WO2004076614A2 - Sequences d'acide nucleique humaines issues de carcinomes de la prostate - Google Patents

Sequences d'acide nucleique humaines issues de carcinomes de la prostate Download PDF

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Publication number
WO2004076614A2
WO2004076614A2 PCT/DE2004/000433 DE2004000433W WO2004076614A2 WO 2004076614 A2 WO2004076614 A2 WO 2004076614A2 DE 2004000433 W DE2004000433 W DE 2004000433W WO 2004076614 A2 WO2004076614 A2 WO 2004076614A2
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WIPO (PCT)
Prior art keywords
protein
nucleic acid
peptide
tissue
substance
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PCT/DE2004/000433
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German (de)
English (en)
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WO2004076614A3 (fr
Inventor
Bernd Hinzmann
Edgar Dahl
André ROSENTHAL
Thomas Specht
Armin Schmitt
Georg Beckmann
Thomas Brümmendorf
Henrik Kinnemann
Stefan Röpcke
Klaus Hermann
Li Xinzhong
Christian Pilarsky
Eike Staub
Original Assignee
Bernd Hinzmann
Edgar Dahl
Rosenthal Andre
Thomas Specht
Armin Schmitt
Georg Beckmann
Bruemmendorf Thomas
Henrik Kinnemann
Roepcke Stefan
Klaus Hermann
Li Xinzhong
Christian Pilarsky
Eike Staub
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Publication date
Priority claimed from DE10309985A external-priority patent/DE10309985A1/de
Priority claimed from DE2003122134 external-priority patent/DE10322134A1/de
Application filed by Bernd Hinzmann, Edgar Dahl, Rosenthal Andre, Thomas Specht, Armin Schmitt, Georg Beckmann, Bruemmendorf Thomas, Henrik Kinnemann, Roepcke Stefan, Klaus Hermann, Li Xinzhong, Christian Pilarsky, Eike Staub filed Critical Bernd Hinzmann
Publication of WO2004076614A2 publication Critical patent/WO2004076614A2/fr
Publication of WO2004076614A3 publication Critical patent/WO2004076614A3/fr

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • C07K14/4701—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals not used
    • C07K14/4748—Tumour specific antigens; Tumour rejection antigen precursors [TRAP], e.g. MAGE

