Chongyun Xing

Long non-coding RNA HOTAIR modulates c-KIT expression through sponging miR-193a in acute myeloid leukemia

HOTAIR is significantly overexpressed in various cancers and facilitates tumor invasion and metastasis. However, whether HOTAIR plays oncogenic roles in acute myeloid leukemia (AML) is still unknown. Here, we report that HOTAIR expression was obviously increased in leukemic cell lines and primary AML blasts. Clinically, AML patients with higher HOTAIR predicted worse clinical outcome compared with those with lower HOTAIR. Importantly, HOTAIR knockdown by small hairpin RNA inhibited cell growth, induced apoptosis, and decreased number of colony formation. Finally, HOTAIR modulated c‐KIT expression by competitively binding miR‐193a. Collectively, our data suggest that HOTAIR plays an important oncogenic role in AML and might serve as a marker for AML prognosis and a potential target for therapeutic intervention.

Honokiol induces proteasomal degradation of AML1-ETO oncoprotein via increasing ubiquitin conjugase UbcH8 expression in leukemia.

ABSTRACT AML1‐ETO is the most common oncoprotein leading to acute myeloid leukemia (AML), in which 5‐year survival rate is only about 30%. However, currently there are no specific therapies for AML patients with AML1‐ETO. Here, we report that AML1‐ETO protein is rapidly degraded by Honokiol (HNK), a natural phenolic compound isolated from the plant Magnolia officinalis. HNK induced the degradation of AML1‐ETO in a concentration‐ and time‐dependent manner in leukemic cell lines and primary AML blasts with t(8;21) translocation. Mechanistically, HNK obviously increased the expression of UbcH8, an E2‐conjugase for the degradation of AML1‐ETO, through triggering accumulation of acetylated histones in the promoter region of UbcH8. Knockdown of UbcH8 by small hairpin RNAs (shRNAs) prevented HNK‐induced degradation of AML‐ETO, suggesting that UbcH8 plays a critical role in the degradation of AML1‐ETO. HNK inhibited cell proliferation and induced apoptotic death without activation of caspase‐3, which was reported to cleave and degrade AML1‐ETO protein. Thus, HNK‐induced degradation of AML1‐ETO is independent of activation of caspase‐3. Finally, HNK reduced the angiogenesis and migration in Kasumi‐1‐injected zebrafish, decreased xenograft tumor size in a xenograft leukemia mouse model, and prolonged the survival time in mouse C1498 AML model. Collectively, HNK might be a potential treatment for t(8;21) leukemia by targeting AML1‐ETO oncoprotein.

The independent association of mean platelet volume with overall survival in multiple myeloma

We retrospectively analyzed the association between mean platelet volume (MPV) and prognosis in 62 newly diagnosed multiple myeloma (MM) patients. The associations between MPV and clinical characteristics were assessed. The log-rank test and the Cox proportional hazards model were used to evaluate the effect of MPV on survival. A MPV value of 8.50 fl was considered to be the optimal cut-off value for prognosis. MPV was associated with IgA isotype (P=0.012), serum creatinine concentration > 176.8 μmol/L (P=0.025) and IgH rearrangement (P=0.008). The log-rank test demonstrated that patients with low MPV experienced a shorter overall survival (OS) (P=0.0397). The multivariate analysis demonstrated that low MPV was an independent prognostic factor for OS [hazard ratio (HR)=2.44, P=0.026]. Therefore, we demonstrated that low MPV predicted an unfavorable prognosis in patients with MM.

Effect of initial absolute monocyte count on survival outcome of patients with de novo non-M3 acute myeloid leukemia.

Abstract Increased absolute monocyte count (AMC) at presentation has recently been associated with clinical outcome in different types of hematological malignancies. This study aimed to assess the prognostic value of AMC on survival in 193 adult patients with de novo non-M3 acute myeloid leukemia (AML). The median AMC for all patients at diagnosis was 0.26 × 109/L, with 41.4, 31.1 and 27.5% of patients showed low (<0.12 × 109/L), normal (0.12–0.80 × 109/L), and high AMC (>0.80 × 109/L), respectively. Univariate analysis revealed that high AMC appeared as a poor prognostic factor for overall survival (OS) (p = 0.0055), but not for disease free survival (DFS) (p = 0.1195). On multivariate analysis, initial high AMC remained an independent predictor of OS (hazard ratio 2.01, p = 0.017). Our results suggest that AMC at diagnosis, which provides additional prognostic information independently from conventional factors related to patient clinical characteristics or tumor biological features, could be a novel prognostic marker for AML.

