Polypeptide derivatives

Abstract

A biologically active peptide selected from growth factors, peptide hormones, interferons and cytokines and analogues and derivatives thereof, and bearing at least one chelating group linked to an amino group of said peptide, the chelating group being capable of complexing a detectable element and such amino group having no significant binding affinity to target receptors, are complexed with a detectable element and are useful as a pharmaceutical, e.g. a radiopharmaceutical for in vivo imaging of target tissues or for therapy.

1 . A biologically active peptide selected from the group consisting of growth factors, insulin, LHRH, gastrin, gastrin releasing peptide, thyrotropin releasing hormone, thyroid stimulating hormone, prolactin, vasoactive intestinal peptide, angiotensin, interferons, IL-1, IL-4 and IL-6, and analogues or derivatives thereof and bearing at least one chelating group linked to an amino group of said peptide, the chelating group being capable of complexing a detectable element and such amino group having no significant binding affinity to target receptors.  
     
     
         2 . A peptide according to  claim 1  wherein the growth factor is EGF, IGF, fibroblast growth factor, tumor necrosis factor, transforming growth factor, platelet derived growth factor, nerve growth factor or bombesin or an analogue or derivative thereof.  
     
     
         3 . A biologically active peptide selected from the group consisting of epidermal growth factor, LHRH, LHRH agonists, LHRH antagonists, bombesin and bombesin antagonists and analogues and derivatives thereof, and bearing at least one chelating group linked to an amino group of said peptide, the chelating group being capable of complexing a detectable element and such amino group having no significant binding affinity to target receptors.  
     
     
         4 . A peptide according to  claim 1  or  3  wherein the chelating group is attached indirectly by means of a divalent bridging group to the amino group.  
     
     
         5 . A peptide according to  claim 1  or  4  wherein the chelating group is attached by an amide bond to the amino group.  
     
     
         6 . A peptide according to  claim 1  wherein the chelating group is a polyaminopolycarboxylic group, a group of formula IIa or IIb,  
       
         
           
           
               
               
           
         
       
       wherein 
 each of R 1 , R 2  and R 3  independently is C 1-6 alkyl, C 6-8 aryl or C 7-9 arylalkyl, each optionally substituted by OH, C 1-4 alkoxy, COOH or SO 3 H,  
 R 4  is  
                     
  the carbon atoms marked with * are attached to no groups,  
 n′ is 1 or 2,  
 i is an integer from 2 to 6, and  
 TT are independently α or β amino acids linked to each other by amide bonds,  
 or a group derived from compounds of formula III  
                     
 wherein  
 each of R 20 , R 20a , R 21 , R 22  and R 23  independently is hydrogen or C 1-4 alkyl,  
 X 2  is either a group capable of reacting with the N-amino group of the peptide, or a group capable of binding with the divalent bridging group and  
 m′ is 2 or 3,  
 from compounds of formula IV  
                     
 wherein  
 X 2  is as defined above,  
 from compounds of formula V  
                     
 wherein  
 each of R 24 , R 25 , R 26 , R 27 , R 28  and R 29  independently is hydrogen or C 1-4 alkyl, and  
 X 2  and m′ are as defined above,  
 from compounds of formula VI  
                     
 wherein  
 X 2  is as defined above,  
 X 3  is C 1-4 alkylene, C 1-4 alkylene substituted by one or two CO 2 R 30 , by CH 2 COR 30 , CONH 2  or CONHCH 2 CO 2 R 30 , phenylene, or phenylene substituted by CO 2 R 30  wherein R 30  is C 1-4 alkyl, and  
 Y 5  is hydrogen or CO 2 R 30 ,  
 from porphyrins or from Desferal.  
 
