U.S. patent application number 14/285577 was filed with the patent office on 2014-11-20 for peptides immunoreactive with autoantibodies from patients suffering from rheumatoid arthritis.
This patent application is currently assigned to STICHTING VOOR DE TECHNISCHE WETENSCHAPPEN. The applicant listed for this patent is Stichting Voor de Technische Wetenschappen. Invention is credited to Rene Michael Antonius HOET, Jozef Maria Hendrik RAATS, Gerardus Antonius SCHELLEKENS, Waltherus Jacobus Wilhelmus VAN VENROOIJ.
Application Number | 20140342378 14/285577 |
Document ID | / |
Family ID | 19763876 |
Filed Date | 2014-11-20 |
United States Patent
Application |
20140342378 |
Kind Code |
A1 |
VAN VENROOIJ; Waltherus Jacobus
Wilhelmus ; et al. |
November 20, 2014 |
PEPTIDES IMMUNOREACTIVE WITH AUTOANTIBODIES FROM PATIENTS SUFFERING
FROM RHEUMATOID ARTHRITIS
Abstract
The invention relates to a peptide derived from an antigen
recognized by autoantibodies, which peptide is reactive with
autoimmune antibodies from a patient suffering from rheumatoid
arthritis. The peptide according to the invention possesses a
modified arginine residue. The invention also relates to antibodies
against the peptide and a method of detecting autoimmune
antibodies.
Inventors: |
VAN VENROOIJ; Waltherus Jacobus
Wilhelmus; (Nijmegen, NL) ; SCHELLEKENS; Gerardus
Antonius; (Den Bosch, NL) ; RAATS; Jozef Maria
Hendrik; (Nijmegen, NL) ; HOET; Rene Michael
Antonius; (Nijmegen, NL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Stichting Voor de Technische Wetenschappen |
Utrecht |
|
NL |
|
|
Assignee: |
STICHTING VOOR DE TECHNISCHE
WETENSCHAPPEN
Utrecht
NL
|
Family ID: |
19763876 |
Appl. No.: |
14/285577 |
Filed: |
May 22, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
11834557 |
Aug 6, 2007 |
8772448 |
|
|
14285577 |
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11059775 |
Feb 16, 2005 |
7335724 |
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11834557 |
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09308150 |
Sep 30, 1999 |
6858438 |
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PCT/NL1997/000624 |
Nov 14, 1997 |
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11059775 |
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Current U.S.
Class: |
435/7.92 ;
530/350; 530/387.9; 530/391.1; 530/391.3 |
Current CPC
Class: |
G01N 2800/102 20130101;
G01N 2800/24 20130101; G01N 33/564 20130101; G01N 2800/10 20130101;
C07K 14/4713 20130101; G01N 33/6893 20130101; C07K 16/18 20130101;
C07K 14/47 20130101; G01N 2333/46 20130101 |
Class at
Publication: |
435/7.92 ;
530/350; 530/387.9; 530/391.3; 530/391.1 |
International
Class: |
G01N 33/68 20060101
G01N033/68; C07K 16/18 20060101 C07K016/18; C07K 14/47 20060101
C07K014/47 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 15, 1996 |
NL |
1004539 |
Claims
1. An artificial antigen consisting of a recombinant or synthetic
polypeptide which comprises a modified amino acid sequence derived
from protein having the amino acid sequence of SEQ ID NO: 17 or a
fragment thereof by replacing at least one arginine residue with a
citrulline residue, which antigen is specifically immunoreactive
with anti-filaggrin antibodies present in the serum of subjects
suffering from rheumatoid arthritis.
2. The antigen of claim 1 wherein one arginine is replaced by
citrulline.
3. The antigen of claim 1 wherein two or more arginines are
replaced by citrulline.
4. A method to detect rheumatoid arthritis by detecting an
autoimmune antibody in the serum of a subject, said method
comprising contacting the antigen of claim 1 with said serum and
detecting the presence or absence of a complex between the antigen
and an antibody, wherein the presence of said complex detects
rheumatoid arthritis.
5. The method of claim 4 wherein the detecting comprises use of an
antihuman antibody.
6. The method of claim 4 wherein the detecting comprises use of an
enzyme-linked immunosorbent assay (ELISA).
