Significance of nucleophosmin1 (NPM1) gene mutation status on acute myeloid leukaemia patients with normal karyotype in South India

AbstractAcute myeloid leukaemia (AML) is a neoplasm of immature myeloid cells characterized by various cytogenetic alterations. The present study showed that in addition to the FLT3-ITD and NPM1 mutation status, telomere length (TL) and telomerase reverse transcriptase (TERT) gene polymorphisms may affect risk and overall survival (OS) in AML. TL was longer in healthy controls than in AML patients and positively correlated with age in the patients, but not in healthy subjects. TL was found to be independently affected by the presence of the FLT3-ITD mutation. As for the TERT gene polymorphism, AML patients with the TERT rs2853669 CC genotype were characterized by significantly shorter OS than patients carrying the T allele. Another observation in our study is the difference in TL and OS in patients belonging to various risk stratification groups related to the FLT3-ITD and NPM1 mutation status. Patients with adverse risk classification (mutation in FLT3-ITD and lack of mutation in NPM1) presented with the shortest telomeres and significantly worse OS. In conclusion, OS of AML patients appears to be affected by TERT gene variability and TL in addition to other well-established factors such as age, WBC count, or FLT3-ITD and NPM1 mutation status.

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Development of a Human Acute Myeloid Leukaemia Screening Panel and Identification of Novel Gene Mutations.

Blood ◽

10.1182/blood.v104.11.2991.2991 ◽

2004 ◽

Vol 104 (11) ◽

pp. 2991-2991

Author(s):

Jude Fitzgibbon ◽

Matthew Smith ◽

Rachael Arch ◽

Lan-Lan Smith ◽

Nigel Bainton ◽

...

Keyword(s):

Acute Myeloid Leukaemia ◽

Myeloid Leukaemia ◽

Candidate Genes ◽

Gene Mutations ◽

Normal Karyotype ◽

Risk Groups ◽

Cyclin D3 ◽

Kit Mutation ◽

Individual Gene ◽

Acute Myeloid

Abstract Mutation studies in Acute Myeloid Leukaemia (AML) are complicated by the existence of distinct morphological and cytogenetic subtypes; consequently although mutations in any one gene may occur in only 5% of AML, the frequency of mutation may differ both between the different FAB-types and cytogenetic risk groups studied. It is important therefore not only to validate the screening methodology used but also the suitability of the patient panel tested. A screening panel was developed allowing detection of novel recurring gene mutations within samples derived from patients with AML. Mutation analysis of 6 previously described genes (RUNX1, FLT3, KIT, CEBPA, PTPN11, NRAS) and 2 candidate genes (CCND3, FES) were carried out in a cohort of 175 AML samples representing all FAB types (except M3) and cytogenetic risk groups using a combination of SSCP, DHPLC and sequence analysis. One hundred and fifteen mutations were identified in 97 (55%) patients comprising 81 patients (46%) with one mutation, 14 patients (8%) with 2 mutations, and 2 patients (1%) with 3 mutations. Fifty-five out of 88 (63%) patients with normal karyotype AML had at least one mutation. There was was a weak negative association between FLT3 ITD and loop mutation (p = 0.095), a positive association between KIT mutation and favourable risk cytogenetics (p = 0.001), CEBPA mutation and intermediate risk/normal cytogenetics (p = 0.045) and PTPN11 mutation and poor risk disease (p = 0.001). The frequency of individual gene mutation was in accordance with previously published studies. Three novel mutations of FLT3 (Y589D, D839G, Y842H) were detected in 4 patients that would have been overlooked by conventional gel electrophoresis techniques. A single in frame 51bp deletion of nucleotide 939 – 990, resulting in a deletion of 17 amino acids at the carboxyl-terminus of the cyclin D3 protein was identified in a single patient. Overall, both the pattern and mutation frequencies reported in this cohort are similar to those in the literature supporting its further use as an investigational tool in the evaluation of candidate genes in the genesis of myeloid malignancy.