     
     
         7 . A peptide according to  claim 1  wherein the chelating group is a group derived from ethylene diaminetetraacetic acid (EDTA), diethylene triamine pentaacetic acid (DTPA), N-hydroxyethyl-N,N′,N′-ethylene diaminetriacetic acid (HEDTA), ethylene glycol-0,0′-bis(2-aminoethyl)-N,N,N′,N′-tetraacetic acid (EGTA), N,N′-bis(hydroxybenzyl)ethylenediamine-N,N′-diacetic acid (HBED), triethylenetetramine hexaacetic acid (TTHA), from substituted EDTA or -DTPA, from 1,4,7,10-tetra-azacyclododecane-N,N′,N″,N′″-tetraacetic acid (DOTA), 1,4,8,11-tetraazacyclotetradecaneN,N′,N″,N′″-tetraacetic acid (TETA), or from compounds of formula Ia, Ib or Ic,  
       
         
           
           
               
               
           
         
       
       wherein 
 R 10  is —CH 2 COOH or a functional derivative thereof, and  
 R 11  is -Alk-X 1  or  
                     
  wherein each of n and m independently i, 1, 2 or 3, Alk is C 1-11 alkylene, X 1  is —NCS or NH 2  optionally substituted by a protecting group and ring A is substituted or unsubstituted.  
 
     
     
         8 . A peptide according to  claim 1  wherein the chelating group is a group derived from EDTA, DTPA, DOTA, N′-p-isothiocyanatobenzyl-diethylene triamine-N,N,N″,N″-tetraacetic acid, N′-p-isothiocyanatophenethyl-diethylene triamine-N,N,N″,N″-tetraacetic acid, N-{2-[bis(carboxymethyl)-amino]ethyl}-N′-{2-[bis(carboxymethyl)amino]-2-(p-isothiocyanatobenzyl)-ethyl}-glycine, a compound of formula Ia, Ib or Ic,  
       
         
           
           
               
               
           
         
         wherein R 10  is —CH 2 COOH or a functional derivative thereof and R 11  is —(CH 2 ) 1-6 -NCS, p-isothiocyanatobenzyl or p-isothiocyanatophenethyl,  
         or a compound of formula Va  
         
           
             
             
                 
                 
             
           
         
       
     
     
         9 . A physiologically peptide selected from the group consisting of growth factors, peptide hormones, interferons and cytokines and analogues or derivatives thereof, and bearing a chelating group linked either directly or indirectly by means of a divalent bridging group to an amino group of said peptide, the chelating group being derived from N′-p-isothiocyanatobenzyl-diethylene triamine-N,N,N″,N″-tetraacetic acid, N′-p-isothiocyanatophenethyl-diethylene triamine-N,N,N″,N″-tetraacetic acid, N-{2-[bis(carboxymethyl)amino]ethyl}-N′-{2-[bis(carboxymethyl)amino]-2-(p-isothiocyanatobenzyl)-ethyl}-glycine, DOTA, C-functionalised tetraazacyclododecane-tetraacetic acids, C-functionalised tetraazacyclotetradecane-tetraacetic acids, C-functionalised triazacyclododecane triacetic acids, C-functionalised triazacyclononane triacetic acids and a compound of formula V  
       
         
           
           
               
               
           
         
       
       wherein 
 each of R 24 , R 25 , R 26 , R 27 , R 28  and R 29  independently is hydrogen or C 1-4 alkyl,  
 m′ is 2 or 3, and  
 X 2  is p-isothiocyanato-benzyl or -phenethyl,  
 the chelating group being capable of complexing a detectable element and said amino group having no significant binding affinity to target receptors.  
 
     
     
         10 . A peptide according to  claim 4  or  9 , wherein the divalent bridging group is a radical of formula (α 1 )  
       Z-R—CO—  (α 1 )  
       wherein 
 R is C 1-11 alkylene, hydroxy substituted C 2-11 alkylene, C 2-11 alkenylene,  
                     
  cyclohexylene, substituted cyclohexylene, or a radical of formula (α 2 )  
                     
  wherein n and m are as defined above, 
 the ring A is substituted or unsubstituted, and  
 R 5  is a residue as attached in Cα of a natural or synthetic α-amino acid, and  
 
 Z is a divalent group derived from a functional moiety capable of covalently reacting with the chelating agent.  
 
