MPN Diagnostics

Bone marrow histology features of MPN

That can be assessed with reasonable consistency:

  • Overall haemopoietic cellularity

  • Relative predominance of increased megakaryocyte numbers

  • Megakaryocyte distribution (not the cytological details)

  • Reticulin pattern


Fibrosis Grade


WHO 0-3

Bain 1-4


Grade of fibrosis predicts overall survival

PT-1 Trial – 5 yr OS – Approx 95% for grade 0/1, Approx 87% for grade 3/4


Phenotypic ‘Driver’ Mutations

mpn dx.png


CML Molecular genetics


Depending on the breakpoints in the BCR and ABL1 genes, different sized proteins will be formed from the fusion gene:


  • e1a2 protein - p190 (seen in ALL > CML)

  • b2a2/b3a2 proteins - p210 (most common in CML)

  • e19a2 protein - p230


This is important diagnostically, and need to know which transcript is present when comes to quantitating its presence by RT-PCR. Different transcripts will need different primers for the lab test —> not all transcripts can be quantified in all labs.


JAK2 & MPL Mutations


Mutated JAK2/MPL is activated spontaneously without the presence of the usually required ligands —> persistent activation of the downstream pathways – e.g. STAT, PI3K, MAPK


JAK2 V617F at CUH Lab

  • Quantitative PCR using a fluorescent probe that is released as the allele replicates

  • One probe for the mutant allele and one for the wild type

  • Compare fluorescence of each to quantify


MPL and JAK2 Exon 12 at CUH

  • High resolution melt profile performed after PCR has produced the protein

  • Difference in the way mutated protein melts at 84 degrees


CALR Mutation


Insertion or deletions in Exon 9 are the most common mutations.

CALR is an endoplasmic protein, mutation leads to abnormal movement of MPL within the cell



  • Primer for the area of interest within the CALR gene

  • The length of the protein produced during replication will tell you if mutated – shorter or longer than the wild type (deletions or insertions)


NGS Panels


Next generation sequencing to test whole panels of genes in a single assay

Sensitive for detecting mutations at low levels

Can test several patients within one assay —> reduces costs


Ion Torrent Analyser used at CUH

  • Currently for MF (not PV or ET)


Significance of results

  • Increasing no. of mutated genes detected in panel —> predicts high molecular risk disease


Future of testing

  • BCR/ABL, JAK2, CALR, MPL may put into a single panel and run all at once

  • May not be quite a sensitive as individual JAK2 assay ?significance of this

Personalised Risk Assessment in MPN

Available online from The Sanger Institute - here