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Zhao, Zi-Ruo (2023) Development of an EGFR-targeted Drug Delivery System for Glioblastoma. MRes thesis, University of Nottingham.

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Abstract

Glioblastoma (GB) is one of the most malignant cancers with a dismal survival of 4 months when no treatment intervenes. After standard of care: surgery, radiotherapy and temozolomide (TMZ) chemotherapy, its median survival has been increased to 16 months. TMZ functions by methylating guanine of GB DNA into O6-methylguanine, which triggers a futile cycle of base excision-reinsertion conducted by DNA mismatch repair mechanism (MMR), eventually leading to DNA-strands breakdown and cell apoptosis. However, TMZ efficacy is largely limited by factors such as poor in-brain accumulation due to low level permeation across the blood-brain barrier, and O6-methylguanine-DNA methyltransferase (MGMT) which reverses the guanine methylation. The natural transportation protein apoferritin (AFt) shapes as a spherical hollow nanocage and has been proven to be a biocompatible drug delivery vehicle. It possesses 3-4 Å surface channels available for the entrance of iron ions and small molecules and allows modification at its N-termini on the exterior surface with different moieties. The overexpression of epidermal growth factor receptor (EGFR) is often observed in GB and can be seen as a biomarker; while the anti-EGFR affibodies, which are small affinity molecules of strong binding-affinity towards specific target, have been developed for many research purposes, an EGFR-affibody directed AFt delivery system is proposed to promote the performance of TMZ against GB.

This study aimed to exploit E. coli protein expression to produce recombinant human heavy chain AFt (human H-AFt) and its derivative with 24 EGFR-affibodies fused at N-termini (EGFRa@AFt) to encapsulate TMZ molecules (~500/cage), constructing the drug delivery systems AFt-TMZ and EGFRa@AFt-TMZ. Upon MTT viability assays (72 h), against EGFR-overexpressed GB cell line U373V, EGFRa@AFt-TMZ system exhibited significantly enhanced growth inhibition with GI50 of 8.18±1.67 pM, which was lower than AFt-TMZ system and free TMZ molecules by ~3 and 7 orders of magnitudes, respectively (P < 0.0001); while the growth of non-tumourigenic control human fibroblast MRC-5 cell line was negligibly compromised at concentrations < 20 nM. These suggested an in vitro targeting ability and enhanced drug activity of EGFRa@AFt-TMZ system. The similar responses between U373V and MGMT- transfected isogenic U373M cell line suggested its potential to overcome MGMT-mediated resistance to TMZ. However, the EGFR-overexpressed and MMR-deficient HCT-116 colon cancer cell line responded to both EGFRa@AFt-TMZ and AFt-TMZ systems at an equivalent level, suggesting that the effectiveness of TMZ might be limited by MMR level.

Item Type:	Thesis (University of Nottingham only) (MRes)
Supervisors:	Thomas, Neil Bradshaw, Tracey
Keywords:	Glioblastoma, oncology, cancer treatment, drug delivery systems
Subjects:	R Medicine > RC Internal medicine > RC 254 Neoplasms. Tumors. Oncology (including Cancer)
Faculties/Schools:	UK Campuses > Faculty of Science > School of Chemistry
Item ID:	72444
Depositing User:	ZHAO, Ziruo
Date Deposited:	26 Jul 2023 04:40
Last Modified:	26 Jul 2023 04:40
URI:	https://eprints.nottingham.ac.uk/id/eprint/72444

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