Anti-ILT4 antibodies and their pharmaceutical uses

Abstract

The present invention relates to anti-ILT4 antibodies and their pharmaceutical uses.

Description

[Technical Field]

[0001] This application claims priority from a Chinese patent application (application number 202211093055.2, filing date September 8, 2022).

[0002] The present disclosure is in the field of biotechnology, and more particularly, the present disclosure relates to anti-ILT4 antibodies and uses thereof. [Background technology]

[0003] Nothing herein necessarily constitutes prior art, but merely provides background information related to the present disclosure.

[0004] Immune checkpoints are a type of molecule expressed in immune cells that can regulate the level of immune responses and play an important role in preventing autoimmune diseases. Immune checkpoints are activated when the body's immune system is stimulated, but not excessively activated. This is because immune checkpoints can apply the brakes in time to keep the immune system within a normal range. Tumor cells can escape immune surveillance by overexpressing and activating certain immune checkpoint ligands, thereby inhibiting immune cell function. Targeted therapies against common T cell immune checkpoints, such as CTLA-4 and PD-1, have already achieved great success. However, due to the existence of other immune checkpoint compensation mechanisms, such inhibitors suffer from low efficacy and drug resistance. Therefore, there is still a need to develop other potential immune checkpoint targets for clinical use.

[0005] Immunoglobulin-like transcript 4 (ILT4, also known as LILRB2) belongs to the leukocyte immunoglobulin-like receptor (LILR) family, which has a total of 11 members and can be further divided into three types depending on whether their intracellular domains contain ITAM or ITIM. ILT4 belongs to the inhibitory LILRB family and is thought to function as an immune checkpoint. ILT4 is expressed in myeloid-derived immune cells such as macrophages, dendritic cells, and granulocytes, and its physiological function is to bind to MHC-I molecules and regulate immune balance and homeostasis. In tumor biology, ILT4 expression levels are positively correlated with the number of myeloid-derived suppressor cells (MDSCs), which is one of the main causes of drug resistance to CTLA-4 and PD-1 antibodies. On the other hand, HLA-G (non-classical MHC-I), one of the ILT4 ligands, has been reported to be widely expressed in various tumor tissues and is thought to be an important mechanism for immune evasion by tumor cells. Targeting ILT4 can induce the conversion of macrophages to a proinflammatory M1 phenotype, enhance antigen presentation by APCs, and ultimately activate T cells. At the same time, ILT4 antibodies can also alter the tumor microenvironment and reduce the inhibition of T cells by MDSCs.

[0006] Currently, Merck's MK-4830 (WO2018187518A1) and Jounce's JTX-8064 (WO2019126514A2) have entered Phase 2 and Phase 1 clinical trials, respectively. According to clinical data published by Merck, the combination of MK-4830 with the PD-1 antibody Keytruda in patients with metastatic or advanced solid tumors achieved an ORR of 24% (34 patients). Of the 11 patients whose disease progressed after PD-1 targeted therapy, 5 (45%) achieved relief, demonstrating the potential of ILT4 antibodies to overcome PD-1 targeting resistance. Meanwhile, adverse events associated with the combination of MK-4830 and Keytruda were similar to those observed with Keytruda alone, suggesting that the ILT4 antibody poses no significant safety concerns.

[0007] ILT4 plays a negative immunoregulatory role, and inhibitory antibodies targeting it can effectively activate immune cells and may be a valuable supplement to existing immune checkpoint therapies. Summary of the Invention

[0008] The present disclosure provides an anti-ILT4 antibody comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3, and a light chain variable region comprising LCDR1, LCDR2 and LCDR3, wherein: a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 184 or 185, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 187 or 186, respectively; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 32, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 33, respectively; or b. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 211, 205, 206, 207, 208, 209, or 210, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in any one of SEQ ID NOs: 212, 213, or 214; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 40, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 41; or c. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of any one of SEQ ID NOs: 181, 176, 177, 178, 179, or 180, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 183 or 182; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of SEQ ID NO: 28, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 29; or d. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 193, 190, 191, or 192, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in any one of SEQ ID NOs: 194, 195, or 196; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 10, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 11; or e. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 163, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 225 or 164, respectively; or f. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 197 or 198, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 199 or 200, respectively; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 36, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 37, respectively; or g. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of any one of SEQ ID NOs: 217, 218, 219, 220, or 221, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 222 or 223; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of SEQ ID NO: 42, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 43; or h. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 34, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 35, respectively; or i. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 18, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 19, respectively; or j. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 20, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 21, respectively; or k. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 24, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 25, respectively; or l. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 44, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 45, respectively; or m. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 46, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 47, respectively; or n. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 48, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 49, respectively; or o. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 50, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 51, respectively; or p. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 4, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 5, respectively; or q. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 6, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 7, respectively; or r. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 8, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 9, respectively; or s. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 12, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 13, respectively; or t. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 14, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 15, respectively; or u. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 16, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 17, respectively; or v. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 22, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 23, respectively; or w. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 26, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 27, respectively; or x. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 30, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 31, respectively; or y. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 38, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 39, respectively.

[0009] In some embodiments, the anti-ILT4 antibody is a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 184 or 185, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 187 or 186, respectively; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 32, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 33, respectively; or b. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 211, 205, 206, 207, 208, 209, or 210, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in any one of SEQ ID NOs: 212, 213, or 214; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 40, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 41; or c. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of any one of SEQ ID NOs: 181, 176, 177, 178, 179, or 180, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 183 or 182; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of SEQ ID NO: 28, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 29; or d. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 193, 190, 191, or 192, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in any one of SEQ ID NOs: 194, 195, or 196; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 10, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 11; or e. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 163, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 225 or 164, respectively; or f. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 197 or 198, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 199 or 200, respectively; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 36, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 37, respectively; or g. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of any one of SEQ ID NOs: 217, 218, 219, 220, or 221, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 222 or 223; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, of SEQ ID NO: 42, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, of SEQ ID NO: 43; or h. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 34, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 35, respectively; or i. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 18, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 19, respectively; or j. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 20, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 21, respectively; or k. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 24, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 25, respectively; or l. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 44, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 45, respectively; or m. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 46, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 47, respectively; or n. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 48, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 49, respectively; or o. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 50, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 51, respectively.

[0010] In some embodiments, the anti-ILT4 antibody is a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 184, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 187; or b. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 211, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 212, respectively; or c. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 181, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 183, respectively; or d. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 193, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 194; or e. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 163, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 225, respectively; or f. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 197, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 199; or g. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 217, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 222, respectively.

[0011] In some embodiments, the anti-ILT4 antibody is a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 184, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 187; or b. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 211, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 212, respectively.

[0012] In some embodiments, the anti-ILT4 antibody is p. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 4, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 5, respectively; or q. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 6, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 7, respectively; or r. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 8, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 9, respectively; or s. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 12, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 13, respectively; or t. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 14, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 15, respectively; or u. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 16, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 17, respectively; or v. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 22, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 23, respectively; or w. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 26, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 27, respectively; or x. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 30, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 31, respectively; or y. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 38, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 39, respectively.

[0013] In some embodiments, in any one of the anti-ILT4 antibodies described above, HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to a similar numbering convention selected from Kabat, IMGT, Chothia, AbM, and Contact. In some embodiments, HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to the Kabat numbering convention. In some embodiments, HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to the IMGT numbering convention. In some embodiments, HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to the Chothia numbering convention. In some embodiments, the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and the LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to the AbM numbering convention. In some embodiments, the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and the LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to the Contact numbering convention.

[0014] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 203, 136, 201, 202, or 204, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; or c. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 111, HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 173, 112, 174, or 175, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 113, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 115, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 116; or d. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 69, HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 189, 70, or 188, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 71, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 61, LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 72; or e. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 165, HCDR2 comprising the amino acid sequence of SEQ ID NO: 166, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 167, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 224 or 168, LCDR2 comprising the amino acid sequence of SEQ ID NO: 169, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 170; or f. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 125, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 126, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 127, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 128; or g. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 141, HCDR2 comprises the amino acid sequence of SEQ ID NO: 215 or 142, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 216 or 143; and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 144, LCDR2 comprises the amino acid sequence of SEQ ID NO: 145, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 146; or h. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 99, HCDR2 comprising the amino acid sequence of SEQ ID NO: 122, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 123, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 124; or i. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 73, HCDR2 comprising the amino acid sequence of SEQ ID NO: 87, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 88, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 61, LCDR2 comprising the amino acid sequence of SEQ ID NO: 89, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 90; or j. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 91, HCDR2 comprising the amino acid sequence of SEQ ID NO: 92, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 93, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 85, LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 94; or k. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 99, HCDR2 comprises the amino acid sequence of SEQ ID NO: 100, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 101; and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 102, LCDR2 comprises the amino acid sequence of SEQ ID NO: 103, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 104; or l. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 147, HCDR2 comprises the amino acid sequence of SEQ ID NO: 148, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 149, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 150, LCDR2 comprises the amino acid sequence of SEQ ID NO: 151, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 152; or m. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 153, HCDR2 comprises the amino acid sequence of SEQ ID NO: 154, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 155, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 133, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; or n. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 58, HCDR2 comprises the amino acid sequence of SEQ ID NO: 156, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 157, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 158, LCDR2 comprises the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 159; or the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 160, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 161; and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 162; or p. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 52, HCDR2 comprises the amino acid sequence of SEQ ID NO: 53, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 54, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 55, LCDR2 comprises the amino acid sequence of SEQ ID NO: 56, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 57; or q. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 58, HCDR2 comprises the amino acid sequence of SEQ ID NO: 59, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 60, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 61, LCDR2 comprises the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 63; or r. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 64, HCDR2 comprising the amino acid sequence of SEQ ID NO: 65, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 66, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 67, LCDR2 comprising the amino acid sequence of SEQ ID NO: 56, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 68; or s. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 73, HCDR2 comprising the amino acid sequence of SEQ ID NO: 74, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 75, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 76, LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 77; or t. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 78, HCDR2 comprises the amino acid sequence of SEQ ID NO: 79, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 80, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 81, LCDR2 comprises the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 82; or u. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO:52, HCDR2 comprising the amino acid sequence of SEQ ID NO:83, and HCDR3 comprising the amino acid sequence of SEQ ID NO:84, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO:85, LCDR2 comprising the amino acid sequence of SEQ ID NO:62, and LCDR3 comprising the amino acid sequence of SEQ ID NO:86; or v. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 95, HCDR2 comprises the amino acid sequence of SEQ ID NO: 96, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 97, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 85, LCDR2 comprises the amino acid sequence of SEQ ID NO: 98, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 68; or w. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 106, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 107, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 108, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 110; or x. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 117, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 118, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; or y. The heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 129, HCDR2 comprising the amino acid sequence of SEQ ID NO: 130, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 131, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 132, LCDR2 comprising the amino acid sequence of SEQ ID NO: 133, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 134.

[0015] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 203, 136, 201, 202, or 204, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; or c. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 111, HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 173, 112, 174, or 175, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 113, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 115, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 116; or d. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 69, HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 189, 70, or 188, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 71, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 61, LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 72; or e. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 165, HCDR2 comprising the amino acid sequence of SEQ ID NO: 166, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 167, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 224 or 168, LCDR2 comprising the amino acid sequence of SEQ ID NO: 169, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 170; or f. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 125, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 126, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 127, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 128; or g. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 141, HCDR2 comprises the amino acid sequence of SEQ ID NO: 215 or 142, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 216 or 143; and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 144, LCDR2 comprises the amino acid sequence of SEQ ID NO: 145, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 146; or h. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 99, HCDR2 comprising the amino acid sequence of SEQ ID NO: 122, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 123, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 124; or i. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 73, HCDR2 comprising the amino acid sequence of SEQ ID NO: 87, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 88, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 61, LCDR2 comprising the amino acid sequence of SEQ ID NO: 89, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 90; or j. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 91, HCDR2 comprising the amino acid sequence of SEQ ID NO: 92, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 93, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 85, LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 94; or k. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 99, HCDR2 comprises the amino acid sequence of SEQ ID NO: 100, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 101; and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 102, LCDR2 comprises the amino acid sequence of SEQ ID NO: 103, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 104; or l. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 147, HCDR2 comprises the amino acid sequence of SEQ ID NO: 148, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 149, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 150, LCDR2 comprises the amino acid sequence of SEQ ID NO: 151, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 152; or m. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 153, HCDR2 comprises the amino acid sequence of SEQ ID NO: 154, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 155, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 133, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; or n. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 58, HCDR2 comprises the amino acid sequence of SEQ ID NO: 156, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 157, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 158, LCDR2 comprises the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 159; or o. The heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 160, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 161, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 162.

[0016] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of SEQ ID NO: 203, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; or c. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 111, HCDR2 comprising the amino acid sequence of SEQ ID NO: 173, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 113, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 115, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 116; or d. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 69, HCDR2 comprising the amino acid sequence of SEQ ID NO: 189, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 71, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 61, LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 72; or e. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 165, HCDR2 comprising the amino acid sequence of SEQ ID NO: 166, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 167, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 224, LCDR2 comprising the amino acid sequence of SEQ ID NO: 169, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 170; or f. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 125, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 126, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 127, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 128; or g. The heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 141, HCDR2 comprising the amino acid sequence of SEQ ID NO: 215, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 216, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 144, LCDR2 comprising the amino acid sequence of SEQ ID NO: 145, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 146.

[0017] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. The heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of SEQ ID NO: 203, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140.

[0018] In some embodiments, the anti-ILT4 antibody of any one of the above, In the heavy chain variable region, HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 105, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 120, and HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 121, and in the light chain variable region, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 114, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 119.

[0019] In some embodiments, the anti-ILT4 antibody of any one of the above, p. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 52, HCDR2 comprises the amino acid sequence of SEQ ID NO: 53, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 54, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 55, LCDR2 comprises the amino acid sequence of SEQ ID NO: 56, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 57; or q. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 58, HCDR2 comprises the amino acid sequence of SEQ ID NO: 59, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 60, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 61, LCDR2 comprises the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 63; or r. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 64, HCDR2 comprising the amino acid sequence of SEQ ID NO: 65, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 66, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 67, LCDR2 comprising the amino acid sequence of SEQ ID NO: 56, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 68; or s. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 73, HCDR2 comprising the amino acid sequence of SEQ ID NO: 74, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 75, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 76, LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 77; or t. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 78, HCDR2 comprises the amino acid sequence of SEQ ID NO: 79, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 80, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 81, LCDR2 comprises the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 82; or u. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO:52, HCDR2 comprising the amino acid sequence of SEQ ID NO:83, and HCDR3 comprising the amino acid sequence of SEQ ID NO:84, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO:85, LCDR2 comprising the amino acid sequence of SEQ ID NO:62, and LCDR3 comprising the amino acid sequence of SEQ ID NO:86; or v. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 95, HCDR2 comprises the amino acid sequence of SEQ ID NO: 96, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 97, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 85, LCDR2 comprises the amino acid sequence of SEQ ID NO: 98, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 68; or w. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 106, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 107, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 108, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 110; or x. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 117, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 118, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; or y. The heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 129, HCDR2 comprising the amino acid sequence of SEQ ID NO: 130, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 131, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 132, LCDR2 comprising the amino acid sequence of SEQ ID NO: 133, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 134.

[0020] In some embodiments, in any one of the above anti-ILT4 antibodies, HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to the Kabat numbering convention.

[0021] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 260, HCDR2 comprising the amino acid sequence of SEQ ID NO: 261, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 262, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 263, LCDR2 comprising the amino acid sequence of SEQ ID NO: 264, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 267, HCDR2 comprising the amino acid sequence of SEQ ID NO: 268, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 269, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 270, LCDR2 comprising the amino acid sequence of SEQ ID NO: 271, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; Among them, the CDR regions are determined by the IMGT numbering convention.

[0022] In some embodiments, the anti-ILT4 antibody of any one of the above, the heavy chain variable region, HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 260, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 261, and HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 262; and the light chain variable region, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 263, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 264, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 119; Among them, the CDR regions are determined by the IMGT numbering convention.

[0023] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 265, HCDR2 comprising the amino acid sequence of SEQ ID NO: 266, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 272, HCDR2 comprising the amino acid sequence of SEQ ID NO: 273, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; Among them, the CDR regions are determined by the Chothia numbering convention.

[0024] In some embodiments, the anti-ILT4 antibody of any one of the above, the heavy chain variable region, HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 265, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 266, and HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 121; and the light chain variable region, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 114, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 119; Among them, the CDR regions are determined by the Chothia numbering convention.

[0025] In some embodiments, the anti-ILT4 antibody of any one of the above is a murine antibody, a chimeric antibody, a humanized antibody, or a fully human antibody. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the antibody is a fully human antibody. In some embodiments, the antibody is a humanized antibody.

[0026] In some embodiments, the anti-ILT4 antibody of any one of the above, which comprises a framework region (FR) of a human antibody.

[0027] In some embodiments, the anti-ILT4 antibody of any one of the above, the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 184 or 185, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 187 or 186; or the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 32, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 33; or b. the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 211, 205, 206, 207, 208, 209 or 210, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 212, 213 or 214. or wherein the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 40 and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 41; or c. the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 181, 176, 177, 178, 179 or 180, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 183 or 182. or the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:28 and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:29; or d. the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 193, 190, 191, or 192, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 194, 195, or 196; or the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 10, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 11; or e. the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 163, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 225 or 164; f. the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 197 or 198, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 199 or 200; or the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 36, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 37; or g. the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 217, 218, 219, 220 or 221, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 222 or 223; or the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 42, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO: 43.

[0028] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 184 or 185, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 187 or 186; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 32, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 33; or b. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 211, 205, 206, 207, 208, 209, or 210, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 212, 213, or 214; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 40, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 41; or c. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 181, 176, 177, 178, 179, or 180, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 183 or 182; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:28 and the light chain variable region comprises the amino acid sequence of SEQ ID NO:29; or d. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 193, 190, 191, or 192, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 194, 195, or 196; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 10, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 11; or e. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 163, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 225 or 164; f. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 197 or 198, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 199 or 200; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 36, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 37; or g. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 217, 218, 219, 220, or 221, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 222 or 223; or The heavy chain variable region comprises the amino acid sequence of SEQ ID NO:42, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:43.

