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Comparative analysis of somatic variant calling on matched FF and FFPE WGS samples

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Abstract
Background Research grade Fresh Frozen (FF) DNA material is not yet routinely collected in clinical practice. Many hospitals, however, collect and store Formalin Fixed Paraffin Embedded (FFPE) tumor samples. Consequently, the sample size of whole genome cancer cohort studies could be increased tremendously by including FFPE samples, although the presence of artefacts might obfuscate the variant calling. To assess whether FFPE material can be used for cohort studies, we performed an in-depth comparison of somatic SNVs called on matching FF and FFPE Whole Genome Sequence (WGS) samples extracted from the same tumor. Methods Four variant callers (i.e. Strelka2, Mutect2, VarScan2 and Shimmer) were used to call somatic variants on matching FF and FFPE WGS samples from a metastatic prostate tumor. Using the variants identified by these callers, we developed a heuristic to maximize the overlap between the FF and its FFPE counterpart in terms of sensitivity and precision. The proposed variant calling approach was then validated on nine matched primary samples. Finally, we assessed what fraction of the discrepancy could be attributed to intra-tumor heterogeneity (ITH), by comparing the overlap in clonal and subclonal somatic variants. Results We first compared variants between an FF and an FFPE sample from a metastatic prostate tumor, showing that on average 50% of the calls in the FF are recovered in the FFPE sample, with notable differences between callers. Combining the variants of the different callers using a simple heuristic, increases both the precision and the sensitivity of the variant calling. Validating the heuristic on nine additional matched FF-FFPE samples, resulted in an average F1-score of 0.58 and an outperformance of any of the individual callers. In addition, we could show that part of the discrepancy between the FF and the FFPE samples can be attributed to ITH. Conclusion This study illustrates that when using the correct variant calling strategy, the majority of clonal SNVs can be recovered in an FFPE sample with high precision and sensitivity. These results suggest that somatic variants derived from WGS of FFPE material can be used in cohort studies.
Keywords
CANCER, FRESH, Somatic variants, FFPE, WGS, Precision oncology, Cohort studies

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MLA
de Schaetzen van Brienen, Louise, et al. “Comparative Analysis of Somatic Variant Calling on Matched FF and FFPE WGS Samples.” BMC MEDICAL GENOMICS, vol. 13, no. 1, 2020, doi:10.1186/s12920-020-00746-5.
APA
de Schaetzen van Brienen, L., Larmuseau, M., Van der Eecken, K., De Ryck, F., Robbe Vandewalle, P., Schuh, A., … Marchal, K. (2020). Comparative analysis of somatic variant calling on matched FF and FFPE WGS samples. BMC MEDICAL GENOMICS, 13(1). https://doi.org/10.1186/s12920-020-00746-5
Chicago author-date
Schaetzen van Brienen, Louise de, Maarten Larmuseau, Kim Van der Eecken, Frederic De Ryck, Pauline Robbe Vandewalle, Anna Schuh, Jan Fostier, Piet Ost, and Kathleen Marchal. 2020. “Comparative Analysis of Somatic Variant Calling on Matched FF and FFPE WGS Samples.” BMC MEDICAL GENOMICS 13 (1). https://doi.org/10.1186/s12920-020-00746-5.
Chicago author-date (all authors)
de Schaetzen van Brienen, Louise, Maarten Larmuseau, Kim Van der Eecken, Frederic De Ryck, Pauline Robbe Vandewalle, Anna Schuh, Jan Fostier, Piet Ost, and Kathleen Marchal. 2020. “Comparative Analysis of Somatic Variant Calling on Matched FF and FFPE WGS Samples.” BMC MEDICAL GENOMICS 13 (1). doi:10.1186/s12920-020-00746-5.
Vancouver
1.
de Schaetzen van Brienen L, Larmuseau M, Van der Eecken K, De Ryck F, Robbe Vandewalle P, Schuh A, et al. Comparative analysis of somatic variant calling on matched FF and FFPE WGS samples. BMC MEDICAL GENOMICS. 2020;13(1).
IEEE
[1]
L. de Schaetzen van Brienen et al., “Comparative analysis of somatic variant calling on matched FF and FFPE WGS samples,” BMC MEDICAL GENOMICS, vol. 13, no. 1, 2020.
@article{8671349,
  abstract     = {{Background Research grade Fresh Frozen (FF) DNA material is not yet routinely collected in clinical practice. Many hospitals, however, collect and store Formalin Fixed Paraffin Embedded (FFPE) tumor samples. Consequently, the sample size of whole genome cancer cohort studies could be increased tremendously by including FFPE samples, although the presence of artefacts might obfuscate the variant calling. To assess whether FFPE material can be used for cohort studies, we performed an in-depth comparison of somatic SNVs called on matching FF and FFPE Whole Genome Sequence (WGS) samples extracted from the same tumor. Methods Four variant callers (i.e. Strelka2, Mutect2, VarScan2 and Shimmer) were used to call somatic variants on matching FF and FFPE WGS samples from a metastatic prostate tumor. Using the variants identified by these callers, we developed a heuristic to maximize the overlap between the FF and its FFPE counterpart in terms of sensitivity and precision. The proposed variant calling approach was then validated on nine matched primary samples. Finally, we assessed what fraction of the discrepancy could be attributed to intra-tumor heterogeneity (ITH), by comparing the overlap in clonal and subclonal somatic variants. Results We first compared variants between an FF and an FFPE sample from a metastatic prostate tumor, showing that on average 50% of the calls in the FF are recovered in the FFPE sample, with notable differences between callers. Combining the variants of the different callers using a simple heuristic, increases both the precision and the sensitivity of the variant calling. Validating the heuristic on nine additional matched FF-FFPE samples, resulted in an average F1-score of 0.58 and an outperformance of any of the individual callers. In addition, we could show that part of the discrepancy between the FF and the FFPE samples can be attributed to ITH. Conclusion This study illustrates that when using the correct variant calling strategy, the majority of clonal SNVs can be recovered in an FFPE sample with high precision and sensitivity. These results suggest that somatic variants derived from WGS of FFPE material can be used in cohort studies.}},
  articleno    = {{94}},
  author       = {{de Schaetzen van Brienen, Louise and Larmuseau, Maarten and Van der Eecken, Kim and De Ryck, Frederic and Robbe Vandewalle, Pauline and Schuh, Anna and Fostier, Jan and Ost, Piet and Marchal, Kathleen}},
  issn         = {{1755-8794}},
  journal      = {{BMC MEDICAL GENOMICS}},
  keywords     = {{CANCER,FRESH,Somatic variants,FFPE,WGS,Precision oncology,Cohort studies}},
  language     = {{eng}},
  number       = {{1}},
  pages        = {{15}},
  title        = {{Comparative analysis of somatic variant calling on matched FF and FFPE WGS samples}},
  url          = {{http://dx.doi.org/10.1186/s12920-020-00746-5}},
  volume       = {{13}},
  year         = {{2020}},
}

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