Objective

Introduction. Immune checkpoint inhibitors (ICIs), i.e. PD1, PDL1 and CTLA4 therapeutic antibodies, have led to long-term survival in many late stage solid tumor patients. Despite their revolutionary clinical impact, the overall response rate is still low. One major roadblock is that compensatory immune inhibitory pathways are turned on to protect tumor cells from being attacked by T cells. Various combinatory ICI treatments have now been investigated in clinical trials to tackle this challenge. We have previously reported the development of immune checkpoint target humanized mice (HuGEMM) through CRISPR-Cas9 knock-in. We have generated PD1 and CTLA4 HuGEMM mice and used them for efficacy assessment of corresponding therapeutic antibodies. We have also generated human PDL1 expressing MC38 cells and PDL1 HuGEMM, and demonstrated robust efficacy of several PDL1 therapeutic antibodies.

Methods. Here we report the establishment of ICI double knock-in (KI) mice, i.e., PD1/PDL1 and PD1/CTLA4 dKI HuGEMM, and use them as tools to test immunotherapies with PD1/PDL1 and PD1/CTLA4 combos.

Results. Similar to clinical benefit of PD1 and CTLA4 combo shown in the melanoma and NSCLC patients, we found combined treatment of Nivolumab and Ipilimumab antibody leads to 103% TGI in the corresponding PD1/CTLA4 dKI-HuGEMM, with complete tumor remission in 80% of treated mice. These mice remained disease free throughout a tumor re-challenge study for over 40 days. We have also developed PD1/PDL1 dKI HuGEMM and tested combined treatment of Atezolizumab and Nivolumab. Interestingly, we observed improved efficacy in the combo treatment group despite the two ICIs are thought to target the same PD1/PDL1 axis. Monotherapies may not fully release the PD1/PDL1 blockade due to complicity of multiple compatible ligands & receptors, including PDL2 and B7-1. We have also tested PD1/OX40 combo treatment with dKI HuGEMM, which is reported separately.

Conclusion. Our dKI HuGEMM models offer robust tests on ICI combinations, as well as combos of ICIs with other anti-tumor therapeutic modalities.