The Impact of Induction Treatment Response on the Prognosis of Pediatric Core Binding Factor-Acute Myeloid Leukemia Patients / 中国实验血液学杂志

OBJECTIVE@#To explore the impact of induction treatment response on the prognosis of pediatric core binding factor-acute myeloid leukemia (CBF-AML).@*METHODS@#The result of induce reaction and survival data of 157 pediatric CBF-AML patients in our hospital from September 2008 to December 2018 were retrospectively analyzed．The survival rate of the patients with different degrees of morphological remission after induction chemotherapy was comparative analyzed.@*RESULTS@#Among the 157 children with CBF-AML, 113 (72.4%) patients achieved morphologic leukemia-free state (MLFS) after the first course of induction chemotherapy, 153 (98.1%) patients achieved MLFS after the second course of induction chemotherapy. The 5-year event-free survival (EFS) rate and 5-year overall survival (OS) rate of patients with non-remission (NR) status after the first course of induction of chemotherapy was significantly lower than the patients achieved MLFS and the patients achieved partial remission (PR). The 5-year EFS rate and 5-year OS rate of the patients with PR status after the second course of induction chemotherapy were lower than the patients achieved MLFS, but the difference was not statistically significant. Multivariable analyze showed that NR after the first course of induction chemotherapy and myeloid sarcoma were the independent risk factors affecting EFS of the patients. There were six patients with NR status after the first course of induction chemotherapy, in which all of them harbored t(8;21), three of them with sex chromosome deletion, two of them with myeloid sarcoma.@*CONCLUSION@#NR status after the first course of induction chemotherapy was the independent risk factor affecting EFS and OS of CBF-AML patients, it can be taken as an indicator for higher risk stratification. PR status after the first course of induction chemotherapy may not be used as a diagnostic criterion for primary drug resistance.

Clinical features and prognosis of core binding factor acute myeloid leukemia in children / 中国当代儿科杂志

OBJECTIVE@#To study the clinical features and prognosis of core binding factor acute myeloid leukemia (CBF-AML) in children.@*METHODS@#A retrospective analysis was performed from the chart review data of children who were newly diagnosed with CBF-AML in the Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Sciences, from August 2009 to November 2015. According to the type of fusion gene, the children were divided into CBFB-MYH11 and AML1-ETO groups. Clinical features and prognosis were analyzed and compared between the two groups.@*RESULTS@#A total of 91 children with CBF-AML were enrolled in this study, among whom there were 74 (81%) in the AML1-ETO group and 17 (19%) in the CBFB-MYH11 group. Additional chromosomal abnormalities were observed in 38 children (42%), and deletion of sex chromosome was the most common abnormality and was observed in 28 children (31%). After the first course of induction treatment, the complete remission rate was 97% (88/91), the recurrence rate was 29% (26/91), the 5-year event-free survival (EFS) rate was 65%±6%, and the 5-year overall survival (OS) rate was 75%±5%. There were no significant differences between the AML1-ETO and CBFB-MYH11 groups in 5-year EFS rate (62%±7% vs 77%±11%, P>0.05) or 5-year OS rate (72%±6% vs 88%±9%, P>0.05).@*CONCLUSIONS@#AML1-ETO is the main type of fusion gene in children with CBF-AML, and deletion of sex chromosome is the most common type of additional chromosomal abnormalities. Children with CBF-AML often have a good prognosis, and the children with AML1-ETO have a similar prognosis to those with CBFB-MYH11.

