Human CDC4 modulates cyclin E degradation
AssigneeThe Scripps Research Institute
InventorsSteve Reed, Charles Spruck, Heimo Strohmaier, Olle Sangfelt
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Abstract
The present invention provides compositions including polypeptides useful for regulating the cell cycle, proliferation, tumorigenesis, and genomic stability, mutants of such polypeptides useful in identifying abnormal cells and diagnosing disease, antibodies useful for identifying the polypeptides, and polynucleotides that encode the polypeptides.
Claims155
1) A composition comprising a purified polypeptide including an amino acid sequence set forth in SEQ ID NO:4.
5) A purified polypeptide comprising an amino acid sequence that is 95% or more identical to SEQ ID NO:2 and includes an activity substantially similar to SEQ ID NO:2.
12) A purified immunogenic composition comprising a polypeptide consisting essentially of 6 to 167 consecutive amino acid residues of SEQ ID NO:4.
16) A composition comprising a purified polypeptide including an amino acid sequence set forth in SEQ ID NO:10.
20) A purified polypeptide comprising an amino acid sequence that is 95% or more identical to SEQ ID NO:6 and includes an activity substantially similar to SEQ ID NO:6.
25) A purified immunogenic composition comprising a polypeptide consisting essentially of 6 to 87 consecutive amino acid residues of SEQ ID NO:6.
29) A purified immunogenic composition comprising a polypeptide consisting essentially of 6 to 49 consecutive amino acid residues of SEQ ID NO:16.
33) A purified immunogenic composition comprising a polypeptide consisting essentially of 6 to 20 consecutive amino acid residues of SEQ ID NO:14.
37) An isolated antibody that specifically binds to an hCdc4 α-exon 1 polypeptide.
41) An isolated antibody that specifically binds to an hCdc4 β-exon 1 polypeptide.
45) An isolated antibody that specifically binds to an hCdc4 γ-exon 1 polypeptide.
49) An isolated antibody that specifically binds to an amino acid sequence consisting essentially of SEQ ID NO:14, or an immunogenic fragment thereof.
51) A purified polynucleotide comprising a nucleic acid sequence encoding a polypeptide including SEQ ID NO:4, or a complement of the nucleic acid sequence.
59) A purified polynucleotide comprising a nucleic acid sequence encoding a polypeptide including SEQ ID NO:10, or a complement of the nucleic acid sequence.
65) A purified polynucleotide comprising a mutant α-hCdc4 polynucleotide including one or more mutations in a codon 124, 367, 371, 465, 472, 479, or 658 of the polynucleotide, wherein a wild-type α-hCdc4 polynucleotide comprises a nucleic acid sequence set forth in SEQ ID NO:1.
67) A purified polypeptide comprising a mutant α-hCdc4 polypeptide including a truncation mutation or a frame shift mutation in a C-terminal domain of the mutant α-hCdc4 polypeptide, wherein a wild-type α-hCdc4 polypeptide includes an amino acid sequence set forth in SEQ ID NO:2, and wherein a wild-type C. terminal domain includes a sequence set forth in SEQ ID NO:14.
72) A purified polynucleotide comprising a mutant β-hCdc4 polynucleotide including a mutation in a codon 23 of the polynucleotide, wherein a wild-type β-hCdc4 polynucleotide comprises a nucleic acid sequence set forth in SEQ ID NO:5.
74) A purified polypeptide comprising a mutant β-hCdc4 polypeptide including a truncation mutation or a frame shift mutation in a C-terminal domain of the mutant β-hCdc4 polypeptide, wherein a wild-type β-hCdc4 polypeptide includes an amino acid sequence set forth in SEQ ID NO:6, and wherein a wild-type C. terminal domain includes a sequence set forth in SEQ ID NO:14.
79) A purified polynucleotide comprising a nucleic acid sequence encoding a cyclin E polypeptide including a T62 mutation, wherein a wild-type cyclin E polypeptide includes an amino acid sequence set forth in SEQ ID NO:31 or SEQ ID NO:32.
86) A method of detecting an α-hCdc4 expression product in a sample, comprising specifically determining that the sample includes an α-exon 1 sequence, wherein the α-exon 1 sequence is a ribonucleic acid sequence or an amino acid sequence.
91) A method of detecting a β-hCdc4 expression product in a sample, comprising specifically determining that the sample includes a β-exon 1 sequence, wherein the β-exon 1 sequence is a ribonucleic acid sequence or an amino acid sequence.
96) A method of detecting a γ-hCdc4 expression product in a sample, comprising specifically determining that the sample includes a γ-exon 1 sequence, wherein the γ-exon 1 sequence is a ribonucleic acid sequence or an amino acid sequence.
101) A diagnostic system, comprising:
- a targeting agent that specifically binds to an α-exon 1 of an hCdc4 expression product and a detecting agent operably linked to the targeting agent.
106) A method of detecting a phosphorylated cyclin E in a sample, comprising:
- a) contacting the sample with an antibody that specifically binds to the phosphorylated cyclin E;
- and
- b) detecting a specific binding of the antibody, thereby detecting the phosphorylated cyclin E in the sample.
115) A purified polypeptide comprising an amino acid sequence CSLIPTPDKEDDDRV (SEQ ID NO:36), wherein the threonine residue at position six includes a phosphorylation.
117) A purified polypeptide comprising an amino acid sequence CSLIPTPDKEDDDRV (SEQ ID NO:37), wherein the threonine residue at position six does not include a phosphorylation.
119) A purified polypeptide comprising an amino acid sequence SGLLTPPQSGKK (SEQ ID NO:38), wherein the threonine residue at position five includes a phosphorylation.
121) A purified polypeptide comprising an amino acid sequence SGLLTPPQSGKK (SEQ ID NO:39), wherein the threonine residue at position five does not include a phosphorylation.
123) A purified polypeptide comprising an amino acid sequence CSGLLTPPQSGKK (SEQ ID NO:40), wherein the threonine residue at position six includes a phosphorylation.
125) A purified polypeptide comprising an amino acid sequence CSGLLTPPQSGKK (SEQ ID NO:41), wherein the threonine residue at position six does not include a phosphorylation.
127) A purified antibody that selectively and specifically binds a phosphorylated cyclin E polypeptide.
128) A purified antibody that selectively and specifically binds a non-phosphorylated cyclin E.
129) A purified antibody that specifically binds to a phosphorylated cyclin E and a non-phosphorylated cyclin E.
132) A purified antibody that specifically binds to a cyclin E polypeptide including a phosphorylated T62 residue, wherein the polypeptide includes 6 or more amino acid residues.
137) A purified antibody that specifically binds to a cyclin E polypeptide including a phosphorylated T380 residue, wherein the polypeptide includes 6 or more amino acid residues.
142) A method of detecting a tumor in a subject;
- comprising:
- a) contacting a sample of the subject with an antibody that specifically binds to a phosphorylated cyclin E;
- and
- b) detecting a specific binding of the antibody, thereby detecting the tumor in the subject.
147) A method of determining a ratio of a phosphorylated cyclin E to a non-phosphorylated cyclin E in a sample, comprising:
- a) measuring a specific binding of an anti-phospho-cyclin E antibody;
- b) measuring a specific binding of an anti-cyclin E antibody;
- and
- c) comparing the specific binding of the anti-phospho-cyclin E antibody to the specific binding of the anti-cyclin E antibody, thereby determining the ratio of the phosphorylated cyclin E to total cyclin E in the sample.
Dependencies are inferred from the claim text.