Nothing Special   »   [go: up one dir, main page]

EP1015633A1 - Procede et kit de diagnostic precoce de l'auto-immunite et du lymphome dans le systeme nerveux central - Google Patents

Procede et kit de diagnostic precoce de l'auto-immunite et du lymphome dans le systeme nerveux central

Info

Publication number
EP1015633A1
EP1015633A1 EP98943596A EP98943596A EP1015633A1 EP 1015633 A1 EP1015633 A1 EP 1015633A1 EP 98943596 A EP98943596 A EP 98943596A EP 98943596 A EP98943596 A EP 98943596A EP 1015633 A1 EP1015633 A1 EP 1015633A1
Authority
EP
European Patent Office
Prior art keywords
seq
cell
group
genes
cells
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP98943596A
Other languages
German (de)
English (en)
Inventor
Laboratories Inc. Medelys
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
MEDELYS LAB Inc
Original Assignee
MEDELYS LAB Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from CA 2216595 external-priority patent/CA2216595A1/fr
Priority claimed from CA 2220245 external-priority patent/CA2220245A1/fr
Application filed by MEDELYS LAB Inc filed Critical MEDELYS LAB Inc
Publication of EP1015633A1 publication Critical patent/EP1015633A1/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material

Definitions

  • the invention relates to a series of B- and T- cell clonality kits for early diagnosis and dif ferential diagnosis of central nervous system (CNS ) autoimmunity and lymphoma in non- immunocompromised or immuno- def icient HIV-associated patients as well as the other neurological diseases .
  • CNS central nervous system
  • MS is the most common CNS disease among young adults. More than a third of a million people in the U.S.A. live with MS and about 50,000 Canadians have MS (information from The National MS Society U.S.A. and National MS Society Canada) . The incidence of MS has risen significantly in the past two decades. MS is a chronic inflammatory disease, characterized by damage of the myelin sheaths and infiltration of mononuclear cell (T-cells, B-cells and macrophages) in white matter of the CNS. The symptom of MS
  • the CNS includes the cerebrum, cerebellum, brain stem and spinal cord.
  • the cerebrum is subdivided into frontal, parietal, occipital and temporal lobes.
  • the symptom of MS is a very variable condition and the symptoms depend on which areas of the CNS have been affected.
  • the systems commonly affected include:
  • ⁇ sexuality ⁇ cognitive function Examples: eye trouble, speech problems, paralysis, weakness (fatigue), shaking of hands, loss of coordination and bladder or bowel control, numbness or pricking feelings, staggering (loss of balance), dragging of feet.
  • MS is a chronic inflammatory disease, characterized by damage of the myelin sheaths in the CNS. It is believed that MS is an autoimmune disease, which results from immunity to myelin.
  • Myelin-specific T-cells recognizing myelin basic protein (MBP) and proteolipid protein (PLP) , could be isolated from both MS patients and healthy individuals.
  • MBP myelin basic protein
  • PGP proteolipid protein
  • T-cells derived from postmortem MS brain plaque are antigen nonspecific (Steinman S, Proc Natl Acad Sci USA 93:2253-2256, 1996).
  • the lack of myelin protein reactive T-cells from postmortem MS brain plaque tissue could be a result of apoptosis of antigen specific T-cells in the CNS.
  • B-cells (plasma cells) with high-affinity for its antigen are generated (Nossal GJV, Cell 68:1-2, 1992; Liu YJ, Johson GD, Gordon J, and MacLennan CM Immunol Today 13:17-21, 1992).
  • CSF cerebrospinal fluid
  • the current technologies that are being used for diagnosis of MS are 1) Medical history, 2) Magnetic Resonance Imaging (MRI) , and 3) Lumbar puncture (CSF analyses) .
