The transduction studies were completed in triplicate for every MOI. cells demonstrated tumorspecific replies against RENCA cells. The restimulated splenocytes gathered from mice treated with or irRC/AdV/GM cells created considerably higher degrees of interleukin2 irRC/SeV/GM, interleukin4, and interferon weighed against their particular handles (P< 0.05). Furthermore, vaccination with irRC/AdV/GM or irRC/SeV/GM cells induced considerably improved recruitment from the cytolytic effectors of Compact disc107a+Compact disc8+T cells and Compact disc107a+organic killer cells into tumors weighed against those induced by their particular handles (P< 0.05). Used together, our outcomes claim that the SeV/dF/G vector is normally a potential applicant for the creation of effective autologous GMCSFtransduced tumor vaccines in scientific cancer immune system gene therapy. (Cancers Sci2008; 99: 23152326) == Abbreviations: == antigenpresenting cell comprehensive moderate cytotoxic T lymphocyte dendritic cell Dulbecco's improved Eagle's moderate draining lymph node fusion fetal bovine serum green fluorescent proteins granulocyte macrophage colonystimulating aspect Hank's buffered sodium alternative interleukin interferon Lewis lung carcinoma main histocompatibility complicated multiplicity of an infection organic killer phosphatebuffered saline renal cell carcinoma Sendai trojan tumorinfiltrating leukocyte tumor necrosis aspect Several studies have got evaluated the capability to augment antitumor immunity using several mouse models and also have proven that GMCSF is among the strongest immunostimulatory cytokines.(1,2,3)GMCSF can be an essential differentiation and maturation aspect for DC,(4)including Langerhans cells to APC,(5)enhancing their capability to provide tumorassociated antigens to activate CTL effectively.(6)Moreover, the cytotoxic activity of NK CTL or cells could be improved or induced by GMCSFrecruited DC. Therefore, GMCSF continues to be postulated to be always a vital mediator of the original antitumor immune system response.(7)Before decade, clinical studies show that autologous GMCSF genetransduced tumor vaccine therapy is feasible, safe and sound, and provides effective antitumor immunomodulating activity against melanoma,(8,9)RCC,(10,11)prostate cancers,(12)pancreatic cancers(13)and nonsmallcell lung cancers.(14)To create effective GMCSFtransduced tumor vaccines, it is vital to efficiently transduce tumor cells also to obtain appropriate appearance from the induced gene. Serotype 5 E1, E3 genedeleted adenovirus encoding individual GMCSF is among the most appealing vectors for tumor vaccines. Nevertheless, adenoviral gene transduction is bound as the receptors for adenovirus serotype 5, including Coxsackie adenovirus receptor, integrin v3, and integrin v5, aren't portrayed on many tumor cells. SeV, a known person in theParamyxoviridaefamily, includes a nonsegmented negativestrand RNA infects and genome via sialic acidity residues on surface area glycoproteins or asialoglycoproteins, which can be found of all mammalian cells.(15,16)Due to the ubiquitous expression from the SeV receptor and high gene transduction capacity, SeV is normally emerging being a appealing gene therapy tool. As SeV possesses a cytoplasmic transcription system, it can transfer exogenous genes to the cytoplasm, where genomic replication and translation are carried out by virally encoded RNA polymerase. This replication system reduces the risk of malignant transformation due to genomic integration of the vector into the hostcell chromosome and increases the safety of this viral vector.(17,18,19,20)To further improve the safety of the SeV vector, we used a newly developed genetically altered temperaturesensitive mutant recombinant vector of nontransmissible SeV (SeV/dF). This vector can selfreplicate but can not be transmitted to adjacent cells due to the lack of the F, thereby increasing the clinical application of this system.