     
     
         11 . A peptide according to  claim 1  which is a LHRH antagonist of formula VII  
       R 33 -A 1 -B 1 -C 1 -D 1 -E 1 -F 1 -G 1 -H 1 -I 1 -K 1 -NH 2   (VII)  
       wherein 
 R 33  is hydrogen, C 1-7 acyl or carbamoyl,  
 A 1  is D-Phe optionally substituted in the phenyl ring by halogen, CF 3 , C 1-3 alkyl and/or C 1-3 alkoxy, α- or β-naphthyl-D-alanine, D-Trp optionally substituted in 5 or 6 position by halogen or C 1-3 alkoxy and/or in 1 position by formyl or acetyl, D- or L-Pro, D- or L-3,4-dehydroproline, D- or L-Ser, D- or L-Thr, D- or L-Ala, D-pyroglutamine, 3-(9-anthryl)-D,L-alanyl, 3-(2-fluorenyl)-D,L-alanyl or 3-(Het)-D,L-alanyl wherein Het is a heterocyclic aryl radical selected from  
                     
  wherein 
 A 2  and A 3  are independently selected from the group consisting of hydrogen, C 1-4 alkyl, chlorine and bromine, and  
 A 4  is O, S or N  
 
 B 1  is D-Phe optionally substituted in the phenyl ring by halogen, NO 2 , C 1-3 alkyl or C 1-3 alkoxy, D-α-methylPhe optionally substituted in 4 position by chlorine, 2,2-diphenylglycine or 3-(2-naphthyl)-D-alanine,  
 C 1  is D-Trp optionally substituted in 5 or 6 position by halogen, NO 2  or C 1-3 alkoxy and/or in 1 position by formyl or acetyl, 3-(2- or 1-(naphthyl)-D-alanine, 3-D-pyridylalanine, D-Tyr, D-Phe optionally substituted by halogen, C 1-3 alkyl and/or C 1-3 alkoxy, D-3-Pz-Ala, D-Tin-Glu or D-Nic-Lys,  
 D 1  is L-Ser,  
 E 1  is Tyr, Phe optionally substituted in the phenyl ring by halogen, C 1-3 alkyl and/or C 1-3 alkoxy, Orn, Lys, Lys-Nic, MPic-Lys, Pic-Lys, DPic-Lys, Mpic-Lys, DMG-Lys, Pmc-Lys, Pzc-Lys, PmACAla, PzACAla, His, Dpo, Arg, 3-(3-pyridyl)-Ala, Trp, N-(3-pyridyl)acetyl-Lys or Glu(pMeO-phenyl), Cit, HOBLys or PzACAla,  
 F 1  is D-Phe optionally substituted in the phenyl ring by halogen, NO 2 , NH 2 , C 1-3 alkyl or C 1-3 alkoxy, D-Trp optionally substituted in 5 or 6 position by halogen, NO 2  and/or C 1-3 alkoxy and/or in 1 position by formyl or acetyl, 3-(2-naphthyl)-L-alanyl, D-Tyr, D-Orn, D-Lys, D-Lys-Nic, D-MNic-Lys, D-MPic-Lys, Pic-Lys, DPic-Lys, D-Pmc-Lys, D-Pzc-Lys, D-Bz-Lys, D-ILys, AnGlu, D-NACAla, D-PzACAla, D-PmACAla, D-3-(3-pyridyl)-Ala, D-His (subst. H or benzyl), D-Arg, D-homo-Arg(Et 2 ), D-Cit, D-HCi, D-Lys-Pic, D-Cit(C 1-3 alkyl), D-HCi(C 1-3 alkyl), D-Glu(AA) or α-amino-ω-ureido-C 2-4 alkanoic acid,  
 G 1  is Leu, Nle, Nval, N-α-methylLeu, Trp, Phe, Met, Tyr, Val, Ile, alloIle, Abu or Ala,  
 H 1  is Arg, IOrn, Lys, ILys or Cyp-Lys  
 I 1  is Pro, hydroxyproline, 3,4-dehydroproline, Pip and  
 K 1  is D-Ala, D-Leu, Gly, D-Ser or Sar,  
 in free form or in salt form.  
 
