The enzymatic activity of 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase is enhanced by NPM-ALK: new insights in ALK-mediated pathogenesis and the treatment of ALCL

Anaplastic large cell lymphoma (ALCL) represent a subset of neoplasms often caused by translocations that juxtapose the anaplastic lymphoma kinase (ALK) to a dimerization partner. The constitutive activation of ALK fusion proteins leads to cellular transformation through a complex signaling network. To further elucidate the ALK pathways sustaining lymphomagenesis and tumor maintenance, we analyzed the tyrosine-kinase protein profiles of ALK-positive cell lines using two complementary proteomic-based approaches, taking advantage of a specific ALK RNA interference (RNAi) or cell-permeable inhibitors. A well-defined set of ALK-associated tyrosine phospho-peptides, including metabolic enzymes, kinases, ribosomal and cytoskeletal proteins was identified. Validation studies confirmed that VASP and ATIC associated with NPM-ALK and their phosphorylation required ALK activity. ATIC phosphorylation was also documented in cell lines and primary tumors carrying ALK proteins and other tyrosine kinases, including TPR-Met and wild type c-Met. Functional analyses revealed that ALK-mediated ATIC phosphorylation enhanced its enzymatic activity, dampering the methotrexate-mediated transformylase activity inhibition. These findings demonstrate that alternative proteomic approaches in well-controlled experimental settings allow the definition of highly-informative proteomic profiles and the discovery of novel ALK-downstream players that contribute to the maintenance of the neoplastic phenotype. Prediction of tumor responses to methotrexate may justify the design of specific molecular-based chemotherapy.