Triple Platform Testing (IHC, FISH and Mutation) To Predict Response to Targeted Therapy in NSCLC
D Lucas, R Camidge, R Doebele, J Haney, M Sugita, L Bemis, W Franklin. University of Colorado at Denver and Health Sciences Center, Aurora, CO
Background: Multiple molecular abnormalities in the EGFR pathway including overexpression of EGFR protein, high gene copy number and mutations in EGFR and Ki-ras genes are reported to correlate with improved survival in response to EGFR blockade. To assess all of these abnormalities in a single predictive profile, our lab developed a triple-platform molecular testing protocol that includes IHC (EGFR, HER2/neu, and ERCC1), FISH (EGFR and HER2/neu), and mutational tests (EGFR, TP53 and Ki-ras) that are combined into a single interpretive report.
Design: 39 patients with NSCLC have been tested to date. IHC was considered positive if the product of the staining intensity (0-3 scale) times the percent of cells stained was >150. High gene copy in multicolor FISH was indicated by >40% of tumor cells with 4 signals/cell. Mutational analysis for Ki-ras (exon 1), EGFR (exons 19-21), and TP53 (exons 5-8) mutations were evaluated in DNA extracted from a 1 mm core of tumor tissue removed from a paraffin block which was amplified by PCR and directly sequenced.
Results: Of the 39 patients evaluated for EGFR and ERCC1 protein overexpression, 20 (51%) and 9 (23%) showed strong expression for EGFR and ERCC1, respectively. Patients were also tested for a high gene copy number and identified 25 (64%) that were both EGFR and HER2-neu FISH-positive. Mutational analysis revealed a total of 21 (54%) somatic mutations which included 7 EGFR (18%), 7 TP53 (18%), and 7 Ki-ras (18%) single mutations. Two patients revealed a double-mutant profile consisting of both a TP53 and Ki-ras mutations. Many of the markers had seemingly contradictory results and at the present time a weighted approach is being developed to interpret the interaction among the markers in regard to prognosis and prediction of treatment response. In this algorithm, mutation is an initial filter followed respectively by FISH and protein expression.
Conclusions: Triple-platform testing provides an integrated approach to biomarker assessment that lends itself to clinical interpretation and documentation for further correlations with outcome. Triple platform testing is expected to aid in patient-specific targeted agent treatment planning.
Tuesday, March 10, 2009 1:00 PM
Poster Session IV # 233, Tuesday Afternoon