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Cytogenetic abnormalities have been proven to be the most valuable parameter for risk stratification of childhood acute lymphoblastic leukemia (ALL). However, studies on the prevalence of cytogenetic abnormalities and their correlation to clinical features in Chinese pediatric patients are limited, especially large-scale studies.We collected the cytogenetics and clinical data of 1541 children newly diagnosed with ALL between 2001 and 2014 in four Chinese hospitals, and retrospectively analyzed their clinical features, prognosis and risk factors associated with pediatric ALL.All of these patients had karyotyping results, and some of them were tested for fusion genes by fluorescence in situ hybridization or reverse-transcription polymerase chain reaction. Overall, 930 cases (60.4%) had abnormal cytogenetics in this study, mainly including high hyperdiploidy (HHD, n=276, 17.9%), hypodiploidy (n=74, 4.8%), t(12;21)/TEL-AML1 (n=260, 16.9%), t(1;19)/E2A-PBX1 (n=72, 4.7%), t(9;22)/BCR-ABL (n=64, 4.2%), and t(v;11q23)/MLL rearrangements (n=40, 2.6%). The distribution of each cytogenetic abnormality was correlated with gender, age, white blood cell count at diagnosis, and immunophenotype. In addition, multivariate analysis suggested that t(v;11q23)/MLL rearrangements (OR: 2.317, 95%CI: 1.219-3.748, P=0.008) and t(9;22)/BCR-ABL (OR: 2.519, 95%CI: 1.59-3.992, P0.001) were independent risk factors for a lower event-free survival (EFS) rate in children with ALL, while HHD (OR: 0.638, 95%CI: 0.455-0.894, P=0.009) and t(12;21)/TEL-AML1 (OR: 0.486, 95%CI: 0.333-0.707, P0.001) were independent factors of a favorable EFS.The cytogenetic characteristics presented in our study resembled other research groups, emphasizing the important role of cytogenetic and molecular genetic classification in ALL, especially in B-ALL.

To investigate the effect of course delay of CCLG-ALL-2008 regimen on the relapse of paediatric B-cell acute lymphoblastic leukemia (B-ALL) patients.Paediatric B-ALL patients newly diagnosed and treated with CCLG-ALL-2008 regimen in the Children's Hospital of Soochow University from January 2011 to December 2014 were retrospectively analyzed to clarify the relationship between chemotherapy course delay and relapse, and explore the causes of course delay which led to relapse. Patients were followed up until July 2019.The correlation between treatment delay (number of weeks) and relapse rate was statistically significant (P=0.034), and hazard ratio indicated that longer than 4 weeks had a significant effect. The effect of positive minimal residual disease (MRD) (1×10Treatment delay exceeding 4 weeks, positive MRD at the 12th week, and myelosuppression are independent prognostic factors for relapse.疗程拖延对儿童急性B淋巴细胞白血病复发的影响.探讨急性B淋巴细胞白血病患儿采用CCLG-ALL-2008方案疗程拖延对疾病复发的影响。.对2011年1月至2014年12月在苏州大学附属儿童医院初诊并接受CCLG-ALL-2008方案治疗的急性B淋巴细胞白血病患儿进行回顾性分析，明确化疗疗程拖延与复发的关系，并探究导致疗程拖延引起复发的原因，随访至2019年7月。.疗程拖延周数对复发率的影响具有统计学意义（P=0.034），风险率比提示大于4周影响显著。第12周微小残留病阳性（1×10-4≤微小残留病≤1×10-2）对复发率的影响亦有统计学意义（P=0.041）。在疗程拖延原因中，单纯骨髓抑制对复发率的影响有统计学意义（P=0.01）。.疗程拖延4周以上、第12周微小残留病阳性和单纯骨髓抑制是影响复发的独立预后因素。.

To observe the efficacy of chimeric antigen receptor T cell (CAR-T) in the treatment of children with refractory/recurrent B acute lymphocytic leukemia (B-ALL).Thirty-two patients with r/r B-ALL were treated by CAR-T, the recurrence and death respectively were the end point events to evaluate the efficacy and safety of CAR-T.The median age of the patients was 7.5 (2-17.5) years old; 40 times CAR-T were received in all patients and the median number of CAR-T was 0.9×107/kg; efficacy evaluation showed that 2 cases died before the first evaluation. Thirty patients showed that 3, 6, and 9-moth RFS was (96.3±3.6)%, (81.4±8.6)% and (65.3±12.5)%, respectively, while 3, 6, and 9-month OS was all 100%, and 12, 24-month OS was (94.7±5.1)% and (76±12.8)%. BM blasts≥36% before reinfusion and ferritin peak≥2 500 ng/ml within two weeks of CAR-T cell reinfusion were associated with recurrence. Adverse reactions mainly included cytokine release syndrome (CRS) and CART-cell-related encephalopathy syndrome (CRES), CRS appeared in 26 patients within a week of CAR-T cell reinfusion. CRES reaction was detected in 12 patients. Eighteen patients received intravenous drip of tocilizumab, among them, 12 combined with glucocorticoid. CRS and CRES reactions were relieved within one week after treatment. Hormone dosage was related to the duration of remission in patients, and the cumulative dose of methylprednisolone≥8 mg/kg showed a poor prognosis.CAR-T is a safe and effective treatment for r/r B-ALL, most CRS and CRES reactions are reversible. BM blasts ≥36% before reinfusion and cumulative dose of methylprednisolone ≥8 mg/kg after reinfusion both affect the therapeutic effect. Ferritin≥2 500 ng/ml within two weeks after reinfusion is related to disease recurrence and is an independent prognostic risk factor.嵌合抗原受体T细胞治疗儿童难治/复发急性B淋巴细胞白血病的疗效.观察嵌合抗原受体（CAR）T细胞治疗儿童难治/复发急性B淋巴细胞白血病（r/r B-ALL）的疗效及不良反应.回顾性分析32例r/r B-ALL患儿行CAR-T细胞治疗的临床资料，分别以复发、死亡为终点事件，评估CAR-T细胞治疗r/r B-ALL的疗效及安全性.2017年7月至2020年1月，共32例r/r B-ALL患儿接受CAR-T细胞治疗，中位年龄7.5（2-17.5）岁。32例患儿共回输CAR-T细胞40次，回输中位CAR-T细胞总量0.9×107/kg（患儿体重）。结果显示，2例患儿在评估前死亡，其余30例患儿3、6、9个月RFS分别为（96.3±3.6）%、（81.4±8.6）%和（65.3±12.5）%；3、6、9个月OS均为100%，12个月OS（94.7±5.1）%，24个月OS（76±12.8）%。回输前骨髓肿瘤负荷≥36%、CAR-T细胞回输2周内铁蛋白峰值≥2 500 ng/ml与复发相关。不良反应主要为细胞因子释放综合征（CRS）和CAR-T治疗相关脑病综合征（CRES），26例在CAR-T细胞回输1周内出现CRS反应，12例出现CRES反应。18例给予托珠单抗静滴，其中12例联合糖皮质激素，CRS反应及CRES反应经治疗后大多1周内缓解。激素使用剂量与患儿持续缓解时间相关，甲泼尼龙累积剂量≥8 mg/kg预后不佳.CAR-T细胞治疗儿童r/r B-ALL有效且安全，CRS及CRES反应经治疗后均可缓解；回输前骨髓肿瘤负荷≥36%、回输后甲泼尼龙累积剂量≥8 mg/kg预后不佳；回输后两周内铁蛋白峰值≥2 500 ng/ml与疾病复发相关，且为独立预后危险因素.

To analyze the efficacy of children with B-cell acute lymphoblastic leukemia (B-ALL) without prognostic fusion genes treated by CCLG-ALL 2008, and investigate the related factors affecting the recurrence of the patients.B-ALL patients without prognostic fusion genes treated by the protocol of CCLG-ALL 2008 in our hospital from March 2008 to December 2012 were retrospectively analyzed. Follow-up time was ended in August 31, 2019. The median follow-up time was 92 months (range 0-136 months). Kaplan-Meier was used to detect the RFS, and COX multivariate regression analysis was employed to identify the independent factors affecting the recurrence of the patients.There were 140 males and 99 females enrolled in this study. The ratio of male to female was 1.41∶1. The median age was 4.4 years old and the median number of WBC at initial stage was 4.98×10Age, initial WBC, and bone marrow MRD at the 12th week were correlated with recurrence in children with B-ALL without prognostic fusion genes, which can be used as prognostic indices of recurrence risk in clinical.无预后标志性基因的急性B淋巴细胞白血病患儿 随访10年复发因素的分析.通过分析CCLG-ALL 2008方案治疗无预后标志性融合基因的急性B淋巴细胞白血病（B-ALL）患儿的疗效，并分析影响复发的相关因素.回顾性分析239例于2008年3月至2012年12月在苏州大学附属儿童医院诊断为B-ALL、多重RT-PCR未检测到预后标志性融合基因，且接受CCLG-ALL 2008方案化疗的患儿，随防到2019年8月31日，中位随访时间92（0-136）个月。Kaplan-Meier分析无复发生存时间（RFS），COX多因素回归分析复发相关的独立因素.239例患儿中男性140例，女性99例，男女比例1.41∶1，中位诊断年龄4.4岁，初诊白细胞中位数是4.98×10年龄、初诊白细胞计数、第12周骨髓MRD与无预后标志性融合基因B-ALL患儿复发相关，在临床工作中可作为评估患儿复发风险的预后指标.