     
     
         12 . A peptide according to  claim 1  which is a bombesin antagonist of formula IXa  
       
         
           
                 
                 
                 
               
                     
                 
                   R 36  - A 6  - B 3  - C 3  - D 3  - E 3  - F 3  - G 3  - I 3  - Q  
                   (IXa) 
                     
                 
                         1    2    3    4    5    6    7    8    9    10 
                 
                     
                 
             
                
                
                
                
               
            
           
         
       
       wherein 
 R 36  is hydrogen, C 1-6 alkyl, C 2-6 alkanoyl, C 4-6 cycloalkoxy-carbonyl or C 1-4 alkoxycarbonyl,  
 A 6  is a direct bond or Gly, Arg, Lys, Phe, Asp, Nal, Pro, β-Ala or Glp,  
 B 3  is a direct bond or Gly, Pro or Asn,  
 C 3  is a direct bond or Lys or D-Nal,  
 D 3  is a direct bond or His, MeHis, EtHis, PrHis, Gln, Glu (OMe)-Glp, Leu, MeLeu, Lys, Pal, Phe, Pro, Arg, Trp or Thr,  
 E 3  is Trp, Val, Nal, Leu, Lys, Pal,  
 F 3  is Ala, MeAla, Aib, Gly, Pro, Leu, Phe, Ser, Val, Nal, Thr, Arg or Glu,  
 G 3  is Val, Aib, Leu, Ile, Thr, Phe or Ser,  
 H 3  is Gly, Sar, Ala, Ser, Aib, Pro, Lys, Asp, Arg, Val, Ac 3 c, Ac 5 c or Ac 6 c,  
 I 3  is His, MeHis, Aib, Val, Leu, MeLeu, Ala, Ile, Met, Pro, Phe, Gln, Lys, Pal, Ser, Thr, Glu, Asp, Trp or Nal, and  
 Q is K 3 -R 37  wherein K 3  is Leu, MeLeu, Ile, MeIle, Aib, Pro, Val, MeVal, Phe, Ape, MeApe, Met, Ser, Gln, Glu or Trp and R 37  is C 1-3 alkylamino, C 1-4 (dialkyl)amino or C 1-3 alkoxy or Q is C 1-6 alkoxy, C 1-10 alkylamino or C 1-10 (dialkyl)amino, or a compound of formula IXb  
                       A 7  - B 4  - Gln -Trp - Ala - val - W - X 6  - Y 6  - T 1            1    2    3    4    5    6    7    8    9    10                         (IXb)                                       
 wherein  
 A 7  is hydrogen, Boc, Lys, Arg,  
 B 4  is a direct bond or Asn, Thr, Glp,  
 W is Gly or Ala,  
 X 6  is a direct bond, His(R 38 ), Phe, Ser or Ala,  
 Y 6  is a direct bond, Leu or Phe,  
 T 1  is amino, NH(CH 2 ) 4 CH 3 , benzylamino, Met-R 39 , Leu-R 39 , Ile-R 39 , Ile-R 39  or Nle-R 39 ,  
 R 38  is hydrogen or benzyl, and  
 R 39  is amino, hydroxy, methoxy or —NHNH 2 ,  
 in free form or in salt form.  
 
     
     
         13 . A peptide according to  claim 1  wherein the chelating group is attached to the terminal amino group.  
     
     
         14 . A peptide according to  claim 1  wherein the chelating group is attached to a side chain amino group.  
     
