Therapeutic lilrb regulation for treating neurological disorders

Abstract

Disclosed herein are antibodies and / or antibody binding fragments having a leukocyte immunoglobulin-like receptor B (LILRB) binding domain. Also provided herein are methods of modulating the production and / or accumulation of amyloid-β (Aβ) in a subject. Further disclosed herein are recombinant antibodies and methods of using thereof. Additionally described herein are methods for treating and / or preventing a neurological disorder in a subject. The present disclosure further provides transgenic non-human animals having diminished amyloid-β (Aβ) pathology. Further provided are methods of identifying compounds or antibodies for treating and / or preventing a neurological disorder in a subject.

Description

[0001] THERAPEUTIC LILRB REGULATION FOR TREATING NEUROLOGICAL DISORDERS

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS

[0003] This application claims priority to, and the benefit of, U.S. Provisional Application No. 63 / 608,965 filed on December 12, 2023, the disclosure of which is hereby expressly incorporated by reference herein in its entirety.

[0004] REFERENCE TO SEQUENCE LISTING

[0005] The sequence listing submitted on December 12, 2024, as an .XML file entitled “10063- 091WOl_ST26” created on December 12, 2024, and having a file size of 372,752 bytes is hereby incorporated by reference pursuant to 37 C.F.R. § 1.52(e)(5).

[0006] BACKGROUND

[0007] Alzheimer’s disease (AD) is a progressive neurodegenerative disorder characterized by pathognomonic deposition of amyloid-P (AP) fragments within the central nervous system (CNS) parenchyma. Ap is directly neurotoxic, therefore a significant effort has been dedicated to understanding the biology of direct neuronal interaction with Ap. Genome-wide association studies (GWAS) also identified microglia as intimate members of AD pathogenesis, redirecting efforts to understanding their contribution to the pathogenesis and modulating their phenotype to mitigate disease burden. Notably, TREM2 is one of the identified risk genes, with a point mutation resulting in increased risk of late-onset Alzheimer’s disease at a level similar to that of patients with the APOE4 allele.

[0008] In Ap-mediated murine models of AD, TREM2 signaling is responsible for regulating the microglial inflammatory and phagocytic response to Ap. TREM2-dependent processes mediate not only microglial localization to sites of Ap deposits, but also compacting plaques and limiting their diffusion, thereby making the plaques less available to engage in neurotoxicity. The compensatory processes mediated by TREM2 signaling are summed into a microglial transcriptional state termed disease-associated microglia (DAM). DAM are a small population of reactive microglia that seemingly attempt to limit pathogenesis in the CNS in various neurological disorders. In the context of AD, DAM exhibit an activated immune transcriptional phenotype that results in a beneficial reactive microgliosis to prevent otherwise enhanced neurodegeneration caused by Ap deposition within the CNS parenchyma. Importantly, the DAM state is controlled by signaling networks initiated by TREM2 activation.

[0009] Increasing the gene dosage of TREM2 has proven to be beneficial in alleviating plaque burden in 5XFAD mice, as does exogenous activation of TREM2 with a ligand that stimulates the receptor or prevents its shedding. Regulating TREM2 expression and function clearly shows utility in diminishing AD pathology. More importantly, enhancing the TREM2-regulated DAM phenotype in the context of AD could prove to be a powerful therapy to enhance microglial plaqueprocessing capacity and thereby limit neurotoxicity of A .

[0010] Leukocyte Immunoglobulin- Like Receptor B (LILRBs) consist of a family of inhibitory receptors (LILRBL5). Similar to their mouse ortholog, PIRB, LILRBs serve as modulators of immune cell function by downregulating inflammatory processes after activation of their immunoreceptor tyrosine-based inhibitory motif (ITIM). LILRB2 was previously introduced in the context of AD as the human homolog of PIRB, and as a functional receptor for Ap on neurons. However, its role in regulating the microglial phenotype in AD has not yet been characterized.

[0011] The compositions and methods disclosed herein address these and other needs.

[0012] SUMMARY

[0013] In various aspects, disclosed herein is a recombinant antibody, wherein the antibody comprises: a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131; and / or a light chain variable region (VL) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

[0014] In some aspects, the recombinant antibody comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25.

[0015] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

[0016] In some aspects, the recombinant antibody has any one of thes structures:

[0017] VH CDR1 is SEQ ID NO: 27;

[0018] VH CDR2 is SEQ ID NO: 28; VH CDR3 is SEQ ID NO: 29; VL CDR1 is SEQ ID NO: 30; VL CDR2 is SEQ ID NO: 31; and VL CDR3 is SEQ ID NO: 32; or

[0019] VH CDR1 is SEQ ID NO: 33; VH CDR2 is SEQ ID NO: 34; VH CDR3 is SEQ ID NO: 35; VL CDR1 is SEQ ID NO: 36; VL CDR2 is SEQ ID NO: 37; and VL CDR3 is SEQ ID NO: 38; or VH CDR1 is SEQ ID NO: 39;

[0020] VH CDR2 is SEQ ID NO: 40;

[0021] VH CDR3 is SEQ ID NO: 41;

[0022] VL CDR1 is SEQ ID NO: 42;

[0023] VL CDR2 is SEQ ID NO: 43; and

[0024] VL CDR3 is SEQ ID NO: 44; or

[0025] VH CDR1 is SEQ ID NO: 45;

[0026] VH CDR2 is SEQ ID NO: 46;

[0027] VH CDR3 is SEQ ID NO: 47;

[0028] VL CDR1 is SEQ ID NO: 48;

[0029] VL CDR2 is SEQ ID NO: 49; and

[0030] VL CDR3 is SEQ ID NO: 50; or

[0031] VH CDR1 is SEQ ID NO: 57;

[0032] VH CDR2 is SEQ ID NO: 58;

[0033] VH CDR3 is SEQ ID NO: 59;

[0034] VL CDR1 is SEQ ID NO: 60;

[0035] VL CDR2 is SEQ ID NO: 61; and

[0036] VL CDR3 is SEQ ID NO: 62; or

[0037] VH CDR1 is SEQ ID NO: 63;

[0038] VH CDR2 is SEQ ID NO: 64;

[0039] VH CDR3 is SEQ ID NO: 65;

[0040] VL CDR1 is SEQ ID NO: 66;

[0041] VL CDR2 is SEQ ID NO: 67; and

[0042] VL CDR3 is SEQ ID NO: 68; or

[0043] VH CDR1 is SEQ ID NO: 69;

[0044] VH CDR2 is SEQ ID NO: 70;

[0045] VH CDR3 is SEQ ID NO: 71;

[0046] VL CDR1 is SEQ ID NO: 72; VL CDR2 is SEQ ID NO: 73; and

[0047] VL CDR3 is SEQ ID NO: 74; or

[0048] VH CDR1 is SEQ ID NO: 75;

[0049] VH CDR2 is SEQ ID NO: 76;

[0050] VH CDR3 is SEQ ID NO: 77;

[0051] VL CDR1 is SEQ ID NO: 78;

[0052] VL CDR2 is SEQ ID NO: 79; and VL CDR3 is SEQ ID NO: 80; or

[0053] VH CDR1 is SEQ ID NO: 81;

[0054] VH CDR2 is SEQ ID NO: 82;

[0055] VH CDR3 is SEQ ID NO: 83;

[0056] VL CDR1 is SEQ ID NO: 84;

[0057] VL CDR2 is SEQ ID NO: 85; and VL CDR3 is SEQ ID NO: 86; or

[0058] VH CDR1 is SEQ ID NO: 87;

[0059] VH CDR2 is SEQ ID NO: 88;

[0060] VH CDR3 is SEQ ID NO: 89;

[0061] VL CDR1 is SEQ ID NO: 90;

[0062] VL CDR2 is SEQ ID NO: 91; and VL CDR3 is SEQ ID NO: 92; or

[0063] VH CDR1 is SEQ ID NO: 93;

[0064] VH CDR2 is SEQ ID NO: 94;

[0065] VH CDR3 is SEQ ID NO: 95;

[0066] VL CDR1 is SEQ ID NO: 96;

[0067] VL CDR2 is SEQ ID NO: 97; and VL CDR3 is SEQ ID NO: 98; or

[0068] VH CDR1 is SEQ ID NO: 99;

[0069] VH CDR2 is SEQ ID NO: 100; VH CDR3 is SEQ ID NO: 101;

[0070] VL CDR1 is SEQ ID NO: 102;

[0071] VL CDR2 is SEQ ID NO: 103; and

[0072] VL CDR3 is SEQ ID NO: 104; or

[0073] VH CDR1 is SEQ ID NO: 105;

[0074] VH CDR2 is SEQ ID NO: 106;

[0075] VH CDR3 is SEQ ID NO: 107;

[0076] VL CDR1 is SEQ ID NO: 108;

[0077] VL CDR2 is SEQ ID NO: 109; and

[0078] VL CDR3 is SEQ ID NO: 110; or

[0079] VH CDR1 is SEQ ID NO: 111 ;

[0080] VH CDR2 is SEQ ID NO: 112;

[0081] VH CDR3 is SEQ ID NO: 113;

[0082] VL CDR1 is SEQ ID NO: 114;

[0083] VL CDR2 is SEQ ID NO: 115; and

[0084] VL CDR3 is SEQ ID NO: 116; or

[0085] VH CDR1 is SEQ ID NO: 117;

[0086] VH CDR2 is SEQ ID NO: 118;

[0087] VH CDR3 is SEQ ID NO: 119;

[0088] VL CDR1 is SEQ ID NO: 120;

[0089] VL CDR2 is SEQ ID NO: 121; and

[0090] VL CDR3 is SEQ ID NO: 122; or

[0091] VH CDR1 is SEQ ID NO: 123;

[0092] VH CDR2 is SEQ ID NO: 124;

[0093] VH CDR3 is SEQ ID NO: 125;

[0094] VL CDR1 is SEQ ID NO: 126;

[0095] VL CDR2 is SEQ ID NO: 127; and

[0096] VL CDR3 is SEQ ID NO: 128; or VH CDR1 is SEQ ID NO: 129;

[0097] VH CDR2 is SEQ ID NO: 130;

[0098] VH CDR3 is SEQ ID NO: 131;

[0099] VL CDR1 is SEQ ID NO: 132;

[0100] VL CDR2 is SEQ ID NO: 133; and

[0101] VL CDR3 is SEQ ID NO: 134.

[0102] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 14.

[0103] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 8.

[0104] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 10.

[0105] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 2.

[0106] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 6.

[0107] In some aspects, the recombinant antibody comprises at least 1 (e.g., at least 2, at least 3, at least 4, at least 5, at least, 6, at least 7, at least 8, at least 9, or at least 10) amino acid substitutions.

[0108] Also described herein are bispecific antibody including: (i) a first binding arm having a first antigen-binding region that specifically binds to LILRB3, and (ii) a second binding arm having a second antigen-binding region that specifically binds to LILRB2.

[0109] In some aspects, the first antigen-binding region is an agonist of LILRB3 and the second antigen binding region is an antagonist of LILRB2.

[0110] In some aspects, the first antigen-binding region comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, or 93, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, or 94, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, or 95.

[0111] In some aspects, the second antigen-binding region comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 101, 107, 113, 119, 125, or 131. In some aspects, the first antigen-binding region comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 1, 3, 5, 7, 9,

[0112] 11, or 13.

[0113] In some aspects, the second antigen-binding region comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 15, 17, 19, 21, 23, or 25.

[0114] In some aspects, the first antigen-binding region comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 2, 4, 6, 8, 10,

[0115] 12, or 14.

[0116] In some aspects, the second antigen-binding region comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 16, 18, 20, 22, 24, or 26.

[0117] In some aspects, the bispecific antibody further includes a transferrin moiety conjugated by a peptide linker.

[0118] In some aspects, the transferrin moiety comprises at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 180.

[0119] In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 163; a first heavy chain comprising a sequence with at least 60% (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) identity to SEQ ID NO: 164; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 166; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 165.

[0120] In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 167; a first heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 168; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 170; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 169.

[0121] In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 181 ; a first heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 182; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 184; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 183.

[0122] In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 185; a first heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 186; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 188; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 187. In some aspects, the first antigen binding region of the bispecific antibody has any one of thes structures:

[0123] VH CDR1 is SEQ ID NO: 27;

[0124] VH CDR2 is SEQ ID NO: 28;

[0125] VH CDR3 is SEQ ID NO: 29;

[0126] VL CDR1 is SEQ ID NO: 30;

[0127] VL CDR2 is SEQ ID NO: 31; and

[0128] VL CDR3 is SEQ ID NO: 32; or

[0129] VH CDR1 is SEQ ID NO: 33;

[0130] VH CDR2 is SEQ ID NO: 34;

[0131] VH CDR3 is SEQ ID NO: 35;

[0132] VL CDR1 is SEQ ID NO: 36;

[0133] VL CDR2 is SEQ ID NO: 37; and

[0134] VL CDR3 is SEQ ID NO: 38; or

[0135] VH CDR1 is SEQ ID NO: 39;

[0136] VH CDR2 is SEQ ID NO: 40;

[0137] VH CDR3 is SEQ ID NO: 41;

[0138] VL CDR1 is SEQ ID NO: 42;

[0139] VL CDR2 is SEQ ID NO: 43; and

[0140] VL CDR3 is SEQ ID NO: 44; or

[0141] VH CDR1 is SEQ ID NO: 45;

[0142] VH CDR2 is SEQ ID NO: 46;

[0143] VH CDR3 is SEQ ID NO: 47;

[0144] VL CDR1 is SEQ ID NO: 48;

[0145] VL CDR2 is SEQ ID NO: 49; and

[0146] VL CDR3 is SEQ ID NO: 50; or

[0147] VH CDR1 is SEQ ID NO: 57;

[0148] VH CDR2 is SEQ ID NO: 58; VH CDR3 is SEQ ID NO: 59;

[0149] VL CDR1 is SEQ ID NO: 60;

[0150] VL CDR2 is SEQ ID NO: 61; and

[0151] VL CDR3 is SEQ ID NO: 62; or

[0152] VH CDR1 is SEQ ID NO: 63;

[0153] VH CDR2 is SEQ ID NO: 64;

[0154] VH CDR3 is SEQ ID NO: 65;

[0155] VL CDR1 is SEQ ID NO: 66;

[0156] VL CDR2 is SEQ ID NO: 67; and

[0157] VL CDR3 is SEQ ID NO: 68; or

[0158] VH CDR1 is SEQ ID NO: 69;

[0159] VH CDR2 is SEQ ID NO: 70;

[0160] VH CDR3 is SEQ ID NO: 71;

[0161] VL CDR1 is SEQ ID NO: 72;

[0162] VL CDR2 is SEQ ID NO: 73; and

[0163] VL CDR3 is SEQ ID NO: 74; or

[0164] VH CDR1 is SEQ ID NO: 75;

[0165] VH CDR2 is SEQ ID NO: 76;

[0166] VH CDR3 is SEQ ID NO: 77;

[0167] VL CDR1 is SEQ ID NO: 78;

[0168] VL CDR2 is SEQ ID NO: 79; and

[0169] VL CDR3 is SEQ ID NO: 80; or

[0170] VH CDR1 is SEQ ID NO: 81;

[0171] VH CDR2 is SEQ ID NO: 82;

[0172] VH CDR3 is SEQ ID NO: 83;

[0173] VL CDR1 is SEQ ID NO: 84;

[0174] VL CDR2 is SEQ ID NO: 85; and

[0175] VL CDR3 is SEQ ID NO: 86; or VHCDR1 is SEQIDNO: 87;

[0176] VHCDR2 is SEQIDNO: 88;

[0177] VH CDR3 is SEQ ID NO: 89;

[0178] VLCDR1 is SEQ ID NO: 90;

[0179] VL CDR2 is SEQ ID NO: 91; and

[0180] VL CDR3 is SEQ ID NO: 92; or

[0181] VHCDR1 is SEQ ID NO: 93;

[0182] VH CDR2 is SEQ ID NO: 94;

[0183] VH CDR3 is SEQ ID NO: 95;

[0184] VLCDR1 is SEQ ID NO: 96;

[0185] VL CDR2 is SEQ ID NO: 97; and VL CDR3 is SEQ ID NO: 98.

[0186] In some aspects, the second antigen binding region of the bispecific antibody has any one of thes structures:

[0187] VH CDR1 is SEQ ID NO: 99;

[0188] VH CDR2 is SEQ ID NO: 100;

[0189] VH CDR3 is SEQ ID NO: 101;

[0190] VLCDR1 is SEQIDNO: 102;

[0191] VL CDR2 is SEQ ID NO: 103; and

[0192] VL CDR3 is SEQ ID NO: 104; or

[0193] VHCDR1 is SEQIDNO: 105;

[0194] VH CDR2 is SEQ ID NO: 106;

[0195] VH CDR3 is SEQ ID NO: 107;

[0196] VLCDR1 is SEQIDNO: 108;

[0197] VL CDR2 is SEQ ID NO: 109; and

[0198] VL CDR3 is SEQ ID NO: 110; or

[0199] VHCDR1 is SEQIDNO: 111;

[0200] VH CDR2 is SEQ ID NO: 112;

[0201] VHCDR3 is SEQIDNO: 113;

[0202] VLCDR1 is SEQIDNO: 114; VL CDR2 is SEQ ID NO: 115; and

[0203] VL CDR3 is SEQ ID NO: 116: or

[0204] VH CDR1 is SEQ ID NO: 117;

[0205] VH CDR2 is SEQ ID NO: 118;

[0206] VH CDR3 is SEQ ID NO: 119;

[0207] VL CDR1 is SEQ ID NO: 120;

[0208] VL CDR2 is SEQ ID NO: 121; and

[0209] VL CDR3 is SEQ ID NO: 122; or

[0210] VH CDR1 is SEQ ID NO: 123;

[0211] VH CDR2 is SEQ ID NO: 124;

[0212] VH CDR3 is SEQ ID NO: 125;

[0213] VL CDR1 is SEQ ID NO: 126;

[0214] VL CDR2 is SEQ ID NO: 127; and

[0215] VL CDR3 is SEQ ID NO: 128; or

[0216] VH CDR1 is SEQ ID NO: 129;

[0217] VH CDR2 is SEQ ID NO: 130;

[0218] VH CDR3 is SEQ ID NO: 131;

[0219] VL CDR1 is SEQ ID NO: 132;

[0220] VL CDR2 is SEQ ID NO: 133; and

[0221] VL CDR3 is SEQ ID NO: 134.

[0222] Also described herein are nucleic acids encoding the recombinant antibodies or bispecific antibodies described herein. Further provided are recombinant expression cassettes or plasmids comprising a sequence to express the recombinant antibodies or bispecific antibodies described herein. Various aspects further include host cells comprising the expression cassette or the plasmid. Also provided are methods of producing an antibody, comprising cultivating or maintaining the host cell under conditions to produce the antibody.

[0223] In various aspects, described herein are methods of treating and / or preventing a neurological disorder in a subject, comprising administering to the subject a therapeutically effective amount of any of the recombinant antibodies described herein. In some aspects, the neurological disorder is Alzheimer’s disease.

[0224] Further disclosed herein is a method of modulating the production and / or accumulation of amyloid-P (A0) in a subject, comprising: administering to the subject a therapeutically effective amount of an antibody or antibody binding fragment having a leukocyte immunoglobulin-like receptor (LILR) binding domain.

[0225] In some aspects, the leukocyte immunoglobulin-like receptor binding domain is a leukocyte immunoglobulin-like receptor B2 (LILRB2) binding domain and / or leukocyte immunoglobulin-like receptor B3 (LILRB3) binding domain.

[0226] In some aspects, the antibody or antibody binding fragment is an agonist of LILRB2 and / or LILRB3. In some aspects, the antibody or antibody binding fragment is a humanized antibody.

[0227] In some aspects, the antibody or antibody binding fragment specifically binds to LILRB2.

[0228] In some aspects, the antibody or antibody binding fragment specifically binds to LILRB3.

[0229] In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131. In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25. In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

[0230] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

[0231] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 14.

[0232] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 8.

[0233] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 10. In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 2.

[0234] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 6.

[0235] In various aspects, disclosed herein is a method of treating a neurological disorder in a subject, comprising: administering to the subject a therapeutically effective amount of an antibody or antibody binding fragment having a leukocyte immunoglobulin-like receptor (LILR) binding region.

[0236] In some aspects, the wherein the neurological disorder comprises dementia, frontotemporal dementia (FTD), Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), Huntington's disease, taupathy disease, Nasu-Hakola disease, and / or multiple sclerosis.

[0237] In some aspects, the leukocyte immunoglobulin-like receptor binding domain is a leukocyte immunoglobulin-like receptor B2 (LILRB2) binding domain and / or leukocyte immunoglobulin-like receptor B3 (LILRB3) binding domain. In some aspects, the antibody or antibody binding fragment is an agonist of LILRB2 and / or LILRB3. In some aspects, the antibody or antibody binding fragment is an antagonist of LILRB2 and / or LILRB3. In some aspects, the antibody or antibody binding fragment is a humanized antibody.

[0238] In some aspects, the antibody or antibody binding fragment specifically binds to LILRB2. In some aspects, the antibody or antibody binding fragment specifically binds to LILRB3.

[0239] In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131.

[0240] In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25.

[0241] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

[0242] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

[0243] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 14.

[0244] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 8.

[0245] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 10.

[0246] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 2.

[0247] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 6. Also described herein is a transgenic non-human animal having diminished amyloid-p (Ap) pathology, wherein the genome of the animal comprises LILRB2 and / or LILRB3 human transgenes. In some aspects, said animal is a mammal. In some aspects, said animal is a rodent.

[0248] In some aspects, said animal is a 5XFAD mouse. In some aspects, the LILRB2 and / or LILRB3 human transgenes comprise bacterial artificial chromosome (BAC) inserted transgenes. In some aspects, said transgenic animal exhibits anti-inflammatory characteristics as compared to a wild-type animal.

[0249] In some aspects, said transgenic animal is a 5XFAD mouse or progeny thereof, and wherein said 5XFAD mouse or progeny thereof exhibits anti-inflammatory characteristics as compared to a 5XFAD mouse not having LILRB2 and LILRB3 human transgenes.

[0250] Also provided herein are cells isolated from any of the the transgenic non-human animals described herein. In some aspects, said cell is a myeloid cell. In some aspects, said cell is a microglia. In some aspects, said cell is a disease-associated microglia (DAM). In some aspects, said DAM exhibits increased expression of Sppl, Lpl, and / or Lilrb4a compared to DAM of a 5XFAD mouse not having LILRB2 and LILRB3 human transgenes.

[0251] In various aspects, described herein is a method of identifying a compound for treating and / or preventing a neurological disorder in a subject, the method comprising: administering the compound to any of the transgenic non-human animals described herein or contacting the compound with any of the isolated cells described herein; and determining a therapeutic and / or prophylactic efficacy of the compound on the neurological disorder. In some aspects, the neurological disorder comprises a neuroinflammatory disorder. In some aspects, the neurological disorder comprises dementia, frontotemporal dementia (FTD), Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), Huntington's disease, taupathy disease, Nasu-Hakola disease, and / or multiple sclerosis. In some aspects, the neurological disorder is Alzheimer’s disease. In some aspects, the subject is a human subject.

[0252] In some aspects, the compound comprises an LILRB- specific antibody or antibody fragment. In some aspects, the compound comprises an LILRB2 antagonist. In some aspects, the compound comprises an LILRB3 agonist. In some aspects, the compound comprises an LILRB2 antagonist. In some aspects, the compound comprises an LILRB3 antagonist.

[0253] Additional advantages will be set forth in part in the description that follows, and in part will be obvious from the description, or may be learned by practice of the aspects described below. The advantages described below will be realized and attained by means of the elements and combinations particularly pointed out in the appended claims. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive.

[0254] BRIEF DESCRIPTION OF DRAWINGS

[0255] The accompanying figures, which are incorporated in and constitute a part of this specification, illustrate several aspects described below.

[0256] Figures 1A-1H show that PIRB and LILRB can modulate TREM2 expression and inflammasome activation Pirb KD in the murine macrophage (Figure 1A; RAW264.7) and microglial (Figure IB; BV2) cell lines diminishes expression of Trem2. (Figure 1C) Under inflammatory stimulus, such that under NLRP3 activating conditions, Trem2 expression is further diminished in BV2 cells with Pirb KD (multiple comparisons were corrected by controlling for FDR). (Figure ID) PIRB7’ mice show a diminishing of TREM2 expression on the cell surface of bone marrow-derived macrophages (CD1 lb+F4 / 80+cells; BMDMs; top panel). Similar to the BV2 cells, under inflammatory conditions, the loss of TREM2 from the cell surface is exacerbated under inflammatory stimuli, such as activation of the NLRP3 inflammasome (middle panel) or under typical Ml-activating culture conditions with lOOng / mL LPS + 20ng / mL IFNy (lower panel; n = 3 mice per group). (Figure IE) After activating the NLRP3 inflammasome in both WT and PIRB’ABMDMs, the PIRB7’ mice show about a 2-fold increase in IL-1 P output. (Figure IF) 5XFAD mice were bred with mice containing BAC-inserted LILRB2 and LILRB3 transgenes (FB2 / 3 mice), and their expression is present on myeloid cells (CDl lb+cells) isolated from peripheral blood of respective mice; gating was done on viable cells -> single cells - CD1 lb+cells. Regular 5XFAD mice do not express human LILRB2 or LILRB 3 and were used as negative controls (top panel). (Figure 1G) Under NLRP3 activating conditions, viable BMDMs (CDl lb+F4 / 80+) from FB2 / 3 mice showed an enhanced expression of TREM2 on the cell surface, and (Figure 1H) diminished secretion of IL- 1 , compared to 5XFAD mice (n = 6 mice per group). Controls (CT) were normalized to 1.0. * p < .05; ** p < 0.005; **** p < 0.0001

[0257] Figures 2A-2J show that microglia from FB2 / 3 have enhanced transcription of DAM genes. (Figure 2A) Microglia from 8-month-old male 5XFAD and FB2 / 3 mice were isolated by Dounce homogenization on ice. Debris was removed by resuspending the cells in a 40% Percoll solution and centrifugation. CD1 lb+cells were then isolated using anti-PE microbeads by Miltenyi Biotec, and qPCR was done on positively selected cells. (Figure 2B) & (Figure 2C) Microglia from FB2 / 3 mice show expression of human LILRB2 and LILRB3, unlike 5XFAD microglia (n = 6 mice per group). Respective DAM genes for LILRB2 (Figure 2B), L1LRB3 (Figure 2C), Trem2 (Figure 2D), Tyrobp (Figure 2E), Itgax (Figure 2F), Cst7 (Figure 2G), Lpl (Figure 2H), Lilrb4a (Figure 21), and Axl (Figure 2J) were measured by qPCR. * p < 0.05; ** p < 0.005; *** p < 0.0005; **** p < 0.0001, by nested Student’s t-test.

[0258] Figures 3A-3H show that LILRB transgenes enhance the expansion of DAM. (Figure 3 A) CD45+cells from male 8mo mice were isolate by Dounce homogenization, n = 2 mice per group. (Figures 3B-3D) Twelve distinct clusters were identified with the Seurat package, three of which represented peripherally infiltrating immune cells. (Figure 3C) None of the clusters were enhanced for ex -vivo activation markers that represents improper tissue handling during isolation (Marsh et al., 2022 #29). (Figure 3E) After pruning the peripheral immune cells from analysis, the remaining microglial cells were reclustered and (Figure 3F) homeostatic microglial populations HM1 and HM2 represented the largest proportions, with DAM stage 1 (DAM1), DAM stage 2 (DAM2), transitioning microglia (TrM), interferon-responsive microglia (IRM), antigen-presenting microglia (AgM), and proliferating microglia (PM) also identified. (Figures 3G-3H) Cluster proportions were altered between 5XFAD and FB2 / 3 mice. (Figure 3H) Statistically significant differences in cluster frequencies are marked in orange, defined by p-value adjusted by FDR < 0.05; gray points are not significant.

