Targeting the phosphatidylinositol-3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways to enhance chemoradiotherapy responsiveness in colorectal cancer
thesisposted on 25.01.2021, 14:36 authored by Aoife Carr
Purpose: Late stage colorectal cancer (CRC) is currently treated with neoadjuvant chemoradiation therapy (NACRT), however, responses are not uniform. The PI3K and MAPK pathways have been implicated in tumourigenesis, poor patient outcomes and resistance to chemoradiation therapy (CRT).
Methods: Whole-exome sequencing and mutational analysis were carried out on pre-treatment tumour biopsies from locally advanced rectal cancer (LARC) patients (n=26). LARC patients (n=66) were monitored for circulating tumour cells (CTCs) and mutational variations in blood and tissue samples taken before, during and post-NACRT. A panel of PI3K/MAPKmut CRC cell lines were treated with the PI3K inhibitor copanlisib, and/ MEK inhibitor refametinib in proliferation assays; and treated with various combinations of radiation, 5-FU chemotherapy, copanlisib and/or refametinib in clonogenic assays. BALB-C mice were implanted with CRC cell lines (n=4) and treated with copanlisib, and / CRT, and monitored for tumour growth.
Results: Whole-exome sequencing detected PI3K and MAPK pathway mutations in 88.5% of samples, including KRAS (15.4%), BRAF (11.5%) and PIK3CA (11.5%); with BRAF and PIK3CA mutations identified exclusively in non-responders. CTCs increased during CRT, with CTCs detected in 40% and 38.1% of samples at week 3 and final week of CRT respectively, compared to 17.1% of pre-treatment samples. In vitro, copanlisib and refametinib were most effective in the PIK3CAmut (IC50=28nm), and KRASmut (IC50=36nm) cell lines respectively. The copanlisib refametinib combination resulted in a synergistic response in 8/10 cell lines. In 3D clonogenic assays, CRC cell growth was significantly reduced when treated with copanlisib CRT (KRASmut), refametinib CRT (KRASmut) and copanlisib refametinib CRT (WT, KRASmut, BRAFmut and PIK3CA KRASmut) compared to CRT alone. In vivo, copanlisib-CRT significantly reduced tumour growth and increased overall survival in KRASmut, PIK3CAmut and WT xenografts compared to untreated controls.
Conclusion: Our results suggest that kinase signalling pathway mutations may modulate treatment responsiveness and clinical outcomes in CRC. Furthermore, treatment with PI3K/MEK inhibitors may enhance CRC patient responsiveness to CRT.
First SupervisorProf. Bryan Hennessy
Second SupervisorDr. Sinead Toomey
Third SupervisorDr. Simon Furney
Fourth SupervisorProf. Elaine Kay & Dr. Brian O’Neill
CommentsA thesis submitted for the degree of Doctor of Philosophy from the Royal College of Surgeons in Ireland in 2020.
Published CitationCarr A. Targeting the phosphatidylinositol-3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways to enhance chemoradiotherapy responsiveness in colorectal cancer [PhD Thesis]. Dublin: Royal College of Surgeons in Ireland; 2020
Date of award31/05/2020
- Doctor of Philosophy (PhD)