OBJECTIVE To evaluate the safety and biodistribution characteristics of mixed activated killer (MAK) immune cells derived from the blood of tumor patients in immunodeficient mice. METHODS Immunodeficient NOG mice were injected with MAK cells 9 times via the tail vein. The mice were observed continuously for 28 d after the last administration. The clinical symptoms, body weight, food intake, hematological and serum biochemical indexes, cytokine indexes and histopathological changes in mice were investigated. In the meanwhile, the distribution characteristics of MAK cells in peripheral blood and various tissues and organs were investigated. RESULTS MAK cells had no significant effect on clinical symptoms, the injection site, body weight and food intake in mice, and could increase the level of human interferon gamma in serum of mice. MAK cells could elevate lymphocytes%, monocytes%, white blood cells and decrease basophils% in peripheral blood. Mix cell aggregates were observed in most organs. Massive expansion of MAK cells was observed in most tissues of mice after administration of MAK cells for 28 d. CONCLUSION MAK cells could proliferate and induce immune responses in immunodeficient NOG mice without obvious toxic reactions. Given the above, the data of this study can provide a reference for the non-clinical safety evaluation of related products.

OBJECTIVE To evaluate the preclinical safety of chimeric antigen receptor T (CAR-T) cell U87 in tumor-bearing immunodeficient mice. METHODS A transplantation tumor model was constructed using NCG mice subcutaneously inoculated with BxPC-3, on which cells U87 (2×10⁶cells each mouse and 10×10⁶ cells each mouse) were administered in a single dose, and continued to be observed until day 84 after administration. Tumor clearance, survival rate, clinical symptoms, body weight changes, hematological indices, lymphocyte subpopulation classification, cytokine indices, and histopathological changes were examined to characterize the mice. RESULTS Single tail vein administration of U87 to hormonal NCG mice significantly prolonged mouse survival and even completely cleared the tumor load. As its pharmacodynamic action, U87 resulted in elevated levels of human-derived IFN-γ, as well as mixed cell aggregation in multiple tissues/organisms, with no local irritation and no apparent systemic toxicity associated with U87. U87 led excessive immune response at high doses, and has certain effects on the values of GLU, TG, TP, ALB, PLT and RET in animals. Mild GvHD-associated histopathological changes were only observed in 2×10⁶ CAR-T (from week 8 after cell administration) and 10×10⁶ CAR-T (from week 4 after cell administration), and the changes correlated with the level of T-cell proliferation. No tumor formation associated with U87 was seen. CONCLUSION U87 does not show any significant risk associated with immunotoxicity and tumorigenicity in BxPC-3 loaded NCG mice and shows some pharmacodynamic effects. The results of this study provide implications for the toxicological evaluation of targeted solid tumors.

OBJECTVIE To provide safety basis for clinical application, and evaluate the toxicity CIK cells by repeated administration of CIK cells in severe immunodeficient NPG mouse model. METHODS The NPG mice were randomized into two groups: the main experimental group and the satellite group. Each group was divided into vehicle control group, low dose group (2×10⁶ cells per mouse) and high dose group (3×10⁷ cells per mouse). The CIK cells were given every two weeks for 6 times with 28-day recovery. During the experiment, the clinical symptoms, body weight and food intake were observed in the main experimental group. Blood samples were dissected and collected at the end of the administration period and recovery period, and the hematology, serum biochemical examination, cytokine detection, organ weighing and pathological examination were performed. The blood samples of satellite animals were collected to investigate the distribution of CIK cells before the first and last administration, 3 h, 1 d, 3 d, 10 d after the administration, and after the recovery period. RESULTS After repeated administration for 6 times at low dose, no obvious toxic reaction was observed in mice. A few CIK cells were detected in the blood at 1 day after the first dose and at some time points (3 h, day 10, day 29) after the last dose. High levels of CIK cells were detected in the blood of mice after repeated administration of 6 times at high dose. The levels of IFN-γ, TNF, IL-10, IL-2 and Il-10 in serum were increased. At the same time, significant graft-versus-host disease (GvHD) reaction was observed in mice, including the changes of clinical symptoms, body weight, food intake, hematology and serum biochemistry, mixed cell aggregation and GvHD-related lesions occurred in multiple organs of animals. CONCLUSION The data from the study indicates that no significant toxic reaction was observed at the dose of 2×10⁶ cells per mouse (the clinical dosage). GvHD associated with xenotransplantation was observed at a dose of 3×10⁷ cells per mouse, and no other related toxicity was observed. These data will facilitate CIK cells to enter into clinical trials.

OBJECTIVE To evaluate the distribution and clearance of tumor infiltrating lymphocytes (TIL) in NOG mice. METHODS Eighty-four NOG mice were used, divided into control group and test group. The mice were given a single tail vein injection. After administration blood was collected and organs were taken at different time points, flow cytometry and real-time quantitative PCR (qPCR) were used to detect the distribution of cells in the blood and different organs. RESULTS After 1-2 d of administration, the number of CD3⁺, CD3⁺CD4⁺, and CD3⁺CD8⁺cells in the blood reached their peaks. At the same time, qPCR showed that TIL was mainly distributed in tissues such as lungs, blood, and liver, with less distribution in other tissues. On day 42 after administration, gene copies of most tissues were below the lower limit of quantitation. CONCLUSION Single tail vein injection of TIL in NOG mice is mainly distributed in the lungs, blood, and liver, and its duration in vivo does not exceed 42 d.