{"id":{"repo_id":"wayne-thes","oai_identifier":"oai:digitalcommons.wayne.edu:oa_dissertations-2204"},"canonical_url":"https://search.dev.ndltd.org/etd/wayne-thes/oai:digitalcommons.wayne.edu:oa_dissertations-2204","repository":{"repo_id":"wayne-thes","name":"Wayne State University","base_url":"https://digitalcommons.wayne.edu/do/oai/"},"display":{"title":"Retroviral-mediated gene transfer of Il-1β?cDNA into human activated T cells","abstract":"Autologous activated T cells (ATC) are being utilized in clinical trials as potential mediators of anti-tumor cytotoxicity in refractory cancer patients who suffer from hematologic malignancies or solid tumors. ATC are readily generated from peripheral blood and possess non-major histocompatibility complex (non-MHC) restricted killing properties. ATC are also powerful targets for genetic modification and hold promise for treatments of cancer, AIDS and T cell disorders. Retrovirus-mediated gene transfer and the fate of proviral gene expression were evaluated in human T cells using 1) immobilized anti-CD3 monoclonal antibody (mAb) plus interleukin-2 (IL-2), or 2) cis costimulation using beads carrying co-immobilized anti-CD3 and anti-CD28 mAbs. By cross-linking the CD3 and CD28 receptors, these mAbs mimic in vivo signaling events, leading to cytokine production and proliferation. A modified human interleukin-1β (IL-Iβ) cDNA inserted into the MFG retroviral vector served as an indicator gene in these studies. Optimized methodologies resulted in T cell transduction frequencies of approximately 50-75%. An important consideration for the success of T lymphocyte-based gene therapy is the relationship between T cell activity and the level of proviral gene expression. Early after mAb stimulation and virus exposure, proviral gene expression was greater at the RNA and protein levels in optimized anti-CD3/anti-CD28 bead-activated cultures, corresponding with augmented endogenous cytokine responses and mitogenesis. Proviral gene expression was not regulated by extrinsic cellular factors present in activated T cell supernatants. Regardless of the mAb stimulation method, proviral IL-lβ expression declined in long-term cultures concomitant with a decrease in cellular cytokines. Restimulation of transduced T cells by either mAb method reinduced both T cell activity and vector expression. The finding that proviral gene regulation is downmodulated in the absence of T cell signaling events suggests that in vivo, high-level proviral gene expression will not be maintained. To overcome the loss of activity in gene-modified T lymphocytes, we have applied a bispecific antibody (BsAb) to redirect cytotoxicity to tumor and enhance T cell activity. For successful cancer adoptive immunotherapy, a coordinated approach of BsAb arming of retrovirus-gene-modified T cells can be utilized for delivery of beneficial, tumoricidal molecules to the tumor site.","abstract_html":"Autologous activated T cells (ATC) are being utilized in clinical trials as potential mediators of anti-tumor cytotoxicity in refractory cancer patients who suffer from hematologic malignancies or solid tumors. ATC are readily generated from peripheral blood and possess non-major histocompatibility complex (non-MHC) restricted killing properties. ATC are also powerful targets for genetic modification and hold promise for treatments of cancer, AIDS and T cell disorders. Retrovirus-mediated gene transfer and the fate of proviral gene expression were evaluated in human T cells using 1) immobilized anti-CD3 monoclonal antibody (mAb) plus interleukin-2 (IL-2), or 2) cis costimulation using beads carrying co-immobilized anti-CD3 and anti-CD28 mAbs. By cross-linking the CD3 and CD28 receptors, these mAbs mimic in vivo signaling events, leading to cytokine production and proliferation. A modified human interleukin-1β (IL-Iβ) cDNA inserted into the MFG retroviral vector served as an indicator gene in these studies. Optimized methodologies resulted in T cell transduction frequencies of approximately 50-75%. An important consideration for the success of T lymphocyte-based gene therapy is the relationship between T cell activity and the level of proviral gene expression. Early after mAb stimulation and virus exposure, proviral gene expression was greater at the RNA and protein levels in optimized anti-CD3/anti-CD28 bead-activated cultures, corresponding with augmented endogenous cytokine responses and mitogenesis. Proviral gene expression was not regulated by extrinsic cellular factors present in activated T cell supernatants. Regardless of the mAb stimulation method, proviral IL-lβ expression declined in long-term cultures concomitant with a decrease in cellular cytokines. Restimulation of transduced T cells by either mAb method reinduced both T cell activity and vector expression. The finding that proviral gene regulation is downmodulated in the absence of T cell signaling events suggests that in vivo, high-level proviral gene expression will not be maintained. To overcome the loss of activity in gene-modified T lymphocytes, we have applied a bispecific antibody (BsAb) to redirect cytotoxicity to tumor and enhance T cell activity. For successful cancer adoptive immunotherapy, a coordinated approach of BsAb arming of retrovirus-gene-modified T cells can be utilized for delivery of beneficial, tumoricidal molecules to the tumor site.","abstract_has_math":false,"creators":["Ruhl, Elizabeth"],"institution":null,"degree_name":"Ph.D.","degree_level":"Open Access Dissertation","degree_discipline":"Molecular Biology and Genetics","degree_department":null,"school":null,"contributors":["Katrina T. Trevor, Ph.D."],"advisors":[],"committee_chairs":[],"committee_members":[],"year":1998,"date_issued":"1998-01-01T08:00:00Z","date_published":"1998-01-01T08:00:00Z","updated_at":"2026-07-24T06:00:09Z","subjects":[],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://digitalcommons.wayne.edu/oa_dissertations/1205","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Katrina T. Trevor, Ph.D."]