worse progression-free survival of ESCC patients after concurrent chemoradiotherapy. Multivariate analysis data revealed that ERCC1 and survivin protein expression were independent predictors of overall survival of ESCC patients after chemotherapy and/or radiotherapy (P < .05).ERCC1 overexpression is an important phenotype that is associated with ESCC lymph node metastasis and advanced tumor clinical stages. ERCC1 expression may also inhibit ESCC cell apoptosis via regulating survivin expression, and ERCC1 and survivin overexpression are independent predictors of prognosis for ESCC patients who receive chemotherapy and/or radiotherapy.

Abstract 5189: Evaluation of thymidylate synthase mRNA expression in circulating tumor cells from non-metastatic colon and rectal patients

Abstract Background: colorectal cancer is the third leading cause of cancer death in the United States. With different behaviors, rectal cancer represents worse prognosis, even in the locally advanced setting, with therapy following three steps, consisting of neoadjuvant chemoradiation, surgery and adjuvant chemotherapy. For non-metastatic colon cancer, the main treatment is surgery and then, patients can or not undergo adjuvant chemotherapy. Studies have shown that circulating tumor cells (CTCs) may be involved in metastatization process. Thymidylate synthase (TYMS) acts in the 5-Fluorouracil metabolism, and when overexpressed, can confer resistance to treatment, which is commonly used for neoadjuvant treatment (rectal cancer) and in combination with oxaliplatin for adjuvant treatment (colon cancer). Excision repair cross complementation Group 1 (ERCC1) is a protein involved in damage DNA repair process that is described to confer resistance to platinum-based chemotherapy. Objective: evaluate the TYMS messenger RNA (mRNA) expression in CTCs isolated from patients with locally advanced colon and rectal cancers; and its predictable value in treatment resistance. For patients with colon cancer, we also evaluated the ERCC1 protein expression in order to verify the adjuvant therapy resistance related to oxaliplatin. Methods: We collected 10 mL of blood before the beginning of treatment and processed it on ISET® (Rarecells) device. RNAscope® Technology is a novel chromogenic in situ hybridization (CISH) assay for detection of target RNA within intact cells. The expression of TYMS target gene was performed using the kit from ACDbio, as described on the protocol, but standardized to cytology. Results: there were included 12 patients of each type of tumor. We analyzed samples from 12 rectal cancer patients, and found 10 with overexpression of TYMS mRNA in CTCs. Curiously, of the two patients who had TYMSneg mRNA expression; one had complete pathological response and the other substantial down-staging. In the positive cases, 50% presented response (partial or complete) and 50% present no response and systemic progression. This result will better understood when we perform the evaluation of the protein TYMS. Among the samples from colon cancer patients, we found 8 out 12 as CTCs TYMSpos mRNA expression. Unfortunately, our recent data does not allow us to do analysis of recurrence. However, one of the positive cases had recurrence. Among the 4 TYMSneg, one had recurrence. This can be explained by the positive immunocytochemical expression of the ERCC1 protein, which confers resistance to oxaliplatin. Our next steps are to expand the CISH experiments in a larger group of patients as well as to evaluate the TYMS protein expression in the same cases. Conclusion: our preliminary results point molecular analysis of CTCs as predictor marker of colon/rectal cancer treatments. Citation Format: Bianca Troncarelli Flores, Emne Ali Abdallah, Virgílio Souza e Silva, Celso Abdon Mello, Samuel Aguiar, Renata Mayumi Takahashi, Vanessa Silva Alves, Bruna Elisa Kupper, Ludmilla T. Chinen. Evaluation of thymidylate synthase mRNA expression in circulating tumor cells from non-metastatic colon and rectal patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5189.