  • Three basic parameters from above examinations have been used to confirm diagnosis of MS: 1) coming and going pattern of clinical neurological symptoms and signs caused by distinct CNS damage, 2) multifocal lesions in the CNS by MRI examination (12), and 3) IgG oligoclonal bands in the CSF.
  • PCNSL Primary central nervous system lymphoma
  • PCNSL and MS both occur in all age groups, from very young to the elderly. Similar to MS, the symptoms of early stage PCNSL is a very variable condition and the symptoms depend on which areas of the CNS have been invaded by lymphpoma cells. It appears to be similar to other NHL of extranodal sites, which lack organized lymphoid tissue (e.g., bone, ovary, and testis) . Lymphoma has a tendency to relapse in the same or different sites, rather than developing as widespread nodal disease. In this circumstance, it can easily be confused with other neurological conditions, such as MS and encephalomyelitis (DeAngelis LM, J Neurooncol .
  • Magnetic resonance image (MRI) and computer topography (CT) are useful diagnostic tools.
  • PCNSL mass is usually identified by CT or MRI scan (a densely enhancing area) , and surgical biopsy. These methods, however, mainly provide indirect evidence or passively demonstrate lesions in the CNS, which usually appear at the late stage of CNS demyelination or lymphoid malignancy. In addition, the prognosis for patients with lymphoid malignancy in CNS diagnosed by MRI or CT is poor, with a median survival of five months .
  • PCNSL The early stage of PCNSL can begin as an infiltrate process, in which no discrete mass is formed and the white matter lesion is often seen in the CNS.
  • the lesions show the tendency of spontaneous disappearance and relapse in same or different area, being called 'ghost tumor' (Vaquero J, Martinez R, Rossi E, and Lopez R, J Neurosurg. 60:174-6, 1984).
  • Figure 1 demonstrates an example of PCNSL indistinguishable from MS (Kuroda Y., Kawasaki T, Haraoka S, Fujiyama F, Kakigi R et al J " Neurol Sci .
  • One aim of the present invention is to provide a series of B- and T-cell clonality assay kits for an earlier diagnosis of multiple sclerosis, brain lymphoma and other CNS diseases.
  • Another aim of the present invention is to provide a series of B- and T-cell clonality assay kits for diagnosis of demyelinating disease such as multiple sclerosis (MS) .
  • MS multiple sclerosis
  • Another aim of the present invention is to provide a sensitive technology for early diagnosis of primary CNS Non Hodgkin's B-cell lymphoma, as well as the systemic lymphoma CNS metastasis.
  • Another aim of the present invention is to provide a specific technology for differential diagnosis between the clonal expansion or/and polyclonal inflammatory diseases.
  • a method for diagnosis of a neurological disorder in a patient comprising the steps of: a) subjecting a nucleic acid sequence from a CNS sample, CSF; brain and spinal cord biopsy specimens or autopsy (necropsy) of said patient to analyze the clonality of the B-cells and T-cells by the amplification of VH, DH, and JH region
  • step b) analyzing sequences amplified in step a) by detecting amplified sequences of said gene regions wherein said amplified sequences are an indication of increased B-cell or T-cell expression and thereby neurological disorder in said patient.
  • a method for diagnosis of a neurological disorder in a patient comprising the steps of : a) subjecting a nucleic acid sequence from a CNS of said patient to enzyme fragmentation of VH, DH, and JH (CDR3) or VL region genes of B-cells and genes of TCR; and b) analyzing fragments produced in step a) by detecting a signal on a southern blot, wherein said signal is an indication of increased B-cell or T-cell expression and thereby neurological disorder in said patient .
  • kits for determining B- and T-cell clonality for diagnosis of a neurological disorder in a patient comprises at least one upstream primer and at least one downstream primer derived from CDR3 genes of a B-cell immunoglobulin heavy chain, from variable region genes of a B-cell immunoglobulin light chain, or from a gene of a T-cell receptor ⁇ .
