Output from the Identify TMB Status workflow

The Identify TMB Status workflow produces a TMB report (Image proteinreport_16_n_p) that contains the TMB score (calculated as the number of mutations per Mb) and TMB confidence values (figure 13.40). The TMB confidence is based on the size of the target regions included in the TMB score calculation, i.e., those with a coverage at least 100X: TMB confidence is low if fewer than 900,000bp of target regions have sufficient coverage, high if more than 1,000,000 bp of target regions have been included in the calculation, and intermediate in between these 2 values.

Image tmbwfreport
Figure 13.40: An example of a TMB report.

Note that it is possible to configure the Calculate TMB Score tool to include TMB status information in the report (see Calculate TMB Score).

In addition, the report lists the adjusted length of the target regions (after removal of the regions whose coverage was below 100X). To estimate how the removal of region of low coverage impacted the original target regions, see Section 1.5 of the "QC for Targeted Sequencing - Coverage Report" output, which offers statistics on the numbers of targets for which all positions are covered by the "Minimum coverage" threshold set in the QC for targeted sequencing dialog (100 by default).

The quality filters statistics of the TMB report recapitulates how many variants are removed by the various filters applied by the tool, and the frequency distributions of input and somatic variants.

Here are two high TMB score reports based on different samples: the first sample (to the left) is a cell line cancer (pure) and the second (to the right) is from tissue (mixed with normal tissue). It can be seen from the distribution of the variants at the different frequencies that the pure sample contains somatic variants at higher frequency compared to the tissue sample where the low frequency variants contribute to the TMB score instead (figure 13.41).

Image frequenciestmb
Figure 13.41: Comparison of high TMB score reports based on different samples (pure cell cancer sample to the left versus mixed tissue sample to the right).

The workflow will also generate a Genome Browser View (Image trackset_16_n_p) as well as the following files:

The difference between the Unfiltered variant track and the Variants passing filters track depends on the following options available in the filtering steps:

The difference between the Variants passing filters track and the TMB Somatic Variant track is that even more stringent filters are applied to exclude variants before calculating the TMB score. For example, only variants with a frequency equal to or higher than 5% will be included in the TMB score. Germline variants, synonymous variants and variants outside of coding regions are also excluded.

The read mapping of the merged UMI groups will let you verify the found variants, and examine why expected variants were not found. The UMI Groups Report gives information about the number of UMI groups found, and how many reads are in each. It includes the following information: