Immunotherapy, which uses immune pathways to treat cancer, is rapidly becoming an accepted cancer treatment after surgery, chemotherapy and radiation. Checkpoint blocking using monoclonal antibodies against cytotoxic T lymphocyte-associated protein 4 (CTLA-4), programmed death receptor 1 (PD-1), or programmed death ligand 1 (PD-L1) has made a significant breakthrough in cancer immunotherapy. However, despite its advantages, immunotherapy is only successful in a small subset of patients, and biomarkers that broadly predict its efficacy are still to be identified. In addition, serious treatment-related toxicities, known as immune-related adverse events (irAE), can occur during treatment, primarily due to an unbalanced immune system.
Therefore, small molecule drugs have returned to people’s vision, because these drugs have many advantages over monoclonal antibodies. In particular, small molecules with shorter half-lives favor acute and reversible effects and may reduce long-lasting systemic side effects. Unlike antibodies, small molecules typically target intracellular proteins and have distinct toxicity profiles, making them suitable candidates for combination therapy. In addition, they can be produced at lower cost than antibodies and can often be administered orally. Therefore, new strategies based on molecular insights from immunological and oncology processes are needed to enhance the potential of small molecules in immunotherapy, with a focus on the promise of small molecules to improve the success of checkpoint blocking for cancer treatment.
Design of small molecules targeting PD-1/PD-L1
The PD-1/PD-L1 receptor-ligand interaction is a classic example of protein-protein interaction (PPI). Therefore, designing inhibitors for these interactions is extremely challenging. This is mainly due to (a) the large contact area of the interaction (1500-3000Å), (b) the existence of a flat interface without deep and well-defined binding pockets suitable for binding ligands with high affinity, and (c) the lack of endogenous small molecule ligands as a reference standard.
At present, there are mainly two different classes of small molecule inhibitors targeting PD-L1. (a) compounds based on biphenyl scaffolds; (b) small amino acid-inspired molecules that mimic the receptor-ligand interface identified in functional assisations.
Biphenyl derivatives
BMS scientists have developed a series of biphenyl derivatives based on the mechanism of the PD-1/PD-L1 interaction.
Peptides
Checkpoint proteins are membrane proteins, most of which come from the B7 family. Most members of the B7 family and their ligands belong to the immunoglobulin superfamily (IgSF). Receptor-ligand interactions of IgSF proteins are mediated through loops, chains, or loops and chains. Peptide design based on these interacting interfaces is a proven strategy for PPI inhibitor design. Several peptide-like molecules have been developed using the above design principles.
Mechanism of action of small molecules targeting PD-1/PD-L1
While much of the molecular design comes from being able to bind to one of the PD-1:PD-L1 complexes and prevent interactions, emerging data suggest that some compounds that antagonize PD-1 signaling have additional complexity.
Induces PD-L1 dimerization and inhibits PD-1/PD-L1 interaction: Small molecule inhibitors of BMS, for example, tend to first interact with one PD-L1 monomer and then form a dimer to gain a stability advantage. Incyte and Arbutus also took advantage of the inherent C2 symmetry in PD-L1 dimers to achieve greater potency by symmetrizing their molecules.
Blocking the export of PD-L1 from the endoplasmic reticulum to the Golgi apparatus: Four N-glycosylation sites (Asn35, Asn192, Asn 200, and Asn 219) present in the extracellular domain of PD-L1 are essential for the stability of the ligand protein, and all other glycosylation sites except Asn35 are critical for its interaction with the receptor. BMS-1166 specifically inhibits partial glycosylation of PD-L1 and inactivates its function by blocking the output of PD-L1 from the endoplasmic reticulum to the Golgi apparatus.
Induce dimerization and internalization of PD-L1: Compound ARB-272572 inhibits the PD-1/PD-L1 axis by inducing dimerization of cell surface PD-L1 through a cis-interacting homodimer that is triggered by rapid internalization into the cytoplasm The rapid loss of PD-L1 on the cell surface prevents further interactions with PD-1-expressing cells.
Binds to PD-L1 without interfering with the formation of PD1: PD-L1 complex: Compared to biphenyl based small molecule inhibitors, CA-170 is highly polar and has a functional antagonistic effect on PD-L1 signaling observed in the cellular environment, while its direct binding to PD-L1 does not destroy PD1: the PD-L1 complex leads to the formation of defective teradic complexes.
Clinical research progress of small molecules targeting PD-1/PD-L1
| Drug | Company | Indications | Current Status |
| CA-170 | Aurigene/Curis | Lymphoma; Advanced solid tumors | Phase 2b/3 |
| INCB-086550 | Incyte Corp | NSCLC urothelial cancer; Renal cell carcinoma | Phase 2 |
| MX-10181 | Maxinovel Pharma | Hepatocellular carcinoma; Melanoma; Advanced solid tumors; Cancer | Phase 1 |
| GS-4224 | Gilead Sciences | Advanced solid tumors; Hepatitis B virus infection | Phase 1 |
| IMMH-010 | Tianjin Chasesun Pharmaceutical Co., LTD | Malignant neoplasms | Phase 1 |
Monoclonal antibody has many inherent disadvantages, including poor oral bioavailability, prolonged tissue retention time and half-life, poor membrane permeability, transportation and storage, and the high cost of antibody drugs is also a problem that cannot be ignored. Therefore, more and more researchers are exploring small molecule drugs as PD-1/L1 inhibitors to circumvent the disadvantages of therapeutic antibodies.
In addition, compared with PD-1/L1 antibodies, oral small-molecule drugs can easily penetrate into tissues, so they can target extracellular and intracellular targets to promote anti-tumor immunity and achieve regulation of the tumor microenvironment. And they typically have a short half-life, reducing the chance of adverse effects.