Structure-Based Design of a Novel Class of Autotaxin Inhibitors Based on Endogenous Allosteric Modulators

Abstract

Autotaxin (ATX) facilitates the hydrolysis of lysophosphatidylcholine to lysophosphatidic acid (LPA), a bioactive phospholipid, which facilitates a diverse range of cellular effects in multiple tissue types. Abnormal LPA expression can lead to the progression of diseases such as cancer and fibrosis. Previously, we identified a potent ATX steroid-derived hybrid (partially orthosteric and allosteric) inhibitor which did not form interactions with the catalytic site. Herein, we describe the design, synthesis, and biological evaluation of a focused library of novel steroid-derived analogues targeting the bimetallic catalytic site, representing an entirely unique class of ATX inhibitors of type V designation, which demonstrate significant pathway-relevant biochemical and phenotypic biological effects. The current compounds modulated LPA-mediated ATX allostery and achieved indirect blockage of LPA 1 internalization, in line with the observed reduction in downstream signaling cascades and chemotaxis induction. These novel type V ATX inhibitors represent a promising tool to inactivate the ATX-LPA signaling axis.

Designing Dual Inhibitors of Autotaxin-LPAR GPCR Axis

“…Lipid-like and non-lipid ATX inhibitors have been reported over the last decade, with co-crystal structures revealed for inhibitors LPA (14:1, PDB: 3NKN , Figure 1 ) and HA-155 (PDB: 2XRG , Figure 2 ) [ 30 , 31 ]. ATX inhibitors can be divided into five different classes based on their binding mode, namely: class I, orthosteric site; class II, hydrophobic pocket; class III, allosteric tunnel; class IV, tunnel-pocket hybrids; and class V, tunnel-active site hybrids, which exhibit a distinct binding mode in the tripartite interacting site in the T-shaped binding pocket of ATX ( Figure 1 ) [ 4 , 32 ]. A significant number of these inhibitors, including HA-155 and PF-8380 (PDB: 5OLB , Figure 2 ), belong to the category of orthosteric-site inhibitors as their mode of action is to completely block the substrate by binding to the active site and the hydrophobic pocket [ 32 , 33 ].…”

“…ATX inhibitors can be divided into five different classes based on their binding mode, namely: class I, orthosteric site; class II, hydrophobic pocket; class III, allosteric tunnel; class IV, tunnel-pocket hybrids; and class V, tunnel-active site hybrids, which exhibit a distinct binding mode in the tripartite interacting site in the T-shaped binding pocket of ATX ( Figure 1 ) [ 4 , 32 ]. A significant number of these inhibitors, including HA-155 and PF-8380 (PDB: 5OLB , Figure 2 ), belong to the category of orthosteric-site inhibitors as their mode of action is to completely block the substrate by binding to the active site and the hydrophobic pocket [ 32 , 33 ]. Hydrophobic pocket ATX inhibitors have been designed to compete with substrate binding without requiring the headgroup aimed for the active site, such as PAT-494 (PDB: 4ZGA, Figure 2 ) [ 4 , 34 ].…”

“…Our group has reported several potent small molecule non-lipid hydrophobic pocket ATX inhibitors, such as 3b and 3f ( Figure 2 ), that exhibit efficacy in vivo [ 1 , 27 , 33 ]. ATX tunnel binders such as UDCA ( Figure 2 ) demonstrate moderate efficacy due to their non-competitive binding to the hydrophobic tunnel [ 32 ]. A recent expansion approach had led to the development of potent tunnel-pocket hybrid inhibitors such as GLPG1690 (PDB: 5MHP , Figure 2 ) [ 2 , 4 , 35 ].…”

“…A recent expansion approach had led to the development of potent tunnel-pocket hybrid inhibitors such as GLPG1690 (PDB: 5MHP , Figure 2 ) [ 2 , 4 , 35 ]. The latest efforts have provided access to the first tunnel-active site hybrid by combining important structural motifs of ATX tunnel binders with that of ATX orthosteric inhibitors (PDB: 7Z0N , Figure 2 ) [ 32 ].…”

“…Clark et al (2022) recently introduced tunnel-pocket hybrid inhibitors (class V) that combine important structural motifs of non-competitive tunnel binders and competitive orthosteric inhibitors (PDB: 7Z0N) [ 32 ]. These inhibitors occupy the tunnel and catalytic site and behave in a competitive manner with the LPC substrate.…”