High incidence of biallelic point mutations in the Runt domain of the AML1/PEBP2αB gene in Mo acute myeloid leukemia and in myeloid malignancies with acquired trisomy 21

Blood ◽  
2000 ◽  
Vol 96 (8) ◽  
pp. 2862-2869 ◽  
Author(s):  
Claude Preudhomme ◽  
Delphine Warot-Loze ◽  
Christophe Roumier ◽  
Nalthalie Grardel-Duflos ◽  
Richard Garand ◽  
...  
Keyword(s):  
Trisomy 21 ◽  
Myeloid Leukemia ◽  
Stop Codon ◽  
Point Mutations ◽  
Chromosome 21 ◽  
Runt Domain ◽  
Myeloid Malignancies ◽  
Aml1 Gene ◽  
Atypical Cml ◽  
Acute Myeloid

The AML1 gene, situated in 21q22, is often rearranged in acute leukemias through t(8;21) translocation, t(12;21) translocation, or less often t(3;21) translocation. Recently, point mutations in the Runt domain of the AML1 gene have also been reported in leukemia patients. Observations for mutations of the Runt domain of the AML1 gene in bone marrow cells were made in 300 patients, including 131 with acute myeloid leukemia (AML), 94 with myelodysplastic syndrome (MDS), 28 with blast crisis chronic myeloid leukemia (CML), 3 with atypical CML, 41 with acute lymphoblastic leukemia (ALL), and 3 with essential thrombocythemia (ET). Forty-one of the patients had chromosome 21 abnormalities, including t(8;21) in 6 of the patients with AML, t(12;21) in 8 patients with ALL, acquired trisomy 21 in 17 patients, tetrasomy 21 in 7 patients, and constitutional trisomy 21 (Down syndrome) in 3 patients. A point mutation was found in 14 cases (4.7%), including 9 (22%) of the 41 patients with AML of the Mo type (MoAML) (none of them had detectable chromosome 21 rearrangement) and 5 (38%) of the 13 myeloid malignancies with acquired trisomy 21 (1 M1AML, 2 M2AML, 1 ET, and 1 atypical CML). In at least 8 of 9 mutated cases of MoAML, both AML alleles were mutated: 3 patients had different stop codon mutations of the 2 AML1 alleles, and 5 patients had the same missense or stop codon mutation in both AML1 alleles, which resulted in at least 3 of the patients having duplication of the mutated allele and deletion of the normal residual allele, as shown by FISH analysis and by comparing microsatellite analyses of several chromosome 21 markers on diagnosis and remission samples. In the remaining mutated cases, with acquired trisomy 21, a missense mutation of AML1, which involved 2 of the 3 copies of the AML1 gene, was found. Four of the 7 mutated cases could be reanalyzed in complete remission, and no AML1 mutation was found, showing that mutations were acquired in the leukemic clone. In conclusion, these findings confirm the possibility of mutations of the Runt domain of the AML1 gene in leukemias, mainly in MoAML and in myeloid malignancies with acquired trisomy 21. AML1 mutations, in MoAML, involved both alleles and probably lead to nonfunctional AML1 protein. As AML1 protein regulates the expression of the myeloperoxidase gene, the relationship between AML1 mutations and Mo phenotype in AML will have to be further explored.

High incidence of biallelic point mutations in the Runt domain of the AML1/PEBP2αB gene in Mo acute myeloid leukemia and in myeloid malignancies with acquired trisomy 21

Blood ◽  
2000 ◽  
Vol 96 (8) ◽  
pp. 2862-2869 ◽  
Author(s):  
Claude Preudhomme ◽  
Delphine Warot-Loze ◽  
Christophe Roumier ◽  
Nalthalie Grardel-Duflos ◽  
Richard Garand ◽  
...  
Keyword(s):  
Trisomy 21 ◽  
Myeloid Leukemia ◽  
Stop Codon ◽  
Point Mutations ◽  
Chromosome 21 ◽  
Runt Domain ◽  
Myeloid Malignancies ◽  
Aml1 Gene ◽  
Atypical Cml ◽  
Acute Myeloid