},{"key":"dc:creator","label":"Author","values":["Ruhl, Elizabeth"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.available","label":"Dc Date Available","values":["2015-10-01T07:00:00Z"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Molecular Biology and Genetics"]},{"key":"thesis:degree_level","label":"Degree Level","values":["Open Access Dissertation"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Ph.D."]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://digitalcommons.wayne.edu/oa_dissertations/1205"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Autologous activated T cells (ATC) are being utilized in clinical trials as potential mediators of anti-tumor cytotoxicity in refractory cancer patients who suffer from hematologic malignancies or solid tumors. ATC are readily generated from peripheral blood and possess non-major histocompatibility complex (non-MHC) restricted killing properties. ATC are also powerful targets for genetic modification and hold promise for treatments of cancer, AIDS and T cell disorders. Retrovirus-mediated gene transfer and the fate of proviral gene expression were evaluated in human T cells using 1) immobilized anti-CD3 monoclonal antibody (mAb) plus interleukin-2 (IL-2), or 2) cis costimulation using beads carrying co-immobilized anti-CD3 and anti-CD28 mAbs. By cross-linking the CD3 and CD28 receptors, these mAbs mimic in vivo signaling events, leading to cytokine production and proliferation. A modified human interleukin-1β (IL-Iβ) cDNA inserted into the MFG retroviral vector served as an indicator gene in these studies. Optimized methodologies resulted in T cell transduction frequencies of approximately 50-75%. An important consideration for the success of T lymphocyte-based gene therapy is the relationship between T cell activity and the level of proviral gene expression. Early after mAb stimulation and virus exposure, proviral gene expression was greater at the RNA and protein levels in optimized anti-CD3/anti-CD28 bead-activated cultures, corresponding with augmented endogenous cytokine responses and mitogenesis. Proviral gene expression was not regulated by extrinsic cellular factors present in activated T cell supernatants. Regardless of the mAb stimulation method, proviral IL-lβ expression declined in long-term cultures concomitant with a decrease in cellular cytokines. Restimulation of transduced T cells by either mAb method reinduced both T cell activity and vector expression. The finding that proviral gene regulation is downmodulated in the absence of T cell signaling events suggests that in vivo, high-level proviral gene expression will not be maintained. To overcome the loss of activity in gene-modified T lymphocytes, we have applied a bispecific antibody (BsAb) to redirect cytotoxicity to tumor and enhance T cell activity. For successful cancer adoptive immunotherapy, a coordinated approach of BsAb arming of retrovirus-gene-modified T cells can be utilized for delivery of beneficial, tumoricidal molecules to the tumor site."]},{"key":"dc:title","label":"Title","values":["Retroviral-mediated gene transfer of Il-1β?cDNA into human activated T cells"]}]}],"canonical_facts":{"dc:contributor":["Katrina T. Trevor, Ph.D."],"dc:creator":["Ruhl, Elizabeth"],"dc:date.available":["2015-10-01T07:00:00Z"],"dc:description.abstract":["Autologous activated T cells (ATC) are being utilized in clinical trials as potential mediators of anti-tumor cytotoxicity in refractory cancer patients who suffer from hematologic malignancies or solid tumors. ATC are readily generated from peripheral blood and possess non-major histocompatibility complex (non-MHC) restricted killing properties. ATC are also powerful targets for genetic modification and hold promise for treatments of cancer, AIDS and T cell disorders. Retrovirus-mediated gene transfer and the fate of proviral gene expression were evaluated in human T cells using 1) immobilized anti-CD3 monoclonal antibody (mAb) plus interleukin-2 (IL-2), or 2) cis costimulation using beads carrying co-immobilized anti-CD3 and anti-CD28 mAbs. By cross-linking the CD3 and CD28 receptors, these mAbs mimic in vivo signaling events, leading to cytokine production and proliferation. A modified human interleukin-1β (IL-Iβ) cDNA inserted into the MFG retroviral vector served as an indicator gene in these studies. Optimized methodologies resulted in T cell transduction frequencies of approximately 50-75%. An important consideration for the success of T lymphocyte-based gene therapy is the relationship between T cell activity and the level of proviral gene expression. Early after mAb stimulation and virus exposure, proviral gene expression was greater at the RNA and protein levels in optimized anti-CD3/anti-CD28 bead-activated cultures, corresponding with augmented endogenous cytokine responses and mitogenesis. Proviral gene expression was not regulated by extrinsic cellular factors present in activated T cell supernatants. Regardless of the mAb stimulation method, proviral IL-lβ expression declined in long-term cultures concomitant with a decrease in cellular cytokines. Restimulation of transduced T cells by either mAb method reinduced both T cell activity and vector expression. The finding that proviral gene regulation is downmodulated in the absence of T cell signaling events suggests that in vivo, high-level proviral gene expression will not be maintained. To overcome the loss of activity in gene-modified T lymphocytes, we have applied a bispecific antibody (BsAb) to redirect cytotoxicity to tumor and enhance T cell activity. For successful cancer adoptive immunotherapy, a coordinated approach of BsAb arming of retrovirus-gene-modified T cells can be utilized for delivery of beneficial, tumoricidal molecules to the tumor site."],"dc:identifier":["https://digitalcommons.wayne.edu/oa_dissertations/1205"],"dc:title":["Retroviral-mediated gene transfer of Il-1β?cDNA into human activated T cells"],"thesis:degree_discipline":["Molecular Biology and Genetics"],"thesis:degree_level":["Open Access Dissertation"],"thesis:degree_name":["Ph.D."]},"updated_at":"2026-07-24T06:00:09Z"}