Validation and Reproducibility of a Microarray-Based Gene Expression Test for Tumor Identification in Formalin-Fixed, Paraffin-Embedded Specimens

Abstract

Tumors whose primary site is challenging to diagnose represent a considerable proportion of new cancer cases. We present validation study results for a gene expression-based diagnostic test (the Pathwork Tissue of Origin Test) that aids in determining the tissue of origin using formalin-fixed, paraffin-embedded (FFPE) specimens. Microarray data files were generated for 462 metastatic, poorly differentiated, or undifferentiated FFPE tumor specimens, all of which had a reference diagnosis. The reference diagnoses were masked, and the microarray data files were analyzed using a 2000-gene classification model. The algorithm quantifies the similarity between RNA expression patterns of the study specimens and the 15 tissues on the test panel. Among the 462 specimens, overall agreement with the reference diagnosis was 89% (95% CI, 85% to 91%). In addition to the positive test results (ie, rule-ins), an average of 12 tissues for each specimen could be ruled out with >99% probability. The large size of this study increases confidence in the test results. A multisite reproducibility study showed 89.3% concordance between laboratories. The Tissue of Origin Test makes the benefits of microarray-based gene expression tests for tumor diagnosis available for use with the most common type of histology specimen (ie, FFPE).

A 92-gene cancer classifier predicts the site of origin for neuroendocrine tumors

“…14 Traditionally, pathologists have used panels of protein-based immunohistochemical stains for tumor classification, but the potential advantage of molecular tests is readily apparent. In recent years, RNA expression-based classification of tumors has become an attractive standardized approach to aiding in the diagnosis and subclassification of tumors [19][20][21][22][23][24][25] and for individualized therapy applications. [26][27][28] In the coming years, gene expressionbased analysis may serve as a complementary approach to genome-wide DNA analysis of tumors for individualized therapy.…”

Cost-effectiveness of using a gene expression profiling test to aid in identifying the primary tumour in patients with cancer of unknown primary

“…34 We used a lifetime horizon and half cycle correction. 35 24 and clinical verification of the test performance 26 (Table 1). We estimated all other distributions in the decision tree and transition probabilities in the Markov models (Table 1) from the observed clinical management and survival outcomes in our study cohort.…”

“…9,10 To date, several gene expression-based tests have demonstrated the potential value of this approach in identifying the primary site. 4,[11][12][13][14][15][16][17][18][19][20][21][22][23][24] One microarray-based test uses 2000-GEP to identify a tumour's primary site using formalin-fixed paraffin-embedded (FFPE) specimens (Tissue of Origin test, Response Genetics, Inc., Los Angeles, CA). 24 The test compares the RNA profile of a tumour FFPE specimen to established RNA profiles of 15 known tissues.…”

“…A maximum SS of 30 or less indicates indeterminate results which might occur if the specimen harbors less than 20% tumor content or if the tumour specimen is not represented by the 15 tissue types included in the test panel. 24 Specimen requirements include a minimum of 20% tumour content, a maximum of 20% necrosis and a minimum tumor area of 0.5 mm 2 .…”

“…24 Based on this analysis, the test was approved by the Food and Drug Administration in 2012 25 and has been available for clinical use in the United states. The 2000-GEP test results are intended for use in the context of the patient's clinicopathologic and radiologic history by a qualified oncologist and pathologist.…”