FR2875512A1 - DIAGNOSTIC BIOPUCE OF THE METASTATIC OR LOCALIZED CHARACTER OF A MAMMARY TUMOR, METHOD AND KIT OF USE - Google Patents

Abstract

The subject of the present invention is a solid support coated with a set of representative fragments of genes whose expression profile for all of these genes in a subject presenting a mammary tumor allows the diagnosis or the prognosis of the metastatic or localized nature. of said tumor, as well as a kit comprising such a support; The subject of the invention is also a process for the in vitro diagnosis or prognosis of the metastatic or localized nature of a mammary tumor, where appropriate also making it possible to characterize whether the cells of said mammary tumor or not express estrogen receptors, said process implementing the use of the solid support.

Description

The present invention relates to a solid support coated with a set of

  representative fragments of genes whose expression profile for all of these genes in a subject having a mammary tumor allows the diagnosis or prognosis of the metastatic or localized nature of said tumor, as well as a kit comprising such a support; the invention

  Another subject of the invention is an in vitro diagnostic or prognostic method for the metastatic or localized character of a mammary tumor, which may also be used to characterize whether or not the cells of said mammary tumor express estrogen receptors, said method using the use of the solid support.

  Cancer can be defined very broadly as a disease linked to the proliferation and uncontrolled spread of abnormal cells of the body: these abnormal cells, called malignant cells, proliferate anarchically from a primitive focus, which can relapse locally after removal, extend to adjacent tissues and spread at a distance (metastases).

  Cancer affects about 10 million people worldwide. More than 4.4 million cases come from Asia. Europe has 2.8 million cases, North America 1.4 million and Africa 627 000 cases. Cancer is one of the leading causes of death in developed countries; in Europe and the United States, it is estimated that one in five people will die of cancer. The incidence of cancer over the last 100 years seems to be increasing, but this observation must be balanced by the fact that the probability of developing cancer increases with age, and that overall the proportion of the elderly population is also increasing .

  2 2875512 In women, breast cancer and the most common: there are thus 43,000 new cases and 11,000 deaths per year in France. Breast cancer is the leading cause of cancer death in women. It should be noted that 99% of breast cancers occurring in women, and 1% in men.

  In Europe, the establishment of means of screening and prevention allows a diagnosis of breast tumors at an early stage, which can then define a therapeutic orientation. An important prognostic factor is to determine whether metastases are present in the lymph nodes; we then speak of ganglionic invasion; in the absence of ganglionic invasion, we speak of localized mammary tumor.

  One of the methods hitherto commonly used to characterize ganglionic invasion is to perform axillary dissection, which consists of removing axillary lymph nodes (armpits) and making sections of these ganglia in hematoxylin and eosin staining. However, this curative technique is currently being questioned: axillary dissection is responsible for most post-surgical functional symptoms of breast cancer, ranging from 10% for lymphedema (large arm syndrome) to more than 80% of discomfort and pain in the mobilization of the upper limb. In addition, in Western countries, the proportion of tumors lacking ganglionic invasion (N-) approaches 80%, which means that nearly 4 in 5 women undergo unnecessary axillary dissection.

  The classification of mammary tumors in the N- (node-free) or N + (ganglionic) stage is all the more accurate as the number of ganglions removed is higher, thus leading to results with a certain risk of error. Standard anatomopathological examination for ganglia of axillary dissection (staining of sections) could be supplemented by immunohistochemistry techniques based on a specific immunological reaction and morphological control of immunoreactive cells. However, the identification of micrometastases by performing serial sections and immunohistochemistry is not feasible in current practice, for reasons of time and cost on all ganglia taken in an axillary dissection.

  An alternative to performing axillary lymph node dissection is Positron Emission Tomography (PET), a nuclear medicine medical imaging technique in which a positron-emitting radioactive marker is injected in vivo (FDG-PET). for (18F) FluoroDeoxyGlucose-PET). This technique, very sensitive, is for the moment too expensive and not very accessible.

