Methods and compositions for inhibiting adam10 biological activities

Abstract

Provided are modified isolated ADAM10 modulating peptides and methods of using the same to modulate ADAM10 biological activities, inhibit ADAM10 biological activities associated with diseases, disorders, or conditions in subjects, including but not limited to decreasing inflammation and inhibiting undesirable cell proliferation. In some embodiments, the modified isolated ADAM10 modulating peptides are based on SEQ ID NO: 3 or SEQ ID NO: 4, and in some embodiments include modifications at or near the N-terminal and/or the C-terminal ends of the disclosed peptides as well as substitutions, insertions, and deletions at one or more amino acid positions of the ADAM10 prodomain peptides disclosed herein.

1 . An ADAM10 modulating peptide, wherein the ADAM10 modulating peptide comprises, consists essentially of, or consists of at least one modification of an amino acid sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 2, wherein the at least one modification is selected from the group consisting of at least one amino acid substitution of at least one furin recognition site, at least one amino acid substitution of at least one meprin recognition site, and at least one modification of cysteine 151, or any combination thereof, such that the ADAM10 modulating peptide is less sensitive to cleavage by furin, less sensitive to cleavage by meprin, less sensitive to oxidation at cysteine 151, or any combination thereof. 
     
     
         2 . The ADAM10 modulating peptide of  claim 1 , wherein:
 (i) the at least one furin recognition site is selected from the group consisting of amino acid positions 26-29 and amino acid positions 52-55 of SEQ ID NOs: 1 or SEQ ID NO: 2; and   (ii) the at least one meprin recognition site is selected from the group consisting of:
 (1) amino acid positions 34-36, amino acid positions 62/63, amino acid positions 88/89, amino acid positions 136-138, amino acid positions 169/170, and amino acid positions 176-178 of SEQ ID NO: 1; or 
 (2) amino acid positions 34-36, amino acid positions 62/63, amino acid positions 88/89, amino acid positions 136-138, and amino acid positions 169/170 of SEQ ID NO: 2. 
   
     
     
         3 . The ADAM10 modulating peptide of  claim 1  or of  claim 2 , wherein cysteine 151 of SEQ ID NO: 1 or SEQ ID NO: 2 is substituted with alanine, serine, glycine, or threonine, or is pegylated. 
     
     
         4 . The ADAM10 modulating peptide of  claim 1 , wherein the ADAM10 modulating peptide comprises, consists essentially of, or consists of an amino acid sequence as set forth in either of SEQ ID NOs: 3 and 4. 
     
     
         5 . The ADAM10 modulating peptide of  claim 4 , wherein the at least one modification is selected from the group consisting of:
 (i) an alanine, serine, glycine, or lysine substitution at one or more of positions 26, 28 and 29 of SEQ ID NO: 3 or SEQ ID NO: 4;   (ii) an alanine, serine, glycine, or lysine substitution at one or more of positions 52, 54 and 55 of SEQ ID NO: 3 or SEQ ID NO: 4;   (iii) an alanine, serine, glycine, or asparagine substitution at one or both of positions 88 and 89;   (iv) an alanine. serine, glycine, or asparagine substitution at one or more of positions 176-178 of SEQ ID NO: 3;   (v) an alanine, serine, glycine, or threonine substitution at cysteine 151; and   (vi) attachment of a polyethylene glycol (PEG) moiety to cysteine 151, or any combination thereof.   
     
     
         6 - 16 . (canceled) 
     
     
         17 . The ADAM10 modulating peptide of  claim 4 , wherein amino acid position 151 of SEQ ID NO: 3 or SEQ ID NO: 4 is pegylated, the N-terminus is pegylated, the C-terminus is pegylated, or any combination thereof. 
     
     
         18 - 21 . (canceled) 
     
     
         22 . The ADAM10 modulating peptide of  claim 1 , wherein the ADAM10 modulating peptide comprises, consists essentially of, or consists of one or more modifications that inactivate a furin recognition site at amino acid positions 26-29 of SEQ ID NOs: 1 or SEQ ID NO: 2, a furin recognition site at amino acid positions 52-55 of SEQ ID NOs: 1 or SEQ ID NO: 2, or both. 
     
