A ternary association of KRas, EphA2, and integrins controls PDAC cell behavior.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers, primarily because late detection, a lack of effective therapies and an abundant desmoplastic reaction result in elevated proliferation, invasion, and immune evasion. KRas is the most frequently mutated driver gene in PDAC and, while drugs are now available that target this GTPase, there remains a need for further pharmacological options to treat the disease. As a first step to identify potential targets, this study aimed to define the molecular environment of KRas using proximity proteomics. A KRasG12V-biotin ligase chimera was stably expressed in several PDAC cell lines and organoids, and associating proteins identified following biotin addition, streptavidin affinity isolation, and mass spectrometry. Among the 126 proteins that were specifically associated in all cell lines, five integrin subunits and the receptor tyrosine kinase ephrin type-A receptor 2 (EphA2) were consistently enriched. Both classes of receptor have been previously implicated in both Ras signaling and desmoplastic responses. A range of protein biochemical and microscopic analyses confirmed the spatial association of integrin β1 and EphA2 with KRas. Cellular KRas levels were reduced following depletion of both integrin β1 and EphA2, and depletion of EphA2 inhibited KRas-dependent migration. Taken together, these findings highlight a ternary crosstalk between receptor tyrosine kinase, integrin, and KRas signaling, which provides insight into PDAC progression and signposts potential therapeutic vulnerabilities.