Description, A stimulated proinflammatory feature of leukemic progenitors facilitates their survival and immune evasion after chemotherapy., Chemotherapy can effectively reduce the leukemic burden and restore immune cell production in most acute myeloid leukemia (AML) cases. Nevertheless, endogenous immunosurveillance usually fails to recover after chemotherapy, permitting relapse. The underlying mechanisms of this therapeutic failure have remained poorly understood. Here, we show that abnormal IL-36 production activated by NF-κB is an essential feature of mouse and human leukemic progenitor cells (LPs). Mechanistically, IL-36 directly activates inflammatory monocytes (IMs) in bone marrow, which then precludes clearance of leukemia mediated by CD8+ T cells and facilitates LP growth. While sparing IMs, common chemotherapeutic agents stimulate IL-36 production from residual LPs via caspase-1 activation, thereby enabling the persistence of this immunosuppressive IL-36–IM axis after chemotherapy. Furthermore, IM depletion by trabectedin, with chemotherapy and PD-1 blockade, can synergistically restrict AML progression and relapse. Collectively, these results suggest inhibition of the IL-36–IM axis as a potential strategy for improving AML treatment.

BACKGROUND Timely reconstitution of a donor-derived immune system is important for recovery and long-term survival of patients after allogeneic hematopoietic stem cell transplantation (HSCT). We describe a case of Wiskott–Aldrich syndrome (WAS) treated by umbilical cord blood transplantation (UCBT) with atypical immune reconstruction. CASE SUMMARY A 1-year-old Chinese male infant was diagnosed with WAS. WAS gene sequencing identified the mutation c.777 + 1G>A (IVS8). On August 8, 2017, he was admitted to our hospital for HSCT. We selected an unrelated Human leukocyte antigen 6/10-matched donor for UCBT. After HSCT, the immune reconstitution process was atypical, the lymphocytes reached 0.5 × 109/L on day 23, and the neutrophils reached 0.5 × 109/L on day 34. The patient’s recovery throughout the year was good. CONCLUSION An increase in lymphocytes (especially T cells) earlier than granulocytes may be a marker of a good prognosis in UCBT.

To analyze the efficacy of CCLG-ALL-2008 protocol and the related factors of treatment failure in children with acute lymphoblastic leukemia (ALL).The clinical data of 400 children newly-diagnosed ALL in Children's Hospital of Soochow University from March 1, 2008 to December 31, 2012 was retrospectively analyzed. All the children accepted CCLG-ALL-2008 protocol, and were followed-up until October 2019. The dates of relapse, death and causes of death were recorded. Treatment failure was defined as relapse, non-relapse death, and secondary tumor.Following-up for 10 years, there were 152 cases relapse or non-relapse death, the treatment failure rate was 38%, including 122 relapse (80.3%), 30 non-relapse deaths (19.7%) which included 7 cases (4 cases died of infection and 3 cases died of bleeding) died of treatment (23.3% of non-relapse deaths), 8 cases died of minimal residual disease (MRD) continuous positive (26.7% of non-relapse deaths) and 15 cases died of financial burden (50% of non-relapse deaths). According to the relapse stage, 37 cases (30%) in very early stage, 38 cases (31%) in early stage, and 47 cases (39%) in late stage, while according to the relapse site, 107 cases relapsed in bone marrow, 3 cases in testis, 3 cases in central nervous system (CNS), 5 cases in bone marrow plus testis and 4 cases in bone marrow plus CNS. Bone marrow relapse was the main cause of death in 89 cases, followed by nervous system. Initially diagnosed WBC count (≥50×10Relapse is the main cause of treatment failure in children with ALL. The initially diagnosed WBC count, immunophenotype and MRD at week 12 were the independent prognostic factors for relapse of the patients. Financial burden accounts for a large proportion of non-relapse death.儿童急性淋巴细胞白血病治疗失败的相关因素分析.分析儿童急性淋巴细胞白血病（ALL）接受CCLG-ALL-2008方案治疗的临床疗效和治疗失败的相关因.回顾性分析2008年3月1日至2012年12月31日在苏州大学附属儿童医院新诊断为ALL并接受CCLG-ALL- 2008方案治疗的400例患儿的临床资料，并随访到2019年10月，记录患儿复发和死亡时间及死亡原因。治疗失败定义为复发、非复发性死亡和二次肿瘤.随访10年，共有152例患儿出现复发或者非复发性死亡，治疗失败率为38%，其中复发122例（80.3%），非复发性死亡30例（19.7%）。非复发性死亡包括治疗相关性死亡7例（占23.3%，感染死亡4例，出血死亡3例），骨髓微小残留病灶（MRD）持续阳性8例（占26.7%），经济原因放弃治疗15例（占50%）。 按照时间构成，极早期复发37例（30%），早期复发38例（31%），晚期复发47例（39%）。按照复发部位构成，单纯骨髓复发107例，单纯睾丸复发3例，单纯中枢神经系统复发3例，骨髓+睾丸复发5例，骨髓+中枢神经系统复发4例。死亡的89例患儿以骨髓复发为主，其次是神经系统。初诊白细胞数≥50×10复发是治疗失败的最主要原因，初诊白细胞数、免疫表型和第12周骨髓MRD是复发的独立预后因素。因经济因素导致治疗失败是无复发治疗失败的主要原因.

To study the difference of long non coding RNA (lncRNA) expression profile in bone marrow specimens of children with acute leukemia (AL) and other hematological disease children with normal bone marrows as controls, to screen the lncRNA related with childhood hematological diseases, and to explore the expression of lncRNA AC002454.1 and its clinical significance in AL children.The microarray gene chip technology was used to statistically analyze the lncRNA in bone marrow cells of newly diagnose AL children and control children. Ninty-seren differentially expressed lncRNAs were selected. The bone marrow specimens of ALL children (21 cases), AML children (22 cases) and control children (21 cases) were verified and compared by using qRT-PCR; then the lncRNA with maximum differential expression-lncRNA AC002454.1 was selected and used to analyze the relation of relative expression level with clinical indicators.The microarray gene chip detection showed that 1 884 differentially expressed lncRNA were found in ALL children, and 4 289 differentically expressed lncRNA were found in AML children. The results confirming these differentically expressed lncRNA by qRT-PCR showed that 9 lncRNA expression were significantly up-regulated in ALL children, and 12 lncRNA expression were significantly up-regulated in AML children. Among these up-regulated lncRNA, the difference of AC002454.1 expression was most significant in ALL and AML children (P＜0.05, P＜0.01). The detection showed that there was a significant difference, in AC002454.1 relative expression level of newly diagnosed T-ALL and B-ALL children (P＜0.01), moreover, this difference also was found in ALL and AML children (P＜0.05). The detection analysis showed that there was no statistical difference in AC002454.1 relative expression level among the different sex, age, WBC count at initial diagnosis, chromosome, fusion gene, and risk stratification (P＞0.05 for all).The lncRNA expression profile of AL children has been gained by using the lncRNA microarray gene chip technicology. AC002454.1 the significantly high expression exist in AL children, which relates with immunotyping and prognosis of AL children in a certain degree.长链非编码RNA AC002454.1在急性白血病患儿中的表达及临床意义.研究长链非编码RNA（long non-coding RNA, lncRNA）在急性白血病（acute leukemia, AL）患儿和骨髓正常的其它血液病患儿（作为对照）标本中的表达谱的差异, 筛选与AL患儿相关的lncRNA, 探讨lncRNA AC002454.1在AL患儿中的表达及临床意义.采用Microarray基因芯片技术对初诊AL儿童和对照儿童骨髓细胞的lncRNA进行统计分析。选取差异表达显著的lncRNA 97个, 对急性淋巴细胞白血病（acute lymphoblastic leukemia, ALL）患儿（21例）、急性髓细胞白血病（acute myelocytic leukemia, AML）患儿（22例）和对照儿童（21例）样本采用实时荧光定量PCR（qRT-PCR）技术进行验证并比较。并选择其中差异最大的lncRNA AC002454.1分析其相对表达量与临床指标的关系.经Microarray基因芯片检测, ALL患儿差异表达的lncRNA 1 884个、 AML患儿差异表达的lncRNA 4 289个。采用qRT-PCR对异常表达lncRNA进行验证的结果显示, ALL组中表达显著上调9例、AML组中表达显著上调12例, 其中AC002454.1在ALL、AML患儿表达差异最显著（P＜0.05, P＜0.01）。AC002454.1的相对表达量在初诊ALL免疫分型T系与B系中有显著差异（P＜0.01）、ALL和 AML 组有显著差异（P＜0.05）, AC002454.1相对表达量在患儿性别、年龄、初诊时WBC计数、染色体、融合基因、危 险度分层之间无显著差异（P值均＞0.05）.用长链非编码RNA芯片分析获得了AL患儿的lncRNA表达谱, AC002454.1在AL患儿中异常高表达, 其表达量与AL患儿的免疫分型及判断患者预后有一定相关性.

Background: Glycolysis, a multi-step enzymatic reaction, is considered to be the root of cancer development and progression. The aim of this study is to figure out the glycolytic enzyme, phosphoglycerate kinase 1 (PGK1) whether participate in the progression of acute myeloid leukemia (AML) and its possible mechanisms. Methods: Four datasets (GSE106096, GSE75086, GSE107968 and GSE106748) containing 30 leukemic blast cell samples of AML at diagnosis, 17 leukemic blast cell samples of AML relapse and 3 bone marrow CD34+ cell samples of healthy donors were downloaded from Gene Expression Omnibus (GEO) database and PGK1 was screened out as a potential survival biomarker in AML. Then we did a series of clinical sample verifications and gene set enrichment analysis (GSEA) focusing on PGK1. We further knocked down expression of PGK1 in myelogenous leukemia cell lines and explored its potential effects. Results: PGK1 expression was up-regulated among AML at diagnosis versus healthy control, AML relapse versus AML at diagnosis and AML relapse versus healthy control datasets. Through a serial of bioinformatic analyses (differentially expressed genes [DEGs] selection, function and pathway enrichments and protein-protein interaction [PPI] network establishment), PGK1 was identified as the most meaningful gene in AML progression. Furthermore, the generally high expression of PGK1 was confirmed in AML samples comparing with healthy controls in our single center and the high-expression PGK1 was associated with a comparatively low complete remission (CR) rate, a significantly high 5-year cumulative incidence of relapse (CIR), a poor 5-year event-free survival (EFS) rate, and a poor 5-year overall survival (OS) rate. The GSEA revealed that high-expression PGK1 in AML was associated with many pathways including cytosolic DNA sensing, pentose phosphate, base excision repair and DNA replication. In vitro, the transfected U937 and K562 cells with PGK1 knock-down showed decreased cell viability and increased apoptotic rate. PGK1 inhibition could greatly decrease the half maximal inhibitory concentrations (IC50) of cytarabine (Ara-C) and daunorubicin (DNR) in U937 and K562 cell lines.Conclusions: High-expression PGK1 was associated with poor prognosis in AML. PGK1 may serve to predict the AML progression and provide a novel therapeutic target for AML.