[0259] Figures 4A-4F show that FB2 / 3 microglia express DAM markers at earlier transition stages. (Figure 4A) TSNE plot of the trajectory analysis output generated by tradeSeq. (Figure 4B) A pseudotime analysis plot using Slingshot is shown in the left panel. The right panel displays clusters HM2, HM1, DAM 1, and DAM 2 compared between 5XFAD and FB2 / 3 mice across pseudotime. (Figure 4C) Apoe (the top TEG) and (Figure 4D) Trem2 expression is presented across pseudotemporal transition from HM2 to DAM 2, compared between 5XFAD and FB2 / 3. (Figure 4E) Differentially expressed genes in the HM2 cluster. (Figure 4F) Differentially expressed genes in the DAM 1 cluster.

[0260] Figures 5A-5I show that FB2 / 3 mice exhibit diminished AD-associated neuropathology. Ap and neurofilament staining was performed in wild-type (WT; Figure 5 A), 5XFAD (Figure 5B), and FB2 / 3 (Figure 5C) mice. Figures 5D -5E show a comparison of cortical (Figure 5D) and hippocampal (Figure 5E) AP immunoreactivity, respectively. * < 0.05 by Student’s t-test with Tukey’s correction for multiple comparisons. Figures 5F-5G show a comparison of cortical (Figure 5F) and hippocampal (Figure 5G) hyperphosphorylated neurofilament immunoreactivity, respectively. * < 0.05 by Student’s t-test. (Figure 5H) Simple linear regression between cortical hyperphosphorylated neurofilament and A immunoreactivity. (Figure 51) Simple linear regression between hippocampal hyperphosphorylated neurofilament and Ap immunoreactivity. Figures 6A-6G show PIRB / _genotyping. Murine immune cells have both PIR receptors on the cell surface (PIRA and PIRB), apart from CD19+cells that only express PIRB. (Figure 6 A) Using CD19+that weren’t stained for PIRA / B as a control, CD19+cells from (Figure 6B) WT and (Figure 6C) PIRB / _mice were stained for PIRA / B. (Figure 6D & Figure 6E) Histograms from respective mice show a loss of PIRA / B staining on the surface of CD19+cells. (Figure 6F) hMDMs were pretreated with respective antibodies and then stimulated with LPS prior to determining TREM2 expression by flow cytometry. (Figure 6G) hMDMs were pretreated with respective antibodies prior to activation of the NLRP3 inflammasome with LPS+ATP. IL- 10 was determined via ELISA.

[0261] Figures 7A-7E show that microglia from FB2 / 3 have enhanced transcription of DAM genes. (Figure 7 A) & (Figure 7B) Representative flow cytometry diagrams showing CDl lb+& CDllb" cells by MACS. Only the CDl lb+cells were used in the qPCRs. (Figure 7C) Ctsb expression was measured by qPCR; n = 5 in the 5XFAD group and n = 4 in the FB2 / 3 group. (Figure 7D) P2ryl2 is a homeostatic marker measured by qPCR, n = 6 per group. (Figure 7E) Irf8 was measured by qPCR in n = 4 mice per group. * p < 0.05; **** p < 0.0001, by nested Student’s t-test.

[0262] Figures 8A-8O show the identification of immune cell subsets. (Figure 8A) Live, singlet, CD45+cells were isolated from each mouse. (Figure 8B & Figure 8C) Activation markers from Marsh et al., 2022 (#29) were measured in the pruned microglial clusters. (Figure 8D) Monocyte / macrophage genes (Ms4a7, Ccr2, and Ms4a4c) were expressed in cluster 9, therefore they were labeled as such. (Figure 8E) B cells are represented in cluster 10, showing highest expression of Cd79a and Ebfl. (Figure 8F) Cluster 11 showed highest expression of Ngp, and therefore was labled as an infiltrating granulocytic cell cluster. (Figure 8G, Figure 8H, Figure 81, Figure 8J) Different definitions of homeostatic microglia were used to identify the HM1 and HM2 populations. (Figure 8K) IRMs were identified using the gene set Ifit, Ifit3, Irf7, and Oasl2. (Figure 8L) Antigen presentation microglia showed highest expression of Cd74, H2-Aa, H2-Abl, Cd52, Cst7 genes. (Figure 8M) Proliferating microglia showed highest expression of Top2a, Mki67, Birc5. (Figure 8N & Figure 80) DAM stage 1 (Apoe, Tyrobp, B2m, Ctsb, Timp2, H2-D1, Fthl, Lyz2, Cstb, Ctsd) & 2 (Ank, Sppl, Axl, Csfl, Cst7, Cd9, Itgax, Lpl, Ctsz, Ctsl, Ccl6, Cadml, Cd63, Ctsa, Serpine2, Ctsz, Cd52, Hifla) gene sets were scored against the remaining clusters to identify TrM, DAM 1 and DAM 2.

[0263] Figures 9A-9I show FB2 / 3 microglia have enhanced DAM gene set expression. (Figure 9 A) Violin plots showing scored expression of the homeostatic microglia gene set in 5XFAD compared to FB2 / 3 mice, using Seurat’ s built-in AddModuleScore feature, (Figure 9B) Violin plots scoring DAM programming expression in the HM1 cluster. (Figure 9C) Violin plots scoring DAM programming in the HM2 cluster. (Figure 9D) Violin plots scoring DAM programming in the DAM 1 cluster. (Figure 9E) Violin plots scoring DAM programming in the DAM 2 cluster. Respective scores from each of the violin plots are shown in Table 1. (Figure 9F, Figure 9G, Figure 9H, Figure 91) Violin plots showing expression of Irf8 in HM1, HM2, DAM 1, and DAM 2 clusters, respectively.

[0264] Figures 10A-10H show microglial trajectory and differential expression. (Figure 10A) Pseudotime analysis using HM1 as the anchoring cluster. Figures 10B-F depict multiple predicted trajectories (Tl, Figure 10B); (T2, Figure IOC); (T3, Figure 10D); (T4, Figure 10E); (T5, Figure 10F) using HM2 as the anchoring cluster. Tl was used as the final projection in Figure 4A. Figures 10G-10H are volcano plots showing differential gene expression between 5XFAD and FB2 / 3 mice in clusters HM1 (Figure 10G) and AgM (Figure 10H) clusters, respectively. Enrichment in the FB2 / 3 cluster is represented as a positive value. Significance is denoted by p-value correction by FDR < 0.05.

[0265] Figures 11A-11H show FB2 / 3 mice exhibit less parenchymal A deposition. Immunohistochemical staining with H32121 for A0 deposition followed by development by DAB in 5XFAD (Figure 11 A) and FB2 / 3 (Figure 1 IB) mice. (Figure 11C & Figure 1 ID) Representative quantification of A0 immunohistochemical staining from Figure 11A & Figure 1 IB. (Figure HE & Figure 1 IF are graphs appropriated from Figure 5D-G, highlighting the sexes of individual mice - orange circles denote male mice; blue circles denote female mice. * < 0.05 by Student’s t-test.

[0266] Figure 12 shows a plot depicting the statistical significant enrichment of gene clusters between 5XFAD and FB2 / 3 models.

[0267] Figures 13A-13E show that basal LILRB2 / 3 expression in myeloid cells and LILRB2 and 3 agonists decrease microglia and MDM inflammation and promote an M2 functional phenotype. (Figure 13 A) LILRB1, B2, B3, B4 and B5 expression in cultured organoid neuron and astrocyte tissue. The results indicate a very low level of LILRB3 expression in neurons and astrocytes. LILRB1 and LILRB4 are also expressed, but at a relatively low level. (Figure 13B) Human monocytes were differentiated into macrophages using 50 ng / mL M-CSF for 7 days, then incubating with 100 ng / mL LPS for 24 hrs, in the presence of control Ig or the agonist antibodies developed. Data represents one of multiple reproducible experiments using macrophages from healthy human donors. TNFa, and IL-ip were measured by ELISA. (Figure 13C) Phagocytosis assay of A filament or fluorescent bead by MDM in the presence of anti-LILRB3 agonist, anti- LILRB2 antagonist, or the combination. The cells were stained with methoxy-X34 and quenched with Trypan blue to detect Ab inside the cells. Average Mean fluorescent intensity (MFI) results from three experiments are presented. (Figure 13D) Cell surface staining for phenotypic changes after antibody treatment for 48 hours. CD206 / CD86 positive population is presented. *** p<0.001 , ** P<0.005, * p<0.05, NS: not significant. (Figure 13E) 2B4 reporter cell system for Ap filament activation through a specific LILRB2 receptor to test effect of anti-LILRB2 antibody on the blocking of coated A filament activation on the reporter cells. One representative of multiple reproducible assays is presented.

[0268] Figure 14 shows a mechanism by which LILRB3 signal disrupts neuroinflammation. Signaling through LILRB3 can diminish NFKB activation and diminish activation of the NLRP3 inflammasome. Additionally, LILRB3 signaling results in the rescuing of Trem2 expression under inflammatory stimuli and enhances expression of disease-associated microglial (DAM) genes responsible for attenuating AD pathogenesis in vivo.

[0269] Figures 15 A- 15C show that AP2 peptide conjugation facilitates blood brain barrier passage of LILRB3 antibody. (Figure 15A) Cy5-labeled LILRB3 antibody (Ab only) and its conjugate to BBB-penetrating peptide AP2 (Ab-AP2) were injected to mice and tested for tissue distribution by IVIS (left panels), *p<0.05 and (Figure 15B) confocal microscopy (right panels, 40x). 4h postinjection, increased amounts of Ab-AP2 compared to Ab only were detected at endothelial layer (CD31+) cells of brain (upper right panels). (Figure 15C) At 48h time-point, a large fraction of Ab-AP2(+) were located within brain parenchyma, while some were still close to endothelial cells of blood vessels (lower right panel).

[0270] Figures 16A-16B show that agonist anti-LILRB3 inhibits inflammatory response in vivo (Figure 16A) The CyTOF analysis further supports that anti-LILRB3 can promote the M-CSF cultured bone marrow myeloid cells’ proliferation and differentiation toward M2 phenotypes in red circle, e.g. upregulation of CD14 / CD16, CD163 expression and downregulation of C64, and CDlc. (Figure 16B) anti-LILRB3 treatment on the inhibition of LPS-mediated pro-inflammatory cytokines production in humanized mouse model. NSG-SGM3 mice were transplanted with 1 x 107human PBMCs at day 0. aLILRB3, or control Ig (150 pg / mouse) were given two injections on days 0 and 3. On day 5, mice were challenged with LPS (2 mg / kg). Blood serums were collected at 2 hours after LPS challenging. Serum pro-inflammatory cytokines were determined by ELISA. The data from two reproducible experiments are presented.

[0271] Figures 17A-17F show a microglial single-cell dynamics by trajectory analysis. (Figure 17A) FB2 / 3 mice microglia signatures are enriched in the MSBB human AD DEGs. Barplot showing the P value significance of the enrichment in mice signatures for CDR (red), CERAD (green), and plaque (blue) associated DEGs in AD brains. (Figure 17B) LILRB2 / 3 and TREM2 microglial expressions co-localize in the red-circled subpopulation. (Figure 17C) Pseudo-temporal microglial trajectory in FAD an FB2 / 3 mice. The subpopulations include homeostatic microglia (HM1 / 2), disease-associated microglia stage 1 and 2 (DAM1 / 2), interferon-responsive microglial (IRM), proliferative microglia (PM), transitioning microglia from HMs to DAMs (TrM) and antigen-presenting microglia (AgM). These cells are color-coded in the legend next to Figure 17E. (Figure 17D) Cell proportions in FAD and FB2 / 3. Significantly enriched population in each model (FDR < 0.05 by Monte-Carlo permutation test) are marked by red dot. (Figure 17E) Trem2 and Apoe expression changes along the microglial trajectory in Figure 17C. (Figure 17F) Spatially resolved transcriptome (SRT) of FB2 / 3 mouse (Left column) and FAD mouse (Right column). Granule neurons are highlighted by dotted rectangles.

[0272] Figure 18 shows the effect of FAD mouse vs. FAD with LILRB2 / 3 trangenic mice (FAD B2 / 3) on beta amyloid phagocytosis by microglia at 9-month-old male mice by using Methoxy- X04 dye staining, which can be used for in vivo Ap labeling. The proportion of phagocytic microglia were obtained around 20 % at 9 month-old 5xFAD mice. The age matched FB2 / 3 mice significantly increase the proportion of phagocytic microglia to around 34%. p value is <0.05.

[0273] Figure 19 shows beta amyloid can activate LILRB2+2B4 reporter cells and antiLILRB2 antagonist antibody can block the beta amyloid mediated reporter cell activation. LILRB2+2B4 reporter cells, which express the LILRB2 extracellular domain with DAP-12-associated NFAT GFP reporter were cultured in the presence beta- amyloid for 16 hours. GFP+cells were enumerated by flow cytometry. Multiple Anti-LILRB2 Ab (aLILRB2) can inhibit beta-amyloid induced LILRB2 activation in RB2+ reporter cells, but not the beta amyloid or control Ig.

[0274] Figure 20 shows human PBMCs were treated with anti-LILRB3 agonist antibodies overnight and then cells were stimulated with LPS for 6 hours, supernatant were analyzed for the the human TNF-alpha by ELISA. The human PBMCs were treated with anti-LILRB3 antibodies overnight and then cells were stimulated with anti-human CD3 for 72 hours, supernatant were analyzed for the human IFN-gamma by ELISA. The anti LILRB3 agonist show the inhibition of TNFalpha and IFNgama secretion.

[0275] Figures 21A-21B. LILRB2 antagonistic mAb (2D6) combine with LILRB3 agonistic mAb (3B1) significantly protect against A-beta amyloid -mediated neuron damage in neuron / macrophage cocultured system. (Figure 21 A) The morphology of neurons which isolated from day 0 newborn FB2 / B3 mice were observed under the microscope after treatment with 500 nM beta-amyloid for 96 hours and in the presence of anti-LILRB2 antagonist, anti-LILRB3 agonist and isotype matched control. Three fields of view were randomly photographed per well, triplicate wells per group. (Figure 2 IB) The phagocytosis of Ab polymers by myeloid cells isolated from FB2 / B3 mice was assessed by FACS after staining with 500 nM Ab-xethoxy-X04 for 24 hours and in the presence of anti-LILRB2 antagonist, anti-LILRB2 agonist, anti-LILRB3 agonist (3B1) and isotype matched control.

[0276] Figures 22A-22C. Screening of LILRB2-binding monoclonal antibodies. (Figure 22A) THP-1 parental cells (gray filled) and THP-1 cells overexpressing LILRB2 (black line) were incubated with Angptl5-His (R&D Systems) followed by staining with anti-His-PE secondary antibody (Miltenyi) per manufacturer’s instructions. THP-1 cells overexpressing LILRB2 (red line) were preincubated with commercial antibodies or in-house generated anti-LILRB2 antibodies for 30 minutes followed by Angptl5-His and then anti-His-PE. (Figure 22 A lower panel) THP-1 cells overexpressing LILRB2 were incubated with streptavidin- APC control (gray filled) or HLA- B APC tetramer (NIH) followed by flow cytometric analyses. To test blocking activities, THP-1 cells overexpressing LILRB2 (red line) were preincubated with commercial or in-house generated anti-LILRB2 antibodies for 30 minutes followed by HLA-B APC tetramer staining. (Figure 22B) The LILRB2 antibody clones can block LILRB2-MHCI receptor- ligand interactions. THP-1 cells transduced with LILRB2 or control vector were stained with titrated PE-conjugated HLA-B tetramer and analyzed by flow cytometry. (Figure 22C) LILRB2+THP-1 cells pre-treated with titrated concentrations of clones A and B were stained with saturating levels of PE-tetramer then analyzed by FACS.

[0277] Figures 23A-23F. Pirb regulates Trem2 expression and NLRP3 activity. Pirb was genetically knocked-down using siRNA in (Figure 23A) RAW264.7 cells (macrophages) and (Figure 23B) BV2 cells (microglia), and TREM2 expression was measured by RT-qPCR compared to control (CT). (Figure 23C) BV2 cells were incubated with 50 ng / mL LPS for 3 hers and 5 mM ATP for the last 30 mins (NLRP3 stimulation). (Figure 23D) BMDMs from mice with total PIRB KO were differentiated with 50 ng / mL M-CSF for 5 days, ± 100 ng / mL LPS for 3 hrs, and (Figure 23E) 5 mM ATP for the last 30 mins (NLRP3) or ± lOOng / mL LPS and 20ng / mL IFNy overnight. TREM2 was measured in viable, singlet, CDl lb+F4 / 80+cells by flow cytometry and IL-ip was measured by ELISA and (Figure 23F) Ml-like conditions (100 ng / mL LPS + 20 ng / mL IFNy) *p<05, **p<005, ***p<.0005 by unpaired Student’s t-test.

[0278] Figures 24A-24C. Human LILRB2 / 3 expression affects TREM2 expression and IL-ip secretion. BMDMs isolated from 5XFAD, and FB2 / 3 mice were cultured for 5 days in 50 ng / mL M-CSF and stimulated with 100 ng / mL LPS for 3 hrs., followed by 5 mM ATP for the last 30 mins. (Figure 24A) Representative blood myeloid cell LILRB2 / 3 expression phenotypes from 5XFAD (top) and FB2 / 3 (bottom) are shown. Gating was done on viable CDl lb+cells. (Figure 24B) ELISA was performed to measure IL- 13 in the supernatant. (Figure 24C) TREM2 was measured by flow cytometry on viable cells CDl lb+F4 / 80+gated cells. * p<.05, **p <0.005 by paired Student’s t-test.

[0279] Figures 25A-25E. LILRBs in microglia enhances DAM- associated gene expression. Microglia were isolated from the brains of 8-month-old WT, B2 / B3 BAC transgenic mice, 5XFAD and FB2 / 3 mice using CDl lb+cell magnetic bead separation method. Gene expression was measured by RT-qPCR. (Figure 25A) TREM2 expression, (Figure 25B) Trem2 dependent DAM genes (Figure 25C) Trem 2 independent genes (Figures 25D-25E) (n = 6 mice per group). Respective DAM genes were measured by qPCR. * p < 0.05; ** p < 0.005; *** p < 0.0005; **** p < 0.0001, by nested Student’s t-test.

[0280] Figures 26A-26H. FB2 / 3 mice exhibit diminished AD-associated neuropathology. (Figure 26A) A0 phagocytic microglia were detected by FACS in 9 months-old male 5xFAD and FB2 / 3mice. 8 mg / kg of Methoxy X-O4 were injected to stain the Ab through intraperitoneal route at 3 hrs before sacrifice. The microglia cells were gated on CD45 medium and CD11 b+ cells. The % of X-O4 positive cells were presented. (Figure 26B) A0 and neurofilament staining was performed in wild type (WT), 5XFAD, and FB2 / 3 mice. (Figure 26C, Figure 26D) Comparison of cortical and hippocampal Ap immunoreactivity, respectively. * < 0.05 by Student’s t-test with Tukey’s correction for multiple comparisons. (Figure 26E, Figure 26F) Comparison of cortical and hippocampal aberrant hyperphosphorylated neurofilament immunoreactivity, respectively. * < 0.05 by Student’s t-test. (Figure 26G) Simple linear regression between aberrant cortical hyperphosphorylated neurofilament and Ap immunoreactivity. (Figure 26H) Simple linear regression between hippocampal hyperphosphorylated neurofilament and Ap immunoreactivity.

[0281] Figures 27A-27D. LILRB2 but not LILRB3 expression on neuron cells. (Figure 27 A) LILRB1, B2, B3, B4 and B5 expression in cultured human organoid neuron and astrocyte tissue. The results indicate a very low level of LILRB3 expression in neurons and astrocytes. Only LILRB5, LILRB1 and LILRB4 have some expression, but at a relatively low level. (Figure 27B) Ap fdament activation through a specific LILRB2 receptor. This system has been used to identify the anti-LILRB2 antibody on the blocking of Ap filament mediated GFP activation on the 2B4 reporter cell system through the co-transfection of DAP 12 fusion construct and NF AT GFP reporter construct in 2B4 cells. (Figure 27C) Ab filament as LILRB2 ligand can activate the reporter cells. The two different clone 1 and 2 of anti-LILRB2 can block the Ab mediated reporter cell activation. (Figure 27D) LILRB2 antagonistic mAb significantly protect against Ab mediated neuron toxicity. One representative of multiple reproducible assays is presented. The morphology of neurons isolated from day 0 newborn FB2 / 3 mice were observed under the microscope after treatment with 5 ug / ml anti-LILRB2 or anti-LILRB3 Ab and isotype matched control for 96 hours. Three fields of view were randomly photographed per well, triplicate wells per group. The cell viability numbers were averaged and calculated by student T test, *** P<0.001.

[0282] Figures 28A-28D. LILRB2 antagonistic mAb (2D6) combined with LILRB3 agonistic mAb (3B1) significantly protect against Ab-mediated neuron damage in the presence of macrophage and neuron co-cultured system. (Figure 28 A) LILRBs gene expression in the microglia cells isolated from B2 / B3 transgenic mice. (Figure 28B) Microglia cells enhance the phagocytosis of Ab in the presence of anti-LILRB3 antibody after the co-staining of Methoxy- X04. (Figure 28C) Combination of anti-LILRB2 and B3 in the presence macrophage to protect the Ab mediated neuron toxicity. The morphology of neurons isolated from day 0 newborn FB2 / 3 mice were observed under the microscope after treatment with / wo 500 nM Ab for 96 hours and in the presence of 5 ug / ml of anti-LILRB2 antagonist, anti-LILRB3 agonist and isotype matched control. Three fields of view were randomly photographed per well, triplicate wells per group. (Figure 28D) The combination of antiLILRB2 and LILRB3 enhance the microglia phagocytosis of Ab in FB2 / 3 mice. 5- and 6-month-old male FB2 / 3 mice were injected with / wo transferrin conjugated anti-LILRB2 / B3 antibody iv. at every four-day interval and total three dosses, then injected with Methoxy x04 for 3 hrs before isolating the microglia from the brain tissues of treated mice. The % of methoxy positive microglia cells were calculated by flow cytometry.

[0283] Figures 29A-29E. Transferrin enhances the antibody penetration through BBB and the combination of anti-LILRB2 antagonist (2D6) with LILRB3 agonist mAb (3B1) significantly protects against Ab accumulation and enhances the microglia infiltration in vivo. (Figure 29 A) Transferrin conjugated anti LILRB2 / LILRB3 antibody vs. native antibody conjugated fluorescent OTT 565 and injected into the FB2 / 3 mice for 48 hours (5-10 mice per group). Frozen tissue sections were co-stained with DAPI and quantified with florescent antibody for penetration into the brain tissue. The positive cells were quantitated by image J. (Figure 29B) 6 months old FB2 / 3 mice were treated with transferrin conjugated IgG, or anti-LILRB2 or anti-LILRB3 or combination at dose 150 ug / mouse for three times and each dose at four days intervals through i.v. The Ab were co-stained with Thioflavin-S and DAPI for nucleus. Random field in the cortex were presented. (Figure 29C) Quantitation of multiple areas with average positive pixels were presented. (Figure 29D) The microglia were co-stained with Iba-1 and Alexa568 and random field from Cortex were presented. (Figure 29E) Quantitation of positive microglia in multiple areas (cell density) were presented and T test was performed.

[0284] Figures 30A-30C. (Figure 30A) Schematic diagram of the bi-functional anti-LILRB3 and anti-LILRB2 with transferrin fusion protein (HMR301) for enhancing microglia and MDM mediated phagocytosis and inhibiting the Ab mediated neuron damage. (Figure 30B) bNDES cross-section showing HMR301 slow elution. (Figure 30C) NDES for intraparenchymal inoculation depiction with trocar insertion method.

[0285] Figures 31 A-31C. (Figure 31 A) 3D IVIS image of intraperitoneal (IP), bolus intratumorally (IT) injection of recombinant protein and nanoparticle (NDES) slow release of AF700-CD40 signal in 4T1 mice, with signal quantification bar graph below. AF700-CD40 signal in (Figure 3 IB) tumor and (Figure 31C) liver on day 7 measured ex vivo via IVIS.

[0286] Figures 32A-32B. (Figure 32A) Barnes Maze assay for assessing 8-month-old, homozygote, and male 5XFAD vs. FB2 / 3 mice or control LILRB2 / B3 transgenic mice or wildtype mice. FB2 / 3 mice spent more time in the target quadrant compared to 5XFAD mice. (Figure 32B) Age-dependent deficits in object location spatial memory in 6-month-old APP mice. Mice were trained with two flasks in set positions (3 trials, 3 min each, 3 min intertrial interval). After 3 min, mice were placed back in the arena, but one object was displaced to the opposite corner. The percent time exploring the displaced object was compared between training and testing phases, which occurred 24 hours after the last training trial. 6 months old no transgenic wildtype mice (NTG) demonstrated object location memory by spending more time with the displaced object (DO) during the testing trial, whereas APP mice did not, suggesting impaired objection location spatial memory. n=6-10-genotype.

[0287] Figures 33A-33B. Agonist anti-LILRB3 inhibits inflammatory response in vivo. (Figure 33A) The CyTOF analysis further supports that anti-LILRB3 can promote the M-CSF cultured bone marrow myeloid cells’ proliferation and differentiation toward M2 phenotypes in red circle, e.g., upregulation of CD14 / CD16, CD163 expression and downregulation of C64, and CDlc. (Figure 33B) anti-LILRB3 treatment on the inhibition of LPS-mediated pro-inflammatory cytokines production in humanized mouse model. NSG-SGM3 mice were transplanted with 1 x 107human PBMCs at day 0. Two injections of aLILRB3 or control Ig (150 pg / mouse) were given on days 0 and 3. On day 5, mice were challenged with LPS (2 mg / kg). Blood serums were collected at 2 hours after LPS challenging. Serum pro-inflammatory cytokines were determined by ELISA. The data from two reproducible experiments are presented. Figures 34A-34J. Microglial single-cell dynamics by trajectory analysis. (Figure 34A) Pseudo-temporal microglial trajectory in FAD and FB2 / 3 mouse. The subpopulations include homeostatic microglia (HM1 / 2), disease- associated microglia stage 1 and 2 (DAM1 / 2), interferonresponsive microglial (IRM), proliferative microglia (PM), transitioning microglia from HMs to DAMs (TrM) and antigen-presenting microglia (AgM). (Figure 34B) Statistically significant differences in cluster frequencies are marked in orange, defined by p-value adjusted by FDR < 0.05; gray points are not significant. (Figures 34C-34D) Apoe and Trem2 expression changes along the microglial trajectory in A. (Figure 34E) Cst7 and Lpl changes in Trem2 dependent DAM trajectory. (Figure 34F) Tyrobp and Cstb changes in Trem2 independent DAM trajectory. (Figures 34G-34J) Volcano plots for DEGs in clusters HM2, DAM 1, DAM2 and AgM.

