Study of GSK-3β at proteinic and molecular level in familial and sporadic breast carcinomas

INTRODUCTION. Breast cancer represents the most common malignancy and cause of cancer -related deaths in female population Worldwide .Although we observe an increase of new-diagnosed cases the death percent is diminuished. This fact is due to new and extensive models of screening ,the new generation of mammography and new therapeutic approaches.This statistic data impones as to expand our researching horizons towards promising fields of reliable diagnostic, therapeutic and prognostic factors. AIM.The aim of this study was the investigation of: Ο σκοπός της παρούσης μελέτης είναι: 1) The expression pattern of protein kinase GSK-3β in primary invasive breast carcinomas(sporadic and familial) , 2) The existence of mutations in the exons 5 and 6(chr. 3q13.3)of the gene GSK-3β . The results will be correlated with the : α) classical clinicopathological parameters(age,menopausal status,tumor size,histological type,nuclear grade of malignancy, stage of disease,hormone receptors) b) overall survival(OS) and progression free survival(PFS) c) biological markers (ER, PR ,β-catenin, AKT, m-TOR). MATERIALS AND METHODSA)The material is consisted of 266 females with primary invasive breast cancer: a)154 sporadic cases, b)112 familial and c) 20 samples of mammary tissue negative to malignancy. The definition of the immunohistochemical expression and of the mutations of the gene GSK-3β will be held in slides of paraffin embedded tissue. B) We applied the three step imunohistochemical method avidin-biotin-hyperoxydase complex. The antibody we used is the rabbit polyclonal GSK-3β (H-76), in dilution 1:70. C) For the study of possible mutations of the gene GSK-3β,exons 5 and6 we used the method of QIAamp DNA Mini Kit/ PCR, with the respective primers,and electrophoresis in agar gel . The fact of bibliographic absence of this kind of mutations forced us to perform trials of acceptable credibility of the method(temperature,time and number of reaction cycles) and then followed the sequencing of PCR in the cases of our study. D) The results and the individual correlations were valuated with statistic methods ,Chi-square test for the biological markers, individual(log rank test) and multivariate( Cox”s hazard regression model) for the valutation of the survival(OS and PFS) . As level of for all the analysis is considered the p