To investigate whether miRNA-30a is involved in the pathogenesis of ITP by affecting the differentiation of Th17 cells, and to explore its possible mechanism of miRNA-30a involved in the pathogenesis of ITP through the verification of the target gene SOCS3 for the prediction of miRNA-30a.Firstly, a chronic ITP mouse model was established. The expression of miRNA-30a and RORγt in the spleen mononuclear cells were detected and their correlation were analyzed. Secondly, the luciferase vector containing 3'UTR of the target gene and green fluorescent vector containing miRNA were constructed. Luciferase fluorescence detection, real-time fluorescent quantitative PCR (qPCR) and Western blot were used to verify whether SOCS3 is the target gene of miRNA-30a.The platelet count of mice in experimental group decreased to below 20% of normal ones after 48 hours of injection of anti-mouse platelet serum (APS), which was maintained for 14 days at least; the expression of miRNA-30a and RORγt in the spleen mononuclear cells in experimental group were higher than those in the control group(P＜0.05), moreover, there was a positive correlation between them (r=0.54); the activity of luciferase in PMDH-GFP-miRNA-30a and pMIR-report-UTR was significantly lower than that in PMDH-GFP empty plasmid and pMIR-report-UTR(P＜0.05); The expression of SOCS3 at mRNA and protein level was not different from that in the control group.Chronic ITP mouse model has been established successfully; miRNA-30a expression in spleen mononuclear cells of ITP mouse increase, and positively correlated with the expression of RORγt, which contribute to the pathogenesis of ITP by affecting the differentiation of Th17 cells; SOCS3 is able to bind to the target site of miRNA-30a, but might not be its functional target gene.miRNA-30a通过介导Th17细胞分化影响免疫性血小板减少性紫癜发病的初步探讨.探讨miRNA-30a是否通过影响Th17细胞分化参与ITP发病；通过对miRNA-30a预测靶基因SOCS3的验证，进一步探讨miRNA-30a参与ITP发病的可能机制.建立慢性ITP小鼠模型，检测其脾脏单个核细胞中miRNA-30a、 RoRγt的表达并分析两者相关性；构建含靶基因mRNA 3′UTR的荧光素酶表达载体及含有miRNA的绿色荧光载体，通过Luciferase荧光检测、实时荧光定量PCR（qPCR）、Western blot验证SOCS3是否为miRNA-30a的靶基因.实验组小鼠血小板计数在注射抗小鼠血小板血清（APS）48 h后下降至正常的20%，且至少可维持14 d。实验组小鼠脾脏单个核细胞中miRNA-30a及RoRγt表达较对照组升高（P＜0.05），且两者存在正性相关（r=0.54）。pMDH-GFP-miRNA-30a与pMIR-report-UTR实验组的荧光素酶的活性明显低于pMDH-GFP空质粒与pMIR-report-UTR的对照组（P＜0.05）。转染miRNA-30a质粒的EL4细胞中SOCS3在mRNA及蛋白水平与对照组均无差异.miRNA-30a在ITP小鼠脾脏单个核细胞中表达升高，且与RORγt表达呈正相关性，它可能通过影响Th17细胞分化参与ITP发病；SOCS3能与miRNA-30a靶位点结合，却不是其功能靶基因.

Gastrointestinal adenocarcinomas (GIAC) of the tubular gastrointestinal (GI) tract including esophagus, stomach, colon, and rectum comprise most GI cancers and share a spectrum of genomic features. However, the unified epigenomic changes specific to GIAC are poorly characterized. Using 907 GIAC samples from The Cancer Genome Atlas, we applied mathematical algorithms to large-scale DNA methylome and transcriptome profiles to reconstruct transcription factor (TF) networks and identify a list of functionally hyperactive master regulator (MR) TF shared across different GIAC. The top candidate HNF4A exhibited prominent genomic and epigenomic activation in a GIAC-specific manner. A complex interplay between the HNF4A promoter and three distal enhancer elements was coordinated by GIAC-specific MRTF including ELF3, GATA4, GATA6, and KLF5. HNF4A also self-regulated its own promoter and enhancers. Functionally, HNF4A promoted cancer proliferation and survival by transcriptional activation of many downstream targets, including HNF1A and factors of interleukin signaling, in a lineage-specific manner. Overall, our study provides new insights into the GIAC-specific gene regulatory networks and identifies potential therapeutic strategies against these common cancers. Significance: These findings show that GIAC-specific master regulatory transcription factors control HNF4A via three distal enhancers to promote GIAC cell proliferation and survival.

BackgroundsGastrointestinal adenocarcinomas (GIACs) of the tubular GI tract including esophagus, stomach, colon and rectum comprise most GI cancers and share a spectrum of genomic features. However, the unified epigenomic changes specific to GIACs are less well-characterized.We applied mathematical algorithms to large-scale DNA methylome and transcriptome profiles to reconstruct transcription factor (TF) networks using 907 GIAC samples from The Cancer Genome Atlas (TCGA). Complementary epigenomic technologies were performed to investigate HNF4A activation, including Circularized Chromosome Conformation Capture (4C), Chromatin immunoprecipitation (ChIP) sequencing, Whole Genome Bisulfite Sequencing (WGBS), and Assay for Transposase-Accessible Chromatin (ATAC) sequencing. In vitro and in vivo cellular phenotypical assays were conducted to study HNF4A functions.ResultsWe identified a list of functionally hyperactive master regulator (MR)TFs shared across different GIACs. As the top candidate, HNF4A exhibited prominent genomic and epigenomic activation in a GIAC-specific manner. We further characterized a complex interplay between HNF4A promoter and three distal enhancer elements, which was coordinated by GIAC-specific MRTFs including ELF3, GATA4, GATA6 and KLF5. HNF4A also self-regulated its own promoter and enhancers. Functionally, HNF4A promoted cancer proliferation and survival by transcriptionally activating many downstream targets including HNF1A and factors of Interleukin signaling in a lineage-specific manner.ConclusionWe use a large cohort of patient samples and an unbiased mathematical approach to highlight lineage-specific oncogenic MRTFs, which provide new insights into the GIAC-specific gene regulatory networks, and identify potential therapeutic strategies against these common cancers.

To investigate the gene mutation of patients with WAS gene defect and its correlation with clinical manifestations.Thirty-one patients consulted in Children's Hospital of Soochow University from January 2013 to February 2018 were enrolled in this study. The hot pot mutations of WAS gene in 31 patients were detected and related clinical phenotypes were analyzed retrospectively.All patients were male. The median onset age was 1 month (range, 0-83 months). Nine mutants were reported as novel mutations among 25 mutants detected in 31 patients, including c.1234_1235dupCC, c.1093-1097delG, c.28-30dupC, c.436GT, c.273 + 10_273 + 11dupCC, c.995_996insG, c.1010TA, c.332_333delCC and c.683CT mutations. There were 25 cases of classic WAS which mutations included missense mutation, deletion mutation, insertion mutation, splicing mutation and nonsense mutation, 2 cases of X-linked thrombocytopenia (XLT) were induced by missense mutation, 1 case of intermittent X-linked thrombocytopenia (IXLT) was induced by splicing mutation, 2 cases of X-linked pancytopenia were induced by missense mutation. Intravenous immunoglobulin (IVIG) and glucocorticoid therapy in IXLT patient was effective, and remission could be sustained, platelets could be increased in the short-term in treated XLT patients, but only a small part of classic WAS patients(8.0%) showed transient response to it, the IVIG and glucocorticoid therapy did not improve the status of platelet in XLP patients. Immune laboratory examination showed that CD3WAS gene defect is an important basis for the diagnosis of WAS and related diseases. IVIG plus glucocorticoid therapy is less effective for fewer patients, the HSCT is an effective treatment for WAS.

To investigate the mechanism of Paris forrestii (Takht.) H. Li (PCT3)-suppressing the proliferation of HL-60, K562, KG-1 and HT-93 cells.cute myeloid leukemia cell lines such as HL-60, K562, KG-1 and HT-93 were treated with Paris forrestii (Takht.) H. Li (PCT3) for 24, 48, and 72 h, and MTT assay was employed to determine the cells proliferation. Meanwhile, the apoptosis of K562, HL-60, KG-1 and HT-93 cells were detected by flow cytometry after PCT3 (Control, 4 μg/ml, 8 μg/ml) treated for 24 h and the Western blot was performed to detect the expression of PARP,Caspase-3, MCL-1, BAX, BCL-2, P53, and P27. GAPDH was used as an internal loading control.MTT assay showed that Paris forrestii (Takht.) H. Li (PCT3) significantly inhibited the proliferation of HL-60, K562, KG-1 and HT-93 cells in concentration and time-dependent manners. Compared with the control group, the leukemia cell viabilities were significantly suppressed (r =0.9436; r =0.8623; r =0.9922; r =0.8918). Paris forrestii (Takht.) H. Li (PCT3) induced apoptosis of leukemia cells in a concentration dependent manner, compared with the control group (P＜0.05 or P＜0.01). Western blot revealed that PARP, a major enzyme in DNA damage repair, and Caspase-3 another one of the major executive apoptotic enzymes were cleaved in cell lines examined, and this cleavage was concentration dependent. Anti-apoptotic proteins such as MCL-1 and BCL-2 were down regulated by Paris forrestii (Takht.) H. Li (PCT3), and Pro-apoptotic protein BAX was upregulated. And the protein of tumor suppressor gene P53 and its downstream signaling protein P27 increased.Paris forrestii (Takht.) H. Li (PCT3) can inhibit the proliferation of leukemia cells by activating endogenous apoptosis pathway, and provide a potential new drug selection for clinical treatment of AML leukemia.

