23例微分化型急性髓系白血病临床分析

[目的]探讨微分化型急性髓系白血病（AML—M0）的临床和细胞核型特点，观察MAVP（米托蒽醌、阿糖胞苷、长春新碱、强的松）的疗效。[方法]形态学、直接双标免疫荧光法（FISH）、直接／间接R显带技术对23例确诊为AML—M0的患者进行形态学、免疫分型等临床和细胞染色体核型分析。23例患者均接受2个疗程以上MAVP方案的诱导缓解化疗。[结果]过氧化酶（POX）、苏丹黑（SB）染色阳性的原始细胞低于3％，免疫表型以MPO、CD13、CD33、CD，38、CD34抗原表达阳性率较高，染色体结构异常较为常见，复杂的染色体核型异常多见。正常细胞核型患者完全缓解率高于异常染色体核型患者。[结论]骨髓细胞形态学、细胞化学、免疫表型是诊断AML—M0的主要依据。染色体改变具有高度的异质性，核型异常患者预后不佳。MAVP是治疗AML—M0的有效化疗方案。     

Treatment of Acute Myeloid Leukemia with the FLT3 Gene Mutation

In acute myeloid leukemia (AML), mutations of the Fms-like tyrosine kinase 3 receptor (FLT3) and its overexpression are related with hyperleukocytosis, higher risk of relapse, and decrease of both disease-free survival and overall survival. It has been suggested that this phenomenon confers proliferative and survival advantages to the malignant blast cells. As a consequence, it is an attractive therapeutic target. As the best treatment strategy for mutated FLT3 AML remains to be defined, the addition of FLT3 inhibitor drugs to chemotherapy or to the bone marrow transplant approach has become a growing strategy. With encouraging results, this combination seems to be an attractive option. Relevant data regarding the current treatment trends on mutated FLT3 AML is reviewed here.     

NOTCH-1 Gene Mutations Influence Survival in Acute Myeloid Leukemia Patients

Background: Although NOTCH-1 gene mutations were reported to contributes to leukemogenesis in lymphocytic leukemias, its role in acute myeloid leukemia (AML) remains unclear. Therefor; this study was designed to determine the prevalence and clinical impact of NOTCH-1 mutations in AML patients. Materials and Methods: In the current study, NOTCH-1 gene mutations were identified in Bone Marrow samples obtained from fifty primary AML patients before start of therapy using Sanger sequencing. Results: NOTCH-1 gene mutations were detected in 6 out of 50 AML cases (12%). The three mutations were (two mutations C7318A in the Pest domain exon 34); (another 2 in the Pest domain Del 7,344, ins C7349, G7356A and the last ones in the HD-N exon-26 (Del A4609). The clinical findings in the mutant AML (mu AML) patients did not significantly different as compared to the un mutated (unmut) AML patients.  There is significant association between CD7 aberrant expression and NOTCH-1 mutations. The complete remission was significantly higher in unmut AML cases as compared to mut AML ones (P=0.024). Multivariate (Age; Gender; Bone Marrow Blast cells; NOTCH-1 mutations) Cox regression analysis revealed that NOTCH-1 mutation is an independent risk factor for AML overall survival (P<0.001). The OS in unmut AML group (21.2 months) was significantly longer as compared to mut AML one (1.2 months) (P<0.001). Conclusion: Our data indicate that NOTCH-1 gene mutations were detected in 12% of AML patients. These mutations displayed bad clinical outcome on AML patients. Therapeutic targeting of NOTCH-1 could be a potentially effective approach to combat master oncogenic drivers in AML.     

Detection of TET2 Mutation in Patients with De Novo Acute Myeloid Leukemia: A Mutation Analysis of 51 Iranian Patients

