Supplementary MaterialsDocument S1. 5?times after tumor shot via either the tail we or vein.p. cavity at two different dosages (low dosage, 1? 106 CAR T?cells; high dosage, 10? 106 CAR T?cells). All untreated mice as well as the cohorts of i.v. and we.p. NT cell remedies showed continuing tumor extension and expired within 45?times of xenograft Cidofovir (Vistide) (Statistics 4B and 4D). Compared, CAR T?cell-treated mice exhibited lower tumor burden and survived longer than did the zero NT or T cell cohorts. i.p. delivery of CAR T?cells led to a far more pronounced treatment impact and success advantage more than i actually.v. delivery (Figures 4C and 4D). In the cohort treated with high-dose i.p. CAR T?cells, the tumors seemed to be almost completely eradicated at 9?days post-xenograft; however, tumor relapse Cidofovir (Vistide) occurred as early as 2?weeks after complete response in most mice. The loss of body weight overall was caused by an increase in tumor burden (Physique?4E). Open in a separate window Physique?4 Efficacy of ICAM-1 CAR T Cells in an Intraperitoneal Xenograft Model (A) Whole-body bioluminescence image of SNU-638-engrafted NSG mice without treatment (no T), or treated with non-transduced T (NT) or low or high doses (LD or HD) of ICAM-1 CAR T?cells. Mice were treated with T?cells 5?days after tumor xenograft either by intravenous or intraperitoneal injection. LD, 1? 106 CAR T?cells; HD, 10? 106 CAR T?cells. (B) Quantitation of total body bioluminescence Cidofovir (Vistide) intensity. Data represent mean? SD (n?= 2C3). (C) Bioluminescence intensities on day 33 following xenograft. LD and HD cohorts were pooled for analysis. An unpaired, two-tailed t test was used for statistical CD79B comparisons. ?p? 0.05, ??p? 0.01. ns, not significant. (D) Kaplan-Meier survival curves. (E) Summary of body weight changes over time. Data represent mean? SD (n?= Cidofovir (Vistide) 2C3). (F) GFP images of tumors and gastrointestinal tracts acquired on day 85 post-xenograft. (G) Histologic images of H&E staining, GFP IHC, and CD3 IHC of tumor or spleen from mice treated with ICAM-1 CAR T?cells. Validation of CAR T Cell Tumor Infiltration images of the gastrointestinal organs further validated the treatment effect of CAR T?cells against SNU-638 peritoneal tumors. In untreated mice, tumors appeared to form multiple lesions along the intestinal tract, identifiable by GFP imaging (Physique?4F). In comparison, tumor lesions in the intestinal tract of CAR T?cell-treated mice were less frequent and smaller. IHC analysis of tumor nodules from the mice treated with ICAM-1 CAR revealed CD3+ T?cells infiltrating GFP+ tumors (Physique?4G). Close inspection of the image revealed the snapshot of CAR T?cell activity against tumors: the area with high CD3 density appeared to be largely devoid of tumor cells, while the area with sparsely distributed CD3 cells still contained a high density of tumor cells. In the spleen of the same mice, CD3+ human T?cells were observed in high abundance even at 80?days after T?cell infusion. Combined Treatment of CAR T Cells with Paclitaxel in an i.p. Xenograft Model Although ICAM-1 CAR T?cells administered i.p. at a high dose appeared to have a survival benefit, the efficacy was modest and short-lived, and most treated animals eventually succumbed to tumor relapse and death. To examine whether the lower tumor burden at the time of CAR T?cell treatment would lead to a better outcome, we first developed peritoneal tumors at different doses of SNU-638 cells (0.1? 106, 0.5? 106, or 3? 106 cells per mouse) and analyzed the survival rate of each cohort. As expected, non-obese diabetic (NOD) severe combined immunodeficiency (SCID) gamma (NSG) mice survived proportionally longer when the xenograft dose was reduced (Physique?5A). In addition to reducing the xenograft dose to 0.1? 106 SNU-638 cells, we explored the combination of CAR T?cell therapy with chemotherapy, which has been shown to sensitize tumor cells to immunotherapy in preclinical studies.31,32 Among chemotherapeutic brokers, we used paclitaxel, which is the Cidofovir (Vistide) most commonly used.