Firstly, in vitro, AUNP-12 displays equipotent antagonism toward PD-L1 and PD-L2 in the rescue of lymphocyte proliferation and effector functions. Also, this compound rescues the proliferation in the mouse splenocyte assay system, with average EC50 values of 17 nM and 16.6 nM against rmPD-L1 and rmPD-L2 respectively.
Nextly, in human PBMC cells, AUNP-12 is also able to significantly rescue recombinant human PD-L1 and PD-L2 mediated inhibition in vitro, with average EC50 values of 63.3 nM and 44.1 nM against PD-L1 and PD-L2 respectively.
AUNP-12 also shows good desirable absorption, distribution, metabolism and excretion (ADME) and pharmacokinetic properties. This compound (6 hours) shows a Plasma protein binding of 93.9% with plasma stability of more than 60%. In addition, AUNP-12 exhibits a half-life of 90 minutes in mouse liver microsomes.
In male Balb/c mice, AUNP-12 subcutaneous injection or intravenous injection as a concentration of 3 mg/kg for its PK study. As a result, AUNP-12 shows a low volume of distribution, and the peak plasma levels reach at 0.2-0.4 hours. The absolute bioavailability of AUNP-12 is about 77% in mice.
Lastly, in different tumor mice models, AUNP-12 shows its potential for tumor treatment alone or a combination.
B16F10 cells are injected intravenously in mice to erect a melanoma model, And from day 1, AUNP-12 treats to mice as a subcutaneous manner for 14 days. AUNP-12 treatment results in a 66% reduction in metastatic nodules when compares to the control group.
Cyclophosphamide is a synthetic alkylating immunosuppressant chemically related to the nitrogen mustards with antineoplastic activity. In the CT26 colon carcinoma model, when cyclophosphamide at a low dose, it can reduce the Treg population and enhance the efficacy of immunomodulatory agents.
AUNP-12 at 3 mg/kg leads to a TGI of 41% alone. However, Cyclophosphamide (100mg/kg) combines AUNP-12 ( 3 mg/kg) result in a TGI of 93%. Besides, the combination of AUNP-12 and cyclophosphamide prolongs the median survival from 34 days to 41 days.
In conclusion, AUNP-12 shows excellent antitumor activities in syngeneic mouse models of cancer such as colon carcinoma (CT26), and melanoma (B16F10). Furthermore, NP-12 exhibits a good tolerance as it lacks any reduction in the body weights during the trial.