Preclinical Efficacy Assessment of Immune Checkpoint Inhibitors

Figure 1. Anti-mPD1 demonstrated significant tumor growth inhibition in both the B6 mouse model inoculated with MC38 (A) and the BALB/c mouse model inoculated with CT26 (B) tumor cells. (A) C57BL/6 mice were inoculated with  murine MC38 colon cancer, followed by i.p. administration of anti-mPD-1 at 1 mg/kg twice a week for a total of 6 times. Tumor volumes were monitored through Day 20. N = 6 mice in each group. (B) BALB/c mice were inoculated with murine CT26 colon cancer, followed by i.p. administration of anti-mPD-1 at 10 mg/kg twice a week for a total of 6 times. Tumor volumes were monitored through Day 20. N = 6 mice per group. Data are shown as mean ± SEM.  A two‑way ANOVA was used, and Šídák's post hoc test was applied for multiple comparisons between each treatment group and the G1 control at endpoint. ***p<0.001

Syngeneic tumor models capture key features of the tumor immune microenvironment, supporting in vivo mechanistic studies of PD-1/PD-L1 antibodies. These models also serve as a robust platform for efficacy assessment, enabling both mechanistic investigation and exploration of combination therapeutic strategies.

Figure 2. In vivo efficacy evaluation of test articles targeting hPD-1 or hPD-L1. (A) C57BL/6-hPD1 mice were inoculated with murine MC38 colon cancer, followed by i.p. administration of Keytruda or Opdivo at 3 mg/kg twice a week for a total of 6 times. Tumor volumes were monitored through Day 25. N = 10 mice in each group. (B) BALB/c-hPD1 mice were inoculated with murine CT26.WT colon cancer, followed by i.p. administration of Keytruda at 5 mg/kg or 10 mg/kg once per three days for a total of 6 times. Tumor volumes were monitored through Day 23. N = 10 mice in each group. (C) C57BL/6-hPD1/hPDL1 mice were inoculated with murine MC38-hPDL1 colon cancer, followed by i.p. administration of Keytruda and Tecentriq at 3 mg/kg once per three days for a total of 6 times. Tumor volumes were monitored through Day 23. N = 8 mice in each group. (D) BALB/c-hPD1/hPDL1 mice were inoculated with murine CT26-hPDL1 colon cancer, followed by i.p. administration of Keytruda and Tecentriq at 10 mg/kg once per three days for a total of 3 times. Tumor volumes were monitored through Day 9. N = 8 mice in each group. Data are shown as mean ± SEM.  A two‑way ANOVA was used, and Tukey's post hoc test was applied for multiple comparisons between each treatment group and the G1 control at endpoint. ***p<0.001

Keytruda, Opdivo, and Tecentriq demonstrated significant anti-tumor effect on C57BL/6J background target humanized mice (B6-hPD1, B6-hPD1/hPDL1) and BALB/c background target humanized mice (BALB/c-hPD1, BALB/c-hPD1/hPDL1). Target-humanized mice provide a reliable in vivo model with strong translational responsiveness to therapeutics targeting human PD-1/PD-L1.

Figure 3. In vivo efficacy evaluation of test articles targeting PD-1 or PD-L1 in human immune cell reconstituted mice bearing CDX tumors. (A) NCG mice were co-inoculated with a LS174T and PBMC admix subcutaneously, followed by i.p. administration of Opdivo at 10 mg/kg once per three days for a total of 6 times. Tumor volumes were monitored through Day 18. N = 6 mice in each group. (B) PBMC reconstituted NCG mice were inoculated with MDA-MB-231 breast cancer, followed by i.p. administration of Tecentriq at 10 mg/kg twice per week for a total of 8 times. Tumor volumes were monitored through Day 38. N = 6 mice in each group. (C) PBMC reconstitution NCG mice were inoculated with RKO colon cancer, followed by i.p. administration of Tecentriq at 10 mg/kg twice per week for a total of 6 times. Tumor volumes were monitored through Day 25. N = 6 mice in each group. (D) HSC reconstitution NCG-IL15 mice were inoculated with HCC827 lung cancer, followed by i.p. administration of Tecentriq at 10 mg/kg twice per week for a total of 6 times. Tumor volumes were monitored through Day 23. N = 5 mice in each group. (E) HSC reconstitution NCG-IL15 mice were inoculated with NCI-H1975 lung cancer, followed by i.p. administration of Anti-PDL1 antibody at 10 mg/kg twice per week for a total of 6 times. Tumor volumes were monitored through Day 23. N = 7 mice in each group. (F) HSC reconstitution NCG-IL15 mice were inoculated with RKO colon cancer, followed by i.p. administration of Tecentriq at 10 mg/kg twice per week for a total of 8 times. Tumor volumes were monitored through Day 33. N = 5 mice in each group.Data are shown as mean ± SEM.  A two‑way ANOVA was used, and Šídák's post hoc test was applied for multiple comparisons between each treatment group and the G1 control at endpoint. *p<0.05, **p<0.01, ***p<0.001

Preclinical efficacy evaluations were conducted to assess the efficacy of PD-1/PD-L1-targeting test agents in humanized immune-reconstituted NCG mice bearing established CDX tumor xenografts. Significant antitumor efficacy was observed for both Opdivo and Tecentriq in PBMC-humanized mice implanted with distinct CDX models, with the therapeutic response exhibiting donor-to-donor variability. In the HSC-NCG-IL15 humanized mouse model, the PD-L1-directed checkpoint inhibitor Tecentriq exhibited robust antitumor responses across different CDX models.