The value of routine bone marrow examination in patients with extranodal NK/T-cell lymphoma staged with PET/CT.

BACKGROUND: Limited evidence supports the omission of routine bone marrow (BM) examination (biopsy and aspiration) in patients with nasal-type extranodal NK/T-cell lymphoma (ENKTCL). This study was aimed at assessing whether BM examination provides valuable information for positron emission tomography/computed tomography (PET/CT)-based staging in this patient population. PATIENTS AND METHODS: Patients newly diagnosed with ENKTCL who underwent initial staging with both PET/CT and BM examination between 2013 and 2020 were retrospectively identified in two Chinese institutions. Overall, 742 patients were included; the BM examination was positive in 67 patients. RESULTS: Compared with BM biopsy alone, the combination of BM biopsy and aspiration assessment did not afford any additional diagnostic value. No patient with a positive BM biopsy was found to have early-stage disease by PET/CT. BM biopsy or PET/CT led to upstaging from stage III to IV as a result of BM involvement in 21 patients. In 135 patients with distant organ involvement, BM involvement was associated with worse overall survival (OS) and progression-free survival (PFS) compared with the corresponding durations in patients without BM involvement (2-year OS: 35.9% vs. 60.4%, p < .001; PFS: 26% vs. 40.7%, p = .003). No difference in survival was noted between groups judged positive based on PET/CT and BM biopsy. CONCLUSION: Compared with aspiration, BM biopsy led to the detection of more BM lesions. Baseline PET/CT can be safely used to exclude BM involvement in early-stage disease. Overall, routine BM examination affords diagnostic or prognostic value over PET/CT in patients with advanced-stage nasal-type ENKTCL.

[Oral Mucosal Diffuse Large B-cell Lymphoma Caused by Long-term Oral Methotrexate:Report of One Case].

A case of primary oral mucosal diffuse large B-cell lymphoma(DLBCL)due to long-term use of methotrexate(MTX)for the treatment of rheumatoid arthritis(RA)was admitted to the Department of Hematology,Fujian Medical University Union Hospital.We analyzed and discussed the clinical features,diagnosis and treatment,and prognosis of specific malignant lymphoma induced by MTX in this RA patient.Our purpose is to improve the awareness and knowledge of other iatrogenic immunodeficiency-associated lymphoproliferative disorders of clinicians and pathologists.This study provides a new reference for the clinical diagnosis and treatment of MTX-associated DLBCL.

[Clinical features and prognosis of children with mature B-cell non-Hodgkin's lymphoma: an analysis of 28 cases].

OBJECTIVE: To study the clinical features and treatment outcome of children with mature B-cell non-Hodgkin's lymphoma (B-NHL). METHODS: A total of 28 previously untreated children with mature B-NHL were enrolled and given the chemotherapy regimen of CCCG-B-NHL-2010. Among them, 20 were given rituximab in addition to chemotherapy. The children were followed up for 31 months (ranged 4-70 months). A retrospective analysis was performed for the clinical features of these children. The Kaplan-Meier method was used for survival analysis. A univariate analysis was performed to investigate the prognostic factors. RESULTS: Among the 28 children, 17 (61%) had Burkitt lymphoma, 8 (29%) had diffuse large B-cell lymphoma (DLBCL), and 3 (11%) had unclassifiable B-cell lymphoma. As for the initial symptom, 13 (46%) had cervical mass, 10 (36%) had maxillofacial mass, 9 (32%) had hepatosplenomegaly, 5 (18%) had abdominal mass, and 5 (18%) had exophthalmos. Of all children, 14 had a lactate dehydrogenase (LDH) level of <500 IU/L, 3 had a level of 500-1 000 IU/L, and 11 had a level of ≥ 1 000 IU/L. After two courses of chemotherapy, 21 children achieved complete remission and 7 achieved partial remission. At the end of follow-up, 24 achieved continuous complete remission and 4 experienced recurrence. The 2-year event-free survival rate was (85.7± 6.6)%. The children with bone marrow infiltration suggested by bone marrow biopsy, serum LDH ≥500 IU/L, and bone marrow tumor cells >25% had a low 2-year cumulative survival rate. CONCLUSIONS: The CCCG-B-NHL 2010 chemotherapy regimen combined with rituximab has a satisfactory effect in the treatment of children with B-NHL. Bone marrow infiltration on bone marrow biopsy is associated with poor prognosis.

[Aberrant Expression of Small Nucleolar RNA SNORA63 and Its Clinical Significance in Patients with Acute Leukemia].

OBJECTIVE: To investigate the expression level of small nucleolar RNA (snoRNA) SNORA63 in bone marrow of patients with acute leukemia (AL) and its significance in the clinical diagnosis, treatment and prognosis of AL patients. METHODS: Bone marrow samples of 53 newly diagnosed AL patients and 29 healthy subjects in the Affiliated Hospital of Guangdong Medical University from March 2018 to December 2021 were collected. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the relative expression level of SNORA63 in bone marrow mononuclear cells of the two groups. The median expression level of SNORA63 in AL patients was used as the boundary value to divide the patients into SNORA63 high and low expression groups, and the relationship between the expression level of SNORA63 and the clinical characteristics, clinical indicators and prognosis of AL patients was analyzed and discussed. RESULTS: The relative expression level of SNORA63 in AL patients was significantly lower than that in healthy control group [0.3018 (0.0244-1.2792) vs 1.0882 (0.2797-1.9889)] (P < 0.01). The expression level of SNORA63 in AL patients without remission after initial treatment was significantly lower than that in healthy controls and the patients who received complete remission (CR) (P < 0.01), while there was no statistical difference in the expression level of SNORA63 between AML and ALL groups (P >0.05). The abnormal low expression of SNORA63 was closely related to fever, hemorrage, poor prognosis, efficacy, platelets (PLT), lactate dehydrogenase (LDH), albumin (ALB), and molecular biological abnormalities of AL patients (P < 0.05), but not significantly correlated with sex, age, AL subtype, pallor, fatigue, extramedullary infiltration, white blood cell count (WBC), hemoglobin (HGB), C-reactive protein (CRP), procalcitonin (PCT), fibrinogen (FIB) or chromosome karyotype (P >0.05). Meanwhile, overall survival (OS) and event-free survival (EFS) of AL patients in SNORA63 high-expression group were significantly higher than those in SNORA63 low-expression group (P < 0.05). Univariate Cox regression analysis showed that SNORA63, molecular biological abnormalities, fever, PLT and LDH were the factors influencing OS and EFS in AL patients (P < 0.05). Multivariate Cox regression analysis indicated that fever, molecular biological abnormalities and LDH were independent factors associated with OS and EFS in AL patients (P < 0.05). CONCLUSION: SNORA63 is significantly down-expressed in AL patients, which is a molecular marker of great clinical value for disease monitoring and prognosis evaluation in AL patients.

LATS1 Promotes B-ALL Tumorigenesis by Regulating YAP1 Phosphorylation and Subcellular Localization.

OBJECTIVE: YAP1 plays a dual role as an oncogene and tumor suppressor gene in several tumors; differentiating between these roles may depend on the YAP1 phosphorylation pattern. The specific function of YAP1 in B cell acute lymphoblastic leukemia (B-ALL), however, is currently unclear. Thus, in the present study, the role of YAP1 in B-ALL was investigated using relevant cell lines and patient datasets. METHODS: The effects of shRNA-mediated knockdown on YAP1 and LATS1 levels in the NALM6 and MOLT-4 cell lines were examined using Western blotting, quantitative real-time polymerase chain reaction, flow cytometry, immunostaining, and nude mouse subcutaneous tumorigenesis experiments. Gene expression levels of Hippo pathway-related molecules before and after verteporfin (VP) treatment were compared using RNA-Seq to identify significant Hippo pathway-related genes in NALM6 cells. RESULTS: Patients with ALL showing high YAP1 expression and low YAP1-Ser127 phosphorylation levels had worse prognoses than those with low YAP1 protein expression and high YAP1-Ser127 phosphorylation levels. YAP1-Ser127 phosphorylation levels were lower in NALM6 cells than in MOLT-4 and control cells; YAP1 was distributed in the nuclei in NALM6 cells. Knockdown of YAP1 inhibited MOLT-4 and NALM6 cell proliferation and arrested the NALM6 cell cycle in the G0/G1 phase. Before and after VP treatment, the expression of the upstream gene LATS1 was upregulated; its overexpression promoted YAP1-Ser127 phosphorylation. Further, YAP1 was distributed in the plasma. CONCLUSION: LATS1 may downregulate YAP1-Ser127 phosphorylation and maintain B-ALL cell function; thus, VP, which targets this axis, may serve as a new therapeutic method for improving the outcomes for B-ALL patients.

A lasso and random forest model using flow cytometry data identifies primary myelofibrosis.

Thrombocythemia (ET), polycythemia vera (PV), primary myelofibrosis (PMF), prefibrotic/early (pre-PMF), and overt fibrotic PMF (overt PMF) are classical Philadelphia-Negative (Ph-negative) myeloproliferative neoplasms (MPNs). Differentiating between these types based on morphology and molecular markers is challenging. This study aims to clarify the application of flow cytometry in the diagnosis and differential diagnosis of classical MPNs. This study retrospectively analyzed the immunophenotypes, clinical characteristics, and laboratory findings of 211 Ph-negative MPN patients, including ET, PV, pre-PMF, overt PMF, and 47 controls. Compared to ET and PV, PMF differed in white blood cells, hemoglobin, blast cells in the peripheral blood, abnormal karyotype, and WT1 gene expression. PMF also differed from controls in CD34+ cells, granulocyte phenotype, monocyte phenotype, percentage of plasma cells, and dendritic cells. Notably, the PMF group had a significantly lower plasma cell percentage compared with other groups. A lasso and random forest model select five variables (CD34+CD19+cells and CD34+CD38- cells on CD34+cells, CD13dim+CD11b- cells in granulocytes, CD38str+CD19+/-plasma, and CD123+HLA-DR-basophils), which identify PMF with a sensitivity and specificity of 90%. Simultaneously, a classification and regression tree model was constructed using the percentage of CD34+CD38- on CD34+ cells and platelet counts to distinguish between ET and pre-PMF, with accuracies of 94.3% and 83.9%, respectively. Flow immunophenotyping aids in diagnosing PMF and differentiating between ET and PV. It also helps distinguish pre-PMF from ET and guides treatment decisions.

[Clinical Features and Prognosis of Secondary Intestinal Diffuse Large B-Cell Lymphoma].

OBJECTIVE: To explore and analyze the clinical features and prognostic factors of secondary intestinal diffuse large B-cell lymphoma (SI-DLBCL), in order to provide reference for the basic research and clinical diagnosis and treatment of secondary lymphoma of rare sites in the field of hematology. METHODS: The clinical data of 138 patients with SI-DLBCL admitted to Fujian Medical University Union Hospital from June 2011 to June 2022 were collected and sorted, the clinical and pathological features, diagnosis, treatment and prognosis were analyzed. Cox regression risk model was used to conduct univariate and multivariate analysis on the prognostic risk factors. RESULTS: Among the 138 patients with SI-DLBCL included in this study, 85 (61.59%) were male, 53 (38.41%) were female, the median age of onset was 59.5 (16-84) years, the clinical manifestations lacked specificity, the first-line treatment regimen was mainly chemotherapy (67.39%), 94 cases (68.12%) received chemotherapy alone, 40 cases (28.98%) were treated with chemotherapy combined with surgery, and 4 cases (2.90%) were treated with surgery alone. The median follow-up time was 72 (1-148) months. Among the 138 patients with SI-DLBCL, 79 (57.25%) survived, 34 (24.64%) died, 25 cases (18.12%) lost to follow-up, the PFS rates of 1-year, 3-year and 5-year were 57.97%, 49.28% and 32.61%, and the OS rates of 1-year, 3-year and 5-year were 60.14%, 54.35% and 34.06%, respectively. The results of univariate Cox regression analysis showed that age, Lugano stage and IPI score were the influencing factors of OS in SI-DLBCL patients, and age, Lugano stage and IPI score were the influencing factors of PFS in SI-DLBCL patients. The results of multivariate Cox analysis showed that Lugano stage was an independent prognostic factor affecting OS and PFS in SI-DLBCL patients. CONCLUSION: Patients with SI-DLBCL are more common in middle-aged and elderly men, and the early clinical manifestations lack specificity, and the first-line treatment regimen is mainly R-CHOP chemotherapy, and Lugano stage is an independent prognostic factor affecting OS and PFS in SI-DLBCL patients.

Targeted agents plus CHOP compared with CHOP as the first-line treatment for newly diagnosed patients with peripheral T-cell lymphoma (GUIDANCE-03): an open-label, multicentre phase 2 clinical trial.

Background: Peripheral T-cell lymphoma (PTCL) is a heterogeneous disease with dismal outcomes. We conducted an open-label, phase 2 nonrandomised, externally controlled study to evaluate the efficacy and safety of targeted agents plus CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisolone) (CHOPX) for PTCL in the front-line setting. Methods: Eligible patients were ≥18 years of age and newly diagnosed PTCL. Patients in the CHOPX group received standard CHOP at Cycle 1. Specific targeted agents were added from Cycle 2, decitabine if TP53 mut, azacytidine if TET2/KMT2D mut, tucidinostat if CREBBP/EP300 mut, and lenalidomide if without mutations above. Patients in the CHOP group received CHOP for 6 cycles. The primary endpoint was the complete response rate (CRR) at the end of treatment (EOT). Secondary endpoints included overall response rate (ORR), progression-free survival (PFS), overall survival (OS), and safety. The study was registered with ClinicalTrials.gov, NCT04480099. Findings: Between July 29, 2020, and Sep 22, 2022, 96 patients were enrolled and included for efficacy and safety analysis with 48 in each group. The study met its primary endpoint. CRR at EOT in the CHOPX group was superior to the CHOP group (64.6% vs. 33.3%, OR 0.27, 95%CI 0.12-0.64; p = 0.004). At a median follow-up of 24.3 months (IQR 12.0-26.7), improved median PFS was observed in the CHOPX group (25.5 vs. 9.0 months; HR 0.57, 95%CI 0.34-0.98; p = 0.041). The median OS was similar between two groups (not reached vs. 30.9 months; HR 0.55, 95%CI 0.28-1.10; p = 0.088). The most common grade 3-4 hematological and non-hematological adverse events in the CHOPX group were neutropenia (31, 65%) and infection (5, 10%). Interpretation: Targeted agents combined with CHOP demonstrated effective and safe as first-line treatment in PTCL. Biomarker-driven therapeutic strategy is feasible and may lead to promising efficacy specifically toward molecular features in PTCL. Funding: This study was supported by the National Key Research and Development Program (2022YFC2502600) and the General Program of the Shanghai Municipal Health Commission (202040400).

