ShallowHRDv2 bioinformatics pipeline
LGA, tumor, ±2
TUMOR
After DNA extraction and whole genome sequencing at low coverage (~1X), read counts profile (bin size ~50 kb) normalized and corrected for GC-content was obtained by ControlFreec [Main steps of CNA processing (Supplementary Fig. Three-way sample quality attribution: CNA profile classification according to tumor content (four categories), intrinsic sWGS noise (three categories) and FFPE noise (four categories) with final integrative classification in three categories: “good”, “fair” and “low” (Supplementary Fig. Noise reduction and optimization of breakpoints in the CNA profile: filtering small segments and assembling segments with small differences or local correlations to the FFPE noise profile (obtained from ~100 normal profiles from FFPE samples, Supplementary Fig. Broad CNA profile characterization by:genome complexity, where the “simple” genome has two most abundant copy number (CN) levels accounting for more than 70% of the genome, otherwise, the genome is classified as “complex” or “complex+” (significant and almost equal contribution of four CN levels);a set of binary attributes, such as a set of parameters characterizing the breakpoints, including the number of large genomic alterations (LGA), that largely contributes to the HRD diagnostics; LGA are defined as CN breaks between genomic segments of more than 9 Mb (segment sizes are rounded to the integer value); a set of ancillary LGA indexes, such as extra-large LGA, telomeric LGA, LGA between two the most abundant CN levels, LGA involving one of the three most abundant CN levels and the number of chromosome arms with LGA.Step-wise HRD diagnostics based on:LGA-score, which is essentially the number of LGA modified by PENALTY and/or BONUS, where PENALTY is defined by binary attributes and is subtracted from the LGA number (PENALTY is set to 0, 5 or 8 if none, one or more than one binary attributes hold true, respectively) and BONUS is defined by the genome complexity and is added to the LGA number (BONUS is set to 5 for “simple” genome and to 0 otherwise);the threshold of 20 with margins (±2) for clear-cut HRD diagnostics, namely 18 and 22, with the LGA-score <18 for “clear-cut” nonHRD and LGA-score >22 for “clear-cut” HRD;LGA-score modification, which is applied to resolve the borderline cases (18 ≤ LGA-score ≤ 22). Briefly, the LGA-score is shifted to 19 if evidence for nonHRD (such as PENALTY > 0, ancillary LGA indexes below the threshold, genome is classified as “complex+”, etc) and to 21 if evidence for HRD (PENALTY = 0, ancillary LGA indexes above the threshold, genome is classified as “simple”) (Supplementary Fig. 