Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
  • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
  • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
  • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
  • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
  • A61K38/1709—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
  • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
  • A61K38/1709—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
  • A61K38/1751—Bactericidal/permeability-increasing protein [BPI]
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P17/00—Drugs for dermatological disorders
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • C07K14/001—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof by chemical synthesis

Definitions

• This invention relates to the treatment of r.ermatological malignancies, such as melanoma. More particularly, this invention relates to the treatment of •iermatological malignancies by administering a biologically active peptide or protein.
• a process for treating a der ⁇ atological malignancy in a host comprising administering to a host at least one biologically active amphiphilic peptide or protein.
• the peptide or protein is an ion channel-forming peptide or protein.
• the peptide or protein is administered in an amount effective to treat a dermatological malignancy in a host.
• treating a dertnatological malignancy means that the peptide or protein prevents, inhibits, or destroys the growth of a dermatological malignancy or skin cancer, and/or reduces the size of or eliminates the dermatological malignancy.
• An ion channel-forming peptide or protein or ionophore is a peptide or protein which increases the permeability for ions across a natural or synthetic lipid membrane.
• an ion channel-forming peptide or ion channel-forming protein is a peptide or protein which has ion channe1-forming properties as determined by the method of Christensen, et al.
• amphiphilic peptide or protein is a peptide or protein which includes both hydrophobic and hydrophilic peptide or protein regions.
• the ion channel-forming peptides employed in the present invention are generally water soluble to a concentration of at least 20 mg/ml at neutral pH in water.
• the structure of such peptide provides for flexibility of the peptide molecule.
• Such peptides are capable of forming an alpha-helix. When the peptide is placed in water, it does not assume an amphiphilic structure. When the peptide encounters an oily surface or membrane, the peptide chain folds upon itself into a rodlike structure.
• such peptides have at least 7 amino acids, and in many cases have at least 20 amino acids. In most cases, such peptides do not have in excess of 40 amino acids.
• the peptides or proteins are administered in an amount effective to treat a dermatological malignancy in a host.
• the peptides or proteins may be administered to a host in vivo, such as,
• the peptides or proteins may be administered intralesionally; i.e., the peptide or protein is injected directly into the dermatological malignancy.
• the peptide or protein may be administered in an amount of from about 50 ⁇ g per injection to about 100 mg per injection, preferably such that peptide dosages are from about 1 mg/kg to about 5 mg/kg body weight.
• the peptides or proteins may also be administered intravenously or intraperitoneally.
• the peptide or protein may be administered in multiple doses. In one embodiment, the peptide or protein may be administered two times a week for a period of U weeks.
• the peptide or protein may be administered topically in an amount of from about 1 mg/kg to about 5 mg/kg of body weight.
• the peptides or proteins are administered in combination with an acceptable pharmaceutical carrier or vehicle such as a filler, non-toxic buffer, or physiological saline solution.
• an acceptable pharmaceutical carrier or vehicle such as a filler, non-toxic buffer, or physiological saline solution.
• the peptides or proteins may also be used in combination with adjuvants, proteas p inhibitors, or compatible drugs.
• Dermatological malignancies which may be treated include, but are not limited to, melanoma, basal cell carcinoma, and squamous cell carcinoma.
• the dermatological malignancy may be metastatic or non-metastatic.
• the peptide is a basic (positively charged) polypeptide having at least sixteen amino acids wherein the polypeptide includes at least eight hydrophobic amino acids and at least eight hydrophilic amino acids.
• hydrophobic amino acids are in groups of two adjacent amino acids, and each group of two hydrophobic amino acids is spaced from another group of two hydrophobic amino acids by at least one amino acid other than a hydrophobic amino acid (preferably at least two amino acids) and generally by no greater than four amino acids, and the amino acids between pairs of hydrophobic amino acids may or may not be hydrophilic.