(21,22) In the present study, we successfully transduced GFP, mouse GMCSF, and human GMCSF cDNA using SeV/dF encoding GFP (SeV/dF/GFP), mouse GMCSF (SeV/dF/mGM), and human GMCSF (SeV/dF/hGM) cDNA, respectively, into numerous tumorcell linesin vitro. Subsequently, the antitumor efficacies of irradiated SeVmediated GMCSFtransduced RENCA (irRC/SeV/GM) cells and irradiated AdVmediated GMCSFtransduced RENCA (irRC/AdV/GM) cells were compared in a RENCAbearing mouse model, demonstrating that irRC/SeV/GM cells and irRC/AdV/GM cells experienced equivalent antitumor effects. == Materials and Methods == Mice.Six to eight week old female immunocompetent BALB/c and C57BL/6 mice were purchased from Clea Japan, (Tokyo, Japan) and.(c) CD107a mobilization of splenic CD8+T cells from mice treated with the indicated tumor vaccinations. were compared in RENCAbearing mice. The results showed that this antitumor effect was comparative between irRC/SeV/GM and irRC/AdV/GM cells, even though the former produced less GMCSFin vitro. The cell numbers of activated (CD80+, CD86+, CD80+CD86+) dendritic cells in lymph nodes from mice treated with irRC/AdV/GM or irRC/SeV/GM cells were increased significantly compared with those of mice treated with the respective controls, at both the earlier Piribedil D8 and later phases. In anin vitrocytotoxicity assay, splenocytes harvested from mice treated with both irRC/SeV/GM and irRC/AdV/GM cells showed tumorspecific responses against RENCA cells. The restimulated splenocytes harvested from mice treated with irRC/SeV/GM or irRC/AdV/GM cells produced significantly higher levels of interleukin2, interleukin4, and interferon compared with their respective controls (P< 0.05). Furthermore, vaccination with irRC/AdV/GM or irRC/SeV/GM cells induced significantly enhanced recruitment of the cytolytic effectors of CD107a+CD8+T cells and CD107a+natural killer cells into tumors compared with those induced Piribedil D8 by their respective controls (P< 0.05). Taken together, our results suggest that the SeV/dF/G vector is usually a potential candidate for the production of effective autologous GMCSFtransduced tumor vaccines in clinical cancer immune gene therapy. (Malignancy Sci2008; 99: 23152326) == Abbreviations: == antigenpresenting cell total medium cytotoxic T lymphocyte dendritic cell Dulbecco's altered Eagle's medium draining lymph node fusion fetal bovine serum green fluorescent protein granulocyte macrophage colonystimulating factor Hank's buffered salt answer interleukin interferon Lewis lung carcinoma major histocompatibility complex multiplicity of contamination natural killer phosphatebuffered saline renal cell carcinoma Sendai computer virus tumorinfiltrating leukocyte tumor necrosis factor Several studies have evaluated the capacity to augment antitumor immunity using numerous mouse models and have shown that GMCSF is one of the most potent immunostimulatory cytokines.(1,2,3)GMCSF is an important maturation and differentiation factor for DC,(4)including Langerhans cells to APC,(5)enhancing their capacity to present tumorassociated antigens to activate CTL effectively.(6)Moreover, the cytotoxic activity of NK cells or CTL can be enhanced or induced by GMCSFrecruited DC. Therefore, GMCSF has been postulated to be a crucial mediator of the initial antitumor immune response.(7)In the past decade, clinical trials have shown that autologous GMCSF genetransduced tumor vaccine therapy is feasible, safe, and has effective antitumor immunomodulating activity against melanoma,(8,9)RCC,(10,11)prostate malignancy,(12)pancreatic malignancy(13)and nonsmallcell lung malignancy.(14)To generate effective GMCSFtransduced tumor vaccines, it is essential to efficiently transduce tumor cells and to obtain appropriate expression of the induced gene. Serotype 5 E1, E3 genedeleted adenovirus encoding human GMCSF is one of the most encouraging vectors for tumor vaccines. However, adenoviral gene transduction is limited because the receptors for adenovirus serotype 5, including Coxsackie adenovirus receptor, integrin v3, and integrin v5, are not expressed on many tumor cells. SeV, a member of theParamyxoviridaefamily, has a nonsegmented negativestrand RNA genome and infects via sialic acid residues on surface glycoproteins or asialoglycoproteins, which are present on most mammalian cells.(15,16)Because of the ubiquitous expression of the SeV receptor and high gene transduction capacity, SeV is usually emerging as a promising gene therapy tool. As SeV possesses a cytoplasmic transcription system, it can transfer exogenous genes to the cytoplasm, where genomic replication and translation are carried out by virally encoded RNA polymerase. This replication system reduces the risk of malignant transformation due to genomic integration of the vector into the hostcell chromosome and increases the safety of this viral vector.