[Show abstract][Hide abstract] ABSTRACT: Severe congenital neutropenia (SCN) could exhibit different modes of inheritance from a genetic perspective, including autosomal dominant, autosomal recessive, X-linked forms and a sporadic one without any hints regarding family history. Investigations in molecular etiology have revealed some specific genes mutation that could account for the different types of SCN respectively. Homozygous HAX1 mutations had been identified in autosomal recessive SCN (Kostmann disease) previously. However, compound heterozygous HAX1 mutations were also confirmed in autosomal recessive SCN by subsequent research. Here we described another patient, also the first Chinese patient, with autosomal recessive SCN carrying a novel compound heterozygous HAX1 mutation consisting of two frame-shift mutations (c. 430-1insG, p.Val144GlyfsX5; c. 655-9del5bp, p. Pro219TrpfsX13) leading to premature stop cordons, who had progressed to chronic myelomonocytic leukemia (CMML) without neurodevelopmental abnormalities. Although genotype-phenotype associations in SCN patients with homozygous HAX1 mutations have been documented previously, by comparing this patient with the other four patients with compound heterozygous HAX1 mutations reported previously, we found that genotype-phenotype associations were much more complicated.
[Show abstract][Hide abstract] ABSTRACT: To assess the prevalence of several tyrosine kinases (TKs) gene mutations including c-Kit, FLT3 and JAK2 V617F in core binding factor related acute myeloid leukemia (CBF-AML), and analyze their impact on clinical characteristics and prognosis.
Mutations of c-Kit, FLT3-ITD and FLT3-TKD were detected by genomic DNA PCR and sequencing, and JAK2 V617F mutation screening by allele-specific PCR in 58 newly diagnosed CBF-AML patients [28 AML with inv(16) and 30 with t(8;21)], and analyze the patients clinical characteristics and prognoses.
c-Kit aberrations were detected in 32.8% cases, including 6 cases mutated in exon 8 (mutKIT8) and 13 mutated in exon 17 (mutKIT17). MutKIT8 was more prominent in inv(16) than in t(8;21) patients (21.4% vs 0, P = 0.009). Only 2 cases had FLT3-ITD and 7 (12.1%) FLT3-TKD mutations. The result of JAK2 V617F mutation screenings in these CBF-AML patients was negative. The frequency of receptor tyrosine kinases(RTK) mutations was 46.6% and only one case had two kinds of missense mutations (mutKIT8 & TKD(+)). Median age of onset was higher for mutKIT17 than for wide-type c-Kit (wtKIT) patients (55 vs 31, P = 0.003). c-Kit mutations were significantly associated with decreased overall survival (OS) and continuous complete remission (CCR) rates (P = 0.053, and 0.048 respectively), and so did more for exon17 mutated patients reduced (P = 0.005, and 0.013 respectively). FLT3-TKD mutation showed no effects on prognosis of CBF-AML patients.
RTK mutations are common in patients with CBF-AML. c-Kit mutations frequently and JAK2V617F mutation rarely appear in CBF-AML. c-Kit mutations, especially mutKIT17 confers higher relapse risk and poorer prognosis.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 10/2011; 32(10):679-83.
[Show abstract][Hide abstract] ABSTRACT: To evaluate the prevalence of TET2 gene mutation in acute myeloid leukemia (AML) patients, and analyze their clinical characteristics and prognosis.
Polymerase chain reaction (PCR) and direct sequencing were used to sequence exon 3 to 11 of TET2 gene.
Among 96 AML patients, TET2 gene mutation was detected in 13 (13.54%) patients (95%CI 6.70% - 20.38%). The median age was 54 years in mutated group and 41 years in unmutated group (P = 0.010). Mutated and unmutated patients did not significantly differ in gender, white blood cells (WBC) count at diagnosis, platelet count, PB and BM blast percentage and chromosome karyotype, excepting for hemoglobin level 84 (70 - 108) g/L in mutated group versus 70 (55 - 87) g/L in unmutated group (P = 0.032). TET2 gene mutation had no significant correlation with C-KIT, FLT3, JAK2V617F mutations, but did with NPM1 mutation. TET2 mutated patients had lower CR1 rate and 2-year overall survival than unmutated in non-M(3) patients (P < 0.05).
