Agonist of spexin-based galanin type 2 receptor and use thereof

Abstract

The present invention relates to spexin-based agonists specific for galanin receptor type 2 and use thereof. More specifically, the present invention provides peptide-based agonists with high specificity for galanin receptor type 2 and improved stability. The peptide-based agonists are involved in the regulation of in vivo physiological functions, such as food intake, anxiety, emotion, and addiction, for which galanin receptors type 2 is responsible, to effectively suppress appetite, help recover from anxiety disorder, and reduce pleasure addiction. Therefore, the peptide-based agonists can be used to prevent or treat galanin receptor type 2-mediated diseases.

1 . A peptide-based galanin receptor type 2 (GALR2) agonist having the amino acid sequence set forth in formula 1: 
       
         
           
                 
                 
               
                     
                   [dN] 1 [W] 2 [T] 3 [P] 4 [N] 5 [A] 6 [A] 7 [L] 8 [Y] 9 [L] 10 [F] 11   
                 
                     
                     
                 
                     
                   [G] 12 [P] 13 [Q] 14 -NH 2  (1) 
                 
             
                
                
                
               
            
           
         
         wherein one or more amino acids in formula 1 are optionally substituted, 
         wherein, in formula 1, [dN] 1  represents D-asparagine and is optionally replaced with one of pyroglutamate (pQ), citrulline (Cit), L-asparagine (N), and glycine (G) or is optionally replaced with asparagine protected with polyethylene glycol (PEG), an acetyl (Ac) group, or Fmoc, 
         [W] 2  is optionally replaced with D-tryptophan (dW) or a 2-naphtyl group, 
         [T] 3  is optionally replaced with alanine (A), lysine (K), or D-threonine (dT), 
         [P] 4  is optionally replaced with leucine (L), glutamate (E), arginine (R), D-alanine (dA), or D-valine (dV), 
         [N] 5  is optionally replaced with glutamine (Q), 
         [A] 6  is optionally replaced with D-alanine (dA) or serine (S), 
         [A] 7  is optionally replaced with methionine (M), 
         [L] 8  is optionally replaced with glycine (G), methionine (M), or glutamine (Q), 
         [Y] 9  is optionally replaced with methionine (M) or D-tyrosine (dY), 
         [L] 10  is optionally replaced with D-leucine (dL), 
         [F] 11  is optionally replaced with leucine (L) or tyrosine (Y), 
         [G] 12  is optionally replaced with D-alanine (dA), 
         [P] 13  is optionally replaced with D-alanine (dA) or alanine (A), and 
         [Q]14 is optionally replaced with D-glutamine (dQ) or histidine (H), and 
         wherein the GALR2 agonist specifically activates galanin receptor type 2. 
       
     
     
         2 - 6 . (canceled) 
     
     
         7 . A method for treating a galanin receptor type 2-mediated disease in a subject in need thereof, comprising administering to the subject a peptide-based galanin receptor type 2 (GALR2) agonist,
 wherein the GALR2 agonist comprises the amino acid sequence   [dN] 1 [W] 2 [T] 3 [P] 4 [N] 5 [A] 6 [A] 7 [L] 8 [Y] 9 [L] 10 [F] 11 [G] 12 [P] 13 [Q] 14  (SEQ ID NO: 1), wherein:   [dN] 1  is D-asparagine, L-asparagine (N), pyroglutamate (pQ), citrulline (Cit), or glycine (G);   [W] 2  is a tryptophan;   [T] 3  is a threonine, an alanine (A), or a lysine (K);   [P] 4  is a proline, a leucine (L), a glutamate (E), an arginine (R), an alanine (A), or a valine (V);   [N] 5  is an asparagine or a glutamine (Q);   [A] 6  is an alanine or a serine (S);   [A] 7  is an alanine or a methionine (M);   [L] 8  is a leucine, a glycine (G), a methionine (M), or glutamine (Q);   [Y] 9  is a tyrosine or a methionine (M);   [L] 10  is a leucine;   [F] 11  is a phenylalanine, a leucine (L), or a tyrosine (Y);   [G] 12  is a glycine or an alanine (A);   [P] 13  is a proline or an alanine (A), and;   [Q]14 is a glutamine or a histidine (H), and   wherein the GALR2 agonist specifically activates galanin receptor type 2.   
     
     
         8 . The method according to  claim 7 , wherein the GALR2 agonist is prepared into a freeze-dried powder or injectable solution. 
     
