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## Introduction
**nf-core/oncoanalyser** is a Nextflow pipeline for the comprehensive analysis of cancer DNA and RNA sequencing data
using the [WiGiTS](https://github.com/hartwigmedical/hmftools) toolkit from the Hartwig Medical Foundation. The pipeline
supports a wide range of experimental setups:
- FASTQ, BAM, and / or CRAM input files
- WGS (whole genome sequencing), WTS (whole transcriptome sequencing), and targeted / panel sequencing1
- Paired tumor / normal and tumor-only samples, and support for donor samples for further normal subtraction
- Purity estimate for longitudinal samples using genomic features of the primary sample from the same patient2
- UMI (unique molecular identifier) processing supported for DNA sequencing data
- Most GRCh37 and GRCh38 reference genome builds
1 built-in support for the [TSO500
panel](https://www.illumina.com/products/by-type/clinical-research-products/trusight-oncology-500.html) with other
panels and exomes requiring [creation of custom panel reference
data](https://nf-co.re/oncoanalyser/usage#custom-panels) 2 for example a primary WGS tissue biospy and longitudinal low-pass WGS ccfDNA sample taken from the
same patient
## Pipeline overview
The pipeline mainly uses tools from [WiGiTS](https://github.com/hartwigmedical/hmftools), as well as some other external
tools. There are [several workflows available](https://nf-co.re/oncoanalyser/usage#introduction) in `oncoanalyser` and
the tool information below primarily relates to the `wgts` and `targeted` analysis modes.
> [!NOTE]
> Due to the limitations of panel data, certain tools (indicated with `*` below) do not run in `targeted` mode.
- Read alignment: [BWA-MEM2](https://github.com/bwa-mem2/bwa-mem2) (DNA), [STAR](https://github.com/alexdobin/STAR) (RNA)
- Read post-processing: [REDUX](https://github.com/hartwigmedical/hmftools/tree/master/redux) (DNA), [Picard MarkDuplicates](https://gatk.broadinstitute.org/hc/en-us/articles/360037052812-MarkDuplicates-Picard) (RNA)
- SNV, MNV, INDEL calling: [SAGE](https://github.com/hartwigmedical/hmftools/tree/master/sage), [PAVE](https://github.com/hartwigmedical/hmftools/tree/master/pave)
- SV calling: [ESVEE](https://github.com/hartwigmedical/hmftools/tree/master/esvee)
- CNV calling: [AMBER](https://github.com/hartwigmedical/hmftools/tree/master/amber), [COBALT](https://github.com/hartwigmedical/hmftools/tree/master/cobalt), [PURPLE](https://github.com/hartwigmedical/hmftools/tree/master/purple)
- SV and driver event interpretation: [LINX](https://github.com/hartwigmedical/hmftools/tree/master/linx)
- RNA transcript analysis: [ISOFOX](https://github.com/hartwigmedical/hmftools/tree/master/isofox)
- Oncoviral detection: [VIRUSbreakend](https://github.com/PapenfussLab/gridss)\*, [VirusInterpreter](https://github.com/hartwigmedical/hmftools/tree/master/virus-interpreter)\*
- Telomere characterisation: [TEAL](https://github.com/hartwigmedical/hmftools/tree/master/teal)\*
- Immune analysis: [LILAC](https://github.com/hartwigmedical/hmftools/tree/master/lilac), [CIDER](https://github.com/hartwigmedical/hmftools/tree/master/cider), [NEO](https://github.com/hartwigmedical/hmftools/tree/master/neo)\*
- Mutational signature fitting: [SIGS](https://github.com/hartwigmedical/hmftools/tree/master/sigs)\*
- HRD prediction: [CHORD](https://github.com/hartwigmedical/hmftools/tree/master/chord)\*
- Tissue of origin prediction: [CUPPA](https://github.com/hartwigmedical/hmftools/tree/master/cuppa)\*
- Pharmacogenomics: [PEACH](https://github.com/hartwigmedical/hmftools/tree/master/peach)
- Summary report: [ORANGE](https://github.com/hartwigmedical/hmftools/tree/master/orange), [linxreport](https://github.com/umccr/linxreport)
For the `purity_estimate` mode, several of the above tools are run with adjusted configuration in addition to the following.
