Allogeneic hematopoietic stem cell transplantation in acute leukemia patients with the SET-NUP214 fusion gene: Efficacy and survival analysis / 中华内科杂志

Objective: To investigate the clinical efficacy of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for patients with acute leukemia who are positive for the SET-NUP214 fusion gene (SET-NUP214+AL). Methods: This was a retrospective case series study. Clinical data of 18 patients with SET-NUP214+AL who received allo-HSCT in the First Affiliated Hospital of Soochow University and Soochow Hongci Hematology Hospital from December 2014 to October 2021 were retrospectively analyzed to investigate treatment efficacy and prognosis. The Kaplan-Meier method was used for survival analysis. Results: Of the 18 patients, 12 were male and 6 were female, and the median age was 29 years (range, 13-55 years). There were six cases of mixed phenotype acute leukemia (three cases of myeloid/T, two cases of B/T, one case of myeloid/B/T), nine cases of acute lymphoblastic leukemia (ALL) (one case of B-ALL and eight cases of T-ALL), and three cases of acute myeloid leukemia. All patients received induction chemotherapy after diagnosis, and 17 patients achieved complete remission (CR) after chemotherapy. All patients subsequently received allo-HSCT. Pre-transplantation status: 15 patients were in the first CR, 1 patient was in the second CR, 1 was in partial remission, and 1 patient did not reach CR. All patients were successfully implanted with stem cells. The median time of granulocyte and platelet reconstitution was +12 and +13 days, respectively. With a median follow-up of 23 (4-80) months, 15 patients survived, while 3 patients died. The cause of death was recurrence of SET-NUP214+AL after transplantation. After allo-HSCT, 5 patients relapsed. The estimated 3-year overall survival (OS) and relapse-free survival (RFS) rates were 83.3%±15.2% and 55.4%±20.7%, respectively. Among the 15 patients who achieved CR before transplantation, there was no significant difference in OS and RFS between haploidentical HSCT and matched sibling donor HSCT (all P>0.05). Conclusions: Allo-HSCT can improve the prognosis and long-term survival rate of patients with SET-NUP214+AL. Disease recurrence is the most important factor affecting long-term survival.

Preliminary exploration of the metabolic profile and metabolic pathways in newly diagnosed multiple myeloma / 中华肿瘤杂志

Objective: To explore the metabolite profile and metabolic pathways of newly diagnosed multiple myeloma (MM). Methods: Gas chromatography-mass spectrometry (GC-MS) was employed for the high-throughput detection and identification of serum samples from 55 patients with MM and 37 healthy controls matched for age and sex from 2016 to 2017 collected at the First Affiliated Hospital of Soochow University. The relative standard deviation (RSD) of quality control (QC) samples was employed to validate the reproducibility of GC-MS approach. The differential metabolites between patients with MM and healthy controls were detected by partial least squares discrimination analysis (PLS-DA), and t-test with false discovery rate (FDR) correction. Metabolomics pathway analysis (MetPA) was employed to construct metabolic pathways. Results: There were 55 MM patients, including 34 males and 21 females. The median age was 60 years old (42-73 years old). There were 30 cases of IgG type, 9 cases of IgA type, 1 case of IgM type, 2 cases of non-secreted type, 1 case of double clone type and 12 cases of light chain type, including 3 cases of kappa light chain type and 9 cases of lambda light chain type. The result of QC sample test showed that the proportion of compounds with the RSD of the relative content of metabolites < 15% was 70.21% obtained by the reproducibility of GC-MS experimental data, which implied that the experimental data were reliable. A total of 17 metabolites were screened differently with the healthy control group, including myristic acid, hydroxyproline, cysteine, palmitic acid, L-leucine, stearic acid, methionine, phenylalanine, glycerin, serine, isoleucine, tyrosine, valine, citric acid, inositol, threonine, and oxalic acid (VIP>1, P<0.05). Metabolic pathway analysis suggested that metabolic disorders in MM patients comprised mainly phenylalanine metabolism, glyoxylic acid and dicarboxylic acid metabolism, phosphoinositide metabolism, cysteine and methionine metabolism, glycerolipid metabolism, glycine, serine, and threonine metabolism. Conclusion: Compared with normal people, patients with newly diagnosed MM have obvious differences in metabolic profiles and metabolic pathways.