TET2 gene mutation is more prevalent in older AML patients and has a certain correlation with clinical characteristics and outcome. It may be a molecular marker for poor prognosis in AML.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 05/2011; 32(5):304-7.
[Show abstract][Hide abstract] ABSTRACT: To investigate the clinical and biological characteristics and prognosis of mixed phenotype acute leukemia (MPAL).
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).
(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.
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.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 01/2011; 32(1):12-6.
[Show abstract][Hide abstract] ABSTRACT: To analyze the clinical and molecular cytogenetic features of hematologic malignancies with idic(20q-).
The clinical data of 10 patients with idic (20q-) were analyzed. Karyotyping analysis was carried out with R banding technique. A CEP20 probe was used to perform single-color fluorescence in situ hybridization (FISH). A subtelomeric probe for 20q and a locus-specific probe for 20q12 were used to perform dual-color FISH. The literatures of hematologic malignancies with idic(20q-) were reviewed.
Of the 10 cases, 2 were diagnosed as acute erythroid leukemia, 1 primary myelofibrosis, 3 myelodysplastic syndromes (MDS) and 4 highly suspected (HS-MDS). Karyotype analysis showed that one of the normal chromosome 20 allele was substituted by one or two metacentric isochromosomes smaller than the normal one in all 10 cases. It was confirmed to be der(20)del(20)(q11q13)idic(20)(p11), i.e., idic(20q-) by FISH assay. Partial cells in 2 of the 10 cases had 20q- as the sole karyotypic anomaly.
Idic(20q-) results from a pre-existing del(20q) and is strongly associated with MDS and acute erythroid leukemia. Idic(20q-) as a recurrent cytogenetic abnormality is helpful for diagnosing HS-MDS in patients with cytopenia but only slight or absent dysplasia.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 01/2011; 32(1):17-20.
[Show abstract][Hide abstract] ABSTRACT: To compare the signal patterns of dual color extra-signal BCR/ABL probe (ES-FISH) and dual color dual fusion BCR/ABL probe (D-FISH) in the fluorescence in situ hybridization (FISH) detection of Ph-positive leukemia, and to explore their diagnostic value.
ES-FISH probe and D-FISH probe were used, respectively, to detect the BCR/ABL fusion gene in 74 cases with typical t(9;22)(q34;q11) and 37 cases with variant t(9;22)(q34;q11) translocation or complex karyotypic abnormalities containing Ph translocation.
The BCR/ABL fusion gene in all cases with typical t(9;22)(q34;q11) could be detected by both FISH probes. D-FISH had a signal pattern of 1O1G2F, while ES-FISH showed a signal pattern of 2O1G1F. ES-FISH enables the minor breakpoint cluster region to be identified in 9 cases (12.2% ) of Ph-positive leukemia, whereas D-FISH could not differentiate the minor breakpoint cluster region from major breakpoint cluster region. D-FISH could distinguish simple ABL gene deletion from simultaneous deletion of the ABL and BCR genes in 8 cases (10.8%) of Ph-positive leukemia patients, but ES-FISH could not. For variant Ph translocation or complex karyotypic abnormalities containing Ph translocation, each FISH probe showed four or six types of signal pattern, most of which were atypical. The exact interpretation was dependent on conventional karyotypic analysis and FISH on metaphases.
ES-FISH and D-FISH probes displayed different signal patterns in Ph-positive leukemia due to their differences in size and covered regions. ES-FISH and D-FISH probes may be selected as better probe for Ph-positive acute lymphocytic leukemia and Ph-positive chronic myeloid leukemia, respectively. When imatinib was used for treatment, there was no preference between ES-FISH and D-FISH probe, because major breakpoint cluster region, minor breakpoint cluster region and partial sequence deletion of derivative chromosome 9, would not affect the prognosis of Ph-positive leukemia. However, considering that ES-FISH probe has a better cost-performance than D-FISH probe does, it is recommended as first choice.
Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics 04/2010; 27(2):166-70. DOI:10.3760/cma.j.issn.1003-9406.2010.02.011
[Show abstract][Hide abstract] ABSTRACT: To determine the pulmonary pathological changes in hematological malignancy patients with pulmonary complications.
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.
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.
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.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 12/2009; 30(12):829-33.
[Show abstract][Hide abstract] ABSTRACT: To establish and characterize a novel human myeloid leukemia cell line SH-2.