Abstract The AML1 gene, situated in 21q22, is often rearranged in acute leukemias through t(8;21) translocation, t(12;21) translocation, or less often t(3;21) translocation. Recently, point mutations in the Runt domain of the AML1 gene have also been reported in leukemia patients. Observations for mutations of the Runt domain of the AML1 gene in bone marrow cells were made in 300 patients, including 131 with acute myeloid leukemia (AML), 94 with myelodysplastic syndrome (MDS), 28 with blast crisis chronic myeloid leukemia (CML), 3 with atypical CML, 41 with acute lymphoblastic leukemia (ALL), and 3 with essential thrombocythemia (ET). Forty-one of the patients had chromosome 21 abnormalities, including t(8;21) in 6 of the patients with AML, t(12;21) in 8 patients with ALL, acquired trisomy 21 in 17 patients, tetrasomy 21 in 7 patients, and constitutional trisomy 21 (Down syndrome) in 3 patients. A point mutation was found in 14 cases (4.7%), including 9 (22%) of the 41 patients with AML of the Mo type (MoAML) (none of them had detectable chromosome 21 rearrangement) and 5 (38%) of the 13 myeloid malignancies with acquired trisomy 21 (1 M1AML, 2 M2AML, 1 ET, and 1 atypical CML). In at least 8 of 9 mutated cases of MoAML, both AML alleles were mutated: 3 patients had different stop codon mutations of the 2 AML1 alleles, and 5 patients had the same missense or stop codon mutation in both AML1 alleles, which resulted in at least 3 of the patients having duplication of the mutated allele and deletion of the normal residual allele, as shown by FISH analysis and by comparing microsatellite analyses of several chromosome 21 markers on diagnosis and remission samples. In the remaining mutated cases, with acquired trisomy 21, a missense mutation of AML1, which involved 2 of the 3 copies of the AML1 gene, was found. Four of the 7 mutated cases could be reanalyzed in complete remission, and no AML1 mutation was found, showing that mutations were acquired in the leukemic clone. In conclusion, these findings confirm the possibility of mutations of the Runt domain of the AML1 gene in leukemias, mainly in MoAML and in myeloid malignancies with acquired trisomy 21. AML1 mutations, in MoAML, involved both alleles and probably lead to nonfunctional AML1 protein. As AML1 protein regulates the expression of the myeloperoxidase gene, the relationship between AML1 mutations and Mo phenotype in AML will have to be further explored.

scholarly journals Asxl1 C-terminal mutation perturbs neutrophil differentiation in zebrafish

Leukemia ◽  
2021 ◽  
Author(s):  
Xiao Fang ◽  
Song’en Xu ◽  
Yiyue Zhang ◽  
Jin Xu ◽  
Zhibin Huang ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Chronic Myelomonocytic Leukemia ◽  
Nonsense Mutations ◽  
Myeloid Malignancies ◽  
A Subunit ◽  
Neutrophil Differentiation ◽  
Polycomb Repressive Complex 1 ◽  
Myelomonocytic Leukemia ◽  
Inhibitor Treatment ◽  
Acute Myeloid

AbstractASXL1 is one of the most frequently mutated genes in malignant myeloid diseases. In patients with myeloid malignancies, ASXL1 mutations are usually heterozygous frameshift or nonsense mutations leading to C-terminal truncation. Current disease models have predominantly total loss of ASXL1 or overexpressed C-terminal truncations. These models cannot fully recapitulate leukemogenesis and disease progression. We generated an endogenous C-terminal-truncated Asxl1 mutant in zebrafish that mimics human myeloid malignancies. At the embryonic stage, neutrophil differentiation was explicitly blocked. At 6 months, mutants initially exhibited a myelodysplastic syndrome-like phenotype with neutrophilic dysplasia. At 1 year, about 13% of mutants further acquired the phenotype of monocytosis, which mimics chronic myelomonocytic leukemia, or increased progenitors, which mimics acute myeloid leukemia. These features are comparable to myeloid malignancy progression in humans. Furthermore, transcriptome analysis, inhibitor treatment, and rescue assays indicated that asxl1-induced neutrophilic dysplasia was associated with reduced expression of bmi1a, a subunit of polycomb repressive complex 1 and a reported myeloid leukemia-associated gene. Our model demonstrated that neutrophilic dysplasia caused by asxl1 mutation is a foundation for the progression of myeloid malignancies, and illustrated a possible effect of the Asxl1-Bmi1a axis on regulating neutrophil development.