Effect of decitabine on immune regulation in patients with acute myeloid leukemia after allogeneic hematopoietic stem cell transplantation / 中国实验血液学杂志

Based on the representative articles in recent years, the different mechanisms of decitabine on immune regulation in patients with acute myeloid leukemia (AML) after allogeneic hematopoietic stem cell transplantation (HSCT) are summarized. Decitabine improves the expression of WT1 gene to stimulate specific cytotoxic T cells which can enhance graft versus leukemia effect (GVL) and improve the expression of FOXP3 gene to stimulate regulatory T cells so as to inhibit the acute graft versus host disease (GVHD). Through the above-mentimed mechanisms, decitabine can improve both therapeutic effect and quality of life in the patients with AML after allogeneic HSCT.

The correlation of cytomegalovirus gB genotype with viral DNA load and treatment time in patients with CMV infection after hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the effect of CMV gB genotypes on viral load and treatment time in patients with CMV infection after hematopoietic stem cell transplantation (HSCT).</p><p><b>METHODS</b>Viral load was detected by real-time (RT) quantitative polymerase chain reaction (PCR) (Q-PCR), CMV gB genotypes by PCR restriction fragment length polymorphism (RFLP) (PCR-RFLP) in 115 patients with CMV infection (CMV-DNA positive) after HSCT during July 2004 and May 2010.</p><p><b>RESULTS</b>(1) The distribution of CMV gB genotypes in HSCT recipients were as following: gB1, 42/115 (36.52%); gB2, 3/115 (2.61%); gB3, 43/115 (37.39%); gB4, 2/115 (1.74%). 20 patients (17.39%) had a combination of 2 different CMV genotypes and 5 patients (4.35%) had a CMV variant that lacked an RsaI digestion site, herein named gB5. (2) The median viral load were 2.7×10(3)(1.81×10(3) ∼ 6.03×10(4)) in gB1, 4.0×10(3) (1.32×10(3) ∼ 6.39×10(4)) in gB3 and 1.2×10(4)(2.28×10(3) ∼ 6.50×10(5)) in mixed gB. There was no statistical difference in viral load between gB1 and gB3 (P > 0.050). There was significantly statistical difference in viral load between single-gB (gB1 or gB3) and mixed-gB (P < 0.05). (3) The median treatment time was 17 days in mixed-gB and 14 days in single-gB. There was significantly statistical difference between two groups (P < 0.05). Conclusion gB genotype may have an impact on CMV DNA load and treatment time in HSCT recipients with CMV infection.</p>

Efficacy analysis of unrelated donor hematopoietic stem cell transplantation for treatment of high risk acute myeloid leukemia / 中国实验血液学杂志