     
         23 . The ADAM10 modulating peptide of  claim 22 , wherein the ADAM10 modulating peptide further comprises, consists essentially of, or consists of one or more modifications that inactivate a meprin recognition site at amino acid positions amino acid positions 34-36 of SEQ ID NO: 1 or SEQ ID NO: 2, amino acid positions 62 and 63 of SEQ ID NO: 1 or SEQ ID NO: 2, amino acid positions 88 and 89 of SEQ ID NO:
 1 or SEQ ID NO: 2, amino acid positions 136-138 of SEQ ID NO: 1 or SEQ ID NO: 2, amino acid positions 169 and 170 of SEQ ID NO: 1 or SEQ ID NO: 2, or amino acid positions 176-178 of SEQ ID NO: 1, or any combination thereof.   
     
     
         24 . (canceled) 
     
     
         25 . The ADAM10 modulating peptide of  claim 1 , wherein the ADAM10 modulating peptide comprises, consists essentially of, or consists of a plurality of modifications that inactivate:
 (i) one or both of the furin recognition sites at amino acid positions 26-29 and 52-55 of SEQ ID NOs: 1 or SEQ ID NO: 2: and   (ii) one or more of the meprin recognition sites at amino acid positions 34-36, 62/63, 88/89, 136-138, 169/170, and 176-178 of SEQ ID NO: 1 or one or more of the meprin recognition sites at amino acid positions 34-36, 62/63, 88/89, 136-138, and 169 and 170 of SEQ ID NO: 2.   
     
     
         26 - 34 . (canceled) 
     
     
         35 . The ADAM10 modulating peptide of  claim 4 , wherein the ADAM10 modulating peptide comprises an amino acid sequence as set forth in any of SEQ ID NOs: 5-71,814. 
     
     
         36 . (canceled) 
     
     
         37 . The ADAM10 modulating peptide of  claim 35 , wherein the ADAM10 modulating peptide further comprises a PEG moiety at the N-terminus, a PEG moiety at the C-terminus, a PEG moiety at amino acid position 151 of SEQ ID NO: 1, or any combination thereof. 
     
     
         38 . The ADAM10 modulating peptide of  claim 35 , wherein the ADAM10 modulating peptide comprises an amino acid sequence at least 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to any of SEQ ID NOs: 5-71,814. 
     
     
         39 . The ADAM10 modulating peptide of  claim 35 , wherein the amino acid sequence comprises one or more amino acid substitutions of SEQ ID NOs: 5-71,814 at an amino acid position selected from the group consisting of amino acid positions 26, 28, 29, 34-36, 52, 54,55, 62, 63, 88, 89, 136-138, 151, 169-178, 182, and 183 as defined in SEQ ID NO: 3 for these amino acid positions. 
     
     
         40 - 46 . (canceled) 
     
     
         47 . An ADAM10 modulating peptide comprising, consisting essentially of, or consisting of an amino acid sequence that is at least 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to any one of SEQ ID NOs: 5-71,814, optionally wherein the ADAM10 modulating peptide is pegylated. 
     
     
         48 - 53 . (canceled) 
     
     
         54 . The ADAM10 modulating peptide of  claim 47 , wherein the amino acid sequence comprises at least one introduction of a cysteine internally, at the N-terminus, at the C-terminus, or any combination thereof within SEQ ID NO: 3 or SEQ ID NO: 4, and further wherein the at least one introduced cysteine results from a substitution of one or more amino acids within SEQ ID NO: 3 or SEQ ID NO: 4, an insertion of a cysteine within SEQ ID NO: 3 or SEQ ID NO: 4, or any combination thereof. 
     
     
         55 . The ADAM10 modulating peptide of  claim 54 , wherein the introduced cysteine is pegylated. 
     