Definitions

  • the invention relates to new human nucleic acid sequences from prostate carcinomas and to proteins or peptides coded thereby, the use of sequences derived therefrom for screening for substances which bind to them, and the use of substances which bind to such nucleic acid sequences and proteins or peptides for diagnosis and / or Treatment of tumor diseases, especially prostate cancer.
  • Prostate cancer is a disease that appears with increasing incidence with increasing age and requires new therapies to combat it.
  • Patients with prostate tumors are currently divided into two groups.
  • the first group includes patients with an operable tumor (approximately 90% of all patients). In these, the tumor is removed as completely as possible by surgery (radical prostatectomy). Prostate removal has significant medical risks and adverse effects on a patient's quality of life, such as incontinence and impotence.
  • the second group (approximately 10% of all patients) are patients with an inoperable tumor. These cannot be treated curatively by surgery because they already have lymph node metastases. These patients are currently being treated palliatively with nti-androgen or radiation therapy.
  • the anti-androgen therapy which is based on the blocking of hormone effects is very often ineffective after a few years, since the tumor becomes hormone-independent, ie continues to grow without hormone effects and forms metastases. Likewise, not all tumor cells can be removed by radiation therapy.
  • the cancer phenomenon is often associated with the over- or under-expression of a large number of genes in the degenerate cells.
  • the identification of tumor-relevant genes is therefore an important starting point for the development of new therapies against prostate cancer (Welsch et al., Cancer Res 61 (16): 5974-5978 (2001)).
  • DNA microarrays were used to search for tumor-related candidate genes in prostate cancer.
  • the analyzed tumor and normal tissue samples can be obtained by microdissection. Using the microdissection, it is possible to precisely, i.e. at the level of individual cell groups. Comparative studies have shown that the use of microdissection can identify differentially expressed genes that are not found in a gene expression study of whole tissue (Ernst et al., Am J Pathol 160 (6): 2169-2180 ( 2002)).
  • the invention is based on the technical problem of pharmaceutical compositions for diagnosis and / or Specify treatment of prostate cancer diseases and means of finding them.
  • the invention first teaches a nucleic acid containing or consisting of one of the disclosed nucleic acid sequences and a peptide or protein containing an amino acid sequence encoded by one of the disclosed nucleic acid sequences or consisting thereof or containing one or consisting of one of the disclosed amino acid sequences.
  • Nucleic acids or proteins or peptides according to the invention can be produced using conventional molecular biological methods.
  • the invention further relates to various uses of the new nucleic acids or peptides or protein, as well as (same) uses of already known nucleic acids. These are:
  • a nucleic acid according to the invention and / or a peptide or protein according to the invention for the detection of prostate cancer or for the detection of a risk of prostate cancer, wherein a prostate tissue sample is examined for over-transcription of the nucleic acid or for over-expression of the protein.
  • a detector substance that preferably binds to the nucleic acid or to the protein or peptide, preferably containing a reporter group, can be used, whereby said nucleic acid and / or said protein or peptide binds to the detector substance semi-quantitatively or is detected quantitatively.
  • the level of expression can also be measured by amplification, for example quantitative PCR.
  • nucleic acid according to the invention or a protein or peptide according to the invention for screening for substances which bind to it, in particular prospective active substances for inhibiting said nucleic acid or said protein or peptide, or prospective detector substances, with a prospective substance or a mixture of such prospective substances with said nucleic acid or said protein or peptide is contacted, whereby binding events are determined with a binding assay, and a binding prospective substance, optionally after deconvolution, is selected.
  • a substance used in the context of the invention can be selected from the group consisting of: a) antisense oligonucleotides, siRNA, and ribozymes against a nucleic acid according to claim 1, b) binding to a peptide or protein according to claim 2, in particular identified according to claim 5 , organic molecule with a molecular weight below
  • aptamer against a protein or peptide according to claim 2 in particular identified according to claim 5, d) (onoclonal) antibody, in particular human or humanized antibody against a protein or peptide according to claim 2, e) anti-idiotypic non-human (monoclonal) Antibodies generated by means of an antibody of subgroup d) and f) above substances derivatized with a reporter group, a cell toxin of an immunostimulating
  • ribozyme for example, a hammerhead ribozyme can be used as the ribozyme.
  • the ribozyme interface is selected with the proviso that the expression of the protein is either prevented by the activity of the ribozyme, 'or an inactive form or an inactive fragment is expressed of the protein.
  • Both can be determined, for example, by contacting this cell system with one or more ribozyme modeled for defined interfaces in a cell system in which a protein according to the invention is expressed at a defined level and determining the expression level or the biological activity of the expressed protein , This is then compared with a negative sample or the results without contact and ribozymes are selected which lead to lower expression or activity. The same can be done in the case of siRNA or antisense nucleic acids.
  • binding substances can be determined by determining the biological activity of the Proteins in a cell system with and without contacting and comparison of the results obtained. Those substances that lead to reduced biological activity are then selected for therapeutic purposes. It is also possible for the biological activity to be determined instead of the binding in the context of a screening method according to the invention; then a validation in the above sense was carried out at the same time as the screening.
  • Biological activity can be determined, for example, by determining natural association partners of the protein and examining their occurrence and form (eg monomer / dimer). Substances arising further downstream in a metabolic cascade can also be used as indicators; these can be identified, for example, by previously analyzing cell components for the cell expressing the protein and comparing them with the same cells, in which, however, the expression has been genetically deleted.
  • Suitable aptamers (c) can easily be identified, for example, by means of the well-known SELEX method, the protein according to the invention being used as the target.
