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Potent Cancer Immunotherapy with Dendritic Cells Generated from Genetically Modified Hematopoietic Stem Cells
CANCER IMMUNOTHERAPY immunocompentent cells, chemotherapy can be used to control tumor growth and provide the critical time needed for the establishment of an effective anti-tumor immune response.
254. Vaccination of Recurrent Glioma Patients with Tumour Lysate-Pulsed Dendritic Cells Elicits Immune Responses: Results of a Clinical Phase I/ II Trial Ryuya Yamanaka,1 Junpei Homma,1 Naoki Yajima,1 Naoto Tsuchiya,1 Ryuichi Tanaka.1 1 Neurosurgery, Brain Research Institute, Niigata University, Niigata, Japan. In this Phase I/II trial, the patient’s peripheral blood dendritic cells were pulsed with an autologous tumour lysate of the glioma. Fourteen patients with glioblastoma and 4 patients with anaplastic glioma, ranging in age from 20 to 73 years, participated in this study. The mean numbers of vaccinations of tumour lysate-pulsed dendritic cells were 4.8 times intradermally close to a cervical lymph node, and 4.2 times intratumourally via an Ommaya reservoir. The percentage of CD56-positive cells in the peripheral blood lymphocytes increased after immunization. There were 1 partial response, 2 minor responses and 7 no change cases evaluated by radiological findings. Dendritic cell vaccination elicited T-cellmediated antitumour activity, as evaluated by ELISPOT assay after vaccination in 2 of 5 tested patients. Three patients showed DTH reactivity to the autologous tumour lysate, two of these had a minor clinical response, and two had an increased ELISPOT result. Intratumoural CD4 and CD8 T-cell infiltration was detected in two patients who underwent re-operation after vaccination. This study demonstrated the safety and antitumour effects of autologous tumour lysate-pulsed dendritic cell therapy for patients with malignant glioma.
255. Potent Cancer Immunotherapy with Dendritic Cells Generated from Genetically Modified Hematopoietic Stem Cells Tsukasa Nabekura,1 Hiromitsu Nakauchi,2 Toshirou Nagasawa,1 Masafumi Onodera.1 1 Department of Hematology, Institute of Clinical Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan; 2Laboratory of Stem Cell Therapy, Institute of Medical Science, University of Tokyo, Minatoku, Tokyo, Japan. Since dendritic cells (DCs) genetically engineered to express tumorassociated antigens (TAAs) can elicit potent anti-tumor immunity, cancer immunotherapy with TAA-transduced DCs would be one of the most attractive and promising approaches to treatments of incurable cancers. However, several obstacles remain to be solved for achievement of clinical applications; low transduction efficiency of DCs resulting in undersupply of the transduced DCs for clinical trials and a rapid decline of the TAA expression due to gene silencing in the transduced DCs. In this study, DCs were generated from hematopoietic progenitor cells transduced with ovalbumin (OVA) by the retroviral vector GCDNsap that is resistant to gene silencing in immature cells. When mouse lineage-/c-KIT+ cells were transduced with OVA, expanded and differentiated into DCs with GM-CSF and IL-4, most of cells in culture highly expressed CD11c, CD40, CD86, and MHC class II and the transduction efficiency determined by EGFP expression was over 90%. The yield of the transduced DCs was about 20-fold more than that of DCs present in normal mouse bone marrow. When mice were inoculated with the OVAtransduced DCs, they rejected OVA-expressing tumor E.G7 completely and its protective effect was equal to that observed in mice inoculated with 40 times more DCs pulsed with OVA peptides. S98
A lysispot assay revealed the mice harbored four times more cytotoxic T cell (CTL) against E.G7 than those inoculated with OVA peptide-pulsed DCs. Furthermore, the mice produced antibody against OVA, suggesting the presence of OVA-specific helper T cells in the mice. Since mice bearing E.G7 eradicated the tumor after inoculation with the OVA-transduced DCs, DCs genetically modified with GCDNsap described here would pave the way to the development of new cancer immunotherapy.