  Another alternative to axillary dissection is the study of the sentinel lymph node. The latter is defined as the first ganglion that receives the lymphatic flow of the primary tumor. The sentinel lymph node technique is therefore an effective method for assessing axillary lymph node status and avoiding axillary dissection in patients with small breast cancer without clinical lymph node involvement. In fact, an unaffected sentinel node excludes in theory a metastatic invasion of the rest of the armpit and thus renders the axillary dissection useless. Although investigations on a single ganglion may be at the origin of axillary staging, the sentinel node technique is not yet standardized and false negative rates are evaluated in the literature from 0 to 15%. This rate can be reduced by the use of molecular biology techniques such as RT-PCR (Reverse Transcription Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction (RT-PCR)). But the study in molecular biology is carried out from frozen tissue, which implies that the pathologist must subtract part of the ganglion from the standard histological study.

  Thus, it is essential to have another diagnostic tool easy to implement, inexpensive and reliable to assess the lymph node status in breast cancer.

  This objective has been achieved by the inventors named in this application. The latter have been able to develop a DNA microarray that defines the lymph node status of patients who have developed a breast tumor. Breast tumors are heterogeneous because they result from the accumulation and combinations of multiple molecular alterations. Therapeutic and diagnostic indications are based on classic prognostic factors (histological and clinical) that do not reveal this heterogeneity. Indeed, this most often corresponds to the study of a single gene or gene product at a time. A global and detailed molecular characterization, obtained using the biochip of the invention, thus makes it possible to refine the diagnosis, the therapeutic indications and therefore the survival of the patients.

  The probes (or representative fragments) deposited on the biochip correspond to genes whose expression levels are modified in mammary tumors, thus allowing the study of parameters such as hormonal receptivity, ganglionic invasion, BRCA1 status. BRCA2, overexpression of erbb2, cell proliferation, cell types present in the tumor. The inventors have been able to identify a set of genes whose overexpression is characteristic of a ganglionic invasion and therefore of a metastatic tumor (N + tumor), and another set of genes whose overexpression is characteristic of the absence of ganglionic invasion. and therefore a localized tumor (N- tumor). These genes are listed in Tables A1, A2 and A3.

  In the present application, the determination of the ganglionic invasion, and therefore of the metastatic or localized nature of a tumor, firstly requires the characterization of the expression of the estrogen receptors by the mammary tumor cells. A mammary tumor is thus classified ER + when it expresses the estrogen receptors, and ER- otherwise.

  Table A1 gathers the discriminating genes for lymph node invasion overexpressed in mammary tumors that express estrogen receptors (ER + tumors).

  Table A2 gathers the ganglionic invasion discriminant genes overexpressed in both mammary tumors that express estrogen receptors (ER + tumors) and those that do not express it (ER- tumors).

  Table A3 collects the discriminating genes for lymph node invasion overexpressed in mammary tumors that do not express estrogen receptors (ER- tumors).

  Thus, the subject of the present invention is a solid support coated with a set of representative fragments of genes whose expression profile for all of these genes in a subject presenting a mammary tumor allows the diagnosis or the prognosis of the metastatic nature. or localized of said tumor, characterized in that said set comprises and / or consists of at least 2, preferably 3 or 4 fragments, each of 2, 3 or 4 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3.

  The expression profile, which is obtained using the solid support according to the present invention, contains the information relating to the expression of all the genes of which a representative fragment has been deposited on said solid support.

  By the expression representative fragment of a gene of Table A1, A2, A3, B or Table C is meant in the present application any coding fragment (mRNA, cDNA) of one of these genes which is long enough for hybridize specifically, under conditions suitable for hybridization, with an mRNA or cDNA fragment present in the test nucleic acid sample or with a fragment present in the reference nucleic acid sample, and thus makes it possible to identify said fragment present in the test or reference nucleic acid sample as being a constitutive fragment of said given gene, said gene therefore being present in the test nucleic acid sample or in the sample of reference nucleic acid; when such a representative fragment of a gene of Table A1, A2, A3, B or of Table C is sequenced, the identified sequence makes it possible to conclude without ambiguity that said fragment belongs to said given gene of Table A1, A2, A3, B or Table C. The term probe is also used in this application to denote a representative fragment.

  Preferably, the representative fragment of a gene according to the invention comprises and / or consists of at least 15, preferably 20, 25, 30, 35, 40, 45, 47, 48, 49 and particularly preferably 50 nucleotides identical to those of the fragment of the test or reference nucleic acid constituting said given gene, said gene being therefore present in the test nucleic acid and in the reference nucleic acid.