Characterization of mutational pattern of patients with core-binding factor acute myeloid leukemia / 中华医学遗传学杂志

OBJECTIVE@#To characterize the mutational profile of patients with core-binding factor acute myeloid leukemia (CBF-AML).@*METHODS@#A total of 81 acute myeloid leukemia patients were recruited, which included 36 cases of CBF-AML and 45 cases of cytogenetically normal acute myeloid leukemia (CN-AML) . Mutations of FLT3-ITD, FLT3-TKD, NPM1, c-KIT, NRAS, KRAS, TET2, IDH1/2, RUNX1, DNMT3A, GATA2, ASjXL1, TP53, PTPN11, JAK2V617F, SETBP1 and CEBPA genes were simultaneously detected by DNA-based PCR and Sanger sequencing.@*RESULTS@#Over all, mutations were detected in 68 patients (83.9%), with the most common ones including double CEBPA mutations (n=17), followed by NPM1 (n=15), c-KIT (n=11), NRAS (n=10), TET2 (n=9), FLT3-TKD (n=9), FLT3-ITD (n=8), IDH1 (n=7), RUNX1 (n=7), KRAS (n=7), DNMT3A (n=6), IDH2 (n=4), and GATA2 (n=4) mutations. AML1-ETO and CBFβ-MYH11 fusions were present in 21 and 15 patients, respectively. Coexistence of ≥2 mutations was more common in CN-AML comparing with CBF-AML. The mutation rate of NPM1, FLT3-ITD, DNMT3A, IDH1 and CEBPA double mutations were higher in patients with CN-AML. NRAS, c-KIT and KRAS mutations were identified more frequently in patients with CBF-AML (P<0.05). Based on the function, aberration of genes involved in DNA methylation, NPM1 proteins and transcription predominated in CN-AML, while tyrosine kinase receptor signaling and RAS pathways have predominated in CBF-AML.@*CONCLUSION@#The genomic landscape of CBF-AML patients has differed from that of CN-AML patients. Synergy of fusion genes with particular mutations may impact the clinical phenotype and prognosis of patients.

Effect of SARI Overexpression on Apoptosis of CBFL Cells and Its Mechanism / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of SARI overexpression on the proliferation and apoptosis of core binding factor leukemia (CBFL) cells and explore the potential molecular mechanisms.@*METHODS@#C-KIT N822K mutation status in Kasumi-1 cell line was detected by exon 17 sequencing. Then the SARI lentiviral vector (pGC-FU-SARI) was constructed, meanwhile Kasumi-1 cells were transfected with the SARI lentiviral vector. Quantitative PCR and Western blot were employed to identify efficacy of SARI overexpression after the transfection of cells. Cells were divided into three groups, including the cells infected with pGC-FU-SARI (OE group), the cells infected with pGC-FU-GFP (NC group) and the untreated cells (blank control group). Cell proliferative activity was tested by MTT assay, cell apoptosis was measured by flow cytometry (FCM) and the expression of apoptosis-related proteins: BCL-2,BAX,Cyto C,Caspase 9,Caspase 3,cleaved-Caspase 3,PARP and cleaved-PARP as well as PI3K/Akt pathway proteins: PI3K（p85）,p-PI3K（p85）,Akt and p-Akt were detected by Western blot.@*RESULTS@#The Kasumi-1 cells were detected to bear c-KIT N822K (T>A) mutation. The Kasumi-1 cells with SARI was overexpression were construeted successfully. Compared with NC group, the cell proliferation was decreased and cell apoptosis was increased; BCL-2 expression was reduced, BAX expression was enharued; cyto C expression appeared; the expression of Caspase 9 and Caspase 3 was down-regulated, the expression of cleaved Caspase 3 was up-regulated; the PARP expression was decreased, cleaved PARP expression was increased; the phosphorylation level of PI3K/Akt pathway proteins: p-PI3K/PI3K, p-Akt/Akt was down-regulated in OE group (P<0.05).@*CONCLUSION@#SARI gene may suppress the proliferation of CBFL cells, and induce their apoptosis through the mitochondrial pathway, which may be related with the inhibition of PI3K/Akt pathway.

