Identifying novel therapeutic leads for neuroblastoma

Supervisor: Dr Andrew Stoker

Neuroblastoma (NB) is the most common extracranial solid tumour in children, underlying 15% of paediatric cancer deaths [1]. High grades of metastatic disease present great clinical challenges, as does disease recurrence. Improved chemical and biological treatments for NB are continually being sought and our group has recently shown that chemical inhibitors derived from the element vanadium can selectively kill 50% of NB cell lines, or induce them to turn into harmless neurons in cell culture. These inhibitors broadly target proteins called tyrosine phosphatases (PTPs). PTPs play key signalling roles during neural development, regulating RTK signalling both positively and negatively [2]. This protein family is also of growing interest in human cancer biology [3 ].

Vanadium-based chemicals have been used in cancer studies for many years [4,5], but these chemicals may not necessarily be ideal themselves for use directly in humans. This proposed project therefore aims to understand the molecular mechanisms underlying the cytotoxicity of vanadium chemicals in neuroblastoma cells, in order to exploit this knowledge for future therapeutic advances.

Main Hypothesis:
We hypothesise that the cytotoxic capacity of BMOV is dependent upon the induction of novel, downstream transcriptional and biochemical changes in NB cells, which in turn either block critical survival-promoting pathways, or directly induce cell death.

Experimental Approach:
We will treat NB cell lines with vanadium compounds and use gene expression profiling and bioinformatics to define key gene expression changes during cell death induction. The genes that underpin the cell-killing potential of the inhibitors will be identified and their functions characterised. We aim to exploit this knowledge in the longer term, to find selective chemical inhibitors of the gene products that can be used therapeutically.

The student will pursue the following plan:
1) mRNA will be extracted from chemically-treated NB cells and controls, and the student will perform a microarray analysis of the transcriptional changes that occur early during the induction of cell death.
2) A bioinformatics analysis will allow the student to extract candidate genes that show significant expression changes during the chemical treatment. Changes in gene expression will be validated using QPCR and immunoblotting.
3) The cellular and biochemical functions of top candidate genes will be extensively investigated using over- and under-expression methodologies and cell signalling analyses in select NB cell lines, as well as in ovo tumour assays.
4) The expression profiles of candidate genes will also be determined both in primary human NB tumours samples and in the developmental context of the sympathoadrenal lineage in human embryos.

References:
1) Brodeur GM (2003) Neuroblastoma: biological insights into a clinical enigma. Nat Rev Cancer 3 (3):203-216
2) Tonks NK (2006) Protein tyrosine phosphatases: from genes, to function, to disease. Nat Rev Mol Cell Biol 7 (11):833-846
3) Julien SG, Dube N, Hardy S, Tremblay ML (2011) Inside the human cancer tyrosine phosphatome. Nature reviews Cancer 11 (1):35-49.
4) Evangelou AM (2002) Vanadium in cancer treatment. Crit Rev Oncol Hematol 42 (3):249-265.
5) Bishayee A, Waghray A, Patel MA, Chatterjee M (2010) Vanadium in the detection, prevention and treatment of cancer: the in vivo evidence. Cancer Lett 294 (1):1-12.