Bone marrow mononuclear cells (BMMNC) isolated from a AML-M2 patient, who failed to obtain complete remission after chemotherapy and allogenic bone marrow transplantation were passed in a long term IMDM culture medium supplemented with 20% fetal calf serum. Stromal cells were retained and rh-IL-3 was added in the culture system. A new human myeloid leukemia cell line SH-2 was successfully established with a cytogenetic characteristics of a loss of Y chromosome (-Y), a derivative chromosome 16 resulting from unbalanced translocation between chromosome 16 and 17, monosome 17, trisomy 19 and p53 alteration. Various methods were employed to characterize SH-2 cell line.
SH-2 cells has been maintained without cytokine and stromal cells for more than 3 years without EB virus and mycoplasma contamination. SH-2 cells had the basically same morphological, immunophenotypic and cytogenetic features as the patient's leukemia cells did, such as myeloid morphology, an immunophenotype of CD13+, CD33+, CD56+, CD16/56+ and a hypodiploid karyotype of 45, X, -Y, der(16)t(16;17)(q24;ql2), -17, +19, which were gradually decreased and replaced by the near-tetraploid cells with a karyotype of 73-102(80), XX, -Y, -Y, del (q131)x2, der(16)t(16;17)(q24;q12)x2, -17, -17, +19, +19. FISH and multiple FISH delineated all the abnormalities and revealed a loss of one p53 allele due to monosomy 17. DNA direct sequencing detected a point mutation of CAG to CAT at codon 576 of exon 5 in another p53 allele. RT-PCR showed that SH-2 cells expressed apoptosis-related genes (bcl-2, Fas, GST-pi and p2) rather than MDR-related genes. Short tandem repeat PCR provided powerful evidence for the derivation of SH-2 cell line from the patient's leukemia cells. SH-2 cells had certain colony formation and tumorigenic capacities in nude and SCID mice.
SH-2 is a new myeloid leukemia cell line with a unique biology background, and will provide a useful tool for leukemia research.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 07/2009; 30(7):458-63.
[Show abstract][Hide abstract] ABSTRACT: To explore the efficacy and toxicity of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for relapsed/refractory acute lymphocytic leukemia (ALL).
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).
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.
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.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 03/2009; 30(2):73-6.
[Show abstract][Hide abstract] ABSTRACT: To evaluate the prevalence of Fms-Like tyrosine kinase 3 (FLT3) mutations including internal tandem duplication (ITD) of juxtamembrane region and point mutation in the second tyrosine kinase domain (TKD) in acute promyelocytic leukemia (APL) and its clinical significance.
Bone marrow mononuclear cells from 160 newly diagnosed APL patients were analyzed. Polymerase chain reaction (PCR) was used to detect FLT3-ITD mutations, FLT3-ITD positive samples were further analyzed for the ITD allelic ratio (ITD-AR, mutant-wild type ratio). The FLT3-TKD mutation was analyzed by PCR amplification of exon 20 followed by EcoR V digestion and sequencing.
Out of 160 patients, 30 (18.75%) patients were FLT3-ITD positive, 17 (10.62%) were FLT3-TKD positive, 2 had both of mutations. The initial WBC count and the ratio of short type PML-RAR alpha isoforms in FLT3-ITD positive and FLT3-TKD positive patients were all higher than that in patients with wild-type FLT3 (FLT3-wt) (P < 0.05). For FLT3-ITD positive patients, the incidences of retinoic acid syndrome (RAS) and disseminated intravascular coagulation (DIC) were 41.7% and 65.4%, respectively, being higher than that of FLT3-wt patients, while their complete remission (CR) rate was lower (69.2% vs 90.3%, P < 0.05). For FLT3-TKD positive patients, the incidence of RAS, DIC and CR rate were not significantly different from that of FLT3-wt patients (P > 0.05). FLT3-ITD positive patients had a shorter overall survival (OS) (P < 0.05), but not disease-free survival (DFS) (P > 0.05) as compared with FLT3-wt patients. There was no significant difference in either OS or DFS between FLT3-TKD positive and FLT3-wt patients. The ITD-AR of 30 FLT3-ITD positive patients varied from 0.11 to 6.55 with a median of 1.0. The initial WBC count, incidence of RAS and DIC, CR rate were not significantly different between the patients with ITD-AR greater than 1.0 and lower than 1.0 (P > 0.05).