Increased ERCC1 expression is linked to chromosomal aberrations and adverse tumor biology in prostate cancer

BackgroundAnimal model experiments have suggested a role of the DNA repair protein ERCC1 (Excision Repair Cross-Complementation Group 1) in prostate cancer progression.MethodsTo better understand the impact of ERCC1 protein expression in human prostate cancer, a preexisting tissue microarray (TMA) containing more than 12,000 prostate cancer specimens was analyzed by immunohistochemistry and data were compared with tumor phenotype, PSA recurrence and several of the most common genomic alterations (TMPRSS2:ERG fusions: deletions of PTEN, 6q, 5q, 3p).ResultsERCC1 staining was seen in 64.7% of 10,436 interpretable tissues and was considered weak in 37.1%, moderate in 22.6% and strong in 5% of tumors. High-level ERCC1 staining was linked to advanced pT stage, high Gleason grade, positive lymph nodes, high pre-operative serum PSA, and positive surgical margin status (p < 0.0001 each). High ERCC1 expression was strongly associated with an elevated risk of PSA recurrence (p < 0.0001). This was independent of established prognostic features. A subgroup analysis of cancers defined by comparable quantitative Gleason grades revealed that the prognostic impact was mostly driven by low-grade tumors with a Gleason 3 + 3 or 3 + 4 (Gleason 4: ≤5%). High ERCC1 expression was strongly associated with the presence of genomic alterations and expression levels increased with the number of deletions present in the tumor. These latter data suggest a functional relationship of ERCC1 expression with genomic instability.ConclusionThe results of our study demonstrate that expression of ERCC1 - a potential surrogate for genomic instability - is an independent prognostic marker in prostate cancer with particular importance in low-grade tumors.

The age-related expression decline of ERCC1 and XPF for forensic age estimation: A preliminary study

The age-related capacity decline of DNA damage repair in human peripheral blood has been demonstrated. Excision repair cross-complementation group1 (ERCC1) and Xeroderma pigmentosum complementation group F (XPF) were rate-limiting enzyme in nucleotide excision repair (NER) which was known as the most important DNA damage repair system. Consequently, we hypothesized that the expression and/or activity of ERCC1 and XPF may be associated with age. However, little was known about the quantitative relationship of ERCC1 and XPF expression levels with age. The aim of the present study was to analyze the correlation of ERCC1 and XPF expression levels with age by detecting the ERCC1 and XPF mRNA levels in peripheral blood mononuclear cells(PBMCs) and protein levels in plasma in healthy ethnic Han Chinese individuals, and finally find new molecular markers for forensic age estimation by establishing the mathematical model between ERCC1 and XPF expression levels and age. The results showed that the ERCC1 and XPF mRNA relative expression levels in PBMCs declined in an age-dependent manner (r = −0.578/−0.844, respectively, P < 0.01). The formula for age estimation based on the ERCC1 and XPF mRNA relative expression levels decline in PBMCs were Y = 3.3E−5x2−0.0261x+1.9175 (R2 = 0.3244, P < 0.01) and Y = 0.0003x2−0.0459x+2.0439 (R2 = 0.729, P < 0.01), respectively. There were no significant differences of the ERCC1 or XPF protein expression levels in plasma between age groups (P > 0.05). Furthermore, there were no significant differences of the ERCC1 or XPF mRNA and/or protein expression levels between males and females(P > 0.05). It suggested that the ERCC1 and XPF mRNA expression levels could be considered as valuable additional tool in individual age estimation, especially in cases where traditional morphologic method was inefficient or absent in forensic practice.

High ERCC1 expression is associated with platinum-resistance, but not survival in patients with epithelial ovarian cancer.

The present study aimed to investigate the association between excision repair cross-complementation group 1 (ERCC1) expression and clinical resistance to platinum-based chemotherapy or clinical characteristics, including survival time, in patients with epithelial ovarian cancer (EOC). ERCC1 expression was determined by immunohistochemical staining in 92 tumor specimens from patients with EOC. The effect of ERCC1 expression on progression-free survival time (PFS) or overall survival time (OS), and its association with clinical resistance to platinum-based chemotherapy was investigated by Kaplan-Meier survival analysis, Cox regression analysis and the χ2 test. Of 92 patients with EOC, 89.13% (82/92) had ERCC1-positive tumors. The positive rate was significantly higher in platinum-resistant patients compared with those who were platinum-responding (P<0.05). The PFS and median OS were 12 and 30 months, respectively, in ERCC1 high expression patients, and 17 and 39 months, respectively, in ERCC1 low expression patients. However, there was no statistically significant difference in PFS (P=0.099) or OS (P=0.103) between the high and low expression groups. Furthermore, it was identified that ERCC1 was not an independent factor affecting the prognosis of patients with EOC based on Cox proportional hazards regression analysis. These results demonstrate that high ERCC1 expression is associated with resistance to platinum-based chemotherapy, but not with survival time, and ERCC1 protein expression is not an independent factor or the only factor affecting the prognosis of patients with EOC.

