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182. New Insights Into the Therapeutic Efficacy of the Pancreatic Cancer-Targeted AV25CDC Oncolytic Adenovirus
CANCER-ONCOLYTIC VIRUSES I were analyzed via qPCR /ELISA. Virus replication was determined by plaque forming unit (PFU) assays. Results: Treatment of intracranial tumors with OV resulted in significant monocyte infiltration into the tumor (p<0.05). Surprisingly, this infiltration was reduced 4.5 fold in tumors treated with the RAMBO virus as compared to rHSVQ1 (p<0.05). Infiltrating monocytes from RAMBO treated tumors had significantly less MHCII, Ly6C, and CD86 than rHSVQ1 treated tumors (p<0.05). The microglia of RAMBO treated tumors had significantly less MHCII and CD206 expression than those treated with rHSVQ1 (p< 0.05). To further examine the interaction of Vstat120 with microglia/monocytes we developed a coculture system with murine microglia and infected human glioma cells. U87DEGFR glioma cells were treated with PBS, rHSVQ1, or RAMBO and cocultured with microglia. We observed a 75-fold induction of TNF-α expression in microglia cultured with rHSVQ1 infected glioma cells compared to untreated microglia (p<0.05). Interestingly, we observed a 3.4fold reduction in microglia TNF-α expression in RAMBO treated cocultures compared to rHSVQ1 cocultures (p<0.01). These results corresponded with ELISA data where we observed a 6.9 decrease in microglia TNF-α secretion in RAMBO treated cocultures (p<0.01). In PFU assays with U251T2 glioma cells and microglia, we observed a 2.6 fold increase in infectious virus in RAMBO treated cocultures as compared to rHSVQ1 cocultures (p<0.01). Treating these cocultures with a microglia (murine) specific TNF-α blocking antibody rescued the differences in viral replication between rHSVQ1 and RAMBO cocultures. There was no difference in virus replication of glioma cells or microglia alone. These results suggest Vstat120 reduces inflammation and enhances OV replication in part through the suppression of TNF-α expression/secretion on microglia/monocytes. This transient suppression of the microglia/monocyte inflammatory response enhances OV replication and promotes anti-tumor efficacy.
181. Efficient Lung Orthotopic Tumor-Growth Suppression of Oncolytic Adenovirus Complexed With RGD-Targeted Bioreducible Polymer
Jaesung Kim,1 Hye Yeong Nam,2 Jung Woo Choi,3 Chae-Ok Yun,3 Sung Wan Kim.1,3 1 Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT; 2Samyang Biopharmaceuticals Corporation, Daejeon, Republic of Korea; 3Bioengineering, Hanyang University, Seoul, Republic of Korea. Oncolytic adenoviruses (Ad) have been developed for the eradication of tumors. Although they hold much promise as a cancer therapy, they have a short blood circulation time and high liver toxicity. An effective strategy to overcome these problems has been complexing Ad with shielding materials. However, the therapeutic efficacy of the Ad complexes has also been an issue because passive accumulation does not allow for sufficient delivery of Ad to the cancer cells. To enhance the therapeutic efficacy of the polymer-coated Ads, the attachment of a targeting moiety to polymer-coated Ad vectors is inescapable. Our lab has previously reported the potential use of ArgGly-Asp (RGD)-targeted bioreducible polymers with a polyethylene glycol (PEG) linker for delivering oncolytic Ads. We have shown the enhanced in vitro transduction efficiency and increased cancer killing effect with producing progeny oncolytic Ad particles. Additionally, we have shown significant tumor-growth inhibition of the polymer-shielded Ad in an in vivo lung orthotopic tumor model. The shielding effect of the Ad surface with the polymers allowed evasion of host immune responses and reduction of liver-toxicity. This data demonstrates that the RGD-conjugated bioreducible polymer for delivering the oncolytic Ad vectors could be utilized for cancer therapy via systemic administration.