AML1 is expressed in skeletal muscle and is regulated by innervation

1994 ◽  
Vol 14 (12) ◽  
pp. 8051-8057
Author(s):  
X Zhu ◽  
J E Yeadon ◽  
S J Burden
Keyword(s):  
Skeletal Muscle ◽  
Acute Myeloid Leukemia ◽  
Electrical Activity ◽  
Myeloid Leukemia ◽  
Structure And Function ◽  
Denervated Muscle ◽  
Aml1 Gene ◽  
Low Levels ◽  
And Function ◽  
Acute Myeloid

Although most skeletal muscle genes are expressed at similar levels in electrically active, innervated muscle and in electrically inactive, denervated muscle, a small number of genes, including those encoding the acetylcholine receptor, N-CAM, and myogenin, are expressed at significantly higher levels in denervated than in innervated muscle. The mechanisms that mediate electrical activity-dependent gene regulation are not understood, but these mechanisms are likely to be responsible, at least in part, for the changes in muscle structure and function that accompany a decrease in myofiber electrical activity. To understand how muscle activity regulates muscle structure and function, we used a subtractive-hybridization and cloning strategy to identify and isolate genes that are expressed preferentially in innervated or denervated muscle. One of the genes which we found to be regulated by electrical activity is the recently discovered acute myeloid leukemia 1 (AML1) gene. Disruption and translocation of the human AML1 gene are responsible for a form of acute myeloid leukemia. AML1 is a DNA-binding protein, but its normal function is not known and its expression and regulation in skeletal muscle were not previously appreciated. Because of its potential role as a transcriptional mediator of electrical activity, we characterized expression of the AML1 gene in innervated, denervated, and developing skeletal muscle. We show that AML1 is expressed at low levels in innervated skeletal muscle and at 50- to 100-fold-higher levels in denervated muscle. Four AML1 transcripts are expressed in denervated muscle, and the abundance of each transcript increases after denervation. We transfected C2 muscle cells with an expression vector encoding AML1, tagged with an epitope from hemagglutinin, and we show that AML1 is a nuclear protein in muscle. AML1 dimerizes with core-binding factor beta (CBF beta), and we show that CGF beta is expressed at high levels in both innervated and denervated skeletal muscle. PEBP2 alpha, which is structurally related to AML1 and which also dimerizes with CBF beta, is expressed at low levels in skeletal muscle and is up-regulated only weakly by denervation. These results are consistent with the idea that AML1 may have a role in regulating gene expression in skeletal muscle.

A novel syndrome of radiation-associated acute myeloid leukemia involving AML1 gene translocations

Blood ◽  
2000 ◽  
Vol 95 (12) ◽  
pp. 4011-4013 ◽  
Author(s):  
Robert Hromas ◽  
Rinah Shopnick ◽  
Hani George Jumean ◽  
Charles Bowers ◽  
Marileila Varella-Garcia ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
World War Ii ◽  
Myeloid Leukemia ◽  
World War ◽  
Nuclear Explosions ◽  
Radiation Induced ◽  
Aml1 Gene ◽  
High Level ◽  
Acute Myeloid

Abstract AML1 is a transcriptional activator that is essential for normal hematopoietic development. It is the most frequent target for translocations in acute leukemia. We recently identified 3 patients in whom pancytopenia developed almost 50 years after high-level radiation exposure from nuclear explosions during or after World War II. In all 3 patients, acute myeloid leukemia (AML) eventually developed that had similar characteristics and clinical courses. Cytogenetics from the 3 patients revealed a t(1;21)(p36;q22), a t(18;21)(q21;q22), and a t(19;21)(q13.4;q22). By fluorescent in situ hybridization (FISH), all 3 translocations disrupted the AML1 gene. Two of theseAML1 translocations, the t(18;21) and the t(19;21), have not been reported previously. It is possible that the AML1 gene is a target for radiation-induced AML.

