Investigating the effect of peptide-functionalized gold nanoparticles on colon cancer cells
- Authors: Ramagoma, Rolivhuwa Bishop
- Date: 2023-12
- Subjects: Colon (Anatomy) -- Cancer -- Research , Colon (Anatomy) -- Cancer -- Treatment , Nanoparticles
- Language: English
- Type: Master's theses , text
- Identifier: http://hdl.handle.net/10948/62564 , vital:72824
- Description: Colon cancer like any other cancer is a condition in which cells grow uncontrollably and may even spread to other regions of the body through metastasis. Colon cancer was ranked the second leading cause of cancer related deaths worldwide in 2018. Research to advance treatment of cancer keeps advancing daily, However, a big challenge is drug-induced side effects due to toxicity against normal body cells. Therefore, the development of controlled release technologies in conjunction with targeted drug delivery may provide a more efficient and less dangerous approach to overcome the limits of traditional chemotherapy. Including the creation of nanoscale delivery vehicles capable of directing the release of chemotherapeutic drugs into cancer cells only. This study aims to investigate p14 peptide that would specifically target colorectal cancer cells and not normal body cells to develop a targeted drug delivery system using gold nanoparticles. This study serves as a pilot study of the primary aim. To achieve this, the effect of the peptide p14 and peptide functionalized gold nanoparticles (p14-AuNP) on colon cancer cells (HT-29) and normal epithelial cells (KMST-6) was determined. Firstly, gold nanoparticles were chemically synthesised and then functionalized with p14 peptide through Polyethylene glycol. Then assessment of their effect through in vitro cytotoxicity assay (MTT) and gene expression analysis (RT-qPCR) was conducted. Nanoparticles’ synthesis and functionalization was performed and confirmed: In vitro cytotoxicity through MTT assay was successfully conducted and p14-AuNP showed toxicity against colon cancer cells and lesser toxicity towards normal cells as compared to 5-Flourouracil (commercially approved drug for colon cancer treatment). Gene expression analysis revealed that apoptosis was induced in both cell lines by p14-AuNP either through upregulation of caspase 3, 7 and/or BCL2. A cell survival gene, AKT1, also had significant effect on this. CDC42 was downregulated which indicates that cell proliferation was inhibited. , Thesis (MSc) -- Faculty of Science, School of Biomolecular & Chemical Sciences, 2023
- Full Text:
- Date Issued: 2023-12
- Authors: Ramagoma, Rolivhuwa Bishop
- Date: 2023-12
- Subjects: Colon (Anatomy) -- Cancer -- Research , Colon (Anatomy) -- Cancer -- Treatment , Nanoparticles
- Language: English
- Type: Master's theses , text
- Identifier: http://hdl.handle.net/10948/62564 , vital:72824
- Description: Colon cancer like any other cancer is a condition in which cells grow uncontrollably and may even spread to other regions of the body through metastasis. Colon cancer was ranked the second leading cause of cancer related deaths worldwide in 2018. Research to advance treatment of cancer keeps advancing daily, However, a big challenge is drug-induced side effects due to toxicity against normal body cells. Therefore, the development of controlled release technologies in conjunction with targeted drug delivery may provide a more efficient and less dangerous approach to overcome the limits of traditional chemotherapy. Including the creation of nanoscale delivery vehicles capable of directing the release of chemotherapeutic drugs into cancer cells only. This study aims to investigate p14 peptide that would specifically target colorectal cancer cells and not normal body cells to develop a targeted drug delivery system using gold nanoparticles. This study serves as a pilot study of the primary aim. To achieve this, the effect of the peptide p14 and peptide functionalized gold nanoparticles (p14-AuNP) on colon cancer cells (HT-29) and normal epithelial cells (KMST-6) was determined. Firstly, gold nanoparticles were chemically synthesised and then functionalized with p14 peptide through Polyethylene glycol. Then assessment of their effect through in vitro cytotoxicity assay (MTT) and gene expression analysis (RT-qPCR) was conducted. Nanoparticles’ synthesis and functionalization was performed and confirmed: In vitro cytotoxicity through MTT assay was successfully conducted and p14-AuNP showed toxicity against colon cancer cells and lesser toxicity towards normal cells as compared to 5-Flourouracil (commercially approved drug for colon cancer treatment). Gene expression analysis revealed that apoptosis was induced in both cell lines by p14-AuNP either through upregulation of caspase 3, 7 and/or BCL2. A cell survival gene, AKT1, also had significant effect on this. CDC42 was downregulated which indicates that cell proliferation was inhibited. , Thesis (MSc) -- Faculty of Science, School of Biomolecular & Chemical Sciences, 2023
- Full Text:
- Date Issued: 2023-12
An investigation into the localization of peptide-gold nanoparticles in an in vitro and in vivo colorectal cancer model
- Authors: Cairncross, Lynn
- Subjects: Colon (Anatomy) -- Cancer -- Treatment , Cancer -- Early detection
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:10351 , http://hdl.handle.net/10948/d1020921