  • Antibodies (d), in particular monoclonal antibodies, can be obtained in a customary manner by immunizing a non-human mammal with a protein according to the invention, a nucleic acid according to the invention (eg cDNA), a cell constitutively expressing a protein according to the invention (cancer cell or, for example, a cell transfected with a nucleic acid according to the invention , such as COS or .NIH3T3), or by means of recombinantly produced protein or peptide according to the invention, for example in E. coli or eukaryotic cells, for example insect cells.
  • Monoclonal antibodies can be obtained by customary selection and stabilization of hybridoma cells.
  • the phage display technology can also be used to generate the antibodies.
  • anti-idiotypic antibodies these can be generated by using an antibody according to the invention, which does not necessarily have to influence the biological activity of the protein according to the invention, in a non-human mammal, a second anti-idiotypic (monoclonal) antibody is generated.
  • this anti-idiotypic antibody When applied to human cells, this anti-idiotypic antibody then simulates the human immune system and becomes an image of the target molecule. recognized as a foreign epitope due to its non-humanized form. Human beings therefore naturally form antibodies against the anti-idiotypic antibody and thus also against the protein or against the protein-expressing cells. This variant of the invention can only be used for therapeutic purposes.
  • the invention further relates to a method for diagnosing a prostate cancer, wherein a pharmaceutical composition according to the invention in the embodiment with a reporter group is applied to the tissue to be examined in vivo or in vitro, the tissue to be examined then being subjected to a detection method step which is sensitive is for the reporter group, and in the case of detection of a defined minimum value of the reporter group in the tissue the tissue is qualified as containing tumor cells, and a method for the treatment of a prostate cancer disease, wherein a pharmaceutical composition according to the invention is administered to a patient in a physiologically effective dose.
  • the invention is based on the knowledge that genes or gene products according to the invention are differentially expressed in prostate tumor tissue, i.e. in prostate tumor tissue the expression is higher or lower, in particular higher, compared to normal cells of the same tissue. On the one hand, this allows, in particular these new genes or
  • a sample from a tissue which is identified as tumor tissue by other methods in advance of treatment with a pharmaceutical composition according to the invention, and to test the tissue sample for expression or overexpression of the gene or gene product according to the invention investigate.
  • a detector substance according to the invention can be used for diagnosis in vivo to test for dependence on the gene or gene product. If expression or overexpression of the gene or gene product with respect to normal tissue of the same type is found, then the use of the pharmaceutical composition according to the invention is indicated.
  • the substance which binds to the gene or the gene product also has a cytotoxic and / or carries immunostimulating component. This ultimately leads to the fact that almost exclusively tumor cells are killed, either by cytotoxicity or by attack by the stimulated immune system, while normal cells are practically completely preserved in the tissue.
  • the binding substance itself does not have to have an inhibitory effect on the gene or gene product, since the binding substance then only has to function as a marker which carries the components to target tumor cells. If a cytotoxic and / or immunostimulating component is used, it can be particularly recommended if the pharmaceutical composition is prepared for local application in tissue containing tumor cells, for example for injection.
  • sequences disclosed and / or claimed in the description are known per se or are parts of previously known sequences
  • sequences disclosed, insofar as they correspond to previously known sequences are the subject of the invention in that they are used only in accordance with the uses described. Revealed and / or claimed sequences, which are parts of previously known sequences, can be so by means of one or more disclaimers in claims be delimited that the previously known sequences are not included.
  • a sequence comprises all human isoforms, known or new, based on nucleic acids or amino acids. These terms also include the short sequences disclosed in the context of this description, which come from isoforms, for example immunization sequences. Also included are homologs, the homology being at least 80%, preferably more than 90%, most preferably more than 95% (calculated using the MEGALIGN, DNASTAR LASERGENE program, in the version current at the time of registration). In the case of nucleic acid sequences, complementary or allelic variants and silent mutations are also included.
  • sequences are included which only represent partial sequences of the explicitly disclosed sequences, for example one or more exons, or complementary sequences thereto, with the proviso that, in the case of nucleic acids, these partial sequences have a length sufficient for hybridization with a nucleic acid according to the invention, at least 50 or 150 bases, up to 1700 bases and more, and in the case of the proteins or peptides bind with at least the same affinity to a protein- or peptide-specific target molecule.
  • all nucleic acids hybridizing with nucleic acids according to the invention are included, namely those which are under stringent conditions (5 ° C. to 25 ° C.
  • the invention also includes expression cassettes, ie one or more of the nucleic acid sequences according to the invention with at least one control or regulatory sequence.
  • Such an expression cassette can also comprise a sequence for a known protein, a fusion protein being formed in the course of the translation from a known protein and a protein or peptide according to the invention.
  • Antisense sequences to the above nucleic acid sequences are also included.
  • RNA as well as correlating DNA and vice versa are included, as are genomic DNA as well as correlated cDNA and vice versa.
  • nucleic acids or protein or peptides include, in addition to the full lengths of the disclosed sequences (see also the preceding paragraph), also partial sequences thereof, with a minimum length of 12 nucleotides, preferably 30 to 90 nucleotides, in the case the nucleic acids and a minimum length of 4 amino acids, preferably 10 to 30 amino acids, in the case of the peptides or proteins.
  • detection and / or treatment of prostate cancer also include the detection and / or treatment of metastases from primary tumors in other tissues.
  • treatment also includes prophylaxis.
  • an inhibitor is a compound or substance which either inhibits the formation of the protein or peptide according to the invention or reduces the activity of the protein or peptide formed, based on the latter Activity in the absence of the inhibitor.
  • an inhibitor can on the one hand be a substance that interferes with the protein or peptide cascade.
