Molecular characterization of breast cancer in patients aged 50 years and older with respect to prognostication, genomic instability, and tumor heterogeneity

Abstract

Despite the majority (70%) of female breast cancer cases being diagnosed in women aged 55 years and older and an increasing population of older breast cancer patients due to demographic change, older patients lack adequate representation in cancer research. Thus, this doctoral thesis focused on breast cancer patients aged 50 years and older, aiming to elucidate the role of intratumor heterogeneity, ploidy, and genomic instability and their influences on disease outcome.

Therefore, a breast cancer collective of 39 patients with short (median 2.4 years) and long survival (median 19 years) were selected. Multiplex interphase fluorescence in situ hybridization (miFISH) was carried out to analyze copy number alterations (CNAs) of eight breast cancer-associated genes for their potential as biomarkers, as well as assessing genomic instability and tumor heterogeneity. Furthermore, image cytometry was performed to detect ploidy and phylogenetic tree modeling to gain more information about tumor development. Supplementary, targeted next-generation sequencing of 563 breast cancer-associated genes was carried out externally, and the obtained mutation status was statistically analyzed, compared to the miFISH results, and interpreted as part of this thesis.

The experimental part of this work revealed several CNAs of breast cancer-specific genes, as well as gene mutations frequently reported in breast carcinomas. The copy number gain of COX2 occurred most frequently (in 72% of the cases), followed by MYC (69%), whereas losses were more common for CDH1 (74%) and TP53 (69%). Comparing aneuploid with diploid tumor samples, significantly higher average signal numbers, CNAs, and instability indices, reflecting the degree of genomic instability, were revealed in the aneuploid tumors. In 16 cases, the signal pattern indicated the formation of an isochromosome 8q and in 14 cases of an isochromosome 17q. Supporting the hypothesis of an isochromosome formation, CNAs of DBC2/MYC and HER2/TP53 significantly co-occurred. Moreover, significant co-occurrence of CNA of HER2/DBC2 was detected, and CNAs for HER2 and PIK3CA mutations and CNAs for CCND1 and PIK3CA mutations were significantly mutually exclusive.

Overall, the distribution of gene mutations of the 563-breast cancer-associated genes, as well as the pattern of CNAs in the eight breast cancer-related genes (miFISH) detected in the 39 patients aged 50 years and older, were comparable to results of the age-unbiased TCGA-cohort. Notably, neither the quantity of CNAs, the tumor ploidy, nor the degree of intratumor heterogeneity revealed an association with the survival time, indicating that for patients above the age of 50, these criteria do not seem to have a substantial effect on disease prognosis.