  The term "appropriate conditions for hybridization" in the present invention refers to conditions of high stringency that permit hybridization when sequences complementary to at least 80, 82, 85, 87, 89, 90, 92, 95 , 97 or 99% are able to hybridize specifically. Conditions of high stringency are widely described in the literature, for example in Sambrook et al., (1989, Molecular cloning, A laboratory Manual, Cold Spring Harbor), Maniatis et al., 1982 (Molecular cloning, A laboratory Manual, Cold Spring Harbor Lab.CSH, NY USA or one of his recent reissues and in Ausubel et al., Eds. (1995) "Current Protocols in Molecular Biology, Chapter 2" (Greene Publishing and Wiley-Interscience, NY), and these conditions can be adapted by those skilled in the art depending on the size of the nucleotide fragments, according to the appropriate teachings known.

  For example, suitable hybridization or washing buffers (Corning) or the following hybridization conditions will be used: (1) prehybridization at 42 ° C. for 3 hours in phosphate buffer (20 mM, pH 7.5) containing 5 × SSC (1X SSC corresponds to a solution 0.15M NaCl + 0. 015M sodium citrate), 50% folinamide, 7% sodium dodecyl sulfate (SDS), 10X Denhardt's solution, 5% dextran sulfate and 1% salmon sperm DNA; (2) hybridization proper for 20 hours at a temperature dependent on the size of the probe (for example 55 C for a probe of size about 50 nucleotides) followed by 2 washes of 20 minutes at 20 C in 2X SSC + 2 % SDS, 1 wash for 20 minutes at 20 C in 0.1X SSC + 0.1% SDS. The last wash is performed in 0.1X SSC + 0.1% SDS for 30 minutes at 60 ° C. for a probe of about 50 nucleotides in size.

  The solid support used for carrying out the present invention, also indifferently referred to in the present application (bio) chip or (bio) microarray, can generally be any miniaturized support of glass, silicon or polymer which may be coated with nucleotide sequences of known sequence, in particular the representative fragments of genes according to the present invention deposited in an ordered arrangement, and whose function is to recognize, in a mixture, their complementary nucleotide sequences. As an example of a solid support, there may be mentioned, but not limited to, membranes such as high density nylon membranes (eg Performa, Genetix, New Milton, UK); micro-amino-silane microarrays are preferably used, especially from Corning (ultraGAPTM, NY). The surface of the support is treated to form a dense and regular network of microsurfaces on which the representative fragments are fixed; the location of each of them is precisely known. Such solid supports are well known to those skilled in the art, who will be able to choose the type of support to be used and the appropriate conditions for a given hybridization test.

  The subject having a mammary tumor is preferably a mammal, and particularly preferably a human. The present invention is particularly suitable for screening for metastatic breast tumors in women, but it can also be used in cases of breast cancer in humans.

  Preferably, the solid support according to the present invention is characterized in that the set comprises and / or consists of at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17 , 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45 or preferably 50 fragments, each of 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45 or 50 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3.

  According to another preference, the solid support according to the present invention is characterized in that the set comprises and / or consists of at least 60, 65, 70, 75, 80, 85, 90, 95 or preferably 100 fragments, each 60, 65, 70, 75, 80, 85, 90, 95 or 100 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3.

  According to yet another preference, the solid support according to the present invention is characterized in that the set comprises and / or consists of at least 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155 , 160 or 165, preferably 150 fragments, each of 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160 or 165 fragments being representative of a distinct gene selected from the genes of the Tables Al, A2 and A3.

  According to a preferred embodiment, the solid support according to the present invention is characterized in that the set comprises and / or consists of 72 fragments, each of the 72 fragments being representative of a distinct gene chosen from the genes of Tables A1 and A2.

  Such a support, since it comprises and / or consists of 72 fragments, each of the 72 fragments being representative of a distinct gene chosen from the genes of Tables A1 and A2 (and thus overexpressed in ER + tumors - cf. supra), allows the detection of ganglion invasion in the case of ER + tumors.

  Preferably, the solid support according to the present invention is characterized in that the 72 fragments are chosen from the following fragments: a) the fragments of sequence SEQ ID N 1 to 55 for Table A1, and the fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, or b) the fragments having at least 70% identity with the fragments as defined in a), or c) the fragments comprising fragments as defined in a) or b), or d) fragments whose sequences are complementary to the sequences of the fragments as defined in a), b) or c).

