Follicular lymphoma accounts for 22.1% of non-Hodgkin lymphoma. It is an indolent lymphoma. The clinical feature is lymphadenopathy with/without marrow involvement. The prognostic factors include follicular lymphoma international prognosis index (FLIPI) and tumor infiltrating immune cells. The fast-line therapy is immunochemotherapy with rituximab. In patients with refractory/refapes disease, stem cell transplantation is the choice.

Unlike children, the outcome of adult acute lymphoblastic leukemia (ALL) has not changed in past decades. There are many risk factors influencing the outcome. The most important factor is the early response to chemotherapy. The outcome of adult ALL is improved when receiving pediatric ALL regimen.Targeting therapy combined with chemotherapy have improved the outcome of Ph+ ALL and ALL expressing CD20+ Allogeneic stem cell transplantation is beneficial to high risk ALL.

DNA and histone lysine methylations are the main epigenetic modulatory elements in the development and progression of acute myeloid leukemia. Methylation-targeted therapy includes DNA methyltransferase, methylation modulatory protein, and histone-lysine methylation inhibitors. Approved DNA methyltransferase inhibitor (demethylating agent) includes azacytidine and dicitabine, which have been used as antileukemic drug clinically. IDH1/2 inhibitor also showed effective and well tolerated in leukemic patients in phaseⅡclinical study. EZH2 inhibitor and LSD1 inhibitor have completed in vitro study and entered clinical trial. Targeting DNA and histone lysine methylations is an alternative approach for leukemia treatment.

BACKGROUND: Cord blood stem cells are one of ideal target cells for gene therapy, but low gene transferring rate is the main difficulty at recent. Janus kinase tyrosine 2 (JAK2) plays an important role in self-renewing of cord blood stem/progenitor cell12s. Therefore, cord blood CD34+ cell line modified by target-amplified JAK2 genes has been developed yet by using gene regulating expression technique in order to overcome low transferring rate of cord blood genes.OBJECTIVE: To investigate the feasibility and reliability of a long-term amplified regulation for cord blood stem/progenitor cells mediated by transgene JAK2. SETTING: Department of Hematology, Beijing Hospital, Ministry of Health.MATERIALS: The experiment was carried out in the Laboratory of Hematological Department, Beijing Hospital, Ministry of Health from June 2003 to April 2006. Cord blood was derived from umbilical cord which was immediately cut from healthy, full-term and natural-parturition infants and was provided by Department of Obstetrics & Gynecology, Beijing Hospital. The experiment was approved by the local ethical committee, and informed consent was obtained from expectant mothers and their relatives for the use of cord blood cells. MiniMACS magnetic separation apparatus and immunomagnetic beads adsorbing CD34 single antibody were provided by Miltenyi Biotec Company, Germany; flow cytometer by FACScalibur, USA; recombinant human stem cell factor (rhSCF), Flt3 ligand (FL), human interleukin-6 (hIL-6), granulocyte macrophage colony stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF) and thrombopoeitin (TPO) by PeproTec Company; nude mice of the SPF level by Animal Center of Beijing Medical University.METHODS: Retroviral vector MGI-F2JAK2, which was composed of functional catalytic domain of JAK2 genes and two site proteins (2xF36v, F2) combined with synthetic drug (AP20187) of target gene of small molecules, was constructed. AP20187 might specially combine with F36v to cause dimerization of JAK2 so as to activate signal conduction in cells. In addition, the vector included green fluorescence protein reporter gene, which was regarded as a label to detect proliferation. MiniMACS magnetic separation apparatus was used to purify and separate cord blood CD34+ cells. While, retrovirus supernatant including JAK2 was used to transfer cord blood CD34+ cells. After transduction, CD34+ cells were cultured with stem cell factor (SCF), Flt3 ligand, TPO and IL-6 and divided into control group (not adding AP20187) and experimental group (AP20187).MAIN OUTCOME MEASURES: ① Flow cytometer was used to detect percentage of green fluorescence protein reporter gene in the CD34+ cells and to determine gene transferring rate. ② Colony culture results of cord blood stem/progenitor cells after amplification. ③ Nude mice were given subcutaneous injection of ten-week cultured cord blood CD34+ cells at costa and neoplasia was observed after 30 days. RESULTS: ① Plentiful amplification of CD34+ cells was observed in both experimental group and control group. With the culture time passing by, positive rate of gel-filtered platelet of amplified CD34+ cells in the experimental group was gradually increased based on the basic level and more than 95% in the 11th week; however, positive rate of green fluorescence protein reporter gene in the control group was gradually decreased below the basic level and disappeared finally. ② Transgenic CD34+ cells in the experimental group still could generate brust forming unit-erythroid (BFU-E), colony-forming units granulocute/monocyte (CFU-GM) and multipotential hematopoietic progenitors (CFU-Mix); especially, CFU-GM was the main cell in hemopoietic progenitor cell (HPC). ③ Nude mice did not have neoplasia. CONCLUSION: Human cord blood CD34+ cells of transferring JAK2 genes may cooperate with other cytokines to amplify cord blood stem/progenitor cells in vitro for long. Therefore, this is potentially valuable for stem cells to treat some hereditary hematologic disease.

