StableLift

• Overview
• How To Run
• Pipeline Steps
  • 1. LiftOver Variant Coordinates
  • 2. Annotate Variants
  • 3. Predict Variant Stability
• Inputs
• Outputs
• Testing and Validation
  • Test Dataset
• References
• Discussions
• Contributors
• License

Overview

StableLift is a machine learning approach designed to predict variant stability across reference genome builds. It addresses challenges in cross-build variant comparison, supplementing LiftOver coordinate conversion with a quantitative "Stability Score" for each variant, indicating the probability of consistent representation across the two most commonly used human reference builds (GRCh37 and GRCh38).

StableLift is implemented as a Nextflow pipeline featuring:

• Robust LiftOver of SNVs, indels, and structural variants
• Variant annotation with external databases
• Variant filtering based on predicted cross-build stability

Pre-trained models are provided along with performance in a whole genome validation set to define the default F1-maximizing operating point and allow for custom filtering based on pre-calibrated specificity estimates.

Supported conversions:

• GRCh37->GRCh38
• GRCh38->GRCh37

Supported variant callers:

• HaplotypeCaller (gSNP)
• MuTect2 (sSNV)
• Strelka2 (sSNV)
• SomaticSniper (sSNV)
• MuSE2 (sSNV)
• DELLY2 (gSV, sSV)

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How To Run

1. Download and extract resource bundle and source code.
2. Download pre-trained model corresponding to variant caller and conversion direction.
3. Copy ./config/template.config (e.g. project.config) and fill in all required parameters.
4. Copy ./input/template.yaml (e.g. project.yaml) and update with input VCF ID and path.
5. Run the pipeline using Nextflow nextflow run -c project.config -params-file project.yaml main.nf.

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Pipeline Steps

1. LiftOver Variant Coordinates

• For SNVs, convert variant coordinates using the BCFtools LiftOver plugin with UCSC chain files.
• For SVs, convert variant breakpoint coordinates using custom R script with UCSC chain files and rtracklayer and GenomicRanges R packages.

2. Annotate Variants

• For SNVs, add dbSNP, GENCODE, and HGNC annotations using GATK's Funcotator. Add trinucleotide context and RepeatMasker intervals with bedtools.
• For SVs, annotate variants with population allele frequency from the gnomAD-SV v4 database.

3. Predict Variant Stability

• Predict variant stability with pre-trained random forest model and the ranger R package.
• Annotate VCF with Stability Score and filter unstable variants.

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Inputs

UCLA pipelines have a hierarchical configuration structure to reduce code repetition:

• config/default.config: Parameters with sensible defaults that may be overridden in project.config.
• config/template.config -> project.config: Required pipeline parameters. Often shared for many inputs.
• input/template.yaml -> project.yaml: Required input-specific parameters.

Input YAML

---
sample_id: ""  # Identifying string for the input VCF
input:
  vcf: ""      # Path to the input VCF file

Input Configuration

Required Parameter	Type	Description
output_dir	path	Path to the directory where the output files are to be saved.
variant_caller	string	Variant calling algorithm used to generate input VCF {HaplotypeCaller, Mutect2, Strelka2, SomaticSniper, Muse2, Delly2}.
rf_model	path	Path to corresponding pre-trained random forest model.
liftover_direction	string	Conversion direction {GRCh37ToGRCh38, GRCh38ToGRCh37}.
fasta_ref_37	path	Path to the GRCh37 reference sequence (FASTA).
fasta_ref_38	path	Path to the GRCh38 reference sequence (FASTA).
chain_file	path	Path to LiftOver chain file between the source and target genome builds (included in resource-bundle.zip).
funcotator_data_source	path	Path to Funcotator data source directory.
repeat_bed	path	Path to bundled RepeatMasker annotation file (included in resource-bundle.zip).
header_contigs	path	Path to header contigs file corresponding to target genome build (included in resource-bundle.zip).
gnomad_rds	path	Path to gnomAD SV data.table for annotation (included in resource-bundle.zip).