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New drug blocks acute myeloid leukaemia gene mutation

The Lancet Oncology ◽

10.1016/s1470-2045(02)00840-9 ◽

2002 ◽

Vol 3 (8) ◽

pp. 455

Author(s):

Kathleen Nelson

Keyword(s):

Acute Myeloid Leukaemia ◽

Myeloid Leukaemia ◽

Gene Mutation ◽

New Drug ◽

Acute Myeloid

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Development of a quantitative real-time polymerase chain reaction method for monitoring CEBPA mutations in normal karyotype acute myeloid leukaemia

British Journal of Haematology ◽

10.1111/j.1365-2141.2006.06001.x ◽

2006 ◽

Vol 0 (0) ◽

pp. 060203080756008

Author(s):

Lan-Lan Smith ◽

Daniel Pearce ◽

Matthew L. Smith ◽

Michael Jenner ◽

T. Andrew Lister ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Acute Myeloid Leukaemia ◽

Myeloid Leukaemia ◽

Normal Karyotype ◽

Polymerase Chain Reaction Method ◽

Chain Reaction ◽

Reaction Method ◽

Polymerase Chain ◽

Cebpa Mutations ◽

Acute Myeloid

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Genes Located within the del(9q) Commonly Deleted Region Are Down-Regualted in Acute Myeloid Leukaemia.

Blood ◽

10.1182/blood.v104.11.2040.2040 ◽

2004 ◽

Vol 104 (11) ◽

pp. 2040-2040

Author(s):

Zheng Zhao ◽

Sarah Daly ◽

John Liu Yin ◽

David Sweetser ◽

Jacqueline Boultwood ◽

...

Keyword(s):

Acute Myeloid Leukaemia ◽

Myeloid Leukaemia ◽

Normal Karyotype ◽

Complex Karyotype ◽

Down Regulation ◽

Expression Levels ◽

Normal Individuals ◽

Low Expression ◽

Acute Myeloid

Abstract Deletions of 9q are recurring cytogenetic abnormalities in acute myeloid leukaemia (AML). In approximately one-third of cases del(9q) occurs in association with t(8;21). We have previously identified a 2.4Mb region located on 9q21.32–21.33 which is deleted in cases of del(9q) AML - the del(9q) commonly deleted region (CDR). This region encodes 11 genes which we have also previously shown not to be mutated in del(9q) AML. In order to further investigate the role of these genes in AML and in particular to elucidate the pathogenesis of del(9q) AML we have examined the expression of these genes in AML. RNA was extracted from the bone marrow or peripheral blood of patients with AML at the time of diagnosis. Patient samples from the following cytogenetic subgroups were included in this study: (1) del(9q) AML (n=8) - this includes 3 patients with associated t(8;21); (2) t(8;21) but no del(9q) (n=15); (3) Normal karyotype (n=6); (4) Complex Karyotype (n=6). Taqman assays were designed for 9 of the 11 genes located within the del(9q) CDR: FRMD3; ENSG00000148057; UBQLN1; GKAP42; Q9UF54; Q8N2B1; Q9H9A7; SLC28A3; NTRK2. For the other 2 genes within the region Taqman assays could not be performed because of uniformly low expression levels (Q8IZ41) and lack of specificity of primer-design (HNRPK). CD34-purified progenitors from normal individuals were used as controls. It was found that 6 of the 9 genes were significantly down-regulated in del(9q) AML (p<0.05): ENSG00000148057; UBQLN1; Q9UF54; Q8N2B1; Q9H9A7; NTRK2. Since del(9q) is commonly associated with t(8;21), cases of t(8;21) in which del(9q) was not present were also analysed for the expression levels of the del(9q) CDR genes. It was found that 5 of the 9 genes were significantly down regulated in t(8;21) AML (ENSG00000148057; Q9UF54; Q8N2B1; Q9H9A7; SLC28A3) (p<0.05). Only two of these genes were found to be down-regulated in AML of normal karyotype (Q9H9A7 and UBQLN1) (p<0.05) and no significant down-regulation was detected in any of these genes in AML of complex karyotype. Our findings indicate that several genes from within the del(9q) AML CDR are down-regulated in del(9q) AML. A similar pattern of down-regulation is found in cases of t(8;21) even in the absence of del(9q) AML. This suggests that down-regulation of one or more of these genes may be important in the pathogenesis of AML. It may therefore be hypothesized that this pattern of gene down-regulation provides a mechanism common to the development of AML with both del(9q) and t(8;21).

