Efficacy and Safety of Decitabine Combined with Modified CAG Regimen in Patients Aged ≥ 70 Years with Newly Diagnosed Acute Myeloid Leukemia / 中国实验血液学杂志

OBJECTIVE@#To evaluate the clinical efficacy and safety of decitabine combined with modified CAG regimen (D-CAG regimen) in patients aged ≥70 years with newly diagnosed acute myeloid leukemia (AML).@*METHODS@#The clinical data of 59 AML patients (≥70 years old) who were newly diagnosed and treated in the Hematology Department of the First Affiliated Hospital of Nanjing Medical University from November 2010 to June 2021 were retrospectively analyzed.@*RESULTS@#Among the 59 AML patients, 28 were males and 31 were females, with a median age of 74 (70-86) years. The complete remission (CR) rate was 69.4% (34/49), and the median duration of CR was 10.7 (0.6-125.4) months after 2 courses of D-CAG treatment. According to the British Medical Research Council (MRC) classification, there was only one patient in the favorable-risk group, and the CR rate was 71.8% (28/39) in the intermediate-risk group, and 55.6% (5/9) in the adverse-risk group, respectively. There was no statistical difference in the CR rate between the intermediate-risk and adverse-risk group. Referring to ELN 2017 genetic risk classification, CR rate was 88.2% (15/17) in the favorable-risk group, 45.5% (5/11) in the intermediate-risk group, and 66.7% (14/21) in the adverse-risk group. There was no significant difference in CR rate between the favorable-risk and adverse-risk categories, but both were significantly higher than that in the intermediate-risk group (P <0.05). Next-generation sequencing (NGS) analysis showed that 11 gene mutations with a frequency of more than 10%, including TET2 mutation (35.6%), ASXL1 mutation (30.5%), NPM1 mutation (28.8%), FLT3-ITD mutation (27.1%), DNMT3A mutation (22.0%), IDH1 mutation (15.3%), CEBPA single mutation (13.6%), TP53 mutation (13.6%), IDH2 mutation (11.9%), RUNX1 mutation (11.9%), and NRAS mutation (10.2%). There were no statistical differences in mutation frequency of these 11 genes between CR group and non-CR group. Compared with normal karyotypes, patients with complex karyotypes were more likely to develop TP53 mutations (P <0.001), while FLT3-ITD and DNMT3A mutations were more likely to occur in patients with normal karyotypes (P =0.04, P =0.047). The median follow-up, overall survival (OS), and event-free survival (EFS) of all the patients was 11.7 (1.5-128.2) months, 12.3 (1.5-128.2) months, and 8.5 (1.5-128.2) months, respectively. The median OS and EFS of CR patients were 19.8 and 13.3 months, respectively, which were significantly longer than 6.4 and 5.7 months in patients experiencing treatment failure (P < 0.001, P =0.009). In regard to genes with mutation frequency >10%, there were no statistical differences in CR rate, median OS, and median EFS between mutated and wild-type patients by Chi-square test and survival analysis. Univariate analysis showed that age, hemoglobin, lactate dehydrogenase, cytogenetics and CR were factors affecting prognosis, while multivariate analysis showed that only CR failure was an independent adverse prognostic factor for OS. The major adverse reactions to D-CAG regimen were grade 3-4 myelosuppression, pulmonary infection, and fever (infection focus was not identified).@*CONCLUSION@#D-CAG regimen is safe and effective in the treatment of AML patients ≥70 years old, and can partially improve the prognosis of elderly and high-risk patients.

The Risk and Survival Analysis of Multiple Malignancies in Hematologic Malignancy Patients: A Single Chinese Center Retrospective Study, 2009 through 2017 / 中国实验血液学杂志

