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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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Malignant ascites inhibits reovirus-induced oncolysis. Ovarian cancer cell lines, Skov-3, OVCA433 and TR175 were infected with reovirus (1 pfu/cell) ± 2.5% ascitic fluid (2 separate samples; AF1 and 2). (a) Cell viability was determined using the Live/dead® viability assay and flow cytometry 48 hr postinfection (p.i.). Graphs show the mean percentage cell death + SEM from three independent experiments. (b) Reovirus replication was determined by plaque assay using L929 cells and fold increase was calculated from initial input reovirus. Graphs show mean fold increase of reovirus titres + SEM at 72 hr p.i. from three independent experiments. (c) Primary ascites-derived ovarian cancer cells from 10 patients were infected ± reovirus (10 pfu/cell) and cell viability was determined 48 hr p.i. using the Live/dead® assay. (d) Four primary samples were infected with reovirus (10 pfu/cell) ± 2.5% autologous ascitic fluid (samples AF3, 4, 5 and 6) for 24, 48 and 72 hr and cell viability determined as above. Graph shows the mean percentage cell death + SEM from four patient samples. (e) Reovirus replication was determined in primary ascites-derived ovarian cancer cells. Cells were infected with reovirus (1 pfu/cell) for 24, 48 and 72 hr ± 2.5% autologous ascitic fluid (samples AF3, 4 and 7) and reovirus replication determined as in (b). Graph shows the mean + SEM from three patient samples (*p < 0.05, **p < 0.01, ***p < 0.001 and ****p < 0.0001).
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fig01: Malignant ascites inhibits reovirus-induced oncolysis. Ovarian cancer cell lines, Skov-3, OVCA433 and TR175 were infected with reovirus (1 pfu/cell) ± 2.5% ascitic fluid (2 separate samples; AF1 and 2). (a) Cell viability was determined using the Live/dead® viability assay and flow cytometry 48 hr postinfection (p.i.). Graphs show the mean percentage cell death + SEM from three independent experiments. (b) Reovirus replication was determined by plaque assay using L929 cells and fold increase was calculated from initial input reovirus. Graphs show mean fold increase of reovirus titres + SEM at 72 hr p.i. from three independent experiments. (c) Primary ascites-derived ovarian cancer cells from 10 patients were infected ± reovirus (10 pfu/cell) and cell viability was determined 48 hr p.i. using the Live/dead® assay. (d) Four primary samples were infected with reovirus (10 pfu/cell) ± 2.5% autologous ascitic fluid (samples AF3, 4, 5 and 6) for 24, 48 and 72 hr and cell viability determined as above. Graph shows the mean percentage cell death + SEM from four patient samples. (e) Reovirus replication was determined in primary ascites-derived ovarian cancer cells. Cells were infected with reovirus (1 pfu/cell) for 24, 48 and 72 hr ± 2.5% autologous ascitic fluid (samples AF3, 4 and 7) and reovirus replication determined as in (b). Graph shows the mean + SEM from three patient samples (*p < 0.05, **p < 0.01, ***p < 0.001 and ****p < 0.0001).

Mentions: Whilst reovirus-induced oncolysis of ovarian cancer cell lines has been previously demonstrated,7 the effects of reovirus-induced oncolysis on primary human ovarian cancer cells and its effects in the context of ascitic fluid have not been examined. In the absence of ascites, reovirus was cytotoxic against three established ovarian cancer cell lines, Skov-3, OVCA433 and TR175 (Fig. 1a) and, importantly, against 10 primary ovarian cancer cell samples freshly isolated from patients’ ascites, albeit with varying sensitivities (Fig. 1c). However, in the presence of 2.5 % ascitic fluid, reovirus-induced cytotoxicity was significantly inhibited in all cell lines and in four out of four patient tumor samples cultured in autologous ascites (Figs. 1a and 1d). Reovirus replication was also examined following infection of ovarian cancer cells ± ascites. In the absence of ascites, reovirus readily replicated in ovarian cancer cell lines (Fig. 1b) and primary ovarian cancer cells (Fig. 1e, although at 10- to 1000-fold lower levels than those detected in cell lines). However, in the presence of ascites, reovirus replication, in line with tumor cell killing, was significantly abrogated (Figs. 1b and 1e).