Acute myeloid leukemia (AML) is a heterogeneous clonal disease that is considered to originate from hematopoietic stem cells, which are characterized by impaired myelopoiesis and blast proliferation. TET oncogene family member 2 (TET2) mutations are frequent in myeloid malignancies and several studies have assessed the clinical importance of TET2 mutations. However, its frequency ratio has not yet been fully clarified. Method: Hence, our study was aimed to analyze TET2mut in patients with de-novo AML and their association with clinical, molecular characteristics and Nucleophosmin 1 (NPM1), Fms-like tyrosine kinase 3 (FLT3), CCAAT Enhancer Binding Protein Alpha (CEBPA) and Wilms’ tumor protein (WT1) gene expression. Fifty-one Iranian patients were screened by polymerase chain reaction (PCR) and direct sequencing to evaluate TET2 mutations frequency. Results: Out of all patients, 10 mutations in 8 patients (15.6%) were detected and closely associated with higher age and higher hemoglobin levels (p-value <0.05). Although FLT3, NPM1 and CEBPA gene expression did not show any significant correlation with TET2mut, cytogenetically normal acute myeloid leukemia (CN-AML)  patients appear to bear TET2mut more frequently with lower platelet counts. Monocyte-lineages leukemia has seemed to be more linked with TET2mut in these patients. Conclusion: Our study suggests the frequency of TET2mut in our study (15.6%) is in line with previous studies and reveals the critical role of TET2 in myeloid transformation, especially in leukemia with monocytic subtypes.     

Lack of Altered BECN1 Gene Expression in Iranian Patients with Acute Myeloid Leukemia

Acute myeloid leukemia (AML), one of the most prevalent leukemia types in adults, demonstrates great heterogeneity in molecular and clinical terms. Hence, there is a necessity to the mechanisms involved in AML generation in order to determine optimal treatment. This cross sectional study aimed to assess changes in BECN1 gene expression in with blood samples from 30 AML patients, compared with samples from 15 healthy persons. RNA was extracted and cDNA was synthesized and Real Time PCR applied to determine BECN1 gene expression. The results showed no significant differences in BECN1 gene expression between patients with AML and normal controls (P > 0.05). It appears that expression of BECN1 does not play a significant role in genesis of AML leukemia.     

SALL4 Gene Expression in Acute Myeloid Leukemia

Objectives: SALL4 gene was aberrantly expressed in many leukemia cell lines and primary leukemia cells of acute myeloid leukemia (AML) and precursor B-cell lymphoblastic leukemia/lymphomas. Its expression may be a useful marker to predict the diagnosis and the risk stratification of patients with AML. Methods: This study aimed to characterize the expression pattern of SALL4 gene in adult patients with acute myeloid leukemia. Quantitative Real-time PCR was used to determine the expression level of the gene in peripheral blood of 52 Egyptian adult AML patients and 10 healthy control cases. Our study was done in the National Cancer Institute during the period of time between December 2014 and June 2015. Results: The observed data revealed that none of the studied controls expressed SALL4 > 1.0 RQ and there was a highly statistically significant difference between cases and controls regarding SALL4 gene expression where all cases showed higher expression of SALL4 than controls with p value SALL4 is one of few genes that bridge the self-renewal properties of ESCs, normal HSCs and LSCs. Their expression is easily determined by real time PCR. They may be useful markers to predict prognosis and help to stratify patients into risk adapted groups. Further studies including increasing patient numbers are essential to understand the relations between SALL4 gene expression and its prognostic impact.     

Association of SDF-1 Gene Polymorphism with Increased Risk of Acute Myeloid Leukemia Patients

Background: Acute myeloid leukemia (AML) is a heterogenous group of disorders that emerge from the malignant transformation of hematopoietic stem cells. Chemokine stromal cell-derived factor 1(SDF-1) and its receptor CXC receptor 4 (CXCR4) has an essential role in dissemination of blast cells. Study aimed to detect CXCR4 expression and the SDF-1 (rs1801157) gene polymorphisms and correlate them with prognosis and outcome in AML patients. Subjects and Methods: The study was conducted on 60 de-novo AML patients, and 60 healthy controls. SDF-1 (rs1801157) gene polymorphisms were detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), and CXCR4 expression was done using flow cytometry analysis. Results: SDF-1 dominant model (AG+AA) had higher risk AML (p 0.002). CXCR4positive cases were associated significantly with toxic manifestations (p 0.019), lower CR rates (p 0.004), and unfavorable cytogenetics (p 0.027). Multivariate analysis showed that combined CXCR4positive with dominant SDF-1 considered as independent prognostic factor for shorter overall survival (OS) in AML patients (p 0.031). Conclusion: SDF-1 dominant model had a higher risk to develop AML, and CXCR4 positive expression predicts poor prognosis in AML patients and it could represent a targeted therapy in AML. In addition, CXCR4 could be easily integrated into the initial routine diagnostic work up of AML.     

