High-throughput analysis of MAPK and JAK-STAT signaling in CT26 tumors using a combination of immunophenotyping and phospho-flow cytometry



David Draper, Alden Wong, Philip Lapinski, Sheri Barnes, Maryland Rosenfeld Franklin, and Scott Wise




AACR Annual Meeting

Introduction and Background

  • The development of robust platforms that enable phospho-protein-based analysis of cell signaling in the tumor microenvironment is pivotal for small molecule drug development. Currently lacking are reliable applications that can simultaneously measure multiple phospho-proteins in both tumor cells and immune cells ex vivo. This can be crucial because many signaling pathways are commonly involved in both pro- and anti-tumor responses in these two cellular compartments.
  • Covance has validated a phospho-flow cytometry-based platform that measures the phospho-protein levels of multiple signaling proteins in separate and distinct subsets simultaneously, which can include solid tumor-derived immune subsets and tumor cells.
  • The Covance phospho-flow platform can be applied to the analysis of treated cell lines and heterogeneous tissue-derived cell cultures in vitro as well as the ex vivo analysis of inflamed tissue from pharmacology studies.
  • In this presentation, we demonstrate how these services can be used to analyze MAPK and JAK-STAT pathway-associated kinases in vitro in cultured MV-4-11 acute myeloid leukemia (AML) cells and activated murine splenocytes, as well as ex vivo in both tumor-derived CD8+ T cells and tumor cells using the CT26 model for colorectal carcinoma.