Radiosensitizing effect of intratumoral interleukin-12 gene electrotransfer in murine sarcoma

Abstract

Background: Interleukin-12 (IL-12) based radiosensitization is an effective way of tumor treatment. Local cytokine production, without systemic shedding, might provide clinical benefit in radiation treatment of sarcomas. Therefore, the aim was to stimulate intratumoral IL-12 production by gene electrotransfer of plasmid coding for mouse IL-12 (mIL-12) into the tumors, in order to explore its radiosensitizing effect after single or multiple intratumoral gene electrotransfer.

Methods: Solid SA-1 fibrosarcoma tumors, on the back of A/J mice, were treated intratumorally by mIL-12 gene electrotransfer and 24 h later irradiated with a single dose. Treatment effectiveness was measured by tumor growth delay and local tumor control assay (TCD(50) assay). With respect to therapeutic index, skin reaction in the radiation field was scored. The tumor and serum concentrations of cytokines mIL-12 and mouse interferon γ (mIFNγ) were measured. Besides single, also multiple intratumoral mIL-12 gene electrotransfer before and after tumor irradiation was evaluated.

Results: Single intratumoral mIL-12 gene electrotransfer resulted in increased intratumoral but not serum mIL-12 and mIFNγ concentrations, and had good antitumor (7.1% tumor cures) and radiosensitizing effect (21.4% tumor cures). Combined treatment resulted in the radiation dose-modifying factor of 2.16. Multiple mIL-12 gene electrotransfer had an even more pronounced antitumor (50% tumor cures) and radiosensitizing (86.7% tumor cures) effect.

Conclusions: Single or multiple intratumoral mIL-12 gene electrotransfer resulted in increased intratumoral mIL-12 and mIFNγ cytokine level, and may provide an efficient treatment modality for soft tissue sarcoma as single or adjuvant therapy to tumor irradiation.

Figure 1

Complete responses of SA-1 tumor bearing mice after combined modality treatment. Single (panel A) and repetitive (panel B) intratumoral mIL-12 gene electrotransfer was used alone or combined with tumor irradiation. Abbreviations: mIL-12 = intratumoral injection of plasmid DNA coding for mIL-12; EGT mIL-12 = intratumoral mIL-12 gene electrotransfer; dsRed = intratumoral injection of non-therapeutic plasmid DNA; EGT dsRed = intratumoral dsRed gene electrotransfer; EP = electric pulse application on the tumors; IR = single dose radiation (10 Gy); All other groups = other therapeutic groups listed in Tables 1 and 2; 3× = triple therapy. Number of animals per treatment group is listed in Tables 1 and 2.

Figure 2

Intratumoral concentration of cytokines mIL-12 (panel A) and mIFNγ (panel B). Panel A: * - Statistically significant difference (p < 0.05) compared to groups mIL-12, mIL-12 + IR. ** - Statistically significant difference (p < 0.05) compared to groups mIL-12, mIL-12 + IR, EGT mIL-12; Panel B: * - Statistically significant difference (p < 0.05) compared to groups mIL-12, mIL-12 + IR. Number of animals per group was 12 (days 0, 3, 7, 10), 18 (day 5), 6 (day 14)

Figure 3

Histology of SA-1 tumors. Histology was evaluated at day 5 after intratumoral mIL-12 gene electrotransfer alone (A and B) or combined with irradiation (C and D). The arrows show infiltrating immune cells (L – lymphocytes, Gr – granulocytes, M – macrophages, P – plasma cells) and giant cells in mitotic arrest (G).

Figure 4

Therapeutic index. Radiation dose response curves for local tumor control of SA-1 sarcoma tumors (panel A) and skin reaction (panel B). Arrows indicate response on dry skin desquamation less than 20% (panel A), and radiation doses with which this skin reaction occurs (panel B). In panel A, 7–10 mice per treatment group were evaluated at each dose point. In panel B, 7–10 mice per treatment group were evaluated at each dose and the average of five highest skin reaction scores for each dose used was plotted.