scholarly journals Translocation breakpoints are clustered on both chromosome 8 and chromosome 21 in the t(8;21) of acute myeloid leukemia

Blood ◽  
1993 ◽  
Vol 81 (3) ◽  
pp. 592-596 ◽  
Author(s):  
JE Tighe ◽  
A Daga ◽  
F Calabi
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Chromosome 8 ◽  
Chromosome 21 ◽  
Novel Gene ◽  
Translocation Breakpoints ◽  
Acute Myeloid

Abstract The t(8;21)(q22;q22) is consistently associated with acute myeloid leukemia (AML) M2. Recent data have suggested that breakpoints on chromosome 21 are clustered within a single intron of a novel gene, AML1, just downstream of a region of homology to the runt gene of D melanogaster. In this report, we confirm rearrangement at the same location in at least 12 of 18 patients with t(8;21). Furthermore, we have isolated recombinant clones spanning the breakpoint regions on both the der(8) and the der(21) from one patient. By using a chromosome 8 probe derived from these clones, we show that t(8;21) breakpoints are also clustered on chromosome 8.

Mutations in the AML1 Gene Define an Unfavorable Subgroup in Acute Myeloid Leukemia with FAB M0.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4340-4340
Author(s):  
Frank Dicker ◽  
Mirjam Klaus ◽  
Torsten Haferlach ◽  
Wolfgang Kern ◽  
Wolfgang Hiddemann ◽  
...  
Keyword(s):  
High Performance ◽  
De Novo ◽  
Mutation Screening ◽  
Point Mutations ◽  
Gene Mutations ◽  
Normal Karyotype ◽  
Runt Domain ◽  
Screening Efficiency ◽  
Genetic Aberration ◽  
Aml1 Gene

Abstract The AML1/RUNX1 gene is the most frequent target for chromosomal translocations in leukemia. Recently point mutations in the AML1 gene have been demonstrated as another mode of genetic aberration. AML1 mutations have been reported in de novo MDS and AML, as well as in therapy related MDS and AML. The AML M0 subtype has been found to be most frequently affected by sporadic AML1 gene mutations. We analysed AML1 gene mutations in a cohort of 49 M0 patients. Mutation screening was performed either with SSCP (n=21) and/or denaturating High Performance Liquid Chromatography (dHPLC) (n=33), 5 cases were analyzed by both methods. SSCP screening of exons 3–5 of the AML1 gene was carried out at the genomic level. These exons cover the socalled Runt domain, which is most frequently mutated. Fragments with aberrant mobility were sequenced. With this method 5 cases were found to be mutated. Subsequently, to improve the screening efficiency an assay using dHPLC was established. Hereby, we screened the cDNA of patient samples for mutations in amino acid codons 1–277 of the AML1b transcript, where the Runt domain is located between codons 49 and 178. All 5 cases detected by SSCP were confirmed by dHPLC. Nine mutations were detected in the cohort of 28 cases (32%) which had not been analyzed by SSCP. In total, 14 of the 49 samples (29%) tested were identified to be mutated, which is a slightly higher frequency than previously reported. In the cohort of 35 AML1 non-mutated cases 20 (57%) had a normal karyotype and 15 (43%) an aberrant karyotypes, whereas only 6 of the 14 AML1 mutated cases (43%) had a normal karyotype (p=0.001). Three of the AML1 mutated cases (21%) also had FLT3 mutations. One had an FLT3-LM, one an FLT3-TKD mutation, and one case both LM and TKD mutations. Clinical follow up data were available for 33 patients (22 AML1 non- mutated, 11 AML1 mutated). The median OS and EFS of the AML1 non-mutated versus the mutated group was 276 days versus 63 days (p = 0.0679) and 276 vs. 63 days (p=0.0630) respectively. Thus the AML1 mutated cases tend to have a worse clinical outcome. When other AML subtypes were screened for AML1 mutations, i.e. M1 (n=26), M2 (n=21) and M4 (n=3), only 1 additional AML1 mutation was detected, confirming the highest prevalence of AML1 mutations in M0. In conclusion, 1) we established a new assay to screen for AML1 mutations. 2) We confirmed the high incidence of AML1 gene mutations in AML M0, both in cases with normal and aberrant karyotype. 3) For the first time we demonstrated that AML1 mutations define an unfavorable subentity in AML M0.

