- a) surgically debulking the mammary carcinoma/tumor; and
- b) administering to said dog a clinically-effective amount of a first nucleic acid molecule comprising a DNA sequence encoding a xenogeneic mammary gland-associated differentiation antigen under the control of a promoter which promotes expression of said xenogeneic mammary gland-associated differentiation antigen in said dog; and wherein canines receiving said nucleic acid molecule exhibit significantly increased overall, disease-free, and/or metastasis-free survival times relative to canines receiving the surgical debulking without subsequent administration of said nucleic acid molecule, thereby treating said carcinoma/tumor.
Her2 DNA vaccine as adjunct treatment for cancers in companion animals
Updated Time 12 June 2019Patent Registration Data
29 April 2015
27 February 2018
Original Assignee (Applicant)
FISCHER, LAURENT BERNARD
This patent contains figures and images illustrating the invention and its embodiment.
The application discloses therapeutic vaccines based upon the “pING” DNA plasmid vector expressing the gene encoding the rat Her2 protein. Vaccines according to the instant disclosure are used as an adjunct treatment for surgery, radiation and/or chemotherapy for dogs and cats with cancers that over express the Her2 antigen, and prolong the post-surgical disease free interval and/or survival time. Also included are therapeutically effective methods of immunization using said vaccines.
1. A method for treating canine mammary carcinoma/tumor in a dog suffering from a canine mammary carcinoma/tumor comprising cells expressing a xenogeneic mammary gland-associated differentiation antigen, comprising: a) surgically debulking the mammary carcinoma/tumor; and b) administering to said dog a clinically-effective amount of a first nucleic acid molecule comprising a DNA sequence encoding a xenogeneic mammary gland-associated differentiation antigen under the control of a promoter which promotes expression of said xenogeneic mammary gland-associated differentiation antigen in said dog; and wherein canines receiving said nucleic acid molecule exhibit significantly increased overall, disease-free, and/or metastasis-free survival times relative to canines receiving the surgical debulking without subsequent administration of said nucleic acid molecule, thereby treating said carcinoma/tumor.
2. The method of claim 1, wherein said nucleic acid molecule is a plasmid.
3. The method of claim 2, wherein said differentiation antigen is a canine Her2/neu antigen, and wherein said xenogeneic antigen is a non-canine Her2/neu antigen.
4. The method of claim 3, wherein said xenogeneic Her2/neu antigen is a rat Her2/neu antigen.
5. The method of claim 4, wherein said plasmid comprises nucleotides 106-3882 of the sequence as set forth in SEQ ID NO: 1, or wherein said plasmid comprises nucleotides encoding the same peptide sequence as nucleotides 106-3882 of the sequence as set forth in SEQ ID NO: 1.
6. The method of claim 1, comprising the steps of: 1) surgically debulking a canine mammary tumor/carcinoma expressing a Her2/neu differentiation antigen; 2) administering a first nucleic acid molecule encoding a xenogeneic Her2/neu; and 3) administering via electrotransfer/electroporation a second nucleic acid molecule; wherein the second nucleic acid molecule is either identical to said first nucleic acid molecule, or is a second, distinct nucleic acid molecule capable of expressing in vivo in a canine a different xenogeneic Her2/neu antigen, including those encoded by SEQ ID NOs:1, 3 or 4, or is a recombinant vector capable of expressing in vivo in said canine any Her2/neu protein, which is capable of eliciting a therapeutically effective immune response against heterologous Her2/neu expressed by said canine mammary tumor/carcinoma.
7. The method of claim 6, wherein the first nucleic acid molecule is a plasmid.
8. The method of claim 6, wherein: 1) the first administration consists of administering to said canine said first nucleic acid molecule without a needle; 2) the first nucleic acid molecule is capable of expressing in vivo in a canine a sequence comprising or consisting of the same sequence as set forth in SEQ ID NO:2; and 3) the second administration comprises or consists of administering said nucleic acid molecule of step 2.
9. The method of claim 8, wherein said first nucleic acid molecule is a plasmid.
10. The method of claim 6, wherein said second administration is provided to surviving canines once every about 3 to about 6 months.
11. The method of claim 8, wherein said second administration is provided to surviving canines once every about 3 to about 6 months.
12. The method of claim 1, which is performed about concurrently with resection of a mammary gland tumor (MGT).
13. The method of claim 4, which is performed about concurrently with resection of a mammary gland tumor (MGT).
14. The method of claim 1, wherein the administration is performed using a needle-free delivery device.
15. The method of claim 1, further comprising at least a second administration, provided at least once or at regular intervals after the initial antigen administration.
16. The method of claim 1, wherein: a) the mean overall survival time is at least about 100 days greater; b) the average disease-free survival time is at least about 200 days greater, and/or c) the metastasis-free survival time is at least about 200 days greater in said canines receiving said nucleic acid molecule, relative to said canines receiving the surgical debulking without subsequent administration of said nucleic acid molecule.
17. The method of claim 16, wherein the three survival times are at least about 100, about 200, and about 200 days greater in the immunized canines relative to said canines receiving the surgical debulking without subsequent administration of said nucleic acid molecule, respectively.
18. The method of claim 9, wherein: a) the mean overall survival time is at least about 100 days greater; b) the average disease-free survival time is at least about 200 days greater, and/or c) the metastasis-free survival time is at least about 200 days greater in said canines receiving said nucleic acid molecule, relative to said canines receiving the surgical debulking without subsequent administration of said first nucleic acid molecule.
19. The method of claim 18, wherein the three survival times are at least about 100, about 200, and about 200 days greater in the immunized canines relative to said canines receiving the surgical debulking without subsequent administration of said first nucleic acid molecule, respectively.
11. A method for treating canine mammary carcinoma/tumor in a dog suffering from a canine mammary carcinoma/tumor comprising cells expressing a xenogeneic mammary gland-associated differentiation antigen, comprising:
2. The method of claim 1, wherein
- said nucleic acid molecule is a plasmid.
6. The method of claim 1, comprising the steps of:
- 1) surgically debulking a canine mammary tumor/carcinoma expressing a Her2/neu differentiation antigen;
- 2) administering a first nucleic acid molecule encoding a xenogeneic Her2/neu; and
- 3) administering via electrotransfer/electroporation a second nucleic acid molecule; wherein the second nucleic acid molecule is either identical to said first nucleic acid molecule, or is a second, distinct nucleic acid molecule capable of expressing in vivo in a canine a different xenogeneic Her2/neu antigen, including those encoded by SEQ ID NOs:1, 3 or 4, or is a recombinant vector capable of expressing in vivo in said canine any Her2/neu protein, which is capable of eliciting a therapeutically effective immune response against heterologous Her2/neu expressed by said canine mammary tumor/carcinoma.
12. The method of claim 1, which is performed about concurrently with resection of a mammary gland tumor (MGT).
14. The method of claim 1, wherein
- the administration is performed using a needle-free delivery device.
15. The method of claim 1, further comprising
- at least a second administration, provided at least once or at regular intervals after the initial antigen administration.
16. The method of claim 1, wherein
- : a) the mean overall survival time is at least about 100 days greater; b) the average disease-free survival time is at least about 200 days greater, and/or c) the metastasis-free survival time is at least about 200 days greater in said canines receiving said nucleic acid molecule, relative to said canines receiving the surgical debulking without subsequent administration of said nucleic acid molecule.