[The Effects and Regulatory Mechanism of Targeting CXC Chemokine Receptor 1/2 Combined with Ara-C on the Malignant Biological Behaviors of U937 Cells of Acute Myeloid Leukemia].

OBJECTIVE: To investigate and analyze the effect of CXC chemokine receptor 1/2 (CXCR1/2) targeting inhibitor Reparixin combined with cytarabine (Ara-C) on the malignant biological behaviors of acute myeloid leukemia cells and its effect on the expression of the CXCR family, while exploring the accompanying molecular mechanism, providing scientific basis and reference for new molecular markers and targeted therapy for AML. METHODS: Acute myeloid leukemia U937 cells were treated with different concentrations of Reparixin, Ara-C alone or in combination, and the cell morphology was observed under an inverted microscope; Wright-Giemsa staining was used to detect cell morphological changes; CCK-8 method was used to detect cell proliferation; the ability of cell invasion was detected by Transwell chamber method; the ability of colony formation was detected by colony formation assay; cell apoptosis was detected by Hoechst 33258 fluorescent staining and Annexin V/PI double-staining flow cytometry; monodansylcadaverine(MDC) staining was used to detect cell autophagy; the expression of apoptosis, autophagy and related signaling pathway proteins was detected by Western blot and the expression changes of CXCR family were detected by real-time quantitative polymerase chain reaction (qRT-PCR). RESULTS: Reparixin could inhibit the proliferation, invasion, migration and clone formation ability of U937 cells. Compared with the single drug group, when U937 cells were intervened by Reparixin combined with Ara-C, the malignant biological behaviors such as proliferation, invasion and colony formation were significantly decreased, and the levels of apoptosis and autophagy were significantly increased (P<0.01). After Reparixin combined with Ara-C intervenes in U937 cells, it can up-regulate the expression of the pro-apoptotic protein Bax and significantly down-regulate the expression of the anti-apoptotic protein Bcl-2, and also hydrolyze and activate Caspase-3, thereby inducing cell apoptosis. Reparixin combined with Ara-C could up-regulate the expressions of LC3Ⅱ and Beclin-1 proteins in U937 cells, and the ratio of LC3Ⅱ/LC3Ⅰ in cells was significantly up-regulated compared with single drug or control group (P<0.01). MDC result showed that the green granules of vesicles increased significantly, and a large number of broken cells were seen (P<0.01). Reparixin combined with Ara-C can significantly inhibit the phosphorylation level of PI3K, AKT and NF-κB signaling molecule, inhibit the malignant biological behavior of cells by inhibiting the activation of PI3K/AKT/NF-κB pathway, and induce programmed cell death. Ara-C intervention in U937 cells had no effect on the expression of CXCR family (P>0.05). The expression of CXCR1, CXCR2, and CXCR4 mRNA could be down-regulated by Reparixin single-agent intervention in U937 cells (P<0.05), and the expression of CXCR2 was more significantly down-regulated than the control group and other CXCRs (P<0.01). When Reparixin and Ara-C intervened in combination, the down-regulated levels of CXCR1 and CXCR2 were more significant than those in the single-drug group (P<0.01), while the relative expressions of CXCR4 and CXCR7 mRNA had no significant difference compared with the single-drug group (P>0.05). CONCLUSION: Reparixin combined with Ara-C can synergistically inhibit the malignant biological behaviors of U937 cells such as proliferation, invasion, migration and clone formation, and induce autophagy and apoptosis. The mechanism may be related to affecting the proteins expression of Bcl-2 family and down-regulating the proteins expression of CXCR family, while inhibiting the PI3K/AKT/NF-κB signaling pathway.

[Clinical Features and Outcomes of the Patients with B-Cell Chronic Lymphoproliferative Disease in the New Drug Era].

