Targeting AKT/mTOR pathway in order to sensitise rectal tumour cells to irradiation using patient derived organoids as a model

Stodolna, Agata Anna (2022). Targeting AKT/mTOR pathway in order to sensitise rectal tumour cells to irradiation using patient derived organoids as a model. University of Birmingham. Ph.D.

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Abstract

Treatment resistance is one of the key contributors to low overall survival rates in Colorectal cancer (CRC) patients. Although both resistance to chemotherapy and radiotherapy is observed in CRC patients, radioresistance of rectal cancer patients is a major issue in the clinic. 40% of patients with rectal cancer that receive neo-adjuvant radiotherapy will have no significant response to treatment, whereas only 10% of patients will have a complete pathological response (pathCR) to treatment. In order to understand pathCR radiosensitivity markers need to be defined. Studying radiotherapy with current models has been challenging due to the nature of the available models and their limitations to recapitulate the patients’ tumour biology. The emerging new 3 dimensional (3D) organoid models show promise in regards to the modelling of cancer and patients’ response with in vitro experiments that will be able to recapitulate tumour microenvironment. The aim of this study was to establish an organoid model for studying irradiation in rectal cancer patients and to identify radiosensitivity and radioresistance drivers in order to understand and tackle the problem of resistance to radiotherapy in clinical patients. Organoid lines were established and characterised for pathological and molecular features. The organoids were subjected to short-course radiotherapy (25Gy dose delivered in 5 fractions over the course of 5 days) and the response to the treatment was measured. Furthermore, the changes caused by the irradiation were investigated by performing whole genome sequencing, DNA methylation arrays, total RNA sequencing, and single-cell sequencing on irradiated and control organoids. Lastly, combination drug and radiotherapy assays were performed using organoids with mTOR and Akt inhibitors in order to sensitise cells to irradiation. The results revealed that irradiation causes changes on a genome-wide scale and disrupts the mTOR/PiK3CA signalling pathway. Combination therapy showed that Rapamycin is not effective in sensitising cells to irradiation, whereas AZD2014 was able to sensitise certain organoid lines to irradiation; the same was found to be the case for MK-2206. In conclusion, the results showed organoids pose as representative models for modelling radiotherapy response, and that blocking mTOR via dual inhibition of mTORC1 and mTORC2 as well as inhibition of Akt can sensitise cells to irradiation. Finally it was found that, in order to sensitise resistant lines, the dual inhibition of mTOR/Akt might be required.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):