Excision Repair Cross-Complementation Group 1 (ERCC1): A Prognostic and Predictive Biomarker in Patients with Colorectal Cancer Receiving Adjuvant Oxaliplatin Based Chemotherapy

Background: Colorectal cancer (CRC) ranks as the third most common cancer and the third most killing cancer worldwide [1]. The addition of oxaliplatin to fluorouracil (FOLFOX) or capecitabine (CAPOX) has become a fundamental component of chemotherapeutic regimens and chemoradiation in adjuvant treatment of CRC cancer. Excision repair cross-complementation group 1 (ERCC1) encodes an enzyme that is essential for the efficient repair of DNA damage induced by platinum compounds including Oxaliplatin. Purpose: This study aims to investigate the role of ERCC1 as a predictive and prognostic marker in colorectal patients receiving oxaliplatin based chemotherapy and chemoradiation. Patients and Methods: 100 annotated stage III CRC patients were prepared as immunohistochemical (IHC) analysis of ERCC1 protein expression. All of the patients received oxaliplatin based chemotherapy. Results: Analysis of data showed that high ERCC1 expression was significantly associated with early treatment failure and disease free survival among patient with stage III CRC. Conclusion: High ERCC1 expression was an independent predictor factor of early treatment failure (P = 0.004).

Abstract PL05-01: ERCC1 alternative readouts

Abstract Platinum-based compounds, including cisplatin and oxaliplatin, trigger cell death by distorting the helix structure and subsequently affecting DNA replication and transcription. The therapeutic efficacy of platinum-based chemotherapy varies among patients and only a portion of them have real benefits. Precision medicine searches for the optimal therapy for each patient. The proteins involved in DNA repair are thus potential biomarkers to select patients more prone to respond to platinum-based chemotherapy. The ERCC1 (excision repair cross-complementation group 1) protein is essential in nucleotide excision repair (NER) and other repair pathways involved in the repair of platinum-induced DNA damages. ERCC1 with its partner XPF (xeroderma pigmentosum complementation group F) form a structure-specific endonuclease that is responsible for 5' incision during DNA repair. ERCC1 has been reported to be a prognostic and predictive biomarker for cisplatin treatments in various cancers. Many studies have shown that the ERCC1 status, including ERCC1 polymorphisms, ERCC1 RNA expression and ERCC1 protein expression, are associated with platinum-based therapy efficacy. However, this correlation between ERCC1 status and the clinical outcome has not always reached a consensus, probably due to inadequate methods to determine ERCC1 expression and unreliability of the current commercial ERCC1 antibodies. Alternative splicing leads to several isoforms of ERCC1. We have reported that only ERCC1-202 mediates highly efficient removal of platinum-GG intrastrand adducts and should therefore be considered as the only functional isoform. Putative roles of the other isoforms remain unknown. The expression of ERCC1 is mostly determined by qRT-PCR or IHC. However, these two methods are not able to discriminate the functional ERCC1-202 isoform from other isoforms. We will present the design of a tailor-made antibody directed against the interacting complex ERCC1/XPF. This new ERCC1/XPF heterodimer-specific antibody permits the detection of the functional ERCC1-202 isoform when it is used in combination with other antibodies in proximity ligation assays (PLA). The applicability of this alternative read out for ERCC1 expression on clinical samples will be discussed. Citation Format: Mei-Shiue Kuo, Ken A. Olaussen, Jean-Charles Soria. ERCC1 alternative readouts. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr PL05-01.