     
         56 . The ADAM10 modulating peptide claim of 55, wherein the pegylated introduced cysteine is located within the first 20 amino acids within SEQ ID NO: 3 or SEQ ID NO: 4; within the last 20 amino acids of within SEQ ID NO: 3 or SEQ ID NO: 4, or a combination thereof. 
     
     
         57 - 65 . (canceled) 
     
     
         66 . The ADAM10 modulating peptide of  1 , wherein the amino acid sequence is at least 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 62 or SEQ ID NO: 63. 
     
     
         67 . The ADAM10 modulating peptide of  claim 1 , wherein ADAM10 modulating peptide comprises one or more modifications selected from the group consisting of conservative amino acid substitutions, non-natural amino acid substitutions, D- or D,L-racemic mixture isomer form amino acid substitutions, amino acid chemical substitutions, carboxy- or amino-terminal modifications, addition of one or more glycosyl groups, and conjugation to a molecule selected from the group consisting of a fatty acid, a PEG, a sugar, a fluorescent molecule, a chromophore, a radionuclide, a bioconjugate, a tag that can be employed for purification and/or isolation of the ADAM10 modulating peptide, and an antibody or a paratope-containing fragment or derivative thereof. 
     
     
         68 . A composition comprising the ADAM10 modulating peptide of  claim 1 . 
     
     
         69 . A pharmaceutical composition comprising the ADAM10 modulating peptide of  claim 1  and a pharmaceutically acceptable carrier or excipient. 
     
     
         70 . The pharmaceutical composition of  claim 69 , wherein the pharmaceutically composition is pharmaceutically acceptable for use in a human. 
     
     
         71 . A polypeptide comprising, consisting essentially of, or consisting of the ADAM10 modulating peptide of  claim 1 , optionally wherein the polypeptide is pegylated. 
     
     
         72 . The polypeptide of  claim 71 , wherein the polypeptide comprises, consists essentially of, or consists of an amino acid sequence as set forth in any of SEQ ID NOs: 5-71,814, optionally wherein the polypeptide is pegylated. 
     
     
         73 . The polypeptide of  claim 71 , wherein the polypeptide further comprises a tag, optionally a His tag or any other peptide or non-peptide tag that can be employed for purification and/or isolation of the polypeptide. 
     
     
         74 . The polypeptide of  claim 73 , wherein the tag is releasable from the polypeptide by proteolytic cleavage. 
     
     
         75 - 81 . (canceled) 
     
     
         82 . A method for modulating an ADAM10 biological activity in vitro or in vivo, the method comprising contacting a solution or a cell comprising an ADAM10 polypeptide with the ADAM10 modulating peptide of  claim 1 , wherein the contacting comprises contacting the ADAM10 polypeptide with an amount of the ADAM10 modulating peptide sufficient to modulate a biological activity of the ADAM10 polypeptide. 
     
     
         83 . (canceled) 
     
     
         84 . (canceled) 
     
     
         85 . A method for inhibiting an ADAM10 biological activity associated with a disease, disorder, or condition in a subject, the method comprising contacting an ADAM10 polypeptide present in the subject with an effective amount of the ADAM10 modulating peptide of  claim 1 , wherein the disease, disorder, or condition is selected from the group consisting of cancer, inflammation, an allergic response, lupus, asthma, an infectious disease, and fibrosis, and further wherein the subject has the disease, disorder, or condition or is predisposed thereto. 
     
     
         86 . (canceled) 
     
     
         87 . (canceled) 
     
     
         88 . A method for inhibiting release of an ADAM10 substrate from a cell, the method comprising contacting the cell with the ADAM10 modulating peptide of  claim 1  in an amount sufficient to inhibit release of the ADAM10 substrate from the cell. 
     
     
         89 . The method of  88 , wherein the ADAM10 substrate is selected from the group consisting of CD23, IL6-R, EGF, Her-2, HB-EGf, betacellulin, jagged-1, Notch receptor 1, Notch receptor 3, RAGE, fractalkine, MICA A, I-TAC, HGFR, GITR, GM-CSF, an IGF soluble receptors, and TGF beta. 
     
     
         90 . The method of  88 , wherein the cell is present in or has been isolated from a subject.