  One (SEQ ID N 71) or the other (SEQ ID N 72) of the two representative fragments of the TUBB gene can be used indifferently in the present invention, just as the two fragments can be used simultaneously ( SEQ ID N 71 and SEQ ID N 72). Thus, preferably, the solid support according to the present invention is characterized in that the set comprises and / or consists of 73 fragments, of which 71 fragments are representative of a distinct gene chosen from the genes of Tables A1 and A2, and 2 fragments (SEQ ID N 71 and 72) are representative of the TUBB gene.

  The percentage of identity between two nucleic acid sequences is intended to denote a percentage of identical nucleotides between the two sequences to be compared, obtained after their better alignment, this percentage being purely statistical and the differences between the two sequences being randomly distributed. and over their entire length. Optimal sequence alignment for comparison can be achieved using mathematical algorithms. In a preferred, nonlimiting manner, mention may be made of the various algorithms mentioned in the document Fundamentals of Database Search (review) Stephen F., Altschul, Bioinformatics: A Trends Guide, 1998: 5: 7-9, in particular Karlin's algorithms and Altschul (1990) Proc. Natl. Acad. Sci. USA 872264, modified in Karlin and Altschul (1993) Proc. Natl. Acad. Sci. USA 90: 5873-5877. In particular, the programs may be used: -BLAST, in particular BLASTN, BLAST2 (Tatusova et al., 1999, "Blast 2 sequences - a new tool for comparing proteins and nucleotide sequences", FEMS Microbiol Lett., 174: 247-250), gapped BLAST (Altschul et al., 1997, Nucleic Acids Res., (17): 3389-3402), -FASTA (Altschul SF et al., 1990, J. Mol Biol., 403-410), -Clustal W (Thompson, JD et al., 1994, Nucleic Acids Res 22, 4673-80), BESTFIT.

  The BLAST algorithm is described in detail on the NCBI website (http: ww.ncbi. Nih.gov).

  The expressions nucleic acid, nucleic or nucleic acid sequence, polynucleotide, oligonucleotide, polynucleotide sequence, nucleotide sequence, may be used indifferently in the present application to designate a specific sequence of nucleotides that can correspond to both DNA than its transcription products (RNA). These nucleic acids are isolated from their natural environment.

  According to another preferred embodiment, the solid support according to the present invention is characterized in that the set comprises and / or consists of 111 fragments, each of the 111 fragments being representative of a distinct gene chosen from the genes of Tables A2. and A3.

  Such a support, because it comprises and / or is constituted by 111 fragments, each of the 111 fragments being representative of a distinct gene selected from the genes of Tables A2 and A3 (and therefore overexpressed in ER tumors - cf. supra) can be used to detect ganglion invasion in the case of ER- tumors.

  Preferably, the solid support according to the present invention is characterized in that the 111 fragments are chosen from the following fragments: a) fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, and the fragments of sequence SEQ ID N 74 to 167 for Table A3, or b) the fragments having at least 70% identity with the fragments as defined in a), or c) the fragments comprising fragments as defined in a) or b), or d) fragments whose sequences are complementary to the sequences of the fragments as defined in a), b) or c).

  Again, one (SEQ ID N 71) or the other (SEQ ID N 72) of the two representative fragments of the TUBB gene may be used indifferently in the present invention, just as the two fragments may be used simultaneous manner (SEQ ID N 71 and SEQ ID N 72). Thus, preferably, the solid support according to the present invention is characterized in that the set comprises and / or consists of 112 fragments, of which 110 fragments are representative of a distinct gene selected from the genes of Tables A2 and A3, and 2 fragments are representative of the TUBB gene (SEQ ID N 71 and 72).

  According to yet another preferred embodiment, the solid support according to the present invention is characterized in that the set comprises and / or consists of 166 fragments, each of the 166 fragments being representative of a distinct gene chosen from the genes of the Tables A1, A2 and A3.

  Such a support, since it comprises and / or consists of 166 fragments, each of the 166 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3 (see above), makes it possible to detect ganglionic invasion in both ER- and ER + tumors. Of course, before this screening, it is necessary to determine whether or not the tumors express the estrogen receptor (respectively ER + or ER- tumors); then, if the tumors are ER +, we will focus only on the expression profile obtained for all or part of the genes listed in Tables A1 and A2, and if the tumors are ER-, we will focus on the expression profile. obtained for all or some of the genes listed in Tables A2 and A3.