OBJECTIVE: To analyze the clinical characteristics of the patients with B-cell chronic lymphoproliferative disease（B-CLPD） in the new drug era and the effect of new drug treatment on efficacy and survival. METHODS: The clinical and laboratory data of 200 cases B-CLPD patients diagnosed between April 2015 and August 2021 were analyzed retrospectively. The clinical efficacy and survival of the patients under different treatments including Bruton tyrosine kinase（BTK） inhibitors， rituximab， and chemotherapy alone were analyzed. The prognostic factors affecting the survival of patients were analyzed by univarite analysis and multivariate analysis. RESULTS: There were 119 male（59.5％） and 81 female（40.5%） in 200 cases B-CLPD patients， the sex ratio(male/female) was 1.5∶1 with median age of 61（30- 91） years old. The distribution of subtypes were as fallows: 51 cases (25.5%) of chronic lymphocytic leukemia/small lymphocytic lymphoma（CLL/SLL）， 64（32.0%） cases of follicular lymphoma（FL）， 40（20.0%） cases mantle cell lymphoma（MCL）， 30（15.0%） cases of marginal zone lymphoma（MZL）， 10（5%） cases of lymphoplasmacytic lymphoma/waldenstrom macroglobulinemia（LPL/WM）， 5（2.5%） cases of B cell chronic lymphoproliferative disorders unclassified（B-CLPD-U） . The main clinical manifestation of 102 patients was lymph node enlargement, 32 cases were complicated with B symptoms. Among CLL/SLL patients， there were 12（23.5%） cases in Binet A and 39（76.5%） cases in Binet B/C. There were 29 patients（20.9%） in Ann Arbor or Lugano stage I-II and 110 cases（79.1%） in stage III-IV of other subtypes. The complete remission（CR） rate was 43.1%（25/58）， 40.2%（39/97）， 7.1%（1/14）， and overaIl response rate（ORR） was 87.9%（51/58）， 62.9%（61/97）， 28.6%（4/14） in the groups of BTK inhibitors， rituximab-based therapy， and chemotherapy alone. The 3-year OS rate and PFS rate in all patients was 79.2% and 72.4% respectively. The 3-year OS rate of patient with MZL, CLL/SLL, FL,WM was 94.7%, 87.7%, 86.8% and 83.3% respectively, while the 3-year OS rate of MCL was only 40.6%, which was significantly lower than other subtypes. The median OS of patients treated with BTK inhibitors and rituximab-based therapy was 20.5 and 18.5 months respectively， and the 3-year OS rate was 97.4% and 90.7%. However， the median PFS of patients receiving chemotherapy alone was 4 months， and the 1-year OS rate was 52.7%， which was statistically significant compared with the other two groups（P<0.05）. Univarite analysis showed that anemia， elevated lactate dehydrogenase， elevated ß2-microglobulin， and splenomegaly were the poor prognostic factors for OS（P<0.05）， elevated lactate dehydrogenase was also poor prognostic factors for PFS（P<0.05）. Multifactor analysis showed that anemia and elevated lactate dehydrogenase were the independent poor prognostic factors for survival（P<0.05）. CONCLUSION: The clinical features of B-CLPD was various， anemia and elevated lactate dehydrogenase are the prognostic factors for poor survival. BTK inhibitors and new immunotherapy can improve the survival and prognosis of patients in the new drug era.

[Experimental Study on the Mechanism of Mangiferin Inhibiting Malignant Biological Characteristics of Multiple Myeloma and Exerting Anticancer Effect].

OBJECTIVE: To investigate the effect of pure Chinese herbal extract Mangiferin on the malignant biological behaviors of multiple myeloma (MM) cells, and to analyze the molecular mechanism of the anti-myeloma effect of Mangiferin, so as to provide experimental basis for MM replacement therapy. METHODS: U266 and RPMI8226 of human MM cell lines were intervened with different concentrations of Mangiferin. Cell proliferation was detected by CCK-8 method. Annexin V/PI double staining flow cytometry was used to detect cell apoptosis. Western blot was used to detect the expression of apoptosis and related signaling pathway proteins, and real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of matrix metalloproteinase (MMP) and CXC chemokine receptor (CXCR) family. RESULTS: Mangiferin could inhibit the proliferation activity of U266 and RPMI8226 cells and induce cells apoptosis. After Mangiferin intervened in U266, RPMI8226 cells for 48 h, the expression of Bcl-2 family pro-apoptotic protein Bax was up-regulated, while the expression of survivin and Bcl-xL proteins was down-regulated and caspase-3 was hydrolyzed and activated to promote cell apoptosis, besides, the expression of Bcl-2 protein in U266 cells was also significantly down-regulated to induce apoptosis (P<0.05). After Mangiferin intervenes in MM cells, it can not only increase the expression level of tumor suppressor p53, but also induce programmed cell death of MM cells by inhibiting the expression of anti-apoptotic molecules and down-regulating the phosphorylation levels of AKT and NF-κB. In addition, after the intervention of Mangiferin, the expressions of CXCR4, MMP2 and MMP9 in U266 cells were down-regulated (P<0.05), while there is no effect on the expressions of CXCR2, CXCR7 and MMP13 (P>0.05). However, the expressions of CXCR4, MMP9, and MMP13 in RPMI8226 cells were down-regulated (P<0.01), the expression of MMP2 was weakly affected, and the expression of CXCR2 and CXCR7 was basically not affected (P>0.05). CONCLUSION: Mangiferin can inhibit the proliferation and induce apoptosis of MM cells, and its mechanism may be related to inhibiting the activation of NF-κB signaling pathway, affecting the expression of Bcl-2 family proteins, and inhibiting the expression of core members of MMP and CXCR family.

[Effect of miR-203/CREB1 Signaling Regulation Mediated by DNA Methylation on the Proliferation and Apoptosis of Multiple Myeloma Cells].

