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MRD

Minimal residual disease (MRD)

  • Patients with acute leukaemia may have 1012 leukaemic cells
    • In a complete clinical remission there may still be up to 1010 malignant cells
    • Responsible for relapse
      • 20% children with ALL
      • 60% adults with ALL

 

  • Variety of methods to detect cells with higher sensitivity than morphology

 

Goal of MRD

  • Tailor treatment
    • Avoid over treatment
    • Reduce risk of relapse
  • Determine quality of stem cell harvests
  • Predict early relapse eg. After allo when DLI might be helpful

 

  • Defined as lowest level of disease detectable in patients with a complete clinical response by the methods available
  • Technique should be
    • Specific
    • Sensitive
      • Detect up to one leukaemic cell in 104 normal cells
    • Reproducible
    • Quantitative
  • 2 main methods
    • Flow cytometry
    • PCR
      • Cytogenetics, FISH, Southern blotting NOT sensitive enough

 

Flow cytometry

  • Identify a specific pattern of antigen expression at presentation
  • Useful in T-ALL where dual expression of TdT and CD3 makes identification of leukaemic cells easier because this phenotype isn’t expressed in normal marrow cells
  • Also been extensively applied to AML
    • AML associated immunophenotypes occur more frequently than cytogenetic abnormalities or their molecular equivalents that can be followed by PCR (80% v. 30%)
  • Advantages
    • Direct measurement of the number of leukaemic cells
    • Measures size of cells
    • Distinguishes between dead and viable cells
    • Quick; 2-3 hours
  • Disadvantage
    • Leukaemic cells may change their immunophenotype
    • Normal cells may be interpreted as malignant because of overlap in antigen expression
    • Variability in technical expertise

 

PCR

  • RT-PCR for DNA
  • Genomic PCR for DNA
    • Can detect gene rearrangement which may not be actively expressed
  • Detect molecular targets that distinguish normal cells from leukaemic cells
    • Eg. Fusion transcripts that result from chromosomal translocations, antigen receptor gene arrangements in lymphoid cells (Ig or TCR)
  • 24-72 hours
  • Integrity of samples is best proven by co-amplifying a control sequence
    • Eg. Normal BCR in a patient with BCR-ABL
  • Sensitivity is determined by selection of primers
  • ALL
    • MRD targets can be identified in 90% of children and 80% of adults
    • In children, MRD analysis at day 28 allows stratification into high/ low risk groups
      • Any detectable disease at 3 months is high risk of relapse
    • Also predictive in adults
      • Conversion from negative to positive PCR or persistence of positive results to the end of treatment is associated with relapse regardless of age
      • Aim to treat these patients before clinical relapse as in APL and CML
  • Widely used in CML monitoring, APL etc
  • FLT-3 not a good target as associated with subclones at presentation, appear de novo


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