This study was purposed to investigate the therapeutic efficacy of unrelated donor hematopoietic stem cell transplantation (URD-HSCT) for patients with high risk and refractory acute myeloid leukemia (AML). Twenty-two patients with high-risk and refractory AML receive URD-HSCT were enrolled in this study. All the patients received myeloablative preconditioning regimen consisting of busulfan/cyclophosphamide (for 20 cases) or total body irradiation/cyclophosphamide (for 2 cases) before URD-HSCT. The cyclosporin A (CsA)/MTX/MMF/ATG were used to prevent the acute graft versus host disease (aGVHD). The results showed that 21 out of 22 patients acquired engraftment with implantation rate 95.5%. The median time of ANC ≥ 0.5×10(9)/L was 12 (10-19) days, and that of Plt ≥ 20×10(9)/L was 14 (5 - 22) days. The median follow-up time post transplantation was 18 (3 to 135.5) months. The 2-year overall survival (OS) and leukemia-free survival (LFS) were (53.9 ± 12.2) % and (49.1 ± 10.7)% respectively. Eight cases developed aGVHD. The cumulative incidence of aGVHD was (39.1 ± 10.6) %. Six patients developed I-II grade of aGVHD and two patients developed III-IV grade of aGVHD. The chronic graft versus host disease (cGVHD) was occurred in 6 patients (4 patients limited, 2 patients extensive) of the 19 evaluable patients. The cumulative incidence was (28.8 ± 9.6)%. Seven cases relapsed, and the cumulative response rate of 2 years was (35.8 ± 11) %. One of 9 patients died from sepsis before hematopoietic reconstruction, one died from lung infection, Six died from relapse and one relapsed patient died from IV grade of aGVHD post chemotherapy and donor lymphocyte infusion (DLI). The univariate analysis revealed that relapse was the major factor for the OS, and the sex, age, preconditioning regimen, aGVHD and infection didn't significantly influence the efficacy of URD-HSCT. The survival of patients with cGVHD was superior to those who didn't have cGVHD (83.3% vs 37%, P = 0.152). It is concluded that URD-HSCT is a safe and effective therapy for high-risk AML patients without related donor. Notably, patients with cGVHD had a better survival. Relapse is an unfavourable factor for the efficacy of URD-HSCT and adoptive immunotherapy such as DLI can prevent it and improve the prognosis to achieve the long-time survival.

Impact of disease status on outcomes of allogeneic hematopoietic stem cell transplantation in patients with refractory and relapsed acute myeloid leukemia / 中国实验血液学杂志

The study was aimed to evaluate the impact of disease status on the outcomes of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in patients with refractory and relapsed acute myeloid leukemia (AML). 32 patients with refractory and relapsed AML received allo-HSCT after myeloablative conditioning regimen, including 17 patients in no-remission (NR) and 15 patients in complete remission (CR) at the time of transplant. Treatment related adverse events, relapse rate and leukemia free survival (LFS) were analyzed. The results showed that the parameters of sex, age, cytogenetic risk and transplant procedures were comparable between the two groups. 30 patients had successful engraftment, except one had graft failure and one died from severe veno-occlusive disease in the NR group. The incidences of aGVHD in NR group and CR group were 47.1% (8 patients) and 33.5% (5 patients) respectively. Out of comparable patients, 5 from 9 patients in NR group developed with cGVHD, and 4 from 11 patients in CR group were subjected to cGVHD. There were no statistic difference in incidences of aGVHD and cGVHD between two group. Compa-red with CR group, NR group had a higher treatment-related mortality (29.4% vs 14.3%, P = 0.392) and relapse rate (42.9% vs 26.7% P = 0.300), but there was no significant difference. With a median follow-up of 13 (1 - 124) months, 6 patients remained alive in both of the two groups, and the 2 year LFS of them were parallel (35.3% vs 40.0%, P = 0.267). Among these 32 patients, overall survival (OS) was better in patients with age < 35 years (P = 0.044) and with the appearance of cGVHD (P = 0.046). It is concluded that allo-HSCT is an effective salvage therapy for patients with refractory and relapsed AML, and the overall outcome seems unrelated to the disease status (NR or CR) before transplantation. As such, for refractory and relapsed AML patients in non-remission, performance of allo-HSCT to achieve long-term survival is feasible.