Comparative Analysis of C-repeat Binding Factors (CBFs) in Tomato and Arabidopsis

Abstract Cold stress is one of the limiting factors of plant production that plants use different mechanisms for cold tolerance. CBF genes play critical role to regulate the cold responsive genes. To better understand of CBF gene functions, the tomato-CBFs and Arabidopsis-CBFs were evaluated using bioinformatics tools, and finally the expression patterns of SlCBF1 gene were analyzed under 10 and 4˚C in two contrasting tomato species (Solanum lycopersicum and S. habrochaites). The different cis regulatory elements were observed in promoter region of SlCBF1 and AtCBF1 genes, and ICE1, COR and HOS1 proteins exhibited high interaction with CBFs. The results of Real time PCR of SlCBF1 exhibited that under 10 and 4 ˚C, SlCBF1 was down regulated in cold sensitive tomato genotype while it was slightly up-regulated in cold tolerant genotype at 4 ˚C. The results showed that the SlCBF1 and AtCBF1 genes have differential expression in cold stress.

Derivation and Differentiation of Bone Marrow Mesenchymal Stem Cells from Osteoarthritis Patients

Osteoarthritis (OA) of the knee is a degenerative joint disease caused by the progressive reduction of the articular cartilage surface that leads to reduced joint function. Cartilage degeneration occurs through gradual loss in extracellular matrix components including type II collagen and proteoglycan. Due to limited inherent self repair capacity of the cartilage, the use of cell-based therapies for articular cartilage regeneration is considered promising. Bone marrow mesenchymal stem cells (BM-MSCs) are multipotent cells and are highly capable of multilineage differentiation which render them valuable for regenerative medicine. In this study, BM-MSCs were isolated from OA patients and were characterized for MSC specific CD surface marker antigens using flowcytometry and their differentiation potential into adipocytes, osteocytes and chondrocytes were evaluated using histological and gene expression studies. BM-MSCs isolated from OA patients showed short spindle shaped morphology in culture and expressed positive MSC related CD markers. They also demonstrated positive staining with oil red O, alizarin red and alcian blue following differentiation into adipocytes, osteocytes and chondrocytes, respectively. In addition, chodrogenic related genes such as collagen type II alpha1, cartilage oligomeric matrix protein, fibromodulin, and SOX9 as well as osteocytic related genes such as alkaline phosphatase, core-binding factor alpha 1, osteopontin and RUNX2 runt-related transcription factor 2 were upregulated following chondrogenic and osteogenic differentiation respectively. We have successfully isolated and characterized BM-MSCs from OA patients. Although BM-MSCs has been widely studied and their potential in regenerative medicine is reported, the present study is the first report in our series of experiments on the BMSCs isolated from OA patients at King Abdulaziz University Hospital, Jeddah, Saudi Arabia.

Manifestation and treatment in a cleidocranial dysplasia patient with a RUNX2 (T420I) mutation / 대한악안면성형재건외과학회지

Cleidocranial dysplasia is an autosomal dominant heritable skeletal disorder. The characteristic features of cleidocranial dysplasia (CCD) may include hypoplasia of the clavicle, delayed closure of frontanelles, late tooth eruption, and other skeletal disorders. This case report describes clinical and radiographic manifestations at the age of 11 and 29 of a CCD patient, investigates the mutation of core-binding factor A1 (CBFA1) based on gene analysis, and illustrates successful oral reconstruction with fixed prosthesis and dental implant after the extraction of multiple teeth.