Monoclonal Antibody Anti-MPO is Useful in Recognizing Minimally Differentiated Acute Myeloid Leukaemia

The enzyme myeloperoxidase (MPO) is the most specific marker of myeloid lineage. The recognition of acute myeloid leukaemia (AML) with minimally differentiation (AML-MO) is established with methods that include myeloid markers CD13/CD33 and detection of MPO in blast cells by immunological techniques or electron microscopy cytochemistry (EM). We have analysed the presence of MPO in leukaemic blast cells by conventional cytochemistry and immunological methods using a monoclonal antibody anti-MPO (CLB-MPO1) in 121 cases of acute leukaemia. The aim of the study was to investigate the sensitivity of this McAb to identify AML-MO, as CD13/CD33 can be expressed in some cases of acute lymphoblastic leukaemia (ALL) and EM cytochemistry is not always available in many laboratories. Anti-MPO was positive in all cases of AML (M1-M5) which were positive by Sudan Black B reaction in similar or higher percentage ratio for each case, although in some of them did not label with CD13/CD33 tested by IF and IPc techniques. Based on the anti-MPO positivity, 5 out of 10 cases called undifferentiated leukaemia (AUL) were reclassified as AML-MO, though 4 cases were CD13/CD33 negative. Furthermore, after analysing the anti-MPO expression among 32 cases of ALL, we had to reclassify four of them as acute biphenotypic leukaemia. We conclude that anti-MPO is a very sensitive and reliable tool in AML diagnosis and has an important role in distinguishing minimally differentiated AML and biphenotypic acute leukaemia from AUL and ALL.     

Treatment Strategies in Patients with Core-Binding Factor Acute Myeloid Leukemia

Core-binding factor acute myeloid leukemias (CBF AML) are characterized by sensitivity to high-dose cytarabine. Due to good prognosis in CBF AML patients, it is important to determine the optimal treatment. Long-term RFS (relapse-free survival) is reported among 40–60%. Experience with FA/FLAG vs. IA/IAG as front-line chemotherapy has been reported by some authors. Other studies, regarding treatment strategies such as high-dose daunorubicin, do not determine survival curves in this precise subgroup of patients. Preliminary data with gemtuzumab ozogamicin plus FLAG has been reported. There are not studies with FLAG using oral fludarabine in acute leukemia patients.     

Cytogenetic and Genetic Mutation Features of de novo Acute Myeloid Leukemia in Elderly Chinese Patients

Objectives: The present study aimed to examine the cytogenetic and genetic mutation features of acutemyeloid leukemia (AML) in elderly Chinese patients. Methods: A retrospective analysis of cytogenetics andgenetic mutations was performed in 113 cases (age range 50-82 years) with de novo AML. Results: The mostfrequent cytogenetic abnormality was t (15;17) (q22;q21), detected in 10.0% (n = 9) of successfully analyzedcases, followed by t (8;21) (q22;q22) in 8.89% (n = 8), and complex karyotypes in 5.56% (n = 5). Those withcomplex karyotypes included 4 cases (4.44%) of monosomal karyotypes. The frequencies of NPM1, FLT3-ITD,c-kit, and CEBPA mutations were 27.4% (31/113), 14.5% (16/110), 5.88% (6/102), and 23.3% (7/30), respectively.The complete remission rates of patients in low, intermediate, and high risk groups were 37.5%, 48.6%, and33.3%, respectively (χ2 = 0.704, P = 0.703) based on risk stratification. Conclusion: Cytogenetics and geneticmutations alone may not be sufficient to evaluate the prognoses of elderly AML patients. The search for a novelmodel that would enable a more comprehensive evaluation of this population is therefore imperative.     