OBJECTIVE@#To explore the risk and location of multiple malignancies in patients with hematologic malignancies who were followed up for 9 years in Jiangsu Province Hospital and to evaluate the impact of the second primary malignancy on survival of patients.@*METHODS@#The incidence and survival of multiple malignancies in 7 921 patients with hematologic malignancies from 2009 to 2017 were analyzed retrospectively.@*RESULTS@#A total of 180 (2.3%, 180/7 921) patients developed second malignancy, of whom 58 patients were diagnosed with hematologic malignancies as the first primary malignancy, and 98 patients developed hematologic malignancies as second primary malignancy, and the other 24 cases were diagnosed with the second malignancy within 6 months after the first primary malignancy was diagnosed, which was difined as multiple malignancies occurring simultaneously. In 180 patients, 18 cases developed two hematologic malignancies successively, and 11 patients developed more than 3 primary cancers (among them, 2 female patients were diagnosed with 4 primary cancers). Patients with lymphoma and multiple myeloma (MM) as the second primary malignancy had poorer survival than patients with lymphoma and MM as the first primary malignancy. Patients with chronic myeloid leukemia as the second primary malignancy were also associated with inferior overall survival.@*CONCLUSION@#In this study, 2.3% of hematologic malignancy patients had multiple mali-gnancies, lymphoma and MM as the second primary malignancy had poor survival.

The Expression and Function of NK Cells in Patients with Acute Myeloid Leukemia / 中国实验血液学杂志

OBJECTIVE@#To explore the expression characteristics of antigens and functional markers of natural killer (NK) cells in patients with acute myeloid leukemia (AML).@*METHODS@#Multi-parameter flow cytometry was used to detect NK cell surface markers and their functional indicators in 56 newly diagnosed AML patients and 24 healthy controls, including activating receptors NKG2D, NKP46, DNAM-1, and killing indicators granzyme B, perforin.@*RESULTS@#Referring to the WHO hematopoiesis and lymph tissue tumor classification criteria, 56 cases were roughly divided into three types: AML M1, M2, and M4/M5. However, there was no differences about NK cells among the three types, so it was no longer subdivided. NK cells were divided into two groups: CD3-CD56hiCD16- (CD56hiNK) and CD3-CD56dimCD16+ (CD56dimNK). Compared with CD56dimNK cell population, except for NKP46, the positive expression levels of NKG2D and other receptors of CD56hiNK cells in AML patients decreased (P<0.001). Compared with healthy controls, the proportion of CD56hiNK cells in AML patients increased, while the number and proportion of NK cells and proportion of CD56dimNK cells significantly decreased (P<0.05). The proportion of perforin in CD56hiNK cells significantly increased (P<0.05). The expression of DNAM-1 in CD56hiNK cells, NKG2D, DNAM-1, and perforin in CD56dimNK cells decreased significantly (P<0.05). There was no statistically significant difference in expression of other functional indexes in AML patients compared with corresponding indexes of healthy controls. In addition, the proportion of CD56hiNK cells was positively correlated with the expression of CD34+ in AML (r=0.303).@*CONCLUSION@#Compared with CD56dimNK, the ratio of CD56hiNK and the expression of functional markers in AML patients are lower. Compared with healthy controls, the number and expression ratio of NK cells in AML patients decrease and the expression of functional markers is abnormal, indicating that its function is impaired.

A Retrospective Analysis of TKI Discontinuation in Patients with CML / 中国实验血液学杂志

OBJECTIVE@#To investigate the clinical characteristics of the patients with chronic myeloid leukemia (CML) discontinued tyrosine kinase inhibitors (TKI) therapy and the outcome of the patients.@*METHODS@#35 cases of CML patients experienced initiative discontinuation of TKI therapy in our hospital from June 1st 2015 to December 31th 2019 were retrospectively analyzed. The TFR of the patients and the factors affecting it were analyzed.@*RESULTS@#The median duration of TKI administration was 72 (range 35-173) months in the 35 patients. Among these patients, 8 had experienced TKI dose reduction or suspension. All the enrolled patients have achieved at least MMR. The median time for these patients achieving MMR was 15 (range 3-75) months after administration of TKI, and for MMR maintenance before TKI suspension was 55 (range 13-164) months. After TKI withdrawal the median follow up time was 20.3 (range 3-57.9) months, 22 out of 35 patients kept TFR, among them, 2 （5.71%） patients restarted TKI after 12 month suspension, and maintained MMR during suspension. 13 （37.1%）patients lost MMR, among them, 9 patients restarted TKI treatment, and 5 of them achieved MR4.0 after the median duration of 3(2-5) month. No patients were found to have disease progression. The estimated TFR rate was 57.8% and 51.8% at 12 and 24 months after discontinuation, respectively. Other clinical characteristic related to relapse were also analyzed, including the cumulative TKI administration duration, cumulative MMR duration, time to achieve MMR, median age at diagnosis, risk stratification by Sokal score, TKI dose reduction and discontinuation history, and second-generation TKI administration before stopping TKI, however, no statistical difference was found.@*CONCLUSION@#TKI discontinuation is practical for CML patients in our center.

