RESUMO
Familial acute myeloid leukemia (AML) pedigrees with germline CCAAT/enhancer-binding protein-α (CEBPA) mutation have been rarely reported due to insufficient knowledge of their clinical features. Here, we report two Chinese families with multiple AML cases carrying germline CEBPA mutations, one of which had 11 cases spanning four consecutive generations. Additionally, we collected clinical data of 57 AML patients from 22 families with germline CEBPA mutations, with 58.3% of them harboring double CEBPA mutations. The first mutation frequently occurred at the N-terminal of CEBP/α (78.6%), resulting in an exclusive expression of p30 of CEBPA (CEBPAp30). The second mutation was mostly found at the C-terminal of CEBP/α (CEBPAothers). Germline CEBPAp30 carriers had higher incidences of AML (80.36% vs. 42.86%) and earlier onset of AML (18 vs. 38.5 years old) compared to germline CEBPAothers carriers. Despite the high rates of relapse, most familial AML cases exhibited favorable overall survival (OS), with germline CEBPAp30 carriers having better survival outcomes (>25 years vs. 11 years for CEBPAothers carriers). Among the 27 healthy germline CEBPA-mutated carriers, we detected a pre-leukemia clone harboring a pathogenic IDH2 variant (R140Q)in one individual. These findings should aid in the genetic counseling and management of AML patients and healthy carriers with germline CEBPA mutations.
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Proteína alfa Estimuladora de Ligação a CCAAT , Leucemia Mieloide Aguda , Humanos , Adulto , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/terapia , Leucemia Mieloide Aguda/patologia , Proteínas Estimuladoras de Ligação a CCAAT/genética , Mutação , Células Germinativas/patologia , PrognósticoRESUMO
Sphingolipids and their metabolic pathways have been implicated in disease development and therapeutic response; however, the detailed mechanisms remain unclear. Using a sphingolipid network focused CRISPR/Cas9 library screen, we identified an endoplasmic reticulum (ER) enzyme, 3-Ketodihydrosphingosine reductase (KDSR), to be essential for leukemia cell maintenance. Loss of KDSR led to apoptosis, cell cycle arrest, and aberrant ER structure. Transcriptomic analysis revealed the indispensable role of KDSR in maintaining the unfolded protein response (UPR) in ER. High-density CRISPR tiling scan and sphingolipid mass spectrometry pinpointed the critical role of KDSR's catalytic function in leukemia. Mechanistically, depletion of KDSR resulted in accumulated 3-ketodihydrosphingosine (KDS) and dysregulated UPR checkpoint proteins PERK, ATF6, and ATF4. Finally, our study revealed the synergism between KDSR suppression and pharmacologically induced ER-stress, underscoring a therapeutic potential of combinatorial targeting sphingolipid metabolism and ER homeostasis in leukemia treatment.
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Oxirredutases do Álcool/metabolismo , Retículo Endoplasmático/fisiologia , Homeostase , Leucemia/patologia , Esfingolipídeos/metabolismo , Resposta a Proteínas não Dobradas , Oxirredutases do Álcool/genética , Apoptose , Proliferação de Células , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Humanos , Leucemia/genética , Leucemia/metabolismo , Células Tumorais CultivadasRESUMO
ABSTRACT: To investigate the prognostic value of the circulating peripheral blood cell counts changes in acute myeloid leukemia (AML) at different time points during induction chemotherapy.We retrospectively analyzed the clinical and laboratory data of 237 newly diagnosed AML patients admitted to Fujian Medical University Union Hospital from January 2011 to December 2014.1. When primitive cells were first removed from the circulating peripheral blood, it was called peripheral blood blast clearance (PBBC). These patients were divided into two groups, according to PBBC. Statistical analysis showed that the day 5 of induction chemotherapy was a better cut-off for PBBC. PBBC≤5âdays is defined as early-blast-clearance, while PBBC >6âdays is delayed-blast-clearance. There was significant difference between the two groups on complete remission (CR) rate (Pâ=â.002), recurrence-free survival (RFS) (Pâ=â.026) and overall survival (OS) (Pâ=â.001). 2. Multivariate analysis suggested PBBC is an independent prognostic factor for CR, RFS, and OS in AML. Receiver operating characteristic(ROC) curve analysis showed the CR rate of patients with white blood cell count less than 1.25â×â109/L was significantly higher than that of patients with white blood cell count more than 1.25â×â10â9/L (Pâ<â.001) at day 5 of induction chemotherapy, but the RFS and OS was no significantly different (Pâ>â.05).The dynamics of peripheral blood blast in AML after initiation of induction chemotherapy, especially the time length to achieve PBBC, has important prognostic value for CR rate, RFS, and OS in AML patients. It is a simple and feasible method to evaluate the efficacy of AML.
