Figure 5.

Direct in vivo immunization with lipoplexes of lipid 1 and MART1 (melanoma antigen)-encoded DNA vaccine (p-CMV-MART1) protects syngeneic C57BL/6J mice from lethal melanoma challenge with remarkable memory response. (a) Six- to eight-week-old female syngeneic C57BL/6J mice (each weighing 20–22 g, n = 5) were immunized (s.c) with lipoplexes of lipid 1 and p-CMV-MART1 and lipoplexes of lipid 1 and p-CMV-β-gal (as negative control) using 150 µl 5% glucose solution containing 15 µg DNA, 4:1 lipid:DNA ratio, three times with 7-day intervals. Two weeks post third immunization, mice were challenged with melanoma tumor by s.c. injection of ~1 × 10⁵ B16F10 cells. Tumor volumes (V = ½ ab² where, a = maximum length of the tumor and b = minimum length of the tumor measured perpendicular to each other) were measured with a slide calipers for up to 30 days. Results represent the means ± SD for n = 5 tumors (*P < 0.005 versus tumor sizes for lipoplexes of lipid 1 and p-CMV- MART1; statistical analysis was performed using students unpaired t-test). (b) Percentage of tumor-free mice immunized (s.c.) with lipoplexes of lipid 1 and p-CMV-MART1 and lipoplexes of lipid 1 and p-CMV-β-gal. (c) All the C57BL/6J mice (n = 5) immunized with lipoplexes of lipid 1 and p-CMV-MART1 which lived a tumor-free life for 100 days post first tumor challenge were challenged a second time with ~1 × 10⁵ B16F10 cells. The short-term antitumor memory responses after this second tumor challenge are shown for 30 days (*P < 0.005 versus tumor sizes for lipoplex of lipid 1 and p-CMV- MART1; statistical analysis was performed using students unpaired t-test). (d) The percentages of tumor-free mice remaining alive up to 180 days after the second melanoma challenge (80%).