Assessment of Multidrug Resistance Genes & Apoptotic Genes by Multiplex PCR and Estimation of Suppresser and Apoptotic Proteins by Immunohistochemistry and Electrochemical Biosensor in Breast Cancer Patients

number: 
2476
English
Degree: 
Imprint: 
Medicine
Author: 
Dr.Mohammed Naji Karim AL-Ajili
Supervisor: 
Dr. Nidhal Abdul Mohymen Mohammed
year: 
2010

Abstract:

Breast cancer (BC) is the most common cause of cancer death among women worldwide. The current rates of increase remain constant; a woman born today has a 1 in 10 chance of developing breast cancer. The response of breast cancer patients to the course of chemotherapy, hormonal therapy and radiotherapy is depending on presence or absence of one or many multidrug resistant genes in their genome. This prospective study was carried out to investigate the possible association of Multidrug resistant genes with resistant to the chemotherapy in BC patients, and detect the apoptotic genes and their correlation with BC
patients and evaluate the efficacy of usage of Electrochemical Biosensor technique "for the first time in the country" to detect the amount of Bcl-2 protein in BC tissues, also we studed the expression of some apoptotic related proteins in BC tissues, and compared all that with disease activity. A total of 50 female patients with BC were studied, their ages range from 21-65 year with a mean age of 48 ± 10.36 years. Two control groups were included in this study for comparison. The first group included (10) patients with benign breast lesions (8 Fibroadenoma and 2 Fibrocystic disease) and (6) apparently healthy individuals. Breast tissue samples were collected from patients and controls, RNA was extracted, and then Reverse transcriptase polymerase chain reaction was performed then applied Multiplex polymerase chain reaction
(MPCR) using a specific sequence primers for multidrug resistant genes (MDR) and apoptotic genes to observe the possible association of some MDR genes (human ApoD, CRYAB, CYR61, G1P2, G1P3, IFITM1
IFITM3, IL1-beta, PDE3A, PEPP2, PLAB and UBB genes) and some Apoptotic genes(human Bcl-2, Bcl-xL, Bax, ICE genes) with BC. Moreover, Immunohistochemistry test was carried out for the estimation of the expression of p53, p27 and P-gp proteins on BC tissues and we used a new development technique called "Electrochemical Biosensor" to estimate the levels of Bcl-2 protein from BC tissues extracted protein and compared the results with that obtained using Bcl-2 enzymelinked immune sorbent assay (ELISA). Results showed a significant differences between the following MDR genes (IFITM1 & PEPP2), and high significant difference of CRYAP, UBB and IFITM3 genes of BC patients as compared with healthy and control patients suggesting that these genes may play an important role in chemotherapy resistant in BC patients.In addition, there was highly significant correlation coefficient in the IFITM1 & PEPP2 genes with the stage and grade of BC patients, and a highly significant correlation coefficient between IFITM1 gene and PEPP2 gene, between PDE3A gene and CYR61 gene, between CRYAP gene with UBB gene, between CRYAP gene and IFITM3 gene, UBB gene and IFITM3 gene were noticed. Also there were correlation coefficients between PEPP2 gene and PDE3A gene, and between PEPP2 gene and CYR61gene. Another interesting finding showed a highly significant of the variation of ICE, Bcl-2 and Bax genes in patients when compared with healthy control. And highly significant correlation coefficient between Bcl- 2 gene with Bcl-xL gene. Furthermore, there were highly significant differences in the expression of p53, p27 and P-gp among the three studed groups. In addition, both Electrochemical Biosensor and ELISA showed a highly significant frequency of Bcl-2 between malignant patients with other two groups. Added to that, there were a highly significant correlation coefficient in the Bcl-2, Bcl-xL genes and P-gp with the tumour stage and grade.
And a significant correlation coefficient in p53 protein expression with tumour stage of breast cancer.