JP2003526317A - Secreted proteins and polynucleotides encoding them - Google Patents

Abstract

(57) Abstract Disclosed are novel polynucleotides and proteins encoded by them.

Description

Detailed Description of the Invention

FIELD OF THE INVENTION The present invention provides novel polynucleotides and proteins encoded by such polynucleotides, as well as therapeutic, diagnostic and research utilities for these polynucleotides and proteins.

BACKGROUND proteinaceous factors invention (e.g., lymphokines, interferons, including cytokine such as CSFs and interleukins) has methods for the purpose of finding this 10
Has matured rapidly over the years. Traditional hybridization and expression cloning methods nowadays use information directly related to the discovered protein (ie,
In the case of hybridization cloning, it depends on the partial DNA / amino acid sequence of the protein; in the case of expression cloning)
The new polypeptide is cloned "directly". Signal sequence cloning (
More recent “indirect” cloning methods, such as the isolation of DNA sequences based on the presence of the now well-recognized secretory leader sequence motif), as well as various PCR or low stringency hybridization cloning methods, , A large number of DNA / amino acid sequences for proteins known to have biological activity due to their secreted nature in the case of cloning of the leader sequence or by the cell or tissue source in the case of the PCR method. Have made progress in the present situation. The present invention is directed to these proteins and the polynucleotides that encode them.

SUMMARY OF THE INVENTION In an embodiment of 1, the invention provides a composition comprising an isolated polynucleotide selected from the group consisting of: (a) comprising the nucleotide sequence of SEQ ID NO: 1. (B) a polynucleotide comprising the nucleotide sequence from nucleotide 257 to nucleotide 622 of SEQ ID NO: 1; (c) the clone ff168 1 deposited under accession number ATCC 98623.
A polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of 2; (d) clone ff168_1 deposited under accession number ATCC 98623.
A polynucleotide encoding the full-length protein encoded by the cDNA insert of 2; (e) clone ff168_1 deposited under accession number ATCC 98623.
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of 2; (f) clone ff168_1 deposited under accession number ATCC 98623.
A polynucleotide encoding a mature protein encoded by the cDNA insert of 2; (g) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 2; (h) a SEQ ID NO: having biological activity : A polynucleotide encoding a protein containing a fragment of the amino acid sequence of SEQ ID NO: 2 (the fragment is 8 of SEQ ID NO: 2).
(I) a polynucleotide which is an allelic variant of the polynucleotide of (a)-(f) above; (j) a species homologue of the protein of (g) or (h) above. A coding polynucleotide; and (k) a polynucleotide capable of hybridizing to any one of the polynucleotides (a) to (h) under stringent conditions. Preferably, such a polynucleotide is nucleotide 257 of SEQ ID NO: 1.
To the nucleotide sequence from nucleotide 622; accession number ATCC 98623
Nucleotide sequence of full-length protein coding sequence of clone ff16812 deposited as :; or clone f deposited under accession number ATCC 98623.
It comprises the nucleotide sequence of the mature protein coding sequence of f16812. In another preferred embodiment, the polynucleotide encodes the full-length or mature protein encoded by the cDNA insert of clone ff16812 deposited under accession number ATCC 98623. In a further preferred embodiment, the invention provides a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 2 having biological activity, said fragment being SEQ ID NO: 2, preferably 8, More preferably, it comprises 20 and most preferably 30 contiguous amino acids, or a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 2 having biological activity (the fragment being SEQ ID NO: 2).
2 amino acid 56 to amino acid 65). Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 1. A further embodiment of the invention provides a method of producing an isolated polynucleotide, which method is selected from the group consisting of: (a) a method comprising the steps of: (i) 65 in 6X SSC. Preparing one or more polynucleotide probes which hybridize at 0 ° C. to a nucleotide sequence selected from the group consisting of: (aa) SEQ ID NO: 1 (polynucleotide at the 3 ′ end of SEQ ID NO: 1) (A) excluding tail); and (ab) clone ff168 12 deposited as ATCC 98623.
The nucleotide sequence of the cDNA insert of (ii) hybridizing the probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by the probe (s); and (B) a method comprising the following steps: (i) preparing one or more polynucleotide primers that hybridize in 6X SSC at 65 ° C. to a nucleotide sequence selected from the group consisting of: ba) SEQ ID NO: 1 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 1); and (bb) clone ff168 12 deposited as ATCC 98623.
Sequence of the cDNA insert of (ii) hybridizing the primer (s) to human DNA; (iii) amplifying the human DNA sequence; and then (iv) the polynucleotide of step (b) (iii). Isolate the product. Preferably, the nucleotide sequence of the polynucleotide isolated by this method corresponds to the cDNA sequence of SEQ ID NO: 1, from the nucleotide sequence corresponding to the 5'end of SEQ ID NO: 1 to the 3'of SEQ ID NO: 1. The poly (A) tail at the 3'end of SEQ ID NO: 1 is excluded, but is continuous up to the nucleotide sequence corresponding to the end. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to nucleotides 257 to 622 of the cDNA sequence of SEQ ID NO: 1, nucleotide 257 of SEQ ID NO: 1.
To the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 622 to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 257 to nucleotide 622 of SEQ ID NO: 1. In another embodiment, the present invention provides a composition comprising a protein, wherein the protein comprises (a) an amino acid sequence of SEQ ID NO: 2; (b) a fragment of the amino acid sequence of SEQ ID NO: 2 (each fragment is (Comprising 8 consecutive amino acids of SEQ ID NO: 2); and (c) clone ff168_1 deposited under accession number ATCC 98623.
A composition containing an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts of 2 and containing substantially no other mammalian proteins is provided. Preferably such protein comprises the amino acid sequence of SEQ ID NO: 2. In a further preferred embodiment, the invention provides
A protein containing a fragment of the amino acid sequence of SEQ ID NO: 2 having biological activity (
Said fragment comprises a contiguous amino acid sequence of SEQ ID NO: 2, preferably 8, more preferably 20, most preferably 30 contiguous amino acids) or a biologically active amino acid sequence of SEQ ID NO: 2. A protein is provided that comprises the amino acid sequence of amino acid 56 to amino acid 65 of SEQ ID NO: 2.

In one embodiment, the invention provides a composition comprising an isolated polynucleotide selected from the group consisting of: (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 3; (B) a polynucleotide comprising the nucleotide sequence from nucleotide 1323 to nucleotide 1829 of SEQ ID NO: 3; (c) a polynucleotide comprising the nucleotide sequence from nucleotide 1539 to nucleotide 1829 of SEQ ID NO: 3; (d) accession number ATCC 98623 A polynucleotide containing the nucleotide sequence of the full-length protein coding sequence of clone ls91 deposited as: (e) c of clone ls91 deposited under accession number ATCC 98623.
A polynucleotide encoding the full-length protein encoded by the DNA insert; (f) a polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of clone ls91 deposited under accession number ATCC 98623; (g) deposit under accession number ATCC 98623 Of cloned clone ls91
A polynucleotide encoding a mature protein encoded by the DNA insert; (h) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 4; (i) a biologically active SEQ ID NO: 4 A polynucleotide encoding a protein containing a fragment of the amino acid sequence of SEQ ID NO: 4
Individual contiguous amino acids); (j) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; (k) a species homologue of the protein of (h) or (i) above. (1) a polynucleotide capable of hybridizing to any one of the polynucleotides shown in (a) to (i) under strict conditions. Preferably, such a polynucleotide is nucleotide 132 of SEQ ID NO: 3.
Nucleotide sequence from nucleotide 3 to nucleotide 1829; nucleotide sequence from nucleotide 1539 to nucleotide 1829 of SEQ ID NO: 3; accession number AT
It contains the nucleotide sequence of the full length protein coding sequence of clone Is91 deposited as CC98623; or the mature protein coding sequence of clone Is91 deposited under accession number ATCC 98623. In another preferred embodiment, the polynucleotide encodes the full-length or mature protein encoded by the cDNA insert of clone ls91 deposited under accession number ATCC 98623. In a further preferred embodiment, the invention provides
A polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 4 having biological activity (the fragment is preferably 8 of SEQ ID NO: 4, more preferably 20 and most preferably 30). Polynucleotide containing a contiguous amino acid sequence, or a protein having a fragment of the amino acid sequence of SEQ ID NO: 4 having biological activity (the fragment is amino acid 79 to amino acid 88 of SEQ ID NO: 4). (Including sequences). Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 3. A further embodiment of the invention provides a method of producing an isolated polynucleotide, which method is selected from the group consisting of: (a) a method comprising the steps of: (i) 65 in 6X SSC. Preparing one or more polynucleotide probes which hybridize at ° C to a nucleotide sequence selected from the group consisting of: (aa) SEQ ID NO: 3 (polynucleotide at the 3'end of SEQ ID NO: 3 (A) excluding tail); and (ab) cd of clone ls91 deposited as ATCC 98623.
Nucleotide sequence of NA insert (ii) hybridizing said probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by said probe (s); and b) a method comprising the steps of: (i) preparing one or more polynucleotide primers that hybridize at 65 ° C. in 6X SSC to a nucleotide sequence selected from the group consisting of: (ba ) SEQ ID NO: 3 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 3); and (bb) cD of clone ls91 deposited as ATCC 98623.
Nucleotide sequence of NA insert (ii) hybridizing said primer (s) to human DNA; (iii) amplifying human DNA sequence; and then (iv) producing polynucleotide of step (b) (iii). Isolate things. Preferably, the nucleotide sequence of the polynucleotide isolated by this method corresponds to the cDNA sequence of SEQ ID NO: 3, from the nucleotide sequence corresponding to the 5'end of SEQ ID NO: 3 to the 3'of SEQ ID NO: 3. The poly (A) tail at the 3'end of SEQ ID NO: 3 is excluded, except the nucleotide sequence corresponding to the end is contiguous. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to nucleotides 1323 to 1829 of the cDNA sequence of SEQ ID NO: 3, nucleotide 1 of SEQ ID NO: 3.
It is a continuous sequence from the nucleotide sequence corresponding to the 5'end of the sequence from 323 to nucleotide 1829 to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 1323 to nucleotide 1829 of SEQ ID NO: 3.
Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method is from nucleotide 1539 to nucleotide 18 of the cDNA sequence of SEQ ID NO: 3.
29, which corresponds to the 5'end of the sequence from nucleotide 1539 to nucleotide 1829 of SEQ ID NO: 3 to nucleotide 3 of the sequence from nucleotide 1539 to nucleotide 1829 of SEQ ID NO: 3.
'It is a continuous sequence up to the nucleotide sequence corresponding to the end. In another embodiment, the present invention provides a composition comprising a protein, wherein the protein comprises (a) an amino acid sequence of SEQ ID NO: 4; (b) a fragment of the amino acid sequence of SEQ ID NO: 4 (each fragment is (Comprising 8 consecutive amino acids of SEQ ID NO: 4); and (c) c of clone ls91 deposited under accession number ATCC 98623.
A composition comprising an amino acid sequence selected from the group consisting of amino acid sequences encoded by DNA inserts and substantially free of other mammalian proteins is provided. Preferably such protein comprises the amino acid sequence of SEQ ID NO: 4. In a further preferred embodiment, the invention provides
A protein containing a fragment of the amino acid sequence of SEQ ID NO: 4 having biological activity (
Said fragment comprises a contiguous amino acid sequence of SEQ ID NO: 4, preferably 8, more preferably 20, most preferably 30 or alternatively a biologically active fragment of the amino acid sequence of SEQ ID NO: 4. A protein is provided that comprises the amino acid sequence of amino acid 79 to amino acid 88 of SEQ ID NO: 4.

In one embodiment, the invention provides a composition comprising an isolated polynucleotide selected from the group consisting of: (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 5; (B) a polynucleotide containing the nucleotide sequence from nucleotide 507 to nucleotide 722 of SEQ ID NO: 5; (c) a polynucleotide containing the nucleotide sequence from nucleotide 615 to nucleotide 722 of SEQ ID NO: 5; (d) accession number ATCC 98623 Clone 1010 deposited as
A polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of 1; (e) clone na1010 deposited under accession number ATCC 98623.
A polynucleotide encoding the full-length protein encoded by the cDNA insert of 1; (f) clone na1010 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of 1; (g) clone na1010 deposited under accession number ATCC 98623.
A polynucleotide encoding a mature protein encoded by the cDNA insert of 1; (h) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 6; (i) a SEQ ID NO having biological activity A polynucleotide encoding a protein containing a fragment of the amino acid sequence of SEQ ID NO: 6 (the fragment is 8 of SEQ ID NO: 6).
Individual contiguous amino acids); (j) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; (k) a species homologue of the protein of (h) or (i) above. (1) a polynucleotide capable of hybridizing to any one of the polynucleotides shown in (a) to (i) under strict conditions. Preferably such polynucleotide is nucleotide 507 of SEQ ID NO: 5.
To nucleotide 722; the nucleotide sequence of nucleotides 615 to 722 of SEQ ID NO: 5; accession number ATCC98
623 containing the nucleotide sequence of the full-length protein coding sequence of clone na10101; or the mature protein coding sequence of clone na10101 deposited with accession number ATCC 98623.
In another preferred embodiment, the polynucleotide has the accession number ATCC 9862.
3 encodes the full-length or mature protein encoded by the cDNA insert of clone na10101. In a further preferred embodiment,
The present invention provides a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 6 having biological activity (the fragment is SEQ ID NO: 6, preferably 8, and more preferably 20). Most preferably it comprises 30 contiguous amino acids, or a polynucleotide encoding a protein comprising a biologically active fragment of the amino acid sequence of SEQ ID NO: 6, said fragment comprising amino acid 31 to amino acid of SEQ ID NO: 6. Including up to 40 amino acid sequences). Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 5. A further embodiment of the invention provides a method of producing an isolated polynucleotide, which method is selected from the group consisting of: (a) a method comprising the steps of: (i) 65 in 6X SSC. Preparing one or more polynucleotide probes which hybridize at 0 ° C. to a nucleotide sequence selected from the group consisting of: (aa) SEQ ID NO: 5 (polynucleotide at the 3'end of SEQ ID NO: 5 (A) excluding tails); and (ab) clone na10101 deposited as ATCC 98623.
The nucleotide sequence of the cDNA insert of (ii) hybridizing the probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by the probe (s); and (B) a method comprising the following steps: (i) preparing one or more polynucleotide primers that hybridize in 6X SSC at 65 ° C. to a nucleotide sequence selected from the group consisting of: ba) SEQ ID NO: 5 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 5); and (bb) clone na10101 deposited as ATCC 98623.
Sequence of the cDNA insert of (ii) hybridizing the primer (s) to human DNA; (iii) amplifying the human DNA sequence; and then (iv) the polynucleotide of step (b) (iii). Isolate the product. Preferably, the nucleotide sequence of the polynucleotide isolated by this method corresponds to the cDNA sequence of SEQ ID NO: 5, from the nucleotide sequence corresponding to the 5'end of SEQ ID NO: 5 to the 3'of SEQ ID NO: 5. The poly (A) tail at the 3'end of SEQ ID NO: 5 is excluded, but is continuous up to the nucleotide sequence corresponding to the end. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to nucleotides 507 to 722 of the cDNA sequence of SEQ ID NO: 5, nucleotide 507 of SEQ ID NO: 5.
To nucleotide 722 from the nucleotide sequence corresponding to the 5'end of the sequence to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 507 to nucleotide 722 of SEQ ID NO: 5. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to nucleotides 615 to 722 of the cDNA sequence of SEQ ID NO: 5, from nucleotide 615 to nucleotide 722 of SEQ ID NO: 5. From the nucleotide sequence corresponding to the 5'end of the sequence to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 615 to nucleotide 722 of SEQ ID NO: 5. In another embodiment, the present invention provides a composition comprising a protein, wherein the protein comprises (a) an amino acid sequence of SEQ ID NO: 6; (b) a fragment of the amino acid sequence of SEQ ID NO: 6 (each fragment is (Comprising 8 consecutive amino acids of SEQ ID NO: 6); and (c) clone na1010 deposited under accession number ATCC 98623.
1. A composition comprising an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA insert of 1 and containing substantially no other mammalian proteins. Preferably such protein comprises the amino acid sequence of SEQ ID NO: 6. In a further preferred embodiment, the invention provides
A protein containing a fragment of the amino acid sequence of SEQ ID NO: 6 having biological activity (
The fragment comprises a contiguous amino acid sequence of SEQ ID NO: 6, preferably 8, more preferably 20, most preferably 30 or an amino acid sequence of SEQ ID NO: 6 having biological activity. And a protein comprising the amino acid sequence of SEQ ID NO: 6 from amino acid 31 to amino acid 40.

[0006] In one embodiment, the invention provides a composition comprising an isolated polynucleotide selected from the group consisting of: (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 7; (B) a polynucleotide containing the nucleotide sequence from nucleotide 673 to nucleotide 978 of SEQ ID NO: 7; (c) a polynucleotide containing the nucleotide sequence from nucleotide 868 to nucleotide 987 of SEQ ID NO: 7; (d) accession number ATCC 98623 A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence of clone nf87 1 deposited as: (e) a polynucleotide encoding the full-length protein encoded by the cDNA insert of clone nf87 1 deposited under accession number ATCC 98623. (F) a polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of clone nf87 1 deposited under accession number ATCC 98623; (g) the mature protein encoded by the cDNA insert of clone nf87 1 deposited under accession number ATCC 98623. (H) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 8; (i) a protein containing a fragment of the amino acid sequence of SEQ ID NO: 8 having biological activity. A polynucleotide encoding (the fragment is 8 of SEQ ID NO: 8)
Individual contiguous amino acids); (j) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; (k) a species homologue of the protein of (h) or (i) above. (1) a polynucleotide capable of hybridizing to any one of the polynucleotides shown in (a) to (i) under strict conditions. Preferably, such a polynucleotide is nucleotide 673 of SEQ ID NO: 7.
To nucleotide 987; nucleotide sequence from nucleotide 868 to nucleotide 987 of SEQ ID NO: 7; accession number ATCC98
Nucleotide sequence of the full-length protein coding sequence of clone nf87 1 deposited as 623; or the mature protein coding sequence of clone nf87 1 deposited as accession number ATCC 98623. In another preferred embodiment, the polynucleotide encodes the full-length or mature protein encoded by the cDNA insert of clone nf87_1 deposited under accession number ATCC 98623. In a further preferred embodiment, the invention provides a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 8 having biological activity (wherein the fragment is SEQ ID NO: 8, preferably 8; More preferably, it comprises 20 consecutive amino acids, most preferably 30 consecutive amino acids, or a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 8 having biological activity (the fragment being SEQ ID NO: 8 amino acid 47 to amino acid 56). Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 7. A further embodiment of the invention provides a method of producing an isolated polynucleotide, which method is selected from the group consisting of: (a) a method comprising the steps of: (i) 65 in 6X SSC. Preparing one or more polynucleotide probes which hybridize at 0 ° C. to a nucleotide sequence selected from the group consisting of: (aa) SEQ ID NO: 7 (polynucleotide at the 3'end of SEQ ID NO: 7 (A) excluding tails); and (ab) c of clone nf87 1 deposited as ATCC 98623.
Nucleotide sequence of the DNA insert: (ii) hybridizing the probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by the probe (s); and b) a method comprising the steps of: (i) preparing one or more polynucleotide primers that hybridize at 65 ° C. in 6X SSC to a nucleotide sequence selected from the group consisting of: (ba ) SEQ ID NO: 7 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 7); and (bb) c of clone nf871 deposited as ATCC 98623.
Nucleotide sequence of the DNA insert (ii) hybridizing the primer (s) to human DNA; (iii) amplifying the human DNA sequence; and then (iv) producing the polynucleotide of step (b) (iii). Isolate things. Preferably, the nucleotide sequence of the polynucleotide isolated by this method corresponds to the cDNA sequence of SEQ ID NO: 7, from the nucleotide sequence corresponding to the 5'end of SEQ ID NO: 7 to the 3'of SEQ ID NO: 7. The poly (A) tail at the 3'end of SEQ ID NO: 7 is excluded, but is continuous up to the nucleotide sequence corresponding to the end. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to nucleotides 673 to 987 of the cDNA sequence of SEQ ID NO: 7, nucleotide 673 of SEQ ID NO: 7.
To the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 987 to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 673 to nucleotide 987 of SEQ ID NO: 7. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to nucleotides 868 to 987 of the cDNA sequence of SEQ ID NO: 7, from nucleotide 868 to nucleotide 987 of SEQ ID NO: 7. From the nucleotide sequence corresponding to the 5'end of the sequence to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 868 to nucleotide 987 of SEQ ID NO: 7. In another embodiment, the present invention provides a composition comprising a protein, wherein the protein comprises (a) an amino acid sequence of SEQ ID NO: 8; (b) a fragment of the amino acid sequence of SEQ ID NO: 8 (each fragment is (Comprising 8 contiguous amino acids of SEQ ID NO: 8); and (c) comprising an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts of clone nf87_1 deposited under accession number ATCC 98623. And a composition that is substantially free of other mammalian proteins. Preferably such protein comprises the amino acid sequence of SEQ ID NO: 8. In a further preferred embodiment, the invention provides
A protein containing a fragment of the amino acid sequence of SEQ ID NO: 8 having biological activity (
Said fragment comprises a contiguous amino acid sequence of SEQ ID NO: 8, preferably 8, more preferably 20, most preferably 30 or alternatively biologically active amino acid sequence of SEQ ID NO: 8. A protein is provided that comprises the amino acid sequence of amino acid 47 to amino acid 56 of SEQ ID NO: 8.

