The hepatic niche leads to aggressive natural killer cell leukemia proliferation through the transferrin-transferrin receptor 1 axis.

Aggressive natural killer cell leukemia (ANKL) is a rare lymphoid neoplasm frequently associated with Epstein-Barr virus, with a disastrously poor prognosis. Owing to the lack of samples from patients with ANKL and relevant murine models, comprehensive investigation of its pathogenesis including the tumor microenvironment (TME) has been hindered. Here we established 3 xenograft mice derived from patients with ANKL (PDXs), which enabled extensive analysis of tumor cells and their TME. ANKL cells primarily engrafted and proliferated in the hepatic sinusoid. Hepatic ANKL cells were characterized by an enriched Myc-pathway and proliferated faster than those in other organs. Interactome analyses and in vivo CRISPR-Cas9 analyses revealed transferrin (Tf)-transferrin receptor 1 (TfR1) axis as a potential molecular interaction between the liver and ANKL. ANKL cells were rather vulnerable to iron deprivation. PPMX-T003, a humanized anti-TfR1 monoclonal antibody, showed remarkable therapeutic efficacy in a preclinical setting using ANKL-PDXs. These findings indicate that the liver, a noncanonical hematopoietic organ in adults, serves as a principal niche for ANKL and the inhibition of the Tf-TfR1 axis is a promising therapeutic strategy for ANKL.

Targeted insertion of conditional expression cassettes into the mouse genome using the modified i-PITT.

BACKGROUND: Transgenic (Tg) mice are widely used in biomedical research, and they are typically generated by injecting transgenic DNA cassettes into pronuclei of one-cell stage zygotes. Such animals often show unreliable expression of the transgenic DNA, one of the major reasons for which is random insertion of the transgenes. We previously developed a method called "pronuclear injection-based targeted transgenesis" (PITT), in which DNA constructs are directed to insert at pre-designated genomic loci. PITT was achieved by pre-installing so called landing pad sequences (such as heterotypic LoxP sites or attP sites) to create seed mice and then injecting Cre recombinase or PhiC31 integrase mRNAs along with a compatible donor plasmid into zygotes derived from the seed mice. PITT and its subsequent version, improved PITT (i-PITT), overcome disadvantages of conventional Tg mice such as lack of consistent and reliable expression of the cassettes among different Tg mouse lines, and the PITT approach is superior in terms of cost and labor. One of the limitations of PITT, particularly using Cre-mRNA, is that the approach cannot be used for insertion of conditional expression cassettes using Cre-LoxP site-specific recombination. This is because the LoxP sites in the donor plasmids intended for achieving conditional expression of the transgene will interfere with the PITT recombination reaction with LoxP sites in the landing pad. RESULTS: To enable the i-PITT method to insert a conditional expression cassette, we modified the approach by simultaneously using PhiC31o and FLPo mRNAs. We demonstrate the strategy by creating a model containing a conditional expression cassette at the Rosa26 locus with an efficiency of 13.7%. We also demonstrate that inclusion of FLPo mRNA excludes the insertion of vector backbones in the founder mice. CONCLUSIONS: Simultaneous use of PhiC31 and FLP in i-PITT approach allows insertion of donor plasmids containing Cre-loxP-based conditional expression cassettes.

Recent advances in extracellular vesicles in gastrointestinal cancer and lymphoma.

Extracellular vesicles (EVs) are intercellular communication agents that transfer microRNAs (miRNAs), other non-coding RNAs (ncRNAs), messenger RNAs (mRNAs), proteins, lipids, metabolites, and other molecules from donor cells (e.g., cancer cells) to recipient cells (e.g., stromal cells). In 2007, miRNAs were reported to be abundant among the ncRNAs present in EVs. Since then, many studies have investigated the functions of miRNAs and have attempted to apply these molecules to aid in the diagnosis and treatment of cancer. Research on EVs has expanded, particularly in the field of cancer, because cancer cells heavily secrete EVs. The cargo of these EVs, especially those in small EVs, such as exosomes, is assumed to work cooperatively and significantly in the tumor microenvironment and to promote metastasis. In this review, we first summarize recent studies on EVs in gastrointestinal cancer and highlight studies on human satellite II RNAs, which are a type of ncRNA found in EVs that possess repetitive sequences. Second, since several recent studies have revealed that phospholipids, which are components of EV membranes, play important roles in intercellular communication and the generation of lipid mediators in the tumor microenvironment, we review the reported roles of these molecules and discuss their potential use in the design of new cancer treatments.