Treatment and prognosis of Fanconi anaemia (FA) and acquired aplastic anaemia (AA) differ. However, delayed and inappropriate treatments are administered in FA due to its similarities to AA in presentation. The objective of the current study was to elucidate differences between the molecular mechanisms underlying FA and AA as well as to identify biomarkers and pathways associated with FA via bioinformatics analyses. Proteomic data were obtained from bone marrow samples of patients with FA and AA. Gene ontology analysis was performed using a Database for Annotation, Visualization and Integrated Discovery. KEGG pathway enrichment analyses were conducted using the ClueGO plug-in in Cytoscape. A DEP-associated protein-protein interaction (PPI) network was constructed using STRING and visualized in Cytoscape. A total of 114 DEPs, including 71 upregulated proteins and 43 downregulated proteins, were present in the FA samples, compared with those in the AA samples. Upregulated proteins were enriched in the nucleosome assembly, canonical glycolysis, glycolytic process, and the glycolysis/gluconeogenesis pathway, whereas downregulated proteins were enriched in relation to immune response, negative regulation of apoptosis, proteolysis and CoA biosynthesis. Eight hub proteins with a high degree of connectivity were obtained as follows: alpha-enolase (ENO1), HSP90AA1, phosphoglycerate kinase 1 (PGK1), HSP90AB1, ACTC1, ACTBL2, EEF1A1 and CFL1. Upregulation of ENO1 and CFL1 in patients with FA was confirmed through a WB experiment, and substantiated by the results of data analyses. Bioinformatics analyses are useful for identification of biomarkers and pathways associated with FA and AA. Some crucial DEPs, such as ENO1, PGK1, ACTC1, ACTBL2, EEF1A1 and CFL1, may play an important role in FA and show potential as serological markers for its early diagnosis.

To evaluate the therapeutic safety and efficacy of VDMP re-induction regimen in Chinese children with relapsed acute lymphoblastic leukemia (ALL).Forty-one patients with relapsed ALL were prospectively enrolled in this study. All the patients were distributed in 3 children's hospitals and treated with VDMP regimen as the first re-induction chemotherapy. Therapeutic efficacy and side-effects were analyzed.The ratio of male to female was 27:14. The median age was 7.9 (2.2-15.4) years old. Patients relapsed at very early, early, and late stage were 7 cases, 11 cases, and 23 cases, respectively.The immunophenotype analysis showed that 38 cases were B-ALL, and 3 cases were T-ALL. All patients suffered from grade 4 of neutropenia and forty(97.6%) cases got infection, of them one case died. Thirty-nine(95.1%) cases had nonhematologic adverse event at least one organ involved grade 3 in 38 out of 41 cases, the VDMP therapy was completed, 34(89.5%) cases achieved a complete remission (CR), 1 case achieved partial remission(PR), and 3 cases didn't get remission. Follow-up data of 38 cases with completing VDMP chemotherapy were obtained, only one case was lost. Among 37 cases available for evaluation, 16 cases received allo-hematopoietic stem cell transplantation(allo-HSCT) after chemotherapy, and 13 patients survived, while 21 cases did not receive allo-HSCT(treated with chemotherapy only), and 8 patients survived.The overall survival rate of allo-HSCT group was significantly higher than that of those treated with chemotherapy only(P0.05).VDMP re-induction regimen is effective and well tolerable for pafients in the treated children with relapsed ALL. After remission, allo-HSCT is recommended with the aim of long survival.

Increasing occurrence of posterior reversible encephalopathy syndrome has been reported in children with acute lymphoblastic leukemia. However, the etiology of posterior reversible encephalopathy syndrome is not clear. To study the possible pathogenetic mechanisms and treatment of this complication, we reported 11 cases of pediatric acute lymphoblastic leukemia who developed posterior reversible encephalopathy syndrome after induction chemotherapy. After appropriate treatment, the clinical symptoms of posterior reversible encephalopathy syndrome in most cases disappeared even though induction chemotherapy continued. During the 1-year follow-up, no recurrence of posterior reversible encephalopathy syndrome was observed. Although the clinical and imaging features of posterior reversible encephalopathy syndrome may be diverse, posterior reversible encephalopathy syndrome should be recognized as a possible important complication of acute lymphoblastic leukemia when neurologic symptoms appear. In line with previous reports, our study also indicated that posterior reversible encephalopathy syndrome was reversible when diagnosed and treated at an early stage. Thus, the occurrence of posterior reversible encephalopathy syndrome should be considered and investigated to optimize the early induction scheme of acute lymphoblastic leukemia treatment.

Histone modification is dysregulated in various types of cancers, including hematological malignancies. However, the expression profile of histone-modifying enzymes in pediatric acute monoblastic leukemia (AML FAB M5) has not been investigated. In this study, we evaluated the mRNA expression profile of 85 genes that encode enzymes involved in histone-modification in 27 pediatric AML FAB M5 samples by using a novel real-time PCR array. We obtained a gene cluster consisting of a total of 28 genes (15 up-regulated genes and 13 down-regulated genes). This gene signature revealed up-regulated expression of putative oncogenes GCN5L2, SETD8, KDM5C, AURKA and AURKB, and downregulated putative tumor suppressor genes (TSGs) EP300, PRMT3, PRMT8 and NOTCH2. We investigated possible biological interactions between differentially expressed genes using ingenuity pathway analysis (IPA) and found 12 significant networks. Among these, gene expression, cancer, and embryonic development showed the highest number of networks with 39 focus molecules and had an associated significance score of 68. Further, Rb, CDKN2C, and E2F1 were found to be upstream regulators of histone-modifying enzymes. This study provides additional insights into the molecular pathogenesis of pediatric AML FAB M5. These genes represent interesting targets with potential for diagnostic, prognostic and therapeutic application in pediatric AML patients.

To investigate the effects and mechanism of CCL2 on the migration and invasion of human leukemia cell THP-1.The CCL2 gene was recombined with the transfer plasmid-PLVX and transfected into THP-1 cells. The CCL2 expression at RNA level was detected by RT-PCR, the CCL2 expression at protein level was determined by Western blot and ELISA, the influence of overexpression of CCL2 recombinant protein and THP-1 cells on the migration and invasion ability of THP-1 cells was analyzed by transwell migration and invasion tests, the PCR-array of migration-related cytokines was used to clarify the patential mechanism.With the Trans-Matrigel assay, the concentration of CCL2 in THP-1 transfected with CCL2 in the upper cells was higher than that in the lower cells, meanwhile, the invasion ability of CCL2-transfected THP-1 cells decreased. Increasing recombinant protein of CCL2 (rpCCL2) in the lower cells promoted migration of THP-1 cells. Migration RT2 profiler PCR array showed that the cells treated with rpCCL2 had higher levels of expression of CCL2, EPX, SPP1, CX3CL1 and CXCL13, as compared with control group.CCL2 affects the migration and invasion of THP-1 by autosecreting a series of inflammatory chemokines.

Polymorphisms in folate pathway genes may influence susceptibility to pediatric acute lymphoblastic leukemia (ALL). This case-control study was undertaken to analyze the association of genetic polymorphisms (677C>T and 1298A>C) of methylenetetrahydrofolate reductase (MTHFR) and reduced folate carrier (RFC1) (80G>A) with the risk of pediatric ALL in China. A total of 176 pediatric ALL patients and 170 matched healthy subjects (as controls) were included and DNA was extracted from the peripheral blood. SNaPshot single nucleotide polymorphism typing was used to determine the genotypes of MTHFR 677C>T, MTHFR 1298A>C, and RFC1 80G>A. All statistical analyses were conducted with SAS software (version 9.2; SAS Institute). There were no significant differences in the genotype and allele frequencies of MTHFR 677C>T, MTHFR 1298A>C, or RFC1 80G>A between patients and controls. No significant correlation was found between the combined genotypes of these polymorphisms and the risk of developing ALL in this study. Furthermore, no significant differences were observed for 677C>T and 1298A>C frequencies between the control and case groups. There was no association between MTHFR 677C>T, MTHFR 1298A>C, or RFC1 80G>A gene polymorphisms and risk of pediatric ALL in the Han Chinese population.

To investigate the percentage of blasts with the CD34The percentage of CD34The median percentage of CD34The percentage of CD34

To investigate the effect of C3AR1 on migration and invasion of HL-60 cells and its mechanism.The HL-60 cells overexpresing C3AR1 was constructed, and the HL-60-C3AR1 cell line was validated by real-time PCR and Western blot. The migration and invasion assay were performed by using transwell system, the PCR Array and flow cytometry were employed to reveal the potential mechanisms.C3AR1 gene was significantly expressed in AL cell lines, especially in acute myeloid leukemia(AML cell lines). C3a and SDF-1 together promoted migration and invasion of C3AR1 over-expressed HL-60 cells. The PCR array detection found that 16 genes (including CCL2) were significantly upregulated and 4 genes were significantly down-regulated. However, the expression of CXCR4 did not significantly change after treated by C3a and SDF-1 together.Over-expression of C3AR1 shows the effects on the migration and invasion of HL-60 cells under the treatment of C3a and SDF-1 together, which may be mediated by the regulation of CCL2 and other genes related to chemotactic factors.

