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Lymphokine-activated killer and dendritic cell carriage enhances oncolytic reovirus therapy for ovarian cancer by overcoming antibody neutralization in ascites.

Jennings VA, Ilett EJ, Scott KJ, West EJ, Vile R, Pandha H, Harrington K, Young A, Hall GD, Coffey M, Selby P, Errington-Mais F, Melcher AA - Int. J. Cancer (2013)

Bottom Line: A major obstacle for effective oncolytic virotherapy is effective delivery of OV to tumor cells.Loading OV onto cell carriers may facilitate virus delivery in the presence of NAb and immune cells which have their own antitumor effector activity are particularly appealing.Reovirus-loaded LAKDC, and to a lesser degree iDC, were able to: (i) protect from NAb and hand-off reovirus for tumor cell killing; (ii) induce a proinflammatory cytokine milieu (IFNɣ, IL-12, IFNα and TNFα) and (iii) generate an innate and specific antitumor adaptive immune response.

View Article: PubMed Central - PubMed

Affiliation: Targeted & Biological Therapies Group, Leeds Institute of Molecular Medicine, University of Leeds, United Kingdom.

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Reovirus-loaded iDC and LAKDC can deliver reovirus for tumor cell killing in the presence of ascites. Reovirus was loaded onto iDC, LAK cells and LAKDC cocultures (1pfu/cell) and either quantified for: (a) Percentage of reovirus retention by plaque assay or (b) Cytotoxicity by Live/dead® viability assay at 48 hr postloading. Graphs show the mean + SEM of three healthy donors. (c) TR175 were treated with neat reovirus or reovirus-loaded onto iDC, LAK cells and LAKDC (iDC-reo, LAK-reo and LAKDC-reo) for 4 hr ± 2.5% ascites. Carrier cells were then removed and replaced with growth media ± ascites; TR175 cell viability was then determined 72 hr p.i. by Live/dead® viability assay. Graph shows mean percentage of cell death + SEM from five healthy donors using two ascitic fluids (AF1 and 7; *p < 0.05).
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fig03: Reovirus-loaded iDC and LAKDC can deliver reovirus for tumor cell killing in the presence of ascites. Reovirus was loaded onto iDC, LAK cells and LAKDC cocultures (1pfu/cell) and either quantified for: (a) Percentage of reovirus retention by plaque assay or (b) Cytotoxicity by Live/dead® viability assay at 48 hr postloading. Graphs show the mean + SEM of three healthy donors. (c) TR175 were treated with neat reovirus or reovirus-loaded onto iDC, LAK cells and LAKDC (iDC-reo, LAK-reo and LAKDC-reo) for 4 hr ± 2.5% ascites. Carrier cells were then removed and replaced with growth media ± ascites; TR175 cell viability was then determined 72 hr p.i. by Live/dead® viability assay. Graph shows mean percentage of cell death + SEM from five healthy donors using two ascitic fluids (AF1 and 7; *p < 0.05).

Mentions: Whilst reovirus has previously been successfully loaded onto human iDC for tumor delivery,18 the ability of LAK cells or LAKDC cocultures to carry and protect reovirus from NAb, has not been investigated. To first identify their suitability as potential viral carriers, iDC, LAK cells and LAKDC cocultures were loaded with reovirus and their levels of reovirus retention and replication determined by plaque assay. Approximately 5% of input virus was retained by all cell populations (Fig. 3a) and no reovirus replication was detected up to 72 hr postloading (data not shown). In addition, for iDC and LAK cells to be effective cell carriers and provide additional antitumor effects, it was important to establish that they remained viable following reovirus-loading. Viability studies showed no significant increase in death 48 hr post-reovirus loading (Fig. 3b). Next, the ability of iDC, LAK cells and LAKDC cocultures to deliver reovirus to tumor cells in the presence of NAb was investigated. Neat reovirus and reovirus-loaded iDC, LAK and LAKDC (iDC-reo, LAK-reo and LAKDC-reo) were cultured with TR175 cells at a 1:1 ratio for 4 hr ± 2.5% ascitic fluid. Carrier cells were removed to exclude cell-mediated TR175 cytotoxicity (no TR175 cell death was detected at 4 hr at this E:T ratio – data not shown), and tumor cells were cultured (± 2.5% ascitic fluid) for a further 72 hr to investigate reovirus-induced cytotoxicity. Figure 3c shows that levels of tumor cell death in the presence of ascites following iDC-reo and LAKDC-reo viral hand-off were significantly greater than for neat reovirus or LAK-reo delivery. These data indicate that iDC or LAKDC cocultures are effective cell carriers for delivering reovirus to tumor cells in the presence of NAb.