7. The method of claim 4 wherein said contacting is with more than
one artificial antigen consisting of a recombinant or synthetic
polypeptide which comprises a modified amino acid sequence derived
from filaggrin by replacing at least one arginine residue with a
citrulline residue of a wild type filaggrin amino acid sequence,
which antigen is specifically immunoreactive with anti-filaggrin
antibodies present in the serum of subjects suffering from
rheumatoid arthritis.
8. An isolated antibody crossreactive with an antibody raised
against an antigen which is a synthetic peptide which comprises an
amino acid sequence that is the same as that of a fragment having
no more than 19 amino acids of the amino acid sequence of SEQ ID
NO:17 and having sufficient length to act as an epitope for
antibody binding, except that said peptide has at least one
arginine residue of SEQ ID NO:17 replaced by a citrulline
residue.
9. The antibody of claim 8 wherein said antigen is in cyclic
form.
10. The antibody of claim 8 wherein said antigen contains at least
two cysteines.
11. The antibody of claim 8 which is a monoclonal antibody.
12. The antibody of claim 9 which is a monoclonal antibody.
13. The antibody of claim 8 which is labeled.
14. The antibody of claim 9 which is labeled.
15. The antibody of claim 8 which is coupled to a carrier.
16. The antibody of claim 9 which is coupled to a carrier.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of U.S. Ser. No.
11/834,557 filed 6 Aug. 2007, now allowed, which is a continuation
of U.S. Ser. No. 11/059,775 filed 16 Feb. 2005 now U.S. Pat. No.
7,335,724, which is a continuation of U.S. Ser. No. 09/308,150
filed 30 Sep. 1999 now U.S. Pat. No. 6,858,438, which is the
national phase of PCT application PCT/NL97/00624 having an
international filing date of 14 Nov. 1997, which claims priority
from Netherlands application 1004539 filed 15 Nov. 1996. The
contents of these documents are incorporated herein by
reference.
REFERENCE TO SEQUENCE LISTING SUBMITTED VIA EFS-WEB
[0002] The entire content of the following electronic submission of
the sequence listing via the USPTO EFS-WEB server, as authorized
and set forth in MPEP .sctn.1730 II.B.2(a)(C), is incorporated
herein by reference in its entirety for all purposes. The sequence
listing is identified on the electronically filed text file as
follows:
TABLE-US-00001 File Name Date of Creation Size (bytes)
595402000103seqlist.txt May 21, 2014 18,575 bytes
TECHNICAL FIELD
[0003] Peptide derived from an antigen recognized by autoantibodies
from patients with rheumatoid arthritis, antibody directed against
said peptide, a combinatorial antigen, and a method of detecting
auto-immune antibodies.
[0004] The present invention relates to a peptide derived from an
antigen recognized by autoantibodies from patients with rheumatoid
arthritis, which peptide is reactive with autoimmune antibodies
from a patient suffering from rheumatoid arthritis.
[0005] Such a peptide is known from the European patent application
0 511 116 (Clonatec S.A.). This application describes an antigen
comprising a filaggrin or profilaggrin fragment. The peptide is
recognized by rheumatoid arthritis-specific autoimmune antibodies.
Rheumatoid arthritis (RA) is a systemic autoimmune disease. It is
the most commonly occurring inflammatory disease of the joints, it
is chronic and may lead to severe physical disablement.
[0006] The object of the present is to provide a peptide which is
reactive with autoimmune antibodies from a patient suffering from
rheumatoid arthritis, which peptide is suitable for diagnostic
research with increased specificity while also being useful for
other purposes such as obtaining (raising, selecting and isolating)
polyand monoclonal antibodies.
[0007] To this end the peptide according to the invention is
characterized in that the derived peptide that is reactive with
autoimmune antibodies, corresponds to a part of a mRNA molecule
coding for the antigen, said part comprising a codon for an
arginine residue, and the arginine residue in the derived peptide,
which is reactive with autoimmune antibodies, is a modified
arginine residue.
[0008] Surprisingly, the peptide according to the invention that
possesses a modified arginine residue, proved to be very suitable
for the specific diagnosis of rheumatoid arthritis.