[0288] DETAILED DESCRIPTION

[0289] The following description of the disclosure is provided as an enabling teaching of the disclosure in its best, currently known embodiments. Many modifications and other embodiments disclosed herein will come to mind to one skilled in the art to which the disclosed compositions and methods pertain having the benefit of the teachings presented in the foregoing descriptions and the associated drawings. Therefore, it is to be understood that the disclosures are not to be limited to the specific embodiments disclosed and that modifications and other embodiments are intended to be included within the scope of the appended claims. The skilled artisan will recognize many variants and adaptations of the aspects described herein. These variants and adaptations are intended to be included in the teachings of this disclosure and to be encompassed by the claims herein.

[0290] Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.

[0291] As can be apparent to those of skill in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has discrete components and features which may be readily separated from or combined with the features of any of the other several embodiments without departing from the scope or spirit of the present disclosure.

[0292] Any recited method can be carried out in the order of events recited or in any other order that is logically possible. That is, unless otherwise expressly stated, it is in no way intended that any method or aspect set forth herein be construed as requiring that its steps be performed in a specific order. Accordingly, where a method claim does not specifically state in the claims or descriptions that the steps are to be limited to a specific order, it is no way intended that an order be inferred, in any respect. This holds for any possible non-express basis for interpretation, including matters of logic with respect to arrangement of steps or operational flow, plain meaning derived from grammatical organization or punctuation, or the number or type of aspects described in the specification.

[0293] All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and / or materials in connection with which the publications are cited. The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided herein can be different from the actual publication dates, which can require independent confirmation.

[0294] It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and is not intended to be limiting. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the disclosed compositions and methods belong. It can be further understood that terms, such as those defined in commonly used dictionaries, should be interpreted as having a meaning that is consistent with their meaning in the context of the specification and relevant art and should not be interpreted in an idealized or overly formal sense unless expressly defined herein.

[0295] Prior to describing the various aspects of the present disclosure, the following definitions are provided and should be used unless otherwise indicated. Additional terms may be defined elsewhere in the present disclosure.

[0296] Definitions

[0297] As used herein, “comprising” is to be interpreted as specifying the presence of the stated features, integers, steps, or components as referred to, but does not preclude the presence or addition of one or more features, integers, steps, or components, or groups thereof. Moreover, each of the terms “by”, “comprising,” “comprises”, “comprised of,” “including,” “includes,” “included,” “involving,” “involves,” “involved,” and “such as” are used in their open, non-limiting sense and may be used interchangeably. Further, the term “comprising” is intended to include examples and aspects encompassed by the terms “consisting essentially of’ and “consisting of.” Similarly, the term “consisting essentially of’ is intended to include examples encompassed by the term “consisting of.” As used in the specification and the appended claims, the singular forms “a,” “an” and “the” include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to “a compound”, “a composition”, or “a disorder”, includes, but is not limited to, two or more such compounds, compositions, or disorders, and the like.

[0298] It should be noted that ratios, concentrations, amounts, and other numerical data can be expressed herein in a range format. It can be further understood that the endpoints of each of the ranges are significant both in relation to the other endpoint, and independently of the other endpoint. It is also understood that there are a number of values disclosed herein, and that each value is also herein disclosed as “about” that particular value in addition to the value itself. For example, if the value “10” is disclosed, then “about 10” is also disclosed. Ranges can be expressed herein as from “about” one particular value, and / or to “about” another particular value. Similarly, when values are expressed as approximations, by use of the antecedent “about,” it can be understood that the particular value forms a further aspect. For example, if the value “about 10” is disclosed, then “10” is also disclosed.

[0299] When a range is expressed, a further aspect includes from the one particular value and / or to the other particular value. For example, where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure, e.g., the phrase “x to y” includes the range from ‘x’ to ‘y’ as well as the range greater than ‘x’ and less than ‘y’. The range can also be expressed as an upper limit, e.g., ‘about x, y, z, or less’ and should be interpreted to include the specific ranges of ‘about x’, ‘about y’, and ‘about z’ as well as the ranges of Tess than x’, less than y’, and Tess than z’. Likewise, the phrase ‘about x, y, z, or greater’ should be interpreted to include the specific ranges of ‘about x’, ‘about y’, and ‘about z’ as well as the ranges of ‘greater than x’ , greater than y ’ , and ‘greater than z’ . In addition, the phrase “about ‘x’ to ‘y’”, where ‘x’ and ‘y’ are numerical values, includes “about ‘x’ to about ‘y’”.

[0300] It is to be understood that such a range format is used for convenience and brevity, and thus, should be interpreted in a flexible manner to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or subranges encompassed within that range as if each numerical value and sub-range is explicitly recited. To illustrate, a numerical range of “about 0.1% to 5%” should be interpreted to include not only the explicitly recited values of about 0.1% to about 5%, but also include individual values (e.g., about 1%, about 2%, about 3%, and about 4%) and the sub-ranges (e.g., about 0.5% to about 1.1%; about 5% to about 2.4%; about 0.5% to about 3.2%, and about 0.5% to about 4.4%, and other possible sub-ranges) within the indicated range. As used herein, the terms “about,” “approximate,” “at or about,” and “substantially” mean that the amount or value in question can be the exact value or a value that provides equivalent results or effects as recited in the claims or taught herein. That is, it is understood that amounts, sizes, formulations, parameters, and other quantities and characteristics are not and need not be exact but may be approximate and / or larger or smaller, as desired, reflecting tolerances, conversion factors, rounding off, measurement error and the like, and other factors known to those of skill in the art such that equivalent results or effects are obtained. In some circumstances, the value that provides equivalent results or effects cannot be reasonably determined. In such cases, it is generally understood, as used herein, that “about” and “at or about” mean the nominal value indicated ±10% variation unless otherwise indicated or inferred. In general, an amount, size, formulation, parameter or other quantity or characteristic is “about,” “approximate,” or “at or about” whether or not expressly stated to be such. It is understood that where “about,” “approximate,” or “at or about” is used before a quantitative value, the parameter also includes the specific quantitative value itself, unless specifically stated otherwise. As used herein, the term “substantially free,” when used in the context of a composition or component of a composition that is substantially absent, is intended to refer to an amount that is then about 1 % by weight or less, e.g., less than about 0.5 % by weight, less than about 0.1 % by weight, less than about 0.05 % by weight, or less than about 0.01 % by weight of the stated material, based on the total weight of the composition.

[0301] “Reduce” or other forms of the word, such as “reducing” or “reduction,” means lowering of an event or characteristic e.g., tumor growth). It is understood that this is typically in relation to some standard or expected value, in other words it is relative, but that it is not always necessary for the standard or relative value to be referred to. For example, “reduces tumor growth” means reducing the rate of growth of a tumor relative to a standard or a control (e.g., an untreated tumor).

[0302] The term “pharmaceutically acceptable” refers to those compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit / risk ratio.

[0303] The term “amino acid sequence” refers to a list of abbreviations, letters, characters, or words representing amino acid residues. The amino acid abbreviations used herein are conventional one letter codes for the amino acids and are expressed as follows: A, alanine; B, asparagine or aspartic acid; C, cysteine; D aspartic acid; E, glutamate, glutamic acid; F, phenylalanine; G, glycine; H histidine; I isoleucine; K, lysine; L, leucine; M, methionine; N, asparagine; P, proline; Q, glutamine; R, arginine; S, serine; T, threonine; V, valine; W, tryptophan; Y, tyrosine; Z, glutamine or glutamic acid.

[0304] The term “antibody” refers to natural or synthetic antibodies that selectively bind a target antigen. The term includes polyclonal and monoclonal antibodies. In addition to intact immunoglobulin molecules, also included in the term “antibodies” are fragments or polymers of those immunoglobulin molecules, and human or humanized versions of immunoglobulin molecules that selectively bind the target antigen.

[0305] As used herein, the term “bispecific antibody” refers to an antibody capable of selectively binding two or more epitopes. Bispecific antibodies generally comprise two different heavy chains with each heavy chain specifically binding a different epitope-either on two different molecules (e.g., antigens) or on the same molecule (e.g., on the same antigen). If a bispecific antibody is capable of selectively binding two different epitopes (a first epitope and a second epitope), the affinity of the first heavy chain for the first epitope will generally be at least one to two or three or four orders of magnitude lower than the affinity of the first heavy chain for the second epitope, and vice versa. The epitopes recognized by the bispecific antibody can be on the same or a different target (e.g., on the same or a different protein). Bispecific antibodies can be made, for example, by combining heavy chains that recognize different epitopes of the same antigen. For example, nucleic acid sequences encoding heavy chain variable sequences that recognize different epitopes of the same antigen can be fused to nucleic acid sequences encoding different heavy chain constant regions and such sequences can be expressed in a cell that expresses an immunoglobulin light chain.

[0306] The term “aptamer” refers to oligonucleic acid or peptide molecules that bind to a specific target molecule. These molecules are generally selected from a random sequence pool. The selected aptamers are capable of adapting unique tertiary structures and recognizing target molecules with high affinity and specificity. A “nucleic acid aptamer” is a DNA or RNA oligonucleic acid that binds to a target molecule via its conformation, and thereby inhibits or suppresses functions of such molecule. A nucleic acid aptamer may be constituted by DNA, RNA, or a combination thereof. A “peptide aptamer” is a combinatorial protein molecule with a variable peptide sequence inserted within a constant scaffold protein. Identification of peptide aptamers is typically performed under stringent yeast dihybrid conditions, which enhances the probability for the selected peptide aptamers to be stably expressed and correctly folded in an intracellular context. The term “carrier” means a compound, composition, substance, or structure that, when in combination with a compound or composition, aids or facilitates preparation, storage, administration, deliver}', effectiveness, selectivity, or any other feature of the compound or composition for its intended use or purpose. For example, a carrier can be selected to minimize any degradation of the active ingredient and to minimize any adverse side effects in the subject.

[0307] The term “chimeric molecule” refers to a single molecule created by joining two or more molecules that exist separately in their native state. The single, chimeric molecule has the desired functionality of all of its constituent molecules. One type of chimeric molecule is a fusion protein.

[0308] The term “fusion protein” refers to a polypeptide formed by the joining of two or more polypeptides through a peptide bond formed between the amino terminus of one polypeptide and the carboxyl terminus of another polypeptide. The fusion protein can be formed by the chemical coupling of the constituent polypeptides, or it can be expressed as a single polypeptide from nucleic acid sequence encoding the single contiguous fusion protein. A single chain fusion protein is a fusion protein having a single contiguous polypeptide backbone. Fusion proteins can be prepared using conventional techniques in molecular biology to join the two genes in frame into a single nucleic acid, and then expressing the nucleic acid in an appropriate host cell under conditions in which the fusion protein is produced.

[0309] The term “identity” refers to sequence identity between two nucleic acid molecules or polypeptides. Identity can be determined by comparing a position in each sequence which may be aligned for purposes of comparison. When a position in the compared sequence is occupied by the same base, then the molecules are identical at that position. A degree of similarity or identity between nucleic acid or amino acid sequences is a function of the number of identical or matching nucleotides at positions shared by the nucleic acid sequences. Various alignment algorithms and / or programs may be used to calculate the identity between two sequences, including FASTA, or BLAST which are available as a part of the GCG sequence analysis package (University of Wisconsin, Madison, Wis.), and can be used with, e.g., default setting. For example, polypeptides having at least 70%, 85%, 90%, 95%, 98% or 99% identity to specific polypeptides described herein and preferably exhibiting substantially the same functions, as well as polynucleotide encoding such polypeptides, are contemplated. Unless otherwise indicated a similarity score will be based on use of BLOSUM62. When BLASTP is used, the percent similarity is based on the BLASTP positives score and the percent sequence identity is based on the BLASTP identities score. BLASTP “Identities” shows the number and fraction of total residues in the high scoring sequence pairs which are identical; and BLASTP “Positives” shows the number and fraction of residues for which the alignment scores have positive values, and which are similar to each other. Amino acid sequences having these degrees of identity or similarity or any intermediate degree of identity of similarity to the amino acid sequences disclosed herein are contemplated and encompassed by this disclosure. The polynucleotide sequences of similar polypeptides are deduced using the genetic code and may be obtained by conventional means, in particular by reverse translating its amino acid sequence using the genetic code.

[0310] The term “nucleic acid” refers to a natural or synthetic molecule comprising a single nucleotide, or two or more nucleotides linked by a phosphate group at the 3’ position of one nucleotide to the 5’ end of another nucleotide. The nucleic acid is not limited by length, and thus the nucleic acid can include deoxyribonucleic acid (DNA) or ribonucleic acid (RNA).

[0311] The term “operably linked to” refers to the functional relationship of a nucleic acid with another nucleic acid sequence. Promoters, enhancers, transcriptional and translational stop sites, and other signal sequences are examples of nucleic acid sequences operably linked to other sequences. For example, operable linkage of DNA to a transcriptional control element refers to the physical and functional relationship between the DNA and promoter such that the transcription of such DNA is initiated from the promoter by an RNA polymerase that specifically recognizes, binds to, and transcribes the DNA.

[0312] The terms “peptide,” “protein,” and “polypeptide” are used interchangeably to refer to a natural or synthetic molecule comprising two or more amino acids linked by the carboxyl group of one amino acid to the alpha amino group of another.

[0313] The term “pharmaceutically acceptable” refers to those compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit / risk ratio.

[0314] The term “protein domain” refers to a portion of a protein, portions of a protein, or an entire protein showing structural integrity; this determination may be based on amino acid composition of a portion of a protein, portions of a protein, or the entire protein.

[0315] A “spacer” as used herein refers to a peptide that joins the proteins comprising a fusion protein. Generally, a spacer has no specific biological activity other than to join the proteins or to preserve some minimum distance or other spatial relationship between them. However, the constituent amino acids of a spacer may be selected to influence some property of the molecule such as the folding, net charge, or hydrophobicity of the molecule. The term “specifically binds”, as used herein, when referring to a polypeptide (including antibodies) or receptor, refers to a binding reaction which is determinative of the presence of the protein or polypeptide or receptor in a heterogeneous population of proteins and other biologies. Thus, under designated conditions (e.g., immunoassay conditions in the case of an antibody), a specified ligand or antibody “specifically binds” to its particular “target” (e.g., an antibody specifically binds to an antigen) when it does not bind in a significant amount to other proteins present in the sample or to other proteins to which the ligand or antibody may come in contact in an organism. Generally, a first molecule that “specifically binds” a second molecule has an affinity constant (Ka) greater than about 105M-1(e.g., 106M-1, 107M-1, 108M-1, 109M-1, IO10M-1, 1011M-1, and 1012M-1or more) with that second molecule.

[0316] The term “subject” refers to any individual who is the target of administration or treatment. The subject can be a vertebrate, for example, a mammal. Thus, the subject can be a human or veterinary patient. The term “patient” refers to a subject under the treatment of a clinician, e.g., physician.

[0317] The term “therapeutically effective” refers to the amount of the composition used is of sufficient quantity to ameliorate one or more causes or symptoms of a disease or disorder. Such amelioration only requires a reduction or alteration, not necessarily elimination.

[0318] The terms “transformation” and “transfection” mean the introduction of a nucleic acid, e.g., an expression vector, into a recipient cell including introduction of a nucleic acid to the chromosomal DNA of said cell.

[0319] The term “treatment” refers to the medical management of a patient with the intent to cure, ameliorate, stabilize, or prevent a disease, pathological condition, or disorder. This term includes active treatment, that is, treatment directed specifically toward the improvement of a disease, pathological condition, or disorder, and also includes causal treatment, that is, treatment directed toward removal of the cause of the associated disease, pathological condition, or disorder. In addition, this term includes palliative treatment, that is, treatment designed for the relief of symptoms rather than the curing of the disease, pathological condition, or disorder: preventative treatment, that is, treatment directed to minimizing or partially or completely inhibiting the development of the associated disease, pathological condition, or disorder; and supportive treatment, that is, treatment employed to supplement another specific therapy directed toward the improvement of the associated disease, pathological condition, or disorder.

[0320] The term “variant” refers to an amino acid or peptide sequence having conservative amino acid substitutions, non-conservative amino acid substitutions (e.g., a degenerate variant), substitutions within the wobble position of each codon (e.g. DNA and RNA) encoding an amino acid, amino acids added to the C-terminus of a peptide, or a peptide having 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99% sequence identity to a reference sequence.

[0321] The term “vector” refers to a nucleic acid sequence capable of transporting into a cell another nucleic acid to which the vector sequence has been linked. The term “expression vector” includes any vector, (e.g., a plasmid, virus, cosmid or phage chromosome) containing a gene construct in a form suitable for expression by a cell (e.g., linked to a transcriptional control element).

[0322] Antibodies

[0323] Further aspects also include a recombinant antibody, wherein the antibody comprises: a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131; and / or a light chain variable region (VL) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

[0324] In some aspects, the recombinant antibody comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25.

[0325] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

[0326] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 14.

[0327] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 8.

[0328] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 10.

[0329] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 2.

[0330] In some aspects, the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 6.

[0331] In some aspects, the recombinant antibody comprises at least 1 (e.g., at least 2, at least 3, at least 4, at least 5, at least, 6, at least 7, at least 8, at least 9, or at least 10) amino acid substitutions.

[0332] In some aspects, the recombinant antibody further includes a transferrin moiety conjugated by a peptide linker.

[0333] In some aspects, the transferrin moiety comprises at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 180.

[0334] Suitable peptide linkers are described in Kumada Y, et al. Biochemical Engineering Journal. 2007 35(2) : 158- 165 ; Albrecht H, et al. J Immunol Methods. 2006 310(1 -2) : 100- 16; Feng J, et al. J Immunol Methods. 2003 282(1 -2):33-43 ; Griffiths AD, et al. Curr Opin Biotechnol. 1998 9(1): 102-8; Huston JS, et al. Methods Enzymol. 1991 203:46-88; Bird RE, et al. Science. 1988 242(4877):423-6; Takkinen K, et al. Protein Eng. 1991 4(7):837-41; Smallshaw JE, et al. Protein Eng. 1999 12(7):623-30; Argos P. J Mol Biol. 1990 211 (4):943-58; and Whitlow M, et al. Protein Eng. 1993 6(8):989-95, each of which are hereby incorporated by reference in their entireties.

[0335] The particular length of the peptide linker used to join the antibodies to the transferrin moiety can be modified to affect half-life, immunogenicity, and activity of the overall construct. In some embodiments, the linker sequence is 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or more amino acids in length. In some cases, the linker comprises 2, 3, 4, 5, or more GGGGS sequences. In some embodiments, the linker sequence comprises GGGGSGGGGSGGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 180). The linker is preferably long enough to not interfere with proper folding and association of the VH-VL chains but not so long as to cause added immunogenicity.

[0336] In some aspects, the transferrin moiety comprises at least 60% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 65% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 70% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 75% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 80% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 85% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 90% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 95% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety is represented by SEQ ID NO: 180.

[0337] In various aspects, disclosed herein are bispecific antibodies. A typical bispecific antibody has two heavy chains each having three heavy chain CDRs, followed by a CHI domain, a hinge, a CH2 domain, and a CH3 domain, and an immunoglobulin light chain that either does not confer antigen-binding specificity but that can associate with each heavy chain, or that can associate with each heavy chain and that can bind one or more of the epitopes bound by the heavy chain antigenbinding regions, or that can associate with each heavy chain and enable binding of one or both of the heavy chains to one or both epitopes. BsAbs can be divided into two major classes, those bearing an Fc region (IgG-like) and those lacking an Fc region, the latter normally being smaller than the IgG and IgG-like bispecific molecules comprising an Fc. The IgG-like bsAbs can have different formats such as, but not limited to, triomab, knobs into holes IgG (kih IgG), crossMab, orth-Fab IgG, Dual-variable domains Ig (DVD-Ig), two-in-one or dual action Fab (DAF), IgG- single-chain Fv (IgG-scFv), or K / .-bodies. The non-IgG-like different formats include tandem scFvs, diabody format, single-chain diabody, tandem diabodies (TandAbs), Dual- affinity retargeting molecule (DART), DART-Fc, nanobodies, or antibodies produced by the dock-and- lock (DNL) method (Gaowei Fan, Zujian Wang & Mingju Hao, Bispecific antibodies and their applications, 8 JOURNAL OF HEMATOLOGY & ONCOLOGY 130; Dafne Muller & Roland E. Kontermann, Bispecific Antibodies, HANDBOOK OF THERAPEUTIC ANTIBODIES 265-310 (2014), the entire teachings of which are herein incorporated). The methods of producing bsAbs are not limited to quadroma technology based on the somatic fusion of two different hybridoma cell lines, chemical conjugation, which involves chemical cross-linkers, and genetic approaches utilizing recombinant DNA technology.

[0338] For example, a bispecific antibody can include: (i) a first binding arm having a first antigenbinding region that specifically binds to LILRB3, and (ii) a second binding arm having a second antigen-binding region that specifically binds to LILRB2. In some aspects, the first antigenbinding region is an agonist of LILRB3 and the second antigen binding region is an antagonist of LILRB2. In some aspects, the first antigen-binding region comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, or 93, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, or 94, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, or 95.

[0339] In some aspects, the second antigen-binding region comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 101, 107, 113, 119, 125, or 131.

[0340] In some aspects, the first antigen-binding region comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 1, 3, 5, 7, 9,

[0341] 11, or 13.

[0342] In some aspects, the second antigen-binding region comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 15, 17, 19, 21, 23, or 25.

[0343] In some aspects, the first antigen-binding region comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 2, 4, 6, 8, 10,

[0344] 12, or 14.

[0345] In some aspects, the second antigen-binding region comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 16, 18, 20, 22, 24, or 26. In some aspects, the bispecific antibody further includes a transferrin moiety conjugated by a peptide linker.

[0346] In some aspects, the transferrin moiety comprises at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 180.

[0347] Suitable peptide linkers are described in Kumada Y, et al. Biochemical Engineering Journal. 2007 35(2) : 158- 165 ; Albrecht H, et al. J Immunol Methods. 2006 310(1 -2) : 100- 16; Feng J, et al. J Immunol Methods. 2003 282(1 -2):33-43 ; Griffiths AD, et al. Curr Opin Biotechnol. 1998 9(1): 102-8; Huston JS, et al. Methods Enzymol. 1991 203:46-88; Bird RE, et al. Science. 1988 242(4877):423-6; Takkinen K, et al. Protein Eng. 1991 4(7):837-41; Smallshaw JE, et al. Protein Eng. 1999 12(7):623-30; Argos P. J Mol Biol. 1990 211 (4):943-58; and Whitlow M, et al. Protein Eng. 1993 6(8):989-95, each of which are hereby incorporated by reference in their entireties.

[0348] The particular length of the peptide linker used to join the antibodies to the transferrin moiety can be modified to affect half-life, immunogenicity, and activity of the overall construct. In some embodiments, the linker sequence is 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or more amino acids in length. In some cases, the linker comprises 2, 3, 4, 5, or more GGGGS sequences. In some embodiments, the linker sequence comprises GGGGSGGGGSGGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 180). The linker is preferably long enough to not interfere with proper folding and association of the VH-VL chains but not so long as to cause added immunogenicity.

[0349] In some aspects, the transferrin moiety comprises at least 60% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 65% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 70% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 75% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 80% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 85% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 90% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety comprises at least 95% identity to SEQ ID NO: 180. In some aspects, the transferrin moiety is represented by SEQ ID NO: 180.

[0350] In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 163; a first heavy chain comprising a sequence with at least 60% (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) identity to SEQ ID NO: 164; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 166; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 165.

[0351] In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 167; a first heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 168; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 170; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 169.

[0352] In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 181 ; a first heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 182; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 184; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 183. In some aspects, the bispecific antibody includes: a first light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 185; a first heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 186; a second light chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 188; and a second heavy chain comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 187.

[0353] In some aspects, the first antigen binding region of the bispecific antibody has any one of thes structures:

[0354] VH CDR1 is SEQ ID NO: 27; VH CDR2 is SEQ ID NO: 28; VH CDR3 is SEQ ID NO: 29; VL CDR1 is SEQ ID NO: 30; VL CDR2 is SEQ ID NO: 31; and VL CDR3 is SEQ ID NO: 32; or

[0355] VH CDR1 is SEQ ID NO: 33; VH CDR2 is SEQ ID NO: 34; VH CDR3 is SEQ ID NO: 35; VL CDR1 is SEQ ID NO: 36; VL CDR2 is SEQ ID NO: 37; and VL CDR3 is SEQ ID NO: 38; or

[0356] VH CDR1 is SEQ ID NO: 39; VH CDR2 is SEQ ID NO: 40; VH CDR3 is SEQ ID NO: 41 ; VL CDR1 is SEQ ID NO: 42; VL CDR2 is SEQ ID NO: 43; and VL CDR3 is SEQ ID NO: 44; or

[0357] VH CDR1 is SEQ ID NO: 45;

[0358] VH CDR2 is SEQ ID NO: 46;

[0359] VH CDR3 is SEQ ID NO: 47;

[0360] VL CDR1 is SEQ ID NO: 48;

[0361] VL CDR2 is SEQ ID NO: 49; and

[0362] VL CDR3 is SEQ ID NO: 50; or

[0363] VH CDR1 is SEQ ID NO: 57;

[0364] VH CDR2 is SEQ ID NO: 58;

[0365] VH CDR3 is SEQ ID NO: 59;

[0366] VL CDR1 is SEQ ID NO: 60;

[0367] VL CDR2 is SEQ ID NO: 61; and

[0368] VL CDR3 is SEQ ID NO: 62; or

[0369] VH CDR1 is SEQ ID NO: 63;

[0370] VH CDR2 is SEQ ID NO: 64;

[0371] VH CDR3 is SEQ ID NO: 65;

[0372] VL CDR1 is SEQ ID NO: 66;

[0373] VL CDR2 is SEQ ID NO: 67; and

[0374] VL CDR3 is SEQ ID NO: 68; or

[0375] VH CDR1 is SEQ ID NO: 69;

[0376] VH CDR2 is SEQ ID NO: 70;

[0377] VH CDR3 is SEQ ID NO: 71;

[0378] VL CDR1 is SEQ ID NO: 72;

[0379] VL CDR2 is SEQ ID NO: 73; and

[0380] VL CDR3 is SEQ ID NO: 74; or

[0381] VH CDR1 is SEQ ID NO: 75;

[0382] VH CDR2 is SEQ ID NO: 76; VH CDR3 is SEQ ID NO: 77;

[0383] VL CDR1 is SEQ ID NO: 78;

[0384] VL CDR2 is SEQ ID NO: 79; and

[0385] VL CDR3 is SEQ ID NO: 80; or

[0386] VH CDR1 is SEQ ID NO: 81 ;

[0387] VH CDR2 is SEQ ID NO: 82;

[0388] VH CDR3 is SEQ ID NO: 83;

[0389] VL CDR1 is SEQ ID NO: 84;

[0390] VL CDR2 is SEQ ID NO: 85; and

[0391] VL CDR3 is SEQ ID NO: 86; or

[0392] VH CDR1 is SEQ ID NO: 87;

[0393] VH CDR2 is SEQ ID NO: 88;

[0394] VH CDR3 is SEQ ID NO: 89;

[0395] VL CDR1 is SEQ ID NO: 90;

[0396] VL CDR2 is SEQ ID NO: 91; and

[0397] VL CDR3 is SEQ ID NO: 92; or

[0398] VH CDR1 is SEQ ID NO: 93;

[0399] VH CDR2 is SEQ ID NO: 94;

[0400] VH CDR3 is SEQ ID NO: 95;

[0401] VL CDR1 is SEQ ID NO: 96;

[0402] VL CDR2 is SEQ ID NO: 97; and

[0403] VL CDR3 is SEQ ID NO: 98.