  • Fig. 1 illustrates an example of PCNSL indistinguishable from MS
  • Fig. 2 illustrates the autopsy finding of diffuse infiltration of lymphoma cells in the cerebral white matter lesions
  • Fig. 3 illustrates the perivascular lymphocyte infiltration of the plaques from MS autopsy that were used for clonality assay
  • Fig. 4 illustrates the diffuse infiltration of malignant lymphocytes of the plaques from PCNSL autopsy that were used for clonality assay
  • Fig. 5 illustrates an ethidium bromide stained 2% agarose gel of CDR3 PCR products from CSF cells
  • Fig. 6 illustrates an ethidium bromide stained 2% agarose gel of CDR3 PCR products from 45 randomly selected cases (unknown diagnosis) ;
  • Fig. 7 illustrates a predominant CDR3 nucleotide sequence of patients with MS or other neurological diseases; and HIV-positive PCNS non- Hodgkin's B-cell lymphoma;
  • Fig. 8 illustrates an ethidium bromide stained 2% agarose gel of CDR3 RT-PCR products from plaques (lesions) of MS.
  • Figs. 9A to 9D illustrate the sequences of the VH gene from a dominant clone of CSF B-cells from patients with MS.
  • PCNS Central Nervous System
  • B- or T-cell clonality assay kits provide a high sensitive, specific, non-invasive technique for the earlier diagnosis and differential diagnosis of neurological diseases. It has been used to distinguish B-cell clonal expanded autoimmune diseases, such as MS, and neoplasm of primary central nervous system non-Hodgkin v s lymphoma and B-cell polyclonal expanded inflammatory diseases, such as encephalomyelitis and other neurological diseases.
  • the B- and T-cell clonality assay kits use the modification of procedure based on amplifying genes of VH, DH and JH region (CDR3) for VH; VL and JL region for VL of B-cells; and genes for various V and J segments of TCR using PCR technique.
  • the procedure involves the instruction of the sample preparation, extraction of the DNA and a set of the primers for amplifying the genes of framework regions (FRs) and J regions of heavy and light chains of immunoglobulin and TCR.
  • the clonality of B- and T-cells can be analyzed following an ethidium bromide staining of agarose gel of PCR products. CONTENTS For fresh cells or frozen tissues:
  • Upstream primers FR2 5 ' -TGG [A/G] TCCG [C/A] CAG [G/C] C [T/C] [T/C] CNNGG-3 '
  • VK variable region genes of B-cell immunoglobulin light chain
  • Upstream primers V ⁇ l 5 ' GACATCC (A/G) G (T/A) TGACCCAGTCTCC (A/T) TC-3 '
  • Upstream primers LII 5' -ATGGCCTGGGCTCTGCTGCTC-3 ' (SEQ ID NO: 14) LIII 5 ' -ATGGCATGGATCCCTCTCTT-3 ' (SEQ ID NO: 15) BL2 5 ' -ATGACCTGCTCCCCTCTCCT-3 ' (SEQ ID NO: 16) 4A 5' -ATGGCCTGGACTCCTCTCTTTCTG-3 ' (SEQ ID NO: 17) LBV 5 ' -ATGGCCTGGGCTCCACTACT-3 ' (SEQ ID NO: 18) Downstream primers :
  • JGT3 5'AGTTACTATGAGC(T/C)TAGTCCC-3 ' (SEQ ID NO:30)
  • JGT4 5'TGTAATGATAAGCTTTGTTCC-3 ' (SEQ ID NO:31)
  • Immunoglobulin heavy chain region genes assay kit has been used to amplify the entire Ig VH genes of B-cells from the neurological disease. After subsequently sequencing PCR products, the differentiation pathway of pathogenic B-cells in the CNS and their origin can be analyzed by studying the antigen-driven somatic mutation on the genes Ig VH region. CONTENTS For fresh cells or frozen tissues:
  • Primer of oligonucleotides specific for each patients-derived dominant VDJ region is designed for analyzing the antigen-driven selected somatic hypermutation on the Ig VH genes derived from dominant clonal B-cells.
  • the B-cell clonality kits have been developed for Ig VH gene amplification and identification of the clonality of B-cells from CSF of the patients with multiple sclerosis and other neurological diseases. These kits can also be used for analysis of surgical, biopsy and autopsy specimens of MS plaques and PCNSL. Patients
