Cannabinoid Receptor Type 2 Antibody and Its Uses

Abstract

The present application provides a CB2 antibody and a construct comprising an immunoglobulin single variable domain (ISVD) that specifically binds to CB2. A method for manufacturing and using such a CB2 antibody is also provided.

Description

Technology Field

[0001] Cross-reference regarding related applications

[0002] The present application claims priority to U.S. Provisional Application No. 63 / 609,666, filed December 13, 2023; U.S. Provisional Application No. 63 / 561,029, filed March 4, 2024; U.S. Provisional Application No. 63 / 645,515, filed May 10, 2024; U.S. Provisional Application No. 63 / 679,061, filed August 2, 2024; and U.S. Provisional Application No. 63 / 716,676, filed November 5, 2024, the contents of which are incorporated herein by reference in their entirety.

[0003] Reference to the electronic sequence list

[0004] The contents of the electronic sequence list (185982000340SEQLIST.xml, size: 291,969 bytes, creation date: December 11, 2024) are incorporated herein by reference in their entirety.

[0005] Statements regarding research or development sponsored by the federal government

[0006] The present invention was made with government support under Project Nos. R43DK125191 and R44DK125191 assigned by the National Institutes of Health, and Project Nos. R43CA241513 and R44CA241513 assigned by the National Institutes of Health. The government holds specific rights to the present invention.

[0007] The present invention relates to an antibody comprising an immunoglobulin single variable domain (ISVD) and specifically binding to cannabinoid receptor type 2 (CB2), and methods for manufacturing and using the same. Background Technology

[0008] Mammals possess two G protein-coupled receptors that mediate the signaling of cannabinoids found in both plants and animals. Cannabinoid receptor 1 (CB1) is most strongly expressed in neurons of the CNS and is primarily responsible for the psychoactive effects of cannabinoids on the body. Nature , 346: 561-564, 1990). CB2 is predominantly expressed in the spleen and hematopoietic cells, showing that this receptor is mainly localized in the periphery (Literature [ Nature , 365: 61-65, 1993], And this is literature[ Pharmacol. Rev. Reviewed in 58(3): 389-462, 2006).

[0009] CB2 agonists are effective in numerous models of acute and chronic pain resulting from chemical, mechanical, and thermal pain stimuli (Literature [ Pharmacol. Ther. 95: 127-135 2002] and literature[ Pharmacol. Rev. (Reviewed in 58(3): 389-462, 2006). In chemotherapy-induced neuropathic pain (CIPN), CB2 activation was shown to inhibit paclitaxel-induced CIPN in rats ( J. Pharmacol. Exp. Ther. 327 : 584-591, 2008). Furthermore, CB2 agonists are taxanes ( in other words , the paclitaxel cited above), vinca alkaloids ( in other words , vincristine, literature[ Br. J. Pharmacol. 152:765-777, 2007]) and platinum origin( in other words , cisplatin, literature[ Mol Pain [ , 8:71]) inhibited CIPN induced by all major classes of chemotherapy agents, including anticancer drugs. CB2 agonists also inhibited rats ( Pharmacology. 82(3):193-200, 2008) and mouse models (literature[ Pain . 2013 Jun;154(6):864-73]; Literature[ Psychopharmacology[(Berl). 233(11):2209-2219, 2016]) It was found to be an effective treatment for diabetic neuropathy in all cases, which further supports the use of CB2 agonists in the treatment of multiple pain indications.

[0010] CB2 activation possesses potent anti-inflammatory activity as part of a natural negative feedback mechanism, where CB2 receptors are induced in damaged or virus-infected cells in an inflammatory environment (Literature[ Gastroenterology . 129:437-453, 2005]; literature[ Am. J. Pathol . 177:187-196, 2010]; literature[ Br. J. Pharmacol. 151:1041-1048, 2007]). CB2 activation subsequently inhibits the secretion of numerous inflammatory factors and signals, including TNFα, IL-6, IL-8, and CCL2 (Literature[ Cell . Mol. Life Sci Reviewed in . 73:4449-4470, 2016). Therefore, CB2 agonists are used for inflammatory conditions of the lungs ( Biomed Res. Int . 2014: 971750, 2014), atherosclerosis( Eur. J. Pharmacol. 649:285-292,2010), inflammatory bowel disease( World J. Gastroenterol . 22:9515-9524, 2016) and virus infection( Virulence It will be useful for the treatment of (9:217-230, 2018).

[0011] CB2 receptor agonism also plays a role in alleviating fibrotic processes in multiple organs. For example, the receptor agonist JWH-133 was shown to reduce leukocyte infiltration and dermal thickening in bleomycin-induced experimental fibrosis ( Arthritis Rheum 60:1129-1136, 2009), alleviates bleomycin-induced pulmonary fibrosis in mice ( Oncotarget.8:103486-103498, 2017). The CB2 receptor has been identified as a potential target for the treatment of systemic sclerosis because it regulates both cutaneous fibroblast proliferation and autoimmune responses ( Am. J. Pathol. 177: 187-196, 2010). In the case of the liver, CB2 action in a mouse model is carbon tetrachloride (CCl4)( Gastroenterology. 128:742-755, 2005) and alcohol( Hepatology It protects against liver fibrosis induced by ( . 54:1217-1226, 2011) and enhances liver regeneration after injury ( Hepatology . 52:1046-1059, 2010). In addition to reducing inflammation, CB2 action reduces the proliferation of myofibroblasts that secrete collagen forming fibrous lesions and induces apoptosis ( Gastroenterology . 128:742-755, 2005). Therefore, CB2 is It can have a beneficial effect on liver disease through multiple mechanisms, such as inflammation and fibrosis.

[0012] There is a need in the industry to develop methods for using CB2 receptor agonists in safe and effective therapy and to reduce the risk of adverse events. CB2 small molecule agonists are rapidly cleared, can have significant CNS permeability, and carry a high risk of drug metabolic effects. Even the most peripherally restricted CB2 small molecules have a CNS penetration rate of 3 to 5% ( Pain Apr 13, 2021). Most CB2 agonists exhibit partial activity at CB1, and therefore, CNS-mediated side effects are a concern. For example, in a Phase 2 study of olorinab, a CB2 small molecule agonist developed for the treatment of irritable bowel syndrome, approximately 20% of treated patients reported some degree of dizziness, drowsiness, or other central signs of CB1 activation ( Higgins , et al 14th Congress of ECCO - Inflammatory Bowel Diseases; March 6-9, 2019; Copenhagen, Denmark).In addition, the short half-life of small molecules adds to the disadvantage in terms of compliance, which makes multiple daily administration inevitable ( J. Behav. Med. 31: 213-224, 2008). The antibody described herein meets these requirements of the industry and also provides related benefits.

[0013] All publications, patents, patent applications, and disclosed patent applications mentioned herein are incorporated herein by reference in their entirety. means of solving the problem

[0014] The present application relates to an anti-CB2 antibody (hereinafter referred to as the 'CB2 antibody') comprising an immunoglobulin single variable domain (ISVD) that specifically binds to CB2, and methods for manufacturing and using the same.

[0015] In one embodiment, the present invention provides a CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD comprises a complementarity determining region (CDR) 1 comprising the amino acid sequence of SEQ ID NO. 1, a CDR2 comprising the amino acid sequence of SEQ ID NO. 2, and a CDR3 comprising the amino acid sequence of SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. Additionally, a CB2 antibody comprising an ISVD that specifically binds to CB2 is provided, wherein the ISVD comprises CDR1, CDR2, and CDR3 of the ISVD comprising the amino acid sequence of SEQ ID NO. 201.

[0016] In some embodiments, ISVD is the heavy chain variable domain (VHH) of a heavy chain antibody. In some embodiments, ISVD is of camelid origin. In some embodiments, ISVD is a chimeric form. In some embodiments, ISVD is humanized.

[0017] In some embodiments, the ISVD comprises one or more amino acid residues selected from the group consisting of V5, Q39, Y58, G65, R71, A74, and L78 in one or more framework regions (FR), wherein the amino acid numbering follows Kabat. In some embodiments, the ISVD comprises amino acid residues V5, Q39, Y58, and G65 in one or more FRs.

[0018] In some embodiments, the ISVD comprises FR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs 4 to 15, FR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs 16 to 30, FR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs 31 to 48 and SEQ ID NO 200, and FR4 having an amino acid sequence selected from the group consisting of SEQ ID NOs 49 to 54. In some embodiments, the ISVD comprises (i) FR1 having an amino acid sequence of SEQ ID NO 14, FR2 having an amino acid sequence of SEQ ID NO 29, FR3 having an amino acid sequence of SEQ ID NO 43, and FR4 having an amino acid sequence of SEQ ID NO 54; (ii) FR1 having an amino acid sequence of SEQ ID NO 15, FR2 having an amino acid sequence of SEQ ID NO 23, FR3 having an amino acid sequence of SEQ ID NO 45, and FR4 having an amino acid sequence of SEQ ID NO 54; or (iii) includes FR1 having the amino acid sequence of SEQ ID NO. 4, FR2 having the amino acid sequence of SEQ ID NO. 16, FR3 having the amino acid sequence of SEQ ID NO. 200, and FR4 having the amino acid sequence of SEQ ID NO. 49.

[0019] In some embodiments, the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs 55 to 77 and 201, or a variant thereof having at least about 85% sequence identity with any one of SEQ ID NOs 55 to 77 and SEQ ID NO 201. In some embodiments, the ISVD comprises the amino acid sequence of SEQ ID NO 200, 74, or 94.

[0020] In some embodiments, the CB2 antibody comprises an Fc region. In some embodiments, the antibody comprises an Fc region of IgG1 or IgG4. In some embodiments, the antibody comprises an IgG4 variant Fc region exhibiting reduced effector function. In some embodiments, the Fc region comprises amino acid substitutions F234A and L235A according to the EU index numbering of Kabat. In some embodiments, the Fc region comprises the amino acid sequence of SEQ ID NO. 130.

[0021] In some embodiments, the antibody includes a hinge region. In some embodiments, the hinge region includes the amino acid sequence of SEQ ID NO. 131.

[0022] In some embodiments, the ISVD is fused to the Fc region through a peptide linker. In some embodiments, the peptide linker comprises the amino acid sequence of SEQ ID NO. 132. In some embodiments, the antibody comprises a polypeptide chain comprising the ISVD, the peptide linker, the hinge region, and the Fc region from the N-terminus to the C-terminus.

[0023] In some embodiments, the CB2 antibody comprises a polypeptide chain comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 78 to 95 and SEQ ID NOs 202 to 203, or a variant thereof having at least about 85% sequence identity with any one of SEQ ID NOs 78 to 95 and SEQ ID NOs 202 to 203. In some embodiments, the CB2 antibody comprises a polypeptide chain comprising the amino acid sequence of SEQ ID NO. 94. In some embodiments, the CB2 antibody comprises a polypeptide chain comprising the amino acid sequence of SEQ ID NO. 203.

[0024] In some embodiments, CB2 is a human, mouse, rat, or cynomolgus monkey CB2. In some embodiments, CB2 is a human CB2.

[0025] In some embodiments, the CB2 antibody is a CB2 agonist. In some embodiments, the CB2 antibody does not act as an agonist for CB1. In some embodiments of the CB2 antibody, (i) the ISVD lowers the forskolin-inducible cyclic adenosine monophosphate (cAMP) level in HEK293 cells overexpressing human CB2 to a half-maximum effective concentration (EC) of less than 55 nM. 50 (ii) reduces and / or lowers forskolin-inducible cyclic adenosine monophosphate (cAMP) levels to less than 30 nM EC in mouse CB2-overexpressing HEK293 cells 50 Reduces to. In some embodiments, ISVD does not specifically bind to cannabinoid receptor type 1 (CB1). In some embodiments, the CB2 antibody is applied to HEK293 cells overexpressing human CB2 at an EC of less than 850 nM. 50 Combines as.

[0026] An isolated nucleic acid encoding any one of the CB2 antibodies described above is further provided. An expression vector comprising said nucleic acid is further provided. A host cell comprising said nucleic acid or said expression vector is further provided.

[0027] A method for producing any one of the CB2 antibodies described above is further provided, the method comprising the step of culturing the host cells described above under conditions in which the CB2 antibodies are produced. In some embodiments, the method further comprises the step of recovering the CB2 antibodies produced by the host cells.

[0028] A pharmaceutical composition comprising any one of the CB2 antibodies described above and a pharmaceutically acceptable carrier is further provided.

[0029] A method acting as an agonist for CB2 on cells is further provided, comprising the step of contacting a cell with any one of the CB2 antibodies described above in an amount sufficient to activate CB2 on cells. In some embodiments, the method in vitro The method is performed. In some embodiments, the method in vivo It is performed.

[0030] A method for treating an individual requiring treatment of a disease or pathological condition is further provided, wherein acting as an agonist for CB2 improves the said disease or pathological condition, and the method comprises the step of administering to the individual an effective amount of any one of the CB2 antibodies described above or the pharmaceutical composition described above. In some embodiments, the disease or pathological condition is selected from the group consisting of chemotherapy-induced peripheral neuropathy (CIPN), diabetic peripheral neuropathy, respiratory infection, hepatic fibrosis, cold sensitivity, inflammatory bowel disease, and endometriosis. In some embodiments, the disease or pathological condition is chemotherapy-induced peripheral neuropathy (CIPN). In some embodiments, CIPN is induced by a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the disease or pathological condition is a respiratory infection. In some embodiments, the respiratory infection is induced by a virus selected from the group consisting of influenza viruses and coronaviruses. In some embodiments, the respiratory infection is induced by SARS-CoV-2. In some embodiments, the respiratory infection is caused by H1N1 influenza. In some embodiments, the disease or condition is hepatic fibrosis. In some embodiments, the disease or condition is diabetic peripheral neuropathy.

[0031] A method for treating cancer in an individual requiring treatment for cancer is further provided, the method comprising the step of administering to the individual an effective amount of a chemotherapy agent and an effective amount of any one of the CB2 antibodies described above.

[0032] A method for alleviating cytokine release syndrome in an individual requiring alleviation of cytokine release syndrome is further provided, comprising the step of administering an effective amount of any one of the CB2 antibodies described above to the individual. In some embodiments, this method reduces the levels of IL-16 and / or IL-8 secreted by the airway cells of the individual.

[0033] In some embodiments of any of the methods described above, the individual is a human.

[0034] A kit comprising any one of the CB2 antibodies described above, the pharmaceutical composition described above, and instructions for use is further provided.

[0035] A CB2 antibody is further provided comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD is i) a complementarity determining region (CDR) 1 having the amino acid formula G-X1-X2-X3-SI-X4-X5, said formula where X1 is selected from D, S, or Q; X2 is selected from G, H, I, or K; X3 is selected from D, E, F, G, H, I, K, N, Q, R, S, T, V, W, or Y; X4 is selected from G, K, M, R, or Y; and X5 is selected from A or G, said CDR 1; and ii) a CDR3 having the amino acid formula C1-C2-C3-IK-C4-C5-C6-C7-C8-C9-C10-I-C11-C12-C13, wherein C1 is selected from F, H, I, K, L, R, S, T, V, or Y; C2 is selected from A, E, F, G, I, L, Q, R, or V; C3 is selected from A, F, G, I, K, L, N, or V; C4 is selected from D, F, N, T, or Y; C5 is selected from A, G, or R; C6 is selected from R, S, or W; C7 is selected from D, G, K, or R; C8 is selected from D, I, L, Q, R, S, or T; C9 is selected from F, L, or T; and C10 is selected from D, E, H, or Q; C11 is selected from K or N; C12 is selected from A, F, G, H, I, K, L, N, P, R, S, V, T or W; and C13 is selected from A, D, F, G, H, T or Y, comprising CDR3. In some embodiments, CDR2 is SEQ ID NO. 2.

[0036] A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2) is further provided, wherein the ISVD comprises a CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 248 to 288, a CDR2 comprising an amino acid sequence of SEQ ID NO. 2, and a CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 289 to 329, and CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs 205 to 212, SEQ ID NOs 214 to 229, and SEQ ID NOs 231 to 247, or a variant thereof having at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs 205 to 212, SEQ ID NOs 214 to 229, and SEQ ID NOs 231 to 247. In some embodiments, the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs 205, 212, 215, 217, 218, 222, 225, 226, 227, 229, 240 and 241, or a variant thereof having at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs 205, 212, 215, 217, 218, 222, 225, 226, 227, 229, 240 and 241.

[0037] In some embodiments, the ISVD is of camelid origin. In some embodiments, the ISVD is a chimeric form. In some embodiments, the ISVD is humanized. In some embodiments, the antibody comprises an Fc region. In some embodiments, the antibody comprises an Fc region of IgG1 or IgG4. In some embodiments, the antibody comprises a variant IgG4 Fc region exhibiting reduced effector function. In some embodiments, the antibody comprises a hinge region. In some embodiments, CB2 is human, mouse, rat, or cynomolgus monkey CB2. In some embodiments, CB2 is human CB2. In some embodiments, the CB2 antibody is a CB2 agonist. In some embodiments, the CB2 antibody does not act as an agonist for CB1.

[0038] An isolated nucleic acid encoding any of the CB2 antibodies described above is additionally provided.

[0039] An expression vector comprising a nucleic acid encoding any of the CB2 antibodies described above is further provided.

[0040] A host cell comprising a nucleic acid encoding any of the CB2 antibodies described above, or an expression vector containing said nucleic acid is further provided.

[0041] A method for producing a CB2 antibody is further provided, comprising the step of culturing any of the host cells described above under conditions in which a CB2 antibody is produced. In some embodiments, the method further comprises the step of recovering the CB2 antibody produced by the host cell.

[0042] A pharmaceutical composition comprising any of the CB2 antibodies described above and a pharmaceutically acceptable carrier is further provided.

[0043] A method acting as an agonist for CB2 on a cell is further provided, comprising the step of contacting a cell with any of the CB2 antibodies described above in an amount sufficient to activate CB2 on the cell. In some embodiments, the method In a test tube The method is performed. In some embodiments, the method In vivo It is performed in.

[0044] A method for treating an individual requiring treatment for a disease or pathological condition is further provided, wherein acting as an agonist for CB2 improves the said disease or pathological condition, and the method comprises the step of administering an effective amount of a CB2 antibody to the individual. In some embodiments, the disease or pathological condition is selected from the group consisting of chemotherapy-induced peripheral neuropathy (CIPN), diabetic peripheral neuropathy, respiratory infection, hepatic fibrosis, cold sensitivity, inflammatory bowel disease, and endometriosis. In some embodiments, the disease or pathological condition is chemotherapy-induced peripheral neuropathy (CIPN). In some embodiments, CIPN is induced by a taxol drug. In some embodiments, the taxol drug is paclitaxel.

[0045] A method for treating cancer in an individual requiring treatment for cancer is further provided, the method comprising the step of administering an effective amount of a chemotherapy agent and an effective amount of a CB2 antibody to the individual. In some embodiments, the disease or condition is a respiratory infection. In some embodiments, the respiratory infection is caused by a virus selected from the group consisting of influenza viruses and coronaviruses. In some embodiments, the respiratory infection is caused by SARS-CoV-2. In some embodiments, the respiratory infection is caused by H1N1 influenza.

[0046] A method for alleviating cytokine release syndrome in an individual requiring alleviation of cytokine release syndrome is further provided, comprising the step of administering an effective amount of any of the CB2 antibodies described above to the individual. In some embodiments, this method reduces IL-16 and / or IL-8 levels secreted by the airway cells of the individual. In some embodiments, the disease or condition is hepatic fibrosis. In some embodiments, the disease or condition is diabetic peripheral neuropathy. In some embodiments, the individual is a human.

[0047] A kit is further provided comprising any of the CB2 antibodies described above, or a pharmaceutical composition comprising any of the CB2 antibodies described above and a pharmaceutically acceptable carrier, and instructions for use.

[0048] A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2) is further provided, wherein the ISVD comprises a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; C6 is selected from Y, A, or F, and CDR3 is defined according to IMGT. In some embodiments, CDR1 is SEQ ID NO. 248 and CDR2 is SEQ ID NO. 2, and CDR1 and CDR2 are defined according to IMGT.

[0049] A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2) is further provided, wherein the ISVD comprises a CDR1 comprising the amino acid sequence of SEQ ID NO. 248, a CDR2 comprising the amino acid sequence of SEQ ID NO. 2, and a CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs. 345 to 359, and CDR1, CDR2 and CDR3 are defined according to IMGT. In some embodiments, the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs. 330 to 344, or a variant thereof having at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs. 330 to 344. In some embodiments, the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343 and 344, or a variant thereof having at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343 and 344.

[0050] In some embodiments, the ISVD is of camelid origin. In some embodiments, the ISVD is a chimeric form. In some embodiments, the ISVD is humanized. In some embodiments, the antibody comprises an Fc region. In some embodiments, the antibody comprises the Fc region of IgG1 or IgG4.

[0051] In some embodiments, the antibody comprises a variant IgG4 Fc region exhibiting reduced effector function. In some embodiments, the antibody comprises a hinge region. In some embodiments, CB2 is human, mouse, rat, or cynomolgus monkey CB2. In some embodiments, CB2 is human CB2. In some embodiments, the CB2 antibody is a CB2 agonist. In some embodiments, the CB2 antibody does not act as an agonist for CB1.

[0052] An isolated nucleic acid encoding any of the CB2 antibodies described above is additionally provided.

[0053] An expression vector comprising any of the nucleic acids described above is additionally provided.

[0054] A host cell comprising any of the nucleic acids described above or any of the expression vectors described above is further provided.

[0055] A method for producing a CB2 antibody is further provided, comprising the step of culturing any of the host cells described above under conditions in which a CB2 antibody is produced. In some embodiments, the method further comprises the step of recovering the CB2 antibody produced by the host cell.

[0056] A pharmaceutical composition comprising any of the CB2 antibodies described above and a pharmaceutically acceptable carrier is further provided.

[0057] A method acting as an agonist for CB2 on a cell is further provided, comprising the step of contacting a cell with any of the CB2 antibodies described above in an amount sufficient to activate CB2 on the cell. In some embodiments, the method In a test tube The method is performed. In some embodiments, the method In vivo It is performed.

[0058] A method for treating an individual requiring treatment for a disease or pathological condition is further provided, wherein acting as an agonist for CB2 improves the said disease or pathological condition, and the method comprises the step of administering an effective amount of any of the CB2 antibodies described above to the individual. In some embodiments, the disease or pathological condition is selected from the group consisting of chemotherapy-induced peripheral neuropathy (CIPN), diabetic peripheral neuropathy, respiratory infection, hepatic fibrosis, cold sensitivity, inflammatory bowel disease, and endometriosis. In some embodiments, the disease or pathological condition is chemotherapy-induced peripheral neuropathy (CIPN). In some embodiments, CIPN is induced by a taxol drug. In some embodiments, the taxol drug is paclitaxel.

[0059] A method for treating cancer in an individual requiring treatment for cancer is further provided, comprising the step of administering to the individual an effective amount of a chemotherapy agent and an effective amount of any of the CB2 antibodies described above. In some embodiments, the disease or condition is a respiratory infection. In some embodiments, the respiratory infection is caused by a virus selected from the group consisting of influenza viruses and coronaviruses. In some embodiments, the respiratory infection is caused by SARS-CoV-2. In some embodiments, the respiratory infection is caused by H1N1 influenza.

[0060] A method for alleviating cytokine release syndrome in an individual requiring alleviation of cytokine release syndrome is further provided, comprising the step of administering an effective amount of any of the CB2 antibodies described above to the individual. In some embodiments, this method reduces IL-16 and / or IL-8 levels secreted by the airway cells of the individual. In some embodiments, the disease or condition is hepatic fibrosis. In some embodiments, the disease or condition is diabetic peripheral neuropathy. In some embodiments, the individual is a human.

[0061] A kit comprising any of the CB2 antibodies described above or any of the pharmaceutical compositions described above, and instructions for use is further provided.

