Stable Analogs of Peptide and Polypeptide Therapeutics

Abstract

The present invention relates to compositions of peptide and polypeptide analogs that are resistant to proteolysis, pharmaceutical uses thereof, and methods of preparation thereof.

1 . A proteinase-resistant analog of a biologically active peptide or polypeptide factor, which peptide or polypeptide factor has an amino acid sequence including a proteinase substrate sequence that is cleaved under physiological conditions by a target proteinase, wherein said analog has an amino acid sequence corresponding to said peptide or polypeptide factor where the P′ 1  residue of said proteinase substrate sequence is replaced with a non-naturally occurring amino acid analog having a tetrasubstituted Cβ carbon, which P′ 1  residue replacement reduces the susceptibility of the analog to cleavage by said target proteinase relative to said peptide or polypeptide factor. 
     
     
         2 . The proteinase-resistant analog of  claim 1 , wherein the non-naturally occurring amino acid analog is represented in Formula II: 
       
         
           
           
               
               
           
         
         wherein 
         R 1  and R 2  are independently selected from a lower alkyl or a halogen; 
         R 3  is selected from lower alkyl, aryl, hydroxyl group, —(CH 2 ) m —COOH, —(CH 2 ) m —NH 2 , —(CH 2 ) m —N—C(═NH)NH 2 , —(CH 2 ) m —C(═O)NH 2 , —SH, or —(CH 2 ) m —S—CH 3 ; and 
         m is 0, 1, or 2. 
       
     
     
         3 . The proteinase-resistant analog of  claim 2 , wherein R 1  and R 2  are independently selected from methyl, ethyl, or propyl. 
     
     
         4 . The proteinase-resistant analog of  claim 2 , wherein R 1  and R 2  are both methyl. 
     
     
         5 . The proteinase-resistant analog of  claim 2 , wherein R 3  is selected from lower alkyl, phenyl, hydroxyphenyl, indole, imidazole, hydroxyl, —COOH, —CH 2 COOH, —CH 2 CH 2 —NC(═NH)NH 2 , —CH 2 C(═O)NH 2 , —CH 2 CH 2 C(═O)NH 2 , —SH, or —CH 2 SCH 3 . 
     
     
         6 . The proteinase-resistant analog of  claim 2 , which retains at least 50 percent of a biological activity of said active peptide or polypeptide factor. 
     
     
         7 . The proteinase-resistant analog of  claim 2 , wherein target proteinase is a serine proteinase, metalloproteinase, aspartic proteinase, or cysteine proteinase. 
     
     
         8 . The proteinase-resistant analog of  claim 2 , wherein said biologically active polypeptide factor is selected from GLP-1, GLP-2, GIP, NPY, PP, and PYY. 
     
     
         9 . The proteinase-resistant analog of  claim 2 , wherein said biologically active polypeptide factor is selected from the group consisting of enkephalin, Leu-enkephalin, Met-enkephalin, angiotensin I, angiotensin II, vasopressin, endothelin, vasoactive intestinal peptide, neurotensin, endorphins, insulin, gramicidin, paracelsin, delta-sleep inducing peptide, gonadotropin-releasing hormone, human parathyroid hormone (1-34), truncated erythropoietin analogues described in Wrighton et al., 1996, Science 273:458-463), specifically EMP-1, Atrial natriuretic peptide (ANP, ANF), human brain natriuretic peptide (hBNP), cecropin, kinetensin, neurophysins, elafin, guamerin, atriopeptin I, atriopeptin II, atriopeptin III, deltorphin I, deltorphin II, vasotocin, bradykinin, dynorphin, dynorphin A, dynorphin B, growth hormone release factor, growth hormone, growth hormone releasing peptide, oxytocin, calcitonin, calcitonin gene-related peptide, calcitonin gene-related peptide II, growth hormone releasing peptide, tachykinin, adrenocorticotropic hormone (ACTH), brain natriuretic polypeptide, cholecystokinin, corticotropin releasing factor, diazepam binding inhibitor fragment, FMRF-amide, galanin, gastric releasing polypeptide, gastric inhibitory polypeptide, gastrin, gastrin releasing peptide, glucagon, glucagon-like peptide- 1, glucagon-like peptide-2, LHRH, melanin concentrating hormone, melanocyte stimulating hormone (MSH), alpha-MSH, morphine modulating peptides, motilin, neurokinin A, neurokinin B, neuromedin 13, neuromedin C, neuromedin K, neuromedin N, neuromedin U, neuropeptide K, neuropeptide Y, pituitary adenylate cyclase activating polypeptide (PACAP), pancreatic polypeptide, peptide YY, peptide histidine-methionine amide (PHM), secretin, somatostatin, substance K, thyrotropin-releasing hormone (TRH), kyotorphin, melanostatin (MIF-1), thrombopoeitin analogs, in particular AF 12505, insulin-like growth factor I (57-70), insulin-like growth factor I (30-41), insulin-like growth factor I (24-41), insulin-like growth factor II (33-40), insulin-like growth factor II (33-40), insulin-like growth factor II (69-84), growth hormone (GH)-releasing peptide-6 (GHRP-6), β-Interleukin 1 (163-171), β-Interleukin II (44-56), Interleukin II (60-70), epidermal growth factor, bivalirudin (Hirulog), hirulog-I, C-type natriuretic peptide, ornipressin (also known as 8-ornithine-vasopressin), octreotide, eptifibatide, calcitonin gene-related peptide (CGRP), endomorphin-1; endomorphin-2, nociceptin, angiotensinogen, adrenomodullin, antiarrhytmic peptide (AA-P), Antagonist G, indolicidin, osteocalcin, cortistatin 29, cortistatin 14, PD-145065, PD-142893, fibrinogen binding inhibitor peptide, leptin, GR 83074, parathyroid hormone related peptide, angiotensinogen, leupeptin, and any modified or truncated analog thereof. 
     
