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MFN2 provides antitumor efficacy in pancreatic cancer cells
Abstracts / Pancreatology 16 (2016) S1eS192
immunohistochemical staining and the particle exclusion assay showed that 4-MU led to decrease of HA rich ECM surrounding the cells. Conclusion: 4-MU inhibits cell proliferation and HA synthesis, and leads to reduce the amount of HA in ECM. 4-MU might be an effective therapeutic agent for pancreatic adenocarcinoma.
S171
signifcantly changed with overexpression of MFN2. Tumor cells' invasion and migration abilities were inhibited. Conclusion: MFN2 provides a novel target for pancreatic cancer. The reexpression of MFN2 may become a effective strategy in pancreatic carcinoma therapy in the future.
P-299.
P-301.
Systemic treatment of mesothelin-targeted oncolytic adenovirus eliminates pancreatic cancer
Dendritic cell vaccine transduced with ubiquitin-mesothelin fusion gene for pancreatic cancer
Mizuho Sato Dahlman 1, Yoshiaki Miura 2, Kari Jacobsen 1, Julia Davydova 1, Masato Yamamoto 1
Motoki Miyazawa, Toshiyasu Ojima, Masahiro Katsuda, Manabu Kawai, Seiko Hirono, Ken-ichi Okada, Atsushi Shimizu, Yuji Kitahata, Hiroki Yamaue
1 2
Department of Surgery, University of Minnesota, USA Internal Medicine (III), University of Toyama, USA
Pancreatic cancer is an aggressive malignant disease. Despite extensive efforts, systemic therapies have provided only limited efficacy for patients with this disease. Oncolytic adenovirus (OAd) is a promising therapeutics for pancreatic cancer. However, the low tumor distribution of injected agents is one of major obstacle for systemic therapy. The better tumor distribution and transduction could overcome the barriers for systemic delivery and enable efficient treatment of spread and/or metastatic lesions of pancreatic cancer. AdML-VTIN was identified as a mesothelin (MSLN) targeted Ad by the Ad library screening system. This virus yielded a selective infectivity and oncolytic effect against MSLN-positive pancreatic cancer (Panc-1) in vitro. To assess the potential of AdML-VTIN for systemic application, we compared organ distribution after intravenous (i.v.) injection to the mice. The liver sequestration of AdML-VTIN was lower than wild type (WT) at 48 hrs after injection. By day seven the viral copy number of AdML-VTIN in the tumor was more than 3 orders of magnitude higher than WT. Anti-tumor effect of AdML-VTIN was assessed with i.v. injection (109 vp/mouse). Panc-1 tumor significantly shrunk only in AdML-VTIN treated group while AdML5WT didn't show any growth suppression. When we compared the therapeutic effect of i.v. and i.t. injections, the tumor volume significant decreased in both groups. Although the viral particle in the tumor with i.v. injection was about half of that with i.t. injection at day 1, the anti-tumor effect of systemic injection group was similar to that of local injection group. In Summary, systemic injection of the targeted OAd showed significantly lower liver sequestration and better tumor accumulation. Both systemic and intratumoral injections resulted in remarkable anti-tumor effect at low dose. An impressive anti-tumor effect of systemic administration indicated that targeted-OAd may embody efficient treatment for cancers which are mostly found with spread or metastatic lesions.
Second Department of Surgery, Wakayama Medical University, Japan Background: We previously reported that the potential of developing future clinical applications of the vaccines using genetically modified dendritic cells (DCs) expressing human mesothelin (MSLN) which is potential target of immunotherapy for pancreatic cancer (Miyazawa et al. Cancer lett. 2011). Ubiquitinated antigens are processed to antigenic peptides by the ubiquitin (Ub)-proteasome system and presented by MHC class I. We investigated whether DCs transduced with Ub gene fused with MSLN gene could induce enhanced anti-tumor immune response. Methods: Adenoviral vector encoding MSLN gene (AdMSLN) and Ubfused MSLN gene (AdUbMSLN) were generated. MSLN specific cytotoxic T lymphocytes (CTLs) were induced from autologous peripheral blood mononuclear cells (PBMCs) from healthy donors by in vitro stimulations with DC-AdMSLN or DC-AdUbMSLN. The cytotoxic activity was tested using a 4-hour 51Cr-release assay. PK1 which is a pancreatic adenocarcinoma cell line endogenously expressing MSLN and lymphoblastoid cell lines (LCLs) transduced with the MSLN gene were used as target cells. The interferongamma production in the culture supernatants were assessed by ELISA. Results: CTLs induced by DC-AdUbMSLN killed both PK1 and MSLN transduced LCLs more effectively than CTLs induced by DC-AdMSLN. The rate of intracellular MSLN expression in DC-AdMSLN was 56% compared with 2% in DC-AdUbMSLN by flow cytometry. The inhibition of MSLN degradation by using a proteasome inhibitor (MG132) increased the rate of MSLN expression to 46% in DC-AdUbMSLN. In addition, the interferongamma production by MSLN-specific CTLs incubated with DC-AdUbMSLN was much higher than MSLN-specific CTLs incubated with DC-AdMSLN under the same condition. Conclusions: These results indicated that DC-AdUbMSLN could induce CTLs more efficiently because ubquitinated MSLN promoted to be processed to peptides by the Ub-proteasome system and presented by DCs as peptide-MHC class I complexes.