[0062] A CB2 antibody is further provided comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD comprises a CDR1 having the amino acid sequence of SEQ ID NO. 248; a CDR2 having the amino acid sequence of SEQ ID NO. 2; and a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; and C6 is selected from Y, A, or F, and CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the ISVD comprises a CDR1 having the amino acid sequence of SEQ ID NO. 248; It includes CDR2 comprising the amino acid sequence of SEQ ID NO. 2; and CDR3 comprising the amino acid sequence selected from the group consisting of SEQ ID NOs. 345 to 359. Brief explanation of the drawing

[0063] Fig. 1a is the agonist activity against cannabinoid receptors (EC) evaluated by cAMP levels expressed as a percentage relative to the control group. 50 The values ​​are presented. For human CB2, the agonist activities of the negative control ('None'), CP55940, CP55940+SR144528, antibody named AB101 (sequence number 55), AB101+SR144528, or SR144528 are presented, and for human CB1, the agonist activities of the negative control ('None'), CP55940, AB101, and negative control VHH are presented. Fig. 1b Is This shows the action of the antibody named AB101 (SEQ No. 55) on human and mouse CB2. . Fig. 2a Is Shows the activity of ABt140 (SEQ No. 94) or the vehicle control in CIPN model mice ('CIPN') or cremophor vehicle control mice ('Healthy'), as evaluated by mechanical avoidance threshold. Fig. 2b Is This shows the activity of Abt140 (SEQ No. 94) or the vehicle control in CIPN model mice ('CIPN') or cremophor vehicle control mice ('Healthy'), evaluated by cold allodynia. Fig. 3a Is ABt140 (sequence number 94) and CP55940 show agonist activity against human CB2. Fig. 3b Is This shows the agonist activity of ABt140 (SEQ No. 94) and control antibodies, evaluated by inhibition of IL6 secretion from LPS-stimulated BEAS2B cells. Fig. 3c Is This shows the agonist activity of ABt140 (SEQ No. 94) and control antibodies, evaluated by inhibition of IL8 secretion from LPS-stimulated BEAS2B cells. Fig. 4a Is This shows the effect of ABt140 (sequence number 94) in a mouse influenza model, evaluated by the improvement of clinical scores. Fig. 4b Is This shows the effect of ABt140 (sequence number 94) in a mouse influenza model, evaluated by 7-day survival rate. Fig. 4c Is This shows the effect of ABt140 (sequence number 94) in a mouse influenza model, evaluated by viral titer. FIGS. 5a to 5b Is After treating serum-starved RAW264.7 mouse macrophages with HU-308 (Onternabez), ABt281, ABt285, or ABt269 (1 μM and 100 nM) for 6 hours, LPS (100 ng / ml; Fig. 5a ) or IL-4(10 ng / ml; Fig. 5b The results are shown after treatment with ) for 12 hours. Gene expression in M1 and M2 macrophages was measured using RTPCR. Two-way ANOVA: * p<0.05, *** p=0.0001, **** p<0.0001. Fig. 6 This represents IL-6 secretion from RAW264.7 cells after treatment with CP-55,940 or the labeled antibody in the presence (+) or absence (-) of LPS. IL-6 concentration in the supernatant was measured using the Perkin Elmer IL-6 LANCE Ultra TR-FRET kit TRF1223C. Fig. 7 silver This represents the cAMP levels of RAW264.7 cells treated with NKH477 and HU308, ABt281, ABt285, or ABt269, as evaluated using the luminescent cAMP kit (cAMP-Glo, Promega, V1501). FIGS. 8a to 8b is mechanical stimulation ( Fig. 8a ) and cold stimulation ( Fig. 8b Shows the activity of Cremophor vehicle control, PBS control, AB269, or ABt281 in the mechanical allodynia assay in mice against ) 6 mice (N=6). Data are mean ± SEM. B: Baseline before paclitaxel administration. Pac: Baseline after paclitaxel administration. *** P For <0.001, statistics for CIPN+antibody (ABt281) versus CIPN+control (ABt269) were analyzed using one-way ANOVA followed by the Bonferroni post hoc test. FIGS. 9a to 9b is mechanical ( Fig. 9a ) and cold( Fig. 9b CB2 in response to ) stimulation In the test of mechanical and cold allodynia in knockout mice Shows the activity of the CIPN+control group (ABt269) versus the CIPN+antibody (ABt281). B: Baseline before paclitaxel administration. Pac: Baseline after paclitaxel administration. Statistics for the CIPN+antibody (ABt281) versus the CIPN+control group (ABt269) were analyzed using Bonferroni post-hoc tests following one-way ANOVA. FIGS. 10a to 10b Is Mechanical stimulation ( Fig. 10a ) and cold stimulation ( Fig. 10b CIPN+control (ABt269) versus CIPN+antibody (ABt281) in the assay of mechanical and cold allodynia in mice activity represents. FIGS. 11a to 11b Mechanical stimulation after AM1710 treatment ( Fig. 11a ) and cold stimulation ( Fig. 11b This shows the activity of CIPN+ antibody (ABt281) versus CIPN+ control (ABt269) in mechanical and cold allodynia tests in mice. FIGS. 12a to 12f Is It demonstrates the effect of the indicated treatment on human liver section model (hPCLS) cells, which is the secretion of collagen 1a1 (Col1a1) into the medium evaluated via ELISA ( Fig. 12a ), and COL1A1( Fig. 12b ), ACTA2( Fig. 12c ), IL6( Fig. 12d ), IL10( Fig. 12e ) and TNFα( Fig. 12f Evaluated via mRNA expression levels for ). Two-way ANOVA; Dunnets multiple comparison test; mean ± standard deviation; *p<0.05, **p<0.005; ns=not significant. FIGS. 13a to 13b Is This represents ERK phosphorylation induced by the indicated agents in RAW264.7 cells, evaluated using the TR-FRET pERK1 / 2 kit. FIGS. 14a to 14b Is Mechanical threshold after the initial injection and two subsequent re-injections ( Fig. 14a ) or cold allodynia ( Fig. 14b This shows the activity of ABt140 (SEQ No. 94), ABt281 (SEQ No. 202), ABt285 (SEQ No. 203), or the vehicle control in CIPN model mice ('CIPN') or cremophor vehicle control mice ('Healthy'), as evaluated by ). Fig. 15 Is This represents the mobilization of beta-arestin 2 induced by the indicated agent in RAW264.7 cells, as evaluated by the HTRF beta-arestin 2 mobilization detection kit. Fig. 16 silver RAW264.7 cells cultured and treated with HU308, CB2 agonist antibody, or a homozygous control (ABt269). Fig. 17 RAW264.7 cells cultured and treated with HU308, CB2 agonist antibody, or a homozygous control (ABt269). FIGS. 18a to 18b Is This represents the mobilization of beta-arestin 2 induced by the indicated agent in RAW264.7 cells, as evaluated by the HTRF beta-arestin 2 mobilization detection kit. Fig. 19 Is This represents the positive hit rate (%) of antibodies generated by the generation model compared to antibodies developed from functional maturation. Fig. 20 represents the probability of a generated antibody having higher activity compared to an antibody with true activity and a seed antibody. Specific details for implementing the invention

[0064] The present disclosure is based, at least in part, on the discovery of a novel antibody that specifically binds to cannabinoid receptor type 2 (CB2) (which may be referred to as “anti-CB2 antibody” or “CB2 antibody” as a synonym) and its antigen-binding fragment. The CB2 antibody comprises at least an immunoglobulin single variable domain (ISVD) that specifically binds to CB2. The ISVD comprises a complementarity determining region 1 sequence (CDR1), a complementarity determining region 2 sequence (CDR2), and a complementarity determining region 3 sequence (CDR3). The ISVD may be the heavy chain variable domain (VHH) of a heavy chain antibody. The ISVD may be, for example, a camelid-derived, chimeric, or humanized ISVD. The ISVDs described herein may specifically bind to CB2 (e.g., human, synomolgus, murine, and / or rat CB2), whereas they may have low affinity or be undetectable for CB1 (e.g., human, synomolgus, murine, and / or rat CB1). The ISVDs described herein are selective agonists for CB2 and have no agonist activity or are undetectable for CB1. In some embodiments, the anti-CB2 antibody is an Fc fragment ( for example Includes human IgG1 Fc, effectorless IgG1 Fc, IgG2 Fc, or IgG4 Fc. Pharmaceutical compositions, kits, and therapeutic methods using these antibodies are also provided.

[0065] I. Definition

[0066] In this institution, the term 'antibody' is used in its broadest sense, and the desired antigen binding activity, in other wordsInsofar as they represent binding to CB2 (e.g., human CB2, cynomolgus CB2, rat CB2 and / or mouse CB2), they encompass various antibody structures including, but not limited to, monoclonal antibodies, quaternary antibodies (e.g., IgG antibodies), heavy-chain antibodies, and antibody fragments thereof. As used herein, the term 'quaternary antibody' is used to refer to an antibody or antigen-binding fragment having two heavy chains and two light chains. The term 'heavy-chain antibody,' also known as 'heavy-chain mono-antibody' or 'HCAb,' refers to a functional antibody containing two heavy chains but lacking the two light chains typically found in quaternary antibodies. Camelid animals (e.g., camels, llamas, or alpacas) are known to produce HCAbs.

[0067] As used herein, the term 'agonist' means that an antibody stimulates the biological function and / or signaling of a target receptor. In the context of CB2 agonists, CB2 agonists can activate endogenous cannabinoid signaling pathways through the activation of CB2. CB2 action includes, but is not limited to, (a) stimulating or activating CB2, and (b) enhancing, increasing, promoting, inducing, or prolonging the activity, function, or presence of CB2 signaling or downstream signaling products.

[0068] The term 'immunoglobulin monovariable domain' (also referred to as 'ISV' or 'ISVD') generally refers to, without interaction with other variable domains ( for exampleIt is used to refer to an immunoglobulin variable domain (which may be a heavy or light chain domain containing a VH, VHH, or VL domain) capable of forming a functional antigen-binding site (without the required VH / VL interaction between the VH and VL domains of a 4-chain monoclonal antibody). Examples of ISVDs include nanobodies (including VHH, humanized VHH, and / or camelized VH (e.g., camelized human VH)), shark IgNAR domains, single-domain antibodies (or dAbs) that are VH domains or derived from VH domains, and single-domain antibodies (or dAbs) that are VL domains or derived from VL domains. ISVDs based on and / or derived from a heavy chain variable domain (e.g., a VH or VHH domain) are generally preferred. In some embodiments, the ISVD is a nanobodi. The term 'nanobody' is generally as defined in WO 2008 / 020079 or WO 2009 / 138519, and thus, in certain embodiments, generally VHH, humanized VHH or camelized VH (e.g., camelized human VH), or generally sequence-optimized VHH (e.g., for example It represents chemical stability and / or solubility, optimized for maximum overlap with known human framework regions and maximum expression.

[0069] "Antibody fragments" comprise a portion of an antibody, preferably the antigen-binding region or variable region of the antibody. Examples of antibody fragments include VHH, single-domain antibodies, Fab, Fab', F(ab')2, and Fv fragments; diabodies; linear antibodies (Example 2 of U.S. Patent No. 5,641,870 and literature [Zapata et al[See Protein Eng. 8(10): 1057-1062

[1995] ]); short-chain antibody molecules; and multispecific antibodies formed from antibody fragments. The term 'constant domain' refers to a portion of an immunoglobulin molecule that has a more conserved amino acid sequence compared to other parts of the immunoglobulin, namely the variable domain containing the antigen-binding site. The constant domain is the C of the heavy chain. H 1, C H 2 and C H 3 domains (collectively referred to as C H ), and light chain CHL (or C L Includes the ) domain.

[0070] In this document, the terms 'Fc region' or 'crystallizable fragment region' are used to define the C-terminal region of an immunoglobulin heavy chain, including the natural sequence Fc region and the variant Fc region. Although the boundaries of the immunoglobulin heavy chain Fc region may vary, the human IgG heavy chain Fc region is typically defined as extending from the amino acid residue at the Cys226 position or Pro230 to its carboxyl terminus. The C-terminal lysine of the Fc region (residue 447 according to the EU numbering system) may be removed, for example, during the production or purification process of the antibody, or by recombinantly manipulating the nucleic acid encoding the heavy chain of the antibody. Thus, an intact antibody composition may comprise a population of antibodies from which all K447 residues have been removed, a population of antibodies from which no K447 residues have been removed, and a population of antibodies having a mixture of antibodies having K447 residues and antibodies not having K447 residues. Natural sequence Fc regions suitable for use in the antibodies described herein include human IgG1, IgG2 (IgG2A, IgG2B), IgG3, and IgG4.

[0071] The term 'monoclonal antibody' as used herein refers to an antibody obtained from a substantially homogeneous population of antibodies, and ,In other words, the individual antibodies constituting that group contain possible naturally occurring mutations and / or post-translational modifications that may be present in trace amounts. (for example, They are identical except for isomerization and amidation. Monoclonal antibodies are highly specific because they target a single antigenic site. In contrast to polyclonal antibody preparations, which generally contain different antibodies targeting different determinants (epitopes), each monoclonal antibody targets a single determinant on the antigen. In addition to this specificity, monoclonal antibodies are advantageous in that they are synthesized by hybridoma cultures without being contaminated by other immunoglobulins. The modifier 'monoclonal' indicates that the antibody was obtained from a population of substantially homologous antibodies and should not be interpreted as requiring the production of the antibody by any specific method. For example, the monoclonal antibody to be used in accordance with this application may be manufactured by various techniques, such as the hybridoma method (e.g., Literature [Kohler and Milstein., Nature , 256:495-97 (1975)]; literature[Hongo et al. , Hybridoma [Harlow , 14 (3): 253-260 (1995)], literature[Harlow et al. , Antibodies : A Laboratory Manual , (Cold Spring Harbor Laboratory Press, 2 nd ed. 1988)]; literature[Hammerling et al. , in: Monoclonal Antibodies and T-Cell Hybridomas 563-681 (Elsevier, NY, 1981)]), recombinant DNA method( for example , U.S. Patent No. 4,816,567 reference ), grip display technology( for example , literature[Clackson et al. , Nature , 352: 624-628 (1991)]; Literature[Marks et al. , J. Mol. Biol.222: 581-597 (1992)]; Literature[Sidhu et al. , J. Mol. Biol. 338(2): 299-310 (2004)]; Literature[Lee et al. , J. Mol. Biol. 340(5): 1073-1093 (2004)]; literature[Fellouse, Proc. Natl. Acad. Sci. USA 101(34): 12467-12472 (2004)]; and literature[Lee et al. , J. Immunol. Methods 284(1-2): 119-132 (2004)] reference )), and technology for producing human or human-like antibodies in animals having part or all of a human immunoglobulin locus, or a gene encoding a human immunoglobulin sequence (e.g., WO 1998 / 24893; WO 1996 / 34096; WO 1996 / 33735; WO 1991 / 10741; literature [Jakobovits et al. , Proc. Natl. Acad. Sci. USA 90: 2551 (1993)]; literature[Jakobovits et al. , Nature 362: 255-258 (1993)]; literature[Bruggemann et al. , Year in Immunol. 7:33 (1993)]; U.S. Patents No. 5,545,807; No. 5,545,806; No. 5,569,825; No. 5,625,126; No. 5,633,425; and No. 5,661,016; Literature [Marks et al. , Bio / Technology 10: 779-783 (1992)]; Literature[Lonberg et al. , Nature 368: 856-859 (1994)]; literature[Morrison, Nature 368: 812-813 (1994)]; literature[Fishwild et al. , Nature Biotechnol. 14: 845-851 (1996)]; literature[Neuberger, Nature Biotechnol. 14: 826 (1996)]; and literature[Lonberg and Huszar, Intern. Rev. Immunol.13: 65-93 (1995)] reference Includes ).

[0072] The term 'variable region' or 'variable domain' refers to a domain of the antibody heavy or light chain involved in binding an antibody to an antigen. The variable domains of the heavy and light chains of natural antibodies (VH and VL, respectively) generally have similar structures, and each domain includes four conserved framework regions (FR) and three hypervariable regions (HVR). for example , literature[Kindt et al. Kuby Immunology, 6th ed., W.H. Freeman and Co., page 91 (2007)] reference A single VH or VL domain may be sufficient to confer antigen-binding specificity. Furthermore, antibodies binding to a specific antigen can be isolated by screening a library of complementary VL or VH domains, respectively, using the VH or VL domain derived from the antibody binding to that antigen. for example , literature[Portolano et al. , J. Immunol. 150:880-887 (1993)] and literature [Clarkson et al. , Nature 352:624-628 (1991)] Refer to .

[0073] As used herein, the terms ‘hypervariable region’ or ‘HVR’ refer to each region in the antibody variable domain that is sequencely hypervariable and / or (‘complementarity determining region’ or ‘CDR’), forms a structurally defined loop and / or (‘hypervariable loop’), or contains antigen contact residues (‘antigen contact region’). Generally, a tetra-chain antibody and its antigen-binding antibody fragment contain six HVRs, namely three in VH (H1, H2, H3) and three in VL (L1, L2, L3). Generally, a heavy-chain antibody contains three HVRs (HVR1, HVR2, HVR3).

[0074] Multiple HVR delimitations are commonly used and are encompassed herein. In the exemplary HVR for a quaternary antibody and its antigen-binding antibody fragment, as described herein, (a) hypervariable loops (Chothia and Lesk) present at amino acid residues 26 to 32 (L1), 50 to 52 (L2), 91 to 96 (L3), 26 to 32 (H1), 53 to 55 (H2), and 96 to 101 (H3). J. Mol. Biol. 196:901-917 (1987)); (b) CDR(Kabat present in amino acid residues 24 to 34 (L1), 50 to 56 (L2), 89 to 97 (L3), 31 to 35b (H1), 50 to 65 (H2), and 95 to 102 (H3). et al. , Sequences of Proteins of Immunological Interest , 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991)); (c) Antigen contact sites (MacCallum) present on amino acid residues 27c to 36 (L1), 46 to 55 (L2), 89 to 96 (L3), 30 to 35b (H1), 47 to 58 (H2), and 93 to 101 (H3). et al. J. Mol. Biol. 262: 732-745 (1996)); and (d) a combination of (a), (b) and / or (c) comprising HVR amino acid residues 46 to 56 (L2), 47 to 56 (L2), 48 to 56 (L2), 49 to 56 (L2), 26 to 35 (H1), 26 to 35b (H1), 49 to 65 (H2), 93 to 102 (H3), and 94 to 102 (H3).

[0075] Unless otherwise specified, HVR residues and other residues within the variable domain ( for example , framework('FR') residue) is at this institution As mentioned earlier Kabat et al. Numbered according to

[0076] The amino acid residues of a single-domain antibody (e.g., VHH), as applied to the camelid VHH domain in the paper by Riechmann and Muyldermans (J. Immunol. Methods 2000 Jun. 23; 240 (1-2): 185-195), Kabat et al. V provided by ('Sequence of proteins of immunological interest', US Public Health Services, NIH Bethesda, Md., Publication No. 91) H They may be numbered according to general numbering for domains. According to this numbering, FR1 of VHH comprises amino acid residues at positions 1 to 30, CDR1 of VHH comprises amino acid residues at positions 31 to 35, FR2 of VHH comprises amino acids at positions 36 to 49, CDR2 of VHH comprises amino acid residues at positions 50 to 65, FR3 of VHH comprises amino acid residues at positions 66 to 94, CDR3 of VHH comprises amino acid residues at positions 95 to 102, and FR4 of VHH comprises amino acid residues at positions 103 to 113. In this regard, V H As is well known in the art regarding domains and VHH domains, it should be noted that the total number of amino acid residues within each CDR may vary and may not correspond to the total number of amino acid residues indicated by Kabat numbering (i.e., one or more positions according to Kabat numbering may not be occupied in the actual sequence, or the actual sequence may contain more amino acid residues than allowed by Kabat numbering).

[0077] 'Framework' or 'FR' residues are variable domain residues other than the HVR residues defined herein.

[0078] The term 'chimeric' antibody refers to an antibody in which a portion of the heavy chain and / or light chain is identical to or derived from a specific source or species, and the remainder of the heavy chain and / or light chain is identical to or derived from a different source or species.

[0079] A 'humanized' antibody is an antibody containing a minimal sequence derived from a non-human antibody. Generally, a humanized antibody is a human immunoglobulin (recipient antibody) in which residues derived from the recipient's hypervariable region are replaced with residues derived from the hypervariable region of a non-human species (donor antibody), such as camelid, mouse, rat, rabbit, or non-human primate, having the desired antibody specificity, affinity, and ability. In certain embodiments, the 'humanized' antibody is a non-human ( for example It refers to a chimeric antibody containing amino acid residues derived from CDR (Camelidae) and amino acid residues derived from human FR. In some cases, the framework domain (FR) residues of human immunoglobulin are replaced with corresponding non-human residues. Furthermore, the humanized antibody may contain residues not found in the recipient or donor antibody. Such modifications are made to further improve antibody performance. Generally, the humanized antibody contains substantially all of at least one, typically two, variable domains, wherein all or substantially all of the hypervariable loop corresponds to the hypervariable loop of non-human immunoglobulin, and all or substantially all of the FR is the FR of the human immunoglobulin sequence. For further details, see the literature [Jones et al. , Nature 321:522-525 (1986)]; Literature[Riechmann et al. See , Nature 332:323-329 (1988)]; and literature [Presta, Curr. Op. Struct. Biol. 2:593-596 (1992)].

[0080] An 'affinity-mature' antibody is an antibody having one or more modifications in one or more CDRs, wherein these modification(s) result in an improvement in the antibody's affinity for an antigen compared to a parent antibody that does not have such modification(s). In some embodiments, the affinity-mature antibody has an affinity for a target antigen at the nanomolar or even picomolar level. Affinity-mature antibodies are produced by procedures known in the art. For example, random mutagenesis of CDRs and / or framework residues is, for example, [Barbas et al. Proc Nat. Acad. Sci. USA 91:3809-3813 (1994)], literature[Schier et al. Gene 169:147-155 (1995)], literature[Yelton et al. J. Immunol. 155:1994-2004 (1995)], literature[Jackson et al. , J. Immunol. 154(7):3310-9 (1995)], and literature [Hawkins et al , J. Mol. Biol. It is described in 226:889-896 (1992).

[0081] "Amino acid sequence identity (%)" or "homology" regarding polypeptide and antibody sequences identified herein is defined as the percentage of amino acid residues in candidate sequences identical to amino acid residues in the polypeptide being compared, after aligning the sequences and considering any conservative substitutions as part of the sequence identity. Alignment for determining amino acid sequence identity (%) may be performed in various ways within the scope of the art, using publicly available computer software such as, for example, BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. A person skilled in the art may determine appropriate parameters for measuring alignment, including any algorithms necessary to achieve maximum alignment over the entire length of the sequences being compared. However, for the purposes of this invention, the amino acid sequence identity (%) value is generated using the sequence comparison computer program ALIGN-2. The ALIGN-2 sequence comparison computer program was written by Genentech, Inc., and its source code, along with user documentation, was submitted to the U.S. Copyright Office at 20559, Washington, D.C., and is registered under U.S. Copyright No. TXU510087. The ALIGN-2 program is publicly available through Genentech, Inc., South San Francisco, California. The ALIGN-2 program must be compiled for use on a UNIX operating system, preferably digital UNIX V4.0D. All sequence comparison parameters are set by the ALIGN-2 program and are not changed.

[0082] As used herein, the terms 'specific binding,' 'binds specifically,' or 'specific to' regarding an epitope on a specific polypeptide or a specific polypeptide target refer to, for example, K for a target D at least about 10 -4 M, alternatively at least about 10-5 M, alternatively at least about 10 -6 M, alternatively at least about 10 -7 M, alternatively at least about 10 -8 M, alternatively at least about 10 -9 M, alternatively at least about 10 -10 M, alternatively at least about 10 -11 M, alternatively at least about 10 -12 It may be represented by M or more molecules. In some embodiments, the term 'specific binding' refers to a molecule binding to a specific polypeptide or an epitope on a specific polypeptide without substantially binding to any other polypeptide or polypeptide epitope. K D This can be determined by methods known in the art, such as ELISA, surface plasmon resonance (SPR), fluorescence-activated cell sorting (FACS), or radioimmunoprecipitation (RIA). Specific binding can be measured by determining the binding of a molecule in comparison to the binding of a control molecule, for example, a molecule of similar structure that generally does not have binding activity. For example, specific binding can be determined through competition with a control molecule similar to the target, for example, an excess of unlabeled targets. In this case, if the binding of the labeled target to the probe is competitively inhibited by an excess of unlabeled targets, it is determined that specific binding exists.

[0083] As used herein, ‘treatment,’ ‘treating,’ or ‘treat’ is defined as an approach to obtain a beneficial or desired outcome, including clinical results. For the purposes of this application, beneficial or desired clinical results include alleviating one or more symptoms of a disease, reducing the severity of the disease, or stabilizing the disease ( for example , preventing or delaying the worsening of the disease), the spread of the disease ( for examplePreventing or delaying (and metastasis), preventing or delaying the recurrence of the disease, delaying or slowing the progression of the disease, improving the disease state, providing (partial or complete) remission of the disease, reducing the dosage of one or more other drugs required for the treatment of the disease, delaying the progression of the disease, increasing or improving the quality of life, increasing weight gain, and / or prolonging survival, one or more of these are included but not limited thereto. 'Treatment' also encompasses the reduction of the pathological consequences of the disease. The methods of this invention consider any one or more of these aspects of treatment. With respect to the treatment of pain, these methods mean pain reduction. Pain reduction includes reducing or improving the occurrence, sensation, perception, and / or effect of pain and / or its other symptoms, reducing the risk of pain occurring in the subject, inhibiting the onset of pain, alleviating pain, in other words Regressing pain or alleviating one or more symptoms of pain, relieving pain, reducing pain, limiting pain, reducing pain including duration and / or intensity of pain, alleviating pain, blocking pain, and inhibiting the propagation of pain are included but not limited to.

[0084] The terms 'effective dose' or 'therapeutic effective dose' for substances (e.g., CB2 antibodies, compounds, or compositions) used herein refer to the minimum concentration required to bring about measurable improvement or prevention of a specific disorder, or to achieve a specific desired outcome or results. In this application, the effective dose may vary depending on factors such as the disease state, the patient's age, sex, and weight, and the ability of the substance to induce a desired response in the individual. The effective dose is also defined as an amount in which the therapeutically beneficial effect outweighs any toxic or harmful effect of the treatment ( in other wordsIt is also a therapeutic index. An effective dose may be administered in one or more doses. As understood in a clinical context, an effective dose may or may not be achieved in combination with another drug, compound, or pharmaceutical composition. Therefore, an effective dose may be considered in the context of administering one or more therapeutic agents, and if a desirable result can or is achieved in combination with one or more other agents, a single agent may be considered to be administered as an effective dose.

[0085] The 'individual' or 'object' is a mammal. Mammals include domesticated animals ( for example , cattle, sheep, cats, dogs, and horses), primates ( for example , humans and non-human primates such as rhesus monkeys and cynomolgus monkeys), rabbits, and rodents ( for example Includes, but not limited to, mice and rats. In some embodiments, the entity or object is a human.