[0029] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 184 or 185, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 187 or 186; or b. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 211, 205, 206, 207, 208, 209, or 210, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 212, 213, or 214; or c. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 181, 176, 177, 178, 179, or 180, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 183 or 182; or d. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 193, 190, 191, or 192, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 194, 195, or 196; or e. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 163, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 225 or 164; f. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 197 or 198, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 199 or 200; or g. The heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 217, 218, 219, 220, or 221, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 222 or 223.

[0030] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 184 and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 187; or b. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:211 and the light chain variable region comprises the amino acid sequence of SEQ ID NO:212; or c. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 181, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 183; or d. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 193, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 194; or e. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 163, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 225; f. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 197, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 199; or g. The heavy chain variable region comprises the amino acid sequence of SEQ ID NO:217, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:222.

[0031] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 184 and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 187; or b. The heavy chain variable region comprises the amino acid sequence of SEQ ID NO:211, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:212.

[0032] In some embodiments, the anti-ILT4 antibody of any one of the above, The heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO:184, and the light chain variable region comprises the amino acid sequence set forth in SEQ ID NO:187.

[0033] In some embodiments, the anti-ILT4 antibody of any one of the above, a. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 32 and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 33; or b. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 40, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 41; or c. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:28 and the light chain variable region comprises the amino acid sequence of SEQ ID NO:29; or d. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 10, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 11; or f. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 36, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 37; or g. The heavy chain variable region comprises the amino acid sequence of SEQ ID NO:42, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:43.

[0034] In some embodiments, the anti-ILT4 antibody described in any one of the above, wherein the heavy chain variable region has FR1, FR2, FR3 derived from IGHV4-31*01 and FR4 derived from IGHJ1*01, and is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 1E, 27S, 30T, 44K, 71R, and 76S, and / or the light chain variable region has FR1, FR2, FR3 derived from IGKV1-39*01 and FR4 derived from IGKJ4*01, and is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 43S, 48V, and 71Y. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region having HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the FR of the heavy chain variable region comprises one or more amino acid substitutions selected from the group consisting of 1E, 27S, 30T, 44K, 71R, and 76S; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 43S, 48V, and 71Y. In some embodiments, the variable region and CDRs are defined according to the Kabat numbering convention.

[0035] In some embodiments, the anti-ILT4 antibody described in any one of the above, wherein the heavy chain variable region has FR1, FR2, FR3 derived from IGHV1-18*01 and FR4 derived from IGHJ6*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 1E, 43E, 69L, 71V, 73K, 76N, and 93T, and / or the light chain variable region has FR1, FR2, FR3 derived from IGKV4-1*01 or IGKV3-11*01 and FR4 derived from IGKJ4*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 4L, 43P, 45K, 58I, 68R, and 85T. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region having HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of SEQ ID NO: 203, 136, 201, 202, or 204, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the FR of the heavy chain variable region comprises one or more amino acid substitutions selected from the group consisting of 1E, 43E, 69L, 71V, 73K, 76N, and 93T; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 4L, 43P, 45K, 58I, 68R, and 85T. In some embodiments, the variable region and CDRs are defined according to the Kabat numbering convention.

[0036] In some embodiments, the anti-ILT4 antibody described in any one of the above, wherein the heavy chain variable region has FR1, FR2, FR3 derived from IGHV1-3*01 and FR4 derived from IGHJ6*01, and is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 1E, 38K, 44S, 69L, 71V, 73K, and 75G, and / or the light chain variable region has FR1, FR2, FR3 derived from IGKV1-39*01 and FR4 derived from IGKJ2*01, and is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 43S, 48V, and 71Y. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region having HCDR1 comprising the amino acid sequence of SEQ ID NO: 111, HCDR2 comprising the amino acid sequence of SEQ ID NO: 173, 112, 174, or 175, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 113, and the FR of the heavy chain variable region comprises one or more amino acid substitutions selected from the group consisting of 1E, 38K, 44S, 69L, 71V, 73K, and 75G; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 115, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 116, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 43S, 48V, and 71Y. In some embodiments, the variable region and CDRs are defined according to the Kabat numbering convention.

[0037] In some embodiments, the anti-ILT4 antibody described in any one of the above, wherein the heavy chain variable region has FR1, FR2, FR3 derived from IGHV3-21*01 and FR4 derived from IGHJ1*01, and is unsubstituted or has a 49A substitution, and / or the light chain variable region has FR1, FR2, FR3 derived from IGKV1-33*01 and FR4 derived from IGKJ4*01, and is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 39R, 42Q, 43S, and 85V. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region, wherein HCDR1 comprises the amino acid sequence of SEQ ID NO: 69, HCDR2 comprises the amino acid sequence of SEQ ID NO: 189, 70, or 188, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 71, and the FR of the heavy chain variable region comprises a 49A substitution; and the light chain variable region, wherein LCDR1 comprises the amino acid sequence of SEQ ID NO: 61, LCDR2 comprises the amino acid sequence of SEQ ID NO: 62, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 72, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 39R, 42Q, 43S, and 85V. In some embodiments, the variable regions and CDRs are defined according to the Kabat numbering convention.

[0038] In some embodiments, the anti-ILT4 antibody described in any one of the above, wherein the heavy chain variable region has FR1, FR2, FR3 derived from IGHV4-31*02 and FR4 derived from IGHJ6*01, and is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 1E, 27Y, 30T, 42E, 44K, and 71R, and / or the light chain variable region has FR1, FR2, FR3 derived from IGKV1-39*01 and FR4 derived from IGKJ4*01, and is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 43S, 48L, and 71Y. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region having HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 125, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 126, and the FR of the heavy chain variable region comprises one or more amino acid substitutions selected from the group consisting of 1E, 27Y, 30T, 42E, 44K, and 71R; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 127, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 128, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 43S, 48L, and 71Y. In some embodiments, the variable region and CDRs are defined according to the Kabat numbering convention.

[0039] In some embodiments, the anti-ILT4 antibody described in any one of the above, wherein the heavy chain variable region has FR1, FR2, FR3 derived from IGHV1-18*01 and FR4 derived from IGHJ6*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 1E, 2I, 44A, 46K, and 76T, and / or the light chain variable region has FR1, FR2, FR3 derived from IGKV4-1*01 or IGKV2-28*01 and FR4 derived from IGKJ4*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 43S and 48L. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region having HCDR1 comprising the amino acid sequence of SEQ ID NO: 141, HCDR2 comprising the amino acid sequence of SEQ ID NO: 215 or 142, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 216 or 143, and the FR of the heavy chain variable region comprises one or more amino acid substitutions selected from the group consisting of 1E, 2I, 44A, 46K, and 76T; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 144, LCDR2 comprising the amino acid sequence of SEQ ID NO: 145, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 146, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 43S and 48L. In some embodiments, the variable region and CDRs are defined according to the Kabat numbering convention.

[0040] In some embodiments, the anti-ILT4 antibody is any one of the above, wherein the anti-ILT4 antibody is an antibody fragment, and in some embodiments, the antibody fragment is selected from Fab, Fab', F(ab')2, Fd, Fv, scFv, dsFv, or dAb.

[0041] In some embodiments, the anti-ILT4 antibody is any one of the above, comprising a heavy chain constant region and a light chain constant region. In some embodiments, the heavy chain constant region is a human IgG1 or IgG4 heavy chain constant region. In some embodiments, the heavy chain constant region has 234A and 235A mutations, wherein the mutations are numbered according to the EU index. In some embodiments, the light chain constant region is a human κ light chain constant region. In some embodiments, the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 240 or 171, and the light chain constant region comprises the amino acid sequence of SEQ ID NO: 172. In some embodiments, the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 240, and the light chain constant region comprises the amino acid sequence of SEQ ID NO: 172.

[0042] In some embodiments, the anti-ILT4 antibody of any one of the above, wherein the anti-ILT4 antibody comprises a heavy chain and a light chain, a. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 228, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 229; or b. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:234, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:235; or c. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 226, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 227; or d. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 230, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 231; or e. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 238, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 239; or f. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 232, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 233; or g. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:236, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:237.

[0043] In some embodiments, the anti-ILT4 antibody of any one of the above, wherein the anti-ILT4 antibody comprises a heavy chain and a light chain, a. the heavy chain comprises the amino acid sequence of SEQ ID NO: 228, and the light chain comprises the amino acid sequence of SEQ ID NO: 229; b. the heavy chain comprises the amino acid sequence of SEQ ID NO: 234, and the light chain comprises the amino acid sequence of SEQ ID NO: 235; c. the heavy chain comprises the amino acid sequence of SEQ ID NO: 226, and the light chain comprises the amino acid sequence of SEQ ID NO: 227; d. the heavy chain comprises the amino acid sequence of SEQ ID NO: 230, and the light chain comprises the amino acid sequence of SEQ ID NO: 231; e. the heavy chain comprises the amino acid sequence of SEQ ID NO: 238, and the light chain comprises the amino acid sequence of SEQ ID NO: 239; f. the heavy chain comprises the amino acid sequence of SEQ ID NO: 232, and the light chain comprises the amino acid sequence of SEQ ID NO: 233; g. The heavy chain comprises the amino acid sequence of SEQ ID NO:236, and the light chain comprises the amino acid sequence of SEQ ID NO:237.

[0044] In some embodiments, the anti-ILT4 antibody of any one of the above, wherein the anti-ILT4 antibody comprises a heavy chain and a light chain, a. the heavy chain comprises the amino acid sequence of SEQ ID NO: 228, and the light chain comprises the amino acid sequence of SEQ ID NO: 229; b. the heavy chain comprises the amino acid sequence of SEQ ID NO:234, and the light chain comprises the amino acid sequence of SEQ ID NO:235.

[0045] In some embodiments, the anti-ILT4 antibody of any one of the above, wherein the anti-ILT4 antibody comprises a heavy chain and a light chain, The heavy chain comprises the amino acid sequence set forth in SEQ ID NO:228, and the light chain comprises the amino acid sequence set forth in SEQ ID NO:229.

[0046] In some embodiments, the present disclosure further provides an isolated anti-ILT4 antibody that binds to human ILT4 or an epitope thereof in competition with any one of the anti-ILT4 antibodies described above.

[0047] In some embodiments, the anti-ILT4 antibodies provided herein specifically bind to human ILT4 but do not bind to other members of the LILR family: ILT2, ILT3, LILRB5, LILRA1, LILRA2, LILRA3, LILRA4, LILRA5 and / or LILRA6.

[0048] In some embodiments, an isolated anti-ILT4 antibody according to the present disclosure has a higher ability to bind to ILT4 protein than a control antibody. Specifically, an isolated anti-ILT4 antibody according to the present disclosure has a binding capacity of 1×10 -8 Less than M (e.g., 8 × 10 -9 Less than M, 5 x 10 -9 Less than M, 2 x 10 -9 Less than M, 1 x 10 -9 Less than M, 8 x 10 -10 Less than M, 5 x 10 -10 Less than M, 2 x 10 -10 Less than M, 1 x 10 -10 Less than M, 8 x 10 -11 Less than M, 5 x 10 -11 It binds to human ILT4 with a KD value of less than 1 M, as measured by Biacore.

[0049] In some embodiments, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively relieve the inhibitory effect of ILT4 and promote the secretion of higher levels of mIL-2 by 3A9 cells.

[0050] In some embodiments, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively promote the conversion of macrophages to the M1 phenotype and secrete high levels of the pro-inflammatory cytokine TNF-α.

[0051] In some embodiments, the humanized anti-ILT4 antibodies provided herein (e.g., huP2-G9) can induce increased levels of pro-inflammatory cytokine or chemokine secretion by dendritic cells in an immunosuppressive environment in the presence of IL-10.

[0052] In some embodiments, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively block the binding of ILT4 to HLA-G.

[0053] In some embodiments, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively block the binding of ILT4 to HLA-G, and the blocking activity is stronger than that of the control antibodies MK and JTX.

[0054] In another aspect, the present disclosure provides a pharmaceutical composition comprising any one of the anti-ILT4 antibodies described above and one or more pharmaceutically acceptable carriers, diluents, or excipients. In some embodiments, the pharmaceutical composition further comprises at least one second therapeutic agent. In some embodiments, the second therapeutic agent is an anti-PD-1 antibody.

[0055] In another aspect, the present disclosure provides an isolated nucleic acid encoding any one of the above anti-ILT4 antibodies.

[0056] In another aspect, the present disclosure provides a vector comprising the nucleic acid molecule.

[0057] In another aspect, the present disclosure provides a host cell comprising any one of the above isolated nucleic acids.

[0058] In another aspect, the present disclosure provides a method of blocking the binding of ILT4 to HLA-G in a human subject in need thereof, said method comprising administering to said human subject an effective amount of any one of the anti-ILT4 antibodies described above.

[0059] In another aspect, the present disclosure provides a method for detecting an ILT4 peptide or a fragment thereof in a sample, said method comprising contacting said sample with any one of the anti-ILT4 antibodies described above and detecting the presence of a complex between said anti-ILT4 antibody and said ILT4 peptide or fragment thereof, wherein detection of said complex indicates the presence of said ILT4 peptide or fragment thereof.

[0060] In another aspect, the present disclosure provides a method of treating a disease, said method comprising the step of administering to a subject an anti-ILT4 antibody or pharmaceutical composition according to any one of the above.

[0061] In another aspect, the present disclosure provides the use of any one of the anti-ILT4 antibodies or pharmaceutical compositions described above in the preparation of a medicament for treating a disease.

[0062] In another aspect, the present disclosure provides an anti-ILT4 antibody or pharmaceutical composition according to any one of the preceding claims for use as a medicament. In some embodiments, the medicament is used in the treatment of a disease.

[0063] In some embodiments, the disease described in any one of the above is a tumor. In some embodiments, the disease is selected from the group consisting of astrocytoma (e.g., anaplastic astrocytoma), glioblastoma, bladder cancer, bone cancer, brain cancer, breast cancer (e.g., breast cancer characterized by BRCA1 and / or BRCA2 mutations), carcinoid, cervical cancer, choroid plexus papilloma, colorectal cancer (e.g., colon and rectal cancer), fallopian tube cancer, ependymoma, gallbladder cancer, gastric cancer, head and neck cancer, idiopathic myelofibrosis, kidney cancer or renal cell carcinoma (e.g., renal rhabdoid tumor and Wilms' tumor), renal pelvis tumor, leukemia, liver cancer (e.g., hepatocellular carcinoma), esophageal cancer (also known as "esophageal cancer," e.g., esophageal squamous cell carcinoma), lung cancer (e.g., non-small cell lung cancer and The disease is selected from small cell lung cancer, lymphoma, medulloblastoma, melanoma, meningioma, Merkel cell carcinoma, mesothelioma, multiple myeloma, neuroblastoma, oligodendroglioma, ovarian cancer, peritoneal tumor, pancreatic cancer, polycythemia vera, primary neuroectodermal tumor, prostate cancer, retinoblastoma, salivary gland cancer, sarcoma (e.g., chondrosarcoma, Ewing's sarcoma, osteosarcoma, rhabdomyosarcoma, synovial sarcoma, and soft tissue sarcoma), small intestine cancer, colon cancer, squamous cell carcinoma (e.g., cutaneous squamous cell carcinoma), thyroid cancer, endometrial cancer, vestibular schwannoma, blastoma, teratoma, pituitary cancer, vulvar cancer, thymoma, testicular cancer, bile duct cancer, pheochromocytoma, paraganglioma, and adenoid cystic carcinoma. In some embodiments, the disease is selected from ovarian cancer, lung cancer, esophageal squamous cell carcinoma, colorectal cancer, renal cell carcinoma, and melanoma.

[0064] In some embodiments, the method for treating the above-mentioned disease further comprises administering a second therapeutic agent. In some embodiments, the second therapeutic agent comprises an immune checkpoint inhibitor. In some embodiments, the second therapeutic agent is an anti-PD-1 antibody.

[0065] In some embodiments, the method for treating the above diseases is provided, wherein the anti-PD-1 antibody comprises a heavy chain variable domain PD-1-VH and a light chain variable domain PD-1-VL, wherein: The PD-1-VH has a PD-1-HCDR1 comprising the amino acid sequence of SEQ ID NO: 250, a PD-1-HCDR2 comprising the amino acid sequence of SEQ ID NO: 251, and a PD-1-HCDR3 comprising the amino acid sequence of SEQ ID NO: 252, and the PD-1-VL has a PD-1-LCDR1 comprising the amino acid sequence of SEQ ID NO: 253, a PD-1-LCDR2 comprising the amino acid sequence of SEQ ID NO: 254, and a PD-1-LCDR3 comprising the amino acid sequence of SEQ ID NO: 255.

[0066] In some embodiments, the method for treating the above diseases is provided, wherein the anti-PD-1 antibody comprises a heavy chain variable domain PD-1-VH and a light chain variable domain PD-1-VL, wherein: The PD-1-VH comprises the amino acid sequence of SEQ ID NO:256, and the PD-1-VL comprises the amino acid sequence of SEQ ID NO:257.

[0067] In some embodiments, the method for treating the above diseases, wherein the anti-PD-1 antibody comprises a heavy chain and a light chain, The heavy chain comprises the amino acid sequence of SEQ ID NO:248, and the light chain comprises the amino acid sequence of SEQ ID NO:249.

[0068] In another embodiment is the use of a combination of an anti-ILT4 antibody and an anti-PD-1 antibody according to any one of the above in the preparation of a medicament for treating a disease.