• the hydrophilic amino acids are generally also in groups of two adjacent amino acids in which at least one of the two amino acids is a basic hydrophilic amino acid, with such groups of two hydrophilic amino acids being spaced from each other by at least one amino acid other than a hydrophilic amino acid (preferably at least two amino acids) and generally no greater than four amino acids, and the amino acids between pairs of hydrophilic amino acids may or may not be hydrophobic.
• the polypeptide comprises a chain of at least four groups of amino acids, with each group consisting of four amino acids. Two of the four amino acids in each group are hydrophobic amino acids, and two of the four amino acids in each group are hydrophilic, with at least one of the hydrophilic amino acids in each group being a basic hydrophilic amino acid and the other being a basic or neutral hydrophilic amino acid.
• the hydrophobic amino acids may be selected from the class consisting of Ala, Cys, Phe, Gly, lie, Leu, Met, Pro, Val, Trp, Tyr, norleucine (Nle), norvaline (Nva), and cyclohexylalanine (Cha).
• the neutral hydrophilic amino acids may be selected from the class consisting of Asn, Gin, Ser, Thr and homoserine (Hse).
• the basic hydrophilic amino acids may be selected from the class consisting of Lys, Arg, His, Orn, homoarginine (Har), 2, 4-diaminobutyric acid (Dbu), and p-aminophenylalanine.
• Each of the groups of four amino acids may be of the sequence ABCD, BCDA, CDAB, or DABC, wherein A and B are each hydrophobic amino acids and may be the same or different, one of C or D is a basic hydrophilic amino acid, and the other of C or D is a basic or neutral hydrophilic amino acid and may be the same or different.
• the polypeptide chain may comprise 5 or 6 groups of this sequence. In each group, each of A, B, C and D may be the same in some or all of the groups or may be different in some or all of the groups.
• the polypeptide chain preferably has at least 20 amino acids, and no greater than 50 amino acids. It is to be understood, however, that the polypeptide does not have to consist entirely of the groups described above.
• the polypeptide may have amino acids extending from either or both ends of the noted groups forming the polypeptide chain and/or there may be amino acids between one or more of the at least four groups and still remain within the scope of the invention.
• the groups of amino acids may be repeating groups of amino acids, or the amino acids in the various groups may vary provided that in each group of the at least four groups of amino acids there are two hydrophobic and two hydrophilic amino acids as hereinabove noted.
• the biologically active polypeptide may comprise a chain including at least four groups of amino acids, each containing four amino acids. Two of the four amino acids in each group are hydrophobic, at least one amino acid is basic hydrophilic, and the remaining one is basic or neutral hydrophilic, with the polypeptide chain preferably having at least 20 amino acids but no greater than 50 amino acids.
• each of the at least four groups of amino acids which are in the peptide chain is of the sequence A-B-C-D, B-C-D-A, C-D-A-B or D-A-B-C wherein A and B are hydrophobic amino acids, one of C or D is a basic hydrophilic amino acid, and the other of C or D is basic or neutral hydrophilic amino acid.
• the resulting polypeptide chain may have one of the following sequences:
• X 2 is A-, D-A- or C-D-A-
• Y is -B, -B-C or B-C-D
• X 3 is B-, A-B-, D-A-B-
• Y 3 is -C, -C-D, -C-D-A
• Y A is -D, -D-A, -D-A-B a is 0 or 1; b is 0 or 1 and n is at least 4.
• the peptide chain may include Amino acids between the hereinabove noted groups of four amino acids provided that the spacing between such groups and the charge on the amino acids does not change the characteristics of the peptide chain which provide amphiphilicity and a positive charge and do not adversely affect the folding characteristics of the chain to that which is significantly different from one in which the hereinabove noted group of four amino acids are not spaced from each other.
• the peptide may have amino acids extending from either end of the chain.
• the chains may have a Ser-Lys sequence before the "Ala” end, and/or an Ala-Phe sequence after the "Lys" end.
• Other amino acid sequences may also be attached to the "Ala” and/or the "Lys" end.