FLT3 mutations (FLT3-ITD or FLT3-TKD) are frequently identified in patients with newly diagnosed APL, both mutations are associated with higher initial WBC and short type PML-RAR alpha isoforms. FLT3-ITD mutation is more frequent than FLT3-TKD mutation, and predicts a poorer prognosis, whereas FLT3-TKD mutation does not show the same unfavorable prognostic effect on APL patients.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 12/2008; 29(11):757-61.
[Show abstract][Hide abstract] ABSTRACT: To compare the clinical outcomes between unrelated donor hematopoietic stem cell transplantation (URD-HSCT) and HLA-haploidentical (Hi)-HSCT.
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.
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.
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.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 02/2008; 29(2):83-6.
[Show abstract][Hide abstract] ABSTRACT: To evaluate the prevalence and impact of NPM1 gene mutations and FLT3 internal tandem duplication (ITD) mutations in acute myeloid leukemia (AML).
Mononuclear cells in bone marrow samples were collected from 86 adult patients with newly diagnosed AML. FLT3 and NPM1 genes were amplified with genomic DNA-PCR. NPM1 exon-12 mutations were evaluated with capillary electrophoresis and FLT3-ITD mutations were determined with agarose electrophoresis.
NPM1 gene mutations were identified in 29 of the 86 (33.7%) patients and FLT3-ITD in 15 of the 86 (17.4%) patients. In AML patients with normal karyotype, the frequencies of the two abnormalities were significantly higher than in those without (46.0% and 24.0%, P < 0.05). Both mutations were related to high peripheral white blood cell counts (P < 0.05). WBC counts in the six of the seven cases with NPM1+/FLT3-ITD+ AML were significantly higher (>50 x 10(9)/L). Compared with NPM1(-)AML patients, NPM1(+)AML patients were associated with lower expression of CD(34) (P < 0.001), higher complete remission (CR) rate after initial therapy (66.7% vs 53.3%, P > 0.05) and better overall survival (OS) (P > 0.05), but compared with FLT3-ITD(-)AML patients, FLT3-ITD+ AML patients were associated with lower CR rate (50.0% vs 58.8%; P > 0.05) and worse OS (P > 0.05). The CR rates were 66.7%, 62.5%, 50.0% and 42.9%, respectively, in NPM1+/FLT3-ITD(-), NPM1(-)/FLT3-ITD(-), NPM1+/FLT3-ITD+ and NPM1(-)/FLT3-ITD+AML groups (P > 0.05), but there was no significant differences in OS for four groups.
NPM1 and FLT3-ITD mutations are common in AML patients with normal karyotype and related with the clinical characteristics and prognosis of AML.
Zhonghua nei ke za zhi [Chinese journal of internal medicine] 11/2007; 46(11):907-10.
[Show abstract][Hide abstract] ABSTRACT: To explore the possible role of CD28/CTLA-4 co-stimulators in immune pathophysiology of acquired aplastic anemia(AA).
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.
(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.
(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.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 09/2007; 28(9):590-3.
[Show abstract][Hide abstract] ABSTRACT: This study was purposed to explore the tumorigenicity of a novel human monocytic leukemic cell line SHI-1 in nude mice and its mechinism. The tumorigenicity in mice was evaluated in sixteen nude mice subcutaneously injected with the SHI-1 cell line. The tumor specimen was studied by the conventional pathologic examination. The mononuclear cells (MNC) of the tumor was assayed by RHG banding, the transcription of MLL-AF6 fusion gene and the VEGF gene was detected by RT-PCR. Gelatin zymography method was used to study the expression of MMP-9 and MMP-2 in the supernatant of the SHI-1 cell line. Matrigel invasion assay was employed for the study of migration of the SHI-1 cell in vitro. The results showed that the tumor masses were found in all sixteen mude mice after subcutaneous injection of SHI-1 cells, the tumor mass was mainly composed of leukemia cells, the transcription of MLL-AF6 fusion gene and VEGF gene was proved by RT-PCR analysis, the expressions of MMP-2 and MMP-9 in the serum-free culture supernatant of the SHI-1 cell line were significantly higher than those in U937, K562, and NB4 cell lines. The SHI-1 cell line exhibited significantly higher in vitro invasiveness than other leukemia cell lines, the blocking antibody of MMP-2 could inhibit the migration of the SHI-1 cell line significantly. It is concluded that the SHI-1 cell line presents higher tumorigenicity in nude mice than other leukemia cell line and the mechanism is associated with p53 gene alteration, high transcription level of VEGF gene, high expression level of MMP, and significantly higher invasiveness.