- Tumor fraction estimation: [WISP](https://github.com/hartwigmedical/hmftools/tree/master/wisp)
## Usage
> [!NOTE]
> If you are new to Nextflow and nf-core, please refer to [this page](https://nf-co.re/docs/usage/installation) on how to set-up Nextflow. Make sure to [test your setup](https://nf-co.re/docs/usage/introduction#how-to-run-a-pipeline) with `-profile test` before running the workflow on actual data.
Create a samplesheet with your inputs (WGS/WTS BAMs in this example):
```csv
group_id,subject_id,sample_id,sample_type,sequence_type,filetype,filepath
PATIENT1_WGTS,PATIENT1,PATIENT1-N,normal,dna,bam,/path/to/PATIENT1-N.dna.bam
PATIENT1_WGTS,PATIENT1,PATIENT1-T,tumor,dna,bam,/path/to/PATIENT1-T.dna.bam
PATIENT1_WGTS,PATIENT1,PATIENT1-T-RNA,tumor,rna,bam,/path/to/PATIENT1-T.rna.bam
```
Launch `oncoanalyser`:
```bash
nextflow run nf-core/oncoanalyser \
-profile \
-revision 2.2.0 \
--mode \
--genome \
--input samplesheet.csv \
--outdir output/
```
> [!WARNING]
> Please provide pipeline parameters via the CLI or Nextflow `-params-file` option. Custom config files including those provided by the `-c` Nextflow option can be used to provide any configuration _**except for parameters**_; see [docs](https://nf-co.re/docs/usage/getting_started/configuration#custom-configuration-files).
For more details and further functionality, please refer to the [usage documentation](https://nf-co.re/oncoanalyser/usage) and the [parameter documentation](https://nf-co.re/oncoanalyser/parameters).
## Pipeline output
To see the results of an example test run with a full size dataset refer to the [results](https://nf-co.re/oncoanalyser/results) tab on the nf-core website pipeline page.
For more details about the output files and reports, please refer to the
[output documentation](https://nf-co.re/oncoanalyser/output).
## Version information
### Extended support
As `oncoanalyser` is used in clinical settings and subject to accreditation standards in some instances, there is a need
for long-term stability and reliability for feature releases in order to meet operational requirements. This is
accomplished through long-term support of several nominated feature releases, which all receive bug fixes and security
fixes during the period of extended support.
Each release that is given extended support is allocated a separate long-lived git branch with the 'stable' prefix, e.g.
`stable/1.2.x`, `stable/1.5.x`. Feature development otherwise occurs on the `dev` branch with stable releases pushed to
`master`.
Versions nominated to have current long-term support:
- TBD
## Known issues
Please refer to [this page](https://github.com/nf-core/oncoanalyser/issues/177) for details regarding any known issues.
## Credits
The `oncoanalyser` pipeline was written and is maintained by Stephen Watts ([@scwatts](https://github.com/scwatts)) from
the [Genomics Platform
Group](https://mdhs.unimelb.edu.au/centre-for-cancer-research/our-research/genomics-platform-group) at the [University
of Melbourne Centre for Cancer Research](https://mdhs.unimelb.edu.au/centre-for-cancer-research).
We thank the following organisations and people for their extensive assistance in the development of this pipeline,
listed in alphabetical order:
- [Hartwig Medical Foundation
Australia](https://www.hartwigmedicalfoundation.nl/en/partnerships/hartwig-medical-foundation-australia/)
- Oliver Hofmann
## Contributions and Support
If you would like to contribute to this pipeline, please see the [contributing guidelines](.github/CONTRIBUTING.md).
For further information or help, don't hesitate to get in touch on the [Slack `#oncoanalyser`
channel](https://nfcore.slack.com/channels/oncoanalyser) (you can join with [this invite](https://nf-co.re/join/slack)).
## Citations
You can cite the `oncoanalyser` Zenodo record for a specific version using the following DOI:
[10.5281/zenodo.15189386](https://doi.org/10.5281/zenodo.15189386)
An extensive list of references for the tools used by the pipeline can be found in the [`CITATIONS.md`](CITATIONS.md)
file.
You can cite the `nf-core` publication as follows:
> **The nf-core framework for community-curated bioinformatics pipelines.**
>
> Philip Ewels, Alexander Peltzer, Sven Fillinger, Harshil Patel, Johannes Alneberg, Andreas Wilm, Maxime Ulysse Garcia,
> Paolo Di Tommaso & Sven Nahnsen.
>
> _Nat Biotechnol._ 2020 Feb 13. doi: [10.1038/s41587-020-0439-x](https://dx.doi.org/10.1038/s41587-020-0439-x).
