TGF-alpha polypeptides, functional fragments and methods of use therefor

Abstract

Disclosed are peptides related to human TGF-α, having TGF-α biological activity, which are useful for many of the indications that full-length TGF-α polypeptide is useful. Also provided are methods of use of such peptides, as well as human TGF-α and biologically related polypeptides. For example, methods for treating or preventing cachexia in subjects are provided as well as methods for stimulating hematopoiesis in patients undergoing cytotoxic chemotherapy. In addition, the use of TGF-α related peptides to related neurodengenerative diseases is also provided. Methods of the invention also provide protection for patients undergoing cytotoxic therapy from side effects such as gastrointestinal (GI) mucositis.

What is claimed:  
     
         1 . A polypeptide comprising a peptide having a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys-COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         2 . The polypeptide of  claim 1 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4: -X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         3 . The polypeptide of  claim 2 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         4 . The polypeptide of  claim 2 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         5 . The polypeptide of  claim 2 , wherein the functional peptide is 18 amino acids in length and X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         6 . A method of treating a subject having or at risk of exhibiting weight-loss comprising administering to the subject a transfonning growth factor-alpha (TGF-α) polypeptide in an effective amount to inhibit or reduce weight-loss in the subject.  
     
     
         7 . The method of  claim 6 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:1.  
     
     
         8 . The method of  claim 6 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:3.  
     
     
         9 . The method of  claim 6 , wherein the polypeptide comprises a peptide having a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         10 . The method of  claim 9 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X6-X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         11 . The method of  claim 10 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         12 . The method of  claim 10 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         13 . The method of  claim 10 , wherein the functional peptide is 18 amino acids in length and X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         14 . The method of  claim 6 , wherein the weight-loss is the result of a disease or disorder.  
     
     
         15 . The method of  claim 6 , wherein the weight-loss is associated with treatment with of the subject with a chemotherapeutic agent.  
     
     
         16 . The method of  claim 15 , wherein the chemotherapeutic agent is selected from the group consisting of carmustine (BCNU), chlorambucil (Leukeran), cisplatin (Platinol), Cytarabine, doxorubicin (Adriamycin), fluorouracil (5-FU), methoxetrate (Mexate), taxol, CPT111, etoposide, and plicamycin (Mithracin).  
     
     
         17 . The method of  claim 15 , wherein the disease or disorder is ARC or AIDS.  
     
     
         18 . The method of  claim 6 , wherein the subject is a mammal.  
     
     
         19 . The method of  claim 18 , wherein the mammal is a human.  
     
     
         20 . A method of increasing the body weight of a subject comprising administering to the subject prior to, simultaneously with, or substantially following weight-loss a transforming growth factor-alpha (TGF-α) polypeptide in an effective amount to increase or maintain the weight of the subject.  
     
     
         21 . The method of  claim 20 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:1.  
     
     
         22 . The method of  claim 20 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:3.  
     
     
         23 . The method of  claim 20 , wherein the polypeptide comprises a peptide having a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         24 . The method of  claim 23 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X6 is Asp or Glu.  
     
     
         25 . The method of  claim 24 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         26 . The method of  claim 24 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         27 . The method of  claim 24 , wherein the functional peptide is 18 amino acids in length and X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         28 . The method of  claim 20 , wherein the weight-loss is the result of a disease or disorder.  
     
     
         29 . The method of  claim 20 , wherein the weight-loss is associated with treatment of the subject with a chemotherapeutic agent.  
     
     
         30 . The method of  claim 29 , wherein the chemotherapeutic agent is selected from the group consisting of carmustine (BCNU), chlorambucil (Leukeran), cisplatin (Platinol), Cytarabine, doxorubicin (Adriamycin), fluorouracil (5-FU), methoxetrate (Mexate), taxol, CPT111, etoposide, and plicamycin (Mithracin).  
     
     
         31 . The method of  claim 28 , wherein the disease or disorder is ARC or AIDS.  
     