Diagnosis and Treatment of Pediatric Acute Myeloid Leukemia / 임상소아혈액종양

Acute myeloid leukemia (AML) is a heterogeneous malignancy that comprises 25-30% of pediatric leukemias in Korea. Several inherited diseases, such as Down syndrome and Fanconi anemia, predispose towards AML leukemogenesis. Subgrouping of AML is a key diagnostic step, previously done with the French-American-British (FAB) classification and recently complemented by that of the World Health Organization (WHO). An important feature of AML is the possibility of chloroma at diagnosis, which, if detected, requires follow-up evaluation to determine treatment response. Numerous genetic abnormalities with prognostic relevance have recently been found, the most important of which include those of the core-binding factor (CBF) leukemias, and FLT3-ITD mutation. These genetic abnormalities, combined with patient response to initial treatment, allow for a scheme of risk stratification, and the current consensus is to treat low risk patients with chemotherapy only, whereas high risk patients may receive allogeneic transplant in first remission, although the benefits of transplant remain inconclusive. Overall, the outcome of children and adolescents with AML has improved significantly so that many clinical trials now report event-free survival of around 60%. However, much of this improvement stems from better supportive care and transplant methods, and the genetics-based diagnostic advances in AML have yet to result in enhanced treatment. New therapeutics, including possibly targeted therapy, are necessary to further improve the outcome of pediatric AML.

Incidences and Prognostic Impact of c-KIT, WT1, CEBPA, and CBL Mutations, and Mutations Associated With Epigenetic Modification in Core Binding Factor Acute Myeloid Leukemia: A Multicenter Study in a Korean Population

BACKGROUND: To identify potential molecular prognostic markers in core binding factor (CBF) AML, we analyzed incidences and prognostic impacts of mutations in c-KIT, WT1, CEBPA, CBL, and a number of epigenetic genes in CBF AML. METHODS: Seventy one and 21 AML patients with t(8;21) and inv(16) were enrolled in this study, respectively. NPM1, CEBPA, c-KIT, IDH1/2, DNMT3A, EZH2, WT1, and CBL mutations were analyzed by direct sequencing. Patients were categorized with respect to c-KIT and WT1 mutation status, and both clinical features and prognoses were compared. RESULTS: The incidences of FLT3 internal tandem duplication (ITD), NPM1, CEBPA, IDH1/2, DNMT3A, EZH2, and CBL mutations were low (< or =5%) in CBF AML patients. However, c-KIT and WT1 mutations occurred frequently (10.9% and 13.8%, respectively). t(8;21) patients with c-KIT mutations showed significantly shorter overall survival (OS) and disease free survival (DFS) periods than those without mutations (P<0.001, for both); however, although the limited number of t(8;21) patients were analyzed, WT1 mutation status did not affect prognosis significantly. Relapse or death during follow-up occurred more frequently in t(8;21) patients carrying c-KIT mutations than in those without the mutation, although the difference was significant only in a specific patient subgroup with no WT1 mutations (P=0.014). CONCLUSIONS: The incidences of mutations in epigenetic genes are very low in CBF AML; however, c-KIT and WT1 mutations occur more frequently than others. The poor prognostic impact of c-KIT mutation in t(8;21) AML patients only applies in a specific patient subgroup without WT1 mutations. The prognostic impact of WT1 mutation in CBF AML is not evident and further investigation is required.

Diagnosis and Treatment of Pediatric Acute Myeloid Leukemia / 임상소아혈액종양

Acute myeloid leukemia (AML) is a heterogeneous malignancy that comprises 25-30% of pediatric leukemias in Korea. Several inherited diseases, such as Down syndrome and Fanconi anemia, predispose towards AML leukemogenesis. Subgrouping of AML is a key diagnostic step, previously done with the French-American-British (FAB) classification and recently complemented by that of the World Health Organization (WHO). An important feature of AML is the possibility of chloroma at diagnosis, which, if detected, requires follow-up evaluation to determine treatment response. Numerous genetic abnormalities with prognostic relevance have recently been found, the most important of which include those of the core-binding factor (CBF) leukemias, and FLT3-ITD mutation. These genetic abnormalities, combined with patient response to initial treatment, allow for a scheme of risk stratification, and the current consensus is to treat low risk patients with chemotherapy only, whereas high risk patients may receive allogeneic transplant in first remission, although the benefits of transplant remain inconclusive. Overall, the outcome of children and adolescents with AML has improved significantly so that many clinical trials now report event-free survival of around 60%. However, much of this improvement stems from better supportive care and transplant methods, and the genetics-based diagnostic advances in AML have yet to result in enhanced treatment. New therapeutics, including possibly targeted therapy, are necessary to further improve the outcome of pediatric AML.

