Fig. 4. Combination immunotherapy controls CFPAC-1 tumor growth in humanized mouse model.

a CFPAC-1 cells were transplanted into the right flank of humanized mice (control, CART alone, CAdTrio alone: n = 5 animals, CAdTrio+CART: n = 6 animals). A total of 1 × 10⁷ vp of CAdTrio (OAd:HD = 1:1) were injected into the tumor. A total of 1 × 10⁶ HER2.CARTs expressing ffLuc were systemically administered 3 days post injection of CAdTrio. Tumor volumes were monitored at different time points. b We collected serum samples from mice at 0, 3, 10, 24, 45, and 66 days post injection of CAdTrio, and measured human Th1 and Th2 cytokine levels by Multiplex. Data are presented as means ± SD. c Bioluminescence of HER2.CARTs was monitored at different time points. Data are presented as means ± SD, p = 0.003. P-values were determined using two-tailed t test (t ratio = 4.213, dF = 8). d CFPAC-1 tumors were harvested from non-humanized and humanized mice (non-humanized mice: n = 4 animals, humanized mice: n = 5 animals) at 3 days post injection of CAdTrio, and total RNA was extracted from whole tumors. Pro-inflammatory genes were quantified and normalized with human β-Actin. Data are presented as means ± SD. P-values were determined using two-tailed t test; p = 0.0093 (t ratio = 3.553, dF = 7), p = 0.0004 (t ratio = 6.352, dF = 7), p = 0.0005 (t ratio = 6.084, dF = 7), p = 0.0045 (t ratio = 4.101, dF = 7), p = 0.014 (t ratio = 3.226, dF = 7), p = 0.027 (t ratio = 2.782, dF = 7). Statistical significance set at p < 0.05, ns > 0.05. e CFPAC-1 tumors were harvested from humanized mice (non-humanized mice: n = 4 animals, humanized mice: n = 5 animals) at 3 days post injection of CAdTrio, and tumor infiltrating human immune cells were analyzed with flow cytometry. Box plot elements: central line, median; box limit, upper and lower quartiles; whisker, 1.5x inter-quartile range; points, outliers. Abbreviations: s.c. subcutaneous, i.t. intratumoral, i.v. intravenous, ND not detectable.