592: Relationship between oxidative DNA damage and p53 mutation in colorectal adenoma and adenocarcinoma

EACR-23 Poster Sessions / European Journal of Cancer 50, Suppl. 5 (2014) S23–S242 591 S100P, a calcium-binding protein, is associated with polypoid tumour growth in colorectal carcinogenesis J. Chiang1 , J. Chen1 . 1 Chang Gung Memorial Hospital, Taipei, Taiwan Introduction: Colorectal cancer (CRC) is a genetically heterogeneous disease with distinct morphological patterns. It has been shown that polypoid and ulcerative CRC displays different genetic alterations. Material and Methods: We set out to investigate genes differentially expressed between ulcerative and polypoid CRC. cDNA microarray was performed to compare gene expression profiling of ulcerative and polypoid CRC with paired normal mucosa. Potential candidates were validated using quantitative real-time polymerase chain reaction, western blot and immunohistochemistry. Epigenetic regulation of gene expression was investigated using methylation-specific PCR (MSP). Results: cDNA microarray identified 11 up-regulated and 14 down-regulated genes differentially expressed in both tumor types compared to matched normal mucosa. Among them, S100P was the only upregulated gene preferentially associated with polypoid CRC (P = 0.032). Polypoid CRC displayed significantly higher S100P protein and mRNA expression level than ulcerative CRC (P < 0.05, respectively). Using semi-quantitative immunohistochemical analyses, strong S100P overexpression was preferentially associated with polypoid CRC (24/30 vs. 14/40, P < 0.001). The relative methylation level determined by MSP was not statistically different between polypoid and ulcerative CRC (43.36% vs. 49.10%, P = 0.168), indicating that promoter hypomethylation was directly related to upregulation of S100P mRNA. Conclusions: Our results show that upregulation of S100P mRNA and protein is a predominant feature in polypoid CRC, whereas ulcerative CRC shows a wide range of expression level, indicating that S100P overexpression is not a key determinant in conferring the proper of invasion. The clinicopathological significance of S100P in CRC requires further investigation in well-controlled studies. No conflict of interest. 592 Relationship between oxidative DNA damage and p53 mutation in colorectal adenoma and adenocarcinoma D.G. Priolli1 , J.R. Scalise1 , J.C. Valdivia2 , N.P. Martinez1 , D.Y. Kanno3 , ˜ Francisco, PostGraduate M.G. Marques4 , I.A. Cardinalli5 . 1 Universidade Sao ˜ Program in Health Science, Bragan¸ca Paulista, Brazil, 2 Universidade Sao Francisco, Medical Course, Bragan¸ca Paulista, Brazil, 3 Sao Francisco 4 Hospital, Surgery, Bragan¸ca Paulista, Brazil, Sao Francisco Hospital, Children Health, Bragan¸ca Paulista, Brazil, 5 Universidade Francisco Hospital, Pathology, Bragan¸ca Paulista, Brazil Introduction: Considering the different mechanisms of carcinogenesis, as well as several epidemiological studies in colorectal cancer, there are still doubts about higher or lower p53 protein mutations frequency. The main function of this protein is to preserve genome integrity of each individual cell, maintaining the same nucleotide sequence among all DNA strands in every cell of the body. For this reason p53 is known as the guardian of the genome. A possible mechanism involved in p53 mutation could be the oxidative DNA damage, resulting from production of reactive oxygen species and organic defense imbalance against these agents, admittedly involved in carcinogenesis. Objective: To relate the oxidative DNA damage to the p53 mutation in colorectal adenomatous lesions. Method: One hundred and fifty-three study subjects underwent colonoscopy, obtaining tumor and non-tumor specimens from patients with adenoma and adenocarcinoma. Normal tissue was obtained from volunteers undergoing colonoscopy for population screening without any morbidity. Conventional histopathological study (H&E) was performed for diagnostic purposes. Immunohistochemistry assay, associated with computer-assisted image analysis was performed to quantify p53. Oxidative DNA damage was obtained by comet assay. Statistical analysis adopted a 5% significance level to test the null hypothesis. Descriptive analysis, measures of central tendency, normality test, analysis of variance, correlation and partial association tests were performed. Results: The p53 expression was higher in non-tumor tissue of colon cancer patients when compared to normal tissue from healthy volunteers (p = 0.01). The p53 expression was higher in tumor tissue compared to non-tumor tissue samples (p = 0.00). Amounts of p53 in normal and tumor tissue are associated (r = 0.459, p = 0.00). Oxidative DNA damage was higher in tumor tissue when compared to non-tumor tissue samples (p = 0.00). There is an association between DNA damage in normal and tumor tissue (rs = 0.439, p = 0.00). The causal relationship between oxidative DNA damage and p53 mutation could be demonstrated by partial correlation (r = 0.393, p = 0.00). It was not possible to demonstrate p53 mutation as a factor promoting oxidative damage. Conclusion: From this study, through partial correlation, it was possible to demonstrate a causal relationship between oxidative DNA damage and p53 mutation and not the contrary. There is no control of DNA oxidative damage by p53. Thus it is reasonable to infer from literature findings and the present results that DNA oxidative damage results in genes mutation, including TP53

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and its corresponding p53 protein. Oxidative DNA damage is related to p53 mutation presenting as an intervening variable in colorectal adenoma and adenocarcinoma. No conflict of interest.