• the chain may have, for example, a C-D sequence before the first A-B-C-D group.
• other amino acid sequences may be attached to the "A" and/or the "D" end of one of these polypeptide chains.
• the peptide may be a magainin peptide.
• a magainin peptide is either a magainin such as magainin I, II or III or an analogue or derivative thereof.
• the magainin peptides preferably include the following basic peptide structure X 17 :
• R ir R i R 12 ⁇ R 13 R ir R 14 "R 12 "R ⁇ " R 14 "R 12 'R i R ir R n "R 14a “(R 15 ) n “R 14a “R 14 '”
• R . is a hydrophobic amino acid
• R._ is a basic hydrophilic amino acid
• R-_ is a hydrophobic, neutral hydrophilic, or basic hydrophilic amino acid
• R-, and j » fl are hydrophobic or basic hydrophilic amino acids
• R _ is glutamic acid or aspartic acid, or a hydrophobic or a basic hydrophilic amino acid
• n is 0 or 1.
• R._ is a hydrophobic or neutral hydrophilic amino acid
• R , is a hydrophobic amino acid
• R.. is glutamic acid or aspartic acid.
• a magainin peptide may include the following structure:
• R. . , R. - , R- , and , ⁇ are as previously defined.
• a magainin peptide may also have the following structure:
• R- where R , is a basic hydrophilic amino acid or asparagine or glutamine.
• R.--R where R _ is a neutral hydrophilic amino acid, a hydrophobic amino acid, or a basic hydrophilic amino acid.
• R _ is a neutral hydrophilic amino acid.
• a magainin peptide may also have the following structure:
• the magainin peptides may also include the following basic peptide structure X,,:
• R are amino acids as hereinabove described. 14a
• the magainin peptide may also include the following structure X-.-Z..; wherein X., is the hereinabove described basic peptide structure and Z trustee is R ll V (R ll n- (R ll n- R 14a n- ( R 15 ) n- ( R 14a n- R 14>n "
• R 16 n- (R 17>n
• R ll' R 14' R 14a' R 15' R 16' ⁇ R 17 are as hereinabove described, and n is 0 or 1, and each n may be the same or different.
• the magainin peptides generally include at least fourteen amino acids and may include up to forty amino acids.
• a magainin peptide preferably has 22 or 23 amino acids. Accordingly, the hereinabove described basic peptide structures of a magainin peptide may include additional amino acids at the amino end or at the carboxyl end, or at both ends.
• magainin peptides having the following primary sequences as given in the accompanying sequence listing as well as appropriate analogues and derivatives thereof:
• Magainin peptides are described in Proc. Natl. Acad Sci. Vol. 84 pp. 5449-53 (Aug. 87).
• the term "magainin peptides" as used herein refers to the basic magainin structure as well as derivatives and analogs thereof, including but not limited to the representative derivatives or analogs.
• the peptide may be a PGLa peptide or an XPF peptide.
• a PGLa peptide is either PGLa or an analogue or derivative thereof.
• the PGLa peptides preferably include the following basic peptide structure X-, :
• the PGLa peptides generally include at least seventeen amino acids and may include as many as forty amino acids. Accordingly, the hereinabove described basic peptide structure for a PGLa peptide may include additional amino acids at the amino end or at the carboxyl end or at both the amino and carboxyl end.
• a PGLa peptide may have the following structure:
• PGLa peptide may also have the following structure:
• a PGLa peptide may also have the following structure:
• a is 0 or 1 and b is 0 or 1.
• An XPF peptide is either XPF or an analogue or derivative thereof.
• the XPF peptides preferably include the following basic peptide structure X .:
• R ll' R 12' R 14' R 15 ⁇ nd R 17 ⁇ re &S P reviousl y defined and R is glutamine or asparagine or a basic lo hydrophilic, or hydrophobic amino acid and, n is 0 or 1.