Detecção de mutações no gene KIT em leucemia mieloide aguda / The detection of KIT mutations in acute myeloid leukemia

OBJETIVO: Descrever a metodologia para detecção de mutações nos éxons 8 e 17 do gene KIT em pacientes portadores de leucemia mieloide aguda, para implementação desse teste no laboratório clínico do Hospital Israelita Albert Einstein. MÉTODOS: Extração do DNA genômico de 54 amostras de sangue periférico ou medula óssea de pacientes com leucemia mieloide aguda para amplificação, por reação em cadeia da polimerase, sequenciamento e análise de fragmentos. RESULTADOS: Dentre as amostras analisadas, quatro apresentaram mutação no éxon 8, duas no éxon 17 e uma amostra apresentou mutação nos dois éxons. CONCLUSÃO: A pesquisa de mutação nos éxons 8 e 17 do gene KIT foi padronizada com sucesso e o teste está em processo de inclusão no menu de exames do laboratório clínico do Hospital Israelita Albert Einstein.

OBJECTIVE: This study describes a new method used in the clinical laboratory at Hospital Israelita Albert Einstein to detect mutations in exons 8 and 17 of the KIT gene in patients with acute myeloid leukemia. METHODS: Genomic DNA extraction was performed on 54 samples of peripheral blood or bone marrow from patients with acute myeloid leukemia. The extracted DNA was amplified by polymerase chain reaction and sequenced, and the fragments were analyzed. RESULTS: Within the analyzed samples, we detected four mutations in exon 8, two mutations in exon 17, and mutations or a double mutation in one sample. CONCLUSION: The tests detecting mutations in exon 8 and 17 on the KIT gene were successfully standardized. The test is now included among the routine diagnostics employed for patients at Hospital Israelita Albert Einstein clinical laboratory.

Clinical and cytogenetic features and their influencing factors of core binding factor acute myeloid leukemia / 中国医学科学院学报

<p><b>OBJECTIVE</b>To discuss the clinical and cytogenetic features of core binding factor (CBF) acute myeloid leukemia (AML) patients and the main factors that influence the prognosis.</p><p><b>METHOD</b>Totally 130 CBF AML patients were followed up and their clinical features, immunophenotype, chromosome karyotype, treatment regimen, overall survival (OS), and relapse-free survival (RFS) were analyzed.</p><p><b>RESULTS</b>The overall complete remission (CR) rate was 96.1%, among which the CR rate after the first treatment course was 77.2%. The overall median OS was 51.64 (0.26-132.5) months, while the median RFS did not reach 1.18-96.62 months. The 3-year OS was 50% and the 5-year OS was 41%; the 3-year RFS was 59% and the 5-year RFS was 54%. Patients who were over 45 years and those with chromosome karyotype of 9q- tended to have poorer prognosis. During the consolidating chemotherapy, patients who had received two or more courses of intermediate-dose Ara-C therapy had better prognosis and longer survival. AML patients with inv (16) /t (16; 16) had a significantly higher OS than those with t (8; 21) (P = 0.046), while the RFS showed an opposite finding (P = 0.038).</p><p><b>CONCLUSIONS</b>Age, chromosomal karyotype, and consolidating chemotherapy are the main factors that influence the survival and prognosis of CBF AML patients. Two or more courses of intermediate-dose Ara-C during consolidating chemotherapy can obviously prolong the OS and RFS of CBF AML patients. AML patients with a chromosomal karyotype of inv (16) /t (16; 16) have longer OS and better prognosis than those with t (8; 21).</p>

Analysis of tyrosine kinases gene mutations in core binding factor related acute myeloid leukemia and its clinical significance / 中华血液学杂志