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Crise Blástica/patologia , Quimioterapia de Indução/métodos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Contagem de Leucócitos/métodos , Adolescente , Adulto , Idoso , China , Feminino , Humanos , Leucemia Mieloide Aguda/mortalidade , Masculino , Pessoa de Meia-Idade , Curva ROC , Recidiva , Estudos Retrospectivos , Análise de Sobrevida , Fatores de Tempo , Adulto JovemRESUMO
This study investigated the therapeutic effects of a water-soluble biphenyl compound, WLP-S-14, in acute-on-chronic liver failure (ACLF). Wistar rats were injected intraperitoneally with porcine serum twice a week for 8 weeks prior to administration of 600 mg/kg D-galactosamine and 50 µg/kg lipopolysaccharide to induce ACLF. Study groups were treated intravenously with saline or with 100 or 200 mg/kg WLP-S-14. WLP-S-14 ameliorated ACLF with significant reductions in the mortality rate and transaminase levels, indicating improved liver function. The mechanism underlying these effects may involve decreased levels of tumor necrosis factor-α and interleukin-6, with associated inhibition of apoptotic pathways.
Assuntos
Compostos de Bifenilo/farmacologia , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Animais , Compostos de Bifenilo/química , Galactosamina/toxicidade , Lipopolissacarídeos/toxicidade , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Estrutura Molecular , Distribuição Aleatória , Ratos , Ratos Wistar , SoroRESUMO
Although hematopoietic stem cell transplantation (HSCT) has been widely used in the treatment of many diseases, graft-versus-host disease (GVHD) remains a major complication after allogeneic HSCT. Butyrophilin-like 2 (BTNL2) protein has been reported to have the ability to inhibit T cell proliferation in vitro; its ability to inhibit T cell responses in vivo has not been determined. We show here that in vivo administration of recombinant BTNL2-IgG2a Fc (rBTNL2-Ig) fusion protein ameliorates GVHD in mice. This is related to the ability of rBTNL2-Ig to inhibit T cell proliferation, activation and Th1/Th17 cytokine production in vivo. Furthermore, rBTNL2-Ig treatment increases the generation of regulatory T cells. Our results suggest that rBTNL2-Ig has the potential to be used in the prevention and treatment of patients with GVHD.