[0009] To this day, no specific serological test is available for
RA. The only test frequently employed is based on the determination
of rheumatoid factors (RF; Ref. 1) which are found in 70 of the RA
patients. However, this test is not very specific and is
characterized by a relatively large number of false positives. For
patients suffering from systemic lupus erythematosus the percentage
of false positives is approximately 20% and for healthy individuals
approximately 5%.
[0010] Preferably the peptide is characterized in that the modified
arginine residue's side chain is a side chain according to Formula
I on the formula sheet, in which
[0011] X=NH.sub.2, CH.sub.3, NHCH.sub.3 or N(CH.sub.3).sub.2;
[0012] Y=o, NH, NHCH.sub.3 or N(CH.sub.3).sub.2;
[0013] Z=O, NH or CH.sub.2; and
[0014] n=2, 3 or 4, on the condition that when X=NH.sub.2 and Z=NH,
Y is not NH; and the modified arginine residue is in particular a
citrulline residue. For citrulline, X=NH.sub.2, Y=0, Z=NH and
n=3.
[0015] A preferred peptide is the peptide selected from the group
of peptides having the Formula II X on the formula sheet.
[0016] By using the peptide according to Formula II, it is possible
to establish the presence of rheumatoid arthritis in about 36% of
patients actually suffering from rheumatoid arthritis, while the
percentage of false positives for other autoimmune diseases and
healthy individuals is less than 2%.
[0017] According to a favourable embodiment the peptide is a cyclic
peptide, for instance, due to the presence of a cystine
residue.
[0018] In some cases such a cyclic peptide exhibits an increased
immunological affinity.
[0019] The preferred cyclic peptide is the peptide having the
Formula XI on the formula sheet.
[0020] Preferably the peptide is a synthetic peptide.
[0021] The reactive peptide according to the invention can be
obtained pure and in large quantities by means of organic
synthesis, making immunological testing on a large scale
possible.
[0022] According to an alternative embodiment, the peptide in
accordance with the invention is characterized in that the peptide
is obtained by the proteolytic treatment of (pro)filaggrin,
separation of peptide fragments formed by proteolysis and
subsequent selection on the presence of a modified arginine residue
in a peptide which was formed during the proteolytic treatment.
[0023] In this manner peptides can be identified which can increase
the sensitivity of a rheumatoid arthritis test. The term
sensitivity is in the present application to be understood to mean
the ability of a test to properly identify a patient suffering from
rheumatoid arthritis.
[0024] According to a favourable embodiment, the antigen is (pro)
filaggrin (SEQ ID NO: 17), and the peptide is reactive with a
rheumatoid arthritis patient's autoimmune antibodies which are
reactive with (pro) filaggrin.
[0025] The peptide has been shown to be very suitable for
high-specificity testing (few false positives) for rheumatism.
[0026] The present invention also relates to an antibody which is
cross-reactive with an antibody raised against a peptide according
to the invention.
[0027] Such an antibody is useful for the indication of rheumatoid
arthritis by analysing sections of biopsy samples and immunological
tests of the sandwich type.
[0028] The antibody is preferably a monoclonal antibody.
[0029] According to another preferred embodiment, the antibody is
obtained by using as antigen a peptide in accordance with the
invention.
[0030] A suitable antibody according to the invention is
characterized in that it is cross-reactive with the antibody as
produced by Escherichia coli TG1 with plasmid RA3, deposited at the
Centraalbureau voor Schimmelcultures, at Baarn, the Netherlands
under accession number CBS143.96.
[0031] The invention further relates to an organic compound
comprising a part that is able to compete with a peptide according
to one of the claims 1 to 9 for binding to an antibody which is
specific for said peptide, wherein at least said part of the
organic compound can be prepared by means of combinationary
chemistry.
[0032] Such organic compounds are found by competitive selection
wherein a peptide of the invention competes for recognition by an
antibody of the invention, such as the antibody produced by E. coli
CBS143.96. The organic compounds, which are often cheaper to
produce than antigens that are prepared solely on the basis of
amino acids that may or may not comprise side chains, are suitable
for immunological kits for diagnosing RA. Also, after coupling to a
solid carrier, said organic compounds could be applied to lower,
through adsorption, the level of autoimmune antibodies in the blood
of patients suffering from RA.