To investigate the apoptotic effects of Hsp90 selective inhibitor 17-AAG on human leukemia HL-60 and NB4 cells and analyse its possible mechanism.CCK-8 assay was used to quantify the growth inhibition of cells after exposure to 17-AAG for 24 hours. Flow cytometrve with annexin V/propidium iodide staining was used to detect apoptosis of leukemia cells. Then Western blot was used to detect the activation of apoptosis related protein caspase-3 and PARP level. Gene expression profile of NB4 cells treated with 17-AAG was analyzed with real-time PCR arrays.The inhibition of leukemia cell proliferation displayed a dose-dependent manner. Annexin V assay, cell cycle analysis and activation of PARP demonstrate that 17-AAG induced apoptosis leukemia cells. Real-time PCR array analysis showed that expression of 56 genes significantly up-regulated and expression of 23 genes were significantly down-regulated after 17-AAG treatment.The 17-AAG can inhibit the proliferation and induce the apoptosis of leukemia cells. After leukemia cells are treated with 17-AAG, the significant changes of apoptosis-related genes occured, and the cell apoptosis occurs via activating apoptosis related signaling pathway.

Background The imbalance of Treg/Th17 cells is an important pathogenic factor for immune thrombocytopenic purpura (ITP). We previously reported miR-125a-5p targeted CXCL13 and participated in the process of ITP. In the present study, the role of miR-125a-5p in regulating Treg/Th17 ratio and its potential molecular mechanism were investigated. Method A total of 30 adults with ITP and 30 healthy subjects were included. MEG3 expression in peripheral blood derived CD4+ T cells from ITP patients and healthy subjects were detected by real-time PCR. In vitro experiments, the effects of inhibiting or overexpressing MEG3 on the expression of miR-125a-5p, Foxp3 and ROTγt in CD4+ T cells were investigated. Results MEG3 expression was increased in CD4+ T cells of patients with ITP. Dexamethasone decreased MEG3 expression level of CD4+ T cells in vitro . MEG3 directly interacted with miR-125a-5p and MEG3 overexpression inhibited miR-125a-5p expression in CD4+ T cells exposed to dexamethasone. MEG3 down-regulation or miR-125a-5p overexpression promoted Foxp3 expression and inhibited RORγt expression. Conclusion MEG3 interacted with miR-125a-5p and inhibited its expression, and MEG3/miR-125a-5p contributed to induce immune imbalance of Treg/Th17 in ITP.

AIM The aim of the study is to investigate the association of interferon gamma (IFN-γ) and interleukin-10 (IL-10) gene single nucleotide polymorphisms with the susceptibility of hemophagocytic lymphohistiocytosis (HLH) in Chinese children without known family history of HLH. PROCEDURE Forty children with HLH and 160 age- and gender-matched healthy controls from Xuzhou Children's Hospital were enrolled in the study. Serum IFN-γ and IL-10 levels were measured by enzyme linked-immunosorbent assay. Polymorphisms of the IFN-γ gene at position +874 and +2109, and IL-10 at position -1082 were analyzed by allele-specific PCR. RESULT Median serum concentrations of IFN -γ and IL-10 were significantly higher in children with HLH compared to healthy controls. The frequencies of IFN-γ +874 T/A and T/T genotypes, as well as T allele, were significantly higher in the HLH group compared with those in the control group. The frequencies of IL-10 -1082 G/A genotype and G allele were significantly increased in HLH patients compared with healthy controls. No significant difference was found in the distribution of IFN-γ +2109G/A genotypes between children with HLH and controls. CONCLUSION This study presents preliminary evidence for the association between IFN +874 T/A, T/T, IL-10 -1082 A/G genotypes, and HLH susceptibility in Chinese children with HLH.

To study the efficacy of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in children with severe aplastic anemia (SAA).A total of 11 children with SAA were treated with HLA matched siblings (n = 7), umbilical cord blood (n = 2) and haploidentical HSCT (n = 2).Among 11 children patients, 10 patients achieved engraftment, but 3 children patients experienced secondary graft failure, after donor lymphocyte infusions (DLI), they achieved engraftment again. One patient received cord blood transplantation and experienced primary graft rejection, but acquired autologous recovery. The median time for neutrophils to reach over 0.5 × 10(9)/L was 14 days (10-19 days) in the 9 children received bone marrow or bone marrow and peripheral blood allo-HSCT, while the median time for platelets to reach over 20 × 10(9)/L was 17 days (8-42 days). For the patient received double cord blood transplantation, the time of neutrophile and platelet level recovery was 16 days and 41 days, respectively.If HLA-matched sibling donor is available, allo-HSCT can be recommended as the first line of treatment for children with SAA. It is feasible for children with SAA to receive allo-HSCT from selective donor, including cord blood and haploidentical HSCT. Donor lymphocyte infusions can improve engraftment.

This study was purposed to explore the effectiveness of mixed transplantation of HLA mismatched bone marrow hematopoietic stem cells(HSC), peripheral blood HSC and umbilical cord blood HSC for treatment of pediatric blood diseases. From August 2012 to December 2012, five children with refractory hematological diseases in our hospital received allogeneic hematopoietic stem cell transplantation. The mixed grafts consisting of HLA-mismatched bone marrow HSC, peripheral blood HSC and umbilical cord blood HSC were used to observe the effects of umbilical cord blood HSC on the time of hematopoietic reconstruction of bone marrow, STR chimeric degrees, incidence of GVHD. and early transplant-associated complications. The results showed that all 5 children patients were grafted successfully with the median grafted time of 11 d for ANC0.5×10(9)/L and 10 d for Plt20×10(9)/L, respectively. On day 30, the STR-PCR test of peripheral blood showed a stable complete chimera. Five cases suffered from mild to moderate symptoms of GVHD, showing with I-II grade of skin GVHD and in which two cases suffered from diarrhea, showing I-II grade of intestinal GVHD. All the 5 patients had no liver function damage. One patient died of severe hemorrhagic cystitis and multi-site infection, and the remaining four cases survived so far on the current median follow-up time of 137 d (130 d-250 d). It is concluded that transplantation of the mixed HLA mismatched bone marrow HSC, peripheral blood HSC, with third-party cord blood HSC can increase the survival rate for pediatric patients with blood disease.

Acute lymphoblastic leukemia (ALL) is the most common childhood cancer, while glucocorticoid (GC) is a critical component in multi-agent chemotherapy protocols currently used for the treatment of ALL. The purpose of this study was to investigate the relationship between the glucocorticoid induction test and the clinical features and the prognosis of Chinese childhood ALL.The study recruited 309 hospitalized patients (187 male and 122 female) with childhood ALL, the sex, age, initial WBC count, immunophenotype, chromosome and gene expression were recorded. After diagnosis, all patients received GC induction test for 7 days. Then they were divided into prednisone good response (PGR) group and prednisone poor response (PPR) group according to the peripheral lymphoblast count on D8. Early responses to chemotherapy and treatment outcomes of the patients in the two groups were also analyzed.Of the 309 patients, 263 belonged to PGR group and 46 belonged to PPR group. Initial WBC count was higher in PPR group than in PGR group (86.30×10(9)/L vs. 30.97×10(9)/L, P0.01) . B lineage ALL showed more sensitive to GC than T-ALL (86.6% vs. 60%, P0.05). Different initial-risk-group's sensitivity to GC differed from one another (high-risk:51.4%, medium-risk: 82.7%, standard risk: 93.7%, P0.0125). There was no significant difference between two groups in chromosomal karyotypes (P0.05). BCR-ABL positive ALL showed lower sensitivity to GC (P0.05) , while MLL, TEL-AML1, E2A-PBX1 positive rates in two groups were of no statistical significance (P0.05). Bone marrow was reviewed on D15 and D33, and the CR rates in PGR group were significantly higher than that in PPR group (D15: 60.5% vs. 32.6%, D33: 94.6% vs. 73.3%, P0.01) ; Minimal residual disease (MRD) levels were examined on D33, W12, and both were much lower in PGR group (D33: P0.01, W12: P0.05). Of the PGR group 215 patients (81.7%) remained continuously in complete remission (CCR) while only 28 cases (60.9%) in PPR group did so. The CCR rate was much higher in PGR group than that in PPR group (P0.01).Closely related to clinical features and the outcomes of treatment, GC induction test is also an important prognostic factor in Chinese childhood ALL.

To investigate the methylation, expression and clinical significance of miR-203 in pediatric acute leukemia.The methylation status of miR-203 promoter CpG islands was detected with methylation-specific polymerase chain reaction. The expression of miR-203 was detected by Taqman real- time quantitative polymerase chain reaction. And the clinical significance of miR-203 in pediatric acute leukemia (ALL) was also analyzed.The promoter of miR-203 was unmethylated in all of 31 pediatric acute lymphoblastic leukemia, all of 15 pediatric acute myeloid leukemia (AML) and all of 23 controls. The relative expression levels of miR-203 in controls, pediatric acute leukemia, ALL and AML were 16.93±6.31, 48.97±10.38, 55.88±12.91, 24.28±9.10 respectively. The results indicated that miR-203 was significantly up- regulated in pediatric acute leukemia (P=0.011) and ALL (P=0.009), not in pediatric AML (P=0.514) compared with control. The expression of miR-203 was significantly related with the gender, immunophenotype, chromosome, fusion gene, BCR-ABL, SIL-TAL1 and prednisone experiment in pediatric ALL and the gender, chromosome, fusion gene, SIL-TAL1 in pediatric acute leukemia (P0.05). And in risk stratification pairwise comparisons, the expression of miR-203 in the medium-risk and high-risk groups appeared significantly different (P=0.022).miR-203 may not be regulated with methylation mechanism in pediatric acute leukemia. miR- 203 may be a protooncogene involved in the formation of pediatric acute leukemia and ALL. Further analyses indicated that high expression of miR-203 may be associated with poor prognosis of pediatric ALL and acute leukemia.