  • an inhibitor can be a substance that binds to the protein or peptide formed, in such a way that further physiological interactions with endogenous substances are at least reduced.
  • Mimicry molecules encompassed by the invention are compounds which emulate the variable region, in particular the binding region of an antibody, and bind to a target molecule in the same place as the underlying antibody.
  • antibody includes polyclonal antibodies, monoclonal antibodies, non-human, human and humanized antibodies, as well as phage display antibodies, but also chimeric antibodies and anti-idiotypic antibodies as well as specific fragments of the light and / or heavy chain of the variable region underlying antibodies of the above type.
  • the production or production of such antibodies with predetermined immunogens is well known to the average person skilled in the art and need not be explained in more detail.
  • the term antibody also includes bispecific antibodies. Bispecific antibodies combine a defined immune cell activity with a specific tumor cell recognition, whereby tumor cells are killed. A bispecific antibody binds on the one hand to a trigger molecule of the immune effector cell (eg CD3, CD16, CD64) and on the other hand to antigens of the tumor target cell.
  • a trigger molecule of the immune effector cell eg CD3, CD16, CD64
  • Counter ions for ionic compounds are, for example, Na + , K + , Li + or cyclohexylammonium.
  • Suitable solid or liquid pharmaceutical preparation forms are, for example, granules, powders, dragees, tablets, (micro) capsules, suppositories, syrups, juices, suspensions, emulsions, drops or injectable solutions (IV, IP, IM) and preparations with a protracted active ingredient -Release, in the production of which conventional auxiliaries such as carriers, explosives, binders, coatings, swelling agents, lubricants or lubricants, flavorings, sweeteners and solubilizers are used.
  • a pharmaceutical composition according to the invention can be produced by mixing at least one substance used according to the invention in a defined dose with a pharmaceutically suitable and physiologically compatible carrier and, if appropriate, other suitable active ingredients, additives or auxiliaries with a defined substance dose and preparing it for the desired dosage form ,
  • Tumor cells express one. Protein differential if normal cells of the same tissue type do not or only slightly express it. Tumor cells overexpress a protein specifically or differentially if the protein is in the Compared to normal cells of the same tissue is expressed at least in twice the amount.
  • Cytotoxic components or groups are compounds which directly or indirectly induce apoptosis or lead to necrosis or at least have an inhibitory effect on growth.
  • radioisotopes for example 188Re, 213Bi, 99mTc, 90Y, 131J, 177Lu
  • groups or compounds can in particular cytostatics Examples of these are: alkylating agents (for example mechlorethamine, ifosfamide, chlorambucil, cyclophosphamide, melphalan, alkylsulfonates, busulphan, nitrosoureas, carmustine, lomustine, semustine, triazenes, dacarbazine), antimetabolites (for example Folic acid antagonists, methotrexate, pyrimidine analogs, fluorouracil, fluorodesoxyuridine, cytarabin, gemcitabine, purine analogs, mercaptopurine), mitotic inhibitors (e.g.
  • antibiotics eg dactinomycin, daunorubicin, idarubicin, anthracycline, bleomycin, L-asparaginase
  • platinum complex compounds eg cisplatin
  • hormones and related compounds eg adrenal gland steroids, aminogluthetimide, progestogens, estrogens, androgens, anti-estrogens, tamoxifen, steroid analogues, flutamide).
  • an immunostimulating component is usually a protein or an effective component thereof, which stimulates cells of the immune system.
  • cytokines such as M-CSF, GM-CSF, G-CSF, interferons such as IFN-alpha, beta, gamma, interleukins such as IL-1 to -16 (except -8), human LIF , Chemokines such as Rantes, MCAF, MIP-1-alpha, -beta, NAP-1 and IL-8.
  • a reporter group is an atom, molecule or a compound which, in conjunction with an assay placed thereon, enables the detection of the reporter group and the compound or substance thus connected to the reporter group.
  • reporter groups and associated detection methods are: 32P labeling and intensity measurement using phosphoimager. Many other examples are known to the person skilled in the art and do not need to be listed in detail.
  • a substance that binds to target molecules can be a substance that binds a target protein or to a target RNA.
  • Definitions expanded in the context of the above definition in relation to the narrow sense of the word also include the specific terms in the narrow sense of the word.
  • Figure 1 Chip analysis for the differential expression of CD24 in prostate tumor tissue, analyzed on 54 normal / tumor tissue samples.
  • FIG. 1 Immunohistochemistry with a CD24 antibody (mouse anti human CD24, clone 24C02), CD24 is expressed much more strongly in primary prostate cancer than in benign tissue.
  • FIG. 3 Immunohistochemistry with a CD24 antibody (mouse anti human CD24, clone 24C02), CD24 is expressed much more strongly in lymph node metastases than in primary prostate carcinoma,
  • Figure 4 Table with information on the sequences according to the invention.
  • RNA is isolated, amplified and labeled from samples from Example 1.
  • the RNA obtained in this way is applied to a gene chip which contains a large number of different oligonucleotides, one (or more, for control purposes) in each case being representative of a defined gene, i.e. has a characteristic partial sequence from it.
  • Both qualitative and quantitative information is obtained as to whether a normal and / or tumor sample in question expresses a gene in question, and also in the tumor / normal ratio. In cases in which a gene is expressed more highly in tumor tissue than in the correlated normal tissue, there is differential expression, i.e. the gene is upregulated in the tumor tissue. On the other hand, if the gene is less expressed in tumor tissue, there is down-regulation. It was found that the sequences according to the invention are differentially upregulated in the tumor tissue. The results can be found in detail in Table 1.
  • Example 3 Examination of expression or overexpression using quantitative PCR.
  • a poly-A + RNA preparation is carried out using a modified protocol according to the poly-a-tract 1000 kit (Amersham, Freiburg, Germany).
  • Tissue samples ' for example, but not necessarily obtained according to example 1, are slowly thawed on ice, crushed and mixed with 300 ⁇ l dilution buffer containing 1% ⁇ -mercaptoethanol and biotinylated oligo-dT primer, and for 5 min. heated to 70 ° C. The samples are then kept for 5 min. kept at 20 ° C and then at 20000g for 10 min. centrifuged. 120 ⁇ l of washed streptavidin-coupled paramagnetic particle (SA-PMP) are added to the supernatant and it was stirred at 20 ° C. for 5 min. incubated. The mRNA was then isolated by magnetic separation. After three washing steps with 0.5x SSC solution, the mRNA is diluted in nuclease-free water, evaporated under vacuum and immediately processed in cDNA.
  • the cDNA synthesis then takes place.
  • the mRNA obtained from 2 is dissolved in 10 ⁇ l of nuclease-free water.
