PhD ThesisIntroduction: Malignant Rhabdoid Tumours (MRT) are unique malignancies caused by
biallelic inactivation of a single gene (SMARCB1). SMARCB1 encodes for a protein that
is part of the SWI/SNF chromatin remodelling complex, responsible for the regulation
of hundreds of downstream genes/pathways. Despite the simple biology of these
tumours, no studies have identified the critical pathways involved in tumourigenesis.
The understanding of downstream effects is essential to identifying therapeutic targets
that can improve the outcome of MRT patients.
Methods: RNA-seq and 450K-methylation analyses have been performed in MRT
human primary malignancies (n > 39) and in 4 MRT cell lines in which lentivirus was
used to re-express SMARCB1 (G401, A204, CHLA-266, and STA-WT1). The MRT cell
lines were treated with 5-aza-2 -deoxycytidine followed by global gene transcription
analysis (RNA-seq and 450K-methylation) to investigate how changes in methylation
lead to tumourigenesis.
Results: We show that primary Malignant Rhabdoid Tumours present a unique and
distinct expression/methylation profile which confirms that MRT broadly constitute a
single and different tumour type from other paediatric malignancies. However, despite
their common cause MRT can be can sub-group by location (i.e. CNS or kidney). We
observe that re-expression of SMARCB1 in MRT cell lines determines
activation/inactivation of specific downstream pathways such as IL-6/TGF beta. We
also observe a direct correlation between alterations in methylation and gene expression
in CD44, GLI2, GLI3, CDKN1A, CDKN2A and JARID after SMARB1 re-expression.
Loss of SMARCB1 also promotes expression of aberrant isoforms and novel transcripts
and causes genome-wide changes in SWI/SNF binding.
Conclusion: Next generation transcriptome and methylome analysis in primary MRT
and in functional models give us detailed downstream effects of SMARCB1 loss in
Malignant Rhabdoid Tumours. The integration of data from both primary and functional
models has provided, for the first time, a genome-wide catalogue of SMARCB1
tumourigenic changes (validated using systems biology). Here we show how a single
V
deletion of SMARCB1 is responsible for deregulation of expression, methylation status
and binding at the promoter regions of potent tumour-suppressor genes. The genes,
pathways and biological mechanisms indicated as key in tumour development may
ultimately be targetable therapeutically and will lead to better treatments for what is
currently one of the most lethal paediatric cancers.NECCR, Children with cancer UK, Brain Trust, Love Oliver,
CCLG, Karen and Iain Wark, The Smiley Ridley Fund, whose financial support made
this project possible