Non-coding RNAs and lipids mediate the function of extracellular vesicles in cancer cross-talk.

Research on extracellular vesicles (EVs) has been expanded, especially in the field of cancer. The cargoes in EVs, especially those in small EVs such as exosomes include microRNAs (miRNAs), mRNA, proteins, and lipids, are assumed to work cooperatively in the tumor microenvironment. In 2007, it was reported that miRNAs were abundant among the non-coding RNAs present in exosomes. Since then, many studies have investigated the functions of miRNAs and have tried to apply these molecules to aid in the diagnosis of cancer. Accordingly, many reviews of non-coding RNAs in EVs have been published for miRNAs. This review focuses on relatively new cargoes, covering long noncoding (lnc) RNAs, circular RNAs, and repeat RNAs, among non-coding RNAs. These RNAs, regardless of EV or cell type, have newly emerged due to the innovation of sequencing technology. The poor conservation, low quantity, and technical difficulty in detecting these RNA types have made it difficult to elucidate their functions and expression patterns. We herein summarize a limited number of studies. Although lipids are major components of EVs, current research on EVs focuses on miRNA and protein biology, while the roles of lipids in exosomes have not drawn attention. However, several recent studies revealed that phospholipids, which are components of the EV membrane, play important roles in the intercommunication between cells and in the generation of lipid mediators. Here, we review the reported roles of these molecules, and describe their potential in cancer biology.

Proteomic and phospholipidomic characterization of extracellular vesicles inducing tumor microenvironment in Epstein-Barr virus-associated lymphomas.

Epstein-Barr virus (EBV) causes malignant carcinomas including B cell lymphomas accompanied by the systemic inflammation. Previously, we observed that phosphatidylserine (PS)-exposing subset of extracellular vesicles (EVs) secreted from an EBV strain Akata-transformed lymphoma (Akata EVs) convert surrounding phagocytes into tumor-associated macrophages (TAMs) via induction of inflammatory response, which is in part mediated by EBV-derived micro RNAs. However, it is still unclear about EV-carried other potential inflammatory factors associated with TAM formation in EBV lymphomas. To this end, we sought to explore proteomic and phospholipidomic profiles of PS-exposing EVs derived from EBV-transformed lymphomas. Mass spectrometric analysis revealed that several immunomodulatory proteins including integrin αLß2 and fibroblast growth factor 2 (FGF2) were highly expressed in PS-exposing Akata EVs compared with another EBV strain B95-8-transformed lymphoma-derived counterparts which significantly lack TAM-inducing ability. Pharmacological inhibition of either integrin αLß2 or FGF2 hampered cytokine induction in monocytic cultured cells elicited by PS-exposing Akata EVs, suggesting the involvement of these proteins in EV-mediated TAM induction in EBV lymphomas. In addition, phospholipids containing precursors of immunomodulatory lipid mediators were also enriched in PS-exposing Akata EVs compared with B95-8 counterparts. Phospholipidomic analysis of fractionated Akata EVs by density gradient centrifugation further demonstrated that PS-exposing Akata EVs might be identical to certain Akata EVs in low density fractions containing exosomes. Therefore, we concluded that a variety of immunomodulatory cargo molecules in a certain EV subtype are presumably conducive to the development of EBV lymphomas.

Anti-HBV drug entecavir ameliorates DSS-induced colitis through PD-L1 induction.

PD-L1-mediated signaling is one of the major processes that regulate local inflammatory responses in the gut. To date, protective effects against colitis through direct Fc-fused PD-L1 administration or indirect PD-L1 induction by probiotics have been reported. We have previously shown that the anti-HBV drug entecavir (ETV) induces PD-L1 expression in human hepatocytes. In the present study, we investigated whether ETV induces PD-L1 expression in intestinal cells and provides a protective effect against DSS-induced colitis. ETV induced PD-L1 expression in epithelial cells, rather than T and B cells, improving the symptoms of colitis. In the mechanistic analysis, Th17 cell differentiation was inhibited and B cell infiltration into the lamina propria was reduced. In addition, PD-L1 expression was positively correlated with Foxp3 or CSF1-R. In conclusion, ETV upregulated PD-L1 expression in epithelial cells and ameliorated inflammation in DSS-induced colitis. These results suggest that ETV may be a potential therapeutic agent as a PD-L1 enhancer for the treatment of human IBD.

Ceramide Metabolism Regulated by Sphingomyelin Synthase 2 Is Associated with Acquisition of Chemoresistance via Exosomes in Human Leukemia Cells.