P-300. MFN2 provides antitumor efficacy in pancreatic cancer cells
P-302.
Qiang Sun, Weilin Wang
Regulation of stemness of pancreatic cancer stem cell by miR-34a through inhibition of epithelial-mesenchymal transition
Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, School of Medicine, Zhejiang University, China Background ＆ aims: In previous studies we confirmed that mitofusion-2 (MFN2) has antitumor efficacy in hepatocellular carcinoma. This study aimed to demonstrate the antitumor efficacy of MFN2 in pancreatic cancer cell lines. Methods: Bxpc-3 and L3.6pl cell lines with MFN2 overexpressed is established by infection of adenovirus. Apoptosis and cell cycle were assessed by flowcytometry and immunoblotting for caspases, cyclins and cyclin-dependent kinases. Cell invasion and migration ability was measured in 24-well transwell plates. Results: Overexpressing MFN2 makes more cells under apoptotic stress with more cleaved caspases than control cells. And cell cycle was not
Gary G. Xiao 1, 2, Kewei Bi 1, WN Paul Lee 2, Vay Liang W. Go 2 1 School of Pharmaceutical Science and Technology, Dalian University of Technology, China 2 Harbor-University of California Los Angeles Research and Education Institute, UCLA School of Medicine, USA
Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers with a five-year survival rate of less than 5%, usually diagnosed in an advanced state and for which there are little or no effective therapies. The ability of PDAC to metastasize in early stages is a primary reason for its lethality. Evidence suggests that this process may be mediated by cancer stem cells (CSCs) as well as the ability of CSCs to undergo epithelialmesenchymal transition (EMT). A line of study showed that miR-34a inhibits stemness of breast cancer stem cells. We then hypothesized that
immunohistochemical staining and the particle exclusion assay showed that 4-MU led to decrease of HA rich ECM surrounding the cells. Conclusion: 4-MU inhibits cell proliferation and HA synthesis, and leads to reduce the amount of HA in ECM. 4-MU might be an effective therapeutic agent for pancreatic adenocarcinoma.
S171
signifcantly changed with overexpression of MFN2. Tumor cells' invasion and migration abilities were inhibited. Conclusion: MFN2 provides a novel target for pancreatic cancer. The reexpression of MFN2 may become a effective strategy in pancreatic carcinoma therapy in the future.
P-299.
P-301.
Systemic treatment of mesothelin-targeted oncolytic adenovirus eliminates pancreatic cancer
Dendritic cell vaccine transduced with ubiquitin-mesothelin fusion gene for pancreatic cancer
Mizuho Sato Dahlman 1, Yoshiaki Miura 2, Kari Jacobsen 1, Julia Davydova 1, Masato Yamamoto 1
Motoki Miyazawa, Toshiyasu Ojima, Masahiro Katsuda, Manabu Kawai, Seiko Hirono, Ken-ichi Okada, Atsushi Shimizu, Yuji Kitahata, Hiroki Yamaue
1 2
Department of Surgery, University of Minnesota, USA Internal Medicine (III), University of Toyama, USA
Pancreatic cancer is an aggressive malignant disease. Despite extensive efforts, systemic therapies have provided only limited efficacy for patients with this disease. Oncolytic adenovirus (OAd) is a promising therapeutics for pancreatic cancer. However, the low tumor distribution of injected agents is one of major obstacle for systemic therapy. The better tumor distribution and transduction could overcome the barriers for systemic delivery and enable efficient treatment of spread and/or metastatic lesions of pancreatic cancer. AdML-VTIN was identified as a mesothelin (MSLN) targeted Ad by the Ad library screening system. This virus yielded a selective infectivity and oncolytic effect against MSLN-positive pancreatic cancer (Panc-1) in vitro. To assess the potential of AdML-VTIN for systemic application, we compared organ distribution after intravenous (i.v.) injection to the mice. The liver sequestration of AdML-VTIN was lower than wild type (WT) at 48 hrs after injection. By day seven the viral copy number of AdML-VTIN in the tumor was more than 3 orders of magnitude higher than WT. Anti-tumor effect of AdML-VTIN was assessed with i.v. injection (109 vp/mouse). Panc-1 tumor significantly shrunk only in AdML-VTIN treated group while AdML5WT didn't show any growth suppression. When we compared the therapeutic effect of i.v. and i.t. injections, the tumor volume significant decreased in both groups. Although the viral particle in the tumor with i.v. injection was about half of that with i.t. injection at day 1, the anti-tumor effect of systemic injection group was similar to that of local injection group. In Summary, systemic injection of the targeted OAd showed significantly lower liver sequestration and better tumor accumulation. Both systemic and intratumoral injections resulted in remarkable anti-tumor effect at low dose. An impressive anti-tumor effect of systemic administration indicated that targeted-OAd may embody efficient treatment for cancers which are mostly found with spread or metastatic lesions.