RUNX1/AML1 DNA Binding Domain and ETO/MTG8 NHR2 Dimerization Domain Are Critical to AML1−ETO9a Leukemogenesis.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 772-772
Author(s):  
Ming Yan ◽  
Scott Hiebert ◽  
Dong-Er Zhang
Keyword(s):  
Acute Myeloid Leukemia ◽  
Dna Binding ◽  
Median Survival ◽  
Myeloid Leukemia ◽  
Binding Domain ◽  
Chromosome 8 ◽  
Chromosome 21 ◽  
Dna Binding Domain ◽  
Dimerization Domain ◽  
Acute Myeloid

Abstract The 8;21 translocation, which involves the gene encoding the RUNX family DNA binding transcription factor AML1 (RUNX1) on chromosome 21 and the ETO (MTG8) gene on chromosome 8, generates AML1−ETO fusion proteins. Previous analyses have demonstrated that full length AML1−ETO blocks AML1 function and requires additional mutagenic events to promote leukemia in mice. More recently, we have identified an alternatively spliced form of AML1−ETO, AML1−ETO9a, from t(8;21) AML patient samples (Yan et al. Nat. Med.12:945–949, 2006). AML1−ETO9a lacks the C−terminal NHR3 and NHR4 domains of AML1−ETO and is highly leukemogenic in mice. Here, we report that the AML1 DNA binding domain and the ETO NHR2 dimerization domain, but not the ETO NHR1 domain are critical for the induction of acute myeloid leukemia by AML1−ETO9a. Using retroviral mediated gene expression and hematopoietic cell transplantation in recipient mice, we examined AML1−ETO9a, AML1−ETO9a without the NHR1 domain [AML1−ETO9a (dNHR1)] or the NHR2 domain [AML1−ETO9a(dNHR2)], without a histone deacetylase/Sin3A interacting domain between NHR1 and NHR2 [AML1−ETO9a(d350–428)], and mutant AML1−ETO9a proteins that have lost the capacity to bind DNA [AML1−ETO9a(L148D)] and [AML1−ETO9a(R173Q)] in leukemogenesis. All of the mice transplanted with AML1−ETO9a (n =11) and AML1−ETO9a(dNHR1) (n = 12) expressing cells developed acute myeloid leukemia with a similar phenotype (Lin−/c−kit+) within 21 weeks. The median survival times of mice with AML1−ETO9a and AML1−ETO9a(dNHR1) are 9.4 weeks and 10.5 weeks, respectively. Furthermore, all of the mice expressing AML1−ETO9a(d350–428) (n = 11) also developed leukemia with a median survival time of 17.2 weeks. Significant numbers of AML1−ETO9a(d350–428) expressing cells are positive for myeloid markers CD11b and Gr1 in these leukemic mice. In contrast, none of the mice with AML1−ETO9a(dNHR2) (n = 14), AML1−ETO9a(L148D) (n = 8), and AML1−ETO9a(R173Q) (n = 8) expressing hematopoietic cells developed leukemia. Taken together, these data suggest that the AML1 DNA binding domain and the ETO NHR2 domain are required for AML1−ETO9a induced leukemia development and the region between amino acids 350 and 428 of AML1−ETO9a also affects the differentiation stage and latency of leukemogenesis.

