In an adult with newly diagnosed advanced/metastatic non‑small‑cell lung carcinoma, what molecular tests should be performed and which first‑line targeted therapies are indicated for each actionable alteration?

Molecular Testing and Targeted Therapy for Advanced/Metastatic NSCLC

All patients with newly diagnosed advanced/metastatic non-small cell lung cancer should undergo broad molecular profiling via next-generation sequencing (NGS) to test for EGFR, ALK, KRAS, ROS1, BRAF, NTRK1/2/3, MET exon 14 skipping, RET, and HER2 alterations, with tissue and/or plasma testing performed concurrently to expedite results before treatment initiation. 1

Required Molecular Testing Panel

Mandatory Biomarkers (All Histologies)

For non-squamous NSCLC (adenocarcinoma, large cell carcinoma, NSCLC-NOS):

• EGFR mutations (Category 1 recommendation) - test exons 18-21 1
• ALK rearrangements (Category 1 recommendation) 1
• KRAS mutations (including G12C) 1, 2
• ROS1 rearrangements 1
• BRAF V600E mutations 1, 2
• NTRK1/2/3 fusions 1
• MET exon 14 skipping mutations 1
• RET fusions 1, 2
• HER2 (ERBB2) mutations 1, 2

For squamous cell carcinoma:

• Consider the same comprehensive panel, as 2-10% harbor actionable alterations 1
• Particularly test in patients <50 years, never/light smokers, or those with >15 years since smoking cessation 1, 2

Additional Essential Testing

PD-L1 expression (Category 1 recommendation) - required upfront regardless of histology to guide immunotherapy decisions if no actionable driver mutations are identified 1

Testing Methodology

Preferred Approach

Use comprehensive NGS panels that combine:

• DNA-based sequencing for mutations and copy number alterations 1, 2
• RNA-based NGS for fusion detection (ALK, ROS1, RET, NTRK, NRG1) - strongly preferred over DNA methods for fusions 2

Concurrent tissue AND plasma testing improves time to results and should be considered based on clinical urgency 1

Critical Technical Specifications

• Sensitivity ≥1% variant allele frequency for mutations 2
• Complete coverage of all specified exons (e.g., EGFR exons 18-21) 1, 2
• Validated bioinformatics pipelines for accurate variant calling 2

Common Pitfall to Avoid

Never perform sequential single-gene testing - this depletes limited tissue, misses actionable targets, and delays treatment 1, 2

First-Line Targeted Therapies by Alteration

EGFR Mutations (Exon 19 deletions, L858R)

Osimertinib is the preferred first-line agent with ~80% response rates 1

• Superior to first/second-generation EGFR TKIs 3
• Excellent CNS penetration with >60% intracranial response rates in brain metastases 3

ALK Rearrangements

Alectinib or lorlatinib are preferred first-line options 3

• Both demonstrate superior intracranial tumor control 3
• Crizotinib yields >60% response rates but is now second-line 3

ROS1 Rearrangements

Crizotinib (FDA-approved) achieves ~70% response rates including complete responses 3

• Occurs in 1-2% of NSCLC, more common in younger women, never-smokers with adenocarcinoma 3

KRAS G12C Mutations

Specific KRAS G12C inhibitors are now available for this smoking-related target 2, 4

BRAF V600E Mutations

BRAF TKIs are available and indicated 2, 4

MET Exon 14 Skipping

MET kinase inhibitors are approved for this indication 1, 4

RET Fusions

RET-specific inhibitors are indicated 4

NTRK Fusions

TRK inhibitors are available for these rare alterations 4

HER2 Mutations

Targeted drugs and antibody-drug conjugates are in development for exon 20 insertions 1, 2

Treatment Timing and Sequencing

When Results Are Pending

If urgent treatment is needed:

• Consider holding immunotherapy for one cycle and using platinum-based chemotherapy alone 1
• Availability of molecular testing before treatment initiation is associated with longer overall survival 1

Critical Warning About Immunotherapy

Avoid combining or sequencing immune checkpoint inhibitors before targeted therapy - particularly osimertinib has higher rates of adverse events (especially pneumonitis) when used after checkpoint inhibitors due to their long half-life 1

If Biomarker Discovered During First-Line Therapy

Either interrupt or complete the planned systemic therapy before switching to targeted therapy 1

• Decision factors: toxicity level and clinical/radiographic response 1

Rationale for Targeted Therapy Over Immunotherapy

When actionable drivers are present, targeted therapies are preferred over first-line immunotherapy regardless of PD-L1 levels because:

• Targeted therapies yield higher response rates (e.g., osimertinib 80% vs. immunotherapy 3.6% in EGFR-mutated disease) 1, 3
• PD-1/PD-L1 inhibitor monotherapy shows inferior efficacy in oncogene-driven NSCLC 3

Testing at Disease Progression

Repeat molecular testing is essential at progression on targeted therapy 1, 2

• Use re-biopsy or cell-free DNA (cfDNA) testing 1, 2
• Identifies resistance mechanisms (e.g., EGFR T790M after first/second-generation TKIs, MET amplification after osimertinib) 1, 2
• Rules out histologic transformation to small cell (~5% of EGFR TKI-resistant tumors) 3

Important Co-Mutations

TP53 co-mutations may be associated with lower efficacy of EGFR, ALK, and ROS1 TKIs 1, 2

• Testing for co-mutations in NGS panels is becoming important for prognostication 1, 2