Finding specific treatments for cancer patients with mutations in the ribosome production/p53 pathway

Keywords: Molecular datasets and disease; Genetic influence on Health; Bioinformatics and personalised medicine
Summary:
BACKGROUND
Broad activity chemotherapeutics are used to treat multiple types of cancer. However, due to the complex and diverse genetic background underlying cancer types and individual tumours, this does not always work and a "personalised medicine" approach is now seen as a way forward.
The TP53 gene, which encodes the tumour suppressor p53, is mutated in more than half of all cancers. Many anti-cancer drugs function through activating p53, leading to cell cycle arrest, apoptosis or senescence.
The majority of anti-cancer chemotherapeutics, including commonly used drugs such as 5FU, are known to block ribosome production (Burger et al., JBC 2010). Sequencing of 12,000 cancers and 1001 cancer cell lines revealed that ribosomal protein (RP) genes are "cancer genes" in 12 of the 30 cancer types studied (including leukaemia, breast, lung and prostate cancers (Iorio et al., Cell 2016).
The primary supervisor's lab demonstrated that defects in ribosome production (e.g. RP gene mutation) leads to the activation of p53 (Sloan et al., Cell Reports 2013). It is, however, currently unclear how p53 activation caused by ribosome production defects would lead to cancer in patients carrying specific mutations in RP genes. Here, we therefore plan to characterise a selection of cancer cell lines, which faithfully recapitulate oncogenic alterations in tumours and carry known mutations in RP genes (Iorio et al., Cell 2016). This approach will allow us to investigate the effect of the mutations on p53 signalling and to determine the sensitivity of these cell lines to a range of chemotherapeutics.

PROJECT
We have selected a series of cancer cell lines containing stop codons/frame-shift mutations early in the ribosomal protein coding sequence to ensure a functional defect. The student will use a wide variety of approaches, including high-throughput screening, to address the following aims:
1) Use a combination of cellular, molecular biology and RNA analysis approaches to determine the impact of RP gene mutations on ribosome production, p53 signalling and cellular growth in the cancer cell lines (primary and secondary supervisors).
2) Use CRISPR/Cas9 editing (established in lab of secondary supervisor) to generate select RP gene cancer mutations in a control cell line (e.g. U2OS) to validate that this mutation, and not other variations in the cancer cell line, is responsible for changes in ribosome production and p53 signaling (secondary and primary supervisors).
3) Use high-throughput screening to test the hypothesis that mutations in RP genes will affect the response of these tumours to anti-cancer chemotherapeutics that impact ribosome production (primary and secondary supervisors and the High-Throughput Screening Facility).
4) Use chemicals (e.g. mTOR activators), to increase general RP production, and long non-coding RNAs (Sineups), to increase expression from the non-mutated RP gene, in cancer cell lines to determine whether this is a valid approach to treat tumours with RP gene mutations (primary supervisor).
This project will form the basis for future collaborations with the NICR including patient-associated material (e.g. Christine Harrison for leukaemia's) and the patient-derived xenograft (PDX) system.