Co-occurrence of JAK2 V617F and an uncommon CALR del (p.K368fs*51) mutation facilitates JAK2/STAT signaling in polycythemia vera.

BCR/ABL-negative myeloproliferative neoplasms (MPNs), which mainly comprise polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF),[1] are characterized by the overexpression of mature blood cells.[2] JAK2 V617F mutation, a valine-tophenylalanine substitution at amino acid position 617, was occurred approximately 96% in patients with PV and 55%–65% in patients with ET and PMF.[3] This mutation caused constitutive activation of JAK2/STAT signaling pathway and facilitated cell proliferation independent of cytokines.[3] Recently, somatic insertions or deletions in exon 9 of CALR, which caused a frameshift to the same alternative reading frame and generated a novel Cterminal peptide in the mutant CALR, were discovered in a significant percentage in ET and PMF patients without JAK2 V617F mutation.[4] Although JAK2 V617F and CALR mutations were mutually exclusive in patients with MPNs,[5] concurrent JAK2 V617F and CALR mutations had been reported in two patients with PMF, two ET, and one PV,[6–9] respectively. Here, we report a single PV patient who bears both JAK2 V617F and an uncommon CALR del (p.K368fs*51) mutation. In October 2014, a 49-year-old man presented to our hospital and reported 12 months-long history of increased weakness and fatigue with splenomegaly. The peripheral blood count indicated a hemoglobin 189 g/L, hematocrit 59%, platelet count 422 10/L, white blood cell count (WBC) 21.4 10/L (neutrophils 71.8%, lymphocytes 14.8%, monocytes 6.1%), and low erythropoietin levels (1.9 IU/L). A bone marrow aspirate showed that red department cells were remarkable proliferation, but no dysmorphic red cells were observed. BCR-ABL fusion gene was negative by fluorescence in situ hybridization (data not shown) and real time PCR (data not shown). The karyotype of this patient was normal. Further, the status of JAK2 V617F and CALR exon 9 mutations was detected by DNA direct sequencing. Heterogenous JAK2 V617F and CALR mutations were both observed in this patient (Fig. 1A and B). Intriguingly, this CALR mutation was not two most cocmmon CALR variants, which are mutation of type 1 (52-bp deletion; c.1092_1143del) and mutation of type 2 (5-bp insertion; c.1154_1155insTTGTC).[4,5] This is an uncommon variant of a 34-bp deletion, which was classified as c.1103–1136del, p.K368fs*51. Like all other reported mutations, CALR del (p.K368fs*51) led to a +1bp shift in the reading frame and thus generated a new C terminus (Fig. 1C). The patient was diagnosed with PV and was administered with interferon a from October 2014 for 9 months, at which time he had a complete hematologic response. Based on the reports that JAK2 V617F and CALR mutations facilitated constitutive activation of JAK2/ STAT signaling, respectively,[3,5] we then determined whether co-occurrence of CALR del (p.K368fs*51) and JAK2 V617F enhanced JAK2/STAT signaling. HEL cells with JAK2 V617F mutation [10] were transfected with retroviral MSCV-Flag-CALR del (p.K368fs*51), wide-type MSCV-CALR, or negative control. As indicated in Fig. 2A, wide-type CALR and CALR del (p.K368fs*51) mutation were ectopically overexpressed in HEL cells by western blotting. Overexpression of CALR del (p.K368fs*51) mutation but not wide-type CALR significantly enhanced the activation of JAK2/STAT signaling in HEL cells (Fig. 2B). Further, we determined whether downstream STAT3/5 targets, such as cyclin D1 [11] and Bcl-2,[12]

Effect of absolute monocyte count post-transplant on the outcome of patients with acute myeloid leukemia undergoing myeloablative allogeneic hematopoietic stem cell transplant with busulfan and cyclophosphamide conditioning

Peripheral monocytes have recently been evaluated as a prognostic factor in different types of hematological malignancies. This study assessed the prognostic value of absolute monocyte count (AMC) post-transplant on the clinical outcomes of 59 patients with acute myeloid leukemia (AML) who had undergone myeloablative conditioning (MAC) allogeneic hematopoietic stem cell transplant (allo-HSCT) with busulfan and cyclophosphamide (Bu/Cy). Kaplan-Meier analysis showed that patients with a high AMC (≥ 0.57 × 109/L) on post-transplant day (PTD) 15 had a significantly worse overall survival (OS) compared to patients with a low AMC (< 0.57 × 109/L) on PTD 15 (P = .0049). Univariate Cox proportional hazard analyses revealed that only high AMC on PTD 15 was a poor prognostic factor for OS (P = .008) and post-relapse survival (P = .030). We conclude that AMC ≥ 0.57 × 109/L on PTD 15 is associated with more deaths in patients with AML who have undergone MAC allo-HSCT with Bu/Cy.

