Brachytherapy (BR, contact radiotherapy) is one of the radiation methods of treating patients with cancer. It destroys cancer cells and also affects the tumor microenvironment (tumor blood vessels and immune cells). A dose of 10Gy acts as an "in situ" vaccination leading to the development of a robust antitumor immune response. Late effect after radiation is characterized by arise of numerous areas of underoxygenation (hypoxia) which weakens the anticancer effect of RT. Imiquimod (IMQ), the toll-like receptor 7 (TLR7) agonist, acts as an immunostimulant and vascular normalizing agent. In this study, we evaluated the effect of IMQ on the long-term antitumor response of BR in a mouse model of melanoma. The combination of IMQ with BR should elicit synergistic effect in cancer treatment.

Mice with well-developed B16-F10 melanoma tumors were treated with IMQ and 5 days later with BR. IMQ was administered subcutaneously at a vascular normalized dose of 50 μg. BR was used at dose of 10Gy. The dose was planned using CT scans on the dedicated commercial platform. Irradiation was performed in the shielded therapeutic room with a high-dose rate afterloader equipped with an iridium-192 radioactive source (Microselectron, Nucletron). The dose per fraction was planned to be specified 2–3 mm from the applicator surface. The time of fraction delivery was adjusted depending on the source activity (3–10 Ci).

Therapeutic anticancer effect of IMQ and BR combination was studied. We observed 70% tumor growth inhibition following monotherapy with the use of IMQ or 10Gy dose radiotherapy at 22th day of therapy. Combined therapy (IMQ with BR) inhibited tumor growth the most effectively compared to the other groups (tumor growth inhibition was 90% to control group and 70% to monotherapies). In a long-term observation we noticed tumor progression after IMQ treatment and large necrosis areas in tumors treated with BR. In mice treated with combination of IMQ+BR we observed prolonged tumor growth inhibition. We also checked the therapy's impact on the hematological system parameters. Combined therapy decreased the number of lymphocytes with increase of granulocytes and eosinophils share in blood of treated mice. The proportion of eosinophils in peripheral blood was 10-times higher after combined therapy, compared to the other groups.

The combination of the vasculature normalizing dose of IMQ with BR elicit synergistic antitumor effect in melanoma treatment. Eosinophils mediated anticancer immunity after combined therapy may be essential for the therapeutic antitumor effect. Our data indicate that it is reasonable to use a drug that will prevent the changes occurring in the tumor microenvironment in combination with radiotherapy. This work was financed by the National Science Centre (Poland), the project no. UMO-2018/31/D/NZ5/01754.