Ps1125 linking microenvironmental signals to metabolic switches and ibrutinib response in chronic lymphocytic leukemia

Background: Chronic lymphocytic leukemia (CLL) is a clinically heterogeneous B cell malignancy, with accumulation of clonal CD5+ B cells in blood, lymph nodes (LN) and bone marrow. CLL has become a paradigm for a cancer that depends on signals from the microenvironment. This is evidenced by the clinical success of the Btk inhibitor ibrutinib, which drives CLL cells out of the LN to eventually die by neglect. In peripheral blood, CLL cells are quiescent, non‐cycling, with low metabolic activity. In contrast, in LN CLL cells receive various signals from surrounding cells, crucial for proliferation, and protection from chemotherapeutic drugs. These acquired advantages are possibly linked with altered metabolism, but it is unclear if and how CLL cells change their metabolism in the LN environment. Aims: We aimed to investigate the altered metabolism of CLL cells in the tumor microenvironment, and what signals determine these changes. Methods: Glucose uptake, glycolysis, mitochondrial mass, oxidative phosphorylation and lipid uptake were measured by flow cytometer or Seahorse XF96 analyzer, and 90 metabolites were analyzed by LC mass‐spectrometry. Results: We first established that the metabolic activity profile of circulating recent LN emigrants (CXCR4 high /CD5 dim PB lymphocytes) resembled that of actual LN residents (obtained by a core biopsy), which showed both higher mitochondrial mass and glucose uptake than paired peripheral blood CLL cells. In vitro stimulation of isolated blood‐derived CLL samples demonstrated that CD40 but not BCR signaling recapitulate these metabolic changes. Next, 10 patient samples were stimulated with/without CD40L for 48 hours and analyzed by mass‐spec for metabolic intermediates. To draw a further parallel with the situation in LN, metabolomics was performed on 13 patients sampled before and after 3 months of ibrutinib treatment. Combined, the data shows that various metabolic pathways are activated in the tumor microenvironment, particularly TCA cycle, pyruvate metabolism, glycolysis, and fatty acid metabolism. Apart from these overall changes, the highest ranking shifts in metabolites point to involvement of amino acids to fuel the TCA cycle. Moreover, aberrant patterns in metabolic changes after ibrutinib were noted in some patients, possibly linked with clinical response. Summary/Conclusion: CD40 signaling leads to increased oxidative phosphorylation and glycolysis in the CLL microenvironment. Opposite changes were observed in ibrutinib treated samples, together providing indirect insight into the metabolism in the CLL LN.