Structural basis of the therapeutic anti-PD-L1 antibody atezolizumab

Abstract

Monoclonal antibodies targeting PD-1/PD-L1 signaling pathway have achieved unprecedented success in cancer treatment over the last few years. Atezolizumab is the first PD-L1 monoclonal antibody approved by US FDA for cancer therapy; however the molecular basis of atezolizumab in blocking PD-1/PD-L1 interaction is not fully understood. Here we have solved the crystal structure of PD-L1/atezolizumab complex at 2.9 angstrom resolution. The structure shows that atezolizumab binds the front beta-sheet of PD-L1 through three CDR loops from the heavy chain and one CDR loop from the light chain. The binding involves extensive hydrogen-bonding and hydrophobic interactions. Notably there are multiple aromatic residues from the CDR loops forming Pi-Pi stacking or cation-Pi interactions within the center of the binding interface and the buried surface area is more than 2000 Å2, which is the largest amongst all the known PD-L1/antibody structures. Mutagenesis study revealed that two hot-spot residues (E58, R113) of PD-L1 contribute significantly to the binding of atezolizumab. The structure also shows that atezolizumab binds PD-L1 with a distinct heavy and light chain orientation and it blocks PD-1/PD-L1 interaction through competing with PD-1 for the same PD-L1 surface area. Taken together, the complex structure of PD-L1/atezolizumab solved here revealed the molecular mechanism of atezolizumab in immunotherapy and provides basis for future monoclonal antibody optimization and rational design of small chemical compounds targeting PD-L1 surface.

Neutralizing PD-L1 and PD-L2 Enhances the Efficacy of Immune Checkpoint Inhibitors in Ovarian Cancer

“…Interestingly, while there were three studies previously published that also generated high-affinity binding sPD-1 clones, none have reported mutations at A140, nor analyzed their mutant molecules for binding towards PD-L2 (32,43,44). Compared to the binding affinity of commercially available anti-PD-L1 antibody atezolizumab (TENCENTRIQ), our sPD-1 mutants have enhanced binding affinity to PD-L1 (kd= 3.406E -10 vs. kd = 9.96E -09 M) under similar experimental conditions (28). In addition, our sPD-1 mutants have enhanced binding towards PD-L2, whereas atezolizumab does not bind to PD-L2 (45).…”

“…The native binding affinity of wild-type PD-1 to both of its ligands is in low micromolar range (28), making wild-type sPD-1 a reasonable antagonist for blocking PD-1/PD-L1 and to a lesser extent PD-1/PD-L2 binding compared with antibodies that are currently in the clinic. To develop a high affinity sPD-1 antagonist capable of neutralizing both PD-L1 and PD-L2, we generated a library of sPD-1 mutants by introducing random mutations using artificial nucleotides and errorprone PCR, throughout the complete extracellular domain of the PD-1 molecule (sPD-1).…”

Immune Checkpoint PD‐1/PD‐L1: Is There Life Beyond Antibodies?

“…Overall, atezolizumab competes with PD-1 for binding to the same surface site of PD-L1 (Figure 1). [32] …”

Structure and Optimization of Checkpoint Inhibitors

“…Atezolizumab and the other licensed anti-PD-L1 antibodies avelumab and durvalumab are IgG1 antibodies, which bind to the front beta-sheet of PD-L1. The heavy chain and light chain regions of these antibodies are involved in binding, with varying buried surface areas on each molecule which may affect their binding affinities [42,43]. These three antibodies have been noted to use all three complementarity determining regions from their heavy chains and two from the light chains [43,44].…”

“…Interestingly, pembrolizumab has a much lower affinity for mouse PD-1, which may be explained by specific amino acid substitutions (Asp 85 to Gly 85 ) which, when mutated in human PD-1, abolish pembrolizumab binding. Atezolizumab has a high binding affinity of 0.4 nM, utilizing specific hot-spot residues on the protein binding surface [42,51], while avelumab and durvalumab have dissociation constants of 42.1 pM [43] and 667 pM [52], respectively.…”