     
         32 . The method of  claim 20 , wherein the subject is a mammal.  
     
     
         33 . The method of  claim 32 , wherein the mammal is a human.  
     
     
         34 . A pharmaceutical composition comprising a polypeptide having a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity, and a pharmaceutically acceptable carrier.  
     
     
         35 . The pharmaceutical composition of  claim 34 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X6-X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         36 . The pharmaceutical composition of  claim 34 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         37 . The pharmaceutical composition of  claim 35 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         38 . The pharmaceutical composition of  claim 35 , wherein the functional peptide is 18 amino acids in length and X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         39 . A method for treating a neurodegenerative disease in a subject comprising administering to the subject a transforming growth factor-alpha (TGF-α) polypeptide in an effective amount to modulate the neurodegenerative disease.  
     
     
         40 . The method of  claim 39 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:1.  
     
     
         41 . The method of  claim 39 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:3.  
     
     
         42 . The method of  claim 39 , wherein the polypeptide has a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         43 . The method of  claim 42 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         44 . The method of  claim 43 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         45 . The method of  claim 43 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         46 . The method of  claim 43 , wherein the functional peptide is 18 amino acids in length wherein X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         47 . A method for treating a CNS disease or disorder comprising administering to a subject a polypeptide having a sequence as set forth in SEQ ID NO:3, or a polypeptide having a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b  -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         48 . The method of  claim 47 , wherein the CNS disease or disorder is selected from the group consisting of CNS ischemia, spinal cord injury, MS, and retinal injury.  
     
     
         49 . The method of  claim 47 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         50 . The method of  claim 49 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         51 . The method of  claim 49 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         52 . The method of  claim 49 , wherein the functional peptide is 18 amino acids in length and wherein X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         53 . A method for enhancing hematopoiesis during cytotoxic or immune-suppressing therapy, comprising administering a TGFα polypeptide in an effective amount to enhance hematopoeisis.  
     
     
         54 . The method of  claim 53 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:1.  
     
     
         55 . The method of  claim 53 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:3.  
     
     
         56 . The method of  claim 53 , wherein the polypeptide has a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys-COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         57 . The method of  claim 56 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         58 . The method of  claim 57 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         59 . The method of  claim 57 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         60 . The method of  claim 57 , wherein the functional peptide is 18 amino acids in length wherein X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         61 . The method of  claim 53 , further comprising administering a second hematopoietic growth factor agent to stimulate the generation of more mature hematopoietic precursor cells, wherein the second hematopoietic growth factor is selected from the group consisting of erythropoietin, thrombopoietin, G-CSF (granulocyte colony stimulating factor), and GM-CSF (granulocyte macrophage colony stimulating factor).  
     
     
         62 . A method for treating or preventing mucositis of the gastrointestinal tract in a subject comprising administering a TGFα polypeptide in an amount effective to inhibit or prevent oral or intestinal mucositis in the subject.  
     
     
         63 . The method of  claim 62 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:1.  
     
     
         64 . The method of  claim 62 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:3.  
     
     
         65 . The method of  claim 62 , wherein the polypeptide has a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         66 . The method of  claim 65 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         67 . The method of  claim 66 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         68 . The method of  claim 66 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         69 . The method of  claim 66 , wherein the functional peptide is 18 amino acids in length wherein X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         70 . The method of  claim 62 , wherein the oral or intestinal mucositis results from cytotoxic or immune-suppressing therapy.  
     
     
         71 . The method of  claim 62 , wherein the mucositis results from ARC or AIDS.  
     
     
         72 . The method of  claim 62 , wherein the subject is a mammal.  
     
     
         73 . The method of  claim 72 , wherein the mammal is a human.  
     
     
         74 . A compound as shown in formula III:  
       loop peptide N-terminus-linker-cyclic C 4 H 8 N 2 -linker-loop peptide N-terminus   (III)  
       wherein the linker moiety is designed to link the N-terminus of the loop peptide to a nitrogen atom of the ring C 4 H 8 N 2  and wherein the loop peptide has a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity, wherein said compound functions as a TGF-α mimetic.  
     