- Description: Background: Colorectal cancer is the third most common cancer and cause of related deaths worldwide. Early colorectal cancer diagnosis is vital in reducing incidence and mortality. There is a need for the development of non-invasive screening tools for enhancing the detection of the disease. Cancer specific peptides are useful cancer targeting agents that can be used to specifically improve early detection strategies. Several cancer targeting peptides have been identified. Previous work investigated the specific binding of three of these peptides (p.C, p.L and p.14) conjugated to quantum dots and were found to bind to colorectal cancer cell lines (HT-29 and Caco-2). However, their uptake, localization and biodistribution in an in vitro and in vivo colorectal cancer model have not been determined. This is essential in gaining an understanding for future diagnostic or therapeutic based applications. Primary Aim: The aim of this study was investigate the localization of three selected peptides p.C, p.L and p.14 conjugated to gold nanoparticles in an in vitro and in vivo colorectal cancer model using HRTEM. Methodology: The AuNP/peptide conjugates were characterized by HRTEM and DLS. For in vitro studies; HT-29, Caco-2 and C3A cells were exposed to the AuNP-p.C, AuNP-p.L and AuNP-p.14, collected and processed for HRTEM to assess targeting and localization. For in vivo studies; the establishment of a colorectal cancer model using the AOM/DSS model 1 and 2 was conducted. Wistar rats were assigned to 6 groups, five experimental and 1 control group. Group 1 received AOM/DSS method 1 and was treated with AuNP-p.L. Group 2 and 3 received AOM/DSS method 2 and were treated with AuNP-p.C and AuNP-p.14. Group 4 and 5 remained healthy and treated with AuNP-p.C and AuNP-p.14. Group 6 remained healthy receiving no nanoparticle treatment. After treatment, rats were sacrificed and tissue was processed for HRTEM. Tissue chosen for HRTEM analysis included: Group 1 (inflamed colon, rectum, pancreatic and kidney), Group 4 (kidney) and Group 5 (liver). Results: results obtained from nanoparticle characterization suggested that nanoparticles were conjugated to their respective peptides and were stable in dispersion. For in vitro studies, results suggested no AuNP targeting and localization in HT-29 cell lines. For in vivo studies, no colorectal cancer tumours were induced. TEM micrographs did not indicate the presence of nanoparticles in colon, rectum, pancreatic, kidney and liver tissue. However, AuNPs were found in the kidney tissue (group 4). Conclusion: Although the overall objectives were not met, this study provided insight into TEM cell preparation and optimization for future nanoparticle cell interaction research. This study also demonstrated the absence of AuNPs in healthy tissue and the presence of AuNPs in healthy kidney tissue through renal clearance, a favourable quality for diagnostic or therapeutic applications.
- Full Text:
- Authors: Cairncross, Lynn
- Subjects: Colon (Anatomy) -- Cancer -- Treatment , Cancer -- Early detection
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:10351 , http://hdl.handle.net/10948/d1020921
- Description: Background: Colorectal cancer is the third most common cancer and cause of related deaths worldwide. Early colorectal cancer diagnosis is vital in reducing incidence and mortality. There is a need for the development of non-invasive screening tools for enhancing the detection of the disease. Cancer specific peptides are useful cancer targeting agents that can be used to specifically improve early detection strategies. Several cancer targeting peptides have been identified. Previous work investigated the specific binding of three of these peptides (p.C, p.L and p.14) conjugated to quantum dots and were found to bind to colorectal cancer cell lines (HT-29 and Caco-2). However, their uptake, localization and biodistribution in an in vitro and in vivo colorectal cancer model have not been determined. This is essential in gaining an understanding for future diagnostic or therapeutic based applications. Primary Aim: The aim of this study was investigate the localization of three selected peptides p.C, p.L and p.14 conjugated to gold nanoparticles in an in vitro and in vivo colorectal cancer model using HRTEM. Methodology: The AuNP/peptide conjugates were characterized by HRTEM and DLS. For in vitro studies; HT-29, Caco-2 and C3A cells were exposed to the AuNP-p.C, AuNP-p.L and AuNP-p.14, collected and processed for HRTEM to assess targeting and localization. For in vivo studies; the establishment of a colorectal cancer model using the AOM/DSS model 1 and 2 was conducted. Wistar rats were assigned to 6 groups, five experimental and 1 control group. Group 1 received AOM/DSS method 1 and was treated with AuNP-p.L. Group 2 and 3 received AOM/DSS method 2 and were treated with AuNP-p.C and AuNP-p.14. Group 4 and 5 remained healthy and treated with AuNP-p.C and AuNP-p.14. Group 6 remained healthy receiving no nanoparticle treatment. After treatment, rats were sacrificed and tissue was processed for HRTEM. Tissue chosen for HRTEM analysis included: Group 1 (inflamed colon, rectum, pancreatic and kidney), Group 4 (kidney) and Group 5 (liver). Results: results obtained from nanoparticle characterization suggested that nanoparticles were conjugated to their respective peptides and were stable in dispersion. For in vitro studies, results suggested no AuNP targeting and localization in HT-29 cell lines. For in vivo studies, no colorectal cancer tumours were induced. TEM micrographs did not indicate the presence of nanoparticles in colon, rectum, pancreatic, kidney and liver tissue. However, AuNPs were found in the kidney tissue (group 4). Conclusion: Although the overall objectives were not met, this study provided insight into TEM cell preparation and optimization for future nanoparticle cell interaction research. This study also demonstrated the absence of AuNPs in healthy tissue and the presence of AuNPs in healthy kidney tissue through renal clearance, a favourable quality for diagnostic or therapeutic applications.
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