This study was aimed to investigate the curative effect and safety of human umbilical cord mesenchymal stem cells (hUCMSC) to treat acute graft-versus-host disease (aGVHD) of children after hematopoietic stem cell transplantation (HSCT). HUCMSC were isolated and cultured by collagenase digestion and passage culture. The 3rd to the 5th passage of hUCMSC were used for clinical treatment. Five cases of children acute leukemia achieved complete remission after chemotherapy. Two cases received HLA 3/6 loci matched haploidentical bone marrow HSCT. One case received HLA-matched sibling bone marrow and peripheral blood HSCT. One case received unrelated HLA 4/6 loci matched umbilical cord blood HSCT. One case received unrelated HLA 5/6 loci matched umbilical cord blood HSCT. The children received immunosuppressive therapy after III-IV aGVHD occurring. They received 0.5×10(6)/kg hUCMSC infusion when conventional therapy was ineffective. The results showed that 5 cases of children acute leukemia achieved hematopoietic reconstitution and developed the III-IV grade aGVHD. The five cases of children were infused with hUCMSC. The rash subsided, the liver function was normalized and the gastrointestinal symptoms were improved. The infusion-related adverse reaction did not happen. At present, the 5 children are in remission. It is concluded that allogeneic HSCT is an effective therapeutic method for children with acute leukemia. HUCMSC infusion can be safely and effectively used for the treatment of refractory aGVHD.

In order to study the potential of Venus, lentiviral vector, applied to acute myeloid leukemia, the recombinant vector Venus-C3aR was transfected into 293T packing cells by DNA-calcium phosphate coprecipitation. All virus stocks were collected and transfected into HL-60, the GFP expression in HL-60 cells was measured by flow cytometry. The expression level of C3aR1 in transfected HL-60 cells was identified by RT-PCR and flow cytometry. The lentiviral toxicity on HL-60 was measured by using CCK-8 method and the ability of cell differentiation was observed. The results indicated that the transfection efficacy of lentiviral vector on HL-60 cells was more than 95%, which meets the needs for further study. C3aR1 expression on HL-60 cells increased after being transfected with recombinant lentiviral vector. Before and after transfection, the proliferation and differentiation of cells were not changed much. It is concluded that the lentiviral vector showed a high efficacy to transfect AML cells and can be integrated in genome of HL-60 cells to realize the stable expression of interest gene. Meanwhile, lentiviral vector can not affect HL-60 cell ability to proliferate and differentiate.

Idiopathic thrombocytopenic purpura (ITP) is an acquired autoimmune disorder. Both impaired platelet production and T cell-mediated effects play a role in ITP thrombocytopenia. A Th1 polarization of the immune response, up-regulation of Th17 cells and decreased number of Treg cells have been demonstrated in ITP patients. High-dose dexamethasone was administered as first-line therapy in adult patients with ITP. However, the mechanism of effects of dexamethasone on ITP is still unclear. In this study, we tested the effectiveness of high-dose dexamethasone as initial treatment in adults with immune thrombocytopenic purpura. PBMCs were isolated from Donors, ITP and Treatment groups. T cell subsets were analyzed by FCM and transcriptional factors were checked by Real-time PCR. We found that dexamethasone returned the ratio of Th1/Th2 and the number of Th17 and Treg cells to the normal levels. Furthermore, we identified that dexamethasone corrected the T cell subset levels through inhibiting GATA3 and FOXp3 expression and promoting RORγt expression. Taken together, we reported a previously unrecognized mechanism on dexamethasone in the ITP treatment.

To investigate thiopurine S-methyltransferase (TPMT) activity and gene promoter polymorphism to probe its significance of individual chemotherapy in acute lymphoblastic leukemia (ALL) children.HPLC method was carried out to determine TPMT activity (n=100), which activity at newly diagnosed. At the same time determination of TPMT activity in healthy children (n=180), these children come from the health care clinic. Using online primer3 software design primers, PCR products were purified. To sequence TPMT gene of the patients with clinical events(n=30). According to the method to analysis of correlation between TPMT activity and toxicity.The average TPMT activities were (31.72±10.31) nmol·g⁻¹Hb·h⁻¹ and (30.70±9.67) nmol·g⁻¹Hb·h⁻¹ in ALL and healthy groups respectively, without gender differences of TPMT activities (P=0.45) in both groups. The TPMT activity with clinical events in newly diagnosed ALL patients (n=30) was (24.07±11.43) nmol·g⁻¹Hb·h⁻¹. There are significant differences of TPMT activities between severe bone marrow suppression [(20.96±7.24) nmol·g⁻¹Hb·h⁻¹] and ALL patients with clinical events groups (P0.05). The TPMT activity of (40.46±8.18) nmol·g⁻¹Hb·h⁻¹ in recurrence children was also significantly different (P0.05). TPMT activity in severe liver toxicity group was not significantly different (P=0. 930). Of TPMT gene sequencing in ALL patients with clinical events, only 3 children were heterozygosity mutations of TPMT*3C, while others homozygous genotype. There were significant differences of TPMT activities between heterozygosity genotype [(11.99±1.32) nmol·g⁻¹Hb·h⁻¹] and homozygous genotype groups [(24.95±11.32) nmol·g⁻¹Hb·h⁻¹] (P0.05). There were five kinds of variations at the vicinity of the promoter region of -100 of tandem repeats (VNTR) polymorphism（*V3/*V3、*V3/*V4、*V4/*V4、*V5/*V5、*V4/*V6）without significant differences of TPMT activities among five kinds (P=0.186).TPMT activity was related to the gene polymorphism. TPMT activity determination had prognostic value and guided individualized treatment.目的 探讨巯基嘌呤甲基转移酶（TPMT）活性和其遗传多态性在儿童急性淋巴细胞白血病（ALL）个体化治疗中的临床意义。方法 运用高效液相色谱法（HPLC）对100例初诊ALL患儿、180名健康儿童进行TPMT活性检测。采用在线Primer3 软件设计引物,对30例具有临床事件的患儿进行TPMT基因DNA测序。分析TPMT活性、遗传多态性与临床不良反应的相关性。TPMT活性以在单位时间(1 h)内单位质量的血红蛋白(Hb,g)催化生成的2-氨基-6-甲基巯基嘌呤（6-MTG）的量(nmol)表示。结果 100例初诊ALL患儿TPMT活性为（31.72±10.31）nmol·g⁻¹Hb·h⁻¹,180名健康儿童TPMT活性为（30.70±9.67）nmol·g⁻¹Hb·h⁻¹,两者TPMT活性差异无统计学意义（P=0.450）。19例发生严重骨髓抑制的患儿TPMT活性为（20.96±7.24）nmol·g⁻¹Hb·h⁻¹,6例ALL复发者初诊时TPMT活性为（40.46±8.18）nmol·g⁻¹Hb·h⁻¹,两者分别与30例具有临床事件的ALL患儿初诊时活性[（24.07±11.43）nmol·g⁻¹Hb·h⁻¹]相比差异具有统计学意义（P值均0.05）。5例化疗后发生严重肝脏毒性者初诊时TPMT活性为（23.60±7.48）nmol·g⁻¹Hb·h⁻¹,与30例ALL患儿初诊时活性相比差异无统计学意义（P=0.930）。对30例具有临床事件的ALL患儿TPMT基因DNA测序, 3例发生TPMT*3C杂合突变,其活性为（11.99±1.32 ）nmol·g⁻¹Hb·h⁻¹,27例呈野生纯合型,其活性为（24.95±11.32）nmol·g⁻¹Hb·h⁻¹,杂合子突变患儿TPMT活性明显低于野生纯合子型患儿,差异有统计学意义（P0.05）。启动子-100附近区域发现5种数目变异的串联重复（VNTR）基因型（*V3/*V3、*V3/*V4、*V4/*V4、*V5/*V5、*V4/*V6）,其平均活性分别为19.35、25.06、22.61、32.16、11.85 nmol·g⁻¹Hb·h⁻¹,各组间活性差异无统计学意义（P=0.186）。结论 TPMT基因多态性与其活性有一定的相关性,TPMT活性高低对于儿童ALL预后的判断和指导个体化用药有重要的临床意义。

