gene and cell therapy
PhD projects for research students
Modelling the t(6:9) translocation and MYB-NFIB gene fusion using the Crispr/Cas9 technology: implications for adenoid cystic carcinoma.
Recent advances in genomic technologies have revealed frequent MYB rearrangements in human malignancies. MYB, MYBL1 and MYBL2 belong to a small gene family encoding transcription factors whose role in oncogenesis has always been suspected, but never fully demonstrated. The main goal of the project is to validate the hypothesis that the t(6:9) chromosomal translocation, and formation of the MYB-NFIB fusion gene, is the leading cause of Adenoid Cystic Carcinoma (ACC), a rare and incurable tumour of exocrine glands that will be used as a model of MYB-addicted malignancy. The establishment of animal and cellular models of ACC will be critical for the validation of MYB targeting molecules. A further aim is the understanding of the signalling pathways downstream of MYB that could lead to the identification of new biomarkers and targets for therapy.
1. To create mouse and cellular models of ACC implementing genomic editing technology. The mouse and cellular models will be used to study the molecular pathogenesis of the disease and in preclinical experiments. 2. To study signalling pathways and kinases activated as a result of oncogenic mutation of MYB that could be used as pharmacological targets. 3. To use genomic approaches to distinguish gene signatures common to mouse and human ACCs that could serve as prognostic biomarkers to predict the clinical outcome of ACC patients. 4. To identify and refine MYB small molecule inhibitors and develop them into drugs for the treatment of ACC and other MYB driven cancers.
Targeting TERT and the MYCN gene network as a novel therapeutic approach for high-risk neuroblastoma
MYCN belongs to a small family of transcription factors implicated in fundamental cellular processes. There are 3 members of the MYC family in mammalian cells, c-MYC, MYCN and L-MYC. They interact with DNA through a consensus sequence called the E-box (CANNTG) and in concert with the partner MAX facilitate gene transcription. The importance of MYC in cell biology is vast. MYC controls key cellular processes including proliferation, survival and metabolism. Critical to this proposal is the fact that one of the genes positively regulated by MYC is the catalytic subunit of the telomerase complex, TERT . A recent study has shown that genomic rearrangements near the TERT gene are frequent in high-risk neuroblastomas and activation of TERT is particularly frequent in MYCN amplified tumours. These results are supported by our own analysis suggesting that the expression of TERT is significantly predictive of poor prognosis in multiple neuroblastoma datasets.
The central objective of the study is to verify whether inhibition of TERT by a clinically viable inhibitor called Imetelstat and using Fluoxetine-Prozac to inhibit the MYCN signalling network can cause synergistic killing of high-risk, MYCN amplified neuroblastoma cells. In vitro studies: neuroblastoma cell lines will be exposed to increasing concentrations of Imetelstat, Prozac and drug combinations for 24-72 hours and the IC50 will be calculated using MTT/MTS assays. The different cell lines will also be subjected to immunofluorescence analysis with activated caspase-3 antibody, propidium iodide staining and FACS analysis to determine the cell cycle and apoptosis status. In vivo studies: it will be verified if Imetelstat can cause regression of human neuroblastoma tumours transplanted into immunocompromised mice.
Metastatic neuroblastoma is a cancer of the nervous system that kills the majority of the affected children. Although there have been tremendous advances in the understanding of the molecular causes of the disease, patients are still treated with highly toxic mixtures of chemicals and radiation. Thus, safer and innovative approaches for this cancer are urgently needed. We propose to evaluate the efficacy of antibodies for neuroblastoma therapeutics. Antibodies are proteins that can bind to cells, triggering their death by engaging the immune system. Antibodies can also be engineered to carry cancer-killing molecules that are delivered specifically to tumour cells while sparing normal tissues. The aim of the project is to pre-clinically validate a newly developed monoclonal antibody for neuroblastoma therapeutics. The anti-LGR5 antibody (called BNC101) has been originally developed to target colon cancer stem cells. We will use the reagent in experiments whose primary objective is to evaluate the ability of the antibody to induce cancer regression in vitro and in vivo, especially in the context of patient-derived xenografts (PDX). A secondary aim is to clarify mechanisms of action, safety and identify biomarkers that predict clinical response. The project is ideally suited to a student with an MD or first or upper second undergraduate degree in cell biology, molecular biology or pharmacology. Familiarity with cancer cell cultures and basic biochemical and molecular techniques is also essential.