Depletion of macrophages reduces the effect of combining ENG-based DNA vaccine with IL-12.

<p>One day after inoculating mice (n = 7–8) with B16-F10 cells, the animals were orally vaccinated (three times at one-week intervals). Additionally, on days 9, 11 and 13 following inoculation with cancer cells, IL-12 was injected directly into tumors (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0191012#sec002" target="_blank">Materials and methods</a>). Moreover, liposomes (‘empty’ liposomes (control) or Clodronate liposomes (Clodr.)) were delivered intraperitoneally at a dose of 10 mg/kg and directly into tumors at a dose of 5 mg/kg, 2 times/week. Depletion of TAMs decreased the growth of control tumors. But in mice treated with combined therapy we observed enhanced tumor growth after TAMs depletion. Photographs were taken on 20^th day of the experiment. Tumors (n = 3) were collected 20 days after challenge and stained with antibodies against CD206 and F4/80. Magnification 20×. *<i>P</i><0.0001, the ANOVA followed by the Tukey’s <i>post hoc</i> test.