Assessment of the primary cancer cell secretome using amino acid-analog labeling.

Abstract

It is well established that reciprocal communication between cancer cells and other cells in the tumor microenvironment plays a crucial role in cancer progression and therapy response. There are multiple ways by which cells communicate, including direct cell-cell contact and the secretion of soluble mediators. The secretome of cancer cells contains valuable information to disentangle the complex conversation that is happening between cancer cells and neighboring or distant cells such as immune cells, fibroblasts and endothelial cells. Here, we provide a workflow of mapping the cancer cell secretome in an unbiased way using amino acid-analog labeling in combination with mass spectrometry. The generation of single cells from fresh tumors, isolation of primary cancer cells from a complex multi-cellular pool, and the detection of newly synthesized proteins that are secreted into the medium is described in detailed protocols. Using this experimental pipeline the secretome of cancer cells across different tumors can be determined, paving the way to unravel cell-cell communication networks in the tumor microenvironment, which may uncover novel therapeutic targets.