Analysis of the Differential Expression of circRNA in Acute Myeloid Leukemia by GEO Database / 中国实验血液学杂志

OBJECTIVE@#To investigate the difference expression of circular RNA (circRNA) in acute myeloid leukemia (AML) by using bioinformatics method.@*METHODS@#The microarray chip data of AML was searched and downloaded from the Gene Expression Omnibus (GEO) of the National Center for Bioinformatics (NCBI). The differences between AML samples and control samples were analyzed by R software. The interaction between deregulated circRNA, miRNA and mRNA were predicted by miranda software and miRTarBase software. The circRNA-miRNA-mRNA regulatory network was constructed by using the cytoHubba plugin based on the Cytoscape software.@*RESULTS@#A total of 203 differential expression of circRNAs were finally collected, including down-regulated 161 circRNAs and up-regulated 42 circRNAs. CircRNA/miRNA/mRNA interaction network was constructed through software prediction. hsa_circ_0001080, hsa_circ_0004511, hsa_circ_0054211, hsa_circ_0001944 may be positively regulated the gene expression in AML.@*CONCLUSION@#Abnormal expression of circRNA in AML may become a new target for AML treatment.

Distribution of Natural Killer Cell in Patients with Newly Diagnosed Myelodysplastic Syndromes and Its Correlation with Prognosis / 中国实验血液学杂志

OBJECTIVE@#To investigate the distribution of peripheral blood lymphocytes and natural killer (NK) cells, and its influence on the prognosis of patients with myelodysplastic syndromes (MDS).@*METHODS@#The lymphocytes proportion, absolute lymphocyte counts (ALC), NK cell proportion and absolute NK cell counts (ANKC) as well as the related data of 95 MDS patients diagnosed between 2013 and 2017 analyzed retrospectively. The correlation of ALC and ANKC with prognosis was also analyzed.@*RESULTS@#As compared with low ALC patients, MDS patients with ALC≥0.885×10/L had a higher overall response rate (66.7% vs 35.8%) (P＜0.01). The ALC of effective patients after treatment significatitly increased in compaison of ALC at diagnosis. Multivariate analysis indicated that patients with ALC≥0.885×10/L had long overall survival (OS) time in comparison with patients with low level (16.4 vs 12.4 months) (P＜0.05). The OS time of patients with ANKC≥0.110×10/L was shorter in comparison with patients with low level (10.9 vs 16.3 months) (P＜0.01). Otherwise, blast, cytogenetic risks and treatment response were also independent risk factors of MDS (P＜0.05). Revised International Prognostic Scoring System (IPSS-R) combined with ANKC could improve predictive accuracy of IPSS-R alone (AUC 0.718 vs 0.674) (P＜0.05).@*CONCLUSION@#Lymphocytes and NK cells are important for the prognosis evaluation of MDS patients.

Lower clearance of sodium tanshinone IIA sulfonate in coronary heart disease patients and the effect of total bilirubin: a population pharmacokinetics analysis / 中国天然药物

This study developed a population pharmacokinetic model for sodium tanshinone IIA sulfonate (STS) in healthy volunteers and coronary heart disease (CHD) patients in order to identify significant covariates for the pharmacokinetics of STS. Blood samples were obtained by intense sampling approach from 10 healthy volunteers and sparse sampling from 25 CHD patients, and a population pharmacokinetic analysis was performed by nonlinear mixed-effect modeling. The final model was evaluated by bootstrap and visual predictive check. A total of 230 plasma concentrations were included, 137 from healthy volunteers and 93 from CHD patients. It was a two-compartment model with first-order elimination. The typical value of the apparent clearance (CL) of STS in CHD patients with total bilirubin (TBIL) level of 10 μmol(L was 48.7 L(h with inter individual variability of 27.4%, whereas that in healthy volunteers with the same TBIL level was 63.1 L(h. Residual variability was described by a proportional error model and estimated at 5.2%. The CL of STS in CHD patients was lower than that in healthy volunteers and decreased when TBIL levels increased. The bootstrap and visual predictive check confirmed the stability and validity of the final model. These results suggested that STS dosage adjustment might be considered based on TBIL levels in CHD patients.