(17,18,19,20)To further improve the safety of the SeV vector, we used a newly developed genetically altered temperaturesensitive mutant recombinant vector of nontransmissible SeV (SeV/dF). This vector can selfreplicate but can not be transmitted to adjacent cells due to the lack of the F, thereby increasing the clinical application of this system.(21,22) In the present study, we successfully transduced GFP, mouse GMCSF, and human GMCSF cDNA using SeV/dF encoding GFP (SeV/dF/GFP), mouse GMCSF (SeV/dF/mGM), and human GMCSF (SeV/dF/hGM) cDNA, respectively, into numerous tumorcell linesin vitro..It could be explained by the statement that irradiation enhanced Rabbit Polyclonal to ARSI the transcription of various genes, including p53 and nuclear factorB,(49)as well as their transfection and transduction efficiencies and transgene integration.(50,51,52)The different effects of irradiation between RENCA and A549 cells are inferred to be dependent on the type or species of tumor cells. treated with both irRC/SeV/GM and irRC/AdV/GM cells showed tumorspecific responses against RENCA cells. The restimulated splenocytes harvested from mice treated with irRC/SeV/GM or irRC/AdV/GM cells produced significantly higher levels of interleukin2, interleukin4, and interferon compared with their respective controls (P< 0.05). Furthermore, vaccination with irRC/AdV/GM or irRC/SeV/GM cells induced significantly enhanced recruitment of the cytolytic effectors of CD107a+CD8+T cells and CD107a+natural killer cells into tumors compared with those induced by their respective controls (P< 0.05). Taken together, our results suggest that the SeV/dF/G vector is Piribedil D8 a potential candidate for the production of effective autologous GMCSFtransduced tumor vaccines in clinical cancer immune gene therapy. (Cancer Sci2008; 99: 23152326) == Abbreviations: == antigenpresenting cell complete medium cytotoxic T lymphocyte dendritic cell Dulbecco's modified Eagle's medium draining lymph node fusion Piribedil D8 fetal bovine serum green fluorescent protein granulocyte macrophage colonystimulating factor Hank's buffered salt solution interleukin interferon Lewis lung carcinoma major histocompatibility complex multiplicity of infection natural killer phosphatebuffered saline renal cell carcinoma Sendai virus tumorinfiltrating leukocyte tumor necrosis factor Several studies have evaluated the capacity to augment antitumor immunity using various mouse models and have shown that GMCSF is one of the most potent immunostimulatory cytokines.(1,2,3)GMCSF is an important maturation and differentiation factor for DC,(4)including Langerhans cells to APC,(5)enhancing their capacity to present tumorassociated antigens to activate CTL effectively.(6)Moreover, the cytotoxic activity of NK cells or CTL can be enhanced or induced by GMCSFrecruited DC. Therefore, GMCSF has been postulated to be a critical mediator of the initial antitumor immune response.(7)In the past decade, clinical trials have shown that autologous GMCSF genetransduced tumor vaccine therapy is feasible, safe, and has effective antitumor immunomodulating activity against melanoma,(8,9)RCC,(10,11)prostate cancer,(12)pancreatic cancer(13)and nonsmallcell lung cancer.(14)To generate effective GMCSFtransduced tumor vaccines, it is essential to efficiently transduce tumor cells and to obtain appropriate expression of the induced gene. Serotype 5 E1, E3 genedeleted adenovirus encoding human GMCSF is one of the most promising vectors for tumor vaccines. Piribedil D8 However, adenoviral gene transduction is limited because the receptors for adenovirus serotype 5, including Coxsackie adenovirus receptor, integrin v3, and integrin v5, are not expressed on many tumor cells. SeV, a member of theParamyxoviridaefamily, has a nonsegmented negativestrand RNA genome and infects via sialic acid residues on surface glycoproteins or asialoglycoproteins, which are present on most mammalian cells.