Clinical utility of plasma Epstein‐Barr virus DNA and ERCC1 single nucleotide polymorphism in nasopharyngeal carcinoma

Single nucleotide polymorphism (SNP) of the excision repair cross-complementing group 1 (ERCC1) gene has been linked with sensitivity to platinum and radiation. The authors hypothesized that the ERCC1 genotype for the SNPs cytosine-to-thymine substitution at codon 118 (C118T) and cytosine-to-adenine substitution at codon 8092 (C8092A) is prognostic in patients with nasopharyngeal carcinoma (NPC) who receive either radiotherapy (RT) or cisplatin plus RT. The authors tested their hypothesis using biomarker screening samples from the Hong Kong NPC Study Group 0502 trial, which was a prospective, multicenter clinical trial that used post-RT plasma Epstein-Bar virus (EBV) DNA (pEBV) levels to screen patients with high-risk NPC for adjuvant chemotherapy. ERCC1 SNPs were analyzed in 576 consecutive patients who were screened by pEBV. In the total biomarker population, there was no significant association of ERCC1 C118T or C8092A genotype with relapse-free survival (RFS) or overall survival (OS). There also was no correlation between ERCC1 genotype and ERCC1 protein or messenger RNA expression in a subset of patients who had available paired biopsies. Post-RT pEBV status was the only independent prognosticator for RFS and OS in multivariate analyses. However, there was a significant interaction between ERCC1 C118T genotype and post-RT pEBV status (RFS, P = .0106; OS, P = .0067). The ERCC1 C118T genotype was significantly associated with both RFS (hazard ratio, 1.67; 95% confidence interval, 1.07-2.61; P = .024) and OS (hazard ratio, 2.31; 95% confidence interval, 1.22-4.40; P = .0106) in the post-RT pEBV-negative population, but not in the pEBV-positive population. The current results prospectively validate pEBV as the most significant prognostic biomarker in NPC that can be used to select high-risk patients for adjuvant therapy. The ERCC1 C118T genotype may help to identify a favorable subgroup (approximately 7%) of pEBV-negative patients with NPC who have an excellent prognosis and can be spared the toxicities of further therapy.

ERCC1 and Thymidylate Synthase as Prognostic Biomarkers in Malignant Mesothelioma

The standard of care for malignant pleural mesothelioma (MPM) is a combination of platinum compounds and Pemetrexed. Still, the median survival of MPM patients is low (about 12 months). Excision Repair Cross Complementing Group 1 (ERCC1) acts by removing DNA adducts formed by platinum compounds, whereas Pemetrexed inhibits Thymidylate Synthase (TS), which is involved in DNA synthesis.Pleural biopsies were collected from 140 MPM patients (98 males and 42 females, mean age 68 years) before treatment initiation. Average disease specific survival was 13 months (range 1‐60). mRNA was extracted from formalin fixed paraffin blocks, reverse transcribed and assayed by qRT‐PCR. Immunohistochemistry with anti‐ERCC1 and anti‐TS mAbs was scored semi‐quantitatively.Of 140 patients, 102 were treated, the remaining were not because of bad performance status or advanced age. Patients with low ERCC1 protein expression survived longer (24.3 vs 12.5 months, P=.03). Likewise, patients with low levels of TS mRNA had better survival (P=.01); there was no advantage, however, in the treated sub‐group, suggesting that Pemetrexed treatment and low TS expression had no additive value.ERCC1 protein expression and TS gene expression both seem important prognostic markers; we suggest their evaluation in routine biopsies from MPM patients.

XRCC1 and ERCC1 polymorphisms are related to susceptibility and survival of colorectal cancer in the Chinese population.

Excision repair cross complementing group 1 (ERCC1) and X-ray repair cross-complementing groups 1 (XRCC1) are DNA repair enzymes. Polymorphisms in DNA repair genes may be important factors affecting cancer susceptibility, prognosis and therapy outcome. The purpose of this study was to investigate the correlation of ERCC1 and XRCC1 polymorphisms with colorectal cancer (CRC) risk, and explore the effect of polymorphisms on event-free, overall survival and oxaliplatin-based therapy in CRC patients. Genotyping was examined with the iMLDR technique. An unconditional logistic regression model was used to estimate the association of certain polymorphisms with CRC risk. The Kaplan-Meier method, log-rank test and Cox regression model were employed to evaluate the effects of polymorphisms on survival analysis. Results showed that Trp/Trp genotype of XRCC1 Arg194Trp and AA genotype of ERCC1 rs2336219 have a significantly increased risk of CRC; Trp allele of XRCC1 Arg194Trp and CC genotype of ERCC1 rs735482 were associated with lower response to oxaliplatin-based chemotherapy, a shorter survival and a higher risk of relapse or metastasis. 194Trp/280Arg/399Arg haplotype was associated with a significant resistance, and the ERCC1 protein expression was statistically higher in tumours with rs735482 CC genotype than with AA genotype. Our studies indicate that XRCC1 and ERCC1 polymorphisms probably affect susceptibility, chemotherapy response and survival of CRC patients.

