Autophilic antibodies and method of making the same

Abstract

Antibodies having noncovalent, autophilic properties are disclosed. The autophilic antibodies are derived from antibodies conjugated with an autophilic peptide. Such autophilic antibodies can promote apoptosis of target cells and enhance therapeutic efficacies in the treatment of patients with diseases or disorders responsive to antibody therapy. Compositions containing the antibodies, and methods of making and using the antibodies are also disclosed.

1 . An autophilic antibody comprising an autophilic peptide conjugated to an immunoglobulin component of a non-autophilic antibody. 
     
     
         2 . An autophilic antibody according to  claim 1 , wherein the autophilic peptide comprises a conserved self-binding sequence. 
     
     
         3 . An autophilic antibody according to  claim 1 , wherein the autophilic peptide is a humanized peptide. 
     
     
         4 . An autophilic antibody according to  claim 1 , wherein the autophilic peptide is selected from the group consisting of T15 peptide, T15-scr2 peptide, R24 peptide, and R24-Charged peptide. 
     
     
         5 . An autophilic antibody according to  claim 1 , wherein the autophilic peptide comprises SEQ ID NO.1. 
     
     
         6 . An autophilic antibody according to  claim 1 , wherein the autophilic peptide comprises SEQ ID NO.4. 
     
     
         7 . An autophilic antibody according to  claim 1 , wherein the autophilic peptide comprises SEQ ID NO.5. 
     
     
         8 . An autophilic antibody according to  claim 1 , wherein the autophilic peptide comprises SEQ ID NO.6. 
     
     
         9 . An autophilic antibody according to  claim 1  comprising one or more functional peptides conjugated to the non-autophilic antibody. 
     
     
         10 . An autophilic antibody according to  claim 9 , wherein the one or more functional peptides are conjugated to the non-autophilic antibody at a site that is different from a site that the autophilic peptide is conjugated to the non-autophilic antibody. 
     
     
         11 . An autophilic antibody according to  claim 9 , wherein the one or more functional peptides comprises an MTS peptide. 
     
     
         12 . An autophilic antibody according to  claim 9 , wherein the one or more functional peptides comprises SEQ ID NO.2. 
     
     
         13 . An autophilic antibody according to  claim 9 , wherein the one or more functional peptides comprises SEQ ID NO.7. 
     
     
         14 . An autophilic antibody according to  claim 1 , wherein the non-autophilic antibody is selected from the group consisting of 5D10, S1C5, rituximab, anti-GM2, trastuzumab, anti-caspase antibodies, humanized TEPC-15, humanized R24, and humanized S107. 
     
     
         15 . A method of producing an autophilic antibody according to  claim 1  comprising attaching the autophilic peptide via photoactivation crosslinking to an immunoglobulin component of the antibody. 
     
     
         16 . A method according to  claim 15 , wherein the autophilic peptide is cross-linked to a nucleotide affinity site of the immunoglobulin component. 
     
     
         17 . A method according to  claim 15 , wherein the autophilic peptide is cross-linked to a heterocyclic compound affinity site of the immunoglobulin component. 
     
     
         18 . A method according to  claim 15 , wherein the autophilic peptide is cross-linked to a carbohydrate site of the Fc portion of the immunoglobulin component. 
     
     
         19 . A method according to  claim 15 , wherein the autophilic peptide is conjugated to an amino or sulfhydryl group of the immunoglobulin component. 
     
     
         20 . A method of producing an autophilic antibody according to  claim 1  comprising expressing the autophilic antibody as a fusion protein containing the autophilic peptide sequence and the non-autophilic antibody. 
     
     
         21 . A method of producing an autophilic antibody according to  claim 9  comprising attaching the one or more functional peptides via photoactivation crosslinking to an immunoglobulin component of the antibody. 
     
     
         22 . A method according to  claim 21 , wherein the one or more functional peptides is cross-linked to a nucleotide affinity site of the immunoglobulin component. 
     
     
         23 . A method according to  claim 21 , wherein the one or more functional peptides is cross-linked to a heterocyclic compound affinity site of the immunoglobulin component. 
     
     
         24 . A method according to  claim 21 , wherein the one or more functional peptides is cross-linked to a carbohydrate site of the Fc portion of the immunoglobulin component. 
     
     
         25 . A method according to  claim 21 , wherein the one or more functional peptides is conjugated to an amino or sulfhydryl group of the immunoglobulin component. 
     
     
         26 . A method according to  claim 21 , wherein the one or more functional peptides is conjugated to the immunoglobulin component at a site that is different from a site of conjugation of the autophilic peptide. 
     
     
         27 . A method of producing an autophilic antibody according to  claim 9  comprising expressing the autophilic antibody as a fusion protein containing the autophilic peptide sequence, the one or more functional peptides, and the non-autophilic antibody. 
     
     
         28 . A composition comprising one or more autophilic antibodies according to  claim 1  and a pharmaceutically acceptable carrier. 
     
     
         29 . A composition comprising one or more autophilic antibodies according to  claim 9  and a pharmaceutically acceptable carrier. 
     
     
         30 . A method of formulating an autophilic antibody composition according to  claim 28  to reduce or mitigate dimerization of the autophilic antibody in solution comprising one or more steps selected from the group consisting of: a. Maintaining a hypertonic salt concentration; b. Adding detergents to the composition; and c. Decreasing the isoelectric point of the antibody composition with succinyl anhydride. 
     