BACKGROUND OF THE INVENTION
This application relates to compositions for treatment of differentiation antigen-dependent cancers and to methods of using such compositions. The invention utilizes compositions containing xenogeneic differentiation antigens, which are associated with cancers to provide effective therapy.
Differentiation antigens are tissue-specific antigens that are shared by autologous and some allogeneic tumors of similar derivation, and on normal tissue counterparts at the same stage of differentiation. Differentiation antigens have been shown to be expressed by a variety of tumor types, including melanoma, leukemia, lymphomas, colorectal carcinoma, breast carcinoma, prostate carcinoma, ovarian carcinoma, pancreas carcinomas, and lung cancers. For example, differentiation antigens expressed by melanoma cells include Melan-A/MART-1, Pmel17, tyrosinase, and gp75. Differentiation antigen expressed by lymphomas and leukemia include CD19 and CD20/CD20 B lymphocyte differentiation markers). An example of a differentiation antigen expressed by colorectal carcinoma, breast carcinoma, pancreas carcinoma, prostate carcinoma, ovarian carcinoma, and lung carcinoma is the mucin polypeptide muc-1. A differentiation antigen expressed by, for example, breast carcinoma is Her2 (synonyms: Her2/neu, ECBB2, ErbB2, c-erb-2), which is a gene coding for a tyrosine kinase receptor that is a member of the family of epidermal growth factor receptors (De Maria et al., 2005). Over expression of Her2 has been demonstrated in mammary gland tumors of both the cat (Winston et al., 2005) and the dog (Rungsipipat et al., 2008). Winston et al. (2005) used existing assay methods (HERCEPTEST™, Dako USA, Carpinteria, Calif.; NCL-CB11, Novocastra, Newcastle, UK) to successfully grade levels of Her2 expression on feline mammary tumors as 0=minimal/absent, 1=weak, 2=moderate, or 3=intense. The HERCEPTEST™ and NCL-CB11 assays identified 27 and 23 cats respectively, out of 30 examined, as having grade 2 or 3 Her2 expression in mammary tumor samples.
In addition to successfully grading levels of Her2 over expression in feline mammary tumors, Winston et al. (2005) used the HERCEPTEST™ to detect low levels of Her2 expression in normal feline epithelial tissues and cell types including: hair follicle, mammary gland, gastric pit, salivary gland duct, renal cortical and medullary tubules, colonic and small intestinal crypt, brain, pancreatic duct and islets, splenic macrophages, adrenal cortex, hepatocytes, and testicular Leydig's cells. Expression of Her2 has been documented on a range of human epithelial cell types including gastro-intestinal, respiratory, reproductive, urinary, skin, mammary and placenta (Press et al., 1990). These findings indicate that the expression of Her2 is common in a range of tissue types of humans and cats. The finding of Her2 over expression in dog mammary tumors suggests this species would share expression characteristics identified in humans and cats. Existing assays and reagents can serve as tools to screen expression levels of Her2 in companion animal cancers in order to justify treatment with the Her2 cancer vaccine.
Unfortunately, in most cases, the immune system of the individual is tolerant of such differentiation antigens, and fails to mount an effective immune response. Several technologies have been considered to address this challenge: (cytokines as genetic adjuvants (Chang et al., 2004), xenogeneic vaccination (Pupa et al., 2005), electrotransfer (Quaglino et al., 2004), combination with chemotherapy (Bernhardt et al., 2002). Although results were encouraging, greater efficacy was required for these approaches to be considered a key component of a first-line therapeutic strategy. Further, recent findings indicate both antibody and cell-mediated immunity are required for tumor eradication post immunization, perhaps explaining, in part, the lack of success in the field (Orlandi et al., 2007). Therefore, for the treatment of cancers where the tumor expresses differentiation antigens therefore, it would be desirable to have a method for stimulating a therapeutically effective immune response against the differentiation antigen in vivo. It an object of the present invention to provide such a method.
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- Chang S Y et al. Enhanced efficacy of DNA vaccination against Her2/neu tumor antigen by genetic adjuvants; IJC vol 111 pages 86-95, 10 Aug. 2004
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All of the above-mentioned applications, patents and references are herein incorporated in their entirety by reference.
SUMMARY OF THE INVENTION
It has now been found that the tolerance of the immune system for self-derived target differentiation antigens can be overcome and an immune response stimulated by administration of a xenogeneic differentiation antigen (wild-type or mutant) of the same type from a species different from the subject being treated (U.S. Pat. No. 6,328,969 & U.S. Pat. No. 7,556,805, to Sloan-Kettering, both incorporated by reference herein). For example, a rat differentiation antigen can be used to stimulate an immune response to the corresponding differentiation antigen in a canine subject. Administration of altered antigens in accordance with the invention results in an effective immunity against the original antigen expressed by the cancer in the treated subject. Thus, in accordance with a first aspect of the invention, there is provided a method for treating in a mammalian subject, comprising the step of administering to the subject an immunologically-effective amount of a xenogeneic mammary gland tumor-associated differentiation antigen.
Therapeutic differentiation antigens based on mammary gland carcinoma/tumor-associated differentiation antigens are used in accordance with the invention to treat, for example, mammary gland carcinoma post-surgical removal of tumors in subjects suffering from said cancers. In one embodiment of the invention, a plasmid comprising a sequence encoding a xenogeneic tyrosine kinase receptor, for example rat tyrosine kinase receptor, under the control of a suitable promoter, is administered to a subject. For example, dogs have been treated using plasmids comprising a DNA sequence encoding rat tyrosine kinase receptor with pronounced clinical benefit.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1A shows overall survival time post-immunization and surgical resection of MGT;
FIG. 1B shows disease-free survival time post-immunization and surgical resection of MGT;
FIG. 1C shows metastasis-free survival time post-immunization and surgical resection of MGT;
FIG. 2 shows a map of the pcDNA3.1 (+/−) plasmid
FIG. 3 shows a sequence for the pINGhumanTyrosinase plasmid, where the coding sequence for the human tyrosinase has been removed. This is where the rat Her2/neu (nucleotides 17-3799 of SEQ ID NO:1) was inserted to produce rHer2/neu-pING of the instant invention;
FIG. 4 shows a map of the pING plasmid.
DETAILED DESCRIPTION OF THE INVENTION
The present invention provides a method for treating mammary gland tumors in a subject by stimulating an immune response to a mammary gland-associated differentiation antigen. The subject is preferably canine or feline, although the invention can be applied to other animal species, preferably mammalian or avian species, as well.
As used in the specification and claims of this application, the term “immune response” encompasses both cellular and humoral immune responses. Preferably, the immune response is sufficient to provide immunoprotection against growth of tumors expressing the target differentiation antigen. The term “stimulate” refers to the initial stimulation of a new immune response or to the enhancement of a pre-existing immune response.
In accordance with the invention, a subject is treated by administering a xenogeneic differentiation antigen of the same type as a target differentiation antigen expressed by mammary gland tumor cells of the subject in an amount effective to stimulate an immune response. Thus, for example, if the target differentiation antigen is the Her2/neu antigen found in mammary cells, the therapeutic antigen is a xenogeneic Her2/neu antigen.