Mean Corpuscular Volume Can Be A Predictor for Therapeutic Response of Patients with Chronic Myeloid Leukemia / 中国实验血液学杂志

<p><b>OBJECTIVE</b>The past studies found that the treatment of chronic myeloid leukemia (CML) with imatinib can induce the macrocytic anemia, moreover the incidence of anemia increases along with enhancement of imatinib concentration. This study was aimed to evaluate the potential relation of erythrocyte mean corpuscular volume (MCV) increase after the treatment with tyrosine kinase inhibitors (TKI) with the therapeutic response in patients with CML-chronic phase (CML-CP).</p><p><b>METHODS</b>The clinical and hematologic data including MCV, molecular and cytogenetic response of 119 patients with CML-CP were collected after treatment with TKIs, and the relation of MCV changes after treatment with the clinical characteristics and therapeutic efficacy for patients with CML-CP was analyzed.</p><p><b>RESULTS</b>The MCV in patients treated with TKIs for 12 months significantly increased as compared with that at initial diagnosis (P<0.05). The proportion of patients with increased MCV in group of complete cytogenetic response (CCyR) was significantly higher than that in group of non-CCyR (P<0.05). As compared with decreased MCV group, the patients in increased MCV group much more easily achieved CCyR after treatment for 6, 12 months (P<0.05, P<0.05) respectively, furthermore, much more easily maintained MMR (P<0.05).</p><p><b>CONCLUSION</b>The MCV as a parameter which is easily acquired may be a new marker for prodecting the therapeutic response of patients treated with TKIs.</p>

Analysis of Clinical and Laboratory Features of Patients with Acute Myeloid Leukemia(AML)-M1/M2 with Cuplike Nuclei Morphology / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To analyze the clinical and laboratory features of acute myeloid leukemia (AML) with cuplike nuclei morphology.</p><p><b>METHODS</b>One hundred and seventy patients diagnosed with AML (M1andM2) between December 2009 and December 2016 were included in the study. Bone marrow smears were prepared for morphologic alanalysis, the immunophenotype was analyzed by flow cytometry and the RHG-banding was for conventional cytogenetic assay (CCA) ,gene mutation was detected by sequencing.</p><p><b>RESULTS</b>Among the 170 AMLpatients, 67 were diagnosed as M1 and 103 patients was diagnosed as M2, 43 patients(25.3%) defined as cuplike nuclei-positive, among them 38patients (88.4%) were M1 while only 5 patients (11.6%) were with M2(P<0.01). No significant value about sex(P> 0.05) between cuplike nuclei-positive and -negative group, while older patients were found in cuplike nuclei-positive group (P<0.05). Higher peroxydas (POX) ratio (P<0.05) and integration (P<0.05) were found in cuplike nuclei- positive group. Furthermore, the patients with cuplike nuclei-positive lack the expressions of CD34 (P<0.01) and HLA-DR(P<0.01) while no other immunophenotype markers were found. Among the 152 patients (89.4%) for genetic analysis ,83.8% karyotype of the cuplike nuclei-positive group were normal while only 54.8 of negative group was normal by CCA. Molecular biology analysis showed that the patients in cuplike nuclei-positive group have significantly highe rNMP1 (P<0.01) and FLT3(P<0.01) mutations as compared with the negative group. Furthermore, the relationship of the ratio o fcuplike nuclei and the type of gene mutations were investigated, and no significant associations were found. However, it was found that the patients with FLT3 mutation displayed more biological nuclear invagination than the patients with NPM1 mutations (P<0/01).</p><p><b>CONCLUSION</b>AML patients with positive cuplike nuclei have characteristic morphological changes, typical immunophenotype with HLA-DR- and CD34, normal karyotype accompanied by NPM1 and FLT3 mutations.</p>

