Monitoring of minimal residual disease in MYCN-amplified neuroblastoma by chromosomal breakpoint recognition

Introduction:

About 25% of all neuroblastoma cases show an amplification of MYCN and half of these relapse after first-line therapy, which implies the survival of neuroblasts, referred to as minimal residual disease (MRD). We show that detection of breakpoints can be used as MRD assay for MYCN-amplified neuroblastoma.

Methods:

We employ a custom hybrid capture-sequencing to recover patient-specific alterations from primary tumors. RQ-PCR and ddPCR are designed from the MYCN amplicon chromosomal breakpoints to assess MRD in bone marrow aspirates throughout therapy.

Results:

As a proof of concept, all MYCN breakpoints (n = 25) in ten neuroblastoma cell lines were successfully recovered. MYCN breakpoints were detectable in single tumor cells among up to 10⁶ reference cells. Moreoever, the assay recovered all MYCN breakpoints (n = 14) in patient tumors (n = 6) and breakpoints persisted up to relapse. Furthermore, we archived parallel detection of MYCN breakpoints and other mutated genes (e.g. ALK, TERT).

Conclusion:

We are establishing prospective MRD for individual high-risk patient follow-ups, to detect residual neuroblasts before their clinical manifestation, which may support therapy decisions.