[0007] In one embodiment, the invention provides a composition comprising an isolated polynucleotide selected from the group consisting of: (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 9; (B) a polynucleotide containing the nucleotide sequence from nucleotide 57 to nucleotide 824 of SEQ ID NO: 9; (c) a polynucleotide containing the nucleotide sequence from nucleotide 114 to nucleotide 824 of SEQ ID NO: 9; (d) accession number ATCC 98623. Clone nh7961 deposited as
A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence of: (e) clone nh796_1 deposited under accession number ATCC 98623
A polynucleotide encoding the full-length protein encoded by the cDNA insert of: (f) clone nh7961 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of: (g) clone nh7961 deposited under accession number ATCC 98623.
(H) a polynucleotide encoding a protein comprising the amino acid sequence of SEQ ID NO: 10; (i) a biologically active SEQ ID NO: A polynucleotide encoding a protein comprising a fragment of the amino acid sequence of 10 (the fragment being SEQ ID NO: 10
(J) an allelic variant of the polynucleotide of (a) to (g) above; (k) a species homology of the protein of (h) or (i) above. A polynucleotide encoding the body; and (l) a polynucleotide capable of hybridizing under stringent conditions to any one of the polynucleotides shown in (a) to (i). Preferably, such a polynucleotide is the nucleotide sequence of SEQ ID NO: 9 from nucleotide 57 to nucleotide 824; SEQ ID NO: 9, nucleotide 1
Nucleotide sequence from 14 to nucleotide 824; accession number ATCC 986
23, the nucleotide sequence of the full-length protein coding sequence of clone nh796 1 deposited as No. 23; or the mature protein coding sequence of clone nh796 1 deposited under accession number ATCC 98623. In another preferred embodiment, the polynucleotide encodes the full-length or mature protein encoded by the cDNA insert of clone nh796_1 deposited under accession number ATCC 98623. In a further preferred embodiment, the invention provides a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 10, which has biological activity, wherein said fragment is SEQ ID NO: 10, preferably 8, More preferably, it comprises 20 consecutive amino acids, most preferably 30 contiguous amino acids, or a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 10 having biological activity (the fragment being SEQ ID NO: 10 amino acids 123 to amino acid 132) are provided). Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 9. A further embodiment of the invention provides a method of producing an isolated polynucleotide, which method is selected from the group consisting of: (a) a method comprising the steps of: (i) 65 in 6X SSC. Preparing one or more polynucleotide probes which hybridize at ° C to a nucleotide sequence selected from the group consisting of: (aa) SEQ ID NO: 9 (polynucleotide at the 3'end of SEQ ID NO: 9 (A) excluding the tail); and (ab) the nucleotide sequence of the cDNA insert of clone nh796_1 deposited as ATCC 98623 (ii) hybridizing the probe (s) to human DNA; then (iii) Isolating a DNA polynucleotide detected by the probe (s); and (b) A method comprising the steps of: (i) preparing one or more polynucleotide primers which hybridize in 6X SSC at 65 ° C. to a nucleotide sequence selected from the group consisting of: (ba) sequences No. 9 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 9); and (bb) the nucleotide sequence of the cDNA insert of clone nh7961 deposited as ATCC 98623. (ii) the primer (multiple (Iii)); (iii) amplifying the human DNA sequence; and then (iv) isolating the polynucleotide product of step (b) (iii). Preferably, the nucleotide sequence of the polynucleotide isolated by this method corresponds to the cDNA sequence of SEQ ID NO: 9, from the nucleotide sequence corresponding to the 5'end of SEQ ID NO: 9 to the 3'of SEQ ID NO: 9. The poly (A) tail at the 3'end of SEQ ID NO: 9 is excluded, but is continuous up to the nucleotide sequence corresponding to the end. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to nucleotides 57 to 824 of the cDNA sequence of SEQ ID NO: 9, from nucleotide 57 to nucleotide 824 of SEQ ID NO: 9. From the nucleotide sequence corresponding to the 5'end of the sequence:
It is a contiguous sequence from nucleotide 57 to nucleotide 824 of SEQ ID NO: 9 up to the nucleotide sequence corresponding to the 3'end of the sequence. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method corresponds to the cDNA sequence of nucleotide 114 to nucleotide 824 of SEQ ID NO: 9, the nucleotide sequence of nucleotide 114 to nucleotide 824 of SEQ ID NO: 9. From the nucleotide sequence corresponding to the 5'end of the sequence, nucleotide 1 of SEQ ID NO: 9
It is a contiguous sequence from 14 to nucleotide 824 up to the nucleotide sequence corresponding to the 3'end of the sequence. In another embodiment, the present invention provides a composition comprising a protein, wherein the protein is (a) an amino acid sequence of SEQ ID NO: 10; (b) a fragment of the amino acid sequence of SEQ ID NO: 10 (each fragment is (Comprising 8 consecutive amino acids of SEQ ID NO: 10); and (c) clone nh796 1 deposited under accession number ATCC 98623.
A composition comprising an amino acid sequence selected from the group consisting of amino acid sequences encoded by the cDNA inserts of 1. and substantially free of other mammalian proteins. In a further preferred embodiment, the invention provides a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 10, which has biological activity, wherein said fragment is SEQ ID NO: 10, preferably 8, more preferably 20, Preferably, it comprises 30 consecutive amino acids), or a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 10 having biological activity (the fragment comprises the amino acid sequence of amino acid 123 to amino acid 132 of SEQ ID NO: 10). Including).

[0008] In one embodiment, the invention provides a composition comprising an isolated polynucleotide selected from the group consisting of: (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 11; (B) a polynucleotide containing the nucleotide sequence from nucleotide 297 to nucleotide 542 of SEQ ID NO: 11; (c) a polynucleotide containing the nucleotide sequence from nucleotide 510 to nucleotide 542 of SEQ ID NO: 11; (d) accession number ATCC 98623. Clone nn229 1 deposited as
A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence of: (e) clone nn229_1 deposited under accession number ATCC 98623
A polynucleotide encoding the full-length protein encoded by the cDNA insert of: (f) clone nn229_1 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of: (g) clone nn229_1 deposited under accession number ATCC 98623.
(H) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 12; (i) a biologically active SEQ ID NO: A polynucleotide encoding a protein comprising a fragment of the amino acid sequence of 12 (the fragment being SEQ ID NO: 12).
(J) an allelic variant of the polynucleotide of (a) to (g) above; (k) a species homology of the protein of (h) or (i) above. A polynucleotide encoding the body; and (l) a polynucleotide capable of hybridizing under stringent conditions to any one of the polynucleotides shown in (a) to (i). Preferably, such a polynucleotide is nucleotide 29 of SEQ ID NO: 11.
7 to nucleotide 542; nucleotide sequence from nucleotide 510 to nucleotide 542 of SEQ ID NO: 11; accession number ATCC
The nucleotide sequence of the full length protein coding sequence of clone nn229_1 deposited as 98623; or the mature protein coding sequence of clone nn229_1 deposited as accession number ATCC 98623.
In another preferred embodiment, the polynucleotide has the accession number ATCC 9862.
3 encodes the full-length or mature protein encoded by the cDNA insert of clone nn229_1. In a further preferred embodiment, the invention provides a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 12 having biological activity, said fragment being SEQ ID NO: 12.
Of the amino acid sequence of SEQ ID NO: 12, which preferably comprises 8, contiguous amino acids of 8, more preferably 20, and most preferably 30 consecutive amino acids. (The fragment comprises the amino acid sequence of amino acid 36 to amino acid 45 of SEQ ID NO: 12). Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 11. A further embodiment of the invention provides a method of producing an isolated polynucleotide, which method is selected from the group consisting of: (a) a method comprising the steps of: (i) 65 in 6X SSC. Preparing one or more polynucleotide probes which hybridize at 0 ° C. to a nucleotide sequence selected from the group consisting of: (aa) SEQ ID NO: 11 (polynucleotide at the 3 ′ end of SEQ ID NO: 11) (A) excluding the tail); and (ab) the nucleotide sequence of the cDNA insert of clone nn229_1 deposited as ATCC 98623. (ii) hybridizing the probe (s) to human DNA; then (iii) Isolating a DNA polynucleotide detected by the probe (s); and ( ) A method comprising the following steps: (i) preparing one or more polynucleotide primers which hybridize in 6X SSC at 65 ° C. to a nucleotide sequence selected from the group consisting of: (ba) SEQ ID NO: 11 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 11); and (bb) the nucleotide sequence of the cDNA insert of clone nn229_1 deposited as ATCC 98623 (ii) the primer for human DNA ( Hybridizing one or more); (iii) amplifying a human DNA sequence; and then (iv) isolating the polynucleotide product of step (b) (iii). Preferably, the nucleotide sequence of the polynucleotide isolated by this method corresponds to the cDNA sequence of SEQ ID NO: 11, 5'of SEQ ID NO: 11.
It is a continuous sequence from the nucleotide sequence corresponding to the end to the nucleotide sequence corresponding to the 3'end of SEQ ID NO: 11, but the poly (
A) The tail is removed. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method is nucleotide 29 of the cDNA sequence of SEQ ID NO: 11.
7 to nucleotide 542, and from the nucleotide sequence corresponding to the 5'end of said sequence from nucleotide 297 to nucleotide 542 of SEQ ID NO: 11, from nucleotide 297 to nucleotide 542 of SEQ ID NO: 11
Up to the nucleotide sequence corresponding to the 3'end of the sequence up to. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method is the cDNA of nucleotide 510 to nucleotide 542 of SEQ ID NO: 11.
From the nucleotide sequence corresponding to the NA sequence and corresponding to the 5'end of the sequence from nucleotide 510 to nucleotide 542 of SEQ ID NO: 11 to 3 of the sequence from nucleotide 510 to nucleotide 542 of SEQ ID NO: 11. 'It is a continuous sequence up to the nucleotide sequence corresponding to the end. In another embodiment, the present invention provides a composition containing a protein, wherein the protein comprises (a) an amino acid sequence of SEQ ID NO: 12; (b) a fragment of the amino acid sequence of SEQ ID NO: 12 (each fragment is (Comprising 8 consecutive amino acids of SEQ ID NO: 12); and (c) clone nn229_1 deposited under accession number ATCC 98623.
A composition comprising an amino acid sequence selected from the group consisting of amino acid sequences encoded by the cDNA inserts of 1. and substantially free of other mammalian proteins. Preferably such protein comprises the amino acid sequence of SEQ ID NO: 12. In a further preferred embodiment, the present invention provides a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 12 having biological activity, wherein said fragment is SEQ ID NO: 12, preferably 8 and more preferably 20.
, Most preferably 30 contiguous amino acids), or a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 12 having biological activity (the fragment comprising amino acid 36 to amino acid 45 of SEQ ID NO: 12). (Including an amino acid sequence).

In one embodiment, the invention provides a composition comprising an isolated polynucleotide selected from the group consisting of: (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 13; (B) a polynucleotide containing the nucleotide sequence from nucleotide 547 to nucleotide 750 of SEQ ID NO: 13; (c) a polynucleotide containing the nucleotide sequence from nucleotide 601 to nucleotide 750 of SEQ ID NO: 13; (d) accession number ATCC 98623 Clone np156 1 deposited as
A polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of: (e) clone np156_1 deposited under accession number ATCC 98623.
A polynucleotide encoding the full-length protein encoded by the cDNA insert of: (f) clone np156_1 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of: (g) clone np156_1 deposited under accession number ATCC 98623.
(H) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 14; (i) a biologically active SEQ ID NO: A polynucleotide encoding a protein comprising a fragment of the amino acid sequence of 14 (the fragment being SEQ ID NO: 14
(J) an allelic variant of the polynucleotide of (a) to (g) above; (k) a species homology of the protein of (h) or (i) above. A polynucleotide encoding the body; and (l) a polynucleotide capable of hybridizing under stringent conditions to any one of the polynucleotides shown in (a) to (i). Preferably, such a polynucleotide is nucleotide 54 of SEQ ID NO: 13.
The nucleotide sequence from 7 to nucleotide 750; the nucleotide sequence from nucleotide 601 to nucleotide 750 of SEQ ID NO: 13; accession number ATCC
Nucleotide sequence of the full-length protein coding sequence of clone np156_1 deposited as 98623; or the mature protein coding sequence of clone np156_1 deposited under accession number ATCC 98623.
In another preferred embodiment, the polynucleotide has the accession number ATCC 9862.
3 encodes the full-length or mature protein encoded by the cDNA insert of clone np156_1. In a further preferred embodiment, the invention provides a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 14 having biological activity, said fragment being SEQ ID NO: 14.
Of the amino acid sequence of SEQ ID NO: 14 having a biologically active sequence of SEQ ID NO: 14), or preferably 8 and more preferably 20 and most preferably 30 consecutive amino acids. (The fragment comprises the amino acid sequence from amino acid 29 to amino acid 38 of SEQ ID NO: 14). Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 13. A further embodiment of the invention provides a method of producing an isolated polynucleotide, which method is selected from the group consisting of: (a) a method comprising the steps of: (i) 65 in 6X SSC. Preparing one or more polynucleotide probes that hybridize at 0 ° C. to a nucleotide sequence selected from the group consisting of: (aa) SEQ ID NO: 13 (polynucleotide at the 3 ′ end of SEQ ID NO: 13) (A) excluding the tail); and (ab) the nucleotide sequence of the cDNA insert of clone np156_1 deposited as ATCC 98623. (ii) hybridizing the probe (s) to human DNA; then (iii) Isolating a DNA polynucleotide detected by the probe (s); and ( ) A method comprising the following steps: (i) preparing one or more polynucleotide primers which hybridize in 6X SSC at 65 ° C. to a nucleotide sequence selected from the group consisting of: (ba) SEQ ID NO: 13 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 13); and (bb) the nucleotide sequence of the cDNA insert of clone np156_1 deposited as ATCC 98623. (ii) the primer to human DNA ( Hybridizing one or more); (iii) amplifying a human DNA sequence; and then (iv) isolating the polynucleotide product of step (b) (iii). Preferably, the nucleotide sequence of the polynucleotide isolated by this method corresponds to the cDNA sequence of SEQ ID NO: 13, 5'of SEQ ID NO: 13.
It is a continuous sequence from the nucleotide sequence corresponding to the end to the nucleotide sequence corresponding to the 3'end of SEQ ID NO: 13, but the poly (
A) The tail is removed. Also preferably, the nucleotide sequence of the polynucleotide isolated by the above method is nucleotide 54 of the cDNA sequence of SEQ ID NO: 13.
7 to nucleotide 750, from the nucleotide sequence corresponding to the 5'end of said sequence from nucleotide 547 to nucleotide 750 of SEQ ID NO: 13, from nucleotide 547 to nucleotide 750 of SEQ ID NO: 13
Up to the nucleotide sequence corresponding to the 3'end of the sequence up to. In another preferred embodiment, the nucleotide sequence of the isolated polynucleotide corresponds to nucleotides 601 to 750 of the cDNA sequence of SEQ ID NO: 13, nucleotides 601 to 750 of SEQ ID NO: 13. Up to the nucleotide sequence corresponding to the 5'end of the sequence up to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 601 to nucleotide 750 of SEQ ID NO: 13. In another embodiment, the invention provides a composition comprising a protein, wherein the protein comprises (a) the amino acid sequence of SEQ ID NO: 14; (b) 8 consecutive amino acids of SEQ ID NO: 14. , A fragment of the amino acid sequence of SEQ ID NO: 14; and (c) clone np156_1 deposited under accession number ATCC 98623.
A composition comprising an amino acid sequence selected from the group consisting of amino acid sequences encoded by the cDNA inserts of 1. and substantially free of other mammalian proteins. Preferably such protein comprises the amino acid sequence of SEQ ID NO: 14. In a further preferred embodiment, the invention provides a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 14 having biological activity, said fragment being SEQ ID NO: 14, preferably 8 and more preferably 20.
, Most preferably 30 contiguous amino acids), or a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 14 having biological activity (the fragment comprising amino acid 29 to amino acid 38 of SEQ ID NO: 14). (Including an amino acid sequence).

In certain preferred embodiments, the polynucleotide is operably linked to expression control sequences. The invention also provides host cells transformed with such polynucleotide compositions, including bacterial, yeast, insect and mammalian cells. Also provided by the present invention are organisms that have enhanced, reduced, or modified expression of the genes corresponding to the polynucleotide sequences disclosed herein. Furthermore, a method for producing a protein, comprising: (a) growing a culture of host cells transformed with such a polynucleotide composition in a suitable medium; and (b) purifying the protein from the culture. A method consisting of is also provided. The protein produced by such a method is also provided by the present invention. Preferred specific examples include those in which the protein produced by such a method is a mature protein. The protein composition of the present invention may further contain a pharmaceutically acceptable carrier. Compositions containing antibodies that specifically react with such proteins are also provided by the present invention. There is also provided a method for the prevention, treatment or amelioration of a medical condition comprising administering to a mammalian subject a therapeutically effective amount of a composition comprising a protein of the invention and a pharmaceutically acceptable carrier.

Detailed Description Isolated Proteins and Polynucleotides The nucleotide and amino acid sequences for each of the clones and proteins disclosed herein are reported below. The actual nucleotide sequence of each clone can be easily determined by sequencing the deposited clones by known methods. The deduced amino acid sequence (both full-length as well as mature) can then be determined from such nucleotide sequences. The amino acid sequences of the proteins encoded by the individual clones can also be determined by expressing the clones in a suitable host cell, collecting the protein, and then determining its sequence. For each of the disclosed proteins, Applicants have identified those determined to be the best identifiable reading frames using the sequence information available at the time of filing. The term "secreted" protein herein refers to those that are transported across the membrane when expressed in a suitable host cell, including transport as a result of a signal sequence in their amino acid sequence. "Secreted" proteins include, but are not limited to, proteins that are totally secreted from the cells in which they are expressed (eg, soluble proteins) or partially secreted (eg, receptors). “Secreted” proteins also include, but are not limited to, those that are transported across the membrane of the endoplasmic reticulum.

Clone "ff168 12" The polynucleotide of the present invention has been identified as clone ff168 12.
ff16812 is an adult testis (tetratocarcinoma NCC) using a method selective for cDNA encoding a secretory protein (see US Pat. No. 5,536,637).
It was isolated from an IT) cDNA library or was identified as encoding a secretory or transmembrane protein based on computer analysis of the amino acid sequence of the encoded protein. ff168 12 is a full-length clone and contains the entire coding sequence for a secreted protein (also referred to herein as “ff168 12 protein”). The nucleotide sequence of ff168 12 determined this time is shown in SEQ ID NO: 1,
It contains a poly (A) tail. What the applicant believes to be the correct reading frame and deduced amino acid sequence of the ff16812 protein corresponding to the above nucleotide sequence is shown in SEQ ID NO: 2. EcoRI / N obtainable from the deposit containing clone ff16812
The otI restriction fragment should be approximately 1600 bp in length. The nucleotide sequence of ff16812 disclosed herein is designated as BLASTN / BLAST
The X and FASTA search protocols were used to search against the GenBank and GenSeq nucleotide sequence databases. ff168 12 is AA025945 (ze91e02.r1 S
oares fetal heart NbHH19W Homo sapiens cDNA clone 366362 5 '), AA156237 (
zl50c09.s1 Soares pregnant uterus NbHPU Homo sapiens cDNA clone 505360 3
'), AA420993 (zu08e09.s1 Soares testis NHT Homo sapiens cDNA clone 73127
2 3 '), N78486 (yz78e03.r1 Homo sapiens cDNA clone 289180 5'), W01843 (za
80a01.r1 Soares fetal lung NbHL19W Homo sapiens cDNA clone 298824 5 '),
And W95777 (ze07e02.r1 Soares fetal heart NbHH19W Homo sapiens cDNA c
It showed at least some similarity to the sequence identified as lone 358298 5 '). Based on sequence similarity, the ff168 12 protein and each similar protein or peptide may share at least some activity.