[0033] Finally, the invention relates to a method of detecting
autoimmune antibodies against rheumatoid arthritis.
[0034] The method according to the invention is characterized in
that in an immunological test at least one immunologically active
molecule selected from the group consisting of i) a peptide
according to the invention; ii) a recombinatory organic molecule
according to the invention; and iii) an antibody according to the
invention is used.
[0035] In addition to increased sensitivity other advantages are
achieved, in particular better reproducibility, quantitative
information and better applicability for prognostic purposes.
[0036] To a person skilled in the art it will be apparent that
there are a number of possible variations to the present invention
as specified by the appended claims. For instance, the peptides
mentioned on the formula sheet may also be part of other
oligopeptides. They may be provided at one or both ends with one or
more other amino acids while also, two or more peptides according
to the invention may be part of one oligopeptide. It is also
possible to shorten the peptides by one or more amino acids,
provided this does not have a significantly adverse effect on the
reactivity. The expert is familiar with the manner in which
peptides and organic compounds according to the invention may
optionally be labelled or be coupled to a carrier, and how on the
basis of such antigens an immunological test may be developed,
using the standard techniques well-known in the field.
[0037] The invention will now be explained in more detail by means
of the following example.
Materials and Methods
[0038] Peptide Synthesis:
[0039] Peptides were selected for synthesis on the basis of amino
acid sequences derived from known cDNA sequences of human
profilaggrin (Ref. 2; Ref. 3). The peptides were synthesized on
solid phase using the method described by Schellekens et al. (Ref.
4). The peptides were at least 95% pure, as determined by the
elution profile by means of reversed phase chromatography and the
relative absorption at 214 nm. The composition of the peptides was
confirmed by means of mass spectrometry (MALDI-MS). All peptides
were synthesized as peptide amides.
TABLE-US-00002 TABLE 1 SEQ ID Name Peptide sequence* NO. cfc1 S H Q
E S T X G R S R G R S G R S G S 1 cfc2 S H Q E S T R G X S R G R S
G R S G S 2 cfc3 S H Q E S T R G R S X G R S G R S G S 3 cfc4 S H Q
E S T R G R S R G X S G R S G S 4 cfc5 S H Q E S T R G R S R G R S
G X S G S 5 cfc6 S H Q E S T X G X S R G R S G R S G S 6 cfc7 S H Q
E S T X G R S X G R S G R S G S 7 cfc8 S H Q E S T X G R S R G X S
G R S G S 8 cfc9 S H Q E S T X G R S R G R S G X S G S 9 cf S H Q E
S T R G R S R G R S G R S G S 11 cfA S H Q E S T A G R S R G R S G
R S G S 12 cfE S H Q E S T E G R S R G R S G R S G S 13 cfQ S H Q E
S T Q G R S R G R S G R S G S 14 nfc1 T G P S T R G R Q G S X H E
15 nf E S S H G W T G P S T R G R Q G S R H E 16 *(A = alanine; G =
glycine; H = histidine; E = glutamic acid; P = proline; R =
arginine; Q = glutamine; S = serine; T = threonine; W = tryptophan;
X = citrulline)
Synthesized Peptides
[0040] The peptide names starting with "cf" are based on the
Cterminal end (amino acids 306-324); and the peptide names staring
with "nf" are based on the sequence near the Nterminal end (amino
acids 18-32 for nfc1).
[0041] Amino acid sequences based on cDNA of a profilaggrin
repeat.
Detection by Means of ELISA
[0042] Via an N-oxysuccinimide surface the peptides were covalently
coupled to the wells of 96-well microtitre plates (Costar amide
binding plates) in an amount of 1 g/well. Coupling took place for
16 hours at 4EC and pH 9.0. The plates were blocked for 1 hour with
2% bovine serum albumin. The sera were diluted 200 times in a
diluent (0.3% BSA, 350 mM NaCl, 10 mM Tris-HCl pH 7.6, 1% vol./vol.
