Development of Gold Nanoparticles for Targeting PD-L1-Presenting Breast Cancers

Simple item page
dc.contributor.authorBlevins, Derek James
dc.contributor.supervisorWulff, Jeremy Earle
dc.date.accessioned2023-07-31T17:28:48Z
dc.date.available2023-07-31T17:28:48Z
dc.date.copyright2023en_US
dc.date.issued2023-07-31
dc.degree.departmentDepartment of Chemistry
dc.degree.levelDoctor of Philosophy Ph.D.en_US
dc.description.abstractOur goal was to selectively target the immunosuppressive programmed cell death protein ligand 1 (PD-L1) presenting cancer cells with gold nanoparticles (AuNP). Initially, small molecules, described as antagonists for the PD-1/PD-L1 interaction, were considered as promising targeting agents for the gold platform. The competitive binding profiles of these molecules were evaluated using surface plasmon resonance (SPR). However, we found that none of the small molecule candidates were capable of disrupting the interaction, despite their apparent cell-based efficacy in other literature. Our findings indicate that the molecules were being mischaracterized as immunomodulators directly blockading the PD-1/PD-L1 interaction. In addition, we found no evidence of direct binding with the small molecules to either PD-1 or PD-L1, indicating they would not be suitable candidates as targeting agents. Instead, a commercially available monoclonal antibody (mAb, αPD-L1; from BioXCell) was found as a suitable alternative for targeting and blockading PD-L1 directly from PD-1. The antibody was conjugated with α-lipoic acid (αLA) through its NHS-ester such that the thiolated mAb may be grafted to functionalize the gold cores. The degree of functionalization on the gold core was quantified in vitro using SPR, where the relationship between valency and molecular weights of the gold core and unbound mAb binding to PD-L1 was studied. This SPR method was primarily used further to optimize the AuNP formulations until they were deemed suitable for cellular work. Gold cores with a surface area coated in 25% αLA-mAb and 75% PEG2000 had shown a near maximal response of binding relative to the unbound mAb, indicating a high degree of functionalized nanoparticles. This formulation was then moved forward into cellular work with naïve human white blood cells (Jurkat) that were stimulated with PHA (to produce PD-1), and the stimulation suppressed by the presence of PD-L1. In addition to AuNP formulation, we sought to investigate controlled release mechanisms indicative of the reducing character of the tumour microenvironment. We developed a series of disulfide tethers with a fluorogenic dye that induces turn-on fluorescence upon disulfide exchange, expected to show selectivity for the higher concentration of glutathione. Instead, we found that the common, but necessary, additive, fetal bovine serum (FBS), was triggering the premature release of our tethered fluorophore, disrupting our intended controlled-release studies. Through systematic investigation, we found that unwanted turn-on fluorescence from our dithiodiacid tethers was ultimately occurring due to esterase activity found in the FBS. This activity was shut down when methyl groups were installed at the α-position to the carbonyl carbon. Future work may use these bulkier dithiodiacid tethers for selective release in other AuNP formulations. Immunotherapeutic efficacy of the functionalized AuNPs was investigated by showing consistent stimulation in the presence of immunosuppressive PD-L1. Naïve Jurkat cells were stimulated with phytohemagglutinin (PHA), where the cell density would significantly increase relative to basal cell growth, and in the presence of solubilized PD-L1, show no change in density attributed to T cell exhaustion. When AuNPs were present, the cell density would reflect that of “uninhibited” stimulation control, regardless of PD-L1 presence, indicating the potential immunotherapeutic benefit of recovery from immune exhaustion.en_US
dc.description.scholarlevelGraduateen_US
dc.identifier.bibliographicCitationBlevins, D. J., Hanley, R., Buldoc, T., Powell, D. A., Gignac, M., Walker, K., Carr, M. D., Hof, F., Wulff, J. E. ACS Med. Chem. Lett. 2019, 10 (8), 1187–1192.en_US
dc.identifier.bibliographicCitationBlevins, D. J., Nazir, R., Dabiri, S. M. H., Akbari, M., Wulff, J. E. J. Drug. Deliv. Sci. Tech. 2022, 78, 103950.en_US
dc.identifier.urihttp://hdl.handle.net/1828/15232
dc.languageEnglisheng
dc.language.isoenen_US
dc.rightsAvailable to the World Wide Weben_US
dc.subjectsurface plasmon resonance (SPR)en_US
dc.subjectgold nanoparticlesen_US
dc.subjectLigand bindingen_US
dc.subjectimmunostimulationen_US
dc.subjectimmunotherapyen_US
dc.subjectin vitro studiesen_US
dc.subjectassay developmenten_US
dc.subjectSPRen_US
dc.subjectAuNPen_US
dc.titleDevelopment of Gold Nanoparticles for Targeting PD-L1-Presenting Breast Cancersen_US
dc.typeThesisen_US

Files

Original bundle
Now showing 1 - 1 of 1
Thumbnail Image
Name:
Blevins_Derek_Chemistry_2023.pdf
Size:
8.69 MB
Format:
Adobe Portable Document Format
Description:
Dissertation
Download
License bundle
Now showing 1 - 1 of 1
No Thumbnail Available
Name:
license.txt
Size:
2 KB
Format:
Item-specific license agreed upon to submission
Description:
Download

Collections

Electronic Theses and Dissertations (ETD)
 
University of Victoria Libraries

We acknowledge and respect the Lək̓ʷəŋən (Songhees and Esquimalt) Peoples on whose territory the university stands, and the Lək̓ʷəŋən and W̱SÁNEĆ Peoples whose historical relationships with the land continue to this day.