In Philadelphia chromosome‑positive acute lymphoblastic leukemia, should measurable residual disease be assessed by next‑generation sequencing or flow cytometry at the end of induction if the initial MRD evaluation was performed with NGS?

MRD Assessment in Ph+ ALL: Continue with NGS After End of Induction

If you initiated MRD monitoring with NGS in Philadelphia chromosome-positive ALL, you should continue using NGS at the end of induction rather than switching to flow cytometry. NGS provides superior sensitivity and prognostic accuracy in Ph+ ALL, particularly when combined with BCR::ABL1 RT-PCR monitoring 1.

Rationale for NGS Continuation

Superior Sensitivity and Prognostic Value

• NGS detects MRD at sensitivities up to 1×10⁻⁶, far exceeding flow cytometry's typical 10⁻⁴ sensitivity 2
• In Ph+ ALL specifically, NGS-based MRD negativity carries stronger prognostic value than RT-PCR alone for predicting relapse-free survival 3, 2
• Recent data show that discordant results where NGS remains positive despite negative flow cytometry consistently predict inferior survival, indicating NGS captures clinically relevant disease that flow cytometry misses 4

Consistency in Methodology

The 2025 Blood Advances expert panel guidelines emphasize that patients should undergo MRD assessment after induction but do not mandate switching assay types 1. Maintaining the same methodology allows for:

• Accurate longitudinal tracking of the same clonal markers
• Avoidance of false-negative results from assay switching
• Better interpretation of MRD kinetics over time

Dual-Method Approach for Ph+ ALL

The optimal strategy combines NGS with BCR::ABL1 RT-PCR, not NGS with flow cytometry 4, 2:

• BCR::ABL1 RT-PCR should be performed in parallel to assess complete molecular response (CMR, defined as transcript <0.01) at 3 months, which is the established prognostic milestone in Ph+ ALL 1
• NGS for IG/TR rearrangements provides deeper sensitivity and can detect emerging clones or clonal evolution that BCR::ABL1 monitoring might miss 3, 4

Important Caveat

In Ph+ ALL, BCR::ABL1 RT-PCR may remain positive despite negative NGS/flow cytometry due to multilineage involvement or CML-like features 4. This discordance requires clinical judgment but does not necessarily indicate treatment failure in the era of potent TKIs and blinatumomab.

Practical Algorithm

At end of induction in Ph+ ALL where NGS was used upfront:

1. Perform NGS for IG/TR rearrangements (same markers tracked from diagnosis)
2. Perform BCR::ABL1 RT-PCR in parallel
3. Interpret results together:
   • NGS-negative + BCR::ABL1 CMR = excellent prognosis, continue protocol
   • NGS-positive (any level) = high-risk regardless of other results, consider treatment intensification 4
   • NGS-negative + BCR::ABL1 positive = continue monitoring, may reflect multilineage disease rather than ALL-specific MRD 4

Flow cytometry can be added as a complementary method but should not replace NGS given its inferior sensitivity in detecting prognostically relevant disease 4, 5, 6.

Timing Considerations

• End of induction is the first critical timepoint 1
• 3-month assessment (approximately post-consolidation) is the most prognostically important timepoint for BCR::ABL1 CMR in Ph+ ALL 1
• Continue every 2-3 months for at least the first year, then every 3-4 months through years 2-3 1

Common Pitfalls to Avoid

• Do not switch from NGS to flow cytometry mid-treatment without compelling reason—this creates interpretive challenges and may miss low-level disease
• Do not rely solely on BCR::ABL1 monitoring in Ph+ ALL—NGS adds independent prognostic information and detects clonal evolution 3, 4
• Do not interpret flow cytometry-negative as truly MRD-negative if NGS was not performed—this may lead to underestimation of relapse risk 4

The evidence strongly supports maintaining NGS as your primary MRD assessment method throughout treatment when initiated upfront, supplemented by BCR::ABL1 RT-PCR for Ph+ ALL-specific monitoring.