Targeting agent antibody conjugates and uses thereof

Abstract

Methods, compositions and uses are provided for bispecific antibodies comprising one or more unnatural amino acids. The bispecific antibodies may bind to two or more different receptors, co-receptors, antigens, or cell markers on one or more cells. The bispecific antibodies may be used to treat a disease or condition (e.g., cancer, autoimmune disease, pathogenic infection, inflammatory disease). The bispecific antibodies may be used to modulate (e.g., stimulate or suppress) an immune response.

That which is claimed is: 
     
         1 . A method of treating cancer in a subject in need thereof using immunotherapy, the method comprising: administering to the subject a composition comprising (a) a non-antibody targeting agent linked to (b) an anti-CD3 antibody or fragment thereof comprising a p-acetylphenylalanine (pAcF) amino acid; wherein the targeting agent is site-specifically linked to the antibody or fragment thereof via the pAcF amino acid; and wherein the targeting agent binds to a cancer cell in the subject, and the anti-CD3 antibody or fragment thereof binds to a CD3 of a cytotoxic effector cell in the subject, thereby bringing the cytotoxic effector cell in proximity of the cancer cell to induce cancer cell elimination in the subject. 
     
     
         2 . The method of  claim 1 , wherein the anti-CD3 antibody or fragment thereof comprises an anti-CD3 Fab fragment. 
     
     
         3 . The method of  claim 1 , wherein the anti-CD3 antibody or fragment thereof comprises a constant region and the pAcF amino acid is located within the constant region. 
     
     
         4 . The method of  claim 1 , wherein the targeting agent binds to a membrane protein of the cancer cell. 
     
     
         5 . The method of  claim 1 , wherein the targeting agent is linked to the pAcF amino acid through an oxime ligation. 
     
     
         6 . The method of  claim 1 , wherein the targeting agent is linked to the pAcF amino acid through an amino-oxy group of the targeting agent or a linker attached thereto. 
     
     
         7 . The method of  claim 1 , wherein the targeting agent is site-specifically linked to the anti-CD3 antibody or fragment thereof through a linker providing between about 10 and about 100 angstrom distance between the targeting agent and the anti-CD3 antibody or fragment thereof. 
     
     
         8 . The method of  claim 1 , wherein the cancer cell is selected from a prostate cancer cell, an epithelial cancer cell, a breast cancer cell, a kidney cancer cell, a lung cancer cell, a colon cancer cell, a colorectal cancer cell, a gastric cancer cell, a brain cancer cell, a glioblastoma cell, a pancreatic cancer cell, a myeloid leukemia cell, a cervical cancer cell, a medullary thyroid carcinoma cell, a stromal ovarian cancer cell, an astrocytoma cell, an endometrial cancer cell, a neuroendocrine cancer cell, a gastroenteropancreatic tumor cell, a non-Hodgkin's lymphoma cell, an exocrine pancreatic cancer cell, an Ewing's sarcoma cell, and a skin cancer. 
     
     
         9 . The method of  claim 2 , wherein the anti-CD3 Fab fragment comprises a variable domain and a constant domain, and the constant domain comprises the pAcF amino acid. 
     
     
         10 . The method of  claim 9 , wherein the targeting agent is linked to the pAcF amino acid through an oxime ligation. 
     
     
         11 . A targeting agent antibody conjugate comprising:
 a. a targeting agent that binds to a cell surface protein or marker of a cancer cell, wherein the targeting agent is not an antibody or fragment thereof;   b. an anti-CD3 antibody or fragment thereof having specificity for a CD3 molecule of a cytotoxic effector cell, wherein the anti-CD3 antibody or fragment thereof comprises a p-acetylphenylalanine (pAcF) amino acid; and   c. a linker molecule linking the targeting agent to the pAcF amino acid.   
     
     
         12 . The targeting agent antibody conjugate of  claim 11 , wherein the cell surface protein or marker is a receptor. 
     
     
         13 . The targeting agent antibody conjugate of  claim 11 , wherein the cell surface protein or marker is a membrane protein. 
     
     
         14 . The targeting agent antibody conjugate of  claim 11 , wherein the targeting agent is a synthesized small molecule. 
     
     
         15 . The targeting agent antibody conjugate of  claim 11 , wherein the anti-CD3 antibody or fragment thereof comprises an anti-CD3 Fab fragment. 
     
     
         16 . The targeting agent antibody conjugate of  claim 11 , wherein the anti-CD3 antibody or fragment thereof comprises a constant region and the pAcF amino acid is located within the constant region. 
     
     
         17 . The targeting agent antibody conjugate of  claim 11 , wherein the targeting agent is linked to the pAcF amino acid through an oxime ligation. 
     
     
         18 . The targeting agent antibody conjugate of  claim 11 , wherein the linker provides between about 10 and about 100 angstrom distance between the targeting agent and the anti-CD3 antibody or fragment thereof. 
     
     
         19 . The targeting agent antibody conjugate of  claim 14 , wherein the anti-CD3 antibody or fragment thereof comprises an anti-CD3 Fab fragment comprising a variable domain and a constant domain, and the constant domain comprises the pAcF amino acid. 
     
     
         20 . A method for treating a cancer in a subject in need thereof, the method comprising administering an effective amount of the targeting agent antibody conjugate of  claim 11 .