Chemistry: natural resins or derivatives; peptides or proteins; – Peptides of 3 to 100 amino acid residues – 15 to 23 amino acid residues in defined sequence
Reexamination Certificate
2000-01-28
2004-09-28
Bugaisky, Gabriele (Department: 1653)
Chemistry: natural resins or derivatives; peptides or proteins;
Peptides of 3 to 100 amino acid residues
15 to 23 amino acid residues in defined sequence
C530S327000, C530S324000, C530S855000, C514S013800, C514S014800
Reexamination Certificate
active
06797808
ABSTRACT:
BACKGROUND OF THE INVENTION
The invention relates to relatively short peptides (termed &agr;-conotoxins herein), about 10-30 residues in length, which are naturally available in minute amounts in the venom of the cone snails or analogous to the naturally available peptides, and which preferably include two disulfide bonds.
The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details respecting the practice, are incorporated by reference, and for convenience are referenced in the following text by author and date and are listed alphabetically by author in the appended bibliography.
The predatory cone snails (Conus) have developed a unique biological strategy. Their venom contains relatively small peptides that are targeted to various neuromuscular receptors and may be equivalent in their pharmacological diversity to the alkaloids of plants or secondary metabolites of microorganisms. Many of these peptides are among the smallest nucleic acid-encoded translation products having defined conformations, and as such, they are somewhat unusual. Peptides in this size range normally equilibrate among many conformations. Proteins having a fixed conformation are generally much larger.
The cone snails that produce these peptides are a large genus of venomous gastropods comprising approximately 500 species. All cone snail species are predators that inject venom to capture prey, and the spectrum of animals that the genus as a whole can envenomate is broad. A wide variety of hunting strategies are used; however, every Conus species uses fundamentally the same basic pattern of envenomation.
Several peptides isolated from Conus venoms have been characterized. These include the &agr;-, &mgr;- and &ohgr;-conotoxins which target nicotinic acetylcholine receptors, muscle sodium channels, and neuronal calcium channels, respectively (Olivera et al., 1985). Conopressins, which are vasopressin analogs, have also been identified (Cruz et al. 1987). In addition, peptides named conantokins have been isolated from
Conus geographus
and
Conus tulipa
(Mena et al., 1990; Haack et al., 1990).
The &agr;-conotoxins are small peptides highly specific for neuromuscular junction nicotinic acetylcholine receptors (Gray et al., 1981; Marshall and Harvey, 1990; Blount et al., 1992; Jacobsen et al., 1997) or highly specific for neuronal nicotinic acetylcholine receptors (Fainzilber et al., 1994; Johnson et al., 1995; Cartier et al., 1996; Luo et al., 1998). The &agr;-conotoxins with specificity for neuromuscular junction nicotinic acetylcholine receptors are used as neuromuscular blocking agents for use in conjunction with surgery, as disclosed in U.S. patent application Ser. No. 09/488,799, filed 21 Jan. 2000, incorporated by reference herein. Additional &agr;-conotoxins and uses for them have been described in U.S. Pat. No. 4,447,356 (Olivera et al., 1984); U.S. Pat. Nos. 5,432,155; 5,514,774, each incorporated herein by reference.
Additional uses for &agr;-conotoxins are described in U.S. Ser. No. 09/219,446, filed 22 Dec. 1998, incorporated herein by reference. In this application, &agr;-conotoxins with specificity for neuronal nicotinic acetylcholine receptors are used for treating disorders regulated at neuronal nicotinic acetylcholine receptors. Such disorders include, but are not limited to, cardiovascular disorders, gastric motility disorders, urinary incontinence, nicotine addiction, mood disorders (such as bipolar disorder, unipolar depression, dysthymia and seasonal effective disorder) and small cell lung carcinoma, as well as the localization of small cell lung carcinoma.
It is desired to provide additional &agr;-conotoxin peptides having uses as described herein.
SUMMARY OF THE INVENTION
The invention relates to relatively short peptides (termed &agr;-conotoxins herein), about 10-30 residues in length, which are naturally available in minute amounts in the venom of the cone snails or analogous to the naturally available peptides, and which preferably include two disulfide bonds.
