Carrier molecules

Abstract

The present invention relates generally to carrier molecules derived from one animal or avian species or strains and which are substantially non-immunogenic when exposed to an immune system from a species or strain of another animal or avian creature. More particularly, the present invention provides deimmunized immunointeractive molecules and even more particular deimmunized antibodies for use in diagnostic and therapeutic applications.

1 . An isolated antibody specific for an epitope on human D-dimer which recognizes cross-linked fibrin but not fibrinogen, wherein said antibody is selected from the group consisting of:
 (i) an antibody that comprises a binding site for said D-dimer epitope, wherein one or more amino acid residues in a T-cell epitope present in the variable (v)-domain of said antibody are mutated to eliminate or reduce the ability of peptide fragments of said v-domain to associate with MHC class II molecules; and   (ii) an antibody that comprises complementary determining regions (CDRs) that define a binding site for said D-dimer epitope, interspersed between framework regions that comprise a human antibody light and heavy chain variable region.   
   
   
       2 . The isolated antibody of  claim 1  wherein said antibody is specific for an epitope recognized by anti-fibrin murine monoclonal antibody 3B6. 
   
   
       3 . The isolated antibody of  claim 1 , wherein the antibody comprises an H-chain selected from the group consisting of Hv5 (3B6DIVHv5; SEQ ID NO:1), Hv6 (3B6DIVHv6; SEQ ID NO:2) and Hv7 (3B6DIVHv7; SEQ ID NO:3). 
   
   
       4 . The isolated antibody of  claim 1 , wherein the antibody comprises an L-chain selected from the group consisting of Kv1 (3B6DIVKv1; SEQ ID NO:4), Kv4 (3B6DIVKv4; SEQ ID NO:5) and Kv7 (3B6DIVKv7; SEQ ID NO:6). 
   
   
       5 . The isolated antibody of  claim 1 , wherein said antibody comprises a combination of an H-chain and an L-chain selected from the group consisting of VHv5/VKv1, VHv6/VKv1, VHv7/VKv1, VHv5/VKv7, VHv6/VKv7, VHv6/VKv4, VHv7/VKv4, VHv7/VKv7 and VHv5/VKv4. 
   
   
       6 . The isolated antibody of  claim 1 , wherein said antibody comprises a combination of an H and an L chain v-domain comprising a combination of amino acid sequences selected from the group consisting of SEQ ID NO:1/SEQ ID NO:4, SEQ ID NO:2/SEQ ID NO:4, SEQ ID NO:3/SEQ ID NO:4, SEQ ID NO:1/SEQ ID NO:6, SEQ ID NO:2/SEQ ID No:6, SEQ ID NO:2/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:6, and SEQ ID NO:1/SEQ ID NO:5 or combinations of amino acid sequences having at least 70% sequence identity to the contiguous sequence of one or both amino acid sequences in each of the above listed pairs. 
   
   
       7 . The antibody of  claim 6 , wherein said antibody is an Fab′ fragment. 
   
   
       8 . The antibody of  claim 1 , wherein said antibody is selected by binding to D-dimer that is not passively bound to a substrate. 
   
   
       9 . The antibody of  claim 1 , wherein said antibody is selected by binding to D-dimer in solution. 
   
   
       10 . A deimmunized antibody specific for an epitope on human D-dimer which recognizes cross-linked fibrin but not fibrinogen, wherein said antibody comprises (a) an H-chain selected from the group consisting of Hv5 (3B6DIVHv5; SEQ ID NO:1), Hv6 (3B6DIVHv6; SEQ ID NO:2) and Hv7 (3B6DIVHv7; SEQ ID NO:3), and (b) an L-chain selected from the group consisting of Kv1 (3B6DIVKv1; SEQ ID NO:4), Kv4 (3B6DIVKv4; SEQ ID NO:5) and Kv7 (3B6DIVKv7; SEQ ID NO:6), and further wherein said antibody is selected by binding to D-dimer in solution. 
   
