Key Points Ibrutinib is the first clinically viable irreversible ITK inhibitor. Ibrutinib inhibits the formation of Th2 but not Th1 immunity.

Aberrant DNA hypermethylation contributes to myeloid leukemogenesis by silencing structurally normal genes involved in hematopoiesis. MicroRNAs (miRNAs) are noncoding RNAs that regulate gene expression by targeting protein-coding mRNAs. Recently, miRNAs have been shown to play a role as both targets and effectors in gene hypermethylation and silencing in malignant cells. In the current study, we showed that enforced expression of miR-29b in acute myeloid leukemia cells resulted in marked reduction of the expression of DNA methyltransferases DNMT1, DNMT3A, and DNMT3B at both RNA and protein levels. This in turn led to decrease in global DNA methylation and reexpression of p15 INK4b and ESR1 via promoter DNA hypomethylation. Although down-regulation of DNMT3A and DNMT3B was the result of a direct interaction of miR-29b with the 3 untranslated regions of these genes, no predicted miR-29b interaction sites were found in the DNMT1 IntroductionDNA methylation consists of an enzymatic addition of a methyl group at the carbon 5 position of cytosine in the context of the sequence 5Ј-cytosine-guanosine (CpG) and is mediated by DNA methyltransferases (DNMTs). 1 The promoter regions of approximately 50% of human genes contain regions of DNA with a cytosine and guanine content greater than expected (so-called CpG islands) that, once hypermethylated, mediate gene transcriptional silencing. 2 Distinct roles in genomic methylation have been reported for DNMT isoforms. Whereas DNMT1 preferentially replicates already existing methylation patterns, DNMT3A and 3B are responsible for establishing de novo methylation. 2 Silencing of structurally normal tumor suppressor genes by aberrant DNA hypermethylation has been reported in hematologic malignancies, including subsets of acute myeloid leukemia (AML). 3,4 Although the mechanisms leading to aberrant DNA hypermethylation remain to be fully elucidated, increased levels of DNMT1 and DNMT3A and 3B have been observed in malignant myeloid blasts compared with normal bone marrow (BM) mononuclear cells (MNCs), suggesting that DNMT overexpression contributes to gene promoter hypermethylation and in turn to leukemogenesis. 4 Growing evidence supports a role for microRNAs (miRNAs) as both targets and effectors in aberrant mechanisms of DNA hypermethylation. 5,6 miRNAs are noncoding RNAs of 19 to 25 nucleotides in length that regulate gene expression by inducing translational inhibition or cleavage of their target mRNAs through base pairing at partially or fully complementary sites. 7 Several groups have shown that miRNAs are altered in human malignancies and can function as tumor suppressor genes or oncogenes through expression regulation of their target genes. 7 Similar to tumor suppressor genes, miRNAs with tumor suppressor activity are often located in deleted genomic areas or are silenced by mutations or promoter hypermethylation in malignant cells. 5,[8][9][10] Saito et al recently demonstrated that miR-127 is silenced by promoter DNA hypermethylation and down-regulated in human bladder ...

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BACKGROUND.Ibrutinib has been shown to have immunomodulatory effects by inhibiting Bruton's tyrosine kinase (BTK) and IL-2-inducible T cell kinase (ITK). The relative importance of inhibiting these 2 kinases has not been examined despite its relevance to immune-based therapies. METHODS.Peripheral blood mononuclear cells from chronic lymphocytic leukemia (CLL) patients on clinical trials of ibrutinib (BTK/ITK inhibitor; n = 19) or acalabrutinib (selective BTK inhibitor; n = 13) were collected serially. T cell phenotype, immune function, and CLL cell immunosuppressive capacity were evaluated. RESULTS. Ibrutinib markedly increased CD4+ and CD8 + T cell numbers in CLL patients. This effect was more prominent in effector/effector memory subsets and was not observed with acalabrutinib. Ex vivo studies demonstrated that this may be due to diminished activation-induced cell death through ITK inhibition. PD-1 and CTLA-4 expression was significantly markedly reduced in T cells by both agents. While the number of Treg cells remained unchanged, the ratio of these to conventional CD4 + T cells was reduced with ibrutinib, but not acalabrutinib. Both agents reduced expression of the immunosuppressive molecules CD200 and BTLA as well as IL-10 production by CLL cells. CONCLUSIONS. Ibrutinib treatment increased the in vivo persistence of activated T cells, decreased the Treg/CD4+ T cell ratio, and diminished the immune-suppressive properties of CLL cells through BTK-dependent and -independent mechanisms. These features provide a strong rationale for combination immunotherapy approaches with ibrutinib in CLL and other cancers. TRIAL REGISTRATION. ClinicalTrials.gov NCT01589302 and NCT02029443. Samples described here were collected per OSU-0025.

Key Points Ibrutinib treatment of CLL enhances the generation of CAR T cells for adoptive immunotherapy. Concurrent ibrutinib therapy improves the engraftment and therapeutic efficacy of anti-CD19 CAR T cells in mouse models.