Abstract. Multidrug resistance (MDR) remains the major cause of disease relapse and poor prognosis in adults with acute myeloid leukemia (AML). Emerging evidence shows that drug resistance not only exists against conventional chemotherapeutic drugs, but also limits the efficacy of new biological agents. Therefore, it is important to elucidate the mechanisms through which AML patients develop drug resistance. MicroRNAs have been shown to play an important role in regulating the chemotherapy resistance in AML. A detailed understanding of the mechanisms of microRNA that are clinically relevant in AML may enhance our ability to predict and overcome drug resistance. Here, we demonstrated, for the first time, that miR-181b was decreased significantly in human multidrug-resistant leukemia cells and relapsed/refractory AML patient samples. Overexpression of miR-181b increased the sensitivity of leukemia cells to cytotoxic chemotherapeutic agents and promoted drug-induced apoptosis. Moreover, miR-181b inhibited HMGB1 and Mcl-1 expression by direct binding to their 3'-untranslated regions. In addition, HMGB1 was expressed at high levels in relapsed/refractory AML patients and suppression of HMGB1 via RNA interference sensitized multidrug-resistant leukemia cells to chemotherapy and induced apoptosis. In conclusion, these results provide a strong rationale for the development of miR-181b-based therapeutic strategies for the enhancement of efficacy in AML treatment.

Acute myeloid leukemia (AML), caused by abnormal proliferation and accumulation of hematopoietic progenitor cells, is one of the most common malignancies in adults. We reported here DYRK1A expression level was reduced in the bone marrow of adult AML patients, comparing to normal controls. Overexpression of DYRK1A inhibited the proliferation of AML cell lines by increasing the proportion of cells undergoing G0/G1 phase. We reasoned that the proliferative inhibition was due to downregulation of c-Myc by DYRK1A, through mediating its degradation. Moreover, overexpression of c-Myc markedly reversed AML cell growth inhibition induced by DYRK1A. DYRK1A also had significantly lower expression in relapsed/refractory AML patients, comparing to newly-diagnosed AML patients, which indicated the role of DYRK1A in chemoresistance of AML. Our study provided functional evidences for DYRK1A as a potential tumor suppressor in AML.

As the pathophysiology of acute myelogenous leukemia (AML) involves a block of myeloid maturation, a desirable therapeutic strategy is to induce leukemic cell maturation to increase the efficacy and to avoid the side effects of traditional chemotherapeutics. Through a compound library screen, 6-benzylthioinosine (6BT) was identified as a promising differentiation-inducing agent. 6BT induces monocytic differentiation of myeloid leukemia cell lines such as HL-60 and OCI-AML3, as well as primary patient samples as evidenced by morphology, immunophenotyping, and nitroblue tetrazolium reduction. Not only can 6BT induce differentiation but a subset of AML cell lines such as MV4-11 and HNT34 instead undergo 6BT-mediated cell death. Despite inducing cell death in some leukemic cells, 6BT exhibits extremely low toxicity on several nonmalignant cells such as fibroblasts, normal bone marrow, and endothelial cells. This toxicity profile may relate to the function of 6BT as an inhibitor of the nucleoside transporter, ent1, which is thought to prevent it from entering many cell types. In contrast, 6BT likely enters at least some leukemic cell lines as shown by its requirement for phosphorylation for its differentiation activity. 6BT is also able to synergize with currently used myeloid differentiation agents such as ATRA and decitabine. Early studies indicate that the mechanism of action of this compound may involve ATP depletion that leads to growth inhibition and subsequent differentiation. Besides in vitro activity, 6BT also shows the ability to impair HL-60 and MV4-11 tumor growth in nude mice. 6BT is a promising new monocytic differentiation agent with apparent leukemic cell-specific activity. [Cancer Res 2008;68(11):4369-76]

Notch and epidermal growth factor receptor (EGFR) signaling play critical roles in cell proliferation, differentiation, and apoptosis, and thereby may contribute to the development of breast cancer. We constitutively overexpressed active Notch1 in human breast cancer cells to explore the consequences of Notch1 signaling on cell growth and to investigate the underlying molecular mechanisms. We found that EGFR expression was increased. Then, using EGFR inhibitor, we found it exhibited an inhibitory role on human breast cancer cells. Overexpression of Notch1 could reverse EGFR inhibitor-induced cell toxicity, suggesting that Notch and EGFR signaling may be positively cross-linked in human breast cancer.