[0404] In some aspects, the second antigen binding region of the bispecific antibody has any one of thes structures:

[0405] VH CDR1 is SEQ ID NO: 99;

[0406] VH CDR2 is SEQ ID NO: 100;

[0407] VH CDR3 is SEQ ID NO: 101;

[0408] VL CDR1 is SEQ ID NO: 102;

[0409] VL CDR2 is SEQ ID NO: 103; and

[0410] VL CDR3 is SEQ ID NO: 104; or VHCDR1 is SEQIDNO: 105;

[0411] VH CDR2 is SEQ ID NO: 106;

[0412] VH CDR3 is SEQ ID NO: 107;

[0413] VLCDR1 is SEQIDNO: 108;

[0414] VL CDR2 is SEQ ID NO: 109; and

[0415] VL CDR3 is SEQ ID NO: 110; or

[0416] VHCDR1 is SEQIDNO: 111;

[0417] VH CDR2 is SEQ ID NO: 112;

[0418] VHCDR3 is SEQIDNO: 113;

[0419] VL CDR1 is SEQ ID NO: 114;

[0420] VL CDR2 is SEQ ID NO: 115; and

[0421] VL CDR3 is SEQ ID NO: 116; or

[0422] VHCDR1 is SEQIDNO: 117;

[0423] VHCDR2is SEQIDNO: 118;

[0424] VHCDR3 is SEQIDNO: 119;

[0425] VLCDR1 is SEQIDNO: 120;

[0426] VL CDR2 is SEQ ID NO: 121; and

[0427] VL CDR3 is SEQ ID NO: 122; or

[0428] VHCDR1 is SEQIDNO: 123;

[0429] VH CDR2 is SEQ ID NO: 124;

[0430] VH CDR3 is SEQ ID NO: 125;

[0431] VLCDR1 is SEQIDNO: 126;

[0432] VL CDR2 is SEQ ID NO: 127; and

[0433] VL CDR3 is SEQ ID NO: 128; or

[0434] VH CDR1 is SEQ ID NO: 129;

[0435] VH CDR2 is SEQ ID NO: 130;

[0436] VHCDR3 is SEQIDNO: 131;

[0437] VLCDR1 is SEQIDNO: 132; VL CDR2 is SEQ ID NO: 133; and

[0438] VL CDR3 is SEQ ID NO: 134.

[0439] In some embodiments, the bispecific antibody may be subjected to an alteration to render it less immunogenic when administered to a human. Such an alteration may comprise one or more of the techniques commonly known as chimerization, humanization, CDR-grafting, deimmunization and / or mutation of framework region amino acids to correspond to the closest human germline sequence (germlining). Bispecific antibodies which have been altered will therefore remain administrable for a longer period of time with reduced or no immune response- related side effects than corresponding bispecific antibodies which have not undergone any such alteration(s). One of ordinary skill in the art will understand how to determine whether, and to what degree an antibody must be altered in order to prevent it from eliciting an unwanted host immune response.

[0440] Also described herein are nucleic acids encoding the recombinant antibodies described herein. Further provided are recombinant expression cassettes or plasmids comprising a sequence to express the recombinant antibodies described herein. Various aspects further include host cells comprising the expression cassette or the plasmid. Also provided are methods of producing an antibody, comprising cultivating or maintaining the host cell under conditions to produce the antibody.

[0441] In various aspects, described herein are methods of treating and / or preventing a neurological disorder in a subject, comprising administering to the subject a therapeutically effective amount of any of the recombinant antibodies described herein. In some aspects, the neurological disorder is Alzheimer’s disease.

[0442] In some aspects, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0443] VH CDR1 is SEQ ID NO: 27;

[0444] VH CDR2 is SEQ ID NO: 28;

[0445] VH CDR3 is SEQ ID NO: 29;

[0446] VL CDR1 is SEQ ID NO: 30;

[0447] VL CDR2 is SEQ ID NO: 31; and

[0448] VL CDR3 is SEQ ID NO: 32; or VHCDR1 is SEQIDNO: 33;

[0449] VH CDR2 is SEQ ID NO: 34;

[0450] VHCDR3 is SEQIDNO: 35;

[0451] VLCDR1 is SEQ ID NO: 36;

[0452] VL CDR2 is SEQ ID NO: 37; and

[0453] VL CDR3 is SEQ ID NO: 38; or

[0454] VH CDR1 is SEQ ID NO: 39;

[0455] VH CDR2 is SEQ ID NO: 40;

[0456] VH CDR3 is SEQ ID NO: 41;

[0457] VLCDR1 is SEQ ID NO: 42;

[0458] VL CDR2 is SEQ ID NO: 43; and

[0459] VL CDR3 is SEQ ID NO: 44; or

[0460] VHCDR1 is SEQ ID NO: 45;

[0461] VH CDR2 is SEQ ID NO: 46;

[0462] VH CDR3 is SEQ ID NO: 47;

[0463] VLCDR1 is SEQ ID NO: 48;

[0464] VL CDR2 is SEQ ID NO: 49; and

[0465] VL CDR3 is SEQ ID NO: 50; or

[0466] VHCDR1 is SEQIDNO: 57;

[0467] VHCDR2 is SEQIDNO: 58;

[0468] VH CDR3 is SEQ ID NO: 59;

[0469] VLCDR1 is SEQ ID NO: 60;

[0470] VL CDR2 is SEQ ID NO: 61; and

[0471] VL CDR3 is SEQ ID NO: 62; or

[0472] VHCDR1 is SEQIDNO: 63;

[0473] VH CDR2 is SEQ ID NO: 64;

[0474] VHCDR3 is SEQIDNO: 65;

[0475] VLCDR1 is SEQ ID NO: 66;

[0476] VL CDR2 is SEQ ID NO: 67; and VL CDR3 is SEQ ID NO: 68; or

[0477] VH CDR1 is SEQ ID NO: 69;

[0478] VH CDR2 is SEQ ID NO: 70;

[0479] VH CDR3 is SEQ ID NO: 71;

[0480] VL CDR1 is SEQ ID NO: 72;

[0481] VL CDR2 is SEQ ID NO: 73; and

[0482] VL CDR3 is SEQ ID NO: 74; or

[0483] VH CDR1 is SEQ ID NO: 75;

[0484] VH CDR2 is SEQ ID NO: 76;

[0485] VH CDR3 is SEQ ID NO: 77;

[0486] VL CDR1 is SEQ ID NO: 78;

[0487] VL CDR2 is SEQ ID NO: 79; and

[0488] VL CDR3 is SEQ ID NO: 80; or

[0489] VH CDR1 is SEQ ID NO: 81;

[0490] VH CDR2 is SEQ ID NO: 82;

[0491] VH CDR3 is SEQ ID NO: 83;

[0492] VL CDR1 is SEQ ID NO: 84;

[0493] VL CDR2 is SEQ ID NO: 85; and

[0494] VL CDR3 is SEQ ID NO: 86; or

[0495] VH CDR1 is SEQ ID NO: 87;

[0496] VH CDR2 is SEQ ID NO: 88;

[0497] VH CDR3 is SEQ ID NO: 89;

[0498] VL CDR1 is SEQ ID NO: 90;

[0499] VL CDR2 is SEQ ID NO: 91; and

[0500] VL CDR3 is SEQ ID NO: 92; or

[0501] VH CDR1 is SEQ ID NO: 93;

[0502] VH CDR2 is SEQ ID NO: 94;

[0503] VH CDR3 is SEQ ID NO: 95; VL CDR1 is SEQ ID NO: 96;

[0504] VL CDR2 is SEQ ID NO: 97; and

[0505] VL CDR3 is SEQ ID NO: 98; or

[0506] VH CDR1 is SEQ ID NO: 99;

[0507] VH CDR2 is SEQ ID NO: 100;

[0508] VH CDR3 is SEQ ID NO: 101;

[0509] VL CDR1 is SEQ ID NO: 102;

[0510] VL CDR2 is SEQ ID NO: 103; and

[0511] VL CDR3 is SEQ ID NO: 104; or

[0512] VH CDR1 is SEQ ID NO: 105;

[0513] VH CDR2 is SEQ ID NO: 106;

[0514] VH CDR3 is SEQ ID NO: 107;

[0515] VL CDR1 is SEQ ID NO: 108;

[0516] VL CDR2 is SEQ ID NO: 109; and

[0517] VL CDR3 is SEQ ID NO: 110; or

[0518] VH CDR1 is SEQ ID NO: 111;

[0519] VH CDR2 is SEQ ID NO: 112;

[0520] VH CDR3 is SEQ ID NO: 113;

[0521] VL CDR1 is SEQ ID NO: 114;

[0522] VL CDR2 is SEQ ID NO: 115; and

[0523] VL CDR3 is SEQ ID NO: 116; or

[0524] VH CDR1 is SEQ ID NO: 117;

[0525] VH CDR2 is SEQ ID NO: 118;

[0526] VH CDR3 is SEQ ID NO: 119;

[0527] VL CDR1 is SEQ ID NO: 120;

[0528] VL CDR2 is SEQ ID NO: 121; and

[0529] VL CDR3 is SEQ ID NO: 122; or

[0530] VH CDR1 is SEQ ID NO: 123;

[0531] VH CDR2 is SEQ ID NO: 124; VH CDR3 is SEQ ID NO: 125;

[0532] VL CDR1 is SEQ ID NO: 126;

[0533] VL CDR2 is SEQ ID NO: 127; and

[0534] VL CDR3 is SEQ ID NO: 128; or

[0535] VH CDR1 is SEQ ID NO: 129;

[0536] VH CDR2 is SEQ ID NO: 130;

[0537] VH CDR3 is SEQ ID NO: 131;

[0538] VL CDR1 is SEQ ID NO: 132;

[0539] VL CDR2 is SEQ ID NO: 133; and VL CDR3 is SEQ ID NO: 134.

[0540] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0541] VH CDR1 is SEQ ID NO: 27;

[0542] VH CDR2 is SEQ ID NO: 28;

[0543] VH CDR3 is SEQ ID NO: 29;

[0544] VL CDR1 is SEQ ID NO: 30;

[0545] VL CDR2 is SEQ ID NO: 31; and

[0546] VL CDR3 is SEQ ID NO: 32.

[0547] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0548] VH CDR1 is SEQ ID NO: 33;

[0549] VH CDR2 is SEQ ID NO: 34;

[0550] VH CDR3 is SEQ ID NO: 35;

[0551] VL CDR1 is SEQ ID NO: 36;

[0552] VL CDR2 is SEQ ID NO: 37; and

[0553] VL CDR3 is SEQ ID NO: 38. In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0554] VH CDR1 is SEQ ID NO: 39;

[0555] VH CDR2 is SEQ ID NO: 40;

[0556] VH CDR3 is SEQ ID NO: 41 ;

[0557] VL CDR1 is SEQ ID NO: 42;

[0558] VL CDR2 is SEQ ID NO: 43; and VL CDR3 is SEQ ID NO: 44.

[0559] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0560] VH CDR1 is SEQ ID NO: 45;

[0561] VH CDR2 is SEQ ID NO: 46;

[0562] VH CDR3 is SEQ ID NO: 47;

[0563] VL CDR1 is SEQ ID NO: 48;

[0564] VL CDR2 is SEQ ID NO: 49; and VL CDR3 is SEQ ID NO: 50.

[0565] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0566] VH CDR1 is SEQ ID NO: 57;

[0567] VH CDR2 is SEQ ID NO: 58;

[0568] VH CDR3 is SEQ ID NO: 59;

[0569] VL CDR1 is SEQ ID NO: 60;

[0570] VL CDR2 is SEQ ID NO: 61; and VL CDR3 is SEQ ID NO: 62.

[0571] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0572] VH CDR1 is SEQ ID NO: 63;

[0573] VH CDR2 is SEQ ID NO: 64;

[0574] VH CDR3 is SEQ ID NO: 65;

[0575] VL CDR1 is SEQ ID NO: 66;

[0576] VL CDR2 is SEQ ID NO: 67; and

[0577] VL CDR3 is SEQ ID NO: 68.

[0578] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0579] VH CDR1 is SEQ ID NO: 69;

[0580] VH CDR2 is SEQ ID NO: 70;

[0581] VH CDR3 is SEQ ID NO: 71 ;

[0582] VL CDR1 is SEQ ID NO: 72;

[0583] VL CDR2 is SEQ ID NO: 73; and

[0584] VL CDR3 is SEQ ID NO: 74.

[0585] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0586] VH CDR1 is SEQ ID NO: 75;

[0587] VH CDR2 is SEQ ID NO: 76;

[0588] VH CDR3 is SEQ ID NO: 77;

[0589] VL CDR1 is SEQ ID NO: 78;

[0590] VL CDR2 is SEQ ID NO: 79; and

[0591] VL CDR3 is SEQ ID NO: 80.

[0592] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0593] VH CDR1 is SEQ ID NO: 81 ; VH CDR2 is SEQ ID NO: 82;

[0594] VH CDR3 is SEQ ID NO: 83;

[0595] VL CDR1 is SEQ ID NO: 84;

[0596] VL CDR2 is SEQ ID NO: 85; and

[0597] VL CDR3 is SEQ ID NO: 86.

[0598] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0599] VH CDR1 is SEQ ID NO: 87;

[0600] VH CDR2 is SEQ ID NO: 88;

[0601] VH CDR3 is SEQ ID NO: 89;

[0602] VL CDR1 is SEQ ID NO: 90;

[0603] VL CDR2 is SEQ ID NO: 91; and

[0604] VL CDR3 is SEQ ID NO: 92.

[0605] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0606] VH CDR1 is SEQ ID NO: 93;

[0607] VH CDR2 is SEQ ID NO: 94;

[0608] VH CDR3 is SEQ ID NO: 95;

[0609] VL CDR1 is SEQ ID NO: 96;

[0610] VL CDR2 is SEQ ID NO: 97; and

[0611] VL CDR3 is SEQ ID NO: 98.

[0612] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0613] VH CDR1 is SEQ ID NO: 99;

[0614] VH CDR2 is SEQ ID NO: 100;

[0615] VH CDR3 is SEQ ID NO: 101;

[0616] VL CDR1 is SEQ ID NO: 102; VL CDR2 is SEQ ID NO: 103; and

[0617] VL CDR3 is SEQ ID NO: 104.

[0618] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0619] VH CDR1 is SEQ ID NO: 105;

[0620] VH CDR2 is SEQ ID NO: 106;

[0621] VH CDR3 is SEQ ID NO: 107;

[0622] VL CDR1 is SEQ ID NO: 108;

[0623] VL CDR2 is SEQ ID NO: 109; and

[0624] VL CDR3 is SEQ ID NO: 110.

[0625] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0626] VH CDR1 is SEQ ID NO: 111;

[0627] VH CDR2 is SEQ ID NO: 112;

[0628] VH CDR3 is SEQ ID NO: 113;

[0629] VL CDR1 is SEQ ID NO: 114;

[0630] VL CDR2 is SEQ ID NO: 115; and

[0631] VL CDR3 is SEQ ID NO: 116.

[0632] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0633] VH CDR1 is SEQ ID NO: 117;

[0634] VH CDR2 is SEQ ID NO: 118;

[0635] VH CDR3 is SEQ ID NO: 119;

[0636] VL CDR1 is SEQ ID NO: 120;

[0637] VL CDR2 is SEQ ID NO: 121; and

[0638] VL CDR3 is SEQ ID NO: 122. In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0639] VH CDR1 is SEQ ID NO: 123;

[0640] VH CDR2 is SEQ ID NO: 124; VH CDR3 is SEQ ID NO: 125; VL CDR1 is SEQ ID NO: 126; VL CDR2 is SEQ ID NO: 127; and VL CDR3 is SEQ ID NO: 128.

[0641] In some embodiments, the recombinant antibody comprises a light chain variable region (VL) that comprises a light chain complementarity determining region (CDRL)l, CDRL2, and CDRL3 and a heavy chain variable region (VH) that comprises a heavy chain complementarity determining region (CDRH)l, CDRH2, and CDRH3, wherein:

[0642] VH CDR1 is SEQ ID NO: 129;

[0643] VH CDR2 is SEQ ID NO: 130;

[0644] VH CDR3 is SEQ ID NO: 131; VL CDR1 is SEQ ID NO: 132;

[0645] VL CDR2 is SEQ ID NO: 133; and VL CDR3 is SEQ ID NO: 134.

[0646] In some aspects, the recombinant antibody comprises at least 1 (e.g., at least 2, at least 3, at least 4, at least 5, at least, 6, at least 7, at least 8, at least 9, or at least 10) amino acid substitutions. In some aspects, the recombinant antibody is genetically engineered to provide enhanced delivery across the blood brain barrier. For example, the recombinant antibodies disclosed herein can be used as a bi-specific antibody with an antibody or aptamer against an endogenous blood-brain barrier (BBB) receptor to facilitate distribution through the central nervous system (CNS). For example, the BBB receptor can include: transferrin receptor (TfR), insulin receptor, insulin-like growth factor receptor (IGF receptor), low density lipoprotein receptor-related protein 8 (LRP8), low density lipoprotein receptor-related protein 1 (LRP1), and heparin-binding epidermal growth factor-like growth factor (HB-EGF). Example methods of forming bi-specific antibodies can be found in Pardridge, W.M., Boado, R.I., Patrick, D.J., Ka-Wai Hui, E. and Lu, J.Z., 2018. Bloodbrain barrier transport, plasma pharmacokinetics, and neuropathology following chronic treatment of the rhesus monkey with a brain penetrating humanized monoclonal antibody against the human transferrin receptor. Molecular pharmaceutics, 15(11), pp.5207-5216 and in US 2017 / 0174778 Al, both of which are incorporated by reference in their entireties. Other methods of enhancing delivery of recombinant antibodies include, for example, mechanical disrupting the BBB using ultrasonic stimulation or encapsulating / conjugating antibodies in nanoparticles having increased BBB penetrating properties. For example, the recombinant antibody can be configured for enhanced delivery using any of the techniques in WO 2021248133 Al, which is herein incorporated by reference in its entirety.

[0647] Also described herein are nucleic acids encoding the recombinant antibodies described herein. Further provided are recombinant expression cassettes or plasmids comprising a sequence to express the recombinant antibodies described herein. Various aspects further include host cells comprising the expression cassette or the plasmid. Also provided are methods of producing an antibody, comprising cultivating or maintaining the host cell under conditions to produce the antibody.

[0648] Methods

[0649] In various aspects, disclosed herein is a method of modulating the production and / or accumulation of amyloid- (A ) in a subject, comprising: administering to the subject a therapeutically effective amount of an antibody or antibody binding fragment having a leukocyte immunoglobulin- like receptor (LILR) binding domain. As used herein, the term “binding domain” refers to a protein, e.g., an immunoglobulin chain or fragment thereof, comprising at least one immunoglobulin variable domain sequence.

[0650] In some aspects, the leukocyte immunoglobulin-like receptor binding domain is a leukocyte immunoglobulin-like receptor B2 (LILRB2) binding domain and / or leukocyte immunoglobulin-like receptor B3 (LILRB3) binding domain.

[0651] In some aspects, the antibody or antibody binding fragment is an agonist of LILRB2 and / or LILRB3. In some aspects, the antibody or antibody binding fragment is a humanized antibody. The term “humanized antibody”, also known as CDR-grafted antibody, refers to an antibody generated by non-human (such as murine) CDR sequences grafted onto human antibody variable region framework, i.e. antibody generated from different types of sequences of human germline antibody framework. Humanized antibody overcomes the strong anti-antibody response induced by chimeric antibody which carries a large amount of non-human (such as murine) components. Such framework sequences can be obtained from public DNA database or published references covering germline antibody gene sequences. For example, germline DNA sequences of human heavy and light chain variable region genes can be found in “VBase” human germline sequence database (available on web www.mrccpe.com.ac.uk / vbase), as well as found in Kabat, E A et al. 1991, Sequences of Proteins of Immunological Interest, the 5th Ed.

[0652] In some aspects, the antibody or antibody binding fragment specifically binds to LILRB2.

[0653] In some aspects, the antibody or antibody binding fragment specifically binds to LILRB3.

[0654] In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131.

[0655] In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25. In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

[0656] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

[0657] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 14.

[0658] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 8.

[0659] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 10.

[0660] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 2.

[0661] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 6.

[0662] In various aspects, disclosed herein is a method of treating a neurological disorder in a subject, comprising: administering to the subject a therapeutically effective amount of an antibody or antibody binding fragment having a leukocyte immunoglobulin-like receptor (LILR) binding region.

[0663] In some aspects, the wherein the neurological disorder comprises dementia, frontotemporal dementia (FTD), Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), Huntington's disease, taupathy disease, Nasu-Hakola disease, and / or multiple sclerosis.

[0664] In some aspects, the leukocyte immunoglobulin-like receptor binding domain is a leukocyte immunoglobulin-like receptor B2 (LILRB2) binding domain and / or leukocyte immunoglobulin-like receptor B3 (LILRB3) binding domain. In some aspects, the antibody or antibody binding fragment is an agonist of LILRB2 and / or LILRB3. In some aspects, the antibody or antibody binding fragment is an antagonist of LILRB2 and / or LILRB3. In some aspects, the antibody or antibody binding fragment is a humanized antibody.

[0665] In some aspects, the antibody or antibody binding fragment specifically binds to LILRB2.

[0666] In some aspects, the antibody or antibody binding fragment specifically binds to LILRB3.

[0667] In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131.

[0668] In some aspects, the antibody or antibody binding fragment comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25.

[0669] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

[0670] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

[0671] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 14. In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 8.

[0672] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 10.

[0673] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 2.

[0674] In some aspects, the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to SEQ ID NO: 6.

[0675] Transgenic Animals

[0676] Also described herein is a transgenic non-human animal having diminished amyloid- f! (A0) pathology, wherein the genome of the animal comprises LILRB2 and / or LILRB3 human transgenes. In some aspects, said animal is a mammal. In some aspects, said animal is a rodent. In some aspects, said animal is a 5XFAD mouse. In some aspects, the LILRB2 and / or LILRB3 human transgenes comprise bacterial artificial chromosome (BAC) inserted transgenes. In some aspects, said transgenic animal exhibits anti-inflammatory characteristics as compared to a wild-type animal.

[0677] In some aspects, said transgenic animal is a 5XFAD mouse or progeny thereof, and wherein said 5XFAD mouse or progeny thereof exhibits anti-inflammatory characteristics as compared to a 5XFAD mouse not having LILRB2 and LILRB3 human transgenes.

[0678] Also provided herein are cells isolated from any of the the transgenic non-human animals described herein. In some aspects, said cell is a myeloid cell. In some aspects, said cell is a microglia. In some aspects, said cell is a disease-associated microglia (DAM). In some aspects, said DAM exhibits increased expression of Sppl, Lpl, and / or Lilrb4a compared to DAM of a 5XFAD mouse not having LILRB2 and LILRB3 human transgenes.

[0679] In various aspects, described herein is a method of identifying a compound for treating and / or preventing a neurological disorder in a subject, the method comprising: administering the compound to any of the transgenic non-human animals described herein or contacting the compound with any of the isolated cells described herein; and determining a therapeutic and / or prophylactic efficacy of the compound on the neurological disorder. In some aspects, the neurological disorder comprises a neuroinflammatory disorder. In some aspects, the neurological disorder comprises dementia, frontotemporal dementia (FTD), Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), Huntington's disease, taupathy disease, Nasu-Hakola disease, and / or multiple sclerosis. In some aspects, the neurological disorder is Alzheimer’s disease. In some aspects, the subject is a human subject.

[0680] In some aspects, the compound comprises an LILRB- specific antibody or antibody fragment. In some aspects, the compound comprises an LILRB2 antagonist. In some aspects, the compound comprises an LILRB 3 agonist.

[0681] SEQUENCES

[0682]

[0683]

[0684]

[0685] EXAMPLES

[0686] The following examples are set forth below to illustrate the methods and results according to the disclosed subject matter. These examples are not intended to be inclusive of all aspects of the subject matter disclosed herein, but rather to illustrate representative methods and results. These examples are not intended to exclude equivalents and variations of the present invention, which are apparent to one skilled in the art.

[0687] Efforts have been made to ensure accuracy with respect to numbers (e.g., amounts, temperature, etc.), but some errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, temperature is in °C or is at ambient temperature, and pressure is at or near atmospheric. There are numerous variations and combinations of reaction conditions, e.g., component concentrations, temperatures, pressures, and other reaction ranges and conditions that can be used to optimize the product purity and yield obtained from the described process. Only reasonable and routine experimentation will be required to optimize such process conditions.

[0688] Example 1.

[0689] Methods

[0690] Cell Lines. BV2 cells were cultured in DMEM (Coming: 10-013-CV) with 10% heat- inactivated fetal bovine serum (FBS; Atlanta Biologicals: S15450), 10,000UI Penicillin / Streptomycin (Gibco: 15140-122), lx MEM NEAA (Gibco: 11140-050), lOmM HEPES (Coming: 25-060-CI), and lx Sodium Pymvate (Gibco: 11360-070). RAW264.7 cells were cultured in RPMI, with 10% heat-inactivated FBS, 10,000UI of Penicillin / Streptomycin, lx MEM NEAA, lOmM HEPES, and lx Sodium Pymvate.

[0691] Bone marrow macrophage culture. Bone marrow cells were isolated from femurs & tibias of respective mice, cultured on 6-well plates at a density 3x105cells / well in 2mL RPMI with 10% heat-inactivated FBS, 10,000UI of Penicillin / Streptomycin, lx MEM NEAA, lOmM HEPES, and lx Sodium Pymvate, and 50ng / mL M-CSF (Peprotech: 315-02) for 5 days. NLRP3 stimulation was done on the 5thday, where applicable. 20ng / mL of LPS (Invivogen: tlrl-eblps) and IFNy (Peprotech: 315-05) treatment were added on day 4 of culture and cells were harvested for experiments on day 5, where applicable.

[0692] Human peripheral blood mononuclear cell isolation. Human PBMCs were obtained from a local blood bank. A 1 : 1 mix of buffy coat to lx PBS was carefully layered on top of 13mL of LymphoPrep (Stem Cell™ Technologies: 07851) in a 50mL conical tube. The samples were then centrifuged at 800xg, 4°C for 30 minutes without any brake. The plasma layer was discarded and the PBMC interface was collected and washed with 50mL of lx PBS and centrifuged at 400xg, 4°C for 10 minutes, with full brake. The PBMC pellet was resuspended in 50mL lx PBS again, and then centrifuged at 120xg, 4°C for 10 minutes with full brake to remove any platelets. The pellet was resuspended in 5mL of ACK Red Blood Cell Lysis Buffer (Thermo Fisher: A1049201), incubated on ice for 5 minutes, then diluted to 50mL in lx PBS. Monocytes were isolated using the Classical Monocyte Isolation Kit (Miltenyi Biotec: 130-117-337), per manufacturer recommendations. 105monocytes were plated in a 96-well plate in RPMI with 10% heat- inactivated FBS, 10,000UI of Penicillin / Streptomycin, lx MEM NEAA, lOmM HEPES, and lx Sodium Pymvate, and 50ng / mL human M-CSF (Peprotech: 300-25) for 5 days. For LPS activation experiments, cells were pretreated with respective antibodies on day 3 of the culture, and lOOng / mL of LPS was added on day 4, prior to analysis on day 5. NLRP3 activation. Murine cells were incubated with lOOng / mL LPS for 3hrs, then 5mM ATP stimulated for the last 30 mins, after which the supernatant was collected, and cells were detached with Accutase (BD Biosciences: 561527) for further experiments. IL-ip in the supernatant was analyzed by ELISA per manufacturer’s instructions (Invitrogen: 88-7013-22). Human monocyte-derived macrophages were treated with lOOng / mL LPS for 3hrs, with subsequent addition of 5mM ATP for 45 minutes. Similarly, supernatants were collected for IL- ip quantification by ELISA (Invitrogen: 88-7261-88) and cells were detached with Accutase for further experiments. siRNA Gene knockdown. Cells were plated and allowed to reach 80% confluence before they were transfected with anti-Pirb siRNA (Santa Cruz Biotechnology: sc-42952). Lyophilized siRNA was reconstituted per manufacturer’s recommendation. The reaction mix consisted of 5 pl of siRNA in 200pL of transfection media (DMEM without any supplements) with 5pL of Lipofectamine RNAiMax (ThermoFisher Scientific: 13778100). The mix was incubated at room temperature for 15-45 minutes. Meanwhile, cells were washed with transfection media (DMEM without any supplements), and then incubated with 800pL of transfection media and 200pL of reagent mix for 5-7hrs at 37 °C. After this initial incubation, ImL of normal culture media (DMEM with all previously mentioned supplements) was added to the cells and incubated for another 18- 24hrs. The media was subsequently exchanged to normal media and respective assays were performed 24-72hrs after transfection. Flow cytometry. Cells were analyzed on a BD LSR Fortessa® X-20. Cells were incubated with TruStain FcX™ (BioLegend: 101320) for 10 minutes prior to staining with their respective antibodies (1 :200 concentration) for 20 minutes, in 4°C, protected from light. Cells were resuspended in 1 :50 DAPI or 1: 1000 Fixable Viability Dye (FVD) prior to analysis on the flow cytometer. For TREM2 quantification on BMDMs, singlet, live (FVD ), CD1 lb+F4 / 80+cells were analyzed. For TREM2 quantification on human monocyte-derived macrophages, singlet, live CD206+cells were treated the same way as the murine BMDMs.

