Modified PD-1 incorporating an unnatural amino acid covalently bound PD-L1 to shrink tumors in vivo.
Major Finding: Modified PD-1 incorporating an unnatural amino acid covalently bound PD-L1 to shrink tumors in vivo.
Concept: Covalent protein drugs combine advantages of small-molecule covalent drugs with proteins' selectivity.
Impact: This proof-of-concept study shows that this novel approach can be used to design new therapeutics.
Small-molecule drugs that bind their targets covalently sometimes have advantages over those that bind noncovalently, such as increased duration of action and complete target inactivation, but a common limitation of small-molecule covalent drugs is off-target reactivity. Some protein-based drugs could hypothetically overcome these off-target effects, but proteins generally do not bind their targets covalently. Li, Chen, Klauser, and colleagues thus sought to develop a means to combine the benefits of drugs that covalently bind their targets with the advantages of protein drugs. As a proof of concept, they focused on developing a protein drug that would covalently bind PD-L1, a protein expressed on cancer cell surfaces. To do this, an unnatural amino acid, fluorosulfate-L-tyrosine (FSY), was incorporated into PD-1, the canonical receptor for PD-L1, using genetic-code expansion, with the idea being that the FSY in the modified PD-1 would react with a proximal histidine residue in PD-L1 upon binding. The modified PD-1 was able to selectively and covalently bind PD-L1 in vitro and in vivo via the intended mechanism. In vitro, the modified PD-1 increased T-cell activation by dendritic cells, whereas wild-type (WT) PD-1 did not. Further, in mice with humanized immune systems, the modified PD-1 exhibited potent antitumor activity; again, WT PD-1 did not produce this effect. Notably, the modified PD-1 performed equivalently to or better than the mainstay immunotherapeutic agent anti–PD-L1. Additionally, the modified PD-1, but not WT PD-1, activated chimeric antigen receptor (CAR) T cells in vitro and increased the CAR T cells' antitumor activity in vivo, promoting enhanced tumor infiltration by the CAR T cells. Together, these results not only demonstrate that the modified PD-1 examined in this work may be worth further study, but also show that this novel method to design covalent protein drugs—dubbed proximity-enabled reactive therapeutics (PERx)—is a promising approach for developing new therapies.
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