Clone "ls9 1" The polynucleotide of the present invention has been identified as clone ls9 1. ls9
1 is a method selective for cDNA encoding a secretory protein (US Pat. No. 553).
6637) and was isolated from an adult brain (substantia nigra) cDNA library,
Alternatively, it was identified as encoding a secretory or transmembrane protein based on computer analysis of the amino acid sequence of the encoded protein. Is91 is a full-length clone containing the entire coding sequence for a secreted protein (also referred to herein as "ls91 protein"). The nucleotide sequence of ls91 as determined this time is shown in SEQ ID NO: 3, which contains a poly (A) tail. This time, the applicant has identified the ls corresponding to the above nucleotide sequence.
What is believed to be the correct reading frame and deduced amino acid sequence for the 91 protein is shown in SEQ ID NO: 4. Amino acids 60 to 72 of SEQ ID NO: 4 are a putative leader / signal sequence, with the putative mature amino acid sequence starting at amino acid 73.
Since the putative leader / signal sequence is hydrophobic, it should act as a transmembrane domain and the putative leader / signal sequence will not be separated from the rest of the ls91 protein. EcoRI / NotI obtainable from the deposit containing clone ls91
The restriction fragment should be approximately 2300 bp in length. The nucleotide sequence of ls91 disclosed herein is designated BLASTA / BLASTX and F
The ASTA search protocol was used to search against the GenBank and GeneSeq databases. ls91 is AA527586 (ng42d05.s1 NCI_CGAP_Co3 Homo sapiens cDN
A clone IMAGE: 937449), AC000119 (Human BAC clone RG104I04 from 7q21-7q22
, complete sequence), T18551 (Human polycystic kidney disease normal PKD
1 gene), Y10196 (H. sapiens PEX gene), and Z94721 (Human DNA sequence
*** SEQUENCING IN PROGRESS *** from clone 167A14; showed at least some similarity to the sequence identified as HTGS phase 1). The deduced amino acid sequence disclosed herein for the ls91 protein was used to generate GenP using the BLASTX search protocol.
The ept and GeneSeq amino acid sequence databases were searched. Estimated ls9 1
The proteins are AB002375 (KIAA0377 [Homo sapiens]) and R95913 (Neural thread
It showed at least some similarity to sequences identified as (protein).
Based on sequence similarity, the ls91 protein and each similar protein or peptide may share at least some activity. The TopPredII computer program predicts additional potential transmembrane domains in the ls91 protein sequence centered around amino acid 40 of SEQ ID NO: 4. The nucleotide sequence of ls91 indicates that it may contain Alu / SVA repeat elements.

Clone "na1010 1" The polynucleotide of the present invention has been identified as clone na1010 1.
na10101 was isolated from an adult brain cDNA library using a method selective for the cDNA encoding the secretory protein (see US Pat. No. 5,536,637), or for computer analysis of the amino acid sequence of the encoded protein. Based on this, it was identified as encoding a secretory protein or a transmembrane protein. na1010 1
Is a full-length clone containing the entire coding sequence for a secreted protein (also referred to herein as "na10101 protein"). The nucleotide sequence of na10101 determined this time is shown in SEQ ID NO: 5,
This includes poly (A) tails. What the applicant believes to be the correct reading frame and deduced amino acid sequence of the na10101 protein corresponding to the above nucleotide sequence is shown in SEQ ID NO: 6. Amino acids 24 to 36 of SEQ ID NO: 6 is a putative leader / signal sequence, with the putative mature amino acid sequence starting at amino acid 37. Since the putative leader / signal sequence is hydrophobic, it should act as a transmembrane domain, and the putative leader / signal sequence will not be separated from the rest of the na10101 protein. EcoRI / N obtainable from the deposit containing clone na10101
The otI restriction fragment should be approximately 1050 bp in length. The nucleotide sequence of na10101 disclosed herein is designated as BLASTA / BLASTX.
And searched against the GenBank and GeneSeq databases using the FASTA search protocol. na10101 is AC002091 (Genomic sequence from Huma
n 17, complete sequence), AC002382 (Human BAC clone RG022J17 from 7q21,
complete sequence), and M26434 (Human hypoxanthine phosphoribosyltran
It showed at least some similarity to sequences identified as sferase (HPRT) genes, complete cds). Based on sequence similarity, the na10101 protein and each similar protein or peptide may share at least some activity. The nucleotide sequence of na10101 indicates that it may contain one or more of the following repeating elements: L1 / A / MIR / SVA / LTRII,
Alu / SVA / A / GAA, or Alu / A / GAAAA.

Clone "nf87 1" The polynucleotide of the present invention has been identified as clone nf87 1. nf
871 is a method selective for cDNA encoding a secretory protein (US Pat.
Isolated from an adult brain (substantia nigra) cDNA library (see No. 536637) or identified as encoding a secretory or transmembrane protein based on computer analysis of the amino acid sequence of the encoded protein. . nf87 1 is a full-length clone and contains the entire coding sequence for a secreted protein (also referred to herein as “nf87 1 protein”). The nucleotide sequence of nf87 1 determined this time is shown in SEQ ID NO: 7, which contains a poly (A) tail. The correct reading frame and deduced amino acid sequence of the nf87 1 protein corresponding to the above nucleotide sequence, which the applicant considers to be, are shown in SEQ ID NO: 8. Amino acids 53 to 65 of SEQ ID NO: 8 are putative leader / signal sequences, with the putative mature amino acid sequence starting at amino acid 66. Because the putative leader / signal sequence is hydrophobic, it should act as a transmembrane domain and the putative leader / signal sequence will not be separated from the rest of the nf87 1 protein. EcoRI / Not obtainable from deposit containing clone nf87 1
The I restriction fragment should be approximately 1200 bp in length. The nf87 1 nucleotide sequence disclosed herein was searched against the GenBank and GeneSeq databases using the BLASTA / BLASTX and FASTA search protocols. nf87 1 is AA358277 (EST67398 Fetal lung III Homo sapie
ns cDNA 5'end similar to similar to interferon-alpha-inducible gene p27
), W52706 (zc55g02.r1 Soares senescent fibroblasts NbHSF Homo sapiens cD
NA clone 326258 5 'similar to SW INI7_HUMAN P40305 INTERFERON-ALPHA INDU
CED 11.5 KD PROTEIN), and at least some similarity to the sequences identified as X67325 (H. sapiens p27 mRNA). The deduced amino acid sequence disclosed herein for nf87 1 was labeled with GenPept and G using the BLASTX search protocol.
A search was made against the eneSeq amino acid sequence database. The putative nf87 1 protein is
It showed at least some similarity to the sequence identified as X67325 (p27 gene product [Homo sapiens]). The gene inducible by interferon-α is located on human chromosome 14q32 and expresses the highly hydrophobic p27 gene product in breast cancer cells. Based on sequence similarity, the nf87 1 proteins and each similar protein or peptide may share at least some activity. The nf87 1 protein was expressed in a COS cell expression system and an expressed protein band of about 16 kDa was detected in the membrane fraction using SDS polyacrylamide gel electrophoresis.

Clone "nh796 1" The polynucleotide of the present invention has been identified as clone nh796 1. n
h7961 is an amino acid sequence of a protein isolated or encoded from an adult brain (hypothalamus) cDNA library using a method selective for a cDNA encoding a secretory protein (see US Pat. No. 5,536,637). Was identified as encoding a secretory protein or a transmembrane protein. nh79
61 is a full-length clone containing the entire coding sequence for a secretory protein (also referred to herein as "nh7961 protein"). The nucleotide sequence of nh7961 as determined this time is set forth in SEQ ID NO: 9, which includes a poly (A) tail. What the applicant believes to be the correct reading frame and deduced amino acid sequence of the nh7961 protein corresponding to the above nucleotide sequence is shown in SEQ ID NO: 10. Amino acids 7 to 19 of SEQ ID NO: 10 are a putative leader / signal sequence, with the putative mature amino acid sequence starting at amino acid 20. Since the putative leader / signal sequence is hydrophobic, it should act as a transmembrane domain, and the putative leader / signal sequence is nh796 1
It will not be separated from the rest of the protein. EcoRI / No obtainable from the deposit containing clone nh7961
The tI restriction fragment should be approximately 1050 bp in length. The nh7961 nucleotide sequence disclosed herein was searched against the GenBank and GeneSeq databases using the BLASTA / BLASTX and FASTA search protocols. nh7961 is AA315985 (EST18772 Lung Homo sapiens cDN
A 5'end), N23239 (yw47b07.s1 Homo sapiens cDNA clone 255349 3 '), N27741
(yw51c06.s1 Homo sapiens cDNA clone 255754 3 '), U69172 (Mus musculus un
known protein mRNA, complete cds), and Z24371 (H. sapiens (D20S195) D
It showed at least some similarity to the sequence identified as NA segment containing (CA) repeat; clone AFM321xc1; single read). The deduced amino acid sequence disclosed herein for nh7961 is GenP using the BLASTX search protocol.
The ept and GeneSeq amino acid sequence databases were searched. The putative nh7961 protein showed at least some similarity to the sequence identified as U69172 (unknown [Mus musculus]). Mouse protein with unknown function (U691
72) is expressed late in palate development. Based on sequence similarity, the nh7961 proteins and each similar protein or peptide may share at least some activity. The nh7961 protein was expressed in the COS cell expression system and an expressed protein band of approximately 25 kDa was detected in the conditioned medium and membrane fractions using SDS polyacrylamide gel electrophoresis.

Clone "nn229 1" The polynucleotide of the present invention has been identified as clone nn229 1. n
n2291 is a protein isolated or encoded from a human fetal kidney (293 cell line) cDNA library using a method selective for cDNAs encoding secretory proteins (see US Pat. No. 5,536,637). It was identified as encoding a secreted or transmembrane protein based on computer analysis of the amino acid sequence. nn2291 is a full-length clone and is a secreted protein (herein “nn229
(Also referred to as "1 protein"). The nucleotide sequence of nn229 1 determined this time is shown in SEQ ID NO: 11,
This includes poly (A) tails. What the applicant believes to be the correct reading frame and deduced amino acid sequence of the nn2291 protein corresponding to the above nucleotide sequence is shown in SEQ ID NO: 12. Amino acids 59 to 71 of SEQ ID NO: 12 are a putative leader / signal sequence and the putative mature amino acid sequence is amino acid 72.
Start with. Since the putative leader / signal sequence is hydrophobic, it should act as a transmembrane domain and the putative leader / signal sequence will not be separated from the rest of the nn229 1 protein. EcoRI / No obtainable from the deposit containing clone nn2291
The tI restriction fragment should be approximately 1050 bp in length. The nucleotide sequence disclosed for nn229 1 disclosed herein was searched against the GenBank and GeneSeq databases using the BLASTA / BLASTX and FASTA search protocols. nn2291 is H24014 (ym49f02.s1 Homo sapiens cDNA clo
ne 51480 3 '), R08508 (ye95h01.r1 Homo sapiens cDNA clone 125521 5' simil
ar to gb | M87910 | HUMALNE34 Human carcinoma cell-derived Alu RNA transcrip
t, (rRNA); gb J02931 TISSUE FACTOR PRECURSOR (HUMAN)), and Z96508 (H.
sapiens telomeric DNA sequence, clone 22QTEL030, read 22QTELOO030.seq) showed at least some similarity to the identified sequence. Based on sequence similarities, the nn2291 protein and each similar protein or peptide may share at least some activity. The TopPredII computer program predicts a potential transmembrane domain in the nn2291 protein sequence centered around amino acid 20 of SEQ ID NO: 12. The nucleotide sequence of nn229_1 indicates that it may contain MER20 repeat elements.

Clone "np156 1" The polynucleotide of the present invention has been identified as clone np156 1. n
p1561 was isolated from a human fetal kidney (293 cell line) cDNA library using a method selective for the cDNA encoding the secretory protein (see US Pat. No. 5,536,637), or of the encoded protein. It was identified as encoding a secreted or transmembrane protein based on computer analysis of the amino acid sequence. np1561 is a full-length clone and is a secreted protein (herein “np1561”
(Also referred to as "1 protein"). The nucleotide sequence of np156 1 determined this time is shown in SEQ ID NO: 13, and
This includes poly (A) tails. What the applicant believes to be the correct reading frame and deduced amino acid sequence of the np156 1 protein corresponding to the above nucleotide sequence is shown in SEQ ID NO: 14. Amino acids 6 to 18 of SEQ ID NO: 14 are a putative leader / signal sequence, with the putative amino acid sequence starting at amino acid 19. Since the putative leader / signal sequence is hydrophobic, it should act as a transmembrane domain and the putative leader / signal sequence will not be separated from the rest of the np156 1 protein. EcoRI / No obtainable from the deposit containing clone np156_1
The tI restriction fragment should be approximately 1200 bp in length. The np156_1 nucleotide sequence disclosed herein was searched against the GenBank and GeneSeq databases using the BLASTA / BLASTX and FASTA search protocols. np156-1 is AA298580 (EST114211 HSC172 cells I Homo
sapiens cDNA 5'end), AA447514 (zw81a05.s1 Soares testis NHT Homo sapien
s cDNA clone 782576 3 '), AC002309 (*** SEQUENCING IN PROGRESS *** Human
Chromosome 22q11 Cosmid Clone 63e9; HTGS phase 1, 3 unordered pieces), A
F007269 (Arabidopsis thaliana BAC IG002N01), and N53641 (yz04g03.r1 H
omo sapiens cDNA clone 282100 5 ') showed at least some similarity to the sequence identified. Based on sequence similarity, the np156_1 proteins and each similar protein or peptide may share at least some activity.

[0019] The deposit of the clone clone ff168 12, ls9 1, na1010 1 , nf87 1,
nh796 1, nn229 1, and np156 1 are American Type Culture Collection (10801) on December 31, 1997 under the Budapest Treaty.
University Boulevard, Manassas, Virginia 20110-2209 USA) as an original deposit, and is given accession number ATCC 98623, from which clones containing individual polynucleotides are available. All restrictions on the public use of deposited material are 37C. F. R. Except for the provision of § 1.808 (b), it will be canceled upon grant of a patent and the deposit period will be 37C. F. R. Will follow § 1.806. Each clone was transfected into separate bacterial cells (E. coli) as a deposit of this complex. EcoRI / NotI digestion (5 'site, EcoRI; 3
Each clone can be taken from the deposited vector by carrying out the'site, NotI) and obtaining a fragment of the appropriate size for such clone. Each clone was deposited in either the pED6 or pNOTs vector shown in Figures 1A and 1B. The pED6dpc2 vector (pED6) was transformed into pED6d by inserting a new polylinker to facilitate cDNA cloning.
It was derived from pc1 (Kaufman et al. (1991). NAR 19: 4485-4490). DHFR
Was deleted, a new polylinker was inserted, and the M13 origin of replication was inserted into the ClaI site to transform the pNOTs vector into pMT2 (Kaufman et al. 1989. Mol.
Cell. Biol. 9: 1741-1750). In some cases, the deposited clones "flip" in the deposited isolate (ie, in the reverse direction). Even in such a case, the cDNA insert can be isolated by digestion with EcoRI and NotI. However, in that case, NotI will generate a 5'site and EcoR to position the cDNA in the correct orientation for expression in the appropriate vector.
I will generate a 3'site. cDNA from the vector in which the cDNA has been deposited
NA may be expressed. Bacterial cells containing individual clones can be obtained from the complex deposits as follows: Oligonucleotide probes or probes should be designed to be for known sequences for individual clones. This sequence can be derived from the sequences herein or a combination of those sequences. The oligonucleotide probe sequences used to isolate each full length clone are shown below and they should be the most reliable in isolating the clone of interest.

Clone probe sequence SEQ ID NO: 21 ff168 12 SEQ ID NO: 15 ls9 1 SEQ ID NO: 16 na1010 1 SEQ ID NO: 17 nf87 1 SEQ ID NO: 18 nh796 1 SEQ ID NO: 19 nn229 1 SEQ ID NO: 20 np156 1 SEQ ID NO : 21 The sequence listed above contains an N at position 2 which is a biotinylated phosphoramidite residue (eg biotin phosphoramidite (1-dimethoxytrityloxy-2
-(N-Biotinyl-4-aminobutyl) -propyl-3-O- (2-cyanoethyl)-(N, N'-diisopropyl) -phosphoramidite (Glen Research Catalog No. 10-1953))). Occupied by. Preferably, the design of the oligonucleotide probe should follow these parameters: (a) it should be designed to be the region of the sequence with the least, if any, ambiguous bases (N); (B) Tm is about 80 ° C (2 ° C for A or T, 4 for G or C)
Should be designed such that Preferably, the oligonucleotide should be labeled with γ- ³² P-ATP (specific activity 6000 Ci / mmole) using T4 polynucleotide kinase, using a widely used method for labeling oligonucleotides. Other labeling methods can also be used. Preferably, unincorporated label should be removed by gel filtration or other established method. The amount of radioactivity incorporated into the probe should be quantified by measuring with a scintillation counter. Preferably,
The specific activity of the resulting probe should be about 4e + 6 dpm / pmole. Preferably, the bacterial culture containing the pool of full length clones should be thawed and 100 μl of the stock should be used to inoculate a sterile culture flask containing 25 ml of sterile L broth containing 100 μg / ml ampicillin. Preferably, the culture should be grown to saturation at 37 ° C., and preferably the saturated culture is fresh L
Should be diluted with broth. Preferably, a portion of these dilutions is plated to give 100 μg / ml ampicillin in a 150 mm Petri dish and 1.
The dilution and volume that yielded 5000 distinct, well-separated colonies when grown overnight at 37 ° C. on solid bacterial medium containing L broth containing 5% agar should be determined. Other known methods for obtaining well-defined and well-separated colonies can also be used. The colonies should then be transferred to a nitrocellulose filter using standard colony hybridization methods, lysed, denatured and then baked. Then, preferably 6X SSC containing 0.5% SDS, 100 μg / ml yeast RNA, and 10 mM EDTA (205.3 stock is 175.3 g).
NaCl / liter, 88.2 g sodium citrate, pH 7.0 with NaOH
(About 10 ml per 150 mm filter) with gentle agitation at 65 ° C. for 1 hour. The probe is then preferably added to the hybridization mix so that the concentration is greater than or equal to 1e + 6 dpm / ml. The filter is then preferably washed with 500 ml of 2X SSC / 0.5% SDS at room temperature without agitation, preferably afterwards at room temperature with 500 ml of 2X SSC / 0 with gentle shaking.
． Wash with 1% SDS. 65 ° C, 30 minutes to 1 hour, 0.1X SSC / 0
． A third wash with 5% SDS is optimal. The filters are then preferably dried and subjected to autoradiography for sufficient time to visualize positives on X-ray film. Other known hybridization methods can also be used. Positive colonies are picked, grown in culture, and then plasmid DNA is isolated using standard procedures. Clones can then be verified by restriction analysis, hybridization analysis or DNA sequencing.

