Ibrutinib therapy downregulates AID enzyme and proliferative fractions in chronic lymphocytic leukemia

SIVINA, MARIELA; BERCA, CATALINA; DI NOIA, JAVIER M.; PABLO MORANDE; SEIJA NOÉ; LANDONI ANA INÉS; OPPEZZO PABLO; URIEPERO, ANGIMAR; FRESIA, PABLO; BURGER, JAN A.

Edimburgo

Workshop; XVIII International Workshop on Chronic Lymphocytic Leukaemia; 2019

iwCLL Consortium

Activation-induced cytidine deaminase (AID) initiates somatic hypermutation and class switchrecombination of the immunoglobulin genes. As a trade-off for its physiological function, AID alsocontributes to tumor development through its mutagenic activity. In chronic lymphocytic leukemia(CLL), AID is over-expressed in the proliferative fractions (PFs) of the malignant B lymphocytesand its anomalous expression has been associated with a clinical poor outcome. Recent preclinicaldata suggested that ibrutinib and idelalisib, two clinically approved kinase inhibitors, increase AIDexpression and genomic instability in normal and neoplastic B cells (Compagno M et al, Nature2017). These results raise concerns about a potential mutagenic risk in patients on long-term therapywith both drugs. To specifically corroborate if ibrutinib up-regulate AID enzyme during CLLtreatment we analyzed AID expression and PFs percentages in a CLL cohort before and duringibrutinib treatment. To further study the effect of ibrutinib on AID expression in CLL, we alsodeveloped in vitro primary cultures; used the MEC-1 cell line; and studied AKT/JAK1/STAT6pathway status by performing phospho-array analysis and Western blot technique.pathway status by performing phospho-array analysis and Western blot technique.MethodsCLL patients were enrolled in an approved clinical trial conducted in the University of Texas MDAnderson Cancer Center (NCT02007044) and treated with ibrutinib 420 mg daily. Peripheral bloodsamples were collected before and after initiation of ibrutinib therapy. MEC-1 cell line was a kindgift of Dr. Caligaris-Cappio. Flow cytometry, PCR and Western blot studies were developed usingprotocols detailed in Morande and Sivina et al, Blood, 2019. For phosphorylation profile studies,we used the AKT/PKB Phospho Antibody Array designed by Full Moon BioSystems For in vitrocultures, cells were incubated in RPMI Medium 1640 supplemented with 10% FBS in the presenceof ibrutinib. For statistical analysis, GraphPad Prism 7 software was used.ResultsOur analyses demonstrated that the distinct CLL PFs, CD19+/CD38+ (i); CD19+/CD86+ (ii);IgM+/IgG+ (iii) and CXCR4low/CD5high (iv), were significantly decreased after 4 weeks ofibrutinib treatment in patients (n=10, p