LILRB2-mediated TREM2 signaling inhibition suppresses microglia functions

Abstract

Background Microglia plays crucial roles in Alzheimer’s disease (AD) development. Triggering receptor expressed on myeloid cells 2 (TREM2) in association with DAP12 mediates signaling affecting microglia function. Here we study the negative regulation of TREM2 functions by leukocyte immunoglobulin-like receptor subfamily B member 2 (LILRB2), an inhibitory receptor bearing ITIM motifs. Methods To specifically interrogate LILRB2-ligand (oAβ and PS) interactions and microglia functions, we generated potent antagonistic LILRB2 antibodies with sub-nanomolar level activities. The biological effects of LILRB2 antagonist antibody (Ab29) were studied in human induced pluripotent stem cell (iPSC)–derived microglia (hMGLs) for migration, oAβ phagocytosis, and upregulation of inflammatory cytokines. Effects of the LILRB2 antagonist antibody on microglial responses to amyloid plaques were further studied in vivo using stereotaxic grafted microglia in 5XFAD mice. Results We confirmed the expression of both LILRB2 and TREM2 in human brain microglia using immunofluorescence. Upon co-ligation of the LILRB2 and TREM2 by shared ligands oAβ or PS, TREM2 signaling was significantly inhibited. We identified a monoclonal antibody (Ab29) that blocks LILRB2/ligand interactions and prevents TREM2 signaling inhibition mediated by LILRB2. Further, Ab29 enhanced microglia phagocytosis, TREM2 signaling, migration, and cytokine responses to the oAβ-lipoprotein complex in hMGL and microglia cell line HMC3. In vivo studies showed significantly enhanced clustering of microglia around plaques with a prominent increase in microglial amyloid plaque phagocytosis when 5XFAD mice were treated with Ab29. Conclusions This study revealed for the first time the molecular mechanisms of LILRB2-mediated inhibition of TREM2 signaling in microglia and demonstrated a novel approach of enhancing TREM2-mediated microglia functions by blocking LILRB2-ligand interactions. Translationally, a LILRB2 antagonist antibody completely rescued the inhibition of TREM2 signaling by LILRB2, suggesting a novel therapeutic strategy for improving microglial functions.

Antibody-mediated targeting of human microglial leukocyte Ig-like receptor B4 attenuates amyloid pathology in a mouse model

“…In addition, by administering the anti-LILRB4 mAb ZM3.1 to a transgenic mouse model with Aβ accumulation expressing human LILRB4 in microglia, we decreased Aβ plaque load by enhancing microglial responses to Aβ. Earlier research indicated that another inhibitory LILR family receptor, LILRB2, binds to Aβ, suppresses TREM2 signaling, and plays a role in neurodegeneration, suggesting that blocking LILRB2 could be a potential therapeutic approach in AD ( 29 , 30 ). LILRB4 appears to act through a mechanism different from that of LILRB2.…”

“…LILRB2, which recognizes MHC class I, was found to bind Aβ, and its mouse homolog, PIRB, was shown to contribute to AD neuropathology ( 29 ), suggesting that blockade of LILRB2 function may be therapeutically beneficial in AD. Another study proposed that LILRB2 and TREM2 are cross-linked through shared ligands like oligomeric Aβ and phosphatidylserine ( 30 ) and that antibody-mediated blockade of LILRB2 enhances TREM2 signaling. Moreover, genetic studies have found DNA polymorphisms within the LILRB2 and LILRA2 genes associated with AD and amyotrophic lateral sclerosis, respectively, corroborating a link between the LILR region and susceptibility to neurodegeneration ( 31 , 32 ).…”

Identification and characterization of the conformation and size of amyloid-β (42) oligomers targeting the receptor LilrB2

“…These low-MW and high-MW AbOs were determined by Hu et al 12 as Ab42 oligomers with MWs of 10-15 kDa (about 2-4 mer) and 50-100 kDa (about 14-48 mer) and mixed with a large number of monomers, 12 indicating that the affinity is dependent on the AbO MW and that high-MW AbO exhibits the higher affinity with the LDD receptor. Zhao et al 36 also reported a strong binding between biotinylated AbO and LilrB2, with a dissociation constant of 6.93 Â 10 À8 M. Moreover, Smith et al 37 found that biotinylated oligomers (4.23 Â 10 À8 M) had a higher affinity for LilrB2 than spherical oligomers (2.07 Â 10 À7 M). Taken together, the dissociation constants measured by the above experimental groups vary widely without any explanations.…”

“…35 It can be seen from the binding energy data (Table 1) that the Ab monomer has the weakest binding ability to the LDD receptor, with binding energy and dissociation constants of À6.1 kcal mol À1 and 3.3 Â 10 À5 M, which are consistent with the experimental observation that the LDD receptor does not bind to the Ab monomer. 11,12,36,37 When the AbO species at the SP1 had an intermolecular b-sheet content of less than 15%, the binding energies of AbO to LDD were À11.1 kcal mol À1 (D), À13.0 kcal mol À1 (T), À14.2 kcal mol À1 (Te), À14.9 kcal mol À1 (P), À16.1 kcal mol À1 (H) and À16.3 kcal mol À1 (O), respectively. When the AbO species at the SP2 had an intermolecular b-sheet content of more than 15%, the binding energies between the AbO and LDD receptor were À13.4 kcal mol À1 (D T ), À14.4 kcal mol À1 (T T ), À15.8 kcal mol À1 (Te T ), À16.1 kcal mol À1 (P T ), À17.1 kcal mol À1 (Te F ) and À20.5 kcal mol À1 (P T ), respectively.…”

A tetravalent TREM2 agonistic antibody reduced amyloid pathology in a mouse model of Alzheimer’s disease

“…www.science.org/doi/10.1126/scitranslmed.abq0095 Supplementary Materials and Methods Figs. S1 to S10 Data files S1 and S2 References (76)(77)(78)(79)(80)(81)(82)(83)(84)(85) View/request a protocol for this paper from Bio-protocol.…”