Development, Implementation and Assessment of Molecular Diagnostics by Next Generation Sequencing in Personalized Treatment of Cancer: Experience of a Public Reference Healthcare Hospital

Abstract

The establishment of precision medicine in cancer patients requires the study of several biomarkers. Single-gene testing approaches are limited by sample availability and turnaround time. Next generation sequencing (NGS) provides an alternative for detecting genetic alterations in several genes with low sample requirements. Here we show the implementation to routine diagnostics of a NGS assay under International Organization for Standardization (UNE-EN ISO 15189:2013) accreditation. For this purpose, 106 non-small cell lung cancer (NSCLC) and 102 metastatic colorectal cancer (mCRC) specimens were selected for NGS analysis with Oncomine Solid Tumor (ThermoFisher). In NSCLC the most prevalently mutated gene was TP53 (49%), followed by KRAS (31%) and EGFR (13%); in mCRC, TP53 (50%), KRAS (48%) and PIK3CA (16%) were the most frequently mutated genes. Moreover, NGS identified actionable genetic alterations in 58% of NSCLC patients, and 49% of mCRC patients did not harbor primary resistance mechanisms to anti-EGFR treatment. Validation with conventional approaches showed an overall agreement >90%. Turnaround time and cost analysis revealed that NGS implementation is feasible in the public healthcare context. Therefore, NGS is a multiplexed molecular diagnostic tool able to overcome the limitations of current molecular diagnosis in advanced cancer, allowing an improved and economically sustainable molecular profiling.

Usefulness of Two Independent DNA and RNA Tissue-Based Multiplex Assays for the Routine Care of Advanced NSCLC Patients

“…Although both techniques (NGS and nCounter) require minimum amounts of genetic material, our dropout percentage increased to 15% when considering those samples for which DNA or RNA testing could not be performed. Some groups, which also included combined DNA and RNA in their sequencing workflows, have reported lower invalid test rates of approximately 2%-3% [18,24]. However, these results do not correspond to the total screening failure rates, which should include samples with insufficient material, the major reason for failure in our series.…”

Impact of Molecular Testing Using Next-Generation Sequencing in the Clinical Management of Patients with Non-Small Cell Lung Cancer in a Public Healthcare Hospital

“…In January 2021, as a consequence of the accreditation of NGS techniques under the UNE-EN ISO 15189:2022 standard [ 14 ], the Lung Cancer Committee of our hospital decided to establish NGS as a routine testing strategy for the molecular profiling of NSCLC. During this period, 128 samples were analyzed using NGS.…”

The importance of precision medicine in modern molecular oncology

“…Several mutant genes (TP53, KRAS, PIK3CA, HER2 and BRAF) in metastatic colorectal cancer (mCRC) are identified by using NGS plateforms. 70,71 Mutated P53 is quite common in advanced CRC and APE1 is overexpressed in CRC. The relationship between P53 and APE1 has been studied recently.…”