{"id":{"repo_id":"buffalo","oai_identifier":"oai:ubir.buffalo.edu:10477/86722"},"canonical_url":"https://search.dev.ndltd.org/etd/buffalo/oai:ubir.buffalo.edu:10477/86722","repository":{"repo_id":"buffalo","name":"Buffalo","base_url":"https://ubir.buffalo.edu/oai/request"},"display":{"title":"Influence of Paracrine Signaling on Neoplastic Induction in Human Oligodendrocyte Progenitor Cells","abstract":"M.S.","abstract_html":"M.S.","abstract_has_math":false,"creators":["Pathak, Khushboo; 0000-0002-6564-6780"],"institution":"State University of New York at Buffalo","degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":["Sim, Fraser","Biological Sciences"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2025,"date_issued":"2025-02-21T21:36:51Z","date_published":"2025-02-21T21:36:51Z","updated_at":"2026-07-27T19:05:34Z","subjects":["neurosciences"],"languages":["eng"],"rights":["Users of works found in University at Buffalo Institutional Repository (UBIR) are responsible for identifying and contacting the copyright owner for permission to reuse. University at Buffalo Libraries do not manage rights for copyright-protected works and cannot assist with permissions.","Copyright retained by author."],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"http://hdl.handle.net/10477/86722","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Sim, Fraser","Biological Sciences"]},{"key":"dc:creator","label":"Author","values":["Pathak, Khushboo; 0000-0002-6564-6780"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2025-02-21T21:36:51Z","2020"]},{"key":"dc:publisher","label":"Institution","values":["State University of New York at Buffalo"]},{"key":"dc:type","label":"Dc Type","values":["Text","Thesis"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["neurosciences"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["eng"]},{"key":"dc:rights","label":"Dc Rights","values":["Users of works found in University at Buffalo Institutional Repository (UBIR) are responsible for identifying and contacting the copyright owner for permission to reuse. University at Buffalo Libraries do not manage rights for copyright-protected works and cannot assist with permissions.","Copyright retained by author."]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["http://hdl.handle.net/10477/86722"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["M.S.","Understanding the origin and cause of tumorigenesis is crucial in the development of new targeted drugs. Multiple mutations are usually required to drive neoplastic transformation which can lead to cancer. Glioblastoma (GBM) is considered to be one of the deadliest type of cancers. WHO classifies GBM as grade IV tumor and has a median survival rate of 12-15 months. It is also the most aggressive form of cancer and accounts for ~80% of malignant brain tumors. Glioma is the most frequent primary brain tumor in adults. While the precise cell of origin remains unknown, studies have shown that oligodendrocyte progenitor cells (OPCs) facilitate malignant glioma. Genomic studies of human glioma have identified three common signaling pathways that are often mutated in GBM. Mutations in three tumor suppressors, p53, PTEN and RB, represent the most common genes that are mutated in each pathway. Previous studies in our lab have shown that over-expression of presumptive dominant negative tumor suppressor (dnTS) proteins alters the morphology of human OPCs (hOPCs) cells and increases the capacity for self-renewal and rate of proliferation in vitro. However, the contribution of individual pathways to hOPC dysregulation and their necessity for tumorigenesis and malignant progression from hOPCs is still unknown. We first examined the effect of expression of individual dnTS proteins on the rate of hOPC proliferation. As shown previously, over-expression of all three dnTS proteins resulted in hyperproliferation of hOPCs in vitro. However, only dominant negative p53 (dnp53) expression was sufficient to induce cell-cycle entry and hyper-proliferation in human OPCs and this occurred regardless of growth factor supplementation. As expression level of dnTS proteins may not be sufficient to result in functional blockade, we decided to take a knockout approach using CRISPR/Cas9. As such, we cloned and produced lentiviral CRISPR/Cas9 vectors that targeted PTEN and RB. Upon transfection with the resultant plasmids, TLCv2-RB1 and TLCv2-PTEN, we were able to achieve induction of these plasmids in HEK 293T cells. We extracted genomic DNA and protein from the transfected cells for sequencing and to check the protein expression respectively. Genomic DNA sequencing and decreased protein expression, compared to the wild type, showed possible mutation in transfected HEK 293T cells. However, due to COVID restrictions, it was not possible to investigate the potential for CRISPR/Cas9-mediated mutation to induce neoplastic transformation by hOPCs. To summarize, we found that hOPCs proliferate at a higher rate when exposed to mutant p53 tumor suppressor gene and developed lentiviral vectors to inactive PTEN and RB1 in hOPCs. More research is required to determine the effectiveness of this approach and investigate the effects of combined inactivation in hOPCs.","**To request an accessible version of the file(s) associated with this item, contact library@buffalo.edu. Please include the item's persistent URL [http://hdl.handle.net/. . .] in your request.