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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 LAKDC prime antitumor immunity in the presence of ascites. Skov-3 cells were infected with either reovirus, iDC-reo or LAKDC-reo in the presence of 2.5% ascites (AF4) for 24 hr. Nonadherent carrier cells/dead tumor cells were removed and cultured with autologous PBMC for a week. CTL were restimulated weekly and tumor specific CTL activity measured in a CD107 assay against specific Skov-3 targets or irrelevant Mel-888 targets. (a) FACS plots are representative of three donors and the percentage of CD8 cells degranulating against targets is shown. (b) Graph shows the mean percentage + SEM of CD8 cells degranulating against Skov-3 targets from three healthy donors (*p < 0.05).
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fig06: Reovirus-loaded LAKDC prime antitumor immunity in the presence of ascites. Skov-3 cells were infected with either reovirus, iDC-reo or LAKDC-reo in the presence of 2.5% ascites (AF4) for 24 hr. Nonadherent carrier cells/dead tumor cells were removed and cultured with autologous PBMC for a week. CTL were restimulated weekly and tumor specific CTL activity measured in a CD107 assay against specific Skov-3 targets or irrelevant Mel-888 targets. (a) FACS plots are representative of three donors and the percentage of CD8 cells degranulating against targets is shown. (b) Graph shows the mean percentage + SEM of CD8 cells degranulating against Skov-3 targets from three healthy donors (*p < 0.05).

Mentions: Finally, to test the ability of reovirus or reovirus-loaded cells to generate a tumor specific adaptive immune response in the presence of ascites, an in vitro priming assay was performed. Skov-3 cells were cultured with reovirus, iDC-reo or LAKDC-reo for 24 hr, after which the nonadherent cells were harvested [containing antigen-loaded antigen presenting cells (APC)[ and cultured with autologous PBMC. CTL were restimulated up to twice more. One week after the final restimulation, tumor-specific cytotoxic activity was measured using CD107 degranulation assays against Skov-3 and irrelevant Mel-888 targets. Both neat reovirus and iDC-reo generated some CTL responses, reflected by low level specific CD8 degranulation against Skov-3 cells (Fig. 6a); however, the highest levels of adaptive response were seen after priming initiated by LAKDC-reo. This is consistent with the ability of LAKDC-reo to kill tumor cells for antigen release in an inflammatory milieu, via both direct oncolysis and innate immune mechanisms, even in the presence of ascitic NAb. Over multiple experiments, LAKDC-reo generated significantly higher proportions of anti-Skov-3 CTL responses than neat reovirus, although the superiority of LAKDC-reo over iDC-reo did not reach statistical significance (Fig. 6b). The ability of LAKDC-reo to generate the highest proportion of anti-Skov-3 CTL may be due to greater killing for antigen release (Fig. 4) and/or the enhanced production of pro-inflammatory cytokines (Supporting Information Fig. 2). Taken together, these data suggest that LAKDC are consummate cell carriers for reovirus, allowing delivery of reovirus for tumor cell oncolysis, additional innate immune-mediated killing, production of proinflammatory cytokines and maturation of iDC to support adaptive antitumor immune priming.


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 LAKDC prime antitumor immunity in the presence of ascites. Skov-3 cells were infected with either reovirus, iDC-reo or LAKDC-reo in the presence of 2.5% ascites (AF4) for 24 hr. Nonadherent carrier cells/dead tumor cells were removed and cultured with autologous PBMC for a week. CTL were restimulated weekly and tumor specific CTL activity measured in a CD107 assay against specific Skov-3 targets or irrelevant Mel-888 targets. (a) FACS plots are representative of three donors and the percentage of CD8 cells degranulating against targets is shown. (b) Graph shows the mean percentage + SEM of CD8 cells degranulating against Skov-3 targets from three healthy donors (*p < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4321045&req=5

fig06: Reovirus-loaded LAKDC prime antitumor immunity in the presence of ascites. Skov-3 cells were infected with either reovirus, iDC-reo or LAKDC-reo in the presence of 2.5% ascites (AF4) for 24 hr. Nonadherent carrier cells/dead tumor cells were removed and cultured with autologous PBMC for a week. CTL were restimulated weekly and tumor specific CTL activity measured in a CD107 assay against specific Skov-3 targets or irrelevant Mel-888 targets. (a) FACS plots are representative of three donors and the percentage of CD8 cells degranulating against targets is shown. (b) Graph shows the mean percentage + SEM of CD8 cells degranulating against Skov-3 targets from three healthy donors (*p < 0.05).
Mentions: Finally, to test the ability of reovirus or reovirus-loaded cells to generate a tumor specific adaptive immune response in the presence of ascites, an in vitro priming assay was performed. Skov-3 cells were cultured with reovirus, iDC-reo or LAKDC-reo for 24 hr, after which the nonadherent cells were harvested [containing antigen-loaded antigen presenting cells (APC)[ and cultured with autologous PBMC. CTL were restimulated up to twice more. One week after the final restimulation, tumor-specific cytotoxic activity was measured using CD107 degranulation assays against Skov-3 and irrelevant Mel-888 targets. Both neat reovirus and iDC-reo generated some CTL responses, reflected by low level specific CD8 degranulation against Skov-3 cells (Fig. 6a); however, the highest levels of adaptive response were seen after priming initiated by LAKDC-reo. This is consistent with the ability of LAKDC-reo to kill tumor cells for antigen release in an inflammatory milieu, via both direct oncolysis and innate immune mechanisms, even in the presence of ascitic NAb. Over multiple experiments, LAKDC-reo generated significantly higher proportions of anti-Skov-3 CTL responses than neat reovirus, although the superiority of LAKDC-reo over iDC-reo did not reach statistical significance (Fig. 6b). The ability of LAKDC-reo to generate the highest proportion of anti-Skov-3 CTL may be due to greater killing for antigen release (Fig. 4) and/or the enhanced production of pro-inflammatory cytokines (Supporting Information Fig. 2). Taken together, these data suggest that LAKDC are consummate cell carriers for reovirus, allowing delivery of reovirus for tumor cell oncolysis, additional innate immune-mediated killing, production of proinflammatory cytokines and maturation of iDC to support adaptive antitumor immune priming.

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