Optional Parameter	Type	Default	Description
target_threshold	numeric	""	Target Stability Score threshold for variant filtering: [0, 1].
target_specificity	numeric	""	Target specificity based on whole genome validation set for variant filtering: [0, 1].
extract_features_cpus	int	4	Number of cpus to use for parallel parsing of large VCFs (>1GB).
work_dir	path	/scratch/$SLURM_JOB_ID	Path of working directory for Nextflow. When included in the sample config file, Nextflow intermediate files and logs will be saved to this directory. With ucla_cds, the default is /scratch and should only be changed for testing/development. Changing this directory to /hot or /tmp can lead to high server latency and potential disk space limitations, respectively.
save_intermediate_files	boolean	false	If set, save output files from intermediate pipeline processes.
min_cpus	int	1	Minimum number of CPUs that can be assigned to each process.
max_cpus	int	SysHelper.getAvailCpus()	Maximum number of CPUs that can be assigned to each process.
min_memory	MemoryUnit	1.MB	Minimum amount of memory that can be assigned to each process.
max_memory	MemoryUnit	SysHelper.getAvailMemory()	Maximum amount of memory that can be assigned to each process.
dataset_id	string	""	Dataset ID to be used as output filename prefix.
blcds_registered_dataset	boolean	false	Set to true when using BLCDS folder structure; use false for now.
ucla_cds	boolean	true	If set, overwrite default memory and CPU values by UCLA cluster-specific configs.

---

Outputs

Output	Description
*_StableLift-${target_build}.vcf.gz	Output VCF in target build coordinates with variant annotations and predicted Stability Scores.
*_StableLift-${target_build}.vcf.gz.tbi	Output VCF tabix index.
*_StableLift-${target_build}_filtered.vcf.gz	Filtered output VCF with predicted "Unstable" variants removed.
*_StableLift-${target_build}_filtered.vcf.gz.tbi	Filtered output VCF tabix index.

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Testing and Validation

Test Dataset

10 whole genomes from The Cancer Genome Atlas (TCGA-SARC) were used to test pipeline outputs and validate model performance. All data was processed using standardized Nextflow pipelines. Somatic VCFs from GRCh37 and GRCh38 alignments are available for the four supported sSNV callers and DELLY2 sSV as release attachments.

Donor ID	Normal Sample ID	Tumour Sample ID
TCGASTSA000008	TCGASTSA000008-N001-B01-P	TCGASTSA000008-T001-P01-P
TCGASTSA000014	TCGASTSA000014-N001-B01-P	TCGASTSA000014-T001-P01-P
TCGASTSA000026	TCGASTSA000026-N002-A01-P	TCGASTSA000026-T001-P01-P
TCGASTSA000041	TCGASTSA000041-N001-B01-P	TCGASTSA000041-T001-P01-P
TCGASTSA000045	TCGASTSA000045-N001-B01-P	TCGASTSA000045-T001-P01-P
TCGASTSA000060	TCGASTSA000060-N001-B01-P	TCGASTSA000060-T001-P01-P
TCGASTSA000064	TCGASTSA000064-N001-B01-P	TCGASTSA000064-T001-P01-P
TCGASTSA000112	TCGASTSA000112-N001-B01-P	TCGASTSA000112-T001-P01-P
TCGASTSA000131	TCGASTSA000131-N001-B01-P	TCGASTSA000131-T001-P01-P
TCGASTSA000200	TCGASTSA000200-N001-B01-P	TCGASTSA000200-T001-P01-P

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References

1. StableLift: Optimized Germline and Somatic Variant Detection Across Genome Builds
2. Metapipeline-DNA: A Comprehensive Germline & Somatic Genomics Nextflow Pipeline
3. uclahs-cds/metapipeline-DNA

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Discussions

• Issue tracker to report errors and enhancement ideas.
• Discussions can take place in pipeline-StableLift Discussions
• pipeline-StableLift pull requests are also open for discussion

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Contributors

Please see list of Contributors at GitHub.

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License

pipeline-StableLift is licensed under the GNU General Public License version 2. See the file LICENSE for the terms of the GNU GPL license.

StableLift is a machine learning approach designed to predict variant stability across reference genome builds, supplementing LiftOver coordinate conversion to increase the portability of variant calls.

Copyright (C) 2024 University of California Los Angeles ("Boutros Lab") All rights reserved.

This program is free software; you can redistribute it and/or modify it under the terms of the GNU General Public License as published by the Free Software Foundation; either version 2 of the License, or (at your option) any later version.

This program is distributed in the hope that it will be useful, but WITHOUT ANY WARRANTY; without even the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public License for more details.