[0693] Animals. 5XFAD {Oakley, 2006 #76} mice were used 6, 8, and 1 Imo of age, as discussed below. Mice were bred and housed in the vivarium within the Houston Methodist Research Institute. 5XFAD mice were purchased from The Jackson Laboratory and cross-bred with BAC- transgenic LILRB2 / LILRB3-containing mice. All animal experiments were conducted in accordance with the animals guidelines and regulations formulated by the Institutional Animal Care and Use Committee (IACUC) of the Houston Methodist Research Institute. C57BL / 6 LILRB2 / 3 transgenic mice were generated via pronuclear injection of bacterial artificial chromosome (BAC) DNA and the offspring inbred to create a stable strain. Each mouse LILRB expression is verified via flow cytometry of peripheral blood cells prior to inclusion in experimental groups. The experiment acquired 5XFAD mice from the Jackson Laboratory, and perform all genotyping according to Jackson Laboratory technical support. PIRB KO mice were a gift from Dr. Toshiyuki Takai of Tohoku University, and they are assessed for PIRB deficiency by flow cytometry of peripheral blood cells. CNS immune cell isolation. Mice were deeply anesthetized with isoflurane, with subsequent cervical dislocation. They were then intracardially perfused with ice cold DBPS, and the brains were isolated and submerged in ice cold DPBS. After removing cerebelli and olfactory bulbs, brains were minced with a scalpel and then homogenized using a Dounce homogenizer, with 15-20 strokes of the loose and tight pestles. Cells were then filtered through a 70pm strainer and transferred to a pre-chilled 15mL conical tube to be pelleted by centrifuging at 500xg for 5 minutes at 4°C. Cells were resuspended in 40% Percoll (Cytiva: 17089101) and centrifuged for 1 hour at 500xg at 4°C. Pelleted cells were washed with ice cold DPBS and centrifuged at 500xg for 5 minutes at 4°C. The supernatant was removed, and the cells were used in the respected downstream analyses. Microglial RT-qPCR. After immune cells were isolated from the CNS as mentioned above, cells were stained with PE-CDl lb (Clone MI / 70), incubated with Anti-PE MicroBeads (Miltenyi Biotec: 130-048-801), and sorted via autoMACS™ Separator per manufacturer recommendations. Sorted CD1 lb+cells were then resuspended in 500pL of TRI-reagent (Thermo: 15596026) and stored in -80°C until use. When ready, RNA was extracted using the chloroform method. cDNA was created using iScript™ cDNA Synthesis Kit (Bio-Rad: 1708890). Primers used for RT-qPCR were:

[0694] Single cell preparation and data processing. Immune cells were isolated as above, and then sorted using a BD FACSAriall. Singlet, viable, PE-CD45+(1 :200, Clone 30-F11) cells were sorted into DMEM (Figure 8A). Isolated CD45+cells from respective mice were prepared via the 10X Genomics Chromium Next GEM Single Cell 3’ v3.1 kit, loaded onto a Chip G, and prepared per manufacturer recommendations. The cells were then sequenced on NovaSeq flow cells at a depth of 20-40,000 reads per cell. FASTQ files were aligned to the mouse genome (mmlO build) using CellRanger from 10X Genomics. Cells were filtered that had <200 genes and >3000 genes; genes that were expressed in <5 cells; and cells that had greater than 15% mitochondrial genes. 35,383 cells were left after quality control.

[0695] Normalization, integration, dimensional reduction. The Seurat R package 4.1.0 { Stuart, 2019 #72;Butler, 2018 #71;Hao, 2021 #73 } was used for scRNAseq analysis and processing. Feature counts were log-normalized, and the top 2,000 variable features were identified from each sample. Two 5XFAD and two FB2 / 3 samples were integrated with 20 dimensions using FindlntegrationAnchors and Integrate Data. The combined dataset was then scaled, and PCA was performed using the 2,000 variable features mentioned above. UMAP was performed using 18 dimensions, based on a quantitative approach and clusters were segmented using FindNeighbors and FindClusters at a resolution of 0.4. FindConservedMarkers was used to differentiate the cluster identities. Clusters 9, 10, and 11 were pruned from analysis and the remaining microglial clusters were normalized, reintegrated, and rescaled, prior to repeating PCA and UMAP with 16 dimensions using the same quantitative approach mentioned above. 34,883 microglia remained for the remainder of the analyses. The pruned populations were reclustered using FindNeighbors and FindClusters at a resolution of 0.4.

[0696] Cluster identification. Clusters were identified using a combination of Seurat’s built in FindConservedMarkers and AddModuleScore features. Cluster definitions were applied for homeostatic, proliferative, interferon-responsive, and disease-associated microglia from Keren- Shaul et al. {Keren-Shaul, 2017 #81 }, Marsh et al. {Marsh, 2022 #109}, and Sala-Frigerio et al. {Sala Frigerio, 2019 #56}. The definition of the antigen-presenting microglia and transitional microglial clusters was inferred based on their top transcripts using FindConservedMarkers. Markers on the violin plots represent the median value.

[0697] Scoring gene set expression. Using Seurat’s built-in AddModuleScore feature, the experiment calculated a score for the gene sets discussed. The average gene expression was compared using Welch’s two sample t-test in R (version 4.1.3). Differentially expressed genes were calculated using Seurat’s built-in FindMarkers function, using the MAST package {Finak, 2015 #75 } to run the test.

[0698] Cell proportion comparison between 5XFAD and FB2 / 3 mice. The study tested the cell proportion change between 5XFAD and FB2 / 3 mice using the Monte-Carlo permutation test {Janssen, 2005 #113 }, assuming the null hypothesis of no difference in proportionality. The alternative hypothesis is there are significant changes in cell proportions between the two groups. FDR correction was applied to p-values and the significance threshold was set to 0.05.

[0699] Trajectory analysis. Slingshot {Street, 2018 #26} (version 2.2.1) R package was used to do the single-cell trajectory analysis and pseudotime inference under R version 4.1.0 {Team, 2021 #116}. The first 20 principal components of cells produced by principal component analysis (PC A) were used as the input, while cluster labels followed previous cell cluster annotation. The PCA analysis was done by implementing the runPCA function of the scatter package {McCarthy, 2017 #114} (version 1.22.0). The HM1 cell cluster and HM2 cell cluster were set as the starting point independently.

[0700] Identification of temporally expressed genes. To identify temporally expressed genes (TEGs), Trade-seq was applied, a statistical framework to model the relationship between cellwise gene expressions and respective pseudotime by generalized additive model (GAM) {Hastie, 1986 #115 }. All genes were ranked by their variance to identify the 1,000 most variable genes. The GAM fitting was applied to the 1000 most variable genes. Then the tested genes were sorted by the p-value of the time-dependent model fit after FDR correction. The significance threshold was set to 0.05.

[0701] Immunohistochemistry. Fixed brains were cryoprotected in 30% sucrose in phosphate- buffered saline, and coronal sections (30 pm) were cut on a sliding microtome (Microm). Serial sections were distributed into ten subseries, each containing every tenth section throughout the rostral-caudal extent of the brain. Each immunostain was performed on one full subseries of sections. Sections underwent 90% formic acid antigen retrieval prior to antibody labeling. Primary antibodies used include rabbit anti-amyloid- 01.43 (ThermoFisher Scientific, 71-5800) and mouse anti-Neurofilament (BioLegend, 837904). Secondary antibodies used include biotinylated goat anti-rabbit secondary antibody (Vector Laboratories, BA- 1000), goat anti -rabbit AlexaFluor 594 (Life Technology, A-11037), and goat anti-mouse AlexaFluor 488 (Life Technology, A-11029). For avidin-biotin / immunoperoxidase immunohistochemistry, ImmPACT Diaminobenzidine EqV (Vector Laboratories, SK-4103) was used as the chromagen. For quantification and analysis, sections were imaged by the RNA In Situ Hybridization Core facility at Baylor College of Medicine. The ImageJ software was used to quantify the percentage area in each region of interest (cortex and hippocampus) covered by a predesignated, fixed optical density threshold. For each mouse, five sections spanning Bregma -1.34mm to -2.54mm were quantified, and then averaged. Data are shown normalized to control groups to illustrate genotype-specific differences. Quantification was performed by an experimenter blinded to the mouse genotype.

[0702] Statistical analyses. Statistical analyses were performed using GraphPad Prism software. Gene expression differences in cell lines were analyzed using unpaired, two-tailed Student’s t-test with correction for false discovery rate. Surface TREM2 and IL- 1 (3 were analyzed using Student’s t-test. Microglial gene expression was analyzed using nested Student’s t-test. Differential A plaque load and neurofilament staining was analyzed using unpaired, two-tailed Student’s t-test, as mentioned above. Simple linear regression was performed between Ap and neurofilament in the cortices and hippocampi to establish their relationship.

[0703] Results

[0704] Pirb regulates Trem2 expression. Because PIRB signaling has an important role in diminishing inflammatory activation {van der Touw, 2018 #1 }, the study tested whether PIRB regulates TREM2 expression. Using the murine macrophage cell line RAW264.7, the experiment knocked down (KD) expression of Pirb using siRNA and observed a concomitant decrease in Trem2 expression (Figure 1A). Similarly, using the murine microglial cell line BV2, KD of Pirb resulted in the same trend of diminishing Trem2 expression (Figure IB). Certain in vitro inflammatory stimuli have been shown to decrease TREM2 {Liu, 2020 #56}. The study further determined whether activation of the NLRP3 inflammasome by LPS and ATP could regulate Trem2 expression in Pirb KD cells. Under NLRP3 activating conditions, Trem2 expression was further diminished in Pirb KD BV2 cells, suggesting that PIRB is responsible for tonic antiinflammatory signaling that maintains Trem2 expression (Figure 1C) {Takai, 2005 #21 }.

[0705] The anti-inflammatory capacity of PIRB signaling was further tested in the myeloid cells of total Pirb knockout Pirb ) mice. There is no available PIRB -detection antibody that reliably differentiates between its paired receptor, PIRA, therefore CD 19+cells were screened for surface PIRB expression by flow cytometry, as CD 19 cells do not express the paired receptor PIRA {Takai, 2005 #21 } (Figures 6A-6E). Bone marrow-derived macrophages (BMDMs) from Pirb ' mice followed a similar trend to that seen in the cell lines - after differentiating bone marrow cells with M-CSF-containing media, BMDMs of Pirb ' mice showed diminished surface TREM2 expression (Figure ID, upper panel). The NLRP3 inflammasome is one of the sources of pathogenic neuroinflammation within AD {Halle, 2008 #35;Yin, 2018 #36}, therefore PIRB signaling was studied to determine whether it plays a role in limiting its activation. Under NLRP3- activating conditions, Pirb' ' BMDMs exhibited decreased surface expression of TREM2 (Figure ID, middle panel), but there was no statistically significant difference in surface TREM2 expression between WT and Pirb'7' BMDMs. Hence, it was believed that this leveling was due to the potent tonic anti-inflammatory role that PIRB serves. Endogenous TREM2 signaling reduces activation of the NLRP3 inflammasome by preventing organization of the NLRP3 inflammasome complex {Sun, 2013 #33 }. The study therefore tested whether the loss of PIRB in myeloid cells would result in the opposite effect. Indeed, activation of the NLRP3 inflammasome in Pirb'7' BMDMs not only decreased surface TREM2 expression, but also increased extracellular levels of IL- 13, a consequence of NLRP3 inflammasome activation {Swanson, 2019 #2} (Figure IE). To further confirm the anti-inflammatory role of PIRB signaling, BMDMs from WT and Pirb'7' mice was stimulated with stereotypical Ml -like culture conditions consisting of LPS and IFNy and observed the same trend in the ability of PIRB to stabilize surface TREM2 expression (Figure ID, bottom panel).

[0706] LILRB transgenes rescue TREM2 expression under inflammatory stimuli in mouse BMDMs. This experiment developed transgenic mice carrying a BAC containing human L1LRB2 and LILRB 3 (LILRB 2 / 3; BAC RP1L634C1) - the human homologs to murine Pirb {Kim, 2013 #3;van der Touw, 2017 #4}- and crossed them with 5XFAD mice, a transgenic mouse model commonly used to study the role of Ap in AD {Oakley, 2006 #76} (FB2 / 3 mice; Figure IF). Upon activation of the NLRP3 inflammasome in BMDMs from 5XFAD and FB2 / 3 mice, those from FB2 / 3 mice showed a lower activation potential, as measured by secreted IL- ip (Figure 1G), suggesting that insertion of LILRB2 and LILRB3 initiates an anti-inflammatory phenotype within myeloid cells. Importantly, the surface TREM2 expression of BMDMs from FB2 / 3 mice was not decreased to the same degree as 5XFAD BMDMs following activation of the NLRP3 inflammasome (Figure 1H). To validate that activation LILRB2 or LILRB3 could have translational results for TREM2 signaling, the study developed antibodies that independently and specifically target human LILRB2 and LILRB 3, acting as functional agonists of the respective receptors. Macrophages derived from peripheral blood monocytes of healthy human donors (hMDMs) were pretreated with respective agonists for 24hrs prior to incubating with 100 ng / mL of LPS overnight. Both LILRB2 and LILRB3 agonists rescued some surface expression of TREM2 after inflammatory stimulus, compared to control IgG (Figure 6F). hMDMS were also further stimulated with 5 mM ATP to oligomerize and activate the NLRP3 inflammasome {Lang, 2018 #34}. Both LILRB2 and LILRB3 agonists also diminished the potential of hMDMs to secrete IL- ip (Figure 6G).

[0707] LILRB transgenes enhance protective transcripts in CNS CDllb+cells from FB2 / 3 mice. Next the effect of LILRB2 / 3 signaling in microglia of 5XFAD mice was investigated, specifically with respect to the disease-associated microglia (DAM) phenotype {Keren-Shaul, 2017 #18}. DAM have a crucial role in limiting A pathology within the CNS parenchyma, and it was believed that microglia would exhibit enhanced Trem2 expression, similar to that of the BMDMs (Figure 1H). After isolating CD1 lb+cells from the brains of 8-month-old mice (Figures 2A-C, Figures 7A-7B), the study found that Trem2 was indeed upregulated in FB2 / 3 mice compared to that of 5XFAD mice (Figure 2D). Importantly, many DAM genes were concurrently upregulated in CNS CDl lb+cells from FB2 / 3 mice, including Trem2 -independent genes such as Tyrobp and Ctsb (Figure 2E, Figure 7C), and Trem2 -dependent genes, such as Itgax, Cst7, Lpl, Lilrb4a, and Ax / (Figures 2F-2J). The experiment used 8-month-old mice as this is the age when the DAM population is most pronounced { Keren-Shaul, 2017 #18 } . Interestingly, the homeostatic microglial marker P2ryl2 was also increased in the CNS CDl lb+cells of FB2 / 3 mice, despite upregulation of many of the DAM markers (Figure 7D). This is reminiscent of human AD- associated microglia described elsewhere, noting that homeostatic microglia markers are upregulated, rather down downregulated in murine AD-associated microglia {Zhou, 2020 #7}. Transcriptomics of human microglial cells reveal an IRF8-driven reactive microglia phenotype similar to that of murine DAM {Masuda, 2012 #37;Zhou, 2020 #7}, therefore the experiment also measured fr 8' expression. CNS CDl lb+cells from FB2 / 3 mice also showed upregulation of Irf8 (Figure 7E), implying that LILRB2 / 3 is involved in enhanced microglial reactivity to Ap pathology.

[0708] LILRB transgenes enhance the expansion of DAM. Different -omics studies have highlighted that microglia are largely heterogeneous within the brain parenchyma {Keren-Shaul, 2017 #18;Sala Frigerio, 2019 #24;Hammond, 2019 #25;Marsh, 2022 #22}. To determine the effects of LILRB2 / 3 signaling at single-cell resolution, single-cell RNA sequencing (scRNAseq) was performed on FAC-sorted immune cells (CD45+) isolated from the CNS of the respective mice (Figures 3A, 8A). To avoid the acquisition of an artificially activated transcriptional state of microglia upon ex vivo preparation under 37 °C dissociation conditions {Marsh, 2022 #22}, the time the samples spent away from ice or a 4°C environment was minimized. Unsupervised clustering of 35,383 cells from 5XFAD and FB2 / 3 mice resulted in 12 clusters that were identified by the top 2000 differentially expressed genes (Figure 3B). To confirm the preparation did not confound differential gene expression, the gene expression pattern representative of artificial ex- vivo activation was measured (Fos, Jun, Duspl, Hspala, and Zfp36{Marsh, 2022 #22}) and did not find appreciable expression in any cluster (Figure 8B-8C). Using Seurat’s built in AddModuleScore function, the study also scored the clusters against the combined gene set, rather than the individual genes, and did not find any high-scoring clusters (Figure 3C).

[0709] The study next identified each of the 12 clusters using predefined signature markers {Marsh, 2022 #22;Keren-Shaul, 2017 #18;Wang, 2020 #9} (Figures 3D, 8G-8K). Clusters 0-8 are microglia, with highest amounts of Tmemll9 and Hexb expression, which was used as canonical microglial markers (Figure 3D); cluster 9 scored highly as an infiltrating monocyte / macrophage cluster, defined by low expression of Tmemll9 and Hexb with highest expression of the monocyte / macrophage gene set Ms4a7, Ccr2, and Ms4a4c (Figure 8D); cluster 10 scores highly as an infiltrating B cell, defined by the highest expression of the gene set containing Cd79a, and Ebfl (Figure 8E); and cluster 11 is an infiltrating neutrophil population, with highest expression of Ngp (Figure 8F). For further analysis, the study pruned the peripheral immune cells from the microglial clusters and performed another round of unsupervised clustering, reducing the populations down to 8 clusters. Each individual cluster with gene sets from previous studies was scored and identified clusters 0 and 1 as homeostatic microglia clusters (P2ryl2, P2ryl3, Serinc3, Cx3crl, Tgfbl, Tmemll9, Txnip, Glut), HM1 and HM2, making up the largest proportion of the microglia populations (-53%) (Figures 3E-3F). The clusters across several gene sets that define the homeostatic state {Keren-Shaul, 2017 #18;Sala Frigerio, 2019 #24;Wang, 2020 #9} were scored and the labels remained the same (Figures 8G-8J). Cluster 5 scored highly as an interferon-responsive microglial (IRM) cluster, defined by high expression of Ifit, lfit3, Irf7, and Oasl2 {Marsh, 2022 #22;Sala Frigerio, 2019 #24} (Figure 8K). Cluster 6 was highly enriched in genes involved with antigen processing and presentation (Cd74, H2-Aa, H2-Abl, Cd52, Cst7) and therefore was labeled as antigen presenting microglia (AgM, Figure 8L). Cluster 7 was highly enriched for proliferation genes (Top2a, Mki67, Birc5) and thus labeled as the proliferative microglia subset (Figure 8M) {Wang, 2020 #9;Marsh, 2022 #22;Sala Frigerio, 2019 #24}. Next, the DAM populations was identified based on definitions from Keren-Shaul et al. {Keren-Shaul, 2017 #18}. DAM stage 1 (Apoe, Tyrobp, B2m, Ctsb, Timp2, H2-D1, Fthl, Lyz2, Cstb, Ctsd) and stage 2 (Ank, Sppl,Axl, Csfl, Cst7, Cd9, Itgax, Lpl, Ctsz, Ctsl, Ccl6, Cadml, Cd63, Ctsa, Serpine2, Ctsz, Cd52, Hifla) marker genes increased through clusters 3 and 2, and was, therefore, labeled them as such (Figures 8N-8O). Cluster 4 mapped separately from other microglial populations and had features of both homeostatic and DAM gene sets, warranting the label of transitioning microglia (Figures 3E, 8G-8J, 8N-8O).

[0710] Comparison of the cluster frequencies between 5XFAD and FB2 / 3 mice revealed a decrease in HM1 with a concomitant increase in HM2; both DAM 1 and DAM 2 populations were increased in FB2 / 3 mice compared to 5XFAD mice; the proportions of TrM, IrM, AgM, and PM were decreased in FB2 / 3 mice (Figure 3G). Quantification of cluster proportion enrichment showed significant reduction of AgM by -60% and HM1 by -40% in FB2 / 3 mice, with a concomitant -20% increase of HM2 & DAM 1, and -60% increase of DAM 2 cells (Figure 3H).

[0711] FB2 / 3 microglia express DAM markers at earlier transition stages. Because CDllb+cells isolated from the CNS showed an increase in DAM programming (Figures 2D-2J), the individual clusters were scored against markers that define the progression into DAM in the 5XFAD and FB2 / 3 groups (Table 1). HM1 ranked higher in the homeostatic microglia signature gene set in the FB2 / 3 group, with diminished DAM-like programming, with p values well below 1014(Figures 9A-9C, Table 1). Without wishing to be bound by theory, it is speculated that the increase in P2ryl2 obtained by qPCR analysis of CD1 lb+cells from the CNS (Figure 7D) was in large part due to the HM1 population, as it represented -30% of the total microglial population (Figure 3G) and likely overwhelmed the expression signal. Interestingly, the HM2 subset followed the opposite trend in that the homeostatic microglia gene set signature was diminished in the FB2 / 3 mice (Figures 9A-9C), while stage 1 and 2 DAM gene set signatures were upregulated, with a more dramatic difference in initiation of stage 1 programming (p < 1015) (Table 1). The identified DAM 1 and DAM 2 populations were compared in their overall programming and found that the DAM 1 population had enhanced expression of the signature gene sets of both stages in the DAM programming in the FB2 / 3 background (p < 1015) (Figure 9D, Table 1). The DAM 2 population showed enhanced expression of the Trem2-independent signature (p < 10"15), but not of the Trem2 -dependent signature (p = 0.5798) (Figure 9E, Table 1). Based on the transcript data that showed lrf8 was upregulated in the FB2 / 3 mice, a phenomenon that is associated with the development of DAM in humans, the study checked whether the individual clusters from FB2 / 3 mice showed upregulation of Irf8 as well. Although lrf8 was not a differentially expressed gene in any cluster, it was upregulated in the HM2, DAM 1, and DAM 2 clusters (Figures 9F-9I). Table 1. Scores of individual clusters against markers defining the progression into DAM in the 5XFAD and FB2 / 3 groups. The study next used Slingshot { Street, 2018 #26 } to infer the pseudo-temporal trajectories of the microglia. It was reasoned that homeostatic microglia constitute the initial cell states from which AD pathogenesis driven microglial states evolve. To this end, the adequacy of utilizing HM1 and HM2 clusters as the initial starting points was examined. Using HM1 as the starting point, pseudotime analysis did not effectively distinguish DAM1, IRM, AgM and PM clusters, and did not infer the transition into the DAM stage 2 cluster (Figure 10A). Therefore, it was inferred HM2 as the trajectory starting point and used it as the anchor (Figure 4A). Microglia followed five distinct trajectories (T1-T5; Figures 10B-10F), of which only T1 and T2 converged on DAM (Figures 4A, 10B-10C). Volcano plots showing differential gene expression between 5XFAD and FB2 / 3 mice in clusters HM1 (Figure 10G) and AgM (Figure 10H) clusters, respectively.The AgM cluster is intermixed within the HM2-DAM2 trajectory, implying these cells expressed activation genes like the Ap-reaclive microglia (Figures 4A-4B). The PM cluster exhibited a distinct point of reference within pseudotime, implying that they may branch off from the homeostatic group (Figure 4B). The TrM cluster did not align well within the pseudotime transitions, suggesting that it wasn’t necessarily part of the trajectory, despite expressing gene sets representative of both homeostatic microglia and DAM (Figures 4A-B, 8M-8O).

[0712] The study further identified significant temporally expressed genes (TEGs) using tradeSeq {Van den Berge, 2020 #70}, a statistical framework used to evaluate significant gene expression changes along a cell trajectory based on the generalized additive model {Hastie, 1986 #58}. Analysis returned 975 significant TEGs after FDR correction, with the top TEG being Apoe (F value = 97021 , p = 0). Importantly, Apoe expression had a more significant time-dependent change in the FB2 / 3 mice and was substantially more enriched earlier in the trajectory, compared to the 5XFAD mice (Figure 4C). Trem2 was also among the top TEGs (F value = 39432, p = 0), in alignment with previous studies that describe its upregulation through AD development {McQuade, 2020 #63}. Like Apoe, Trem2 was expressed more in FB2 / 3 mice earlier in the trajectory (Figure 4D), further supporting that LILRB2 / 3 enhanced microglial activation.

[0713] The study isolated the clusters within the HM2-DAM2 transition and stratified them by 5XFAD and FB2 / 3 groups - in every transcriptional state apart from the DAM2 cluster, the FB2 / 3 microglia were further along in pseudotime (Figure 4B), suggesting that LILRB2 / 3 signaling is involved in enhancing the microglial reaction to A0. The difference within the DAM stage 2 state followed the same trend seen in the gene set enrichment analysis presented in Figure 9E, where the Trem2 -dependent gene set was not statistically different between the 5XFAD and FB2 / 3 mice (Table 1). Thus, the transition from the homeostatic microglial state to DAM state was enhanced in the context of LILRB2 / LILRB3, displaying earlier initiation of the transcriptional signatures that respond to Ap pathology and a higher proportion of cells converted to the rescuing phenotype.

