PREFERENTIAL TRANSCRIPTION OF THE MUTATED ALLELE IN NPM1 MUTATED ACUTE MYELOID LEUKAEMIA
Author(s): ,
Graham Bailey
Affiliations:
Academic Haematology,University of Nottingham,Nottingham,United Kingdom
,
Aranga Baskar
Affiliations:
Academic Haematology,Univeristy of Nottingham,Nottingham,United Kingdom
,
Louise Smith
Affiliations:
Academic Haematology,Univeristy of Nottingham,Nottingham,United Kingdom
,
Nigel Russell
Affiliations:
Academic Haematology,University of Nottingham,Nottingham,United Kingdom
Claire Seedhouse
Affiliations:
Academic Haematology,Univeristy of Nottingham,Nottingham,United Kingdom
EHA Library. Bailey G. Jun 15, 2019; 266639; PS1022
Dr. Graham Bailey
Dr. Graham Bailey
Contributions
Abstract

Abstract: PS1022

Type: Poster Presentation

Presentation during EHA24: On Saturday, June 15, 2019 from 17:30 - 19:00

Location: Poster area

Background
Nucleophosmin (NPM1) is a multi-functional protein which shuttles continually between cellular compartments. Under normal circumstances NPM1 is located predominantly in the nucleolus. NPM1 expression has been shown to be upregulated in many cancers including acute myeloid leukaemia (AML). In addition to overexpression, NPM1 is also commonly mutated in AML.  Mutation of NPM1 causes a frameshift in the 3’ region of the gene resulting in a protein with an abnormal C-terminus, causing aberrant cytoplasmic localisation. Whilst NPM1 mutations are always heterozygous, the ability of the mutant protein to oligomerise with wild type protein results in both wild type and mutant variants located in the cytoplasm.

NPM1 has a number of different splice variants with the major variant encoded by exons 1-9 and 11-12 (NPM1.1). Further variants include NPM1.2 which lacks exons 8 and 10 and NPM1.3 which comprises exons 1-10 (and so lacks the region of sequence mutated in AML). 

Aims
We aimed to study the expression of NPM1.1, NPM1.2 and NPM1.3 in AML samples with and without NPM1 mutations and also assess the level of expression from the wildtype and mutant alleles in variants NPM1.1 and NPM1.2.

Methods
Primers were designed to discriminate between NPM1.1, NPM1.2 and NPM1.3 variants as well as between wild type and mutated alleles. Appropriate preliminary testing demonstrated the ability of the primers to distinguish between variants using sybr green real time PCR, following which AML samples were assessed (n=56 wild type; n=36 mutated).

Results
Total expression of variants NPM1.2 and NPM1.3 was similar in NPM1 mutated and wild type samples, however levels of NPM1.1 were significantly increased in NPM1 mutated AML (normalised NPM1.1 mean values: WT samples 0.115, MUT samples 0.245 p=<0.0001). This difference was attributed to preferential expression of the NPM1.1 mutated allele; levels of mutated NPM1.1 transcripts were twice those of wild type NPM1.1 (mutated NPM1.1 allele; 0.164, wildtype NPM1.1 allele; 0.081 p=0.0007). No differences in the levels of mutated and wildtype NPM1.2 were apparent.

Conclusion
These results suggest that transcription of NPM1 from the mutated and wildtype alleles are not equal, with the mutated allele being preferentially transcribed.  In NPM1 mutated AML, aberrant cytoplasmic localisation of NPM1 is responsible for disease maintenance. It is likely that preferential transcription of the mutated allele results in an increased population of mutant protein compared to wild type protein, with consequential effects on disease maintenance.  Targeted silencing of the mutated allele could therefore provide an effective target for therapeutic interventions.

Session topic: 3. Acute myeloid leukemia - Biology & Translational Research

Keyword(s): Acute myeloid leukemia, Alternative splicing, Transcriptional regulation

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