Second Department of Surgery, Wakayama Medical University, Japan Background: We previously reported that the potential of developing future clinical applications of the vaccines using genetically modified dendritic cells (DCs) expressing human mesothelin (MSLN) which is potential target of immunotherapy for pancreatic cancer (Miyazawa et al. Cancer lett. 2011). Ubiquitinated antigens are processed to antigenic peptides by the ubiquitin (Ub)-proteasome system and presented by MHC class I. We investigated whether DCs transduced with Ub gene fused with MSLN gene could induce enhanced anti-tumor immune response. Methods: Adenoviral vector encoding MSLN gene (AdMSLN) and Ubfused MSLN gene (AdUbMSLN) were generated. MSLN specific cytotoxic T lymphocytes (CTLs) were induced from autologous peripheral blood mononuclear cells (PBMCs) from healthy donors by in vitro stimulations with DC-AdMSLN or DC-AdUbMSLN. The cytotoxic activity was tested using a 4-hour 51Cr-release assay. PK1 which is a pancreatic adenocarcinoma cell line endogenously expressing MSLN and lymphoblastoid cell lines (LCLs) transduced with the MSLN gene were used as target cells. The interferongamma production in the culture supernatants were assessed by ELISA. Results: CTLs induced by DC-AdUbMSLN killed both PK1 and MSLN transduced LCLs more effectively than CTLs induced by DC-AdMSLN. The rate of intracellular MSLN expression in DC-AdMSLN was 56% compared with 2% in DC-AdUbMSLN by flow cytometry. The inhibition of MSLN degradation by using a proteasome inhibitor (MG132) increased the rate of MSLN expression to 46% in DC-AdUbMSLN. In addition, the interferongamma production by MSLN-specific CTLs incubated with DC-AdUbMSLN was much higher than MSLN-specific CTLs incubated with DC-AdMSLN under the same condition. Conclusions: These results indicated that DC-AdUbMSLN could induce CTLs more efficiently because ubquitinated MSLN promoted to be processed to peptides by the Ub-proteasome system and presented by DCs as peptide-MHC class I complexes.
P-300. MFN2 provides antitumor efficacy in pancreatic cancer cells
P-302.
Qiang Sun, Weilin Wang
Regulation of stemness of pancreatic cancer stem cell by miR-34a through inhibition of epithelial-mesenchymal transition
Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, School of Medicine, Zhejiang University, China Background ＆ aims: In previous studies we confirmed that mitofusion-2 (MFN2) has antitumor efficacy in hepatocellular carcinoma. This study aimed to demonstrate the antitumor efficacy of MFN2 in pancreatic cancer cell lines. Methods: Bxpc-3 and L3.6pl cell lines with MFN2 overexpressed is established by infection of adenovirus. Apoptosis and cell cycle were assessed by flowcytometry and immunoblotting for caspases, cyclins and cyclin-dependent kinases. Cell invasion and migration ability was measured in 24-well transwell plates. Results: Overexpressing MFN2 makes more cells under apoptotic stress with more cleaved caspases than control cells. And cell cycle was not
Gary G. Xiao 1, 2, Kewei Bi 1, WN Paul Lee 2, Vay Liang W. Go 2 1 School of Pharmaceutical Science and Technology, Dalian University of Technology, China 2 Harbor-University of California Los Angeles Research and Education Institute, UCLA School of Medicine, USA
Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers with a five-year survival rate of less than 5%, usually diagnosed in an advanced state and for which there are little or no effective therapies. The ability of PDAC to metastasize in early stages is a primary reason for its lethality. Evidence suggests that this process may be mediated by cancer stem cells (CSCs) as well as the ability of CSCs to undergo epithelialmesenchymal transition (EMT). A line of study showed that miR-34a inhibits stemness of breast cancer stem cells. We then hypothesized that