Define LOH and LOH tract length.

<p>Case 1) LOH composed of 1 HET->HOM event. The minimum LOH tract length is 1 bp. Case 2) LOH composed of ≥2 HET->HOM events. The minimum LOH tract length is the distance between leftmost and rightmost informative SNPs.</p

Total LOH tract length (≥2 HET->HOM events) adjusted for chromosome arm length.

<p>a) Td, b) Tt of 31 IDC patients. Overlaid scattering points are the actually value for each patient colored by T stage of the patient’s Tt sample. The scale of the Y axis was adjusted to magnify LOH patterns. Scatter points outside of the Y axis top boundary were represented by points at the top of the boundary to show their existence. c) LOH tract length (≥2 HET->HOM events) of 548 IDC patients stratified by T stage. Bar height and error bar represent the median, first and third quartile values across patients.</p

Genome-Wide Analysis of Loss of Heterozygosity in Breast Infiltrating Ductal Carcinoma Distant Normal Tissue Highlights Arm Specific Enrichment and Expansion across Tumor Stages

<div><p>Studies have shown concurrent loss of heterozygosity (LOH) in breast infiltrating ductal carcinoma (IDC) and adjacent or distant normal tissue. However, the overall extent of LOH in normal tissue and their significance to tumorigenesis remain unknown, as existing studies are largely based on selected microsatellite markers. Here we present the first autosome-wide study of LOH in IDC and distant normal tissue using informative loci deduced from SNP array-based and sequencing-based techniques. We show a consistently high LOH concurrence rate in IDC (mean = 24%) and distant normal tissue (m = 54%), suggesting for most patients (31/33) histologically normal tissue contains genomic instability that can be a potential marker of increased IDC risk. Concurrent LOH is more frequent in fragile site related genes like WWOX (9/31), NTRK2 (10/31), and FHIT (7/31) than traditional genetic markers like BRCA1 (0/23), BRCA2 (2/29) and TP53 (1/13). Analysis at arm level shows distant normal tissue has low level but non-random enrichment of LOH (topped by 8p and 16q) significantly correlated with matched IDC (Pearson r = 0.66, p = 3.5E-6) (topped by 8p, 11q, 13q, 16q, 17p, and 17q). The arm-specific LOH enrichment was independently observed in tumor samples from 548 IDC patients when stratified by tumor size based T stages. Fine LOH structure from sequencing data indicates LOH in low order tissues non-randomly overlap (∼67%) with LOH that usually has longer tract length (the length of genomic region affected by LOH) in high order tissues. The consistent observations from multiple datasets suggest progressive LOH in the development of IDC potentially through arm-specific pile up effect with discernible signature in normal tissue. Our finding also suggests that LOH detected in IDC by comparing to paired adjacent or distant normal tissue are more likely underestimated.</p></div

Gene with top concurrent LOH in IDC and distant normal tissue.

a<p>LOH supported by ≥2 HET->HOM events in both Td and Tt in same gene region but do not require genomic overlap.</p>b<p>LOH supported by ≥2 HET->HOM events in both Td and Tt and have genomic overlap of at least 1 bp in same gene region.</p>c<p>Number of patients that have at least 1 heterozygote SNP in the corresponding gene region.</p

Numbers of concurrent and independent LOH (≥1 HET->HOM event(s)) for three patients with deep sequencing data.

<p>See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0095783#pone-0095783-t003" target="_blank">Table 3</a> for results using stringent criterion (≥2 HET->HOM events).</p

Count and length of concurrent LOH from patient A7-A0CE (#1).

<p>a) Count of concurrent LOH. Exa for longer LOH tract length in Td; Exb for longer LOH tract length in Tt; Pa for partial overlap. b) LOH tract length of different categories of LOH. Exa for extension length in Td; Exb for extension length in Tb; Equal for length of equal LOH; Pa-Ov for length of the overlapping part of the partial overlap; Pa-NOv for length of the non-overlapping part of the partial overlap.</p

Overlap pattern of concurrent LOH.

a<p>Extension length of Exa-d + Exa-s. Number in parentheses is the percentage of total LOH tract length.</p>b<p>Extension length of Exb-d + Exb-s. Number in parentheses is the percentage of total LOH tract length.</p>c<p>(Total - Equal)/Total.</p>d<p>Use LOH supported by ≥1 | ≥2 HET->HOM events.</p

Basic components of the iterative workflow as compared to a standard NGS whole genome analysis.

<p>Basic components of the iterative workflow as compared to a standard NGS whole genome analysis.</p

Evaluation of SNVs and Indels called by the iterative and standard workflow.

<p>Evaluation of SNVs and Indels called by the iterative and standard workflow.</p

Concordance of SNP data with variants from standard and iterative workflows for sample NA12878.

<p>Concordance of SNP data with variants from standard and iterative workflows for sample NA12878.</p