Posters
Presentation Type
Poster
Discipline Track
Biomedical Science
Abstract Type
Research/Clinical
Abstract
Background: Glioblastoma is the most common malignant brain tumor characterized by its aggressive and highly proliferative nature. Although GBM rarely metastasizes to distant organs, it presents unique characteristics that challenge chemo- and radio-therapeutic efforts. Previous studies have attributed glioblastoma's prominent level of resistance to its involvement in multiple signaling pathways. To counter this, current promising treatments, such as Niclosamide and Camptothecin (CPT), have been evaluated and subsequently combined for their synergistic effect, targeting the multiple pathways that glioblastoma upregulates. The current study provides further insight into the duration of effect in with the usage of such promising treatments. The lasting effects the treatment has on the U87 cells will show the more long-term effects of Niclosamide, CPT, and combined treatments have on the cells. It is important to understand how U87 cells recover post-treatment and how long the treatment lasts at inducing cell cytotoxicity.
Methods: Western blot analysis investigated the different components of the suspected pathways affected by Niclosamide and CPT. Long lasting effects of Niclosamide and CPT were studied after 24 hours post individual or combined treatment on cell cycle pathways, autophagy pathway, ER stress pathway, ubiquitin pathway, and MAPK signaling pathway. Cells with each corresponding treatment group: Control, CPT, Niclosamide, and a combination of CPT and Niclosamide, were homogenized and assessed for protein concentration with BCA. 20-30 μg of total protein were then loaded on 4-20% SDS-PAGE, transferred onto nitrocellulose membranes, and blocked with 5% BSA or non-fat dried milk dissolved in TBST. The membranes were then incubated overnight at 4°C with their corresponding primary antibody, washed the next day with TBST, incubated for 1 hour at room temperature with HRP-conjugated secondary antibodies. Protein bands were then imaged with iBright 1500 and quantified using the ImageJ software.
Results: The present study results demonstrate that 24-hour recovery from treatment with CPT, Niclosamide, and combined treatment continue to induce levels of cell apoptosis similar to what has been previously reported. Western blot analysis shows that the methods of cell apoptosis that contribute to cell death 24-hours post removal of the drug include ER stress, cell cycle regulation, and autophagy. Additionally, the 24-hour recovery from treatment continues to show that Niclosamide induced major cell death through inhibition of pro-survival signaling pathways, ER stress and autophagy. Furthermore, CPT induces cell apoptosis through activation of pro-apoptotic cell signaling pathways.
Conclusions: The findings of the study suggest that post 24-hour recovery from the combined treatment of Niclosamide and CPT continue to suppress cell cycle survival and proliferation. The synergism of Niclosamide with CPT helps induce at least a 24-hour long effect in inducing cell cytotoxicity. These results provide new insight into the long-lasting effects of Niclosamide, CPT, and combined treatments on U87 cells.
Academic/Professional Position
Medical Student
Mentor/PI Department
Neuroscience
Recommended Citation
Fang, X.; Cheng, B.; Lara, R.; Osho, J. B.; and Fonseca, Josue, "Investigating the Post-Treatment Mechanisms underlying Prolonged Effect of Niclosamide and Camptothecin on Glioblastoma Cells" (2024). Research Symposium. 65.
https://scholarworks.utrgv.edu/somrs/2023/posters/65
Included in
Investigating the Post-Treatment Mechanisms underlying Prolonged Effect of Niclosamide and Camptothecin on Glioblastoma Cells
Background: Glioblastoma is the most common malignant brain tumor characterized by its aggressive and highly proliferative nature. Although GBM rarely metastasizes to distant organs, it presents unique characteristics that challenge chemo- and radio-therapeutic efforts. Previous studies have attributed glioblastoma's prominent level of resistance to its involvement in multiple signaling pathways. To counter this, current promising treatments, such as Niclosamide and Camptothecin (CPT), have been evaluated and subsequently combined for their synergistic effect, targeting the multiple pathways that glioblastoma upregulates. The current study provides further insight into the duration of effect in with the usage of such promising treatments. The lasting effects the treatment has on the U87 cells will show the more long-term effects of Niclosamide, CPT, and combined treatments have on the cells. It is important to understand how U87 cells recover post-treatment and how long the treatment lasts at inducing cell cytotoxicity.
Methods: Western blot analysis investigated the different components of the suspected pathways affected by Niclosamide and CPT. Long lasting effects of Niclosamide and CPT were studied after 24 hours post individual or combined treatment on cell cycle pathways, autophagy pathway, ER stress pathway, ubiquitin pathway, and MAPK signaling pathway. Cells with each corresponding treatment group: Control, CPT, Niclosamide, and a combination of CPT and Niclosamide, were homogenized and assessed for protein concentration with BCA. 20-30 μg of total protein were then loaded on 4-20% SDS-PAGE, transferred onto nitrocellulose membranes, and blocked with 5% BSA or non-fat dried milk dissolved in TBST. The membranes were then incubated overnight at 4°C with their corresponding primary antibody, washed the next day with TBST, incubated for 1 hour at room temperature with HRP-conjugated secondary antibodies. Protein bands were then imaged with iBright 1500 and quantified using the ImageJ software.
Results: The present study results demonstrate that 24-hour recovery from treatment with CPT, Niclosamide, and combined treatment continue to induce levels of cell apoptosis similar to what has been previously reported. Western blot analysis shows that the methods of cell apoptosis that contribute to cell death 24-hours post removal of the drug include ER stress, cell cycle regulation, and autophagy. Additionally, the 24-hour recovery from treatment continues to show that Niclosamide induced major cell death through inhibition of pro-survival signaling pathways, ER stress and autophagy. Furthermore, CPT induces cell apoptosis through activation of pro-apoptotic cell signaling pathways.
Conclusions: The findings of the study suggest that post 24-hour recovery from the combined treatment of Niclosamide and CPT continue to suppress cell cycle survival and proliferation. The synergism of Niclosamide with CPT helps induce at least a 24-hour long effect in inducing cell cytotoxicity. These results provide new insight into the long-lasting effects of Niclosamide, CPT, and combined treatments on U87 cells.