Study on the clinical characteristics of 32 patients with mixed phenotype acute leukemia / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the clinical and biological characteristics and prognosis of mixed phenotype acute leukemia (MPAL).</p><p><b>METHODS</b>Thirty two patients were diagnosed as MPAL by bone marrow examination, immunophenotyping, cytogenetic and molecular assay and were treated with combined chemotherapy regimens for both acute lymphoblastic and acute myeloid leukemia. Two cases were received allogeneic hematopoietic stem cell transplantation (allo-HSCT).</p><p><b>RESULTS</b>(1) The incidence of MPAL in acute leukemias was 2.6%. There were 16 cases (50.0%) of mixed myeloid and B-lymphoid (M/B), 14(43.8%) myeloid and T-lymphoid (M/T), one each (3.1%) of trilineage (M/B/T) and B- and T-lymphoid (B/T) phenotype. (2) The positive rates of CD34 and HLA-DR were 87.5% and 62.5%, respectively. (3) Abnormal karyotypes were detected in 70.0% of 30 MPAL patients, which were structural and numerical abnormalities including t(9;22), 11q23 and complex karyotypes. (4) The total complete remission (CR) rate was 75.0% and the overall survival (OS) and disease-free survival (DFS) at 2 years were 14.8% and 14.2% respectively. The CR rates for M/B and M/T cases were 75.0% and 71.4% respectively. No statistical difference was observed in OS and DFS between M/B and M/T cases.</p><p><b>CONCLUSIONS</b>MPAL is a rare type of acute leukemia with a high heterogeneity. The unfavorable indicators of MPAL may be factors such as abnormal karyotypes, high expression of CD34 and extramedullary infiltration. Combined regimens and more intensive therapy including allo-HSCT might contribute to improving survival.</p>

Analysis of high risk factors for relapse of leukemia after allogeneic hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To screen the high risk factors for relapse after allogeneic hematopoietic stem cell transplantation (allo-HSCT) in acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML) and chronic myeloid leukemia (CML) respectively, then to compare the contribution of each risk factor to relapse and investigate the relevant mechanisms.</p><p><b>METHODS</b>A retrospective study from single center involved in 262 evaluable cases of leukemia received allo-HSCT over the past 8 years, of them 69 cases with ALL, 90 AML (except APL) and 103 CML. Cox proportional hazard regression model was used for univariate and multivariate analysis to screen the high risk factors.</p><p><b>RESULTS</b>The risk factors significantly affecting relapse in ALL included: Cytogenetic risk classification, the cycles of initial induction chemotherapy; AML: Cytogenetic risk classification, minimal residual disease (MRD) level before transplant, reconstitution of WBC, and CD4(+)/CD8(+) lymphocyte ratio in the graft; CML: disease stage before transplant.</p><p><b>CONCLUSIONS</b>The relapse risk after HSCT of ALL mainly depends on the grade of malignancies, and the relapse risk of AML is closely related to the course of transplant. Chronic phase of CML favors a good prognosis after HSCT. Cytogenetic risk classification is the most relevant predictor of relapse after HSCT.</p>

Advance of research on relationship between ABCG2 and drug resistance in hematologic malignancies - review / 中国实验血液学杂志

ABCG2, a half-transporter concerning with the endo and exon-toxin-efflux, plays an important role in protecting the normal tissues from the toxin-hurt as well as mediating the multidrug resistance, because many of the chemotherapeutic drugs are the substrate of ABCG2. In this paper, the advance of research about this gene's single nucleotide polymorphisms (SNPs) was explained concisely. The relationship among ABCG2, the stem cells and the tyrosine kinase inhibitor (TKI) was reviewed. The research about drug resistance related-progress in hematologic malignancies was analyzed retrospectively and the present problems and the perspective in the future were discussed.

