Genomic Analyses of Breast Cancer Progression Reveal Distinct Routes of Metastasis Emergence

A main controversy in cancer research is whether metastatic abilities are present in the most advanced clone of the primary tumor or result from independently acquired aberrations in early disseminated cancer cells as suggested by the linear and the parallel progression models, respectively. The genetic concordance between different steps of malignant progression is mostly unexplored as very few studies have included cancer samples separated by both space and time. We applied whole exome sequencing and targeted deep sequencing to 26 successive samples from six patients with metastatic estrogen receptor (ER)-positive breast cancer. Our data provide support for both linear and parallel progression towards metastasis. We report for the first time evidence of metastasis-to-metastasis seeding in breast cancer. Our results point to three distinct routes of metastasis emergence. This may have profound clinical implications and provides substantial novel molecular insights into the timing and mutational evolution of breast cancer metastasis.


Supplementary
. Venn diagrams of the mutational concordance of non-synonymous, splicing and synonymous somatic mutations between the different steps of malignant progression of the six studied patients. Figure S9. BubbleTree analysis graph and copy number plot of P8 synchronous axillary lymph node metastasis. The BubbleTree analysis is based on exome sequencing data. In the upper panel, the R-HDS plot, the R score indicates the copy number ratio between the tumor and matched normal sample, and HDS is the heterozygous-deviation score. The tree branches represent the integer allelespecific copy numbers. The lower panel depicts the copy number events. The tumor cell content of each sample is estimated, indicated by the first figure in the purity estimate. The following figures represent the frequency of major subclones.

metastasis.
The BubbleTree analysis is based on exome sequencing data. In the upper panel, the R-HDS plot, the R score indicates the copy number ratio between the tumor and matched normal sample, and HDS is the heterozygous-deviation score. The tree branches represent the integer allele-specific copy numbers.
The lower panel depicts the copy number events. The tumor cell content of each sample is estimated, indicated by the first figure in the purity estimate. The following figures represent the frequency of major subclones.
lymph node metastasis. The BubbleTree analysis is based on exome sequencing data. In the upper panel, the R-HDS plot, the R score indicates the copy number ratio between the tumor and matched normal sample, and HDS is the heterozygous-deviation score. The tree branches represent the integer allelespecific copy numbers. The lower panel depicts the copy number events. The tumor cell content of each sample is estimated, indicated by the first figure in the purity estimate. The following figures represent the frequency of major subclones. metastasis. The Bubble Tree analysis is based on exome sequencing data. In the upper panel, the R-HDS plot, the R score indicates the copy number ratio between the tumor and matched normal sample, and HDS is the heterozygous-deviation score. The tree branches represent the integer allele-specific copy numbers.
The lower panel depicts the copy number events. The tumor cell content of each sample is estimated, indicated by the first figure in the purity estimate. The following figures represent the frequency of major subclones.
metastasis. The BubbleTree analysis is based on exome sequencing data. In the upper panel, the R-HDS plot, the R score indicates the copy number ratio between the tumor and matched normal sample, and HDS is the heterozygous-deviation score. The tree branches represent the integer allele-specific copy numbers. synchronous axillary lymph node metastasis. The BubbleTree analysis is based on exome sequencing data. In the upper panel, the R-HDS plot, the R score indicates the copy number ratio between the tumor and matched normal sample, and HDS is the heterozygous-deviation score.
The tree branches represent the integer allele-specific copy numbers. The lower panel depicts the copy number events. The tumor cell content of each sample is estimated, indicated by the first figure in the purity estimate. The following figures represent the frequency of major subclones.
Supplementary Figure S23. Overview of copy number aberrations based on micro array analysis on the primary tumor (region 1), synchronous axillary lymph node metastasis and asynchronous distant metastasis of P4.