192P - Prognostic Biomarkers in Malignant Pleural Mesothelioma

ABSTRACT Aim: The standard of care for malignant pleural mesothelioma (MPM) includes combination of platinum compounds and pemetrexed. Despite this, the median survival of MPM patients is still bad (12 months). Excision Repair Cross Complementing Group 1 (ERCC1) acts removing DNA adducts formed by platinum compounds, whereas Pemetrexed inhibits Thymidylate Synthase (TS), a folic enzyme also involved in ex novo synthesis of DNA. This study aims to analyse retrospectively, in a large series of MPM specimens, the ERCC1 and TS gene and protein expression to investigate their prognostic and predictive role. Methods: Pleural biopsies were collected from140 consecutive MPM patients (98 males and 42 females, mean age 68 years, range 27-90). Histologic subtypes were: epithelioid (#103, 73%), sarcomatoid (#19, 14%) and biphasic (#18, 13%). At the end of the study 113 patients were died (average disease specific survival (DSS) 13 months; range 1-60); mRNA was extracted from formalin fixed paraffin embedded blocks and reverse transcribed in cDNA to perform gene expression assay by qRT-PCR. Immunohistochemistry was performed by anti-ERCC1 and anti-TS mAbs and the results were scored from 0 to 3 depending on the percentage of positive tumour cells and staining intensity. The results were statistically correlated with the clinical features and with the DSS of the patients respectively by Fisher's exact test and Logrank test. Results: Of 140 patients 102 received platinum and pemetrexed (in single or in combination), the remaining were untreated due to bad performance status, advanced age or refusing therapy. Statistically significant correlation was found between the lack of protein expression of ERCC1 and DSS in the whole cohort (P = .03). Low levels of TS mRNA were significantly correlated with the DSS in the whole cohort (P = .01) and in the untreated subgroup (including patients not receiving pemetrexed) (P = .04), whereas did not in the subgroup of pemetrexed treated patients Conclusions: ERCC1 protein expression, scored by IHC, and TS gene expression, evaluated by mRNA analysis, seem both important prognostic markers; we suggest their evaluation in routine biopsies from patients with MPM. Disclosure: All authors have declared no conflicts of interest.

Cell cycle association and hypoxia regulation of excision repair cross complementation group 1 protein (ERCC1) in tumor cells of head and neck cancer.

Excision repair cross complementation group 1 (ERCC1) is a key component of homologous recombination-based repair of interstrand DNA cross-links (ICLs). As a consequence, ERCC1 mediates resistance to mitomycin C (MMC) and platinum chemotherapeutic agents and may predict treatment failure. Clinical response to MMC or cisplatin (CDDP)-based radiochemotherapy (RCT) was assessed in 106 head and neck squamous cell carcinoma (HNSCC) patients and correlated with cell nuclear immunoreactivity of the mouse monoclonal (clone: 8 F1) ERCC1 antibody in tumor tissue samples. BEAS-2B epithelial and Detroit 562 pharyngeal squamous carcinoma cells were treated with CDDP, MMC, and 5-fluorouracil (5-FU) at 50 % growth inhibitory (IC-50) concentrations. ERCC1 protein synthesis was compared with cell cycle distribution using combined immunocytochemistry and flow cytometry. ERCC1 messenger RNA (mRNA) and protein expression was investigated in normoxic and hypoxic conditions in Detroit 562 cells. Clinically, the nonresponder revealed significantly lower HNSCC tissue ERCC1 immunoreactivity than the responder (p = 0.0064) or control normal mucosa, which led to further mechanistic investigations. In vitro, control cells and cells treated with cytotoxic agents showed increasing ERCC1 levels from the G1 through S and G2 phases of the cell cycle. In CDDP-treated cells, ERCC1 mRNA and protein expression increased. Under hypoxic conditions, ERCC1 gene expression significantly decreased. Although ERCC1+ cells show increased chemoresistance, they might be particularly radiosensitive, representing G2 cell cycle phase and less hypoxic. ERCC1 expression might be indirectly related with some conditions important for RCT treatment, but it is not a clear predictor for its failure in HNSCC patients.Electronic supplementary materialThe online version of this article (doi:10.1007/s13277-014-2001-2) contains supplementary material, which is available to authorized users.