  Preferably, the solid support according to the present invention is characterized in that the 166 fragments are chosen from the following fragments: a) the fragments of sequence SEQ ID N 1 to 55 for Table A1, the fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, and fragments of sequence SEQ ID N 74 to 167 for Table A3, or b) fragments having at least 70% identity. with the fragments as defined in a), or c) the fragments comprising the fragments as defined in a) or b), or d) the fragments whose sequences are complementary to the sequences of the fragments as defined in a), goat).

  Again, one (SEQ ID N 71) or the other (SEQ ID N 72) of the two representative fragments of the TUBB gene may be used indifferently in the present invention, just as the two fragments may be used simultaneous manner (SEQ ID N 71 and SEQ ID N 72). Thus, preferably, the solid support according to the present invention is characterized in that the set comprises and / or consists of 167 fragments, of which 165 fragments are representative of a distinct gene chosen from the genes of Tables A1, A2 and A3. and 2 fragments are representative of the TUBB gene (SEQ ID N 71 and 72).

  According to another preferred embodiment, the solid support according to the present invention is characterized in that the set comprises and / or consists of at least one representative fragment of a gene chosen from the genes of Table B. 14 2875512 Preferably, the solid support according to the present invention is characterized in that the clearance comprises and / or is constituted by at least 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 310 or 315 fragments, each of 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 75, 100, 125 , 150, 175, 200, 225, 250, 275, 300, 310 or 315 fragments being representative of a gene selected from the genes of Table B. Particularly preferably, the solid support according to the present invention is characterized in that the set comprises and / or is further constituted by at least one representative gene fragment for all the genes of Table B. According to a variant of re alisation, the solid support according to the present invention is characterized in that at least one representative fragment of a gene selected from the genes of Table A1, A2, A3 or B is chosen from the following fragments: a) fragments of sequence SEQ ID N 1 to 55 for Table A1, the fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, the fragments of sequence SEQ ID N 74 to 167 for Table A3, and fragments of sequence SEQ ID N 168 to 192, SEQ ID N 193 or 194, SEQ ID N 195 to 201, SEQ ID N 202 or 203, and SEQ ID N 204 to 483 for Table B, or b) fragments having at least 70% identity with the fragments as defined in a), or c) fragments comprising the fragments as defined in a) or b), or d) fragments whose sequences are complementary sequences fragments as defined in a), b) or c).

  In this variant, it is understood that one (SEQ ID N 71) or the other (SEQ ID N 72) of the two representative fragments of the TUBB gene can be used indifferently in the present invention, as well as the two fragments can be used simultaneously (SEQ ID N 71 and SEQ ID N 72). Similarly, one (SEQ ID N 193) or the other (SEQ ID N 194) of the two representative fragments of the BCAR1 gene can be used indifferently in the present invention, just as the two fragments can be used simultaneous manner (SEQ ID N 193 and SEQ ID N 94). Finally, one (SEQ ID NO: 202) or the other (SEQ ID NO: 203) of the two representative fragments of the CCNE1 gene can be used indifferently in the present invention, just as the two fragments can be used simultaneous (SEQ ID N 202 and SEQ ID N 203).

  According to a preferred variant, the solid support according to the present invention is characterized in that at least one representative gene fragment for all the genes of Table A1, A2, A3 or B is chosen from the following fragments: a) the fragments of sequence SEQ ID N 1 to 55 for Table A1, fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, fragments of sequence SEQ ID N 74 to 167 for Table A3, and fragments of sequence SEQ ID N 168 to 192, SEQ ID N 193 or 194, SEQ ID N 195 to 201, SEQ ID N 202 or 203, and SEQ ID N 204 to 483 for Table B, or b) fragments having at least 70% identity with the fragments as defined in a), or c) fragments comprising fragments as defined in a) or b), or d) fragments with the sequences are complementary to the sequences of the fragments as defined in a), b) or c).