Molecular Therapy Volume 22, Supplement 1, May 2014 Copyright © The American Society of Gene & Cell Therapy
182. New Insights Into the Therapeutic Efficacy of the Pancreatic Cancer-Targeted AV25CDC Oncolytic Adenovirus Helga Weber,1,2 Manuel Gidekel,2 Santiago Werbajh,1 Edgardo Salvatierra,1 Cecilia Rotondaro,1 Leonardo Sganga,1 David T. Curiel,3 Eduardo Cafferata,1 Osvaldo L. Podhajcer.1 1 Laboratory of Molecular and Cellular Therapy, Instituto Leloir, IIBBA-CONICET, Argentina, Buenos Aires, Argentina; 2Office of the Vice-Rector for Research and Post-Graduate Studies, Universidad de La Frontera, Temuco, Chile; 3The Division of Cancer Biology, Washington University School of Medicine, St. Louis.
Pancreatic Cancer is the fourth leading cause of cancer death in men and women and is characterized by dismal prognosis with no available therapeutics. We constructed a novel oncolytic adenovirus, AV25CDC, whose replication was driven by a 0.5 Kb of the cdc25B promoter placed upstream of E1A. The 468 bp cdc25B fragment extended from -426 to +42 and includes two SP1 sites, a TATA box and an INR motif. Cdc25B is overexpressed in more than 70 % of human pancreatic primary and disseminated cancers while faint or no expression was observed in chronic pancreatitis and pancreatic normal cells. After previous assessment of different fiber modifications we decided to improve viral lytic efficacy by pseudotyping AV25CDC with a chimeric fiber of serotypes 5/3. We investigated the in vitro lytic effect and the in vivo therapeutic efficacy, in combination or not with gemcitabine, on human pancreatic tumor xenografts orthotopically growing in nude mice and in immunocompetent tumors in Syrian hamsters. We also assessed biochemical markers of hepatic toxicity and tumor biomarkers levels. AV25CDC exhibited a strong in vitro lytic effect on pancreatic, gastric and colon cancer cells, even at MOIs of 0.1, while it showed a highly attenuated effect on normal fibroblasts. Moreover, it showed a synergistic effect with gemcitabine, in highly resistant, SW1990 pancreatic cancer cells. These in vitro lytic effects correlated with cdc25B expression in the different cell types. Mice harboring 15-days old orthotopic human SW1990 and MiaPaca 2 pancreatic tumors, treated intratumorally with AV25CDC combined or not with gemcitabine, exhibited 70%-80% reduction in tumor size at day 65 compared to control mice. Systemic treatment with AV25CDC combined with gemcitabine of orthotopic SW1990, MiaPaCa2 and Syrian Hamster Hap-T1 pancreatic tumors also showed in average 80% inhibition in tumor growth. At the end of the study in Syrian hamsters we observed 2.5 X 105 viral particles per ng of DNA in Hap-T1 tumors, while lungs, kidneys and spleen showed no viral particles and the liver exhibited only 1% of the viral particles observed in the tumor mass. The chemo-virotherapy treatment induced a return to normal levels of biochemical parameters of hepatic toxicity such as levels of alanine transaminase, aspartate aminotransferase and pancreatic amylase; moreover, mice treated with chemo-virotherapy also exhibited more than 90% reduction in CA19.9 serum levels compared to control mice. Further studies demonstrated that viral treatment disrupted the tumor stroma facilitating gemcitabine activity. These data demonstrates that AV25CDC is an effective oncolytic agent and a potential candidate for pancreatic cancer chemo-virotherapy.
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181. Efficient Lung Orthotopic Tumor-Growth Suppression of Oncolytic Adenovirus Complexed With RGD-Targeted Bioreducible Polymer
Jaesung Kim,1 Hye Yeong Nam,2 Jung Woo Choi,3 Chae-Ok Yun,3 Sung Wan Kim.1,3 1 Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT; 2Samyang Biopharmaceuticals Corporation, Daejeon, Republic of Korea; 3Bioengineering, Hanyang University, Seoul, Republic of Korea. Oncolytic adenoviruses (Ad) have been developed for the eradication of tumors. Although they hold much promise as a cancer therapy, they have a short blood circulation time and high liver toxicity. An effective strategy to overcome these problems has been complexing Ad with shielding materials. However, the therapeutic efficacy of the Ad complexes has also been an issue because passive accumulation does not allow for sufficient delivery of Ad to the cancer cells. To enhance the therapeutic efficacy of the polymer-coated Ads, the attachment of a targeting moiety to polymer-coated Ad vectors is inescapable. Our lab has previously reported the potential use of ArgGly-Asp (RGD)-targeted bioreducible polymers with a polyethylene glycol (PEG) linker for delivering oncolytic Ads. We have shown the enhanced in vitro transduction efficiency and increased cancer killing effect with producing progeny oncolytic Ad particles. Additionally, we have shown significant tumor-growth inhibition of the polymer-shielded Ad in an in vivo lung orthotopic tumor model. The shielding effect of the Ad surface with the polymers allowed evasion of host immune responses and reduction of liver-toxicity. This data demonstrates that the RGD-conjugated bioreducible polymer for delivering the oncolytic Ad vectors could be utilized for cancer therapy via systemic administration.