  • the CSF cell samples were obtained from each of twelve (12) MS patients and fifteen (15) patients with other neurological diseases (Table 1) as well as 47 random selected cases (unknown diagnosis) . Seven of 12 MS patients were considered as at early stage of disease, CSF was collected from initial onset of clinical symptom. MS diagnosis was confirmed on the second-episode. MS patients including 11 females and 1 male with clinically or laboratory- supported definite MS diagnosis with a mean age of 34 11 years. The average duration of disease was 2 ⁇ 0.4 years. The mean expanded disability status score (EDSS) was 3.2 ⁇ 1.3. All MS patients were categorized as having relapsing-remitting disease.
  • Oligoclonal bands of all CSF samples were analyzed for presence of oligoclonal immunoglobulin bands by isoelectric focussing or agarose electrophoresis .
  • MRI scan demonstrated that the lesion (s) varying from no lesion to multi lesions and areas were affected from spinal cord to brain individually (Table 1) .
  • the control patients were divided into inflammatory (I) (3 of 15 cases) : 2 patients with subacute illnesses diagnosed as acute disseminated encephalomyelitis and 1 with a remote history of Herpes Zoster encephalitis.
  • ONDs amyotrophic lateral sclerosis
  • pseudotumer a progressive neurotrophic lateral sclerosis
  • spinocerebellar degeneration spinal cord infarct
  • myopathy headache and neuropathy.
  • ONDs other neurological diseases
  • DNA samples were also collected from paraffin embedded specimens from plaques of four (4) patients with MS (Fig 3) and two (2) patients with PCNSL (Fig 4) . Extraction of RNA or DNA from CSF
  • Protocol 1 Preparation of samples
  • RNA of the frozen CSF cell pellets and Specimens are extracted using RNAeasyTM Kit (Qiagen Inc. Chatsworth, Ca) .
  • Table 1 summarizes the clinical and laboratory data for the patients under study.
  • OB oligoclonal bands ;
  • Cell number the total numbers of white blood cells in the specimen (2 -8 ml) ; + multiple plaques ;
  • SL spinal lesion;
  • NL normal ;
  • ADEM acute disseminated encephalomyelitis ;
  • ALS amyotrophic lateral sclerosis ;
  • HZEM Herpes Zoster encephalitis .
  • UN unknown .
  • the f irst step is to use B-cell clonality assay kit for screening the CDR3 region of Ig gene rearrangement derived from B-cells in the CNS of patients with neurological diseases.
  • the method of the present invention is simple, convenient, specific, highly sensitive and needs only a small sample of CSF with mild or no side effects and biopsy and autopsy specimens. It provides a clear-cut information to support a diagnosis or a differential diagnosis.
  • the method has provided us an important tool to study the pathogenesis of B-cells and T-cells in CNS.
  • the results from B-cells derived from CSF and the plaques of MS brain using said kits showed the high frequency of clonal B-cell evolution in the CNS these patients (Figs. 3 and 4) .
  • the utility of this B- and T-cell clonality assay kits has been demonstrated with high sensitivity in patients with early multiple sclerosis; and other neurological diseases, as well as CNS lymphoid malignancy (Figs. 5 and 6) who have no MRI change or oligoclonal bands (Table 1) .
  • the PCR was performed in a final volume of 50 ⁇ l reaction buffer that contain 2.5 units of recombinant Taq Polymerase, 50 mM/L Tris-HCL, PH 9.0 at 25°C: 20 mM/L (NH4) 2 S0 4 ; 3.0 mM/L MgCl 2 , 0.1 mM/L each of deoxyadenosine triphosphate, deoxycytidine triphosphate, deoxyguanosine triphosphate, and thymidine triphosphate, 25 pmol of each primer, and 50 to 200 ng of DNA.
  • reaction buffer that contain 2.5 units of recombinant Taq Polymerase, 50 mM/L Tris-HCL, PH 9.0 at 25°C: 20 mM/L (NH4) 2 S0 4 ; 3.0 mM/L MgCl 2 , 0.1 mM/L each of deoxyadenosine triphosphate, deoxycytidine triphosphate, deoxygua