To explore clinical and experimental features of 28 cases of childhood acute myeloid leukemia (AML) with 11q23/MLL gene rearrangements.Karyotypes of 234 cases of de novo childhood AML were analyzed using short-term culture of bone marrow cells and R-banding. The fusion transcripts involving MLL gene and partial tandem duplication of MLL (MLL-PTD) were detected by multiple reverse transcription polymerase chain reaction (RT-PCR) assay. Two cases with 11q23 translocation by karyotypic analysis but with negative result of multiple RT-PCR were studied with MLL-dual-color fluorescence in situ hybridization (D-FISH).R-banding karyotypic analysis has revealed 20 cases with 11q23 translocation (14 cases with M5, 4 cases with M4, 2 cases with M2), including 12 cases with t(9;11)(p22;q23), 3 cases with t(1;11)(q21;q23), 2 cases with t(6;11)(q27;q23), 1 case with t(11;19)(q23;p13), 1 with t(5;11)(q31;q23), and 1 with t(X;11)(q24;q23). Eighteen cases with 11q23 translocation having fusion transcripts involving MLL genes were confirmed with multiple RT-PCR; 2 cases showed negative results, but they were confirmed to have MLL rearrangements by D-FISH. MLL-PTD was also detected in 8 cases (4 cases M5, 2 cases M4, M2 and M6, one case each) from the other childhood AML cases. The total incidence of 11q23/MLL gene rearrangements was 11.97% (28/234), and most of patients(85.7%, 24/28) were M4/M5. The complete remission (CR) rate after treatment for the 28 cases with MLL rearrangements was 53.8%, the difference was significant by statistics (P0.05) compared with 90.5% for the control group (M4/M5 childhood AML with other karyotypic abnormalities or normal karyotype). Of them, 2 cases receiving intensive chemotherapy survived for 81 and 66 months, respectively, 4 cases receiving allogeneic stem cell transplantation survived for 21, 20, 16 and 11 months, respectively, and are still alive with CR. The medium survival (MS) time for 28 cases with 11q23/MLL rearrangements was 11 months, whereas the MS for control group was 15 months. The difference was not statistically significant(P0.05).The 11q23/MLL rearrangements is highly correlated with the occurrence of monocytic leukemia (M4 and M5). The 11q23 translocation and MLL-PTD are mutually exclusive, though both are indicative of poor prognosis. Intensive chemotherapy and allogeneic stem cell transplantation may ameliorate the clinical outcome. Multiple RT-PCR combined with karyotypic analysis and D-FISH are useful for screening the 11q23/MLL rearrangements in childhood AML.

To investigate the relationship between genetic polymorphism in exon 12 C1236T, exon 21 G2677T/A and exon 26 C3435T of the multidrug resistance 1 (MDR1) gene and the risk of childhood acute lymphocytic leukemia (ALL).A total of 176 patients with ALL and a cohort of 170 matched healthy subjects were included. SNaPshot SNP typing was used to determine the genotypes of MDR1 C1236T, G2677T/A, C3435T. Based on the clinical data, the relationship between genetic polymorphism of MDR1 and the risk of childhood ALL was analyzed.There was significant difference in the distribution of genotype of MDR1 C3435T between the group of controls and cases. The mutant homozygous TT genotype was found to be associated with occurrence of ALL (P = 0.000; OR = 4.504). The data show evidence of pairwise linkage disequilibrium between the three common SNPs (C1236T-G2677T/A-C3435T). The haplotypes of TTT, TGC, CGC and CAC were predominant. The haplotype CGT distributed significantly differently between the groups of controls and cases (P = 0.034). The frequency of the haplotype TTT/TTT in the high risk group was higher than the other groups (P = 0.037).The present findings suggest that 3435C→T polymorphism in MDR1 gene may be a genetic susceptibility factor for ALL. The haplotype of MDR1 (C1236T-G2677T/A-C3435T) could be the clinical parameter at diagnosis.

To explore the effect of down-regulation of insulin-like growth factor binding protein 7 (IGFBP7) on the proliferation and invasiveness of leukemia cell line U937 cells.Three pairs of double-strand siRNA targeting IGFBP7 gene were transfected into SMMC7721 cells to select the most efficient one for U937 cells. qRT-PCR and Western blot were used to detect the expression of IGFBP7 in U937 cells after transiently transfected with siRNA of IGFBP7. Cell proliferation, adhesion, trans-endothelial migration and invasion were performed in transfected cells and control groups.After transfected with siRNA of IGFBP7 in U937 cells, the ability of cell proliferation was significantly decreased at 24 h (0.580 ± 0.159) compared to that of parental cells and scramble negative control (1.049 ± 0.274, 0.946 ± 0.195, respectively) (P0.01). Adhesion of U937 cells transfected with IGFBP7 gene specific siRNA to ECV304 cells was significantly lower than that of the control groups (0.247 ± 0.031 vs 0.406 ± 0.023 and 0.395 ± 0.011) (P0.01). Transendothelial membrane of U937 cells into the bottom of the 24-well plate for experimental group were less than those in the control groups \[(0.387 ± 0.021)×10(5) vs (1.017 ± 0.031)×10(5) and (0.908 ± 0.027)×10(5)\]. Cells adherent to the matrigel for experimental group were less than those in the control groups \[(0.197 ± 0.098)×10(5) vs (0.493 ± 0.067)×10(5) and (0.469 ± 0.083)×10(5)\]. The difference was significant (P0.01).IGFBP7 gene plays a contributing role in leukemogenesis involving in leukemic cells' proliferation and interaction with endothelial cells through adhesion, invasion and migration.

The aim of this study was to construct venus vector carrying the gene encoding insulin-like growth factor binding protein 7 (IGFBP7), which provides an effective platform for exploring the function of this gene in leukemia. After digestion by restriction endonuclease, the IGFBP7 gene was recombined with the transfer plasmid. The venus particles were packaged using 293T cells to transfect K562 cells, and identification was performed by means of flow cytometry, RT-PCR and Western blot. The results showed that the sequence of cloned IGFBP7 gene was the same as that in GenBank. The size of product restricted by BamHI was same as the predicted one. GFP expression was observed in 293T and K562 cells with the fluorescent microscopy and flow cytometry. The expression level of mRNA and protein of IGFBP7 was confirmed by RT-PCR and Western blotting in K562 cells. It is concluded that venus vector carrying IGFBP7 gene has been successfully constructed and provides basis for exploring function of IGFBP7 in K562 cells.

Background Treatment failure for breast cancer is frequently due to lymph node metastasis and invasion to neighboring organs. The aim of the present study was to investigate invasion- and metastasis-related genes in breast cancer cells in vitro and in vivo. Identification of new targets will facilitate the developmental pace of new techniques in screening and early diagnosis. Improved abilities to predict progression and metastasis, therapeutic response and toxicity will help to increase survival of breast cancer patients. Methods Differential protein expression in two breast cancer cell lines, one with high and the other with low metastatic potential, was analyzed using two-dimensional liquid phase chromatographic fractionation (Proteome Lab PF 2D system) followed by matrix-assisted laser desorption/time-of-flight mass spectrometry (MALDI-TOF/MS). Results Up regulation of α-subunit of ATP synthase was identified in high metastatic cells compared with low metastatic cells. Immunohistochemical analysis of 168 human breast cancer specimens on tissue microarrays revealed a high frequency of ATP synthase α-subunit expression in breast cancer (94.6%) compared to normal (21.2%) and atypical hyperplasia (23%) breast tissues. Levels of ATP synthase expression levels strongly correlated with large tumor size, poor tumor differentiation and advanced tumor stages (P < 0.05). ATP synthase α-subunit over-expression was detected on the surface of a highly invasive breast cancer cell line. An antibody against the ATP synthase α-subunit inhibited proliferation, migration and invasion in these breast cancer cells but not that of a non-tumor derived breast cell line. Conclusions Over-expression of ATP synthase α-subunit may be involved in the progression and metastasis of breast cancer, perhaps representing a potential biomarker for diagnosis, prognosis and a therapeutic target for breast cancer. This finding of this study will help us to better understand the molecular mechanism of tumor metastasis and to improve the screening, diagnosis, as well as prognosis and/or prediction of responses to therapy for breast cancer.

Background Treatment failure for esophageal carcinoma is frequently due to lymph node metastasis and invasion to neighboring organs. The aim of the present study was to investigate invasion- and metastasis-related genes in esophageal carcinoma cells in vitro and in vivo. Methods A metastasis model using a Matrigel invasion clonal selection approach was employed to establish a highly invasive subline EC9706-P4 from the esophageal carcinoma cell (ESCC) line EC9706. The differentially expressed genes of the subline and the parental cells determined by gene microarrays were further analyzed by RT-PCR and Western blotting. Results We identified sphingosine kinase 1 (SPHK1) as an invasion and metastasis-related gene of esophageal cancer. SPHK1 was overexpressed in the EC9706-P4 subline with high invasive capacity. Among six ESCC lines tested, KYSE2 and KYSE30 cells showed the highest SPHK1 mRNA and protein expressions as well as the most invasive phenotype. By Western blotting, in 7/12 cases (58%), SPHK1 expression was higher in esophageal carcinomas than in the companion normal tissue. In 23/30 cases (76%), SPHK1 protein expression was upregulated in the tumors compared to matched normal tissue by immunohistochemistry (IHC). Esophageal carcinoma tissue microarray analysis indicated that SPHK1 expression correlated with the depth of tumor invasion (P < 0.0001) and lymph node metastasis (P = 0.016). By Kaplan-Meier analysis, strong SPHK1 expression was significantly associated with clinical failure (P < 0.01), suggesting the involvement of SPHK1 in aggressiveness of human esophageal carcinoma. SPHK1 overexpression significantly increased the invasiveness of EC9706 cells in vitro and also increased EC9706 cell growth and spontaneous metastasis in vivo, promoting significant increases in tumor growth, tumor burden and spontaneous lung metastasis in nude mice. SPHK1 expression significantly correlated with the expression of many EGFR pathway genes associated with invasion of cancer cells. SPHK1 protein expression also significantly correlated with the phosphorylation of EGFR. Conclusion In summary, our data implicate SPHK1 in the metastasis of esophageal cancer. Our study also identified downstream mediators of SPHK1 in esophageal cancer cells that may mediate enhanced malignant behavior, and several of these mediators may be useful as therapeutic targets.

