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Figure S2. Global tissue expression profiles of selected kinases.

Table S1. Mutational status of all specimens regarding KRAS and EGFR.

Legends for Supplementary Figures and Tables

Table S4. Comparison of dasatinib and sunitinib targets between NSCLC, leukemia and melanoma cell lines.

Table S7. Mass spectrometry data of phosphoproteomics experiments with eluates from c-dasatinib and c-sunitinib affinity chromatography eluates across all samples from primary NSCLC tissues, cell lines and cell line-derived mouse xenografts.

Figure S1. Cross-docking of sunitinib and dasatinib in NQO2 and PDXK.

Table S2. Mass spectrometry data of chemical proteomics experiments with c-dasatinib and c-sunitinib in primary NSCLC tissue samples.

A growing number of gene mutations, which are recognized as cancer drivers, can be successfully targeted with drugs. The redundant and dynamic nature of oncogenic signaling networks and complex interactions between cancer cells and the microenvironment, however, can cause drug resistance. While these challenges can be addressed by developing drug combinations or polypharmacology drugs, this benefits greatly from a detailed understanding of the proteome-wide target profiles. Using mass spectrometry-based chemical proteomics, we report the comprehensive characterization of the drug–protein interaction networks for the multikinase inhibitors dasatinib and sunitinib in primary lung cancer tissue specimens derived from patients. We observed in excess of 100 protein kinase targets plus various protein complexes involving, for instance, AMPK, TBK1 (sunitinib), and ILK (dasatinib). Importantly, comparison with lung cancer cell lines and mouse xenografts thereof showed that most targets were shared between cell lines and tissues. Several targets, however, were only present in tumor tissues. In xenografts, most of these proteins were of mouse origin suggesting that they originate from the tumor microenvironment. Furthermore, intersection with subsequent global phosphoproteomic analysis identified several activated signaling pathways. These included MAPK, immune, and integrin signaling, which were affected by these drugs in both cancer cells and the microenvironment. Thus, the combination of chemical and phosphoproteomics can generate a systems view of proteins, complexes, and signaling pathways that are simultaneously engaged by multitargeted drugs in cancer cells and the tumor microenvironment. This may allow for the design of novel anticancer therapies that concurrently target multiple tumor compartments. Mol Cancer Ther; 13(11); 2751–62. ©2014 AACR.

Table S3. Mass spectrometry data of chemical proteomics experiments with c-dasatinib and c-sunitinib in H23 and H292 cell line samples.

Table S8. Pathway analysis of all identified phosphoproteins enriched by c-dasatinib and c-sunitinib affinity chromatography.

Table S5. Mass spectrometry data of chemical proteomics experiments with c-dasatinib and c-sunitinib in mouse xenograft samples of H23 and H292 cell lines.

Table S9. Comparison of dNSAF label-free quantitation values from chemical proteomics experiments in NSCLC cells with in vitro kinase assay parameters.