A Study On Molecular Biomarkers Associated With Bladder Cancer And Breast Cancer

This PhD degree project is composed of two parts of study, on molecular markers for bladder cancer and breast cancer.Part I:Study on DNA copy number aberrations in the genome as molecular biomarkers of bladder urothelial carcinomaAbnormal change of the DNA structure is common in the carcinogenesis and malignant progression of bladder. These molecular genetic aberrations provide an insight for the molecular mechanism on development of the bladder cancer; and also benefit for discovery and identification of biomarkers for the disease. The purpose of this study was to identify the characteristic DNA copy number aberrations （CNA） in the genome that could serve as molecular biomarkers of bladder urothelial carcinoma.We previously generated a genome-wide DNA copy number aberration profile in bladder cancer by using the array comparative genome hybridization （array CGH） approach. In this study, we selected11DNA fragments/genes （A₁4_P139169, CEP63, CLK1, FOSL2, GHR, LILRA3, PAQR6, PARVA, SFRS8, ZFAND3and ZNF76） with high frequencies of CNA in the genome of bladder cancer, by re-mining the array-CGH data. The copy number aberration of the11genes was firstly examined by real time polymerase chain reaction in56tumor samples that were used for the array CGH analysis. Among the11genes,5（CEP63, FOSL2, GHR, PAQR6and ZFAND3） were verified, for further validation. Three independent sample sets of bladder cancer were applied for the validation; they were58freshly frozen tumor tissues derived from patients of bladder cancer with treated partial cystectomy,62formalin-fixed paraffin-embedded （FFPE） tumor tissues, and43freshly frozen tumor tissues derived from patients treated with transurethral resection of bladder tumor （TURBT）. Comparing with30peripheral leukocyte genomic DNA samples of healthy individuals, gain of copy number of both CEP63and PAQR6was detected in all three sample sets of bladder cancer tissues; whereas for FOSL2and GHR, gain of copy number was only observed in the ’FFPE’ samples. While loss of copy number for ZFAND3was found in the ’FFPE’ samples and ’TURBT’ samples. It is noteworthy that, the copy number aberrations of all5genes （CEP63, FOSL2, GHR, PAQR6and ZFAND3） were identified by real-time PCR in the ’FFPE’ samples consisting with that of array CGH analysis. Based on data derived from the tumor tissues, we examined5genes in urine sediment samples of123patients with bladder cancer, and found gain of copy number of PAQR6and FOSL2in the bladder cancer group, comparing with patients with tumor of head and neck.In conclusion, copy number aberration of the5candidate genes could be common molecular events, and can be detected in urine sediments of bladder cancer patients. However, further multicentre study is needed for developing them as diagnostic biomarkers for bladder cancer. Part Ⅱ:Study on molecular biomarkers of histopathological response to neoadjuvant chemotherapy for breast cancerNeoadjuvant chemotherapy （NAC） has become an integral part in the comprehensive treatment for breast cancer. Those patients achieve complete pathological response or significant pathological response have a longer disease-free survival and overall survival. To properly estimate pathological response to neoadjuvant chemotherapy is important for choosing therapeutic schedule. The purpose of this study was to identify appropriate molecular biomarkers of histopathological response to neoadjuvant chemotherapy for breast cancer.A panel of protein markers composed of ER, PR, HER2, CK5/6, EGFR, cyclin B1,14-3-3σ and14-3-3ζ was examined by immunohistochemical （IHC） staining, on core needle biopsy tissue samples collected from215patients with invasive breast cancer, prior to neoadjuvant chemotherapy. With Miller&Payne （MP） criterion, the histopathologic response to NAC was classified upon the tissue section of surgical resected tumor after neoadjuvant chemotherapy. The results of the IHC analysis showed that positive staining rates were, ER42.8%, PR42.8%, HER234.0%, CK5/627.9%, EGFR55.8%, cyclin B147.9%,14-3σ28.8%and14-3-3ζ79.5%. Among them, HER2mainly expressed in the postmenopausal group; ER and PR mainly expressed in the tumors with low grade, whereas CK5/6mainly expressed in the tumors with high grade. Between the groups of MP good response and poor response, there were significantly differential expression of ER （P<0.001）, PR （P=0.001）, CK5/6（P<0.001）, EGFR （P=0.024）, and14-3-3σ （P=0.017）. The logistic regression analysis showed that only ER and14-3-3σ were included in the prediction model for histopathological response to NAC. Joint detection of ER and14-3-3σ have more power to estimate the pathological response than that any single protein marker did.