     
         75 . The compound of  claim 74 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4: -X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         76 . The compound of  claim 75 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         77 . The compound of  claim 75 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         78 . The compound of  claim 75 , wherein the functional peptide is 18 amino acids in length wherein X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         79 . The compound of  claim 74 , wherein the linker group is independently selected from the group consisting of substituted or unsubstituted C 1-6  alkyl, substituted or unsubstituted C 2-6  alkenyl, substituted or unsubstituted C 1-6  alkoxy, xylenyl, wherein the substitutions are selected from the group consisting of oxo, epoxyl, hydroxyl, chloryl, bromyl, fluoryl, and amino.  
     
     
         80 . A method for treating inflammatory bowel disease, colitis, and Chron's Disease of the gastrointestinal tract, comprising administering a TGFα polypeptide in an effective amount to inhibit or ameliorate the disease.  
     
     
         81 . The method of  claim 80 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:1.  
     
     
         82 . The method of  claim 80 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:3.  
     
     
         83 . The method of  claim 80 , wherein the polypeptide has a sequence NH 2 -X 1a -Cys-His-Ser-X 1b - X 2 -X 1a -X 1b -X 1a - X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         84 . The method of  claim 83 , wherein at least one or more of the following amino acids are linked to the C-terminal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X 6  is Asp or Glu.  
     
     
         85 . The method of  claim 84 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         86 . The method of  claim 84 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         87 . The method of  claim 84 , wherein the functional peptide is 18 amino acids in length and wherein X 1a  is Val, X 1b  is Gly, X 1c  is Ala, and X 4  is Gly.  
     
     
         88 . A method for treating or preventing an inflammatory reaction of an autoimmune disease in a subject comprising administering a TGFα polypeptide to the subject in an effective amount to inhibit or ameliorate the inflammatory reaction.  
     
     
         89 . The method of  claim 88 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:1.  
     
     
         90 . The method of  claim 88 , wherein the polypeptide has an amino acid sequence as set forth in SEQ ID NO:3.  
     
     
         91 . The method of  claim 88 , wherein the polypeptide has a sequence NH 2 -X 1a -Cys-His-Ser-X 1b -X 2 -X 1a -X 1b -X 1a -X 3 -Cys COOH (SEQ ID NO:4) wherein X 1a  and X 1b  are independently Val, Gly or Ala; X 2  is Tyr or Phe; X 3  is Arg or Lys; and the two Cys moieties are linked via a disulfide bond to form an at least 11-amino acid functional peptide having TGF-α activity.  
     
     
         92 . The method of claim  91 , wherein at least one or more of the following amino acids are linked to the C-terninal Cys moiety of SEQ ID NO:4:-X 4 -His-X 1c -X 4 -X 5 -X 6 -X 1c  (SEQ ID NO:5) wherein X 4  is Glu or Asp; X 5  is Leu or Ile; and X6 is Asp or Glu.  
     
     
         93 . The method The method of claim  92 , wherein X 1a  is Val, X 1b  is Gly and X 1c  is Ala.  
     
     
         94 . The method of claim  92 , wherein X 2  is Tyr and X 3  is Arg.  
     
     
         95 . The method of claim  92 , wherein the functional peptide is 18 amino acids in length wherein X 1a  is Val, X 1b  is Gly, X 1c  is Ala and X 4  is Gly.  
     
     
         96 . The method of  claim 88 , wherein the autoimmune diseases are selected from the group consisting of Type II (Juvenile) Diabetes, rheumatoid arthritis, lupus, glomerular nephritis, and multiple sclerosis.  
     
     
         97 . The method of  claim 88 , wherein the subject is a mammal.  
     
     
         98 . The method of claim  97 , wherein the mammal is a human.