Activity-Based Proteomics Reveals Heterogeneous Kinome and ATP-Binding Proteome Responses to MEK Inhibition in KRAS Mutant Lung Cancer

Abstract

One way cancer cells can escape from targeted agents is through their ability to evade drug effects by rapidly rewiring signaling networks. Many protein classes, such as kinases and metabolic enzymes, are regulated by ATP binding and hydrolysis. We hypothesized that a system-level profiling of drug-induced alterations in ATP-binding proteomes could offer novel insights into adaptive responses. Here, we mapped global ATP-binding proteomes perturbed by two clinical MEK inhibitors, AZD6244 and MEK162, in KRAS mutant lung cancer cells as a model system harnessing a desthiobiotin-ATP probe coupled with LC-MS/MS. We observed strikingly unique ATP-binding proteome responses to MEK inhibition, which revealed heterogeneous drug-induced pathway signatures in each cell line. We also identified diverse kinome responses, indicating each cell adapts to MEK inhibition in unique ways. Despite the heterogeneity of kinome responses, decreased probe labeling of mitotic kinases and an increase of kinases linked to autophagy were identified to be common responses. Taken together, our study revealed a diversity of adaptive ATP-binding proteome and kinome responses to MEK inhibition in KRAS mutant lung cancer cells, and our study further demonstrated the utility of our approach to identify potential candidates of targetable ATP-binding enzymes involved in adaptive resistance and to develop rational drug combinations.

Proteomics Insights into Autophagy

“…The findings of the study by Kim et al. align with MEK‐inhibitors having been previously reported as inducing protective autophagy in KRAS mutant non‐small cell lung cancer (NSCLC), and suggests that upregulation of autophagy in KRAS mutant lung cancer may be kinase dependent . When validated, this may provide a reasonable basis for combination chemotherapy with kinase inhibitors.…”

“…Cells were treated with two MEK‐inhibitors, lysed, and the ATP‐binding kinome enriched, using a kinase enrichment kit, for subsequent digestion and LC‐MS/MS analysis. The study found that changes in the ATP‐binding proteome were cell type dependent and not KRAS mutation dependent . Furthermore, p53/LKB1 status was not determined to be a major factor in changes in the ATP‐binding proteome.…”

“…This implies that the sensitivity of the lung adenocarcinoma cells used in the study to erlotinib may be autophagy dependent . The phosphorylation action of kinases is known to be regulated by ATP binding and hydrolysis . Kinases are also thought to be involved in the ability of cancer cells to evade chemotherapy.…”

Activation of tumor suppressor protein PP2A inhibits KRAS-driven tumor growth

“…Small-molecule inhibitors that target protein kinases in the RAS pathway have been developed and are FDA approved (1). However, potential resistance to inhibition of KRAS signaling can involve mutations in targets or activation of compensatory mechanisms (28)(29)(30). Inhibiting multiple pathways concurrently with the MEK and AKT inhibitors, AZD6244 and MK2206, respectively, successfully abrogates tumor growth in preclinical models (31).…”

Approaches to identify kinase dependencies in cancer signalling networks

“…Chemical biology methods to measure kinase activation rely on the assumption that only active kinases may bind to either ATP or kinase inhibitors immobilised on beads (e.g. kinobeads and multiplex kinase inhibitor beads ).…”