Distribution of Type 9 T Helper Cells and Expression of Transcriptional Factors in Acute Myeloid Leukemia / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the distribution of T helper (Th9) cells and its relationship with clinical characteristics of patients with acute myeloid leukemia (AML), and to analyze the activating levels of different transcriptional factors in Th9 cells.</p><p><b>METHODS</b>The peripheral blood specimens of 102 AML patients and 83 healthy persons as controls were collected, then the T cells of peripheral blood in AML patients and controls were isolated by using CD3 magnetic beads, the mRNA expression of IL-9 was detected by real-time quantitative PCR, the Th9(CD4IL-9) cell levels in diffrent stages and activating level of Th9 coexpression with IL-9 were detected by flow cytometry.</p><p><b>RESULTS</b>The mRNA expression of IL-9 in peripheral blood of AML M2 and M3 patients was significantly higher than that in control groups (P<0.01), at same time the CD4IL-9cell rate was significantly higher than that in control group also(P<0.01). The results of dynamically monitoring the distribution of Th9 cells in AML-M2 and M3 patients showed that the Th9 cell rate and the mRNA expression of IL-9 in newly diagnosed M2 and M3, and relapsed M2 groups were significantly higher than those of M2 and M3 in remission (P<0.01); the detection results of IL-9co-expression with transcriptional factors (SMAD3, IRF-1and IRF-4) indicated that the percantage of Th9 pSMAD3cells in peripheral blood of newly diagnosed and relapsed M2 and M3 patients was significantly higher than that in M2 and M3 patients in remission (P<0.01); on the contrary, the percentage of Th9 IRF-1cells in peripheral blood of M2 and M3 patients in remission was significantly higher than that in newly diagnosed M2 and M3 patients (P<0.01).</p><p><b>CONCLUSION</b>The distribution of T helper cells in peripheral blood of AML-M2 and M3 patients significantly increases, moreover, correlates with disease status. The prediction of Th9 cell functions should be performed in combination with it transcriptional factors which have inmportant significance for microenvironment of tumors in AML patients.</p>

Heparin-derived oligosaccharide inhibits vascular intimal hyperplasia in balloon-injured carotid artery / 中国天然药物

The aims of the present study were to determine the effects of heparin-derived oligosaccharides (HDOs) on vascular intimal hyperplasia (IH) in balloon-injured carotid artery and to elucidate the underlying mechanisms of action. An animal model was established by rubbing the endothelia within the common carotid artery (CCA) in male rabbits. The rabbits were fed a high-cholesterol diet. Arterial IH was determined by histopathological changes to the CCA. Serum lipids were detected using an automated biochemical analysis. Expressions of mRNAs for vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), vascular cell adhesion molecule-1 (VCAM-1), monocyte chemoattractant protein-1 (MCP-1), scavenger receptor class B type I (SR-BI), and ATP-binding cassette transporter A1 (ABCA-1) were analyzed using reverse transcription polymerase chain reaction assays. Expressions of VEGF, VCAM-1, MCP-1, SR-BI and ABCA-1 proteins were analyzed by Western blotting. Enzyme-linked immunosorbent assays were used to quantify expression levels of VEGF and bFGF. Our results showed that administration of HDO significantly inhibited CCA histopathology and restenosis induced by balloon injury. The treatment with HDOs significantly decreased the mRNA and protein expression levels of VEGF, bFGF, VCAM-1, MCP-1, and SR-BI in the arterial wall; however, ABCA-1 expression level was elevated. HDO treatment led to a reduction in serum lipids (total cholesterol, triglycerides, high-density and low-density lipoproteins). Our results from the rabbit model indicated that HDOs could ameliorate IH and underlying mechanism might involve VEGF, bFGF, VCAM-1, MCP-1, SR-BI, and ABCA-1.

Nilotinib treatment for patients with imatinib-resistant or intolerant chronic myeloid leukemia / 中国实验血液学杂志

This study was purposed to evaluate the efficacy and safety of nilotinib for treating patients with imatinib-resistant or intolerant chronic myeloid leukemia (CML). A total of 23 patients with imatinib-resistant or intolerant CML were enrolled in this study. These patients received nilotinib orally 600-800 mg every day, their curative efficacy, tolerance and overal survival were evaluated. The results showed that all the patients treated with nilotinib obtained complete hematologic remission (CHR), out of them 82.6% patients achieved complete cytogenetic remission (CCyR) and 56.5% patients achieved complete molecular remission (CMR), their adverse events mostly were mild to moderate, generally were transient and easily cured; the median treatment time with nilotinib was 13.5 (1-44) months, and the median follow-up time was 40 (12-102) months. It is concluded that nilotinib has been confirmed to be effective for patients with imatinib-resistant or intolerant CML, and may be selected as a second generation of tyrosine kinase inhibitor (TKI).