[0069] In some embodiments, the medicament is used to treat a disease, preferably the disease is a tumor, more preferably the disease is selected from the group consisting of astrocytoma (e.g., anaplastic astrocytoma), glioblastoma, bladder cancer, bone cancer, brain cancer, breast cancer (e.g., breast cancer characterized by BRCA1 and / or BRCA2 mutations), carcinoid, cervical cancer, choroid plexus papilloma, colorectal cancer (e.g., colon and rectal cancer), fallopian tube cancer, ependymoma, gallbladder cancer, gastric cancer, head and neck cancer, idiopathic myelofibrosis, kidney cancer or renal cell carcinoma (e.g., renal rhabdoid tumor and Wilms' tumor), renal pelvic tumor, leukemia, liver cancer (e.g., hepatocellular carcinoma), esophageal cancer (also known as "esophageal cancer", e.g., squamous esophageal skin cancer), lung cancer (e.g., non-small cell lung cancer and small cell lung cancer), lymphoma, medulloblastoma, melanoma, meningioma, Merkel cell carcinoma, mesothelioma, multiple myeloma, neuroblastoma, oligodendroglioma, ovarian cancer, peritoneal tumor, pancreatic cancer, polycythemia vera, primary neuroectodermal tumor, prostate cancer, retinoblastoma, salivary gland cancer, sarcoma (e.g., chondrosarcoma, Ewing's sarcoma, osteosarcoma, rhabdomyosarcoma, synovial sarcoma, and soft tissue sarcoma), small intestine cancer, colon cancer, squamous cell carcinoma (e.g., cutaneous squamous cell carcinoma), thyroid cancer, endometrial cancer, vestibular schwannoma, blastoma, teratoma, pituitary cancer, vulvar cancer, thymoma, testicular cancer, bile duct cancer, pheochromocytoma, paraganglioma, and adenoid cystic carcinoma. In some embodiments, the medicament is used to treat a disease selected from ovarian cancer, lung cancer, esophageal squamous cell carcinoma, colorectal cancer, renal cell carcinoma, and melanoma.

[0070] In some embodiments, the use of a combination of an anti-ILT4 antibody and an anti-PD-1 antibody in the preparation of a medicament for treating a disease, wherein the anti-ILT4 antibody and the anti-PD-1 antibody are in the same or different containers.

[0071] In some embodiments, there is provided a use of a combination of an anti-ILT4 antibody and an anti-PD-1 antibody in the preparation of a medicament for treating a disease, wherein the anti-PD-1 antibody comprises a heavy chain variable domain PD-1-VH and a light chain variable domain PD-1-VL, wherein: The PD-1-VH has a PD-1-HCDR1 comprising the amino acid sequence of SEQ ID NO: 250, a PD-1-HCDR2 comprising the amino acid sequence of SEQ ID NO: 251, and a PD-1-HCDR3 comprising the amino acid sequence of SEQ ID NO: 252, and the PD-1-VL has a PD-1-LCDR1 comprising the amino acid sequence of SEQ ID NO: 253, a PD-1-LCDR2 comprising the amino acid sequence of SEQ ID NO: 254, and a PD-1-LCDR3 comprising the amino acid sequence of SEQ ID NO: 255.

[0072] In some embodiments, there is provided a use of a combination of an anti-ILT4 antibody and an anti-PD-1 antibody in the preparation of a medicament for treating a disease, wherein the anti-PD-1 antibody comprises a heavy chain variable domain PD-1-VH and a light chain variable domain PD-1-VL, wherein: The PD-1-VH comprises the amino acid sequence of SEQ ID NO:256, and the PD-1-VL comprises the amino acid sequence of SEQ ID NO:257.

[0073] In some embodiments, use of a combination of an anti-ILT4 antibody and an anti-PD-1 antibody in the preparation of a medicament for treating a disease, wherein the anti-PD-1 antibody comprises a heavy chain and a light chain, The heavy chain comprises the amino acid sequence of SEQ ID NO:248, and the light chain comprises the amino acid sequence of SEQ ID NO:249.

[0074] In another aspect, the combination of any one of the above anti-ILT4 antibodies and anti-PD-1 antibodies is for treating a disease, preferably wherein the disease is a tumor, more preferably wherein the disease is selected from the group consisting of astrocytoma (e.g., anaplastic astrocytoma), glioblastoma, bladder cancer, bone cancer, brain cancer, breast cancer (e.g., breast cancer characterized by BRCA1 and / or BRCA2 mutations), carcinoid, cervical cancer, choroid plexus papilloma, colorectal cancer (e.g., colon and rectal cancer), fallopian tube cancer, ependymoma, gallbladder cancer, gastric cancer, head and neck cancer, idiopathic myelofibrosis, kidney cancer or renal cell carcinoma (e.g., renal rhabdoid tumor and Wilms' tumor), renal pelvis tumor, leukemia, liver cancer (e.g., hepatocellular carcinoma), esophageal cancer ("esophageal cancer"), and the like. Cancer, also referred to as "cancer," is a disease selected from, for example, esophageal squamous cell carcinoma), lung cancer (e.g., non-small cell lung cancer and small cell lung cancer), lymphoma, medulloblastoma, melanoma, meningioma, Merkel cell carcinoma, mesothelioma, multiple myeloma, neuroblastoma, oligodendroglioma, ovarian cancer, peritoneal tumor, pancreatic cancer, polycythemia vera, primary neuroectodermal tumor, prostate cancer, retinoblastoma, salivary gland cancer, sarcoma (e.g., chondrosarcoma, Ewing's sarcoma, osteosarcoma, rhabdomyosarcoma, synovial sarcoma, and soft tissue sarcoma), small intestine cancer, colon cancer, squamous cell carcinoma (e.g., cutaneous squamous cell carcinoma), thyroid cancer, endometrial cancer, vestibular schwannoma, blastoma, teratoma, pituitary cancer, vulvar cancer, thymoma, testicular cancer, bile duct cancer, pheochromocytoma, paraganglioma, and adenoid cystic carcinoma. In some embodiments, the medicament is used to treat a disease selected from ovarian cancer, lung cancer, esophageal squamous cell carcinoma, colorectal cancer, renal cell carcinoma, and melanoma. [Brief explanation of the drawings]

[0075] [Figure 1A] 1 shows the blocking effect of the anti-ILT4 antibody huP2-E3 on ILT4 and its downstream signaling pathway. [Figure 1B] 1 shows the blocking effect of the anti-ILT4 antibody huP2-G9 on ILT4 and its downstream signaling pathway. [Figure 1C] 1 shows the blocking effect of anti-ILT4 antibody huP2-C6 on ILT4 and its downstream signaling pathway. [Figure 1D] 1 shows the blocking effect of the anti-ILT4 antibody P2m on ILT4 and its downstream signaling pathway. [Figure 2A] 1 shows the effect of the anti-ILT4 antibody huP2-G9 on the secretion of the cytokine TNF-α by macrophages. [Figure 2B] 1 shows the effect of the anti-ILT4 antibody huP2-C6 on the secretion of the cytokine TNF-α by macrophages. [Figure 2C] 1 shows the effect of the anti-ILT4 antibody huP2-E8 on the secretion of the cytokine TNF-α by macrophages. [Figure 2D] 1 shows the effect of the anti-ILT4 antibody P2m on the secretion of the cytokine TNF-α by macrophages. [Figure 3A] 1 shows the combined effect of anti-ILT4 antibody huP2-E3 and anti-PD-1 antibody in an allogeneic MLR. [Figure 3B] 1 shows the combined effect of anti-ILT4 antibody huP2-G9 and anti-PD-1 antibody in an allogeneic MLR. [Figure 3C] 1 shows the combined effect of anti-ILT4 antibody huP2-C6 and anti-PD-1 antibody in an allogeneic MLR. [Figure 3D] 1 shows the combined effect of anti-ILT4 antibody huP2-E8 and anti-PD-1 antibody in an allogeneic MLR. [Figure 3E] 1 shows the combined effect of anti-ILT4 antibody P2m and anti-PD-1 antibody in allogeneic MLR. DETAILED DESCRIPTION OF THE INVENTION

[0076] term In order that the present disclosure may be more readily understood, the following provides an explanation of some technical and scientific terms. Unless expressly defined otherwise herein, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art.

[0077] As used in the specification and claims, the singular forms "a," "an," and "the above" include plural referents unless the context clearly dictates otherwise.

[0078] Unless the context clearly indicates otherwise, in the patent specification and claims, the words "comprises," "has," "includes," and the like are to be understood in the sense of "including, but not limited to," rather than in the exclusive or exhaustive sense.

[0079] The term "cytokine" is a general term for proteins released by one cell population that act on other cells as intercellular mediators. Examples of such cytokines include lymphokines, monokines, chemokines, and traditional polypeptide hormones. Exemplary cytokines include mIL-2, IFNγ, TNFα, CCL-2, and IL-6.

[0080] The term "and / or" is intended to be inclusive. For example, the phrase "A, B and / or C" is intended to cover each of A, B and C, A, B or C, A or C, A or B, B or C, A and C, A and B, B and C, A (alone), B (alone), and C (alone).

[0081] The three-letter and one-letter codes for amino acids used in this disclosure are as described in J. Biol. Chem, 243, p. 3558 (1968).

[0082] The term "ILT4" refers to the full-length ILT4 protein, with human ILT4 having the amino acid sequence of SEQ ID NO: 3. The amino acid sequences of ILT4 molecules from non-human species (e.g., mouse, monkey, rabbit, dog, pig, etc.) are available from public sources.

[0083] The term "αPD-1" refers to an anti-PD-1 antibody.

[0084] The term "amino acid" refers to naturally occurring amino acids, synthetic amino acids, and amino acid analogs and amino acid mimetics that act in a manner similar to naturally occurring amino acids. Naturally occurring amino acids are those encoded by the genetic code and those that are later modified, such as hydroxyproline, γ-carboxyglutamate, and O-phosphoserine. Amino acid analogs refer to compounds that have the same basic chemical structure as a naturally occurring amino acid (i.e., an α-carbon bonded to a hydrogen, a carboxy group, an amino group, and an R group), such as homoserine, norleucine, methionine sulfoxide, and methionine methylsulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid. Amino acid mimetics refer to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that act in a manner similar to a naturally occurring amino acid.

[0085] The term "amino acid mutation" includes amino acid substitution (also called amino acid replacement), deletion, insertion, and modification. Any combination of substitution, deletion, insertion, and modification can be used to achieve the final construct, provided that the final construct possesses the desired properties, such as reduced binding to Fc receptors. Deletions and insertions in the amino acid sequence include deletions and insertions at the amino and / or carboxy termini of the polypeptide chain. A specific amino acid mutation may be an amino acid substitution. In one embodiment, the amino acid mutation is a non-conservative amino acid substitution, i.e., replacing one amino acid with another amino acid having different structural and / or chemical properties. Amino acid substitutions include replacement with non-naturally occurring amino acids or derivatives of the 20 naturally occurring amino acids (e.g., 4-hydroxyproline, 3-methylhistidine, ornithine, homoserine, 5-hydroxylysine). Amino acid mutations can be generated by genetic or chemical methods known in the art. Genetic methods may include site-directed mutagenesis, PCR, gene synthesis, etc. It is anticipated that methods for modifying amino acid side groups other than genetic engineering, such as chemical modification, may also be utilized. In this specification, the same amino acid mutation may be referred to by various names. In this specification, the amino acid residue at a specific position may be referred to by the format of position + amino acid residue, for example, 102S indicates that the amino acid residue at position 102 is S. C102S indicates that the amino acid residue at position 102 is mutated from the original C to S. When an amino acid sequence is defined in the claims by the format of position + residue, it should be understood that the amino acid at that position before mutation does not limit the technical solution.

[0086] The term "antibody" is used in the broadest sense and covers a variety of antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, monospecific antibodies, multispecific antibodies (e.g., bispecific antibodies), full-length antibodies, and antibody fragments (or antigen-binding fragments, or antigen-binding portions), provided they exhibit the desired antigen-binding activity.

[0087] A "native antibody" is a naturally occurring immunoglobulin molecule. For example, a native IgG antibody is a heterotetrameric glycoprotein of approximately 150,000 daltons, composed of two light chains and two heavy chains linked by disulfide bonds. From the N-terminus to the C-terminus, each heavy chain contains one variable region (VH), also called a variable heavy chain domain or heavy chain variable region, followed by a heavy chain constant region, which generally contains three constant domains (CH1, CH2, and CH3). Similarly, from the N-terminus to the C-terminus, each light chain contains one variable region (VL), also called a variable light chain domain or light chain variable domain, followed by a constant light chain domain (light chain constant region, CL). The terms "full-length antibody," "complete antibody," and "whole antibody" may be used interchangeably herein and refer to antibodies having a structure substantially similar to that of a native antibody or having heavy chains of the Fc region as defined herein. The light chain of a native intact antibody comprises a light chain variable region VL and a constant region CL, with VL at the amino terminus of the light chain, and the light chain constant region comprises a kappa chain and a lambda chain; the heavy chain comprises a variable region VH and constant regions (CH1, CH2, and CH3), with VH at the amino terminus of the heavy chain and the constant region at the carboxy terminus, of which CH3 is closest to the carboxy terminus of the polypeptide; and the heavy chain may belong to any isotype, including IgG (including IgG1, IgG2, IgG3, and IgG4 subtypes), IgA (including IgA1 and IgA2 subtypes), IgM, and IgE.

[0088] The term "variable region" or "variable domain" of an antibody refers to the domain in the heavy or light chain of the antibody that is involved in binding to an antigen. As used herein, the heavy chain variable region (VH) and light chain variable region (VL) of an antibody each contain four conserved framework regions (FR) and three complementarity-determining regions (CDR). The term "complementarity-determining region" or "CDR" refers to the region in the variable domain that primarily promotes antigen binding, and "framework" or "FR" refers to the variable domain residues excluding the CDR residues. VH contains three CDR regions, HCDR1, HCDR2, and HCDR3, while VL contains three CDR regions, LCDR1, LCDR2, and LCDR3. Each VH and VL consists of three CDRs and four FRs, arranged in the following order from the amino terminus (also referred to as the N-terminus) to the carboxy terminus (also referred to as the C-terminus): FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.

[0089] The boundaries of the amino acid sequences of CDRs can be determined by various known methods, such as the "Kabat" numbering convention (see Kabat et al. (1991), "Sequences of Proteins of Immunological Interest", 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD), the "Chothia" numbering convention, the "ABM" numbering convention, the "contact" numbering convention (see Martin, ACR. Protein Sequence and Structure Analysis of Antibody Variable Domains [J]. 2001), and the ImMunoGenTics (IMGT) numbering convention (Lefranc, MP et al., Dev. Comp. Immunol., 27, 55-77 (2003); Front Immunol. 2018 Oct 16; 9: 2278). The correspondence between various numbering systems is well known to those skilled in the art and is shown, for example, in Table 1 below. [Table 1]

[0090] Unless otherwise specified, the variable regions and CDRs in the examples of the present disclosure are all governed by the "Kabat" numbering convention. In specific embodiments, one numbering system (e.g., Kabat) is used to define amino acid residues, but technical solutions corresponding to other numbering systems are considered equivalent technical solutions.

[0091] The term "antibody fragment" refers to a molecule distinct from a complete antibody and includes a portion of the complete antibody that binds to the same antigen as the complete antibody. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab')2, single domain antibodies, single-chain Fab (scFab), diabodies, linear antibodies, single-chain antibodies (e.g., scFv), and multispecific antibodies composed of antibody fragments.

[0092] The term "Fc region" or "fragment crystallizable region" is used to define the C-terminal region of an antibody heavy chain, and includes native Fc regions and modified Fc regions. In some embodiments, the Fc region comprises two subunits, which may be the same or different. In some embodiments, the Fc region of a human IgG heavy chain is defined to extend from the amino acid residue at position Cys226, or from Pro230 to its carboxy terminus. Suitable Fc regions for use in the antibodies described herein include the Fc regions of human IgG1, IgG2 (IgG2A, IgG2B), IgG3, and IgG4. In some embodiments, the boundaries of the Fc region may be altered, for example, by deletion of the C-terminal lysine (residue 447 according to the EU numbering system) of the Fc region, or deletion of the C-terminal glycine and lysine (residues 446 and 447 according to the EU numbering system) of the Fc region. Unless otherwise specified, the numbering convention for the Fc region is the EU numbering system, also known as the EU index.

[0093] The term "chimeric" antibody refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, while the remainder of the heavy and / or light chain is derived from another, different source or species.

[0094] The term "humanized" antibody refers to an antibody that retains the reactivity of a non-human antibody while having relatively low immunogenicity in humans, which may be achieved, for example, by retaining the non-human CDR regions and replacing the remainder of the antibody with their human counterparts (i.e., the constant and variable region framework region portions).

[0095] The terms "human antibody," "humanized antibody," "fully human antibody," and "fully human antibody" may be used interchangeably and refer to antibodies in which the variable and constant regions are human sequences. The term covers antibodies derived from human genes but with altered sequences, e.g., to reduce potential immunogenicity, increase affinity, or remove cysteines or glycosylation sites that may cause undesired folding. The term also covers antibodies produced recombinantly in non-human cells (which may confer glycosylation not characteristic of human cells). The term also covers antibodies produced in transgenic mice containing some or all human immunoglobulin heavy and light chain loci. The term specifically excludes humanized antibodies.

[0096] The term "affinity" refers to the overall strength of non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding ligand (e.g., an antigen). Unless otherwise specified, as used herein, binding "affinity" refers to internal binding affinity, reflecting a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen). The affinity of a molecule X for its ligand Y can generally be expressed as a dissociation constant (KD). Affinity can be measured by conventional methods known in the art, including those described herein.

[0097] As used herein, the terms "kassoc" or "ka" refer to the association rate of a particular antibody-antigen interaction, and the terms "kdis" or "kd" refer to the dissociation rate of a particular antibody-antigen interaction. The term "KD" refers to the dissociation constant, which is obtained from the ratio of kd to ka (i.e., kd / ka) and is expressed as a molar concentration (M). The KD value of an antibody can be measured by methods known in the art. For example, the affinity can be measured by a biosensor system such as a surface plasmon resonance (e.g., Biacore) system, or by measuring the affinity in solution by solution equilibrium titration (SET).

[0098] The term "surface plasmon resonance" refers to an optical phenomenon that analyzes real-time interactions by detecting changes in protein concentration in a biosensor matrix, for example, using a BIAcore™ system (Biacore LifeSciences division of GE Healthcare, Piscataway, NJ).

[0099] The term "effector function" refers to a biological activity that can be attributed to an antibody Fc region (a native sequence Fc region or an amino acid sequence mutated Fc region) and varies depending on the antibody isotype. Examples of antibody effector functions include, but are not limited to, C1q binding and complement-dependent cytotoxicity, Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, down-regulation of cell surface receptors (e.g., B cell receptors), and activation of B cells.

