Conjugates between a variant staphylococcal enterotoxin e superantigen and a targeting antibody that binds to a cancer-associated cell surface structure

Abstract

The present invention relates to compositions and methods of use, wherein the composition comprises a conjugate of a bacterial superantigen and an antibody moiety. More particularly, the bacterial superantigen has been modified to decrease seroreactivity with retained superantigen activity.

1 - 92 . (canceled) 
     
     
         93 . A conjugate comprising a bacterial superantigen and an antibody moiety, wherein:
 the superantigen is a variant of Staphylococcal enterotoxin E, reference SEQ ID NO: 7, and differs from Staphylococcal enterotoxin E in comprising amino acid substitutions as follows, wherein the positions of the amino acid substitutions are relative to the amino acid positions in reference SEQ ID NO: 7:   (i) amino acid position 20 is glycine or a conserved variant thereof, amino acid position 21 is threonine or a conserved variant thereof, amino acid position 24 is glycine or a conserved variant thereof, amino acid position 27 is lysine or a conserved variant thereof, and amino acid position 227 is serine or alanine, or a conserved variant thereof; and   (ii) wherein at least one amino acid in a region C is substituted with a different amino acid, such that the superantigen variant has reduced seroreactivity compared to the seroreactivity of Staphylococcal enterotoxin having the amino acid sequence of SEQ ID NO: 7, and the position of the amino acid substitution in region C is selected from the group consisting of amino acid positions 74, 75, 78, 79, 81, 83 and 84;   
       and wherein the antibody moiety is a full length antibody, or a molecule-binding antibody fragment, that binds a cancer-associated cell surface antigen. 
     
     
         94 . The conjugate of  claim 93 , wherein the antibody moiety is an Fab fragment. 
     
     
         95 . The conjugate of  claim 93 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         96 . The conjugate of  claim 93 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         97 . The conjugate of  claim 93 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         98 . The conjugate of  claim 93 , wherein the substitution at amino acid position 227 is serine. 
     
     
         99 . The conjugate of  claim 93 , wherein the superantigen variant further comprises an amino acid substitution in a region E, wherein at least one amino acid in the region E is substituted with a different amino acid, such that the variant has reduced seroreactivity compared to the seroreactivity of Staphylococcal enterotoxin having the amino acid sequence of SEQ ID NO: 7, and the position of the amino acid substitution in region E is selected from the group consisting of amino acid positions 217, 220, 222, 223, 225 and 227. 
     
     
         100 . The conjugate of  claim 99 , wherein the superantigen variant comprises SEQ ID NO: 2. 
     
     
         101 . The conjugate of  claim 99 , wherein the antibody moiety is an Fab fragment. 
     
     
         102 . The conjugate of  claim 99 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         103 . The conjugate of  claim 99 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         104 . The conjugate of  claim 99 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         105 . The conjugate of  claim 99 , wherein the substitution at amino acid position 227 is serine. 
     
     
         106 . The conjugate of  claim 93 , wherein the substituted amino acid in region C comprises an amino acid selected from the group consisting of threonine or a conserved variant thereof at position 74, alanine or a conserved variant thereof at position 75, serine or a conserved variant thereof at position 78, glutamic acid or a conserved variant thereof at position 79, glutamic acid or a conserved variant thereof at position 81, serine or a conserved variant thereof at position 83, serine or a conserved variant thereof at position 84. 
     
     
         107 . The conjugate of  claim 106 , wherein the antibody moiety is an Fab fragment. 
     
     
         108 . The conjugate of  claim 106 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         109 . The conjugate of  claim 106 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         110 . The conjugate of  claim 106 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         111 . The conjugate of  claim 106 , wherein the substitution at amino acid position 227 is serine. 
     
     
         112 . The conjugate of  claim 99 , wherein the substituted amino acid in region E comprises an amino acid selected from the group consisting of threonine or a conserved variant thereof at position 217, serine or a conserved variant thereof at position 220, threonine or a conserved variant thereof at position 222, serine or a conserved variant thereof at position 223, and serine or a conserved variant thereof at position 225. 
     
     
         113 . The conjugate of  claim 112 , wherein the antibody moiety is an Fab fragment. 
     