Ceramide levels controlled by the sphingomyelin (SM) cycle have essential roles in cancer cell fate through the regulation of cell proliferation, death, metastasis, and drug resistance. Recent studies suggest that exosomes confer cancer malignancy. However, the relationship between ceramide metabolism and exosome-mediated cancer malignancy is unclear. In this study, we elucidated the role of ceramide metabolism via the SM cycle in exosomes and drug resistance in human leukemia HL-60 and adriamycin-resistant HL-60/ADR cells. HL-60/ADR cells showed significantly increased exosome production and release compared with parental chemosensitive HL-60 cells. In HL-60/ADR cells, increased SM synthase (SMS) activity reduced ceramide levels, although released exosomes exhibited a high ceramide ratio in both HL-60- and HL-60/ADR-derived exosomes. Overexpression of SMS2 but not SMS1 suppressed intracellular ceramide levels and accelerated exosome production and release in HL-60 cells. Notably, HL-60/ADR exosomes conferred cell proliferation and doxorubicin resistance properties to HL-60 cells. Finally, microRNA analysis in HL-60 and HL-60/ADR cells and exosomes showed that miR-484 elevation in HL-60/ADR cells and exosomes was associated with exosome-mediated cell proliferation. This suggests that intracellular ceramide metabolism by SMS2 regulates exosome production and release, leading to acquisition of drug resistance and enhanced cell proliferation in leukemia cells.

The RNA-Binding Protein ELAVL1 Regulates Hepatitis B Virus Replication and Growth of Hepatocellular Carcinoma Cells.

Previous RNA immunoprecipitation followed by proteomic approaches successfully demonstrated that Embryonic Lethal, Abnormal Vision, Drosophila-Like 1 (ELAVL1) interacts with hepatitis B virus (HBV)-derived RNAs. Although ELAVL family proteins stabilize AU-rich element (ARE)-containing mRNAs, their role in HBV transcription remains unclear. This study conducted loss-of-function assays of ELAVL1 for inducible HBV-replicating HepAD38 cells and HBx-overexpressed HepG2 cells. In addition, clinicopathological analyses in primary hepatocellular carcinoma (HCC) surgical samples were also conducted. Lentivirus-mediated short hairpin RNA knockdown of ELAVL1 resulted in a decrease in both viral RNA transcription and production of viral proteins, including HBs and HBx, probably due to RNA stabilization by ELAVL1. Cell growth of HepAD38 cells was more significantly impaired in ELAVL1-knockdown than those in the control group, with or without HBV replication, indicating that ELAVL1 is involved in proliferation by factors other than HBV-derived RNAs. Immunohistochemical analyses of 77 paired HCC surgical specimens demonstrated that diffuse ELAVL1 expression was detected more frequently in HCC tissues (61.0%) than in non-tumor tissues (27.3%). In addition, the abundant expression of ELAVL1 tended to affect postoperative recurrence in HBV-related HCC patients. In conclusion, ELAVL1 contributes not only to HBV replication but also to HCC cell growth. It may be a potent therapeutic target for HBV-related HCC treatment.

Extracellular vesicles secreted by HBV-infected cells modulate HBV persistence in hydrodynamic HBV transfection mouse model.

Hepatitis B, a viral infection that affects the liver, is thought to affect over 257 million people worldwide, and long-term infection can lead to life-threatening issues such as cirrhosis or liver cancer. Chronic hepatitis B develops by the interaction between hepatitis B virus (HBV) and host immune response. However, questions of how HBV-infected cells thwart immune system defenses remain unanswered. Extracellular vesicles (EVs) are used for cellular communication, carrying cargoes such as RNAs, proteins, and lipids and delivering them intracellularly after being endocytosed by target cells. HBV-infected liver cells secrete several types of EVs into body fluids such as complete and incomplete virions, and exosomes. We previously demonstrated that monocytes that incorporated EVs moved to immunoregulatory phenotypes via up-regulation of PD-L1, an immunocheckpoint molecule, and down-regulation of CD69, a leukocyte activation molecule. In this study, we transfected mice with HBV using hydrodynamic injection and studied the effects of EVs secreted by HBV-infected liver cells. EVs secreted from cells with HBV replication strongly suppressed the immune response, inhibiting the eradication of HBV-replicating cells in the mice transfected with HBV. EVs were systemically incorporated in multiple organs, including liver, bone marrow (BM), and intestine. Intriguingly, the BM cells that incorporated EVs acquired intestinal tropism and the dendritic cell populations in the intestine increased. These findings suggest that the EVs secreted by HBV-infected liver cells exert immunosuppressive functions, and that an association between the liver, bone marrow, and intestinal tract exists through EVs secreted from HBV-infected cells.