  • I ⁇ l T7-dT24- (GGCCAG) primer (100 pmol / ⁇ l) is added and it was kept at 70 ° C. for 5 min. heated. Then the sample was placed on ice and 4 ⁇ l 5x first strand buffer (Invitrogen), 2 ⁇ l DTT (0.1M), l ⁇ l dNTP's (lOmM), and 14U anti-RNAse (Ambion) were added, followed by incubation for 2 min , at 37 ° C. Then 1 ⁇ l of Superscript II reverse transcriptase (Invitrogen) are added, followed by an incubation for 1 h at 37 ° C.
  • the second strand synthesis and DNA purification then take place.
  • 91 ⁇ l water, 30 ⁇ l 5x second strand buffers, 3 ⁇ l dNTP's (10mm), 10U E. coli DNA ligase, 40U DNA polymerase I and 2U RNAse H (all from Invitrogen) added and the mixture is incubated at 16 ° C. for 2 h.
  • 10U T4 DNA polymerase (Invitrogen) are added and a further 5 min. incubated.
  • the reaction is stopped by adding 10 ⁇ l of 0.5 mM EDTA.
  • the DNA is purified in accordance with the GFX PCR DNA and Gel Band Purification Kit (A ersham).
  • the second round of in vitro transcription is then carried out.
  • the second round of reinforcements is carried out with only minor deviations from the first round.
  • the synthesis of the first strand is done with random hex primer (250ng / ⁇ l). After incubation for 60 min. the cRNA-cDNA hybrid for 20 min. incubated with 2U RNase H, followed by a 2 minute inactivation step at 37 ° C.
  • the first strand is synthesized using the cRNA from the preceding stage.
  • Ing cDNA are used for amplification with 2.5 ⁇ l lOx SYBR®Green PCR buffer, 3 ⁇ l magnesium chloride (25mM), 2 ⁇ l dNTP's (with dUTP; 12.5mM) and 0, 625U Ampli Taq Gold in a reaction volume of 25 ⁇ l.
  • the reaction is carried out in a GeneAmp 5700 Sequence Detection System (Applied Biosystems, Rothstadt,
  • the conditions are: 2 min. 50 ° C, 10 min. 95 ° C, 15 s 95 ° C, 1 min. 60 ° C, the last two phases in 40 cycles.
  • the appropriate forward or reverse primers are used.
  • the evaluation is carried out using the ⁇ Ct method according to the manufacturer's instructions.
  • the Ct value of beta actin was measured at a limit of 0.1.
  • the Ct value of the beta actin is subtracted from the Ct value of the gene under investigation. In the case of tumor tissues, this normalized Ct value is related or normalized to the normal tissues, whereby the ⁇ Ct is obtained.
  • this value is used as a power to base 2, a relative size of the over- or under-expression in tumor tissue compared to the normal tissue of the same patient is obtained. As a result, it can be determined whether a specific tumor tissue of a particular patient is sensitive to a treatment according to the invention. This method can also be used to determine whether unclassified tissue should be classified as containing tumor cells. In the latter case, a comparison is made with reference values or classified normal tissue from the same patient or from other people.
  • Example 4 Differential expression measured using the gene chip technology, using the example of CD24.
  • Example 2 The result of experiments according to Example 2 is shown in FIG. 1 using CD24 as an example. It can be seen that CD24 is up-regulated in a significant number of samples from 54 patients. Analogous results were obtained for the further sequences according to the invention, which are not shown for the sake of clarity.
  • Example 5 Detection of an overexpressed gene using antibodies.
  • tumor cells by an antibody directed against a protein according to the invention in vivo (mouse model) is described.
  • an antibody according to the invention is labeled with a marker molecule (e.g. radioisotope).
  • a marker molecule e.g. radioisotope
  • Human cells transfected with a gene according to the invention are transplanted into NMRI nude mice. After a defined period of time, for example 30 days, the labeled antibody is injected into the mice. The control animals are treated with an irrelevant antibody. A few hours after the antibody application, the animals are sacrificed and tissue sections are made from all organs. These sections are examined for the presence of labeled antibody.
  • the antibodies can be polyclonal antibodies against human protein, conjugated to a carrier protein, raised in rabbits and affinity-purified with the specific immobilized peptides.
  • Suitable immunization peptides are formed, for example, from partial sequences of a protein according to the invention.
  • Cells which are transfected with cDNA of the gene or partial sequences thereof, such as, for example, COS cells or NIH3T3 cells, can also be used as immunogens. Tumor cells that express the protein endogenously are also suitable.
  • recombinantly produced protein or partial sequences thereof, which are expressed in producer cells such as E. coli or eukaryotic cells such as insect cells can also be used for the immunization.
  • producer cells such as E. coli or eukaryotic cells such as insect cells
  • Example 6 Immunohistochemical detection of tumor cells.
  • Tissue is isolated from a patient with cancer or suspected to be cancer and prepared as paraffin or frozen sections. These sections are examined with an antibody directed against a protein according to the invention for the overexpression of the protein in tumor cells.
  • the immunohistological examination with the antibody shows higher expression of the protein in the tumor cells compared to surrounding normal tissue.
  • the investigation is carried out in detail by incubation with the antibody as the primary antibody, a biotinylated secondary anti-rabbit antibody and a streptavidin-coupled horseradish peroxidase.
  • the coloring is carried out with DASS as a chromogenic substrate (brown coloring).
  • the counterstaining is done with hemalaun solution (blue staining).
  • Malignant and non-malignant cells can be distinguished, the malignant cells being strongly stained, i.e. have a high content of protein according to the invention, while the non-malignant cells are only moderately stained.
  • FIGS. 2 and 3 show exemplary results using CD24.
  • a CD24 antibody mouse, anti human CD24, clone 24C02
  • left benign tissue was used. It can be seen that benign atrophy shows strong staining in the apical membrane.
  • primary prostate cancer was used.
  • FIG. 3 shows a comparison of primary tumors (left) with lymph node metastases (right) using the same antibodies. Comparatively higher expression can be seen in the lymph node metastases.
  • a monoclonal antibody Abi is generated in a manner which is able to specifically recognize the protein and bind to it. It is immaterial whether a functional domain or another accessible area is recognized.
  • a second anti-idiotypic non-humanized, for example mouse, monoclonal antibody aABl is generated in the same way, which is suitable for the manufacture of a pharmaceutical composition for the treatment of prostate tumors.
  • the function of the antibody aABl is based on the fact that it simulates the human immune system an image of the (human) protein antigen, the immune system recognizing the antibody aABl as foreign to the body due to its lack of humanization. The human body consequently forms its own antibodies, which are directed against aABl and thus also against the human protein or tumor cell expressing it.