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)

Malignant ascites inhibits reovirus-induced oncolysis. Ovarian cancer cell lines, Skov-3, OVCA433 and TR175 were infected with reovirus (1 pfu/cell) ± 2.5% ascitic fluid (2 separate samples; AF1 and 2). (a) Cell viability was determined using the Live/dead® viability assay and flow cytometry 48 hr postinfection (p.i.). Graphs show the mean percentage cell death + SEM from three independent experiments. (b) Reovirus replication was determined by plaque assay using L929 cells and fold increase was calculated from initial input reovirus. Graphs show mean fold increase of reovirus titres + SEM at 72 hr p.i. from three independent experiments. (c) Primary ascites-derived ovarian cancer cells from 10 patients were infected ± reovirus (10 pfu/cell) and cell viability was determined 48 hr p.i. using the Live/dead® assay. (d) Four primary samples were infected with reovirus (10 pfu/cell) ± 2.5% autologous ascitic fluid (samples AF3, 4, 5 and 6) for 24, 48 and 72 hr and cell viability determined as above. Graph shows the mean percentage cell death + SEM from four patient samples. (e) Reovirus replication was determined in primary ascites-derived ovarian cancer cells. Cells were infected with reovirus (1 pfu/cell) for 24, 48 and 72 hr ± 2.5% autologous ascitic fluid (samples AF3, 4 and 7) and reovirus replication determined as in (b). Graph shows the mean + SEM from three patient samples (*p < 0.05, **p < 0.01, ***p < 0.001 and ****p < 0.0001).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig01: Malignant ascites inhibits reovirus-induced oncolysis. Ovarian cancer cell lines, Skov-3, OVCA433 and TR175 were infected with reovirus (1 pfu/cell) ± 2.5% ascitic fluid (2 separate samples; AF1 and 2). (a) Cell viability was determined using the Live/dead® viability assay and flow cytometry 48 hr postinfection (p.i.). Graphs show the mean percentage cell death + SEM from three independent experiments. (b) Reovirus replication was determined by plaque assay using L929 cells and fold increase was calculated from initial input reovirus. Graphs show mean fold increase of reovirus titres + SEM at 72 hr p.i. from three independent experiments. (c) Primary ascites-derived ovarian cancer cells from 10 patients were infected ± reovirus (10 pfu/cell) and cell viability was determined 48 hr p.i. using the Live/dead® assay. (d) Four primary samples were infected with reovirus (10 pfu/cell) ± 2.5% autologous ascitic fluid (samples AF3, 4, 5 and 6) for 24, 48 and 72 hr and cell viability determined as above. Graph shows the mean percentage cell death + SEM from four patient samples. (e) Reovirus replication was determined in primary ascites-derived ovarian cancer cells. Cells were infected with reovirus (1 pfu/cell) for 24, 48 and 72 hr ± 2.5% autologous ascitic fluid (samples AF3, 4 and 7) and reovirus replication determined as in (b). Graph shows the mean + SEM from three patient samples (*p < 0.05, **p < 0.01, ***p < 0.001 and ****p < 0.0001).
Mentions: Whilst reovirus-induced oncolysis of ovarian cancer cell lines has been previously demonstrated,7 the effects of reovirus-induced oncolysis on primary human ovarian cancer cells and its effects in the context of ascitic fluid have not been examined. In the absence of ascites, reovirus was cytotoxic against three established ovarian cancer cell lines, Skov-3, OVCA433 and TR175 (Fig. 1a) and, importantly, against 10 primary ovarian cancer cell samples freshly isolated from patients’ ascites, albeit with varying sensitivities (Fig. 1c). However, in the presence of 2.5 % ascitic fluid, reovirus-induced cytotoxicity was significantly inhibited in all cell lines and in four out of four patient tumor samples cultured in autologous ascites (Figs. 1a and 1d). Reovirus replication was also examined following infection of ovarian cancer cells ± ascites. In the absence of ascites, reovirus readily replicated in ovarian cancer cell lines (Fig. 1b) and primary ovarian cancer cells (Fig. 1e, although at 10- to 1000-fold lower levels than those detected in cell lines). However, in the presence of ascites, reovirus replication, in line with tumor cell killing, was significantly abrogated (Figs. 1b and 1e).

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