(15,16)Because of the ubiquitous expression of the SeV receptor and high gene transduction capacity, SeV is emerging as a promising gene therapy tool. As SeV possesses a cytoplasmic transcription system, it can transfer exogenous genes to the cytoplasm, where genomic replication and translation are carried out by virally encoded RNA polymerase. This replication system reduces the risk of malignant transformation due to genomic integration of the vector into the hostcell chromosome and increases the safety of this viral vector.(17,18,19,20)To further improve the safety of the SeV vector, we used a newly developed genetically modified temperaturesensitive mutant recombinant vector of nontransmissible SeV (SeV/dF). This vector can selfreplicate but can not be transmitted to adjacent cells due to the lack of the F, thereby increasing the clinical application of this system.(21,22) In the present study, we successfully transduced GFP, mouse GMCSF, and human GMCSF cDNA using SeV/dF encoding GFP (SeV/dF/GFP), mouse GMCSF (SeV/dF/mGM), and human GMCSF (SeV/dF/hGM) cDNA, respectively, into various tumorcell linesin vitro. Subsequently, the antitumor efficacies of irradiated SeVmediated GMCSFtransduced RENCA (irRC/SeV/GM) cells and irradiated AdVmediated GMCSFtransduced RENCA (irRC/AdV/GM) cells were compared in a RENCAbearing mouse model, demonstrating that irRC/SeV/GM cells and irRC/AdV/GM cells had equivalent antitumor effects. == Materials and Methods == Mice.Six to eight week old female immunocompetent BALB/c and C57BL/6 mice were purchased from Clea Japan, (Tokyo, Japan) and housed in the Animal Maintenance Facility at Kyushu University under specific pathogenfree conditions. All animal experiments were approved by the Committee of the Ethics on Animal Experiments in the Faculty of Medicine, Kyushu University and carried out following the Guidelines for Animal Experiments in the Faculty of Medicine, Kyushu University, Fukuoka, Japan and The Law and Notification of the Government. Mouse experiments were carried out at least twice to confirm results. Tumor cell lines.WEHI3B, a mouse myelomonocytic leukemia cell line, was kindly provided by Dr D. Metcalf (University.The transduction studies were completed in triplicate for every MOI. cells demonstrated tumorspecific replies against RENCA cells. The restimulated splenocytes gathered from mice treated with or irRC/AdV/GM cells created considerably higher degrees of interleukin2 irRC/SeV/GM, interleukin4, and interferon weighed against their particular handles (P< 0.05). Furthermore, vaccination with irRC/AdV/GM or irRC/SeV/GM cells induced considerably improved recruitment from the cytolytic effectors of Compact disc107a+Compact disc8+T cells and Compact disc107a+organic killer cells into tumors weighed against those induced by their particular handles (P< 0.05). Used together, our outcomes claim that the SeV/dF/G vector is normally a potential applicant for the creation of effective autologous GMCSFtransduced tumor vaccines in scientific cancer immune system gene therapy. (Cancers Sci2008; 99: 23152326) == Abbreviations: == antigenpresenting cell comprehensive moderate cytotoxic T lymphocyte dendritic cell Dulbecco's improved Eagle's moderate draining lymph node fusion fetal bovine serum green fluorescent proteins granulocyte macrophage colonystimulating aspect Hank's buffered sodium alternative interleukin interferon Lewis lung carcinoma main histocompatibility complicated multiplicity of an infection organic killer phosphatebuffered saline renal cell carcinoma Sendai trojan tumorinfiltrating leukocyte tumor necrosis aspect Several studies have got evaluated the capability to augment antitumor immunity using several mouse models and also have proven that GMCSF is among the strongest immunostimulatory cytokines.(1,2,3)GMCSF can be an essential differentiation and maturation aspect for DC,(4)including Langerhans cells to APC,(5)enhancing their capability to provide tumorassociated antigens to activate CTL effectively.(6)Moreover, the cytotoxic activity of NK CTL or cells could be improved or induced by GMCSFrecruited DC. Therefore, GMCSF continues to be postulated to be always a vital mediator of the original antitumor immune system response.