• the XPF peptides generally include at least nineteen amino acids and may include up to forty amino acids. Accordingly, the hereinabove described basic peptide structure of XPF may include additional amino acids at the amino end, or at the carboxyl end or at both the amino and carboxyl ends.
• an XPF peptide may include the following structure:
• An XPF peptide may include the following structure:
• An XPF peptide may also have the following structure:
• X.-, Y., and Z. are as previously defined: a is 0 or 1 and b is 0 or 1.
• XPF or PGLa peptides which are characterized by the following primary amino acid sequences as given in the accompanying sequence listing: PGLa : (SEQ ID NO: 12) (NH 2 )
• the peptide may be a CPF peptide or appropriate analogue or derivative thereof.
• CPF peptides as well as analogues and derivatives thereof are herein sometimes referred to collectively as CPF peptides.
• the CPF peptide may be one which includes the following basic peptide structure X- n :
• R_ is a hydrophobic amino acid
• R Dust amount is a hydrophobic amino acid or a basic hydrophilic amino acid
• R_ is a basic hydrophilic amino acid
• R Pain is a hydrophobic or neutral hydrophilic amino acid
• R__ is a basic or neutral hydrophilic amino acid.
• the hereinabove basic structure is hereinafter symbolically indicated as X ?f ..
• the hydrophobic amino acids are Ala, Cys, Phe, Gly, lie, Leu, Met, Val, Trp, Tyr, norleucine (Nle), norvaline (Nva), and cyclohexylalanine (Cha).
• the neutral hydrophilic amino acids are Asn, Gin, Ser, Thr, and homoserine (Hse).
• the basic hydrophilic amino acids are Lys, Arg, His, Orn, homoarginine (Har), 2,4-diarainobutyric acid (Dbu), and p-aminophenylalanine.
• the CPF peptide may include only the hereinabove noted amino acids or may include additional amino acids at the amino and/or carboxyl end or both the amino and carboxyl end. In general, the peptide does not include more than 40 amino acids.
• the CPF peptides including the above basic structure preferably have from 1 to 4 additional amino acids at the amino end.
• R 22 _R 25 ' (ii) R 22 _R 25 '; or (iii) R 2l “R 22 ⁇ R 25 ; ° r (iv) R 22 "R 2l 'R 22 “R 25 ; P referabl y Glycine - R 21 "R 2 2 ⁇ R 25" wherein R_-, R 2 hail and R_,- are as previously defined.
• the carboxyl end of the basic peptide structure may also have additional amino acids which may range from 1 to 13 additional amino acids.
• the basic structure may have from 1 to 7 additional amino acids at the carboxyl end, which may be represented as follows:
• X is the hereinabove defined basic peptide structure and Z 2Q is
• R 21 -R 21 -R 24 -R 24 -R 26 -Gln-Gln wherein R 2J and R , are as previously defined, and R , is proline or a hydrophobic amino acid.
• Preferred peptides may be represented by the following structural formula
• X 2n> Y ?n and Z_. are as previously defined and a is 0 or 1 and b is 0 or 1.
• CPF peptides which may be employed, some of which have been described in the literature, include the following sequences as given in the accompanying sequence listing:
• the peptide may include one of the following basic structures X_- through X__ wherein:
• X 31 is "[R 3 R 32 "R 32 'R 33 "R 3l 'R 32 “R 32 ]” n
• the basic hydrophilic amino acids may be selected from the class consisting of Lys, Arg, His, Orn, homoarginine (Har), 2,4-diaminobutyric acid (Dbu), and p-aminophenylalanine.
• the hydrophobic amino acids may be selected from the class consisting of Ala, Cys, Phe, Gly, lie, Leu, Met, Pro, Val, Trp and Tyr.norleucine (Nle), norvaline (Nva), and cyclohexylalanine (Cha).
• the neutral hydrophilic amino acids may be selected from the class consisting of Asn, Gin, Ser, Thr, and h ⁇ moserine (Hse).
• the peptide when the peptide includes the structure X-., the peptide may include the following structure:
• the peptide when the peptide includes the structure Xo, .