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Abstract

L'invention concerne de nouvelles séquences d'acide nucléique humaines issues de carcinomes de la prostate, les protéines et peptides ainsi codés et leur utilisation en liaison avec le diagnostic et/ou le traitement du cancer de la prostate.
PCT/DE2004/000433 2003-02-27 2004-02-22 Sequences d'acide nucleique humaines issues de carcinomes de la prostate WO2004076614A2 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
DE10309985A DE10309985A1 (de) 2003-02-27 2003-02-27 Humane Nukleinsäuresequenzen aus Prostatakarzinomen
DE10309985.9 2003-02-27
DE10322134.4 2003-05-14
DE2003122134 DE10322134A1 (de) 2003-05-14 2003-05-14 Humane Nukleinsäuresequenzen aus Prostatakarzinomen

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WO2004076614A3 WO2004076614A3 (fr) 2005-08-18

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Cited By (14)

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WO2006090389A2 (fr) * 2005-02-24 2006-08-31 Compugen Ltd. Nouveaux marqueurs diagnostiques, en particulier pour l'imagerie in vivo, et dosage et procedes d'utilisation associes
WO2005103712A3 (fr) * 2004-04-20 2006-12-14 Sphingotec Gmbh Utilisation de precurseurs de tachykinines et/ou de fragments de celles-ci dans un diagnostic medical
WO2007074341A1 (fr) * 2005-12-28 2007-07-05 Randox Laboratories Ltd Detection du cancer de l'oesophage
WO2005113816A3 (fr) * 2004-05-07 2007-08-09 Jackson H M Found Military Med Methodes de diagnostic ou de traitement du cancer de la prostate au moyen du gene erg, seul ou combine a d'autres genes surexprimes ou sous-exprimes dans le cancer de la prostate
WO2009027527A2 (fr) * 2007-08-30 2009-03-05 Santaris Pharma A/S Composes antagonistes d'arn permettant la modulation de fabp4/ap2
US8148093B2 (en) 2003-08-15 2012-04-03 Diadexus, Inc. Pro108 antibody compositions and methods of use and use of Pro108 to assess cancer risk
US8173434B2 (en) 2006-04-04 2012-05-08 Diadexus, Inc. PCan065 antibody compositions and methods of use
US8288511B2 (en) 2004-12-03 2012-10-16 Medtrain Technologies, Llc Nuclear targeting sequence
US8377443B2 (en) 2010-08-27 2013-02-19 Gilead Biologics, Inc. Antibodies to matrix metalloproteinase 9
CN104583422A (zh) * 2012-06-27 2015-04-29 博格有限责任公司 标志物在诊断和治疗前列腺癌中的用途
EP3192521A1 (fr) * 2007-10-25 2017-07-19 Toray Industries, Inc. Inducteur de réponse immunitaire
US9732156B2 (en) 2012-02-29 2017-08-15 Gilead Biologics, Inc. Methods of treating rheumatoid arthritis using antibodies to matrix metalloproteinase 9
IT201600127428A1 (it) * 2016-12-16 2018-06-16 Cusani Alberto Bartorelli Nuova proteina ricombinante uk 114 in forma stabile polimerica per uso nella terapia, nella diagnostica e nella prevenzione di neoplasie maligne
US10539566B2 (en) 2014-12-08 2020-01-21 Berg Llc Use of markers including filamin A in the diagnosis and treatment of prostate cancer