A High Resolution Analysis of Chromosome 21 Amplification In Myeloid Malignancies Reveals An Association with a Specific Cytogenetic Subgroup and Enhanced ERG Gene Expression.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1687-1687
Author(s):  
Hideki Makishima ◽  
Hideki Muramatsu ◽  
Asahito Hama ◽  
Ramon V. Tiu ◽  
Yuka Sugimoto ◽  
...  
Keyword(s):  
Gene Expression ◽  
Down Syndrome ◽  
High Resolution ◽  
Copy Number ◽  
Trisomy 21 ◽  
Chromosome 21 ◽  
Mrna Levels ◽  
Myeloid Malignancies ◽  
Runx1 Gene ◽  
Cd34 Cells

Abstract Abstract 1687 Genetic alterations including chromosomal translocation, somatic mutation, and gene amplification are thought to play a key role in oncogenesis. Gains of whole or segmental chromosome 21 (Ch21) are observed in many types of myeloid malignancies and are often associated with acute megakaryoblastic leukemia (AMKL). In Down syndrome, transient abnormal myelopoiesis and acute lymphoblastic leukemia can be observed, but the prevalence of AMKL is striking. In rare Down syndrome patients, a subcytogenetic Ch21 minimal amplified region is observed and always found to include ERG as well as the RUNX1 gene locus. Recently, gain of ERG gene copy number has been demonstrated to induce leukemia in mouse models and mutations in RUNX1 have been reported in patients with myeloid malignancies with somatic trisomy 21. The pathogenic gene(s) driving malignant disease in congenital and/or somatic gain of Ch21 are poorly understood. We applied high resolution single nucleotide polymorphism array (SNP-A) to study whether small copy number gains are present on Ch21, which cannot be seen by metaphase cytogenetics. We also tested for potential synergistic karyotypic abnormalities in the patients with gain of Ch21 gene segments. We screened a large cohort of 522 patients with myeloid malignancies by SNP-A platform, and detected 36 events that included whole or partial amplification of Ch21 in 32 cases (6%). The affected length was between 215,063 and 46,944,323 bp and the average was 30,732,002. These include 13 congenital lesions (AMKL evolving in Down syndrome), and 23 somatic alterations. Among the AMKL cohort of 34 cases, gains of Ch21 were observed in 15/25 (60%) juvenile and 2/9 (22%) adult cases. A minimal consensus amplification region was defined from nt38637816 to nt38852879 on Ch21 and this region included ERG. Amplification of ERG was identified in 30/36 of the Ch21 gain lesions studied. Although we sequenced all exons of the ERG gene in all cases with Ch21 gain, no mutation was detected. Based on the possibility that gene amplification leads to increased gene expression, ERG mRNA levels were investigated. CD34+ cells showed the highest ERG expression among hematopoietic cell types. When CD34+ cells from acute myeloid leukemia (AML) patients with somatic trisomy 21, with normal copy of Ch21 and healthy donors were investigated by real time PCR, relative expression of ERG was the highest in trisomy 21 patients among three groups. Based on our previous work and that of others, we tested the mutational status of RUNX1 in the 23 cases with Ch21 amplification that included RUNX1. Mutations were found in 2/23 (9%) accompanied by trisomy 21. No mutation was found in patients with Down syndrome. In one mutant case, a homozygous missense mutation, (L56S) was identified and associated with uniparental trisomy that included RUNX1. The second mutant case (W106L) was in a patient with a 45,XY,-7,i(21)(q10) karyoptype. The mutation was duplicated but was not associated with loss of heterozygosity (LOH). When RUNX1 gene expression in the cases with and without trisomy 21 using CD34 positive bone marrow cells was investigated, no significant difference in relative RUNX1 mRNA levels between trisomy 21 and cases with diploid Ch21 was found. Finally, we evaluated whether additional chromosomal lesions were associated with a gain of Ch21 gene segments. Recurrent losses were detected on chromosome 1, 2, 3, 5, 7, 9, and 17. Deletions of 5q were frequent in the cases with somatic gain of Ch21 (47%; 8/17), while no del5q was detected in the cases with Down syndrome. Conversely, LOH17p (3 uniparental disomies (UPDs) and 2 deletions) was found in both somatic and congenital cases (5/32), with one case of deletion17p associated with a hemizygous p53 mutation. In addition, UPD11q was accompanied by a CBL homozygous mutation in a RAEB case with somatic trisomy 21. Del7q was also observed in both groups (4 in somatic and 3 in congenital cases), including a 7q36.1 microdeletion associated with EZH2 in AMKL with Down syndrome. In sum, our study demonstrates that high resolution SNP-A analysis focused on Ch21 gene segments revealed frequent cryptic somatic gain lesions and a uniparental trisomy. ERG was the sole gene located in the minimally shared gain lesions and is overexpressed in a wild type form in AML cases with somatic trisomy 21. RUNX1 mutations were found in 3 or 2 identical alleles of somatic trisomy 21 cases but are absent in most cases of trisomy 21. Disclosures: No relevant conflicts of interest to declare.