Combination of TRAP1 and ERCC1 Expression Predicts Clinical Outcomes in Metastatic Colorectal Cancer Treated with Oxaliplatin/5-Fluorouracil

PurposeThe novel heat shock protein tumor necrosis factor receptor-associated protein 1 (TRAP1) is associated with multidrug resistance in colorectal cancer (CRC) cells in vitro. Excision repair cross-complementation group 1 (ERCC1) expression levels in tumor tissues also predict clinical outcomes in metastatic CRC patients receiving combination oxaliplatin and 5-fluorouracil treatment. We investigated whether TRAP1 and ERCC1 protein expression by immunohistochemistry predict clinical outcomes in CRC patients.Materials and MethodsThe study population consisted of 56 patients with metastatic CRC who received first-line oxaliplatin/5-fluorouracil therapy. Clinical response and overall survival (OS) by levels of the markers TRAP1 and ERCC1 were evaluated.ResultsThe rates of TRAP1 and ERCC1 expression were 21% and 52%, respectively. Patients negative for ERCC1 expression showed a tendency to respond to chemotherapy (p=0.066). Median OS was significantly longer in patients negative for TRAP1 than those positive for TRAP1 (p=0.023). Patients negative for ERCC1 expression also had a better OS than those positive for ERCC1 (p=0.021). The median OS was 30.9 months for patients negative for TRAP1 and ERCC1 compared to 13.2 months for those positive for TRAP1 and/or positive for ERCC1 expression (p=0.006). The combination of TRAP1 and ERCC1 expression was significantly associated with the response to chemotherapy (p=0.046) and independently predicted median OS in multivariate analysis (hazard ratio, 2.98; 95% confidence interval, 1.18 to 7.49).ConclusionThe present study demonstrates that the combination of TRAP1 and ERCC1 expression predicts the survival of metastatic CRC patients who were treated with oxaliplatin/5-fluorouracil.

Meta-analysis of Excision Repair Cross-complementation Group 1 (ERCC1) Association with Response to Platinum-based Chemotherapy in Ovarian Cancer

Recent studies suggested that the ovarian cancers with negative excision repair cross-complementation group 1 enzyme (ERCC1) expression have a better response to platinum-based chemotherapy than those with positive ERCC1 expression. The objective of this study was to evaluate whether ERCC1 expression is associated with response to platinum-based chemotherapy in ovarian cancers. MEDLINE, PubMed, Web of Science and CNKI databases were used for searching studies relating to ERCC1 protein expression and response to platinum-based chemotherapy in ovarian cancers. Statistical analysis was based on the method for a fixed effects meta-analysis. Pooled odds ratios (ORs) with 95% confidence intervals for ERCC1 protein expression and response to platinum- based chemotherapy were generated. Publication bias was investigated with Begg's test. Five studies involving 306 patients with ovarian cancer were included. Compared to patients with positive ERCC1 expression, those with negative ERCC1 expression had a better response to platinum-based chemotherapy. The pooled OR was 5.264 (95% CI: 2.928 - 9.464, P < 0.001) and publication bias was not found (P = 0.904). The result was similar in both in Asians and Caucasians (P < 0.001 and P = 0.028, respectively). ERCC1 protein expression status is significantly associated with response to platinum-based chemotherapy in ovarian cancers.