(7)Before decade, clinical studies show that autologous GMCSF genetransduced tumor vaccine therapy is feasible, safe and sound, and provides effective antitumor immunomodulating activity against melanoma,(8,9)RCC,(10,11)prostate cancers,(12)pancreatic cancers(13)and nonsmallcell EPZ-5676 (Pinometostat) lung cancers.(14)To create effective GMCSFtransduced tumor vaccines, it is vital to efficiently transduce tumor cells also to obtain appropriate appearance from the induced gene. Serotype 5 E1, E3 genedeleted adenovirus encoding individual GMCSF is among the most appealing vectors for tumor vaccines. Nevertheless, adenoviral gene transduction is bound as the receptors for adenovirus serotype 5, including Coxsackie adenovirus receptor, integrin v3, and integrin v5, aren't portrayed on many tumor cells. SeV, a known person in theParamyxoviridaefamily, includes a nonsegmented negativestrand RNA infects and genome via sialic acidity residues on surface area glycoproteins or asialoglycoproteins, which can be found of all mammalian cells.(15,16)Due to the ubiquitous expression from the SeV receptor and high Rabbit Polyclonal to RhoH gene transduction capacity, SeV is normally emerging being a appealing gene EPZ-5676 (Pinometostat) therapy tool. As SeV possesses a cytoplasmic transcription system, it can transfer exogenous genes to the cytoplasm, where genomic replication and translation are carried out by virally encoded RNA polymerase. This replication system reduces the risk of malignant transformation due to genomic integration of the EPZ-5676 (Pinometostat) vector into the hostcell chromosome and increases the safety of this viral vector.(17,18,19,20)To further improve the safety of the SeV vector, we used a newly developed genetically altered temperaturesensitive mutant recombinant vector of nontransmissible SeV (SeV/dF). This vector can selfreplicate but can not be transmitted to adjacent cells due to the lack of the F, thereby increasing the clinical application of this system.(21,22) In the present study, we successfully transduced GFP, mouse GMCSF, and human GMCSF cDNA using SeV/dF encoding GFP (SeV/dF/GFP), mouse GMCSF (SeV/dF/mGM), and human GMCSF (SeV/dF/hGM) cDNA, respectively, into numerous tumorcell linesin vitro. Subsequently, the antitumor efficacies of irradiated SeVmediated GMCSFtransduced RENCA (irRC/SeV/GM) cells and irradiated AdVmediated GMCSFtransduced RENCA (irRC/AdV/GM) cells were compared in a RENCAbearing mouse model, demonstrating that irRC/SeV/GM cells and irRC/AdV/GM cells experienced equivalent antitumor effects. == Materials and Methods == Mice.Six to eight week old female immunocompetent BALB/c and C57BL/6 mice were purchased from Clea Japan, (Tokyo, Japan) and.(c) CD107a mobilization of splenic CD8+T cells from mice treated with the indicated tumor vaccinations. were compared in RENCAbearing mice. The results showed that this antitumor effect was comparative between irRC/SeV/GM and irRC/AdV/GM cells, even though the former produced less GMCSFin vitro. The cell numbers of activated (CD80+, CD86+, CD80+CD86+) dendritic cells in lymph nodes from mice treated with irRC/AdV/GM or irRC/SeV/GM cells were increased significantly compared with those of mice treated with the respective controls, at both the earlier and later phases. In anin vitrocytotoxicity assay, splenocytes harvested from mice treated with both irRC/SeV/GM and irRC/AdV/GM cells showed tumorspecific responses against RENCA cells. The restimulated splenocytes harvested from mice treated with irRC/SeV/GM or irRC/AdV/GM cells produced significantly higher levels of interleukin2, interleukin4, and interferon compared with their respective controls (P< 0.05). Furthermore, vaccination with irRC/AdV/GM or irRC/SeV/GM cells induced significantly enhanced recruitment of the cytolytic effectors of CD107a+CD8+T cells and CD107a+natural killer cells into tumors compared with those induced by their respective controls (P< 0.05). Taken together, our results suggest that the SeV/dF/G vector is usually a potential candidate for the production of effective autologous GMCSFtransduced tumor vaccines in clinical cancer immune gene therapy. (Malignancy Sci2008; 99: 23152326) == Abbreviations: == antigenpresenting cell total medium cytotoxic T lymphocyte dendritic cell Dulbecco's altered Eagle's medium draining lymph node fusion fetal bovine serum green fluorescent protein granulocyte macrophage colonystimulating factor Hank's buffered salt answer interleukin interferon Lewis lung carcinoma major histocompatibility complex multiplicity of contamination natural killer phosphatebuffered saline renal cell carcinoma Sendai computer virus tumorinfiltrating leukocyte tumor necrosis factor Several studies have evaluated the capacity to augment antitumor immunity using numerous mouse models and have shown that GMCSF is one of the most potent immunostimulatory cytokines.