New rapid and robust transcriptome-based methods for cellular characterization of the tumor microenvironment and biomarker discovery are required to improve cancer prognosis and treatment. However, challenges with current approaches for the above applications include high sample requirements, poor sensitivity, low dynamic range, and limited throughput. To address these limitations, we have developed a unique approach for targeted transcriptome profiling using validated targeted primers that leverages the sensitivity of multiplex RT-PCR with the throughput of Next-Generation Sequencing (NGS) technology. By combining these methods, just 10-100 ng of total RNA is sufficient to quantify over 5 orders of magnitude variation in gene expression levels. Further, the use of targeted primers enables direct analysis of total RNA isolate and obviates the need for globin depletion from whole blood samples. Finally, using a defined set of amplicons to assess expression levels of all protein-coding genes facilitates and simplifies data analysis and allows more precise sample-to-sample normalization. We will present profiling results that demonstrate how this assay can be used to analyze the level of immune cell infiltration, assess intact and deficient immune mechanisms, and generally elucidate the tumor microenvironment of breast cancer samples.
Citation Format: Alex Chenchik, Andrey Komarov, Michael Makhanov, Sunitha Sastry, Costa Frangou. Targeted expression and molecular profiling assay for tumor microenvironment [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr B094.
- ©2016 American Association for Cancer Research.