To investigate the effects of targeting elimination of the leukemic CD34+ progenitor cells by using cytotoxic T lymphocytes (CTLs) specifically against Wilms tumor gene (WT1)-derived peptide.A 9-mer WT1 peptide (CMTWNQMNL) containing HLA-A * 0201-binding anchor motifs was synthesized. The suspended cells were collected from the culture of the peripheral blood mononuclear cells and divided into 2 groups: Group D (pure T cells) and Group C (IL2 + T cells). The dendritic cells (DCs) generated from the peripheral blood mononuclear cells of an HLA-A* 0201-positive healthy donor were repeatedly loaded with WT1 peptide so as to elicit cytotoxic T cells (CTLs) specifically for WT1 peptide, and restricted by HLA-A * 0201 (Group A). The specific lysis effects of WT1 peptide specific CTLs upon leukemic bone marrow CD34+ progenitor cells positively expressing WT1 (3 being HLA-A * 0201(+) and 3 being HLA-A*0201(-)), peripheral blood CD34+ cells from healthy persons (2 being HLA-A * 0201(+) and 1 being HLA-A * 0201(-)), and leukemia cells of the lines of NB4 (HLA-A * 0201(+)/WT1(+)), U937 (HLA-A * 0201(+)/WT1(-)), and K562 (HLA-A * 0201(-)/WT1(+)) were measured by methyl thiazolyl tetrazolium (MTT) chromatometry assay. The colony-forming unit-granulocyte macrophage (CFU-GM) forming capability of the leukemic bone marrow CD34+ progenitor cells and peripheral blood CD34+ cells from healthy persons treated with WT1 peptide specific CTLs were also determined by methylcellulose medium colony-forming unit assay. The CTLs elicited by DCs without WT1 peptide loading(Group B)and T cells cultured with IL-2 (Group C) were taken as controls.The CTLs specific for WT1 peptide and restricted by HLA-A * 0201 (Group A) exerted a specific lysis upon the 3 cases with HLA-A * 0201(+)/WT1(+) leukemic bone marrow CD34+ progenitor cells and NB4 leukemia cells, the specific lysis levels were 55.3% +/- 2.8%, 67.1% +/- 3.2%, 49.4% +/- 3.8% and 55.0% +/- 3.7% respectively, all significantly higher than those upon the 3 cases with HLA-A * 0201(-)/WT1(+) leukemic bone marrow CD34+ progenitor cells, normal healthy peripheral blood CD34+ cells of the healthy persons, and leukemia cell of the lines K562 and U937 (the specific lysis levels were all20%). The specific lysis level of Group A CTLs was significantly higher than those of Group B and Group C CTLs (both P0.01). The relative colony formation rates of WT1-CTLs in Group A upon the 2 cases with HLA-A * 0201(+)/WT1(+) leukemic CD34+ progenitor cells were 17.8% +/- 4.0% and 20.8% +/- 3.41% respectively, both significantly lower than those of the none WT1-CTLs in Group B (88.9% +/- 3.4% and 91.8% +/- 5.7% respectively, both P0.01). There was no significant difference in the relative colony formation rate upon HLA-A * 0201(+)/WT1(-) leukemic bone marrow CD34+ progenitor cells and the HLA-A * 0201(-)/WT1(-) or HLA-A * 0201(+)/WT1(-) normal peripheral blood CD34+ progenitor cells between the CTLs of Groups A and B.The CTLs specific for WT1 and restricted by HLA-A * 0201 can exert specific lysis upon leukemic CD34+ progenitor cells highly expressing WT1 gene, and can inhibit the CFU-GM colony formation of such leukemic CD34+ progenitor cells. The product of expression of WT1 gene may be used as a target in the leukemic CD34+ cells.

The objective of study was to investigate tissue-specific transcriptional activity of WT1 (Wilms' tumor gene) promoter and enhancer in cell lines with diverse tissue origin for leukemic gene therapy depending on WT1 transcriptional regulation elements. WT1 promoter and enhancer were ligated into pEGFP-1 to construct a recombinant vectors with EGFP gene as a reporter. By using electroporation or lipofectamine, the resultant constructs were transfected into 13 cell lines including WT1-expressing hematopoietic cell lines (K562, NB4, THP-1 and SHI-1), WT1-nonexpressing hematopoietic cell lines (U937 and Jurkat), WT1-expressing nonhematopoietic cell lines (MCF-7, T47D and 293) and WT1-nonexpressing nonhematopoietic cell lines (ECV304, SMMC7721, HT-29 and SHG44). The mean fluorescence intensity (MFI) of EGFP representing the transcriptional activities of promoter and/or enhancer was analyzed by using flow cytometry in the transfected cells which stably expressed EGFP. The results indicated that the vectors, pEWP containing WT1 promoter and pEWPA containing both WT1 enhancer and promoter, were constructed by recombinant DNA technique. Among nonhematopoietic cell lines, pEWP induced the highest EGFP expression in ECV304 (16.54 +/- 2.45 times as high as pEGFP-1), mildly higher in MCF-7 and SHG44 (9.46 +/- 1.10 and 7.29 +/- 0.73 times of pEGFP-1 level), and lowest in HT-29 (0.99 +/- 0.02 times as much as pEGFP-1) respectively. Among hematopoietic cell lines, EGFP expression was highest in K562 cell line (2.93 +/- 0.27 times of pEGFP-1), which was statistically higher than those in Jurkat and SHI-1 cell lines (0.74 +/- 0.03 and 0.84 +/- 0.09 times of pEGFP-1 level) respectively. pEWPA, with WT1 enhancer inserted at Afl II site near SV40 polyA, increased basal transcription levels of the WT1 promoter in HT-29, SHI-1 and K562 cells by 4.81, 3.06 and 1.01-fold respectively. It is concluded that the transcriptional activities of WT1 promoter in the recombinant vector seem unrelated to the constitutional expression level of endogenous WT1 gene. The WT1 enhancer promotes the transcriptional activities of WT1 promoter in some of the cell lines regardless of the hematopoietic tissue origin.

To investigate the clinical significance of Wilms tumor gene (WT1) expression levels in the bone marrow (BM) of chronic myelogenous leukemia patients.Real-time quantitative retroversion polymerase chain reaction (RQ-RT-PCR) method was established for detecting WT1 and GAPDH expression levels in the BM of 109 CML and 23 non-leukemic patients with Light Cycler. Normalized WT1 expression level (WT1(N)) was determined as a ratio between WT1 and GAPDH expression times 10(4).The median expression levels of WT1(N) in 23 CML patients of accelerate phase (CML-AP) and 22 CML patients of blast crisis (CML-BC) were statistically higher than those of 64 CML patients of chronic phase (CML-CP) and 23 non-leukemic controls (103.71 and 129.44 vs 3.44 and 1.47). No statistic differences were found between the CML-CP group and control group, nor between the CML-AP group and CML-BC group. Furthermore, the WT1(N) levels were correlated to the BCR/ABL fusion gene expression levels. Dynamic change of WT1(N) levels in 7 CML patients before or after allogeneic bone marrow transplantation showed that WT1 expression levels could predict relapse of leukemia.WT1 expression levels in CML patients in accelerate phase or blast crisis were strikingly higher than those in non-leukemic patients or CML patients in chronic phase, in whom WT1 expression was undetectable or showed low expression. WT1 gene could be an useful marker for detecting MRD and evaluating therapeutic efficacy in CML.

In order to investigate the feasibility of using EGFP as a reporter gene in WT1 transcriptional regulation study. WT1 promoter and enhancer were ligated into the vector pEGFP-1 by recombinant DNA technique and confirmed by restriction enzymes digestion. The resultant constructs were transfected into K562 cell line by DMRIE-C reagent and the function of these WT1 gene elements was detected by using a fluorescent microscope after transfection for 48 hours. The results indicated that the recombinant vectors, pEWP containing WT1 promoter, and pEWPE, pEWPA and pEWPD harboring both WT1 enhancer and promoter, had been successfully constructed. Fluorescence was observed in K562 cells transfected by pEWP, pEWPE, pEWPA and pEWPD, while no fluorescence could be detected in cells transfected by pEGFP-1. It is concluded that EGFP gene as a reporter gene can be applied to the WT1 transcriptional regulation study, which provides the basis for gene therapy.

To explore the mechanism of action of RbAp46 gene on leukemic cells.K562 leukemic cells and SHG44 glioma cells were transfected with eukaryotic expression vector carrying full-length cDNA of RbAp46 driven by the cytomegalovirus promoter mediated by lipofectamine transfection reagent. Empty vector were transfected at the same time as control. G418-resistant colonies were selected after 3 weeks culturing. Series genes were amplified using RT-PCR. Growth curve and colony formation assays were performed.The number of K562/RbAp46 and K562/CMV cells were (90.00 +/- 8.40) x 10(4) and (119.58 +/- 9.87) x 10(4), respectively after 4 days growth, and SHG44/RbAp46 and SHG44/CMV cells were (89.13 +/- 4.88) x 10(4) and (149.42 +/- 10.83) x 10(4), respectively after 5 days growth. Seven-day yields of K562/RbAp46 and K562/CMV cell colonies were 131.67 +/- 15.57 and 250.33 +/- 26.31, respectively (P0.01), while those of SHG44/RbAp46 and SHG44/CMV cells were 50.78 +/- 6.77 and 206.67 +/- 37.18, respectively (P0.01). The fraction of K562/RbAp46 and K562/CMV cells in S phase was (48.88 +/- 4.35)% and (62.78 +/- 4.78)% (P0.01), and in G(0)/G(1) phase was (29.10 +/- 4.14)% and (22.40 +/- 2.43)%, respectively (P0.05), and that of SHG44/RbAp46 and SHG44/CMV cells in G(0)/G(1) phase was 65.6% and 48.8%, and in S phase was 22.6% and 38.4%, respectively. Insulin-like growth factor binding protein-related protein-1 (IGFBP-rP1) gene was induced to express only in the RbAp46-over expressing K562 cells and was not in SHG44 cells.A regulatory pathway between RbAp46 and IGFBP-rP1 genes might exit in K562 leukemic cells.