Triton X-100, 0.5% w./vol. Na-deoxycholate, 0.1% SDS) supplemented
with 10% normal rabbit serum, and incubated for one hour at room
temperature. After washing the plates (3 times with PBS containing
0.05 by vol. of Tween.RTM.20), 100 l of antihuman IgG conjugated
with peroxidase (Dako P214), 1000 times diluted in dilution buffer,
was added to the wells. After incubation for 1 hour at room
temperature, the plates were washed 3 times with PBS/Tween.RTM.,
and bound antibodies were detected with tetramethyl benzidine as a
substrate. After 10 minutes the reaction was stopped by adding 100
l of 2 M sulphuric acid per well. Readout occurred at 450 nm. Sera
having an OD450 of 0.2, after deduction of the blank for the
respective serum (a well without a coupled peptide), were
considered to be positive.
Results
[0043] The results are listed in Table 2. In total, 288 sera from
patients suffering from rheumatoid diseases were used, 132 of which
were from patients suffering from rheumatoid arthritis.
TABLE-US-00003 TABLE 2 Results with peptide cfc1 to cfc9 (Formula
II to X of the formula sheet) control PM/ RA sera sera.sup.1
SLE.sup.2 SSC.sup.3 pSS.sup.4 DM.sup.5 (%) (%) (%) (%) (%) (%)
Peptide* (n = 134) (n = 154) (n = 50) (n = 50) (n = 50) (n = 50)
cfc1 49 (36) 1 (0.6) 1 (2) 0 0 0 cfc2 27 (20) 4 (2.6) 1 (2) 0 1 (2)
1 (2) cfc3 37 (28) 2 (0.6) 0 0 1 (2) 1 (2) cfc4 32 (24) 2 (1.3) 0 0
0 0 cfc5 64 (48) 1 (0.6) 0 1 (2) 2 (4) 1 (2) cfc6 65 (48) 1 (0.6) 0
0 2 (4) 1 (2) cfc7 60 (45) 1 (0.6) 0 0 1 (2) 1 (2) cfc8 55 (41) 1
(0.6) 0 0 1 (2) 1 (2) cfc9 57 (42) 1 (0.6) 0 0 2 (4) 0
.sup.1)Control sera are from patients suffering from rheumatic
diseases other than RA. .sup.2)SLE is systemic lupus erythematosus.
.sup.3)pSS is primary Sjogren's syndrome. .sup.4)SSC is systemic
scleroderma. .sup.5)PM/DM is polymyositis/dermatomyositis.
[0044] Of the total of 134 RA sera from patients suffering from
rheumatoid arthritis, 102 were positive with at least one peptide
from the cfc1 to cfc9 series. Therefore, when using these peptides,
the sensitivity was 76% ( 102/134). Of the total of 354 control
sera, 13 sera were positive on at least one peptide from the cfc1
to cfc9 series. Therefore the test sensitivity, expressed as
percentage of true positives, was 96%. Of the 37 sera that were
reactive with cfc3, none were not recognized by peptide cfc1 or
cfc23. Of the sera that were reactive with cfc7, cfc8 and cfc9,
none were not recognized by cfc1, cfc2, cfc4, cfc5 or cfc6. This
means that cfc2, cfc7, cfc8 and cfc9 do not contribute to the test
sensitivity and a test sensitivity of 76% may be realized by using
the combination of the peptides cfc1, cfc2, cfc4, cfc5 and
cfc6.
[0045] It should be noted that these percentages depend on the
specificity-threshold value applied by applicants. The same data
(from the ELISA experiments) can be interpreted as a sensitivity of
approximately 80-85% by choosing a slightly lower sensitivity,
which incidentally, is still much better than the one obtainable
when using the known rheumatoid factor test (Ref. 1).
[0046] Sera from patients suffering from various infectious
diseases (Borrelia, syphilis, malaria, endocarditis, Legionella,
tuberculosis, mycoplasma, Yersinia, salmonella, parvovirus B19,
Epstein-Barr virus, rubella, schistosomiasis, Toxoplasma,
leishmaniasis, Chagas' disease) were tested for the presence of
antibodies reactive with cfc1. Of the 308 tested sera 9 were
positive. This means that the specificity was 97%, a considerable
improvement compared with the RF test.
[0047] Variants of cfc1 wherein citrulline was replaced by a
neutral (alanine; cfA), acid (glutamic acid; cfE) or amide
(glutamine; cfQ) residue, did not seem to be immunologically
reactive. The same applies to the control peptide cf, which does
not possess a modified arginine residue.
