Bromma, Kyle W2023-04-192023-04-1920232023-04-19http://hdl.handle.net/1828/14958One of the major issues in cancer radiotherapy (RT) is normal tissue toxicity. Introduction of radiosensitizers like gold nanoparticles (GNPs) into cancer cells to enhance the local RT dose is a promising technique that is being explored. However, a large portion of experimentation involving GNPs has been done in simple two-dimensional (2D) monolayer models that cannot properly encapsulate the complex heterogeneous interactions that occur in vivo. By introducing an in vitro three-dimensional (3D) model that better mimics the tumour microenvironment (TME), we can more rapidly facilitate a quicker translation of various treatment technologies like GNPs to the clinic. Further, clinical trials show that the chemotherapy drug docetaxel (DTX) given in conjunction with RT can improve survival in high-risk cancers. Addition of GNPs to this current DTX/RT protocol is expected to further improve therapeutic benefits. Elucidation of a combined therapy of GNPs, DTX, and RT to optimize treatment can better improve patient outcome and reduce normal tissue toxicity by specifically targeting tumours and is completely novel research. The work in this dissertation explores the application of GNPs to various elements that are present in a TME. Many cell types are present in TME and contribute in different ways to the proliferation of cancer. One of these cell lines, cancer associated fibroblasts (CAFs), which can promote tumour growth and metastasis, was compared to cancer epithelial cells and normal fibroblasts (FBs). Hence, we used FBs and CAFs to evaluate the differences in GNP uptake and resulting radiation induced damage. It was found that the CAFs had a much larger uptake of GNPs relative to the other cells, with on average 265% more GNPs relative to cervical cancer cells while FBs had only 7.55% the uptake of the tumour cells and 2.87% the uptake of CAFs. This translated to increases in 53BP1-related DNA damage foci in CAFs (13.5%) and tumour cells (9.8%) along with FBs (8.8%), compared to control with RT treatment. This difference in DNA damage due to selective targeting of cancer associated cells over normal cells may allow GNPs to be an effective tool in future cancer RT to battle normal tissue toxicity while improving local RT dose to the tumour. To expedite a quicker clinical translation, 3D tumor spheroid models were optimized and compared to 2D monolayer. The uptake of various sizes of GNPs was tested on monolayer and spheroids to evaluate the differences between a 2D and 3D model in similar conditions. Moreover, combined treatment of GNPs with DTX was introduced and how they effect the uptake of the GNPs was elucidated.iv In the 2D monolayer model, the addition of DTX induced a small increase of uptake of GNPs of between 13% and 24%, while in the 3D spheroid model, DTX increased uptake by between 47% and 186%. It was observed that the more complex spheroid, which introduces an extracellular matrix, had larger uptake and penetration of smaller GNPs (15 nm) relative to larger GNPs (50 nm). Moreover, while the addition of DTX had a beneficial effect on the uptake of GNPs into cells, it also synchronized the cells into a radiosensitive cell cycle phase. This translated to a larger effect when radiation was introduced, in a combined treatment modality with GNP, DTX, and RT. In spheroids, the addition of GNPs to the treatment regime decreased the surviving tumour cells by 16-32% compared to samples not treated with GNPs. Further, the addition of DTX seems to synergistically increase damage in some cancer cell lines. This work highlights the necessity to optimize GNP treatment conditions in a more realistic tumor-like environment. A 3D spheroid model can capture important details which are absent from a simple 2D monolayer model.enAvailable to the World Wide WebGold nanoparticlesRadiotherapySpheroidsCancerNanotechnologyThesisBromma, K., Cicon, L., Beckham, W., Chithrani, DB., Gold nanoparticle mediated radiation response among key cell components of the tumour microenvironment for the advancement of cancer nanotechnology. Scientific Reports. https://doi.org/10.1038/s41598-020-68994-0 (2020)Bromma, K., Alhussan, A., Perez, M.M., Howard, P., Beckham, W., Chithrani, D.B., Three‑Dimensional Tumor Spheroids as a Tool for Reliable Investigation of Combined Gold Nanoparticle and Docetaxel Treatment. Cancers. https://doi.org/10.3390/cancers13061465 (2021)Bromma, K., Dos Santos, N., Bartra, I., Alexander, A., Beckham, W., Krishnan, S., Chithrani, D.B., Enhancing nanoparticle accumulation in two dimensional, three dimensional, and xenograft mouse cancer cell models in the presence of docetaxel. Scientific Reports. https://doi.org/10.1038/s41598-022-17752-5 (2022)