ObjectiveTo explore the cytogenetic characteristics of multiple myeloma (MM) patients,to evaluate the effect of a long-term culture stimulated by cytokines on cytogenetic study of MM,and to investigate the clinical detection value of RB1 and P53 deletion in interphase plasma cells by using fluorescence in situ hybridization (FISH).MethodsKaryotype analysis was performed in 81 MM patients by using the short-term culture of bone marrow cell and G-banding technique.Among the 81 MM patients,28 patients used two culture methods:one was the short-term culture and the other was to culture cells for 6 days with recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF) (40 μg/L) and IL-6 (10 μg/L).RB1 and P53 deletion were detected on interphase plasma cells by using FISH in 31 patients.ResultsAmong the 81 patients,75 had enough metaphases for analysis.Among the 75 patients,31 (41.3％)had clonal karyotypic abnormalities including 4 numeric abnormalities,11structural abnormalities and 16 both abnormalities.Among the 28 patients using two culture methods,the clonal karyotypic abnormalities were detected in 6 patients(25.0％ ) in the group of cultured for 24 hours,and 14 patients (51.9％) in 6-day culture group with a significant difference (P =0.026).RB1 deletion and P53 deletion were detected in 10 patients (32.3％ ) and 11 patients(35.5％ ),respectively,with both RB1 and P53 deletions be detected in 5 patients ( 16.1％ ).ConclusionsMore than half of the tested MM patients have both numeric and structural chromosome abnormalities.The karyotype analysis using banding technique is basic cytogenetic study.Extended culture in the presence of IL-6 and GM-CSF could improve the efficiency of cytogenetic analysis to MM.Interphase FISH is a sensitive method of clinical application significance to detect the gene deletion of MM.

Objective To investigate the clinical efficacy of different treatment in 35 chronic lymphocytic leukemia (CLL) patients. Methods Patients were treated with different regimen according to Binet stage. Patients at stage A were subcutaneously injected with interferon (3-6) MU/day, consecutive for 5 days every week. The dosage could be reduced to 2-3 times a week in long term maintenance phase after the 6 months loading treatment if there was no disease progression. Those at stage B or C were initially treated with chemotherapy regimen of FC/FC -R or CHOP/COP, and interferon were administered during chemotherapy interval, after complete remission (CR) or partially remission (PR) as maintenance therapy.Results Twenty patients were at stage A and treated with interferon, with 5 patients(25%) achieving partial remission (PR), 14 patients at stable status while no patients acquiring CR. Three of the 5 patients who achieved PR collapsed after 36.3 months at average. Eight of the 14 patients at stable status deteriorated to stage B and received chemotherapy after mean 74 months interferon maintenance treatment. In total, 27 patients in the current observation were finally included at stage B or C. Patients at stage B or C in FC/FC-R chemotherapy regimen achieved CR at 38.9% and total effective rate 77.8%, which were superior to that of CHOP/COP prescription (CR 11.1 %). The mean survival time for patients at stage A, B and C were 155.2,97.5 and 82.9 months, respectively and were statistically significant via Kaplan-Meier analysis method (P =0.032). Ten patients died in this observation, 2 at stage A, 4 at stage B and C, respectively, among whom 9died of infection and 1 for gastric cancer bleeding. The side effects of interferon were generally mild during the long term treatment. Conclusion Patients with CLL need to be individually treated with different regimen by considering disease stage and other prognosis criteria. Interferon could be applied at early phase of CLL and may reduce occurrence of infection after long term treatment.

Objective To evaluate the characteristics of immunophenotyping in adults with acute myeloid leukemia (AML) using muhi-eolor flow cytometry. Methods Immunophenotyping was performed by three color flow cytometry using CD45/SSC gating. Results In 126 patients with AML, the myeloid antigen of CD13, CD33 and CD117 was highly expressed. The positive rate was 86.4 %, 70.2 % and 90.4 %, respectively. The CD34 and HLA-DR were lowerly expressed as 63.5 % and 61.7 %, respectively. About 34.2 % of lymphoid-assoeiated antigen expression of all the AML patients. The lymphoid-associated antigens of CD7 and CD19 expression in patients with AML was 23.6 % and 2.3 %, respectively. Conclusion Multi-color flow cytometry is an important method for diagnosis and prognosis for AML.