Effects of multiple-trough sampling design and algorithm on the estimation of population and individual pharmacokinetic parameters / 药学学报

The purpose of this study is to investigate the effects of multiple-trough sampling design and nonlinear mixed effect modeling (NONMEM) algorithm on the estimation of population and individual pharmacokinetic parameters. Oxcarbazepine and tacrolimus were used as one-compartment and two-compartment model drugs, respectively. Seven sampling designs were investigated using various number of trough concentrations per individual ranging from 1-4. Monte Carlo simulations were performed to produce state-steady trough concentrations. One-compartment model was used to fit simulated data from oxcarbazepine and tacrolimus. The accuracy and precision of the estimated parameters were evaluated using the median prediction error (PE), the median absolute PE and boxplot. The results indicated that trough concentrations could yield reliable estimates of apparent clearance (CL/F). For oxcarbazepine, as the number of trough concentrations per subject increased, the accuracy and precision of CL/F, between-subject variability (BSV) of CL/F and residual variability (RUV) tended to be improved. For tacrolimus, however, although no improvement were observed in the accuracy of CL/F and BSV of CL/F, the PE distribution ranges were significantly narrowed and the RUV estimates were less bias and imprecise. In terms of algorithm, Monte Carlo importance sampling (IMP) and IMP assisted by mode a posteriori estimation (IMPMAP) were consistently better than other methods. Additionally, the sampling design had no significant effects on the individual parameter estimates, which were only depended on the interaction between BSV and RUV in various algorithms. Decreased in BSV and RUV levels can improve the accuracy and precision of the estimation for both population and individual pharmacokinetic parameter estimates.

Efficacy of dasatinib in treatment of imatinib-resistant BCR/ABL positive leukemia / 中国实验血液学杂志

This study was aimed to evaluate the efficacy and safety of dasatinib in BCR/ABL positive leukemia patients with primary or secondary resistance to imatinib. 27 patients with primary or secondary imatinib-resistant chronic myelogenous leukemia (CML) or Philadelphia chromosome positive acute lymphocytic leukemia (Ph(+) ALL) received 100 - 140 mg/d dasatinib orally. Their overall survival and tolerance were evaluated. The results showed that the median duration of dasatinib therapy was 8 (1-66) months in the 27 imatinib-resistant BCR/ABL positive leukemia cases, with a median follow-up of 54 (3-75) months. After the dasatinib treatment, 88.8% of all the 27 cases achieved complete hematologic response (CHR), 29.6% of them achieved major cytogenetic response (mCyR), 37% of all achieved complete cytogenetic response (CCyR) and 18.5% cases achieved major molecular response (MMR). Patients who received dasatinib in progress of disease (CML-AP, CML-BC and bone marrow relapse Ph(+) ALL) had a lower CCyR rate than those in stable disease (CML-CP and bone marrow remission Ph(+) ALL) (P = 0.0377), and 3 - 4 grade adverse events occurred more frequently in progress of disease than that in stable disease. Overall survival of the patients who achieved CCyR after dasatinib therapy was statistically longer than those who did not achieve CCyR (63 m vs 9 m, P = 0.0126). The most common grade 3 - 4 adverse events during dasatinib therapy including hematology events such as thrombocytopenia (51.8%), neutropenia (48.1%), anemia (33.3%), and non-hematologic events such as pleural effusion (18.5%), pulmonary infection (18.5%), pericardial effusion (11.1%). The 3-4 grade adverse events occurred within 12 months from dasatinib therapy, and were mainly observed in patients with progress of disease. It is concluded that dasatinib is an effective drug in imatinib-resistant BCR/ABL positive leukemia patients, the better curative effect and better tolerance has been observed in patients who received dasatinib in stable disease.

Effects of matrine on the expression of NKG2D ligands in leukemia cells / 中国实验血液学杂志

This study was aimed to analyze the expression of NKG2D ligands in human leukemic cells and to investigate the effects of matrine on NKG2D ligand expression. The expressions of NKG2D ligand MICA/B, ULBP1-3 in several human leukemia cell lines (K562, OUN-1, U937 and K562/AO2), as well as primary leukemic cells isolated from malignant leukemia patients were analyzed by flow cytometry. After treatment with different doses of matrine, the expression level of NKG2D ligands in these leukemic cells was detected by FCM. The results indicated that NKG2D ligand expression was detected in both the leukemia cell lines and primary malignant leukemic cells. Generally, the expression of ULBP was high or obviously higher than that of MICA/B in leukemia cell lines and primary leukemic cells. The expression pattern of NKG2D ligands was different among these cells, possibly due to the different types of leukemia. Not all the expression of NKG2D ligands was upregulated after matrine treatment. Much higher expressions of ULBP2 and ULBP3 were found in K562 cells, compared to the other cell lines, which partly contributes to the higher sensitivity of K562 cells to NK cytotoxicity as target cells. It is concluded that there is universal expression of NKG2D ligand in leukemia cells. The high ULBP expression is prevalent in human leukemia cells. Matrine has the potential to induce the expression of NKG2D ligands in leukemia cells.