OBJECTIVE: To investigate the effect of miR-203/CREB1 signaling regulation mediated by DNA methylation on the proliferation, invasion and apoptosis of multiple myeloma (MM) cells. METHODS: The methylation level of miR-203 in the RPMI 8226 cells was detected by bisulfite sequcucing polymerase chain reaction (BSP). The mRNA expression of miR-203 was measured by quantitative real-time polymerase chain reaction. RPMI 8226 cells were treated with DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (5-Aza-CdR). The miR-203 mimic in MM cell line RPMI 8226 was transfected to establish overexpressed miR-203 cell. The proliferation, invasion ability and apoptosis of RPMI 8226 cell was detected by CCK-8 assay, Transwell, and flow cytometry, respectively. The targeting relationship between miR-203 and CREB1 was verified by double luciferase report assay. Western blot was used to detect the expression of CREB1 protein. RESULTS: Hypermethylation of miR-203 promoter region and low expression level of miR-203 mRNA were detected in the RPMI 8226 cells, which showed that demethylation could induce the expression of miR-203. The proliferation and invasion ability of RPMI 8226 cells after treated by 5-Aza-CdR were inhibited, and showed statistical significance as compared with blank control group (both P<0.05)，while the apoptosis rate was promoted (P<0.05). The proliferation, invasion ability and apoptosis of overexpressed miR-203 were the same as the demethylation group. Double luciferase report assay confirmed that CREB1 was the direct target of miR-203. The protein level of CREB1 was inhibited by demethylation and showed statistical significance as compared with control group (P<0.05). CONCLUSION: MiR-203 targeting CREB1 mediated by DNA methylation leads to maintain the malignant biological behaviors of MM cells.

[The Value of T Cell subsets and Cytokine Levels Changes in the Clinical Diagnosis, Treatment and Prognosis Evaluation of Multiple Myeloma].

OBJECTIVE: To explore the correlation between the changes of T lymphocyte subsets and cytokines in patients with MM and immune function status, biochemical indicators, and their relationships with clinical stage and prognosis, which is expected to provide a scientific basis for the prognosis analysis and condition monitoring of MM patients. METHODS: The clinical data of 89 MM patients in two hospitals were collected, and 36 healthy people without tumor or infectious diseases were selected as the control group. Flow cytometry and enzyme-linked immunosorbent assay (ELISA) were used to detect the changes of core members of peripheral blood T lymphocyte subsets and cytokine levels, respectively. At the same time, automatic biochemical analyzer and automatic blood cell analyzer were used to detect serum ß2-microglobulin (ß2-MG), lactate dehydrogenase (LDH), albumin (ALB), creatinine (CRE) and hemoglobin (HGB) levels, and the relationship between T lymphocyte subsets and the above indexes and their clinical significance were analyzed. RESULTS: The proportions of NK cells and CD8+T lymphocytes in the peripheral blood of MM patients were significantly higher than that of the control group (P<0.01), the proportion of CD4+T and the ratio of CD4+/CD8+ were lower than those of the control group (P<0.05); however, there was no significant difference in the numbers CD3+T cells compared with the control group (P>0.05). The proportion of CD4+T and ratios of CD4+/CD8+ in MM patients were lower than those of normal controls, and were negatively correlated with MM staging (r=-0.964, r=-0.653), that is, the later the MM staging, the more obvious their levels were reduced, while CD8+T and NK cells were positively correlated with MM staging (r=0.891, r=0.728), that is, the later the MM staging, the more significant their levels increased. The levels of Treg cells (CD4+CD25highCD127low/-T cells/CD4+T cells) of MM patients in the disease stage Ⅰ, Ⅱ and Ⅲ were (5.87±0.92)%, (7.97±1.32)%, (11.52±4.71)% respectively, the difference was statistically significant compared with control group (P<0.05), and the level of Treg cells in MM patients with stage III was significantly higher than that in controls and patients with other disease stages (P<0.01). The proportion of Treg cells (CD4+CD25highCD127low/-T cells/CD4+T cells) in MM patients was positively correlated with the concentration of ß2-MG and LDH (r=0.793, r=0.536), but had no significant correlation with HGB, ALB and CRE. The serum levels of IL-6, IL-10 and TNF-α in MM patients were significantly higher than those in the control group (P<0.05), which were closely related to MM staging(r=0.839, r=0.917, r=0.746), that is, the later the MM staging, the higher the levels; The serum IFN-γ level was negatively correlated with the stage of MM (r=-0.689), and its level gradually decreased with the increase of the disease stage and degree (P<0.01). There was no significant correlation between the levels of IL-2 and IL-4 and the disease stage, but they were all up-regulated compared with the control group (P<0.05). CONCLUSION: The abnormal regulation of the core members of T lymphocyte subsets and the levels of various cytokines are closely related to the disease progression and poor prognosis of MM patients, which is an effective indicator for the disease monitoring of MM patients.

[Effect of Body Mass Index on the Prognosis of Mantle Cell Lymphoma].

