A second-generation microRNA-based assay for diagnosing tumor tissue origin

Abstract

Abstract Background: Cancer of unknown primary (CUP) constitutes 3%~5% of all newly diagnosed cancer cases. It presents a major diagnostic challenge due to the therapeutic implications of the tissue origin of the cancer. Accurate tumor of origin (ToO) assignment could help guide use of specific therapy which may impact survival. MicroRNAs are a family of noncoding, regulatory RNA genes that are involved in development, differentiation and cell cycle and were shown to be involved in oncogenesis. MicroRNAs are highly tissue specific and therefore ideal candidates for CUP diagnosis. We have previously presented a biologically-motivated classification assay to identify the tissue of origin using a set of 48 microRNAs measured on a qRT-PCR platform. This assay can differentiate 25 different tumor types from 17 origins. We present here the development and validation of a second generation assay that can identify 43 different tumor types using a custom array platform. The assay is based on the normalized expression of less than 70 microRNAs. We will present performance data including inter-laboratory concordance from the validation of the assay. Methods: Over 1200 primary and metastatic tumor FFPE samples were used for the training and development of the next generation assay. High-quality RNA, including the well-preserved microRNA fraction, was extracted from the FFPE blocks using proprietary protocols previously described. Expression levels of all known and additional Rosetta's proprietary microRNAs were profiled using a custom array platform. MicroRNAs containing information about the ToO were identified and two classifiers were developed to utilize the information of the microRNA expression patterns for the diagnosis of ToO. The assay reports a ToO based on these two classifications, where in most cases the assay result is a single ToO reported. Results: The developed assay is able to accurately identify more than 40 tumor types based on the expression of only several dozens microRNAs. The results of a comprehensive validation study using hundreds of independent tumor and metastases FFPE samples will be presented, including inter-laboratory concordance data. Conclusions: Previous studies have highlighted the tissue-specificity of microRNA expression, and have demonstrated their potential use for classification of human malignancies. An assay utilizing microRNA expression in a biopsy for identifying the tissue of origin was developed. Here we present the development and the performance of a second generation assay, designed to identify over 40 tumor types. This assay will provide an important new tool for diagnosing tumor tissue origin.