Somatic Mode Options

Inputs a pair of FASTQ files into the mapper aligner and somatic variant caller. You can use these options with OTHER FASTQ options to run in tumor-normal mode. For example:

dragen -f -r /staging/human/reference/hg19/hg19.fa.k_21.f_16.m_149 \
--tumor-fastq1 <TUMOR_FASTQ1> \
--tumor-fastq2 <TUMOR_FASTQ2> \
--RGID-tumor <RG0-tumor> -–RGSM-tumor <SM0-tumor> \
-1 <NORMAL_FASTQ1> \
-2 <NORMAL_FASTQ2> \
--RGID <RG0> –RGSM <SM0> \
--enable-variant-caller true \
--output-directory /staging/examples/ \
--output-file-prefix SRA056922_30x_e10_50M

Inputs a list of FASTQ files into the mapper aligner and somatic variant caller. You can use these options with other FASTQ options to run in tumor-normal mode. For example:

dragen -f \
-r /staging/human/reference/hg19/hg19.fa.k_21.f_16.m_149 \
‑‑tumor-fastq-list <TUMOR_FASTQ_LIST> \
--fastq-list <NORMAL_FASTQ_LIST> \
--enable-variant-caller true \
--output-directory /staging/examples/ \
--output-file-prefix SRA056922_30x_e10_50M

Inputs a mapped BAM or CRAM file into the somatic variant caller. You can use these options with other BAM/CRAM options to run in tumor-normal mode.

The --vc-target-vaf is used to select the variant allele frequencies of interest. The variant caller will aim to detect variants with allele frequencies larger than this setting. We recommend adding a small safety factor, e.g. to ensure variants in the ballpark of 1% are detected, the minimum vc-target-vaf can be specified as 0.009 (0.9%). This setting will not apply a hard threshold, and it is possible to detect variants with allele frequencies lower than the selected threshold. On high coverage and clean datasets, a lower target-vaf may help increase sensitivity. On noisy samples (like FFPE) a higher target-vaf (like 0.03) maybe help reduce false positives. Using a low target-vaf may also increase runtime. Set the vc-target-vaf to 0 to disable this feature. When this feature is disabled the variant caller will require at least 2 supporting reads to discover a candidate variant. Valid range=[0, 1], default=0.01.

The minimum BQ that will be used at any stage in the SNV caller.

Specifies the minimum read quality (MAPQ) to be considered for variant calling. The default value is 3 for tumor-normal analysis or 20 for tumor-only analysis.

Specifies the callability threshold for tumor samples. The somatic callable regions report includes all regions with tumor coverage above the tumor threshold. The default value is 15. For more information on the somatic callable regions report, see Somatic Callable Regions Report

Specifies the callability threshold for normal samples, if present. If applicable, the somatic callable regions report includes all regions with normal coverage above the normal threshold. The default value is 5. For more information on the somatic callable regions report, see Somatic Callable Regions Report.

Use --vc-somatic-hotspots to specify a hotspots input VCF. The variants in the hotspots VCF indicate positions where more somatic mutations are expected. For example, a hotspots input VCF can be derived from COSMIC. DRAGEN genotyping priors are boosted for all positions specified in the VCF, so you can call a variant at one of these sites with fewer supporting reads. If you do not provide a value for vc-somatic-hotspots, DRAGEN automatically selects a reference-specific hotspots VCF file from /opt/edico/config/somatic_hotspots*. If you provide a vc-somatic-hotspots VCF, the VCF always takes precedence over the default hotspots VCF.

To disable the hotspots feature, use vc-use-somatic-hotspots=false. Neither the default VCF nor a provided VCF is considered.

If using --vc-use-somatic-hotspots, use vc-hotspotlog10-prior-boost to control the size of the adjustment. The default value is 4 (log10 scale) corresponding to an increase of 40 Phred.

In a tumor-normal analysis, DRAGEN accounts for tumor-in-normal (TiN) contamination by running liquid tumor mode. Liquid tumor mode is disabled by default. When liquid tumor mode is enabled, DRAGEN is able to call variants in the presence of TiN contamination up to a specified maximum tolerance level. --vc-enable-liquid-tumor-mode enables liquid tumor mode with a default maximum contamination TiN tolerance of 0.15. If using the default maximum contamination TiN tolerance, somatic variants are expected to be observed in the normal sample with allele frequencies up to 15% of the corresponding allele in the tumor sample. vc-tin-contam-tolerance enables liquid tumor mode and allows you to set the maximum contamination TiN tolerance.

Liquid tumor mode is not equivalent to liquid biopsy. Liquid tumors in liquid tumor mode refer to hematological cancers, such as leukemia. For liquid tumors, it is not feasible to use blood as a normal control because the tumor is present in the blood. Skin or saliva is typically used as the normal sample. However, skin and saliva samples can still contain blood cells, so that the matched normal control sample contains some traces of the tumor sample and somatic variants are observed at low frequencies in the normal sample. If the contamination is not accounted for, it can severely impact sensitivity by suppressing true somatic variants.

Liquid tumor mode typically uses a library that is WGS or WES with medium depth for example (100x T/ 40xN), and the lowest VAF detected for these types of depths is ~5%. Liquid biopsy typically uses a targeted gene panel (eg, 500 genes), with very high raw depth, and uses UMI indexing (collapsing down to a depth of >2000x) to enable sensitivity at VAF down to 0.1 % in some cases (the limit of detection will vary depending on coverage and data quality).

If using different sequencing systems or different library preparation methods for tumor and normal samples, DRAGEN recommends setting this option to false. In tumor-normal mode, DRAGEN estimates a set of PCR error parameters separately for each of the tumor and normal samples. By default, DRAGEN ignores the tumor-sample parameters and uses normal-sample parameters for analysis of both samples. This default prevents overestimation of tumor-sample error rates that can occur if the somatic variant rate is high. For example, the error rates can occur in germline admixture data sets where the somatic variant rate is equal to the germline variant rate.

These options control the Sample-specific NTD Error Bias Estimation feature.