[0048] With the aid of the above-described ELISA, a cyclic variant
(with the Formula XI on the formula sheet, in which two cysteine
residues (C) are bound by means of a sulphur bridge) of cfc1 was
tested for 134 RA sera. This cyclic variant was shown to be
reactive with 85 sera (63%), signifying an increase in sensitivity.
Of the 154 sera of patients suffering from rheumatic diseases other
than RA, 3 were shown to be positive (specificity 98%). The
priority document of the present application reports 5 falsely
determined positives. However, it has been shown that in two of
these cases the patients did indeed suffer from RA. Not one serum
from 59 healthy individuals was positive with this cyclic peptide,
nor with any of the peptides cfc1 to cfc9. The cyclic peptide
variant was shown to be reactive with 4 sera of the 200 additional
control sera (50 SLE, 50 SCC, 50 pSS, 50 PM/DM) so that the
specificity in respect of these sera was 98 k. Of the sera from
patients suffering from various infectious diseases (308 sera as
described above), 7 sera were shown to be positive with the cyclic
peptide variant so that in this case also the specificity in
respect of these sera was 98%. The use of the cyclic peptide
variant thus enhances the sensitivity compared with the individual
linear peptide variants, but the specificity is also enhanced due
to an improved signal/noise ratio in the described ELISA test.
[0049] A second citrulline-substituted peptide (nfc1) was shown to
be specifically reactive with 10% of the RA sera, but not with the
control peptide nf, which does not comprise citrulline. Of the RA
sera reactive with nfc1, some were not reactive with cfc1 to cfc9.
Therefore, it is possible to increase the sensitivity of a test for
rheumatoid arthritis by applying more peptides comprising a
modified arginine residue.
[0050] Obviously, a peptide may comprise several modified arginine
residues, but the peptide may also comprise one or more
non-modified arginine residues.
[0051] Applicants believe that modified amino acids, in particular
those derived from arginine residues, could possibly also play a
role in other autoimmune diseases. For this reason, the invention
is also aimed at peptides comprising modified amino acids that are
reactive with auto-antibodies from patients suffering from
autoimmune diseases other than RA. This relates especially to
peptides comprising a modified arginine residue wherein X=NHCH3
(wherein Y=NH or NCH.sub.3) or NH(CH.sub.3).sub.2 is, which
peptides will be useful for the detection of autoimmune diseases
such as SLE, scleroderma, primary Sjogrens syndrome and
polymyositis/dermatomyositis, in which nuclear autoantigens play a
role. Said peptides are useful for the development of monoclonal
antibodies against these diseases as well as for diagnosing the
respective autoimmune diseases, in particular for the detection of
autoimmune antibodies in body fluid such as blood, plasma and serum
of patients who are suspected of suffering from the autoimmune
disease. Again the peptides and antibodies offer the possibility of
developing an organic compound with the aid of combinatorial
chemistry, which compound is comprised within the scope of the
invention.
[0052] The recombinant monoclonal antibody described by applicants
is reactive with peptide cfc1 but not with the control peptides
cfA, cfE, cfQ or cf. The commercially available monoclonal antibody
AKH1 (Ref. 5), directed against human filaggrin, is not reactive
with any of the peptides described herein and is therefore not
crossreactive with an antibody raised against a peptide according
to the invention. The polyclonal serum anti-54 kD (Ref. 5), raised
against filaggrin, is not reactive with any of the peptides
described herein and is therefore not cross-reactive with an
antibody reactive with a peptide according to the invention. This
suggests that in a normal immune reaction antibodies that are
cross-reactive with an antibody raised against a peptide according
to the invention, are not necessarily formed.
REFERENCES
[0053] 1) Smolen, J. S., (1996) Autoantibodies in rheumatoid
arthritis, in Manual of biological markers of disease (W. J. van
Venrooij and R. N. Maini, red.) vol. C Chapter 1.1 pp. 1-18. Kluwer
Scientific Publishers, Dordrecht. [0054] 2) McKinley-Grant, L. J.,
Idler, W. W., Bernstein, I. A., Parry, D. A. D., Cannizzaro, L.,
Croce, C. M., Huebner, K., Lessin, S. R. & Steinert, P. M.