     
         15 . A peptide which is DTPA-O-Ala-mEGF, [DTPA-β-Ala-Trp 14 ]-tetragastrin, acetyl-DPhe(pCl)-DPhe(pCl)-DTrp-Ser-Tyr-DLys(DTPA)-Leu-Arg-Pro-DAla-NH 2 , DTPA-DNal-(2) 1 -DPhe(p-Cl)-DTrp-Ser-Tyr-DLys(CH 2 —CHOH—CH 2 OH)-Leu-Lys(CH 2 —CHOH—CH 2 OH)-Pro-DAla-NH 2  and acetyl-His-Trp-Ala-Val-DAla-Lys(β-Ala-DTPA)-Leu-OEt.  
     
     
         16 . A process for the preparation of a peptide according to  claim 1  or  9 , comprising 
 a) removing at least one protecting group which is present in a peptide bearing a chelating group, or  
 b) linking together by an amide bond two peptide fragments each of them containing at least one amino acid in protected or unprotected form and one of them containing the chelating group, wherein the amide bond is in such a way that the desired amino acid sequence is obtained, and then effecting optionally stage a) of the process, or  
 c) linking together a chelating agent and the desired peptide in protected or unprotected form in such a way that the chelating group is fixed on the desired amino group of the peptide, and then effecting optionally stage a), or  
 d) removing a functional group of an unprotected or a protected peptide bearing a chelating group or converting it into another functional group so that another unprotected or a protected peptide bearing a chelating group is obtained and in the latter case effecting stage a) of the process,  
 and recovering the peptide thus obtained in free form or in salt form.  
 
     
     
         17 . A peptide according to  claim 1  or  9  in free form or in pharmaceutically acceptable salt form for use as a pharmaceutical.  
     
     
         18 . A pharmaceutical composition comprising a peptide of  claim 1  or  9 , in free form or in pharmaceutically acceptable salt form in association with a pharmaceutically carrier or diluent.  
     
     
         19 . A chelate which comprises a peptide as defined in  claim 1  or  9 , complexed with a detectable element, in free form or in pharmaceutically acceptable salt form.  
     
     
         20 . A chelate according to  claim 19  wherein the detectable element is a fluorescent or a α-, β- or γ-emitting element.  
     
     
         21 . A chelate according to  claim 19 , in free form or in pharmaceutically acceptable salt form, for use as a pharmaceutical.  
     
     
         22 . A chelate according to  claim 19 , in free form, for use as an imaging agent when the detectable element is a fluorescent or γ-emitting element, or for use in therapy when the detectable element is an α- or β-emitting element.  
     
     
         23 . A pharmaceutical composition comprising a chelate according to  claim 19 , in free form or in pharmaceutically acceptable salt form in association with a pharmaceutically carrier or diluent.  
     
     
         24 . A process for the production of a chelate according to  claim 19 , comprising reacting said peptide, in free form or in salt form, with a detectable element yielding compound.  
     
     
         25 . A package containing unit dosages of a peptide as defined in  claim 1  or  9  and of a detectable element with instructions for mixing them and for the use as imaging agent or therapeutic agent.  
     
     
         26 . A compound of formula XI  
       A-Z-R-Z 2 -X 7   (XI)  
       wherein 
 A is a chelating group,  
 Z is a divalent group derived from a functional moiety capable of covalently reacting with a chelating agent,  
 R is C 1-11 alkylene, hydroxy substituted C 2-11 alkylene, C 2-11 alkenylene,  
                     
  cyclohexylene, substituted cyclohexylene, or a radical of formula (α 2 )  
                     
  wherein each of n and m independently is 0, 1, 2 or 3, 
 the ring A is substituted or unsubstituted, and  
 R 5  is a residue as attached in Cα of a natural or synthetic α-amino acid, and  
 
 Z 2  is COOH or a functional group of a carboxy function,  
 or a compound of formula XII  
 A-Z′ 1 l-CO—X 7   (XII)  
 wherein  
 A is as defined above,  
 Z′ 1  is either a direct bond or -Z-R wherein Z and R are as defined above, and  
 X 7  is —NH—NH 2  in protected or unprotected form or —N 3 .  
 
     
     
         27 . Use of a peptide according to  claim 1  in free or pharmaceutically acceptable salt form, in the preparation of a diagnostic agent for imaging target tissues.