Clinical study of allogeneic hematopoietic stem cell transplantation for relapsed/refractory acute lymphocytic leukemia / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the efficacy and toxicity of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for relapsed/refractory acute lymphocytic leukemia (ALL).</p><p><b>METHODS</b>Forty-seven patients with relapsed/refractory ALL received allo-HSCT, which containing 19/47 from HLA-identical sibling donors (sib-HSCT), 18/47 from HLA-identical unrelated donors (URD-HSCT) and 10/47 from haplo-identical donors (Hi-HSCT). Conditioning regimens included "TBI plus Cyclophosphamide (Cy) (42/ 47)" or "busulfan (Bu) plus Cy (5/47)". Cyclosporine (CsA) combined with a short-course Methotrexate (MTX) were used for graft versus host disease (GVHD) prophylaxis. In addition, patients receiving URD-HSCT or Hi-HSCT were given mycophenolate mofetil (MMF) and anti-thymocyte immunoglobulin (ATG). Patients with molecular or cytogenetic relapse tendency on minimal residual disease (MRD) monitoring received donor lymphocyte infusion (DLI).</p><p><b>RESULTS</b>All patients tolerated the therapy well except for mucositis. Renal dysfunction occurred in 2 patients on CsA therapy. Epilepsy occurred in 1 patient, fatal infectious complications in 9 (including 3 interstitial pneumonia), grade III-IV acute GVHD (aGVHD) in 7, chronic GVHD (cGVHD) in 22 and hemorrhagic cystitis (HC) in 4 patients. Thirteen patients relapsed after transplantation. The median time of hematopoietic reconstitution was + 17 ds. Nineteen patients received DLI, and 6 of them had no disease progression. With a median follow-up duration of 43 (10-77) months, the estimated 5-year overall survival (OS) and disease free survival (DFS) rates were 49.65% and 46.55%, respectively.</p><p><b>CONCLUSION</b>Allo-HSCT is an effective therapy for relapsed/refractory ALL. Relapse after transplantation, fatal infection, and severe acute GVHD are the main causes for failure. DLI might decrease the relapse rate after transplantation.</p>

Outcomes of CAG regimen for refractory biphenotypic acute leukemia patients / 中国医学科学杂志(英文版)

<p><b>OBJECTIVE</b>To evaluated the efficiency of low-dose cytosine arabinoside plus aclarubicin with concurrent administration of granulocyte colony-stimulating factor (CAG) regimen for refractory biphenotypic acute leukemia (BAL).</p><p><b>METHODS</b>We treated 5 refractory BAL patients by CAG regimen (10 mg x m(-2) cytosine arabinoside subcutaneously administrated every 12 hours, day 1-14; 5-7 mg x m(-2) aclarubicin intravenously administrated daily, day 1-8; and concurrently used 200 microg x m(-2) x d(-1) granulocyte colony-stimulating factor subcutaneously) from November 2002 to April 2007. The efficacy of the regimen was evaluated by response rate, and the side effects were also measured.</p><p><b>RESULTS</b>The complete remission rate was 80%, median duration of absolute neutrophil count < 5.0 x 10(8)/L and platelet count < 2.0 x 10(10)/L was day 13 and day 1, respectively; and the infection rate was low (III-IV infection rate, 20.00%).</p><p><b>CONCLUSION</b>CAG regimen as remission induction chemotherapy for BAL patients is effective with a high remission rate and low toxicity.</p>

Surgical treatment and pathological findings of hematological malignancies patients complicated with lung diseases / 中华血液学杂志

<p><b>OBJECTIVE</b>To determine the pulmonary pathological changes in hematological malignancy patients with pulmonary complications.</p><p><b>METHODS</b>17 hematological malignancy patients underwent surgical treatment were evaluated retrospectively. The pathological changes of all the surgical specimens were examined postoperatively by standard hematoxylin and eosin (HE) staining.</p><p><b>RESULTS</b>Pathological examination confirmed: aspergillus infection in 9 patients, sub-acute inflammation (fibrosis and hematoma formation) in 3, and each in 1 of pulmonary infarction with granulomatous tissue in the periphery; granulomatous inflammation with calcified tubercle; alveolar dilation and hemorrhage, interstitial fibrosis and focal vasculitis; intercostal neurilemmoma; and moderate-differentiated adenocarcinoma accompanied by intrapulmonary metastasis. And several operative complications (1 case of fungal implantation, 3 pleural effusion and adhesions and 2 pulmonary hematoma) were occurred. The coincidence rate of pre- and post-operative diagnosis was 9/14 (64.3%). After surgery, 8 patients were received hematopoietic stem cell transplantation (HSCT, allo-gene or autologous), with 7 succeeded. On effective secondary antifungal prophylaxis, 4 of 5 patients of aspergillosis succeeded in transplantation with free from mycotic relapse, one patient died from fungal relapse.</p><p><b>CONCLUSION</b>Hematological malignancies with persistent and/or resistant pulmonary infection, hemoptysis, or unexplained lung diseases, should be treated in time by surgery operation to effectively eliminate residual disease and obtain a definitive diagnosis, so as to create a prerequisite condition for the following treatments. Moreover, the secondary antifungal prophylaxis can provide active roles for patients scheduled for chemotherapy and/or HSCT.</p>

