Clinical Implications

Lung cancer is a leading cause of cancer death worldwide. Patients with non-small cell lung cancer (NSCLC) often present with advanced disease and treatment benefit with standard chemotherapy is modest. The introduction of therapies that target the epidermal growth factor receptor (EGFR) pathway in NSCLC, such as the tyrosine kinase inhibitors erlotinib (Tarceva®), osimertinib (Tagrisso®) and gefitinib (Iressa®) has begun to improve outcomes in patients with advanced NSCLC. Especially those NSCLC cases harboring activating mutations in the EGFR gene show response to anti-EGFR tyrosin kinase inhibitors. The highest response rates have been shown for exon 19 deletions, G719A/C, L858R and L861Q. Therefore, it is important to identify EGFR mutations in NSCLC.

Treatment with first- or second-generation EGFR-tyrosine kinase inhibitors (TKIs) is effective for NSCLC patients harboring a sensitizing EGFR mutation. However, resistance is often acquired after 9–14 months. The most common mechanism of resistance to first- and second-generation EGFR-TKIs in the first-line setting is the EGFR T790M mutation, which accounts for approximately 60% of cases. Third-generation EGFR-TKIs targeting EGFR T790M mutation (e.g. osimertinib), are reported to be highly active against T790M-positive NSCLC. To detect the T790M mutation in patients progressing during EGFR-TKI treatment, tumor re-biopsy is necessary. As re-biopsies with invasive procedures (bronchoscopy or needle biopsy) are often infeasible in standard care of NSCLC patients, circulating tumor DNA (ctDNA) detected in plasma is recognized as a noninvasive biomarker for the molecular analysis of NSCLC. The cobas® EGFR Mutation Test (Roche Diagnostics K.K., Switzerland.) is a companion diagnostic test for the detection of EGFR mutations in plasma specimens and has been approved to identify such patients with NSCLC. T790M monitoring in plasma ctDNA of patients receiving EGFR-TKIs could yield valuable clinical information.

Specimen Requirements

FFPE:

  • Formalin-fixed, paraffin-embedded (FFPE) NSCLC tissue specimens with a fixation time of 6‑48 hours

Plasma:

  • Whole blood or plasma derived from (K2) EDTA anti-coagulated peripheral whole blood

Volume

FFPE:

  • One representative paraffin block is preferred. Alternatively, 3 unstained tissue sections are requested. Of interest, if additional ALK FISH analysis is required (confirmation of ALK IHC positive case), two 3 µm thick unstained sections are required for ALK FISH analysis.

Plasma:

  • 8ml of whole blood or 2ml of plasma is required.

Storage and Shipment Instructions

FFPE:

  • Maintain and ship FFPE specimens at ambient temperature.

Plasma:

  • Maintain and ship whole blood specimens at ambient temperature.
  • Maintain plasma specimens at -70°C or colder and ship on dry ice.

Limitations

FFPE:

  • Insufficient tumor content may not allow the detection of EGFR mutations (< 10%); tumor content is evaluated prior to analysis and macrodissection is performed. Fixatives other than formalin or prolonged fixation time may give rise to inadequate results.

Plasma:

  • Samples tested at high concentrations ( >105 copies/mL) may generate false results.

General:

  • The Cobas® EGFR Mutation Test v2 shows cross-reactivity (results of “Mutation Detected”) to the exon 19 L747S mutation, a rare acquired mutation that may confer resistance to TKI treatment.
  • Detection of a mutation is dependent on the number of copies present in the specimen and may be affected by sample integrity, amount of isolated DNA, and the presence of interfering substances. The presence of PCR inhibitors may cause false negative or invalid results.

Turn-Around Time

Five to seven business days for plasma and slides and paraffin blocks, respectively.