Composition and method for modifying polypeptides

Abstract

The present disclosure provides methods for site-selectively crosslinking payloads to antibodies and other proteins. This can be accomplished using traceless affinity labels designed to label target proteins with bio-orthogonally reactive entities (ORE) using the compositions and methods described herein.

1 - 14 . (canceled) 
     
     
         15 . A linker-antibody conjugate of Formula III 
       
         
           
           
               
               
           
         
         or a pharmaceutically acceptable salt thereof, 
         wherein 
         R 1  and R 2  are, each independently, a reactive handle; 
         q1, q2, and q3 are each, independently, 0-40; 
         r1, r2, and r3 are each, independently, 1-20; 
         s is 1-6; 
         m is 1-2; and 
         AB is an antibody. 
       
     
     
         16 . The linker-antibody conjugate of  claim 15 , wherein the linker is site selectively attached to a lysine on AB. 
     
     
         17 . The linker-antibody conjugate of  claim 16 , wherein the lysine is lysine 188. 
     
     
         18 . The linker-antibody conjugate of  claim 15 , wherein the linker is between 50-100% site-selective for lysine 188. 
     
     
         19 . The linker-antibody conjugate of  claim 15 , wherein the linker is between 75-100% site-selective for lysine 188. 
     
     
         20 . The linker-antibody conjugate of  claim 15 , wherein the linker is at least 90% site-selective for lysine 188. 
     
     
         21 . The linker-antibody conjugate of  claim 15 , wherein R 1  and R 2  are, each independently, selected from the group consisting of alkyne, cycloalkyne (e.g. cyclooctyne, mono- or difluorinated cyclooctyne, di methoxyazacyclooctyne, di benzocyclooctyne, dibenzoazacyclooctyne, biarylazacyclooctynone, bicyclononyne, 2,3,6,7-tetramethoxydibenzocyclooctyne, sulfonylated di benzocyclooctyne, carboxymethyl monobenzocyclooctyne, mono- or difluorinated monobenzocyclooctyne, pyrrolocyclooctyne), azide, benzyl azide, 1,3-diene, nitrile oxide, nitrone, tetrazine, trans-cyclooctene or their structurally related derivatives capable of undergoing bio-orthogonal cycloaddition to an appropriate corresponding reaction entity 
     
     
         22 . The linker-antibody conjugate of  claim 15 , wherein R 1  and R 2  are, each independently, selected from trans-cyclooctene or tetrazine. 
     
     
         23 . The linker-antibody conjugate of  claim 15 , wherein R 1  and R 2  are azide. 
     
     
         24 . A payload-linker-antibody conjugate of Formula IV 
       
         
           
           
               
               
           
         
         or a pharmaceutically acceptable salt thereof, 
         wherein 
         q1, q2, and q3 are each, independently, 0-40; 
         r1, r2, and r3 are each, independently, 1-20; 
         s is 1-6; 
         m is 1-2; 
         L 1  and L 2  are linking moieties, which are the product of a click reaction; 
         Y and Z are each, independently, selected from a toxin, a drug, and a chelator payload; and 
         AB is an antibody. 
       
     
     
         25 . The payload-linker-antibody conjugate of  claim 24 , wherein L 1  and L 2  are the product of a click reaction between an azide and a cycloalkyne group. 
     
     
         26 . The payload-linker-antibody conjugate of  claim 24 , wherein the L 1  or L 2  group formed are the product of reacting an azide on Y or Z of Formula IV. 
     
     
         27 . The payload-linker-antibody conjugate of  claim 24 , wherein the L 1  or L 2  group formed are the product of reacting a cycloalkyne on Y and/or Z of Formula IV. 
     
     
         28 . The payload-linker-antibody conjugate of  claim 24 , wherein L 1  or L 2  are, each independently, selected from —C(O)C 1 -C 30 -heteroaromatic- or -C 1 -C 30 -heteroaromatic-C(O)—; 
     
     
         29 . The payload-linker-antibody conjugate of  claim 24 , wherein L 1  or L 2  are a linking moiety of the structure: 
       
         
           
           
               
               
           
         
       
       wherein either (A) or (B) are attached to the toxin, drug, or chelator payload. 
     
     
         30 . The payload-linker-antibody conjugate of  claim 24 , wherein the linker is site selectively attached to lysine. 
     
     
         31 . (canceled) 
     
     
         32 . The payload-linker-antibody conjugate of  claim 30  wherein the linker is between 50-100% site-selective for lysine 188. 
     
     
         33 . The payload-linker-antibody conjugate of  claim 30 , wherein the linker is between 75-100% site-selective for lysine 188. 
     
     
         34 . The payload-linker-antibody conjugate of  claim 15 , wherein AB is selected from the group consisting of abciximab, adalimumab, adalimumab-atto, ado-trastuzumab emtansine, alemtuzumab, alirocumab, atezolizumab, avelumab, basiliximab, belimumab, bevacizumab, bezlotoxumab, blinatumomab, brentuximab vedotin, brodalumab, canakinumab, capromab pendetide, certolizumab pegol, cetuximab, daclizumab (Zenapax), daclizumab (Zinbryta), daratumumab, denosumab, dinutuximab, dupilumab, durvalumab, eculizumab, elotuzumab, evolocumab, golimumab, golimumab, ibritumomab tiuxetan, idarucizumab, infliximab, infliximab-abda, infliximab-dyyb, ipilimumab ixekizumab, mepolizumab, natalizumab, necitumumab, nivolumab, obiltoxaximab, obinutuzumab, ocrelizumab, ofatumumab, olaratumab, omalizumab,palivizumab, panitumumab, pembrolizumab, pertuzumab, ramucirumab, ranibizumab, raxibacumab, reslizumab, rituximab, secukinumab, siltuximab, tocilizumab, tocilizumab, trastuzumab, ustekinumab, vedolizumab, sarilumab, rituximab, hyaluronidaseguselkumab, inotuzumab ozogamicin, adalimumab-adbm, gemtuzumab ozogamicin, bevacizumab-awwb, benralizumab, emicizumab-kxwh, trastuzumab-dkst, ibalizumab-uiyk, tildrakizumab-asmn, burosumab-twza, and erenumab-aooe. 
     
