External quality assessment (EQA) for tumor mutational burden: results of an international IQN path feasibility pilot scheme.

Tumor mutational burden (TMB) has recently been approved as an agnostic biomarker for immune checkpoint inhibitors. However, methods for TMB testing have not yet been standardized. The International Quality Network for Pathology (IQNPath) organized a pilot external quality assessment (EQA) scheme for TMB testing. The aim of this program was the validation of the materials and the procedures for the EQA of this complex biomarker. Five formalin-fixed paraffin-embedded (FFPE) cell lines were selected to mimic the various TMB values observed in clinical practice. The FFPE samples were tested with the FoundationOne CDx (F1CDx) assay as the reference test and three commercially available targeted sequencing panels. Following this internal validation, the five cell lines were sent to 29 laboratories selected on the basis of a previous survey. Nineteen of the 23 laboratories that submitted results (82.6%) used targeted sequencing for TMB estimation. Only two laboratories performed whole exome sequencing (WES) and two assessed TMB by clinical exome. A high variability in the reported TMB values was observed. The variability was higher for samples with the highest TMB value according to the F1CDx test. However, good reproducibility of the TMB score was shown by laboratories using the same panel. The majority of laboratories did not indicate a TMB cut-off value for clinical interpretation. In conclusion, this pilot EQA scheme suggests that it is feasible to run such an EQA program for TMB assessment. However, the results of our pilot highlight the numerous challenges for the standardization of this test.

Assessment of high-sensitive methods for the detection of EGFR mutations in circulating free tumor DNA from NSCLC patients.

AIM: We assessed the ability of the Therascreen(®) kit (plasma-Therascreen) and of a peptide nucleic acids (PNA)-clamp approach to detect EGFR mutations in plasma-derived circulating-free tumor DNA (cftDNA) from non-small-cell lung cancer patients. MATERIALS & METHODS: cftDNA from 96 patients was analyzed for exon 19 deletions and the p.L858R mutation, using both plasma-Therascreen and PNA-clamp-based assays. RESULTS: None of the 70 EGFR wild-type patients showed EGFR mutations in cftDNA with both techniques (specificity: 100%). In 17/26 EGFR-mutant patients, plasma-Therascreen analysis confirmed the mutation identified in the primary tumor (analytical sensitivity: 65.4%). Similar results were obtained with the PNA-clamp method. CONCLUSION: Both approaches were specific and sensitive for EGFR mutational analysis of cftDNA in non-small-cell lung cancer patients.