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995029245A2 (fr) * 1994-04-25 1995-11-02 Association Pour Le Developpement De La Recherche En Genetique Moleculaire (A De Re Ge M) Proteines du complexe btf2 et leurs applications

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995029245A2 (fr) * 1994-04-25 1995-11-02 Association Pour Le Developpement De La Recherche En Genetique Moleculaire (A De Re Ge M) Proteines du complexe btf2 et leurs applications

Non-Patent Citations (9)

* Cited by examiner, † Cited by third party
Title
BONKHOFF H ET AL: "DIFFERENTIAL EXPRESSION OF THE PS2 PROTEIN IN THE HUMAN PROSTATE AND PROSTATE CANCER: ASSOCIATION WITH PREMALIGNANT CHANGES AND NEUROENDOCRINE DIFFERENTIATION" HUMAN PATHOLOGY, SAUNDERS, PHILADELPHIA, PA, US, Bd. 26, Nr. 8, August 1995 (1995-08), Seiten 824-828, XP009010130 ISSN: 0046-8177 *
DATABASE EMBL [Online] 16. November 2001 (2001-11-16), "TFIIH basal transcription factor complex p44 subunit(BTF2-p44)" XP002304612 gefunden im EBI accession no. Q13888 Database accession no. Q13888 *
DATABASE EMBL [Online] 6. Juni 1996 (1996-06-06), EGLY: "Transcription factor BTF2 complex p44 subunit" XP002304611 gefunden im EBI accession no. AAR88225 Database accession no. AAR88225 *
FIGUEROA J A ET AL: "DIFFERENTIAL EXPRESSION OF INSULIN-LIKE GROWTH FACTOR BINDING PROTEINS IN HIGH VERSUS LOW GLEASON SCORE PROSTATE CANCER" JOURNAL OF UROLOGY, BALTIMORE, MD, US, Bd. 159, Nr. 4, April 1998 (1998-04), Seiten 1379-1383, XP000971791 ISSN: 0022-5347 *
HUMBERT S ET AL: "P44 AND P34 SUBUNITS OF THE BTF2/TFIIH TRANSCRIPTION FACTOR HAVE HOMOLOGIES WITH SSL1 A YEAST PROTEIN INVOLVED IN DNA REPAIR" EMBO JOURNAL, OXFORD UNIVERSITY PRESS, SURREY, GB, Bd. 13, Nr. 10, 1994, Seiten 2393-2398, XP002047712 ISSN: 0261-4189 *
LEE DONG KUN ET AL: "Androgen receptor interacts with the positive elongation factor P-TEFb and enhances the efficiency of transcriptional elongation" JOURNAL OF BIOLOGICAL CHEMISTRY, Bd. 276, Nr. 13, 30. M{rz 2001 (2001-03-30), Seiten 9978-9984, XP002304609 ISSN: 0021-9258 *
LEE DONG KUN ET AL: "From androgen receptor to the general transcription factor TFIIH. Identification of cdk activating kinase (CAK) as an androgen receptor NH2-terminal associated coactivator" JOURNAL OF BIOLOGICAL CHEMISTRY, Bd. 275, Nr. 13, 31. M{rz 2000 (2000-03-31), Seiten 9308-9313, XP002304610 ISSN: 0021-9258 *
NG A Y K ET AL: "DIFFERENTIAL EXPRESSION OF HSIM2 IN PROSTATE CANCER" PROCEEDINGS OF THE ANNUAL MEETING OF THE AMERICAN ASSOCIATION FOR CANCER RESEARCH, NEW YORK, NY, US, Bd. 40, M{rz 1999 (1999-03), Seite 232, XP000986062 ISSN: 0197-016X *
THOMAS R ET AL: "Differential expression of placental bone morphogenetic protein (PLAB) gene during prostate cancer progression to bone" PROCEEDINGS OF THE 91ST ANNUAL MEETING OF THE AMERICAN ASSOCIATION FOR CANCER RESEARCH. SAN FRANCISCO, CA, APRIL 1 - 5, 2000, PROCEEDINGS OF THE ANNUAL MEETING OF THE AMERICAN ASSOCIATION FOR CANCER RESEARCH, PHILADELPHIA, PA : AACR, US, Bd. VOL. 41, 1. April 2000 (2000-04-01), Seite 587, XP002180666 *