Assuntos
Butirofilinas/metabolismo , Butirofilinas/farmacologia , Doença Enxerto-Hospedeiro/prevenção & controle , Animais , Butirofilinas/imunologia , Doença Enxerto-Hospedeiro/metabolismo , Transplante de Células-Tronco Hematopoéticas/métodos , Humanos , Fragmentos Fc das Imunoglobulinas/imunologia , Imunoglobulina G/imunologia , Imunoglobulina G/farmacologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/farmacologia , Linfócitos T Reguladores/imunologia , Células Th1/imunologia , Células Th17/imunologia , Transplante HomólogoRESUMO
BACKGROUND: BCL-2 Associated X (BAX) is an important modulator of apoptosis. The associations between BAX gene polymorphism and cancer susceptibility and prognosis in different ethnic groups and types of cancer have yielded controversial results. To reconcile the results, a systematic review followed by meta-analysis was performed to assess the associations. METHODS: A systematic search of Medline database (PubMed), EMBASE, China Biology Medicine disc, China National Knowledge Infrastructure, Wanfang databases for publications on BAX polymorphisms, and susceptibility and prognosis was carried out until July 2017. Retrieved 14 articles met the inclusions. Summary odds ratios (ORs) and hazard ratios (HRs) with their 95% confidence intervals (CIs) were harnessed to determine the strength of correlation between BAX polymorphisms and cancer susceptibility and prognosis, which were combined using fixed- or random-effects models as appropriate. RESULTS: A total of 12 trials involving 3321 cases and 3209 controls were included in our pooled analysis regarding the polymorphisms and the susceptibility of cancers. Overall, results of the present meta-analysis demonstrated that there was no significant association between BAX polymorphisms and susceptibility of cancers (ORâ=â1.052, 95% CI: 0.827-1.339, Pâ=â.679, A vs G). Even in a stratified analysis by ethnicity and the sources of control groups, the results were consistent. Four retrospective studies of 549 cases qualified for meta-analysis were identified to set forth the associations of the polymorphisms with cancer prognosis. Our results suggested that BAX gene polymorphisms were significantly associated with unfavorable prognosis (HRâ=â1.735, 95% CI: 1.368-2.202, Pâ=â.000, GG vs GA/AA). CONCLUSION: There is no significant association between BAX gene polymorphism and cancer susceptibility, but it probably contributes to increased adverse prognosis to cancer.
Assuntos
Neoplasias/genética , Proteína X Associada a bcl-2/genética , Predisposição Genética para Doença , Humanos , Polimorfismo Genético , PrognósticoRESUMO
DHX15 plays a role in leukaemogenesis and leukaemia relapse. However, the mechanism underlying the transcriptional regulation of DHX15 in ALL has not been elucidated. Our present study aimed to explore the functional promoter region of DHX15 and to investigate the transcription factors controlling the transcription of this gene. A luciferase assay performed with several truncated constructs identified a 501-bp region as the core promoter region of DHX15. Site-directed mutagenesis, electrophoretic mobility shift and chromatin immunoprecipitation assays showed that ETS1 and SP1 occupied the DHX15 promoter. Furthermore, knockdown of ETS1 and SP1 resulted in suppression of DHX15, whereas the overexpression of these genes led to up-regulation of DHX15. Interestingly, in samples obtained from patients with ALL at diagnosis, both ETS1 and SP1 correlated positively with DHX15 expression. Additionally, differences in methylation of the DHX15 core promoter region were not observed between the patients and controls. In conclusion, we identified the core promoter region of DHX15 and demonstrated that ETS1 and SP1 regulated DHX15 expression in ALL.
Assuntos
Regulação Leucêmica da Expressão Gênica , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Proteína Proto-Oncogênica c-ets-1/metabolismo , RNA Helicases/genética , Fator de Transcrição Sp1/metabolismo , Pareamento de Bases/genética , Sequência de Bases , Sítios de Ligação , Linhagem Celular Tumoral , Ilhas de CpG/genética , Metilação de DNA/genética , Humanos , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Regiões Promotoras Genéticas , Ligação Proteica , Proteína Proto-Oncogênica c-ets-1/genética , RNA Helicases/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Sítio de Iniciação de Transcrição , Transcrição GênicaRESUMO
Genetic heterogeneity is the basis of clinical heterogeneity among different subtypes of AML. We have successfully cloned a gene related to AML termed FAMLF from a FAB-M2 patient's sample of a second largest AML pedigree. Then we revealed at least three splice variants, named as FAMLF-1, FAMLF-2 and FAMLF-3, and found miR181a1/b1 in the second intron of FAMLF gene family. Higher expression of FAMLF-1 was related to a higher complete remission (CR) rate, but shorter relapse free survival (RFS) in AML. We further found that the FAMLF-1 single nucleotide polymorphism (SNP) haplotype and its expression were positively correlated to clinical parameters of acute myeloid leukemia partially differentiated (FAB-M2) patients, but not FAB non-M2 patients or Acute Monocytic Leukemia (FAB-M5) patients. GTAGG SNP haplotype of FAMLF gene might increase FAB-M2 susceptibility in Han population and act as a useful candidate biomarker for FAB-M2 screening. We also demonstrated that FAMLF-1 gene silencing in FAB-M2 cells could lead to proliferation inhibition, cell cycle G0/G1 phase arrest, and differentiation promotion independent of its intronic miR-181a1, which might be related to Akt/c-Myc pathway. These findings reveal a role of FAMLF-1 as a potential pathogenic gene for FAB-M2.