Detection and clinical features of MLL gene rearrangement in adult patients with acute leukemia / 中国实验血液学杂志

This study was purposed to investigate the incidence of mixed lineage leukemia (MLL) gene rearrangement and partner gene types as well as the clinical features and prognosis of acute leukemia (AL) with this rearrangement through detection in adult AL using combination of 3 techniques, and to evaluate the clinical value of this combination detection. The MLL gene rearrangement in 183 cases of adult AL was detected by combination of conventional cytogenetics, split signal FISH and multiplex nested PCR. The results showed that the incidence of MLL rearrangements in adult patients with AL was low (8.2%), and MLL-AF4 fusion gene was most common and predominant in acute lymphoblastic leukemia (ALL), while the MLL-AF6 and MLL-AF9 were most frequent in acute myeloid leukemia (AML). Extramedullary involvements were found in 40% of MLL-rearranged AL patients, and 33.3% of patients with MLL-rearranged AL reached to complete remission within 30 days during induction chemotherapy. In addition, in this cohort of MLL-rearranged adult AL patients, the 3-month relapse rate and 6-month overall survival rate were 50.0% and 50.0% respectively. It is concluded that the rate of missed diagnosis of CC technique for patients with MLL-rearranged AL reached to 60% in this study, while the combination of CC, FISH and multiplex nested PCR has been confirmed to have important significance for evaluating prognosis and conducting clinical therapy of patients with MLL-rearranged AL.

Clinical study on high-dose etoposide with granulocyte colony-stimulating factor for mobilization of autologous peripheral blood stem cells in patients with hematologic malignancies / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the effectivity and safety of single high-dose (HD) etoposide (Vp16) with granulocyte colony-stimulating factor (G-CSF) for mobilization of autologous peripheral blood stem cells (PBSC) in patients with hematologic malignancies.</p><p><b>METHODS</b>80 patients of hematologic malignancies including 20 patients with acute leukemia (AL), 23 with multiple myeloma (MM), 35 with non-Hodgkin's lymphoma (NHL) and 2 with Hodgkin's lymphoma (HL) received Vp16 (1.6 g/m(2)) continuous intravenous infusion for 10 hrs on day 1. G-CSF at 10 µg/kg once daily subcutaneous injection began to use on day of ANC lower than 1×10(9)/L and continued until PBSC collection was completed. Autologous PBSC (APBSC) was collected on day of WBC greater than 5×10(9)/L and continuing until the collection goal was met (target value: MNC ≥ 6.0×10(8)/kg and CD34(+) ≥ 2.0×10(6)/kg). The patients received APBSC after conditioning regimen. The number of the cells collection, time of hematopoietic reconstruction, adverse effect and so on were observed during the course of stem cell mobilization and collection.</p><p><b>RESULTS</b>PBSC was collected on day 11 (range: 7 - 25 days) of after Vp16 administration with a median collection time of 2 (range 1 - 5). 3/80 patients with AML got stem cell mobilization failure. 5 of 6 patients who failed to mobilize before got successful stem cell mobilization, 1/6 patient with AML-M(5) got a second failure after the mobilization of VP16 whose first time's mobilization using Ara-C did not succeed. The median number of CD34(+) cells collected in 77 patients who got successful mobilization was 4×10(6)/kg \[range (1.59 - 24.68)×10(6)/kg\]. The collection of 20 patients with AL and 23 with MM were got detection for minimal residual disease, no pollution of tumor cells were happened. All patients could tolerate the whole course of stem cell mobilization. 29/80 (36.25%) patients got a 4 grade leucopenia, 19/80 (23.75%) patients got infection.</p><p><b>CONCLUSION</b>Single high-dose etoposide with G-CSF for mobilization of APBSC has a higher achievement ratio, a controllable adverse effect, a promising hematopoiesis recovery, which is an effective and safe mobilizing regimen for patients with hematologic malignancies.</p>