  • each sample was amplified with upstream primers FR2 or FR3 , respectivelly, and downstream primer LJH, for 40 cycles of 50 seconds at o o o
  • the second step in accordance with one embodiment of the present invention is sequencing of PCR products for further identification of clonality of the CSF B-cells.
  • the PCR products were purified by electrophoresis and recovered from the gel by treatment with gelase (CalbioChem, San Diego, CA. ) .
  • gelase CalbioChem, San Diego, CA.
  • the recovered DNA was ligated into the pGEMTM T vector (Promega, La Jolla, CA.), which was used to transfect Escherichia coli DH5a.
  • Six (6) to eight (8) white colonies were picked up at random and grown overnight in 3 ml of LB (Luria-Bertani) medium.
  • the double- stranded DNA template from the colonies was sequenced by the method of Sanger et al .
  • the CDR3 gene sequences of the CSF B-cells were analyzed using the FASTA program, the sequences of expressed D segments were compared with those of the published germline D and DIR segments. The most common sequence from each patient was considered as being derived from the dominantly clonal B-cells. Based on the complete sequence, oligo-nucleotide primer specific for each patient -derived dominant VH, DH and JH region were designed.
  • the third step is to use Immunoglobulin heavy chain variable region genes assay kit for investigating the role of the antigen-driven selection in the dominant clonal expansion of the CSF B-cells.
  • a semi -nested PCR was performed:
  • the first round, five 250 ng of DNA was amplified with 6 VH family primers and LJH for 40 cycles of 94°C for 1 min, 55°C for 1 min and 72°C for 1 min.
  • the second round, 2 to 5 ⁇ l aliquot of the initial PCR product was reamplified with primers 6 VH family primers and the primer of oligonucleotides specific from each patients-derived dominant, for 35 cycles of 94 C for 1 min; 59 C for 1 min and 72 °C for 1 min. Aliquots of the final reaction product were analyzed by electrophoresis in a 1% agarose gel (Sigma) containing ethidium bromide. The amplified VH product was sequenced as described above. All sequences were confirmed by sequencing in both orientations . Assignment of mutations
  • CDR3 VDJ region
  • PCR products from all MS and I-OND patients, 3 NI-OND as well as 8 of 45 randomly selected (unknown diagnosis) controls showed high density bands suggesting a possible limited clonality pattern.
  • PCR products from 9 NI-OND cases (cases 4 to 12) and 37 of 45 unknown diagnosis cases showed only a faint smear or invisible bands, reflecting size differences in the VH, DH, and JH regions in a polyclonal population of B-cells.
  • Monoclonal PCR band also demonstrated from the plaques from 3 of 4 MS patients and 2 PCNSL patients (Fig. 7) .
  • Fig. 7 shows the dominant CDR3 sequences of CSF B- cells. Dominant sequences from each patients are grouped and subdivided into VH, N, D, N, and JH regions. Names of the germline D and JH genes with maximum homology to the segments used in the VDJ joining are shown in parentheses in the appropriate rows .
  • CSF B-cells from patients with MS and OND as well as plaques which showed a possible or probable clonal band of PCR products on ethidium- bromide stained agarose gels was determined by sequencing PCR products of CDR3 genes.
  • CSF B-cells from 10 of 12 MS patients, including the both cases of first clinical onset or multi-episodes as well as MS plaques had dominated clonal or dual clonal expansion.
  • Four (4) of sixteen (16) controls were found to have dominant B-cell clones (Fig. 8) .
  • the case from inflammatory ONDs group showed that CSF B-cell clonal expansion has history of systemic cancer.