RESUMO
The role of DHX15, a newly identified DEAH-box RNA helicase, in leukemogenesis remains elusive. Here, we identified a recurrent mutation in DHX15 (NM_001358:c.664C>G: p.(R222G)) in one familial AML patient and 4/240 sporadic AML patients. Additionally, DHX15 was commonly overexpressed in AML patients and associated with poor overall survival (OS) (P=0.019) and relapse-free survival (RFS) (P=0.032). In addition, we found a distinct expression pattern of DHX15. DHX15 was highly expressed in hematopoietic stem cells and leukemia cells but was lowly expressed in mature blood cells. DHX15 was down-regulated when AML patients achieved disease remission or when leukemia cell lines were induced to differentiate. DHX15 silencing greatly inhibited leukemia cell proliferation and induced cell apoptosis and G1-phase arrest. In contrast, the restoration of DHX15 expression rescued cell viability and reduced cell apoptosis. In addition, we found that DHX15 was down-regulated when cell apoptosis was induced by ATO (arsenic trioxide); overexpression of DHX15 caused dramatic resistance to ATO-induced cell apoptosis, suggesting an important role for DHX15 in cell apoptosis. We further explored the mechanism of DHX15 in apoptosis and found that overexpression of DHX15 activated NF-kB transcription. Knockdown of DHX15 inhibited the nuclear translocation and activation of the NF-kB subunit P65 in leukemia cells. Several downstream targets of the NF-kB pathway were also down-regulated, and apoptosis-associated genes CASP3 and PARP were activated. In conclusion, this study represents the first demonstration that DHX15 plays an important role in leukemogenesis via the NF-kB signaling pathway and may serve as an independent prognostic marker for AML.
RESUMO
Recent reports have described a new post-transcriptional regulation that RNA transcripts can crosstalk with each other by competing for their common microRNAs. These RNA transcripts termed competing endogenous RNAs (ceRNAs) regulate the distribution of miRNAs on their targets. One corollary from ceRNA interaction is that chromosomal translocation in acute promyelocytic leukemia (APL) would perturb ceRNA regulation due to altered expression of 3'UTRs. In our study, we demonstrate that expression of PML/RARα, the APL-associated fusion oncogene is repressed by c-Myc mRNA transcript independent of protein-coding function but dependent upon microRNA. Attenuation of c-Myc transcript results in PML/RARα-degraded cellular phenotypes in APL cells, but these Myc reduction-associated cell phenotypes are sufficient to abrogate in a microRNA dependent manner. We also show that let-7 microRNA family members promote differentiation of All-Trans-Retinoic Acid (ATRA)-induced NB4 cells and their activities are affected by expression levels of both c-Myc and PML/RARα through altering miRNA targets. These results indicate that c-Myc mRNA represses PML/RARα expression via altering the distribution of let-7 miRNAs on their targets. Our findings reveal a previously unrecognized role of c-Myc as a potential ceRNA for PML/RARα in APL.
