Abstract 3387: A complete workflow for high throughput isolation of serum microRNAs and downstream analysis by qRT-PCR: application to cancer biomarker discovery
Authors: Zeringer Emily M. , Rai Alex J. , DeCastro Joel , Qu Luming , Gonzalez Marie , Chapman Laura , Vlassov Alexander V. , Magdaleno Susan M. , Liu Chunmei , Hu Fangqi , Dong Shoulian , Wong Linda 
Published: August 2015

Biomarkers have become invaluable tools in the clinical space for cancer detection, diagnosis, patient prognosis and treatment selection. An ideal biomarker should be easily assayed with minimally invasive medical procedures but possess high sensitivity and specificity. Although many candidate biomarkers for various diseases have been proposed in the literature, very few have made their way to clinical use, and for certain types of cancer there are no reliable biomarker options.

The use of prostate specific antigen (PSA) has led to a dramatic increase in the diagnosis of prostate cancer - over 230,000 new cases in 2014. However, most of these cases are of low risk disease that is unlikely to cause significant morbidity and mortality, leading to the accepted notion that prostate cancer is overdiagnosed. On the other hand, while it is a sensitive test, less than ¼ of patients undergoing a prostate biopsy based on PSA testing are actually found to have the disease. The identification of biomarkers that are more senstitive and specific predictors for the presence of prostate cancer, as well as for the aggressiveness of the disease, could decrease the complications and morbidity related to prostate biopsies and definitive treatment.

The circulating miRNA in blood could potentially fill this void, providing biomarkers suitable for real-time detection and application to the management of prostate cancer patients. The main advantages of miRNAs versus longer molecules (mRNA, lncRNA) include: (i) smaller number of sequences &lt;2000 and thus ease of analysis (ii) stability due to small size ∼16-27 nt and thus robustness of detection.

The objective of this project was to develop a set of reagents and tools for efficient recovery and analysis of circulating miRNAs. Our team has built a complete workflow to provide: (1) high throughput isolation of total serum RNA using a new MagMaxTM mirVanaTM reagent kit for the KingFisherTM Flex and Duo instruments, and (2) downstream analysis of microRNA profiles by qRT-PCR using next generation TaqMan® microRNA assays.

The effectiveness of this workflow is exemplified by analysis of a panel of miRNAs in samples derived from serum of patients with metastatic prostate cancer versus control groups. Following this fast and easy workflow suitable for serum and other body fluids, disease-specific RNA signatures can be identified and used as biomarkers.

TaqMan® is a registered Trademark of Roche Molecular Systems, Inc. used under permission and license.

Citation Format: Emily M. Zeringer, Alex J. Rai, Joel DeCastro, Luming Qu, Marie Gonzalez, Laura Chapman, Alexander V. Vlassov, Susan M. Magdaleno, Chunmei Liu, Fangqi Hu, Shoulian Dong, Linda Wong. A complete workflow for high throughput isolation of serum microRNAs and downstream analysis by qRT-PCR: application to cancer biomarker discovery. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3387. doi:10.1158/1538-7445.AM2015-3387