Study of the metabolic reprogramming of chronic lymphocytic leukaemia cells in an ex-vivo model of the lymphatic niche
Chronic Lymphocytic Leukemia (CLL), the most prevalent leukemia in adults, is marked by a monoclonal accumulation of tumor and quiescent mature B lymphocytes in the blood. In advanced stages (Binet stage C), marrow invasion and lymphocyte proliferation occurs in secondary lymphoid organs, resulting in adenopathy. The primary complication is the transformation into high-grade lymphoma (Richter's lymphoma), which is detectable by positron emission tomography (PET) and reveals increased binding of a glucose derivative (18FDG) by cancer cells. Our data from the stage C CLL patient cohort at the University Hospital of Nice indicate hyperfixation in lymph node regions, suggesting that the tumor microenvironment promotes proliferation and metabolic changes in CLL cells, which exhibit considerable resistance to current therapies.
Developing an ex-vivo tumoroid model that simulates the lymph node microenvironment is crucial for identifying and testing new therapeutic targets because many patients experience relapse or are refractory to existing treatments. Our objectives were: i) to create an ex-vivo cell co-culture model mimicking the lymph node, ii) to metabolically characterize this resistant lymph node, and iii) to assess the efficacy of an antimetabolic therapy along with current treatment (Ibrutinib).
We established an ex-vivo model of the lymphatic niche using a co-culture of peripheral blood mononuclear cells (PBMC) from CLL patients, stimulated with CpG ODN (a TLR9 agonist) and IL2 (activating CD25 on B cell surfaces), and primary human lymph node fibroblasts (HLF). This novel co-culture model replicates the lymphatic niche of CLL patients ex-vivo. Using biochemical and flow cytometry techniques, we validated this model, which demonstrated increased viability, proliferation, and antiapoptotic proteins MCL1, and BCL2 at 48h of co-culture. The model also revealed elevated lactate production, glucose consumption, and metabolic activation detected by the Seahorse analyzer, along with the production of glycolytic enzymes like hexokinase 2 (HK2).
HK 2 inhibition disrupts the glycolytic metabolic pathway, which is essential for CLL cell energy supply in co-culture, reducing energy availability for CLL cell survival and heightening vulnerability to treatment. Moreover, glucose flux disruption may affect intracellular signaling pathways and biological processes related to CLL progression.
The efficacy of antimetabolic therapy targeting HK 2, combined with current treatments like ibrutinib presents new opportunities to enhance clinical outcomes for patients with relapsed or refractory CLL. The promising results from our ex-vivo lymph node model could serve as a foundation for developing novel targeted therapeutic strategies aimed at disrupting CLL cell energy metabolism and overcoming resistance to existing treatments.
Metabolism, CLL, microenvironment, lymphatic niche, ex-vivo model.
Dr Patrick AUBERGER, DR Inserm, C3M, Université Côte d’Azur
Pr Myriam LABALETTE, PU-PH, INFINITE - U1286 Faculté de Médecine, Université de Lille
Pr Fanny BARAN-MARSZAK, PU-PH, Université Sorbonne Paris Nord-UFR SMBH
Pr Loïc YSEBAERT, PU-PH, Centre de Recherches en Cancérologie de Toulouse (CRCT), Université Paul Sabatier
Thesis supervisor :
Dr Marcel DECKERT, DR Inserm, C3M, Université Côte d’Azur
Invited members :
Dr Ghislaine BERNARD, MCU, Université Côte d’Azur
Pr Thomas CLUZEAU, PU-PH, Université Côte d’Azur