[0100] The term "monoclonal antibody" refers to a population of essentially homogeneous antibodies, i.e., the amino acid sequences of the antibody molecules within the population are the same except for possible minor natural mutations. In contrast, polyclonal antibodies typically include a plurality of different antibodies with different amino acid sequences in their variable domains, which are generally specific for different epitopes. "Monoclonal" should not be construed as requiring the antibody to be produced by any particular method. In some embodiments, the antibodies provided by the present disclosure are monoclonal antibodies.

[0101] The term "antigen" refers to a molecule or portion of a molecule that can be selectively bound by, for example, an antigen-binding protein (including, for example, an antibody). An antigen may have one or more epitopes that can interact with different antigen-binding proteins (e.g., antibodies).

[0102] The term "epitope" refers to an area or region on an antigen capable of specific binding to an antibody or antigen-binding fragment thereof. An epitope can be formed by a contiguous string of amino acids (linear epitope) or can include non-contiguous amino acids that are spatially adjacent, for example, due to folding of the antigen (conformational epitope). Conformational and linear epitopes differ in that antibody binding to conformational epitopes is lost in the presence of denaturing solvents. An epitope comprises at least 3, at least 4, at least 5, at least 6, at least 7, or 8-10 amino acids in a unique spatial conformation. Screening for antibodies that bind to a specific epitope (i.e., antibodies that bind to the same epitope) can be performed by conventional methods in the art, including, but not limited to, alanine scanning, peptide blotting, peptide cleavage analysis, epitope excision, epitope extraction, chemical modification of the antigen (see Prot. Sci. 9 (2000) 487-496), and cross-blocking.

[0103] The terms "capable of specifically binding," "specifically binds," or "binding" refer to the ability of an antibody to bind to an antigen or epitope thereof with greater affinity than to other antigens or epitopes. Generally, antibodies bind to antigens with a greater affinity than other antigens or epitopes. -7 M or less (e.g., about 1 × 10 -8 M, 1 x 10 -9 M, 1 x 10 -10 M, 1 x 10 -11 An antibody binds to an antigen or its epitope with an equilibrium dissociation constant (KD) of M or less. In some embodiments, the KD of an antibody for binding to an antigen is 10% or less (e.g., 1%) of the KD for binding to a non-specific antigen (e.g., BSA, casein). KD can be measured by known methods, for example, by BIACORE® surface plasmon resonance assay. However, antibodies that specifically bind to an antigen or its epitope may be cross-reactive with other related antigens, for example, with corresponding antigens from other species (homologs) (e.g., humans or monkeys such as cynomolgus monkeys (Macaca fascicularis) (cynomolgus, cyno), chimpanzees (Pan troglodytes) (chimpanzee, chimp), or common marmosets (Callithrix jacchus) (marmoset)).

[0104] The terms "anti-ILT4 antibody" and "antibody that binds ILT4" refer to an antibody that is capable of binding to ILT4 or an epitope thereof with sufficient affinity.

[0105] The terms "antibody-dependent cellular cytotoxicity," "antibody-dependent cell-mediated cytotoxicity," or "ADCC" refer to a mechanism of cell death that relies on the interaction of antibody-coated target cells with lytic effector cells (e.g., natural killer cells (NK), monocytes, macrophages, and neutrophils) via Fcγ receptors (FcγR) expressed on the effector cells. For example, NK cells express FcγRIIIa, while monocytes express FcγRI, FcγRII, and FcγRIIIa. The ADCC activity of the antibodies provided herein can be assessed in vitro using antigen-expressing cells as target cells and NK cells as effector cells. Cytolysis is detected by a marker (e.g., a radioactive substrate, a fluorescent dye, or a natural intracellular protein) released from the lysed cells.

[0106] The term "antibody-dependent cellular phagocytosis (ADCP)" refers to the mechanism by which antibody-coated target cells are removed by internalization by phagocytes (eg, macrophages or dendritic cells).

[0107] The term "complement-dependent cytotoxicity" or "CDC" refers to a mechanism for inducing cell death in which the Fc effector domain of a target-binding antibody binds to and activates complement component C1q, which further activates the complement cascade, resulting in the death of the target cell. Complement activation can also lead to the deposition of complement components on the target cell surface, which promote CDC by binding to complement receptors (e.g., CR3) on leukocytes.

[0108] The term "nucleic acid" may be used interchangeably with the term "polynucleotide" herein and refers to deoxyribonucleotides or ribonucleotides and polymers thereof in single- or double-stranded form. The term covers nucleic acids containing known nucleotide analogs or modified backbone residues or linkers, including synthetic, naturally occurring, and non-naturally occurring nucleic acids, which have similar binding properties as the reference nucleic acid and are metabolized in a manner similar to the reference nucleotide. Examples of such analogs include, but are not limited to, phosphorothioates, phosphoramidates, methyl phosphonates, chiral-methyl phosphonates, 2-O-methyl ribonucleotides, and peptide-nucleic acids (PNAs).

[0109] An "isolated" nucleic acid refers to a nucleic acid molecule separated from a component of its natural environment. An isolated nucleic acid encoding a polypeptide refers to one or more nucleic acid molecules encoding a polypeptide, including one or more such nucleic acid molecules in a single vector or separate vectors, and one or more such nucleic acid molecules present in one or more locations in a host cell. Unless otherwise specified, a particular nucleic acid sequence implicitly covers its conservatively modified variants (e.g., degenerate codon substitutions) and complementary sequences, as well as the explicitly stated sequence. Specifically, as detailed below, degenerate codon substitutions can be obtained by generating sequences in which the third position of one or more selected (or all) codons is substituted with mixed-base and / or deoxyinosine residues.

[0110] The terms "polypeptide" and "protein" may be used interchangeably herein to refer to a polymer of amino acid residues. The terms apply to amino acid polymers in which one or more amino acid residues are artificial chemical mimetics of corresponding naturally occurring amino acids, and apply to naturally occurring and non-naturally occurring amino acid polymers. Unless otherwise specified, a particular polypeptide sequence also implicitly covers conservatively modified variants thereof.

[0111] The term "sequence identity" refers to the degree (percentage) of homology between two sequences at equivalent amino / nucleic acid positions when the two sequences are optimally aligned, introducing gaps as necessary to achieve the maximum percentage of sequence identity, and not counting any conservative substitutions as part of the sequence identity. To measure the percentage of sequence identity, alignment can be achieved using techniques known in the art, such as publicly available computer software such as BLAST, BLAST-2, ALIGN, ALIGN-2, or Megalign (DNASTAR) software. Those skilled in the art can determine the parameters to be applied for measurement and alignment, including any algorithms necessary to achieve maximum alignment over the entire length of the sequences being compared.

[0112] The term "vector" refers to a polynucleotide molecule capable of transporting another polynucleotide linked to it. One type of vector is a "plasmid," which refers to a circular double-stranded DNA circle that can be ligated to an additional DNA segment. Another type of vector is a viral vector, such as an adeno-associated viral vector (AAV or AAV2), in which another DNA segment can be ligated to the viral genome. Some vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors with a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) can integrate into the genome of a host cell after introduction and thereby replicate along with the host genome. The term "expression vector" or "expression construct" refers to a vector capable of transforming a host cell and containing nucleic acid sequences that direct and / or control (with the host cell) the expression of one or more heterologous coding regions operably linked thereto. Expression constructs may include, but are not limited to, sequences that affect or control transcription, translation, and, when present, RNA splicing of the coding region operably linked to it.

[0113] The terms "host cell," "host cell line," and "host cell culture" may be used interchangeably and refer to cells into which exogenous nucleic acid has been introduced, including the progeny of such cells. Host cells include "transformants" and "transformed cells," and include the original transformed cell and its derived progeny, without regard to the number of passages. Progeny may not be completely identical in nucleic acid content to the parent cell, but may contain mutations. Included herein are mutant progeny that have the same function or biological activity as the cells screened or selected for in the original transformed cell. Host cells include prokaryotic and eukaryotic host cells, of which eukaryotic host cells include, but are not limited to, mammalian cells, insect cell lines, plant cells, and fungal cells. Mammalian host cells include human, mouse, rat, dog, monkey, pig, goat, bovine, horse, and hamster cells, including, but not limited to, Chinese hamster ovary (CHO) cells, NSO, SP2 cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS), human hepatocellular carcinoma cells (e.g., Hep G2), A549 cells, 3T3 cells, and HEK-293 cells.Fungal cells include yeast and filamentous fungal cells, such as Pichia pastoris, Pichia finlandica, Pichia trehalophila, Pichia koclamae, Pichia membranaefaciens, Pichia minuta (Ogataea minuta, Pichia lindneri), Pichia opuntiae, Pichia thermotolerans, Pichia salictaria, Pichia guercuum, Pichia pijperi, and Pichia stiptis. stiptis, Pichia methanolica, Pichia spp., Saccharomyces cerevisiae, Saccharomyces spp., Hansenula polymorpha, Kluyveromyces spp., Kluyveromyces lactis, Candida albicans, Aspergillus nidulans, Aspergillus niger, Aspergillus oryzae, Trichoderma reesei, Chrysosporium lucknowense, Fusarium sp., Fusarium gramineum gramineum, Fusarium venenatum, Physcomitrella patens, and Neurospora crassa.Including Pichia, any Saccharomyces, Hansenula polymorpha, any Kluyveromyces, Candida albicans, any Aspergillus, Trichoderma reesei, Chrysosporium lucknowense, any Fusarium, Yarrowia lipolytica, and Neurospora crassa.

[0114] "Optionally" or "optionally" means that the subsequently described event or circumstance may occur, but need not occur, and the description includes cases where the event or circumstance occurs and cases where it does not occur.

[0115] The term "pharmaceutical composition" refers to a mixture comprising one or more anti-ILT4 antibodies described herein and other chemical components, such as physiologically / pharmaceutically acceptable carriers and excipients.

[0116] The term "pharmaceutically acceptable carrier, diluent, or excipient" refers to an ingredient in a pharmaceutical formulation that is different from the active ingredient and is non-toxic to a subject. Pharmaceutically acceptable carriers, diluents, or excipients include, but are not limited to, buffers, fillers, stabilizers, or preservatives.

[0117] The term "subject" or "individual" includes humans and non-human animals. Non-human animals include all vertebrates (e.g., mammals and non-mammals), such as non-human primates, sheep, dogs, cows, chickens, amphibians, and reptiles. Unless indicated otherwise, the terms "patient" or "subject" may be used interchangeably herein. In some embodiments, the individual or subject is a human.

[0118] "Administration" or "giving," as applied to an animal, human, experimental subject, cell, tissue, organ, or biological fluid, means contacting an exogenous agent, therapeutic agent, diagnostic agent, or composition with the animal, human, subject, cell, tissue, organ, or biological fluid.

[0119] The term "sample" refers to a collection (e.g., fluid, cell, or tissue) isolated from a subject, as well as fluids, cells, or tissues present within a subject. Exemplary samples are biological fluids, such as blood, serum and serous fluid, plasma, lymph, urine, saliva, cyst fluid, tears, feces, sputum, mucosal secretions of secretory tissues or organs, vaginal secretions, ascites, pleura, pericardium, peritoneum, fluids of the abdominal cavity and other body cavities, fluids collected from bronchial washings, synovial fluid, liquid solutions that have come into contact with a subject or biological source, such as culture media (including conditioned media), lavage fluids, tissue biopsies, fine needle aspirates, surgically resected tissues, organ cultures, or cell cultures.

[0120] "Treatment" and "treatment" (and grammatical variations thereof) refer to clinical intervention in the individual being treated and can be performed prophylactically or during the clinical pathological process. Desired effects of treatment include, but are not limited to, prevention of disease onset or recurrence, alleviation of symptoms, reduction / reduction of any direct or indirect pathological consequences of the disease, prevention of metastasis, slowing of disease progression, amelioration or alleviation of the disease state, and regression or improvement of prognosis. In some embodiments, the antibodies of the present disclosure are used to delay the formation of disease or slow the progression of disease.

[0121] An "effective amount" is generally an amount sufficient to reduce the severity and / or frequency of symptoms, eliminate those symptoms and / or their underlying cause, prevent the onset of symptoms and / or their underlying cause, and / or ameliorate or ameliorate damage caused by or associated with a disease state (e.g., pulmonary disease). In some embodiments, the effective amount is a therapeutically effective amount or a prophylactically effective amount.

[0122] A "therapeutically effective amount" is an amount sufficient to treat a disease state or symptom, particularly a condition or symptom associated with the disease state, or to prevent, inhibit, delay, or reverse the progression of the disease state or any other undesirable symptom associated in any way with the disease, in other ways. A "prophylactically effective amount" is an amount that, when administered to a subject, provides a desired prophylactic effect, such as preventing or delaying the onset (or recurrence) of the disease state, or reducing the likelihood of the onset (or recurrence) of the disease state or associated symptoms. Complete treatment or prevention does not necessarily occur after administration of a single dose, but may occur after administration of a series of doses. Thus, a therapeutically or prophylactically effective amount can be administered in one or multiple doses. "Therapeutically effective amounts" and "prophylactically effective amounts" can vary depending on various factors, such as the individual's disease state, age, sex, and weight, and the ability of the therapeutic agent or combination of therapeutic agents to elicit a desired response in an individual. Exemplary indicators of an effective therapeutic agent or combination of therapeutic agents include, for example, improved health status of the patient.

[0123] The term "immune checkpoint" refers to a group of molecules on the cell surface of CD4 T cells and CD8 T cells. These molecules can effectively function as "brakes" that downregulate or inhibit antitumor immune responses. Immune checkpoint molecules include, but are not limited to, programmed death-1 (PD-1), cytotoxic T-lymphocyte antigen 4 (CTLA-4), B7H1, B7H4, OX-40, CD137, CD40, and LAG-3, which directly inhibit immune cells.

[0124] The immune checkpoint inhibitor used in the present disclosure is a substance that inhibits the function of an immune checkpoint molecule. Immune checkpoint inhibitors are not particularly limited, as long as they are capable of inhibiting the function (signal) of an immune checkpoint molecule. Immune checkpoint inhibitors that can be used in the methods of the present disclosure include, but are not limited to, inhibitors of PD-1, PD-L2, CTLA-4, TIM-3, LAG-3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4, CEACAM (e.g., CEACAM-1, CEACAM-3, and / or CEACAM-5), and / or TGFRβ. Inhibition of an inhibitory molecule can be achieved by inhibition at the DNA, RNA, or protein level. In embodiments, inhibitory nucleic acids (e.g., dsRNA, siRNA, or shRNA) can be used to inhibit the expression of an inhibitory molecule. In other embodiments, the inhibitor of an inhibitory signal is a polypeptide that binds to the inhibitory molecule, such as a soluble ligand or an antibody. Examples of immune checkpoint inhibitors used in the present disclosure include, but are not limited to, anti-PD-1 antibodies, anti-PD-L1 antibodies, and anti-CTLA-4 antibodies, and preferably, anti-PD-1 antibodies.

[0125] target molecule The term "ILT4" should be understood broadly and is intended to cover various forms of ILT4 molecules at various stages in the mammalian body, including, but not limited to, molecules produced during the amplification, replication, transcription, splicing, processing, translation, and modification processes of the ILT4 gene (e.g., precursor ILT4, mature ILT4, membrane-expressed ILT4, ILT4 splice variants, modified ILT4, or fragments thereof), and the term also covers artificially prepared or in vitro expressed ILT4.

[0126] Disclosed anti-ILT4 antibodies The present disclosure provides anti-ILT4 antibodies with many advantageous properties, such as good in vitro killing activity, therapeutic activity, safety, pharmacokinetic properties, and druggability (e.g., yield, purity, stability, etc.).

[0127] Exemplary Anti-ILT4 Antibodies Examples of the present disclosure disclose the antibody series P1-B4, P1-B9, P1-D10, P2-E8, P2-C10, P2-F4, P2-B3, P2-E7, P2-G4, P2-C4, P2-E11, P1-F7, P2-E3, P2-F10, P2-G9, P1-C4, P1-G7, P2-C7, P2-C6, P1-D7, P1-E11, P2-B9, P2-C2, and P2-E10. The antibodies of the present disclosure are described below using antibodies P2-G9 and P2-C6 as examples.

[0128] Exemplary embodiments of the present disclosure include an anti-ILT4 antibody comprising a heavy chain variable region and a light chain variable region, wherein: a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 184 or 185, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 187 or 186, respectively; or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 32, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 33, respectively; or b. The HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 211, 205, 206, 207, 208, 209, or 210, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in any one of SEQ ID NOs: 212, 213, or 214, or the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 40, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 41.

[0129] Exemplary embodiments of the present disclosure include an anti-ILT4 antibody comprising a heavy chain variable region and a light chain variable region, wherein: a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 184, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 187; or b. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 211, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 212, respectively.

[0130] Exemplarily, in the anti-ILT4 antibodies of the present disclosure, the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and the LCDR1, LCDR2, and LCDR3 of the light chain variable region are defined according to a similar numbering convention selected from Kabat, IMGT, Chothia, AbM, and Contact.

[0131] Exemplary embodiments of the present disclosure include an anti-ILT4 antibody comprising a heavy chain variable region and a light chain variable region, wherein: In the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 121, and in the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119.

[0132] Exemplary embodiments of the present disclosure include an anti-ILT4 antibody comprising a heavy chain variable region and a light chain variable region, wherein: the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of SEQ ID NO: 203, 136, 201, 202 or 204, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; Preferably, the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of SEQ ID NO: 203, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140.

[0133] Exemplarily, in the anti-ILT4 antibodies of the present disclosure, HCDR1, HCDR2 and HCDR3 of the heavy chain variable region and LCDR1, LCDR2 and LCDR3 of the light chain variable region are defined according to the Kabat numbering convention.

[0134] Exemplary anti-ILT4 antibodies according to the present disclosure include those comprising: a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 260, HCDR2 comprising the amino acid sequence of SEQ ID NO: 261, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 262, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 263, LCDR2 comprising the amino acid sequence of SEQ ID NO: 264, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 267, HCDR2 comprising the amino acid sequence of SEQ ID NO: 268, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 269, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 270, LCDR2 comprising the amino acid sequence of SEQ ID NO: 271, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; Among them, the CDR regions are determined by the IMGT numbering convention.