[0086] 'Antibody effector function' refers to biological activity attributed to the antibody's Fc region (natural sequence Fc region or amino acid sequence variant Fc region) and varies depending on the antibody isoform. Examples of antibody effector functions include C1q binding and complement-dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; cell surface receptors ( for exampleDownregulation of (, B cell receptor); and B cell activation are included. 'Reduced or minimized' antibody effector function means reduced by at least 50% (alternatively 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%) compared to wild-type or unmodified antibodies. The determination of antibody effector function can be readily determined and measured by a person skilled in the art. In preferred embodiments, antibody effector functions of complement binding, complement-dependent cytotoxicity, and antibody-dependent cytotoxicity are affected. In some embodiments, the effector function is a mutation within the invariant region from which glycosylation has been removed, for example It is eliminated via an 'effector-deficient mutation'. In one embodiment, the effector-deficient mutation is C H It is an N297A or DANA mutation (D265A+N297A) in region 2. Literature [Shields et al. , J. Biol. Chem. Refer to 276 (9): 6591-6604 (2001). Alternatively, additional mutations resulting in a reduction or elimination of effector function include K322A and L234A / L235A (LALA). Alternatively, mutations resulting in a reduction or elimination of effector function include F234A and L235A (FALA) based on the EU index. Alternatively, mutations resulting in a reduction or elimination of effector function include L234A, L235A, and P329G (LALAPG) based on the EU index. Alternatively, mutations resulting in a reduction or elimination of effector function include M252Y, S254T, and T256E (YTE) based on the EU index. Alternatively, effector function in non-glycosylated host cells ( for example , E. coli[ E. coli It can be reduced or eliminated through production techniques such as expression in ]), or expression in host cells resulting in altered glycosylation patterns that are ineffective or less effective in promoting effector function ( for example , literature[Shinkawa et al. , J. Biol. Chem. 278(5): 3466-3473 (2003)]).

[0087] As used herein, the terms 'pharmacologically acceptable' or 'pharmacologically suitable' mean a substance that is not biologically or otherwise undesirable, and for example The substance may be incorporated into a pharmaceutical composition administered to a patient without causing any significant undesirable biological effects or interacting in a harmful manner with any of the other components of the composition containing it. Pharmaceutically acceptable carriers or excipients preferably meet the required standards for toxicological and manufacturing tests and / or are included in the Inactive Ingredient Guide prepared by the U.S. Food and Drug Administration.

[0088] Half-maximum effective concentration (EC 50 EC₀ is a measure indicating the efficacy of a substance (e.g., an antibody) that induces a specific biological or biochemical function. It represents how much of a specific drug or other substance (e.g., an antibody) is required to achieve a response midway between the baseline and the maximum after a specific exposure time in a given biological process. This value is generally expressed as molar concentration. EC₀ 50 'IC' for antagonist drugs or other substances 50 It is comparable to EC 50 or IC 50 It can be measured by bioassays such as ligand binding inhibition by FACS analysis (competitive binding assay), cell-based cytokine release assay, or amplified luminescence proximity assay (AlphaLISA).

[0089] The 'isolated' nucleic acid molecules encoding the composition, antibody, or antigen-binding fragment thereof described herein are nucleic acid molecules identified and separated from at least one contaminating nucleic acid molecule typically associated with the environment in which they were produced. Preferably, the isolated nucleic acid is not associated with any components associated with the environment in which it was produced. The isolated nucleic acid molecules encoding the polypeptide and antibody described herein exist in a form different from that found in nature or in the environment. Thus, the isolated nucleic acid molecules are distinguished from the nucleic acids encoding the polypeptide and antibody described herein that naturally exist within the cell. The isolated nucleic acid includes nucleic acid molecules existing within the cell that typically contain the corresponding nucleic acid molecules, but which exist outside the chromosome or at a chromosomal location different from their natural chromosomal location.

[0090] The term 'regulatory sequence' refers to a DNA sequence required for the expression of a coding sequence operably linked in a specific host organism. Regulatory sequences suitable for prokaryotes include, for example, a promoter, optionally an operator sequence, and a ribosome binding site. Eukaryotic cells are known to utilize promoters, polyadenylation signals, and enhancers.

[0091] A nucleic acid is 'operably linked' when it is placed in a functional relationship with another nucleic acid sequence. For example, DNA for a presequence or secretion leader is operably linked to DNA for a polypeptide if it is expressed as a precursor protein participating in the secretion of the polypeptide; a promoter or enhancer is operably linked to a coding sequence if it influences the transcription of the sequence; and a ribosome binding site is operably linked to a coding sequence if it is positioned to facilitate translation. Generally, 'operably linked' means that the DNA sequences being linked are adjacent, and in the case of a secretion leader, they are adjacent and in the reading phase. However, enhancers do not need to be adjacent. Linkage is achieved through ligation at a convenient restriction site. Where such sites are absent, synthetic oligonucleotide adapters or linkers are used according to common practice.

[0092] As used herein, the term "vector" refers to a nucleic acid molecule capable of amplifying another nucleic acid linked to itself. This term includes vectors as self-replicating nucleic acid constructs as well as vectors incorporated into the genome of a host cell into which they are introduced. Certain vectors can direct the expression of nucleic acids operably linked to themselves. Such vectors are referred to herein as "expression vectors."

[0093] As used herein, the terms 'transfected,' 'transformed,' or 'transduced' refer to the process in which exogenous nucleic acids are delivered or introduced into a host cell. 'Transfected,' 'transformed,' or 'transduced' cells refer to cells that have been transfected, transformed, or introduced with exogenous nucleic acids. These cells include the original target cell and its progeny.

[0094] The terms 'host cell', 'host cell line', and 'host cell culture' are used interchangeably and refer to cells into which exogenous nucleic acids have been introduced, including the progeny of such cells. Host cells include 'transformers' and 'transformed cells', which include the original transformed cells and the progeny derived therefrom, regardless of the number of passages. The progeny may not be exactly identical to the parent cells in terms of nucleic acid content but may contain mutations. Mutant progeny having the same function or biological activity as that screened or selected from the original transformed cells are included herein.

[0095] The terms ‘pharmaceutical preparation’ or ‘pharmaceutical composition’ refer to a preparation that has a form that enables the biological activity of an active ingredient to be effective and does not contain additional ingredients that are unacceptably toxic to the subject to whom the preparation is to be administered. Such preparations are sterile. A ‘sterile’ preparation is sterile or free of all living microorganisms and their spores.

[0096] In this document, when a value or parameter is referred to as "approximately," it refers to the usual range of error for said value that is readily known to a person skilled in the art. In this document, when a value or parameter is referred to as "approximately," it refers to said value or parameter Self It includes (and describes) aspects regarding. For example, a description mentioning 'about X' includes a description of 'X'.

[0097] It is understood that the aspects and embodiments of the present application include aspects and embodiments of the 'comprising', 'consisting of', and 'essentially made of' forms.

[0098] The singular forms 'one', 'one', and 'above' as used in the present and appended claims include multiple references unless the context clearly indicates otherwise.

[0099] The term 'and / or' used herein in phrases such as 'A and / or B' is intended to include both A and B; A or B; A (alone); and B (alone). Likewise, the term 'and / or' used herein in phrases such as 'A, B and / or C' is intended to include each of the embodiments corresponding to A, B and C; A, B or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0100] II. Anti-CB2 Antibody

[0101] In various embodiments, an antibody that specifically binds to CB2 ( in other words , 'anti-CB2 antibody', 'CB2 antibody', 'antibody targeting CB2') are described herein. The CB2 antibody described herein may comprise three CDRs derived from an immunoglobulin monovariable domain (ISVD), comprise an ISVD, or be an ISVD. For example, in some embodiments, the CB2 antibody described herein is the one taught herein, for example Table 1 It includes a binding domain comprising three CDRs derived from the ISVD as listed in [ ]. In the embodiments below, the structure and functionality of the CB2 antibody are presented modularly as anti-CB2 ISVD. Describing only anti-CB2 ISVD is not intended to limit the scope, and this is a CB2 antibody comprising, in whole or in part, three CDRs derived from anti-CB2 ISVD or anti-CB2 ISVD in a non-limiting manner ( for example , Table 1 It encompasses the CDR presented in ).

[0102] The CB2 antibody described herein comprises an anti-CB2 ISVD. In some embodiments, the anti-CB2 ISVD specifically recognizes human CB2. In some embodiments, the anti-CB2 ISVD does not specifically recognize CB1, e.g., human CB1. In some embodiments, the anti-CB2 ISVD does not act as an agonist for CB1, e.g., human CB1. In some embodiments, the anti-CB2 ISVD specifically recognizes an epitope within human CB2. In some embodiments, the CB2 antibody comprises an ISVD that is the heavy chain variable domain (VHH) of a heavy chain antibody. In some embodiments, the CB2 antibody comprises an ISVD that is a camelid antibody. In some embodiments, the CB2 antibody comprises an ISVD that is in a chimeric form. In some embodiments, the CB2 antibody comprises a humanized ISVD. In some embodiments, the CB2 antibody comprises a partially humanized ISVD.

[0103] In some embodiments, the ISVD comprises CDR1, CDR2, and / or CDR3 of the ISVD comprising the amino acid sequence of SEQ ID NO. 55. In some embodiments, the ISVD comprises CDR1, CDR2, and / or CDR3 of the ISVD comprising the amino acid sequence of SEQ ID NO. 55, wherein CDR1, CDR2, and CDR3 are defined according to IMGT (for a description of IMGT, see literature [Lefranc et al. , Comparat. Immunol. 27:55-77, 2003] and literature[Ruiz et al. , Nucleic Acids Res.[See 29(1):207-9, 2001]). In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 1. In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR2 having the amino acid sequence of SEQ ID NO. 2. In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR3 having the amino acid sequence of SEQ ID NO. 3. In some embodiments, the ISVD comprises a CDR1, CDR2 and / or CDR3 of an ISVD having the amino acid sequence of SEQ ID NO. 55, where CDR1, CDR2 and CDR3 are defined according to Kabat. In some embodiments, the ISVD comprises a CDR1, CDR2 and / or CDR3 of an ISVD having the amino acid sequence of SEQ ID NO. 55, where CDR1, CDR2 and CDR3 are defined according to Chothia.

[0104] In some embodiments, a CB2 antibody comprising an ISVD that specifically binds to CB2 is provided herein, wherein the ISVD comprises a CDR1 comprising the amino acid sequence of SEQ ID NO. 1, a CDR1 comprising the amino acid sequence of SEQ ID NO. 2, and a CDR1 comprising the amino acid sequence of SEQ ID NO. 3, and CDR1, CDR2 and CDR3 are defined according to IMGT.

[0105] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 96 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 103 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0106] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 96 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 204 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0107] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 97 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 104 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0108] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 105 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0109] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 106 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0110] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 110 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 107 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0111] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 108 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0112] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 100 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 109 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0113] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 101 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 110 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0114] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 111 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0115] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 101 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 112 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0116] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 113 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0117] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 115 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0118] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 116 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0119] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 98 defined according to Kabat, a CDR2 having the amino acid sequence of SEQ ID NO. 117 defined according to Kabat, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Kabat.

[0120] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 118 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0121] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 119 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0122] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 120 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0123] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 121 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0124] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 122 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0125] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 123 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0126] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 124 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0127] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 125 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0128] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined by Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 126 defined by Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined by Chothia.

[0129] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 127 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0130] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 102 defined according to Chothia, a CDR2 having the amino acid sequence of SEQ ID NO. 128 defined according to Chothia, and a CDR3 having the amino acid sequence of SEQ ID NO. 129 defined according to Chothia.

[0131] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 345 defined according to IMGT.

[0132] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 346 defined according to IMGT.

[0133] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 347 defined according to IMGT.

[0134] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 348 defined according to IMGT.

[0135] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 349 defined according to IMGT.

[0136] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 350 defined according to IMGT.

[0137] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 351 defined according to IMGT.

[0138] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 352 defined according to IMGT.

[0139] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 353 defined according to IMGT.

[0140] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 354 defined according to IMGT.

[0141] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 355 defined according to IMGT.

[0142] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 356 defined according to IMGT.

[0143] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 357 defined according to IMGT.

[0144] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 358 defined according to IMGT.

[0145] In some embodiments, the CB2 antibody comprises an ISVD comprising a CDR1 having the amino acid sequence of SEQ ID NO. 248 defined according to IMGT, a CDR2 having the amino acid sequence of SEQ ID NO. 2 defined according to IMGT, and a CDR3 having the amino acid sequence of SEQ ID NO. 359 defined according to IMGT.

[0146] In some embodiments, the CB2 antibody is as follows Table 1 AB101, ABt107, ABt108, ABt109, ABt110, ABt111, ABt112, ABt113, ABt114, ABt115, ABt116, ABt117, ABt118, ABt119, ABt120, ABt121, ABt124, ABt247, ABt250, ABt251, ABt252, ABt253, ABt254, ABt105, ABt125, ABt126, ABt127, ABt128, ABt129, ABt130, ABt131, ABt132, ABt133, ABt134, ABt135, ABt136, ABt137, ABt138, ABt139, ABt140, as defined in It includes an ISVD comprising CDR1, CDR2, and CDR3 of antibodies named ABt143, ABt276, ABt281, ABt285, ABhit_0426, ABhit_0427, ABhit_0428, ABhit_0429, ABhit_0430, ABhit_0431, ABhit_0432, ABhit_0433, ABhit_0434, ABhit_0435, ABhit_0436, ABhit_0437, ABhit_0438, ABhit_0439, or ABhit_0440.

[0147] The CDR sequence of the exemplary CB2 antibody mentioned herein is as follows: Table 1 Provided in and defined according to IMGT, Kabat, or Chothia. In some embodiments of the teachings provided herein, antibodies may be referred to by a leading zero ( for example, ABt285=ABt0285). A person skilled in the art will readily understand that a CB2 antibody predicted using an algorithm that includes a CDR or variable domain sequence derived from the antibody described herein, but where said CDR or variable domain sequence is not one of the IMGT, Kabat or Chothia systems, is also within the scope of the present invention.

[0148]

[0149]

[0150]

[0151]

[0152]

[0153]

[0154]

[0155]

[0156]

[0157]

[0158]

[0159]

[0160]

[0161]

[0162]

[0163]

[0164]

[0165]

[0166] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD comprises a CDR1 comprising the amino acid sequence of SEQ ID NO. 1, or a variant thereof comprising up to three amino acid substitutions (e.g., any number of one, two, or three); a CDR2 comprising the amino acid sequence of SEQ ID NO. 2, or a variant thereof comprising up to three amino acid substitutions (e.g., any number of one, two, or three); and a CDR3 comprising the amino acid sequence of SEQ ID NO. 3, or a variant thereof comprising up to three amino acid substitutions (e.g., any number of one, two, or three), and CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the CB2 antibody comprises any one of the CDR1 sequences described herein ( for example It comprises an ISVD having one, two, or three amino acid substitutions at , SEQ ID NO. 1). In some embodiments, the ISVD comprises one, two, or three amino acid substitutions at positions 27, 28, 29, 31, 32, or 33 with respect to SEQ ID NO. 1, and the amino acid numbering follows IMGT. In some embodiments, the CB2 antibody comprises any one of the CDR3 sequences described herein ( for example, SEQ ID NO. 3) comprises an ISVD having one, two, or three amino acid substitutions. In some embodiments, the ISVD comprises A or P at position 15, F or W at position 52, A or S at position 54, A or G at position 55, S or W at position 103, and / or S or W at position 118, wherein the amino acid numbering follows IMGT. In some embodiments, the CB2 antibody comprises an ISVD, wherein the ISVD is zero, one, two, or three amino acid substitutions at one or more IMGT positions selected from positions 15, 52, 54, 55, 103, and 118, wherein the substitutions are A or P at position 15, F or W at position 52, A or S at position 54, A or G at position 55, S or W at position 103, and S or W at position 118 (where each is present); any one of the CDR3 sequences ( for example , at sequence no. 3), zero, one, two, or three substitutions where present, none of which are substitutions to C or M; and zero, one, two, or three amino acid substitutions where present, none of which are substitutions to C or M, at positions 27, 28, 29, 31, 32, or 33 based on sequence no. 1 as defined by IMGT.

[0167] In some embodiments, the CB2 antibody comprises a binding domain that specifically binds to CB2, wherein the binding domain comprises a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; and C6 is selected from Y, A, or F. In some embodiments, the CB2 antibody comprises the CDR1 of SEQ ID NO. 248. In some embodiments, the CB2 antibody comprises the CDR2 of SEQ ID NO. 2.

[0168] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD comprises a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; and C6 is selected from Y, A, or F. In some embodiments, the CB2 antibody comprises the CDR1 of SEQ ID NO. 248. In some embodiments, the CB2 antibody comprises the CDR2 of SEQ ID NO. 2.

[0169] In some embodiments, the CB2 antibody comprises a binding domain that specifically binds to CB2, wherein the binding domain comprises a CDR1 having the amino acid sequence of SEQ ID NO. 248; a CDR2 having the amino acid sequence of SEQ ID NO. 2; and a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; and C6 is selected from Y, A, or F, and CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the ISVD comprises a CDR1 having the amino acid sequence of SEQ ID NO. 248; a CDR2 having the amino acid sequence of SEQ ID NO. 2; It includes a CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 345 to 359.

[0170] In some embodiments, a CB2 antibody is provided comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to CB2, wherein the ISVD comprises a CDR1 having the amino acid sequence of SEQ ID NO. 248; a CDR2 having the amino acid sequence of SEQ ID NO. 2; and a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; and C6 is selected from Y, A, or F, and CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the ISVD comprises a CDR1 having the amino acid sequence of SEQ ID NO. 248; a CDR2 having the amino acid sequence of SEQ ID NO. 2; It includes a CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 345 to 359.

[0171] The CB2 antibody described herein comprises an ISVD comprising four framework regions (FR). The CB2 antibody described herein may comprise any suitable sequence for the framework regions. In some embodiments, the ISVD comprises any one framework region 1 (FR1) of SEQ ID NOs 4 to 15. In some embodiments, the ISVD comprises any one FR2 of SEQ ID NOs 16 to 30. In some embodiments, the ISVD comprises any one FR3 of SEQ ID NOs 31 to 48. In some embodiments, the ISVD comprises any one FR4 of SEQ ID NOs 49 to 54. In some embodiments, the ISVD comprises the FR3 of SEQ ID NO. 200. In some embodiments, FR1, FR2, FR3, and FR4 are defined according to IMGT.

[0172] In some embodiments, the ISVD comprises any one of SEQ ID NOs 4 to 15 framework region 1 (FR1), or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of about 1, 2, 3, 4, or 5). In some embodiments, the ISVD comprises any one of SEQ ID NOs 16 to 30 FR2, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of about 1, 2, 3, 4, or 5). In some embodiments, the ISVD comprises any one of SEQ ID NOs 31 to 48 FR3, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of about 1, 2, 3, 4, or 5). In some embodiments, ISVD comprises FR3 of SEQ ID NO. 200, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of about 1, 2, 3, 4, or 5). In some embodiments, ISVD comprises any one FR4 of SEQ ID NOs 49 to 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of about 1, 2, 3, 4, or 5). In some embodiments, FR1, FR2, FR3, and FR4 are defined according to IMGT.

[0173] In some embodiments, the ISVD comprises any one framework 1 region (FR1) of SEQ ID NOs 4 to 15, any one FR2 of SEQ ID NOs 16 to 30, any one FR3 of SEQ ID NOs 31 to 48, and any one FR4 of SEQ ID NOs 49 to 54. In some embodiments, the ISVD comprises any one framework 1 region (FR1) of SEQ ID NOs 4 to 15, any one FR2 of SEQ ID NOs 16 to 30, the FR3 of SEQ ID NO. 200, and any one FR4 of SEQ ID NOs 49 to 54. In some embodiments, the ISVD comprises any one FR1 of SEQ ID NOs 4 to 15, or a variant thereof comprising up to five amino acid substitutions (e.g., any number of one, two, three, four, or five); It comprises any one FR2 of SEQ ID NOs 16 to 30, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); any one FR3 of SEQ ID NOs 31 to 48 or 200, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and any one FR4 of SEQ ID NOs 49 to 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5).

[0174] In some embodiments, the CB2 antibody is as follows Table 2AB101, ABt107, ABt108, ABt109, ABt110, ABt111, ABt112, ABt113, ABt114, ABt115, ABt116, ABt117, ABt118, ABt119, ABt120, ABt121, ABt124, ABt247, ABt250, ABt251, ABt252, ABt253, ABt254, ABt105, ABt125, ABt126, ABt127, ABt128, ABt129, ABt130, ABt131, ABt132, ABt133, ABt134, ABt135, ABt136, ABt137, ABt138, ABt139, ABt140, as defined in It includes an ISVD comprising FR1, FR2, FR3, and FR4 of the antibody named ABt143, ABt276, ABt281, or ABt285.

[0175] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 4, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 16, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 31, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 49, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0176] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 4, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 16, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 200, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 49, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0177] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 14, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 29, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 43, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0178] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 15, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 23, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 45, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0179] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 15, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 23, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 45, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0180] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 15, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 23, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 46, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0181] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 15, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 29, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 47, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0182] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD is FR1 of SEQ ID NO. 15, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR2 of SEQ ID NO. 29, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); FR3 of SEQ ID NO. 45, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5); and FR4 of SEQ ID NO 54, or a variant thereof comprising up to 5 amino acid substitutions (e.g., any number of 1, 2, 3, 4, or 5), wherein FR1, FR2, FR3, and FR4 conform to IMGT.

[0183] In some embodiments, the CB2 antibody comprises an ISVD comprising FR1, FR2, FR3 and FR4, wherein the ISVD comprises one or more amino acid residues selected from the group consisting of V5, Q44, Y66, G74, R80, A83 and L87 in one or more of the framework regions, and the amino acid numbering follows IMGT.

[0184] In some embodiments, the framework region of the CB2 antibody is (1) a substitution at amino acid position 5 ( for example , substitution with V); (2) Substitution at amino acid position 44( for example , substitution with Q); (3) Substitution at amino acid position 66( for example , substitution with Y); (4) Substitution at amino acid position 74( for example , substitution with G); (5) Substitution at amino acid position 80( for example , substitution with R); (6) Substitution at amino acid position 83( for example , substitution with A); and (7) substitution at amino acid position 87 ( for exampleIt includes one or more amino acid substitutions selected from the group consisting of (substitution with L), and the amino acid positions are in accordance with IMGT numbering. In some embodiments, the CB2 antibody includes one or more amino acid substitutions in the framework region, said one or more amino acid substitutions include M5V, and the amino acid positions are in accordance with IMGT numbering. In some embodiments, the CB2 antibody includes one or more amino acid substitutions in the framework region, said one or more amino acid substitutions include M5V, R44Q, and I66Y, and the amino acid positions are in accordance with IMGT numbering. In some embodiments, the CB2 antibody includes one or more amino acid substitutions in the framework region, said one or more amino acid substitutions include M5V, R44Q, I66Y, and D65G, and the amino acid positions are in accordance with IMGT numbering. In some embodiments, the CB2 antibody includes one or more amino acid substitutions in the framework region, said one or more amino acid substitutions include M5V, R44Q, and D65G, and the amino acid positions are in accordance with IMGT numbering. In some embodiments, the CB2 antibody comprises one or more amino acid substitutions in the framework region, said one or more amino acid substitutions include M5V and I66Y, and the amino acid positions are in accordance with IMGT numbering. In some embodiments, the CB2 antibody comprises one or more amino acid substitutions in the framework region, said one or more amino acid substitutions include M5V, I66Y and D74G, and the amino acid positions are in accordance with IMGT numbering. In some embodiments, the CB2 antibody comprises one or more amino acid substitutions in the framework region, said one or more amino acid substitutions include N80R, S83A and A87L, and the amino acid positions are in accordance with IMGT numbering.In some embodiments, the CB2 antibody comprises CDR1 according to SEQ ID NO. 1, CDR2 according to SEQ ID NO. 2, and CDR3 according to SEQ ID NO. 3, and comprises a framework region comprising amino acid V at position 5, and the amino acid numbering follows IMGT. In some embodiments, the CB2 antibody comprises CDR1 according to SEQ ID NO. 1, CDR2 according to SEQ ID NO. 2, and CDR3 according to SEQ ID NO. 3, and comprises a framework region comprising V at position 5, Q at position 44, and Y at position 66, and the amino acid numbering follows IMGT. In some embodiments, the CB2 antibody comprises CDR1 according to SEQ ID NO. 1, CDR2 according to SEQ ID NO. 2, and CDR3 according to SEQ ID NO. 3, and comprises a framework region comprising V at position 5, Q at position 44, Y at position 66, and G at position 74, and the amino acid numbering follows IMGT. In some embodiments, the CB2 antibody comprises CDR1 according to SEQ ID NO. 1, CDR2 according to SEQ ID NO. 2, and CDR3 according to SEQ ID NO. 3, and comprises a framework region comprising V at position 5, Q at position 44, and G at position 74, and the amino acid numbering follows IMGT. In some embodiments, the CB2 antibody comprises CDR1 according to SEQ ID NO. 1, CDR2 according to SEQ ID NO. 2, and CDR3 according to SEQ ID NO. 3, and comprises a framework region comprising V at position 5 and Y at position 66, and the amino acid numbering follows IMGT. In some embodiments, the CB2 antibody comprises CDR1 according to SEQ ID NO. 1, CDR2 according to SEQ ID NO. 2, and CDR3 according to SEQ ID NO. 3, and comprises a framework region comprising V at position 5, Y at position 66, and G at position 74, and the amino acid numbering follows IMGT.In some embodiments, the CB2 antibody comprises CDR1 according to SEQ ID NO. 1, CDR2 according to SEQ ID NO. 2, and CDR3 according to SEQ ID NO. 3, and comprises a framework region comprising R at position 80, A at position 83, and L at position 87, and the amino acid numbering follows IMGT.

[0185]

[0186]

[0187]

[0188]

[0189]

[0190] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 55.

[0191] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 201.

[0192] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 56, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 56.

[0193] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 57.

[0194] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 58.

[0195] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 59, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 59.

[0196] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 60.

[0197] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 61.

[0198] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 62.

[0199] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 63.

[0200] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 64.

[0201] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 65, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 65.

[0202] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 66.

[0203] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 67, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 67.

[0204] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 68.

[0205] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 69, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 69.

[0206] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70.

[0207] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 330.

[0208] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 331.

[0209] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 332.

[0210] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 333.

[0211] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 334.

[0212] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 335.

[0213] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 336.

[0214] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 337.

[0215] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 338.

[0216] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 339.

[0217] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 340.

[0218] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 341.

[0219] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 342.

[0220] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 343.

