Figure 6.
Decrease in Cell Viability by AA-LF132 In Vitro and Its Influence on Tumor Growth In Vivo
(A and B) In vitro assessment of toxicity on KB cells at 48 hr post-transfection with AA-LF132, AAstop-LF132, or treatment with 2% sucrose (vehicle control). (A) Representative pictures of KB cells transfected with 100 ng cmRNA. (B) CellTiter-Glo Luminescence Viability Assay. Cell viability was proportional to the measured luminescence. Data is presented as mean in % ± SEM of untransfected control cells (UT, dotted line). Statistical significance versus AAstop-LF132 was assessed by two-way ANOVA adjusted for multiple comparisons, with ****p < 0.0001 and n = 3. (C) Luciferase activity. 5 × 106 KB cells were injected into the flank of immuno-deficient NMRI-nu mice. 10 μg of lipid nanoparticle formulated cmRNA coding for firefly luciferase was injected intratumorally on days 9, 11, and 13 after injection of tumor cells. On day 14, bioluminescence was determined. (D and E) In vivo anti-tumor activity of AA-LF132. 5 × 106 KB cells were injected into the flank of immuno-deficient NMRI-nu mice. 10 μg of AA-LF132, 10 μg of AAstop-LF132 or 2% sucrose were injected intratumorally on days 9, 11, 13, and 18 after injection of tumor cells. (D) Tumor volume was measured in vivo throughout the experiment using a caliper. Arrows display days of treatment. Data represent means ± SEM (left) or individual values of each mouse. n = 7 for AA-LF132; n = 10 for AAstop-LF132 and 2% sucrose. (E) Tumor volume was determined ex vivo on day 21 after injection of tumor cells. Data represent means ± SEM. Statistical significance was assessed by Kruskal-Wallis test adjusted for multiple comparisons, with *p < 0.05, ***p < 0.001, n = 7 for AA-LF132, n = 10 for AAstop-LF132 and 2% sucrose.