Assuntos
Regulação Neoplásica da Expressão Gênica , Leucemia Promielocítica Aguda/genética , MicroRNAs/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Mensageiro/metabolismo , Regiões 3' não Traduzidas/genética , Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Leucemia Promielocítica Aguda/patologia , Proteínas Proto-Oncogênicas c-myc/genética , Interferência de RNA , RNA Mensageiro/genética , RNA Interferente Pequeno/metabolismo , Translocação Genética , Tretinoína/farmacologiaAssuntos
Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/genética , Proteínas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Feminino , Regulação Leucêmica da Expressão Gênica , Estudos de Associação Genética , Humanos , Leucemia Mieloide Aguda/mortalidade , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Prognóstico , Resultado do TratamentoRESUMO
OBJECTIVES: Although additional sex comb-like 1 (ASXL1) gene mutations have long been reported in myelodysplastic syndromes (MDSs) and chronic myelomonocytic leukemia (CMML), the prognostic significance has been controversial. Therefore, a meta-analysis to study the impact of ASXL1 mutations on patients with MDS and CMML is useful. METHODS: The identified articles were retrieved from some common databases. We extracted hazard ratios (HRs) for overall survival (OS) and leukemic-free survival (LFS) and P-value of some clinical parameters, which compared AXSL1 mutations to those without from the available studies. Each individual HR and P-value was used to calculate the pooled HR and P-value. RESULTS: Six studies covering 1689 patients were selected for this meta-analysis. The pooled HRs for OS and LFS were 1.45 (95% confidential interval (CI), 1.24-1.70) and 2.20 (95% CI, 1.53-3.17), respectively. When considering CMML patients alone the HR for OS was 1.50 (95% CI, 1.18-1.90). Additionally, ASXL1 mutations were more frequently found in male (P = 0.008), older (P = 0.019), and patients with lower platelets (P = 0.009) or hemoglobin level (P = 0.0015) and associated with other mutations such as EZH2, IDH1/2, RUNX1, and TET2. DISCUSSION: Although our analysis has its limitation, it showed that ASXL1 mutations had significant inferior impact on OS and LFS for French-American-British-defined MDS patients. However, the influence of different types of ASXL1 mutations on patients with MDS still needs illustrating. CONCLUSION: ASXL1 mutations were associated with poor prognosis in MDS, which may contribute to risk stratification and prognostic assessment in the disease.
Assuntos
Leucemia Mielomonocítica Crônica/genética , Síndromes Mielodisplásicas/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , PrognósticoRESUMO
An inherited predisposition to acute myeloid leukaemia (AML) is exceedingly rare, but the investigation of these families will aid in the delineation of the underlying mechanisms of the more common, sporadic cases. Three AML predisposition genes, RUNX1, CEBPA and GATA2, have been recognised, but the culprit genes in the majority of AML pedigrees remain obscure. We applied a combined strategy of linkage analysis and next-generation sequencing (NGS) technology in an autosomal-dominant AML Chinese family with 11 cases in four generations. A genome-wide linkage scan using a 500K SNP genotyping array was conducted to identify a previously unreported candidate region on 20p13 with a maximum multipoint heterogeneity LOD (HLOD) score of 3.56 (P=0.00005). Targeted NGS within this region and whole-exome sequencing (WES) revealed a missense mutation in TGM6 (RefSeq, NM_198994.2:c.1550T>G, p.(L517W)), which cosegregated with the phenotype in this family, and was absent in 530 healthy controls. The mutated amino acid was located in a highly conserved position, which may be deleterious and affect the activation of TGM6. Our results strongly support the candidacy of TGM6 as a novel familial AML-associated gene.
Assuntos
Leucemia Mieloide Aguda/genética , Mutação de Sentido Incorreto , Transglutaminases/genética , Adulto , Estudos de Casos e Controles , Criança , Feminino , Humanos , Leucemia Mieloide Aguda/diagnóstico , Escore Lod , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , Polimorfismo de Nucleotídeo ÚnicoRESUMO
This research was conducted to compare differences in colon cancer lymphatic vessel invasion (LVI) with D2-40 antibody labeling and regular HE staining, blood vessel invasion (BVI) with CD34 antibody labeling and HE staining and to assess the possibility of using D2-40-LVI/CD34-BVI in combination for predicting stage II colon cancer prognosis and guiding adjuvant chemotherapy.Anti-D2-40 and anti-CD34 antibodies were applied to tissue samples of 220 cases of stage II colon cancer to label lymphatic vessels and small blood vessels, respectively. LVI and BVI were assessed and multivariate COX regression analysis was performed for associations with colon cancer prognosis. Regular HE staining proved unable to differentiate lymphatic vessels from blood vessels, while D2-40 selectively labeled lymphatic endothelial cell cytosol and CD34 was widely expressed in large and small blood vessels of tumors as well as normal tissues. Compared to regular HE staining, D2-40-labeling for LVI and CD34-labeling for BVI significantly increased positive rate (22.3% vs 10.0% for LVI, and 19.1% vs 9.1% for BVI). Multivariate analysis indicated that TNM stage, pathology tissue type, post-surgery adjuvant chemotherapy, D2-40-LVI, and CD34-BVI were independent factors affecting whole group colon cancer prognosis, while HE staining-BVI, HE staining-LVI were not significantly related. When CD34-BVI/D2-40-LVI were used in combination for detection, the risk of death for patients with two or one positive results was 5.003 times that in the LVI(-)andBVI(-) group (95% CI 2.365 - 9.679). D2-40 antibody LVI labeling and CD34 antibody BVI labeling have higher specificity and accuracy than regular HE staining and can be used as molecular biological indicators for prognosis prediction and guidance of adjuvant chemotherapy for stage II colon cancer.