[0221] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 70 or the amino acid sequence of SEQ ID NO. 344.

[0222] In some embodiments, the CB2 antibody is as follows Table 3AB101, ABt107, ABt108, ABt109, ABt110, ABt111, ABt112, ABt113, ABt114, ABt115, ABt116, ABt117, ABt118, ABt119, ABt120, ABt121, ABt124, ABt247, ABt250, ABt251, ABt252, ABt253, ABt254, ABt105, ABt125, ABt126, ABt127, ABt128, ABt129, ABt130, ABt131, ABt132, ABt133, ABt134, ABt135, ABt136, ABt137, ABt138, ABt139, ABt140, as defined in Includes the amino acid sequence of a CB2 antibody named ABt143, ABt276, ABt281, ABt285, ABhit_0426, ABhit_0427, ABhit_0428, ABhit_0429, ABhit_0430, ABhit_0431, ABhit_0432, ABhit_0433, ABhit_0434, ABhit_0435, ABhit_0436, ABhit_0437, ABhit_0438, ABhit_0439, or ABhit_0440.

[0223]

[0224]

[0225]

[0226]

[0227]

[0228]

[0229]

[0230] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 71.

[0231] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 72.

[0232] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 73.

[0233] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 74, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 74.

[0234] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 75, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 75.

[0235] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 76.

[0236] In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises the amino acid sequence of SEQ ID NO. 77, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 77.

[0237] In some embodiments, the CB2 antibody described herein comprises an Fc region. In some embodiments, the CB2 antibody comprises the Fc region of IgG1, IgG2, IgG3, or IgG4, or a variant thereof. In some embodiments, the CB2 antibody comprises the Fc region of IgG1. In some embodiments, the CB2 antibody comprises the Fc region of IgG4. In some embodiments, the CB2 antibody comprises an Fc variant exhibiting reduced effector function. In some embodiments, reduced effector function reduces ADCC activity. In some embodiments, the antibody comprises an IgG1 Fc region exhibiting reduced effector function. In some embodiments, the antibody comprises a variant IgG1 Fc region containing one or more variant amino acids. In some embodiments, one or more variant amino acids of the variant IgG Fc region described herein may be additions, substitutions, or deletions. Such variant IgG1 Fc regions have a conserved Fc sequence, for example It is based on the preserved human Fc sequence.

[0238] In some embodiments, the antibody comprises an IgG1 Fc region exhibiting reduced effector function, the Fc region comprises amino acid substitutions L234A and L235A, and the numbering follows the EU index of Kabat. In some embodiments, the antibody comprises an IgG4 Fc region exhibiting reduced effector function. In some embodiments, the antibody comprises an IgG4 Fc region exhibiting reduced effector function, the Fc region comprises amino acid substitution F234A / L235A (FALA), and the numbering follows the EU index of Kabat. In some embodiments, the variant IgG1 Fc region comprises amino acid substitutions selected from the group consisting of (1) LALA, (2) YTE, and (3) LALAPG. In some embodiments, the variant IgG1 Fc region comprises a LALAPG variant. In some embodiments, the variant IgG1 Fc region comprises a LALAPG variant and a YTE variant. In some embodiments, the variant IgG1 Fc region includes LALA variants and YTE variants. The LALAPG variants are known as L234A, L235A, and P329G, and their residues are numbered according to the EU index. The LALA variants are known as L234A and L235A, and their residues are numbered according to the EU index. The YTE variants are known as M252Y, S254T, and T256E, and their residues are numbered according to the EU index.

[0239] In some embodiments, the CB2 antibody comprises an Fc region, the Fc region comprises the amino acid sequence of SEQ ID NO. 130. In some embodiments, the CB2 antibody comprises an Fc region, the Fc region comprises the amino acid sequence of SEQ ID NO. 363. In some embodiments, the CB2 antibody comprises an Fc region, the Fc region comprises the amino acid sequence of SEQ ID NO. 364. In some embodiments, the CB2 antibody comprises an Fc region, the Fc region comprises the amino acid sequence of SEQ ID NO. 365. In some embodiments, the CB2 antibody further comprises a hinge region. In some embodiments, the hinge region comprises the amino acid sequence of SEQ ID NO. 131. In some embodiments, the ISVD is fused to the Fc region via a peptide linker. In some embodiments, the peptide linker comprises the amino acid sequence GGGGSGGGGSGGGGS (SEQ ID NO. 132). In some embodiments, the CB2 antibody comprises a polypeptide chain including an ISVD, a peptide linker, a hinge region, and an Fc region extending from the N-terminus to the C-terminus. In some embodiments, the CB2 antibody does not include an Fc region. An exemplary hinge is provided as AESKYGPPCPPCP (SEQ ID NO. 131).

[0240] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 78, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 78.

[0241] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 79, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 79.

[0242] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 80, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 80.

[0243] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 81, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 81.

[0244] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 82, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 82.

[0245] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 83, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 83.

[0246] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 84, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 84.

[0247] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 85, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 85.

[0248] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 86, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 86.

[0249] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 87, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 87.

[0250] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 88, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 88.

[0251] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 89, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 89.

[0252] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 90, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 90.

[0253] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 91, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 91.

[0254] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 92, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 92.

[0255] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 93, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 93.

[0256] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 94, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 94.

[0257] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 95, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 95.

[0258] In some embodiments, the CB2 antibody described herein comprises a CB2 antibody that specifically binds to CB2 competitively with any one of the other CB2 antibodies described herein. In some embodiments, competitive binding may be determined using an ELISA assay. In some embodiments, the CB2 antibody specifically binds to CB2 competitively with a CB2 antibody comprising CDR1 of SEQ ID NO. 1, CDR2 of SEQ ID NO. 2, and CDR3 of SEQ ID NO. 3. In some embodiments, the CB2 antibody specifically binds to CB2 competitively with a CB2 antibody comprising any one of the amino acid sequences of SEQ ID NOs. 55 to 95. In some embodiments, the CB2 antibody Table 1 It specifically binds to CB2 competitively with CB2 antibodies according to the CB2 antibodies. In some embodiments, the binding K between the competing CB2 antibody and CB2. D is about 10-7 M to about 10 -12 M (e.g., about 10 -8 M to about 10 -12 M, or about 10 -9 M to about 10 -11 M) is. In some embodiments, the CB2 antibody cross-reacts with CB2 from non-human mammals. In some embodiments, the competing CB2 antibody is a camelid, chimera, human, partially humanized, or fully humanized antibody.

[0259] In some embodiments, the CB2 antibody described herein comprises a CB2 antibody that has undergone functional maturation. In some embodiments, the CB2 antibody comprises a CDR1 having the amino acid formula G-X1-X2-X3-SI-X4-X5, wherein X1 is selected from D, S, or Q; X2 is selected from G, H, I, or K; X3 is selected from D, E, F, G, H, I, K, N, Q, R, S, T, V, W, or Y; X4 is selected from G, K, M, R, or Y; and X5 is selected from A or G. In some embodiments, the antibody comprises CDR2 and CDR3 as defined herein. In some embodiments, CDR2 is SEQ ID NO. 2 according to IMGT. In some embodiments, the CB2 antibody comprises a binding domain that specifically binds to CB2, wherein the binding domain comprises CDR1 having the amino acid formula G-X1-X2-X3-SI-X4-X5, said formula where X1 is selected from D, S, or Q; X2 is selected from G, H, I, or K; X3 is selected from D, E, F, G, H, I, K, N, Q, R, S, T, V, W, or Y; X4 is selected from G, K, M, R, or Y; and X5 is selected from A or G. In some embodiments, the antibody comprises CDR2 and CDR3 as defined herein. In some embodiments, CDR2 is SEQ ID NO. 2 according to IMGT.

[0260] In some embodiments, the CB2 antibody comprises a CDR3 having the amino acid formula C1-C2-C3-IK-C4-C5-C6-C7-C8-C9-C10-I-C11-C12-C13, wherein C1 is selected from F, H, I, K, L, R, S, T, V, or Y; C2 is selected from A, E, F, G, I, L, Q, R, or V; C3 is selected from A, F, G, I, K, L, N, or V; C4 is selected from D, F, N, T, or Y; C5 is selected from A, G, or R; C6 is selected from R, S, or W; C7 is selected from D, G, K, or R; C8 is selected from D, I, L, Q, R, S, or T; C9 is selected from F, L, or T; and C10 is selected from D, E, H, or Q; C11 is selected from K or N; C12 is selected from A, F, G, H, I, K, L, N, P, R, S, V, T or W; and C13 is selected from A, D, F, G, H, T or Y. In some embodiments, the antibody comprises CDR1 and CDR2 as defined herein. In some embodiments, CDR2 is SEQ ID NO. 2 according to IMGT.In some embodiments, the CB2 antibody comprises a binding domain that specifically binds to CB2, wherein the binding domain comprises a CDR3 having the amino acid formula C1-C2-C3-IK-C4-C5-C6-C7-C8-C9-C10-I-C11-C12-C13, wherein C1 is selected from F, H, I, K, L, R, S, T, V, or Y; C2 is selected from A, E, F, G, I, L, Q, R, or V; C3 is selected from A, F, G, I, K, L, N, or V; C4 is selected from D, F, N, T, or Y; C5 is selected from A, G, or R; C6 is selected from R, S, or W; C7 is selected from D, G, K, or R; C8 is selected from D, I, L, Q, R, S, or T; and C9 is selected from F, L, or T; C10 is selected from D, E, H, or Q; C11 is selected from K or N; C12 is selected from A, F, G, H, I, K, L, N, P, R, S, V, T, or W; and C13 is selected from A, D, F, G, H, T, or Y. In some embodiments, the antibody comprises CDR1 and CDR2 as defined herein. In some embodiments, CDR2 is SEQ ID NO. 2 according to IMGT.

[0261] In some embodiments, the CB2 antibody comprises a CDR1 having the amino acid formula G-X1-X2-X3-SI-X4-X5, wherein X1 is selected from D, S, or Q; X2 is selected from G, H, I, or K; X3 is selected from D, E, F, G, H, I, K, N, Q, R, S, T, V, W, or Y; X4 is selected from G, K, M, R, or Y; and X5 is selected from A or G. In some embodiments, the CB2 antibody comprises a CDR3 having the amino acid formula C1-C2-C3-IK-C4-C5-C6-C7-C8-C9-C10-I-C11-C12-C13, wherein C1 is selected from F, H, I, K, L, R, S, T, V, or Y; C2 is selected from A, E, F, G, I, L, Q, R, or V; C3 is selected from A, F, G, I, K, L, N, or V; C4 is selected from D, F, N, T, or Y; C5 is selected from A, G, or R; C6 is selected from R, S, or W; C7 is selected from D, G, K, or R; C8 is selected from D, I, L, Q, R, S, or T; C9 is selected from F, L, or T; C10 is selected from D, E, H, or Q; C11 is selected from K or N; C12 is selected from A, F, G, H, I, K, L, N, P, R, S, V, T, or W; and C13 is selected from A, D, F, G, H, T, or Y. In some embodiments, CDR2 is the CDR defined herein. In some embodiments, CDR2 is sequence number 2 according to IMGT.In some embodiments, the CB2 antibody comprises a binding domain that specifically binds to CB2, wherein the binding domain comprises a CDR1 having the amino acid formula G-X1-X2-X3-SI-X4-X5, said formula where X1 is selected from D, S, or Q; X2 is selected from G, H, I, or K; X3 is selected from D, E, F, G, H, I, K, N, Q, R, S, T, V, W, or Y; X4 is selected from G, K, M, R, or Y; and X5 is selected from A or G. In some embodiments, the CB2 antibody comprises a CDR3 having the amino acid formula C1-C2-C3-IK-C4-C5-C6-C7-C8-C9-C10-I-C11-C12-C13, said formula where C1 is selected from F, H, I, K, L, R, S, T, V, or Y; C2 is selected from A, E, F, G, I, L, Q, R, or V; C3 is selected from A, F, G, I, K, L, N, or V; C4 is selected from D, F, N, T, or Y; C5 is selected from A, G, or R; C6 is selected from R, S, or W; C7 is selected from D, G, K, or R; C8 is selected from D, I, L, Q, R, S, or T; C9 is selected from F, L, or T; C10 is selected from D, E, H, or Q; C11 is selected from K or N; C12 is selected from A, F, G, H, I, K, L, N, P, R, S, V, T, or W; and C13 is selected from A, D, F, G, H, T, or Y. In some embodiments, CDR2 is the CDR defined herein. In some embodiments, CDR2 is sequence number 2 according to IMGT.

[0262] In some embodiments, a CB2 antibody is provided comprising CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs 248 to 288, CDR2 having an amino acid sequence of SEQ ID NO. 2, and CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs 289 to 329, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, CDR1, CDR3, and CDR3 are Table 4 It is a set provided according to

[0263] In some embodiments, the CB2 antibody is as follows Table 4 ABhit_0231, ABhit_0232, ABhit_0233, ABhit_0234, ABhit_0235, ABhit_0236, ABhit_0238, ABhit_0239, ABhit_0241, ABhit_0242, ABhit_0243, ABhit_0244, ABhit_0245, as defined in ABhit_0246, ABhit_0247, ABhit_0248, ABhit_0249, ABhit_0250, ABhit_0251, ABhit_0252, ABhit_0253, ABhit_0254, ABhit_0255, ABhit_0256, ABhit_0258, ABhit_0259, It comprises the amino acid sequence of one of the CB2 antibodies named ABhit_0260, ABhit_0261, ABhit_0262, ABhit_0263, ABhit_0264, ABhit_0266, ABhit_0267, ABhit_0268, ABhit_0269, ABhit_0270, ABhit_0271, ABhit_0272, ABhit_0320, ABhit_0321, or ABhit_0322. In some embodiments, the following Table 4ABhit_0231, ABhit_0232, ABhit_0233, ABhit_0234, ABhit_0235, ABhit_0236, ABhit_0238, ABhit_0239, ABhit_0241, ABhit_0242, ABhit_0243, ABhit_0244, ABhit_0245, as defined in ABhit_0246, ABhit_0247, ABhit_0248, ABhit_0249, ABhit_0250, ABhit_0251, ABhit_0252, ABhit_0253, ABhit_0254, ABhit_0255, ABhit_0256, ABhit_0258, ABhit_0259, A CB2 antibody comprising a CDR sequence of one of ABhit_0260, ABhit_0261, ABhit_0262, ABhit_0263, ABhit_0264, ABhit_0266, ABhit_0267, ABhit_0268, ABhit_0269, ABhit_0270, ABhit_0271, ABhit_0272, ABhit_0320, ABhit_0321, or ABhit_0322 is provided.

[0264]

[0265]

[0266]

[0267]

[0268]

[0269]

[0270]

[0271] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 205, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 205.

[0272] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 206, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 206.

[0273] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 207, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 207.

[0274] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 208, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 208.

[0275] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 209, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 209.

[0276] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 210, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 210.

[0277] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 211, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 211.

[0278] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 212, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 212.

[0279] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 214, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 214.

[0280] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 215, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 215.

[0281] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 216, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 216.

[0282] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 217, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 217.

[0283] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 218, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 218.

[0284] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 219, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 219.

[0285] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 220, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 220.

[0286] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 221, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 221.

[0287] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 222, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 222.

[0288] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 223, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 223.

[0289] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 224, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 224.

[0290] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 225, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 225.

[0291] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 226, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 226.

[0292] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 227, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 227.

[0293] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 228, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 228.

[0294] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 229, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 229.

[0295] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 231, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 231.

[0296] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 232, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 232.

[0297] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 233, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 233.

[0298] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 234, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 234.

[0299] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 235, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 235.

[0300] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 236, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 236.

[0301] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 237, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 237.

[0302] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 238, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 238.

[0303] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 239, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 239.

[0304] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 240, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 240.

[0305] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 241, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 241.

[0306] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 242, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 242.

[0307] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 243, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 243.

[0308] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 244, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 244.

[0309] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 245, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 245.

[0310] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 246, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 246.

[0311] In some embodiments, the CB2 antibody comprises the amino acid sequence of SEQ ID NO. 247, or a variant thereof having at least 85%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 247.

[0312] CB2

[0313] Cannabinoid receptor type 2 (CB2) is a G protein-coupled receptor belonging to the cannabinoid receptor family. In humans, this CNR2 It is encoded by a gene. This is closely related to cannabinoid receptor type 1 (CB1). CB1 action is primarily responsible for the psychoactive effects of cannabis, particularly tetrahydrocannabinol (THC). Due to the psychoactive effects of CB1 action, an antibody that specifically binds to CB2 but does not exhibit specific binding to CB1 is preferred. Accordingly, in one embodiment, the CB2 antibody provided herein is specific to CB2, does not exhibit significant affinity for CB1, and does not bind specifically to it. The CB2 receptor comprises seven transmembrane domains and includes a glycosylated N-terminus and an intracellular C-terminus.

[0314] The terms 'cannabinoid receptor type 2', 'CB2', 'CB2 antigen', and 'CB2 epitope' are used interchangeably and include variants, isoforms, species homologs of human CB2, and analogs having at least one common epitope with CB2. In some embodiments, CB2 is human CB2, and the CB2 antibody specifically binds to human CB2. CB2 gene and polypeptide sequence ( for example , human CNR2 Gene and polypeptide sequences are known in the art. An exemplary human CB2 sequence is presented in GenBank accession number P34972. In some embodiments, the human CB2 may have an amino acid sequence that is at least about 90%, 95%, 96%, 97%, 98%, or 99% identical to the human CB2 of GenBank accession number P34972. In some embodiments, the human CB2 sequence has about 10 or fewer amino acid differences from the human CB2 of GenBank accession number P34972. In some embodiments, the human CB2 may exhibit five or fewer, four or fewer, three or fewer, two or fewer, or one or fewer amino acid differences from the human CB2 of GenBank accession number P34972. In some embodiments, the human CB2 sequence may differ from the human CB2 of GenBank accession number P34972 by having, for example, a conserved mutation or a mutation in a non-conserved region, and such CB2 has substantially the same biological function as the human CB2 of GenBank accession number P34972.

[0315] In some embodiments, the CB2 antibody described herein specifically recognizes a CB2 polypeptide having at least about 90% amino acid sequence identity with human CB2 of GenBank accession number P34972. In some embodiments, the CB2 antibody recognizes CB2 of non-human species (e.g., rat, murine, and / or synomolgus CB2), or other proteins structurally related to human CB2 ( for example It can cross-react with human CB2 homologs. In some embodiments, the CB2 antibody does not specifically bind to CB1.

[0316] biological activity

[0317] The CB2 antibody described herein is ISVD ( for example Including , VHH or camelid), and (a) that ISVD is an agonist for CB2; (b) that ISVD does not act as an agonist for CB1; (c) that ISVD is at a half-maximum effective concentration (EC) of less than 55 nM in HEK293 cells overexpressing human CB2 50 (d) reduction of forskolin-inducible cyclic adenosine monophosphate (cAMP) levels by ); (d) reduction of EC10 nM or less by ISVD in HEK293 cells overexpressing mouse CB2 50 (e) reducing forskolin-inducible cyclic adenosine monophosphate (cAMP) levels; (f) ISVD not specifically binding to CB1; and (f) CB2 antibody at EC levels of less than 850 nM in HEK293 cells overexpressing human CB2 50 It has one or more characteristics of combining.

[0318] The CB2 antibody described herein binds to CB2 with high affinity and specificity. In some embodiments, the CB2 antibody is about 0.001 nM to about 10 nM ( for example K of , 10 nM or less, 1 nM or less, 0.1 nM or less, 0.01 nM or less, or 0.001 nM or less). D It binds to human CB2, and the values ​​include any value or range between the exemplified values. In some embodiments, the CB2 antibody is about 0.001 nM to about 10 nM ( for example , 10 nM or less, 1 nM or less, 0.1 nM or less, 0.01 nM or less) or K of 0.001 nM or less DIt binds to synomolgus CB2, and the values ​​include any value or range between the exemplified values. In some embodiments, the CB2 antibody is about 0.001 nM to about 10 nM ( for example K of , 10 nM or less, 1 nM or less, 0.1 nM or less, 0.01 nM or less, or 0.001 nM or less). D It binds to rat CB2, and the values ​​include any value or range between the exemplified values. In some embodiments, the CB2 antibody is about 0.001 nM to about 10 nM ( for example K of , 10 nM or less, 1 nM or less, 0.1 nM or less, 0.01 nM or less, or 0.001 nM or less). D It binds to murine CB2, and the value includes any value or range between the exemplified values.

[0319] In some embodiments, the CB2 antibody is a selective CB2 agonist. In some embodiments, the CB2 antibody is a selective CB2 agonist if it exhibits detectable CB2 agonist activity and does not exhibit detectable CB1 agonist activity. In some embodiments, the CB2 antibody is a selective CB2 agonist if it exhibits detectable CB2 agonist activity and does not exhibit detectable CB1 agonist activity when evaluated by a decrease in forskolin-induced cyclic adenosine monophosphate (cAMP) production in HEK293 cells that overexpress CB2 (for evaluating CB2 action) or overexpress CB1 (for evaluating CB1 action). In some embodiments, if the decrease in cAMP compared to the control is not statistically significant (e.g., p is not less than 0.05), the agonist activity is not detectable. In some embodiments, if the measured degree of activity differs by an order of magnitude between CB2 activity and CB1 activity, CB1 agonist activity is not detectable. In some embodiments, CB2 antibody activity (e.g., activity against CB1 or CB2) is evaluated at a concentration of about 1 μM.

[0320] In some embodiments, the CB2 antibody selectively binds to CB2 and does not selectively bind to CB1. In some embodiments, the CB2 antibody is greater than about 1,000 nM, e.g., greater than about 1,000 nM, greater than 2,000 nM, greater than 3,000 nM, greater than 4,000 nM, greater than 5,000 nM, or greater than K D When binding to CB1, it does not selectively bind to CB1, and the values ​​include values ​​and ranges between the exemplified values. In some embodiments, CB1 is human, rat, murine, or synomolgus CB1.

[0321] K of the CB2 antibody provided herein against human CB2, rat CB2, murine CB2 and / or synomolgus CB2 D and koff any method known in the industry, for example It may be determined by methods including, but not limited to, ELISA, fluorescence-activated cell sorting (FACS) analysis, radioimmunoprecipitation (RIA), and surface plasmon resonance (SPR). In some embodiments, K of the CB2 antibody provided herein against human CB2, murine CB2, rat CB2, and / or synomolgus CB2. D and / or k off is determined via SPR. In some embodiments, K of the CB2 antibody provided herein D and / or k off is determined through surface plasmon resonance (SPR).

[0322] The characteristics of the CB2 antibody described herein are based on well-known methods, for example It can be evaluated using the method used in the following examples. In some embodiments, the CB2 antibody binds to CB1, e.g., human CB1 ( for example , does not specifically bind)

[0323] The biological activity of the CB2 antibody described herein is determined by its half-maximum effective concentration (EC 50 It can be determined by measuring ), which is a measure indicating the effectiveness of the antibody binding to the target. For example, EC 50 It can be used to indicate the effective concentration of CB2 antibodies required to bind to 50% of CB2 on the cell surface. EC 50 Eun also In vivo Indicates the plasma concentration required to obtain 50% of the maximum effect. EC 50 It can be measured by bioassays known in the art, such as FACS binding assay, ligand binding inhibition by FACS assay (competitive binding assay), cell-based cytokine release assay, or amplified luminescence proximity assay (AlphaLISA).

[0324] For example, the agonist activity of a CB2 antibody can be studied by measuring the reduction in forskolin-induced cAMP levels in HEK293 cells overexpressing human CB2 (except CB1 when evaluating CB2 activity) or human CB1 (except CB2 when evaluating CB1 activity), as described in Example 2 below. The results of this study can demonstrate the ability of the CB2 antibody to act as an agonist for CB2 but not for CB1, and can compare the CB2 antibody with other CB1 or CB2 agonists.

[0325] agonist EC of binding to CB2 by CB2 antibody 50 It can be studied by evaluating the binding of CB2 antibodies to CB2-expressing HEK293 stable cells. Similarly, the agonist EC of binding to CB1 by CB2 antibodies 50 This can be studied by evaluating the binding of CB2 antibodies to CB1-expressing HEK293 stable cells. As discussed in Example 2 below, after incubating CB1-expressing or CB2-expressing HEK293 stable cells with CB2 antibodies in the presence of forskolin, the cells can be lysed by adding a tracer and detection reagent. After incubation, the quantification of cAMP levels and EC2 levels via FRET are performed. 50 Calculation of is possible. In some embodiments, detectable agonist activity of CB1 (e.g., human, mouse, synomolgus and / or rat CB1) by the CB2 antibody described herein is not exhibited.

[0326] (II) Fusion Works

[0327] The present application covers the CB2 antibody or antigen-binding fragment described herein ( for exampleThe present invention further provides an anti-CB2 fusion construct comprising any one of , ISVD, or VHH, and a second polypeptide, such as the second antibody or its antigen-binding fragment, or the Fc fragment of an immunoglobulin. In some embodiments, the anti-CB2 construct comprises two or more polypeptides other than the CB2 antibody. These additional polypeptide(s) may or may not alter or otherwise affect the biological properties of the CB2 antibody, and may or may not impart additional functionality to the CB2 antibody. In some embodiments, the second polypeptide imparts one or more desired properties or functionalities to the CB2 antibody.

[0328] In some embodiments, the anti-CB2 construct comprises a second antibody or its antigen-binding fragment (e.g., ISVD, scFv, Fab, full-length antibody, etc.) that specifically recognizes the second epitope. In some embodiments, the second epitope originates from CB2. In some embodiments, the second epitope does not originate from CB2. In some embodiments, the second antibody specifically recognizes the same epitope on CB2 that is recognized by the CB2 antibody described herein. In some embodiments, the second antibody specifically recognizes an epitope on CB2 that is different from that recognized by the CB2 antibody described herein.