(1989) Characterization of a cDNA clone encoding human filaggrin
and localization of the gene to chromosome region lq21. Proceedings
of the National Academy of Science U.S.A. 86, pp. 4848-4852. [0055]
3) Gan, S. Q., McBride, O. W., Idler, W. W., Nedialka, M. &
Steinert, P. M. (1990) Organization, structure, and polymorphisms
of the human profilaggrin gene. Biochemistry 29, pp. 9432-9440.
[0056] 4) Schellekens, G. A., Lasonder, E., Feijlbrief, M.,
Koedijk, D. G. A. M., Drijfhout, J. W., Scheffer, A. J.,
Welling-Wester, S & Welling, G. W. (1994) Identification of the
core residues of the epitope of a monoclonal antibody raised
against glycoprotein D of herpes simplex virus 1 by screening of a
random peptide library. The European Journal of Immunology 24, pp.
3188-3193. [0057] 5) Hoet, R. M. A., Boerbooms, A. A. Th., Arends,
M., Ruiter, D. J., van Venrooij, W. J. (1991) Antiperinuclear
factor, a marker autoantibody for rheumatoid arthritis:
colocalisation of the perinuclear factor and prof ilaggrin. Annals
of the Rheumatic diseases 50, pp. 611-618.
Sequence CWU 1
1
17119PRTArtificial SequenceDerived from known cDNA sequences of
human profilaggrin 1Ser His Gln Glu Ser Thr Xaa Gly Arg Ser Arg Gly
Arg Ser Gly Arg1 5 10 15 Ser Gly Ser219PRTArtificial
SequenceDerived from known cDNA sequences of human profilaggrin
2Ser His Gln Glu Ser Thr Arg Gly Xaa Ser Arg Gly Arg Ser Gly Arg1 5
10 15 Ser Gly Ser319PRTArtificial SequenceDerived from known cDNA
sequences of human profilaggrin 3Ser His Gln Glu Ser Thr Arg Gly
Arg Ser Xaa Gly Arg Ser Gly Arg1 5 10 15 Ser Gly
Ser419PRTArtificial SequenceDerived from known cDNA sequences of
human profilaggrin 4Ser His Gln Glu Ser Thr Arg Gly Arg Ser Arg Gly
Xaa Ser Gly Arg1 5 10 15 Ser Gly Ser519PRTArtificial
SequenceDerived from known cDNA sequences of human profilaggrin
5Ser His Gln Glu Ser Thr Arg Gly Arg Ser Arg Gly Arg Ser Gly Xaa1 5
10 15 Ser Gly Ser619PRTArtificial SequenceDerived from known cDNA
sequences of human profilaggrin 6Ser His Gln Glu Ser Thr Xaa Gly
Xaa Ser Arg Gly Arg Ser Gly Arg1 5 10 15 Ser Gly
Ser719PRTArtificial SequenceDerived from known cDNA sequences of
human profilaggrin 7Ser His Gln Glu Ser Thr Xaa Gly Arg Ser Xaa Gly
Arg Ser Gly Arg1 5 10 15 Ser Gly Ser819PRTArtificial
SequenceDerived from known cDNA sequences of human profilaggrin
8Ser His Gln Glu Ser Thr Xaa Gly Arg Ser Arg Gly Xaa Ser Gly Arg1 5
10 15 Ser Gly Ser919PRTArtificial SequenceDerived from known cDNA
sequences of human profilaggrin 9Ser His Gln Glu Ser Thr Xaa Gly
Arg Ser Arg Gly Arg Ser Gly Xaa1 5 10 15 Ser Gly
Ser1021PRTArtificial SequenceDerived from known cDNA sequences of
human profilaggrin 10His Gln Cys His Gln Glu Ser Thr Xaa Gly Arg