A comparison of clinical outcomes between unrelated donor and HLA-haploidentical donor hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To compare the clinical outcomes between unrelated donor hematopoietic stem cell transplantation (URD-HSCT) and HLA-haploidentical (Hi)-HSCT.</p><p><b>METHODS</b>Twenty-five patients with hematologic malignancies received URD-HSCT and thirty patients received Hi-HSCT. The conditioning regimen consisted of modified BUCY or modified total body irradiation (TBI) plus CY. Acute graft-versus-host disease (aGVHD) prophylaxis consisted of cyclosporin ( CsA), short-term methotrexate (MTX), mycophenolate mofetil (MMF), or the combination of CsA, MTX and MMF plus antithymocyte globulin (ATG) or antilymphocyte globulin (ALG), or the combination of CsA, MTX, MMF, ATG/ ALG and CD25 monoclonal antibody.</p><p><b>RESULTS</b>All patients in the URD-HSCT group and 29 patients in the Hi-HSCT group were engrafted successfully. The median follow-up duration was 7 (2 -59) months for URD-HSCT group and 7.3 (1 - 35) months for Hi-HSCT group. The 3-year probabilities of disease-free survival (DFS) for URD-HSCT and Hi-HSCT group were (54.1 +/- 11.9)% and (43.1 +/- 9.1)%, respectively (P =0.13). Grade III - IV aGVHD occurred in 10 patients in URD-HSCT group and 11 in Hi-HSCT group (the cumulative incidence 40.0% vs 37.9%, P > 0.05), respectively. Ten patients (40.0%) died of transplantation-related mortality (TRM) in URD-HSCT group and 17 (56.7%) in Hi-HSCT group (P >0. 5). Two patients relapsed in each group (the rate of relapse 8.0% vs 6.0%, P >0.05). The primary causes of death included severe aGVHD with infection,severe pulmonary infection and relapse.</p><p><b>CONCLUSION</b>Both URD-HSCT and Hi-HSCT are effective and curable treatment for refractory or high-risk hematologic malignancies. The optimal donor should be chose individually. The severe aGVHD and consequent infection are still the main cause of TRM.</p>

Transcriptional activity of WT1 gene promoter and enhancer in diverse cell lines / 中国实验血液学杂志

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.

Application of EGFP as reporter gene in study of WT1 regulation elements / 中国实验血液学杂志

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.