     
         9 . The method according to  claim 7 , wherein the galanin receptor type 2-mediated disease comprises an attention deficit hyperactivity disorder (ADHD), bipolar disorder, body dysmorphic disorder, bulimia nervosa and other eating disorders, cataplexy, dysthymia, general anxiety disorder, hypersexuality, irritable bowel syndrome, impulse-control disorder (MDD), kleptomania, migraine, major depressive disorder, narcolepsy, obsessive-compulsive disorder, oppositional-defiant disorder, panic disorder, post-traumatic stress disorder (PTSD), premenstrual dysphoric disorder (PMDD), social anxiety disorder, chronic pain, intermittent explosive disorder, pathological gambling, personality disorder, pyromania, substance abuse and addiction, trichotillomania, or Alzheimer's disease. 
     
     
         10 . A polynucleotide encoding the GALR2 agonist of  claim 1 . 
     
     
         11 . A method of making a GALR2 agonist, comprising chemically synthesizing the GALR2 agonist of  claim 1 . 
     
     
         12 . The method of  claim 7 , wherein the amino acid sequence of the GALR2 agonist is attached to NH 2  on the C-terminus. 
     
     
         13 . The method of  claim 7 , wherein [A] 7  is A and [F] 11  is F. 
     
     
         14 . The method of  claim 7 , wherein [N] 5  is N, [A] 7  is A, and [F] 11  is F. 
     
     
         15 . The method of  claim 7 , wherein [N] 5  is N, [A] 7  is A, [F] 11  is F, and [P] 13  is P. 
     
     
         16 . The method of  claim 7 , wherein the GALR2 agonist comprises the amino acid sequence set forth in SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 27, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 50, SEQ ID NO: 51; SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, or SEQ ID NO: 59. 
     
     
         17 . The method of  claim 7 , wherein [P] 4  is dA or R. 
     
     
         18 . The method of  claim 7 , wherein [dN] 1  is replaced with N, and wherein the N is protected with a polyethylene glycol (PEG), acetyl (Ac) group, or Fmoc. 
     
     
         19 . A method of preparing a peptide-based galanin receptor type 2 (GALR2) agonist that specifically activates GALR2, comprising modifying one or more amino acids of a human spexin peptide, such that the GALR2 agonist comprises the amino acid sequence NWTPQAMLYLKGAQ (SEQ ID NO: 8) with one or more of the following amino acid modifications:
 (1) a substitution of asparagine (N) at position 1 with a pyroglutamate (pQ), a citrulline (Cit), or a glycine (G);   (2) a substitution of the tryptophan (W) at position 2 with a 2-naphtyl group;   (3) a substitution of the threonine (T) at position 3 with an alanine (A) or a lysine (K);   (4) a substitution of the proline (P) at position 4 with a leucine (L), a glutamate (E), an arginine (R), an alanine (A), or a valine (V);   (5) a substitution of the glutamine (Q) at position 5 with an asparagine (N);   (6) a substitution of the alanine (A) at position 6 with a serine (S);   (7) a substitution of the methionine (M) at position 7 with an alanine (A);   (8) a substitution of the leucine (L) at position 8 with a glycine (G), a methionine (M), or a glutamine (Q);   (9) a substitution of the tyrosine (Y) at position 9 with a methionine (M);   (10) a substitution of the lysine (K) at position 11 with a phenylalanine (F), leucine (L), or tyrosine (Y);   (11) a substitution of the glycine (G) at position 12 with an alanine (A);   (12) a substitution of the alanine (A) at position 13 with a proline (P);   (13) a substitution of the glutamine (Q) at position 14 with a histidine (H); or   (14) any combination of (1) to (13).   
     
     
         20 . The method of  claim 19 , wherein the GALR2 agonist comprises:
 (i) a substitution of the M at position 7 with A;   (ii) a substitution of the K at position 11 with F; or   (iii) both (i) and (ii).   
     
     
         21 . The method of  claim 20 , wherein the GALR2 agonist comprises a substitution of the Q at position 5 with N. 
     
     
         22 . The method of  claim 21 , wherein the GALR2 agonist comprises a substitution of the A at position 13 with P. 
     
     
         23 . The method of  claim 19 , wherein the GALR2 agonist comprises the amino acid sequence set forth in SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 27, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 50, SEQ ID NO: 51; SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, or SEQ ID NO: 59. 
     
     
         24 . The method of  claim 19 , wherein the N at position 1 is protected with a polyethylene glycol (PEG), an acetyl (Ac) group, or a Fmoc. 
     
     
         25 . The method of  claim 19 , wherein the amino acid sequence of the GALR2 agonist is attached to NH2 on the C-terminus end.