• the peptide may include the following structure:
• the peptide may include the following structure:
• the peptide may include the following structure:
• the peptide when the peptide includes the structure X , the peptide may include the following structure:
• the peptide may include the following structure:
• the peptide when the peptide includes the structure X- > - j. the peptide may include the following structure:
• R 32 (i) R 32 ; (ii) R 31 -R 32 ; (iii) R 32 - R 31 - R 32 ; (iv) R 32 -R 32 - R 31 - 32 ;
• R 3l R 32 "R 32 “R 3r R 32 ; ° r (vi) 3 3 - 31 - 3 2 - R 32- R 3 R 32' wherein R 3 1> R 32' " ⁇ R _ are as hereinabove described.
• the peptide when the peptide includes the structure X.,-,- the peptide may include the following structure:
• the peptide may include the following structure:
• the peptide when the peptide includes the structure X , the peptide may include the following structure:
• the peptide when the peptide includes the structure X- .
• the peptide may include the following structure:
• X 3_4,-Z3_4, wherein X3_4, is as hereinabove described, and Z_. is: 34
• the peptide may include the following structure:
• the peptide when the peptide includes the structure X _, the peptide may include the following structure:
• Y is as hereinabove described, and Y 35 is:
• the peptide when the peptide includes the structure X,_, the peptide may include the following structure:
• the peptide may include the following structure:
• the peptide when the peptide includes the structure X : , the peptide may
• Jo include the following structure:
• the peptide when the peptide includes the structure X--., the peptide may include the following structure:
• the peptide may include the following structure:
• the peptide when the peptide includes the structure X, 7 , the peptide may includes the structure Y__-X__, wherein X route is as hereinabove described, and Y,_ is: (l) R 32 ; (ID R 31 -R 32 ; (iii) R 33 - 31 - R 32 ; (iv) R 32 - R 33 - R 31 - R 32 ;
• the peptide when the peptide includes the structure X__, the peptide may include the following structure:
• the peptide may include the following structure:
• n 3
• peptide is of one of the following structures as given in the accompanying sequence listing:
• Lys He Ala (Lys He Ala Gly Xaa He Ala) 3 (SEQ ID NO:68). Lys He Ala (Lys He Ala Gly Lys He Ala) (SEQ ID NO:69) In (SEQ ID NO:67) and (SEQ ID NO:68), Xaa is p-aminophenylalanine.
• amphiphilic peptide includes the following basic structure X, • 40
• the peptide may include the following structure:
• the peptide may include the following structure:
• peptide has the following structural formula as given in the accompanying sequence listing:
• the peptide has the following structural formula as given in the accompanying sequence listing: (SEQ ID NO: 71)
• the peptide has one of the one of the following structural formulae as given in the accompanying sequence listing:
• the peptide may include the following structural formula:
• n is from 2 to 5.
• n is 3, and the peptide has the following structural formula:
• the peptide may include the following structural formula:
• n is from 2 to 5.
• n 3
• the peptide has the following structural formula:
• the peptide may include the following structural formula:
• n is from 2 to 5.
• n is 3, and the peptide has the following structural formula:
• the peptide may be selected from the group consisting of the following structural formulae as given in the accompanying sequence listing:
• the peptide may be a cecropin or sarcotoxin.
• cecropins includes the basic structure as well as analogues and derivatives thereof.
• the cecropins and analogues and derivatives thereof are described in Ann. Rev. Microbiol. 1987, Vol. 41, pages 103-26, in particular page 108, and in Christensen, et al., PNAS Vol. 85, pgs. 5072-76, which are hereby incorporated by reference.
• sarcotoxins includes the basic materials as well as analogues and derivatives thereof.
• the sarcotoxins and analogues and derivatives thereof are described in Molecular Entomology, pages 369-78, in particular page 375, Alan R. Liss, Inc. (1987), which is hereby incorporated by reference.