     
         10 . A proteinase-resistant analog of a biologically active peptide or polypeptide factor, which peptide or polypeptide factor has an amino acid sequence including a proteinase substrate sequence that is cleaved under physiological conditions by a target post-proline cleaving proteinase, wherein said analog has an amino acid sequence corresponding to said peptide or polypeptide factor where the P′ 1  residue of said proteinase substrate sequence is replaced with a non-naturally occurring amino acid analog of Formula I: 
       
         
           
           
               
               
           
         
         wherein 
         R 1  and R 2  are independently selected from lower alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, alkoxyl, carboxyl, carboxamide, carbonyl, halogen, hydroxyl, amine, or cyano, or R 1  and R 2  taken together form a ring of 4-7 atoms; 
         R 3  is selected from lower alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, amino, alkoxyl, halogen, carboxyl, carboxamide, carbonyl, cyano, thioalkyl, acylamino, amido, cyano, nitro, azido, sulfate, sulfonate, sulfonamido, —(CH 2 ) m —R 4 , —(CH 2 ) m OH, —(CH 2 ) m COOH, —(CH 2 ) m O-lower alkyl, —(CH 2 ) m O-lower alkenyl, —(CH 2 ) m O(CH 2 ) m —R 4 , —(CH 2 ) m SH, —(CH 2 ) m S-lower alkyl, —(CH 2 ) m S-lower alkenyl, —(CH 2 ) n S(CH 2 ) m —R 4 ; —(CH 2 ) m NH 2 , —(CH 2 ) m N—C(═NH)NH 2 , —(CH 2 ) m C(═O)NH 2 , or —(CH 2 ) m NH 2 ; 
         R 4  represents, independently for each occurrence, aryl, aralkyl, cycloalkyl, cycloalkenyl, or heterocycle; 
         m is 0, 1, or 2; and 
         n is 0, 1, or 2. 
       
     
     
         11 . A pharmaceutical preparation comprising the proteinase-resistant analog of  claim 1 . 
     
     
         12 . A packaged pharmaceutical preparation comprising the proteinase-resistant analog of  claim 1 , in a pharmaceutically acceptable excipient, and
 a label and/or instructions for administering to a patient.   
     
     
         13 . A packaged veterinary preparation comprising the proteinase-resistant analog of  claim 1 , in an acceptable excipient, and a label and/or instructions for administering to an animal. 
     
     
         14 . A method for treating or preventing a disease in which administration of a proteinase-resistant analog would be beneficial, comprising administering a biological active peptide or polypeptide factor of  claim 1  is beneficial. 
     
     
         15 . The method of  claim 14 , wherein the proteinase-resistant analog is an analog of GLP-1, GIP, or PYY, and the disease is selected from one or more of insulin resistance, glucose intolerance, heart-related ailments, hyperglycemia, hyperinsulinemia, obesity, hyperlipidemia, hyperlipoproteinemia, abnormalities of digestive tract mucosa, food intake disorder, and gastrointestinal disorder. 
     