In one embodiment, the inventive method may include the following steps: (1) immunization to an animal in need of a xenogeneic antigen, for example, the rat Her2/neu as set forth in SEQ ID NO:2 and encoded by nucleotides 106-3885 of the sequence as set forth in SEQ ID NO:1, (2) needle-free priming of immune responses, (3) electrotransfer-based booster, and (4) vaccination after tumor debulking by surgical primary therapy.
In another embodiment, the inventive method is carried out on subjects, including companion animals, without metastasis (i.e. in relatively early stages of mammary carcinoma disease progression).
In some embodiments, the boost comprises administering plasmids encoding xenogeneic antigens, for example those encoding rat Her2 protein (SEQ ID NO:2).
In some embodiments, the xenogeneic antigen is encoded by a nucleotide having favorable nucleotide substitutions with respect to the sequence as set forth in SEQ ID NO:1. Favorable substitutions include any changes that result in improved immune response against the Her2/neu expressed by the cells of the mammary tumor/carcinoma. Substitutions can include existing sequences, such as murine Her2 (SEQ ID NO:3), human Her2 (SEQ ID NO:4), or any other xenogeneic Her2 sequence, or fragment thereof, capable of eliciting a therapeutically effective immune response in a target animal against a Her2-associated mammary carcinoma.
In some embodiments, the boost comprises administering a xenogeneic differentiation antigen.
In other embodiments, the boost comprises administering a syngeneic differentiation antigen.
Xenogeneic differentiation antigen may be administered as a purified differentiation antigen derived from the source organism. Proteins can be purified for this purpose from cell lysates using column chromatography procedures. Proteins for this purpose may also be purified from recombinant sources, such as bacterial or yeast clones or mammalian or insect cell lines expressing the desired product.
Administration of the xenogeneic differentiation antigen can be accomplished by several routes. First, the xenogeneic differentiation antigen may be administered as part of a vaccine composition which may include one or more adjuvants such as alum, QS21, TITERMAX or its derivatives, incomplete or complete Freund's and related adjuvants, and cytokines such as granulocyte-macrophage colony stimulating factor, flt-3 ligand, interleukin-2, interleukin-4 and interleukin-12 for increasing the intensity of the immune response. The vaccine composition may be in the form of a xenogeneic differentiation antigen in a solution or a suspension, or the therapeutic differentiation antigen may be introduced in a lipid carrier such as a liposome. Such compositions will generally be administered by subcutaneous, intradermal or intramuscular route. Vaccine compositions containing expressed xenogeneic differentiation antigen are administered in amounts which are effective to stimulate an immune response to the target differentiation antigen in the subject. The preferred amount to be administered will depend on the species of the subject and on the specific antigen, but can be determined through routine preliminary tests in which increasing doses are given and the extent of antibody formation or T cell response is measured by ELISA or similar tests. T cell responses may also be measured by cellular immune assays, such as cytotoxicity, cytokine release assays and proliferation assays.
The xenogeneic differentiation antigen may also be introduced in accordance with the invention using a DNA immunization technique in which DNA encoding the antigen is introduced into the subject such that the xenogeneic differentiation antigen is expressed by the subject. cDNA encoding the differentiation antigen is combined with a promoter which is effective for expression of the nucleic acid polymer in mammalian cells. This can be accomplished by digesting the nucleic acid polymer with a restriction endonuclease and cloning into a plasmid containing a promoter such as the SV40 promoter, the cytomegalovirus (CMV) promoter or the Rous sarcoma virus (RSV) promoter. The resulting construct is then used as a vaccine for genetic immunization. The nucleic acid polymer could also be cloned into plasmid and viral vectors that are known to transduce mammalian cells. These vectors include retroviral vectors, adenovirus vectors, vaccinia virus vectors, pox virus vectors and adenovirus-associated vectors.
The nucleic acid constructs containing the promoter and the antigen-coding region can be administered directly or they can be packaged in liposomes or coated onto colloidal gold particles prior to administration. Techniques for packaging DNA vaccines into liposomes are known in the art, for example from Murray, ed. “Gene Transfer and Expression Protocols” Humana Pres, Clifton, N.J. (1991). Similarly, techniques for coating naked DNA onto gold particles are taught in Yang, “Gene transfer into mammalian somatic cells in vivo”, Crit. Rev. Biotech. 12: 335-356 (1992), and techniques for expression of proteins using viral vectors are found in Adolph, K. ed. “Viral Genome Methods” CRC Press, Florida (1996).
For genetic immunization, the vaccine compositions are preferably administered intradermally, subcutaneously or intramuscularly by injection or by gas driven particle bombardment, and are delivered in an amount effective to stimulate an immune response in the host organism. The compositions may also be administered ex vivo to blood or bone marrow-derived cells (which include APCs) using liposomal transfection, particle bombardment or viral infection (including co-cultivation techniques). The treated cells are then reintroduced back into the subject to be immunized. While it will be understood that the amount of material needed will depend on the immunogenicity of each individual construct and cannot be predicted a priori, the process of determining the appropriate dosage for any given construct is straightforward. Specifically, a series of dosages of increasing size, starting at about 0.1 μg is administered and the resulting immune response is observed, for example by measuring antibody titer using an ELISA assay, detecting CTL response using a chromium release assay or detecting TH (helper T cell) response using a cytokine release assay.
Once tolerance is broken through the administration of the xenogeneic differentiation antigen, subsequent treatments with syngeneic differentiation may be employed to maintain and in some cases enhance the immune response. (See, Weber, et al., “Tumor immunity and autoimmunity induced by immunization with homologous DNA.” J Clin Invest 102 (6):1258 (1998).) Thus, in one embodiment of the invention, the subject is first treated by administration of a xenogeneic differentiation antigen (for example for three treatment cycles), and subsequently by administration of a syngeneic differentiation antigen (for example for an additional three treatment cycles). As an alternative to treatment cycles using different therapeutic agents, one can use a single therapeutic agent containing both xenogeneic and syngeneic differentiation antigens. Thus, for example, a mixture of the rHer2-pING and hHer2-pING vectors, or a single vector encoding both rat and human Her2/neu under the control of a promoter such that they are expressed in a canine subject can be employed for the treatment of mammary gland tumor in canines. Vectors are available commercially, for example from Stratagene and other companies, which can express two independent genes. Commonly, these vectors use an internal ribosomal entry site, or IRES, between the two genes. This approach has the advantage of requiring approval for only a single therapeutic agent.
All documents cited herein are herein incorporated by reference in their entirety.
The invention will now be further described with reference to the following, non-limiting examples.
Example 1—Her2/neu Expression Plasmid Construction
The extracellular domain of rat HER2/neu (nucleotides 17-3799 of SEQ ID NO:1) was amplified by PCR from the pCMVneuNT (Amici et al., 1998) plasmid using the primers forward: 5′-CGAAGCTTACCATGGAGCTGGCGGCCTGG-3′ (SEQ ID NO:6) and reverse: 5′-CGGAATTCTTATGTCACCGGGCTGGC-3′ (SEQ ID NO:7). The HindIII-EcoRI fragment was cloned into pcDNA3.1(+) (Invitrogen, Carlsbad, Calif.; and FIG. 2). The original sequence of the rat neu cDNA was described previously (Bargmann et al., 1986), and is herein set forth in SEQ ID NO:1, with the coding sequence from nucleotides 17 to 3799. The rat HER2/neu coding sequence was then subcloned into the pING vector (Bergman et al., Clin Cancer Res, 9: 1284-1290, 2003, backbone depicted in FIG. 3; map depicted in FIG. 3A; and sequence as set forth in SEQ ID NO:5), to yield rat HER2/neu-pING.
