Dataset Information

A critical spotlight on the paradigms of FFPE-DNA sequencing.

ABSTRACT: In the late 19th century, formalin fixation with paraffin-embedding (FFPE) of tissues was developed as a fixation and conservation method and is still used to this day in routine clinical and pathological practice. The implementation of state-of-the-art nucleic acid sequencing technologies has sparked much interest for using historical FFPE samples stored in biobanks as they hold promise in extracting new information from these valuable samples. However, formalin fixation chemically modifies DNA, which potentially leads to incorrect sequences or misinterpretations in downstream processing and data analysis. Many publications have concentrated on one type of DNA damage, but few have addressed the complete spectrum of FFPE-DNA damage. Here, we review mitigation strategies in (I) pre-analytical sample quality control, (II) DNA repair treatments, (III) analytical sample preparation and (IV) bioinformatic analysis of FFPE-DNA. We then provide recommendations that are tested and illustrated with DNA from 13-year-old liver specimens, one FFPE preserved and one fresh frozen, applying target-enriched sequencing. Thus, we show how DNA damage can be compensated, even when using low quantities (50 ng) of fragmented FFPE-DNA (DNA integrity number 2.0) that cannot be amplified well (Q129 bp/Q41 bp = 5%). Finally, we provide a checklist called 'ERROR-FFPE-DNA' that summarises recommendations for the minimal information in publications required for assessing fitness-for-purpose and inter-study comparison when using FFPE samples.

SUBMITTER: Steiert TA

PROVIDER: S-EPMC10415133 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

A critical spotlight on the paradigms of FFPE-DNA sequencing.

Steiert Tim A TA Parra Genís G Gut Marta M Arnold Norbert N Trotta Jean-Rémi JR Tonda Raúl R Moussy Alice A Gerber Zuzana Z Abuja Peter M PM Zatloukal Kurt K Röcken Christoph C Folseraas Trine T Grimsrud Marit M MM Vogel Arndt A Goeppert Benjamin B Roessler Stephanie S Hinz Sebastian S Schafmayer Clemens C Rosenstiel Philip P Deleuze Jean-François JF Gut Ivo G IG Franke Andre A Forster Michael M

Nucleic acids research 20230801 14

In the late 19th century, formalin fixation with paraffin-embedding (FFPE) of tissues was developed as a fixation and conservation method and is still used to this day in routine clinical and pathological practice. The implementation of state-of-the-art nucleic acid sequencing technologies has sparked much interest for using historical FFPE samples stored in biobanks as they hold promise in extracting new information from these valuable samples. However, formalin fixation chemically modifies DNA ...[more]

PMID: 37351572

Similar Datasets

Project description:Formalin-fixed, paraffin-embedded (FFPE) tissue specimens are routinely used in pathological diagnosis, but their large number of artifactual mutations complicate the evaluation of companion diagnostics and analysis of next-generation sequencing data. Identification of variants with low allele frequencies is challenging because existing FFPE filtering tools label all low-frequency variants as artifacts. To address this problem, we aimed to develop DEEPOMICS FFPE, an AI model that can classify a true variant from an artifact. Paired whole exome sequencing data from fresh frozen and FFPE samples from 24 tumors were obtained from public sources and used as training and validation sets at a ratio of 7:3. A deep neural network model with three hidden layers was trained with input features using outputs of the MuTect2 caller. Contributing features were identified using the SHapley Additive exPlanations algorithm and optimized based on training results. The performance of the final model (DEEPOMICS FFPE) was compared with those of existing models (MuTect filter, FFPolish, and SOBDetector) by using well-defined test datasets. We found 41 discriminating properties for FFPE artifacts. Optimization of property quantification improved the model performance. DEEPOMICS FFPE removed 99.6% of artifacts while maintaining 87.1% of true variants, with an F1-score of 88.3 in the entire dataset not used for training, which is significantly higher than those of existing tools. Its performance was maintained even for low-allele-fraction variants with a specificity of 0.995, suggesting that it can be used to identify subclonal variants. Different from existing methods, DEEPOMICS FFPE identified most of the sequencing artifacts in the FFPE samples while retaining more of true variants, including those of low allele frequencies. The newly developed tool DEEPOMICS FFPE may be useful in designing capture panels for personalized circulating tumor DNA assay and identifying candidate neoepitopes for personalized vaccine design. DEEPOMICS FFPE is freely available on the web ( http://deepomics.co.kr/ffpe ) for research.

Dataset Information

A critical spotlight on the paradigms of FFPE-DNA sequencing.

Publications

A critical spotlight on the paradigms of FFPE-DNA sequencing.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets