Detection of emerging ibrutinib resistance by flow cytometry in patients with chronic lymphocytic leukemia

Abstract

Background: Bruton ’ s tyrosine kinase inhibitor ibrutinib has revolutionized the treatment of chronic lymphocytic leukemia (CLL). Although ibrutinib is a highly effective drug, continuous treatment is required to maintain remission, which may lead to acquired ibrutinib resistance. Early detection of acquired resistance preceding clinical disease progression is an important issue. This is why our aim was to investigate several phenotypic markers on CLL cells to reveal changes in their expression during ibrutinib treatment in sensitive and clinically resistant patients. Materials and methods: In our study 28 (treatment naive, ibrutinib sensitive, clinically ibrutinib resistant) peripheral blood (PB), and 6 paired PB and bone marrow (BM) samples from CLL patients were examined. The expression of several surface markers (CD69, CD184, CD86, CD185, CD27) was assessed by flow cytometry in each sample. Furthermore, the presence of the BTK C481S resistance mutation was tested using digital droplet PCR (ddPCR) in samples from ibrutinib sensitive and resistant cases. In addition, we investigated the changes of CLL cells ’ phenotype during ibrutinib treatment in one patient with acquired ibrutinib resistance. Results: We found that the expression of CD27 decreased during ibrutinib therapy but increased again at the onset of clinical resistance. Expressions of CD69 and CD86 were also elevated at the onset of clinical ibrutinib resistance. Furthermore, the expression of CD86 showed correlation between PB and BM samples. Relapsed cases with high CD86 expression were positive for BTK C481S mutation. In addition, our prospective study showed that the increases in the expression of CD27, CD69 and CD86 were detectable up to several months before the onset of clinical resistance. Conclusion: Our research suggests that the flow cytometric measurements of certain markers, especially CD86, may predict development of ibrutinib resistance, however, confirmatory experiments are still required. Monitoring CD86 expression on peripheral blood CLL cells during ibrutinib treatment may become a potential new method to detect acquired ibrutinib resistance in the near future.