Development and analytical validation of a next-generation sequencing based microsatellite instability (MSI) assay

Area of Science:

• Oncology
• Genomics
• Molecular Diagnostics

Background:

• Microsatellite instability (MSI) is a key biomarker in solid tumors.
• Accurate MSI classification is crucial for guiding cancer treatment decisions.
• Current MSI testing methods can be complex and time-consuming.

Purpose of the Study:

• To develop and validate a novel next-generation sequencing (NGS) assay for classifying MSI in formalin-fixed paraffin-embedded (FFPE) tumor specimens.
• To establish a rapid and reliable method for MSI detection without the need for matched-normal tissue.

Main Methods:

• Utilized DNA-sequencing (DNA-seq) to evaluate insertion/deletion (indel) variability at 29 genomic loci.
• Developed the assay on a training set (n=94) and validated on an independent test set.
• Assessed assay performance across diverse FFPE samples, histologies, and pre-analytical/analytical variables.

Main Results:

• The NGS assay demonstrated high accuracy, reproducibility, and precision compared to gold-standard PCR methods.
• Achieved a clinically relevant five-day turnaround time using as little as 20 ng of DNA.
• Validated through NYS CLEP, confirming its reliability for clinical use.

Conclusions:

• This validated NGS assay offers a robust and reproducible method for MSI testing in solid tumors.
• Eliminates the need for matched-normal tissue, streamlining clinical decision-making.
• Empowers clinicians with reliable genomic data for personalized cancer therapy.