Abstract 4530: A novel blood-based method for evaluating KRAS in circulating microvesicles from colorectal cancer patients

Circulating microvesicles (cMV) are lipid-encapsulated bodies that are secreted from various tissues and can be detected in a number of body fluids, including plasma. Once collected from plasma, they can be exploited diagnostically for their protein and RNA signatures. Mutations in KRAS are diagnostically important for predicting response to chemotherapy and prognosis. A blood-based method of assessing KRAS mutation status would be helpful for patients with colorectal cancer (CRC). Traditional methods of KRAS detection examine the genomic DNA sequence. We developed a method to sequence exon 3 from KRAS using Pyrosequencing. The limit of detection for the assay was 0.78 ng/uL of mutant cDNA for both Pyrosequencing and Sanger sequencing. To further enhance mutation detection, we sorted cMV from CRC patients by first capturing cMV with a CRC associated membrane protein and then sorting for CD63 and CD24 positive events. A Taqman gene expression assay was not sensitive enough to detect KRAS in the sorted samples. However, Pyrosequencing was sufficient to identify mutant and wild-type sequences in patient plasma samples. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4530. doi:1538-7445.AM2012-4530