OBJECTIVE: To explore the correlation between different body mass index (BMI) and prognosis of mantle cell lymphoma (MCL). METHODS: The clinical characteristics and biological indices of 108 patients with MCL treated in Fujian Medical University Union Hospital were retrospectively analyzed, and the effects of different BMI on overall survival (OS) and progression-free survival (PFS) were analyzed. The correlation between BMI and B symptoms, LDH and Ki-67 was further observed. Furthermore，the differences of BMI between Autologous peripheral blood stem cell transplantation(Auto-PBSCT) and conventional chemotherapy groups were explored. RESULTS: Among 108 patients, the median age at diagnosis was 59（25-79） years old, and the male to female ratio was 4.4∶1. 88.89% of patients with Ann Arbor staging III-IV, 63.89% with bone marrow involvement, and 49.07% with splenic infiltration. Patients with BMI ≥ 24 kg/m2 were divided into two groups: the high BMI group and the low BMI group. The 5-year PFS and OS of patients in the low BMI group were 31.9% and 47.0%, respectively, while those in the high BMI group were 64.6% and 68.7%, respectively. The incidence of death in the high BMI group was lower than that of the low BMI group (P＜0.01). In multivariate analysis, BMI was an independent predictor of PFS (HR=0.282; 95% CI: 0.122-0.651; P=0.003) and an independent predictor of OS (HR=0.299; 95% CI: 0.129-0.693; P=0.005). Also, patients with B symptoms had a lower BMI than those without B symptoms (P=0.01), but BMI had no effect on patients' LDH and Ki-67. The prognosis of 16 patients treated with Auto-PBSCT was significantly better than that of the conventional chemotherapy group. There was no significant difference in BMI between Auto-PBSCT group and conventional chemotherapy group. CONCLUSION: BMI is an independent prognostic factor for PFS and OS in MCL, and may be influenced by the effect of B symptoms on BMI.

Erratum to: Genistein-induced Anticancer Effects on Acute Leukemia Cells Involve the Regulation of Wnt Signaling Pathway Through H4K20me1 Rather Than DNA Demethylation.

The article "Genistein-induced Anticancer Effects on Acute Leukemia Cells Involve the Regulation of Wnt Signaling Pathway Through H4K20me1 Rather Than DNA Demethylation", written by Hua-rong ZHOU, Jian-zhen SHEN, Hai-ying FU, Feng ZHANG was originally published electronically on the publisher's internet portal on October 2021 without open access. With the author(s)' decision to opt for Open Choice, the copyright of the article is changed to © The Author(s) 2021 and the article is forthwith distributed under a Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ ), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.The original article has been corrected.

[The Effect of MicroRNA-143 on the Proliferation and Apoptosis of Leukemia Cell Line U-937].

OBJECTIVE: To investigate the expression of microRNA-143 (miR-143) in patients with acute myeloid leukemia (AML), and the effect of miR-143 over-expression on the proliferation and apoptosis of AML cells. And to verify the targeting relationship between miR-143 and long non-coding RNA MALAT-1. So as to explore the possible regulatory mechanism of miR-143 in the pathogenesis of AML. METHODS: The expression of miR-143 in bone marrow cells of AML patients was detected by RT-qPCR. After miR-143 was over-expressed in U-937 cell lines, the proliferation of U-937 cell lines was detected by CCK-8, clone formation assay, and flow cytometry. In addition, cell apoptosis was detected by Hoechst 33258 fluorescence staining and flow cytometry. At the same time, bioinformatics, RT-qPCR and dual luciferase reporter gene assay were used to predicted and verified the targeting relationship between miR-143 and MALAT-1. RESULTS: Expression of miR-143 in AML patients was significantly lower than those in normal controls. Over-expressed miR-143 could inhibit the proliferation of U-937 cells and promote the apoptosis of the cell. The miR-143 binding site was located on the MALAT-1 RNA sequence, and MALAT-1 was down-regulated in U-937 cells after over-expressed miR-143. However, the expression of miR-143 showed no significantly changed after MALAT-1 silencing. CONCLUSION: Expression of miR-143 in AML patients is lower than that in normal controls. Over-expression of miR-143 can inhibit the proliferation of U-937 cells and promote its apoptosis. And its mechanism may be related to its targe regulation of MALAT-1.

Genistein-induced Anticancer Effects on Acute Leukemia Cells Involve the Regulation of Wnt Signaling Pathway Through H4K20me1 Rather Than DNA Demethylation.

OBJECTIVE: To investigate the effects and mechanisms of genistein on the gene expression in the Wnt pathway in acute leukemia (AL) cells. METHODS: The expression of Wnt pathway genes and cell cycle-related genes were analyzed in two AL cell lines. Pyrophosphate sequencing was performed to determine the methylation degree. Then, the enrichment of H4K20me1 and H3K9ac was determined using ChIP-qPCR. Flow cytometry was used to analyze the cell cycle. RESULTS: The IC50 of genistein in the two AL cell lines was lower than that for the bone marrow mesenchymal stem cell line. Genistein upregulated H4K20me1, KMT5A and Wnt suppressor genes, including Wnt5a, and downregulated the downstream target genes of Wnt, such as c-myc and ß-catenin. The methylation degree and H3K9ac enrichment in the Wnt5a promoter region remained unchanged. However, the enrichment of H4K20me1 in the Wnt5a promoter and coding regions increased. In addition, genistein upregulated Phospho-cdc2, Myt1, Cyclin A, Cyclin E2, p21 and Phospho-histone H3, but downregulated Phospho-wee1. Cell cycle arrest was induced in the G2/M phase. CONCLUSION: Genistein inhibits the activation of the Wnt pathway by promoting the expression of Wnt5a through the activation of KMT5A and enrichment of H4K20me1 in the Wnt5a gene promoter and coding regions, rather than demethylation. Genistein also blocks the cell cycle in the G2/M phase. Therefore, genistein is a potential anti-leukemia drug.