     
         114 . The conjugate of  claim 112 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         115 . The conjugate of  claim 112 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         116 . The conjugate of  claim 112 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         117 . The conjugate of  claim 112 , wherein the substitution at amino acid position 227 is serine. 
     
     
         118 . A composition comprising the conjugate of  claim 93  and a carrier. 
     
     
         119 . The composition of  claim 118 , wherein the antibody moiety is an Fab fragment. 
     
     
         120 . The composition of  claim 118 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         121 . The composition of  claim 118 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         122 . The composition of  claim 118 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         123 . The composition of  claim 118 , wherein the substitution at amino acid position 227 is serine. 
     
     
         124 . The composition of  claim 118 , wherein the carrier is aqueous. 
     
     
         125 . The composition of  claim 118 , wherein the composition is lyophylized. 
     
     
         126 . The composition of  claim 118 , wherein the superantigen variant further comprises an amino acid substitution in a region E, wherein at least one amino acid in the region E is substituted with a different amino acid, such that the variant has reduced seroreactivity compared to the seroreactivity of Staphylococcal enterotoxin having the amino acid sequence of SEQ ID NO: 7, and the position of the amino acid substitution in region E is selected from the group consisting of amino acid positions 217, 220, 222, 223, 225 and 227. 
     
     
         127 . The composition of  claim 126 , wherein the superantigen variant comprises SEQ ID NO: 2. 
     
     
         128 . The composition of  claim 126 , wherein the antibody moiety is an Fab fragment. 
     
     
         129 . The composition of  claim 126 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         130 . The composition of  claim 126 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         131 . The composition of  claim 126 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         132 . The composition of  claim 126 , wherein the substitution at amino acid position 227 is serine. 
     
     
         133 . The composition of  claim 126 , wherein the carrier is aqueous. 
     
     
         134 . The composition of  claim 126 , wherein the composition is lyophylized. 
     
     
         135 . The composition of  claim 118 , wherein the substituted amino acid in region C comprises an amino acid selected from the group consisting of threonine or a conserved variant thereof at position 74, alanine or a conserved variant thereof at position 75, serine or a conserved variant thereof at position 78, glutamic acid or a conserved variant thereof at position 79, glutamic acid or a conserved variant thereof at position 81, serine or a conserved variant thereof at position 83, serine or a conserved variant thereof at position 84. 
     
     
         136 . The composition of  claim 135 , wherein the antibody moiety is an Fab fragment. 
     
     
         137 . The composition of  claim 135 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         138 . The composition of  claim 135 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         139 . The composition of  claim 135 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         140 . The composition of  claim 135 , wherein the substitution at amino acid position 227 is serine. 
     
     
         141 . The composition of  claim 135 , wherein the carrier is aqueous. 
     
     
         142 . The composition of  claim 135 , wherein the composition is lyophylized. 
     
     
         143 . The composition of  claim 126 , wherein the substituted amino acid in region E comprises an amino acid selected from the group consisting of threonine or a conserved variant thereof at position 217, serine or a conserved variant thereof at position 220, threonine or a conserved variant thereof at position 222, serine or a conserved variant thereof at position 223, and serine or a conserved variant thereof at position 225. 
     
     
         144 . The composition of  claim 143 , wherein the antibody moiety is an Fab fragment. 
     
     
         145 . The composition of  claim 143 , wherein the cancer-associated cell surface antigen is associated with a cancer selected from the group consisting of lung, breast, colon, kidney, pancreatic, ovarian, stomach, cervix and prostate cancer. 
     
     
         146 . The composition of  claim 143 , wherein the cancer-associated cell surface antigen is selected from the group consisting of carcinoembryonic antigen, prostate specific antigen, urinary tumor associated antigen, fetal antigen, tyrosinase (p97), gp68, TAG-72, HMFG, Sialyl Lewis Antigen, MucA, MucB, PLAP, estrogen receptor, laminin receptor, erb B and p155. 
     
     
         147 . The composition of  claim 143 , wherein the substitution at amino acid position 227 is alanine. 
     
     
         148 . The composition of  claim 143 , wherein the substitution at amino acid position 227 is serine. 
     
     
         149 . The composition of  claim 143 , wherein the carrier is aqueous. 
     
     
         150 . The composition of claim  1432 , wherein the composition is lyophylized.