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Title: Pathobiology of oestrogen receptor negative invasive breast cancer
Author: Benhasouna, Ahmed Abdulsalam Mohammed
Awarding Body: University of Nottingham
Current Institution: University of Nottingham
Date of Award: 2012
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Breast cancer, which is the most common cancer in women and second leading cause of cancer-related death worldwide, is a heterogeneous disease. Breast cancer patients' risk stratification and prediction of usefulness of different therapeutic protocols currently depend on patient/tumour clinico-pathological criteria in addition to few clinically validated biomarkers. However, substantial proportions of patients with apparently similar clinico-pathological features may show distinctive outcomes and variable responses to therapy. In addition, the continuous improvement in the field of systemic and targeted therapy and the increasing need for a personalised and tailored therapy have emphasised the importance of refinement of the current classification systems and identifying novel prognostic/predictive factors. It is now well-documented that oestrogen receptor (ER)-negative breast cancers are a group of clinically aggressive tumours with poor prognosis and fewer cancer prevention and treatment strategies compared with ER-positive tumours. The aims of this thesis were: 1) to investigate the biological and molecular features of the ER-negative tumours and 2) to assess the role of some important pathways involved in the pathogenesis and progression of ER-negative tumours to better understand their behaviour. Material and Methods: Protein expreSSlOn of a large panel of relevant markers was assessed immunohistochemically using a large well-characterised series of primary invasive breast cancer with long term follow-up. This series included a large number of ER-negative tumours (no=776) and tumours from patients with BRCA I-germline mutated tumours. Moreover, a novel molecular assay (High Resolution Melting (HRM) analysis) was used to screen mutations of TP53 gene in genomic DNA extracted from formalin fixed paraffin embedded (FFPE) and matched fresh frozen tissues. Results: This study usmg well-defined clinico-pathological prognostic variables and a large panel of relevant biomarkers has identified the apoptosis related marker BCL2 as an independent prognostic marker in the ER-negative VI j .1 ,:1 tumours together with lymph node stage, primary tumour size. In clinically aggressive triple-negative (ER-, PR- and HER2-negative) subgroup, only Bcl2 expression and lymph node stage were independent predictors of outcome. Interestingly, in the triple-negative/lymph node-negative tumours, BCL2 was the only predictor of outcome. Due to the importance of DNA repair defects in ER-negative tumours, the expression of different genes involved in DNA repair mechanisms were assessed. The results showed that unlike ER-positive breast cancer, BRCA 1- associated tumours and ER-negative sporadic tumours frequently show abnormal BRCAl protein expression, lack nuclear expression and positive cytoplasmic expression. The BRCAl transcriptional suppressor ID4 was associated withBRCAI protein expression in the triple-negative subgroup but not in the ER-positive or HER2 positive tumours. The majority of breast cancers showed expression of the single-stand DNA break repair (SSBR) marker PARPl including the non-cleaved (PARPlnc) and the cleaved (P ARP 1 c) forms. P ARP 1 nc expression was frequentl y expressed in premenopausal younger patients and in tumours showing higher tumour grade with nuclear pleomorphism and high Ki67 expression. P ARP 1 expression was positively associated with other DNA repair proteins including the doublestrand DNA breast repair proteins BRCAl and RAD5I, and SSBR proteins Ku70/80, CHK1, and PIASy and it was negatively associated with ID4. PARPlc but not PARPlnc showed an association with expression of ER. There was no significant association between P ARP 1 (cleaved and non-cleaved) and outcome in the whole series or in the ER-negative subgroup. Furthermore, characterisation of BR CA I-associated tumours indicated that these tumours are more likely to be of high grade, highly proliferating, steroid receptor negative, more often expressing p53 protein and of basal phenotype. The results demonstrated that triple-negative that express basal markers (basal cytokeratins and/or EGFR), which is defined as basal-like breast cancer (BLBC), and those that do not express such markers (non-basal phenotype) are molecularly and clinical distinct classes of ER-negative tumour despite the lack of morphological difference. Also, the expression of the calcium-dependent VII --~---------------- 7 cysteine protease calpastatin and calpain-l were frequently expressed in ERnegative tumours and their expression was significantly associated with clinicopathologic variables characteristic of aggressive behaviour and with an adverse breast cancer-specific survival. HRM analysis of the TP53 gene has identified mutations in its genomic DNA however; a significant difference was identified between TP53 mutations in ER-negative tissue samples from FFPE as compared to the gold standard fresh frozen tissues. Conclusions: This study provided further evidence for the biological and molecular heterogeneity of ER-negative breast cancer and identifies BCL2 as a strong prognostic marker in ER-negative and triple negative classes. Several molecular pathways are involved in the pathogenesis and progression of ERnegative tumours with DNA repair pathways play major roles. Immunohistochemical assessment of these cellular and molecular derangements is a cost-effective, reproducible, and clinically applicable. Therefore, targeting these molecular allies, through discovering specific newer promising therapies will aid in the prevention and/or treatment of ER-negative breast cancer. More studies are warranted to translate the histopathological and molecular investigation into clinical management of breast cancer.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available