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MicroRNA Expression in Acute Myeloid Leukaemia and Their Effects on the Differentiation of Acute Myeloid Leukaemia Cells

Blood ◽

10.1182/blood.v118.21.2430.2430 ◽

2011 ◽

Vol 118 (21) ◽

pp. 2430-2430

Author(s):

Catalina A Palma ◽

Elise Tonna ◽

Adam J Bryant ◽

Vivek Jayaswal ◽

David Agapiou ◽

...

Keyword(s):

Transcription Factors ◽

Acute Myeloid Leukaemia ◽

Myeloid Leukaemia ◽

Normal Karyotype ◽

Microrna Expression ◽

Luciferase Reporter ◽

Maturation Arrest ◽

Over Expression ◽

Qrt Pcr ◽

Acute Myeloid

Abstract Abstract 2430 Acute myeloid leukaemia (AML) is typified by an aberrant halt in maturation of myeloid progenitor cells, leading to uncontrolled proliferation of immature blasts. In the majority of cases of AML with normal karyotype, the underlying cause of the maturation arrest remains unclear. MicroRNAs, small inhibitory RNAs, are known to be dysregulated in cancers and have been postulated to play a causative role in leukaemogenesis. We aimed to investigate the contribution of aberrant microRNA expression to the inhibition of maturation in AML cells. MicroRNA expression profiling was performed on the bone marrow of 28 AML patients with normal karyotype and 8 normal controls. Differential expression was confirmed by qRT-PCR. We found that the expression of a panel of 12 microRNAs was able to accurately separate AML-M1 and AML-M5 subtypes. The AML-M1 subtype represents a more immature cell population when compared to the monoblast morphology observed in AML-M5. Four candidate microRNAs, miR-181a, -146a, -130a and -135b were selected for further investigation based in their putative targeting of key monocytic transcription factors as determined by in silico modelling followed by luciferase assays. In vitro monocyte and macrophage differentiation of HL60 and NB4 cell lines with 1,25-dihydroxyvitamin D3 and/or phorbol-12-myristate-13-acetate treatment resulted in a significant decrease in all four candidate microRNAs (measured by qRT-PCR), supporting the hypothesis that candidate microRNA over-expression in AML-M1 may contribute to its maturation arrest. Over-expression of the candidate microRNAs by transfection with Pre-miR microRNA precursor molecules (Ambion) into the HL60 and NB4 monocyte differentiation model, resulted in the significant suppression of CD14 (Figure, *p <0.05 compared to Scrambled control) and CD15 expression, markers of monocytes and granulocytes respectively. Conversely, knockdown of miR-181a, -146a, -130a and -135b with miRCURY LNA microRNA knockdown probes (Exiqon) induced an increase in CD14 expression in HL60 cells compared to non-targeting Scrambled control. An important regulatory role of these microRNAs in myeloid/monocytic differentiation is strongly suggested by their targeted suppression of the key transcription factors KLF4, MAFB, IRF8, HOXA10, MCL1 and PU.1 which was confirmed by luciferase reporter assay. This study provides evidence that the over-expressed microRNAs discovered in our profiling work between AML-M1 and AML-M5 are biologically relevant microRNAs, which have the potential to affect the maturation potential of AML cells. Disclosures: No relevant conflicts of interest to declare.

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