# Implementation of an ISO 15189 accredited next generation sequencing service for cell-free total nucleic acid (cfTNA) analysis to facilitate driver mutation reporting in blood: the experience of a clinical diagnostic laboratory

## TL;DR

This paper describes the implementation of a reliable, fast, and minimally invasive NGS service for detecting cancer-related mutations in blood, improving patient care when tissue samples are not available.

## Contribution

The paper presents the first ISO 15189 accredited NGS service for cfTNA analysis with reduced turnaround time and high accuracy.

## Key findings

- High sensitivity (>83%) and specificity were achieved between plasma and tissue results.
- A sequencing limit of detection of 1.2% was achieved with >22,000× coverage.
- Turnaround time for liquid biopsy results was reduced by over 68% compared to reference labs.

## Abstract

Next generation sequencing (NGS) on tumour tissue is integral to the delivery of personalised medicine and targeted therapy. NGS on liquid biopsy, a much less invasive technology, is an emerging clinical tool that has rapidly expanded clinical utility. Gene mutations in cell-free total nucleic acids (cfTNA) circulating in the blood are representative of whole tumour biology and can reveal different mutations from different tumour sites, thus addressing tumour heterogeneity challenges.

The novel Ion Torrent Genexus NGS system with automated sample preparation, onboard library preparation, templating, sequencing, data analysis and Oncomine Reporter software was used. cfTNA extracted from plasma was verified with the targeted pan-cancer (~50 genes) Oncomine Precision Assay (OPA). Assessment criteria included analytical sensitivity, specificity, limits of detection (LOD), accuracy, repeatability, reproducibility and the establishment of performance metrics.

An ISO 15189 accredited, minimally invasive cfTNA NGS diagnostic service has been implemented. High sensitivity (>83%) and specificity between plasma and tissue were observed. A sequencing LOD of 1.2% was achieved when the depth of coverage was >22 000×. A reduction (>68%) in turnaround time (TAT) of liquid biopsy results was achieved: 5 days TAT for in-house analysis from sample receipt to a final report issued to oncologists as compared with >15 days from reference laboratories.

Tumour-derived somatic variants can now be reliably assessed from plasma to provide minimally invasive tumour profiling. Successful implementation of this accredited service resulted in:

Appropriate molecular profiling of patients where tumour tissue is unavailable or inaccessible.

Rapid TAT of plasma NGS results.