[A Case-Control Study on Receptor Gene Polymorphism and Risk Suffering from Adult Acute Leukemia in Fujian Area].

OBJECTIVE: To investigate the correlation of receptor gene (P2X7, VDR and SLC19A1) polymorphisms with risk suffering from acute leukemia (AL) in Fujian area. METHODS: Ninety-three cases of newly diagnosed AL as AL group and 90 persons not suffered from hematologic and other tumors as control group were selected and used for comparative analysis of receptor gene polymorphisms and risk suffering from AL between case and control groups. The bone marrow and peripheral blood were collected, from which the DNA was extracted. The PCR-RFLP was used to detect 8 SNP sites (P2X7: rs208294, rs2230911, rs3751143; VDR: rs2228570, rs7975232; SLC194A1: rs1051266, rs1131596, rs3788200) of receptor genes related with the environment response, and the genotypes analysis was used to the correlation of receptor gene polymorphisms with risk suffering from adult AL. RESULTS: The unvariate logistic analysis showed that as compared with control group, P2X7 rs208294 T>C mutation and rs3751143 A>C mutation in codominant model, dominant model and over-dominant model were higher in case group, moreover the differences were statistically significant (P<0.01, P<0.05 and P<0.05, respectively), which suggested that they could reduce the risk suffering from AL. The recessive inheritance model showed that SLC1941 rs1131596 G>A mutation could increase the risk suffering from AL (P<0.05). The stepwise multivariate logistic regression analysis showed that there was still statistically significant difference in P2X7 rs208294 mutation between case group and control group (P<0.05), moreover, the heterozygous mutation (CT) could decrease the risk suffering from AL, showing the better protective effect, compared with homozygous mutation(CC). CONCLUSION: The P2X7 rs208294 T>C mutation is one of protective factors against adult acute leukemia.

[The significance of methylation status of secreted frizzled-related protein gene promoter in acute leukemia].

OBJECTIVE: To investigate the relationship between promoter hypermethylation of secreted frizzled-related protein (SFRP) gene and acute leukemia (AL). METHODS: We examined the promoter methylation status of SFRP1, 2, 4 and 5 in primary or relapsed AL patients, cell lines (HL60, NB4, Molt-4 and Jurkat) and peripheral blood mononuclear cells from healthy people with methylation-specific PCR (MSP). RESULTS: None of the normal mononuclear cells showed methylation of any SFRP genes. The frequencies of aberrant methylation among the samples were 33.9% (20/59) for SFRP1, 23.7% (14/59) for SFRP2, 6.8% (4/59) for SFRP4 and 10.2% (6/59) for SFRP5 in acute myelocytic leukemia (AML), and 39.3% (11/28) for SFRP1, 28.6% (8/28) for SFRP2, 25.0% (7/28) for SFRP4 and 32.1% (9/28) for SFRP5 in acute lymphoblastic leukemia (ALL). Hypermethylation of SFRP1, 2, 5 genes were present in the 4 AL cell lines. SFRP4 was methylated in NB4, Molt-4 and Jurkat cell lines. However, methylation and unmethylation of SFRP4 were both detected in HL60. CONCLUSIONS: Hypermethylation of SFRP genes is a common event in the evolution of AL. Methylation of SFRP genes might serve as potential independent biomarkers for early detection of AL.

[The establishment and evaluation of diagnostic accuracy of AllGlo(TM) probe-based techniques for invasive aspergillosis.].

OBJECTIVE: To establish and evaluate a diagnostic technique based on the AllGlo(TM) probe for the invasive aspergillosis. METHODS: With the self-designed AllGlo(TM) probes and primers and the standards, two standard curves of the real-time PCR based on AllGlo(TM) probes were established respectively for A. flavus and A. fumigatus, then its specificity, sensitivity and reproducibility were evaluated respectively. RESULTS: The findings indicated that the standard curve of A. flavus was Y = -3.003X + 36.825, and A.fumigatus' was Y = -3.052X + 38.016, and their interassay coefficient of variation respectively were 15.60% and 12.94%, suggesting a good reproducibility. The lowest spore concentration they could be detected was 10 CFU/ml, which equated to 100 - 1000 copies of internal transcribed spacer (ITS)2 genes, suggesting a good sensibility. They didn't have cross-positive reaction with other fungus, human genome and bacteria, which indicated a good specificity. CONCLUSION: The diagnostic technique based on the AllGlo(TM) probe for the invasive aspergillosis possessed a good sensitivity, good specificity and deadly accuracy.