Example 2—Immunization of Mammary Gland Tumor (MGT)-Positive Canines with pING-rHer2
In this trial, 10 dogs with MGT were enrolled and immunized with 100 μg of pING-rHer2 DNA per dose. The signalment for these dogs is set forth in Table 1 and the tumor staging is set forth in Table 2.
As indicated, this group included five stage I and five stage III dogs, which all received three doses of vaccine at two week intervals. The first and second doses were administered with the VITAJET™ transdermal device and the third dose by intramuscular injection concurrent with electroporation. Vaccination was initiated following surgical removal of the MGT with concurrent ovariohysterectomy (OHE). All dogs were negative for regional lymph node and pulmonary metastasis. Disease free survival and overall survival times were calculated using day of surgery as day 0 with results presented in Table 3.
A group of 19 dogs was identified as historical control cases. All control dogs underwent surgical removal of MGT with concurrent OHE and were negative for regional lymph node and pulmonary metastasis. This group included 7 stage I, 3 stage II, and 9 stage III dogs. Disease free and overall survival times were calculated for these dogs using day of surgery as day 0. The signalment for these dogs is set forth in Table 4 and tumor staging for each dog is set forth in Table 5. Disease free and overall survival times were calculated for the control group and are presented in FIGS. 1A-1C.
Philibert et al. (2003) reviewed survival statistics for 97 dogs with MGT and reported median survival times for 41 dogs with MGT less than 3 cm in diameter to be 22 months (˜666 days) versus 14 months (˜424 days) for 56 dogs with MGT greater than 3 cm in diameter. In the absence of lymph node involvement or metastasis, tumor size less than 3 cm correlates with stage I disease and greater than 3 cm correlates with stage II or higher disease status. They did not find a difference in survival time for dogs in stages II, III or IV.
Overall median survival time for all dogs treated with the pING-rHer2 vaccine is 678 days. This was significantly higher as compared to the historical data from the 19 dogs provided by NTU indicating a median overall survival time of 300 days, and to the data published by Philibert et al. (2003) indicating 424 days overall survival time for dogs with stage II or greater MGT.
The pING-rHer2 DNA vaccine will target dogs and cats with tumors shown to over express the Her2 antigen based upon tumor tissue analysis using existing Her2 tissue expression assays. The vaccine will be administered using the Vetjet™ transdermal device to deliver 100 μg of DNA into the medial thigh of dogs or lateral thigh of cats, at two week intervals for four doses. Dogs and cats that survive will receive a booster dose every six months.
The invention will now be described by the following non-limiting claims.
<160> NUMBER OF SEQ ID NOS: 4
<210> SEQ ID NO: 1
<211> LENGTH: 4727
<212> TYPE: DNA
<213> ORGANISM: Rattus norvegicus
<400> SEQENCE: 1
gcgccccttc ccaggcggcc ccttccggcg ccgcgcctgt gcctgccctc gccgcgcccc 60
gcgcccgcag cctggtccag cctgagccat ggggccggag ccgcaatgat catcatggag 120
ctggcggcct ggtgccgctg ggggttcctc ctcgccctcc tgccccccgg aatcgcgggc 180
acccaagtgt gtaccggcac agacatgaag ttgcggctcc ctgccagtcc tgagacccac 240
ctggacatgc tccgccacct gtaccagggc tgtcaggtag tgcagggcaa cttggagctt 300
acctacgtgc ctgccaatgc cagcctctca ttcctgcagg acatccagga agttcagggt 360
tacatgctca tcgctcacaa ccaggtgaag cgcgtcccac tgcaaaggct gcgcatcgtg 420
agagggaccc agctctttga ggacaagtat gccctggctg tgctagacaa ccgagatcct 480
caggacaatg tcgccgcctc caccccaggc agaaccccag aggggctgcg ggagctgcag 540
cttcgaagtc tcacagagat cctgaaggga ggagttttga tccgtgggaa ccctcagctc 600
tgctaccagg acatggtttt gtggaaggac gtcttccgca agaataacca actggctcct 660
gtcgatatag acaccaatcg ttcccgggcc tgtccacctt gtgcccccgc ctgcaaagac 720
aatcactgtt ggggtgagag tccggaagac tgtcagatct tgactggcac catctgtacc 780
agtggttgtg cccggtgcaa gggccggctg cccactgact gctgccatga gcagtgtgcc 840
gcaggctgca cgggccccaa gcattctgac tgcctggcct gcctccactt caatcatagt 900
ggtatctgtg agctgcactg cccagccctc gtcacctaca acacagacac ctttgagtcc 960
atgcacaacc ctgagggtcg ctacaccttt ggtgccagct gcgtgaccac ctgcccctac 1020
aactacctgt ctacggaagt gggatcctgc actctggtgt gtcccccgaa taaccaagag 1080
gtcacagctg aggacggaac acagcgttgt gagaaatgca gcaagccctg tgctcgagtg 1140
tgctatggtc tgggcatgga gcaccttcga ggggcgaggg ccatcaccag tgacaatgtc 1200
caggagtttg atggctgcaa gaagatcttt gggagcctgg catttttgcc ggagagcttt 1260
gatggggacc cctcctccgg cattgctccg ctgaggcctg agcagctcca agtgttcgaa 1320
accctggagg agatcacagg ttacctgtac atctcagcat ggccagacag tctccgtgac 1380
ctcagtgtct tccagaacct tcgaatcatt cggggacgga ttctccacga tggcgcgtac 1440
tcattgacac tgcaaggcct ggggatccac tcgctggggc tgcgctcact gcgggagctg 1500