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US8148093B2 (en) 2003-08-15 2012-04-03 Diadexus, Inc. Pro108 antibody compositions and methods of use and use of Pro108 to assess cancer risk
WO2005103712A3 (fr) * 2004-04-20 2006-12-14 Sphingotec Gmbh Utilisation de precurseurs de tachykinines et/ou de fragments de celles-ci dans un diagnostic medical
US9188591B2 (en) 2004-04-20 2015-11-17 Sphingotec Gmbh Use of precursors of tachykinins and/or their fragments in medical diagnostic
US7838245B2 (en) 2004-04-20 2010-11-23 Sphingotec Gmbh Use of precursors of tachykinins and/or their fragments in medical diagnostic
WO2005113816A3 (fr) * 2004-05-07 2007-08-09 Jackson H M Found Military Med Methodes de diagnostic ou de traitement du cancer de la prostate au moyen du gene erg, seul ou combine a d'autres genes surexprimes ou sous-exprimes dans le cancer de la prostate
US9347101B2 (en) 2004-05-07 2016-05-24 The Henry M. Jackson Foundation For The Advancement Of Military Medicine, Inc. Methods of diagnosing or treating prostate cancer using the ERG gene, alone or in combination with other over or under expressed genes in prostate cancer
US9868993B2 (en) 2004-05-07 2018-01-16 The Henry M. Jackson Foundation For The Advancement Of Military Medicine, Inc. Methods of diagnosing or treating prostate cancer using the ERG gene, alone or in combination with other over or under expressed genes in prostate cancer
US10066268B2 (en) 2004-05-07 2018-09-04 The Henry M. Jackson Foundation For The Advancement Of Military Medicine, Inc. Methods of diagnosing or treating prostate cancer using the ERG gene, alone or in combination with other over or under expressed genes in prostate cancer
US11236395B2 (en) 2004-05-07 2022-02-01 The Henry M. Jackson Foundation For The Advancement Of Military Medicine, Inc. Methods of diagnosing or treating prostate cancer using the ERG gene, alone or in combination with other over or under expressed genes in prostate cancer
US8288511B2 (en) 2004-12-03 2012-10-16 Medtrain Technologies, Llc Nuclear targeting sequence
WO2006090389A3 (fr) * 2005-02-24 2007-11-29 Compugen Ltd Nouveaux marqueurs diagnostiques, en particulier pour l'imagerie in vivo, et dosage et procedes d'utilisation associes
US7741433B2 (en) 2005-02-24 2010-06-22 Compugen Ltd. Diagnostic markers, especially for in vivo imaging and assays and methods of use thereof
US7488813B2 (en) 2005-02-24 2009-02-10 Compugen, Ltd. Diagnostic markers, especially for in vivo imaging, and assays and methods of use thereof
WO2006090389A2 (fr) * 2005-02-24 2006-08-31 Compugen Ltd. Nouveaux marqueurs diagnostiques, en particulier pour l'imagerie in vivo, et dosage et procedes d'utilisation associes
WO2007074341A1 (fr) * 2005-12-28 2007-07-05 Randox Laboratories Ltd Detection du cancer de l'oesophage
US8173434B2 (en) 2006-04-04 2012-05-08 Diadexus, Inc. PCan065 antibody compositions and methods of use
WO2009027527A3 (fr) * 2007-08-30 2009-07-02 Santaris Pharma As Composes antagonistes d'arn permettant la modulation de fabp4/ap2
WO2009027527A2 (fr) * 2007-08-30 2009-03-05 Santaris Pharma A/S Composes antagonistes d'arn permettant la modulation de fabp4/ap2
EP3192521A1 (fr) * 2007-10-25 2017-07-19 Toray Industries, Inc. Inducteur de réponse immunitaire
US9260532B2 (en) 2010-08-27 2016-02-16 Gilead Biologics, Inc. Antibodies to matrix metalloproteinase 9
US9120863B2 (en) 2010-08-27 2015-09-01 Gilead Sciences, Inc. Nucleic acids encoding antibodies to matrix metalloproteinase 9
US8501916B2 (en) 2010-08-27 2013-08-06 Gilead Biologics, Inc. Antibodies to matrix metalloproteinase 9
US8377443B2 (en) 2010-08-27 2013-02-19 Gilead Biologics, Inc. Antibodies to matrix metalloproteinase 9
US9732156B2 (en) 2012-02-29 2017-08-15 Gilead Biologics, Inc. Methods of treating rheumatoid arthritis using antibodies to matrix metalloproteinase 9
EP2867375A4 (fr) * 2012-06-27 2016-06-01 Berg Llc Utilisation de marqueurs dans le diagnostic et le traitement du cancer de la prostate
US9797905B2 (en) 2012-06-27 2017-10-24 Berg Llc Use of markers in the diagnosis and treatment of prostate cancer
CN104583422A (zh) * 2012-06-27 2015-04-29 博格有限责任公司 标志物在诊断和治疗前列腺癌中的用途
US10539566B2 (en) 2014-12-08 2020-01-21 Berg Llc Use of markers including filamin A in the diagnosis and treatment of prostate cancer
IT201600127428A1 (it) * 2016-12-16 2018-06-16 Cusani Alberto Bartorelli Nuova proteina ricombinante uk 114 in forma stabile polimerica per uso nella terapia, nella diagnostica e nella prevenzione di neoplasie maligne
WO2018109096A1 (fr) * 2016-12-16 2018-06-21 Alberto Bartorelli Cusani Nouvelle protéine recombinante uk 114 sous forme de polymère stable destinée à être utilisée dans le traitement, le diagnostic et la prévention de tumeurs malignes solides et systémiques
US11352398B2 (en) 2016-12-16 2022-06-07 Alberto Bartorelli Cusani Recombinant protein UK 114 in stable polymer form for use in the treatment, diagnosis and prevention of malignant solid and systemic tumours

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