[0714] FB2 / 3 microglia show enhanced activation. The study further analyzed the differentially expressed genes in the clusters that had contrasting enrichment - HM1, HM2, DAM 1, DAM2, and AgM (Figures 3H, 4E-4F, 10G-10H). In comparison to the 5XFAD mice, these clusters from FB2 / 3 mice displayed an enhanced activation signature and stronger reactivity to A . In the FB2 / 3 mice, the HM2 cluster showed a decrease in homeostatic markers, such as Crybbl {Grubman, 2021 #23 } , Ecscr {Jin, 2021 #68 } , and Ms4a6b { Schwabe, 2020 #69 } , with a concomitant increase in activation transcripts, such as Ifitm3, and Ifi27l2a {Hammond, 2019 #25 }. Importantly the HM2 cluster from FB2 / 3 mice was already enriched in transcripts that define the DAM state - it showed an upregulation of the Trem2-independent genes Apoe, and Lyz2, and upregulation of a Trem2- dependent gene, Lpl (Figure 4E). The HM2 cluster also showed modulation of other genes involved in immune activation, such as decreased Tmeml76b expression, which is otherwise associated with decrease microglial viability {Melchior, 2010 #65 }; and a decrease in Fcgr2b, which has been shown to polarize myeloid cells toward a “M2-like” functional state {Chauhan, 2017 #66}. The DAM stage 1 cluster followed a similar trend - the FB2 / 3 mice showed a decrease in homeostatic Crybbl expression and Lag3, inhibition of which has been shown to induce neurogenesis {Rimmerman, 2021 #67}. More importantly, however, is that the FB2 / 3 mice showed an upregulation of the Trem2 -dependent transcripts Sppl, Lpl, and Lilrb4a, supporting that LILRB2 / 3 pushes the DAM stage 1 trajectory further along in pseudotime towards the DAM stage 2 transcriptional phenotype (Figure 4F). Although enriched proportionally in the FB2 / 3 mice, the DAM2 cluster did not display any differentially expressed genes among the 5XFAD and FB2 / 3 mice (Figure 3H).

[0715] FB2 / 3 mice exhibit diminished AD-associated neuropathology. The study next evaluated the effect of LILRB2 / 3 insertion on amyloid deposition within the brain parenchyma. It was believed that the increase in the protective microglial populations in the FB2 / 3 mice would result in a concomitant decrease in Ap pathology. Immunofluorescence analysis was performed on brains of 11 -month-old mice - when disease burden begins to peak { Forner, 2021 #27 } - by staining for A . It was found that the FB2 / 3 mice had a -48% (p < 0.05) reduction in cortical plaque load and a -51 % (p < 0.05) reduction in hippocampal plaque load, compared to 5XFAD mice (Figures 5A- 5E). The study also quantified plaque load after DAB staining and observed similar decreases in cortical Ap plaque load (Figures 11A-11D). Since Ap is directly neurotoxic {Ciudad, 2020 #28}, the experiment also quantified the amount of accumulated neuronal damage. The experiment concurrently measured neuritic dystrophy on the same sections by staining against extensively phosphorylated neurofilaments, which accumulates in dystrophic neurites in brains of both AD patients and mice {Masliah, 1996 #77}. Compared to 5XFAD mice, the brains from FB2 / 3 mice showed a -53% reduction in cortical neurofilament staining (p < 0.05) (Figure 5F), and -41% reduction in hippocampal neurofilament staining, although not statistically significant (Figure 5G). To further validate this model and show that the reduction of Ap plaque load in FB2 / 3 mice corresponded with diminishing neuronal damage, a simple regression was performed between Ap immunoreactivity and neurofilament immunoreactivity. The study found that there was a direct relationship between Ap load and neurofilament immunoreactivity in the cortex, with R = 0.8521 (p < 0.005) (Figure 5H), as well as in the hippocampus, with R = 0.7533 (p < 0.05) (Figure 51). Discussion

[0716] Although LILRB2 was previously defined as a neuronal receptor for Ap, there have not been any investigations into the effect of LILRB2 / 3 on brain-resident immune cells in a mouse model of AD. This data showed that LILRB2 / 3 had a rescuing capacity on pathogenic AD- associated neuroinflammation. The experiement found that the murine homolog to LILRB, PIRB, regulated myeloid cell expression of Trem2 and activation of the NLRP3 inflammasome (Figures 1A-1E). The study chose the NLRP3 activation model in particular because of its association with pathogenesis in AD {Yin, 2018 #36} {Zhang, 2020 #59}. Furthermore, NLRP3 activation can serve as a metric for activation of signaling downstream of TREM2, as TREM2 activation can prevent oligomerization of the inflammasome {Qu, 2018 #60} {Jairaman, 2022 #61 }. Tonic signaling through LILRB 2 / 3 enhanced surface TREM2 expression in murine macrophages, while simultaneously limiting their potential to activate the NLRP3 inflammasome (Figures 1A-1E), suggesting that LILRB2 / 3 prevents internalization and cessation of TREM2 signaling. After developing a mouse model of AD, which includes BAC-inserted LILRB2 / 3 in the 5XFAD background, the experiment showed that LILRB2 / 3 enhanced the neuroprotective DAM transcription signature in CDllb+cells isolated from the CNS and in microglia analyzed by scRNAseq. Age-matched male mice were used for all transcriptomic analyses.

[0717] The study showed that the insertion of the BAC transgenes resulted in an expansion of neuroprotective microglia subsets with a simultaneous reduction in a homeostatic microglia population that exhibited lower expression of the gene sets associated with the DAM phenotype. Trajectory prediction using pseudotime analysis showed a gradual transition from the HM2 state, through HM1, DAM 1, and DAM 2, with other reactive microglial clusters spread across the transition. After stratifying the pseuodotime trajectory by genotype, a clear difference in microglial trajectory was discovered - clusters HM2, HM1, and DAM 1 from FB2 / 3 mice were further along in the development towards the Trem2 -dependent DAM 2 phenotype, when compared to 5XFAD mice, suggesting that LILRB2 / 3 signaling enhances microglial reactivity to Ap and their differentiation into support cells. Enhanced expression of A -processing and activation genes in microglia of FB2 / 3 mice further support the beneficial role of LILRB2 / 3 signaling (Figures 4E- 4F). Although further along in pseudotime than HM2, HM1 did not express a greater DAM signature in FB2 / 3 mice (Figures 9A-9C). However, there was -40% reduction in the HM1 proportion in FB2 / 3 mice (Figure 3H), therefore the reactive phenotype changes of HM2 and DAM 1 most likely played a larger role in neuroprotection. The additional -50% enrichment of DAM 2 cells in FB2 / 3 mice was most probably the greatest source of increased neuroprotective transcripts seen by qPCR (Figure 2A). Trem2 -dependent DAM 2 cells represent a crucial sink for Ap clearance and processing {Wang, 2015 #20;Keren-Shaul, 2017 #18;Ulland, 2017 #29;Joshi, 2021 #62;McQuade, 2020 #63}, thus the expansion of the population under the induction of LILRB2 / 3 signaling is an important discovery.

[0718] Additionally, the experiment showed that the expanded neuroprotective microglia from FB2 / 3 mice also differentially expressed Trem2 -dependent genes that are responsible for Ap processing and tissue remodeling, with simultaneous downregulation of homeostatic markers. With this, it was shown that LILRB2 / 3 in vivo signaling initiated a broad network to lessen the pathological A -mediated burden in 5XFAD mice. Without wishing to be bound by theory, it is believed that lrf8 may be a mediator in the signaling network.

[0719] Moreover, the signaling initiated by LILRB2 / 3 culminated into a dramatic reduction of Ap deposits - about half that seen in age-matched 5XFAD mice (Figures 5D-5E). A similar reduction was observed in neurofilament accumulation (Figures 5F-5G), indicating that the programing set in place by LILRB2 / 3 reduced the neuritic dystrophy typically observed in 5XFAD mice {Lazic, 2020 #40}. For tissue pathology, mice from both sexes were analyzed. While changes due to sex could not be discounted, sex was not determined to be an appreciable confounder for the parameters that were measured (Figures 11E-H), in alignment with the comprehensive study performed by Oblak et al. that showed no difference in plaque radio tracing {Oblak, 2021 #38}.

[0720] The question remains which of the inserted LILRB receptors is most critical in expanding the neuroprotective capacity of microglia - whether it is LILRB2, LILRB3, or the combination of both. Both receptors have been described to exert potent anti-inflammatory effects via activation of their ITIM {van der Touw, 2017 #4;Deng, 2021 #39}, therefore both receptors have the potential to serve as regulators of Trem2 -dependent transcriptional states. Furthermore, murine ligands that interact with either of the human receptors are yet to be identified. Components of HLA serve as ligands to the majority of LILRB family of proteins, other than the still orphan receptor, LILRB3 {van der Touw, 2017 #4}. Without wishing to be bound by theory, it is believed that murine MHC is similar enough in homology to act as a functional ligand to human LILRB2. However, signal transduction through LILRB3 may also operate in a similar manner. The homology of murine PIRB is about 50% to that of LILRB2 and LILRB3 {Kim, 2013 #3 }, and therefore it was believed that the added gene dosage from the inserted LILRB members creates additional reactive signaling via activation by MHC proteins that is powerful enough to drive the exhibited phenotype. Af> also serves as a functional ligand to LILRB2 { Kim, 2013 #3 } , but in vitro culture of BMDMs in the absence of A[3 already showed anti-inflammatory capacity (Figures 1A- IE), therefore it is unlikely that this is the sole activation source, as are Nogo {Takeda, 2017 #47} or Angiopoietin- like proteins {Person, 2015 #46}.

[0721] In summary, the study exemplified that LILRB2 / 3 transgenes on the 5XFAD background diminished the inflammatory capacity of macrophages, enhanced Trem2 expression along with dependent signaling, and increased expression of other microglia-derived neuroprotective genes. This experiment indicates that LILRB2 / 3 can serve as an upstream regulator to many signaling cascades important in diminishing Ap-mediated pathology.

[0722] Example 2.

[0723] LILRB3 agonist antibodies affect gene expression profiles and phagocytosis. LILRB2 is highly expressed on CDl lb+cells isolated from FB2 / 3 brains (Figures 2B-2E). Neuronal organoids derived from human iPSCs also express high levels of LILRB2 while LILRB3 expression is undetectable (Figure 13A). This study developed antibodies specific to LILRB3, B 1 , B2, B4, without cross-reactivity to other members of the LILR family, that function as agonists of their respective receptors. Since neurons do not express LILRB3, the study aimed to specifically target LILRB3 on microglia and MDM in order to avoid potential toxicity (Figure 13A) through the use of aLILRB3 agonist therapy. Alternatively, the experiment uses a combination of aLILRB3 with aLILRBl or LILRB4 to quantify potential synergistic effects, as neurons exhibit very low surface expression of these members, or in combination with aLILRB2 antagonist blockade antibody. Activation of the respective receptors results in reduced secretion of TNFa and IL-ip in human monocyte-derived macrophages cultured with LPS (Figure 13B). The study further evaluated the effect of agonist aLILRB3 vs. antagonist aLILRB2 or combination of MDM mediated phagocytosis ability on Ab filament and fluorescent beads uptake. The mean fluorescent intensities were compared. Treating MDM with an agonist antibody enhanced both intracellular Ap filament and fluorescent bead uptake significantly, but not antagonist (Figure 13C), while promoting expression of the CD206 mannose receptor, M2 markers (Figure 13D). LILRB2 reporter cells have been constructed using the LILRB2 extracellular domain with a DAP 12 activation signal for NFAT. Specific ligand engagement with this receptor trigger NF AT activation with subsequent induction of GFP or YFP expression. The study further confirmed that a antagonist LILRB2 antibody, can block A mediated activation on reporter cells (Figure 13E) thus indicating that aLILRB2 can block AP-and receptor interaction, if it may blockade Ab mediated neuron toxicity. LILRB signaling can regulate immune activation stereotypical for AD. Specifically, LILRB3 signaling can diminish activation of the NLRP3 inflammasome and promote TREM2 activation to enhance the neuroprotective function of microglia and MDM. To enhance the signaling effect of LILRB, antibodies have been developed for targeting the individual LILRB receptors that activate the intracellular anti-inflammatory networks, as exemplified in Figure 14. By interfering with Ap-mediated inflammation seen in AD, AD pathogenesis may be attenuated and rescue the associated learning and memory deficits (Figures 5A-5I).

[0724] Determination of the effects of LILRB3 agonism on TREM2 expression, NLRP3 activation, microglia phagocytosis, and inflammatory responses. Experimental data shows that LILRB signaling enhances TREM2 expression and has strong anti-inflammatory effects. Compared to regular 5XFAD mice, BMDMs from 5XFAD mice with BAC-inserted LILRB2 / LILRB3 human genes (FB2 / 3) exhibited attenuated activation of the NLRP3 inflammasome, as assessed by lower IL- 13 production (Figures 1F-1H). Under inflammatory in vitro conditions, BMDMs from FB2 / 3 mice exhibited rescue of TREM2, which serves as an endogenous inhibitor of the NLRP3 inflammasome assembly (Figures 1F-1H, 2B-2J). Moreover, microglia isolated from FB2 / 3 mice showed increased expression of TREM2 and DAM-associated genes compared to 5XFAD control mice by RT-PCR and scRNAseq (Figures 2B-2J, 3G, 12, 4F, 4E). The experimental data suggests that LILRB3 can regulate DAM function and the pathogenic NLRP3 inflammasome in AD and serve as a new therapeutic strategy for limiting neuroinflammation.

[0725] Delineating the network by which LILRB3 rescues TREM2 expression, enhances phagocytosis and reducing inflammation through the use of agonist antibodies targeting LILRB3. TREM2 has been shown to be a crucial receptor not only in limiting inflammation in AD but also in maintaining overall microglial health and phagocytic capacity. Under inflammatory conditions in vitro, surface TREM2 expression is decreased due to upregulation of miRNA-34, secondary to NFKB activation43. With the presence of BAC-inserted LILRB2 / 3, TREM2 expression was rescued through an unknown mechanism. Using the same 6 month old WT, 5XFAD, and BAC B2 / B3 murine BMDMs, LILRB 2 / 3 transduced murine BV2 microglia cell line, and human microglia cell line HMC3, the network responsible for rescuing TREM2 expression using a combination of RNA sequencing (RNAseq), RPPA (reverse phase protein array), and immunoblotting can be investigated to detect changes in NFKB activation. Bulk RNAseq and protein array allows the identification of the precise perturbations caused by the immunoinhibitory pathway activated by LILRB, especially concerning reduced miRNA-34a. LILRB3-specific agonist and antagonist antibodies can be used to activate and inhibit LILRB3 signaling, respectively. RNAseq data from agonist vs. antagonist treated groups, in addition to Ig controls, are compared. Moreover, pathway analysis can also allow the discovery of novel therapeutic targets to enhance the anti-inflammatory capability of LILRB activation. Several LILRB3 and TREM2 prexpression reporter cells have been generated (GFP color vs. YFP color), as shown in Figures 13A-13E, which are co-transfected into human HMC3 cell or mouse BV2 cells microglia to measure the LILRB3 antagonist vs. agonist effect on the activation or blocking LILRB3 effect for TREM2 activation and myeloid cells’ functional activity change e.g. phagocytosis, cytokine secretion and DAM1 vs. DAM2 marker changes.

[0726] Determining whether LILRB3 interferes with NLRP3 activation through TREM2 and diminishes NLRP3 activation in response to stimulation by LILRB3 agonist. In AD, the NLRP3 inflammasome is activated specifically in response to Ap accumulation in the parenchymal space37,46. Ap serves as the danger signal that activates microglia, causing NFKB-dependent upregulation and assembly of the components of the NLRP3 inflammasome - ASC, caspase- 1, NLRP3, and pro-IL-ip, resulting in the release of mature IL-ip47. In vitro, the NLRP3 inflammasome can be activated in BMDMs and other myeloid cells via ligation of HMGB1 or LPS to TLR4 to initiate transcription of the inflammasome components, with the subsequent addition of ATP or A to trigger oligomerization of the complex, activation of caspase- 1, and cleavage of the pro-IL-ip precursor to active IL-ip4849.

[0727] It has been demonstrated that antagonism of LILRB2 signaling can induce NFKB activation29and similar effects have been shown via other LILRB members44, offering one possible mechanism of regulation. Additionally, TREM2 has also been shown to act as a direct inhibitor of NLRP3 activation, specifically by preventing oligomerization of the inflammasome components33. Using primary myeloid cells and the murine LILRB3 transduced microglial cell line, BV2, with antiLILRB3 agonist vs. antagonist vs. control Ig, the precise mechanism through which LILRB interferes with NLRP3 activation can be delineated by quantifying the individual components necessary for inflammasome assembly, as well as NFKB components p65 and p50, and IKB via immunoblotting. Additionally, a rescuing effect of TREM2 expression has been discovered in the BMDM from primary cultured BAC-inserted LILRB2 / 3 (FB2 / 3) mice (Figures 1F-1H). Therefore, inflammasome activation of TREM2’s intracellular adaptor protein, TYR0BP / DAP12 w / wo LILRB3 engagement can be quantified. Activation of TYR0BP / DAP12 by immunoprecipitating the adaptor protein with TREM2, followed by probing with an a- phosphotyrosine detection antibody23can then be distinguished. Similarly, after gene silencing of TREM2 with siRNA or complete KO, the anti-inflammatory effect of LILRB3 can be observed to determined whether it is dependent on TREM2 expression. With these data, it is possible to specifically distinguish whether LILRB’s effect on diminishment of NLRP3 activation is due to inhibition of inflammasome component expression or assembly through TREM2 activation.

[0728] Determining the effect of LILRB3 agonist on TREM2 activity, microglia phagocytosis and reduction of inflammation in vivo. Evaluation of the agonist antiLILRB3 antibody’s effect on increasing microglia functional activity in vivo is performed. However, antibody penetration of the blood-brain barrier (BBB) is limited, even in the presence of BBB degeneration seen in AD23. To facilitate the LILRB3-specific antibody’s ability to penetrate the BBB, the study employed an antibody conjugated with angiopep-2 (AP2) peptide, the synthetic low-density lipoprotein receptor-related protein 1 (LRP-1). AP2 conjugation to antibody has been shown to help antibodies penetrate the BBB50. The study conjugated AP2 with the Cy5-labeled aLILRB3 antibody (red color) and showed that this antibody can more effectively penetrate the BBB thus accessing the brain with continuous increase / homing to the brain, with decreased presence in other organs (Figures 15A-15C). The study administers AP-2-agonist anti-LILRB3 or AP-2 control Ig (40 mg / kg / every 3 days) to 8 month old FAD, FB2 / 3, and control WT or LILRB2 / B3 mice for 7 days and 14 days, with some mice infused with methoxy X-4 10 mg / kg after 3 hours to check the status of Ap and phagocytosis of A in microglia and MDM, with some mice sacrificed for pathology, western blot for TREM2 activity and cytokine measurement.

[0729] Determining the mechanism by which LILRB3 activation enhances the DAM state and exerts its neuroprotective function. The effect of the anti-LILRB3 agonist can be evaluated on the anti-inflammatory network initiated by the BAC-inserted LILRB2 / LILRB3 transgenes at a single-cell resolution and correlate the signaling pathways with available bioinformatics of 2000 human AD specimens. TREM2 has an important role in clearing Ap plaques from the CNS parenchyma in AD. With the genetic insertion of human LILRB2 / LILRB3 transgenes into 5XFAD mice, the study found significant upregulation of TREM2 expression, both in vitro and ex vivo (Figures 1F-1H, 2B-2J), resulting in the enhancement of neuroprotective microglial genes (Figures 12, 17A-17F) and diminishment of Ap plaque-load in the parenchymal space (Figures 5A-5I). Thus, the effects of LILRB3 signaling on the entirety of the immunological landscape of the CNS in the murine model of AD using scRNAseq can be defined in conjunction with LILRB3- specific agonist antibody1,40. The respective roles of microglia and peripheral immune components are compared, such as MDMs, to previous studies within the 5XFAD model22. Additionally, the anti-inflammatory network initiated by the agonist antibody in BAC-inserted LILRB2 / LILRB3 transgenic 5XFAD (FB2 / 3) can be evaluated and used to correlate these disease progression networks with human AD single-cell information40,51.

[0730] Determining the pathways affected by LILRB3 in microglia and MDM and further agonize this network with agonist antibodies targeting LILRB3. Despite being a small population of cells, DAM and MDM are importan in curtailing the progression of AP-mediated neurological deficits1,52. Central to their development is TREM2, which initiates the expression of a network of genes that are targeted toward processing A , such as Tyrobp, Lilrb4a, Lpl, and Axl (Figures 2B-2J)53. The experimental data shows many DAM genes are enhanced in the presence of human Lilrb2 / 3 transgenes in 5XFAD mice by RT-qPCR and single cell analysis (Figures 2B- 2J, 12); therefore, scRNAseq can be used to further characterize the immune landscape. After enriching for CD45+cells from the CNS, scRNAseq allows the capture of the individual immune components involved in the Ap model of AD. The scRNAseq study showed an enhancement of the DAM phenotype in FB2 / 3 mice. Therefore, further evaluation of the signaling network initiated by aLILRB3 agonist therapy within the gene signatures of D AMs and infiltrating MDMs is explored. To do so, the study treats WT, 5XFAD and FB2 / 3 mice with AP2-conjugated LILRB3- specific agonist antibody (vs. control Ig) at 40 mg / kg body weight weekly for two months once the mice reach 5-6 months of age. Following treatment, scRNAseq is performed on CD45+cells isolated from the brains of the respective mice to intimately characterize the immune landscape affected by aLILRB3 agonist therapy. A minimum of 5-6 age-matched mice per group is used, including both sexes. The study has demonstrated that agonist anti-LILRB3 antibodies have achieved a significant effect in promoting maturation toward CD 14 / CD 16 / CD 163 positive cells, i.e. M2 like phenotype (Figure 16A). Furthermore, the experiment tested the anti-inflammatory effect of LILRB3 agonist in vivo. Anti-LILRB3 could significantly reduce LPS-mediated inflammatory cytokines in the mouse circulation, as assessed by TNFa and IL-6 in serum using humanized NSG-SGM3 mice from repeated experiments (Figure 16B). It has also been found that agonist aLILRB3 antibodies in conjunction with an aLILRBl or aB4 agonist could induce a synergistic effect on the inhibition of myeloid cell-mediated T cell proliferation and promote maturation toward CD14+ / CD16+ / CD163+myeloid cells, i.e. M2-like phenotype, and suppress inflammatory cytokines.

[0731] Correlating single-cell RNA sequencing information with in situ identification of active cells with imaging mass cytometry and spatial transcriptomics. The DAM transcriptional state is initiated by the AP-mediated pathology seen in AD. Microglia assume this state in an attempt to mitigate further AP-mediated neurotoxicity. The data show that FB2 / 3 mice exhibit upregulation of signature transcripts that resemble HM2, DAM1 and DAM2 (Figure 12), implying that LILRB signaling may enhance these neuroprotective microglia. To gain a complete understanding of the CNS microenvironment, the DAM transcriptional signature can be quantified on a protein level while maintaining spatial and tissue architecture. Unlike homeostatic microglia, DAM cluster closer to AP plaques, seemingly in an attempt to shield neurons and process plaques1. Because LILRB -expressing myeloid or microglia cells have an enhanced DAM profile and phagocytic activity upon treatement with LILRB3 agonist antibody (Figures 12, 13A-13E), more microglia may cluster around and engage A plaques, as measured by staining with the aAp 3D6 antibody, limiting potential neurotoxicity. While preserving spatial information through immunofluorescence and Imaging Mass Cytometry (IMC), the microglial and MDM presence surrounding Ap plaques can be quantified and characterized41. IMC staining procedure is well established in the immune assessment core55,56,57. Furthermore, the added reduction in plaque formation after treating mice with aLILRB3 agonist antibodies can be quantified in a similar manner as shown in Figures 5A-5I. aLILRB3 treated FB2 / 3 mice exhibit a greater reduction in plaque presence compared to regular 5XFAD or control Ig-treated mice. Gene expression level changes are further evaluated through single-cell spatial transcriptomics and follow the localization of DAM and MDM populations. Added changes in the gene signatures are compared after treating mice with aLILRB3 agonist therapy.

[0732] Correllating disease progression with the human AD database to compare therapeutic outcomes at a single cell level. To have a complete understanding of the CNS microenvironment, the DAM transcriptional signature on a protein-level is quantified, while maintaining tissue architecture and spatial data. Unlike homeostatic microglia, DAM localizes closer to Ap plaques, seemingly in an attempt to shield neurons and process the plaques as seen in human AD brain tissues1. Because LILRB -expressing microglia have an enhanced DAM profile upon LILRB3- activation, localization of the protective DAM around Ap plaques are enhanced, thus limiting potential neurotoxicity, especially in the presence of LILRB2 blockade. Through immunofluorescence and Imaging Mass Cytometry, the LILRB-activated microglial presence surrounding Ap plaques can be quantified and characterized.

[0733] Evaluating the benefit of LILRB3 / B2 genes on diminishing AD-related pathology. First, the concordance between the FB2 / 3 mouse and human AD brains is evaluated to justify the utility of the murine model. Leveraging a large cohort of postmortem brains from AD and control subjects in the Mount Sinai Brain Bank (MSBB), the study generated RNA-seq, whole-genome sequencing, and proteomics data in 4 brain regions from over 2,000 brain tissues1>2. It was found that microglia- specific differentially expressed genes (DEGs) in FB2 / 3 vs FAD mice were enriched for upregulated genes in advanced AD, as defined by cognitive dementia rating (CDR), CERAD, and plaque (Figure 17A) via gene set enrichment analysis (GSEA)13. The shared genes between human AD brains and the transgenic mice included Clqa, Clqb, Clqc, Ctss, Lat2, Trem2, and Tyrobp. Using a human AD single nucleus RNA-seq (snRNA-seq) dataset by Mathys et al.14(80,660 cells from 24 AD individuals), the study also observed co-localization of LILRB3 and TREM2, a key DAM marker, in a microglial subpopulation (Figure 17B), suggesting the relevance of LILRB3 modulation in DAM. These preliminary results support the mechanism of LILRB- activated microglia in the transgene model.

[0734] Evaluating spatio-temporal molecular changes modulated by active LILRB signaling in microglia. LILRB 3 -specific agonist antibodies (aLILRB3, compared against control Ig) are used to activate LILRB signaling and quantify changes in the DAM signature using scRNAseq, thus identifying the specific pathways altered through LILRB3 signaling in the murine model. To this end, cell-level dynamics such as molecular kinetics of spliced and unspliced reads (i.e. RNA velocity)3,4and pseudo-temporal trajectories (using slingshot software)5are inferred. The correlation between molecular changes associated with aLILRB3 treatment (i.e. aLILRB3 microglial DEGs) and the pseudotime along the trajectory is evaluated by TradeSeq8, a generalized additive model (GAM) framework to assess non-linear associations with pseudotime. Using the workflow, single-cell transcriptome of murine microglia from FAD and FB2 / 3 mice are analyzed to understand LILRB -mediated microglial pseudo-temporal dynamics in AD pathogenesis. The analysis revealed 8 distinct clusters that over-expressed predefined signature markers3-5(Figure 17C) including homeostatic microglia (HM1 / 2: P2ryl2, P2ryl3, Serinc3, Cx3crl, Tgfbl, Tmemll9, Txnip, Glut), disease-associated microglia stage 1 (DAM1 : Apoe, Tyrobp, B2m, Ctsb, Timp2, H2-D1, Fthl, LyZ2, Cstb, Ctsd) and 2 (DAM2: Ank, Sppl, Axl, Csfl, Cst7, Cd9, Itgax, Lpl, Ctsz, Ctsl, Ccl6, Cadml, Cd63, Ctsa, Serpine2, Ctsz, Cd52, Hifla). By Monte-Carlo permutation test6, the FB2 / 3 model was enriched for DAM1 and DAM2 cells (Figure 17D), compared to the FAD mice. The inferred cell trajectory also showed subsequent emergence of DAM1 / 2 cells following the homeostatic cells, and these correlated with gradual increases in key DAM markers such as Apoe and Trem2 (Figure 17E).