Molecular Therapy Volume 22, Supplement 1, May 2014 Copyright © The American Society of Gene & Cell Therapy
182. New Insights Into the Therapeutic Efficacy of the Pancreatic Cancer-Targeted AV25CDC Oncolytic Adenovirus Helga Weber,1,2 Manuel Gidekel,2 Santiago Werbajh,1 Edgardo Salvatierra,1 Cecilia Rotondaro,1 Leonardo Sganga,1 David T. Curiel,3 Eduardo Cafferata,1 Osvaldo L. Podhajcer.1 1 Laboratory of Molecular and Cellular Therapy, Instituto Leloir, IIBBA-CONICET, Argentina, Buenos Aires, Argentina; 2Office of the Vice-Rector for Research and Post-Graduate Studies, Universidad de La Frontera, Temuco, Chile; 3The Division of Cancer Biology, Washington University School of Medicine, St. Louis.
Pancreatic Cancer is the fourth leading cause of cancer death in men and women and is characterized by dismal prognosis with no available therapeutics. We constructed a novel oncolytic adenovirus, AV25CDC, whose replication was driven by a 0.5 Kb of the cdc25B promoter placed upstream of E1A. The 468 bp cdc25B fragment extended from -426 to +42 and includes two SP1 sites, a TATA box and an INR motif. Cdc25B is overexpressed in more than 70 % of human pancreatic primary and disseminated cancers while faint or no expression was observed in chronic pancreatitis and pancreatic normal cells. After previous assessment of different fiber modifications we decided to improve viral lytic efficacy by pseudotyping AV25CDC with a chimeric fiber of serotypes 5/3. We investigated the in vitro lytic effect and the in vivo therapeutic efficacy, in combination or not with gemcitabine, on human pancreatic tumor xenografts orthotopically growing in nude mice and in immunocompetent tumors in Syrian hamsters. We also assessed biochemical markers of hepatic toxicity and tumor biomarkers levels. AV25CDC exhibited a strong in vitro lytic effect on pancreatic, gastric and colon cancer cells, even at MOIs of 0.1, while it showed a highly attenuated effect on normal fibroblasts. Moreover, it showed a synergistic effect with gemcitabine, in highly resistant, SW1990 pancreatic cancer cells. These in vitro lytic effects correlated with cdc25B expression in the different cell types. Mice harboring 15-days old orthotopic human SW1990 and MiaPaca 2 pancreatic tumors, treated intratumorally with AV25CDC combined or not with gemcitabine, exhibited 70%-80% reduction in tumor size at day 65 compared to control mice. Systemic treatment with AV25CDC combined with gemcitabine of orthotopic SW1990, MiaPaCa2 and Syrian Hamster Hap-T1 pancreatic tumors also showed in average 80% inhibition in tumor growth. At the end of the study in Syrian hamsters we observed 2.5 X 105 viral particles per ng of DNA in Hap-T1 tumors, while lungs, kidneys and spleen showed no viral particles and the liver exhibited only 1% of the viral particles observed in the tumor mass. The chemo-virotherapy treatment induced a return to normal levels of biochemical parameters of hepatic toxicity such as levels of alanine transaminase, aspartate aminotransferase and pancreatic amylase; moreover, mice treated with chemo-virotherapy also exhibited more than 90% reduction in CA19.9 serum levels compared to control mice. Further studies demonstrated that viral treatment disrupted the tumor stroma facilitating gemcitabine activity. These data demonstrates that AV25CDC is an effective oncolytic agent and a potential candidate for pancreatic cancer chemo-virotherapy.
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