Role of CD28/CTLA-4 co-stimulators in immune pathophysiology of aplastic anemia / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the possible role of CD28/CTLA-4 co-stimulators in immune pathophysiology of acquired aplastic anemia(AA).</p><p><b>METHODS</b>By FACS, the percentages of CD28, CTLA-4 expressing CD3+ CD4+ T cells and the level of Th1, Th2 in bone marrow were detected in 23 AA patients at active phase, 10 at recovery phase and 15 normal controls. The relationship between the co-stimulators, Th1, Th2, and absolute neutrophil counts (ANC) was evaluated.</p><p><b>RESULTS</b>(1) The percentage of CD28 and CTLA-4 expressing CD3+ CD4+ T cells in bone marrow, and CD28+/CTLA-4+ ratios were (31.40 +/- 10.83)%, (2.45 +/- 1.30)% , and 17.02 +/- 13.44 in normal controls respectively, (39.84 +/- 10.89)%, (1.43 +/- 0.67)%, and 43.04 +/- 37.61 in AA at active phase, respectively, (22 +/- 9.08)%, (3.46 +/- 2.26)%, and 10.49 +/- 7.8 in AA at recovery phase, respectively. The percentage of CD28 and CD28+/CTLA-4+ ratio were significantly higher, while CTLA-4 were lower in active phase AA patients than in normal controls (P < 0.05). These values in recovery phase AA were comparable to those in normal controls. (The Th1, Th2, and Th1/Th2 in bone marrow were (4.21 +/- 2.11)%, (1.99 +/- 1.27)%, and 2.46 +/- 1.28 in normal controls respectively, (11.13 +/- 4. 96)%, (2.46 +/- 1.65)%, and 5.20 +/- 1.98 in active phase AA and (5.39 +/- 4.2)9%, (2.53 +/- 2.41)%, and 2.87 +/- 1.43 in recovery phase AA, respectively. The percentage of Th1 and Th1/Th2 ratio were significantly higher in AA patients at active phase than in normal controls (P < 0.05). (3) The CD28+/CTLA-4+ ratio was positively related to the Th1+ /Th2+ ratio (P < 0.05). ANC was negatively related to CD3+ CD4+ CD28+ T cells (P < 0.01), and positively to CD3 + CD4 ' CTLA-4' T cells (P < 0.01) respectively.</p><p><b>CONCLUSION</b>(1) The expression of CD28 was increased while CTLA-4 decreased on the membranes of CD3+ CD4+ T cells in bone marrow of AA patients. (2) The abnormal expression of CD28 costimulator promoted the shift of immune balance to Thl type. (3) The unbalance of CD28+ / CTLA-4+ is important for the immune pathophysiology of AA.</p>

Abnormal expression of cCD79a/cCD22 in acute myeloid leukemia with t (8;21) / 中华血液学杂志

<p><b>OBJECTIVE</b>To report abnormal expression of cCD79a/cCD22 in four cases of acute myeloid leukemia (AML) with t (8;21).</p><p><b>METHODS</b>The characteristics of morphology, immunophenotype, chromosome karyotype (MIC) and clinical manifestations of 4 AML patients with t (8;21) expressing cCD79a/cCD22 were analyzed.</p><p><b>RESULTS</b>The features of the 4 patients were: (1) no difference in gender; (2) young age; (3) exmedullary infiltration may be present; (4) normal number of white blood cells in peripheral blood; (5) morphology showed acute myeloid leukemia with high percentage of blast cells; (6) B-lymphoid and myeloid immunophenotype, and high expression of CD34; (7) frequent depletion of Y chromosome and complex changes of chromosomes; (8) positive for AML1/ETO fusion gene; (9) response well to chemotherapy regimen which simultaneously treated myeloid and lymphocytic leukemia.</p><p><b>CONCLUSION</b>Abnormal expression of cCD79a/cCD22 in AML with t (8;21) (q22;q22) suggested that this kind of leukemia might be related with abnormal expression gene of B cell.</p>

Application of sequential and quantitative monitoring of chimerism in allogeneic hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To establish multiple short tandem repeat (STR) amplification by fluorescence labeling polymerase chain reaction (PCR) combined with capillary electrophoresis for quantitative determination of chimerism, and to evaluate the status of engraftment and predict the outcome of allogeneic hematopoietic stem cell transplantation (allo-HSCT).</p><p><b>METHODS</b>Thirty-one patients received bone marrow transplantation (BMT) or nonmyeloablative allogeneic stem cell transplantation (NST) were evaluated. Peripheral blood and bone marrow were co-llected before and after transplantation in different period. Nine different STR markers were co-amplified in a single reaction by using a commercial AmpF/STR Profiler Plus PCR amplification kit. Separation of the PCR products and fluorescence detection were performed by ABI prism 310 Genetic Analyzer with capillary electrophoresis. The Genescan and Genotype software were used for size calling and quantification of peak areas. The formula to calculate donor chimerism values was based on the different allelic distribution type between donor and recipient.</p><p><b>RESULTS</b>48.4% of the patients received sex-matched transplantation and the quantification of donor chimerism could only be performed by STR-PCR method. Comparison of values obtained by FISH analysis with that by STR-PCR in patients transplanted from sex-mismatched donors showed an excellent correlation. The median number of informative alleles was 6.7 (range 2 - 10). The donor's alleles appeared in all the patients on day 7 post-transplant. The median values of donor chimerism in BMT group were inferior to that in NST group on day 7, day 14 and 1 month post-transplant. However the difference disappeared in the midterm or later period of transplant. On day 21, all of the 31 patients had stable engraftment and the percentage of donor chimerism was more than 92%. Median follow-up was 17 (3.5 - 29.0) months after transplantation. Twenty-six of 31 patients had durable engraftment and donor chimerism ratio was more than 90%. So for all of them survived leukemia-freely. Four of the 31 patients had unstable mixed chimerism and relapsed within 6 months post allo-HSCT. Another patient with unstable mixed chimerism appeared graft rejection. Decreasing values of donor chimerism were detected prior to the occurrence of graft rejection and disease relapse. The incidence of GVHD was much higher in the group of full donor chimerism.</p><p><b>CONCLUSION</b>Sequential and quantitative monitoring of STR is a valuable tool for studying engraftment dynamics, graft rejection, and relapse and for predicting GVHD. Furthermore it can provide a basis for early intervention of clinical treatment.</p>