ggcagtggat tggctctgat tcaccgcaac gcccatctct gctttgtaca cactgtacct 1560
tgggaccagc tcttccggaa cccacatcag gccctgctcc acagtgggaa ccggccggaa 1620
gaggattgtg gtctcgaggg cttggtctgt aactcactgt gtgcccacgg gcactgctgg 1680
gggccagggc ccacccagtg tgtcaactgc agtcatttcc ttcggggcca ggagtgtgtg 1740
gaggagtgcc gagtatggaa ggggctcccc cgggagtatg tgagtgacaa gcgctgtctg 1800
ccgtgtcacc ccgagtgtca gcctcaaaac agctcagaga cctgctttgg atcggaggct 1860
gatcagtgtg cagcctgcgc ccactacaag gactcgtcct cctgtgtggc tcgctgcccc 1920
agtggtgtga aaccggacct ctcctacatg cccatctgga agtacccgga tgaggagggc 1980
atatgccagc cgtgccccat caactgcacc cactcctgtg tggatctgga tgaacgaggc 2040
tgcccagcag agcagagagc cagcccggtg acattcatca ttgcaactgt agtgggcgtc 2100
ctgctgttcc tgatcttagt ggtggtcgtt ggaatcctaa tcaaacgaag gagacagaag 2160
atccggaagt atacgatgcg taggctgctg caggaaactg agttagtgga gccgctgacg 2220
cccagcggag caatgcccaa ccaggctcag atgcggatcc taaaagagac ggagctaagg 2280
aaggtgaagg tgcttggatc aggagctttt ggcactgtct acaagggcat ctggatccca 2340
gatggggaga atgtgaaaat ccccgtggct atcaaggtgt tgagagaaaa cacatctcct 2400
aaagccaaca aagaaattct agatgaagcg tatgtgatgg ctggtgtggg ttctccgtat 2460
gtgtcccgcc tcctgggcat ctgcctgaca tccacagtac agctggtgac acagcttatg 2520
ccctacggct gccttctgga ccatgtccga gaacaccgag gtcgcctagg ctcccaggac 2580
ctgctcaact ggtgtgttca gattgccaag gggatgagct acctggagga cgtgcggctt 2640
gtacacaggg acctggctgc ccggaatgtg ctagtcaaga gtcccaacca cgtcaagatt 2700
acagatttcg ggctggctcg gctgctggac attgatgaga cagagtacca tgcagatggg 2760
ggcaaggtgc ccatcaaatg gatggcattg gaatctattc tcagacgccg gttcacccat 2820
cagagtgatg tgtggagcta tggagtgact gtgtgggagc tgatgacttt tggggccaaa 2880
ccttacgatg gaatcccagc ccgggagatc cctgatttgc tggagaaggg agaacgccta 2940
cctcagcctc caatctgcac cattgatgtc tacatgatta tggtcaaatg ttggatgatt 3000
gactctgaat gtcgcccgag attccgggag ttggtgtcag aattttcacg tatggcgagg 3060
gacccccagc gttttgtggt catccagaac gaggacttgg gcccatccag ccccatggac 3120
agtaccttct accgttcact gctggaagat gatgacatgg gtgacctggt agacgctgaa 3180
gagtatctgg tgccccagca gggattcttc tccccggacc ctaccccagg cactgggagc 3240
acagcccata gaaggcaccg cagctcgtcc accaggagtg gaggtggtga gctgacactg 3300
ggcctggagc cctcggaaga agggcccccc agatctccac tggctccctc ggaaggggct 3360
ggctccgatg tgtttgatgg tgacctggca atgggggtaa ccaaagggct gcagagcctc 3420
tctccacatg acctcagccc tctacagcgg tacagcgagg accccacatt acctctgccc 3480
cccgagactg atggctatgt tgctcccctg gcctgcagcc cccagcccga gtatgtgaac 3540
caatcagagg ttcagcctca gcctccttta accccagagg gtcctctgcc tcctgtccgg 3600
cctgctggtg ctactctaga aagacccaag actctctctc ctgggaagaa tggggttgtc 3660
aaagacgttt ttgccttcgg gggtgctgtg gagaaccctg aatacttagt accgagagaa 3720
ggcactgcct ctccgcccca cccttctcct gccttcagcc cagcctttga caacctctat 3780
tactgggacc agaactcatc ggagcagggg cctccaccaa gtaactttga agggaccccc 3840
actgcagaga accctgagta cctaggcctg gatgtacctg tatgagacgt gtgcagacgt 3900
cctgtgcttt cagagtgggg aaggcctgac ttgtggtctc catcgccaca aagcagggag 3960
agggtcctct ggccacatta catccagggc agacggctct accaggaacc tgccccgagg 4020
aacctttcct tgctgcttga atcctgagtg gttaagaggg ccctgcctgg ctgggagaga 4080
tggcactgga cggcctctgg attacagacc ctgccctgac agactatagg gtccagtggg 4140
tatcatggcc atggcttctt gcctggcctg gctctcttgg ttctgaggac tgaggaaagc 4200
tcagcctaga agggaagagg tctggaggga acatcctggg aacaggacaa gccactagga 4260
ctgagacaca tgcatcccaa cagggggctg cactttcatc cagaccagtc tttgtacaga 4320
gtgtattttg ttctgttttt acttttgctt ttttttttaa aaaaagatga aataaggaca 4380
cggagggaga gtggatgtta gggaatggtg tccctctttc ttcatttaca atgagatttg 4440
taaaatagct gggccccagc ctatgcctgg gagtggtccc aggctagacc ttactgctca 4500
cctgacacac agctcctcct tgagttgagt gtgtagaagt tttccaaaag tttgagatgg 4560
tttggctttg gggttgaggg actgggaagt taggatcctt tctgagggcc ctttggcaac 4620
aggatcattc ttcattggac gcactcattc caaggctacc cctagaatga agtccttccc 4680
tcccagtggg agagtggccc ttgaaaggag cactgtcaca tgactca 4727
<210> SEQ ID NO: 2
<211> LENGTH: 1259
<212> TYPE: PRT
<213> ORGANISM: Rattus norvegicus
<400> SEQENCE: 2
Met Ile Ile Met Glu Leu Ala Ala Trp Cys Arg Trp Gly Phe Leu Leu
1 5 10 15
Ala Leu Leu Pro Pro Gly Ile Ala Gly Thr Gln Val Cys Thr Gly Thr
20 25 30
Asp Met Lys Leu Arg Leu Pro Ala Ser Pro Glu Thr His Leu Asp Met
35 40 45
Leu Arg His Leu Tyr Gln Gly Cys Gln Val Val Gln Gly Asn Leu Glu
50 55 60
Leu Thr Tyr Val Pro Ala Asn Ala Ser Leu Ser Phe Leu Gln Asp Ile
65 70 75 80
Gln Glu Val Gln Gly Tyr Met Leu Ile Ala His Asn Gln Val Lys Arg
85 90 95
Val Pro Leu Gln Arg Leu Arg Ile Val Arg Gly Thr Gln Leu Phe Glu
100 105 110
Asp Lys Tyr Ala Leu Ala Val Leu Asp Asn Arg Asp Pro Gln Asp Asn
115 120 125
Val Ala Ala Ser Thr Pro Gly Arg Thr Pro Glu Gly Leu Arg Glu Leu
130 135 140
Gln Leu Arg Ser Leu Thr Glu Ile Leu Lys Gly Gly Val Leu Ile Arg
145 150 155 160
Gly Asn Pro Gln Leu Cys Tyr Gln Asp Met Val Leu Trp Lys Asp Val
165 170 175