**"]},{"key":"dc:format","label":"Dc Format","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Influence of Paracrine Signaling on Neoplastic Induction in Human Oligodendrocyte Progenitor Cells"]}]}],"canonical_facts":{"dc:contributor":["Sim, Fraser","Biological Sciences"],"dc:creator":["Pathak, Khushboo; 0000-0002-6564-6780"],"dc:date":["2025-02-21T21:36:51Z","2020"],"dc:description":["M.S.","Understanding the origin and cause of tumorigenesis is crucial in the development of new targeted drugs. Multiple mutations are usually required to drive neoplastic transformation which can lead to cancer. Glioblastoma (GBM) is considered to be one of the deadliest type of cancers. WHO classifies GBM as grade IV tumor and has a median survival rate of 12-15 months. It is also the most aggressive form of cancer and accounts for ~80% of malignant brain tumors. Glioma is the most frequent primary brain tumor in adults. While the precise cell of origin remains unknown, studies have shown that oligodendrocyte progenitor cells (OPCs) facilitate malignant glioma. Genomic studies of human glioma have identified three common signaling pathways that are often mutated in GBM. Mutations in three tumor suppressors, p53, PTEN and RB, represent the most common genes that are mutated in each pathway. Previous studies in our lab have shown that over-expression of presumptive dominant negative tumor suppressor (dnTS) proteins alters the morphology of human OPCs (hOPCs) cells and increases the capacity for self-renewal and rate of proliferation in vitro. However, the contribution of individual pathways to hOPC dysregulation and their necessity for tumorigenesis and malignant progression from hOPCs is still unknown. We first examined the effect of expression of individual dnTS proteins on the rate of hOPC proliferation. As shown previously, over-expression of all three dnTS proteins resulted in hyperproliferation of hOPCs in vitro. However, only dominant negative p53 (dnp53) expression was sufficient to induce cell-cycle entry and hyper-proliferation in human OPCs and this occurred regardless of growth factor supplementation. As expression level of dnTS proteins may not be sufficient to result in functional blockade, we decided to take a knockout approach using CRISPR/Cas9. As such, we cloned and produced lentiviral CRISPR/Cas9 vectors that targeted PTEN and RB. Upon transfection with the resultant plasmids, TLCv2-RB1 and TLCv2-PTEN, we were able to achieve induction of these plasmids in HEK 293T cells. We extracted genomic DNA and protein from the transfected cells for sequencing and to check the protein expression respectively. Genomic DNA sequencing and decreased protein expression, compared to the wild type, showed possible mutation in transfected HEK 293T cells. However, due to COVID restrictions, it was not possible to investigate the potential for CRISPR/Cas9-mediated mutation to induce neoplastic transformation by hOPCs. To summarize, we found that hOPCs proliferate at a higher rate when exposed to mutant p53 tumor suppressor gene and developed lentiviral vectors to inactive PTEN and RB1 in hOPCs. More research is required to determine the effectiveness of this approach and investigate the effects of combined inactivation in hOPCs.","**To request an accessible version of the file(s) associated with this item, contact library@buffalo.edu. Please include the item's persistent URL [http://hdl.handle.net/. . .] in your request.**"],"dc:format":["application/pdf"],"dc:identifier":["http://hdl.handle.net/10477/86722"],"dc:language":["eng"],"dc:publisher":["State University of New York at Buffalo"],"dc:rights":["Users of works found in University at Buffalo Institutional Repository (UBIR) are responsible for identifying and contacting the copyright owner for permission to reuse. University at Buffalo Libraries do not manage rights for copyright-protected works and cannot assist with permissions.","Copyright retained by author."],"dc:subject":["neurosciences"],"dc:title":["Influence of Paracrine Signaling on Neoplastic Induction in Human Oligodendrocyte Progenitor Cells"],"dc:type":["Text","Thesis"]},"updated_at":"2026-07-27T19:05:34Z"}