   
       11 . The deimmunized antibody of  claim 10 , wherein said antibody comprises a combination of an H and an L chain v-domain comprising a combination of amino acid sequences encoded by a combination of nucleotide sequences selected from the group consisting of SEQ ID NO:7/SEQ ID NO:10, SEQ ID NO:8/SEQ ID NO:10, SEQ ID NO:9/SEQ ID NO:10, SEQ ID NO:7/SEQ ID NO:12, SEQ ID NO:8/SEQ ID NO:12, SEQ ID NO:8/SEQ ID NO:11, SEQ ID NO:9/SEQ ID NO:11, SEQ ID NO:9/SEQ ID NO:12, and SEQ ID NO:7/SEQ ID NO:11. 
   
   
       12 . The deimmunized antibody of  claim 10 , wherein said antibody comprises a combination of an H and an L chain v-domain comprising a combination of amino acid sequences selected from the group consisting of SEQ ID NO:1/SEQ ID NO:4, SEQ ID NO:2/SEQ ID NO:4, SEQ ID NO:3/SEQ ID NO:4, SEQ ID NO:1/SEQ ID NO:6, SEQ ID NO:2/SEQ ID No:6, SEQ ID NO:2/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:6, and SEQ ID NO:1/SEQ ID NO:5. 
   
   
       13 . The deimmunized antibody of  claim 10 , wherein said antibody is an Fab′ fragment. 
   
   
       14 . An isolated antibody wherein said antibody comprises human constant regions, and variable regions specific for an epitope recognized by anti-fibrin murine monoclonal antibody 3B6. 
   
   
       15 . The antibody of  claim 14 , wherein said antibody comprises a combination of heavy and light chain v-regions from VHv5 and VKv1. 
   
   
       16 . The antibody of  claim 15 , wherein said antibody comprises a heavy chain v-region comprising amino acid residues 26-36, 51-71 and 99-107 of SEQ ID NO:1. 
   
   
       17 . The antibody of  claim 15 , wherein said antibody comprises a light chain v-region comprising amino acid residues 24-34, 50-56 and 89-97 of SEQ ID NO:4. 
   
   
       18 . The antibody of  claim 15 , wherein said antibody comprises a heavy chain v-region comprising amino acid residues 26-36, 51-71 and 99-107 of SEQ ID NO:1 and a light chain v-region comprising amino acid residues 24-34, 50-56 and 89-97 of SEQ ID NO:4. 
   
   
       19 . An isolated antibody that comprises human constant regions specific for an epitope recognized by anti-fibrin murine monoclonal antibody 3B6, and variable regions specific for an epitope recognized by anti-fibrin murine monoclonal antibody 3B6. 
   
   
       20 . A deimmunized antibody specific for an epitope on human D-dimer which recognizes cross-linked fibrin but not fibrinogen, wherein said antibody comprises a combination of an H and an L chain v-domain comprising a combination of amino acid sequences selected from the group consisting of SEQ ID NO:1/SEQ ID NO:4, SEQ ID NO:2/SEQ ID NO:4, SEQ ID NO:3/SEQ ID NO:4, SEQ ID NO:1/SEQ ID NO:6, SEQ ID NO:2/SEQ ID No:6, SEQ ID NO:2/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:6, and SEQ ID NO:1/SEQ ID NO:5, or combinations of amino acid sequences having a conservative amino acid substitution in one or both amino acid sequences in each of the above listed pairs. 
   
   
       21 . A conjugate comprising an imaging tag and an antibody specific for an epitope on human D-dimer which recognizes cross-linked fibrin but not fibrinogen wherein said antibody is selected from the group consisting of:
 (i) an antibody that comprises a binding site for said D-dimer epitope, wherein one or more amino acid residues in a T-cell epitope present in the v-domain of said antibody are mutated to eliminate or reduce the ability of peptide fragments of said v-domain to associate with MHC class II molecules; and   (ii) an antibody that comprises complementary determining regions (CDRs) that define a binding site for said D-dimer epitope interspersed between framework regions that comprise a human antibody light and heavy chain variable region   
   
   
       22 . The conjugate of  claim 21  wherein said antibody is specific for an epitope recognized by anti-fibrin murine monoclonal antibody 3B6. 
   
