Somatic Mode Options

The --tumor-fastq1 and --tumor-fastq2 options are used to input a pair of FASTQ files into the mapper aligner and somatic variant caller. These options can be used 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

The --tumor-fastq-list option is used to input a list of FASTQ files into the mapper aligner and somatic variant caller. This option can be used 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

The --tumor-bam-input or --tumor-cram-input option is used to input a mapped BAM or CRAM file into the somatic variant caller. These options can be used with other BAM/CRAM options to run in tumor-normal mode.

The --vc-min-tumor-read-qual option 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.

The --vc-callability-tumor-thresh option specifies the callability threshold for tumor samples, and the --vc-callability-normal-thresh option specifies the callability threshold for normal samples, if present. The somatic callable regions report includes all regions with tumor coverage above the tumor threshold and if applicable, normal coverage above the normal threshold. For more information on the somatic callable regions report, see Somatic Callable Regions Report. The default value for the tumor-only threshold is 15 and the tumor-normal threshold is 5.

The somatic hotspots option allows an input VCF to specify the positions where the risk for somatic mutations are assumed to be significantly elevated. DRAGEN genotyping priors are boosted for all postions specified in the VCF, so it is possible to call a variant at one of these sites with fewer supporting reads. The cosmic database in VCF format can be used as one source of prior information to boost sensitivity for known somatic mutations. The size of the adjustment can be controlled via vc-hotspot-log10-prior-boost, which has a default value of 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. The maximum contamination TiN tolerance must be greater than zero. For example, vc-tin-contam-tolerance=-0.1.

Liquid tumors in liquid tumor mode refers 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.

In typical use cases of liquid tumor mode, the library is WGS or WES, with medium depth for example (100x T/ 40xN), and the lowest VAF detected for these types of depths is ~5%. In typical use cases of liquid biopsy, the library is a targeted gene panel (eg 500 genes), with very high raw depth (> 2000-5000x), and uses UMI indexing to enable sensitivity at VAF down to 0.5 % LoD.