     
         16 . The method of  claim 14 , wherein proteinase-resistant analog is an analog of GLP-2, and the disease is selected from one or more Crohn's disease and inflammatory bowel disease. 
     
     
         17 . The proteinase-resistant analog of  claim 1 , further comprising the C-terminal amino acid sequence of Exendin-4. 
     
     
         18 . The proteinase-resistant analog of  claim 17 , wherein the C-terminal amino acid sequence of Exendin-4 is PSSGAPPPS (SEQ ID NO: 36). 
     
     
         19 . A proteinase-resistant analog of a biologically active peptide or polypeptide factor, which peptide or polypeptide factor has the following amino acid sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 35) 
                 
                     
                   HAXGTFTSDVSSYLEGQAAKEFIAWLVKGRPSSGAPPPS-NH 2   
                 
             
                
                
               
            
           
         
         wherein X is a non-naturally occurring amino acid analog of Formula II: 
       
       
         
           
           
               
               
           
         
         wherein 
         R 1  and R 2  are independently selected from a lower alkyl or a halogen; 
         R 3  is selected from lower alkyl, aryl, hydroxyl group, —(CH 2 ) m —COOH, —(CH 2 ) m —NH 2 , —(CH 2 ) m —N—C(═NH)NH 2 , —(CH 2 ) m —C(═O)NH 2 , —SH, or —(CH 2 ) m —S—CH 3 ; and 
         m is 0, 1, or 2. 
       
     
     
         20 . A method for treating or preventing a disease or condition in which administration of a proteinase-resistant analog would be beneficial, comprising administering a proteinase-resistant analog of a biologically active peptide or polypeptide factor, which peptide or polypeptide factor has the following amino acid sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 35) 
                 
                     
                   HAXGTFTSDVSSYLEGQAAKEFIAWLVKGRPSSGAPPPS-NH 2   
                 
             
                
                
               
            
           
         
         wherein X is a non-naturally occurring amino acid analog of Formula II: 
       
       
         
           
           
               
               
           
         
         wherein 
         R 1  and R 2  are independently selected from a lower alkyl or a halogen; 
         R 3  is selected from lower alkyl, aryl, hydroxyl group, —(CH 2 ) m —COOH, —(CH 2 ) m —NH, —(CH 2 ) m —N—C(═NH)NH 2 , —(CH 2 ) m —C(═O)NH 2 , —SH, or —(CH 2 ) m —S—CH 3 ; and 
         m is 0, 1, or 2. 
       
     
     
         21 . The method of  claim 20 , wherein the disease or condition is selected from heart-related ailments, hyperglycemia, obesity, hyperlipidemia, diabetic complications, obesity-related hypertension, osteoporosis, depression, schizoaffective disorders, sleep apnea, attention deficit syndromes with poor concentration, memory loss, forgetfulness, narcolepsy, and gastrointestinal diseases. 
     
     
         22 . The method of  claim 21 , wherein the disease or condition is a heart-related ailment selected from myocardial infarction, ischemia-reperfusion injury, congestive heart failure, and cardiac arrest. 
     
     
         23 . The method of  claim 21 , wherein the disease or condition is selected from depression schizoaffective disorders, sleep apnea, attention deficit syndromes, memory loss, forgetfulness, and narcolepsy. 
     
     
         24 . The method of  claim 21 , wherein the disease or condition is a gastrointestinal disease selected from regional enteritis (Crohn's disease) and inflammatory bowel disease. 
     
     
         25 . A method for the modification of glucose metabolism, comprising administering a proteinase-resistant analog of a biologically active peptide or polypeptide factor, which peptide or polypeptide factor has the following amino acid sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO: 35) 
                 
                     
                   HAXGTFTSDVSSYLEGQAAKEFIAWLVKGRPSSGAPPPS-NH 2   
                 
             
                
                
               
            
           
         
         wherein X is a non-naturally occurring amino acid analog of Formula II: 
       
       
         
           
           
               
               
           
         
         wherein 
         R 1  and R 2  are independently selected from a lower alkyl or a halogen; 
         R 3  is selected from lower alkyl, aryl, hydroxyl group, —(CH 2 ) m —COOH, —(CH) m —NH 2 , —(CH 2 ) m —N—C(═NH)NH 2 , —(CH 2 ) m —C(═O)NH 2 , —SH, or —(CH 2 ) m —S—CH 3 ; and 
         m is 0, 1, or 2.