[0329] In some embodiments, an anti-CB2 construct is provided comprising a plurality of (e.g., 2, 3, 4 or more) of the CB2 antibodies described herein. In some embodiments, these plurality of CB2 antibodies are fused together through a linker (e.g., a peptide linker). The plurality of CB2 antibodies may be identical or different.

[0330] In some embodiments, the CB2 antibody comprises a second polypeptide that, compared to the CB2 antibody alone, improves and / or improves the half-life, solubility, and / or uptake of the CB2 antibody, reduces and / or immunogenicity or toxicity, eliminates or attenuates / or imparts other advantageous properties to the CB2 antibody, or reduces other undesirable properties of the CB2 antibody. Some non-limiting examples of such polypeptides include serum proteins, such as human serum albumin (HSA, For example Enter , see WO 2000 / 027435) or haptenic molecules ( for example , hapten recognized by circulating antibodies, For example Enter , WO 98 / 22141 reference ) is included. Fragments of immunoglobulin (e.g., V H It has been shown that antibody half-life can be increased by linking the domain) to serum albumin or its fragment ( For example Enter (See WO 00 / 27435 and WO 01 / 077137). Accordingly, in some embodiments, the CB2 antibody comprises a CB2 antibody fused to serum albumin or a fragment thereof via an optionally suitable linker (e.g., a peptide linker). In some embodiments, the serum albumin comprises at least domain III (PCT / EP2007 / 002817 reference ).

[0331] CB2 heavy chain monoclonal antibody (HCAb)

[0332] In some embodiments, the CB2 antibody is a heavy chain monoclonal antibody (HCAb) comprising the CB2 ISVD described herein. In some embodiments, the CB2 ISVD comprises one or more C H 2 and / or C H 3 domains, for example Fc is fused to the fragment. In some embodiments, C H 2 and / or C HThe 3 domain is derived from human immunoglobulin. In some embodiments, the CB2 antibody is C via a peptide linker. H 2 and / or C H It is fused into 3 domains. C H 2 and / or C H 3 domains are In vivo It can increase the half-life of CB2 antibodies.

[0333] Accordingly, in some embodiments, an isolated CB2 HCAb is provided comprising the CB2 ISVD described herein (meaning an ISVD that specifically binds to CB2) fused to an Fc fragment of an immunoglobulin, such as IgA, IgD, IgE, IgG, or IgM. In some embodiments, the CB2 HCAb comprises an Fc fragment of IgG, such as IgG1, IgG2, IgG3, or IgG4. In some embodiments, the Fc fragment is a human Fc, such as human IgG1 (hIgG1) Fc, hIgG2 Fc, or hIgG4 Fc. In some embodiments, the Fc fragment is in an effector-deficient state, in which antibody effector functions such as ADCC, CDC, and / or ADCP (antibody-dependent phagocytosis) are reduced, minimized, or eliminated. In some embodiments, the effector-deficient Fc is C HRegion 2 contains N297A or DANA mutations (D265A+N297A). In some embodiments, the effector-deficient Fc contains K322A and L234A / L235A (LALA) mutations. In some embodiments, the effector-deficient Fc contains F234A and L235A (FALA) mutations. In some embodiments, the Fc fragment is effector-deficient IgG1 Fc, e.g., effector-deficient hIgG1 Fc. In some embodiments, the Fc fragment is human IgG4 Fc (S228P). In some embodiments, the Fc fragment is IgG4 Fc (FALA). In some embodiments, CB2 HCAb is monomeric. In some embodiments, CB2 HCAb is dimeric. In some embodiments, CB2 HCAb is multispecific and multivalent (e.g., bispecific and divalent), for example It comprises two or more different CB2 antibodies described herein. In some embodiments, the CB2 HCAb is monospecific and multivalent ( for example , 2gaim), for example It contains two or more copies of the same CB2 ISVD. An exemplary IgG4 Fc as follows is provided.

[0334] APEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISK AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG (SEQ ID NO: 130)

[0335] In addition, the following exemplary IgG1 Fc sequence is provided.

[0336] EPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALGAP IEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 363)

[0337] EPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAP IEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 364)

[0338] EPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALGAP IEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 365)

[0339] In some embodiments, the CB2 antibody and the Fc fragment are fused to each other through a peptide linker. In some embodiments, the peptide linker is (G4S)3 (SEQ ID NO. 132).

[0340] Accordingly, in some embodiments, an isolated CB2 HCAb comprising an ISVD that specifically recognizes CB2 is provided, wherein the CB2 antibody comprises a CDR1 comprising the amino acid sequence of SEQ ID NO. 1, or a variant thereof comprising up to 3 amino acid substitutions (e.g., any number of 1, 2, or 3); a CDR2 comprising the amino acid sequence of SEQ ID NO. 2, or a variant thereof comprising up to 3 amino acid substitutions (e.g., any number of 1, 2, or 3); and a CDR3 comprising the amino acid sequence of SEQ ID NO. 3, or a variant thereof comprising up to 3 amino acid substitutions (e.g., any number of 1, 2, or 3), and the CB2 ISVD is fused to the Fc fragment of the immunoglobulin. In some embodiments, the ISVD comprises (1) any one FR1 of SEQ ID NOs 4 to 15; (2) any one FR2 of SEQ ID NOs 16 to 30; (3) any one FR3 of SEQ ID NOs 31 to 48; and (4) any one FR4 of SEQ ID NOs 49 to 54 are included. In some embodiments, an isolated CB2 HCAb is provided comprising an ISVD that specifically recognizes CB2, wherein the ISVD comprises any one amino acid sequence of SEQ ID NOs 55 to 77, or a variant thereof having at least about 80% sequence identity with any one of SEQ ID NOs 55 to 77 (e.g., at least about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or any value of 99%), and the CB2 antibody is fused to the Fc fragment of the immunoglobulin. In some embodiments, an isolated CB2 HCAb comprising an ISVD that specifically recognizes CB2 is provided, wherein the ISVD comprises any one amino acid sequence of SEQ ID NOs 55 to 77, and a CB2 antibody is fused to the Fc fragment of the immunoglobulin. In some embodiments, the CB2 antibody is fused to the Fc fragment via a peptide linker.In some embodiments, CB2 HCAb is monomeric. In some embodiments, the Fc fragment is human IgG1 Fc, effector-deficient human IgG1 Fc, hIgG2 Fc, human IgG4 Fc, IgG4 Fc (F234A / L235A), or IgG1 (L234A / L235A, numbering according to the EU index of Kabat).

[0341] In some embodiments, an isolated CB2 HCAb is provided comprising any one amino acid sequence of SEQ ID NOs 78 to 95, or a variant thereof having at least about 80% sequence identity with any one of SEQ ID NOs 78 to 95 (e.g., at least about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or any value of 99%). In some embodiments, a polypeptide comprising any one amino acid sequence of SEQ ID NOs 78 to 95 is provided.

[0342] Additionally, an isolated CB2 HCAb is provided that specifically binds to CB2 competitively with any one of the isolated CB2 HCAb or CB2 antibody described herein.

[0343] peptide linker

[0344] Various domains and components within a CB2 antibody construct, such as a CB2 ISVD (meaning an ISVD that specifically binds to CB2), an Fc fragment, a first antigen-binding portion, and a second antigen-binding portion, may be fused together via a suitable linker, such as a peptide linker. The length, degree of flexibility, and / or other characteristics of the peptide linker(s) used in the CB2 antibody may affect characteristics including, but not limited to, affinity, specificity, or binding strength for one or more specific antigens or epitopes. For example, a longer peptide linker may be selected to ensure that two adjacent domains do not sterically interfere with each other. In some embodiments, the peptide linker includes flexible residues (e.g., glycine and serine) to allow adjacent domains to move freely relative to each other. For example, a glycine-serine doublet may be a suitable peptide linker.

[0345] The peptide linker may have any suitable length. In some embodiments, the peptide linker has a length consisting of at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 50, 75, 100 amino acids or more. In some embodiments, the peptide linker has a length consisting of any number of amino acids, or fewer than about 100, 75, 50, 40, 35, 30, 25, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, or 5. In some embodiments, the length of the peptide linker is any of about 1 to about 10 amino acids, about 1 to about 20 amino acids, about 1 to about 30 amino acids, about 5 to about 15 amino acids, about 10 to about 25 amino acids, about 5 to about 30 amino acids, about 10 to about 30 amino acids, about 30 to about 50 amino acids, about 50 to about 100 amino acids, or about 1 to about 100 amino acids.

[0346] Peptide linkers can have naturally occurring or non-naturally occurring sequences. For example, a sequence derived from the hinge region of a heavy-chain monoclonal antibody can be used as a linker. For example, WO1996 / 34103 Refer to In some embodiments, the linker is a (G4S)3 linker (Sequence No. 132).

[0347] (III) Antibody variants

[0348] In some embodiments, amino acid sequence variants of the CB2 antibody provided herein are considered. For example, it may be desirable to improve the binding affinity and / or other biological properties of the antibody. Amino acid sequence variants of the antibody may be prepared by introducing appropriate modifications to the nucleic acid sequence encoding the antibody, or by peptide synthesis. Such modifications include, for example, deletion, insertion, and / or substitution of residues within the amino acid sequence of the antibody. The final product having the desired properties, for example As long as antigen binding is present, such final constructs can be derived by any combination of deletions, insertions, and substitutions.

[0349] a) Substitutions, insertions, deletions, and variants

[0350] In some embodiments, an antibody variant having one or more amino acid substitutions is provided. Sites of interest for substitution mutagenesis include CDR and FR. Conservative substitutions are Table 5 It is presented under the heading 'Desirable Substitution'. More substantial changes Table 5 Under the heading 'Exemplary Substitutions', and in relation to amino acid side chain classes, as further described below. Amino acid substitutions can be introduced into the antibody of interest, and the product has the desired activity, for example It can be screened for maintained / improved antigen binding, reduced immunogenicity, or modified ADCC or CDC.

[0351]

[0352] Amino acids can be classified according to common side chain characteristics into (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 affecting chain orientation: Gly, Pro; and (6) aromatic: Trp, Tyr, Phe.

[0353] Non-conservative substitution involves exchanging a member of one of these classes for another.

[0354] In some embodiments, the CB2 antibody provided herein comprises an ISVD comprising an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with any one of SEQ ID NOs 55 to 77. In some embodiments, the ISVD sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity has a substitution ( for example , conservative substitution), including insertions or deletions, but CB2 antibodies containing such sequences are CB2 ( for example It retains the ability to bind to human CB2, murine CB2, rat CB2, and / or synomolgus CB2). In some embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted from any of SEQ ID NOs 55 to 77. In some embodiments, the substitution, insertion, or deletion is in a region outside the CDR ( in other words Occurs in , FR). In some embodiments, the CB2 antibody comprises the ISVD presented in any one of SEQ ID NOs 55 to 77, which includes a post-translational modification of the sequence.

[0355] One type of substitution variant is the parent antibody ( for example It involves substituting one or more hypervariable domain residues of a humanized antibody or human antibody. Generally, the resulting variant(s) selected for further study have specific biological characteristics ( for example Modification in (, increased affinity, decreased immunogenicity) for exampleIt will have (improvement) or substantially retain specific biological characteristics of the parent antibody. An exemplary substitution variant is an affinity-mature antibody, which can be conveniently produced.

[0356] A useful method for identifying antibody residues or regions that can be targets for mutagenicity is [Cunningham and Wells (1989)] Science It is referred to as 'alanine scanning mutagenesis' as described in [ , 244:1081-1085]. In this method, to determine whether the interaction between the antibody and the antigen is affected, a residue or a target residue group ( for example Identify charged residues such as , Arg, Asp, His, Lys, and Glu, and neutral or negatively charged amino acids ( for example It is replaced with (alanine or polyalanine). Additional substitutions may be introduced at amino acid positions that exhibit functional sensitivity to the initial substitution. Alternatively or additionally, the crystal structure of the antigen-antibody complex is used to identify contact points between the antibody and the antigen. These contact residues and neighboring residues can be targeted as substitution candidates or eliminated. Variants may be screened to determine whether they possess the desired characteristics.

[0357] Amino acid sequence insertion includes amino-terminal and / or carboxyl-terminal fusions ranging in length from one residue to polypeptides containing 100 or more residues, as well as the insertion of single or multiple amino acid residues into the sequence. Examples of terminal insertion include antibodies having an N-terminal methionyl residue. Other insertion variants of antibody molecules include the N-terminus or C-terminus of the antibody, which are enzymes ( for example This includes fusing to a polypeptide that increases the serum half-life of the antibody (for ADEPT) or antibody.

[0358] b) Glycosylated variant

[0359] In some embodiments, the isolated CB2 antibody provided herein is modified to increase or decrease the degree to which the corresponding composition is glycosylated. The addition or deletion of glycosylation sites to the antibody can be conveniently achieved by modifying the amino acid sequence so that one or more glycosylation sites are created or removed.

[0360] CB2 antibody in Fc region[ for example , CB2 antibody-Fc fusion protein( for example If [, HCAb)] is included, the attached carbohydrate may be altered. Natural antibodies produced by mammalian cells generally contain branched double-antenna type oligosaccharides, which are typically connected to the C of the Fc region by N-linking. H It is attached to Asn297 of the 2 domain. For example Enter , literature[Wright et al. TIBTECH Refer to [15:26-32 (1997)]. The corresponding oligosaccharides are various carbohydrates, for example It may include mannose, N-acetylglucosamine (GlcNAc), galactose, and sialic acid, as well as fucose attached to GlcNAc in the 'stem' of the double-antenna type oligosaccharide structure. In some embodiments, modifications of the oligosaccharides in the CB2 antibody of the present application may be made to produce antibody variants having specific improved properties.

[0361] In some embodiments, a CB2 antibody variant having a carbohydrate structure lacking fucose attached (directly or indirectly) to the Fc region is provided. For example, the amount of fucose in such antibodies may be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. The amount of fucose is, for example, when measured by MALDI-TOF mass spectrometry as described in WO 2008 / 077546, all glycostructures attached to Asn297 ( for exampleIt is determined by calculating the average amount of fucose within the sugar chain at Asn297 relative to the sum of the complex, hybrid, and gomannose structures. Asn297 refers to an asparagine residue located at approximately position 297 of the Fc region (EU numbering of Fc region residues), but due to minute sequence variations in the antibody, Asn297 is within approximately 3 amino acids upstream or downstream of position 297, in other words It may be located between positions 294 and 300. These fucosylated variants may have improved ADCC capabilities. for example See U.S. Patent Publication US 2003 / 0157108 (Presta, L.) and U.S. Patent Publication US 2004 / 0093621 (Kyowa Hakko Kogyo Co., Ltd). Examples of literature related to 'defucosylated' or 'fucose-deficient' antibody variants include US 2003 / 0157108, WO 2000 / 61739, WO 2001 / 29246, US 2003 / 0115614, US 2002 / 0164328, US 2004 / 0093621, US 2004 / 0132140, US 2004 / 0110704, US 2004 / 0110282, US 2004 / 0109865, WO 2003 / 085119, WO 2003 / 084570, WO 2005 / 035586, WO 2005 / 035778, WO2005 / 053742, WO2002 / 031140, Literature [Okazaki et al.J. Mol. Biol. 336:1239-1249 (2004)], literature[Yamane-Ohnuki et al. Examples of cell lines capable of producing defucosylated antibodies include Lec13 CHO cells deficient in protein fucosylation (Literature [Ripka et al. Biochem. Biophys. 249:533-545 (1986)]; U.S. Patent Application US 2003 / 0157108 A1 (Presta, L); and WO 2004 / 056312 A1 (Adams et al.)(particularly Example 11)), and knockout cell lines, e.g., alpha-1,6-fucosyltransferase gene FUT8 Knockout CHO cells ( for example , literature[Yamane-Ohnuki et al. Bioeng. 87: 614 (2004)]; Literature[Kanda, Y. et al. Includes , Bioeng., 94(4):680-688 (2006)]; and WO2003 / 085107).

[0362] A CB2 antibody variant having a bisected oligosaccharide is additionally provided, and for example This is a bidentate oligosaccharide attached to the Fc region of the antibody by GlcNAc. These antibody variants may have reduced fucosylation and / or enhanced ADCC function. Examples of these antibody variants are, for example , WO 2003 / 011878(Jean-Mairet et al. ), U.S. Patent No. 6,602,684 (Umana et al. ), and US 2005 / 0123546(Umana et al. It is described in ). An antibody variant having at least one galactose residue in an oligosaccharide attached to the Fc region is also provided. This antibody variant may have an enhanced CDC function. This antibody variant, for example , WO 1997 / 30087(Patel et al. It is described in WO 1998 / 58964 (Raju, S.), and WO 1999 / 22764 (Raju, S.).

[0363] c) Fc region variant

[0364] In some embodiments, one or more amino acid modifications may be introduced into the Fc region of the CB2 antibody provided herein to generate an Fc region variant. The Fc region variant has amino acid modifications at one or more amino acid positions ( for example Human Fc region sequences containing , substitutions ( for exampleIt may include human IgG1, IgG2, IgG3, or IgG4 Fc regions.

[0365] In some embodiments, the present application considers a CB2 antibody comprising an Fc region variant having some, but not all, effector function, wherein such antibody In vivo While the half-life of CB2 antibodies is important, specific effector functions (e.g., complement and ADCC) make them desirable candidates for unnecessary or harmful applications. To confirm the reduction / depletion of CDC and / or ADCC activity In a test tube and / or In vivo Cytotoxicity assays may be performed. For example, Fc receptor (FcR) binding assays may be performed to confirm that the antibody lacks FcR binding (and is therefore likely to lack ADCC activity) but retains FcRn binding ability. NK cells, the major cells mediating ADCC, express only FcγRIII, whereas monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression in hematopoietic cells [Ravetch and Kinet, Annu. Rev. Immunol. It is summarized in Table 2 on page 464 of [9:457-492 (1991)]. To evaluate the ADCC activity of the molecule of interest In a test tube A non-limiting example of testing is U.S. Patent No. 5,500,362 ( for example , literature[Hellstrom, I. et al. Proc. Nat'l Acad. Sci. USA 83:7059-7063 (1986)] reference ) and literature[Hellstrom, I et al. , Proc. Nat'l Acad. Sci. USA 82:1499-1502 (1985)], No. 5,821,337 (Literature [Bruggemann, M. et al.It is described in [see , Med. 166:1351-1361 (1987)]. Alternatively, non-radioactive assay methods may be used [e.g., ACTI’s non-radioactive cytotoxicity assay for flow cytometry (CellTechnology, Inc., Mountain View, California), and CytoTox 96 ® Non-radiotoxicity assay (Promega, Madison, Wisconsin) reference Effector cells useful for these assays include peripheral blood mononuclear cells (PBMCs) and natural killer (NK) cells. Alternatively or additionally, the ADCC activity of the molecule of interest is In vivo , for example Literature [Clynes et al. Proc. Nat'l Acad. Sci. USA It can be evaluated in animal models such as that disclosed in [95:652-656 (1998)]. A C1q binding assay can also be performed to confirm that the antibody cannot bind to C1q and thus lacks CDC activity. for example , refer to the C1q and C3c binding ELISAs of WO 2006 / 029879 and WO 2005 / 100402. To evaluate complement activation, a CDC assay may be performed ( for example , literature[Gazzano-Santoro et al. , Methods 202:163 (1996)], Literature[Cragg, MS et al. , Blood 101:1045-1052 (2003)], and literature[Cragg, MS and MJ Glennie, Blood [Refer to 103:2738-2743 (2004)]). FcRn binding and In vivo The determination of clearance / half-life can also be performed using methods known in the art ( for example , literature[Petkova, SB et al. [See , Immunol. 18(12):1759-1769 (2006)]).

[0366] Antibodies having reduced effector function include antibodies having one or more substitutions of Fc region residues 238, 265, 269, 270, 297, 327, and 329 (U.S. Patent No. 6,737,056). These Fc mutants include Fc mutants having substitutions at two or more of amino acid positions 265, 269, 270, 297, and 327, for example, the so-called 'DANA' Fc mutant in which residues 265 and 297 are substituted with alanine (U.S. Patent No. 7,332,581). Fc mutants are [Schlothauer et al. Protein Eng Des Sel. 2016 Oct;29(10):457-466], literature[Pejchal et al. Antibodies . 2023; 12(3):54], literature[Dall'Acqua et al. J Biol Chem. 2006 Aug 18;281(33):23514-24], literature[Ramdani et al. International Journal of Molecular Sciences Described in . 2022; 23(17):9604], which are incorporated herein by reference.

[0367] Specific antibody variants having improved or reduced binding to FcR are described ( for example , U.S. Patent No. 6,737,056, WO 2004 / 056312, and literature [Shields et al. , J. Biol. Chem. [See 9(2): 6591-6604 (2001)]).

[0368] In some embodiments, the CB2 antibody variant has one or more amino acid substitutions that reduce ADCC activity, for example Includes an Fc region having substitutions at locations 298, 333 and / or 334 of the Fc region (residual numbers according to EU numbering).

[0369] In some embodiments, modified ( in other words Changes resulting in C1q binding and / or complement-dependent cytotoxicity (CDC) (improved or reduced) occur in the Fc region, which is for exampleU.S. Patent No. 6,194,551, WO 99 / 51642, and literature [Idusogie et al.J. Immunol. As described in 164: 4178-4184 (2000).

[0370] In some embodiments, a CB2 antibody is provided comprising a variant Fc region comprising one or more amino acid substitutions that increase the half-life and / or improve binding to the neonatal Fc receptor (FcRn). An antibody having an increased half-life and improved binding to the neonatal Fc receptor (FcRn) responsible for transferring maternal IgG to the fetus (Reference [Guyer et al. , J . Immunol. 117:587 (1976)] and literature[Kim et al. , J. Immunol. 24:249 (1994)]) is US2005 / 0014934A1(Hinton et al. It is described in ). Such antibodies comprise an Fc region having one or more internal substitutions that improve the binding of the Fc region to FcRn. These Fc variants include variants having substitutions on one or more Fc region residues, for example Variants having a substitution of Fc region residue 434 are included (U.S. Patent No. 7,371,826).

[0371] For other examples of Fc region variants, see the literature [Duncan & Winter, Nature See also 322:738-40 (1988)], U.S. Patent No. 5,648,260, U.S. Patent No. 5,624,821, and WO 94 / 29351.

[0372] A CB2 antibody comprising any of the Fc variants described herein, or a combination thereof, is considered.

[0373] d) Antibody derivative

[0374] In some embodiments, the CB2 antibody provided herein may be further modified to include additional non-protein moiety that is known in the art and readily available. Moiety suitable for derivatization of the antibody includes, but is not limited to, water-soluble polymers. Non-limiting examples of water-soluble polymers include polyethylene glycol (PEG), ethylene glycol / propylene glycol copolymer, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene / maleic anhydride copolymer, polyamino acids (homogeneous or random copolymer), and dextran or poly(n-vinylpyrrolidone)polyethylene glycol, propropylene glycol homopolymer, prolypropylene oxide / ethylene oxide copolymer, polyoxyethylated polyol ( for example Polyethylene glycol propionaldehyde includes, but is not limited to, glycerol, polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde may have a manufacturing advantage due to its stability in water. Polymers may have any molecular weight and may be branched or unbranched. The number of polymers attached to the antibody may vary, and if two or more polymers are attached, they may be the same or different molecules. Generally, the number and / or type of polymers used for derivatization depends on the specific characteristics or functions of the antibody to be improved, and whether the antibody derivative will be used in therapy under specified conditions. etc. It may be determined based on considerations including but not limited to.

[0375] In some embodiments, a conjugate of a CB2 antibody and a non-protein moiety is provided, which can be selectively heated by radiation exposure. In some embodiments, the non-protein moiety is a carbon nanotube (Kam et al. , Proc. Natl. Acad. Sci. USA102: 11600-11605 (2005)). Radiation may have any wavelength and includes, but is not limited to, wavelengths that do not harm normal cells but heat the non-protein moiety to a temperature at which cells near the antibody-non-protein moiety die.

[0376] In some embodiments, the CB2 antibody provided herein may be further modified to include one or more biologically active proteins, polypeptides, or fragments thereof. As used interchangeably herein, 'bioactive' or 'biological activity' means exhibiting biological activity in the body to perform a specific function. For example, this includes proteins, DNA etc. It may imply binding to specific biomolecules such as, and subsequently promoting or inhibiting the activity of such biomolecules. In some embodiments, the bioactive protein or its fragment includes proteins and polypeptides administered to a patient as active drug substances for the prevention or treatment of diseases or pathological conditions, as well as proteins and polypeptides used for diagnostic purposes, such as diagnostic tests or In a test tube It includes enzymes used for testing, and proteins and polypeptides administered to patients to prevent disease, such as vaccines. In some embodiments, the bioactive protein or its fragment has immunostimulatory / immunomodulatory, membrane transport, or enzymatic activity. In some embodiments, the bioactive protein, polypeptide, or its fragment is an enzyme, hormone, growth factor, cytokine, or a mixture thereof. In some embodiments, the bioactive protein, polypeptide, or fragment can specifically recognize a target peptide (e.g., an antigen or another protein).

[0377] In some embodiments, the bioactive protein or fragment thereof that may be included in the CB2 antibody described herein is a protein-binding protein. In some embodiments, the bioactive protein or fragment thereof that may be included in the CB2 antibody described herein is an antibody mimic, which is an engineered small protein containing an antigen-binding domain similar to that of an antibody (Reference [Geering and Fussenegger, Trends Biotechnol., 33(2):65-79, 2015]). These molecules are derived from existing human scaffold proteins and comprise a single polypeptide. Exemplary antibody mimics that may be included in the CB2 antibody described herein may be, but are not limited to, a designed ankyrin repeat protein (DARPin; comprising 3 to 5 fully synthetic ankyrin repeats adjacent to N-terminal and C-terminal Cap domains), an avidity multimer [avimer, a high-affinity protein comprising multiple A domains, each domain having low affinity for a target], or an anticalin (based on a scaffold of lipocalin, having 4 accessible loops, the sequence of each loop may be randomized). In some embodiments, a bioactive protein or fragment thereof that may be included in the CB2 antibody described herein is an armadillo repeat protein [ for example[β-catenin, α-impotin, placoglobin, adenomatous polyposis (APC) protein], which contains an armadillo repeat unit (a characteristic, repetitive amino acid sequence of about 40 residues in length). Each armadillo repeat unit consists of a pair of alpha helices forming a hairpin structure. Multiple copies of the repeat unit form a so-called alpha solenoid structure. Armadillo repeat proteins can bind to different types of peptides by relying on a specific binding mode of the peptide backbone, without requiring interaction with specific conserved side chains or the free N-terminus or C-terminus of the peptide. The ability to recognize peptides at the residue level, combined with the unique modularity of the repeat protein, makes armadillo repeat proteins promising candidates for general scaffold designs for peptide binding.