Phe Arg Lys Asn Asn Gln Leu Ala Pro Val Asp Ile Asp Thr Asn Arg
180 185 190
Ser Arg Ala Cys Pro Pro Cys Ala Pro Ala Cys Lys Asp Asn His Cys
195 200 205
Trp Gly Glu Ser Pro Glu Asp Cys Gln Ile Leu Thr Gly Thr Ile Cys
210 215 220
Thr Ser Gly Cys Ala Arg Cys Lys Gly Arg Leu Pro Thr Asp Cys Cys
225 230 235 240
His Glu Gln Cys Ala Ala Gly Cys Thr Gly Pro Lys His Ser Asp Cys
245 250 255
Leu Ala Cys Leu His Phe Asn His Ser Gly Ile Cys Glu Leu His Cys
260 265 270
Pro Ala Leu Val Thr Tyr Asn Thr Asp Thr Phe Glu Ser Met His Asn
275 280 285
Pro Glu Gly Arg Tyr Thr Phe Gly Ala Ser Cys Val Thr Thr Cys Pro
290 295 300
Tyr Asn Tyr Leu Ser Thr Glu Val Gly Ser Cys Thr Leu Val Cys Pro
305 310 315 320
Pro Asn Asn Gln Glu Val Thr Ala Glu Asp Gly Thr Gln Arg Cys Glu
325 330 335
Lys Cys Ser Lys Pro Cys Ala Arg Val Cys Tyr Gly Leu Gly Met Glu
340 345 350
His Leu Arg Gly Ala Arg Ala Ile Thr Ser Asp Asn Val Gln Glu Phe
355 360 365
Asp Gly Cys Lys Lys Ile Phe Gly Ser Leu Ala Phe Leu Pro Glu Ser
370 375 380
Phe Asp Gly Asp Pro Ser Ser Gly Ile Ala Pro Leu Arg Pro Glu Gln
385 390 395 400
Leu Gln Val Phe Glu Thr Leu Glu Glu Ile Thr Gly Tyr Leu Tyr Ile
405 410 415
Ser Ala Trp Pro Asp Ser Leu Arg Asp Leu Ser Val Phe Gln Asn Leu
420 425 430
Arg Ile Ile Arg Gly Arg Ile Leu His Asp Gly Ala Tyr Ser Leu Thr
435 440 445
Leu Gln Gly Leu Gly Ile His Ser Leu Gly Leu Arg Ser Leu Arg Glu
450 455 460
Leu Gly Ser Gly Leu Ala Leu Ile His Arg Asn Ala His Leu Cys Phe
465 470 475 480
Val His Thr Val Pro Trp Asp Gln Leu Phe Arg Asn Pro His Gln Ala
485 490 495
Leu Leu His Ser Gly Asn Arg Pro Glu Glu Asp Cys Gly Leu Glu Gly
500 505 510
Leu Val Cys Asn Ser Leu Cys Ala His Gly His Cys Trp Gly Pro Gly
515 520 525
Pro Thr Gln Cys Val Asn Cys Ser His Phe Leu Arg Gly Gln Glu Cys
530 535 540
Val Glu Glu Cys Arg Val Trp Lys Gly Leu Pro Arg Glu Tyr Val Ser
545 550 555 560
Asp Lys Arg Cys Leu Pro Cys His Pro Glu Cys Gln Pro Gln Asn Ser
565 570 575
Ser Glu Thr Cys Phe Gly Ser Glu Ala Asp Gln Cys Ala Ala Cys Ala
580 585 590
His Tyr Lys Asp Ser Ser Ser Cys Val Ala Arg Cys Pro Ser Gly Val
595 600 605
Lys Pro Asp Leu Ser Tyr Met Pro Ile Trp Lys Tyr Pro Asp Glu Glu
610 615 620
Gly Ile Cys Gln Pro Cys Pro Ile Asn Cys Thr His Ser Cys Val Asp
625 630 635 640
Leu Asp Glu Arg Gly Cys Pro Ala Glu Gln Arg Ala Ser Pro Val Thr
645 650 655
Phe Ile Ile Ala Thr Val Val Gly Val Leu Leu Phe Leu Ile Leu Val
660 665 670
Val Val Val Gly Ile Leu Ile Lys Arg Arg Arg Gln Lys Ile Arg Lys
675 680 685
Tyr Thr Met Arg Arg Leu Leu Gln Glu Thr Glu Leu Val Glu Pro Leu
690 695 700
Thr Pro Ser Gly Ala Met Pro Asn Gln Ala Gln Met Arg Ile Leu Lys
705 710 715 720
Glu Thr Glu Leu Arg Lys Val Lys Val Leu Gly Ser Gly Ala Phe Gly
725 730 735
Thr Val Tyr Lys Gly Ile Trp Ile Pro Asp Gly Glu Asn Val Lys Ile
740 745 750
Pro Val Ala Ile Lys Val Leu Arg Glu Asn Thr Ser Pro Lys Ala Asn
755 760 765
Lys Glu Ile Leu Asp Glu Ala Tyr Val Met Ala Gly Val Gly Ser Pro
770 775 780
Tyr Val Ser Arg Leu Leu Gly Ile Cys Leu Thr Ser Thr Val Gln Leu
785 790 795 800
Val Thr Gln Leu Met Pro Tyr Gly Cys Leu Leu Asp His Val Arg Glu
805 810 815
His Arg Gly Arg Leu Gly Ser Gln Asp Leu Leu Asn Trp Cys Val Gln
820 825 830
Ile Ala Lys Gly Met Ser Tyr Leu Glu Asp Val Arg Leu Val His Arg
835 840 845
Asp Leu Ala Ala Arg Asn Val Leu Val Lys Ser Pro Asn His Val Lys
850 855 860
Ile Thr Asp Phe Gly Leu Ala Arg Leu Leu Asp Ile Asp Glu Thr Glu
865 870 875 880
Tyr His Ala Asp Gly Gly Lys Val Pro Ile Lys Trp Met Ala Leu Glu
885 890 895
Ser Ile Leu Arg Arg Arg Phe Thr His Gln Ser Asp Val Trp Ser Tyr
900 905 910
Gly Val Thr Val Trp Glu Leu Met Thr Phe Gly Ala Lys Pro Tyr Asp
915 920 925
Gly Ile Pro Ala Arg Glu Ile Pro Asp Leu Leu Glu Lys Gly Glu Arg
930 935 940
Leu Pro Gln Pro Pro Ile Cys Thr Ile Asp Val Tyr Met Ile Met Val
945 950 955 960
Lys Cys Trp Met Ile Asp Ser Glu Cys Arg Pro Arg Phe Arg Glu Leu
965 970 975
Val Ser Glu Phe Ser Arg Met Ala Arg Asp Pro Gln Arg Phe Val Val
980 985 990
Ile Gln Asn Glu Asp Leu Gly Pro Ser Ser Pro Met Asp Ser Thr Phe
995 1000 1005
Tyr Arg Ser Leu Leu Glu Asp Asp Asp Met Gly Asp Leu Val Asp
1010 1015 1020
Ala Glu Glu Tyr Leu Val Pro Gln Gln Gly Phe Phe Ser Pro Asp
1025 1030 1035
Pro Thr Pro Gly Thr Gly Ser Thr Ala His Arg Arg His Arg Ser
1040 1045 1050
Ser Ser Thr Arg Ser Gly Gly Gly Glu Leu Thr Leu Gly Leu Glu
1055 1060 1065
Pro Ser Glu Glu Gly Pro Pro Arg Ser Pro Leu Ala Pro Ser Glu
1070 1075 1080
Gly Ala Gly Ser Asp Val Phe Asp Gly Asp Leu Ala Met Gly Val
1085 1090 1095
Thr Lys Gly Leu Gln Ser Leu Ser Pro His Asp Leu Ser Pro Leu
1100 1105 1110
Gln Arg Tyr Ser Glu Asp Pro Thr Leu Pro Leu Pro Pro Glu Thr
1115 1120 1125
Asp Gly Tyr Val Ala Pro Leu Ala Cys Ser Pro Gln Pro Glu Tyr
1130 1135 1140
Val Asn Gln Ser Glu Val Gln Pro Gln Pro Pro Leu Thr Pro Glu
1145 1150 1155
Gly Pro Leu Pro Pro Val Arg Pro Ala Gly Ala Thr Leu Glu Arg