Quantification of Excision Repair Cross-Complementing Group 1 and Survival in p16-Negative Squamous Cell Head and Neck Cancers

Multimodality treatment of squamous cell carcinoma of the head and neck (SCCHN) often involves radiotherapy and cisplatin-based therapy. Elevated activity of DNA repair mechanisms, such as the nucleotide excision repair (NER) pathway, of which ERCC1 is a rate-limiting element, are associated with cisplatin and possibly RT resistance. We have determined excision repair cross-complementing group 1 (ERCC1) expression in human papillomavirus (HPV)-negative SCCHN treated with surgery [± adjuvant radiotherapy/chemoradiation (CRT)]. We assessed ERCC1 protein expression in archival tumors using immunofluorescence staining and automatic quantitative analysis (AQUA) with three antibodies to ERCC1 (8F1, FL297, and HPA029773). Analysis with Classification and Regression Tree (CART) methods ascertained the cutoff points between high/low ERCC1 expression. Multivariable analysis adjusted for age, T, and N stage. Kaplan-Meier curves determined median survival. ERCC1 expression at initial tumor presentation and in recurrent disease were compared. Performance characteristics of antibodies were assessed. ERCC1 low/high groups were defined on the basis of AQUA analysis with 8F1/2009, FL297, and HPA029773. Among patients treated with surgery plus adjuvant radiotherapy/CRT, longer median survival was observed in ERCC1-low versus ERCC1-high tumors (64 vs. 29 months; P = 0.02; HPA029773). Data obtained with HPA029773 indicated no survival difference among patients treated only with surgery. Recurrent cancers had lower ERCC1 AQUA scores than tumors from initial presentation. Extensive characterization indicated optimal specificity and performance by the HPA029773 antibody. Using AQUA, with the specific ERCC1 antibody HPA029773, we found a statistical difference in survival among high/low-ERCC1 tumors from patients treated with surgery and adjuvant radiotherapy.

Protective effect of ginsenoside Rb1 on ultraviolet B-induced damage and its possible mechanisms

Objective To estimate the effect of ginsenoside Rb1 on the production and clearance of cyclobutane pyrimidine dimer (CPD) as well as on the expression of two nucleotide excision repair-associated proteins,xeroderma pigmentosum group C (XPC) and excision repair cross-complementing group 1 (ERCC1),by ultraviolet B (UVB)-irradiated murine epidermal cells and human HaCaT keratinocytes.Methods Totally,42 BALB/c mice were shaved on the back and divided into four groups: untreated group (n =6),UVB group irradiated with UVB only (n =12),low-dose and high-dose Rb1 group (both n =12) treated with Rb1 of 0.5 g/L and 2g/L (100 μl/cm2) respectively two hours before UVB irradiation.The dose of UVB in the animal experiment was 180 mJ/cm2.Half of the mice in each group were killed at 0.5 and 16 hours respectively after the irradiation,then,the back skin was resected and subjected to the determination of CPD levels in the epidermis by immunohistochemical SP method.Some cultured HaCaT cells were divided into several groups to be treated with different concentrations (5,20,50 mg/L) of Rb1 before or after different doses (15 and 30 mnJ/cm2) of UVB irradiation,and cells were collected at 0.5 and 12 hours after the irradiation.Subsequently,genomic DNA was extracted and CPD was detected by dot blot hybridization.Some HaCaT cells were cultured with or without the presence of Rb1 (50 mg/L) and irradiated with UVB (30 mJ/cm2),then,the cells were collected immediately or at 0.5,2,4 and 12 hours after the irradiation,and total protein was extracted and subjected to immunoblot analysis for the quantification of XPC and ERCC1 proteins.Results There was a high level of CPD in the epidermis of mice at 0.5 hour after the irradiation,with no significant differences between these groups (P ＞ 0.05).The number of CPD-positive cells per high power field (× 400) in the murine epidermis at 16 hours was statistically lower in the low-and high-dose Rb1 group than in the UVB group (32.1 ± 8.5 and 14.6 ± 4.1 vs.67.3 ± 11.2,both P ＜0.01).The CPD level in HaCaT cells was similar between these groups at 0.5 hour after UVB irradiation,but was markedly decreased at 12 hours in Rb1-treated groups.After UVB irradiation,the protein expressions of XPC and ERCC1 decreased with time in untreated HaCaT cells but increased with time in Rb1 (50 mg/L)-treated HaCaT cells.In detail,the XPC/glyceraldehyde-3-phosphate dehydrogenase (GAPDH) protein ratio in untreated HaCaT cells was 0.68 ± 0.11 immediately after the irradiation,significantly higher than that at 0.5 hour (0.47 ± 0.09,P＜0.05),2 hours (0.45 ± 0.08,P＜0.05),4 hours (0.37 ± 0.06,P＜0.01),and 12 hours (0.18 ± 0.03,P ＜0.01),and that in Rb1-treated HaCaT cells was 0.56 ± 0.07 immediately after the irradiation,compared to 0.48 ± 0.14 at 0.5 hour (P＞ 0.05),0.68 ± 0.15 at 2 hours (P＞ 0.05),0.97 ± 0.20 at 4 hours (P＜0.01),and 0.79 ± 0.12 at 12 hours (P ＜0.05).The ERCC1/GAPDH protein ratio in untreated HaCaT cells was 0.28 ± 0.03 immediately after the irradiation,higher than that at 0.5 hour (0.25 ± 0.03,P ＞ 0.05),2 hours (0.21 ± 0.02,P＜0.05),4 hours (0.14 ± 0.02,P＜0.01) and 12 hours (0.11 ± 0.01,P＜0.01),and that in Rb1-treated HaCaT cells was 0.27 ± 0.04 immediately after the irradiation,compared to 0.24 ± 0.04 at 0.5 hour (P＞ 0.05),0.29 ± 0.05 at 2 hours (P＞ 0.05),0.35 ± 0.05 at 4 hours (P＜0.05),0.39 ± 0.05 at 12 hours (P ＜0.01).Conclusions Ginsenoside Rb1 shows no obvious effect on the UVB-induced production of CPD,but markedly accelerates the clearance of CPD,which may be partly associated with the upregulation of XPC and ERCC1 protein expression. Key words: GINSENOSIDE; Ultraviolet rays; Cyclobutane pyrimidine dimers; Keratinocytes