• amphiphilic peptide includes the following basic structure X cn :
• R 4 is a hydrophobic amino acid, and R 4 is a basic hydrophilic or neutral hydrophilic amino acid.
• the peptide includes the basic structure Y_ -X c - wherein X_ ⁇ is as hereinabove described 50 50 and Y 50 is:
• R is leucine.
• R,_ is lysine.
• Representative examples of peptides in accordance with this aspect of the present invention include those having the following structures: (SEQ ID NO: 95 ) (SEQ ID NO: 96) (SEQ ID NO: 97) (SEQ ID NO: 98)
• the amphiphilic peptide includes the following basic structure X
• R is a hydrophobic amino acid and R 4 is a basic hydrophilic or neutral hydrophilic amino acid.
• R. - is leucine.
• R,_ is lysine.
• the peptide includes the basic structure Y_ -X. , wherein X_ ? is as hereinabove described, and Y_ is:
• the peptide may have the following structure
• the peptide includes the basic structure X_ breathe - Z_. isolate , wherein X-.,. is as hereinabove described, and Z_. is:
• the peptide may have the following structure:
• the peptide may include the structure:
• the peptide includes the following basic structure X_,:
• 41 42 4J is a netural hydrophilic amino acid.
• the peptide may have the following structure:
• the peptide may have the following structure:
• the peptide includes the following basic structure X cfi :
• the peptide may include the following structure Y_,-X_,, wherein X c - is the basic
• JO O O peptide structure hereinabove described, and Y_, is: (i) -R 41 ( ii) - R 41 - R 4 f.
• the peptide may include the structure:
• the peptide may have one of the following structures:
• the peptide may have the structure (YJ c O,)a-XJ,-,O- . ⁇ (ZJ c O,).D, wherein XJ_O,, YJ r O_ , and ZJ_O, are as hereinabove described, a is 0 or 1, and b is 0 or 1.
• the peptide includes the following basic structure cR :
• the peptide may include the structure Y, R -X_ R , wherein X. ⁇ is as hereinabove described, and Y CD is:
• the peptide includes the structure X J_o-Z5-8 repeat, wherein X5 r 8 ⁇ is as hereinabove described, and Z_ R is:
• the peptide has the following structure:
• the peptide may have the structure
• the peptide includes the following basic structure X fin ;
• the peptide may have the following structure:
• the peptide may include the structure X ⁇ n 'Z ⁇ r ,, wherein X-., is as hereinabove oU oU oU described, and Z, n is:
• the peptide has a structure selected from the group consisting of:
• the peptide has the structure
• the peptide has the structure (b), and a representative example of such a structure is (SEQ ID NO: 108), which is given in the accompanying sequence listing.
• the peptide has the structure (c), and a representative example of such a structure is (SEQ ID NO: 109) as given the accompanying sequence listing.
• the peptide has the structure (d), and a representative example of such a structure is (SEQ ID NO: 110) as given in the accompanying sequence listing.
• the peptide has the structure (e), and representative examples of such a structure are (SEQ ID NO: 111) and (SEQ ID NO: 112) as given in the accompanying sequence listing.
• the peptide has the following structural formula:
• the peptide is elittin.
• Melittin is an amphipathic peptide consisting of 26 amino acid residues, and is isolated from honeybee (Apis mellifera) venom. The peptide is known to be cytolytic.
• Melittin has the following structural formula as represented by the three-letter amino acid code: Gly He Gly Ala Val Leu Lys Val Leu
• the peptide purified in accordance with the present invention is an apidaecin.
• apidaecin as used herein includes the basic structure as well as analogues and derivaties thereof. Apidaecins are further described in European Patent Application No. 299,828.
• the peptide may be an amide - or carboxy-terminated peptide represented by the following structural formula, and the numbers below each amino acid residue refer to the position of the residue in the peptide:
• the peptide may be an analogue of such peptide wherein at least one of amino acid residues 1 through 7, 9, 11, 12, 14, 16, or 18 is deleted from the peptide.
