An Anti-Programmed Death-1 Antibody (αPD-1) Fusion Protein That Self-Assembles into a Multivalent and Functional αPD-1 Nanoparticle

Abstract

Cancer immune checkpoint therapy has achieved remarkable clinical successes in various cancers. However, current immune checkpoint inhibitors block the checkpoint of not only the immune cells that are important to cancer therapy but also the immune cells that are irrelevant to the therapy. Such an indiscriminate blockade limits the efficacy and causes the autoimmune toxicity of the therapy. It might be beneficial to use a carrier to target immune checkpoint inhibitors to cancer-reactive immune cells. Here, we explore a method to load the inhibitors into carriers. We used the anti-programmed death-1 antibody (αPD-1) as a model immune checkpoint inhibitor. First, we generated a recombinant single-chain variable fragment (scFv) of αPD-1. Then, we designed and generated a fusion protein consisting of the scFv and an amphiphilic immune-tolerant elastin-like polypeptide (iTEP). Because of the amphiphilic iTEP, the fusion was able to self-assemble into a nanoparticle (NP). The NP was proved to block the PD-1 immune checkpoint in vitro and in vivo. Particularly, the NP exacerbated diabetes development in non-obese diabetic mice as effectively as natural, intact αPD-1. In summary, we successfully expressed αPD-1 as a recombinant protein and linked αPD-1 to a NP, which lays a foundation to develop a delivery system to target αPD-1 to a subpopulation of immune cells.

Depletion of PD-1-positive cells ameliorates autoimmune disease

“…We used an additional method to establish the role of PD-1 in the binding and internalization of αPD-1-ABD-PE to PD-1 + cells. Here, we utilized a fusion protein of the ligand 1 of programmed death 1 protein and a human IgG Fc domain (PD-L1-Fc) 27 and performed a competitive binding assay (Figure 1g, Figure S3). We found concurrent incubation of αPD-1-ABD-PE and PD-L1-Fc with PD-1 + cells abolished the binding and internalization of αPD-1-ABD-PE into PD-1 + cells.…”

“…Normally, αPD-1 accelerates T1D progression in NOD mice because it allows autoreactive PD-1 + cells to proliferate 19,[24][25][26][27] . In this study, αPD-1 accelerated the onset of T1D in NOD mice that were pretreated with PBS and the mixture control: the median T1D-free survivals were 15 and 13 days, respectively ( Figure 4i).…”

“…The three groups were treated intraperitoneally with αPD-1-ABD-PE (5 mg/kg), a mixture of αPD-1 (scFv, 2.5 mg/kg) and ABD-PE (2.5 mg/kg), or PBS every other day for totally five treatments. Two days after the last dosing, 0.5 mg/mouse αPD-1 IgG (clone RMP1-14) 59 ) was administered intraperitoneally to all the mice. Two days later, all these mice received four additional doses of αPD-1 IgG with a two-day interval at a dose level of 0.25 mg/ mouse/injection.…”

“…Here, we describe αPD-1-ABD-PE as a tool for PD-1 + cell depletion. This immunotoxin consists of a single-chain variable fragment (scFv) of αPD-1 27 , an albumin-binding domain (ABD) 28,29 , and a Pseudomonas exotoxin (PE) 30,31 . The αPD-1 scFv serves as a targeting moiety.…”

Dendritic Cells and Programmed Death-1 Blockade: A Joint Venture to Combat Cancer

“…Today, we are learning how to recognize and manage immune-related adverse events and toxicities and gaining knowledge on which therapeutic combinations could be applied best at what time point ( 120 , 121 ). As an alternative to human(ized) mABs, different blocking moieties with advanced target specificity and affinity and reduced toxicity profiles are under investigation, including chimeric fusion proteins (AMP-224, extracellular domain of PD-L2, and an Fc portion of IgG) and nanotechnologies [nanoparticles ( 122 ) and nanobodies (( 123 ), Theravectys, Ablynx)]. Although research in this area is limited, these alternative blockers have interesting features because of their size, stability, and pharmacodynamical properties ( 124 ), which might pave the way for implementation in combination therapy with DCs.…”