[0135] Exemplary anti-ILT4 antibodies according to the present disclosure include those comprising: the heavy chain variable region, HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 260, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 261, and HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 262; and the light chain variable region, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 263, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 264, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 119; Among them, the CDR regions are determined by the IMGT numbering convention.

[0136] Exemplary anti-ILT4 antibodies according to the present disclosure include those comprising: a. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 265, HCDR2 comprising the amino acid sequence of SEQ ID NO: 266, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 272, HCDR2 comprising the amino acid sequence of SEQ ID NO: 273, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; Among them, the CDR regions are determined by the Chothia numbering convention.

[0137] Exemplary anti-ILT4 antibodies according to the present disclosure include those comprising: the heavy chain variable region, HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 265, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 266, and HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 121; and the light chain variable region, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 114, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 119; Among them, the CDR regions are determined by the Chothia numbering convention.

[0138] Exemplary anti-ILT4 antibodies of the present disclosure are murine, chimeric, humanized, or fully human. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the antibody is a fully human antibody. In some embodiments, the antibody is a humanized antibody.

[0139] Exemplary, an anti-ILT4 antibody according to the present disclosure, which comprises a framework region (FR) of a human antibody.

[0140] Exemplary anti-ILT4 antibodies according to the present disclosure include those comprising: a. the heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 184, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 187; or b. The heavy chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:211, and the light chain variable region comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO:212.

[0141] Exemplary embodiments of the anti-ILT4 antibody of the present disclosure include a heavy chain variable region having FR1, FR2, and FR3 derived from IGHV4-31*01 and FR4 derived from IGHJ1*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 1E, 27S, 30T, 44K, 71R, and 76S, and / or a light chain variable region having FR1, FR2, and FR3 derived from IGKV1-39*01 and FR4 derived from IGKJ4*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 43S, 48V, and 71Y. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region having HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, HCDR2 comprising the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 121, and the FR of the heavy chain variable region comprises one or more amino acid substitutions selected from the group consisting of 1E, 27S, 30T, 44K, 71R, and 76S; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 119, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 43S, 48V, and 71Y. In some embodiments, the variable region and CDRs are defined according to the Kabat numbering convention.

[0142] Exemplary embodiments of the anti-ILT4 antibody of the present disclosure include a heavy chain variable region having FR1, FR2, and FR3 derived from IGHV1-18*01 and FR4 derived from IGHJ6*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 1E, 43E, 69L, 71V, 73K, 76N, and 93T, and / or a light chain variable region having FR1, FR2, and FR3 derived from IGKV4-1*01 or IGKV3-11*01 and FR4 derived from IGKJ4*01, which is unsubstituted or has one or more amino acid substitutions selected from the group consisting of 4L, 43P, 45K, 58I, 68R, and 85T. In some embodiments, the anti-ILT4 antibody comprises the heavy chain variable region having HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, HCDR2 comprising the amino acid sequence of SEQ ID NO: 203, 136, 201, 202, or 204, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the FR of the heavy chain variable region comprises one or more amino acid substitutions selected from the group consisting of 1E, 43E, 69L, 71V, 73K, 76N, and 93T; and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 140, and the FR of the light chain variable region comprises one or more amino acid substitutions selected from the group consisting of 4L, 43P, 45K, 58I, 68R, and 85T. In some embodiments, the variable region and CDRs are defined according to the Kabat numbering convention.

[0143] Exemplary anti-ILT4 antibodies of the present disclosure include those in which the anti-ILT4 antibody is an antibody fragment, and in some embodiments, the antibody fragment is selected from Fab, Fab', F(ab')2, Fd, Fv, scFv, dsFv, or dAb.

[0144] Exemplary embodiments include an anti-ILT4 antibody according to the present disclosure, which comprises a heavy chain constant region and a light chain constant region. In some embodiments, the heavy chain constant region is a human IgG1 or IgG4 heavy chain constant region. In some embodiments, the heavy chain constant region has 234A and 235A mutations, wherein the mutations are numbered according to the EU index. In some embodiments, the light chain constant region is a human κ light chain constant region. In some embodiments, the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 240 or 171, and the light chain constant region comprises the amino acid sequence of SEQ ID NO: 172.

[0145] Exemplary embodiments of the present disclosure provide an anti-ILT4 antibody, the anti-ILT4 antibody comprising a heavy chain and a light chain, wherein: a. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 228, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98%, or 99%) sequence identity to SEQ ID NO: 229; or b. the heavy chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO:234, and the light chain comprises an amino acid sequence having at least 70% (e.g., at least 70%, 80%, 85%, 90%, 95%, 98% or 99%) sequence identity to SEQ ID NO:235.

[0146] Exemplary embodiments of the present disclosure provide an anti-ILT4 antibody, the anti-ILT4 antibody comprising a heavy chain and a light chain, wherein: a. the heavy chain comprises the amino acid sequence of SEQ ID NO: 228 and the light chain comprises the amino acid sequence of SEQ ID NO: 229, or b. the heavy chain comprises the amino acid sequence of SEQ ID NO:234, and the light chain comprises the amino acid sequence of SEQ ID NO:235.

[0147] Illustratively, the present disclosure further provides an isolated anti-ILT4 antibody that binds to human ILT4 or an epitope thereof in competition with any one of the anti-ILT4 antibodies described above.

[0148] Illustratively, the anti-ILT4 antibodies provided in the present disclosure specifically bind to human ILT4 but do not bind to other members of the LILR family, ILT2, ILT3, LILRB5, LILRA1, LILRA2, LILRA3, LILRA4, LILRA5 and / or LILRA6.

[0149] Illustratively, an isolated anti-ILT4 antibody according to the present disclosure has a higher binding ability to ILT4 protein than a control antibody. Specifically, an isolated anti-ILT4 antibody according to the present disclosure has a binding ability to ILT4 protein of 1×10 -8 Less than M (e.g., 8 × 10 -9 Less than M, 5 x 10 -9 Less than M, 2 x 10 -9 Less than M, 1 x 10 -9 Less than M, 8 x 10 -10 Less than M, 5 x 10 -10 Less than M, 2 x 10 -10 Less than M, 1 x 10 -10 Less than M, 8 x 10 -11 Less than M, 5 x 10 -11 It binds to human ILT4 with a KD value of less than 1 M, as measured by surface plasmon resonance (Biacore).

[0150] Illustratively, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively relieve the inhibitory effect of ILT4 and promote the secretion of higher levels of mIL-2 by 3A9 cells.

[0151] Illustratively, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively promote the conversion of macrophages to the M1 phenotype and secrete high levels of the pro-inflammatory cytokine TNF-α.

[0152] Illustratively, the humanized anti-ILT4 antibodies provided in the present disclosure (e.g., huP2-G9) can induce higher levels of pro-inflammatory cytokine or chemokine secretion by dendritic cells in an immunosuppressive environment in the presence of IL-10.

[0153] Illustratively, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively block the binding of ILT4 to HLA-G.

[0154] Illustratively, the humanized anti-ILT4 antibodies provided in the present disclosure can effectively block the binding of ILT4 to HLA-G, and the blocking activity is stronger than that of the control antibodies MK and JTX.

[0155] Antibody structure In certain embodiments, the antibodies provided herein are full-length antibodies.

[0156] In certain embodiments, the antibodies provided herein are antibody fragments.

[0157] In one embodiment, the antibody fragment is a Fab, Fab', Fab'-SH, or F(ab')2 fragment, particularly a Fab fragment. "Fab" is a monovalent fragment consisting of the VL, VH, CL, and CH1 domains. "Fab fragments" may be produced by lysing an antibody with papain. "Fab'" contains the VL, CL, and VH and CH1 domains, and also contains the region between the CH1 and CH2 domains, allowing interchain disulfide bonds to form between the two heavy chains of two Fab' fragments to form F(ab')2 molecules. "Fab'-SH" is a Fab' fragment in which cysteine ​​residues in the constant regions have free sulfhydryl groups. "F(ab')2" is a bivalent fragment containing two Fab fragments linked by disulfide bonds at the hinge region.

[0158] In another embodiment, the antibody fragment is a diabody, a triabody, or a tetrabody. A diabody is an antibody fragment having two antigen-binding sites, the fragment comprising a VH and a VL linked in the same polypeptide chain (VH-VL). Using a linker that is too short prevents pairing between the two domains on the same chain, forcing them to pair with complementary domains on another chain, thereby generating two antigen-binding sites, where the two antigens may be the same or different.

[0159] In another embodiment, the antibody fragment is a single-chain Fab fragment. A "single-chain Fab fragment" or "scFab" is a polypeptide consisting of VH, CH1, VL, CL, and a linker, wherein the antibody domains and linker are arranged in one of the following orders from N-terminus to C-terminus: a) VH-CH1-linker-VL-CL, b) VL-CL-linker-VH-CH1, c) VH-CL-linker-VL-CH1, or d) VL-CH1-linker-VH-CL. In one embodiment, the linker is a polypeptide having at least 30 amino acids. In another embodiment, the linker is a polypeptide having 32 to 50 amino acids. The single-chain Fab fragment is stabilized by a native disulfide bond between CL and CH1. Additionally, these single-chain Fab molecules can be further stabilized by inserting cysteine ​​residues (e.g., position 44 in the heavy chain variable region and position 100 in the light chain variable region, Kabat numbering) to create interchain disulfide bonds.

[0160] In another embodiment, the antibody fragment is an Fv fragment consisting of the VH and VL domains of a single arm of an antibody.

[0161] In another embodiment, the antibody fragment is a single-chain variable fragment (scFv). An "scFv" is a fusion protein comprising at least one antibody fragment comprising a light chain variable region and at least one antibody fragment comprising a heavy chain variable region, wherein the light chain variable region and the heavy chain variable region are linked by a short, flexible peptide linker, and can be expressed as a single-chain polypeptide, and wherein the scFv retains the specificity of the intact antibody from which it is derived. Unless otherwise specified, herein, an scFv may have the VL and VH variable regions in any order; for example, relative to the N-terminus and C-terminus of the polypeptide, the scFv may comprise VL-linker-VH or VH-linker-VL.

[0162] In another embodiment, the antibody fragment is a dsFv, which is obtained by linking polypeptides in which one amino acid residue in each of VH and VL is substituted with a cysteine ​​residue via a disulfide bond between the cysteine ​​residues. The amino acid residue substituted with the cysteine ​​residue can be selected based on prediction of the three-dimensional structure of an antibody by a known method (Protein Engineering. 7:697 (1994)).

[0163] In another embodiment, the antibody fragment is a single domain antibody (dAb). A single domain antibody is an antibody fragment that comprises all or part of the heavy chain variable domain, or all or part of the light chain variable domain of an antibody.

[0164] In certain embodiments, the antibodies provided herein are chimeric antibodies. In one example, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non-human primate such as a monkey) and a human constant region. In another example, a chimeric antibody is a "class-switched" antibody in which the class or subclass has been changed from that of the parent antibody.

[0165] In certain embodiments, the antibody is a humanized antibody. Humanization of a non-human antibody typically reduces its immunogenicity to humans while retaining the specificity and affinity of the parent non-human antibody. Generally, a humanized antibody comprises one or more variable regions in which CDRs or portions thereof are derived from a non-human antibody and FRs or portions thereof are derived from a human antibody. Optionally, a humanized antibody may further comprise a portion of a human constant region. In some embodiments, some FR residues in a humanized antibody may be replaced with corresponding residues from a non-human antibody (e.g., the antibody providing the CDR sequences).

[0166] Humanized antibodies and methods for their production are generally described in, for example, Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008), and further described in Riechmann et al., Nature 332:323-329 (1988), Queen et al., Proc. Nat'l Acad. Sci. USA 86:10029-10033 (1989), U.S. Patent Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409, Kashmiri et al., Methods 36:25-34 (2005) (describing grafting of specificity determining regions (SDRs)), Padlan, Mol. Immunol. 28:489-498 (1991) (describing "resurfacing"), Dall'Acqua et al., Methods 36:43-60 (2005) (describing "FR shuffling"), and Osbourn et al., Methods 36:61-68 (2005) and Klimka et al., Br. J. Cancer 83:252-260 (2000) (describing "guided selection" methods of FR shuffling).

[0167] Human framework regions that can be used for humanization include framework regions selected by the "best-fit" method (see, e.g., Sims et al., J. Immunol. 151:2296 (1993)), framework regions derived from consensus sequences of human antibodies of a particular subclass of light chain variable region or heavy chain variable region (see, e.g., Carter et al., Proc. Natl. Acad. Sci. USA, 89:4285 (1992) and Presta et al., J. Immunol., 151:2623 (1993)), human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008)), and framework regions obtained by screening FR libraries (see, e.g., Baca et al., J. Immunol. 151:2623 (1993)). al., J. Biol. Chem. 272:10678-10684 (1997) and Rosok et al., J. Biol. Chem. 271:22611-22618 (1996)).

[0168] Anti-ILT4 antibody variants In certain embodiments, amino acid sequence variants of the anti-ILT4 antibodies provided herein are provided. For example, these variants may be expected to improve the binding affinity and / or other biological properties of the antibody. Amino acid sequence variants of the antibody can be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody or by peptide synthesis. Such modifications include, for example, deletions, insertions, and / or substitutions of residues in the amino acid sequence of the anti-ILT4 antibody. Any combination of deletions, insertions, and substitutions can be made to obtain the final construct, provided that the final construct possesses the desired properties, such as antigen-binding properties.

[0169] Mutant substitutions, insertions and deletions In one embodiment, antigen variants with one or more amino acid substitutions are provided. Sites of interest for substitutional mutagenesis include CDRs and FRs. Conservative substitutions are shown in Table 2 under the heading "Preferred Substitutions." More substantial changes are provided in Table 2 under the heading "Exemplary Substitutions," and are as further described below with reference to amino acid side chain classifications. Amino acid substitutions can be introduced into an antibody of interest, and the products screened for a desired activity, such as, for example, retained / improved antigen binding, reduced immunogenicity, or improved ADCC or CDC. [Table 2]

[0170] Amino acids can be grouped according to common side chain properties as follows: (1) Hydrophobic: Norleucine, Met, Ala, Val, Leu, Ile, (2) Neutral, hydrophilic: Cys, Ser, Thr, Asn, Gln, (3) Acidic: Asp, Glu, (4) Basic: His, Lys, Arg, (5) Residues that affect chain orientation: Gly, Pro, (6) Aromatic: Trp, Tyr, Phe.

[0171] Non-conservative substitutions involve replacing a member in one of these classes with a member in another class.

[0172] One type of substitutional variant involves substituting one or more CDR residues of a parent antibody (e.g., a humanized or human antibody). Generally, the resulting variant selected for further study has altered (e.g., improved) biological properties relative to the parent antibody (e.g., improved affinity, decreased immunogenicity) and / or essentially retains certain biological properties of the parent antibody. One exemplary substitutional variant is an affinity-matured antibody, which can be readily produced, for example, by phage display-based affinity maturation techniques (e.g., techniques such as those described herein). Briefly, one or more CDR residues are mutated, the mutant antibodies are displayed on phage, and screened for a particular biological activity (e.g., binding affinity). For example, changes (e.g., substitutions) can be made in the CDRs to improve antibody affinity. Such changes can be made to CDR "hot spots," i.e., residues encoded by codons frequently mutated during somatic maturation and / or residues that contact the antigen, and the resulting variant VH or VL can be tested for binding affinity. In some affinity maturation embodiments, diversity is introduced into the variable genes selected for maturation by any of a variety of methods (e.g., error-prone PCR, chain shuffling, or oligonucleotide-directed mutagenesis). A secondary library is then generated. The library is then screened to identify any antibody variants with the desired affinity. Another method for introducing diversity involves CDR-oriented methods, in which several CDR residues (e.g., 4-6 residues at a time) are randomized. For example, alanine scanning mutagenesis or modeling can specifically identify CDR residues involved in antigen binding. HCDR3 and LCDR3 are particularly commonly targeted.

[0173] In certain embodiments, substitutions, insertions, or deletions may be made in one or more CDRs, so long as such changes do not substantially reduce the ability of the antibody to bind to the antigen. For example, conservative changes (e.g., conservative substitutions as provided herein) can be made to a CDR that do not substantially reduce binding affinity. Such changes may, for example, be outside of the antigen-contacting residues in the CDR. In certain embodiments of the variant VH and VL sequences provided above, each CDR is unchanged or contains no more than one, two, or three amino acid substitutions.

[0174] A method that can be used to identify potential target residues or regions for mutagenesis in antibodies is called "alanine scanning mutagenesis." In this method, a residue or group of target residues (e.g., charged residues such as Arg, Asp, His, Lys, and Glu) is identified and substituted with neutral or negatively charged amino acids (e.g., Ala or polyalanine) to determine whether the antibody-antigen interaction is affected. Further substitutions can be introduced at amino acid positions that demonstrate functional sensitivity to the initial substitution. Additionally, contact points between the antibody and antigen can be identified by examining crystal structures of antigen-antibody complexes. These contact residues and adjacent residues can be targeted or removed as substitution candidates. Mutants can be screened to determine whether they contain desired properties.

[0175] Amino acid sequence insertions include amino- and / or carboxy-terminal fusions of polypeptides ranging in length from one residue to 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues. An example of a terminal insertion is an antibody with an N-terminal methionyl residue. Other insertional variants of the antibody molecule include the fusion to the N- or C-terminus of the antibody of an enzyme or a polypeptide which increases the serum half-life of the antibody.

[0176] Recombinant methods Anti-ILT4 antibodies can be produced by recombinant methods. For these methods, one or more isolated nucleic acids encoding the anti-ILT4 antibody are provided.

[0177] In one embodiment, the present disclosure provides isolated nucleic acids encoding the anti-ILT4 antibodies as described above. Such nucleic acids can each independently encode any one of the polypeptide chains described above. In another aspect, the present disclosure provides one or more vectors (e.g., expression vectors) comprising such nucleic acids. In another aspect, the present disclosure provides host cells comprising such nucleic acids. In one embodiment, a method of preparing a polypeptide is provided, the method comprising culturing a host cell comprising a nucleic acid encoding the polypeptide under conditions suitable for expression, as provided above, and optionally recovering the anti-ILT4 antibody from the host cell (or host cell medium).