More specifically, the present invention is directed to &agr;-conotoxin peptides having the general formula I:
Xaa
1
-Xaa
2
-Xaa
3
-Xaa
4
-Xaa
5
-Cys-Cys-Xaa
6
-Xaa
7
-Xaa
8
-Xaa
9
-Cys-Xaa
10
-Xaa
11
-Xaa
12
-Cys-Xaa
13
(SEQ ID NO1:), wherein Xaa
1
is des-Xaa
1
, Ile, Leu or Val; Xaa
2
is des-Xaa
2
, Ala or Gly; Xaa
3
is des-Xaa
3
, Gly, Trp (D or L), neo-Trp, halo-Trp or any unnatural aromatic amino acid; Xaa
4
is des-Xaa
4
, Asp, Phe, Gly, Ala, Glu, &ggr;-carboxy-Glu (Gla) or any unnatural aromatic amino acid; Xaa
5
is Glu, Gla, Asp, Ala, Thr, Ser, Gly, Ile, Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr or any unnatural hydroxy containing amino acid; Xaa
6
is Ser, Thr, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys or any unnatural basic amino acid; Xaa
7
is Asp, Glu, Gla, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys or any unnatural basic amino acid; Xaa
8
is Ser, Thr, Asn, Ala, Gly, Arg, Lys, ornithine, homoarginine, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys, any unnatural basic amino acid, His, halo-His, Pro or hydroxy-Pro; Xaa
9
is Thr, Ser, Ala, Asp, Pro, hydroxy-Pro, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys or any unnatural basic amino acid; Xaa
10
is Gly, Ser, Thr, Ala, Asn, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys or any unnatural basic amino acid; Xaa
11
is Gln, Leu, His, halo-His, Trp (D or L), halo-Trp, neo-Trp, Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys, any unnatural basic amino acid or any unnatural aromatic amino acid; Xaa
12
is Asn, His, halo-His, Ile, Leu, Val, Gln, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys or any unnatural basic amino acid; Xaa
13
is des-Xaa
13
, Val, Ile, Leu, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys or any unnatural basic amino acid. The C-terminus may contain a free carboxyl group or an amide group. The halo is chlorine, bromine or iodine, preferably iodine for Tyr and His and preferably bromine for Trp. The Cys residues may be in D or L configuration and may optionally be substituted with homocysteine (D or L). The Tyr residues maybe substituted with the 3-hydroxyl or 2-hydroxyl isomers and corresponding O-sulpho- and O-phospho-derivatives. The acidic amino acid residues may be substituted with any synthetic acidic bioisoteric amino acid surrogate, e.g., tetrazolyl derivatives of Gly and Ala.
More specifically, the present invention is directed to &agr;-conotoxin peptides having the general formula 11:
Xaa
1
-Xaa
2
-Xaa
3
-Xaa
4
-Cys-Cys-Xaa
5
-Xaa
6
-Xaa
7
-Xaa
8
-Cys-Xaa
9
-Xaa
10
-Xaa
11
-Xaa
12
-Xaa
13
-Xaa
14
-Cys-Xaa
15
-Xaa
16
-Xaa
17
(SEQ ID NO:2), wherein Xaa
1
is des-Xaa
1
, Asp, Glu or &ggr;-carboxy-Glu (Gla); Xaa
2
is des-Xaa
2
, Gln, Ala, Asp, Glu, Gla; Xaa
3
is des-Xaa
3
, Gly, Ala, Asp, Glu, Gla, Pro or hydroxy-Pro; Xaa
4
is des-Xaa
4
, Gly, Glu, Gla, Gln, Asp, Asn, Pro or hydroxy-Pro; Xaa
5
is Ser, Thr, Gly, Glu, Gla, Asn, Trp (D or L), neo-Trp, halo-Trp, Arg, ornithine, homoarginine, Lys, N-methyl-Lys, N,N-dimethyl-Lys, N,N,N-trimethyl-Lys, any unnatural basic amino acid, Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr or any unnatural hydroxy containing amino acid; Xaa
6
is Asp, Asn, His, halo-His, Thr, Ser, Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr or any unnatural hydroxy containing amino acid; Xaa
7
is Pro or hydroxy-Pro; Xaa
8
is Ala, Ser, Thr, Asp, Val, Ile, Pro, hydroxy-Pro, Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr or any unnatural hydroxy containing amino acid; Xaa
9
is Gly, Ile, Leu, Val, Ala, Thr, Ser, Pro, hy
Hillyard David R.
Jones Robert M.
McIntosh J. Michael
Olivera Baldomero M.
Watkins Maren
Bugaisky Gabriele
Rothwell Figg Ernst & Manbeck
University of Utah Research Foundation
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