PD-L1 is induced on the hepatocyte surface via CKLF-like MARVEL transmembrane domain-containing protein 6 up-regulation by the anti-HBV drug Entecavir.

Chronic hepatitis B is now controllable when treated with nucleoside reverse transcriptase inhibitors (NRTIs), which inhibit hepatitis B virus (HBV) replication. However, once the NRTIs are discontinued, most patients relapse, necessitating lifelong NRTIs treatment. HBV infection relapse is assumed to be caused by the persistent existence of covalently closed circular DNA (cccDNA) in the nuclei of infected hepatocytes. The mechanism by which cccDNA-positive hepatocytes escape immune surveillance during NRTIs treatment remains elusive. Entecavir (ETV), a commonly used NRTI, post-transcriptionally up-regulates programmed cell death-ligand 1 (PD-L1), an immune checkpoint molecule, on the cell surface of hepatocytes regardless of HBV infection. Up-regulation by ETV depends on up-regulation of CKLF-like MARVEL transmembrane domain-containing 6, a newly identified potent regulator of PD-L1 expression on the cell surface. ETV-treated hepatic cells suppressed the activity of primary CD3 T cells and programmed cell death protein-1 (PD-1)-over-expressed Jurkat cells. Finally, ETV induces PD-L1 in primary hepatocytes infected by HBV. These results provide evidence that ETV considerably up-regulates PD-L1 on the cell surface of infected hepatocytes, which may be one of the mechanisms by which infected hepatocytes subvert immune surveillance.

Role of exosomes as a proinflammatory mediator in the development of EBV-associated lymphoma.

Epstein-Barr virus (EBV) causes various diseases in the elderly, including B-cell lymphoma such as Hodgkin's lymphoma and diffuse large B-cell lymphoma. Here, we show that EBV acts in trans on noninfected macrophages in the tumor through exosome secretion and augments the development of lymphomas. In a humanized mouse model, the different formation of lymphoproliferative disease (LPD) between 2 EBV strains (Akata and B95-8) was evident. Furthermore, injection of Akata-derived exosomes affected LPD severity, possibly through the regulation of macrophage phenotype in vivo. Exosomes collected from Akata-lymphoblastoid cell lines reportedly contain EBV-derived noncoding RNAs such as BamHI fragment A rightward transcript (BART) micro-RNAs (miRNAs) and EBV-encoded RNA. We focused on the exosome-mediated delivery of BART miRNAs. In vitro, BART miRNAs could induce the immune regulatory phenotype in macrophages characterized by the gene expressions of interleukin 10, tumor necrosis factor-α, and arginase 1, suggesting the immune regulatory role of BART miRNAs. The expression level of an EBV-encoded miRNA was strongly linked to the clinical outcomes in elderly patients with diffuse large B-cell lymphoma. These results implicate BART miRNAs as 1 of the factors regulating the severity of lymphoproliferative disease and as a diagnostic marker for EBV+ B-cell lymphoma.

Heat-Killed Fusobacterium nucleatum Triggers Varying Heme-Related Inflammatory and Stress Responses Depending on Primary Human Respiratory Epithelial Cell Type.

Fusobacterium nucleatum (Fn) is generally an opportunistic oral pathogen that adheres to mammalian mucosal sites, triggering a host inflammatory response. In general, Fn is normally found within the human oral cavity; however, it was previously reported that Fn is a risk factor for certain respiratory diseases. Surprisingly, this was never fully elucidated. Here, we investigated the virulence potential of heat-killed Fn on primary human tracheal, bronchial, and alveolar epithelial cells. In this study, we measured the secretion of inflammatory- (IL-8 and IL-6), stress- (total heme and hydrogen peroxide), and cell death-related (caspase-1 and caspase-3) signals. We established that the inflammatory response mechanism varies in each epithelial cell type: (1) along tracheal cells, possible Fn adherence would trigger increased heme secretion and regulated inflammatory response; (2) along bronchial cells, potential Fn adherence would simultaneously initiate an increase in secreted H2O2 and inflammatory response (ascribable to decreased secreted heme amounts); and (3) along alveolar cells, putative Fn adherence would instigate the increased secretion of inflammatory responses attributable to a decrease in secreted heme levels. Moreover, regardless of the epithelial cell-specific inflammatory mechanism, we believe these are putative, not harmful. Taken together, we propose that any potential Fn-driven inflammation along the respiratory tract would be initiated by differing epithelial cell-specific inflammatory mechanisms that are collectively dependent on secreted heme.