Degradation of TRAF6 by Bortezomib-Induced Autophagy Contributes to Cell Death in Acute Myeloid Leukemia and Myelodysplastic Syndrome

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2452-2452
Author(s):  
Jing Fang ◽  
Lyndsey Bolanos ◽  
Garrett Rhyasen ◽  
Carmen Rigolino ◽  
Agostino Cortelezzi ◽  
...  
Keyword(s):  
Cell Death ◽  
Acute Myeloid Leukemia ◽  
Myelodysplastic Syndrome ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
26S Proteasome ◽  
Negative Regulator ◽  
Myeloid Malignancies ◽  
Chromosome 5Q ◽  
Acute Myeloid

Abstract Abstract 2452 Deletion of chromosome 5q in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) patients results in loss of miR-146a, which is a negative regulator of the innate immune pathway by targeting TNF receptor associated factor-6 (TRAF6). Therefore, MDS and AML patients with reduced miR-146a expression concomitantly exhibit elevated TRAF6 protein. TRAF6 is an E3 ubiquitin ligase that catalyzes K63-linked polyubiquitin chains on substrates that lead to pathway activation, one of which includes NF-kB. Mice lacking miR-146a, or with overexpression of TRAF6, develop AML- and MDS-like features. Bortezomib (Velcade©), which shows promise alone or in combination with chemotherapy in certain groups of MDS and AML patients, is a selective and reversible inhibitor of the 26S proteasome. Studies on the mechanism of action of Bortezomib have shown that pro-apoptotic proteins are stabilized following proteasome inhibition and contribute to the anti-cancer effect. In this report, paradoxically, we find that Bortezomib induces rapid and complete degradation of TRAF6 protein, but not mRNA, in MDS/AML cell lines and human CD34+ cells. A similar finding was observed when AML cells were treated with MG132, another proteasome inhibitor, indicating that degradation of TRAF6 is secondary to proteasomal inhibition. Interestingly, the reduction in TRAF6 protein coincides with Bortezomib-induced autophagy, as indicated by conversion of LC3B-I to LC3B-II and degradation of SQSTM1/p62, and subsequently with apoptosis in MDS/AML cells. Addition of an autophagy inhibitor (3-methyladenine [3-MA]) to Bortezomib-treated AML cells maintained TRAF6 protein expression and enhanced cell viability. Similarly, TRAF6 degradation was blocked by 3-MA when cells were treated with Rapamycin, an mTOR inhibitor and inducer of autophagy. These findings suggest that a mechanism of Bortezomib-induced cell death in myeloid malignancies involves elimination of TRAF6 protein by autophagosomes. Forced expression of TRAF6 in two AML cell lines partially blocked the cytotoxic effect of Bortezomib, suggesting that TRAF6 is an important target of Bortezomib. To determine whether loss of TRAF6 is sufficient to impede growth of MDS and AML, we used a genetic approach to inhibit TRAF6 in MDS/AML cell lines and bone marrow cells from MDS patients with deletion of chromosome 5q. RNAi-mediated depletion of TRAF6 in MDS and AML samples resulted in impaired malignant hematopoietic stem/progenitor function and rapid apoptosis. To uncover the molecular consequences following loss of TRAF6, we applied gene expression profiling and identified genes relevant to the survival of MDS and AML cells. In summary, these findings implicate TRAF6 in Bortezomib-induced cell death and in the maintenance of myeloid malignancies, and reveal a novel mechanism of TRAF6 regulation through autophagic degradation. Disclosures: Oliva: Celgene: Consultancy.

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