[0378] In some embodiments, the biologically active protein or fragment thereof that may be included in the CB2 antibody described herein is a ligand, such as a lymphokine and a cell factor that interacts with a specific cell receptor. A lymphokine is a low molecular weight protein secreted by a T cell when an antigen or lectin stimulates T cell growth.

[0379] III. Pharmaceutical Composition

[0380] The present application further provides a pharmaceutical composition comprising any one of the CB2 antibodies described herein and optionally a pharmaceutically acceptable carrier. The pharmaceutical composition may be prepared in the form of a lyophilized formulation or an aqueous solution by mixing the CB2 antibody described herein having a desired purity with any pharmaceutically acceptable carrier, excipient, or stabilizer [Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980)].

[0381] It is preferable that the pharmaceutical composition be a stable composition in which the CB2 antibody described herein essentially maintains its physical and chemical stability and integrity during storage. Various analytical techniques for measuring protein stability are available in the art, and literature [ Peptide and Protein Drug Delivery , 247-301, Vincent Lee Ed., Marcel Dekker, Inc., New York, NY, Pubs. (1991)] and literature[Jones, A. Adv. Drug Delivery Rev. [10: 29-90 (1993)] is reviewed. Stability may be measured at a selected temperature for a selected period. For rapid screening, the formulation may be maintained at 40°C for 2 weeks to 1 month, at which point stability is measured. If the formulation is stored at 2 to 8°C, it should generally be stable for at least 1 month at 30°C or 40°C and / or for at least 2 years at 2 to 8°C. If the formulation is stored at 30°C, it should generally be stable for at least 2 years at 30°C and / or for at least 6 months at 40°C. For example, the degree of aggregation during storage may be used as an indicator of protein stability. In some embodiments, the stable formulation of the CB2 antibody described herein may contain less than about 10% (preferably less than about 5%) of the CB2 antibody present in the formulation in aggregate form.

[0382] Acceptable carriers, excipients, or stabilizers are non-toxic to the recipient at the dosage and concentration used, and include buffers, ascorbic acid, methionine, vitamin E, antioxidants including sodium metabisulfite; preservatives, isotonic agents ( for example , sodium chloride), stabilizer, metal complex ( for example , Zn-protein complex); includes chelating agents such as EDTA and / or nonionic surfactants.

[0383] Examples of physiologically acceptable carriers include buffers such as phosphates, citrates, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (e.g., octadecyldimethylbenzylammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl, or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins such as serum albumin, gelatin, or immunoglobulin; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrin; and chelating agents such as EDTA; Sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium; metal complexes ( for example , Zn-protein complex); and / or nonionic surfactants such as TWEEN, polyethylene glycol (PEG) and PLURONICS, or polyethylene glycol (PEG) are included.

[0384] Buffers are used to adjust the pH to a range that optimizes therapeutic efficacy, particularly when stability is pH-dependent. Buffers are preferably present at a concentration ranging from about 50 mM to about 250 mM. Buffers suitable for use in this application include organic and inorganic acids and their salts. Examples include citrate, phosphate, succinate, tartrate, fumarate, gluconate, oxalate, lactate, and acetate. Additionally, buffers may include trimethylamine salts such as histidine and Tris.

[0385] Preservatives are added to delay microbial growth and are typically present in the range of 0.2% to 1.0% (w / v). The addition of preservatives can, for example, facilitate the production of multi-use (multiple-dose) formulations. Preservatives suitable for use in this application include octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; and benzalkonium halide ( for example Includes , chloride, bromide, iodide), benzethonium chloride; thimerosal, phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol, 3-pentanol and m-cresol.

[0386] Isotonic agents, also referred to as 'stabilizers,' are present to adjust or maintain the liquid properties within the composition. Isotonic agents are commonly referred to as 'stabilizers' because, when used with large, charged biomolecules such as proteins and antibodies, they can reduce the potential for intermolecular and intramolecular interactions by interacting with the charged groups of amino acid side chains. Isotonic agents may be present in any amount from 0.1% to 25% by weight, preferably from 1% to 5% by weight, taking into account the relative amounts of other components. Preferred isotonic agents include polyvalent sugar alcohols, preferably trivalent or higher sugar alcohols such as glycerin, erythritol, arabitol, xylitol, sorbitol, and mannitol.

[0387] Additional excipients include agents that can act as one or more of (1) fillers, (2) solubility enhancers, (3) stabilizers, and (4) agents that prevent denaturation or adhesion to the container wall. These excipients include polysaccharide alcohols (listed above); amino acids such as alanine, glycine, glutamine, asparagine, histidine, arginine, lysine, ornithine, leucine, 2-phenylalanine, glutamic acid, threonine, etc.; sucrose, lactose, lactitol, trehalose, stachyose, mannose, sorbose, xylose, ribose, ribitol, myoinisitose, myoinisitol, galactose, galactitol, glycerol, cyclitol ( for example Organic sugars or sugar alcohols such as polyethylene glycol (, inositol); sulfur-containing reducing agents such as urea, glutathione, thioctic acid, sodium thioglycolate, thioglycerol, α-monothioglycerol, and sodium thiosulfate; low molecular weight proteins such as human serum albumin, bovine serum albumin, gelatin, or other immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; monosaccharides ( for example , xylose, mannose, fructose, glucose); disaccharides( for example It includes , lactose, maltose, sucrose); trisaccharides such as raffinose; and polysaccharides such as dextrin or dextran.

[0388] Nonionic surfactants or detergents (also known as 'wetting agents') are present not only to aid in the solubilization of the therapeutic agent but also to protect the therapeutic protein from agitation-induced aggregation, which also allows the formulation to be exposed to shear surface stress without causing denaturation of the active therapeutic protein or antibody. Nonionic surfactants are present in the range of about 0.05 mg / ml to about 1.0 mg / ml, preferably about 0.07 mg / ml to about 0.2 mg / ml.

[0389] Suitable nonionic surfactants include polysorbates (20, 40, 60, 65, 80, etc.), polyoxamers (184, 188, etc.), and pluronic® Polyol, TRITON ® Includes polyoxyethylene sorbitan monoethers (TWEEN®-20, TWEEN®-80, etc.), lauromacrogol 400, polyoxyl 40 stearate, polyoxyethylene hydrogenated castor oil 10, 50, and 60, glycerol monostearate, sucrose fatty acid esters, methyl cellulose, and carboxymethyl cellulose. Anionic detergents that may be used include sodium lauryl sulfate, dioctyl sodium sulfosuccinate, and dioctyl sodium sulfonate. Cationic detergents include benzalkonium chloride or benzethonium chloride.

[0390] A pharmaceutical composition In vivo It must be sterilized to be used for administration. The pharmaceutical composition may be sterilized by filtration through a sterile filter membrane. The pharmaceutical composition of the present invention is generally contained in a container having a sterile access port, for example, an intravenous solution bag or vial having a stopper that can be punctured with a subcutaneous injection needle.

[0391] The route of administration is a known and accepted method, e.g., a single or multiple bolus or injection over a long period in a suitable manner, for example It follows injection or infusion by subcutaneous, intravenous, intra-abdominal, intramuscular, intra-arterial, intralesional, or intra-articular routes, local administration, inhalation, or sustained release or extended release means.

[0392] A sustained-release formulation can be manufactured. Suitable examples of sustained-release formulations include a semipermeable matrix of a solid hydrophobic polymer containing an antagonist, and this matrix is ​​a molded article, for exampleIt is in the form of a film or microcapsule. Examples of sustained-release matrices include polyesters, hydrogels [e.g., poly(2-hydroxyethyl-methacrylate) or poly(vinyl alcohol)], polylactide (U.S. Patent No. 3,773,919), copolymers of L-glutamic acid and ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as LUPRON DEPOTZE (injectable microspheres composed of a lactic acid-glycolic acid copolymer and leuprolide acetate), and poly-D-(-)-3-hydroxybutyric acid.

[0393] The pharmaceutical composition of the present invention may also contain two or more active compounds as needed for the specific indication being treated, preferably active compounds having complementary activities that do not negatively affect each other. Alternatively or additionally, the composition may include cytotoxic agents, chemotherapy agents, cytokines, immunosuppressants, or growth inhibitors. These molecules are suitably present in combination in amounts effective for the intended purpose.

[0394] The active ingredient may also be encapsulated, for example, by coacervation technology or interfacial polymerization, into microcapsules prepared, for example, from hydroxymethylcellulose or gelatin microcapsules and poly-(methyl methacrylate) microcapsules, respectively, within colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles, and nanocapsules), or within macroemulsions. Such technology is described in the literature [ Remington's Pharmaceutical Sciences It is disclosed in the 18th edition.

[0395] In some embodiments, the pharmaceutical composition is contained in a single-use vial, such as a single-use sealed vial. In some embodiments, the pharmaceutical composition is contained in a multi-use vial. In some embodiments, the pharmaceutical composition is contained in bulk in a container. In some embodiments, the pharmaceutical composition is cryopreserved.

[0396] IV. Treatment Methods

[0397] The CB2 antibody and its compositions (e.g., pharmaceutical compositions) described herein are useful for various uses, such as diagnosis, molecular assays, and therapy.

[0398] In some embodiments, a method acting as an agonist for CB2 on a cell is provided, the method comprising the step of contacting a cell with any one of the CB2 antibodies described herein in an amount sufficient to activate CB2 on the cell. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the method In a test tube The method is performed. In some embodiments, the method In vivo It is performed. In some embodiments, acting as an agonist on CB2 on cells treats or improves a disease or pathological condition. The action of CB2 on the subject can trigger acute and sustained anti-algesic effects.

[0399] In some embodiments, a method for treating a disease or condition in a subject is provided, wherein the disease or condition is improved by acting as an agonist for CB2, and the method comprises the step of administering to the subject an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the disease or condition is chemotherapy-induced peripheral neuropathy (CIPN). In some embodiments, the disease or condition is a respiratory infection. In some embodiments, the disease or condition is hepatic fibrosis. In some embodiments, the disease or condition is cold sensitivity. In some embodiments, the disease or condition is inflammatory bowel disease. In some embodiments, the disease or condition is endometriosis.

[0400] Chemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of chemotherapy for cancer treatment. CIPN is generally associated with neurotoxic chemotherapy agents. CIPN is a painful and dose-limiting side effect that can limit treatment tolerance, impair patient prognosis, reduce treatment adherence, and lower the patient's quality of life. Taxol drugs, such as paclitaxel, are commonly associated with CIPN. The CB2 antibody described herein is useful for treating or improving CIPN, including CIPN induced by taxol drugs.

[0401] In some embodiments, a method for treating chemotherapy-induced peripheral neuropathy (CIPN) in a subject receiving a chemotherapy agent is provided, the method comprising the step of administering to the subject an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, and CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the chemotherapy agent is a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the chemotherapy agent is vincristine. In some embodiments, the chemotherapy agent is cisplatin.

[0402] In some embodiments, a method for treating cancer in a subject requiring treatment for cancer is provided, the method comprising the step of administering to the subject an effective amount of a chemotherapy agent and an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the chemotherapy agent is a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the chemotherapy agent is vincristine. In some embodiments, the chemotherapy agent is cisplatin. In some embodiments, the chemotherapy agent causes chemotherapy-induced peripheral neuropathy (CIPN).

[0403] A significant number of cancer patients treated with neurotoxic chemotherapy experience chemotherapy-induced peripheral neuropathy. CIPN, among other adverse effects, can affect a subject's health-related quality of life, treatment adherence, and appetite, and can exacerbate existing pain. Taken together, these effects limit the dose of neurotoxic chemotherapy that a subject can tolerate; if there were no such limitation, higher doses would have been more effective in treating the cancer. CIPN can be measured by appropriate means known in the art, including the administration of questionnaires or the application of diagnostic protocols. Exemplary questionnaires and protocols for CIPN include the Functional Assessment of Cancer Therapy / Gynecologic Oncology Group Neurotoxicity Questionnaire (FACT / GOG-Ntx), the Chemotherapy-induced Peripheral Neuropathy Assessment Tool (CIPNAT), the European Organization for Research & Treatment in Cancer Quality of Life Questionnaire-CIPN 20 (EORTC QLQ-CIPN 20), the Modified Total Neuropathy Score (mTNS), the Total Neuropathy Score, clinical version (TNSc), and the 5-item reduced Total Neuropathy Score (TNSr 5-item).Other scales may also be used to evaluate the symptoms of CIPN, such as the Fullerton Advanced Balance Scale (FABS), Timed Up and Go (TUG), Activities Specific Balance Confidence Scale (ABC), Balance Evaluation Systems Test (BESTest), Berg Balance Scale (BBS), Repeated Sit to Stand test, Functional Reach test, Short Physical Performance Battery (SPB), and the grooved peg board test. The methods described herein encompass any appropriate means for evaluating CIPN.

[0404] In some embodiments, a method for improving chemotherapy-induced peripheral neuropathy (CIPN) is provided, the method comprising the step of administering an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, CIPN is induced by a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the chemotherapy agent is vincristine. In some embodiments, the chemotherapy agent is cisplatin.

[0405] In some embodiments, a method for treating adverse effects associated with chemotherapy in cancer patients is provided, the method comprising the step of administering an effective amount of any one of the CB2 antibodies described herein to the patient. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the adverse effect is CIPN or an adverse effect associated with CIPN. In some embodiments, the method further comprises the step of administering a chemotherapy agent. In some embodiments, the chemotherapy agent is a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the chemotherapy agent is vincristine. In some embodiments, the chemotherapy agent is cisplatin.

[0406] In some embodiments, a method for improving the quality of life of a subject receiving chemotherapy is provided, comprising the step of administering an effective amount of any one of the CB2 antibodies described herein to the patient. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a CDR1 comprising SEQ NO. 1, a CDR2 comprising SEQ NO. 2, and a CDR3 comprising SEQ NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. The quality of life may be assessed by suitable means known in the art, including a Quality of Life Questionnaire (QoLQ). The QoLQ may be completed by the patient or by a proxy. The method described herein encompasses any suitable QoLQ. In some embodiments, the method further comprises the step of administering a chemotherapy agent. In some embodiments, the chemotherapy agent is a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the chemotherapy agent is vincristine. In some embodiments, the chemotherapy agent is cisplatin.

[0407] In some embodiments, a method for improving compliance of a subject receiving chemotherapy is provided, the method comprising the step of administering an effective amount of any one of the CB2 antibodies described herein to a patient. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the method further comprises the step of administering a chemotherapy agent. In some embodiments, the chemotherapy agent is a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the chemotherapy agent is vincristine. In some embodiments, the chemotherapy agent is cisplatin.

[0408] In some embodiments, a method for increasing the dose of a chemotherapy regimen administered to a subject is provided, the method comprising the step of administering an effective amount of any one of the CB2 antibodies described herein to the subject, and the dose of the chemotherapy regimen is increased after administration of the CB2 antibody. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, and CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the method further comprises the step of administering a chemotherapy agent. In some embodiments, the chemotherapy agent is a taxol drug. In some embodiments, the taxol drug is paclitaxel. In some embodiments, the chemotherapy agent is vincristine. In some embodiments, the chemotherapy agent is cisplatin.

[0409] In some embodiments, a method for treating an individual having a respiratory infection is provided, the method comprising the step of administering to the individual an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the respiratory infection is an influenza infection, such as an influenza infection caused by an influenza A virus or an influenza B virus. In some embodiments, the influenza infection is caused by an influenza A virus subtype H1N1. In some embodiments, the respiratory infection is caused by a coronavirus, such as SARS-CoV-2 (in which case the respiratory infection is COVID-19).

[0410] In some embodiments, a method for alleviating cytokine release syndrome in an individual requiring alleviation of cytokine release syndrome is provided, the method comprising the step of administering to the individual an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. In some embodiments, the method reduces the levels of IL-16 and / or IL-8 secreted by the airway cells of the individual. In some embodiments, the method reduces IL-16 secretion by airway cells by about 20% or more, e.g., about 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, and 90% or more, and values ​​and ranges between these. In some embodiments, the method reduces IL-8 secretion by airway cells by about 20% or more, e.g., about 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, and 90% or more, and values ​​and ranges between these. In some embodiments, the method reduces the secretion of IL-8 and IL-16 by airway cells by about 20% or more, e.g., about 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, and 90% or more, and values ​​and ranges between these.

[0411] In some embodiments, a method for treating liver fibrosis is provided, the method comprising the step of administering an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein to an individual. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. Liver fibrosis may be evaluated by suitable means known in the art, including non-invasive means such as liver biopsy or elastography and / or serum biochemical testing.

[0412] In some embodiments, a method for treating cold sensitivity and / or allodynia is provided, the method comprising the step of administering an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein to an individual requiring said treatment. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to IMGT. Cold sensitivity and / or allodynia may be evaluated by suitable means known in the art, including questionnaires and / or patient-reported grades for cold sensitivity or allodynia.

[0413] In some embodiments, a method for treating inflammatory bowel disease is provided, the method comprising the step of administering an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein to an individual requiring said treatment. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, and CDR1, CDR2, and CDR3 are defined according to IMGT.

[0414] In some embodiments, a method for treating endometriosis is provided, the method comprising the step of administering an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein to an individual requiring said treatment. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, wherein the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, and CDR1, CDR2, and CDR3 are defined according to IMGT.

[0415] In some embodiments, a method for treating diabetic peripheral neuropathy (DPN) is provided, comprising the step of administering an effective amount of a pharmaceutical composition comprising any one of the CB2 antibodies described herein to an individual requiring said treatment. In some embodiments, the CB2 antibody comprises an ISVD that specifically binds to CB2, and the ISVD comprises a CDR1 comprising SEQ ID NO. 1, a CDR2 comprising SEQ ID NO. 2, and a CDR3 comprising SEQ ID NO. 3, wherein CDR1, CDR2, and CDR3 are defined according to the IMGT. The dosage and desired drug concentration of the pharmaceutical composition of this application may vary depending on the specific intended use. Determination of an appropriate dosage or route of administration can be readily performed by a person skilled in the art. Animal studies provide reliable guidance for determining an effective dose for human therapy. Interspecies scaling of the effective dose [Mordenti, J. and Chappell, W. "The Use of Interspecies Scaling in Toxicokinetics," In Toxicokinetics and New Drug Development , Yacobi et al. It can be performed according to the principles presented in [ , Eds, Pergamon Press, New York 1989, pp. 42-46].

[0416] of the CB2 antibody described herein (including a pharmaceutical composition comprising the CB2 antibody described herein) In vivoWhen administration is used, the usual dosage may vary from about 10 ng / kg to about 100 mg / kg of mammalian body weight depending on the route of administration. It is within the scope of this application that different agents may be effective for different treatments and different disorders, and that administration to treat a specific organ or tissue may require a different mode of delivery than delivery to other organs or tissues. Additionally, the dosage may be administered as one or more individual doses or as a continuous infusion. In cases of repeated administration over several days or more depending on the pathological condition, treatment continues until the desired suppression of disease symptoms occurs. However, other administration regimens may be useful. The progress of this therapy is easily monitored by conventional techniques and tests.

[0417] In some embodiments, the pharmaceutical composition is administered once ( for example , bolus injection). In some embodiments, the pharmaceutical composition is administered multiple times (e.g., 2, 3, 4, 5, 6, or any number thereof). In the case of multiple administration, such administration may be performed via the same or different routes and may be performed at the same or different sites. The pharmaceutical composition may be administered daily or once a year. The administration interval may be any value between about 24 hours and one year. The administration interval is ( for example , depending on tumor progression) may be irregular. In some embodiments, there is no break in the administration schedule. The optimal dosage and treatment regimen for a specific patient can be easily determined by a person skilled in the medical field who monitors the patient for signs of the disease and adjusts the treatment accordingly.

[0418] The pharmaceutical compositions of the present application, including but not limited to reconstituted formulations and liquid formulations, are administered to an individual requiring treatment, preferably a human, by a route of intramuscular, intraperitoneal, cerebrospinal, subcutaneous, intravenous (iv), intra-articular, synovial, or spinal canal, or orally, locally, or by inhalation, according to known methods, e.g., by intravenous administration as a bolus or by continuous infusion over a period of time. The reconstituted formulation may be prepared by dissolving the lyophilized CB2 antibody described herein in a diluent so that the protein is dispersed throughout. Exemplary pharmaceutically acceptable (safe and non-toxic for administration to humans) diluents suitable for use in the present application include sterile water, bacteriostatic water for injection (BWFI), and pH buffer solution ( for example , phosphate-buffered saline), sterile saline solution, Ringer's solution or dextrose solution, or aqueous solutions of salt and / or buffer are included, but not limited to.

[0419] In some embodiments, the pharmaceutical composition is subcutaneous ( in other words It is administered to an individual by administration (under the skin). For this purpose, the pharmaceutical composition may be injected using a syringe. However, other devices for administering the pharmaceutical composition, such as injection devices, syringe pens, auto-injector devices, needleless devices, and subcutaneous patch delivery systems, are also available. In some embodiments, the pharmaceutical composition is administered intravenously to an individual. In some embodiments, the pharmaceutical composition is administered to an individual by infusion, such as intravenous infusion. Infusion techniques for immunotherapy are known in the art ( for example , literature[Rosenberg et al. , New Eng. J. of Med. 319: 1676 (1988)] reference ).

[0420] In some embodiments, administration to an individual is made by injection. In some embodiments, the pharmaceutical composition is injected multiple times. In some embodiments, the pharmaceutical composition is re-injected once, twice, three times, four times, five times or more. In some embodiments, the pharmaceutical composition is re-injected after about 20 days and after about 25 days.

[0421] V. Manufacturing Method

[0422] The CB2 antibody described herein may be prepared using any method known in the art or the method described herein. In some embodiments, a method for producing a CB2 antibody is provided, the method comprising: (a) culturing a host cell containing an isolated nucleic acid or vector encoding the CB2 antibody described herein under conditions effective for the expression of the encoded CB2 antibody; and (b) obtaining the expressed CB2 antibody from the host cell. In some embodiments, the method of step (a) further comprises the step of producing a host cell containing an isolated nucleic acid or vector encoding the CB2 antibody described herein.

[0423] Methods for preparing antibodies containing ISVD are known in the art. For example, the literature [Els Pardon et al. , Nature Protocol , 2014; 9(3): 674] Refer to . sdAb(e.g., V H H) is a method known in the art, for example Camelidae A hybridoma can be obtained by immunizing a species (e.g., camel or llama) and obtaining the hybridoma therefrom, or by cloning a library of single-domain antibodies using molecular biology techniques known in the art and then selecting by ELISA using individual clones of the non-selective library or by using phage display.

[0424] For the recombinant production of ISVDs, the nucleic acid encoding the antibody is isolated and inserted into a replicatable vector for further cloning (DNA amplification) or expression. The DNA encoding the single-domain antibody is produced using conventional procedures ( for example , using oligonucleotide probes capable of specifically binding to genes encoding the heavy and light chains of antibodies) are easily isolated and sequenced. A number of vectors are available. The choice of vector depends in part on the host cell to be used. Generally, the preferred host cell is of prokaryotic or eukaryotic (typically mammalian) origin.

[0425] In some embodiments, a vector comprising the nucleic acid described herein ( for example , expression vector) is provided. In some embodiments, a host cell containing such nucleic acid or vector is provided. In some embodiments, the host cell is a eukaryotic cell, for example Chinese hamster ovary (CHO) cells, Expi293 cells, or lymphatic cells ( for example , Y0, NS0, Sp20 cells). In some embodiments, the host cell is a prokaryotic cell, for example E. coli It is a cell. In some embodiments, a method for producing a CB2 antibody is provided, the method comprising the steps of: culturing a host cell containing a nucleic acid encoding the antibody under conditions suitable for the expression of the antibody as provided above; and optionally recovering the antibody from the host cell (or host cell culture medium).

[0426] For the recombinant production of CB2 antibodies, for example As described above, nucleic acids encoding antibodies are isolated and inserted into a vector for further cloning and / or expression in host cells. These nucleic acids are obtained using conventional procedures ( for example, can be easily isolated and sequenced using an oligonucleotide probe that can specifically bind to genes encoding the heavy and light chains of antibodies.

[0427] Host cells suitable for the cloning or expression of antibody-coding vectors include the prokaryotic or eukaryotic cells described herein. For example, antibodies may be produced in bacteria, particularly where glycosylation and Fc effector functions are not required. Regarding the expression of antibody fragments and polypeptides in bacteria, for example , see U.S. Patents No. 5,648,237, 5,789,199 and 5,840,523( E. coli Literature describing antibody fragment expression in [Charlton, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ, 2003), pp. 245-254] Also reference After expression, the antibody can be isolated from the bacterial cell paste within the soluble fraction and further purified.

[0428] In addition to prokaryotes, eukaryotic microorganisms such as filamentous fungi or yeasts are suitable cloning or expression hosts for antibody-coding vectors, including fungal and yeast strains in which glycosylation pathways are 'humanized' to produce antibodies having partially or fully human glycosylation patterns. Literature [Gerngross, Nat. Biotech. 22:1409-1414 (2004)] and literature[Li et al. , Nat. Biotech. 24:210-215 (2006)] Refer to .