Lymphokine-activated killer and dendritic cell carriage enhances oncolytic reovirus therapy for ovarian cancer by overcoming antibody neutralization in ascites.

Jennings VA, Ilett EJ, Scott KJ, West EJ, Vile R, Pandha H, Harrington K, Young A, Hall GD, Coffey M, Selby P, Errington-Mais F, Melcher AA - Int. J. Cancer (2013)

Reovirus-loaded iDC and LAKDC can deliver reovirus for tumor cell killing in the presence of ascites. Reovirus was loaded onto iDC, LAK cells and LAKDC cocultures (1pfu/cell) and either quantified for: (a) Percentage of reovirus retention by plaque assay or (b) Cytotoxicity by Live/dead® viability assay at 48 hr postloading. Graphs show the mean + SEM of three healthy donors. (c) TR175 were treated with neat reovirus or reovirus-loaded onto iDC, LAK cells and LAKDC (iDC-reo, LAK-reo and LAKDC-reo) for 4 hr ± 2.5% ascites. Carrier cells were then removed and replaced with growth media ± ascites; TR175 cell viability was then determined 72 hr p.i. by Live/dead® viability assay. Graph shows mean percentage of cell death + SEM from five healthy donors using two ascitic fluids (AF1 and 7; *p < 0.05).
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fig03: Reovirus-loaded iDC and LAKDC can deliver reovirus for tumor cell killing in the presence of ascites. Reovirus was loaded onto iDC, LAK cells and LAKDC cocultures (1pfu/cell) and either quantified for: (a) Percentage of reovirus retention by plaque assay or (b) Cytotoxicity by Live/dead® viability assay at 48 hr postloading. Graphs show the mean + SEM of three healthy donors. (c) TR175 were treated with neat reovirus or reovirus-loaded onto iDC, LAK cells and LAKDC (iDC-reo, LAK-reo and LAKDC-reo) for 4 hr ± 2.5% ascites. Carrier cells were then removed and replaced with growth media ± ascites; TR175 cell viability was then determined 72 hr p.i. by Live/dead® viability assay. Graph shows mean percentage of cell death + SEM from five healthy donors using two ascitic fluids (AF1 and 7; *p < 0.05).
Mentions: Whilst reovirus has previously been successfully loaded onto human iDC for tumor delivery,18 the ability of LAK cells or LAKDC cocultures to carry and protect reovirus from NAb, has not been investigated. To first identify their suitability as potential viral carriers, iDC, LAK cells and LAKDC cocultures were loaded with reovirus and their levels of reovirus retention and replication determined by plaque assay. Approximately 5% of input virus was retained by all cell populations (Fig. 3a) and no reovirus replication was detected up to 72 hr postloading (data not shown). In addition, for iDC and LAK cells to be effective cell carriers and provide additional antitumor effects, it was important to establish that they remained viable following reovirus-loading. Viability studies showed no significant increase in death 48 hr post-reovirus loading (Fig. 3b). Next, the ability of iDC, LAK cells and LAKDC cocultures to deliver reovirus to tumor cells in the presence of NAb was investigated. Neat reovirus and reovirus-loaded iDC, LAK and LAKDC (iDC-reo, LAK-reo and LAKDC-reo) were cultured with TR175 cells at a 1:1 ratio for 4 hr ± 2.5% ascitic fluid. Carrier cells were removed to exclude cell-mediated TR175 cytotoxicity (no TR175 cell death was detected at 4 hr at this E:T ratio – data not shown), and tumor cells were cultured (± 2.5% ascitic fluid) for a further 72 hr to investigate reovirus-induced cytotoxicity. Figure 3c shows that levels of tumor cell death in the presence of ascites following iDC-reo and LAKDC-reo viral hand-off were significantly greater than for neat reovirus or LAK-reo delivery. These data indicate that iDC or LAKDC cocultures are effective cell carriers for delivering reovirus to tumor cells in the presence of NAb.

Bottom Line: A major obstacle for effective oncolytic virotherapy is effective delivery of OV to tumor cells.Loading OV onto cell carriers may facilitate virus delivery in the presence of NAb and immune cells which have their own antitumor effector activity are particularly appealing.Reovirus-loaded LAKDC, and to a lesser degree iDC, were able to: (i) protect from NAb and hand-off reovirus for tumor cell killing; (ii) induce a proinflammatory cytokine milieu (IFNɣ, IL-12, IFNα and TNFα) and (iii) generate an innate and specific antitumor adaptive immune response.

View Article: PubMed Central - PubMed

Affiliation: Targeted & Biological Therapies Group, Leeds Institute of Molecular Medicine, University of Leeds, United Kingdom.

Show MeSH
Related in: MedlinePlus