<p><b>OBJECTIVE</b>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.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSIONS</b>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.</p>

Clinical value of interphase fluorescent in situ hybridization in diagnosis of core-binding factor acute myelocytic leukemia / 中国实验血液学杂志

The purpose of this study was to evaluate the clinical value of interphase fluorescence in situ hybridization (I-FISH) in diagnosis of core-binding factor acute myelocytic leukemia (CBF AML). The cytogenetic characteristics in leukemia cells from 82 cases of AML-M(2) and 43 cases of AML-M(4)/M(5) were detected by using I-FISH with AML1-ETO double color double fusion probe and double color break point isolated gene probe CBFβ-MYH11, and the detected results were compared with results detected by conventional cytogenetic R banding technique (CC). The results indicated that AML1-ETO fusion gene was detected in 30.5% cases (25/82) by FISH, and t(8;21)(q22;q22) karyotypic aberrations was found in 28.0% cases (23/82) by CC method. Among 25 FISH positive cases, typical FISH positive signal pattern (1R1G2F) was displayed in 22 cases and atypical signal pattern (1R2G1F and 2R1G2F) was found in the other 3 cases. Among all 43 AML-M(4)/M(5) cases, the CBFβ-MYH11 fusion gene was detected in 23.3% cases (10/43) by FISH, which sensitivity was significant higher than that by CC method (2/43) (p < 0.05). It is concluded that some insufficiency of CC technique can be compensated by FISH, and combination of I-FISH with CC technique play a crucial role in diagnosis of CBF AML and in monitoring of minimal residual disease.

Expression of core binding factor a1, bone morphogenetic proteins and osteopontin in the developing periodontal tissues of mice / 华西口腔医学杂志

<p><b>OBJECTIVE</b>To study the expression and interaction of core binding factor a1 (Cbfa1), bone morphogenetic proteins (BMPs) and osteopontin (OPN) in the developing periodontal tissues of mice.</p><p><b>METHODS</b>A mice developing periodontal tissues study model was created histologically by 5-27 day postnatal BALB/c mice, then the immunohistochemical localization of Cbfa1, BMPs and OPN in different developing stages were undertaken.</p><p><b>RESULTS</b>In early stage of postnatal mice periodontal tissues development, only BMPs expressed in dental follicle cells, though the signal was weak. When root was forming, all of them were expressed in periodontal ligament cells and cementoblasts, while only OPN in acellular cementum, cellular cementum and the surface of alveolar bone, Cbfa1 only in cellular cementum and BMPs was seen in neither acellular cementum nor cellular cementum.</p><p><b>CONCLUSION</b>Cbfa1, BMPs and OPN all involve in the development of periodontal tissues, while OPN is crucial for cementum.</p>

Expression of core-binding factor a1 by human skin fibroblasts induced in vitro / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the probabilities of core-biding factor a1 (Cbfa1) expression by human skin fibroblasts induced in vitro.</p><p><b>METHODS</b>The fibroblasts were isolated, purified from human skin, and were grown in incubation in the media of TNF-alpha, BMP-2, and combined TNF-alpha and BMP-2 at certain concentrations, respectively. The changes in biological features of these fibroblasts correlated with osteogenesis were detected by immunohistochemistry and RT-PCR assay.</p><p><b>RESULTS</b>TNF-alpha could switch phenotype of collagen in fibroblasts from Type I and III to Type I and induce fibroblasts to express Ras and BMP type I receptor (BMPR-IA). TNF-alpha in combination with BMP-2 could induce fibroblasts to express Cbfa1 and osteocalcin mRNA.</p><p><b>CONCLUSION</b>Human skin fibroblast could be induced into pro-osteoblast expressing Cbfa1, an osteoblast-specific transcription factor and a regulation of osteoblast differentiation, and combined use of TNF-alpha and BMP-2 was one of the regulating factors.</p>