[0429] Host cells suitable for the expression of glycosylated antibodies also originate from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant cells and insect cells. Along with insect cells, in particular Spodoroptera prugiferda ( Spodoptera frugiperdaA number of baculovirus strains that can be used for the transfection of ) cells have been identified.

[0430] Plant cell cultures can also be used as hosts. For example Enter , U.S. Patents No. 5,959,177, 6,040,498, 6,420,548, 7,125,978 and 6,417,429 (PLANTIBODIES for producing antibodies from transgenic plants TM Refer to the description of the technology.

[0431] Vertebrate cells can also be used as hosts. For example, mammalian cell lines adapted to grow in suspension may be useful. Other examples of useful mammalian host cell lines include the monkey kidney CV1 cell line (COS-7) transformed by SV40; the human embryonic kidney cell line (293 or 293 cells, for example Literature [Graham et al. , J. Gen Virol. Described in 36:59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells ( for example Literature [Mather, Biol. Reprod. TM4 cells as described in 23:243-251 (1980); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); dog kidney cells (MDCK); buffalo rat liver cells (BRL 3A); human lung cells (W138); human liver cells (Hep G2); mouse mammary gland tumor (MMT 060562); for example Literature [Mather et al. , Annals NY Acad. Sci. There are TRI cells as described in [383:44-68 (1982)]; MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include DHFR - CHO cells (literature [Urlaub et al. , Proc. Natl. Acad. Sci. USAChinese hamster ovary (CHO) cells, including [77:4216 (1980)]]; and myeloma cell lines such as Y0, NS0, and Sp2 / 0. A review of specific mammalian host cell lines suitable for antibody production is for example Literature [Yazaki and Wu, Methods in Molecular Biology, Vol. 248 See [BKC Lo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003)].

[0432] In some embodiments, the CB2 antibody described herein may undergo functional maturation or be the result thereof. As described herein, functional maturation involves introducing variability into the antibody backbone using a variable region of the library. In some embodiments, the antibody backbone is ABt285. In some embodiments, the library comprises a sequence of ABt251 having one or more variations of one of two amino acids at each of the six toggle positions, a randomization of one of the six CDR1 (SEQN 1) positions, or a randomization of two of the six CDR3 (SEQN 3) positions. In some embodiments, the six toggle positions are IMGT position 15 (A or P), position 52 (F or W), position 54 (A or S), position 55 (A or G), position 103 (S or W), and position 118 (S or W). In some embodiments, the mutated positions within CDR1 (SEQ No. 1) are IMGT positions 27, 28, 29, 31, 32, or 33 ( in other words , positions 2 to 4 and 6 to 7 of sequence number 1). In some embodiments, any amino acid other than C and M may be placed at randomized positions within CDR1 and CDR3.

[0433] The human CB2 receptor was expressed on the yeast cell membrane and linked to the yeast growth pathway. CB2 receptor-expressing yeast was transformed with a VHH library of ABt285 variants. Stronger variants (with superior titers and / or efficacy) were induced to grow under conditions different from those of ABt285. Agent antibody clones were identified through DNA sequencing of antibody-coding nucleic acids within the transformed yeast.

[0434] VI. Manufactured Products and Kits

[0435] Kits and manufactured products are further provided comprising any of the isolated CB2 antibodies described herein, pharmaceutical compositions comprising any of the CB2 antibodies described herein, isolated nucleic acids or vectors encoding them, or isolated host cells comprising isolated nucleic acids or vectors encoding the CB2 antibodies described herein. In some embodiments, a kit is provided comprising any one of the pharmaceutical compositions described herein and preferably providing instructions for use thereof.

[0436] The kit of the present application is contained in suitable packaging materials. Suitable packaging materials include vials, bottles, jars, and flexible packaging materials ( for example Includes, but is not limited to, sealed Mylar or plastic bags. The kit may optionally provide additional components, such as buffers and interpretation information. Accordingly, the present application also provides manufactured products, which include vials (e.g., sealed vials), bottles, jars, flexible packaging materials, etc.

[0437] The manufactured product may include a container and a label or package insert attached to or enclosed on the container. Suitable containers include, for example, bottles, vials, and syringes. etc.This is included. The container may be formed from various materials such as glass or plastic. Generally, the container holds a composition effective for treating the disease or disorder described herein (e.g., cancer) and may have a sterile access port (e.g., the container may be an intravenous solution bag, or a vial having a stopper perforable with a subcutaneous injection needle). A label or package insert indicates that the composition is used to treat a specific pathological condition in an individual. The label or package insert further includes instructions on how to administer the composition to an individual. The label may contain instructions for reconstitution and / or use. The container holding the pharmaceutical composition may be a multi-dose vial, thereby allowing for repeated administration of the reconstituted preparation ( for example (2 to 6 administrations) may be administered. Package inserts refer to instructions typically included in the commercial packaging of a therapeutic product, which contain information regarding indications, usage, dosage, administration, contraindications, and / or warnings concerning the use of such therapeutic product. Additionally, the product may further comprise a second container containing a pharmaceutically acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate-buffered saline, Ringer's solution, and dextrose solution. It may further comprise other materials desirable from a commercial and user perspective, such as other buffers, diluents, filters, needles, and syringes.

[0438] The kit or product may include multiple unit doses of a pharmaceutical composition and instructions for use, and is packaged in sufficient quantities for storage and use in pharmacies, e.g., hospital pharmacies and dispensing pharmacies.

[0439] Examples

[0440] The following examples are purely for illustrating the invention and should not be construed as limiting the invention in any way. The following examples and detailed description are provided as examples and not as limitations.

[0441] Example 1: Production of CB2 antibody.

[0442] Antibodies functionally activating CB2 were screened using Abalone Bio’s proprietary Functional Antibody Screening Technology (FAST) system, which is described in more detail in US 2021 / 0198806 A1, the contents of which are incorporated herein by reference for all purposes. Briefly, the human CB2 receptor was expressed on the yeast cell membrane and linked to the pheromone response pathway, which induces the expression of HIS3 upon agonist binding in a histidine-deficient medium (H-medium). his3 - It induces the growth of yeast cells. These CB2 receptor-expressing yeasts were transformed with a VHH library encoded in a plasmid, which induces the pericocytic expression of each VHH linked to the cell membrane via the YPS1 glycophosphatidylinositol fixation domain (as further described in US 2021 / 0198806 A1). After selecting yeast clones growing in H-medium, the antibody-encoding plasmids in each of these clones were DNA sequenced to identify the agonist antibodies, thereby identifying unique agonist clones, which were further tested as presented in subsequent examples. One of the identified VHHs was named AB101 (VHH according to SEQ ID NO. 55, the above Table 3 reference).

[0443] Example 2. Quantification of cAMP levels for measuring agonist activity at CB1 and CB2 receptors.

[0444] CB2 receptor ( for example , human CB2 receptor) and CB1 receptor ( for exampleAgonist antibody activity against the human CB1 receptor was measured using PerkinElmer’s LANCE ultra-cAMP kit (catalog number TRF0262; PerkinElmer, Boston, Massachusetts) according to the manufacturer’s instructions. All assays were performed at room temperature using 384-OptiPlate (catalog number 6007299; PerkinElmer). Briefly, HEK cells (HEK CB2 and HEK CB1) stably transfected with CB2 or CB1 receptors were harvested, pelleted, and resuspended in 1X stimulation buffer (1X Hanks equilibrium salt solution, 5 mM HEPES, 0.5 mM IBMX, 0.1% bovine serum albumin (BSA), pH 7.4, freshly prepared on the day of the experiment). Cells were incubated in 1X stimulation buffer at 37°C, 5% CO2, and humidified air conditions for 1 hour, then transferred to 384-OptiPlate (500 cells per well) and subsequently stimulated for 5 minutes with the appropriately prepared drug / compound in 1X stimulation buffer and forskolin (final concentration 2 μM). Subsequently, cells were lysed by adding Eu-cAMP tracer and Ulight anti-cAMP detection reagent (both 4X solutions, freshly prepared under low light conditions in the lysis buffer provided with the kit). The reaction mixture was incubated at room temperature under low light for 1 hour. The plates were then read on an Enspire plate reader (PerkinElmer) in TR FRET mode (340 nm excitation, 665 nm / 615 nm emission). These measurements were used to determine EC50 values ​​for CB2 receptor activity. The EC50 value of AB101 was found to be 56 nM for human CB2 and 30 nM for mouse CB2, and no activity was observed for human CB1. Fig. 1 ).

[0445] Fig. 1aAs presented in [figure], 1 μM AB101 reduced forskolin-induced cAMP levels in HEK293 cells overexpressing human CB2 and non-overexpressing CB1, whereas 1 μM CP55940, a small molecule agonist potently activating CB1 and CB2, reduced cAMP in both cell types. Data were normalized for forskolin-stimulated and non-stimulated responses. Co-treatment with the CB2-specific inverse agonist SR144528 eliminated the effects of CP55940 and AB101. The 1 μM control anti-GFP antibody had no effect on cAMP. ****: P<0.0001 compared to untreated control; **: P=0.002 compared to untreated control; (All comparisons: one-way ANOVA). Fig. 1b As shown in [figure], AB101 is a potent agonist for human and mouse CB2. Normalized cAMP levels (n=3) after treatment with AB101 of cells overexpressing human (gray squares) or mouse (black circles) CB2. EC50: hsCB2: 56 nM; mmCB2: 30 nM. All values ​​presented are mean ± SEM, and n=3 to 4.

[0446] Example 3A. Paclitaxel model of chemotherapy-induced peripheral neuropathy (CIPN).

[0447] 4 mg / kg paclitaxel ('CIPN') or Cremophor vehicle control ('Healthy') was injected intraperitoneally (ip) into C57BL / 6 mice on days 1, 3, 5, and 7. On day 16, when allodynia was established and stable, ABt140 (25 mg / kg ip) or vehicle was injected. Mechanical allodynia was evaluated by measuring the avoidance threshold (g) twice for each paw using an electronic von Frey sensory system. Cold allodynia was assessed on paws responsive to acetone stimulation ( in other words The reaction time(s) taken to lift, lick, bite, or shake the paw was evaluated by measuring it three times. The data Fig. 2a (Mechanical) and Fig. 2bPresented in (Cold). 6 female mice (N=6). Data are mean ± SEM. BL: Baseline before paclitaxel administration. PacBL: Baseline after paclitaxel administration. *** P <0.001, CIPN+antibody versus CIPN+veh, statistics were analyzed using Bonferroni post-hoc test after one-way ANOVA.

[0448] also, Fig. 3a As shown in [Figure], ABt140 (SEQ No. 94) acts as an agonist for human CB2. cAMP levels (n=2) after treating cells overexpressing human CB2 with ABt140 or CP55940 (CB1 / CB2 small molecule agonist). EC50: ABt140 (square) 30 nM; CP55940 (circle) 9.5 nM.

[0449] Example 3B. Paclitaxel model of chemotherapy-induced peripheral neuropathy (CIPN).

[0450] ABt269 is an anti-GFP VHH-Fc antibody used as a control in some of the following experiments and has the following sequence.

[0451] QVQLVESGGALVQPGGSLRLSCAASGFPVNRYSMRWYRQAPGKEREWVAGMSSAGDRSSYEDSVKGRFTISRDDARNTVYLQMNSLKPEDTAVYYCNVNVGFEYWGQGTQVTVSSGGGGSGGGGSGGGGSEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKDTLMISRTPEVTCVVVDV SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRD ELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 204)

[0452] C57BL / 6 mice were injected intraperitoneally (ip) with 4 mg / kg of paclitaxel ('CIPN') or a Cremophor vehicle control ('Healthy') on days 1, 3, 5, and 7. When allodynia was established and stable on day 16, ABt281 (5 or 25 mg / kg ip), the antibody control ABt269 (25 mg / kg ip), or the vehicle (PBS) were injected. Mechanical allodynia was evaluated by measuring the avoidance threshold (g) twice for each paw using an electronic von Frey sensory system. Cold allodynia was assessed on paws responsive to acetone stimulation ( in other words The reaction time(s) taken to lift, lick, bite, or shake the paw was evaluated by measuring it three times. The data Fig. 8a (Mechanical) and do 8b Presented in (Cold). 6 female mice (N=6). Data are mean ± SEM. BL: Baseline before paclitaxel administration. PacBL: Baseline after paclitaxel administration. *** P <.001, CIPN + antibody vs. CIPN + control (ABt269). Statistical analysis was performed using Bonferroni post-hoc test following one-way ANOVA.

[0453] Figs. 9a and 9b This shows the same experiment performed with CB2 knockout mice. In CB2 knockout mice, there is no difference between the CIPN+ antibody (ABt281) and the CIPN+ control (ABt269), which confirms the requirement of the CB2 receptor for antibody specificity and antibody activity.

[0454] Fig. 10a and Fig. 10b In this study, either the antibody or the control was administered daily for 5 days, and mechanical allodynia and cold allodynia were evaluated 2 hours after daily administration. Superior ABt281 activity was demonstrated at a lower dose of ABt281 (5 mg / kg) compared to a higher dose (25 mg / kg).

[0455] Each group was treated with the CB2-specific agonist AM1710 24 hours after administration of ABt281 or PBS. In the PBS group, as expected, the CB2 agonist activity induced by AM1710 increased the mechanical threshold ( Fig. 11a ) and reaction time reduction( Fig. 11b ...did result in improved efficacy. In contrast, AM1710 treatment did not further improve the response in mice treated with ABt281, indicating the absence of free CB2 receptors for AM1710 activation and an increased response to CB2 action. This experiment provides further evidence that antibody-mediated CB2 depletion occurs upon ABt281 treatment. AM1710 is (3-(1,1-dimethyl-heptyl)-1-hydroxy-9-methoxy-benzo(c)cromen-6-one) and is a cannabilactone cannabinoid receptor type 2 (CB2) agonist (reference [Khanolkar et al. [ , J. Med. Chem. 2007 Dec 27;50(26):6493-500] see reference).

[0456] Example 4. Reduction of IL-6 and IL-8 secretion by BEAS2B human airway cells.

[0457] BEAS2B human airway epithelial cells (ATCC CRL-9609) were cultured according to the guidelines, and Fig. 3b and Fig. 3c Treatment was performed with 10 micrograms / mL LPS for 24 hours in the presence of ABt140 at the concentrations indicated or control anti-lysozyme IgG4 (control antibody). IL-6 and IL-8 concentrations in the supernatant were measured using the Invitrogen ELISA kit (EH2IL6). ABt140 [induced] IL-6 secretion by BEAS2B human airway cells ( Fig. 3b ) and IL-8 secretion ( Fig. 3c ) reduced both (n=2). Two-way ANOVA: * p<0.05, *** p=0.0001, **** p<0.0001.

[0458] Example 5. ABt140 improves clinical scores and reduces mortality in an H1N1 (PR8) influenza infection mouse model.

[0459] On Day 0, adult female BALB / c mice were challenged with 38 TCID50 A / PR / 8 / 34(H1N1)(Charles River), and on Days 3 and 5, treated with vehicle (n=3), 25 mg / kg of control antibody IgG4 (n=10), or 25 mg / kg of ABt140 (SEQ No. 94) (n=10). The results were FIGS. 4a to 4c It is presented in.

[0460] Fig. 4a This shows the area under the curve (AUC) of clinical scores (1 to 5) for coat condition, posture, respiration, motility, and eye secretions / eye closure over 7 days. **** p<0.0001, ** p=0.0018. ABt140 improved the clinical scores of mice.

[0461] Fig. 4b This shows the 7-day survival rate of mice challenged with 38 TCID50 A / PR / 8 / 34(H1N1). ABt140 increased the 7-day survival rate of mice compared to the vehicle control or antibody control.

[0462] Fig. 4c This shows the viral load. Adult female BALB / c mice were challenged with 89 TCID50 A / PR / 8 / 34 (H1N1) on Day 0, and treated with vehicle (n=3), 25 mg / kg of control antibody IgG4 (n=10), or 25 mg / kg of ABt140 (n=10) at 2 hours and on Day 2. Viral load was determined from lung homogenates. Data are presented as 50% tissue culture infection dose (TCID50) per group (n=10). Data were analyzed by log-rank (Mantel-Cox) test compared to the control antibody. ABt140 treatment did not reduce viral titers.

[0463] Example 6. Stability mutation and test.

[0464] Several mutations were tested to improve manufacturability, reduce immunogenicity, and enhance stability. The M5V mutation is considered beneficial because it eliminates methionine, a potential side chain prone to oxidation. The R39Q mutation was selected because Q is conserved at this position in humans, and the original R is not found in the majority of VHH sequences. I58Y was selected because Y is more conserved at this position and was found in the original AB101 primordial clone during screening. Finally, D65G was selected due to greater conservation in known VHH sequences. Combinations of these four mutations were tested to evaluate manufacturability and VHH activity.

[0465] Using a VHH clone with a pET-28b expression vector (Sigma-Aldrich 69865) modified to include a pelB secretory signal E. coliThe vector was expressed in the peripheral cytoplasm. The vector was transformed into BL21(DE3) cells (Sigma-Aldrich 70235) according to the manufacturer's instructions. A single colony from the transformation was grown in LB+kanamycin until the OD600 reached 0.4 to 0.6, induced with IPTG to 1 mM, and then grown overnight at 18°C. After 16 hours of induction, the culture was centrifuged at 4750 rpm for 20 minutes at 4°C, and the supernatant was discarded. The pellet was washed with 200 mM Tris-HCl and centrifuged at 4750 rpm. The supernatant was discarded, and the pellet was resuspended in 200 mM Tris-HCl (pH 7.4) containing 20% ​​sucrose, 500 µg / mL lysozyme, and 1 mM PMSF at 25°C for 15 minutes. Three volumes of water were added, and the solution was shaken at 4 °C for 15 minutes. The solution was centrifuged at 4750 rpm for 20 minutes, and the supernatant was transferred to a small flask for antibody purification. The obtained VHH antibodies were purified using HisPur Cobalt Resin (ThermoFisher 89964) according to the manufacturer's recommendations for the purification of his-tagged proteins using a gravity flow column. The obtained antibodies were tested for thermal stability using the UNCLE system (Unchained Labs). 9 µL of each antibody was loaded into Uni cuvettes in three replicates. Temperature scans were performed from 25 °C to 95 °C at a scan rate of 0.3 °C per minute. Tm values ​​were calculated using the centroidal mean assay.

[0466]

[0467] Example 7. CB2 activation reduces LPS or IL-4-induced inflammatory responses in macrophages.

[0468] Serum-starved RAW264.7 mouse macrophages (ATCC TIB-71) were cultured and exposed to HU-308 (Onternaves), ABt281, ABt285, or ABt269 (1 uM and 100 nM) for 6 hours, followed by LPS (100 ng / ml; Fig. 5a) or IL-4(10ng / ml; Fig. 5b They were exposed to ) for 12 hours. M1 and M2 macrophage gene expression was measured using RTPCR. The CB2-specific agonist (HU-308) and CB2-specific agonist antibodies (ABt281 and ABt285) significantly reduced the LPS or IL-4-induced inflammatory response compared to the buffered control, LPS alone, or IL-4 alone.

[0469] Example 8. CB2 activation reduces IL-6 secretion by RAW264.7 mouse macrophages.

[0470] RAW264.7 cells were serum-starved for 4 hours and exposed to 1 µg / ml LPS for 12 hours in the presence of a CB2 agonist (CP55,940), a CB2-specific agonist antibody (ABt281 or ABt285), or the homologous control ABt269. IL-6 concentrations in the supernatant were measured using the Perkin Elmer IL-6 LANCE Ultra TR-FRET Kit (TRF1223C). CP-55,940, ABt281, and ABt269 reduced IL-6 secretion by RAW264.7 mouse macrophages in a dose-dependent manner. Fig. 6 ).

[0471] Example 9. Of the CB2 receptor In a test tube Activation inhibits cAMP signaling in RAW264.7 mouse macrophages.

[0472] RAW264.7 cells were cultured and treated with the cAMP activator NKH477 to stimulate cAMP production. Subsequently, cells were treated with the CB2 agonist HU308 or CB2-specific agonist antibodies (ABt281, ABt285) or a homozygous control (ABt269) ( Fig. 7The efficacy of CB2-mediated Gi-cAMP signaling was measured using the cAMP-Glo ​​assay (Promega), where luminescence is inversely proportional to cAMP concentration. The application of HU308, ABt281, and ABt285 reduced NKH477-stimulated cAMP production. The EC50 values ​​for HU308, ABt281, and ABt285 were 32 nM, 4.5 nM, and 1.5 nM, respectively.

[0473] Example 10. CB2 agonist antibody reduces fibrotic and inflammatory responses in a precision-cut human liver section model.

[0474] The human liver section model (hPCLS) preserves all cell types within its natural structure, uniquely capturing endogenous cell-to-cell interactions. Using PCLS obtained from de-identified resection samples from two damaged human males, the activity of ABt281 and ABt285 (100 nM and 1 μM), ABt269 isomorphic control (1 μM), Alk5i (positive control), or PBS (vehicle) was tested after 24 and 48 hours of incubation. Alk5i is a TGF-beta type 1 receptor kinase (ALK5) inhibitor (CAS 446859-33-2).

[0475] Human precise-cut liver sections (hPCLS) were generated from the waste residue of surgically resected human livers. Normal-looking resection margins surrounding the HCC were selected for hPCLS generation. The ischemic time from hepatectomy to PCLS generation was 3 to 4 hours. Cores with a diameter of 8 mm were generated, and intact liver sections (thickness 200 μm) were collected from them from a buffer tray using a soft-bristle brush and transferred to 6-well tissue culture plates (3 sections per well) filled with warm William's E GlutaMAX medium at 37 °C supplemented with 25 mM glucose and 50 μg / ml gentamicin. The sections were pre-incubated for 24 hours to equilibrate the tissues. After pre-incubation, fresh medium containing ABt281 (100 nM, 1 μM), ABt285 (100 nM, 1 μM), ABt269 isomorphic control (1 μM), Alk5i (10 μM), or PBS was added for an additional 24 and 48 hours.

[0476] To evaluate efficacy, the secretion of collagen 1a1 (Col1a1) into the medium at the end of drug treatment under each condition was evaluated by ELISA ( Fig. 12a At 48 hours, CB2, agonist antibodies, and Alk5i all significantly reduced Col1a1 compared to the PBS control group. The ABt269 isomorphic control group was not significantly different from the PBS control group.

[0477] In addition, fragments were used for total mRNA extraction at the end of drug treatment for each condition. Collagen 1a1 (COL1A1, Fig. 12b) , alpha smooth muscle actin (ACTA2, Fig. 12c ), Interleukin 6 (IL6, Fig. 12d ), Interleukin 10 (IL10, Fig. 12e ) and tumor necrosis factor alpha (TNFα, Fig. 12fmRNA levels of ) were quantified using RT-qPCR. For COL1A1, ACTA2, IL6, and IL10, treatment with ABt281, ABt285, or the positive control Alk5i resulted in a statistically significant reduction of these inflammatory or fibrotic factors, whereas no significant changes were observed with treatment with ABt269 (homologous control antibody) or PBS (vehicle, negative control). For NFα, treatment with 1 mm of ABt281 or ABt285 led to a reduction of this highly inflammatory factor, whereas no significant changes were observed in the other groups.

[0478] Example 11. Of the CB2 receptor In a test tube Activation activates pERK1 / 2 signaling in RAW264.7 mouse macrophages.

[0479] RAW264.7 cells were cultured and serum-starved for 4 hours, then exposed overnight to the CB2-specific agonist HU-308, CB2-specific agonist antibodies (ABt281 or ABt285) or a homozygous control (ABt269), and the pERK activator (PMA). CB2-mediated pERK1 / 2 signaling efficacy was measured using the TR-FRET pERK1 / 2 Kit (LANCE Ultra pERK1 / 2 Kit, Perking Elmer, TRF4000). Application of PMA, HU308, ABt281, and ABt285 increased TR-FRET signaling in a dose-dependent manner, whereas no ERK phosphorylation was observed in the homozygous control ABt269. EC1D for HU-308, ABt281, ABt285, and PMA 50 The values ​​were 53 nM, 98 nM, 159 nM, and 25 nM, respectively. Fig. 13a This experiment was repeated to obtain EC50 values ​​of 120 nM, 95 nM, 99 nM, and 109 nM for HU-308, ABt281, ABt285, and PMA, respectively ( Fig. 13bThis demonstrates consistent CB2 receptor-mediated activation of pERK1 / 2 signaling in RAW264.7 mouse macrophages.

[0480] Example 12. Paclitaxel model of chemotherapy-induced peripheral neuropathy (CIPN).

[0481] C57BL / 6 mice were injected intraperitoneally (ip) with 4 mg / kg of paclitaxel (to induce CIPN) or Cremophor vehicle control on days 0, 2, 4, and 6. When allodynia was established and stable on day 15, ABt281 (5 mg / kg ip), ABt285 (5 or 25 mg / kg ip), ABt140 (5 or 25 mg / kg ip), or vehicle (PBS) were injected. The same antibody doses were re-injected on days 20 and 25. Mechanical allodynia was evaluated by measuring the avoidance threshold (g) twice for each paw using an electronic von Frey sensory system. Cold allodynia was assessed on paws responsive to acetone stimulation ( in other words The reaction time(s) taken to lift, lick, bite, or shake the paw was evaluated by measuring it three times. The data Fig. 15a (Mechanical) and Fig. 15b Presented in (Cold), the study involved 6 mice (N=6), and data are ± SEM (B: Baseline before paclitaxel administration; PAC: Baseline after paclitaxel administration; mechanical and cold stimuli were applied on Day 0, Day 4, Day 7, Day 11, and Day 15, and after antibody treatment FIGS. 15a to 15b (Evaluated at the time indicated).

[0482] Example 13. A CB2 agonist antibody recruits B-arrestin 2 in RAW264.7 mouse macrophages.