Sunday 6 July 2014 Poster Session

Translational Research I 593 High incidence of EGFR gene mutations in Serbian female lung adenocarcinoma patients M. Cavic1 , A. Krivokuca1 , K. Brotto1 , E. Malisic1 , S. Radulovic1 , R. Jankovic1 . 1 Institute for Oncology and Radiology of Serbia, Experimental Oncology, Belgrade, Serbia Background: In the last decade lung cancer has become a serious health issue in Serbia, with approximately 5200 new cases diagnosed and 4600 deaths occurring annually. In the same period, disease and mortality rates were two times higher in females than in males. The epidermal growth factor receptor (EGFR) gene mutations confer sensitivity to targeted therapy with tyrosine kinase inhibitors (TKI). EGFR mutation testing of Serbian nonsmall cell lung cancer patients became obligatory in 2011. Our laboratory as the referent one for this analysis in Serbia was certified by The European Molecular Genetics Quality Network. The aim of this study was to determine the frequency and types of EGFR mutations in Serbian lung cancer patients and explore potential correlations between the EGFR mutation status and patient and tumor characteristics. Material and Methods: The patient group consisted of 400 patients with primary lung cancer from various cancer centers in Serbia in the period from July 2011 to May 2012. The group comprised 245 males (61.2%) and 155 females (38.8%), all of Caucasian descent. Patient age range was 20−84 years, with a median of 60 years. The majority of the analyzed samples were adenocarcinoma (97%), while large-cell and planocellular carcinoma were less abundant (2.5% and 0.5% respectively). Detection of mutations was performed by real-time PCR. Chi-square, Fisher’s exact or Wilcoxon’s test were used for the analysis of associations between the EGFR mutation status and patient and tumor characteristics. Two-sided p values of less than 0.05 were considered as statistically significant. Results: Mutations were detected in 54 patients (13.5%), of which 50.8% were deletions in exon 19, 30.9% were L858R, 7.3% were G719X (G719S, G719A or G719C), 5.5% were L861Q, 5.5% were insertions in exon 20 and the S768I mutation was not detected. In the group of mutated samples, 63% mutations were detected in females and 37% in males, and the obtained difference was statistically significant (c2 = 15.42, p < 0.001). In the female group, the total mutation rate was 21.9%. The mutations were detected only patients with lung adenocarcinoma. Conclusions: Our results showed a positive correlation between the presence of EGFR mutations and the female gender and adenocarcinoma. An ongoing study including more patients will soon show whether this data might help further stratification of Serbian lung cancer patients most likely to benefit from targeted treatment with TKI. No conflict of interest. 595 The miR21/10b ratio as a prognostic marker in metastasis-free clear cell renal cell carcinoma patients H. Fritz1 , D. Lindgren2 , B. Ljungberg3 , H. Axelson2 , B. Dahlback ¨ 1 . 1 Lund ¨ Sweden, 2 Lund University, Molecular University, Laboratory Medicine, Malmo, Tumor Biology, Lund, Sweden, 3 Umea˚ University, Surgical and Perioperative ˚ Sweden Sciences, Umea, Introduction: Clear Cell Renal Cell Carcinoma (ccRCC) is the most common type of cancer in the adult kidney, and the prognosis of metastatic ccRCC remains poor with high mortality. In ccRCC, microRNAs (miRs) differentially expressed in tumor tissue have been identified and have been proposed to predict prognosis. The purpose of this study was to evaluate candidate miR markers identified from analysis of The Cancer Genome Atlas (TCGA) datasets in a large RCC cohort and to elucidate whether a ratio of miRs provided additional prognostic information. Materials and Methods: Deep sequencing data from TCGA datasets were analyzed using biostatistical methods to identify miRs that correlate with survival and stage of disease. Candidate miRs were analyzed by RT-qPCR in a cohort of 198 RCC tumor patients (ccRCC, n = 152) and 50 normal kidney samples. We analyzed the data using Kaplan–Meier analysis and Log-rank test for survival outcomes, and the Cox proportional hazard regression model for multivariate analysis. Results and Discussion: Four candidate miRs (miR-10b, miR-21, miR101, and miR-223) were selected from the TCGA analysis and analyzed in our cohort. Of these, miR-21 and miR-10b were differentially expressed in