[0178] To produce an anti-ILT4 antibody recombinantly, the nucleic acid encoding the protein is isolated and inserted into one or more vectors for further cloning and / or expression in a host cell. Such nucleic acids can be readily isolated and sequenced using routine procedures, or produced by recombinant methods, or obtained by chemical synthesis.

[0179] Suitable host cells for cloning or expressing vectors encoding anti-ILT4 antibodies include prokaryotic or eukaryotic cells as described herein. For example, they can be produced in bacteria, particularly where glycosylation and Fc effector functions are not required. After expression, they can be isolated from bacterial cell paste in a soluble fraction and further purified.

[0180] In addition to prokaryotes, eukaryotic microbes, such as filamentous fungi and yeast, are also suitable cloning or expression hosts for protein-encoding vectors, including fungal and yeast strains. Suitable host cells for protein expression may be derived from multicellular organisms (invertebrates and vertebrates), with examples of invertebrate cells including plant and insect cells. Many baculovirus strains have been identified and may be used in conjunction with insect cells, particularly for transfection of Spodoptera frugiperda cells. Plant cell cultures may also be used as hosts, as in, for example, US Pat. No. 5,959,177, US Pat. No. 6,040,498, US Pat. No. 6,420,548, US Pat. No. 7,125,978, and US Pat. No. 6,417,429, or vertebrate cells, such as mammalian cell lines suitable for growth in suspension, may also be used as hosts. Other examples of suitable mammalian host cell lines are SV40-transformed monkey kidney CV1 (COS-7), human embryonic kidney (293 or 293T) cells, baby hamster kidney (BHK) cells, mouse Sertoli (TM4) cells, monkey kidney (CV1) cells, African green monkey kidney (VERO-76) cells, human cervical carcinoma (HELA) cells, canine kidney (MDCK) cells, buffalo rat hepatocytes (BRL3A), human lung (W138) cells, human hepatocytes (Hep G2), mouse mammary tumor (MMT 060562) cells, TRI cells, MRC 5 cells, and FS4 cells. Other suitable mammalian host cell lines include Chinese hamster ovary (CHO) cells, including DHFR-CHO cells, and myeloma cell lines such as Y0, NS0, and Sp2 / 0. For a review of several mammalian host cell lines suitable for antibody production, see, e.g., Yazaki, P. and Wu, AM, Methods in Molecular Biology, Vol. 248, Lo, BKC (ed.), Humana Press, Totowa, NJ (2004), pp. 255-268.

[0181] measurement The anti-ILT4 antibodies provided herein can be identified, screened, or characterized for their physical / chemical characteristics and / or biological activity by various assays known in the art. In one embodiment, the activity of an anti-ILT4 antibody of the present disclosure is tested by known methods, such as, for example, ELISA, Western blotting, etc.

[0182] Treatment methods and administration routes Any anti-ILT4 antibody provided herein can be used in a method of treatment. In another aspect, the present disclosure provides a use of an anti-ILT4 antibody in the manufacture or preparation of a medicament. In some embodiments, the disease is an ILT4-associated disease or condition. In some embodiments, the disease is selected from the group consisting of astrocytoma (e.g., anaplastic astrocytoma), glioblastoma, bladder cancer, bone cancer, brain cancer, breast cancer (e.g., breast cancer characterized by BRCA1 and / or BRCA2 mutations), carcinoid, cervical cancer, choroid plexus papilloma, colorectal cancer (e.g., colon and rectal cancer), fallopian tube cancer, ependymoma, gallbladder cancer, gastric cancer, head and neck cancer, idiopathic myelofibrosis, kidney cancer or renal cell carcinoma (e.g., renal rhabdoid tumor, Wilms tumor), renal pelvis tumor, leukemia, liver cancer (e.g., hepatocellular carcinoma), esophageal cancer (e.g., esophageal squamous cell carcinoma), lung cancer (e.g., non-small cell lung cancer and small cell lung cancer), lymphoma, medulloblastoma, melanoma, meningioma, Merkel cell carcinoma, and the like. In some embodiments, the disease is selected from the group consisting of ovarian cancer, lung cancer, esophageal cancer, mesothelioma, multiple myeloma, neuroblastoma, oligodendroglioma, ovarian cancer, peritoneal tumor, pancreatic cancer, polycythemia vera, primary neuroectodermal tumor, prostate cancer, retinoblastoma, salivary gland cancer, sarcoma (e.g., chondrosarcoma, Ewing's sarcoma, osteosarcoma, rhabdomyosarcoma, synovial sarcoma, soft tissue sarcoma), small intestine cancer, colon cancer, squamous cell carcinoma (e.g., cutaneous squamous cell carcinoma), thyroid cancer, endometrial cancer, vestibular schwannoma, blastoma, teratoma, pituitary cancer, vulvar cancer, thymoma, testicular cancer, bile duct cancer, pheochromocytoma, paraganglioma, or adenoid cystic carcinoma, thrombocytosis, and in some embodiments, the disease is selected from ovarian cancer, lung cancer, esophageal squamous cell carcinoma, colorectal cancer, renal cell carcinoma, or melanoma.

[0183] In a further aspect, there is provided a pharmaceutical composition comprising the anti-ILT4 antibody described above, e.g., for use in any of the pharmaceutical uses or methods of treatment described above. In one embodiment, the pharmaceutical composition comprises any of the anti-ILT4 antibodies provided herein and a pharmaceutically acceptable carrier. In another embodiment, the pharmaceutical composition further comprises at least one additional therapeutic agent.

[0184] The anti-ILT4 antibodies of the present disclosure can be used alone or in combination with other agents for treatment, for example, the antibodies of the present disclosure may be administered with at least one additional therapeutic agent.

[0185] The anti-ILT4 antibodies (and any additional therapeutic agents) of the present disclosure may be administered by any suitable means, including parenteral, pulmonary, and intranasal administration, and, where localized treatment is required, intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. Dosing can be by any suitable route, for example, by injection, such as intravenous or subcutaneous injection, which will depend in part on whether administration is brief or chronic. Various dosing schedules are contemplated herein, including, but not limited to, a single dose or multiple doses at multiple time points, bolus administration, and pulse infusion.

[0186] The anti-ILT4 antibodies of the present disclosure are formulated, dosed, and administered in a manner consistent with good medical practice. Factors to consider in this context include the specific condition being treated, the specific mammal being treated, the clinical condition of the individual patient, the etiology of the condition, the site of delivery of the reagent, the method of administration, the administration schedule, and other factors known to medical professionals. The anti-ILT4 antibodies may or may not be formulated with one or more other reagents currently used to prevent or treat the condition. The effective amount of such other reagents will depend on the amount present in the pharmaceutical composition, the type of condition or treatment, and other factors. They will generally be used in the same dosages and by the same routes of administration as described herein, or at about 1% to 99% of the dosages described herein, or at alternative dosages and by any route determined empirically / clinically appropriate.

[0187] The appropriate dose of an anti-ILT4 antibody of the present disclosure (whether used alone or in combination with one or more other therapeutic agents) for preventing or treating a disease will depend on the type of disease being treated, the type of therapeutic molecule, the severity and course of the disease, whether the administration is prophylactic or therapeutic, previous treatments, the patient's clinical history and response to the therapeutic molecule, and the judgment of the attending physician. The therapeutic molecule is administered to the patient at one time or over a series of treatments, as appropriate.

[0188] product Another aspect of the present disclosure provides an article of manufacture (e.g., a product prepared in the form of a kit) containing materials usable for the treatment, prevention, and / or diagnosis of the above-described conditions. The article of manufacture includes a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, syringes, IV solution bags, etc. The container may be formed from a variety of materials, such as glass or plastic. The container holds a composition that effectively treats, prevents, and / or diagnoses a disease, alone or in combination with another composition, and may have a sterile access port (e.g., the container may be an intravenous solution bag or vial with a plug pierceable by a hypodermic needle). At least one active agent in the composition is an anti-ILT4 antibody of the present disclosure. The label or package insert indicates that the composition is used to treat the selected condition. The article of manufacture may also include (a) a first container containing a composition comprising an anti-ILT4 antibody of the present disclosure and (b) a second container containing a composition comprising another cytotoxic or other therapeutic agent. The product in this embodiment of the present disclosure may further include a package insert indicating that the composition is applicable to the treatment of a particular medical condition. Alternatively, or in addition, the product may further include a second (or third) container containing a pharmaceutically acceptable buffer. From a business and user perspective, it may further include other necessary materials, including other buffers, diluents, filters, needles, and syringes. [Example]

[0189] The present disclosure will be further explained below with reference to examples and test examples, but these examples and test examples do not limit the scope of the present disclosure. Experimental methods for which specific conditions are not specified in the examples and test examples of the present disclosure generally follow standard conditions, such as those in the Cold Spring Harbor Antibody Technology Laboratory Manual and the Molecular Cloning Manual, or conditions recommended by manufacturers of raw materials or products. Reagents for which specific sources are not specified are standard commercially available reagents.

[0190] Example 1: Preparation of ILT4 antigen The amino acid sequences of the antigens and detection proteins used in this disclosure were designed using UniProt LIRB2_HUMAN (human LILRB2 protein, Uniprot code: Q8N423) as a template for ILT4. Different tags, such as His or Fc, were selectively fused to the ILT4 protein. Each was cloned into the pTT5 vector (Biovector, CAT#102762), transiently transfected in 293 cells, expressed, and purified to obtain the immunization and detection antigens described in this disclosure.

[0191] 1. Human-IgG1-Fc tagged ILT4 protein extracellular domain: ILT4-Fc was used as the immunization antigen.

[0192] [Table 3]

[0193] 2. His-tagged extracellular domain of ILT4 protein: ILT4-His was used as the detection antigen.

[0194] [Table 4]

[0195] CHO-S cells overexpressing the full-length ILT4 protein (CHO-S / ILT4) were used as detection cells.

[0196] [Table 5]

[0197] Example 2: Purification of recombinant proteins related to ILT4 1. Purification step of His-tagged recombinant protein The cell expression supernatant sample was centrifuged at high speed to remove impurities, and the buffer was replaced with PBS, with imidazole added to a final concentration of 5 mM. A nickel column was equilibrated with PBS containing 5 mM imidazole and washed with 2 to 5 column volumes. The replaced cell supernatant sample was applied to a Ni Sepharose Excel column (GE, 17-3712-02). 280 The column was washed with PBS containing 5 mM imidazole until the readings were reduced to baseline. The chromatography column was then washed with PBS + 10 mM imidazole to remove nonspecifically bound heteroproteins, and the flow-through was collected. The target protein was then eluted with PBS containing 300 mM imidazole, and the elution peak was collected. The collected eluate was concentrated and further purified using a Superdex 200 gel chromatograph (GE, 28-9893-35) with a PBS mobile phase. The dimer peak was removed, and the elution peak was collected. The resulting protein was identified as accurate by electrophoresis, peptide mapping, and LC-MS, and then aliquoted for use. His-tagged ILT4-His was isolated and used as a detection antigen for the antibody of the present disclosure.

[0198] 2. Purification step of Fc-tagged recombinant protein or antibody The cell expression supernatant sample was centrifuged at high speed to remove impurities, and the supernatant was subjected to affinity chromatography using MabSelect Sure (GE, 17-5438-01). The MabSelect Sure chromatography column was first regenerated with 0.2 M NaOH, washed with pure water, and then equilibrated with PBS. After binding the supernatant, the column was then eluted with PBS. 280 The sample was washed with PBS until the readings were reduced to baseline. The target protein was eluted with 0.1 M acetate buffer at pH 3.5 and neutralized with 1 M Tris-HCl. After appropriate concentration, the eluted sample was further purified using a gel chromatograph Superdex 200 (GE, 28-9893-35) equilibrated with PBS. The receiving tubes containing the target protein were combined and concentrated to an appropriate concentration. This method was used to purify the ILT4-Fc fusion protein, but it may also be used to purify the humanized antibody protein of the present disclosure.

[0199] Example 3: Obtaining anti-human ILT4 antibody 1. Immunization of mice Anti-human ILT4 monoclonal antibodies were produced by immunization of mice. Female SJL white mice, 6-8 weeks old (Shanghai SLAC Laboratory Animal Co., Ltd., Animal Production License Number: SCXK(Hu)2017-0005) were used for the experiments. The breeding environment was SPF grade. After purchasing, the mice were housed in a laboratory environment for one week, with a 12 / 12 h light / dark cycle, a temperature of 20-25°C, and humidity of 40%-60%. Once acclimated, the mice were immunized according to the following schedule.

[0200] Mice were immunized with ILT4-Fc protein. Cross-immunization was performed with the adjuvants TiterMax® Gold Adjuvant (Sigma Cat No. T2684) and Thermo Imject® Alum (Thermo Cat No. 77161). The antigen to adjuvant (TiterMax® Gold Adjuvant) ratio was 1:1, and the antigen to adjuvant (Thermo Imject® Alum) ratio was 3:1. The doses were 50 μg / mouse / primary immunization and 25 μg / mouse / booster immunization. The antigen was emulsified and immunized on days 0, 14, and 34. Blood samples were collected on days 25 and 41 to measure serum antibody titers. After the third immunization, mice with high and stable serum antibody titers were selected for single B cell screening. Three days before screening, booster immunization was performed by intraperitoneal (ip) injection of 25 μg / mouse of an antigen solution prepared in physiological saline.

[0201] 2. Single B Cell Screening Single B cells were screened primarily using positive and negative screening, 10*genomics, and bioinformatics techniques to obtain a series of antibody variable region sequences. The resulting sequences were then subjected to CDR typing, and the mouse variable region sequences were selected and linked to human antibody constant region sequences to express chimeric antibodies. Screening was performed using methods such as ILT4 protein binding, cell binding, and ILT4 / HLA-G blocking to select antibody sequences that specifically bind to ILT4. The heavy and light chain variable region sequences of the antibodies obtained through screening are as follows:

[0202] [Table 6] [Table 7] [Table 8] [Table 9] [Table 10] [Table 11]

[0203] The heavy and light chain CDR region sequences of the mouse antibody are shown in Table 3 below. [Table 12-1] [Table 12-2] [Table 12-3]

[0204] 3. Phage Library Screening Since the screening was aimed at obtaining antibody sequences that bind only to ILT4, a method combining negative and positive screening was used. First, phages capable of binding to biotin-labeled human ILT2 were removed from the phage library (negative screening), and then the remaining phages were screened for those capable of binding to biotin-labeled human ILT4 (positive screening). The specific process was as follows: The phage library (5 x 10 12(1 / pfu) was suspended in 1 mL of PBS containing nonfat dry milk (BD, 232100), and 2 μg / mL of biotin-labeled human ILT2-His protein was added and incubated with inversion, followed by 100 μL of Dynabeads® M-280 Streptavidin (Thermofisher, 11206D). The tube was placed on a magnetic rack for 30 seconds, the supernatant was removed, and 2 μg / mL of biotin-labeled human ILT4-His protein was added and incubated with inversion, followed by 100 μL of Dynabeads. The tube was placed on a magnetic rack for 30 seconds, the supernatant was removed, and the Dynabeads were washed several times with 1 mL of PBS containing Tween-20 before being eluted with 0.5 mL of trypsin (Sigma, T1426-250MG). The eluted phages were infected into Escherichia coli SS320, titered, amplified, and concentrated for subsequent elutriation. Screening was performed three times according to this method, and the monoclones obtained were inoculated into 96-well plates (Sangon Biotech, F600582-0001) for phage packaging. Phages capable of binding to human ILT4 protein but not human ILT2 protein were detected by ELISA, and the corresponding monoclones were sequenced and analyzed. The variable region sequences of the finally obtained antibodies are as follows:

[0205] [Table 13]

[0206] The sequences of the heavy and light chain CDR regions of the fully human antibody are shown in Table 4 below.

[0207] [Table 14]

[0208] 4. Construction of Chimeric Antibodies Primer PCR was designed to obtain VH / VK gene fragments of each mouse antibody. These fragments were then homologously recombined with the expression vector pTT5 (which contains the signal peptide and constant region gene (CH1-FC / CL) fragments and was constructed in the laboratory) to construct the full-length antibody expression vector VH-CH1-FC-pTT5 / VK-CL-pTT5, and used to express an anti-human ILT4 chimeric antibody in the form of IgG4 (LALA). Chimeric antibodies were obtained by combining the variable region sequences of mouse anti-ILT4 antibodies ChP1-C4, ChP1-B4, ChP1-D10, ChP2-C7, ChP1-G7, ChP1-B9, ChP2-C6, ChP1-F7, ChP2-E8, ChP2-E3, ChP2-C10, ChP2-F4, ChP2-F10, ChP2-C4, ChP2-G9, ChP2-B3, ChP2-E7, ChP2-G4, ChP2-E11, ChP1-D7, ChP1-E11, ChP2-B9, ChP2-C2, and ChP2-E10 with the constant regions set forth in SEQ ID NOs: 171 and 172. Exemplary, ChP1-C4 represents a chimeric antibody comprising the MP1-C4 mouse heavy chain variable region, light chain variable region, and the above-described constant region. The sequences of the heavy and light chain constant regions of the chimeric antibody are as follows:

[0209] [Table 15]

[0210] [Table 16]

[0211] Example 4: Humanization of anti-human ILT4 mouse antibody By aligning the IMGT human antibody heavy and light chain variable region germline genes database with the MOE software, the heavy and light chain variable region germline genes with high homology to MP2-E3, MP2-G9, MP2-E8, MP1-G7, MP2-C6, and MP1-D7 were selected as templates, and the CDRs of these six mouse antibodies were grafted onto the corresponding human templates to form the variable region sequences in the order FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4, where amino acid residues were annotated according to the Kabat numbering system.

[0212] 1.M Humanization of P2-E3 For the humanized MP2-E3 antibody, IGKV1-39*01 was selected as the template for the light chain framework region, and IGHV1-3*01 was selected as the template for the heavy chain framework region. The CDR regions of the mouse antibody were grafted onto the selected humanized template, and back-mutations and point mutations were performed using MOE software to identify buried residues, residues that directly interact with the CDR regions, and high-risk hotspots and sites with immunogenicity risks, to design different light and heavy chain variable regions of the humanized antibody.