Mobilization of peripheral blood stem cells with mitoxantrone and high-dose cytarabine chemotherapy and rhG-CSF in patients with hematopoietic malignancies / 中华血液学杂志

<p><b>OBJECTIVE</b>To evaluate the efficacy of mitoxantrone combined high dose of cytarabine and recombinant human granulocyte colony-stimulating factor (MAG) regimen for mobilizing autologous peripheral blood stem cells (APBSC) in patients with hematopoietic malignancies.</p><p><b>METHODS</b>From December 1995 to April 2003, 14 lymphoma and 29 acute leukemia patients were treated with high-dose cytarabine (2 g/m2 every 12 h, days 1 and 2) and mitoxantrone (10 mg/m2, days 2 and 3), followed by 300 microgram recombinant human granulocyte colony-stimulating factor per day (rhG-CSF 300 microg/d) i.e, the MAG regimen as mobilization regimen of peripheral blood stem cells. rhG-CSF was given subcutaneously when the white blood cell (WBC) count below 1.0 x 10(9)/L following the MA chemotherapy, APBSC were harvested when WBC count increased using Baxter CS3000plus or Cobe Spectra.</p><p><b>RESULTS</b>Mobilization was successful in 13 of 14 lymphoma patients with MNC (3.91 +/- 2.70) x 10(8)/kg, CD34+ cells (17.79 +/- 12.90) x 10(6)/kg. Meanwhile, mobilization was successful in 24 of 29 acute leukemia patients with average of 2.13 times for apheresis. The median MNC and CD34+ cells yielded were 3.62 x 10(8)/kg and 7.37 x 10(6)/kg respectively, rhG-CSF was used for a median time of 7 days. Excepting for grade I-II gastrointestinal toxicity in 8 and infection in 14 cases, no major side effects were observed. There was no mobilization-related mortality. Minimal residual diseases became undetectable after mobilization in some patients.</p><p><b>CONCLUSION</b>MAG is a safe and highly effective mobilization regimen in patients with lymphoma and acute leukemia.</p>

Pulmonary disease caused by nontuberculous mycobacteria after allogeneic hematopoietic stem cell transplantation--a case report and literature review / 中华血液学杂志

<p><b>OBJECTIVE</b>To report a case with pulmonary disease caused by nontuberculous mycobacteria (NTM) after allogeneic hematopoietic stem cell transplantation (allo-HSCT), with a literature review.</p><p><b>METHODS</b>Case report and literature review.</p><p><b>RESULTS</b>A patient with acute non-lymphocytic leukemia was treated by allo-HSCT. Her NTM lung disease developed after HSCT was successfully treated with a 3 antimicrobials combination of clarithromycin, levofloxacin and capreomycin for 10 months.</p><p><b>CONCLUSION</b>NTM infections are infrequent in allo-HSCT recipients and have a good clinical prognosis if correctly treated.</p>