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Anticorpos Monoclonais Murinos , Antígenos CD34/imunologia , Neoplasias do Colo/patologia , Metástase Linfática/patologia , Neoplasias Vasculares/secundário , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais , Neoplasias do Colo/mortalidade , Neoplasias do Colo/cirurgia , Citosol/imunologia , Células Endoteliais/imunologia , Feminino , Humanos , Vasos Linfáticos/patologia , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/patologia , Prognóstico , Estudos RetrospectivosAssuntos
Leucemia Promielocítica Aguda/complicações , Leucemia/etiologia , Adolescente , Feminino , Humanos , Masculino , Adulto JovemRESUMO
This study was purposed to investigate the expression of RPL36A (ribosomal protein 36a) in the newly diagnosed acute myeloid leukemia (AML) cells and its mechanism at the molecular level. The RPL36A mRNA expression in the newly diagnosed AML cells, U937 cells and normal MNCs was determined by RT-PCR. Small interfering RNA (siRNA) targeting to RPL36A was transfected into U937 cells by Lipofectamine 2000 system. Proliferation, cell cycle, apoptosis of U937 were observed through MTT assay, flow cytometry, acridine orange/ethidium bromide (AO/EB) double staining, TUNEL and Annexin V/FITC respectively. RPL36A mRNA and protein expression levels were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot analysis respectively. The results showed that RPL36A expression in the newly diagnosed AML cells and U937 cells was significantly upregulated. The average OD value of U937 cells transfected with RPL36A siRNA was significantly lower as compared with 3 control groups. The cell percentage in G2-and S-phase increased, which indicated the inhibition effect of RPL36A siRNA on cell proliferation. Remarkable cell apoptosis in U937 cells treated with RPL36A siRNA was observed by AO/EB, TUNEL analysis and Annexin V/FITC assay; RPL36A mRNA and protein expression level of U937 cells treated with siRNA were significantly declined in a time-dependent manner (r=0.9813 and 0.9537). It is concluded that the RPL36A expression in the AML cells is significantly enhanced and the RPL36A gene may be involved in regulation of cell cycle and cell apoptosis of AML, which promotes proliferation of AML cells and inhibits apoptosis of cells.