[0483] RAW264.7 cells were cultured and transfected with B-Arr2 plasmid (20 ng, Revvity, PWTBARR2) and pcDNA3 plasmid (130 ng) via Lipofectamine 2000. Transfected cells were treated with the CB2-specific agonist HU308, CB2 agonist antibodies (ABt0281, ABt0285), or a homozygous control (ABt269). CB2-mediated B-Arr2 mobilization was measured using the TR-FRET B-Arr2 mobilization kit (HTRF B-Arr2 mobilization kit, Revvity, 62BDBARR2PEB). Application of HU308 or CB2 agonist antibodies increases the TR-FRET signal.

[0484] Example 14. Functional maturation of CB2 antibody.

[0485] Antibodies that functionally activate CB2 more potently than ABt285 were screened using Abalone Bio’s proprietary Functional Antibody Screening Technology (FAST) system, which is described in more detail in US 2021 / 0198806 A1, the contents of which are incorporated herein by reference for all purposes.

[0486] To further improve antibody stability, a library was designed based on the ABt251 sequence (SEQN 74). Each library member in this experiment possesses the ABt251 sequence, except for a variant in which one of two amino acids is used at each of the six toggle positions, one of the six CDR1 (SEQN 1) positions is randomized, and two of the sixteen CDR3 (SEQN 3) positions are randomized. The six toggle positions are IMGT position 15 (A or P), position 52 (F or W), position 54 (A or S), position 55 (A or G), position 103 (S or W), and position 118 (S or W). Mutated positions within CDR1 (SEQN 1) are IMGT positions 27, 28, 29, 31, 32, or 33 ( in other words, positions 2 to 4 and 6 to 7 of SEQ ID NO. 1. All CDR3 (SEQ ID NO. 3) positions are included in the randomization strategy as described above. In the randomized positions within CDR1 and CDR3, all amino acids except C and M are allowed. The exception is IMGT position 31, where the parent Abt251 sequence (SEQ ID NO. 74) is M. Therefore, in the randomization of IMGT position 31, M is allowed and only C is excluded.

[0487] Briefly, the variable region of the library described above was transplanted onto the ABt285 backbone. The human CB2 receptor was expressed on the yeast cell membrane and linked to the yeast growth pathway. CB2 receptor-expressing yeast was transformed into the VHH library of ABt285 variants. Stronger variants (with superior titers and / or efficacy) were induced to grow under conditions different from those of ABt285. Agent antibody clones were identified by DNA sequencing of antibody-coding plasmids within the selected clones. These clones were validated and scored based on their growth characteristics in yeast, and further tests were conducted as presented in subsequent examples. Table 7 ABt285 variants identified from FAST functional maturation have growth characteristics ( for example It shows how it was scored based on the size and / or number of colonies. The parent (ABt285) growth score was defined as 1, and the negative control growth score as 0.

[0488]

[0489]

[0490] Example 15. A functionally matured CB2 antibody has enhanced efficacy and potency.

[0491] by CB2 receptor agonist antibodies In a test tubeActivation inhibits cAMP signaling in RAW264.7 mouse macrophages and THP-1 human monocyte cells differentiated into macrophages. RAW264.7 cells are cultured with the CB2 agonist HU308 (Onternavez) or the CB2 positive control agonist antibody (ABt285, SEQ ID NO. 203), the isomorphic control (ABt269, SEQ ID NO. 204), or ABhit231-272 ( Table 7 THP-1 cells were treated with (see reference). Prior to treatment with the CB2 agonist, THP-1 cells were differentiated into macrophages by treating them with 200 nM phorbol 12-myristate 13-acetate (PMA). ABhit231-272 represents the apex functional maturation CB2 agonist antibody (Example 14). For 2 to 3 concentrations of the antibody, CB2-mediated Gi-cAMP signaling efficacy was measured using the cAMP-Glo ​​assay (Promega, V1501), where luminescence is inversely proportional to the cAMP concentration ( Fig. 16 The application of HU308 or ABt285 increased the luminescence signal in a dose-dependent manner. A low response was observed with the negative control antibody ABt269. Several ABhit antibodies exhibited dose-dependent changes in luminescence consistent with CB2 action ( Fig. 17 , Tables 8 and 9 ).

[0492]

[0493]

[0494] Example 16. A CB2 agonist antibody that has undergone functional maturation recruits B-arrestin 2 in RAW264.7 mouse macrophages.

[0495] RAW264.7 and THP-1 cells were cultured and transfected with B-Arr2 plasmid (20 ng, Revvity, PWTBARR2) and pcDNA3 plasmid (130 ng) via Lipofectamine 2000. THP-1 cells were differentiated into macrophages by treating them with 200 nM Forbol 12-Myristate 13-Acetate (PMA) prior to CB2 agonist treatment. Transfected cells were treated with the CB2-specific agonist HU308, a CB2 agonist antibody (ABt0285), or a homozygous control (ABt269). CB2-mediated B-Arr2 mobilization was measured using the TR-FRET B-Arr2 mobilization kit (HTRF B-Arr2 mobilization kit, Revvity, 62BDBARR2PEB). Application of HU308 or the CB2 agonist antibody increased the TR-FRET signal in a dose-dependent manner ( Fig. 17 , Tables 8 and 9 ).

[0496] Example 17. Analysis of allowable mutations in ABt285 CDR1 and CDR3.

[0497] The sequences of the enriched antibodies were analyzed over the functional maturation rounds, and the amino acid usage frequency at each position was calculated. To quantify how close a given site is to fixation, the Shannon entropy H = -∑(pi·log2(pi)) was calculated. A cutoff of 0.2 for fixed amino acids was determined based on sequencing noise identified at the fixed library positions (CDR1, positions 1 and 5) and small variations in the selection process. Four positions are fixed: CDR1-position 6 is S, CDR3-position 4 is I, CDR3-position 5 is K, and CDR3-position 13 is I. Five positions have entropy exceeding the cutoff and are one-predominant, allowing for two or three amino acids: CDR1-Position 2 is S / Q, CDR3-Position 7 is G / A, CDR3-Position 9 is G / K, CDR3-Position 11 is F / L, and CDR3-Position 14 is K / N. The results of this analysis Table 10 and Table 11It is provided in.

[0498]

[0499] Table 11 . Amino acid frequency at each CDR3 position.

[0500]

[0501] Table 11 (continued).

[0502]

[0503] Example 18. Production of a CB2 antibody with increased functional activity.

[0504] Historically, antibody discovery and design have relied on labor-intensive biopanning techniques, chance-dependent screening libraries, or complex rational manipulation approaches. Even when effective, these methods often proved to be time-consuming, costly, and limited in scope, sometimes failing to find any hits for difficult targets. The recent emergence of Protein Language Models (PLMs)—models based on Large Language Models (LLMs) originally developed for natural language—has brought about a paradigm shift and revolutionized the way antibody sequences are explored, manipulated, and generated. Trained on vast datasets of antibody sequences and associated characteristics such as binding, stability, and immunogenicity, PLMs now offer an attractive prospect of accelerating antibody discovery and design with unprecedented speed, precision, and creativity.

[0505] PLM is an antibody that, beyond specifically binding to a target, also modulates the target's signaling activity for a therapeutic effect. in other wordsIt has not previously been applied to the problem of generating 'functional antibodies'. PLMs are primarily trained to predict the next amino acid (AA) given the sequence of an AA, or to fill in a masked set of AAs within a larger sequence. By training with a large corpus of observed sequences, PLMs can learn sequence-structure relationships. Given functional information regarding a sequence, a PLM pre-trained using language modeling tasks on a large sequence corpus can be fine-tuned to classify the sequence as likely active or inactive. A key obstacle for PLMs in the discovery and ultimate design of functional antibodies is the lack of large-scale, high-quality functional datasets.

[0506] Abalone Bio's Functional Antibody Selection Technology (FAST) platform can generate large-scale, high-quality functional datasets of antibody sequence-function relationships for targets such as GPCRs. Data generated from an agonist antibody selection campaign for the receptor CB2 were combined. Pre-trained antibody PLMs were fine-tuned to become binary classifiers of antibody functionality using a 10-fold cross-validation subset of this data. Test set sequences not presented in the models were scored using the trained models. Using an ensemble of these 10 models, novel sequences generated by modifying the seed sequence—the highest-scoring sequence in the training set—were scored. The novel sequences derived from FAST selection, high-entropy regions and amino acids, in other wordsAfter selection, variants were generated by identifying sites exhibiting mutations from the starting sequence. Based on the identified sites and residues, all possible variants of the seed sequence were scored using a model ensemble. All sequences with a mean log odds ratio (logit) greater than 2.5 were clustered into 256 clusters based on sequence identity using the k-means algorithm. To ensure sequence diversity, the top four sequences from each cluster were selected as candidate sequences for experimental verification. Since the amount of sequences with more edits is greater than the amount of sequences with fewer edits, sequences with fewer than six edits were largely absent from the candidate set; therefore, additional sequence sets were sampled from this edit distance radius. A complete candidate set of novel sequences, along with negative controls (low logit scores), was synthesized and evaluated using the FAST platform. Among the candidate set, multiple sequences exhibiting agonist activity in the CB2 FAST yeast strain were identified, demonstrating a higher hit rate (3.5%) than that of functionally mature variants (0.18%). Fig. 19 ). In addition, the candidate set included antibody sequences that exhibited greater activity than the seed sequence (Fig. 20 ).

[0507] Example 19. A CB2 agonist antibody produced by GenAI induces growth in yeast expressing human CB2.

[0508] Antibodies functionally activating CB2 were screened using Abalone Bio’s proprietary Functional Antibody Screening Technology (FAST) system, which is described in more detail in US 2021 / 0198806 A1 and is incorporated herein by reference for all purposes. Briefly, sequences generated by genAI were selected, and the coding regions were synthesized and cloned into the same backbone as ABhit_0231. The human CB2 receptor was expressed on the yeast cell membrane and ligated to the yeast growth pathway. CB2 receptor-expressing yeast was transformed with the genAI VHH library. VHHs with agonist activity were selected under different conditions. The agonists induced growth under selective conditions. Stronger agonists (higher titers and / or efficacy) induced growth under conditions different from those of ABhit_0231. Agonist antibody clones were identified by DNA sequencing of antibody-coding plasmids within the selected clones. These clones were validated based on growth characteristics in yeast. Table 12 is the growth characteristics of genAI variants identified from FAST ( for example It shows how it was scored based on the size and / or number of colonies. The seed (ABhit_0231) growth score was defined as 5 in Example 14, the ABt285 growth score as 1, and the negative control growth score as 0.

[0509]

Claims

Claim 1 A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD comprises a complementarity determining region (CDR) 1 comprising the amino acid sequence of SEQ ID NO. 1, a CDR2 comprising the amino acid sequence of SEQ ID NO. 2, and a CDR3 comprising the amino acid sequence of SEQ ID NO. 3, wherein the CDR1, CDR2, and CDR3 are defined according to IMGT. Claim 2 A CB2 antibody comprising an ISVD that specifically binds to CB2, wherein the ISVD comprises CDR1, CDR2, and CDR3 of the ISVD having the amino acid sequence of SEQ ID NO.

201. Claim 3 A CB2 antibody according to claim 1 or 2, wherein the ISVD comprises one or more amino acid residues selected from the group consisting of V5, Q39, Y58, G65, R71, A74 and L78 in one or more framework regions (FR), and the amino acid numbering is according to Kabat. Claim 4 In paragraph 3, the ISVD is a CB2 antibody comprising amino acid residues V5, Q39, Y58, and G65 in one or more FRs. Claim 5 A CB2 antibody according to any one of claims 1 to 4, wherein the ISVD comprises: FR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs 4 to 15; FR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs 16 to 30; FR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs 31 to 48 and 200; and FR4 having an amino acid sequence selected from the group consisting of SEQ ID NOs 49 to 54. Claim 6 In claim 5, the ISVD comprises (i) FR1 comprising the amino acid sequence of SEQ ID NO. 14, FR2 comprising the amino acid sequence of SEQ ID NO. 29, FR3 comprising the amino acid sequence of SEQ ID NO. 43, and FR4 comprising the amino acid sequence of SEQ ID NO. 54; (ii) FR1 comprising the amino acid sequence of SEQ ID NO. 15, FR2 comprising the amino acid sequence of SEQ ID NO. 23, FR3 comprising the amino acid sequence of SEQ ID NO. 45, and FR4 comprising the amino acid sequence of SEQ ID NO. 54; or (iii) FR1 comprising the amino acid sequence of SEQ ID NO. 4, FR2 comprising the amino acid sequence of SEQ ID NO. 16, FR3 comprising the amino acid sequence of SEQ ID NO. 200, and FR4 comprising the amino acid sequence of SEQ ID NO. 49, a CB2 antibody. Claim 7 A CB2 antibody according to any one of claims 1 to 6, wherein the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs 55 to 77 and 201, or a variant thereof having at least about 85% sequence identity with any one of SEQ ID NOs 55 to 77 and 201. Claim 8 In claim 7, the ISVD is a CB2 antibody comprising the amino acid sequence of SEQ ID NO. 201, 70, or 74. Claim 9 A CB2 antibody comprising a polypeptide chain comprising, in any one of claims 1 to 8, an amino acid sequence selected from the group consisting of SEQ ID NOs 78 to 95 and 202 to 203, or a variant thereof having at least about 85% sequence identity with any one of the amino acid sequences SEQ ID NOs 78 to 95 and 202 to 203. Claim 10 In claim 9, a CB2 antibody comprising a polypeptide chain comprising the amino acid sequence of SEQ ID NO.

203. Claim 11 A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD is i) a complementarity determining region (CDR) 1 having the amino acid formula G-X1-X2-X3-SI-X4-X5, wherein X1 is selected from D, S, or Q; X2 is selected from G, H, I, or K; X3 is selected from D, E, F, G, H, I, K, N, Q, R, S, T, V, W, or Y; X4 is selected from G, K, M, R, or Y; and X5 is selected from A or G, CDR1; and ii) a CDR3 having the amino acid formula C1-C2-C3-IK-C4-C5-C6-C7-C8-C9-C10-I-C11-C12-C13, wherein C1 is selected from F, H, I, K, L, R, S, T, V, or Y; C2 is selected from A, E, F, G, I, L, Q, R, or V; C3 is selected from A, F, G, I, K, L, N, or V; C4 is selected from D, F, N, T, or Y; C5 is selected from A, G, or R; C6 is selected from R, S, or W; C7 is selected from D, G, K, or R; C8 is selected from D, I, L, Q, R, S, or T; C9 is selected from F, L, or T; and C10 is selected from any of D, E, H, or Q; A CB2 antibody comprising CDR3, wherein C11 is selected from K or N; C12 is selected from A, F, G, H, I, K, L, N, P, R, S, V, T or W; and C13 is selected from A, D, F, G, H, T or Y. Claim 12 In paragraph 11, CDR2 is the CB2 antibody of sequence number 2. Claim 13 A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD comprises a CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs 248 to 288, a CDR2 having an amino acid sequence of SEQ ID NO. 2, and a CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs 289 to 329, wherein the CDR1, CDR2, and CDR3 are defined according to IMGT. Claim 14 A CB2 antibody according to any one of claims 11 to 13, wherein the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs 205 to 212, 214 to 229 and 231 to 247, or a variant thereof having at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs 205 to 212, 214 to 229 and 231 to 247. Claim 15 A CB2 antibody according to claim 11, wherein the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NO 205, SEQ ID NO 212, SEQ ID NO 215, SEQ ID NO 217, SEQ ID NO 218, SEQ ID NO 222, SEQ ID NO 225, SEQ ID NO 226, SEQ ID NO 227, SEQ ID NO 229, SEQ ID NO 240 and SEQ ID NO 241, or a variant thereof having at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NO 205, SEQ ID NO 212, SEQ ID NO 215, SEQ ID NO 217, SEQ ID NO 218, SEQ ID NO 222, SEQ ID NO 225, SEQ ID NO 226, SEQ ID NO 227, SEQ ID NO 229, SEQ ID NO 240 and SEQ ID NO 241. Claim 16 A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD comprises a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; and C6 is selected from Y, A, or F, and the CDR3 is defined according to IMGT. Claim 17 In claim 16, CDR1 is SEQ ID NO. 248 and CDR2 is SEQ ID NO. 2, and said CDR1 and CDR2 are CB2 antibodies defined according to IMGT. Claim 18 A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD comprises a CDR1 having the amino acid sequence of SEQ ID NO. 248, a CDR2 having the amino acid sequence of SEQ ID NO. 2, and a CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs. 345 to 359, wherein the CDR1, CDR2, and CDR3 are defined according to IMGT. Claim 19 A CB2 antibody according to any one of claims 16 to 18, wherein the ISVD comprises an amino acid sequence selected from the group consisting of SEQ ID NOs 330 to 344, or a variant thereof having at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NOs 330 to 344. Claim 20 In claim 16, the said ISVD has at least about 85% sequence identity with an amino acid sequence selected from the group consisting of SEQ ID NO. 330, SEQ ID NO. 331, SEQ ID NO. 332, SEQ ID NO. 333, SEQ ID NO. 334, SEQ ID NO. 335, SEQ ID NO. 336, SEQ ID NO. 337, SEQ ID NO. 338, SEQ ID NO. 339, SEQ ID NO. 340, SEQ ID NO. 341, SEQ ID NO. 342, SEQ ID NO. 343 and SEQ ID NO. 344, or an amino acid sequence selected from the group consisting of SEQ ID NO. 330, SEQ ID NO. 331, SEQ ID NO. 332, SEQ ID NO. 333, SEQ ID NO. 334, SEQ ID NO. 335, SEQ ID NO. 336, SEQ ID NO. 337, SEQ ID NO. 338, SEQ ID NO. 339, SEQ ID NO. 340, SEQ ID NO. 341, SEQ ID NO. 342, SEQ ID NO. 343 and SEQ ID NO.

344. A CB2 antibody containing his variant. Claim 21 A CB2 antibody comprising an immunoglobulin monovariable domain (ISVD) that specifically binds to cannabinoid receptor type 2 (CB2), wherein the ISVD comprises: a CDR1 having the amino acid sequence of SEQ ID NO. 248; a CDR2 having the amino acid sequence of SEQ ID NO. 2; and a CDR3 having the amino acid formula C1-C2-C3-IKYGSG-C4-FDIK-C5-C6, wherein C1 is selected from L, I, V, Y, or F; C2 is selected from F, A, or V; C3 is selected from A, I, or V; C4 is selected from D or L; C5 is selected from G, T, or V; and C6 is selected from Y, A, or F, wherein the CDR1, CDR2, and CDR3 are defined according to IMGT. Claim 22 In claim 21, the ISVD comprises a CB2 antibody comprising: a CDR1 having the amino acid sequence of SEQ ID NO. 248; a CDR2 having the amino acid sequence of SEQ ID NO. 2; and a CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs. 345 to 359. Claim 23 In any one of claims 1 to 22, the ISVD is a CB2 antibody that is the heavy chain variable domain (VHH) of a heavy chain antibody. Claim 24 In paragraph 23, the above ISVD is a CB2 antibody of camel family origin. Claim 25 In paragraph 23, the above ISVD is a CB2 antibody in the form of a chimeric form. Claim 26 In paragraph 23, the above ISVD is a humanized CB2 antibody. Claim 27 In any one of claims 1 to 26, the antibody is a CB2 antibody comprising an Fc region. Claim 28 In claim 27, the antibody is a CB2 antibody comprising the Fc region of IgG1 or IgG4. Claim 29 In claim 28, the antibody is a CB2 antibody comprising a variant IgG4 Fc region exhibiting reduced effector function. Claim 30 In claim 29, the CB2 antibody wherein the Fc region comprises amino acid substitutions F234A and L235A, and the numbering is according to the EU index of Kabat. Claim 31 A CB2 antibody according to claim 27 or 28, wherein the Fc region comprises the amino acid sequence of SEQ ID NO.

130. Claim 32 In paragraph 28, the antibody is a CB2 antibody comprising a variant IgG1 Fc region exhibiting reduced effector function. Claim 33 In claim 32, the CB2 antibody wherein the variant IgG1 Fc region comprises amino acid substitutions L234A, L235A, M252Y, S254T, and T256E, and the residues are numbered according to the EU index. Claim 34 In claim 33, the CB2 antibody wherein the variant IgG1 Fc region comprises the amino acid sequence of SEQ ID NO.

363. Claim 35 In paragraph 32, the CB2 antibody wherein the variant IgG1 Fc region comprises amino acid substitutions L234A, L235A, and P329G, and the residues are numbered according to the EU index. Claim 36 In claim 35, the CB2 antibody wherein the variant IgG1 Fc region comprises the amino acid sequence of SEQ ID NO.

364. Claim 37 In claim 32, the CB2 antibody wherein the variant IgG1 Fc region comprises amino acid substitutions L234A, L235A, M252Y, S254T, T256E and P329G, and the residues are numbered according to the EU index. Claim 38 In claim 37, the CB2 antibody wherein the variant IgG1 Fc region comprises the amino acid sequence of SEQ ID NO.

365. Claim 39 In any one of claims 27 to 38, the antibody is a CB2 antibody comprising a hinge region. Claim 40 In claim 39, the hinge region comprises the amino acid sequence of SEQ ID NO. 131, a CB2 antibody. Claim 41 A CB2 antibody according to any one of claims 27 to 40, wherein the ISVD is fused to the Fc region through a peptide linker. Claim 42 In claim 41, the peptide linker comprises the amino acid sequence of SEQ ID NO. 132, a CB2 antibody. Claim 43 A CB2 antibody according to claim 41 or 42, wherein the antibody comprises a polypeptide chain comprising the ISVD, a peptide linker, a hinge region, and an Fc region in the direction from the N-terminus to the C-terminus. Claim 44 In any one of claims 1 to 43, the CB2 is a CB2 antibody in which the CB2 is human, mouse, rat, or cynomolgus monkey CB2. Claim 45 In paragraph 44, the above CB2 is a CB2 antibody, which is human CB2. Claim 46 In any one of claims 1 to 45, the CB2 antibody is a CB2 antibody that is a CB2 agonist. Claim 47 In paragraph 46, the above CB2 antibody is a CB2 antibody that does not act as an agonist against CB1. Claim 48 In paragraph 46 or 47, (i) said ISVD is less than half-maximum effective concentration (EC₀) of 55 nM in HEK293 cells overexpressing human CB2 50 (ii) to reduce forskolin-inducible cyclic adenosine monophosphate (cAMP) levels and / or; or (ii) said ISVD reduces EC levels of less than 30 nM in HEK293 cells overexpressing mouse CB2. 50 A CB2 antibody that reduces forskolin-inducible cyclic adenosine monophosphate (cAMP) levels. Claim 49 In any one of claims 1 to 48, the ISVD is a CB2 antibody that does not specifically bind to cannabinoid receptor type 1 (CB1). Claim 50 In any one of claims 1 to 49, the CB2 antibody is less than 850 nM EC in HEK293 cells overexpressing human CB2 50 CB2 antibody that binds to Claim 51 Isolated nucleic acid encoding the CB2 antibody of any one of claims 1 to 50. Claim 52 An expression vector comprising the nucleic acid of claim 51. Claim 53 A host cell comprising the nucleic acid of claim 51 or the expression vector of claim 52. Claim 54 A method for producing a CB2 antibody, comprising the step of culturing the host cell of claim 53 under conditions in which the CB2 antibody is produced. Claim 55 A method according to claim 54, further comprising the step of recovering the CB2 antibody produced by the host cell. Claim 56 A pharmaceutical composition comprising a CB2 antibody of any one of claims 1 to 50 and a pharmaceutically acceptable carrier. Claim 57 A method of acting as an agonist for CB2 on a cell, comprising the step of contacting the cell with an amount of a CB2 antibody of any one of claims 1 to 50 sufficient to activate CB2 on the cell. Claim 58 In paragraph 57, the above method in vitro A method performed in. Claim 59 In paragraph 57, the above method In vivo Method being performed. Claim 60 A method for treating an individual requiring treatment for a disease or pathological condition, wherein acting as an agonist for CB2 improves the said disease or pathological condition, and the method comprises the step of administering an effective amount of a CB2 antibody according to any one of claims 1 to 50 to said individual. Claim 61 In paragraph 60, the above disease or condition is selected from the group consisting of chemotherapy-induced peripheral neuropathy (CIPN), diabetic peripheral neuropathy, respiratory infection, hepatic fibrosis, cold sensitivity, inflammatory bowel disease, and endometriosis. Claim 62 In paragraph 61, the above disease or condition is chemotherapy-induced peripheral neuropathy (CIPN). Claim 63 In paragraph 62, the above CIPN is induced by the drug Taxol. Claim 64 In paragraph 63, the above taxol drug is paclitaxel, method. Claim 65 A method for treating cancer in an individual requiring treatment for cancer, comprising the step of administering an effective amount of a chemotherapy agent and an effective amount of a CB2 antibody according to any one of claims 1 to 50 to said individual. Claim 66 In paragraph 61, the above disease or condition is a respiratory infection. Claim 67 In paragraph 66, the method wherein the respiratory infection is caused by a virus selected from the group consisting of influenza viruses and coronaviruses. Claim 68 In paragraph 66, the method wherein the respiratory infection is caused by SARS-CoV-2. Claim 69 In paragraph 66, the method wherein the respiratory infection is caused by H1N1 influenza. Claim 70 A method for alleviating cytokine release syndrome in an individual requiring alleviation of cytokine release syndrome, comprising the step of administering an effective amount of a CB2 antibody according to any one of claims 1 to 50 to said individual. Claim 71 In paragraph 70, the method reduces the levels of IL-16 and / or IL-8 secreted by airway cells in the individual. Claim 72 In paragraph 61, the above disease or condition is liver fibrosis. Claim 73 In paragraph 61, the above disease or condition is diabetic peripheral neuropathy. Claim 74 A method in which, in any one of paragraphs 60 to 73, the individual is a human. Claim 75 A kit comprising a CB2 antibody of any one of claims 1 to 50 or the pharmaceutical composition of claim 49, and instructions for use.

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