Also included in the present invention are fragments of the proteins of the present invention capable of exhibiting biological activity. The protein fragment may be linear, or, for example, HUSaragovi.
, et al., Bio / Technology 10, 773-778 (1992) and RSMc Dowell, et al., J.
Amer. Chem. Soc. 114, 9245-9253 (1992) (both references are considered to be herein incorporated by reference) and may be cyclized using known methods. . For many purposes, including increasing the valency of protein binding sites,
Such fragments may be fused to carrier molecules, such as immunoglobulins.
For example, a protein fragment may be fused to the Fc portion of an immunoglobulin via a "linker". For a bivalent form of the protein, such a fusion may be performed on the F of an IgG molecule.
It can be done for part c. Other immunoglobulin isotypes may be used to obtain such fusions. For example, a protein-IgM fusion will yield a decavalent form of the protein of the invention. The invention also provides the disclosed proteins in full length and mature form. Full length forms of such proteins have been identified in the sequence listing by translation of the nucleotide sequences of the disclosed clones. Mature forms of such proteins may be obtained by expressing the disclosed full length polynucleotides (preferably those deposited with the ATCC) in a suitable mammalian cell or other host cell. The sequence of the mature form of the protein may be determined from the full length amino acid sequence.

The invention also provides genes corresponding to the cDNA sequences disclosed herein. "
A "corresponding gene" is a region of the genome that is transcribed to produce mRNA, the m of which
CDNA is derived from RNA and is flanked by genomic regions (including coding sequences, 5'and 3'untranslated regions) required for regulated expression of such genes, or spliced exons, introns, promoters, enhancers, and It also includes silencer or suppressor elements. The sequence information disclosed herein can be used to isolate the corresponding gene. The sequence information disclosed herein can be used to isolate the corresponding gene. Such methods involve preparing probes or primers from the disclosed sequence information to identify and / or amplify genes in suitable genomic libraries or other sources of genomic material. An "isolated gene" is a gene that is separated from the flanking coding sequences (if any) that are present in the genome of the organism in which it was isolated. Chromosomal locations corresponding to the polynucleotide sequences disclosed herein may be determined, for example, by hybridizing an appropriately labeled polynucleotide of the invention to the chromosome in situ. Corresponding chromosomal positions of the disclosed polynucleotides may be determined by identifying significantly similar nucleotide sequences in public databases, such as expressed sequence tags (ESTs) that have already been mapped to specific chromosomal positions. For at least some of the polynucleotide sequences disclosed herein, public database sequences that have at least some similarity to the polynucleotides of the present invention are listed by database accession number. A search of these public database sequences using the GenBank accession number to identify UniGene clusters of overlapping sequences is described in the National Center for Biotechnology I
It can be done on the internet site provided by nformation, the address is http://www.ncbi.nlm.nih.gov/UniGene/. Many UniGene clusters so identified have already been mapped to specific chromosomal locations.

Organisms having enhanced, decreased, or altered expression of the genes corresponding to the polynucleotide sequences disclosed herein are provided. Desirable changes in gene expression are achieved by using antisense polynucleotides or by using ribozymes that bind and / or cleave mRNA transcribed from the gene (Albert and Morris, 1994, Trends Pharmacol. Sci. 15). (7): 250-2
54; Lavarosky et al., 1997, Biochem. Mol. Med. 62 (1): 11-22; and Hampel,
1998, Prog. Nucleic Acid Res. Mol. Biol. 58: 1-39; all of which are considered herein incorporated by reference). A transgenic animal having a multicopy gene corresponding to a polynucleotide disclosed herein, preferably obtained by transforming a cell with a genetic construct that is stably maintained in transformed cells and their progeny. Transgenic animals are provided. Also provided are transgenic animals having altered genetic control regions that increase or decrease gene expression levels or alter temporal or spatial patterns of gene expression (see European Patent No. 0 649 464 B1; All of these are considered to be described herein by reference). Furthermore, insertion of an external sequence or deletion of all or part of the corresponding gene provides an organism in which the gene corresponding to the polynucleotide sequences disclosed herein is incompletely or completely inactivated. By insertion, preferably by incorrect post-excision insertion of the transposable element (Plasterk, 1992, Bioessays 14 (9): 629-633; Zwaal et al., 19
93, Proc. Natl. Acad. Sci. USA 90 (16): 7431-7435; Clark et al., 1994, Pr.
oc. Natl. Acad. Sci. USA 91 (2): 719-722; all of which are considered to be described herein by reference), or by homologous recombination, preferably positive / negative genetic selection. By homologous recombination detected by the method (Mansour et a
l., 1988, Nature 336: 348-352; US Patent Nos. 5,464,764; 5,487,992; 5,
627,059; 5,631,153; 5,614, 396; 5,616,491; and 5,679,523; all of which are considered as described herein by reference), incomplete or complete gene inactivation can be performed. These organisms with altered gene expression are
Eukaryotes are preferred, and mammals are more preferred. Such organisms are useful in the development of non-human models for the study of diseases associated with the corresponding gene, as well as in the development of assay systems for the identification of molecules that interact with the protein product from the corresponding gene. When the proteins of the invention are membrane bound (eg, receptors), the invention also provides soluble forms of such proteins. In such forms, some or all of the intracellular and transmembrane domains of the protein are removed so that the protein is fully secreted from the host in which it is expressed. The intracellular and transmembrane domains of the protein of the invention can be identified by known techniques for determining such domains from sequence information. For example, TopP
The position of the transmembrane domains in the amino acid sequence can be predicted using the redII computer program, which domains are described by the central position of the transmembrane domain,
With at least 10 transmembrane amino acids on either side of the listed central residue. The proteins and protein fragments of the invention comprise at least 25% (more preferably at least 50%, most preferably at least 75%) of the amino acid sequence of the disclosed proteins.
At least 60% sequence identity to the disclosed protein (more preferably at least 75% identity, most preferably at least 90%).
Or 95% identity). Sequence identity is determined by comparing the amino acid sequences of proteins when the sequences are aligned to maximize overlap and identity while minimizing sequence gaps. There is preferably at least 75% sequence identity (more preferably at least 85% identity, most preferably at least 95% identity) to any of the disclosed protein segments, preferably 8 or more ( More preferably 20 or more, most preferably 30 or more) proteins or protein fragments containing segments containing contiguous amino acids are also included in the invention.

In detail, WU-BLAST (Washington University BLAST) version 2
． Sequence software may be used to determine sequence identity, which software is based on NCBI-BLAST version 1.4 based WU, which is publicly available for use without limitation.
-Building on BLAST (Altschul and Gish, 1996, Local ali
gnment statistics, Doolittle ed., Methods in Enzymology 266: 460-480; Al
tschul et al., 1990, Basic local alignment search tool, Journal of Molec
ular Biology 215: 403-410; Gish and States, 1993, Identification of prot
ein coding regions by database similarity search, Nature Genetics 3: 266
-272; Karlin and Altschul, 1993, Applications and statistics for multipl
e high-scoring segments in molecular sequences, Proc. Natl. Acad. Sci. U
SA 90: 5873-5877, which is hereby incorporated by reference for all these documents. WU for some UNIX platforms
-BLAST version 2.0 executable program ftp://blast.wustl.edu/bl
It can be downloaded from ast / executables. In addition to some support programs, a complete set of search programs (BLASTP, BLASTN, B
LASTX, TBLASTN and TBLASTX) are provided at this site. WU-BLAST 2.0 is copyrighted and may not be sold or redistributed in any form without permission of the author, but is not for commercial use or for research use You can use it freely. Set (BLASTP
, BLASTN, BLASTX, TBLASTN and TBLASTX), the gap alignment routine (gapp
ed alignment routines) are integrated into the database search itself, resulting in highly sensitive and highly selective, easily interpretable results. Optimally, you can turn off gapping in all of these programs. The default penalty (Q) for the length gap is BL
Q = 9 for ASTP and Q = 10 for BLASTN,
1 to 8, 9, 10, 11, 12 to 20, 21 to 50, including zero
, 51 to 100, or any other integer value. The default penalty per residue (R) for expanding the gap is protein and BLA.
R = 2 for STP and R = 10 for BLASTN, but 0
It may be changed to an integer value such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 to 20, 21 to 50, 51 to 100. To compare the sequences side-by-side to maximize overlap and identity, and to minimize sequence gaps, Q and R
Any combination of may be used. The default amino acid comparison matrix is BLOSUM62, but other amino acid comparison matrices such as PAM can also be used.

Species homologues of the disclosed polynucleotides and proteins are also provided by the present invention. The term "species homolog" as used herein refers to a protein or polynucleotide for a species different from the origin of the protein or polynucleotide, but as determined by one of skill in the art,
It refers to those having significant sequence similarity. Preferably, the polynucleotide species homologue has at least 60% (more preferably at least 75%, most preferably at least 90%) sequence identity to the particular polynucleotide, and the protein species homologue is At least 30% (more preferably at least 45%)
%, Most preferably at least 60%). Here, sequence identity is determined by comparing the nucleotide sequences of polynucleotides or the amino acid sequences of proteins that are aligned to maximize overlap and identity and minimize sequence gaps. Species homologs may be isolated and identified by making appropriate probes or primers from the sequences provided herein and then screening for an appropriate source of nucleic acid from the desired species. Preferably, the species homolog is one isolated from a mammalian species. Most preferably, the species homologue is, for example, Pan troglodyte.
s, Gorilla gorilla, Pongo pygmaeus, Hylobates concolor, Macaca mulatta,
Papio papio, Papio hamadryas, Cercopithecus aethiops, Cebus capucinus, A
otus trivirgatus, Sanguinus oedipus, Microcebus murinus, Mus musculus, R
attus norvegicus, Cricetulus griseus, Felis catus, Mustela vison, Canis
Genomes of genes in one species have been isolated from specific mammals, such as familiaris, Oryctolagus cuniculus, Bos taurus, Ovis aries, Sus scrofa and Equus caballus, and have been genetically mapped. It is possible to identify the relationship of gene sequence order between organization and genomic organization of related genes in another species (O'Brien and Seuanez, 1988, Ann. Rev. Genet. 22: 323- 351;
O'Brien et al., 1993, Nature Genetics 3: 103-112; Johansson et al., 199.
5, Genomics 25: 682-690; Lyons et al., 1997, Nature Genetics 15: 47-56;
O'Brien et al., 1997, Trends in Genetics 13 (10): 393-399; Carver and Stu
bbs, 1997, Genome Research 7: 1123-1137, all of which are considered herein incorporated by reference).

The invention also provides allelic variants of the disclosed polynucleotides, ie, naturally occurring variants of an isolated polynucleotide that encodes a protein that is the same, homologous, or related to the protein encoded by the disclosed polynucleotide. Includes forms. Preferably, the allelic variant is at least 60% (
(More preferably at least 75%, most preferably at least 90%) identity, wherein the sequence identities are aligned such that the overlaps and identities are maximized and the sequence gaps are minimized. Is determined by comparing the nucleotide sequences of Allelic variants may be isolated and identified by making suitable probes or primers from the sequences described herein and screening for a suitable source of nucleic acid from an individual of the appropriate species. The present invention also includes a polynucleotide having a sequence complementary to the sequence of the polynucleotide disclosed herein.

The invention also provides a polynucleotide capable of hybridizing to a polynucleotide described herein under reduced stringency conditions, more preferably under stringent conditions, and most preferably under very stringent conditions. It also includes nucleotides. The following table shows examples of stringency conditions. Very strict conditions are, for example, at least as strict as conditions A to F, strict conditions are, for example, at least as strict as conditions G to L, and conditions with reduced strictness are: For example, it is at least as strict as the conditions MR.

[Table 1] a): Hybrid length is the expected length for the hybridizing region (s) of the hybridizing polynucleotide. When hybridizing a polynucleotide to a target polynucleotide of unknown sequence, hybrid length is assumed to be the length of the hybridizing polynucleotide. When a polynucleotide of known sequence hybridizes, the hybrid length can be determined by aligning the polynucleotide sequences and identifying the region (s) of optimal sequence complementarity. b): SSPE (1X in hybridization and wash buffers
SSPE is 0.15M NaCl, 10 mM NaH ₂ PO ₄ , and 1.25 mM
EDTA, pH 7.4) can be replaced with SSC (1 × SSC is 0.15 M NaCl and 15 mM sodium citrate). After the hybridization is completed, washing is performed for 15 minutes. * T _B -T _R: The hybridization temperature for hybrids anticipated to shorter than 50 base pairs in length, than the hybrid melting temperature (T _m) 5 to 10
℃ should be lower. The following equation determines T _m . For hybrids less than 18 base pairs in length, T _m (° C) = 2 (number of A + T bases) + 4 (number of G + C bases). For a hybrid of 18 to 49 base pairs in length, T _m (° C) = 81.5 + 16.6 (log ₁₀ [Na ⁺ ]) + 0.41 (% G + C)-(600 / N), where N is the hybrid Base number, sodium ion concentration in the hybridization buffer ([Na ⁺ ] = 0.165M for 1 × SSC). Further examples of stringency conditions for polynucleotide hybridization are Sambrook, J., EF Fritsch, and T. Maniatis, 1989, Molecular Cloning:
A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Ha
rbor, NY, chapters 9 and 11, and Current Protocols in Molecular Biology,
1995, FM Ausubel et al., Eds., John Wiley & Sons, Inc., sections 2.10
and 6.3-6.4, and are hereby incorporated by reference. Preferably, each such hybridizing polynucleotide comprises at least 25% of the polynucleotide of the invention to hybridize.
(More preferably at least 50%, most preferably at least 75%) and has at least 60% sequence identity (more preferably at least 75%) to the polynucleotide of the invention to be hybridized. Identity, most preferably at least 90% or 95% identity). Sequence identity is determined by comparing the amino acid sequences of proteins when the sequences are aligned to maximize overlap and identity while minimizing sequence gaps.

An isolated polynucleotide encoding the protein of the present invention has been isolated from Kaufman et al., Nucl.
The protein may be recombinantly produced by operably linking it to an expression control sequence such as the pMT2 or pED expression vector disclosed in eic Acids Res. 19, 4485-4490 (1991). Many suitable expression control sequences are known in the art. Many suitable expression control sequences are known in the art. General methods for recombinant protein expression are also known and are described by R. Kaufman, Methods in Enzymology 185, 537-566.
(1990) is a typical example. The term "operably linked" as defined herein means that the isolated polynucleotide of the present invention and the expression control sequence are placed in a vector or cell and transformed (transfected) with the linked polynucleotide / expression control sequence. It is meant to be expressed by the host cell. Many types of cells can act as suitable host cells for the expression of the proteins of the invention.
Mammalian cells include, for example, monkey COS cells, Chinese hamster ovary (CHO) cells, human kidney 293 cells, human epidermal A431 cells, human Colo2.
05 cells, 3T3 cells, CV-1 cells, other transformed primate cell lines, normal diploid cells, cell lines obtained from in vitro culture of primary tissues, primary explants, HeLa cells, mouse cells, BHK. , HL-60, U937HaK or Ju
Includes rkat cells. Alternatively, it is possible to produce the protein in lower eukaryotic cells such as yeast or prokaryotic cells such as bacteria. A potentially suitable yeast strain is Saccharomyces ce
It includes revisiae, Schizosaccharomyces pombe, Kluyveromyces strains, Candida, or yeast strains capable of expressing heterologous proteins. A potentially suitable bacterial strain is Escherichia
coli, Bacillus subtilis, Salmonella typhimurium, or bacterial strains capable of expressing heterologous proteins. If the protein is produced in yeast or bacteria, it may be necessary to modify the protein produced therein, for example by phosphorylation or glycosylation of suitable sites to obtain a functional protein. Such covalent attachment may be accomplished using known chemical or enzymatic methods. The isolated polynucleotide of the invention may be operably linked to appropriate control sequences in one or more insect expression vectors and the protein obtained by using an insect expression system. Materials and methods for baculovirus / insect cell expression systems are described, for example, in In
It is commercially available in kit form (MaxBac ^R kit) from Invitrogen, San Diego, California, USA, and such methods are well known in the art and described by Summers and Smith, Texas, Agricultural Experiment Station Bulletin No. 1555 (1987).
) (Considered to be described herein by reference). As used herein, insect cells capable of expressing the polynucleotides of the present invention have been "transformed". The protein of the present invention may be produced by culturing the transformed host cell under culture conditions suitable for expressing the recombinant protein. The resulting expressed protein may then be purified from such cultures (ie, media or cell extracts) using known purification processes such as gel filtration and ion exchange chromatography. Also, purification of proteins is accomplished by an affinity column containing an agent that will bind to the protein; one or more column steps with an affinity column such as Concanavalin A-Agarose, Heparin-Toyopearl ^R or Cibacrom blue 3GA Sepharose ^R ;
One or more steps using hydrophobic interaction chromatography with a resin such as phenyl ether, butyl ether, or propyl ether; or immunoaffinity chromatography. Alternatively, the protein of the invention may be expressed in an easily purified form. For example, maltose binding protein (MBP), glutathione-S-tonspherase (GST)
Alternatively, it may be expressed as a fusion protein such as a fusion protein with thioredoxin (TRX). Kits for expressing and purifying such fusion proteins are available in New En
Commercially available from glan BioLab (Beverly, MA), Pharmacia (Piscataway, NJ) and InVitrogen. The protein can also be tagged with an epitope and then purified by using specific antibodies directed against such epitope. One such epitope (“flag”) is commercially available from Kodak (New Haeven, CT). Finally, the protein is further purified using one or more reversed phase high quality liquid chromatography (RP-PLC) steps using a hydrophobic RP-HPLC medium such as silica gel with pendant methyl or other aliphatic groups. be able to. A combination of some or all of the above purification steps can also be used in combination to obtain a substantially homogeneous isolated recombinant protein. The protein thus purified is substantially free of other mammalian proteins and is defined herein as an "isolated protein". The protein of the invention is expressed as a product of a transgenic animal, for example, as a component of transgenic bovine, goat, porcine, or ovine milk characterized by somatic or germ cells containing the nucleotide sequence encoding the protein. May be. The protein may be produced by known conventional chemical synthesis. The method for constructing the protein of the present invention by synthetic means is known to those skilled in the art. Synthetically constructed protein sequences may have common biological properties, including protein activity, by sharing primary, secondary or tertiary structure and / or conformational properties. Thus, they may be used as biological activity or immunological substitutes for naturally occurring purified proteins in immunological processes for screening therapeutic compounds and generating antibodies.

The proteins presented herein are characterized by an amino acid sequence similar to that of the purified protein, but may be modified therein either naturally or by derivatization. For example, modification of a peptide or DNA sequence can be done by one of ordinary skill in the art using known methods. Desired modifications in a protein sequence include changes, substitutions, replacements, insertions or deletions of selected amino acid residues in the coding sequence. For example, one or more cysteine residues may be deleted or replaced with another amino acid to alter the conformation of the molecule. Methods for such alterations, substitutions, replacements, insertions or deletions are well known to those of skill in the art (eg, US Pat. No. 415).
8584). Preferably, such changes, substitutions, replacements, insertions or deletions retain the desired activity of the protein. Other fragments and derivatives of protein sequences that are expected to retain protein activity in whole or in part, and thus may be useful in screening or other immunological methods, can be made by one of skill in the art from the disclosure herein. . It is believed that such modifications are covered by the present invention.

Uses and Biological Activities The polynucleotides and proteins of the invention are believed to exhibit one or more of the following uses or biological activities, including those associated with the assays cited herein. To be The uses or activities described with respect to the proteins of the invention may be due to the administration or use of such proteins, or to the administration or use of polynucleotides encoding such proteins (eg, vectors suitable for gene therapy or DNA transfer). Can be provided.

