Dendritic cells infected with a vaccinia virus interleukin-2 vector secrete high levels of IL-2 and can become efficient antigen presenting cells that secrete high levels of the immunostimulatory cytokine IL-12

Abstract

Dendritic cell (DC) therapies using DC presenting tumor antigen/s can induce CD8 + CTL that mediate tumor eradication, nonetheless many patients remain unresponsive. Thus, cytokine gene vectors applied to DC may amplify these responses. Herein, we examined the responses that monocyte-derived DC (at different maturational stages) make when infected with a vaccinia virusinterleukin-2 (VV-IL-2) vector in vitro. VV-IL-2-infected DC secreted significant levels of bioactive IL-2 and maintained their antigen presentation function. However, we show that DC are exquisitely sensitive to their local antigenic microenvironment, and that responses generated by one antigen can be altered by another. VV-IL-2 infection of immature DC led to DC activation (upregulation of CD80, CD86 and class II surface molecules) when the virus was propagated through xenogeneic, but not syngeneic, mammalian cells; these DC secreted IL-10 and tumor necrosis factor-a (TNF-a), but not IL-12. In contrast, after VV-IL-2 infection (regardless of their mammalian cellular context), IFNg/LPS-matured DC inevitably downregulated their antigen presenting machinery. In conclusion, immunostimulatory DC can be generated by VV-IL-2, but this depends upon (i) infecting immature DC only, (ii) the mammalian cells through which the virus is prepared and (iii) individual donors; hence donors must be screened to assess their specific responses.

Immune properties of recombinant vaccinia virus encoding CD154 (CD40L) are determined by expression of virally encoded CD40L and the presence of CD40L protein in viral particles

“…This is consistent with another report demonstrating a diminished IL-12 response upon direct infection of DC with a recombinant VV expressing IL-2 produced in syngeneic cells. 38 This report suggested that DC were highly sensitive to the local environment and characteristics of the VV vectors used for infection. Using sucrose-purified viral particles, we observed stimulation of IL-12 synthesis by PS/UV-inactivated rV-CD40L, which was blocked by anti-CD40 Ab indicating that rV-CD40L particles contained biologically active CD40L protein.…”

Cytokine-armed vaccinia virus infects the mesothelioma tumor microenvironment to overcome immune tolerance and mediate tumor resolution

“…VV. Furthermore, we have earlier shown that these VV vectors prevent DC maturation, 13 whereas others have shown that DC viability is compromised after phagocytosing-killed neoplastic cells in the presence of MVA viral particles. 50 Similarly, our data show that high doses of the VV-HA and VV-IL-12 constructs produce increased DC death.…”

“…As we have already shown that the VV constructs readily infect 143BTK À cells, 13 we next examined the ability of the VV-HA viral vector and its cytokine constructs to Cytokine-armed vaccinia can cure mesothelioma C Jackaman and DJ Nelson infect 1 Â 10 6 mesothelioma tumor cells in vitro. AE17 mesothelioma cells were incubated with 5 Â 10 6 or 1 Â 10 6 pfus of the VV constructs for 24, 48, or 72 h (Figures 2a-c).…”

“…However, we found that this approach downregulated antigen-presenting function in mature DC and was likely to be unreliable. 13 Thus, cytokine-armed DC may not be an effective alternative therapy and, as suggested by studies in mesothelioma and other cancers, regardless of the vector used, the direct i.t. approach remains a viable proposition.…”

The “Trojan Horse” Approach to Tumor Immunotherapy: Targeting the Tumor Microenvironment

“…However, our in vitro studies showed that mesothelioma tumor cells were highly permissive to infection by VVcytokine vectors resulting in significant cytokine production and impaired proliferation, whilst macrophages secreted low levels of IL-2 suggesting resistance to overt infection [123]. We have also shown that human dendritic cells are readily infected with the VVIL-2 vector and that that they become efficient antigen presenting cells that secrete high levels of the immunostimulatory cytokine IL-12 [124]. Our murine proof-of-principle in vivo studies of murine mesotheliomas showed that i.t.…”