ObjectiveTo evaluate the application of 18fluoro-deoxyglucose positron emission tomography (FDG-PET) to the staging and predicting outcome in patients with lymphoma.Methods 41 patients with newly diagnosed lymphoma(median age 57 years)were explored with FDG-PET prior to and after 4 cycles of chemotherapy.With a median follow-up of 30 months (range 10-68 months),the value of FDG-PET to staging and predicting clinical outcome was assessed. Results The maximum standardized uptake value (SUVmax) of nodal and extranodal lesions was 9.7±6.9 and 8.4±6.8 respectively prior to treatment.There were significant difference (P＜0.05) in aggressive non-Hodgkin's lymphoma and indolent non-Hodgkin's lymphoma,no significant difference(P＞0.05)in non-Hodgkin's lymphoma and Hodgkin's lymphoma(HL), B-cell neoplasms and T-cell neoplasms,germinal center B-cell-like DLBCL and activated B-cell-like DLBCL. In 41 patients, 22 patients (54 ％)were detected extranodal focus by FDG-PET before chemotherapy. FDG-PET imaging upstaged in 6(15％)of initial lymphoma patients.There were 15 patients (37 ％) in stage Ⅰ and Ⅱ and 26 patients(63 ％)in stage Ⅲ and Ⅳ by FDC-PET scan.1 patient (7 ％) in stage Ⅰ and Ⅱ,6 patient (23 ％) in stage Ⅲ and Ⅳ died of disease progression during follow-up.After 4 cycles of chemotherapy,the FDG-PET was negative in 41％(17/41),positive in 59 ％(24/41) respectively.1 patient(6 ％)died of disease relapse among 17 patients who were FDG-PET negative, 6 patient (25 ％)died of disease progression among 24 patients who were FDG-PET positive during follow-up. Conclusion FDG-PET scanning plays an important role in the pretreatment staging and prediction of the prognosis after 4 cycles of chemotherapy in patients with lymphoma.Thus it may offer the potential for change in treatment paradigms.

AIM:To explore the feasibility and biological characterization of long-term regulated expansion of JAK2 transduced human CD34+ cord blood cells in vitro.METHODS: A retrovirus (RV) vector which contains JAK2 catalytic domain and two binding sites for a chemical inducer, dimerization (AP20187), was cloned (designated MGI-F2JAK2). CD34+cells were enriched from cord blood with a MiniMACS system. The purified CD34+cells were transfected with supernatant from the retrovirus packaging cell line that expressed JAK2. Following transduction, cells were expanded into four groups: AP20187 alone, FL alone, TPO, alone, AP20187+FL+TPO, respectively. The expanded cells were monitored by GFP expression, immunophenotyping, progenitor colony assay, karyotype analysis as well as tumorigenesis in nude mice. RESULTS: The purity of selected CD34+ cells was over 91% and gene transfer rate was 49.32%?6.21%. Only the group of AP20187 +FL+ TPO was obtained a significant sustained outgrowth of the transduced CD34+ cord blood cells. The percentage of GFP+ cells consistently produced a rise to the 90% peak level by the end of 8th week of culture. Flow cytometry analysis showed that the phenotype of the expanded cells was CD33+, CD61+ and Gly-A+ partial positive; CD38+ and HLA-DR+ strong positive, while CD2, CD7 and CD19 were almost negative. Colony assays performed in methycelluos, which can give rise to BFU-E, CFU-GM and CFU-Mix, the CFU-GM was predominantly in all colonies. The tumor was not observed in nude mice and the karyotype analysis was normal from expanded cells.CONCLUSION: The results demonstrate that AP20187-mediated activation of JAK2 signaling is capable of stimulating expansion JAK2 transduced CB CD34+ cells in combination with FL and TPO. This system may have applications for studies in signaling transduction, hematopoiesis, and for gene and cell therapy.

Objective To explore the cytogenetic characteristics of acute myeloid leukemia(AML) patients.Methods The karyotype analysis was performed in 178 AML using the short-term culture of bone marrow cell and G-banding technique.Results Among the 178 patients,171 had enough metaphases for analysis and 128(74.9%)had clonal karyotypic abnormalities.Twenty-seven patients were secondary to myelodysplastic syndrome (MDS-AML),with 25 (92.6%) patients carrying clonal karyotypic abnormalities.Among the remaining 144 patients of de novo AML,103(71.5%)had clonal karyotypic abnormalities.The rate of abnormal clonal karyotype was higher in MDS-AML than that of de novo AML (P=0.021).Among the 171 patients,41(24.0%)were in favorable risk group,80(46.8%)in intermediate risk group and 50(29.2%)in adverse risk group.t(15;17)was the most common chromosomal aberration.The maiority intermediate risk chromosomal aberration was;normal karyotype.The most common cytogenetic abnormality among adverse group was a complex karyotype.Adverse cytogenetic aberrations,such as -5/5q-,-7/7q-,frequently occurred in conjunction with one another as part of a complex karyotype.Totally 75 patients were 60 years or older,among them,16.0%were in favorable risk group,48.0%in intermediate risk group and 36.0%in adverse risk group.Among 96 younger patients,30.2%were in favorable risk group.45.8%in intermediate risk group and 24.0%in adverse risk group.The rate of favorable risk chromosomal aberration was lower in elder patients than in younger(P=0.03 1).The rate of adverse risk chromosomal aberration and the rate of monosomal karyotype were higher in MDSAML than in de novo AML patients(P<0.001).Conclusions The most common favorable,intermediate and adverse chromosomal aberrations were t(15;17),normal karyotype and complex karyotype respectively.The karyotype was poor in MDS-AML and elder AML patients.