[Exosomes in the hematopoietic system].

The secretion of extracellular vesicles (EVs) from cells has been observed. Recently, because EVs were found to contain functional molecules such as micro RNAs (miRNAs) and possess the ability to transfer them to other cells, its functions were expanded as an "intracellular communicator." The exosome is one such EV that has been extensively investigated, particularly in cancer research because cancer cells abundantly secrete exosomes, suggesting their potential as promising diagnostic markers. Research on exosomes in the hematopoietic system has just begun. We recently reported that the exosome secreted from the EBV-infected lymphoma cells has critical functions in lymphomagenesis and maintenance. Moreover, EVs in HBV infection are now being investigated to generalize their functions.

miRNAs in Normal and Malignant Hematopoiesis.

Lineage specification is primarily regulated at the transcriptional level and lineage-specific transcription factors determine cell fates. MicroRNAs (miRNAs) are 18-24 nucleotide-long non-coding RNAs that post-transcriptionally decrease the translation of target mRNAs and are essential for many cellular functions. miRNAs also regulate lineage specification during hematopoiesis. This review highlights the roles of miRNAs in B-cell development and malignancies, and discusses how miRNA expression profiles correlate with disease prognoses and phenotypes. We also discuss the potential for miRNAs as therapeutic targets and diagnostic tools for B-cell malignancies.

A single miRNA complements transcription factor deficiency through the TGF pathway at time of B-cell lineage determination.

Cell fate and lineage are primarily regulated at the transcriptional level. However, the transcriptional level alone does not appear to control all aspects of cellular functioning, suggesting the presence of other, as-yet-unknown, mechanisms. miR-126 induced B-cell differentiation in MLL-AF4 acute lymphoblastic leukemia (ALL) without upregulation of TCF3/E2A, EBF1, and PAX5. Early B-cell factor 1 (EBF1) which are critical transcription factor involved in B lymphopoiesis. To challenge the conventional wisdom that cell fate is solely governed by transcription factors, we investigated whether microRNAs (miRNAs) could induce complete B-cell lineage commitment in cells lacking EBF1. miR-126 upregulated B220 in EBF1-deficient hematopoietic progenitor cells (HPCs). Moreover, miRNA-195 (miR-195) induced CD19 expression in EBF1-deficient HPCs, suggesting that these cells were committed to the B-cell lineage. The functional target genes of miR-195 involved in this process belonged to the TGF beta family, a potent inhibitor of B-cell differentiation. These results suggest that some miRNAs could function as alternatives to transcription factors.

Novel functional small RNAs are selectively loaded onto mammalian Ago1.

Argonaute (Ago) proteins function in RNA silencing as components of the RNA-induced silencing complex (RISC). In lower organisms, the small interfering RNA and miRNA pathways diverge due in part to sorting mechanisms that direct distinct small RNA (sRNA) duplexes onto specific Ago-RISCs. However, such sorting mechanisms appear to be lost in mammals. miRNAs appear not to distinguish among Ago1-4. To determine the effect of viral infection on the sorting system, we compared the content of deep-sequenced RNA extracted from immunoprecipitation experiments with the Ago1 and Ago2 proteins using Epstein-Barr virus (EBV)-infected cells. Consistent with previous observations, sequence tags derived from miRNA loci in EBV and humans globally associate in approximately equivalent amounts with Ago1 and Ago2. Interestingly, additional sRNAs, which have not been registered as miRNAs, were associated with Ago1. Among them, some unique sequence tags derived from tandem loci in the human genome associate exclusively with Ago1 but not, or rarely, with Ago2. This is supported by the observation that the expression of the unique sRNAs in the cells is highly dependent on Ago1 proteins. When we knocked down Ago1, the expression of the Ago1-specific sRNAs decreased dramatically. Most importantly, the Ago1-specific sRNAs bound to mRNAs and regulated target genes and were dramatically upregulated, depending on the EBV life cycle. Therefore, even in mammals, the sorting mechanism in the Ago1-4 family is functional. Moreover, the existence of Ago1-specific sRNAs implies vital roles in some aspects of mammalian biology.

MicroRNA-126-mediated control of cell fate in B-cell myeloid progenitors as a potential alternative to transcriptional factors.