Zhongguo shi yan xue ye xue za zhi / Zhongguo bing li sheng li xue hui = Journal of experimental hematology / Chinese Association of Pathophysiology 09/2007; 15(4):683-6.
[Show abstract][Hide abstract] ABSTRACT: To investigate the frequency and clinical implication of JAK2V617F mutation in Chinese patients with myeloproliferative disorders (MPD).
Genomic DNA from bone marrow or blood mononuclear cells of 137 cases of MPD was screened with allele-specific polymerase chain reaction (PCR) and JAK2V617F mutation was detected with gel electrophoresis. There were 57 cases with polycythemia vera (PV), 68 with essential thrombocythemia (ET), 12 with idiopathic myelofibrosis (IMF).
JAK2V617F mutation was detected in 42 (73.7%) of the 57 patients with PV, 40 (58.8%) of the 68 with ET and 8 (66.7%) of the 12 with IMF. Sequence analysis of PCR products from selected patients confirmed the coexistence of both mutant and wild-type alleles. A higher prevalence was observed in elderly patients with MPD (P < 0.05). Cytogenetic analysis was performed in 115 of the 137 patients. Among the 108 patients with normal karyotype, JAK2V617F mutation was detected in 74 patients (68.5%) as compared with 5 of the 7 patients with karyotypic abnormalities (71.4%).
JAK2V617F mutation occurs in a significant percentage of Chinese patients with the myeloproliferative disorders. There is a higher prevalence in elderly patients.
Zhonghua nei ke za zhi [Chinese journal of internal medicine] 05/2007; 46(4):271-3.
[Show abstract][Hide abstract] ABSTRACT: To evaluate the prevalence of nucleophosmin (NPM1) gene exon 12 mutations in adults with acute myeloid leukemia (AML) and its clinical characteristics.
Genomic DNAs from 101 AML adults were screened by PCR and sequencing or capillary electrophoresis (CE) for NPMI mutations.
NPM1 exon 12 mutations were present in 31.7% of the overall cohort, including 1/1 (100%) of M0, 9/17(52.9%) of M1 , 7/25 (28.0%) of M2, 0/23(0%) of M3, 2/7 (28.6%) of M4 and 13/25 (52.0% ) of M5. NPM1 gene mutations were more prevalent in patients with normal karyotype (27/59, 45.8%) compared with that in those with karyotypic abnormalities (5/42, 11.9% ) (P < 0.001). NPM1 mutant cases were significantly associated with old age (P < 0.05), high peripheral white cell count (P < 0.05) and low expression of CD34 (P < 0.05) and CD17 (P<0.05). Sequence analysis of these NPM1 mutant cases revealed 5 known mutations (type A, B, D, N(M), and P(M)) and 1 novel variant (named as type S).
NPM1 exon 12 mutations occur with a considerable percentage in AML patients with normal karyotype, M1/M5 subtype and older age, and are associated with higher peripheral white cell count and lower expression of CD34 and CD117.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 05/2007; 28(5):289-93.
[Show abstract][Hide abstract] ABSTRACT: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the most effective method to treat malignant hematological disease. However the human leukocyte antigen (HLA)-matched sibling donors can be found for only 25%-30% patients. The probability of finding an unrelated matched donor is 1/50000-1/100000, or even lower. If we choose haploid HLA-matched hematopoietic stem cell transplantation instead, we may be able to find donors for 90% patients. Our present study was to explore the feasibility of allo-HSCT by using haploid HLA-matched donor in treatment for malignant hematological disease.
Twenty-five patients with malignant hematological disease received allo-HSCT with HLA 2 or 3 antigen mismatched related donors. All patients were treated with intensive immunosuppression, granulocyte colony stimulating factor (G-CSF) mobilization, antithymocyte globulin (ATG) and combination of bone marrow and peripheral blood stem cell transplantation. The conditioning regimen was intensified and prolonged by using the combination of cyclosporine (Cs) A, MMF, ATG and anti-CD25 antibody for graft-versus-host disease (GVHD) prophylaxis.