     
         31 . A method of formulating an autophilic antibody composition as claimed in  claim 30 , wherein the detergents are SDS or anionic detergents. 
     
     
         32 . A method of formulating an autophilic antibody composition as claimed in  claim 30 , wherein the salt concentration is equal to or greater than 0.5M. 
     
     
         33 . A method of selecting an antigen target in vitro for an autophilic antibody according to  claim 1  comprising administering the autophilic antibody to one or more cell lines expressing antigens, and assaying for apoptosis of the one or more cell lines. 
     
     
         34 . A method of identifying an autophilic antibody according to  claim 1  for use in humans comprising administering the autophilic antibody to SCID or nude mice having human tumor xenografts. 
     
     
         35 . A method of synthesizing a peptide sequence having enhanced autophilic binding properties comprising providing a plurality of peptides, each having one or more amino acid substitutions compared to a template peptide, and comparing self-binding properties of the peptides relative to those of the template peptide. 
     
     
         36 . The method of  claim 35 , wherein the template peptide is selected from the group consisting of T15 peptide, T15-scr2 peptide, R24 peptide, and R24-Charged peptide. 
     
     
         37 . A method of treating a patient suffering from a disease or disorder responsive to antibody therapy comprising administering to the patient one or more autophilic antibodies according to  claim 1 , or a composition comprising one or more autophilic antibodies according to  claim 1  and a pharmaceutically acceptable carrier, in a therapeutically effective amount to alleviate symptoms of the disease or disorder. 
     
     
         38 . The method of  claim 37 , wherein the disease or disorder is a malignancy, an auto-immune disorder, transplantation rejection, Alzheimer's disease, a neuro-degenerative condition, atheroschlerosis, or any other condition responsive to antibody therapy. 
     
     
         39 . The method of  claim 37 , wherein the autophilic antibody is administered in one or more dose amounts substantially identical to amounts used to administer non-autophilic antibodies. 
     
     
         40 . The method of  claim 37 , wherein an initial dose is about 250 mg per day and a later dose is about 100 mg per week. 
     
     
         41 . The method of  claim 37 , wherein a maintenance dose is about 50 mg per week. 
     
     
         42 . A method of potentiating apoptosis, complement fixation, or cell-mediated killing of selected cells in a patient comprising administering to the patient one or more autophilic antibodies according to  claim 1 . 
     
     
         43 . A method according to  claim 42  further comprising administering to the patient a second antibody directed to the autophilic peptide of the autophilic antibody. 
     
     
         44 . A method of restoring autophilic activity to an antibody that has lost its autophilic activity comprising conjugating an autophilic peptide to the antibody. 
     
     
         45 . A method according to  claim 44 , wherein the autophilic peptide is selected from the group consisting of T15 peptide, T15-scr2 peptide, R124 peptide, and 124-Charged peptide. 
     
     
         46 . A method according to  claim 44 , wherein the autophilic peptide is conjugated to the antibody via photoactivation crosslinking to an immunoglobulin component of the antibody. 
     
     
         47 . A method according to  claim 44 , wherein the autophilic peptide is cross-linked to a nucleotide affinity site of the immunoglobulin component. 
     
     
         48 . A method according to  claim 44 , wherein the autophilic peptide is cross-linked to a heterocyclic compound affinity site of the immunoglobulin component. 
     
     
         49 . A method according to  claim 44 , wherein the autophilic peptide is cross-linked to a carbohydrate site of the Fc portion of the immunoglobulin component. 
     
     
         50 . A method according to  claim 44 , wherein the autophilic peptide is conjugated to an amino or sulfhydryl group of the immunoglobulin component. 
     
     
         51 . A method according to  claim 44  comprising expressing the autophilic antibody as a fusion protein containing the autophilic peptide sequence and the antibody. 
     
     
         52 . A peptide comprising SEQ ID NO. 4. 
     
     
         53 . A peptides comprising SEQ ID NO. 5. 
     
     
         54 . A peptide comprising SEQ ID NO. 6. 
     
     
         55 . A peptide comprising SEQ ID NO.7. 
     
     
         56 . An antibody conjugate according to  claim 1  comprising S1C5 antibody conjugated with T15 peptide. 
     
     
         57 . An antibody conjugate according to  claim 1  comprising 5D10 antibody conjugated with T15 peptide. 
     
     
         58 . An antibody conjugate comprising anti-caspase 3 antibody conjugated with MTS peptide. 
     
     
         59 . An antibody conjugate according to  claim 1  comprising anti-CD20 antibody conjugated with T 5 peptide. 
     
     
         60 . An antibody conjugate according to  claim 1  comprising rituximab conjugated with T15 peptide. 
     
     
         61 . An antibody conjugate according to  claim 1  comprising 1F5 conjugated with T15 peptide. 
     
     
         62 . An antibody conjugate according to  claim 1  comprising tositumomab conjugated with T15 peptide. 
     
     
         63 . An antibody conjugate according to  claim 1  comprising anti-GM2 conjugated with T15 peptide. 
     
     
         64 . An antibody conjugate according to  claim 1  comprising a non-human antibody to GM2 glycolipid conjugated with an autophilic peptide. 
     
     
         65 . An antibody conjugate according to  claim 1  comprising Herceptin conjugated with an autophilic peptide.