(1,2,3)GMCSF is an important maturation and differentiation factor for DC,(4)including Langerhans cells to APC,(5)enhancing their capacity to present tumorassociated antigens to activate CTL effectively.(6)Moreover, the cytotoxic activity of NK cells or CTL can be enhanced or induced by GMCSFrecruited DC. Therefore, GMCSF has been postulated to be a crucial mediator of the initial antitumor immune response.(7)In the past decade, clinical trials have shown that autologous GMCSF genetransduced tumor vaccine therapy is feasible, safe, and has effective antitumor immunomodulating activity against melanoma,(8,9)RCC,(10,11)prostate malignancy,(12)pancreatic malignancy(13)and nonsmallcell lung malignancy.(14)To generate effective GMCSFtransduced tumor vaccines, it is essential to efficiently transduce tumor cells and to obtain appropriate expression of the induced gene. Serotype 5 E1, E3 genedeleted adenovirus encoding EPZ-5676 (Pinometostat) human GMCSF is one of the most encouraging vectors for tumor vaccines. However, adenoviral gene transduction is limited because the receptors for adenovirus serotype 5, including Coxsackie adenovirus receptor, integrin v3, and integrin v5, are not expressed on many tumor cells. SeV, a member of theParamyxoviridaefamily, has a nonsegmented negativestrand RNA genome and infects via sialic acid residues on surface glycoproteins or asialoglycoproteins, which are present on most mammalian cells.(15,16)Because of the ubiquitous expression of the SeV receptor and high gene transduction capacity, SeV is usually emerging as a promising gene therapy tool. As SeV possesses a cytoplasmic transcription system, it can transfer exogenous genes to the cytoplasm, where genomic replication and translation are carried out by virally encoded RNA polymerase. This replication system reduces the risk of malignant transformation due to genomic integration of the vector into the hostcell chromosome and increases the safety of this viral vector.(17,18,19,20)To further improve the safety of the SeV vector, we used a newly developed genetically altered temperaturesensitive mutant recombinant vector of nontransmissible SeV (SeV/dF). This vector can selfreplicate but can not be transmitted to adjacent cells due to the lack of the F, thereby increasing the clinical application of this system.(21,22) In the present study, we successfully transduced GFP, mouse GMCSF, and human GMCSF cDNA using SeV/dF encoding GFP (SeV/dF/GFP), mouse GMCSF (SeV/dF/mGM), and human GMCSF (SeV/dF/hGM) cDNA, respectively, into numerous tumorcell linesin vitro..It could be explained by the statement that irradiation enhanced the transcription of various genes, including p53 and nuclear factorB,(49)as well as their transfection and transduction efficiencies and transgene integration.(50,51,52)The different effects of irradiation between RENCA and A549 cells are inferred to be dependent on the type or species of EPZ-5676 (Pinometostat) tumor cells. treated with both irRC/SeV/GM and irRC/AdV/GM cells showed tumorspecific responses against RENCA cells. The restimulated splenocytes harvested from mice treated with irRC/SeV/GM or irRC/AdV/GM cells produced significantly higher levels of interleukin2, interleukin4, and interferon compared with their respective controls (P< 0.05). Furthermore, vaccination with irRC/AdV/GM or irRC/SeV/GM cells induced significantly enhanced recruitment of the cytolytic effectors of CD107a+CD8+T cells and CD107a+natural killer cells into tumors compared with those induced by their respective controls (P< 0.05). Taken together, our results suggest that the SeV/dF/G vector is a potential candidate for the production of effective autologous GMCSFtransduced tumor vaccines in clinical cancer immune gene therapy. (Cancer Sci2008; 99: 