• amino acid residues 1, 3, 7, 9, 11, 12, 14, 16, or 18 is deleted from the peptide.
• amino acid residues 1 through 3, 1 through 4, 1 through 5, 1 through 6, and 1 through 7 are deleted from the peptide.
• amino acid residues 1 through 3 or 1 through 4 are deleted from the peptide, and such peptides have the following structural formulae: (SEQ ID NO: 116) (SEQ ID NO: 117)
• the peptide includes the following structural formula X-_:
• R 4 R 42 _R 42 ' wherein R, - is a hydrophobic amino acid, and R,_ is a basic hydrophilic or neutral hydrophilic amino acid.
• R, is leucine, and in another embodiment, R, is lysine.
• the peptide has the following structure:
• the peptide includes the following structural formula X,,:
• R 4l R 41, wherein R,- is a hydrophobic amino acid, and R,_ is a basic hydrophilic or neutral hydrophilic amino acid.
• R 4 is leucine, and in another embodiment, R,. is lysine.
• the peptide has the following structural formula:
• the peptide includes the following structural formula X,,: oo
• R 4 R 4 R 2 "R 2 "R R 2 'R 2 "R R 4 R 42 “R 42 “R 4l “R r wherein R 4 _ is a hydrophobic amino acid, and R 42 is a basic hydrophilic or neutral hydrophilic amino acid.
• the peptide may include the following structure:
• R,. is leucine
• R, 2 is lysine
• the peptide has the following structural formula: Leu Leu Lys Lys Leu Lys Lys Leu Leu Lys Lys Leu Leu Lys Lys Leu Leu Lys
• amphiphilic peptide or protein may be an ion channel-forming peptide or protein.
• Ion channel-forming proteins or peptides which may be employed include defensins, also known as human neutrophil antimicrobial peptides (HNP), major basic protein (MBP) of eosinophils, bactericidal permeability-increasing protein (BPI), and a pore-forming cytotoxin called variously perforin, cytolysin, or pore-forming protein.
• HNP human neutrophil antimicrobial peptides
• MBP major basic protein
• BPI bactericidal permeability-increasing protein
• a pore-forming cytotoxin called variously perforin, cytolysin, or pore-forming protein.
• Defensins are described in Selsted, et al., J. Clin. Invest. , Vol. 76, pgs. 1436-1439 (1985).
• MBP proteins are described in Wasmoen, et al., J. Biol. Chem..
• BPI proteins are described in Ooi, et al, > Biol. Chem.. Vol. 262, pgs. 14891-14894 (1987). Perforin is described in Henkart, et al., J. Exp. Med., 160: 75 (1984), and in Podack, et al., J. Exo. Med.. 160:695 ( 1984) . The above articles are hereby incorporated by reference.
• ion channel-forming proteins includes the basic structures of the ion channel-forming proteins as well as analogues and derivatives.
• the peptides may be produced by known techniques and obtained in substantially pure form.
• the peptides may be synthesized on an automatic peptide synthesizer. Journal of the American Chemical Society. Vol. 85, pgs. 2149-54 (1963). It is also possible to produce such peptides by genetic engineering techniques.
• the codons encoding the amino acids are known to those skilled in the art, and thus one may construct DNA encoding any of the peptides by accepted techniques, and clone such DNA into an expression vehicle such as, for example, a plasmid, and transfeet such an expression vehicle into a cell which will express the peptide.
• transfected cells which include an expression vehicle to a host containing DNA encoding ion channel-forming pe ⁇ tide(s) or protein(s) such as those hereinabove described.
• peptides or proteins may be administered in combination with one another.
• n accor ance w ano er em o men , ne pep e or protein may be administered in combination with an ion having pharmacological properties.
• An ion having pharmacological properties is one which when introduced into a target cell, inhibits and/or prevents and/or destroys the growth of the target cell.
• Such an ion having pharmacological properties is one which in the absence of an ion channel-forming peptide or protein is unable to cross a natural or synthetic membrane; in particular a cell membrane, in sufficient amounts to affect a cell adversely.