Research applications and utilities The polynucleotides provided by the present invention may be used by the research population for a variety of purposes. For analysis, for characterization or therapeutic use, as a marker for tissues in which the corresponding protein is preferentially expressed (constitutively or at a particular stage of tissue differentiation or development, or in a disease state), and As a molecular weight marker in Southern gels, as a chromosomal marker or tag for identification of chromosomes or mapping of related gene positions (if labeled), to identify potential genetic diseases compared to the patient's endogenous DNA , Known sequences in the process of discovering other novel polynucleotides as an information source for obtaining PCR primers for genetic fingerprinting in order to discover novel related DNA sequences for use as hybridizing probes. As a probe for deduction, a "gene chip" or other To select and make oligomers for binding to the carrier, to test expression patterns, to generate anti-protein antibodies using DNA immunization, as well as to generate anti-DNA antibodies, or further immunization. Polynucleotides can be used as antigens to induce a response. If it encodes a protein that binds or potentially binds to another protein (eg, as in a receptor-ligand interaction), identifies another protein that binds, or an inhibitor of the binding interaction. The polynucleotides can be used in an interaction trap assay to identify E. coli (eg, as described in Gyuris et al., Cell 75: 791-803 (1993)). The proteins provided by the invention include assays for the determination of biological activity, including a panel of large numbers of proteins for high throughput screening; to induce antibody production or other immune responses; As a reagent (including a labeling reagent) in an assay designed to quantitatively determine the level of a protein (or its receptor) in a biological fluid; the corresponding protein is preferentially expressed (constitutively) , Or as a marker of tissue (expressed at a particular stage of tissue differentiation or development, or in a disease state); and, of course, may be used to isolate the relevant receptor or ligand. If a protein binds or potentially binds to another protein, the protein can be used to identify other proteins that bind, or to identify inhibitors of binding interactions. Proteins involved in these binding interactions can also be used to screen peptides or small molecule inhibitors or agonists for binding interactions. Any or all of these research applications can be developed into reagent grade or kit formats for commercialization as research products. Methods for performing the above uses are well known to those of skill in the art. The literature disclosing such a method is "Molecular Cloning: A Laboratory Manual", 2nd ed., Cold Sp.
ring Harbor Laboratory Press, Sambrook, J., EF Fritsch and T. Maniatis
eds., 1989, and "Methods in Enzymology: Guide to Molecular Cloning Tec
hniques ", Academic Press, Berger, SL and AR Kimmel eds., 1987, but is not limited to these.

Use as nutrition The polynucleotide and protein of the present invention can be used as a nutrient source or an additive. Such uses include, but are not limited to, use as a protein or amino acid additive, use as a carbon source, use as a nitrogen source and use as a carbohydrate source. In such cases, the protein or polynucleotide of the invention may be added as a bait for a particular organism or it may be administered as a separate solid liquid formulation such as in the form of a powder, pill, solution, suspension or capsule. . In the case of a microorganism, the protein or polynucleotide of the present invention can be added to the medium in which the microorganism is cultured.

Cytokines and Cell Proliferation / Differentiation Activities The proteins of the invention may exhibit cytokine activity, cell proliferation activity (induction or inhibition) or cell differentiation activity (induction or inhibition), or the production of other cytokines in certain cell populations. Can be induced. Many proteinaceous factors that have been found to date, including all known cytokines, show activity in one or more factor-dependent cell proliferation assays, thus making the assay a convenient way to confirm cytokine activity. Serve as. The activity of the protein of the present invention is determined by cell lines (32D, DA
2, DA1G, T10, B9, B9 / 11, BaF3, MC9 / G, M + (pr
eBM +), 2E8, RB5, DA1, 123, T1165, HT2, CTL
Confirmed by any of the many conventional factor-dependent cell proliferation assays for L2, TF-1, Mo7e and CMK, including but not limited to. The activity of the proteins of the invention may be measured, inter alia, by the following methods: Assays for T cell or thymocyte proliferation are described in Current Protocols in I.
mmunology, Ed by JE Coligan, AM Kruisbeek, DH Margulies, EM She
vach, W Strober, Pub. Greene Publishing Associates and Wiley-Interscienc
e (Chapter 3, In Vitro assays for Mouse Lymphocyte Function 3.1-3.19; Ch
apter 7, Immunologic studies in Humans); Takai et al., J. Immunol. 137: 3.
494-3500, 1986; Bertagnolli et al., J. Immunol. 145: 1706-1712, 1990; Ber.
tagnolli et al., Cellular Immunology 133: 327-341, 1991; Bertagnolli, et
al., J. Immunol. 149: 3778-3783, 1992; Bowman et al., J. Immunol. 152: 17
56-1761, 1994, but is not limited thereto. Assays for cytokine production and / or proliferation of spleen cells, lymph node cells or thymocytes are described in Polyclonal T cell stimulation, Kruisbeek, AM an.
d Shevach, EM In Current Protocols in Immunology. JEea Coligan eds
.Vol 1 pp. 3.12.1-3.12.14, John Wiley and Sons, Toronto. 1994; and Meas
urement of mouse and human Interferon g, Schreiber, RD In Current Pro
tocols in Immunology. JEea Coligan eds. Vol 1 pp. 6.8.1-6.8.8, John
Includes, but is not limited to, those described in Wiley and Sons, Toronto. 1994. Assays for proliferation and differentiation of hematopoietic and lymphogenic cells are described in Measurement of Human and Murine Interleukin 2 and Interleukin 4, Bottoml.
y, K., Davis, LS and Lipsky, PE In Current Protocols in Immunology.
JEea Coligan eds. Vol 1 pp. 6.3.1-6.3.12, John Wiley and Sons, Toron
to. 1991; deVries et al., J. Exp. Med. 173: 1205-1211, 1991; Moreau et al.
., Nature 336: 690-692, 1988; Greenberger et al., Proc. Natl. Acad. Sci.
USA 80: 2931-2938, 1983; Measurement of mouse and human interleukin 6
-Nordan, R. In Current Protocols in Immunology. JEea Coligan eds. V
ol 1 pp. 6.6.1-6.6.5, John Wiley and Sons, Toronto. 1991; Smith et al.,
Proc. Natl. Acad. Sci. USA 83: 1857-1861, 1986; Measurement of human I
nterleukin 11-Bennett, F., Giannotti, J., Clark, SC and Turner, K. J
In Current Protocols in Immunology. JEea Coligan eds. Vol 1 pp.
6.15.1 John Wiley and Sons, Toronto. 1991; Measurement of mouse and hum
an Interleukin 9-Ciarletta, A., Giannotti, J., Clark, SC and Turner,
KJ In Current Protocols in Immunology. JEea Coligan eds. Vol 1 p
Includes, but is not limited to, those described in p. 6.13.1, John Wiley and Sons, Toronto. 1991. Assays for the response of T cell clones to antigens (specifically identifying APC-T cell interactions and direct T cell effects by measuring proliferation and cytokine production) are described in Current Protocols in Immunology, Ed by JE.
Coligan, AM Kruisbeek, DH Margulies, EM Shevach, W Strober, Pub.
Greene Publishing Associates and Wiley-Interscience (Chapter 3, In Vitro
assays for Mouse Lymphocyte Function; Chapter 6, Cytokines and their ce
llular receptors; Chapter 7, Immunologic studies in Humans); Weinberger
et al., Proc. Natl. Acad. Sci. USA 77: 6091-6095, 1980; Weinberger et al.
, Eur. J. Immun. 11: 405-411, 1981; Takai et al., J. Immunol. 137: 3494-35.
00, 1986; Takai et al., J. Immunol. 140: 508-512, 1988, but is not limited thereto.

Immunostimulatory or Suppressive Activity The protein of the present invention may also exhibit immunostimulatory or immunosuppressive activity including, but not limited to, the activity in the assays described herein. Protein is
It may be useful in a variety of deficiencies and disorders, including severe immunodeficiency complications (SCIDs), eg, in up- or down-regulating the proliferation of T and / or B lymphocytes, and N
It may be useful in potentiating the cytolytic activity of K cells and other cell populations. These immune deficiencies may be inherited and may also be viral (
For example, HIV) and bacterial or fungal infections, or autoimmune diseases. More particularly, infectious diseases caused by viruses, bacteria, fungi or other infectious agents, such as various fungal infections such as HIV, hepatitis virus, herpes virus, mycobacteria, leishmania, malaria and Candida species, It can be treated with the protein of the present invention. Of course, in this regard, the proteins of the invention may be used where a boost to the immune system is required, ie in the treatment of cancer. Autoimmune diseases that may be treated using the protein of the present invention include, for example, multiple sclerosis,
Systemic deep erythematosus, rheumatoid arthritis, autoimmune pneumonia, Guillain-Barre syndrome, autoimmune thyroiditis, insulin-dependent diabetes mellitus, myasthenia gravis, graft-versus-host disease and autoimmune inflammatory Diseases of the eye. Such proteins of the invention may be used to treat allergic reactions and conditions such as asthma or other respiratory disorders. Other conditions in which immunosuppression is desired, including asthma (particularly allergic asthma) and related respiratory problems, may be treated with the proteins of the invention. Other conditions in which immunosuppression is desired, including organ transplantation
May be treated with the protein of the present invention. The proteins of the invention can also be used to enable an immune response in many ways. Down-regulation may be in the form of inhibiting or blocking an already ongoing immune response, or may involve interfering with the induction of an immune response. The function of activated T cells may be inhibited by suppressing the T cell response, or by inducing specific resistance in the T cells, or both. In general, immunosuppression of T cell responses is an active, non-antigen-specific process that requires continuous exposure of T cells to suppressors. Tolerance refers to non-responsiveness or anergy in T cells and differs from immunosuppression in that it is generally antigen-specific and persists after termination of exposure to tolerizing agents.
Operationally, resistance can be demonstrated by the lack of a T cell response upon exposure to a specific antigen in the absence of tolerizing agent. Down-regulating or interfering with one or more antigenic functions, including but not limited to B lymphocyte antigenic functions (such as B7), eg high levels of lymphokines by activated T cells. Interfering with synthesis may be useful in tissue, skin and organ transplantation and graft versus host disease (GVHD). For example, blocking T cell function should reduce tissue destruction in tissue transplants. Typically, in tissue transplantation, graft rejection is initiated by the recognition of the tissue piece as foreign by T cells, followed by an immune reaction that destroys the graft. Of a monomeric form having the activity of a molecule (another B lymphocyte antigen (eg, B7-1, B7-3)) that inhibits or blocks the interaction of the B7 lymphocyte antigen with its native ligand on immune cells. Mixed with peptides,
Alternatively, pre-implantation administration of a soluble, monomeric peptide having B7-2 activity alone) induces binding of the molecule to its native ligand on immune cells without transducing a responsive costimulatory signal. there is a possibility. In this regard, blocking B lymphocyte antigen function interferes with cytokine synthesis by immune cells such as T cells, thus acting as an immunosuppressant. Moreover, lack of costimulation may be sufficient to abolish T cell responses. Induction of long-term tolerance by B lymphocyte antigen blocking reagents may avoid the need for repeated administration of these blocking reagents. It may also be necessary to block the function of the B lymphocyte antigen combination in order to achieve sufficient immunosuppression or tolerance in the subject. The effectiveness of individual blocking reagents in interfering with organ transplant rejection or GVHD can be evaluated using animal models that can predict efficacy in humans. Examples of suitable systems that can be used include allogeneic heart transplants in rats and xenogeneic pancreatic islet cell transplants in mice, both of which have been used to test the immunosuppressive effects of CTLA4Ig fusion proteins in vivo. (Lensch
ow et al., Science 257: 789-792 (1992) and Turka et al., Proc Natl. Acad.
Sci. USA, 89: 11102-11105 (1992)). In addition, a mouse model of GVHD (Paul ed., Fundamental Immunology, Ravan Press, New York, 1
989, pp. 846-847) can be used to determine the blocking effect of B lymphocyte antigen function on the progression of the disease in vivo. Also, blocking antigen function may be therapeutically useful for treating autoimmune diseases. Many autoimmune diseases are the result of inappropriate activation of T cells that are responsive to self tissues and promote the production of cytokines and autoantibodies involved in the pathology of the disease. Interfering with the activation of autoreactive T cells can reduce or eliminate the symptoms of disease. Autoantibodies or T Cell Derivatives That May Be Involved in Disease Processes by Inhibiting T Cell Activation Using Administration of Reagents That Block T Cell Co-Stimulation by Disrupting B Lymphocyte Antigen Receptor: Ligand Interactions Can interfere with the production of other cytokines. In addition, blocking reagents can induce antigen-specific tolerance of autoreactive T cells that can lead to long-term remission of the disease. The effectiveness of blocking agents in preventing or ameliorating autoimmune disease can be determined using a number of well-characterized animal models for human autoimmune disease. Examples are murine experimental autoimmune encephalitis, MRLlpr / lpr mice or N.
Systemic deep-seated lupus erythematosus in ZB hybrid mice, murine autoimmune collagen arthritis, diabetes in NOD mice and BB rats, and murine experimental myasthenia gravis (Paul ed., Fundamental Immunology, Raven Press,
New York, 1989, pp.840-856). Upregulation of antigen function (preferably B lymphocyte antigen function) as a means for upregulating an immune response is also therapeutically useful.
Upregulation of the immune response may be in the form of promoting an existing immune response or eliminating the initial immune response. For example, promoting an immune response by stimulating B lymphocyte antigen function may be useful in the case of viral infection.
In addition, systemic viral diseases such as influenza, common cold, and encephalitis may be ameliorated by systemic administration of stimulatory forms of B lymphocyte antigens. Alternatively, T cells are taken from a patient and co-stimulated in vitro with TCP-loaded viral antigens loaded with ACPs expressing the peptides of the invention, or combined with a stimulatory form of the soluble peptides of the invention, and then in vitro. Re-introduction of T cells activated by P. coli into the patient may promote an anti-viral immune response in infected patients. Another way to promote an antiviral immune response is to
Infected cells are taken from a patient and the nucleic acids encoding the proteins of the invention described herein are transfected into them so that the cells express all or part of the protein on their surface, and the transfected cells are then transferred to the patient. Will be re-introduced to. The infected cell would then be able to transmit a costimulatory signal to the T cell in vivo, thereby activating the T cell. In another application, upregulation or promotion of antigen function, preferably B lymphocyte antigen function, may be useful in inducing tumor immunity. Tumor-specific in a subject by administering to the subject tumor cells (eg, sarcoma, melanoma, lymphoma, leukemia, neuroblastoma, carcinoma) transfected with a nucleic acid encoding at least one peptide of the invention Tolerance can be overcome. If desired, tumor cells can be transfected to express the peptide combination. For example, an expression vector that directs expression of tumor cells obtained from a patient in combination with a peptide having B7-2 like activity alone or in combination with a peptide having B7-1 like activity and / or B7-3 like activity, It can be transfected ex vivo. The transfected tumor cells are returned to the patient to express the peptide on the surface of the transfected cells. Alternatively, gene therapy methods can be used to target tumor cells for transfection in vivo. The presence of peptides of the invention having the activity of B lymphocyte antigens on the surface of tumor cells provides T cells with the necessary costimulatory signals to induce T cell mediated immune responses against transfected tumor cells. To do. Furthermore, MHC
Tumor cells lacking class I or MHC class II molecules, or sufficient amount of MHC
Tumor cells that are unable to re-express class I or MHC class II molecules can be transformed into whole or part of MHC class I α chain protein and β ₂ microglobulin protein or MHC class II α chain and MHC class II β chain protein (eg, cytoplasmic truncated proteins). Domain), which allows the expression of class I or class II MHC proteins on the cell surface. Expression of a suitable class I or class II HMC in combination with a peptide having activity of a B lymphocyte antigen (eg, B7-1, B7-2, B7-3) is a T cell mediated transfected tumor. Induces an immune response against cells. Optionally, a gene encoding an antisense construct that blocks the expression of MHC class II binding proteins, such as invariant chain, is co-transfected with DNA encoding a peptide having B lymphocyte antigen activity. It can also facilitate the presentation of tumor-associated antigens and induce tumor-specific immunity. Thus, induction of a T cell-mediated immune response in a human subject may be sufficient to overcome tumor-specific resistance in the subject.

The activity of the proteins of the invention may be measured, in particular, by the following method: Suitable assays for cytotoxicity of thymocytes or spleen cells are Current Protocols.
in Immunology, Ed by JE Coligan, AM Kruisbeek, DH Margulies, EM
Shevach, W Strober, Pub. Greene Publishing Associates and Wiley-Intersc
ience (Chapter 3, In Vitro assays for Mouse Lymphocyte Function 3.1-3.19
; Chapter 7, Immunologic studies in Humans); Herrmann et al., Proc. Natl
Acad. Sci. USA 78: 2488-2492, 1981; Herrmann et al., J. Immunol. 128:
1968-1974, 1982; Handa et al., J. Immunol. 135: 1564-1572, 1985; Takai e.
t al., J. Immunol. 137: 3494-3500, 1986; Takai et al., J. Immunol. 140:
508-512, 1988; Herrmann et al., Proc. Natl. Acad. Sci. USA 78: 2488-2492.
, 1981; Herrmann et al., J. Immunol. 128: 1968-1974, 1982; Handa et al.,
J. Immunol. 135: 1564-1572, 1985; Takai et al., J. Immunol. 137: 3494-3.
500, 1986; Bowman et al., J. Virology 61: 1992-1998; Takai et al., J. Imm.
unol. 140: 508-512, 1988; Bertagnolli et al., Cellular Immunology 133: 3
27-341, 1991; Brown et al., J. Immunol. 153: 3079-3092, 1994, but is not limited thereto. Assays for T cell-dependent immunoglobulin responses and isotype switching, particularly those that modulate T cell-dependent antibody responses and identify proteins that affect the Th1 / Th2 profile, have been described by Maliazewski, J. Immunol. 144: 3028-3033, 1990, including but not limited to assays for B cell function, such as In vitro antibody production, Mond, JJand Brunswick, M. In Current.
Protocols in Immunology JEColigan eds. Val 1 pp.3.8.1-3.8.16, John Wile
y and Sons, Tronto 1994. The mixed lymphocyte reaction (MLR) assay (especially identifying proteins that primarily generate Th1 and CTL responses) is described in Current Protocols in Immunology, Ed by.
JE Coligan, AM Kruisbeek, DH Margulies, EM Shevach, W Strober,
Pub. Greene Publishing Associates and Wiley-Interscience (Chapter 3, In
Vitro assays for Mouse Lymphocyte Function 3.1-3.19; Chapter 7, Immunol
ogic studies in Humans); Takai et al., J. Immunol. 137: 3494-3500, 1986;
Takai et al., J. Immunol. 140: 508-512, 1988; Bertagnolli et al., J. Im
munol. 149: 3778-3783, 1992, but is not limited to these. Dendritic cell-dependent assays, particularly identifying proteins expressed by dendritic cells that activate naive T cells, have been described by Guery et al., J. Immunol. 134: 536-544.
, 1995; Inaba et al., Journal of Experimental Medicine 173: 549-559, 199.
1; Macatonia et al., Journal of Immunology 154: 5071-5079, 1995; Porgado
r et al., Journal of Experimental Medicine 182: 255-260, 1995; Nair et a.
l., Journal of Virology 67: 4062-4069, 1993; Huang et al., Science 264:
961-965, 1994; Macatonia et al., Journal of Experimental Medicine 169:
1255-1264, 1989; Bhardwaj et al., Journal of Clinical Investigation 94:
797-807, 1994; and Inaba et al., Journal of Experimental Medicine 172:
631-640, 1990, but is not limited to these. Lymphocyte survival / apoptosis assays (especially identifying proteins that prevent apoptosis after hyperantibody induction, as well as proteins that regulate lymphocyte homeostasis) are reported by Darzynkiewicz et al., Cytometry 13: 795-808, 1992; Gorczyca et al.
, Leukemia 7: 659-670, 1993; Gorczyca et al., Cancer Research 53: 1945-1.
951, 1993; Itoh et al., Cell 66: 233-243, 1991; Zacharchuk, Journal of I.
mmunology 145: 4037-4045, 1990; Zamai et al., Cytometry 14: 891-897, 199.
3; including but not limited to the assays described in Gorczyca et al., International Journal of Oncology 1: 639-648, 1992. Early stage T cell commitment and developmental assays include:
Antica et al., Blood 84: 111-117,1994; Fine et al., Cellular Immunology 15
5: 111-122, 1994; Galy et al., Blood 85: 2770-2778, 1995; Toki et al., Pr
oc. Natl. Acad. Sci. USA 88: 7548-7551, 1991, but is not limited thereto.

