Intratumoral Immunization by p19Arf and Interferon-β Gene Transfer in a Heterotopic Mouse Model of Lung Carcinoma

Abstract

Therapeutic strategies that act by eliciting and enhancing antitumor immunity have been clinically validated as an effective treatment modality but may benefit from the induction of both cell death and immune activation as primary stimuli. Using our AdRGD-PG adenovector platform, we show here for the first time that in situ gene transfer of p19Arf and interferon-β (IFNβ) in the LLC1 mouse model of lung carcinoma acts as an immunotherapy. Although p19Arf is sufficient to induce cell death, only its pairing with IFNβ significantly induced markers of immunogenic cell death. In situ gene therapy with IFNβ, either alone or in combination with p19Arf, could retard tumor progression, but only the combined treatment was associated with a protective immune response. Specifically in the case of combined intratumoral gene transfer, we identified 167 differentially expressed genes when using microarray to evaluate tumors that were treated in vivo and confirmed the activation of CCL3, CXCL3, IL1α, IL1β, CD274, and OSM, involved in immune response and chemotaxis. Histologic evaluation revealed significant tumor infiltration by neutrophils, whereas functional depletion of granulocytes ablated the antitumor effect of our approach. The association of in situ gene therapy with cisplatin resulted in synergistic elimination of tumor progression. In all, in situ gene transfer with p19Arf and IFNβ acts as an immunotherapy involving recruitment of neutrophils, a desirable but previously untested outcome, and this approach may be allied with chemotherapy, thus providing significant antitumor activity and warranting further development for the treatment of lung carcinoma.

Gene-based Interventions for Cancer Immunotherapy

“…In a mouse model of melanoma, we have confirmed the induction of an antitumor immune response in vaccine and immunotherapy settings, with critical involvement of NK cells, CD4+ and CD8+ T cells [112]. In a mouse model of lung carcinoma, we have shown that in situ gene therapy can bring about an antitumor immune response with critical involvement of neutrophils [113]. Together these studies show that our gene transfer approach is an effective immunotherapy [114,115].…”

Harnessing combined p19Arf and interferon-beta gene transfer as an inducer of immunogenic cell death and mediator of cancer immunotherapy

“…10, 11 We propose that our approach may provide advantages not seen with existing immunotherapies. For example, gene transfer is not associated with strong adverse reactions, such as the cardiotoxicity seen with a bona fide ICD inducer like doxorubicin.…”

p19Arf sensitizes B16 melanoma cells to interferon-β delivered via mesenchymal stem cells in vitro

“…In our previous studies, we have shown that direct intratumoral delivery of an adenoviral vector encoding IFNb inhibits tumor progression (12,13). Moreover, the association of IFNb gene transfer along with p19Arf (alternative reading frame of the Cdkn2a gene, p19Arf in mice, p14ARF in humans) brought the added benefit of immunogenic cell death and the combined, but not individual, treatments served as an immunotherapy (12)(13)(14). The role of p19Arf is to block the interaction of Mdm2 with p53, sparing p53 from ubiquitination and subsequent degradation, thus promoting its activation and tumor suppressor function (15).…”