Chemotherapeutic Resistance in Egyptian Acute Myeloid Leukemia Patients

Background: Acute Myeloid Leukemia (AML) is a heterogeneous disorder with variable genetic abnormalities andcytogenetic alterations which provide a significant disease prognosis and determine response to therapy. Purpose: Weaim to investigate the expression of the MDR1 gene in 100 Egyptian AML patients, to identify their role on both theprogression and chemotherapeutic refractoriness together with assessment of known prognostic molecular markers;FLT3-ITD and NPM1 mutations. Methodology: Quantitative assessment of MDR1 gene expression was performedby quantitative RT-PCR. Additional prognostic molecular markers were determined as internal tandem duplications ofthe FLT 3 gene and nucleophosmin gene mutation A. Results: MDR1 gene expression levels and FLT3/ITD mutationswere significantly higher in AML patients with resistant disease with P value NPM1 was insignificantly higher in patients with CR P-value 0.14. In MDR positive group, wild FLT3/ITD with orwithout NPM1 mutation was favorable in achieving CR with p value 0.02. MDR negative group, wild FLT3/ITD withor without NPM1 mutation showed insignificantly higher CR rates with p value (0.35). Kaplan-Meier curves revealedstatistically significant difference between MDR1-negative and MDR1-positive patients regarding their DFS and OSbetween the two groups where DFS and OS were higher in MDR1-negative patients with p value 0.004 and 0.01,respectively. Conclusion: The results obtained by the current work together with the previous researches concerningthe study of multidrug resistance genes in AML patients provide additional evidence of the role played by these genesas predictors of chemoresistance and poor treatment outcome.     

Treatment of Elderly Acute Myeloid Leukemia Patients

Older patients with acute myelogenous leukemia (AML) fare much less well than younger patients with the same disease due to a combination of comorbidities and intrinsic disease resistance. Likely due to aging of the US population, the median age of AML patients at diagnosis has increased from 68 to 72 years. AML is a heterogeneous disease, particularly in older patients, making therapeutic decisions challenging. Older patients who are ‘fit’ for intensive chemotherapy and would have a reasonable chance to benefit based on host and disease characteristics should receive standard induction chemotherapy with 7 days of continuous infusion of cytarabine and at least 60 mg/m² daunorubicin daily for 3 days. Therapeutic options for patients who are not candidates for or are not likely to respond to intensive therapy include clofarabine, low intensity chemotherapy such as low dose cytarabine, hypomethylating agents, or investigational agents. For older AML patients in complete remission, post-remission or consolidation chemotherapy with repeat induction or modified high dose cytarabine may offer a small chance for long term disease-free survival. Selected older patients who achieve remission by any means should be considered for reduced-intensity stem cell transplantation which may offer improved chances of cure and survival compared with standard post-remission chemotherapy.     

Defining Acute Myeloid Leukemia Ontogeny in Older Patients

BackgroundAcute myeloid leukemia (AML) in elderly patients is associated with poor outcomes and often arises from antecedent hematologic disorders (AHD), classified as secondary AML (sAML).Patients and MethodsTo validate the use of somatic mutations to determine AML ontogeny in the elderly population, we identified 178 elderly (> 70 years) patients with AML with NexGen Sequencing data. Patients were divided clinically into primary AML (pAML) or sAML based on prior history of AHD. Patients were then reclassified into 4 groups based on somatic mutations and cytogenetics as suggested by Lindsley et al: group 1 (pAML) with CBF rearrangements, 11q23/MLL, and NPM1 mutation (MT); group 2 (sAML) with SRSF2, SF3B1, U2AF1, ZRSR2, ASXL1, EZH2, BCOR, or STAG2 MT; group 3 with TP53 MT; and group 4 as not otherwise specified (NOS).ResultsBased on clinical criteria, 95 patients were classified as pAML and 82 patients as sAML. Based on the AML ontogeny proposed, 8 patients were classified as pAML, 72 patients as sAML, 28 patients had TP53 MT, and 70 patients were classified as NOS. The median overall survival was 22.4,14, 2.8, and 11.2 months, respectively. Clinical versus molecular classification was discordant where 25% (n = 2) of patients classified as pAML by molecular signature had a history of AHD, whereas 44% (n = 32) of patients classified molecularly as sAML had no prior AHD. In the TP53 MT and NOS categories, 37% (n = 28) and 43% (n = 70) of patients had AHD, respectively.ConclusionOur data shows that molecular annotation of elderly patients with AML reclassifies a significant proportion of patients as sAML, which may have therapeutic implications.     