  • case 1 has been diagnosed as a primary central nervous non-Hodgkin' s lymphoma, in this study, that serves as a control of monoclonal B-cells, • case 2 suffered from headache band for more than 6 months, using current diagnostic technologies no diagnosis was made, was found B-cell clonal expansion occurring in the CSF; case 3 with initially diagnosed as spinal cord infarct, was found B-cell clonal expansion in the CSF. Similar symptom has been reported in the patients who had systemic lymphoma CNS metastasis. Using our novel technology, 8 in 45 randomly selected cases (unknown diagnosis) were identified as having high-density clonal band.
  • PCR products illustrated high density of clonal band and sequence analysis demonstrate a dominant B-cell clonal expansion.
  • the polyclonal and monoclonal expansion was detected from the surgical biopsy specimens of encephalomyelitis (Fig. 7) .
  • the B-cell clonal expansion can be identified at very early stage of MS as well as in the late stage of autopsy MS plaques.
  • the invented technology permits examination of a wider array of patients then can be performed on surgical biopsy (invasive diagnosis) and autopsy tissue, and is particularly suited for patients at relatively early stages of disease. Somatic hypermutation
  • the average R:S ratio in the framework regions was 0.7 (expected ⁇ 2.9) .
  • the average R:S ratio in CDRl and CDR2 was 4.0.
  • a higher CDR R:S mutation ratio reflects the positive selective pressure of an antigen on those gene products which come into close contact with antigen, while a lower FR R:S mutation ratio reflects the negative pressure for mutant selection applied to structural components that need to be conserved. This pattern is consistent with the notion of an antigen- driven selection of antibodies with high-affinity antigen-binding sites.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Wood Science & Technology (AREA)
  • Analytical Chemistry (AREA)
  • Zoology (AREA)
  • Genetics & Genomics (AREA)
  • Engineering & Computer Science (AREA)
  • Pathology (AREA)
  • Immunology (AREA)
  • Microbiology (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention a trait à la mise au point d'une série de kits de détermination de la clonalité des lymphocytes B et T. Les données exposées dans cette demande montrent l'importance des kits de détermination de la clonalité des lymphocytes B et T dans le diagnostic précoce et le diagnostic différentiel de la sclérose en plaques, le lymphome primaire du système nerveux central et autres troubles neurologiques. En comparaison avec des technologies actuelles de diagnostic (cytologie, bandes oligoclonales, tomodensitomètre, imagerie par résonance magnétique et biopsie chirurgicale), la technologie de l'invention s'avère être positive comme technique très sensible, spécifique, non invasive et économique, et peut être appliquée à différents échantillons (cellules du fluide cérébrospinal, biopsie chirurgicale et autopsie). L'utilisation de cette technologie dans l'étude de la sclérose en plaques a montré que le clonage des lymphocytes B s'étend principalement chez les patients atteints de sclérose en plaques, ces lymphocytes ayant subi une mutation somatique induite par un antigène dans les centres germinaux des tissus lymphoïdes périphériques. On a ainsi découvert un important mécanisme pathogène dans l'évolution de la démyélinisation dans le système nerveux central du patient atteint de sclérose en plaques. De plus, cette technologie permet d'établir une banque d'ARN spécifique clonale à partir des cellules pathogènes du système nerveux central des patients, ce qui est important pour comprendre ensuite le rôle du ou des antigènes dans le déclenchement de l'extension clonale des lymphocytes B, et vis à vis du développement de la stratégie thérapeutique spécifique de l'antigène.
EP98943596A 1997-09-19 1998-09-17 Procede et kit de diagnostic precoce de l'auto-immunite et du lymphome dans le systeme nerveux central Withdrawn EP1015633A1 (fr)