[0213] [Table 17]

[0214] [Table 18]

[0215] The variable region sequences of humanized antibody P2-E3 are as follows:

[0216] [Table 19]

[0217] [Table 20]

[0218] 2.M Humanized modification of P2-G9 For the humanized MP2-G9 antibody, IGKV1-39*01 was selected as the template for the light chain framework region, and IGHV4-31*01 was selected as the template for the heavy chain framework region. The CDR regions of the mouse antibody were grafted onto the selected humanized template, and back-mutations and point mutations were performed using MOE software to identify buried residues, residues that directly interact with the CDR regions, and high-risk hotspots and sites with immunogenicity risks, to design different light and heavy chain variable regions of the humanized antibody.

[0219] [Table 21]

[0220] The variable region sequences of humanized antibody P2-G9 are as follows: [Table 22]

[0221] [Table 23] [Table 24] [Table 25] [Table 26]

[0222] 3.M Humanization of P2-E8 For the humanized MP2-E8 antibody, IGKV1-33*01 was selected as the template for the light chain framework region, and IGHV3-21*01 was selected as the template for the heavy chain framework region. The CDR regions of the mouse antibody were grafted onto the selected humanized template, and back-mutations and point mutations were performed using MOE software to identify buried residues, residues that directly interact with the CDR regions, and high-risk hotspots and sites with immunogenicity risks, to design different light and heavy chain variable regions of the humanized antibody.

[0223] [Table 27]

[0224] [Table 28]

[0225] The variable region sequences of humanized antibody P2-E8 are as follows:

[0226] [Table 29]

[0227] [Table 30]

[0228] 4. Humanized modification of M P1-G7 For the humanized MP1-G7 antibody, IGKV1-39*01 was selected as the template for the light chain framework region, and IGHV4-31*02 was selected as the template for the heavy chain framework region. The CDR regions of the mouse antibody were grafted onto the selected humanized template, and back-mutations and point mutations were performed using MOE software to identify buried residues, residues that directly interact with the CDR regions, and high-risk hotspots and sites with immunogenicity risks, to design different light and heavy chain variable regions of the humanized antibody.

[0229] [Table 31]

[0230] The variable region sequences of humanized antibody P1-G7 are as follows:

[0231] [Table 32]

[0232] [Table 33]

[0233] 5.M Humanization of P2-C6 For the humanized P2-C6 antibody, IGKV4-1*01 and IGKV3-11*01 were selected as templates for the light chain framework region, and IGHV1-18*01 was selected as a template for the heavy chain framework region. The CDR regions of a mouse antibody were grafted onto the selected humanized template, and back-mutations and point mutations were performed using MOE software to identify buried residues, residues that directly interact with the CDR regions, and high-risk hotspots and sites with immunogenicity risks, to design different light and heavy chain variable regions of the humanized antibody. [Table 34]

[0234] [Table 35]

[0235] The variable region sequences of humanized antibody P2-C6 are as follows: [Table 36] [Table 37]

[0236] [Table 38] [Table 39] [Table 40] [Table 41]

[0237] 6.M Humanized modification of P1-D7 For the humanized MP1-D7 antibody, IGKV4-1*01 and IGKV2-28*01 were selected as templates for the light chain framework region, and IGHV1-18*01 was selected as a template for the heavy chain framework region. The CDR regions of the mouse antibody were grafted onto the selected humanized template, and back-mutations and point mutations were performed using MOE software to identify buried residues, residues that directly interact with the CDR regions, and high-risk hotspots and sites with immunogenicity risks, to design different light and heavy chain variable regions of the humanized antibody.

[0238] [Table 42]

[0239] [Table 43]

[0240] The variable region sequences of humanized antibody P1-D7 are as follows:

[0241] [Table 44]

[0242] [Table 45]

[0243] 7. Engineering of the Fully Human Antibody P2 High-risk hotspots selectively added the N30Q point mutation to the VL, and immunogenic risk sites selectively added T76S and A81E point mutations to the VL.

[0244] [Table 46]

[0245] [Table 47]

[0246] 8. Construction and Expression of the Full Form of the Anti-human ILT4 Humanized Antibody Primer PCR was designed to construct each humanized antibody VH / VK gene fragment, which was then homologously recombined with the expression vector pTT5 (which contains the signal peptide and constant region gene (CH1-FC / CL) fragments and was constructed in the laboratory) to construct the full-length antibody expression vector VH-CH1-Fc-pTT5 / VK-CL-pTT5. The heavy and light chain sequences of the humanized antibodies huP2-E3, huP2-G9, huP2-E8, huP1-G7, huP2-C6, and huP1-D7, as well as the fully human antibody P2m, were finally determined by SPR and FACS detection of ILT4 binding. They are shown below.

[0247] [Table 48]

[0248] [Table 49]

[0249] [Table 50]

[0250] [Table 51]

[0251] [Table 52]

[0252] [Table 53]

[0253] [Table 54]

[0254] The sequences of the heavy and light chain constant regions of the humanized antibody are as follows:

[0255] [Table 55]

[0256] [Table 56]

[0257] [Table 57]

[0258] The positive control molecules used in this disclosure are MK and JTX.

[0259] MK sequence origin: WO2018187518A1 The theoretical light and heavy chain amino acid sequences of MK are as follows (single underlined: variable region, double underlined: CDR, dotted underlined: constant region).

[0260] [Table 58]

[0261] Origin of JTX sequence: WO2019126514A2 The theoretical light and heavy chain amino acid sequences of JTX are as follows (single underlined: variable region, double underlined: CDR, dotted underlined: constant region).

[0262] [Table 59]

[0263] [Table 60]

[0264] Example 5: Detection of protein binding of anti-ILT4 antibodies by ELISA Human ILT4-His protein was diluted to 1 μg / mL in PBS and coated onto a 96-well plate (Corning, CLS3590-100EA) and incubated overnight at 4°C. After washing, the plate was blocked with 1% BSA at 37°C for 1 hour. After washing, different concentrations of anti-ILT4 antibody were added and incubated at 37°C for 1 hour. After washing, HRP-goat anti-human F(ab′)2 secondary antibody (Jackson, 109-036-097) was added and incubated at 37°C for 1 hour. After washing, TMB chromogenic substrate (KPL, 52-00-03) was added and incubated at room temperature for 5–10 minutes. The reaction was stopped with 1 M H2SO4, and the absorbance was read at 450 nm using a VERSAmax plate reader (Molecular Devices). The results are shown in Table 22.

[0265] [Table 61]

[0266] The experimental results show that the anti-ILT4 chimeric antibodies all have higher binding ability to ILT4 protein than the control antibody JTX.

[0267] Example 6: Detection of protein binding of anti-ILT4 antibodies by Biacore The affinity of the humanized antibodies of the present disclosure for ILT4-His protein was tested using Biacore. After affinity capture of the antibodies using a Protein A biosensor chip (Cytiva, 29127556), ILT4-His was applied to the chip surface, and the reaction signal was detected in real time by the instrument to obtain binding-dissociation curves. After the dissociation period for each experimental cycle was completed, the biosensor chip was regenerated by washing with 10 mM Glycine-HCl pH 1.5 (Cytiva, BR-1003-54). A 1:1 model was used for data fitting. The binding and dissociation profiles of each antibody are shown in Table 23.

[0268] [Table 62]

[0269] Experimental results show that the humanized and fully human antibodies of the present disclosure have a higher ability to bind to ILT4 protein than the control antibody.

[0270] Example 7: Detection of cell binding of anti-ILT4 antibodies by FACS The affinity of the humanized antibodies of the present disclosure to CHO-S / ILT4 cells was tested by FACS. 6 After blocking CHO-S / ILT4 cells / mL with 1% BSA, different concentrations of anti-ILT4 antibody samples were added and incubated at 4°C for 1 hour. After washing twice with 1% BSA, F(ab')2-goat anti-human IgG Fc secondary antibody and PE (Invitrogen, H10104) were added and incubated at 4°C for 0.5 hours. After washing twice with 1% BSA, the fluorescent signal values ​​were read by FACS. The results are shown in Table 24.

[0271] [Table 63]

[0272] Experimental results show that the humanized and fully human antibodies of the present disclosure have binding ability to ILT4-positive cells that is equal to or greater than that of the control antibody.

[0273] Example 8: Anti-ILT4 antibodies do not bind to other members of the LILR family Human ILT2-His (the method used to prepare ILT4-His in Examples 1 and 2 was the same as in Examples 1 and 2, and the amino acid sequence is shown in SEQ ID NO: 245), ILT3-His (the method used to prepare ILT4-His in Examples 1 and 2 was the same as in Examples 1 and 2, and the amino acid sequence is shown in SEQ ID NO: 246), ILT4-His (the method used to prepare ILT4-His in Examples 1 and 2), LILRB5-His (Acro, CDC-H5220), LILRA1-His (Acro, LI1-H82E8), and LILR-His (Acro, LI1-H82E8) were prepared using PBS. A2-His (Acro, LI2-H82E9), LILRA4-His (SinoBiological, 16058-H08H), LILRA5-His (Acro, LI5-H82E1), LILRA6-His (Acro, LI6-H82E2), or LILRA3-His (Acro, LI3-H82E0) was diluted to 1 μg / mL and coated onto a 96-well plate (Corning, CLS3590-100EA) and incubated overnight at 4°C. After washing, the plate was blocked with 1% BSA at 37°C for 1 hour. After washing, 10 μg / mL of anti-ILT4 antibody was added and incubated at 37°C for 1 hour. After washing the plate, HRP-goat anti-human F(ab')2 secondary antibody (Jackson, 109-036-097) was added and incubated at 37°C for 1 hour. After washing the plate, TMB chromogenic substrate (KPL, 52-00-03) was added and incubated at room temperature for 5-10 minutes. The reaction was stopped by adding 1M H2SO4, and the absorbance was read at 450 nm using a VERSAmax plate reader (Molecular Devices). The results are shown in Table 25.

[0274] [Table 64]

[0275] [Table 65]

[0276] [Table 66]

[0277] Experimental results show that both the humanized and fully human antibodies of the present disclosure do not bind to other members of the LILR family.

[0278] Example 9: Blocking effect of anti-ILT4 antibodies on the binding of ILT4 to HLA-G The cell blocking activity of anti-ILT4 antibodies was detected by flow cytometry. 6 CHO-S / ILT4 cells (100 cells / mL) were blocked with 1% BSA, and then various concentrations of anti-ILT4 antibodies diluted in 1% BSA and CF647-labeled HLA-G tetramers (novoprotein, SIGMA, MX647S100-1KT) were added and incubated for 1 h at 4°C. The cells were centrifuged at 500g for 4 min, and the supernatant was discarded. After washing twice with 1% BSA, the fluorescent signal was read by FACS.

[0279] [Table 67]

[0280] The experimental results show that the antibodies huP2-E3, huP2-G9, huP2-C6, and P2m according to the present disclosure can all effectively block the binding of ILT4 to HLA-G, and their blocking activity is stronger than that of the control antibodies MK and JTX.

[0281] Example 10: Blocking effect of anti-ILT4 antibodies on ILT4 and its downstream signaling pathways The blocking ability of anti-ILT4 antibodies to ILT4 was detected using the 3A9 / ILT4 reporter system. 1 μg / mL of mCD3 antibody (BioLegend, 100238) was seeded into a 96-well plate (Corning, 3599) and incubated at 37°C for 2 h. The supernatant was then discarded. 3A9 cells (ATCC, CRL-3293) overexpressing full-length ILT4 protein were harvested and resuspended in RPMI 1640 (Meilun, PWL004) + 5% FBS (Gibco, 10099-141) + 0.05 mM 2-B-mercaptoethanol (Sigma, M3148-25mL) to a density of 8.3 × 10 cells. 5 The cells were adjusted to a concentration of 1000 cells / mL and added to the 96-well plate. At the same time, different concentrations of anti-ILT4 antibody were added and the cells were cultured at 37°C and 5% CO for 24 hours. After 24 hours, the cell supernatant was collected. mIL-2 secretion was detected using an mIL-2 kit (Xinbosheng, EMC002.96.2). The results are shown in Figure 1A-1D.

[0282] The experimental results show that the antibodies according to the present disclosure can more effectively relieve the inhibitory effect of ILT4 than the control antibody MK and promote higher levels of mIL-2 secretion by 3A9 cells.

[0283] Example 11: Effect of anti-ILT4 antibodies on cytokine secretion by macrophages CD14 MicroBeads (Miltenyi Biotec, 130-050-201) were used to detect CD14 from healthy human PBMCs. + Monocytes were isolated and obtained by CD14 immunization using RPMI 1640 (Meilun, PWL004) + 10% FBS (Gibco, 10099-141) + 50 ng / mL M-CSF (Peprotech, 300-25). + Resuspend the cells to a cell density of 2 x 10 6 After 7 days of differentiation, macrophages were obtained. The macrophages were resuspended in RPMI 1640 + 10% FBS to a density of 1 × 10 6The cells were adjusted to cells / mL and inoculated into a 96-well plate (Jet, 3788). At the same time, LPS (Sigma, L3024-5MG) and different concentrations of anti-ILT4 antibody were added, and the cells were incubated at 37°C and 5% CO for 24 h. The supernatants were collected and TNFα was detected using a HUMAN TNFα KIT (Youningwei, 62HTNFAPEG). The results are shown in Figure 2A-2D.

[0284] Experimental results show that the antibodies of the present disclosure are able to more effectively promote the conversion of macrophages to the M1 phenotype and secrete higher levels of the pro-inflammatory cytokine TNF-α than the control antibody MK.

[0285] Example 12: Combination effect of anti-ILT4 antibody and anti-PD-1 antibody in allogeneic MLR CD14 MicroBeads (Miltenyi Biotec, 130-050-201) were used to detect CD14 from healthy human PBMCs. + Monocytes were isolated and obtained by CD14 immunization using RPMI 1640 (Meilun, PWL004) + 10% FBS (Gibco, 10099-141) + 50 ng / mL M-CSF (Peprotech, 300-25). + Resuspend the cells to a cell density of 2 x 10 6 After 7 days of differentiation, macrophages were obtained. The macrophages were resuspended in RPMI 1640 + 10% FBS to a density of 2 × 10 5 The concentration was adjusted to cells / mL and inoculated into a 96-well plate (Jet, 3788).

[0286] Pan T cells were isolated from PBMCs of another donor using a T cell sorting kit (Miltenyi, 130-096-535) and resuspended in RPMI 1640 + 10% FBS at a density of 2 × 10 6The cells were then added to the 96-well plate at a final concentration of 10 μg / mL. Anti-ILT4 antibody and / or anti-PD-1 antibody (anti-PD-1 antibody Hu23-11.IgG4AA, prepared in accordance with patent WO2020156509 A1 and with the following sequence) were added simultaneously to a final concentration of 10 μg / mL, and the cells were cultured at 37°C and 5% CO2 for 6 days. The cell supernatant was collected and IFNγ was detected using a HUMAN IFNγ KIT (Yuningwei, 62HIFNGPEG). The results are shown in Figures 3A-3E.

[0287] [Table 68]

[0288] [Table 69]

[0289] [Table 70]

[0290] The experimental results show that the antibody of the present disclosure, when combined with an anti-PD-1 antibody in an MLR experiment, is clearly more effective at promoting INFγ release than the combination of the positive antibody MK or JTX with an anti-PD-1 antibody.

[0291] Example 13: Promoting effect of anti-ILT4 antibody on dendritic cell activation CD14 MicroBeads (Miltenyi Biotec, 130-050-201) were used to detect CD14 from healthy human PBMCs. + Monocytes were isolated and obtained by CD14 immunization using RPMI 1640 (Meilun, PWL004) + 10% FBS (Gibco, 10099-141) + 100 ng / mL GM-CSF (Peprotech, 300-03-100 μg) + 100 ng / mL IL-4 (R&D, 217-IL-010). + The cells were resuspended. After differentiation for 5-6 days, dendritic cells were obtained. The density of dendritic cells was 1 × 10 6The cells were adjusted to a concentration of 10 cells / mL and inoculated into a 96-well plate (Jet, 3788). IL-10 (R&D, 217-IL-010) was added at the same time to a final concentration of 10 ng / mL, and anti-ILT4 antibody was added at a final concentration of 10 μg / mL. The cells were then cultured at 37°C, 5% CO2 for 48 hours. The cell supernatant was collected and CCL-2 (Xinbosheng, EHC.113.96.2) and IL-6 (Cisbio, 62HIL06PEG) were detected using ELISA kits. The upregulation of each cytokine or chemokine is shown in Table 28. [Table 71]

[0292] Experimental results show that in an immunosuppressive environment in the presence of IL-10, the humanized antibody huP2-G9 of the present disclosure can still induce higher levels of secretion of pro-inflammatory cytokines or chemokines by dendritic cells.

[0293] Example 14: Inhibitory effect of anti-ILT4 antibody on subcutaneously transplanted tumors in human breast cancer-bearing MDA-MB-231 / HLA-G mice MDA-MB-231 cells (ATCC, HTB-26) overexpressing HLA-G in logarithmic growth phase: T cells: Mφ cells = (3 × 10 6 ):(5×10 5 ):(5×10 5 The mixture was mixed at a ratio of 6:1:1 and inoculated subcutaneously into the right flank of mice (Weiting Lihua) at 200 μL (containing 100 μL of Matrigel) per mouse, with 6–8 mice per group. The vehicle control group was not inoculated with Mφ or T cells, and the day of grouping was defined as day 0 of the experiment. The remaining PBMCs were discontinued and continued to be cultured. After 1 week, 5 × 10 6 Tumor-bearing mice were intraperitoneally injected with 200 μL of cells per mouse. Anti-ILT4 antibody was intraperitoneally injected twice a week from day 0 at a dose of 20 mpk. Tumor volume was monitored twice a week and the data were recorded.

[0294] The tumor volumes of animals in each group were expressed as mean±SEM and plotted using Graphpad Prism 8 software, followed by statistical analysis using two-way analysis of variance.

[0295] The formula for calculating tumor volume (V) is V = 1 / 2 × a × b 2 In this case, a and b indicate the length and width, respectively.

