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Clinical framework for next generation sequencing based analysis of treatment predictive mutations and multiplexed
Kajsa Ericson Lindquist1, Anna Karlsson2, Per Levéen1
1Department of Pathology, Regional Laboratories Region Skåne, Lund SE 22185, Sweden.
Abstract:
Precision medicine requires accurate multi-gene clinical diagnostics. We describe the implementation of an Illumina TruSight Tumor (TST) clinical NGS diagnostic framework and parallel validation of a NanoString RNA-based ALK, RET, and ROS1 gene fusion assay for combined analysis of treatment predictive alterations in non-small cell lung cancer (NSCLC) in a regional healthcare region of Sweden (Scandinavia). The TST panel was clinically validated in 81 tumors (99% hotspot mutation concordance), after which 533 consecutive NSCLCs were collected during one-year of routine clinical analysis in the healthcare region (~90% advanced stage patients). The NanoString assay was evaluated in 169 of 533 cases. In the 533-sample cohort 79% had 1-2 variants, 12% >2 variants and 9% no detected variants. Ten gene fusions (five ALK, three RET, two ROS1) were detected in 135 successfully analyzed cases (80% analysis success rate). No ALK or ROS1 FISH fusion positive case was missed by the NanoString assay. Stratification of the 533-sample cohort based on actionable alterations in 11 oncogenes revealed that 66% of adenocarcinomas, 13% of squamous carcinoma (SqCC) and 56% of NSCLC not otherwise specified harbored ≥1 alteration. In adenocarcinoma, 10.6% of patients (50.3% if including KRAS) could potentially be eligible for emerging therapeutics, in addition to the 15.3% of patients eligible for standard EGFR or ALK inhibitors. For squamous carcinoma corresponding proportions were 4.4% (11.1% with KRAS) vs 2.2%. In conclusion, multiplexed NGS and gene fusion analyses are feasible in NSCLC for clinical diagnostics, identifying notable proportions of patients potentially eligible for emerging molecular therapeutics.
Insights
Multiplexed next-generation sequencing (NGS) and NanoString gene fusion assays are feasible for non-small cell lung cancer (NSCLC) diagnostics. These methods identify numerous patients eligible for targeted therapies and emerging molecular treatments.
Area of Science:
- Oncology
- Genetics
- Molecular Diagnostics
Background:
- Precision medicine in non-small cell lung cancer (NSCLC) necessitates accurate multi-gene clinical diagnostics.
- Implementing advanced molecular profiling is crucial for identifying actionable alterations and guiding treatment decisions.
Purpose of the Study:
- To describe the clinical implementation and validation of an Illumina TruSight Tumor (TST) NGS panel and a NanoString RNA-based assay for gene fusions (ALK, RET, ROS1) in NSCLC.
- To assess the utility of these combined diagnostic approaches for identifying patients eligible for targeted therapies in a Swedish healthcare region.
Main Methods:
- Clinical validation of the Illumina TruSight Tumor (TST) panel on 81 NSCLC tumors.
- Routine clinical analysis of 533 consecutive NSCLCs using the TST panel over one year.
- Parallel validation of a NanoString RNA-based assay for ALK, RET, and ROS1 gene fusions in 169 cases.
- Stratification of identified alterations against actionable targets for emerging and standard therapeutics.
Main Results:
- The TST panel showed 99% hotspot mutation concordance in initial validation.
- In the 533-sample cohort, 79% had 1-2 variants, and 9% had no detected variants.
- Ten gene fusions (five ALK, three RET, two ROS1) were detected in 135 analyzed cases (80% success rate).
- No ALK or ROS1 FISH-positive cases were missed by the NanoString assay.
- Actionable alterations were found in 66% of adenocarcinomas, 13% of squamous cell carcinomas, and 56% of NSCLC not otherwise specified.
- Up to 50.3% of adenocarcinoma patients (including KRAS) and 11.1% of squamous cell carcinoma patients (including KRAS) could be eligible for emerging therapeutics.
Conclusions:
- Multiplexed NGS and gene fusion analyses are clinically feasible for NSCLC diagnostics.
- These assays identify a significant proportion of NSCLC patients eligible for emerging molecular therapeutics.
- The combined approach enhances the molecular characterization of NSCLC, supporting personalized treatment strategies.