Prognostic potential of ERCC1 protein expression and clinicopathologic factors in stage III/N2 non-small cell lung cancer

BackgroundPathological stage III/N2 non-small cell lung cancer (NSCLC) is heterogeneous, and the optimal prognostic marker for survival remains unclear in Chinese patients. The aim of the present study was to assess the prognostic value of the clinicopathologic features and excision repair cross-complementing group-1 (ERCC1) in resected p-stage III/N2 NSCLC patients that received cisplatin-based adjuvant chemotherapy.MethodsClinical data concerning 115 patients with histopathologically confirmed stage III/N2 NSCLC who underwent a complete resection were reviewed retrospectively. All patients received cisplatin-based adjuvant chemotherapy. The protein expression levels for ERCC1 were immunohistochemically examined in 115 patients. The relationship between the ERCC1 protein expression level and the clinical outcomes of the patients was then observed.ResultsThe 5-year survival rate and median survival time of patients with pathological stage III/N2 NSCLC after surgery and postoperative chemotherapy was 27.0% and 28.0 months, respectively. Survival of patients with ERCC1 negative tumors was significantly longer than those with ERCC1 positive tumors (p = 0.004). However, it was not entirely clear whether adjuvant chemotherapy with cisplatin-based agents was beneficial for ERCC1-negative patients with p-stage III/N2. A multivariate analysis of survival in patients with stage III/N2 NSCLC showed that surgical procedure (pneumonectomy vs. lobectomy; p = 0.001), number of involved lymph nodes (≤5 vs. >5; p = 0.001) and ERCC1 protein expression (negative vs. positive; p = 0.012) were significant prognostic factors. In addition, the prognosis of patients with skip mediastinal lymph node metastasis showed a tendency for improved survival, but this was no significant (p = 0.432).ConclusionsFindings from this retrospective study suggested that the number of involved lymph nodes and the type of pulmonary resection are significant and independent prognosis factors in patients with p-stage III/N2 NSCLC. In addition, it was found that ERCC1 protein expression might play an important role in the prognosis of p-stage III/N2 NSCLC patients treated with cisplatin-based adjuvant chemotherapy.