Clonal Analysis of Multiple Point Mutations in the N-ras Gene in Patients with Acute Myeloid Leukemia

We have screened mutations of the N- ras gene at codons 12, 13, and 61 in leukemia cells obtained from 100 patients with acute myeloid leukemia (AMD, and found mutated N-ras alleles in 9 patients. We further analyzed the polyclonality of multiple N- ras gene mutations in 4 AML patients. One patient, who had the monoclonal karyotype, t(11;17), had two types of double missense mutations at codons 13 and 61 in the same allele. Each of the remaining three patients, one of whom had t(15;17) with a monoclonal rearrangement of the retinoic acid receptor alpha and PML genes, carried two missense mutations in a relatively small population of leukemia cells. We have demonstrated that multiple clonality of the N- ras gene is occasionally observed in leukemia with a monoclonal karyotype. These findings indicate that the N-ras mutations may not always be characterized simply by an accumulative process and that the activated N- ras gene alone is not sufficient to cause leukemia.     

Prognostic Value of a CYP2B6 Gene Polymorphism in Patients with Acute Myeloid Leukemia

Background: The objectives of this study aimed to detect a CYP2B6 polymorphism in de novo cases of acutemyeloid leukemia patients and identify any role in disease progression and outcome. Materials and Methods:DNA was isolated from peripheral blood of 82 newly diagnosed acute myeloid leukemia cases and the CYP2B6G15631T gene polymorphism was assayed by PCR restriction fragment length polymorphism (PCR-RFLP).Results: The frequency of the GG genotype (wild type) was 48 (58.5%) and that of the mutant type T allele was 34(41.9%). GT genotype heterozygous variants were found in 28 (34%), and TT genotype homozygous variants in6 (7.3%) cases. We found no significant association between the CYP2B6 G15631T polymorphism and completeresponse (CR) (p-value=0.768), FAB classification (p-value=0.51), cytogenetic analysis (p-value=0.673), and overallsurvival (p-value=0.325). Also, there were no significant links with early toxic death (p-value=0.92) or progressionfreesurvival (PFS) (p-value=0.245). Conclusions: Our results suggest that the CYP2B6 polymorphism has norole in disease progression, therapeutic outcome, patient free survival, early toxic death and overall survival inacute myeloid leukemia patients.     

Risk Effects of GST Gene Polymorphisms in Patients with Acute Myeloid Leukemia: A Prospective Study

Glutathione S-transferase (GST) enzyme levels are associated with risk of many cancers, including hematologictumours. We here aimed to investigate the relationships between GSTM1, GSTT1 and GSTP1 polymorphisms andthe risk of AML. Genotyping of GSTs was based upon duplex polymerase-chain-reactions with the confrontingtwo-pair primer (PCR-CTPP) method in 163 cases and 204 controls. Individuals carrying null GSTT1 genotypehad a 1.64 fold risk of acute leukemia relative to a non-null genotype (P<0.05). A heavy risk was observed inthose carrying combination of null genotypes of GSTM1 and GSTT1 and GSTP1 Val allele genotypes whencompared with those carrying wild genotypes, with an OR (95% CI) of 3.39 (1.26-9.26) (P<0.05). These findingsindicate that genetic variants of GST and especially the GSTT1 gene have a critical function in the developmentof AML. Our study offers important insights into the molecular etiology of AML.     

急性髓系白血病患者HOX11基因的表达及意义

目的 探讨HOX11基因在急性髓系白血病(acute myeloid leukemia,AML)中的表达及对预后的影响,为个体化治疗提供依据.方法 应用多重巢式RT-PCR方法对73例初诊AML患者的融合基因进行检测,对HOX11基因表达阳性和阴性的患者进行标准治疗后的疗效进行分析.结果 在预后良好组、预后中等组及预后不良组AML患者的标准治疗中,HOX11基因阳性表达患者的第一疗程完全缓解率(complete remission rate)并不高于阴性表达的患者(P＞0.05),而复发或死亡率(relapse or mortality rate)与HOX11基因阴性表达的患者比较差异有统计学意义(P＜0.05).结论 HOX11基因的表达可能影响AML患者的预后.