256. Importance of Temporal Control of HSVtk Gene Therapy in Combination with T Cell Based Therapies for Cancer Luis A. Sanchez-Perez,1 Michael Gough,1 Jill Thompson,1 Richard G. Vile.1,2 1 Molecular Medicine Program; 2Department of Immunology, Mayo Clinic College of Medicine, Rochester, MN. Many cancer gene therapy strategies rely upon the ability to deliver genes to tumors in vivo leading to the direct killing of tumor cells. We have shown previously, using the B16/HSVtk melanoma model, that in situ tumor cell death can be immunogenic due to recruitment, loading and activation of antigen presenting cells with released tumor antigens. We have now focused upon combining gene therapy with other therapeutic modalities to increase the efficacy of either alone in an attempt to compensate for the poor efficiency of gene delivery. Therefore, we have studied the effects of killing tumors with the HSVtk system to generate an inflammatory environment in vivo to enhance the recruitment of adoptively transferred T cells to the tumor site. We hypothesized that a local inflammatory response at the tumor site should incorporate production of cytokines and chemokines that will enhance the recruitment of adoptively transferred activated T cells. To our surprise, adoptive transfer of ova-specific activated CD8 T cells during the time of GCV treatment did not influence tumor progression. In vitro studies revealed that the supernatant from GCV-treated tumor cells caused death of activated T cells. For this reason, in vivo experiments were repeated with adoptive transfer of ova-specific activated CD8 T two days after GCV treatment. Under these conditions adoptive transfer enhanced the survival of GCV-treated mice. These studies indicate that additive effects can be obtained from the combination of adoptive transfer and local inflammation achieved via gene transfer of the HSVtk gene into tumors. However, our studies also reveal that the HSVtk/GCV strategy may have potentially adverse effects on infiltrating immune cells. Therefore, careful timing of the application of gene therapy with T cell adoptive therapy will be required to optimize the benefits of the combination.
257. AAV Vector-Loading DC with an Autoimmune Domain-Depleted HCV Core Gene Stimulates High Anti-Core and Low Anti-Self CTL Maurizio Chiriva-Internati,1 Yong Liu,2 Rongcheng Luo,2 Changxuan You,2 Paul L. Hermonar.2 1 Microbiology and Immunology, Texas Tech University, Lubbock, TX; 2Internal Medicine, University of Arkansas for Medical Sciences, Little Rock, AR. Background: Many cases of chronic hepatitis C virus (HCV) infection are resistant to conventional therapies. Aims: Such cases might be treated by cell-mediated immunotherapy as cytotoxic T lymphocytes (CTL) are the main mechanism by which viral infections are cleared. The HCV core gene, being well conserved among HCV types, may be an appropriate antigen for targeting HCV-infected cells. However, two regions within core can stimulate autoimmunity.
Molecular Therapy Volume 9, Supplement 1, May 2004 Copyright © The American Society of Gene Therapy
254. Vaccination of Recurrent Glioma Patients with Tumour Lysate-Pulsed Dendritic Cells Elicits Immune Responses: Results of a Clinical Phase I/ II Trial Ryuya Yamanaka,1 Junpei Homma,1 Naoki Yajima,1 Naoto Tsuchiya,1 Ryuichi Tanaka.1 1 Neurosurgery, Brain Research Institute, Niigata University, Niigata, Japan. In this Phase I/II trial, the patient’s peripheral blood dendritic cells were pulsed with an autologous tumour lysate of the glioma. Fourteen patients with glioblastoma and 4 patients with anaplastic glioma, ranging in age from 20 to 73 years, participated in this study. The mean numbers of vaccinations of tumour lysate-pulsed dendritic cells were 4.8 times intradermally close to a cervical lymph node, and 4.2 times intratumourally via an Ommaya reservoir. The percentage of CD56-positive cells in the peripheral blood lymphocytes increased after immunization. There were 1 partial response, 2 minor responses and 7 no change cases evaluated by radiological findings. Dendritic cell vaccination elicited T-cellmediated antitumour activity, as evaluated by ELISPOT assay after vaccination in 2 of 5 tested patients. Three patients showed DTH reactivity to the autologous tumour lysate, two of these had a minor clinical response, and two had an increased ELISPOT result. Intratumoural CD4 and CD8 T-cell infiltration was detected in two patients who underwent re-operation after vaccination. This study demonstrated the safety and antitumour effects of autologous tumour lysate-pulsed dendritic cell therapy for patients with malignant glioma.