Lineage specification is thought to be largely regulated at the level of transcription, where lineage-specific transcription factors drive specific cell fates. MicroRNAs (miR), vital to many cell functions, act posttranscriptionally to decrease the expression of target mRNAs. MLL-AF4 acute lymphocytic leukemia exhibits both myeloid and B-cell surface markers, suggesting that the transformed cells are B-cell myeloid progenitor cells. Through gain- and loss-of-function experiments, we demonstrated that microRNA 126 (miR-126) drives B-cell myeloid biphenotypic leukemia differentiation toward B cells without changing expression of E2A immunoglobulin enhancer-binding factor E12/E47 (E2A), early B-cell factor 1 (EBF1), or paired box protein 5, which are critical transcription factors in B-lymphopoiesis. Similar induction of B-cell differentiation by miR-126 was observed in normal hematopoietic cells in vitro and in vivo in uncommitted murine c-Kit(+)Sca1(+)Lineage(-) cells, with insulin regulatory subunit-1 acting as a target of miR-126. Importantly, in EBF1-deficient hematopoietic progenitor cells, which fail to differentiate into B cells, miR-126 significantly up-regulated B220, and induced the expression of B-cell genes, including recombination activating genes-1/2 and CD79a/b. These data suggest that miR-126 can at least partly rescue B-cell development independently of EBF1. These experiments show that miR-126 regulates myeloid vs. B-cell fate through an alternative machinery, establishing the critical role of miRNAs in the lineage specification of multipotent mammalian cells.

[Pathogenesis and novel therapy for EBV-related B-cell lymphoma].

Epstein-Barr virus (EBV), a type of γ-herpes virus, is known to be a tumor virus. About 90% of adults were found to be persistently infected with EBV and this infection is responsible for Burkitt lymphoma (BL), extranodal NK/T cell lymphoma, Hodgkin lymphoma (HL), acquired Immune deficiency syndrome (AIDS)-associated lymphoma, and a portion of diffuse large B cell lymphomas (DLBCL). EBV-positive DLBCL in the elderly, a disease recognized in Japan, is described in the WHO classification as a new category of DLBCLs. Clinical studies of DLBCLs have since accumulated. We herein describe our clinicopathological study of EBV-positive DLBCL in the elderly in the rituximab era, and review EBV-positive B cell lymphoma cases. A potentially promising novel therapy for EBV-positive B cell lymphoma, anti-PD-1 antibody, is then introduced. Finally, we briefly discuss our unpublished study of EBV-positive B cell lymphoma and its microenvironment.

In vivo leukemogenic potential of an interleukin 7 receptor α chain mutant in hematopoietic stem and progenitor cells.

Somatic gain-of-function mutations in interleukin 7 receptor α chain (IL7Rα) have been described in pediatric T and B acute lymphoblastic leukemias (T/B-ALLs). Most of these mutations are in-frame insertions in the extracellular juxtamembrane-transmembrane region. By using a similar mutant, a heterozygous in-frame transmembrane insertional mutation (INS), we validated leukemogenic potential in murine hematopoietic stem/progenitor cells, using a syngeneic transplantation model. We found that ectopic expression of INS alone in hematopoietic stem/progenitor cells caused myeloproliferative disorders, whereas expression of INS in combination with a Notch1 mutant led to the development of much more aggressive T-ALL than with wild-type IL7Rα. Furthermore, forced expression of INS in common lymphoid progenitors led to the development of mature B-cell ALL/lymphoma. These results demonstrated that INS has significant in vivo leukemogenic activity and that the lineage of the resulting leukemia depends on the developmental stage in which INS occurs, and/or concurrent mutations.

Imatinib mesylate directly impairs class switch recombination through down-regulation of AID: its potential efficacy as an AID suppressor.

Activation-induced cytidine deaminase (AID) is essential for class switch recombination and somatic hypermutation. Its deregulated expression acts as a genomic mutator that can contribute to the development of various malignancies. During treatment with imatinib mesylate (IM), patients with chronic myeloid leukemia often develop hypogammaglobulinemia, the mechanism of which has not yet been clarified. Here, we provide evidence that class switch recombination on B-cell activation is apparently inhibited by IM through down-regulation of AID. Furthermore, expression of E2A, a key transcription factor for AID induction, was markedly suppressed by IM. These results elucidate not only the underlying mechanism of IM-induced hypogammaglobulinemia but also its potential efficacy as an AID suppressor.