Ser Arg Gly Arg Cys1 5 10 15 Gly Arg Ser Gly Ser 20
1119PRTArtificial SequenceDerived from known cDNA sequences of
human profilaggrin 11Ser His Gln Glu Ser Thr Arg Gly Arg Ser Arg
Gly Arg Ser Gly Arg1 5 10 15 Ser Gly Ser1219PRTArtificial
SequenceDerived from known cDNA sequences of human profilaggrin
12Ser His Gln Glu Ser Thr Ala Gly Arg Ser Arg Gly Arg Ser Gly Arg1
5 10 15 Ser Gly Ser1319PRTArtificial SequenceDerived from known
cDNA sequences of human profilaggrin 13Ser His Gln Glu Ser Thr Glu
Gly Arg Ser Arg Gly Arg Ser Gly Arg1 5 10 15 Ser Gly
Ser1419PRTArtificial SequenceDerived from known cDNA sequences of
human profilaggrin 14Ser His Gln Glu Ser Thr Gln Gly Arg Ser Arg
Gly Arg Ser Gly Arg1 5 10 15 Ser Gly Ser1514PRTArtificial
SequenceDerived from known cDNA sequences of human profilaggrin
15Thr Gly Pro Ser Thr Arg Gly Arg Gln Gly Ser Xaa His Glu1 5 10
1620PRTArtificial SequenceDerived from known cDNA sequences of
human profilaggrin 16Glu Ser Ser His Gly Trp Thr Gly Pro Ser Thr
Arg Gly Arg Gln Gly1 5 10 15 Ser Arg His Glu 20 17324PRTHomo
sapiensVARIANT(3)...(3)Xaa = Y or I 17Phe Leu Xaa Gln Val Ser Thr
His Glu Gln Ser Xaa Ser Xaa His Gly1 5 10 15 Xaa Xaa Xaa Xaa Ser
Thr Xaa Xaa Arg Gln Gly Ser Xaa His Xaa Gln 20 25 30 Ala Xaa Asp
Ser Ser Xaa His Ser Xaa Ser Gln Xaa Gly Gln Asp Thr 35 40 45 Ile
Xaa Xaa Xaa Pro Gly Xaa Xaa Xaa Xaa Gly Arg Xaa Gly Xaa Xaa 50 55
60 Xaa Glu Gln Xaa Xaa Xaa Xaa Xaa Gly His Xaa Gly Ser Xaa His
Xaa65 70 75 80 His Thr Thr Xaa Gln Gly Arg Ser Asp Ala Ser Xaa Gly
Xaa Ser Gly 85 90 95 Ser Arg Ser Xaa Ser Arg Xaa Thr Xaa Xaa Xaa
Glu Gln Ser Gly Asp 100 105 110 Gly Xaa Arg His Ser Gly Ser Xaa His
His Glu Ala Ser Xaa Xaa Ala 115 120 125 Xaa Ser Ser Xaa His Ser Xaa
Xaa Xaa Gln Gly Xaa Ser Ser Gly Xaa 130 135 140 Arg Xaa Ser Arg Xaa
Xaa Gly Ser Ser Xaa Ser Gln Asp Xaa Asp Ser145 150 155 160 Xaa Xaa
Xaa Xaa Glu Asp Ser Glu Arg Xaa Ser Xaa Ser Ala Ser Arg 165 170 175
Asn His Xaa Gly Ser Xaa Xaa Glu Gln Xaa Arg Asp Xaa Ser Arg His 180
185 190 Pro Xaa Ser His Xaa Glu Asp Xaa Ala Xaa His Xaa Xaa Ser Xaa
Xaa 195 200 205 Xaa Xaa Xaa Xaa Ser Gly Xaa Xaa Xaa Xaa Xaa Xaa Ser
Ser Xaa Xaa 210 215 220 Gln Xaa Ala Ser Ser Xaa Glu Gln Ala Arg Ser
Xaa Xaa Gly Xaa Arg225 230 235 240 His Gly Ser Xaa Xaa Gln Gln Ser
Ala Asp Ser Ser Arg His Xaa Xaa 245 250 255 Xaa Gly Xaa Gly Gln Ala
Ser Xaa Ala Val Xaa Asp Xaa Gly His Arg 260 265 270 Gly Xaa Xaa Gly
Ser Gln Ala Xaa Asp Xaa Glu Gly His Ser Glu Xaa 275 280 285 Ser Asp
Xaa Gln Ser Val Xaa Xaa Xaa Xaa Gln Ala Xaa Xaa Xaa Xaa 290 295 300
Xaa Xaa His Gln Glu Ser Xaa Arg Gly Xaa Ser Xaa Xaa Xaa Ser Gly305
310 315 320 Arg Ser Gly Ser
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