255. Potent Cancer Immunotherapy with Dendritic Cells Generated from Genetically Modified Hematopoietic Stem Cells Tsukasa Nabekura,1 Hiromitsu Nakauchi,2 Toshirou Nagasawa,1 Masafumi Onodera.1 1 Department of Hematology, Institute of Clinical Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan; 2Laboratory of Stem Cell Therapy, Institute of Medical Science, University of Tokyo, Minatoku, Tokyo, Japan. Since dendritic cells (DCs) genetically engineered to express tumorassociated antigens (TAAs) can elicit potent anti-tumor immunity, cancer immunotherapy with TAA-transduced DCs would be one of the most attractive and promising approaches to treatments of incurable cancers. However, several obstacles remain to be solved for achievement of clinical applications; low transduction efficiency of DCs resulting in undersupply of the transduced DCs for clinical trials and a rapid decline of the TAA expression due to gene silencing in the transduced DCs. In this study, DCs were generated from hematopoietic progenitor cells transduced with ovalbumin (OVA) by the retroviral vector GCDNsap that is resistant to gene silencing in immature cells. When mouse lineage-/c-KIT+ cells were transduced with OVA, expanded and differentiated into DCs with GM-CSF and IL-4, most of cells in culture highly expressed CD11c, CD40, CD86, and MHC class II and the transduction efficiency determined by EGFP expression was over 90%. The yield of the transduced DCs was about 20-fold more than that of DCs present in normal mouse bone marrow. When mice were inoculated with the OVAtransduced DCs, they rejected OVA-expressing tumor E.G7 completely and its protective effect was equal to that observed in mice inoculated with 40 times more DCs pulsed with OVA peptides. S98
A lysispot assay revealed the mice harbored four times more cytotoxic T cell (CTL) against E.G7 than those inoculated with OVA peptide-pulsed DCs. Furthermore, the mice produced antibody against OVA, suggesting the presence of OVA-specific helper T cells in the mice. Since mice bearing E.G7 eradicated the tumor after inoculation with the OVA-transduced DCs, DCs genetically modified with GCDNsap described here would pave the way to the development of new cancer immunotherapy.
256. Importance of Temporal Control of HSVtk Gene Therapy in Combination with T Cell Based Therapies for Cancer Luis A. Sanchez-Perez,1 Michael Gough,1 Jill Thompson,1 Richard G. Vile.1,2 1 Molecular Medicine Program; 2Department of Immunology, Mayo Clinic College of Medicine, Rochester, MN. Many cancer gene therapy strategies rely upon the ability to deliver genes to tumors in vivo leading to the direct killing of tumor cells. We have shown previously, using the B16/HSVtk melanoma model, that in situ tumor cell death can be immunogenic due to recruitment, loading and activation of antigen presenting cells with released tumor antigens. We have now focused upon combining gene therapy with other therapeutic modalities to increase the efficacy of either alone in an attempt to compensate for the poor efficiency of gene delivery. Therefore, we have studied the effects of killing tumors with the HSVtk system to generate an inflammatory environment in vivo to enhance the recruitment of adoptively transferred T cells to the tumor site. We hypothesized that a local inflammatory response at the tumor site should incorporate production of cytokines and chemokines that will enhance the recruitment of adoptively transferred activated T cells. To our surprise, adoptive transfer of ova-specific activated CD8 T cells during the time of GCV treatment did not influence tumor progression. In vitro studies revealed that the supernatant from GCV-treated tumor cells caused death of activated T cells. For this reason, in vivo experiments were repeated with adoptive transfer of ova-specific activated CD8 T two days after GCV treatment. Under these conditions adoptive transfer enhanced the survival of GCV-treated mice. These studies indicate that additive effects can be obtained from the combination of adoptive transfer and local inflammation achieved via gene transfer of the HSVtk gene into tumors. However, our studies also reveal that the HSVtk/GCV strategy may have potentially adverse effects on infiltrating immune cells. Therefore, careful timing of the application of gene therapy with T cell adoptive therapy will be required to optimize the benefits of the combination.
257. AAV Vector-Loading DC with an Autoimmune Domain-Depleted HCV Core Gene Stimulates High Anti-Core and Low Anti-Self CTL Maurizio Chiriva-Internati,1 Yong Liu,2 Rongcheng Luo,2 Changxuan You,2 Paul L. Hermonar.2 1 Microbiology and Immunology, Texas Tech University, Lubbock, TX; 2Internal Medicine, University of Arkansas for Medical Sciences, Little Rock, AR. Background: Many cases of chronic hepatitis C virus (HCV) infection are resistant to conventional therapies. Aims: Such cases might be treated by cell-mediated immunotherapy as cytotoxic T lymphocytes (CTL) are the main mechanism by which viral infections are cleared. The HCV core gene, being well conserved among HCV types, may be an appropriate antigen for targeting HCV-infected cells. However, two regions within core can stimulate autoimmunity.
Molecular Therapy Volume 9, Supplement 1, May 2004 Copyright © The American Society of Gene Therapy