Assuntos
Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Leucemia Mieloide Aguda/patologia , RNA Interferente Pequeno/farmacologia , Proteínas Ribossômicas/farmacologia , Adolescente , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Interferência de RNA , RNA Interferente Pequeno/genética , Proteínas Ribossômicas/genética , Células U937 , Adulto JovemRESUMO
OBJECTIVE: To clone the full-length cDNA of a novel gene related to familial acute myelogenous leukemia (AML) and to demonstrate its molecular mechanisms on the gene level. METHODS: Bone marrow specimen was obtained from a patient of familial AML, male, aged 11, and peripheral blood samples were obtained from 23 AML patients outside this family, 9 normal persons in this family, and 23 normal persons outside this family. Based on the EST sequence zywb87 (GenBank accession number: CV973101) from a subtractive cDNA library of differential expressed genes constructed in familial AML, SMART-rapid amplification of cDNA ends (SMART-RACE) was applied to clone the full-length cDNA of the novel gene, and bioinformatics was used to predict its biological function, the expression of the novel gene in AML was detected by One-Step RT-PCR. RESULTS: A full-length cDNA of 2313 bp was obtained from the bone marrow specimen of the familial AML patient with complete open reading frame (ORF) of 249 bp. Localized on 1q31.3 of human chromosome, it coded a 82-amino acid polypeptide with signal peptide, leucine-rich repeat (LRR_SD22), and intrinsic disorder functional domain. BLAST analysis confirmed this gene as a novel gene designated with the accession number: (nucleotide) EF413001 and (protein) ABN58747 by GenBank and was named as Homo sapiens familial acute myelogenous leukemia related factor (FAMLF). The FAMLF expression level of the AML patients outside this family was (2.61 +/- 0.66), significantly higher than that of the normal persons outside this family (0.97 +/- 0.51, P < 0.01). CONCLUSION: A full-length cDNA of the novel gene FAMLF related to familial AML has been obtained. The FAMLF gene is expressed highly in AML and may present biological function on the progress of AML.
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Predisposição Genética para Doença , Leucemia Mieloide Aguda/genética , Proteínas de Neoplasias/genética , Proteínas/genética , Adolescente , Adulto , Idoso , Criança , Clonagem Molecular , DNA Complementar/genética , Feminino , Biblioteca Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Fases de Leitura Aberta , Análise de Sequência de DNARESUMO
OBJECTIVE: To clone the full-length cDNA of a novel gene of familial acute myelogenous leukemia and to explain the molecular mechanism of the disease at the gene level. METHODS: Based on a EST sequence (zywb4) from a subtractive cDNA library of specially or differentially expressed genes constructed in familial acute myelogenous leukemia, electronic cDNA cloning and SMART-rapid amplification of cDNA ends (SMART-RACE) were used to clone the full-length cDNA of a novel associated gene of familial acute myelogenous leukemia. RESULTS: One sequence of 2257 bp was obtained, which obeyed Kozak rules, and contained an open reading frame (ORF) and a polyA tail. BLAST analysis showed that it may be a novel gene. The new sequence was submitted to GenBank with the accession number: DQ359746 and designated as ELF2C. CONCLUSION: A full- length cDNA of a novel gene (ELF2C) related to familial acute myelogenous leukemia is obtained, which is helpful to further investigation of its function in familial acute myelogenous leukemia.
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Predisposição Genética para Doença , Leucemia Mieloide Aguda/genética , Fatores de Transcrição/genética , Criança , Clonagem Molecular , DNA Complementar/química , DNA Complementar/genética , Biblioteca Gênica , Humanos , Masculino , Dados de Sequência Molecular , Análise de Sequência de DNARESUMO
OBJECTIVE: To isolate genes expressed differentially from the bone marrow cells of patients with familial acute myelocytic leukemia and to explain the molecular mechanisms of the disease at the gene level. METHODS: Bone marrow cells were obtained from a family with cases of familial acute myelocytic leukemia for successive 4 generations. Super SMART cDNA synthesis and suppression subtractive hybridization (SSH) were performed to set up a subtractive cDNA library of specially or highly expressed genes. The white clones were screened and identified by modified differential screening technique in which the probes were labeled with digoxin/deoxyuridine 5'-triphosphate. Positive clones were sequenced and compared with the known sequences in the public databases of GenBank/EMBL/DDBJ using NCBI BLAST for homology analysis. The unknown fragments were then submitted to GenBank. RESULTS: A subtractive library of differentially expressed genes for the family with cases of familial acute myelocytic leukemia was constructed successfully. After differential screening, 28 effective sequences were confirmed to be positive differentially expressed gene fragments and were submitted to GenBank for homology analysis. Then 17 sequences were identified as a part of known genes, and the rests were verified as unknown fragments. CONCLUSION: SSH is an effective approach to isolate differentially expressed genes. Some genes related to cell proliferation and differentiation has been acquired by SSH. Eleven novel expressed sequence tags related to library have been successfully isolated and accepted by GenBank.