23152326) == Abbreviations: == antigenpresenting cell complete medium cytotoxic T lymphocyte dendritic cell Dulbecco's modified Eagle's medium draining lymph node fusion fetal bovine serum green fluorescent protein granulocyte macrophage colonystimulating factor Hank's buffered salt solution interleukin interferon Lewis lung carcinoma major histocompatibility complex multiplicity of infection natural killer phosphatebuffered saline renal cell carcinoma Sendai virus tumorinfiltrating leukocyte tumor necrosis factor Several studies have evaluated the capacity to augment antitumor immunity using various mouse models and have shown that GMCSF is one of the most potent immunostimulatory cytokines.(1,2,3)GMCSF is an important maturation and differentiation factor for DC,(4)including Langerhans cells to APC,(5)enhancing their capacity to present tumorassociated antigens to activate CTL effectively.(6)Moreover, the cytotoxic activity of NK cells or CTL can be enhanced or induced by GMCSFrecruited DC. Therefore, GMCSF has been postulated to be a critical mediator of the initial antitumor immune response.(7)In the past decade, clinical trials have shown that autologous GMCSF genetransduced tumor vaccine therapy is feasible, safe, and has effective antitumor immunomodulating activity against melanoma,(8,9)RCC,(10,11)prostate cancer,(12)pancreatic cancer(13)and nonsmallcell lung cancer.(14)To generate effective GMCSFtransduced tumor vaccines, it is essential to efficiently transduce tumor cells and to obtain appropriate expression of the induced gene. Serotype 5 E1, E3 genedeleted adenovirus encoding human GMCSF is one of the most promising vectors for tumor vaccines. However, adenoviral gene transduction is limited because the receptors for adenovirus serotype 5, including Coxsackie adenovirus receptor, integrin v3, and integrin v5, are not expressed on many tumor cells. SeV, a member of theParamyxoviridaefamily, has a nonsegmented negativestrand RNA genome and infects via sialic acid residues on surface glycoproteins or asialoglycoproteins, which are present on most mammalian cells.(15,16)Because of the ubiquitous expression of the SeV receptor and high gene transduction capacity, SeV is emerging as a promising gene therapy tool. As SeV possesses a cytoplasmic transcription system, it can transfer exogenous genes to the cytoplasm, where genomic replication and translation are carried out by virally encoded RNA polymerase. This replication system reduces the risk of malignant transformation due to genomic integration of the vector into the hostcell chromosome and increases the safety of this viral vector.(17,18,19,20)To further improve the safety of the SeV vector, we used a newly developed genetically modified temperaturesensitive mutant recombinant vector of nontransmissible SeV (SeV/dF). This vector can selfreplicate but can not be transmitted to adjacent cells due to the lack of the F, thereby increasing the clinical application of this system.(21,22) In the present study, we successfully transduced GFP, mouse GMCSF, and human GMCSF cDNA using SeV/dF encoding GFP (SeV/dF/GFP), mouse GMCSF (SeV/dF/mGM), and human GMCSF (SeV/dF/hGM) cDNA, respectively, into various tumorcell linesin vitro. Subsequently, the antitumor efficacies of irradiated SeVmediated GMCSFtransduced RENCA (irRC/SeV/GM) cells and irradiated AdVmediated GMCSFtransduced RENCA (irRC/AdV/GM) cells were compared in a RENCAbearing mouse model, demonstrating that irRC/SeV/GM cells and irRC/AdV/GM cells had equivalent antitumor effects. == Materials and Methods == Mice.Six to eight week old female immunocompetent BALB/c and C57BL/6 mice were purchased from Clea Japan, (Tokyo, Japan) and housed in the Animal Maintenance Facility at Kyushu University under specific pathogenfree conditions. All animal experiments were approved by the Committee of the Ethics on Animal Experiments in the Faculty of Medicine, Kyushu University and carried out following the Guidelines for Animal Experiments in the Faculty of Medicine, Kyushu University, Fukuoka, Japan and The Law and Notification of the Government. Mouse experiments were carried out at least twice to confirm results. Tumor cell lines.WEHI3B, a mouse myelomonocytic leukemia cell line, was kindly provided by Dr D. Metcalf (University.