Hematopoiesis-regulating activity The proteins of the present invention are useful in the regulation of hematopoiesis and therefore in the treatment of bone marrow and lymphocytic deficiency. Even the minimal biological activity that supports colony forming cells or factor-dependent cell lines has been implicated in the regulation of hematopoiesis.
For example, erythroid progenitor cells in support of proliferation of erythroid progenitor cells only, or in combination with other cytokines, eg, showing utility in the treatment of various anemias, or in combination with radiation / chemotherapy And / or stimulating the production of erythroid cells; supporting the proliferation of bone marrow cells such as granulocytes and monocytes / macrophages (ie traditional CSF activity), eg in combination with chemotherapy, followed by Useful for preventing the myelosuppression that occurs; useful for supporting the growth of megakaryocytes, and then for the growth of platelets, and thereby preventing or treating various platelet diseases such as thrombocytopenia, It is also commonly used in place of or as a complement to platelet infusion; and / or hematopoietic stem cells (matured above Becomes all of hematopoietic cells, therefore, a variety of stem cell disease (
Diseases that are usually treated by transplantation, such as those found to be useful in aplastic anemia and paroxysmal nocturnal hemoglobinuria), and also in vivo or ex vivo. It is also useful in the repopulation of stem cell compartments as normal cells after radiation / chemotherapy (ie, in combination with bone marrow transplant) or after being genetically engineered for gene therapy. In particular, the activity of the protein of the present invention may be measured by the following method. Assays suitable for proliferation and differentiation of various hematopoietic cell lines have already been cited. Embryonic stem cell differentiation assays, in particular to identify proteins that affect embryonic hematopoiesis, are described by Johansson et al. Cellular Biology 15: 141-151, 1995; Keller et al.
, Molecular and Cellular Biology 13: 473-486, 1993; McClanahan et al., B
lood 81: 2903-2915, 1993, but is not limited to these. Stem cell survival and differentiation assays, particularly identifying proteins that regulate lympho-hematopoiesis, are described in Methylcellulose colony forming assays, Freshney, MG In Cultu.
re of Hematopoietic Cells. RI Freshney, et al. eds. Vol pp. 265-268,
Wiley-Liss, Inc., New York, NY. 1994; Hirayama et al., Proc. Natl. Acad.
Sci. USA 89: 5907-5911, 1992; Primitive hematopoietic colony forming ce
lls with high proliferative potential, McNiece, IK and Briddell, RA
In Culture of Hematopoietic Cells. RI Freshney, et al. Eds. Vol pp. 2
3-39, Wiley-Liss, Inc., New York, NY. 1994; Neben et al., Experimental H
ematology 22: 353-359, 1994; Cobblestone area forming cell assay, Ploema
cher, RE In Culture of Hematopoietic Cells. RI Freshney, et al. eds.
Vol pp. 1-21, Wiley-Liss, Inc .., New York, NY. 1994; Long term bone ma
rrow cultures in the presence of stromal cells, Spooncer, E., Dexter, M.
and Allen, T. In Culture of Hematopoietic Cells. RI Freshney, et al.
eds. Vol pp. 163-179, Wiley-Liss, Inc., New York, NY. 1994; Long term c
ulture initiating cell assay, Sutherland, HJ In Culture of Hematopoiet
ic Cells. RI Freshney, et al. eds. Vol pp. 139-162, Wiley-Liss, Inc.,
Includes, but is not limited to, the assays described in New York, NY. 1994.

Tissue Proliferation Activity The protein of the present invention is used in the composition for the growth or regeneration of bone, cartilage, key, ligament and / or nerve tissue, as well as wound healing and tissue repair, and in the treatment of burns, lacerations and ulcers. Has utility. The proteins of the present invention, which induce cartilage and / or bone growth in environments where bone is not normally formed, find use in the healing of bone fractures and cartilage damage or defects in humans and other animals. Such formulations using the proteins of the invention are useful in reducing closed as well as open fractures and also in improving fixation of artificial joints. De novo bone formation induced by osteogenic agents contributes to the repair of congenital, traumatic, or oncological resection-induced craniofacial defects and is also useful in cosmetic surgery. The proteins of the invention may be used in the treatment of periodontal disease and other tooth repair processes. Such agents provide an environment that attracts osteogenic cells, stimulates the proliferation of osteogenic cells,
Alternatively, it may induce differentiation of osteogenic cell precursors. The protein of the present invention blocks osteoporosis or osteoarthritis by, for example, blocking a process of tissue destruction (collagenase activity, osteoclast activity, etc.) mediated by stimulation of bone and / or cartilage repair or by inflammation or an inflammatory process. May also be useful in the treatment of Another category of tissue developmental activity that may be attributed to the proteins of the present invention is key / ligament formation. Proteins of the invention that induce the formation of key / ligament-like tissues or other tissues in an environment in which other tissues are not normally formed, are useful for preventing key or ligament lacerations, deformities and other key or ligament defects in humans and other animals. Applied to healing. Such formulations using the key / ligament-like tissue-inducing protein may be used to prevent damage to the key or ligament tissue as well as improve the fixation of the key or ligament to bone or other tissue. De novo key / ligament-like tissue formation induced by the composition of the present invention contributes to the repair of congenital, traumatic, or oncological resection-induced craniofacial defects, and further key or ligament attachment. It is also useful in cosmetic surgery for repair. The composition of the present invention provides an environment for attracting key- or ligament-forming cells, stimulates proliferation of key- or ligament-forming cells, and induces differentiation of key- or ligament-forming cell precursors. Or induce ex vivo key / ligament cell or progenitor proliferation to effect tissue repair when returned to in vivo.
The compositions of the present invention are also useful for key inflammation, carpal tunnel syndrome and other key or ligament defects. The compositions of the present invention may include a suitable matrix and / or sequestering agents such as carriers well known in the art. The protein of the present invention is involved in nerve cell proliferation and nerve and brain tissue regeneration, that is, including central and peripheral nervous system diseases and neuropathy as well as mechanical diseases and traumatic diseases (degeneration, death or trauma to nerve cells or nerve tissue. ) May also be useful in the treatment of More specifically, peripheral nervous system disorders such as peripheral nerve injury, peripheral neuropathy and localized neuropathy, as well as Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral myelosclerosis and Shy-Drager.
The protein may be used to treat central nervous system diseases such as syndromes. Further conditions which may be treated according to the present invention include mechanical and traumatic disorders such as spinal cord disorders, head trauma and disorders of the cerebrovascular system such as stroke. Peripheral neuropathy caused by chemotherapy or other medical therapies can be treated with the proteins of the invention. The protein of the present invention is also useful for promoting more preferable and rapid closure of non-healing wounds including (but not limited to) pressure ulcers, ulcers associated with vascular dysfunction, surgical and traumatic wounds, etc. It is possible. Further, the protein of the present invention can be used in other tissues such as organs (including pancreas, liver, intestine, kidney, skin, endothelium), muscles (smooth muscle, skeletal muscle or myocardium), and blood vessels (including vascular endothelium) tissues. Or the activity of promoting the growth of cells constituting such a tissue. Some of the desired effects may be the regeneration of normal tissue by inhibiting fibrotic scarring. The proteins of the invention may also exhibit angiogenic activity. The proteins of the invention may also be useful in the protection or regeneration of the intestine and in the treatment of conditions caused by lung or liver fibrosis, reperfusion injury of various tissues, and systemic cytokine damage. The protein of the present invention promotes or suppresses differentiation of the above tissues from precursor tissues or cells;
Alternatively, it may be useful for suppressing the growth of the above tissues. The activity of the protein of the present invention may be measured, inter alia, by the following method: The tissue regeneration activity assay is carried out according to International Publication WO95 / 16035 (bone, cartilage, key).
International publication WO95 / 05846 (nerve, neuron); International publication WO91 / 0
7491 (skin, endothelium). An assay for wound healing activity is described in Winter, Epidermal Wound Healing, pps. 71-112.
(Maibach, HI and Rovee, DT, eds.), Year Book Medical Publishers, Inc.,
Includes, but is not limited to, those described in Chicago (modified by Eaglstein and Mertz, J. Invest. Dermatol 71: 382-384 (1978)).

Activin / Inhibin Activity The proteins of the invention may also exhibit activin- or inhibin-related activity. Inhibins are characterized by their ability to suppress the release of follicle-stimulating hormone (FSH),
Activins are characterized by their ability to stimulate the release of follicle stimulating hormone (FSH). Thus, the protein of the present invention, alone or in the form of a heterodimer with a member of the inhibin α family, reduces infertility in female mammals and is a fertility drug based on the ability of inhibin to reduce spermatogenesis in male mammals. Can be useful as Administration of a sufficient amount of other inhibins can induce infertility in these mammals. Alternatively, the protein of the invention, as a homodimer or as a heterodimer with a subunit of another protein of the inhibin-β group, is based on the ability of the activin molecule to stimulate FSH release from anterior pituitary cells, based on fertility. It may be useful as a therapeutic agent for inducing For example, US Pat.
See 98885. The proteins of the invention may also be useful in improving reproduction in sexually immature mammals, resulting in increased fertility during the life of livestock such as cattle, sheep and pigs. The activity of the proteins of the invention may be measured, inter alia, by the following method: An assay for activin / inhibin activity is Vale et al., Endocrinology 91:
562-572, 1972; Ling et al., Nature 321: 779-782, 1986; Vale et al., Nat
ure 321: 776-779, 1986; Mason et al., Nature 318: 659-663,1985; Forage e.
Tal., Proc. Natl. Acad. Sci. USA 83: 3091-3095, 1986, but is not limited thereto.

Chemotactic / Chemokinetic Activity The protein of the present invention has the chemotactic or chemokinetic activity of mammalian cells such as monocytes, neutrophils, T cells, mast cells, eosinophils and / or endothelial cells (eg, chemokinetic activity). May act as a chemokine). Chemotactic and chemokinesis proteins can be used to recruit or attract a desired cell population to a desired site of action. Chemotactic or chemokinesis proteins are particularly advantageous for the treatment of tissue damage and other trauma as well as local infections. For example, the attraction of lymphocytes, monocytes or neutrophils to a tumor or site of infection may improve the immune response to the tumor or infectious agent. Certain proteins or peptides have chemotactic activity for particular cell populations and, when stimulating, directly or indirectly direct the direction or movement of such cell populations. Preferably, the protein or peptide has the ability to directly stimulate the directed movement of cells. Whether a particular protein has chemotactic activity on a cell population can be readily determined by using such proteins or peptides in known assays of cell chemotaxis. The activity of the proteins of the invention may be measured, inter alia, by the following methods: An assay for chemotaxis activity (an assay for identifying a protein that induces or interferes with chemotaxis) induces migration across a cell's membrane. Assay to determine the ability of the protein to induce, as well as the ability of the protein to induce the attachment of one cell population to another. Suitable assays for transfer and attachment are Current Protocols in Immunology, Ed by JE
Coligan, AM Kruisbeek, DH Margulies, EM Shevach, W. Strober, Pub. G
reene Publishing Associates and Wiley-Interscience (Chapter 6.12, Measur
ement of alpha and beta Chemokines 6.12.1-6.12.28; Taub et al. J. Clin.
Invest. 95: 1370-1376, 1995; Lind et al. APMIS 103: 140-146, 1995; Mulle
r et al Eur. J. Immunol. 25: 1744-1748; Gruber et al. J. of Immunol. 152
: 5860-5867, 1994; Johnston et al. J. of Immunol. 153: 1762-1768, 1994, but is not limited thereto.

Hemostatic and thrombolytic activity Further , the protein of the present invention may exhibit hemostatic or thrombolytic activity. As a result, such proteins are useful in the treatment of various coagulation disorders, including inherited disorders such as hemophilia, or in the treatment of injuries caused by trauma, surgery or other causes. Can be promoted. The proteins of the invention may be useful in the treatment and prevention of thrombolysis or inhibition of thrombus formation and the conditions resulting therefrom, such as myocardial infarction and central nervous system vascular infarction (eg stroke). The activity of the proteins of the invention may be measured, inter alia, by the following methods: Assays for hemostatic and thrombolytic activity are performed by Linet et al., J. Clin. Pharmacol.
26: 131-140, 1986; Burdick et al., Thrombosis Res. 45: 413-419, 1987; Hum
phrey et al., Fibrinolysis 5: 71-79 (1991); Schaub, Prostaglandins 35: 4
67-474, 1988, but is not limited to these.

Receptor / Ligand Activity The protein of the present invention may also exhibit activity as an inhibitor or an agonist of a receptor, a receptor ligand or a receptor / ligand interaction. Examples of such receptors and ligands include cytokine receptors and their ligands, receptor kinases and their ligands, receptor phosphatases and their ligands, cells-
Receptors involved in cell interactions and their ligands (such as cell adhesion molecules (such as selectins, integrins and their ligands) and receptors / ligands involved in antigen presentation, antigen recognition, generation of cellular and humoral immune responses) (Including pairs and the like), but not limited to these. Receptors and ligands are also useful in screening potential peptide or small molecule inhibitors for related receptor / ligand interactions. The proteins of the present invention, including but not limited to fragments of receptors and ligands, may themselves be useful as inhibitors of receptor / ligand interactions. The activity of the proteins of the invention may be measured, inter alia, by the following methods: Suitable assays of receptor-ligand activity are Current Protocols in Immunolog
y, Ed by JE Coligan, AM Kruisbeek, DH Margulies, EM Shevach,
W. Strober, Pub. Greene Publishing Associates and Wiley-Interscience (C
hapter 7.28, Measurement of Cellular Adhesion under static conditions 7.
28.1-7.28.22), Takai et al., Proc. Natl. Acad. Sci. USA 84: 6864-6868, 1
987; Bierer et al., J. Exp. Med. 168: 1145-1156, 1988; Rosenstein et al.
, J. Exp. Med. 169: 149-160 1989; Stoltenborg et al., J. Immunol. Metho
Ds 175: 59-68, 1994; Stitt et al., Cell 80: 661-670, 1995, but is not limited thereto.

Anti-inflammatory activity The protein of the present invention may exhibit anti-inflammatory activity. Anti-inflammatory activity is the activity of cells-cell interactions (such as adhesion) by stimulating the cells involved in the stimulus response.
By inhibiting or promoting the chemotaxis of cells involved in the inflammatory process, by inhibiting or promoting cell extravasation,
Alternatively, it is exerted by stimulating or suppressing the production of other factors that more directly inhibit or promote the inflammatory response. Inflammation symptoms (
Includes chronic or acute symptoms, mediated by infection-related inflammation (such as septic shock, sepsis or systemic inflammatory response syndrome (SIRS)), ischemia-reperfusion injury, fatal endotoxin injury, arthritis, complement Hyperacute rejection reaction,
(Including but not limited to nephritis, lung injury induced by cytokines or chemokines, inflammatory bowel disease, Crohn's disease or diseases resulting from overproduction of cytokines such as TNF or IL-1). . The proteins of the invention may also be useful in the treatment of anaphylaxis and hypersensitivity to antigenic substances or materials.

Cadherin / Tumor Invasion Suppressing Activity Cadherin is a calcium-dependent adhesion molecule and appears to play a major role during development, specifically in determining specific cell types. Loss or alteration of normal cadherin expression can induce alterations in cell adhesion properties associated with tumor growth and metastasis. Cadherin dysfunction is also involved in other human diseases such as pemphigus vulgaris and pemphigus foliaceus (autoimmune skin blistering disease), Crohn's disease, and some developmental disorders. The cadherin superfamily includes over 40 members, each with different expression patterns. All members of the superfamily have a common conserved extracellular repeat (cadherin domain), but structural differences are found in other parts of the molecule. Calcium is required for their attachment because the cadherin domain binds to calcium to form their tertiary structure. Since a few amino acids in the first cadherin domain provide the basis for homophilic attachment, modification of this recognition site may alter the specificity of cadherin, thus recognizing only itself. Instead, the mutant molecule will also be able to bind to a different cadherin. Moreover, some cadherins are also involved in heterophilic attachment with other cadherins. E-cadherin, a member of the cadherin superfamily, is expressed in epithelial cell types. Pathologically, when E-cadherin expression is lost in the tumor, the malignant cells become invasive and the cancer metastasizes. Transfecting E-cadhedrin-expressing polynucleotides into cancer cell lines restores altered cell morphology, restores cell-cell attachment to other and other substrates, reduces cell growth rate, and reduces scaffolding. By dramatically reducing the proliferation of dependent cells,
Restored the changes associated with cancer. Thus, reintroducing E-cadhedrin expression returns the carcinoma to a less advanced stage. Other cadhedrin may also have the same invasion suppressive role in carcinomas from other tissue types. Therefore, cancer can be treated using the protein of the present invention having cadhedrin activity, and the polynucleotide of the present invention encoding such a protein. Introducing such proteins or polynucleotides into cancer cells may reduce or eliminate the cancerous alterations observed in these cells by providing normal cadhedrin expression. It is also known that cancer cells express cadherin in a tissue type different from the original, so these cancer cells can invade different tissues and can metastasize. The protein of the present invention having cadherin activity and the polynucleotide of the present invention encoding such a protein substitute for cadhedrin which is inappropriately expressed in these cells, restore normal cell adhesion properties, and reduce the metastatic tendency of cells. It can be removed or removed. Furthermore, the protein of the present invention having cadherin activity and the polynucleotide of the present invention encoding such a protein can be used to obtain an antibody that recognizes cadherin and binds to it. Such antibodies can also be used to block the attachment of inappropriately expressed tumor cell cadhedrin, preventing cells from forming tumors elsewhere. Such anti-cadhedrin antibodies can be used as markers for cancer grade, pathological type, and prognosis. That is, as the cancer progresses, the expression of cadhedrin decreases, and this decrease in cadhedrin expression can be detected by using a cadhedrin-binding antibody. A fragment of a protein of the invention having cadhedrin activity, preferably a decapeptide at the cadhedrin recognition site, and a polynucleotide of the invention which encodes such a protein fragment are used to bind to cadherin and prevent cadherin binding which results in undesirable effects. Can also block cadherin function. Further, using a fragment of the protein of the present invention having cadherin activity, preferably a truncated soluble cadhedrin fragment known to be stable in the circulatory system of cancer patients, and a polynucleotide encoding such a protein fragment, It can also disrupt correct cell-cell attachment. An assay for cadhedrin adhesion and entry inhibition activity was performed by Hortsch et al. J Bio
l Chem 270 (32): 18809-18817, 1995; Miyaki et al. Oncogene 11: 2547-2552.
, 1995; Ozawa et al. Cell 63: 1033-1038, 1990.

Tumor inhibitory activity In addition to the above-mentioned activities relating to immunotherapy or prevention of tumors, the protein of the present invention may exhibit other antitumor activity. Certain proteins directly or indirectly (e.g.,
(Via ADCC). Certain proteins act on tumor tissue or tumor precursor tissue to inhibit the formation of tissue necessary to support tumor growth (eg, by inhibiting angiogenesis) and to inhibit tumor growth in others. Tumor inhibitory activity may be exhibited by causing the production of a factor, agent or cell type, or by removing or inhibiting a factor, agent or cell type that promotes tumor growth.

Other Activities The proteins of the present invention may exhibit one or more of the following additional activities or effects: of infectious agents including but not limited to bacteria, viruses, fungi and other parasites. Sacrifice; height, weight, hair color, eye color, pigmentation of skin or other tissues, or size or morphology of organs or body parts (eg breast augmentation or vice versa)
Effects on body characteristics including (suppressing or promoting); effects on digestion of ingested fat, protein or carbohydrate; appetite, libido, stress, cognition (including cognitive disorders), depression (including depressive disorders) ) And behavioral traits including but not limited to violent behavior; providing analgesic or other pain-relieving effects; promoting differentiation and proliferation of embryonic stem cells in non-hematopoietic lineages; hormones or endocrine Activity; in the case of enzymes,
Correction of enzyme deficiency and treatment of related disorders; treatment of hyperproliferative disorders (such as psoriasis); immunoglobulin-like activity (eg, ability to bind antibody or complement);
And the ability to act as an antigen in a vaccine composition to generate an immune response to such protein or other substances or entities that cross-react with such protein.