1160 1165 1170
Pro Lys Thr Leu Ser Pro Gly Lys Asn Gly Val Val Lys Asp Val
1175 1180 1185
Phe Ala Phe Gly Gly Ala Val Glu Asn Pro Glu Tyr Leu Val Pro
1190 1195 1200
Arg Glu Gly Thr Ala Ser Pro Pro His Pro Ser Pro Ala Phe Ser
1205 1210 1215
Pro Ala Phe Asp Asn Leu Tyr Tyr Trp Asp Gln Asn Ser Ser Glu
1220 1225 1230
Gln Gly Pro Pro Pro Ser Asn Phe Glu Gly Thr Pro Thr Ala Glu
1235 1240 1245
Asn Pro Glu Tyr Leu Gly Leu Asp Val Pro Val
<210> SEQ ID NO: 3
<211> LENGTH: 3768
<212> TYPE: DNA
<213> ORGANISM: Mus musculus
<400> SEQENCE: 3
atggagctgg cggcctggtg ccgttggggg ttcctcctcg ccctcctgtc ccccggagcc 60
gcgggtaccc aagtgtgtac cggtaccgac atgaagttgc gactccctgc cagtcctgag 120
acccacctgg acatgcttcg ccacctctac cagggctgtc aggtggtgca gggcaatttg 180
gagcttacct acctgcccgc caatgccagc ctctcattcc tgcaggacat ccaggaagtc 240
cagggataca tgctcatcgc tcacaaccga gtgaaacacg tcccactgca gaggttgcgc 300
atcgtgagag ggactcagct ctttgaggac aagtatgccc tggctgtgct agacaaccga 360
gaccctttgg acaacgtcac caccgccgcc ccaggcagaa ccccagaagg gctgcgggag 420
ctgcagcttc gaagtctcac agagatcttg aagggaggag ttttgatccg tgggaaccct 480
cagctctgct accaggacat ggttttgtgg aaggatgtcc tccgtaagaa taaccagctg 540
gctcctgtcg acatggacac caatcgttcc cgggcctgtc caccttgtgc cccaacctgc 600
aaagacaatc actgttgggg tgagagtcct gaagactgtc agatcttgac tggcaccatc 660
tgtactagtg gctgtgcccg gtgcaagggc cggctgccca ctgactgttg ccatgagcag 720
tgtgctgcag gctgcacggg tcccaagcat tctgactgcc tggcctgcct ccacttcaat 780
catagtggta tctgtgagct gcactgcccg gccctcatca cctacaacac agacaccttc 840
gagtccatgc tcaaccctga gggtcgctac acctttggtg ccagctgtgt gaccacctgc 900
ccctacaact acctctccac ggaagtggga tcctgcactc tggtctgtcc cccgaacaac 960
caagaggtca cagctgagga cggaacacag cggtgtgaga aatgcagcaa gccctgtgct 1020
ggagtatgct atggtctggg catggagcac ctccgagggg cgagggccat caccagtgac 1080
aatatccagg agtttgctgg ctgcaagaag atctttggga gcctggcatt tttgccggag 1140
agctttgatg ggaacccctc ctccggcgtt gccccactga agccagagca tctccaagtg 1200
ttcgaaaccc tggaggagat cacaggttac ctatacattt cagcatggcc agagagcttc 1260
caagacctca gtgtcttcca gaaccttcgg gtcattcggg gacggattct ccatgatggt 1320
gcttactcat tgacgttgca aggcctgggg attcactcac tggggctacg ctcactgcgg 1380
gagctgggca gtggattggc tctcattcac cgcaacaccc atctctgctt tgtaaacact 1440
gtaccttggg accagctctt ccggaacccg caccaggccc tactccacag tgggaaccgg 1500
ccagaagagg catgtggtct tgagggcttg gtctgtaact cactgtgtgc ccgtgggcac 1560
tgctgggggc cagggcccac ccagtgtgtc aactgcagtc agttcctccg gggccaggag 1620
tgtgtggagg agtgccgagt atggaagggg ctccccaggg agtatgtgag gggcaagcac 1680
tgtctgccat gccaccccga gtgtcagcct caaaacagct cggagacctg ctatggatcg 1740
gaggctgacc agtgtgaggc ttgtgcccac tacaaggact catcttcctg tgtggctcgc 1800
tgccccagtg gtgtgaagcc agacctctcc tacatgccta tctggaagta cccggatgag 1860
gagggcatat gtcagccatg ccccatcaac tgcacccact catgtgtgga cctggacgaa 1920
cgaggctgcc cagcagagca gagagccagc ccagtgacat tcatcattgc aactgtggtg 1980
ggcgtcctgt tgttcctgat catagtggtg gtcattggaa tcctaatcaa acgaaggcga 2040
cagaagatcc ggaagtatac catgcgtagg ctgctgcagg agaccgagct ggtggagccg 2100
ctgacgccca gtggagctgt gcccaaccag gctcagatgc ggatcctaaa ggagacagag 2160
ctaaggaagc tgaaggtgct tgggtcagga gccttcggca ctgtctacaa gggcatctgg 2220
atcccagatg gggagaacgt gaaaatcccc gtggccatca aggtgttgag ggaaaacaca 2280
tctcctaaag ctaacaaaga aatcctagat gaagcgtacg tcatggctgg tgtgggttct 2340
ccatatgtgt cccgcctcct gggcatctgc ctgacatcca cagtgcagct ggtgacacag 2400
cttatgccct atggctgcct tctggaccat gtccgagaac accgaggtcg cttaggctcc 2460
caggacctgc tcaactggtg tgttcagatt gccaagggga tgagctacct ggaggaagtt 2520
cggcttgttc acagggacct agctgcccga aacgtgctag tcaagagtcc caaccacgtc 2580
aagattaccg acttcgggct ggcacggctg ctggacattg atgagactga ataccatgca 2640
gatgggggca aggtgcccat caagtggatg gcattggaat ctattctcag acgccggttc 2700
acccatcaga gtgatgtgtg gagctatggt gtgactgtgt gggagctgat gacctttggg 2760
gccaaacctt acgatgggat cccagctcgg gagatccctg atttgctgga gaagggagaa 2820
cgcctacctc agcctccaat ctgcaccatc gacgtctaca tgatcatggt caaatgttgg 2880
atgattgact ccgaatgtcg cccgagattc cgggagttgg tatcagaatt ctcccgtatg 2940
gcaagggacc cccagcgctt tgtggtcatc cagaacgagg acttaggccc ctccagcccc 3000
atggacagca ccttctaccg ttcactgctg gaggatgatg acatggggga gctggtcgat 3060
gctgaagagt acctggtacc ccagcaggga ttcttctccc cagaccctgc cctaggtact 3120
gggagcacag cccaccgcag acaccgcagc tcgtcggcca ggagtggcgg tggtgagctg 3180
acactgggcc tggagccctc ggaagaagag ccccccagat ctccactggc tccctccgaa 3240
ggggctggct ccgatgtgtt tgatggtgac ctggcagtgg gggtaaccaa aggactgcag 3300
agcctctctc cacatgacct cagccctcta cagcggtaca gtgaggatcc cacattacct 3360
ctgccccccg agactgatgg ctacgttgct cccctggcct gcagccccca gcccgagtat 3420
gtgaaccagc cagaggttcg gcctcagtct cccttgaccc cagagggtcc tccgcctccc 3480
atccgacctg ctggtgctac tctagaaaga cccaagactc tctctcctgg gaaaaatggg 3540
gttgtcaaag acgtttttgc ctttgggggt gctgtggaga accctgaata cttagcaccc 3600
