Inactive extra-chromosomal MYCN amplicons co-localize to the nuclear lamina in therapy-induced senescent neuroblastoma cells

Introduction: Neuroblastoma (NB) is the most common solid tumour in childhood. MYCN amplification, frequently extrachromosomally, is associated with aggressive tumour behaviour. Metronomic topotecan (TPT) and other drugs can induce cellular senescence, a stable cell cycle arrest, in NB in vitro and in vivo leading to a reduction of MYCN gene copies and abrogation of MYCN expression, however the underlying mechanisms are poorly understood.

Methods: Co-localisation analysis of MYCN FISH and lamin immunofluorescence stainings; Lamin A/C knock-down with subsequent Western Blot analysis of MYCN and p21^WAF1/CIP1 and FACS proliferation analysis; ChIP-qPCR experiments for repressive histone marks.

Results: We demonstrate an increased localisation of MYCN amplicons at the nuclear lamina in senNB cells, which also show a high lamin A/C expression in contrast to non-senescent NB cells. Lamin A/C silencing in TPTsen NB cells did not restore MYCN expression, but alleviated cell cycle arrest. Further, H3K9 and H3K27 ChIP experiments showed enrichment of repressive histone marks at the MYCN promoter region in senNB.

Conclusion: We propose that MYCN copies are recruited to the nuclear lamina leading to transcriptional inactivation of the oncogene by repressive histone marks, followed by their expulsion from the nucleus and subsequent degradation.