Administration and Dose The protein of the invention, which may be from any source, including but not limited to recombinant and non-recombinant methods, is mixed with a pharmaceutically acceptable carrier. May be used in a pharmaceutical composition. Such compositions may contain (in addition to protein and carrier) diluents, fillers, salts, buffers, stabilizers, solubilizers, and other materials well known in the art. The term "pharmaceutically acceptable" means
A non-toxic substance that does not interfere with the effectiveness of the biological activity of the active ingredient. The characteristics of the carrier will depend on the route of administration. The pharmaceutical composition of the present invention comprises M-CSF, GM-CSF, T
NF, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-
7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13,
IL-14, IL-15, IFN, TNF0, TNF1, TNF2, G-CSF
, Meg-CSF, thrombopoietin, stem cell factor, and cytokines such as erythropoietin, lymphokines, or other hematopoietic factors. The pharmaceutical composition may further contain other agents which enhance protein activity or supplement its activity or utility in therapy. Such additional factors and / or agents may be included in the pharmaceutical composition to exert a synergistic effect with the protein of the invention or to minimize side effects. On the contrary, a specific cytokine, lymphokine, other hematopoietic factor, thrombolytic factor or antithrombotic factor, or anti-inflammatory agent formulation containing the protein of the present invention, cytokine, lymphokine, other hematopoietic factor, thrombolytic factor or The side effects of antithrombotic factors, or anti-inflammatory agents, may be minimized. The protein of the present invention may be active in the form of a multimer (eg, heterodimer or homodimer) or a complex with itself or another protein. Consequently, the pharmaceutical composition of the present invention may contain the protein of the present invention in the form of such a multimer or complex. The pharmaceutical composition of the present invention may be in the form of a complex of the protein of the present invention and a protein or peptide antigen. Protein and / or peptide antigens can be applied to B lymphocytes and T
It will deliver a stimulatory signal to lymphocytes. B lymphocytes will respond to antigen through their surface immunoglobulin receptors. T lymphocytes will respond to antigen through the T cell receptor (TCR) after antigen presentation by MHC proteins.
Structurally related proteins, including those encoded by MHC and host cell class I and class II MHC genes, will serve to present peptide antigens to T lymphocytes. The antigenic component can be supplied as a purified MHC-peptide complex alone or with a costimulatory molecule that can directly signal T cells. Alternatively, surface immunoglobulin on B cells and T on T cells
Antibodies capable of binding CR and other molecules can also be mixed with the pharmaceutical compositions of the invention. The pharmaceutical composition of the present invention may be in the form of liposomes, in which the protein of the present invention is an amphipathic agent such as another pharmaceutically acceptable carrier or lipid (in the form of an aggregate such as a micelle in an aqueous solution). , Present as an insoluble monolayer, liquid crystal or lamellar layer). Suitable lipids for liposome formulations include, but are not limited to, monoglycerides, diglycerides, sulfatizides, lysolecithins, phospholipids, saponins, bile acids and the like. The manufacture of such liposome formulations is well within the level of ordinary skill in the art, see, eg, US Pat. Nos. 4,235,87, 4501728, 4837028,
And 4737323, which are hereby incorporated by reference in their entirety.

The term “therapeutically effective amount” as used herein refers to each of the pharmaceutical compositions or methods sufficient to exhibit a significant patient benefit, eg, amelioration of symptoms of such symptoms, cure, increased rate of healing. It means the total amount of active ingredients. When applied to an individual active ingredient, administered alone, the term refers to that ingredient alone. When used as a combination of active ingredients, it refers to the total amount of active ingredients that produce a therapeutic effect regardless of sequential administration or simultaneous administration. In practicing the therapeutic methods of this invention, a therapeutically effective amount of a protein of this invention is administered to a mammal having the condition to be treated. According to the method of the present invention, the protein of the present invention may be administered alone or in combination with other therapeutic agents such as cytokines, lymphokines or other hematopoietic factors. When co-administered with one or more cytokines, lymphokines or other hematopoietic factors, the proteins of the invention may be administered simultaneously or sequentially with cytokines, lymphokines, other hematopoietic factors, thrombolytic or antithrombotic factors. When administered sequentially, the attending physician will decide on the appropriate sequence of administration of the protein of the invention in combination with cytokines, lymphokines, other hematopoietic factors, thrombolytic or antithrombotic factors. Administration of the protein of the present invention in the pharmaceutical composition of the present invention or used in the practice of the present invention can be carried out by various conventional methods, such as oral feeding, inhalation, or intradermal, subcutaneous, or intravenous injection. Intravenous injection into the patient is preferred. When a therapeutically effective amount of a protein of the invention is administered orally, the protein of the invention will be in the form of tablets, capsules, powders, solutions or elixirs. When administered in tablet form, the pharmaceutical composition of the present invention may further contain a solid carrier such as gelatin or an adjuvant. Tablets, capsules, and powders contain about 5 to 95% of the protein of the invention,
It preferably contains about 25 to 90% of the protein of the present invention. When administered in liquid form, water, petroleum, fats of animal or vegetable origin, such as peanut oil, mineral oil, soybean oil, or sesame oil, or synthetic fats and oils may be added. The liquid form of the pharmaceutical composition may further contain physiological saline solution, dextrose or other saccharide solution, or glycols such as ethylene glycol, propylene glycol or polyethylene glycol. When administered in liquid form, the pharmaceutical composition contains from about 0.5 to 90% by weight of the protein of the invention, preferably from about 1 to 50%. When a therapeutically effective amount of the protein of the present invention is administered by intravenous, intradermal or subcutaneous injection,
The protein of the present invention will be in the form of a pyrogen-free, parenterally acceptable aqueous solution. The preparation of such parenterally acceptable protein solutions with appropriate pH, isotonicity and stability is within the skill of the art. Preferred pharmaceutical compositions for intravenous, intradermal, or subcutaneous injection include sodium chloride for injection, Ringer's solution, dextrose for injection, dextrose and sodium chloride for injection, lactate-containing Ringer's solution for injection, in addition to the protein of the present invention. Or it should contain other carriers known in the art. The pharmaceutical compositions of the present invention may contain stabilizers, preservatives, buffers, antioxidants, or other additives known to those of skill in the art. The amount of the protein of the invention in the pharmaceutical composition of the invention depends on the nature and severity of the condition to be treated,
As well as the nature of the treatment the patient has already received. In the end, the doctor in charge
Each patient will determine the amount of protein of the invention to be treated. First, the attending physician administers a low dose of the protein of the present invention and observes the patient's response. Higher doses of the protein of the invention may be administered until the optimal therapeutic effect is obtained, and the dose is not further increased at the time when the optimal therapeutic effect is obtained. Various pharmaceutical compositions for carrying out the method of the present invention include about 0.01 μg to about 100 mg of the protein of the present invention (preferably,
It should contain from about 0.1 μg to about 10 mg / kg body weight, more preferably from 0.1 μg to about 1 mg / kg body weight of the protein of the present invention. The duration of intravenous treatment with the compositions of the present invention will vary depending on the severity of the disease to be treated and the condition and response of each patient. Each application period of the protein of the present invention is considered to be in the range of 12 to 24 hours in the case of continuous intravenous administration. Ultimately, the attending physician will decide the duration of treatment by intravenous administration with the pharmaceutical composition of the present invention.

The protein of the present invention may be used to immunize animals to obtain polyclonal and monoclonal antibodies that specifically react with the protein of the present invention. Whole antibodies or fragments thereof may be used as immunogens to obtain such antibodies. The peptide immunogen may further include a cysteine residue at the carboxy terminus, which binds to a hapten such as keyhole limpet hemocyanin (KLH). Methods of synthesizing such peptides are known in the art, eg, RP Merrifield, J. Amer. Che.
m. Soc. 85, 2149-2154 (1963); JL Krstenansky, et. al., FEBS Lett. 211.
, 10 (1987). A monoclonal antibody that binds to the protein of the present invention may be a useful diagnostic agent for immunodetection of the protein of the present invention. The neutralizing antibody that binds to the protein of the present invention may be useful as a therapeutic agent for conditions associated with the protein of the present invention,
Furthermore, it may be useful in the treatment of certain tumors in the treatment of some forms of cancer in which aberrant expression of the proteins of the invention is implicated. For cancer cells or white blood cells,
A neutralizing monoclonal antibody against the protein of the present invention may be useful for detecting and preventing metastatic spread of cancer cells which can be mediated by the protein of the present invention. With respect to the composition of the present invention useful for the regeneration of bone, cartilage, tendon or ligament, the method of treatment comprises
It includes administering the composition locally, systemically, or locally, such as as an implant or device. When administered, the therapeutic compositions used in this invention are, of course, in a pyrogen-free, physiologically acceptable form. In addition, the composition may desirably be encapsulated or injected as a viscous form for delivery to the site of bone, cartilage or tissue damage. Topical administration is suitable for wound healing and tissue repair. A therapeutically useful agent other than the protein of the present invention, which may be contained in the above composition, may alternatively or additionally be administered simultaneously or sequentially with the composition in the method of the present invention. Preferably, for bone and / or cartilage formation, the composition delivers the protein-containing composition to the site of bone and / or cartilage damage, provides a structure for bone and cartilage development, and more optimally Will contain a matrix that can be absorbed by the body. Such matrices may be made of the materials currently used in other implanted medical devices. The choice of matrix material is based on biocompatibility, biodegradability, mechanical properties, cosmetic appearance and interface properties. The particular formulation of a composition will determine the appropriate formulation. Possible matrices for the composition are biodegradable and may be the chemically known calcium sulfate, tricalcium phosphate, hydroxyapatite, polylactic acid, polyglycolic acid and polyanhydrides. Other materials that can be used are biodegradable, well known biologically, such as bone or skin collagen. In addition, the matrix may contain pure proteins or extracellular matrix components.
Other possible matrices are chemically known, not biodegradable, such as sintered hydroxyapatite, bioglass, aluminates, or other ceramics. The matrix may comprise a combination of materials of the above type, for example polylactic acid and hydroxyapatite or collagen and tricalcium phosphate. The bioceramics may be modified in the composition, e.g. calcium-aluminate-phosphate and processed to produce pore size, particle size,
The particle shape and biodegradability may be varied. A 50:50 (molar weight) copolymer of lactic acid and glycolic acid in the form of porous particles having a diameter in the range of 150 to 800 microns is presently preferred. In some applications, it may be useful to use a sequestering agent such as carboxymethyl cellulose or autologous blood clots to prevent dissociation of the protein composition from the matrix. A preferred family of sequestrants is cellulosic materials such as alkylcelluloses (including hydroxyalkylcelluloses), including methylcellulose, ethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, and carboxymethylcellulose, carboxymethylcellulose (CMC). Most preferred are the cationic salts of
Other preferred sequestrants include hyaluronic acid, sodium alginate, poly (ethylene glycol), polyoxyethylene oxide, carboxyvinyl polymers and poly (vinyl alcohol). The amount of sequestrant useful in the present invention is 0.5 to 20% by weight, preferably 1 to 10% by weight, based on the total formulation weight, which amount is
An amount that prevents desorption of the protein from the polymer matrix and allows for proper handling of the composition, but provides an opportunity for the progenitor cells to infiltrate the matrix, thereby promoting the osteogenic activity of the progenitor cells. Not too much to be granted to.

In a further composition, the protein of the invention may be mixed with other agents that are beneficial in the treatment of bone and / or cartilage defects, wounds, or tissues. These agents include epidermal growth factor (EGF), platelet-derived growth factor (PDGF), transforming growth factors (TGF-α and TGF-β), and insulin-like growth factor (IGF). Also, therapeutic compositions are currently of value in veterinary applications. In detail,
In addition to humans, livestock and thoroughbred horses are desirable patients for such treatment with the proteins of the invention. The administration rules of the protein-containing pharmaceutical composition used for tissue regeneration are various factors that change the action of protein, such as the tissue weight desired to be formed, the site of damage, the condition of damaged tissue, the size of wound, It will be determined by the physician in view of the type of tissue required (eg, bone), patient age, sex, and diet, severity of infection, duration of administration, and other clinical factors. The dose may vary depending on the type of matrix used for reconstitution and the inclusion of other proteins in the pharmaceutical composition. For example, IGF
The addition of other known growth factors such as I (insulin-like growth factor I) to the final composition can also affect the dose. The progress can be monitored by periodic assessment of tissue / bone growth and / or repair, for example by X-rays, histomorphological examination and tetracycline labeling. The polynucleotide of the present invention can also be used for gene therapy. Such polynucleotides can be introduced into cells in vivo or ex vivo and expressed in a mammalian subject. The polynucleotides of the present invention may be administered by other known methods for introducing nucleic acids into cells or organisms (including but not limited to in a viral vector or in the form of naked DNA). ). Cells may be cultured ex vivo in the presence of the protein of the invention to proliferate, or to produce a desired effect on such cells, or to produce a desired activity in such cells. The treated cells can then be introduced in vivo for therapeutic purposes. The patents and publications cited herein are hereby incorporated by reference in their entirety.

[0050]

[Brief description of drawings]

1A and 1B: pED used to deposit the clones disclosed herein.
6 is a schematic representation of the 6 and pNOTs vectors, respectively.

[Sequence list]

─────────────────────────────────────────────────── ─── Continued Front Page (51) Int.Cl. ⁷ Identification Code FI Theme Coat (Reference) A61P 7/06 A61P 15/16 4H045 11/00 15/18 15/16 17/02 15/18 19/02 17/02 19/08 19/02 19/10 19/08 25/00 19/10 29/00 25/00 101 29/00 35/00 101 37/06 35/00 43/00 105 37/06 C07K 14 / 47 43/00 105 C12P 21/02 C C07K 14/47 C12Q 1/68 A C12N 5/10 C12N 15/00 ZNAA C12P 21/02 5/00 B C12Q 1/68 A61K 37/02 (81) Designated country EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE), OA (BF, BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, K E, LS, MW, SD, SZ, UG, ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), AL, AM, AT, AU, AZ, BA, BB, BG, BR, BY, CA, CH, CN, CU, CZ, DE, DK, EE, ES, FI, GB, GE, GH, GM, HR, HU, ID , IL, IS, JP, KE, KG, KP, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MD, MG, MK, MN, MW, MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, TJ, TM, TR, TT, UA, UG, UZ, VN, YU, ZW (72) Inventor John M. McCoy United States 01867 Inventor, 56 (72), Howard Street, Reading, Massachusetts Doward Earl Larry United States 01451 Ann Lee Road, Harvard, Massachusetts 113 (72) Inventor Lisa A. Collins-Lacy United States 01720 Acton, Massachusetts 124 (72) Inventor David Marburg United States 01720 Orchard Drive, Acton, Mass. 2 (72) Inventor Maurice Tracy United States 02167 Chestnut Hill, Massachusetts Walcot Rod 93 (72) Inventor Michael Jay Agostino United States 01810 Andover, Massachusetts Walcott Avenue No. 26 (72) Inventor Robert Jay Steininger The Second United States 02140 100 Reed Street, Cambridge, Massachusetts, USA No. F term (reference) 4B024 AA01 BA21 CA04 DA02 EA04 GA11 HA01 4B063 QA01 QQ43 QQ49 QR32 QR55 QS34 4B064 AG02 CA10 CA19 CC24 DA01 4B065 AA90X AA99Y AB01 AC14 BA02 ZA51 ZA53 ZA51 BA53 CA02 BA53 BA02 BA08 BA02 BA02 BA08 BA02 BA08 BA02 BA08 BA02 BA08 BA02 BA08 ZA67 ZA86 ZA89 ZA94 ZA96 ZA97 ZB02 ZB11 ZB13 ZB15 ZB26 4H045 AA10 AA30 BA10 CA40 DA01 EA28 FA74

Claims (32)

