Background:
Neuroblastoma (NB) accounts for ~15% of total childhood cancer mortality as high-risk NB patients have particularly poor overall survival outcomes. MYCN oncogene amplification confers much poorer outcomes.
MicroRNAs (miRNAs) are a class of non-coding RNAs which repress multiple target genes via base-pairing at a seed sequence. Recently several individual miRNAs have been shown to be oncogenic or tumour suppressive in NB, this research represents the first comprehensive genome-wide analysis in any cancer.
Methods:
A genome-wide functional screen of >1200 miRNAs using both miRNA mimics (overexpression) and antisense inhibitors was carried out in both a highly MYCN amplified cell line (Kelly) and a non-MYCN amplified NB cell line (Shep). Cell viability was quantified in the presence of vincristine, doxorubicin and without chemotherapy.
In my project I aim to pursue two classes of miRNAs arising from this screen: (1) those exhibiting a synthetic lethal interaction with chemotherapy, which synergise with chemotherapy and have no phenotype without chemotherapy. (2) miRNAs that are lethal to NB independent of chemotherapy. These miRNAs will first be validated in a secondary screen which includes two additional MYCN-amplified NB cell lines. Validated hits will then be examined for mechanisms of action including direct target genes of the miRNA. We also intend to assess these miRNAs in vivo as potential therapeutics. Alternatively, there is potential for synthetic lethal miRNAs to predict patient response to chemotherapy based on their expression in patient tissue. In addition we can also search for over-represented target genes and pathways.
Results:
In the genome-wide functional screen of miRNA mimics 130 lethals were discovered (viability fold change <0.25) for the MYCN amplified Kelly cell line. 24 out of the 30 lethals found in the non-MYCN amplified Shep cell line were lethals in Kelly.
Furthermore, 22 miRNA mimcs were synthetic lethal in Kelly with either vincristine or doxorubicin (media alone >0.7 and chemotherapy < half of media viability). No synthetic lethals were found in Shep cells however.