[0296] The relative tumor growth rate T / C (%) = (T-T0) / (C-C0) × 100%, where T and C are the tumor volumes of the treatment and control groups at the end of the experiment, and T0 and C0 are the tumor volumes at the start of the experiment.

[0297] Tumor inhibition rate TGI(%)=1-T / C(%).

[0298] The tumor inhibition rate for each group is shown in Table 29 below. [Table 72]

[0299] The experimental results showed that on day 13, compared to the PBS solvent control group (vehicle group), the tumor inhibition rates of the T+Mφ+negative antibody-20mpk, MK-20mpk and huP2-G9-20mpk groups were 33.0%, 46.2% and 47.9%, respectively; and on day 13, compared to the control group T+Mφ+negative antibody-20mpk group, the tumor inhibition rates of the MK-20mpk and huP2-G9-20mpk groups were 19.6% and 22.2%, respectively, indicating that the efficacy of the huP2-G9 single-drug administration group was equivalent to that of the positive MK single-drug administration group, and therefore the TGI value was slightly higher.

[0300] Example 15: Inhibitory effect of anti-ILT4 antibody on subcutaneously transplanted tumors in double transgenic ILT4-ILT5 mice bearing MC38 / HLA-G colon cancer Double transgenic human ILT4-ILT5 mice were cultured in a suspension of MC38 cells (Chemical Biology, CBP60825) overexpressing HLA-G (as in Example 9) (5 × 10 5100 μL of the solution was inoculated subcutaneously into the right armpit of the mouse until the tumor cells were approximately 80–100 mm 3 After selecting tumors that had grown to a volume of approximately 1000 mg / mL, the mice were randomly assigned to groups of 8 mice per group. Anti-ILT4 antibody and anti-PD-1 antibody (self-produced; the amino acid sequences of the heavy and light chains of anti-PD-1 antibody (αPD-1) are set forth in SEQ ID NO: 258 and SEQ ID NO: 259, respectively) were intraperitoneally injected twice a week at doses of 30 mpk and 0.3 mpk, respectively. Tumor volume was monitored twice a week, and data were recorded.

[0301] The tumor volumes of animals in each group were expressed as mean±SEM and plotted using Graphpad Prism 8 software, followed by statistical analysis using two-way analysis of variance.

[0302] The formula for calculating tumor volume (V) is V = 1 / 2 × a × b 2 In this case, a and b indicate the length and width, respectively.

[0303] The relative tumor growth rate T / C (%) = (T-T0) / (C-C0) × 100%, where T and C are the tumor volumes of the treatment and control groups at the end of the experiment, and T0 and C0 are the tumor volumes at the start of the experiment.

[0304] Tumor inhibition rate TGI(%)=1-T / C(%).

[0305] The tumor inhibition rate for each group is shown in Table 30 below. [Table 73]

[0306] [Table 74]

[0307] The experimental results showed that compared with the control antibody MK, single-agent administration of huP2-G9 and huP2-C6 had better efficacy, single-agent administration of anti-PD-1 antibody had relatively strong efficacy, and the combined use of huP2-G9 or huP2-C6 groups had slightly stronger efficacy than the corresponding single-agent groups.

[0308] It is to be clearly understood that while the foregoing invention has been described in detail by way of illustration and example, such descriptions and examples should not be construed as limiting the scope of the present disclosure. The disclosures of all patent and scientific literature cited herein are hereby incorporated in their entirety and expressly by reference.

Claims

1. An anti-ILT4 antibody comprising a heavy chain variable region comprising HCDR1, HCDR2 and HCDR3, and a light chain variable region comprising LCDR1, LCDR2 and LCDR3, a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 184 or 185, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 187 or 186, respectively; or HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 32, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 33, respectively; or b. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 211, 205, 206, 207, 208, 209, or 210, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in any one of SEQ ID NOs: 212, 213, or 214; or HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 40, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 41, respectively; or c. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 181, 176, 177, 178, 179, or 180, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 183 or 182; or HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO:28, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO:29, respectively; or d. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 193, 190, 191, or 192, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in any one of SEQ ID NOs: 194, 195, or 196; or HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 10, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 11, respectively; or e. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 163, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 225 or 164; or f. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 197 or 198, respectively, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 199 or 200, respectively; or HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 36, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 37, respectively; or g. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in any one of SEQ ID NOs: 217, 218, 219, 220, or 221, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 222 or 223; or HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO: 42, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO: 43, respectively; or h. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 34, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 35; or i. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 18, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 19; or j. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 20, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 21; or k. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 24, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 25; or l. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 44, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 45; or m. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 46, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 47; or n. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 48, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 49; or o. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 50, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 51; Preferably, a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 184, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 187; or b. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO:211, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO:212; or c. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 181, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 183; or d. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 193, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 194; or e. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 163, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 225; or f. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 197, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 199; or g. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 217, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 222; More preferably, a. the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3, respectively, in SEQ ID NO: 184, and the LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3, respectively, in SEQ ID NO: 187; or b. HCDR1, HCDR2, and HCDR3 of the heavy chain variable region comprise the amino acid sequences of HCDR1, HCDR2, and HCDR3 in SEQ ID NO:211, respectively, and LCDR1, LCDR2, and LCDR3 of the light chain variable region comprise the amino acid sequences of LCDR1, LCDR2, and LCDR3 in SEQ ID NO:212, respectively; Anti-ILT4 antibody.

2. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 121, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; or b. the heavy chain variable region has an HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, an HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 203, 136, 201, 202, or 204, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has an LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; or the heavy chain variable region has an HCDR1 comprising the amino acid sequence of SEQ ID NO: 111, an HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 173, 112, 174, or 175, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 113, and the light chain variable region has an LCDR1 comprising the amino acid sequence of SEQ ID NO: 114, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 115, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 116; or d. the heavy chain variable region has an HCDR1 comprising the amino acid sequence of SEQ ID NO: 69, an HCDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 189, 70, or 188, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 71, and the light chain variable region has an LCDR1 comprising the amino acid sequence of SEQ ID NO: 61, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 72; or e. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 165, HCDR2 comprising the amino acid sequence of SEQ ID NO: 166, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 167, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 224 or 168, LCDR2 comprising the amino acid sequence of SEQ ID NO: 169, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 170; or f. the heavy chain variable region comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 125, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 126, and the light chain variable region comprises an LCDR1 comprising the amino acid sequence of SEQ ID NO: 127, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 128; or g. the heavy chain variable region has HCDR1 comprising the amino acid sequence of SEQ ID NO: 141, HCDR2 comprising the amino acid sequence of SEQ ID NO: 215 or 142, and HCDR3 comprising the amino acid sequence of SEQ ID NO: 216 or 143, and the light chain variable region has LCDR1 comprising the amino acid sequence of SEQ ID NO: 144, LCDR2 comprising the amino acid sequence of SEQ ID NO: 145, and LCDR3 comprising the amino acid sequence of SEQ ID NO: 146; or h. the heavy chain variable region comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO:99, an HCDR2 comprising the amino acid sequence of SEQ ID NO:122, and an HCDR3 comprising the amino acid sequence of SEQ ID NO:123, and the light chain variable region comprises an LCDR1 comprising the amino acid sequence of SEQ ID NO:114, an LCDR2 comprising the amino acid sequence of SEQ ID NO:109, and an LCDR3 comprising the amino acid sequence of SEQ ID NO:124; or i. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO:73, HCDR2 comprises the amino acid sequence of SEQ ID NO:87, and HCDR3 comprises the amino acid sequence of SEQ ID NO:88, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO:61, LCDR2 comprises the amino acid sequence of SEQ ID NO:89, and LCDR3 comprises the amino acid sequence of SEQ ID NO:90; or j. the heavy chain variable region comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 91, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 92, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 93, and the light chain variable region comprises an LCDR1 comprising the amino acid sequence of SEQ ID NO: 85, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 94; or k. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO:99, HCDR2 comprises the amino acid sequence of SEQ ID NO:100, and HCDR3 comprises the amino acid sequence of SEQ ID NO:101, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO:102, LCDR2 comprises the amino acid sequence of SEQ ID NO:103, and LCDR3 comprises the amino acid sequence of SEQ ID NO:104; or the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 147, HCDR2 comprises the amino acid sequence of SEQ ID NO: 148, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 149, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 150, LCDR2 comprises the amino acid sequence of SEQ ID NO: 151, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 152; or m. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 153, HCDR2 comprises the amino acid sequence of SEQ ID NO: 154, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 155, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 133, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; or n. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO:58, HCDR2 comprises the amino acid sequence of SEQ ID NO:156, and HCDR3 comprises the amino acid sequence of SEQ ID NO:157, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO:158, LCDR2 comprises the amino acid sequence of SEQ ID NO:62, and LCDR3 comprises the amino acid sequence of SEQ ID NO:159; or the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 160, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 161, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 162; Preferably, a. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 121, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; b. the heavy chain variable region has an HCDR1 comprising the amino acid sequence of SEQ ID NO: 135, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 203, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 137, and the light chain variable region has an LCDR1 comprising the amino acid sequence of SEQ ID NO: 138, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 139, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 140; or the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 111, HCDR2 comprises the amino acid sequence of SEQ ID NO: 173, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 113, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 115, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 116; or d. the heavy chain variable region comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 69, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 189, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 71, and the light chain variable region comprises an LCDR1 comprising the amino acid sequence of SEQ ID NO: 61, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 62, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 72; or e. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 165, HCDR2 comprises the amino acid sequence of SEQ ID NO: 166, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 167, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 224, LCDR2 comprises the amino acid sequence of SEQ ID NO: 169, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 170; or f. the heavy chain variable region comprises an HCDR1 comprising the amino acid sequence of SEQ ID NO: 105, an HCDR2 comprising the amino acid sequence of SEQ ID NO: 125, and an HCDR3 comprising the amino acid sequence of SEQ ID NO: 126, and the light chain variable region comprises an LCDR1 comprising the amino acid sequence of SEQ ID NO: 127, an LCDR2 comprising the amino acid sequence of SEQ ID NO: 109, and an LCDR3 comprising the amino acid sequence of SEQ ID NO: 128; or g. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 141, HCDR2 comprises the amino acid sequence of SEQ ID NO: 215, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 216, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 144, LCDR2 comprises the amino acid sequence of SEQ ID NO: 145, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 146; More preferably, a. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 105, HCDR2 comprises the amino acid sequence of SEQ ID NO: 120, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 121, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 114, LCDR2 comprises the amino acid sequence of SEQ ID NO: 109, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 119; b. the heavy chain variable region, HCDR1 comprises the amino acid sequence of SEQ ID NO: 135, HCDR2 comprises the amino acid sequence of SEQ ID NO: 203, and HCDR3 comprises the amino acid sequence of SEQ ID NO: 137, and the light chain variable region, LCDR1 comprises the amino acid sequence of SEQ ID NO: 138, LCDR2 comprises the amino acid sequence of SEQ ID NO: 139, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 140; The anti-ILT4 antibody of claim 1.

3. The anti-ILT4 antibody according to claim 1 or 2, which is a murine antibody, a chimeric antibody, a humanized antibody or a fully human antibody, preferably a humanized antibody.

4. the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 184 or 185, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 187 or 186, or the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 32, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 33; or b. the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 211, 205, 206, 207, 208, 209, or 210, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 212, 213, or 214; or the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:40, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:41; or c. the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 181, 176, 177, 178, 179, or 180, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 183 or 182; or the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:28, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:29; or d. the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 193, 190, 191, or 192, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 194, 195, or 196; or the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 10, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 11; or e. the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 163, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 225 or 164; f. the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 197 or 198, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 199 or 200; or the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 36, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 37; or g. the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 217, 218, 219, 220, or 221, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 222 or 223; or the heavy chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 42, and the light chain variable region comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 43; Preferably, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 184 or 185, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 187 or 186, or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 32, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 33; or b. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 211, 205, 206, 207, 208, 209, or 210, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 212, 213, or 214; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:40, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:41; or c. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 181, 176, 177, 178, 179, or 180, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 183 or 182; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:28, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:29; or d. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 193, 190, 191, or 192, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 194, 195, or 196; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 10, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 11; or e. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 163, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 225 or 164; f. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 197 or 198, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 199 or 200; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 36, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 37; or g. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 217, 218, 219, 220, or 221, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 222 or 223; or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 42, and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 43; The anti-ILT4 antibody according to any one of claims 1 to 3.

5. The anti-ILT4 antibody according to any one of claims 1 to 4, wherein the anti-ILT4 antibody is an antibody fragment, and preferably, the antibody fragment is Fab, Fab', F(ab')2, Fd, Fv, scFv, dsFv or dAb.

6. the anti-ILT4 antibody comprises a heavy chain constant region and a light chain constant region; Preferably, the heavy chain constant region is a human IgG1 or IgG4 heavy chain constant region and the light chain constant region is a human κ light chain constant region; More preferably, the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 240 or 171, and the light chain constant region comprises the amino acid sequence of SEQ ID NO:

172. The anti-ILT4 antibody according to any one of claims 1 to 4.

7. The anti-ILT4 antibody comprises a heavy chain and a light chain, a. the heavy chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 228, and the light chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 229, or b. the heavy chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:234, and the light chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:235, or c. the heavy chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 226, and the light chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 227, or d. the heavy chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 230, and the light chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 231; or e. the heavy chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 238, and the light chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 239, or f. the heavy chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 232, and the light chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO: 233, or g. the heavy chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:236, and the light chain comprises an amino acid sequence having at least 70% sequence identity to SEQ ID NO:237; Preferably, a. the amino acid sequence of the heavy chain is set forth in SEQ ID NO:228 and the amino acid sequence of the light chain is set forth in SEQ ID NO:229, or b. the amino acid sequence of the heavy chain is set forth in SEQ ID NO:234 ​​and the amino acid sequence of the light chain is set forth in SEQ ID NO:235; or c. the amino acid sequence of the heavy chain is set forth in SEQ ID NO:226 and the amino acid sequence of the light chain is set forth in SEQ ID NO:227, or d. the amino acid sequence of the heavy chain is set forth in SEQ ID NO:230 and the amino acid sequence of the light chain is set forth in SEQ ID NO:231; or e. the amino acid sequence of the heavy chain is set forth in SEQ ID NO:238 and the amino acid sequence of the light chain is set forth in SEQ ID NO:239, or f. the amino acid sequence of the heavy chain is set forth in SEQ ID NO:232 and the amino acid sequence of the light chain is set forth in SEQ ID NO:233; or g. The amino acid sequence of the heavy chain is set forth in SEQ ID NO:236, and the amino acid sequence of the light chain is set forth in SEQ ID NO:

237. The anti-ILT4 antibody of claim 6.

8. 8. A pharmaceutical composition comprising the anti-ILT4 antibody of any one of claims 1 to 7 and one or more pharmaceutically acceptable carriers, diluents, or excipients, and preferably further comprising at least one second therapeutic agent, more preferably the second therapeutic agent is an anti-PD-1 antibody.

9. An isolated nucleic acid encoding the anti-ILT4 antibody of any one of claims 1 to 7.

10. A host cell comprising the isolated nucleic acid of claim 9.

11. A method for blocking the binding of ILT4 to HLA-G in a human subject in need thereof, comprising administering to the human subject an effective amount of an anti-ILT4 antibody of any one of claims 1 to 7.

12. 10. A method for detecting an ILT4 peptide or a fragment thereof in a sample, the method comprising contacting the sample with an anti-ILT4 antibody according to any one of claims 1 to 7 and detecting the presence of a complex between the anti-ILT4 antibody and the ILT4 peptide or fragment thereof, wherein detection of the complex indicates the presence of the ILT4 peptide or fragment thereof.

13. A method for treating a disease, the method comprising administering to a subject a therapeutically effective amount of the anti-ILT4 antibody of any one of claims 1 to 7 or the pharmaceutical composition of claim 8; Preferably, the disease is a tumor, and more preferably, the disease is selected from the group consisting of astrocytoma, glioblastoma, bladder cancer, bone cancer, brain cancer, breast cancer, carcinoid, cervical cancer, choroid plexus papilloma, colorectal cancer, fallopian tube cancer, ependymoma, gallbladder cancer, gastric cancer, head and neck cancer, idiopathic myelofibrosis, kidney cancer, renal pelvic tumor, leukemia, liver cancer, esophageal cancer, lung cancer, lymphoma, medulloblastoma, melanoma, meningioma, Merkel cell carcinoma, mesothelioma, and the like. tumor, multiple myeloma, neuroblastoma, oligodendroglioma, ovarian cancer, peritoneal tumor, pancreatic cancer, polycythemia vera, primary neuroectodermal tumor, prostate cancer, retinoblastoma, salivary gland cancer, sarcoma, small intestine cancer, colon cancer, squamous cell carcinoma, thyroid cancer, endometrial cancer, vestibular schwannoma, blastoma, teratoma, pituitary cancer, vulvar cancer, thymoma, testicular cancer, bile duct cancer, pheochromocytoma, paraganglioma and adenoid cystic carcinoma; Most preferably, the disease is selected from ovarian cancer, lung cancer, esophageal cancer, colorectal cancer, renal cell carcinoma and melanoma.

14. The method for treating a disease according to claim 13, further comprising using a second therapeutic agent, preferably wherein the second therapeutic agent comprises an immune checkpoint inhibitor, more preferably wherein the second therapeutic agent is an anti-PD-1 antibody.

15. The anti-PD-1 antibody comprises a heavy chain variable region PD-1-VH and a light chain variable region PD-1-VL, wherein: the PD-1-VH has a PD-1-HCDR1 comprising the amino acid sequence of SEQ ID NO: 250, a PD-1-HCDR2 comprising the amino acid sequence of SEQ ID NO: 251, and a PD-1-HCDR3 comprising the amino acid sequence of SEQ ID NO: 252; and the PD-1-VL has a PD-1-LCDR1 comprising the amino acid sequence of SEQ ID NO: 253, a PD-1-LCDR2 comprising the amino acid sequence of SEQ ID NO: 254, and a PD-1-LCDR3 comprising the amino acid sequence of SEQ ID NO: 255; Preferably, the PD-1-VH comprises the amino acid sequence of SEQ ID NO:256, and the PD-1-VL comprises the amino acid sequence of SEQ ID NO:

257.

15. A method for treating the disease of claim 14.

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