Applications Claiming Priority (5)

Application Number Priority Date Filing Date Title
CA 2216595 CA2216595A1 (fr) 1997-09-19 1997-09-19 Nouvelle methode en trois etapes pour un diagnostic precoce de l'autoimmunite et du lymphome a cellules b du systeme nerveux central primaire
CA2216595 1997-09-19
CA 2220245 CA2220245A1 (fr) 1997-11-04 1997-11-04 Nouvelle methode de diagnostique precoce de l'autoimmunite et du lymphome du systeme nerveux central
CA2220245 1997-11-04
PCT/CA1998/000873 WO1999015696A1 (fr) 1997-09-19 1998-09-17 Procede et kit de diagnostic precoce de l'auto-immunite et du lymphome dans le systeme nerveux central

Publications (1)

Publication Number Publication Date
EP1015633A1 true EP1015633A1 (fr) 2000-07-05

Family

ID=25679649

Family Applications (1)

Application Number Title Priority Date Filing Date
EP98943596A Withdrawn EP1015633A1 (fr) 1997-09-19 1998-09-17 Procede et kit de diagnostic precoce de l'auto-immunite et du lymphome dans le systeme nerveux central

Country Status (3)

Country Link
EP (1) EP1015633A1 (fr)
AU (1) AU9148498A (fr)
WO (1) WO1999015696A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107345240A (zh) * 2016-05-12 2017-11-14 眭维国 T细胞抗原受体β链CDR3的处理方法

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
ATE421998T1 (de) * 2003-12-15 2009-02-15 Pasteur Institut Ermittlung des repertoires von b-lymphozyten populationen
US20070160994A1 (en) * 2003-12-15 2007-07-12 Institut Pasteur Repertoire determination of a lymphocyte b population

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1990004648A1 (fr) * 1988-10-20 1990-05-03 Alexander Alan Morley Methode pour le diagnostic de la monoclonalite dans la leucemie et le lymphome
US5298396A (en) * 1989-11-15 1994-03-29 National Jewish Center For Immunology And Respiratory Medicine Method for identifying T cells disease involved in autoimmune disease
ES2225816T5 (es) * 1990-02-01 2014-08-21 Siemens Healthcare Diagnostics Products Gmbh Producción y utilización de bancos de genes de anticuerpos humanos ("bibliotecas de anticuerpos humanos")
ATE140733T1 (de) * 1990-05-01 1996-08-15 Univ Leland Stanford Junior Stoffgemische zur anwendung in einem verfahren zur behandlung eines an multipler sklerose leidenden menschen
JPH07503132A (ja) * 1991-12-17 1995-04-06 ジェンファーム　インターナショナル，インコーポレイティド 異種抗体を産生することができるトランスジェニック非ヒト動物

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO9915696A1 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107345240A (zh) * 2016-05-12 2017-11-14 眭维国 T细胞抗原受体β链CDR3的处理方法

Also Published As

Publication number Publication date
AU9148498A (en) 1999-04-12
WO1999015696A1 (fr) 1999-04-01

Similar Documents

Publication Publication Date Title
US20210310076A1 (en) Methods of monitoring conditions by sequence analysis
EP2663864B1 (fr) Procédé d'évaluation de la biodiversité et son utilisation
WO2010011894A1 (fr) Signature de codon de vh4 pour sclérose multiple
US20170088895A1 (en) Immunorepertoire normality assessment method and its use
JP2024036563A (ja) 筋痛性脳脊髄炎／慢性疲労症候群（ｍｅ／ｃｆｓ）のバイオマーカー
Haubold et al. B‐lymphocyte and plasma cell clonal expansion in monosymptomatic optic neuritis cerebrospinal fluid
Mitsunaga et al. Deep characterization of the human antibody response to natural infection using longitudinal immune repertoire sequencing
Muraro et al. Rapid identification of local T cell expansion in inflammatory organ diseases by flow cytometric T cell receptor Vβ analysis
EP1015633A1 (fr) Procede et kit de diagnostic precoce de l'auto-immunite et du lymphome dans le systeme nerveux central
US11041208B2 (en) Urine biomarkers for detecting graft rejection
RU2715633C2 (ru) Способ идентификации в иммунном репертуаре cdr3 участков, ассоциированных с заболеваниями
CA2303978A1 (fr) Procede et kit de diagnostic precoce de l'auto-immunite et du lymphome dans le systeme nerveux central
CA2220245A1 (fr) Nouvelle methode de diagnostique precoce de l'autoimmunite et du lymphome du systeme nerveux central
ES2415583B1 (es) Métodos y composiciones para la determinación de la diversidad del repertorio de linfocitos t de un individuo
CA2216595A1 (fr) Nouvelle methode en trois etapes pour un diagnostic precoce de l'autoimmunite et du lymphome a cellules b du systeme nerveux central primaire
WO2016127113A1 (fr) Procédés pour diagnostiquer la sclérose en plaques à l'aide de gènes d'anticorps vh4

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 20000406

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): AT BE CH DE DK FR GB IE IT LI NL SE

RIN1 Information on inventor provided before grant (corrected)

Inventor name: QIN, YUFEN

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN

18D Application deemed to be withdrawn

Effective date: 20020403