All patients achieved sustained, full donortype engraftment. Acute GVHD occurred in 21 of 25 patients. Eight of them were grade I aGVHD, six grade II aGVHD, two grade III aGVHD and five grade IV aGVHD. The cumulative incidence of grade II-IV aGVHD was 48%,and grade III-IV aGVHD was 28.57%. Chronic GVHD was observed in 12 of 25 patients and none of them developed extensive cGVHD. Sixteen patients were alive and disease free, with 64.0+/- 2.98% 1 year disease-free survival rate. One year overall survival rate was 64.0+/-3.08%. Nine patients died, 1 from relapse and 8 from transplantation related mortality.
Haploid HLA-matched allo-HSCT is a relatively efficient method for the treatment of patients with malignant hematological disease, who have no related matched donors. Nevertheless, strict administration should be carried out since it's a high risk approach.
Ai zheng = Aizheng = Chinese journal of cancer 08/2006; 25(8):1019-22. · 2.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To establish a novel human monocytic leukemic cell line and characterize its biological features.
Mononuclear cells isolated from the bone marrow of an acute monocytic leukemia (AML-M(5b)) patient at relapse were inoculated in a liquid culture system. And the biologic features of the established cell line SHI-1 were characterized by morphology, cytochemical staining, flow cytometry, karyotypic analysis, polymerase chain reaction (PCR), reverse transcriptase PCR (RT-PCR), fluorescence in situ hybridization (FISH), tumorigenicity in nude mice, quantitative fluorescent polymerase chain reaction, broth medium culture, short tandem repeating sequences-PCR (STR-PCR), multiplex-FISH (M-FISH), and (3)H-thymidine incorporation assay.
A human acute monocytic leukemia cell line, SHI-1, was established and has proliferated continuously in vitro for over one year. The cell line presented typical morphology and immuno-profile of monocytic lineage with the original t(6;11)(q27;q23) and del(17)(p11) abnormalities. The MLL-AF6 fusion transcript was detected by RT-PCR. The rearrangement of MLL gene, deletion of p53 gene, and translocation between chromosomes 6 and 11 were revealed by FISH. A point mutation of ATC-->ACC at exon 6 of the p53 gene was found by sequencing of the PCR products. The clonality and the high tumorigenicity of the SHI-1 cell line were confirmed. Infections of EBV and mycoplasma were excluded. A derivative chromosome 7 resulting from a translocation between chromosomes 7 and 13, monosomy 18 and a minute derived from chromosome 8 in addition to t(6;11) and deletion(17)(p11) were detected by M-FISH in SHI-1 cells passaged to March 2003. Cell line authentication by STR-PCR confirmed the identity to the original leukemic cells of the patient. (3)H-thymidine incorporation assay showed that IL-4 and IL-15 had proliferative effects, while IFN-gamma, TNFalpha, IL-2, PDGF, and IL-7 had inhibitory effects on the cell line.
SHI-1 is a novel acute monocytic leukemia-derived cell line carrying t(6;11)(q27;q23) and p53 gene alteration and having high tumorigenicity in nude mice. It provides a new useful tool for leukemia research.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 02/2005; 26(2):94-9.
[Show abstract][Hide abstract] ABSTRACT: To explore the value of fluorescence in situ hybridization (FISH) in the detection of mixed lineage leukemia (MLL) rearrangement and to assess the incidence and prognostic significance of MLL gene rearrangement in AML-M4/M5.
Bone marrow chromosome preparation of 23 cases of acute myeloid leukemia(AML) consisting of 19 cases with M5 and 4 cases with M4 was made using direct method or short-term culture. Karyotypic analysis was carried out by R-banding technique. Dig labeled 11q23 probe which spans the breakpoint cluster region in MLL was used to detect the MLL rearrangement in the above 23 cases.
R-banding karyotyping analysis revealed 11q23 translocation in 7 cases, while FISH analysis detected MLL rearrangement in 12 cases including the above 7 cases.
Interphase FISH was more sensitive in detecting the MLL rearrangement in AML-M4/M5, compared with the conventional cytogenetic method. MLL rearrangement is highly related to AML-M4/M5; it is an indication of poor prognosis.
Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics 07/2004; 21(3):288-90.