Proteomics of resistance to Notch1 inhibition in acute lymphoblastic leukemia reveals targetable kinase signatures

Notch1 is a crucial oncogenic driver in T-cell acute lymphoblastic leukemia (T-ALL), making it an attractive therapeutic target. However, the success of targeted therapy using γ-secretase inhibitors (GSIs), small molecules blocking Notch cleavage and subsequent activation, has been limited due to development of resistance, thus restricting its clinical efficacy. Here, we systematically compare GSI resistant and sensitive cell states by quantitative mass spectrometry-based phosphoproteomics, using complementary models of resistance, including T-ALL patient-derived xenografts (PDX) models. Our datasets reveal common mechanisms of GSI resistance, including a distinct kinase signature that involves protein kinase C delta. We demonstrate that the PKC inhibitor sotrastaurin enhances the anti-leukemic activity of GSI in PDX models and completely abrogates the development of acquired GSI resistance in vitro. Overall, we highlight the potential of proteomics to dissect alterations in cellular signaling and identify druggable pathways in cancer. NOTCH1 is a driver of T-cell acute lymphoblastic leukemia that can be inhibited by γ-secretase inhibitors (GSIs), but their clinical efficacy is limited. Here, the authors compare the phosphoproteomes of GSI resistant and sensitive models, and identify potential kinase targets to overcome GSI resistance.