• the peptide or protein and ion having pharmacological properties may be administered as a single composition or in separate compositions, and the single or separate compositions may include additional materials, active and/or inactive, in addition to the peptide or protein and ion having pharmacological properties.
• additional materials active and/or inactive, in addition to the peptide or protein and ion having pharmacological properties.
• ions having pharmacological properties there may be mentioned fluoride, peroxide, bicarbonate, silver, zinc, mercury, copper, platinum, antimony, gold, nickel, selenium, and bismuth ions.
• the peptide or protein and ion having pharmacological properties are employed in amounts effective to inhibit and/or prevent and/or destroy the growth of the .. dermatological malignancy.
• the ion potentiates the action of the peptide, i.e., the amount of the ion is effective to reduce the maximum effective concentration of the peptide or protein for inhibiting growth of a dermatological malignancy.
• the ion having pharmacological properties when used systemically (e.g., intravenously, intraperitoneally, intralesionally), is generally employed in an amount of from 1 mg to 10 mg per kg of host weight.
• Peptide or protein dosages may be within the ranges hereinabove described.
• the ion may be administered in the form of a salt such as, for example, sodium fluoride.
• peptides or proteins may be administered in combination with chemotherapeutic agents such as, but not limited to, cyclophosphamide, cisplatin, d ⁇ xorubicin, hexamethylamine, and VP-16.
• chemotherapeutic agents such as, but not limited to, cyclophosphamide, cisplatin, d ⁇ xorubicin, hexamethylamine, and VP-16.
• WM239-A metal melanoma
• WM278 primary melanoma
• WM793 primary melanoma
• WM1158 metal melanoma
• Peptides SEQ ID NO: 121), (SEQ ID NO: 122), D-(SEQ ID NO: 121), in which each amino acid residue is a D-amino residue or a glycine residue, and a "control" peptide having the structure of Magainin II (SEQ ID NO:7), in which the Phe and Lys residues are D-amino acid residues, were frozen at -20°C in 100 ml aliquots at a concentration of 25 mg/ml.
• Each peptide was then dissolved in sterile distilled water at a dilution of 1:1,000 or 1:5,000 to provide peptide concentrations of 25 ⁇ g/ml or 5 ⁇ g/ml, respectively.
• the peptides at the 25 ⁇ g/ral or 5 ⁇ g/ml concentrations were then added to the plates containing the melanoma cells. The amount of
• Peptides (SEQ ID NO: 117) and D-(SEQ ID NO: 117), in which each amino acid residue is a D-amino acid residue, are tested for % growth reduction of melanoma cell lines WM852 (grown in 2% tumor media or PF media), WM983A (grown in 2% tumor media or PF media), WM793 (grown in 2% tumor media), and WM1985 (grown in PF media) according to the procedure of Example 1, except that only a concentration of 25 ⁇ g/ml was employed for (SEQ ID NO: 117), and concentrations of 1.0, 2.5, 5.0, 10.0, and 25.0 ⁇ g/ml were employed for D-(SEQ ID NO: 117). The results are given in Table II below.
• the following melanoma cell lines are seeded in 200 ul of 2.0%
• ADDRESSEE Carella, Byrne, Bain, Gilfillan,
• Gly lie Gly Lys Phe Leu His Ser Ala Gly
• NAME/KEY Magainin II peptide.
• NAME/KEY magainin peptide
• NAME/KEY magainin peptide
• NAME/KEY magainin peptide

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• 1993
  • 1993-12-03 JP JP6513508A patent/JPH08507749A/ja active Pending
  • 1993-12-03 EP EP94903514A patent/EP0671939A1/en not_active Withdrawn
  • 1993-12-03 CA CA002150946A patent/CA2150946A1/en not_active Abandoned
  • 1993-12-03 AU AU57431/94A patent/AU5743194A/en not_active Abandoned
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