Germline DNA Copy Number Aberrations Identified as Potential Prognostic Factors for Breast Cancer Recurrence

Abstract

Breast cancer recurrence (BCR) is a common treatment outcome despite curative-intent primary treatment of non-metastatic breast cancer. Currently used prognostic and predictive factors utilize tumor-based markers, and are not optimal determinants of risk of BCR. Germline-based copy number aberrations (CNAs) have not been evaluated as determinants of predisposition to experience BCR. In this study, we accessed germline DNA from 369 female breast cancer subjects who received curative-intent primary treatment following diagnosis. Of these, 155 experienced BCR and 214 did not, after a median duration of follow up after breast cancer diagnosis of 6.35 years (range = 0.60–21.78) and 8.60 years (range = 3.08–13.57), respectively. Whole genome CNA genotyping was performed on the Affymetrix SNP array 6.0 platform. CNAs were identified using the SNP-Fast Adaptive States Segmentation Technique 2 algorithm implemented in Nexus Copy Number 6.0. Six samples were removed due to poor quality scores, leaving 363 samples for further analysis. We identified 18,561 CNAs with ≥1 kb as a predefined cut-off for observed aberrations. Univariate survival analyses (log-rank tests) identified seven CNAs (two copy number gains and five copy neutral-loss of heterozygosities, CN-LOHs) showing significant differences (P<2.01×10−5) in recurrence-free survival (RFS) probabilities with and without CNAs.We also observed three additional but distinct CN-LOHs showing significant differences in RFS probabilities (P<2.86×10−5) when analyses were restricted to stratified cases (luminal A, n = 208) only. After adjusting for tumor stage and grade in multivariate analyses (Cox proportional hazards models), all the CNAs remained strongly associated with the phenotype of BCR. Of these, we confirmed three CNAs at 17q11.2, 11q13.1 and 6q24.1 in representative samples using independent genotyping platforms. Our results suggest further investigations on the potential use of germline DNA variations as prognostic markers in cancer-associated phenotypes.

A Genome-Wide Association Study to Identify Potential Germline Copy Number Variants for Sporadic Breast Cancer Susceptibility

“…Germline DNA was extracted from peripheral blood samples of both cases and controls, using commercially available Qiagen TM (Mississauga, Ont., Canada) DNA isolation kits, as described earlier [Sehrawat et al, 2011;Sapkota et al, 2013a]. Whole genome genotyping was carried out using the Affymetrix Genome-Wide Human SNP Array 6.0, following the guidelines provided by the manufacturer.…”

“…Further, we have demonstrated that copy-neutral losses of heterozygosity are also an important class of structural variations and serve as biomarkers of prognostic value in a BC setting [Sapkota et al, 2013a]. Large CNVs have an appreciable impact on many human diseases, including non-small-cell lung cancer, HIV infection, developmental delay, schizophrenia, autism, and idiopathic learning disability [Glessner et al, , 2010Hollox and Hoh, 2014;Li et al, 2014].…”

“…Herein, we sought to investigate the role of germline CNVs in sporadic BC susceptibility using 422 invasive BC cases and 348 healthy individuals from Alberta, Canada, which were genotyped employing Affymetrix GenomeWide Human SNP Array 6.0 [Sapkota et al, 2013a]. Currently, there are no consensus guiding principles on the use of segmentation algorithms, nor ideal SNP or CNV arrays from any manufacturer to identify CNVs associated with disease susceptibility.…”

Germline DNA Variations in Breast Cancer Predisposition and Prognosis: A Systematic Review of the Literature

“…With the advent of SNP genotyping platforms that can measure both CNVs and CN-LOHs, it is now possible to investigate potential roles of these large chromosomal defects as genetic determinants for complex diseases using germline DNA. A recent study conducted by us that aimed to evaluate the role of germline CNVs and CN-LOHs in breast cancer recurrence identified multiple copy number aberrations (2 copy number gains and 5 CN-LOHs) showing statistically significant differences between recurrence-free survival probabilities with and without these aberrations [Sapkota et al, 2013a]. Of these, 3 CNLOHs were successfully validated by qPCR.…”

“…As an alternative source of genetic heritability for breast cancer, structural variations such as copy number variations (CNVs) and copy-neutral loss of heterozygosity (CN-LOH) are being interrogated for their associations with breast cancer predisposition as well as for disease recurrence [Wellcome Trust Case Control Consortium, 2010;Sapkota et al, 2013a]. GWAS literature has also witnessed few reports providing evidence for genetic interactions as another source to explain the remaining genetic risk of breast cancer [Milne et al, 2010;Campa et al, 2011;Nickels et al, 2013;Sapkota et al, 2013b].…”