[0735] Using lOx Visium platform, spatially resolved transcriptome (SRT) data from the murine model, a well established technique9 10, is concomitantly analyzed to identify spatial localization patterns of LILRB3-driven microglia population. Using Giotto workflow, an R package dedicated to SRT analyses, coherent spatial expression patterns of key cell type markers (excitatory neuron: Nrgn, inhibitory neuron: Gadl, microglia: Tyrobp, astrocyte: Gfap, Oligodendrocyte: Plpl, oligodendrocyte progenitor: Vcan, endothelial: Fltl), DAM signatures, and P-amyloid precursor protein (App) are examined to identify cell populations co-localizing with Ap plagues. Furthermore, changes in cellular neighborhoods in aLILRB3-treated mice are evaluated by comparing cell type compositions in adjacent cells / spots using Hidden Markov Random Field (HMRF) model implemented in Giotto7. The section-wise transcriptomes are integrated using Harmony10to identify robust spatial clusters across different sections. Using this workflow, the study analyzed the spatial transcriptome of the FAD and FB2 / 3 transgene models (Figure 17F), and identified distinct localized expression patterns of cortex / hippocampus, dentate gyrus with granule neurons, mesencephalon, oligodendrocytes and vasculatures in both models. Interestingly, the FB2 / 3 section shows active granule neuron localization, compared to the loss of the granule neuronal patterns in the FAD model, suggestive of impaired firing properties in dentate gyrus8. In tandem with the activation of DAM signatures in the FB2 / 3 model single-cell transcriptome, these further support the protective role of LILRB in AD pathogenesis.

[0736] Evaluating the ability of LILRB2 / LILRB3 therapy to ameliorate AD-related pathology, behavioral, and memory deficits. The 5XFAD murine model of AD exemplifies the role of excess Ap production in driving changes in neuroimmunology with simultaneous neurodegeneration. The data shows a decreased A burden in FB2 / 3 mice compared to regular 5XFAD mice. It is therefore beneficial to characterize the ability of the LILRB2 / LILRB3 transgene to ameliorate AD-related behavioral and memory deficits. WT, 5XFAD, and FB2 / 3 mice are treated with aLILRB3 agonist therapy then quantify the rescuing effects initiated by LILRB signaling in a double-blind manner. Importantly, mice from both sexes are included and 8-10 mice per group and repeat experiments at least three times with mice of the same age.

[0737] Evaluating the therapeutic capacity of aLILRB3 agonist in the absence or presence of LILRB2 blockade therapy for reducing the Aft pathology in AD mice. Ap-dependent deficits in hippocampal learning and memory correlate well with alterations in calcium- and synaptic activity -related proteins in granule cells of the dentate gyrus (DG), a brain region critically involved in learning and memory. These functionally relevant molecular alterations include reductions in the calcium-binding protein calbindin-D28K; the immediate-early gene products Arc and Fos. The levels of these molecules are affected and have been demonstrated in multiple different familial AD-mutant hAPP (hAPPFAD)64'66.

[0738] The study further evaluates the therapeutic capacity of the LILRB 3 -agonist w / wo LILRB2 antagonist blockade of Ap in enhancing the neuroprotective role of DAM and MDM to reduce the Ap burden and rescue the memory response and cognitive dysfunction. This therapy may augment the functional capacity of myeloid cells to clear A (quantified by AP-specific immunostaining24) from the CNS parenchyma as well as reduce neurotoxicity, as measured by N-terminal APP immunohistochemistry23. The synaptic integrity and markers of neuronal function in the hippocampus, a brain region critical for memory and particularly vulnerable in AD can also be assessed. These genes / markers are known to correspond with spatial memory64-66. Quantifying the molecular markers of neuronal functionality is complementary to the neuroimmunology assessment by scRNAseq, in addition to the Ap pathology analysis and behavioral studies, creating a complete picture of the therapeutic capacity of LILRBs within the 5XFAD mouse model. Such markers include immunohistochemistry of cFos, calbindin, delta-FosB, which correlate strongly with memory in AD mice and are altered in several different transgenic mouse lines including Line J20, Tg2576, and APP / PS1 mice64; also synaptophysin as a marker of synaptic density, which is reduced in many models of AD but has improved upon therapeutic treatments. The immunostaining techniques and quantitation of neuronal functionality are well established. Staining and quantifications is performed in a double-blind manner. For these studies, the mice are stratified into 6 different groups - age-matched WT mice, 5XFAD, FB2 / 3 mice, + AP2- conjugated control Ig treatment, AP2-conjugated aLILRB3 agonist antibody. aLILRB3 agonist significantly reduces inflammation and enhances Ap phagocytosis which is superior to the aLILRB2 antagonist (Figures 13A-13E). Treatments is administered weekly in mice of both sexes at 4-5 months vs. 7-8 months of age at the early stage of FAD mice. At least 8-10 mice per group are followed for longitudinal analysis, and are sacrificed at respective time points 1-2 months post initial treatment. One hemibrain is fixed in 4% paraformaldehyde for immunohistochemistry and pathology, and the other hemibrain is snap-frozen for biochemistry including western blot analysis of the hippocampus. Some mice are infused in methoxy X-4 to stain the Ap to measure the reduction and improved degradation of Ap. The data show that FB2 / 3 mice have diminished deposition of Ap assemblies compared to regular 5XFAD mice (Figures 5A-5I), and can be further enhanced by aLILRB3 therapy, compared to 5XFAD mice and the control Ig-treated group. In addition, the reduction of Ap may be accompanied by the restoration of cFos, calbindin, and synaptophysin expression in the hippocampus. These molecular alterations are critical for synaptic plasticity and memory (cFos, calbindin) and serve as a marker of synaptic density and integrity (synaptophysin), which can be concomitant with the reduction of Ap and improvement in memory. FB2 / 3 mice with aLILRB3 therapy may also have less severe symptoms compared to regular 5XFAD mice or FB2 / 3 mice with control Ig treatment. Alternatively, the combination of aLILRB3 with antagonist LILRB2, which may inhibit the neuron toxicity, can be tested and compared using FB2 / 3 mice vs. FB1 / 2 / 3 / 4 to establish a dose-response comparison of the combined effects of therapy and neuropathology. The study the characterizes any potential for protection of neurotoxicity based on a dose-response relationship with antibody therapy.

[0739] Evaluating the ability of LILRB3 / 2 modulation to ameliorate behavioral and memory deficit characteristics of 5XFAD mice. Genetic insertion of LILRB2 / 3 shows a dramatic decrease in myeloid cell response to inflammatory stimuli in murine models (Figures 1F-1H), therefore, this extends to the rescuing of associated cognitive deficits in 5XFAD mice (Figures 5A-5I). Recent evidence suggests that evaluating behavioral phenotypes of 5XFAD mice can be confounded by their inherent hyperactivity, especially in tasks that are reliant on motor activity (10.3389 / fnagi.2021.713726). Behavioral phenotypes are therefore quantified in WT, 5XFAD, and FB2 / 3 mice using spontaneous alternation tasks, such as the Y-maze. At least 8 mice per group are used to achieve statistical significance, and Anova test is used to measure the average time spent exploring each arm. Three repeated experiments are performed and compared. The study systematically evaluates the age-dependent effects of the amyloid burden in 5XFAD mice and FB2 / 3 mice by analyzing the mice at 4, 6, 8, and 10 months of age in a double-blind manner. The study also evaluates the potential added benefit for AP2 conjugated aLILRB3 agonist w / wo anti LILRB2 antagonist antibody or antiLILRB2 antagonist alone of treatment, compared to IgG control. TREM2 expression on myeloid cells may be responsible for diminishing the plaque load within the CNS21,24and increased gene dosage of TREM2 ameliorates AD-associated deficits in 5XFAD mice25, therefore the study determines whether anti LILRB -dependent increases in TREM2 expression can serve as a novel therapeutic regulator for neuroprotection. To validate the importance of the LILRB-TREM2 signaling axis, FB2 / 3 mice are crossed with TREM2 KO mice or use nanoparticles to deliver aTREM2 siRNA59. Additionally, the respective roles of microglia and infiltrating peripheral myeloid cells are differentiated using aCCR2 blockade58or nanoparticle delivery of CCR2 siRNA.

[0740] FB2 / 3 mice show an increased proportion of neuroprotective microglia, in addition to the enhanced expression of neuroprotective transcripts, associated with decreased A0 accumulation in their brains. FB2 / 3 also shows mitigated cognitive decline, compared to 5XFAD age-matched controls. Likewise, FB2 / 3 mice treated with aLILRB3 agonist therapy have additional rescue of cognitive deficits, with an undetectable effect in regular 5XFAD mice lacking LILRB3 expression. Since aLILRB2 antagonist may prevent neuron toxicity, it may lead to a synergistic effect to improve outcome when used in combination with aLILRB3 agonist. The rescuing capacity of LILRB signaling is followed throughout the course of the disease and quantify behavioral phenotypes at 4, 6, 8, and 10 months of age. Alternatively, spatial memory can be evaluated by assessing behavioral differences in the Morris water maze64and the object location memory task68,69, which are well established techniques.

[0741] Superior beta-amyloid uptake by micropglia in F2 / B3 mice when compared to those from FAD mice. Since the FB2 / B3 mice exhibited significantly reduced plague load and neurofilaman damage, the study futher compared the beta-amyloid uptake by microglia from the FB2 / B3 vs. FAD mice. The mice were intravenously injected with Methoxy-X04 dye to label beta-amyloid in vivo. Four hours later, the mice were sacrified, microgia were isolatedand beta-amyloid uptake by microglia was assessed flow cytometry. The results support that microglia of the FB2 / B3 mice can take up signiciantly higher amounts of beta- amyloid than those of FAD mice (Figure 18).

[0742] Anti-LILRB2 antibody can inhibit the beta amyloid mediated reporter cell activation. The study futher evaluated the effect of LILRB2 antagonist antibodies on the inhibition of betaamyloid polymer mediated activation of the 2B4 reporter cells that express the LILRB2 extracellular domain with DAP-12-associated NFdsdAT GFP reporter (Figure 19). The results indicate that only cetain LILRB2 antagonist antibody clones can block the beta-amyloid mediated activation of the LILRB2+ 2B4 reporter cells.

[0743] Idenitification of agonist anti-LILRB3 antibodies. The study also identified agonist anti- LILRB3 antibodies that can inhibit the LPS mediated TNFalpha secretion and OKT3 mediated T cell proliferation in multiple donors (Figure 20). These agonist antobody can promote the M2 macrophage differentiation and reduce the inflammation cytokine secretion.

[0744] The synergistic effect of anti-LILRB3 agonist and LILRB2 antagonist on the inhibition of beta amyloid mediated neuron toxicity. Since anti-LILRB3 and LILRB2 enhanced macrophage phagocytosis of Ab, the experiment further tested the combination effect in protection against neuron toxicity mediated by Ab in the absence or presence of macrophages. Control Ig and anti-LILRB3 failed to protect neuron cells from toxicity mediated by Ab polymers. Anti- LILRB2 (2D6) alone in the presence of macrophage mediated a significant protection. Importantly the combination of anti-LILRB3 agonist 3B1 and anti-LILRB2 antagonist 2D6 in the presence of macrophages further enhanced the protective effect (Figures 21A-21B). The result supports the synergistic effect of neuron protection mediate4d by anti-LILRB2, 2D6 and anti- LILRB 3, 3B1 in the presence of macrophages.

[0745] Anti-LILRB2 / B3 combination reduces the A / l accumulation and enhances the microglia infiltration in the FB2 / 3 mice. The study further evaluated the BBB penetration of transferrin conjugated vs. normal antibody with OTT 565 dye conjugated antibody in a large cohort of FB2 / 3 mice within 48 hours after i.v. injection. The results indicated that transferrin conjugated antibody can more effectively penetrate (7-fold increase) into the brain than control antibodies (Fig. 23A). The data indicate that combination of anti-LILRB3 and B2 were more efficient in reducing the Ap accumulation (Fig. 23B, Fig. 23C) and increasing the penetration or maturation of microglia in the Cortex (Fig. 23D, Fig. 23E). It is believed that LILRB3 signaling can regulate the immune activation seen in AD. Specifically, LILRB3 signaling can diminish activation of the NLRP3 inflammasome and promote TREM2 expression to enhance the neuroprotective function of microglia and MDM. To enhance LILRB3 signaling, the study developed antibodies targeting individual LILRB receptors that activate the intracellular antiinflammatory networks through TRAM2 dependent and independent pathways, which prevent AD progression in both early and late stages of AD patients. Additional experiments may be conducted to evaluate the combined effect with / without LILRB2 antagonist in the blockade of A mediated neuron toxicity.

[0746] Example 3. antiLILRB2 blocks ANGPTL5 and MHC tetramer binding

[0747] Experiments were conducted that showed that the presently described antibodies, but not commercial LILRB2 antibodies, are antagonists that can block HLA tetramer binding (Fig. 22A, lower panel) and ANGPTL-5 interaction (Fig. 22A, upper panel). These antibodies are specific, since they only bind to LILRB 2, but not other LILRB members. The identified antibodies that recognize LILRB2 have been screened. These observations suggest that these LILRB2-binding antagonistic antibodies are sufficient to enhance LPS- dependent classical activation in monocytes and can interfere its ligands binding. Immunostimulatory LILRB2 antibodies are LILRB2-MHCI receptor-ligand antagonists. Using fluorescently labeled human MHC-I tetramers, the study determined whether these tetramers could specifically bind LILRB2- expressing cells using flow cytometry. Dose-response experiments demonstrated that human HLA-A / B tetramer specifically bound LILRB2-transduced cells in a dose-dependent manner while no binding was detected on non-transduced parental cells (Fig. 22B). The study subsequently examined whether LILRB2 antibody clones A and B could block the LILRB2- MHC-I interaction. It was found that treatment of LILRB2-expressing THP-1 cells with antibody A or B was sufficient to block tetramer binding in a dose-dependent manner (Fig. 22C). These clones were significantly able to outcompete tetramer binding as determined by the IC50 at the 10'9to IO10M range. This data strongly suggest that the presently disclosed antibodies are unique to LILRB2 in that they can block LILRB2 MHC-I and ANGPTL-2 / 5 binding. Example 4.

[0748] The present example describes the generation and evaluation of the kinetics and biodistribution of a bispecific LILRB3 / B2 transferrin fusion protein (HMR 301) with intravenous (IV.), or intranasal (IN.) and / or biodegradable nanofludic drug-eluting seed (NDES) nanodevices for brain delivery. The collected results shows that transferrin fusion antibody delivery can achieve certain success with i.v. injection of transferrin-conjugated antibodies.

[0749] Bispecific antibody (anti-LILRB2 and anti-LILRB3) are designed with transferrin fusion proteins (named HMR301) (Fig. 30A). IV. vs. IN. or biodegradable NDES delivery systems are compared through intracranial implantation procedures with slow release of HMR301 into the brain. It has been previously shown that IN. delivery of mRNA or drug delivery through IN. can be favorable for the drug delivery to the brain. A nanofluidic drug-eluting seed (bNDES) is a small implant that uses a biodegradable nanoporous implant composed of FDA-approved therapeutic materials, Poly(e-caprolactone) (PCL) and Poly (D, L-lactic-co-glycolic acid) (PLGA) (Fig. 30B). Drugs are loaded within the central hollow core, where sustained and controlled release is achieved through diffusion across the nanoporous fibers. The current size of NDES is 1.3 mm x4 mm; however, it is also fabricate the device to a size of 1.3 mm x2 mm. In the present example, the antibody amount can be up to 0.8 mg per / device and last for a minimum of 60 days.

[0750] Evaluating the functional activity of HMR301 in vitro in the presence of neuron and macrophage co-culture. The neuron-killing activity of HMR301 is compared with transferrin conjugated control Ig, anti-LILRB2 or anti-LILRB3 or combination in the cocultured neuron, macrophage w / wo antibody (Ab) to follow the neuron cell proliferation, survival, and damage. Various doses of antibody (e.g., 0, 0.25, 0.5, 1, 2, 4 mg / ml) are compared and titrated. The combined effect of increased and prolonged neuron survival and inhibition of neuron death is analyzed as shown in Figs. 26A-26H and 27A-27D . The study has designed the bispecifc antibody and the single chain of anti-LILRB3 and anti-LILRB2 for CAR-T therapy with good functional activity. Alternatively, it is possible to compare the bispecific vs. the single chain antibody with tandem repeat on the inhibition efficacy of Ab neuron toxicity and promotion of the macrophage-mediated phagocytosis. Without wishing to be bound by theory, it is believed that the single reagent HMR301 (the bispecific antibody and transferrin fusion protein) has significantly stronger macrophage-mediated phagocytosis and inhibition of Ab-mediated neuron killing, even at a lower dose of antibody and with the highest neuron cell survival due to the close vicinity of antibody interaction. Assessing the persistence and kinetics of HMR301-mediated sustained IV. vs. IN. or NDES delivery kinetics on anti-neuron inflammation and biodistribution. To investigate in vivo biodistribution of HMR301, this study uses fluorescence imaging by in vivo imaging system (IVIS) and histology to track spatiotemporal biodistribution of AlexaFluor 700 (AF700)- tagged HMR301 as compared to the original AF700-tagged control Ig, anti-LILRB3 and anti- LILRB2 w / wo transferrin conjugation in FB2 / 3 vs. FAD or B2 / B3 control male or female mice at 6 months of age. In a previous nanoparticle biodistribution study, it was shown that nanoparticle (NDES) injected in the tumor site can slow release-AF700-CD40 recombinant proteins with sustained localization in the tumor for 7 days as opposed to a declining signal detected in tumors of intraperitoneal (IP, 100 mg) and intratumorally (IT, 50 mg) injected groups throughout the 7-day study (Fig. 31A, 31B). For fluorescence quantification, the highest signal in each group was calculated as 100% and each group was evaluated separately, as described. IP injected mice demonstrated drug localization in the liver at study termination on day 7 (Fig. 31C). Therefore, biodistribution of HMR301 mediated biodistribution can be determined and compared with the original antibody as a benchmark.

[0751] Experimental design. 6 months old LILRB2 / B3 transgenic C57BL and FB2 / 3 female or male mice (n=6) are randomized into six groups with a total of 800 ug dose of HMR301 delivered via IV., or IN. or NDES nanoparticle through the intraparenchymal injection vs. anti- LILRB3 and anti-LILRB2 AF700-tagged fusion protein w / wo transferrin for three times (0, 50, 100, 200 mg / mouse, each dose at one week interval, a total of four injections, NDES loaded only one time for a single injection at antibody dose of 200, 400, 800 mg). The antibody is conjugated to AF700 via N-hydroxysuccinimide ester using established protocols. The study has validated that conjugation does not affect Ab binding ability. Fluorescent tagging allows for the ability to monitor antibody localization in the brain and pass through BBB in a dynamic and noninvasive manner using IVIS Spectrum thrice a week. Detection of fluorophore-co-expressed Ab is not affected by internalization or recycling.

[0752] Drug distribution assessment. IVIS Spectrum live animal imaging is performed at baseline pre-injection, as well as at 5 mins, 1, 3, 7, 10, 13, 16, 20 and 30-days post-injection. The fluorescent signal of AF700-labeled Ab by nanoparticle delivery vs. IV. or IN. injection with the same amount of total dose is measured at an excitation wavelength of 675-1150 nm and an emission of 280-720 nm. The fluorescence intensity measured as radiance photons (photons / second / cm2 / steradian) is quantified at the brain region of interest (ROI) and other organs at each time point with Live Image software and used as a surrogate for protein concentration. Background / autofluorescence from control mice is then subtracted. Fluorescence data for each mouse is graphically plotted as a surrogate marker of antibody localization. Ab and microglia cell infiltration is measured daily for 14 days, at which point animals are euthanized, or earlier if humane endpoints are reached (IACUC, ACURO). On 1, 3, 7, 10, 14, 20, and 30 days, five mice per group are euthanized, blood drawn at each time point for Olink measurement on inflammation cytokines, and tissues collected for analysis and weighed. At the end time point, the brain, spleen, lymph nodes, lung, liver, heart, bladder, ovarian, kidney and bone are harvested and imaged for protein localization via IVIS. Ex vivo IVIS imaging of organs allow the comparison of fluorescent signal intensity across treatment groups at different time points.

[0753] Sustained transferrin conjugated antibody through IN. or IV. or NDES delivery may achieve localization with high concentrations in the brain and may last longer with NDES with minimal dispersion to other organs throughout the study. If higher or lower release rates are desired, the different doses of HMR301 can be adjusted by changing the amount of antibody or pore size. Without wishing to be bound by theory, it is believed that HMR301 has a higher and relatively longer-lasting circulation in the brain through IN. and NDES delivery in vivo than IV. due to the bivalent and local effects. The reduced Ab and increased microglia infiltration and reduced inflammatory cytokines in the treated mice blood can correlate to the amount of antibody distribution.

[0754] Evaluating the ability of LILRB2 / LILRB3 therapy to ameliorate AD-related neuron pathology, behavioral, and memory deficits. Collected data show a decreased Ap burden in FB2 / 3 mice compared to regular 5XFAD mice Figs. 26A-26H, The anti-LILRB3 and B2 combination can significantly reduce the Ab and increase the microglia infiltration (Fig. 29A- 29E). Therefore, the present example sought to characterize the ability of the LILRB2 / LILRB3 transgene to ameliorate AD-related behavioral and memory deficits. WT, 5XFAD, LILRB2 / 3, and FB2 / 3 mice are treated with HMR301, aLILRB3-Tf alone or aLILRB2-Tf alone therapy vs. control Ig-Tf and quantify the rescuing effects initiated by LILRB3 or B2 or combination signaling in a double-blind manner.

[0755] Evaluating the therapeutic capacity of aLILRB3 agonist in the absence or presence of LILRB2 blockade therapy for reducing A neuron pathology in AD mice. A -dependent deficits in hippocampal learning and memory correspond well with alterations in calcium- binding and synaptic activity-related proteins in granule cells of the dentate gyrus, a brain region critically involved in learning and memory. These functionally relevant molecular alterations include reductions in the calcium-binding protein calbindin-D28K; and the immediate-early gene products Arc and Fos. The levels of these molecules are affected and have been demonstrated in multiple different familial AD-mutant hAPP. The therapeutic capacity of the LILRB 3 -agonist with or without LILRB2 blockade of Ap may be evaluated in enhancing the neuroprotective role of DAM and MDM to reduce the A burden and rescue the memory response and cognitive dysfunction. Without wishing to be bound by theory, it is believed that this therapy can augment the functional capacity of myeloid cells to clear Ap (quantified by Ap-specific immunostaining, 4G8 or 6E10, Thioflavin-S (ThS)) from the CNS parenchyma as well as reduce neurotoxicity, as measured by immunohistochemistry of dystrophic neurites (as in Figs. 26A-26H). For these studies, the mice are stratified into 3 different age- and sex-matched groups: WT, 5XFAD, LILRB2 / 3, and FB2 / 3, treated with transferrin-conjugated control Ig or aLILRB3 agonist antibody or aLILRB2 or HMR301 (or a combination of anti-LILRB3 and B2). As has been shown, the aLILRB3 agonist significantly reduces inflammation and enhances Ap phagocytosis which is superior to the aLILRB2 antagonist (Fig. 28A-28D). Treatments are administered weekly through IN. at a dose of 100 mg / mouse every week, a total of 800 mg / mouse, or implant NEDS at a maximal dose of 800 mg / device into intraparenchymal at a single injection in mice of both sexes at three stages of disease progression and pathology: 3-4 months (early stage) vs. 5-6 months vs. 7-8 months of age. 10 mice or more per group are followed for each age group, and they are sacrificed 2 months after initial treatment. One hemibrain is fixed in 4% paraformaldehyde for immunohistochemistry and pathology, and the other is snap-frozen for biochemistry including western blot analysis of the hippocampus. Some mice are infused with methoxy X-4 to stain A to assess its reduction and improved degradation. The microglia are stained with Iba-1 (Wako, Cat 019-19741) with the secondary Alexa 568 (Fisher, cat Al 136).

[0756] Without wishing to be bound by theory, it is believed that this reduction of neuron pathological effect can be further enhanced by HMR301 and aLILRB3-Tf, and to a lower degree, by aLILRB2-Tf therapy. Additionally, the reduction of Ap may be accompanied by restoration of cFos, calbindin, and synaptophysin expression in the hippocampus. These molecular alterations are significant for synaptic plasticity and memory (cFos, calbindin) and serve as a marker of synaptic density and integrity (synaptophysin), which may be concomitant with the reduction of Ap and improvement in memory. FB2 / 3 mice with HMR301 and aLILRB3 therapy may have less severe symptoms compared to regular 5XFAD mice or FB2 / 3 mice with control Ig treatment. Alternatively, the combination of aLILRB3 with antagonist LILRB2 (HMR301) may inhibit neurotoxicity, and this can be compared using FB2 / 3 mice vs. APP / PS-1 / LILRB2 / B3 mice to establish a dose-response comparison of the combined effects of therapy and neuropathology. It is further possible to characterize any potential for protection of neurotoxicity based on a dose-response relationship with antibody therapy and route of delivery.

[0757] Evaluating the efficacy of HMR301 (or a combination of anti-LILRB3 / B2 antibodies conjugated with transferrin) in ameliorating behavioral and memory deficit characteristics of FB2 / 3 mice. Genetic insertion of LILRB2 / 3 shows a dramatic decrease in myeloid cell response to inflammatory stimuli in murine models (Figs. 24A-24C); therefore, it was believed that this could extend to the rescuing of associated cognitive and behavioral deficits in 5XFAD mice (Fig. 27A-27D). Indeed, the collected results suggest FB2 / 3 mice have improved spatial memory in the Barnes maze task compared to 5XFAD mice (Fig. 32). To evaluate the degree of hyperactivity of FB2 / B3 mice and whether it is reduced by HMR301 (or a combination of aLILRB3-TI' and aLILRB2-Tf), aLILRB3-Tf, or aLILRB2-Tf treatment, the study also assesss mice in the open field. This allows the ability to quantify the timeframe in which each mouse habituates to the open field arena, a measure that is also influenced by hippocampal function. Prior to the assessment of memory and hyperactivity, each mouse undergos a SHIRPA (SmithKline, Harwell, Imperial College, Royal Hospital Phenotypic Assessment) exam. This exam is a battery of rapid tests that assess basic neurological and sensorimotor function, which are sensitive to detecting phenotypes in several neurogenerative disease models. The SHIRPA exam complements cognitive testing and enables the detection of whether treatment strategies cause even subtle changes in behavior. Behavioral phenotypes are classified in 8 groups: WT, 5XFAD, LILRB2 / 3, and FB2 / 3 mice, treated with transferrin-conjugated control Ig or HMR301 (or a combination of aLILRB3-Tf and aLILRB2-Tf) for two months in a double-blind manner including both male and female mice for the study. At least 12 mice per sex are used for each group. The study assesses one group of mice at 3-4 months of age and another group at 5-6 months of age to examine whether treatment is effective at both early and later stages of disease progression and to evaluate age-dependent effects of the amyloid burden. The study also evaluates the potential added benefit of anti-LILRB2 antagonist antibody and the effect of anti- LILRB2 antagonist alone without LILRB3 -agonist, compared to IgG control in FB2 / B3 mice. TREM2 expression on myeloid cells may be responsible for diminishing the plaque load within the CNS, and increased gene dosage of TREM2 ameliorates AD-associated deficits in 5XFAD mice; therefore, it is possible to determine whether anti-LILRB-dependent increases in TREM2 expression can serve as a novel therapeutic regulator for neuroprotection. The TREM2 (R47H) mutant Knock-in (Jackson lab# 037497) or KO mice (Jackson Lab, # 027197) and cross-bred with B2 / B3 transgenic mice are further tested for the therapeutic effect with HMR301 vs. control Ig. If HMR301 (or a combination of anti-LILRB3 and anti-LILRB2) can still improve the AD symptoms in both pathology and behavior assessments, it may support that HMR301 mediated TREM-2 independent pathway plays an important role in the functional activity of LILRBs. Without wishing to be bound by theory, it is believed that FB2 / 3 mice treated with HMR301 therapy have an additional rescue of cognitive / behavioral deficits, with an undetectable effect in regular 5XFAD mice lacking LILRB3 expression. Since aLILRB2 antagonist may prevent neurotoxicity, it may lead to a synergistic effect to improve outcome when used in combination with aLILRB3 agonist. The rescuing capacity of LILRB signaling is followed throughout the course of the disease and quantify behavioral phenotypes at 4, 6, 8, and 10 months of age.

