hla-neo-nf

Pipeline to predict neoantigens from WGS of T/N pairs

Usage

#using a tn_pairs file
nextflow run iarcbioinfo/hla-neo-nf -r v1.0  \
-profile singularity --ref chr6.mhc.fa \
--tn_file cohort_neoantigen.tsv --cram_dir cram \
--vcf_dir vcfs --vep_dir vep-db-111 --output_folder results_hla_neo

Dependencies

1. This pipeline is based on nextflow. As we have several nextflow pipelines, we have centralized the common information in the IARC-nf repository. Please read it carefully as it contains essential information for the installation, basic usage and configuration of nextflow and our pipelines.
2. External software:
   • xHLA
   • VEP
   • pvactools
     • vatools

You can avoid installing all the external software by only installing Docker or singularity. See the IARC-nf repository for more information.

Input (mandatory)

Type	Description
--tn_file	[file] File containing list of T/N bam/cram files to be processed
--cram_dir	[dir] directory where the BAM or CRAM file are stored
--vcf_dir	[dir] directory where the VCF files are stored
--vep_dir	[dir] directory containing VEP database for annotation [hg38, GENCODE 33]
--ref	[file] fasta file of chr6 of reference genome [chr6-hg38.fa], shold be indexed with BWA-mem

Example of cohort_neoantigen.tsv file (--tn_file)

A text file tabular separated, with the following header:

id      vcf     normal_cram     normal_id       tumor_id
sample1	sample1.vcf.gz	sample1_N.cram	NORMAL1	TUMOR1
sample2	sample2.vcf.gz	sample2_N.cram	sample2_N.cram	None	TUMOR2
sample3	sample3.vcf.gz	sample3_N.cram	sample3_N.cram	NORMAL3	TUMOR3_1,TUMOR3_2

Note that in case the vcf contains variants confidently called as somatic, the normal_id can be omitted (put "None" instead).

Optional parameters

Name	type	Description
--pvactools_predictors	[string]	predictions tools to compute neoantigens [def:all_class_i,all_class_ii or NetMHCpan,NetMHCIIpan]
--bam	[flag]	active bam mode [def:cram]
--output_folder	[string]	name of output folder
--cpu	[Integer]	Number of CPUs[def:20]
--mem	[Integer]	Max memory [def:8Gb]

Output

results
└── xHLA                        # HLA-typing predictions
│   ├── report-MESO_001-hla.json
│   ├── report-MESO_002-hla.json
│   ├── report-MESO_003-hla.json
│   ├── report-MESO_004-hla.json
│   ├── report-MESO_005-hla.json
│   ├── report-MESO_006-hla.json
│   ├── report-MESO_007-hla.json
│   ├── report-MESO_008-hla.json
│   ├── report-MESO_009-hla.json
├── VEP				# Annotation of variant impact
│   ├── MESO_001.vep.vcf
│   ├── MESO_002.vep.vcf
│   ├── MESO_003.vep.vcf
│   ├── MESO_004.vep.vcf
│   ├── MESO_005.vep.vcf
│   ├── MESO_006.vep.vcf
|
├── pVACTOOLS					#pvactools predictions
│   ├── MESO_002.pvactools.log			#pvactools log
│   ├── MESO_002_T1_pvactools				
│   │   ├── combined				 # neoatigens for class I and class II
│   │   │   ├── B00JALW.all_epitopes.aggregated.tsv    # raw predictions aggregated
│   │   │   ├── B00JALW.all_epitopes.tsv               # raw predictions 
│   │   │   ├── B00JALW.filtered.tsv	               # filtered predictions
│   │   │   └── B00JALW.aggregated.filtered.tsv	       # filtered aggregated
│   │   ├── MHC_Class_I				#neoatigens for class I
│   │   │   ├── B00JALW.all_epitopes.aggregated.tsv
│   │   │   ├── B00JALW.all_epitopes.tsv
│   │   │   ├── B00JALW.filtered.tsv
│   │   │   └── log
│   │   │       └── inputs.yml
│   │   └── MHC_Class_II			#neoatigens for class II
│   │       ├── B00JALW.all_epitopes.aggregated.tsv
│   │       ├── B00JALW.all_epitopes.tsv
│   │       ├── B00JALW.fasta
│   │       ├── B00JALW.filtered.tsv
│   │       └── log
|	         └── inputs.yml
├── nf-pipeline_info # Nextflow information directory
    ├── hla-neo_report.html
    ├── hla-neo_timeline.html
    ├── hla-neo_trace.txt
    └── run_parameters_report.txt # Custom file providing info for software versions and calling parameters

Common errors

vep-db

The first time is necesary to get a local copy of the vep database, you can achieve this by running the following command within the vep singularity container:

#get the singularity container
singularity pull docker://docker.io/iarcbioinfo/ensembl-vep:latest
#open a shell and run the folloing command
singularity shell ensembl-vep_latest.sif
#get a local copy of the vep cache database (gencode v33)
vep_install -a cf -s homo_sapiens -y GRCh38 -c vep-db-111 --CONVERT

chr6 BWA index

To perform the HLA-typing this pipeline extract reads from the CRAM/BAM file and remap them to the chr6 of GRCh38 using BWA-MEM (0.7.15-r1140), hence the chr6.fa of GRCh38 should be indexed (BWA index) to perform the mapping of the reads.

Singularity

The first time that the container is built from the docker image, the TMPDIR should be defined in a non parallel file-system, you can set this like:

export TMPDIR=/tmp

Running error

The mhccons module may occasionally crash and require a re-run (see griffithlab/pVACtools#325 ). If this happens too much, you can use increase hte maximum numbers of retries in the nextflow config file.

Contributions

Name	Email	Description
Matthieu Foll*	follm@iarc.who.int	Developer to contact for support
Alex Di Genova		Developer
Nicolas Alcala	alcalan@iarc.who.int	Developer