[Claims] 1. (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1; (b) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1 from nucleotide 257 to nucleotide 622; (c) a deposit under accession number ATCC 98623. Clone ff168 1
A polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of 2; (d) clone ff168_1 deposited under accession number ATCC 98623.
A polynucleotide encoding the full-length protein encoded by the cDNA insert of 2; (e) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 2; (f) a SEQ ID NO: having biological activity : A polynucleotide encoding a protein containing a fragment of the amino acid sequence of SEQ ID NO: 2 (the fragment is 8 of SEQ ID NO: 2).
(G) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(f) above; and (h) is shown in (a)-(f) under stringent conditions. An isolated polynucleotide selected from the group consisting of polynucleotides capable of hybridizing to any one of the polynucleotides. 2. The polynucleotide of claim 1 operably linked to at least one expression control sequence. 3. A host cell transformed with the polynucleotide of claim 2. 4. The host cell of claim 3, which is a mammalian cell. 5. A polynucleotide encoded by claim 2 comprising: (a) growing a culture of the host cell of claim 3 in a suitable medium; and (b) purifying the protein from the culture. A method for producing a protein. 6. A protein produced by the method of claim 5. 7. An isolated polynucleotide encoding the protein of claim 6. 8. A clone ff168 deposited as ATCC 98623.
The polynucleotide of claim 7 comprising 12 cDNA inserts. 9. (a) an amino acid sequence of SEQ ID NO: 2; (b) a fragment of the amino acid sequence of SEQ ID NO: 2 (each fragment containing 8 consecutive amino acids of SEQ ID NO: 2); and (c ) Clone ff168 1 deposited under accession number ATCC 98623
A protein containing an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts of 2 and containing substantially no other mammalian proteins. 10. The protein of claim 9 wherein said protein comprises the amino acid sequence of SEQ ID NO: 2. 11. A composition comprising the protein of claim 9 and a pharmaceutically acceptable carrier. 12. A method for producing an isolated polynucleotide, which method comprises the following: (a) a method comprising the following steps: (i) selected from the group consisting of the following at 65 ° C. in 6X SSC: Preparing one or more polynucleotide probes that hybridize to the nucleotide sequence; (aa) SEQ ID NO: 1 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 1); and (ab) ATCC 98623. Clone ff168 12 deposited as
The nucleotide sequence of the cDNA insert of (ii) hybridizing the probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by the probe (s); and (B) a method comprising the following steps: (i) preparing one or more polynucleotide primers that hybridize in 6X SSC at 65 ° C. to a nucleotide sequence selected from the group consisting of: ba) SEQ ID NO: 1 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 1); and (bb) clone ff168 12 deposited as ATCC 98623.
Sequence of the cDNA insert of (ii) hybridizing the primer (s) to human DNA; (iii) amplifying the human DNA sequence; and then (iv) the polynucleotide of step (b) (iii). The isolated polynucleotide selected from the group consisting of isolating the product, wherein the isolated polynucleotide is: (v) corresponding to the cDNA sequence of SEQ ID NO: 1, 5 of SEQ ID NO: 1
It is a continuous sequence from the nucleotide sequence corresponding to the “end to the nucleotide sequence corresponding to the 3 ′ end of SEQ ID NO: 1, but the poly (A
And (w) nucleotides 257 to 6 of the cDNA sequence of SEQ ID NO: 1.
22 from the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 257 to nucleotide 622 of SEQ ID NO: 1 to 3 of the sequence from nucleotide 257 to nucleotide 622 of SEQ ID NO: 1. 'A method comprising a nucleotide sequence selected from the group consisting of nucleotide sequences that are contiguous up to the nucleotide sequence corresponding to the end. 13. An isolated polynucleotide produced by the method of claim 12. 14. An isolated polynucleotide comprising the polynucleotide of claim 13. 15. (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 3; (b) a polynucleotide comprising the nucleotide sequence of nucleotides 1323 to 1829 of SEQ ID NO: 3; (c) a sequence of SEQ ID NO: 3 A polynucleotide comprising the nucleotide sequence of nucleotides 1539 to 1829; (d) a polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of clone ls91 deposited under accession number ATCC 98623; (e) deposited under accession number ATCC 98623 Clone of clone ls91
A polynucleotide encoding the full-length protein encoded by the DNA insert; (f) a polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of clone ls91 deposited under accession number ATCC 98623; (g) deposit under accession number ATCC 98623 Of cloned clone ls91
A polynucleotide encoding a mature protein encoded by the DNA insert; (h) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 4; (i) a biologically active SEQ ID NO: 4 A polynucleotide encoding a protein containing a fragment of the amino acid sequence of SEQ ID NO: 4
Individual contiguous amino acids); (j) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; (k) a species homologue of the protein of (h) or (i) above. An encoding polynucleotide; and (l) an isolated polynucleotide selected from the group consisting of polynucleotides capable of hybridizing to any one of the polynucleotides (a) to (i) under stringent conditions. . 16. (a) an amino acid sequence of SEQ ID NO: 4; (b) a fragment of the amino acid sequence of SEQ ID NO: 4 (each fragment containing 8 consecutive amino acids of SEQ ID NO: 4); and (c ) C of clone ls91 deposited under accession number ATCC 98623
A protein comprising an amino acid sequence selected from the group consisting of amino acid sequences encoded by DNA inserts, which is substantially free of other mammalian proteins. 17. A method for producing an isolated polynucleotide, which method comprises: (a) a method comprising the following steps: (i) selected from the group consisting of the following at 65 ° C. in 6X SSC: Preparing one or more polynucleotide probes that hybridize to the nucleotide sequence; (aa) SEQ ID NO: 3 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 3); and (ab) ATCC 98623. Of clone ls91 deposited as
Nucleotide sequence of NA insert (ii) hybridizing said probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by said probe (s); and b) a method comprising the steps of: (i) preparing one or more polynucleotide primers that hybridize at 65 ° C. in 6X SSC to a nucleotide sequence selected from the group consisting of: (ba ) SEQ ID NO: 3 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 3); and (bb) cD of clone ls91 deposited as ATCC 98623.
Nucleotide sequence of NA insert (ii) hybridizing said primer (s) to human DNA; (iii) amplifying human DNA sequence; and then (iv) producing polynucleotide of step (b) (iii). Wherein the isolated polynucleotide is selected from the group consisting of: (v) corresponding to the cDNA sequence of SEQ ID NO: 3, 5 of SEQ ID NO: 3
The sequence from the nucleotide sequence corresponding to the “end to the nucleotide sequence corresponding to the 3 ′ end of SEQ ID NO: 3 is continuous, but the poly (A
) A nucleotide sequence in which the tails are excluded; (w) a sequence corresponding to nucleotides 1323 to 1829 of the cDNA sequence of SEQ ID NO: 3, of the sequence of nucleotides 1323 to 1829 of SEQ ID NO: 3 A nucleotide sequence that is a sequence from the nucleotide sequence corresponding to the 5'end to the nucleotide sequence corresponding to the 3'end of the sequence from nucleotide 1323 to nucleotide 1829 of SEQ ID NO: 3; and (x) SEQ ID NO: 3 from the nucleotide sequence corresponding to nucleotides 1539 to 1829 of the cDNA sequence of SEQ ID NO: 3 and corresponding to the 5'end of the sequence from nucleotide 1539 to nucleotide 1829 of SEQ ID NO: 3 to nucleotide 1539 of SEQ ID NO: 3 Or The method is intended to include a nucleotide sequence selected from the group consisting of continuous those nucleotide sequences to the nucleotide sequence corresponding to the 3 'end of the sequence to nucleotide 1829. 18. (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 5; (b) a polynucleotide comprising the nucleotide sequence of nucleotides 507 to 722 of SEQ ID NO: 5; (c) a sequence of SEQ ID NO: 5 A polynucleotide comprising the nucleotide sequence of nucleotides 615 to 722; (d) clone na1010 deposited under accession number ATCC 98623.
A polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of 1; (e) clone na1010 deposited under accession number ATCC 98623.
A polynucleotide encoding the full-length protein encoded by the cDNA insert of 1; (f) clone na1010 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of 1; (g) clone na1010 deposited under accession number ATCC 98623.
A polynucleotide encoding a mature protein encoded by the cDNA insert of 1; (h) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 6; (i) a SEQ ID NO having biological activity A polynucleotide encoding a protein containing a fragment of the amino acid sequence of SEQ ID NO: 6 (the fragment is 8 of SEQ ID NO: 6).
(A) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; and (k) is shown under (a)-(i) under stringent conditions. An isolated polynucleotide selected from the group consisting of polynucleotides capable of hybridizing to any one of the polynucleotides. 19. (a) an amino acid sequence of SEQ ID NO: 6; (b) a fragment of the amino acid sequence of SEQ ID NO: 6 (each fragment comprising 8 consecutive amino acids of SEQ ID NO: 6); and (c ) Clone na1010 deposited under accession number ATCC 98623
1. A protein containing an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts of 1 and containing substantially no other mammalian proteins. 20. A method for producing an isolated polynucleotide, which comprises the following method: (a) a method comprising the following steps: (i) selected from the group consisting of the following at 65 ° C. in 6X SSC: Preparing one or more polynucleotide probes that hybridize to the nucleotide sequence; (aa) SEQ ID NO: 5 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 5); and (ab) ATCC 98623. Clone na10101 deposited as
The nucleotide sequence of the cDNA insert of (ii) hybridizing the probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by the probe (s); and (B) a method comprising the following steps: (i) preparing one or more polynucleotide primers that hybridize in 6X SSC at 65 ° C. to a nucleotide sequence selected from the group consisting of: ba) SEQ ID NO: 5 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 5); and (bb) clone na10101 deposited as ATCC 98623.
Sequence of the cDNA insert of (ii) hybridizing the primer (s) to human DNA; (iii) amplifying the human DNA sequence; and then (iv) the polynucleotide of step (b) (iii). The isolated polynucleotide selected from the group consisting of isolating the product is as follows: (v) corresponds to the cDNA sequence of SEQ ID NO: 5, 5 of SEQ ID NO: 5
The sequence from the nucleotide sequence corresponding to the “end to the nucleotide sequence corresponding to the 3 ′ end of SEQ ID NO: 5 is continuous, but the poly (A
) Nucleotide sequence without tails; (w) nucleotides 507 to 7 of the cDNA sequence of SEQ ID NO: 5
22 corresponding to the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 507 to nucleotide 722 of SEQ ID NO: 5 to 3 of the sequence from nucleotide 507 to nucleotide 722 of SEQ ID NO: 5. 'A nucleotide sequence that is a sequence up to the nucleotide sequence corresponding to the end; and (x) nucleotides 615 to 7 of the cDNA sequence of SEQ ID NO: 5.
22 from the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 615 to nucleotide 722 of SEQ ID NO: 5 to 3 of the sequence from nucleotide 615 to nucleotide 722 of SEQ ID NO: 5. 'A method comprising a nucleotide sequence selected from the group consisting of nucleotide sequences that are contiguous up to the nucleotide sequence corresponding to the end. 21. (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 7; (b) a polynucleotide comprising the nucleotide sequence of nucleotide 673 to nucleotide 978 of SEQ ID NO: 7; (c) the nucleotide sequence of SEQ ID NO: 7. A polynucleotide comprising the nucleotide sequence from nucleotide 868 to nucleotide 987; (d) a polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of clone nf87_1 deposited under accession number ATCC 98623; (e) deposited under accession number ATCC 98623 A polynucleotide encoding the full-length protein encoded by the cDNA insert of clone nf87 1; (f) the mature protein coding sequence of clone nf87 1 deposited under accession number ATCC 98623 (G) a polynucleotide containing the amino acid sequence of SEQ ID NO: 8; (g) a polynucleotide containing a cleotide sequence; (g) a polynucleotide encoding the mature protein encoded by the cDNA insert of clone nf87_1 deposited under accession number ATCC 98623; (I) a polynucleotide encoding a protein containing a fragment of the amino acid sequence of SEQ ID NO: 8 having biological activity (the fragment being 8 of SEQ ID NO: 8);
(A) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; and (k) is shown under (a)-(i) under stringent conditions. An isolated polynucleotide selected from the group consisting of polynucleotides capable of hybridizing to any one of the polynucleotides. 22. (a) an amino acid sequence of SEQ ID NO: 8; (b) a fragment of the amino acid sequence of SEQ ID NO: 8 (each fragment containing 8 consecutive amino acids of SEQ ID NO: 8); and (c ) A protein comprising an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts of clone nf87_1 deposited under accession number ATCC 98623, which is essentially free of other mammalian proteins. 23. A method for producing an isolated polynucleotide, which method comprises: (a) a method comprising the following steps: (i) selected from the group consisting of the following at 65 ° C. in 6X SSC: Preparing one or more polynucleotide probes that hybridize to the nucleotide sequence; (aa) SEQ ID NO: 7 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 7); and (ab) ATCC 98623. C of clone nf87 1 deposited as
Nucleotide sequence of the DNA insert: (ii) hybridizing the probe (s) to human DNA; and (iii) isolating the DNA polynucleotide detected by the probe (s); and b) a method comprising the steps of: (i) preparing one or more polynucleotide primers that hybridize at 65 ° C. in 6X SSC to a nucleotide sequence selected from the group consisting of: (ba ) SEQ ID NO: 7 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 7); and (bb) c of clone nf871 deposited as ATCC 98623.
Nucleotide sequence of the DNA insert (ii) hybridizing the primer (s) to human DNA; (iii) amplifying the human DNA sequence; and then (iv) producing the polynucleotide of step (b) (iii). Wherein the isolated polynucleotide is selected from the group consisting of: (v) corresponding to the cDNA sequence of SEQ ID NO: 7, 5 of SEQ ID NO: 7
The sequence from the nucleotide sequence corresponding to the “terminal to the nucleotide sequence corresponding to the 3 ′ terminal of SEQ ID NO: 7 is continuous, but the poly (A
) Nucleotide sequence without tails; (w) nucleotides 673 to 9 of the cDNA sequence of SEQ ID NO: 7
87 to the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 673 to nucleotide 987 of SEQ ID NO: 7 to 3 of the sequence from nucleotide 673 to nucleotide 987 of SEQ ID NO: 7. 'A nucleotide sequence that is contiguous up to the nucleotide sequence corresponding to the end; and (x) nucleotides 868 to 9 of the cDNA sequence of SEQ ID NO: 7.
87 to the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 868 to nucleotide 987 of SEQ ID NO: 7 to 3 of the sequence from nucleotide 868 to nucleotide 987 of SEQ ID NO: 7. 'A method comprising a nucleotide sequence selected from the group consisting of nucleotide sequences that are contiguous up to the nucleotide sequence corresponding to the end. 24. (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 9; (b) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 9 from nucleotide 57 to nucleotide 824; (c) a SEQ ID NO: 9 A polynucleotide comprising the nucleotide sequence of nucleotides 114 to 824; (d) clone nh796_1 deposited under accession number ATCC 98623.
A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence of: (e) clone nh796_1 deposited under accession number ATCC 98623
A polynucleotide encoding the full-length protein encoded by the cDNA insert of: (f) clone nh7961 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of: (g) clone nh7961 deposited under accession number ATCC 98623.
(H) a polynucleotide encoding a protein comprising the amino acid sequence of SEQ ID NO: 10; (i) a biologically active SEQ ID NO: A polynucleotide encoding a protein comprising a fragment of the amino acid sequence of 10 (the fragment being SEQ ID NO: 10
8 consecutive amino acids); (j) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; and (k) (a)-(i) under stringent conditions. An isolated polynucleotide selected from the group consisting of polynucleotides capable of hybridizing to any one of the polynucleotides shown in. 25. (a) an amino acid sequence of SEQ ID NO: 10; (b) a fragment of the amino acid sequence of SEQ ID NO: 10 (each fragment comprising 8 consecutive amino acids of SEQ ID NO: 10); and (c ) Clone nh7961 deposited under accession number ATCC 98623
A protein containing an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts, which is substantially free of other mammalian proteins. 26. A method for producing an isolated polynucleotide, the method comprising: (a) a method comprising the following steps: (i) selected from the group consisting of the following at 65 ° C. in 6X SSC: Preparing one or more polynucleotide probes that hybridize to the nucleotide sequence; (aa) SEQ ID NO: 9 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 9); and (ab) ATCC 98623. Sequence of the cDNA insert of clone nh7961 deposited as (ii) hybridizing said probe (s) to human DNA; and (iii) a DNA polynucleotide detected with said probe (s). And (b) a method comprising the following steps: (i) 6 in 6X SSC Preparing one or more polynucleotide primers that hybridize at ° C to a nucleotide sequence selected from the group consisting of: (ba) SEQ ID NO: 9 (polynucleotide at the 3'end of SEQ ID NO: 9 (A) excluding tail); and (bb) nucleotide sequence of cDNA insert of clone nh796_1 deposited as ATCC 98623 (ii) hybridizing the primer (s) to human DNA; (iii) human DNA Amplifying the sequence; then (iv) isolating the polynucleotide product of step (b) (iii), wherein the isolated polynucleotide is: (v) SEQ ID NO: 9 5 corresponding to the cDNA sequence of SEQ ID NO: 9
The sequence from the nucleotide sequence corresponding to the “end to the nucleotide sequence corresponding to the 3 ′ end of SEQ ID NO: 9 is continuous, but the poly (A
) Nucleotide sequence without tails; (w) nucleotide 57 to nucleotide 82 of the cDNA sequence of SEQ ID NO: 9.
4 corresponds to the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 57 to nucleotide 824 of SEQ ID NO: 9
9 a nucleotide sequence that is contiguous from nucleotide 57 to nucleotide 824 of 9 to the nucleotide sequence corresponding to the 3'end of the sequence; and (x) cD from nucleotide 114 to nucleotide 824 of SEQ ID NO: 9.
The nucleotide sequence corresponding to the NA sequence and corresponding to the 5'end of the sequence from nucleotide 114 to nucleotide 824 of SEQ ID NO: 9 to 3 of the sequence from nucleotide 114 to nucleotide 824 of SEQ ID NO: 9. 'A method comprising a nucleotide sequence selected from the group consisting of nucleotide sequences that are contiguous up to the nucleotide sequence corresponding to the end. 27. (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 11; (b) a polynucleotide comprising the nucleotide sequence of nucleotides 297 to 542 of SEQ ID NO: 11; (c) a nucleotide sequence of SEQ ID NO: 11. A polynucleotide comprising the nucleotide sequence of nucleotides 510 to 542; (d) clone nn229_1 deposited under accession number ATCC 98623.
A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence of: (e) clone nn229_1 deposited under accession number ATCC 98623
A polynucleotide encoding the full-length protein encoded by the cDNA insert of: (f) clone nn229_1 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of: (g) clone nn229_1 deposited under accession number ATCC 98623.
(H) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 12; (i) a biologically active SEQ ID NO: A polynucleotide encoding a protein comprising a fragment of the amino acid sequence of 12 (the fragment being SEQ ID NO: 12).
8 consecutive amino acids); (j) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; and (k) (a)-(i) under stringent conditions. An isolated polynucleotide selected from the group consisting of polynucleotides capable of hybridizing to any one of the polynucleotides shown in. 28. (a) the amino acid sequence of SEQ ID NO: 12; (b) a fragment of the amino acid sequence of SEQ ID NO: 12 (each fragment containing 8 consecutive amino acids of SEQ ID NO: 12); and (c ) Clone nn229 1 deposited under accession number ATCC 98623
A protein containing an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts, which is substantially free of other mammalian proteins. 29. A method for producing an isolated polynucleotide, the method comprising: (a) a method comprising the following steps: (i) selected from the group consisting of the following at 65 ° C. in 6X SSC: Preparing one or more polynucleotide probes that hybridize to the nucleotide sequence; (aa) SEQ ID NO: 11 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 11); and (ab) ATCC 98623. Sequence of the cDNA insert of clone nn229_1 deposited as (ii) hybridizing said probe (s) to human DNA; and (iii) a DNA polynucleotide detected with said probe (s). And (b) a method comprising the following steps: (i) in 6X SSC Preparing one or more polynucleotide primers that hybridize at 65 ° C. to a nucleotide sequence selected from the group consisting of: (ba) SEQ ID NO: 11 (at the 3 ′ end of SEQ ID NO: 11) (Bb) except the poly (A) tail); and (bb) the nucleotide sequence of the cDNA insert of clone nn229_1 deposited as ATCC 98623. (ii) hybridizing the primer (s) to human DNA; (iii) human Selected from the group consisting of amplifying a DNA sequence; and then isolating the polynucleotide product of (iv) step (b) (iii), wherein the isolated polynucleotide is: (v) SEQ ID NO: 11 corresponds to the cDNA sequence of SEQ ID NO: 11,
From the nucleotide sequence corresponding to the 5'end of SEQ ID NO: 11 to the nucleotide sequence corresponding to the 3'end of SEQ ID NO: 11, but excluding the poly (A) tail at the 3'end of SEQ ID NO: 11 (W) the nucleotide sequence corresponding to nucleotides 297 to 542 of the cDNA sequence of SEQ ID NO: 11, and corresponding to the 5'end of the sequence from nucleotide 297 to nucleotide 542 of SEQ ID NO: 11. From the nucleotide sequence, nucleotides 297 to 542 of SEQ ID NO: 11 3 ′ of the sequence.
A nucleotide sequence that is continuous up to the nucleotide sequence corresponding to the end;
And (x) c from nucleotide 510 to nucleotide 542 of SEQ ID NO: 11
From the nucleotide sequence corresponding to the 5'end of the sequence from nucleotide 510 to nucleotide 542 of SEQ ID NO: 11 to the nucleotide sequence of nucleotides 510 to 542 of SEQ ID NO: 11 '
A method comprising a nucleotide sequence selected from the group consisting of nucleotide sequences that are contiguous up to the nucleotide sequence corresponding to the terminus. 30. (a) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 13; (b) a polynucleotide comprising the nucleotide sequence of nucleotides 547 to 750 of SEQ ID NO: 13; (c) the nucleotide sequence of SEQ ID NO: 13 A polynucleotide comprising the nucleotide sequence of nucleotides 601 to 750; (d) clone np156_1 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the full length protein coding sequence of: (e) clone np156_1 deposited under accession number ATCC 98623.
A polynucleotide encoding the full-length protein encoded by the cDNA insert of: (f) clone np156_1 deposited under accession number ATCC 98623
A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence of: (g) clone np156_1 deposited under accession number ATCC 98623.
(H) a polynucleotide encoding a protein containing the amino acid sequence of SEQ ID NO: 14; (i) a biologically active SEQ ID NO: A polynucleotide encoding a protein comprising a fragment of the amino acid sequence of 14 (the fragment being SEQ ID NO: 14
8 consecutive amino acids); (j) a polynucleotide which is an allelic variant of a polynucleotide of (a)-(g) above; and (k) (a)-(i) under stringent conditions. An isolated polynucleotide selected from the group consisting of polynucleotides capable of hybridizing to any one of the polynucleotides shown in. 31. (a) the amino acid sequence of SEQ ID NO: 14; (b) a fragment of the amino acid sequence of SEQ ID NO: 14 comprising 8 consecutive amino acids of SEQ ID NO: 14; and (c) accession number ATCC 98623. Clone np156 1 deposited as
A protein containing an amino acid sequence selected from the group consisting of the amino acid sequences encoded by the cDNA inserts, which is substantially free of other mammalian proteins. 32. A method for producing an isolated polynucleotide, which method comprises: (a) a method comprising the following steps: (i) selected from the group consisting of the following at 65 ° C. in 6X SSC: Preparing one or more polynucleotide probes that hybridize to the nucleotide sequence; (aa) SEQ ID NO: 13 (excluding the poly (A) tail at the 3'end of SEQ ID NO: 13); and (ab) ATCC 98623. Sequence of the cDNA insert of clone np156_1 deposited as (ii) hybridizing said probe (s) to human DNA; and (iii) a DNA polynucleotide detected by said probe (s). And (b) a method comprising the following steps: (i) in 6X SSC Preparing one or more polynucleotide primers that hybridize at 65 ° C. to a nucleotide sequence selected from the group consisting of: (ba) SEQ ID NO: 13 (at the 3 ′ end of SEQ ID NO: 13) (Bb) except the poly (A) tail); and (bb) the nucleotide sequence of the cDNA insert of clone np156_1 deposited as ATCC 98623. (ii) hybridizing the primer (s) to human DNA; (iii) human Selected from the group consisting of amplifying a DNA sequence; and then isolating the polynucleotide product of (iv) step (b) (iii), wherein the isolated polynucleotide is: (v) SEQ ID NO: 13 corresponds to the cDNA sequence of SEQ ID NO: 13
From the nucleotide sequence corresponding to the 5'end of SEQ ID NO: 13 to the nucleotide sequence corresponding to the 3'end of SEQ ID NO: 13, but excluding the poly (A) tail at the 3'end of SEQ ID NO: 13 (W) the nucleotide sequence corresponding to nucleotides 547 to 750 of the cDNA sequence of SEQ ID NO: 13 and corresponding to the 5'end of the sequence of nucleotides 547 to 750 of SEQ ID NO: 13 From the nucleotide sequence, nucleotides 547 to 750 of SEQ ID NO: 13 3 ′ of the sequence
A nucleotide sequence that is continuous up to the nucleotide sequence corresponding to the end;
And (x) from the nucleotide sequence corresponding to nucleotides 601 to 750 of the cDNA sequence of SEQ ID NO: 13, corresponding to the 5'end of said sequence of nucleotides 601 to 750 of SEQ ID NO: 13, 3'of the sequence from nucleotide 601 to nucleotide 750 of SEQ ID NO: 13
A method comprising a nucleotide sequence selected from the group consisting of nucleotide sequences that are contiguous up to the nucleotide sequence corresponding to the terminus.