Notch signaling plays a critical role in embryonic vascular development and tumor angiogenesis. The present study was conducted to investigate the prognostic role of the angiogenesis-related Notch ligand and the receptor in acute myeloid leukemia (AML) and assess whether their expression correlates with that of the vascular endothelial growth factor (VEGF) and angiopoietin (Ang)-2. Bone marrow mononuclear cells from 60 untreated AML patients and 40 healthy controls were obtained. Real-time RT-PCR was performed to evaluate the mRNA expression of δ-like ligand 4 (Dll4), Notch1, VEGF, VEGF receptor (VEGFR)-1, VEGFR-2, Ang-1, Ang-2 and Tie2. Western blot analysis was used to determine the protein levels of Dll4 and Notch1. The results demonstrated that Dll4, Notch1, VEGF, VEGFR-2 and Ang-2 expression were significantly higher in untreated AML patients than in the controls. Univariate analysis of factors associated with the overall survival showed a significantly shorter survival in patients with the unfavorable karyotype, higher Dll4 expression, higher Notch1 expression, higher VEGF expression or higher Ang-2 expression. Furthermore, multivariate analysis revealed that the karyotype and expression levels of Notch1, Dll4, VEGF and Ang-2 were independent prognostic factors for overall survival. Additionally, the prognostic value of Dll4 expression (but not Notch1) was more significant in the subgroup consisting of patients with intermediate-risk cytogenetics. Subgroup analysis showed that Notch1 and Dll4 expression levels had a prognostic impact on patients with high VEGF or Ang-2 levels. Taken together, our data provide evidence that the activation of the Notch pathway may indicate an unfavorable prognosis in AML. In particular, Dll4 may be a relevant prognostic marker in intermediate-risk AML.

Inspite of effective treatment with imatinib (IM), chronic myeloid leukemia (CML) is still an incurable disease. Some patients became refractory because of IM resistance. Bone marrow mesenchymal stem cells (BMSCs) have been implicated a role in promoting CML cells' resistance against IM treatment. The detailed molecular mechanisms, however, remain largely unknown. In this study, we found that BMSCs increased the expression of FZD7 and activated Wnt/β-catenin signaling pathway in CML cells. BMSCs from CML patients showed increased efficiency to accelerate CML cell proliferation, enhance the drug resistance of K562 cells and up-regulate the expression of FZD7. Antagonism of FZD7 expression by shRNA significantly suppressed proliferation and increased IM sensitivity of CML cells co-cultured with BMSCs cells. Our findings suggest that FZD7, involved in canonical Wnt signaling pathway, plays a critical role in mediating BMSCs-dependent protection of CML cells, and potentially provide a novel therapeutic target for CML.

Background: Acute myeloid leukemia (AML) is a hematopoietic malignancy with a prognosis that varies with genetic heterogeneity of hematopoietic stem/progenitor cells (HSPCs). Induction chemotherapy with cytarabine and anthracycline has been the standard care for newly diagnosed AML, but about 30% of patients have no response to this regimen. The resistance mechanisms require deeper understanding.Methods: In our study, using single-cell RNA sequencing, we analyzed the heterogeneity of bone marrow CD34 + cells from newly diagnosed patients with AML who were then divided into sensitive and resistant groups according to their responses to induction chemotherapy with cytarabine and anthracycline. We verified our findings by TCGA database, GEO datasets, and multiparameter flow cytometry. Results:We established a landscape for AML CD34 + cells and identified HSPC types based on the lineage signature genes. Interestingly, we found a cell population with CRIP1 high LGALS1 high S100As high showing features of granulocyte-monocyte progenitors was associated with poor prognosis of AML. And two cell populations marked by CD34 + CD52 + or CD34 + CD74 + DAP12 + were related to good response to induction therapy, showing characteristics of hematopoietic stem cells. Conclusion:Our study indicates the subclones of CD34 + cells confers for outcomes of AML and provides biomarkers to predict the response of patients with AML to induction chemotherapy.

BackgroundWe previously demonstrated that 6-benzylthioinosine (6-BT) could induce the differentiation of a subset of acute myeloid leukemia (AML) cell lines and primary AML cells regardless of their cytogenetics. In this study we investigated whether Wnt signaling pathways played roles in 6-BT-induced differentiation of AML cells.MethodsWe induced differentiation of HL-60 leukemic cells and primary AML cells in vitro using 6-BT. Real-time PCR (qPCR), western blot, and luciferase assays were used to examine the molecules’ expression and biological activity in canonical and noncanonical Wnt signaling pathways. AML cell differentiation was measured by the Nitroblue tetrozolium (NBT) reduction assay.Results6-BT regulated the expression of both canonical and non-canonical Wnt signaling molecules in HL-60 cells. Both 6-BT and all-trans-retinoic-acid (ATRA) reduced canonical Wnt signaling and activated noncanonical Wnt/Ca2+ signaling in HL-60 cells. Pre-treatment of HL-60 cells with an inhibitor of glycogen synthase kinase-3β (GSK-3β), which activated canonical Wnt signaling, partly abolished the differentiation of HL-60 cells induced by 6-BT. Pre-treatment of HL-60 cells with an inhibitor of protein kinase C (PKC), resulting in inactivation of non-canonical Wnt/Ca2+ signaling, abolished 6-BT-induced differentiation of HL-60 cells. Several molecules in the non-canonical Wnt/Ca2+ pathway were detected in bone marrow samples from AML patients, and the expression of FZD4, FZD5, Wnt5a and RHOU were significantly reduced in newly diagnosed AML samples compared with normal controls.ConclusionsBoth canonical and non-canonical Wnt signaling were involved in 6-BT-induced differentiation of HL-60 cells, and played opposite roles in this process. Wnt signaling could be involved in the pathogenesis of AML not only by regulating self-renewal of hematopoietic stem cells, but also by playing a role in the differentiation of AML cells.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-886) contains supplementary material, which is available to authorized users.