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Validation of a Targeted Next-Generation Sequencing Panel for Tumor Mutation Burden Analysis: Results from the
Francesca Fenizia1, Ilaria Alborelli2, Jose Luis Costa3
1Cell Biology and Biotherapy Unit, Istituto Nazionale Tumori-IRCCS-Fondazione G. Pascale, Naples, Italy.
Abstract:
Tumor mutation burden (TMB) is evaluated as a biomarker of response to immunotherapy. We present the efforts of the Onconetwork Immuno-Oncology Consortium to validate a commercial targeted sequencing test for TMB calculation. A three-phase study was designed to validate the Oncomine Tumor Mutational Load (OTML) assay at nine European laboratories. Phase 1 evaluated reproducibility and accuracy on seven control samples. In phase 2, six formalin-fixed, paraffin-embedded samples tested with FoundationOne were reanalyzed with the OTML panel to evaluate concordance and reproducibility. Phase 3 involved analysis of 90 colorectal cancer samples with known microsatellite instability (MSI) status to evaluate TMB and MSI association. High reproducibility of TMB was demonstrated among the sites in the first and second phases. Strong correlation was also detected between mean and expected TMB in phase 1 (r2 = 0.998) and phase 2 (r2 = 0.96). Detection of actionable mutations was also confirmed. In colorectal cancer samples, the expected pattern of MSI-high/high-TMB and microsatellite stability/low-TMB was present, and gene signatures produced by the panel suggested the presence of a POLE mutation in two samples. The OTML panel demonstrated robustness and reproducibility for TMB evaluation. Results also suggest the possibility of using the panel for mutational signatures and variant detection. Collaborative efforts between academia and companies are crucial to accelerate the translation of new biomarkers into clinical research.
Insights
The Oncomine Tumor Mutational Load (OTML) assay shows high reproducibility and accuracy for calculating tumor mutation burden (TMB), a key immunotherapy biomarker. This validated assay supports reliable TMB assessment in clinical research.
Area of Science:
- Oncology
- Genomics
- Biomarker Validation
Background:
- Tumor mutation burden (TMB) is a critical biomarker for predicting immunotherapy response.
- Standardization of TMB assessment is essential for clinical application.
Purpose of the Study:
- To validate the Oncomine Tumor Mutational Load (OTML) assay for accurate and reproducible TMB calculation.
- To assess the OTML assay's concordance with other methods and its performance in diverse sample types.
Main Methods:
- A three-phase study involving nine European laboratories.
- Validation using control samples, formalin-fixed paraffin-embedded samples, and colorectal cancer patient samples.
- Assessment of reproducibility, accuracy, concordance, and correlation with microsatellite instability (MSI) status.
Main Results:
- High reproducibility and accuracy of TMB measurements across multiple sites.
- Strong correlation between mean and expected TMB values (r² = 0.998 in Phase 1, r² = 0.96 in Phase 2).
- Accurate detection of actionable mutations and expected TMB patterns in relation to MSI status.
Conclusions:
- The OTML panel is a robust and reproducible tool for TMB evaluation.
- The assay shows potential for mutational signature analysis and variant detection.
- Collaborative efforts are vital for biomarker translation in immuno-oncology research.