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Lysophosphatidylserine (LysoPS) is an endogenous pan-agonist of three G-protein coupled receptors (GPCRs): LPS 1 /GPR34, LPS 2 /P2Y 10 , and LPS 3 /GPR174, and we previously reported a series of LysoPS-based agonists of these receptors. Interestingly, we found that LPS 1 agonist activity was very sensitive to structural change at the hydrophobic fatty acid moiety, whereas LPS 2 agonist activity was not. Here, to probe the molecular basis of LPS 2 agonist binding, we developed a new class of hydrophobic fatty acid surrogates having a biphenyl-ether scaffold. The LPS 2 agonist activity of these compounds proved sensitive to molecular modification of the hydrophobic skeleton. Thus, we next constructed an LPS 2 model by homology modeling and docking/molecular dynamics (MD) simulation, and validated it by means of SAR studies together with point mutations of selected receptor amino-acid residues. The putative ligand-binding site of LPS 2 is Γ-shaped, with a hydrophilic site horizontally embedded in the receptor transmembrane helix bundles and a perpendicular hydrophobic groove adjoining transmembrane domains 4 and 5 that is open to the membrane bilayer. The binding poses of LPS 2 agonists to this site are consistent with easy incorporation of various kinds of fatty acid surrogates. Structural development based on this model afforded a series of potent and selective LPS 2 full agonists, which showed enhanced in vitro actin stress fiber formation effect. [Display omitted] • A biphenyl-ether skeleton of LPS 2 -selective agonist was found to amplify the activity change upon structural modifications of the hydrophobic moiety. • Docking studies and point mutation of the receptor elucidated LPS 2 agonist binding mode as representing a hydrophilic site parallel to membrane bilayer and a perpendicular hydrophobic groove between transmembrane helices 4 & 5. • Binding model-guided-ligand design led to the development of potent and selective LPS 2 agonists with improved actin stress fiber formation activity. [ABSTRACT FROM AUTHOR]

The blockade of the PD-1/PD-L1 immune checkpoint pathway with small molecules is an emerging immunotherapeutic approach. A novel series of 4-phenylindoline derivatives were synthesized, and their inhibitory activity against the PD-1/PD-L1 protein-protein interaction (PPI) was evaluated through a homogenous time-resolved fluorescence (HTRF) assay. Among them, A20 and A22 exhibited potent activity with IC 50 values of 17 nM and 12 nM, respectively. Furthermore, A20 showed the promising inhibitory activity against the PD-1/PD-L1 interaction with the EC 50 value of 0.43 μM in a co-culture model of PD-L1/TCR Activator-expressing CHO cells and PD-1-expressing Jurkat cells. Besides, the structure−activity relationships (SAR) of the novel synthesized 4-phenylindoline derivatives was concluded, and the binding mode of A22 with the PD-L1 dimer was analyzed by molecular simulation and docking, demonstrating that the N-atom in the side chain of indoline fragment could interact with the amino acid residue of the PD-L1 protein to lead to the potent inhibitory activity. This study provided a new insight for further drug design. Image 1 • Several indoline derivatives targeting the PD-1/PD-L1 pathway were developed. • A22 inhibited the interaction of PD-1/PD-L1 potently (IC 50 = 12 nM). • A20 and A22 exhibited low toxicity in a co-culture model. • A20 inhibited the binding of PD-1/PD-L1 in a co-culture model (EC 50 = 0.43 μM). [ABSTRACT FROM AUTHOR]

Aberrant activation of the endosomal Toll-like receptor 7 (TLR7) has been implicated in myriad autoimmune diseases and is an established therapeutic target in such conditions. Development of diverse TLR7 antagonists is mainly accomplished through random screening. To correlate human TLR7 (hTLR7) antagonistic activity with the structural features in different chemotypes, we derived a hypothetical binding model based on molecular docking analysis along with molecular dynamics (MD) simulations study. The binding hypothesis revealed different pockets, grooves and a central cavity where ligand-receptor interaction with specific residues through hydrophobic and hydrogen bond interactions take place, which correlate with TLR7 antagonistic activity thus paving the way for rational design using varied chemotypes. Based on the structural insight thus gained, TLR7 antagonists with quinazoline were designed to understand the effect of engagement of these pockets as well as boundaries of the chemical space associated with them. The newly synthesized most potent hTLR7 antagonist, i.e. compound 63 , showed IC 50 value of 1.03 ± 0.05 μM and was validated by performing primary assay in human plasmacytoid dendritic cells (pDC) (IC 50 pDC: 1.42 μM). The biological validation of the synthesized molecules was performed in TLR7-reporter HEK293 cells as well as in human plasmacytoid dendritic cells (pDCs). Our study provides a rational design approach thus facilitating further development of novel small molecule hTLR7 antagonists based on different chemical scaffolds. Image 1 • hTLR7 antagonist binding hypothesis based on docking analysis of known antagonists. • Significance of engaging different pockets and grooves for hTLR7 antagonism. • MD simulation showed ligand binding stability and protein conformational change. • Newly designed ligands based on binding model showed potent hTLR7 antagonism. • Interaction pattern can correlate the structural change in molecule with the activity. [ABSTRACT FROM AUTHOR]