     
         35 . The payload-linker-antibody conjugate of  claim 29 , wherein (A) or (B) is a chelator. 
     
     
         36 . The payload-linker antibody conjugate of  claim 35 , wherein the chelator is bound to a radionuclide. 
     
     
         37 . The payload-linker-antibody conjugate of  claim 35  wherein the chelator is macropa or DOTA. 
     
     
         38 . The payload-linker-antibody-conjugate of  claim 36 , wherein the radionuclide is actinium-225 or lutetium-177. 
     
     
         39 . The payload-linker-antibody-conjugate of  claim 36 , wherein the chelator is macropa and the radionuclide is actinium-225. 
     
     
         40 . The payload-linker-antibody-conjugate of  claim 36 , wherein the chelator is DOTA and the radionuclide is lutetium-177. 
     
     
         41 - 53 . (canceled) 
     
     
         54 . A linker-antibody conjugate of Formula III′ 
       
         
           
           
               
               
           
         
       
       or a pharmaceutically acceptable salt thereof, 
       wherein
 R 1  and R 2  are each independently a reactive handle; 
 AB is an antibody; 
 Y 1  is (Z 1A )-(J 1 )-(Z 1B ); 
 Y 3  is (Z 3A )-(J 3 )-(Z 3B ); 
 J 1  is either —(CH 2 CH 2 O) q1 (CH 2 ) r1 — or —(CH 2 ) r1 (OCH 2 CH 2 ) q1 —; 
 J 3  is either —(CH 2 CH 2 O) q2 (CH 2 ) 3 — or —(CH 2 ) r3 (OCH 2 CH 2 ) q3 —; 
 Z 1A , Z 1B , Z 3A , and Z 3B  are each independently a bond or C 1-6  alkyl, wherein 1, 2, 3, or 4 —CH 2 — groups of C 1-6  alkyl are optionally and independently replaced with —O—, —N(H)—, —C(O)—, —C(O)N(H)—, —N(H)C(O)—, —C(O)O—, —OC(O)—, —OC(O)O—, —N(H)C(O)O—, and —OC(O)N(H)—; 
 q1, q2, and q3 are each independently 0-40; 
 r1, r2, and r3 are each independently 1-20; and 
 m is 1-2. 
 
     
     
         55 . The linker-antibody conjugate of  claim 54 , wherein Z 1B  and Z 3B  are each independently —(CH 2 ) s C(O)N(H)— or —(CH 2 ) s N(H)C(O)—, and wherein each s is independently 0-6. 
     
     
         56 . The linker-antibody conjugate of  claim 54 , wherein the structure is represented by Formula III′a 
       
         
           
           
               
               
           
         
         or a pharmaceutically acceptable salt thereof. 
       
     
     
         57 . The linker-antibody conjugate of  claim 54 , wherein the structure is represented by Formula III′b 
       
         
           
           
               
               
           
         
         or a pharmaceutically acceptable salt thereof. 
       
     
     
         58 . The linker-antibody conjugate of  claim 54 , wherein R 1  and R 2  are each either a trans-cyclooctene or tetrazine. 
     
     
         59 . The linker-antibody conjugate of  claim 54 , wherein R 1  and R 2  are each either an azide or cycloalkyne. 
     
     
         60 . The linker-antibody conjugate of  claim 54 , wherein R 1  or R 2  are an azide, cycloalkyne, trans-cyclooctene, or tetrazine. 
     
     
         61 . A payload-linker-antibody conjugate of Formula IV′ 
       
         
           
           
               
               
           
         
         or a pharmaceutically acceptable salt thereof, 
       
       wherein
 m is 1-2; 
 Y 1  is (Z 1A )-(J 1 )-(Z 1B ); 
 Y 3  is (Z 3A )-(J 3 )-(Z 3B ); 
 J 1  is either —(CH 2 CH 2 O) q1 (CH 2 ) r1  or —(CH 2 ) r1 (OCH 2 CH 2 ) q1 —; 
 J 3  is either —(CH 2 CH 2 O) q3 (CH 2 ) r3 — or —(CH 2 ) r3 (OCH 2 CH 2 ) q3 —; 
 q1, q2, and q3 are each, independently, 0-40; 
 r1, r2, and r3 are each, independently, 1-20; 
 Z 1A , Z 1B , Z 3A , and Z 3B  are each independently a bond or C 1-6  alkyl, wherein 1, 2, 3, or 4 —CH 2 — groups of C 1-6  alkyl are optionally and independently replaced with —O—, —N(H)—, —C(O)—, —C(O)N(H)—, —N(H)C(O)—, —C(O)O—, —OC(O)—, —OC(O)O—, —N(H)C(O)O—, and —OC(O)N(H)—; 
 L 1  and L 2  are linking moieties, which are the product of a click reaction; 
 Y′ and Z′ are each, independently, selected from a toxin, a drug, and a chelator payload; and 
 AB is an antibody.