Long noncoding RNA HOTAIR promotes the self-renewal of leukemia stem cells through epigenetic silencing of p15

Acute myeloid leukemia (AML) is a heterogeneous hematopoietic disorder initiated from a small subset of leukemia stem cell (LSC), which presents unrestricted self-renewal and proliferation. Long non-coding RNA HOTAIR is abundantly expressed and plays oncogenic roles in solid cancer and AML. However, whether HOTAIR regulates the self-renewal of LSC is largely unknown. Here, we reported that the expression of HOTAIR was increased in LSC than in normal hematological stem and progenitor cells (HSPCs). HOTAIR inhibition by short hairpin RNAs (shRNAs) decreased colony formation in leukemia cell lines and primary AML blasts. We then investigated the role of HOTAIR in leukemia in vivo. HOTAIR knockdown extends the survival time in U937-transplanted NSG mice. Furthermore, HOTAIR knockdown reduced infiltration of leukemic blasts, decreased frequency of LSC, and prolonged overall survival in MLL-AF9-induced murine leukemia, suggesting that HOTAIR is required for the maintenance of AML. Mechanistically, HOTAIR inhibited p15 expression through zeste homolog 2 (EZH2)-enrolled tri-methylation of Lys 27 of histone H3 (H3K27me3) in p15 promoter. In addition, p15 partially reversed the decrease of colony and proliferation induced by HOTAIR knockdown, suggesting that p15 plays an important role in the leukemogenesis by HOTAIR. In conclusion, our study suggests that HOTAIR facilitates leukemogenesis by enhancing self-renewal of LSC. HOTAIR might be a potential target for anti-LSC therapy.

Down-regulation of CD19 expression inhibits proliferation, adhesion, migration and invasion and promotes apoptosis and the efficacy of chemotherapeutic agents and imatinib in SUP-B15 cells: Effect of down-regulation of CD19 in Ph+ ALL

The survival rate of childhood acute lymphoblastic leukemia (ALL) has increased while that of Philadelphia‐positive (Ph+) ALL remains low. CD19 is a B‐cell specific molecule related to the survival and proliferation of normal B cells. However, there is little information available on the effects of CD19 on the biological behavior of Ph+ ALL cells. In this study, we explored a lentiviral vector‐mediated short hairpin RNA (shRNA) expression vector to stably reduce CD19 expression in Ph+ ALL cell line SUP‐B15 cells and investigated the effects of CD19 downregulation on cell proliferation, apoptosis, drug sensitivity, cell adhesion, cell migration and cell invasion in vitro. CD19 mRNA and protein expression levels were inhibited significantly by CD19 shRNA. Down‐regulation of CD19 could inhibit cell proliferation, adhesion, migration and invasion, and increase cell apoptosis and the efficacy of chemotherapeutic agents and imatinib in SUP‐B15 cells. Moreover, we found that down‐regulation of CD19 expression inhibits cell proliferation and induces apoptosis in SUP‐B15 cells in a p53‐dependent manner. Taken together, our results suggest that lentiviral vector‐mediated RNA interference of CD19 gene may be a promising strategy in the treatment of Ph+ ALL.

Prognostic significance of leukopenia during the induction phase in adult B cell acute lymphoblastic leukemia

The association between chemotherapy-induced leukopenia and clinical outcome has been reported for several types of cancer. The objective of the current study was to evaluate the association of chemotherapy-induced leukopenia during the induction phase with the clinical outcome of adult B cell acute lymphoblastic leukemia (B-ALL). Fifty-one cases of B-ALL, age ≥14 years, were reviewed. The variables under consideration included age, sex, the initial white blood cell (WBC) count (WBC-0), as well as the WBC counts on days 8 (WBC-8), 15 (WBC-15), and 22 (WBC-22) during induction therapy, early bone marrow responses on day 15 during induction therapy, immunophenotype, and cytogenetics. Univariate analysis revealed that WBC-15 ≥0.40×109/L was significantly associated with inferior event-free survival (EFS) (hazard ratio [HR]=2.95, P=0.004) and overall survival (OS) (HR=2.92, P=0.015). On multivariate analysis, high WBC-15 (≥0.40×109/L) remained an independent prognostic factor for EFS (HR=3.29, P=0.014) and OS (HR=3.29, P=0.038). Our results suggested that WBC-15 may contribute to refinements in the current risk stratification algorithms for adult B-ALL.