agagcaggca ctgcctctca gccccaccct tctcctgcct tcagcccagc ctttgacaac 3660
ctctattact gggaccagaa ctcatcggag cagggtcctc caccaagtac ctttgaaggg 3720
acccccactg cagagaaccc tgagtaccta ggcctggatg tgccagta 3768
<210> SEQ ID NO: 4
<211> LENGTH: 3765
<212> TYPE: DNA
<213> ORGANISM: homo sapiens
<400> SEQENCE: 4
atggagctgg cggccttgtg ccgctggggg ctcctcctcg ccctcttgcc ccccggagcc 60
gcgagcaccc aagtgtgcac cggcacagac atgaagctgc ggctccctgc cagtcccgag 120
acccacctgg acatgctccg ccacctctac cagggctgcc aggtggtgca gggaaacctg 180
gaactcacct acctgcccac caatgccagc ctgtccttcc tgcaggatat ccaggaggtg 240
cagggctacg tgctcatcgc tcacaaccaa gtgaggcagg tcccactgca gaggctgcgg 300
attgtgcgag gcacccagct ctttgaggac aactatgccc tggccgtgct agacaatgga 360
gacccgctga acaataccac ccctgtcaca ggggcctccc caggaggcct gcgggagctg 420
cagcttcgaa gcctcacaga gatcttgaaa ggaggggtct tgatccagcg gaacccccag 480
ctctgctacc aggacacgat tttgtggaag gacatcttcc acaagaacaa ccagctggct 540
ctcacactga tagacaccaa ccgctctcgg gcctgccacc cctgttctcc gatgtgtaag 600
ggctcccgct gctggggaga gagttctgag gattgtcaga gcctgacgcg cactgtctgt 660
gccggtggct gtgcccgctg caaggggcca ctgcccactg actgctgcca tgagcagtgt 720
gctgccggct gcacgggccc caagcactct gactgcctgg cctgcctcca cttcaaccac 780
agtggcatct gtgagctgca ctgcccagcc ctggtcacct acaacacaga cacgtttgag 840
tccatgccca atcccgaggg ccggtataca ttcggcgcca gctgtgtgac tgcctgtccc 900
tacaactacc tttctacgga cgtgggatcc tgcaccctcg tctgccccct gcacaaccaa 960
gaggtgacag cagaggatgg aacacagcgg tgtgagaagt gcagcaagcc ctgtgcccga 1020
gtgtgctatg gtctgggcat ggagcacttg cgagaggtga gggcagttac cagtgccaat 1080
atccaggagt ttgctggctg caagaagatc tttgggagcc tggcatttct gccggagagc 1140
tttgatgggg acccagcctc caacactgcc ccgctccagc cagagcagct ccaagtgttt 1200
gagactctgg aagagatcac aggttaccta tacatctcag catggccgga cagcctgcct 1260
gacctcagcg tcttccagaa cctgcaagta atccggggac gaattctgca caatggcgcc 1320
tactcgctga ccctgcaagg gctgggcatc agctggctgg ggctgcgctc actgagggaa 1380
ctgggcagtg gactggccct catccaccat aacacccacc tctgcttcgt gcacacggtg 1440
ccctgggacc agctctttcg gaacccgcac caagctctgc tccacactgc caaccggcca 1500
gaggacgagt gtgtgggcga gggcctggcc tgccaccagc tgtgcgcccg agggcactgc 1560
tggggtccag ggcccaccca gtgtgtcaac tgcagccagt tccttcgggg ccaggagtgc 1620
gtggaggaat gccgagtact gcaggggctc cccagggagt atgtgaatgc caggcactgt 1680
ttgccgtgcc accctgagtg tcagccccag aatggctcag tgacctgttt tggaccggag 1740
gctgaccagt gtgtggcctg tgcccactat aaggaccctc ccttctgcgt ggcccgctgc 1800
cccagcggtg tgaaacctga cctctcctac atgcccatct ggaagtttcc agatgaggag 1860
ggcgcatgcc agccttgccc catcaactgc acccactcct gtgtggacct ggatgacaag 1920
ggctgccccg ccgagcagag agccagccct ctgacgtcca tcatctctgc ggtggttggc 1980
attctgctgg tcgtggtctt gggggtggtc tttgggatcc tcatcaagcg acggcagcag 2040
aagatccgga agtacacgat gcggagactg ctgcaggaaa cggagctggt ggagccgctg 2100
acacctagcg gagcgatgcc caaccaggcg cagatgcgga tcctgaaaga gacggagctg 2160
aggaaggtga aggtgcttgg atctggcgct tttggcacag tctacaaggg catctggatc 2220
cctgatgggg agaatgtgaa aattccagtg gccatcaaag tgttgaggga aaacacatcc 2280
cccaaagcca acaaagaaat cttagacgaa gcatacgtga tggctggtgt gggctcccca 2340
tatgtctccc gccttctggg catctgcctg acatccacgg tgcagctggt gacacagctt 2400
atgccctatg gctgcctctt agaccatgtc cgggaaaacc gcggacgcct gggctcccag 2460
gacctgctga actggtgtat gcagattgcc aaggggatga gctacctgga ggatgtgcgg 2520
ctcgtacaca gggacttggc cgctcggaac gtgctggtca agagtcccaa ccatgtcaaa 2580
attacagact tcgggctggc tcggctgctg gacattgacg agacagagta ccatgcagat 2640
gggggcaagg tgcccatcaa gtggatggcg ctggagtcca ttctccgccg gcggttcacc 2700
caccagagtg atgtgtggag ttatggtgtg actgtgtggg agctgatgac ttttggggcc 2760
aaaccttacg atgggatccc agcccgggag atccctgacc tgctggaaaa gggggagcgg 2820
ctgccccagc cccccatctg caccattgat gtctacatga tcatggtcaa atgttggatg 2880
attgactctg aatgtcggcc aagattccgg gagttggtgt ctgaattctc ccgcatggcc 2940
agggaccccc agcgctttgt ggtcatccag aatgaggact tgggcccagc cagtcccttg 3000
gacagcacct tctaccgctc actgctggag gacgatgaca tgggggacct ggtggatgct 3060
gaggagtatc tggtacccca gcagggcttc ttctgtccag accctgcccc gggcgctggg 3120
ggcatggtcc accacaggca ccgcagctca tctaccagga gtggcggtgg ggacctgaca 3180
ctagggctgg agccctctga agaggaggcc cccaggtctc cactggcacc ctccgaaggg 3240
gctggctccg atgtatttga tggtgacctg ggaatggggg cagccaaggg gctgcaaagc 3300
ctccccacac atgaccccag ccctctacag cggtacagtg aggaccccac agtacccctg 3360
ccctctgaga ctgatggcta cgttgccccc ctgacctgca gcccccagcc tgaatatgtg 3420
aaccagccag atgttcggcc ccagccccct tcgccccgag agggccctct gcctgctgcc 3480
cgacctgctg gtgccactct ggaaaggccc aagactctct ccccagggaa gaatggggtc 3540
gtcaaagacg tttttgcctt tgggggtgcc gtggagaacc ccgagtactt gacaccccag 3600
ggaggagctg cccctcagcc ccaccctcct cctgccttca gcccagcctt cgacaacctc 3660
tattactggg accaggaccc accagagcgg ggggctccac ccagcacctt caaagggaca 3720
cctacggcag agaacccaga gtacctgggt ctggacgtgc cagtg 3765
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