Expression characteristics of isoforms of Ikaros and Helios in patients with leukemia and their mechanism / 中国实验血液学杂志

This study was aimed to investigate the expression characteristics of two transcriptional factors in Ikaros family, Ikaros and Helios isoforms and their mechanism, as well as their correlation with clinical parameters, which play important roles in transcriptional regulation of hematopoiesis. Expression of Ikaros and Helios isoforms in a total of 163 patients with leukemia and correlations between Ikaros and Helios isoforms were analyzed by PCR. The results showed that different expression patters of Ikaros and Helios isoforms existed in leukemia patients, that is, Ikaros isoform (Ik-6) was predominantly expressed in acute lymphoblastic leukemia (ALL) with BCR/ABL fusion gene, while Helios isoform (He-i) was overexpressed in T-cell ALL patients. The results of cloning and sequencing demonstrated that the isoforms of Ikaros and Helios had different genetic alterations. The statistical correlation between these two isoforms not was found in this study, although interaction between Ikaros and Helios has been reported. It is concluded that although Ikaros and Helios belong to the same family with similar structure of zinc fingers, their isoforms have different expression profile, specific genetic alterations, and different clinical relevance in patients with leukemia. The connection and interaction between Ik-6 and He-i needs further research.

Comparison of conventional cytogenetics and interphase fluorescence in situ hybridization in diagnosis of acute promyelocytic leukemia / 中国实验血液学杂志

The aim of this study was to explore cytogenetic characteristics of acute promyelocytic leukemia (APL) and compare the interphase fluorescence in situ hybridization (I-FISH) with conventional cytogenetic (CC) analysis. A total number of 157 APL patients were recruited in this study, and the I-FISH and CC were applied to analyze cytogenetic features. Chromosome samples of bone marrow cells were prepared by short-term culture. Out of all 157 cases, 136 were observed with CC assay, 66 with I-FISH, of which 45 samples were analyzed with both methods. The results showed that among all 136 CC samples, t(15;17)(q22;q21) was found in 120 cases, of which 107 cases was isolated t(15;17)(q22;q21) abnormality, 13 cases was complex abnormalities and 12 case without mitotic figure. Among all 66 cases of I-FISH group, PMI/RARα fusion gene was found in 64 cases (97.0%), suggesting that I-FISH group was more sensitive than CC group (p = 0.041). It is concluded that combination of I-FISH and CC techniques plays a pivotal role for diagnosis and detection of minimal residual disease in APL.

Analysis of DNMT3a gene mutations in acute myelogenous leukemia / 中国实验血液学杂志

This study was purposed to investigate the mutational status of DNA methyltransferase (DNMT3a) gene and the clinical features of AML patients with DNMT3a mutations. Using PCR combined with directly sequencing, the somatic mutations of DNMT3a involving residue of amino acid 882 were detected in 77 AML patients. Furthermore, the clinical features of these patients were also studied. The results showed that the DNMT3a mutation were detected in 7 out of 59 patients with de novo AML (11.9%), which included 4 patients with DNMT3a R882C, 2 patients with DNMT3a R882H and 1 patient with DNMT3a Y874C. Morphology examination indicated that 2 patients were M(2), 1 patient was M(4) and 4 patients were M(5). Cytogenetic analysis revealed that karyotype in 5 out of 7 patients with DNMT3a mutation were normal. In total of 27 patients with normal karyotype 5 patients (22.7%) were found harboring DNMT3a mutation, while no DNMT3a mutation was found in 21 patients with abnormal karyotype. The mutation rate in patients with positive CEBPA was obviously higher than that in patients with negative CEBPA (p = 0.002). Immunophenotype analysis showed that 4 patients (4/7, 57.1%) with DNMT3a mutation expressed lymphoid antigens including CD4 or/and CD7. There were no statistical significance in age, gender, blast cells of bone marrow, white blood cell and platelet counts, hemoglobin level, ratio of CR, mutations of FLT3-ITD, NPM1 and c-kit between patients with DNMT3a mutation and patients with wild DNMT3a (p > 0.05). It is concluded that the DNMT3a mutations are more prevalent in AML patients with normal karyotype accompanying with positive NPM1 and/or CEBPA mutation, the role of DNMT3a mutation in AML prognosis needs to be further studied.