[0758] Determining the mechanism / pathway by which HMR301 vs. anti-LILRB3 activation enhances the DAM state and exerts its neuroprotective function.

[0759] Next, the study sought to evaluate the effect of HMR301 on the anti-inflammatory network initiated by the FB2 / B3 vs. 5X FAD mice using RNAseq at single-cell resolution, proteomics and correlate the signaling pathways with available bioinformatics and protein biomarkers from human AD specimens. The study compares the roles of microglia and peripheral immune components, such as MDMs, to previous studies within the 5XFAD model. Additionally, the anti-inflammatory network initiated by the HMR301 in FB2 / 3 mice is evaluated and this disease progression is correlated with human AD information.

[0760] Determining the synergistic pathways affected by HMR301 in microglia and MDM recruitment, maturation and changes in inflammation profile. Despite being a small population of cells, DAM and MDM are crucial in curtailing the progression of AP-mediated neurological deficits. Central to their development is TREM2, which initiates the expression of a network of genes that are targeted toward processing A , such as Tyrobp, Lilrb4a, Lpl, and Axl (Figs. 25A-25E). Collected data show many DAM genes are enhanced in the presence of human Lilrb2 / 3 trans genes in 5XFAD mice by RT-qPCR and single cell analysis (Figs. 25A-25E, 34A- 34J); therefore, the study takes advantage of scRNAseq to further characterize the immune landscape change w / wo antibody treatment. After enriching for both CD45+and CD45" cells from the CNS, scRNAseq allow the capture of the individual immune components involved in the Ap model of AD. scRNAseq study showed an enhancement of the DAM phenotype in FB2 / 3 vs. 5XFAD mice (Figs. 34A-34J). Thus, the study sought further evaluation of the signaling network initiated by HMR301 therapy within the gene signatures of DAMs and infiltrating MDMs. WT, 5XFAD, and FB2 / 3 mice are treated with a single injection of the HMR 301 or control Ig (200 ug / mouse) when the mice reach 4, 6 and 8 months of age. Following treatment, scRNAseq is performed on CD45+cells isolated from the brains of the respective mice to characterize the immune landscape affected by HMR301 therapy. The delivery method with the highest antibody distribution in the brain is used, e.g., IN. or NDES intraparenchymal injection. The study has demonstrated that agonist anti-LILRB3 antibodies significantly promote maturation and expansion toward CD14+ / CD16+ / CD163+positive cells, i.e. M2 like phenotype (Fig. 33A), and significantly reduce LPS-mediated inflammatory cytokines in the circulation of humanized NSG-SGM3 mice, as assessed by serum TNFa and IL-6 levels (Fig. 33B) and suppress the production of multiple inllammatory cytokines.

[0761] The phenotype of circulating immune cells in brain tissues is determined using CyTOF from the immune monitoring core. The study also measures HMR301 -induced microglia, macrophages and other immune cell profile and their functional markers. The kinetics of CDllb+CD45+ leukocyte recruitment are identified, e.g., microglia, macrophages derived from circulating monocytes, neutrophils after 1, 3, 5 and 7 days in AD mice post-HMR301, anti- LILRB3-Tf or anti-LILRB2-Tf treatment or with control Ig. Mice are perfused with heparinized saline, and the ipsilateral brain cortex is harvested. The tissue is minced with a razor blade and incubated with a digestion solution for 20 min at 37°C. A suspension is set at room temperature for 15 min, passed through a 70 m cell strainer, and rinsed with PBS. Cells are incubated with the following primary antibodies: GDI lb, CD45, and F4 / 80. Alternatively, brain dissociates were isolated, and flow cytometry was used to gate for microglia (CD1 lb+, CD45lowcells), monocytes (CDl lb+, CD45hlgh, F4 / 80+ cells), and G-MDSC, M-MDSC or neutrophils (CDl lb+, CD45hlgh, Ly6G+ / Ly6C cells), T cell subsets e.g. Thl, Th2, Treg, Thl7 and B cells to measure their recruitment to the brain. The study investigates if HMR301 or anti-LILRB3 can affect the activation, expansion of microglia cells and macrophages in the brain. Similar pro-inflammatory protein expression are analyzed in the brain by imaging mass cytometry (IMC) with cellular markers, and the major histocompatibility complex class II (MHC-II) expression of CDllb+microglia, Iba-1, CD68, TREMs, MS4A4A, MS4A6A and MS4A7A (membranespanning 4-domains subfamily A) is explored, which have significant associations with sTREM2 and Inflammasome NLRP3 activation, and CD 11b antibodies, NADPH oxidase (N0X2) neutrophil, monocytes, leukocytes, and macrophage infiltration using MPO, F4 / 80, Ly-6G, CD14, CD45, CD115, and CDl lb antibodies), neurons (NeuN), and astrogliosis (using GFAP, NG2 antibodies). Quantitative methods are applied using unbiased stereology analysis to determine the total positive cell counts in the lesioned cortical and hippocampus regions. The study measures the 48 inflammatory cytokines / chemokines by Olink, which only requires 1 ml of cerebrospinal fluid (CSF) or plasma according to the manufacturer’s instructions.

[0762] Without wishing to be bound by theory, it is believed that FB2 / 3 mice exhibit a distinct immune landscape in the CNS compartment compared to that of regular 5XFAD or WT mice, especially in the presence of HMR301 or agonist aLILRB3 antibodies. Agonist antibodies may further enhance the DAM phenotype and have the added benefit of reducing Ap pathology. MDMs and BMDMs may have anti-inflammatory capacity after treatment with HMR301 or aLILRB3 agonist antibody.

[0763] Correlating single-cell RNA sequencing information with in situ identification of active cells with imaging mass cytometry and spatial transcriptomics. The DAM transcriptional state is initiated by the A -mediated pathology seen in AD. Microglia assume this state to mitigate further AP-mediated neurotoxicity. Collected results show that FB2 / 3 mice exhibit upregulation of signature transcripts that resemble DAM1, DAM2 and AgM and microglial trajectory gene profile change, e.g., Trem2, Apoe, and DAM gene profile change in TREM2 dependent and independent and in Volcano plots (Figs. 34A-34J), implying that LILRB signaling may enhance these neuroprotective microglia. Gene expression level changes may be further evaluated through single-cell spatial transcriptomics and follow the localization of DAM and MDM populations. The study identifies changes in the gene signatures resulting from treating mice with control Ig, HMR301, aLILRB3 agonist, or aLILRB2 therapy.

[0764] Without wishing to be bound by theory, it is believed that aLILRB3 treated FB2 / 3 mice may exhibit an enhancement of the neuroprotective DAM population in addition to more immune suppressive like MDMs, culminating in the reduction of Ap plaque accumulation. As regular 5XFAD mice lack the LILRB3 receptor, this effect may be specific to FB2 / 3 mice treated with HMR301. To differentiate the respective contributions of resident microglia vs. peripheral myeloid cell infiltrate, the study also treats with aCCR2 blocking antibody (clone MC-21, Creative Biolab. Inc.) or nanoparticle delivery of CCR2 siRNA to block myeloid cell migration to the CNS. Alternatively, the adaptive transfer of microglia linage tracing (C57BL Tmeml l9-2A-EGFP knock-in mice, Jackson Lab 031823), microglia and MDM linage tracing (Cx3crlD2knock-in / knock-out mice express Dendra2 green / red photo switchable monomeric fluorescent protein in myeloid cells, Jackson Lab 037016) is used to confirm the migration vs. in situ maturation of myeloid cells that contributes the DAM1, DAM2 and AgM population.

[0765] Using 12 samples (4 wildtype, 4 FAD and 4 FB2 / 3 mice w / wo HMR301) at 11,000 cells per sample, there is an estimate of 94.7% probability of identifying true DEGs as statistically significant. As 4 mice are sequenced in each treatment group, there is sufficient power to perform the analysis and expect to identify robust disease- associated murine microglia subpopulations modulated by HMR301 treatment. The study further curates and analyzes scRNA-seq and snRNA-seq datasets from human brain tissues to determine cell type-specific signatures of human AD, which are compared with the signatures of HMR301 vs. control Ig treatment in mice. A more significant overlap between cell type-specific signatures of human AD and the signatures of improvement with HMR301 treatment in mice may be observed. For spatial transcriptomic scRNAseq, 16 sections (4 FAD, 4 FB2 / 3) w / wo HMR301 treatment are generated to complement scRNA-seq. SRT will identify cell types whose spatial patterns and interactions with neighboring cells are affected by HMR301 treatment. Along with the major cell type markers, the study uses cell cluster markers from scRNA-seq to further characterize non- conventional cell populations such as heterogeneous microglial subpopulations. Alternatively, different ages of mice are used to include various stages of AP-mediated pathology. Without wishing to be bound by theory, it is believed that younger mice benefit most from HMR301 therapy and older mice may have improvements in DEGs. Additionally, the results are compared with aLILRB3 and aLILB2 alone to prevent the deleterious effects of Ap on neuron toxicity and characterize synergistic effects.

[0766] Determining the proteomic inflammation biomarkers from treated mice to correlate the disease stages from clinical AD patients. To facilitate future clinical translation, the study evaluates the benefit of HMR301 on diminishing AD-related pathology and correlate the improvement of HMR301 therapeutic effect with the stages of clinical AD patient’s progression and identify the potential protein profile changes as a biomarker for early diagnosis and evaluation of treatment outcome from different ages of treated AD mice.

[0767] Inflammatory biomarkers are measured in blood plasma, and cerebrospinal fluid (CSF) is collected from different time points at 0, 1-, 2-, 3-, and 7-days post HMR301, aLILRB3 and aLILB2 alone and control Ig treatments at dose 100 ug / mouse through IN. or NDES delivery route. The Olink assay is very sensitive, only requires 1-2 uL of fluid and can detect 48-45000 protein biomarkers. 2, 4, 6, 8 and 10 months old of WT, LILRB2 / B3 mice, 5X FAD and FB2 / 3 mice, 5 mice in each treatment group, is collected and measured for kinetics of patients’ blood plasma profile changes. This information is correlated with different stages (early, mid, and late) of AD patients’ blood samples.

[0768] In this regard, it is possible to determine whether the treatment can reduce the bloodstream or CSF cytokine expression in AD mice following HMR301 therapy. Treatment that can significantly revert the inflammatory stage in the FB2 / 3 mice but not significantly improve in the 5XFAD mice, illustrates supports of the efficacy of HMR301 on LILRB receptor effect. The WT and LILRB2 / B3 transgenic mice may show no effect due to low inflammation. The key cytokine profile changes from high throughput screening is further validated by ELISA. IL- 1 p, TNF-a, IFN-g, IL-6, and TGF-P expression is measured by ELISA. The potential protein biomarkers may be identified to correlate the age and the treatment outcome. Alternatively, different AD mice e.g. APP / PS-lx B2 / B3 mice are evaluated and compared.

[0769] The following patents, applications and publications as listed below and throughout this document are hereby incorporated by reference in their entirety herein.

[0770] - Zhao, Peng, et al. "LILRB2-mediated TREM2 signaling inhibition suppresses microglia functions." Molecular neurodegeneration 17.1 (2022): 44.

[0771] Smith, Levi M., et al. "Systematic and standardized comparison of reported amyloid- P receptors for sufficiency, affinity, and Alzheimer's disease relevance." Journal of Biological Chemistry 294.15 (2019): 6042-6053.

[0772] Ma, Xiaohong, et al. "Recognition of Ap oligomer by LilrB2 acceptor: a tetracoordinated zipper mechanism." Journal of Molecular Modeling 28.10 (2022):

[0773] Other advantages which are obvious, and which are inherent to the invention, will be evident to one skilled in the art. It will be understood that certain features and sub-combinations are of utility and may be employed without reference to other features and sub-combinations. This is contemplated by and is within the scope of the claims. Since many possible embodiments may be made of the invention without departing from the scope thereof, it is to be understood that all matter herein set forth or shown in the accompanying drawings is to be interpreted as illustrative and not in a limiting sense.

Claims

CLAIMSWhat is claimed is:

1. A recombinant antibody, wherein the antibody comprises: a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131 ; and / or a light chain variable region (VL) with a CDR1 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

2. The recombinant antibody of claim 1, wherein the recombinant antibody comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25.

3. The recombinant antibody of any one of claims 1-2, wherein the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

4. The recombinant antibody of any one of claims 1-3, wherein:VH CDR1 is SEQ ID NO: 27;VH CDR2 is SEQ ID NO: 28;VH CDR3 is SEQ ID NO: 29;VL CDR1 is SEQ ID NO: 30;VL CDR2 is SEQ ID NO: 31; andVL CDR3 is SEQ ID NO: 32; orVHCDR1 is SEQIDNO: 33;VH CDR2 is SEQ ID NO: 34;VHCDR3 is SEQIDNO: 35;VLCDR1 is SEQ ID NO: 36;VL CDR2 is SEQ ID NO: 37; andVL CDR3 is SEQ ID NO: 38; orVH CDR1 is SEQ ID NO: 39;VH CDR2 is SEQ ID NO: 40;VH CDR3 is SEQ ID NO: 41;VLCDR1 is SEQ ID NO: 42;VL CDR2 is SEQ ID NO: 43; andVL CDR3 is SEQ ID NO: 44; orVHCDR1 is SEQ ID NO: 45;VH CDR2 is SEQ ID NO: 46;VH CDR3 is SEQ ID NO: 47;VLCDR1 is SEQ ID NO: 48;VL CDR2 is SEQ ID NO: 49; andVL CDR3 is SEQ ID NO: 50; orVHCDR1 is SEQIDNO: 57;VHCDR2 is SEQIDNO: 58;VH CDR3 is SEQ ID NO: 59;VL CDR1 is SEQ ID NO: 60;VL CDR2 is SEQ ID NO: 61; andVL CDR3 is SEQ ID NO: 62; orVHCDR1 is SEQIDNO: 63;VH CDR2 is SEQ ID NO: 64;VHCDR3 is SEQIDNO: 65;VL CDR1 is SEQ ID NO: 66;VL CDR2 is SEQ ID NO: 67; andVL CDR3 is SEQ ID NO: 68; orVH CDR1 is SEQ ID NO: 69;VH CDR2 is SEQ ID NO: 70;VH CDR3 is SEQ ID NO: 71;VL CDR1 is SEQ ID NO: 72;VL CDR2 is SEQ ID NO: 73; and VL CDR3 is SEQ ID NO: 74; orVH CDR1 is SEQ ID NO: 75;VH CDR2 is SEQ ID NO: 76;VH CDR3 is SEQ ID NO: 77;VL CDR1 is SEQ ID NO: 78;VL CDR2 is SEQ ID NO: 79; and VL CDR3 is SEQ ID NO: 80; orVH CDR1 is SEQ ID NO: 81;VH CDR2 is SEQ ID NO: 82;VH CDR3 is SEQ ID NO: 83;VL CDR1 is SEQ ID NO: 84;VL CDR2 is SEQ ID NO: 85; and VL CDR3 is SEQ ID NO: 86; orVH CDR1 is SEQ ID NO: 87; VH CDR2 is SEQ ID NO: 88; VH CDR3 is SEQ ID NO: 89;VL CDR1 is SEQ ID NO: 90;VL CDR2 is SEQ ID NO: 91; and VL CDR3 is SEQ ID NO: 92; orVH CDR1 is SEQ ID NO: 93;VH CDR2 is SEQ ID NO: 94;VH CDR3 is SEQ ID NO: 95;VL CDR1 is SEQ ID NO: 96;VL CDR2 is SEQ ID NO: 97; andVL CDR3 is SEQ ID NO: 98; orVH CDR1 is SEQ ID NO: 99;VH CDR2 is SEQ ID NO: 100;VH CDR3 is SEQ ID NO: 101;VL CDR1 is SEQ ID NO: 102;VL CDR2 is SEQ ID NO: 103; andVL CDR3 is SEQ ID NO: 104; orVH CDR1 is SEQ ID NO: 105;VH CDR2 is SEQ ID NO: 106;VH CDR3 is SEQ ID NO: 107;VL CDR1 is SEQ ID NO: 108;VL CDR2 is SEQ ID NO: 109; andVL CDR3 is SEQ ID NO: 110; orVH CDR1 is SEQ ID NO: 111;VH CDR2 is SEQ ID NO: 112;VH CDR3 is SEQ ID NO: 113;VL CDR1 is SEQ ID NO: 114;VL CDR2 is SEQ ID NO: 115; andVL CDR3 is SEQ ID NO: 116; orVH CDR1 is SEQ ID NO: 117;VH CDR2 is SEQ ID NO: 118;VH CDR3 is SEQ ID NO: 119;VL CDR1 is SEQ ID NO: 120;VL CDR2 is SEQ ID NO: 121; andVL CDR3 is SEQ ID NO: 122; orVH CDR1 is SEQ ID NO: 123;VH CDR2 is SEQ ID NO: 124;VH CDR3 is SEQ ID NO: 125;VL CDR1 is SEQ ID NO: 126;VL CDR2 is SEQ ID NO: 127; and VL CDR3 is SEQ ID NO: 128; orVH CDR1 is SEQ ID NO: 129;VH CDR2 is SEQ ID NO: 130;VH CDR3 is SEQ ID NO: 131;VL CDR1 is SEQ ID NO: 132;VL CDR2 is SEQ ID NO: 133; and VL CDR3 is SEQ ID NO: 134.

5. The recombinant antibody of any one of claims 1-4, wherein the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 14.

6. The recombinant antibody of any one of claims 1-4, wherein the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 8.

7. The recombinant antibody of any one of claims 1-4, wherein the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 10.

8. The recombinant antibody of any one of claims 1-4, wherein the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60%identity to SEQ ID NO: 2.

9. The recombinant antibody of any one of claims 1-4, wherein the recombinant antibody comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 6.

10. The recombinant antibody of any one of claims 1-9, wherein the recombinant antibody comprises at least 1 amino acid substitution.

11. The recombinant antibody of any one of claims 1-10, further comprising a transferrin moiety conjugated by a peptide linker.

12. The recombinant antibody of claim 11 , wherein the transferrin moiety comprises at least 60% identity to SEQ ID NO: 180.

13. A bispecific antibody comprising:(i) a first binding arm having a first antigen-binding region that specifically binds to LILRB3, and(ii) a second binding arm having a second antigen-binding region that specifically binds to LILRB2.

14. The bispecific antibody, wherein the first antigen- binding region is an agonist of LILRB3 and the second antigen binding region is an antagonist of LILRB2.

15. The bispecific antibody of any one of claims 13-14, wherein the first antigen-binding region comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, or 93, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, or 94, and a CDR3 comprising asequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, or 95.

16. The bispecific antibody of any one of claims 13-15, wherein the second antigen-binding region comprises a heavy chain variable region (VH) with a CDR1 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity (e.g., at least 65% identity, at least 70% identity, at least 75% identity, at least 80% identity, at least 85% identity, at least 90% identity, at least 95% identity) to any one of SEQ ID NOS: 101, 107, 113, 119, 125, or 131.

17. The bispecific antibody of any one of claims 13-16, wherein the first antigen-binding region comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, or 13.

18. The bispecific antibody of any one of claims 13-17, wherein the second antigen-binding region comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 15, 17, 19, 21, 23, or 25.

19. The bispecific antibody of any one of claims 13-18, wherein the first antigen-binding region comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, or 14.

20. The bispecific antibody of any one of claims 13-19, wherein the second antigen-binding region comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 16, 18, 20, 22, 24, or 26.

21. The bispecific antibody of any one of claims 13-20, further comprising a transferrin moiety conjugated by a peptide linker.

22. The bispecific antibody of claim 21 , wherein the transferrin moiety comprises at least 60% identity to SEQ ID NO: 180.

23. The bispecific antibody of any one of claims 13-22, comprising: a first light chain comprising a sequence with at least 60% identity to SEQ ID NO: 163; a first heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 164; a second light chain comprising a sequence with at least 60% identity to SEQ ID NO:166; and a second heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 165.

24. The bispecific antibody of any one of claims 13-23, comprising: a first light chain comprising a sequence with at least 60% identity to SEQ ID NO: 167; a first heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 168; a second light chain comprising a sequence with at least 60% identity to SEQ ID NO: 170; and a second heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 169.

25. The bispecific antibody of any one of claims 13-24, comprising: a first light chain comprising a sequence with at least 60% identity to SEQ ID NO: 181; a first heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 182; a second light chain comprising a sequence with at least 60% identity to SEQ ID NO:184; and a second heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 183.

26. The bispecific antibody of any one of claims 13-25, comprising: a first light chain comprising a sequence with at least 60% identity to SEQ ID NO: 185; a first heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 186;a second light chain comprising a sequence with at least 60% identity to SEQ ID NO: 188; and a second heavy chain comprising a sequence with at least 60% identity to SEQ ID NO: 187.

27. An isolated nucleic acid encoding the recombinant antibody of any one of claims 1-12, the bispecific antibody of any one of claims 13-26.

28. A vector comprising the isolated nucleic acid of claim 27.

29. A cell comprising the vector of claim 28.

30. A composition comprising the recombinant antibody of any one of claims 1-12, the bispecific antibody of any one of claims 13-26.

31. A method of treating and / or preventing a neurological disorder in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the recombinant antibody of any one of claims 1-12, the bispecific antibody of any one of claims 13- 26.

32. The method of claim 31 , where the neurological disorder is Alzheimer’ s disease.

33. A method of modulating the production and / or accumulation of amyloid- P (A ) in a subject, comprising: administering to the subject a therapeutically effective amount of an antibody or antibody binding fragment having a leukocyte immunoglobulin-like receptor (LILR) binding domain.

34. The method of claim 33, wherein the leukocyte immunoglobulin- like receptor binding domain is a leukocyte immunoglobulin- like receptor B2 (LILRB2) binding domain and / or leukocyte immunoglobulin-like receptor B3 (LILRB3) binding domain.

35. The method of any one of claims 33-34, wherein the antibody or antibody binding fragment is an antagonist of LILRB2 and / or agonist of LILRB3.

36. The method of any one of claims 33-35, wherein the antibody or antibody binding fragment is a humanized antibody.

37. The method of any one of claims 33-36, wherein the antibody or antibody binding fragment specifically binds to LILRB2.

38. The method of any one of claims 33-37, wherein the antibody or antibody binding fragment specifically binds to LILRB3.

39. The method of any one of claims 33-38, wherein the antibody or antibody binding fragment comprises a heavy chain variable region (VH) having a complementary determining region (CDR)l comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 27, 33, 39, 45, 57, 63, 69, 75, 81, 87, 93, 99, 105, 111, 117, 123, or 129, a CDR2 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 28, 34, 40, 46, 58, 64, 70, 76, 82, 88, 94, 100, 106, 112, 118, 124, or 130, and a CDR3 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 29, 35, 41, 47, 59, 65, 71, 77, 83, 89, 95, 101, 107, 113, 119, 125, or 131.

40. The method of any one of claims 33-39, wherein the antibody or antibody binding fragment comprises a heavy chain variable region (VH) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, or 25.

41. The method of any one of claims 33-40, wherein the antibody or antibody binding fragment comprises a light chain variable region (VL) having a CDR1 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 30, 36, 42, 48, 60, 66, 72, 78, 84, 90, 96, 102, 108, 114, 120, 126, or 132, a CDR2 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 31, 37, 43, 49, 61, 67, 73, 79, 85, 91, 97, 103, 109, 115, 121, 127, or 133, and a CDR3 comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 32, 38, 44, 50, 62, 68, 74, 80, 86, 92, 98, 104, 110, 116, 122, 128, or 134.

42. The method of any one of claims 33-41, wherein the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to any one of SEQ ID NOS: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, or 26.

43. The method of any one of claims 33-42, wherein the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 13 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 14.

44. The method of any one of claims 33-43, wherein the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 7 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 8.

45. The method of any one of claims 33-44, wherein the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 9 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 10.

46. The method of any one of claims 33-45, wherein the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 1 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 2.

47. The method of any one of claim 33-46, wherein the antibody or antibody binding fragment comprises a light chain variable region (VL) comprising a sequence with at least 60% identity to SEQ ID NO: 5 and a heavy chain variable region (VH) comprising a sequence with at least 60% identity to SEQ ID NO: 6.

48. A transgenic non-human animal having diminished amyloid-P (AP) pathology, wherein the genome of the animal comprises an LILRB2 and / or an LILRB3 human transgene.

49. The transgenic non-human animal of claim 42, wherein said animal is a mammal.

50. The transgenic non-human animal of any one of claims 48-49, wherein said animal is a rodent.

51. The transgenic non-human animal of any one of claims 48-50, wherein said animal is a 5XFAD mouse.

52. The transgenic non-human animal of any one of claims 48-51 , wherein the LILRB2 and / or the LILRB3 human transgene comprises bacterial artificial chromosome (BAC) inserted transgenes.

53. The transgenic non-human animal of any one of claims 48-52, wherein said transgenic animal exhibits anti-inflammatory characteristics as compared to a wild-type animal.

54. The transgenic non-human animal of any one of claims 48-53, wherein said transgenic animal is a 5XFAD mouse or progeny thereof, and wherein said 5XFAD mouse or progeny thereof exhibits anti-inflammatory characteristics as compared to a 5XFAD mouse not having the LILRB2 and / or the LILRB3 human transgene.

55. An isolated cell from the transgenic non-human animal of any one of claims 48-54,56. The isolated cell of claim 55, wherein said cell is a myeloid cell.

57. The isolated cell of claim 55, wherein said cell is a microglia.

58. The isolated cell of claim 55, wherein said cell is a disease-associated microglia (DAM).

59. The isolated cell of claim 58, wherein said DAM exhibits increased expression of Sppl,Lpl, and / or Lilrb4a compared to DAM of a 5XFAD mouse not having the LILRB2 and / or the LILRB3 human transgene.

60. A method of identifying a compound for treating and / or preventing a neurological disorder in a subject, the method comprising: administering the compound to the transgenic non-human animal of any one of claims 46-54 or contacting the compound with the isolated cell of any one of claims 55-59; and determining a therapeutic and / or prophylactic efficacy of the compound on the neurological disorder.

61. The method of claim 60, wherein the neurological disorder comprises a neuroinflammatory disorder.

62. The method of any one of claims 60-61, wherein the neurological disorder comprises dementia, frontotemporal dementia (FTD), Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), Huntington's disease, taupathy disease, Nasu-Hakola disease, and / or multiple sclerosis.

63. The method of claim 62, wherein the neurological disorder is Alzheimer’s disease.

64. The method of any one of claims 60-63, wherein the subject is a human subject.

65. The method of any one of claims 60-64, wherein the compound comprises an LILRB- specific antibody or antibody fragment.

66. The method of any one of claims 60-65, wherein the compound comprises an LILRB2 antagonist.

67. The method of any one of claims 60-66, wherein the compound comprises an LILRB3 agonist.

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