   
       23 . The conjugate of  claim 21 , wherein said antibody comprises an H-chain selected from the group consisting of Hv5 (3B6DIVHv5; SEQ ID NO:1), Hv6 (3B6DIVHv6; SEQ ID NO:2), and Hv7 (3B6DIVHv7; SEQ ID NO:3). 
   
   
       24 . The conjugate of  claim 21 , wherein said antibody comprises an L-chain selected from the group consisting of Kv1 (3B6DIVKv1; SEQ ID NO:4), Kv4 (3B6DIVKv4; SEQ ID NO:5), and Kv7 (3B6DIVKv7; SEQ ID NO:6). 
   
   
       25 . The conjugate of  claim 21 , wherein said antibody comprises a combination of an H-chain and an L-chain selected from the group consisting of VHv5/VKv1, VHv6/VKv1, VHv7/VKv1, VHv5/VKv7, VHv6/VKv7, VHv6/VKv4, VHv7/VKv4, VHv7/VKv7 and VHv5/VKv4. 
   
   
       26 . The conjugate of  claim 21 , wherein said antibody comprises a combination of an H and an L chain v-domain comprising a combination of amino acid sequences selected from the group consisting of SEQ ID NO:1/SEQ ID NO:4, SEQ ID NO:2/SEQ ID NO:4, SEQ ID NO:3/SEQ ID NO:4, SEQ ID NO:1/SEQ ID NO:6, SEQ ID NO:2/SEQ ID No:6, SEQ ID NO:2/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:5, SEQ ID NO:3/SEQ ID NO:6, and SEQ ID NO:1/SEQ ID NO:5 or combinations of amino acid sequences having at least 70% sequence identity to the contiguous sequence of one or both amino acid sequences in each of the above listed pairs. 
   
   
       27 . The conjugate of  claim 21  wherein said imaging tag is an x-ray, MRI, ultrasound-, CT-, or nuclear medicine-type tag. 
   
   
       28 . The conjugate of  claim 27  wherein the imaging tag is selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       29 . The conjugate of  claim 28  wherein the imaging tag is  99m Tc. 
   
   
       30 . The conjugate of  claim 27 , wherein the imaging tag is a gamma scintigraphy or PET-type tag. 
   
   
       31 . A conjugate comprising a deimmunized antibody according to  claim 10 , and an imaging tag selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       32 . A conjugate comprising a deimmunized antibody according to  claim 11 , and an imaging tag selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       33 . A conjugate comprising a deimmunized antibody according to  claim 12 , and an imaging tag selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       34 . A conjugate comprising a deimmunized antibody according to  claim 13 , and an imaging tag selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       35 . A conjugate comprising an antibody according to  claim 14 , and an imaging tag selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       36 . A conjugate comprising an antibody according to  claim 19 , and an imaging tag selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       37 . A conjugate comprising an antibody according to  claim 20 , and an imaging tag selected from the group consisting of  99m Tc,  18 F,  64 Cu,  67 Ga,  68 Ga,  77 Br,  97 Ru,  111 In,  123 I,  124 I,  131 I and  188 Re. 
   
   
       38 . The conjugate of  claim 31  wherein the imaging tag is  99m Tc. 
   
   
       39 . The conjugate of  claim 32  wherein the imaging tag is  99m Tc. 
   
   
       40 . The conjugate of  claim 33  wherein the imaging tag is  99m Tc. 
   
   
       41 . The conjugate of  claim 34  wherein the imaging tag is  99m Tc. 
   
   
       42 . The conjugate of  claim 35  wherein the imaging tag is  99m Tc. 
   
   
       43 . The conjugate of  claim 36  wherein the imaging tag is  99m Tc. 
   
   
       44 . The conjugate of  claim 37  wherein the imaging tag is  99m Tc. 
   
   
       45 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 21  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       46 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 25  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       47 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 26  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       48 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 31  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       49 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 33  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       50 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 34  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       51 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 38  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       52 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 40  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       53 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 41  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present. 
   
   
       54 . A method for detecting a blood clot in a human patient, said method comprising introducing into said patient a conjugate according to  claim 44  under conditions whereby said conjugate disseminates through the circulatory system, and subjecting said patient to a planar or computer assisted tomographic nuclear medicine scan to visualize a clot if present.