  In this other variant, it is also understood that one (SEQ ID N 71) or the other (SEQ ID N 72) of the two representative fragments of the TUBB gene 2875512 can be used indifferently in the present invention, the same two fragments can be used simultaneously (SEQ ID N 71 and SEQ ID N 72). Similarly, one (SEQ ID N 193) or the other (SEQ ID N 194) of the two representative fragments of the BCAR1 gene can be used indifferently in the present invention, just as the two fragments can be used simultaneous (SEQ ID N 193 and SEQ ID N 94). Finally, one (SEQ ID No. 202) or the other (SEQ ID Na 203) of the two representative fragments of the CCNE1 gene can be used indifferently in the present invention, just as the two fragments can be used simultaneous (SEQ ID N 202 and SEQ ID N 203).

  Fragments as defined in point b) of the above embodiments which remain representative of the given genes, result for example from a nucleotide polymorphism or a mutation (deletion, insertion, substitution ...).

  A second aspect of the present invention relates to a method for diagnosing or prognosing in vitro the metastatic or localized character of a mammary tumor from a sample of said tumor in a subject, characterized in that said method comprises a step in which is deposited a sample of a nucleic acid extract, where appropriate after retro-transcription, from said tumor sample, on a solid support according to the present invention, and in that the amount of nucleic acid having hybridized with the set of representative fragments deposited on said support.

  Preferably, the method of diagnosis or in vitro prognosis of the metastatic or localized character of a mammary tumor is characterized in that it comprises the following steps: a) extraction of a test nucleic acid sample from the tumor sample, b) the labeling of the test nucleic acid of the sample obtained in step a) with the aid of a first marker, c) in parallel, the marking of a sample of reference nucleic acid using a second marker different from the first marker of step b), d) bringing the labeled test nucleic acid sample obtained in step b) into contact with each other, and labeled reference nucleic acid sample obtained in step c) with the support according to the present invention under conditions suitable for hybridization, each representative fragment of a solid support gene being able to hybridize with a fragment of the gene present in the sample of labeled test nucleic acid and a fragment of the gene present in the reference nucleic acid sample, e) normalizing, for each representative fragment of gene deposited on the support, the hybridization intensity obtained for the sample of nucleic acid. labeled nucleic acid and that obtained for the labeled reference nucleic acid sample, f) comparing, for each representative fragment of gene deposited on the support, the standardized hybridization intensity obtained for the sample of test nucleic acid labeled with that obtained for the labeled reference nucleic acid sample, so as to obtain an expression profile of the genes whose representative fragments have been deposited on the solid support, g) the classification of the tumor mammary as being metastatic or localized after analysis of the expression profile obtained in step f).

  The extraction of a test nucleic acid sample from the tumor sample as defined in step a) of the prediction method according to the present invention is a technique commonly used in molecular biology. In general, for carrying out the present invention, the test nucleic acid sample is either an mRNA sample or a cDNA sample obtained after retrotranscription of this mRNA; the nucleic acid (mRNA or cDNA) contained in such a sample is in sufficient quantity to characterize the expression profile of all the genes of which a representative fragment has been deposited on the solid support. The molecular biology techniques employed for carrying out the present invention are well known to those skilled in the art, and are widely described in the literature (see, for example, Sambrook, Russell et al., "Molecular cloning: A laboratory manual" , Vol.3, January 15, 2001, Cold Spring Harbor Laboratory, Nucleic Acid Hybridization, BD Haines, SJ Higgins (Eds), 1985, IRL Press). Thus, those skilled in the art will be able to apply the procedure necessary for extracting a sample of the test nucleic acid from the tumor sample.

  The labeling of the test nucleic acid sample in step b) and that of the reference nucleic acid sample in step c) allows the emission of a detectable and quantifiable signal characteristic of the hybridization intensity obtained when a representative fragment of a given gene hybridizes on the one hand with a fragment present in the test nucleic acid sample, on the other hand with a fragment present in the sample of nucleic acid reference. The intensity of hybridization obtained is proportional to the level of expression of the gene represented by the fragment deposited on the solid support.

  The labeling may consist of a radio-labeling (such as 32P, 33P, 35S, which may be detected using, for example, a gamma counter or a scintillator, autoradiography, etc.), enzymatic labeling (biotin, digoxigenin, etc.). .), a chemiluminescent label (luminol, dioxetane, luciferase, luciferin) or a fluorescent label. When radioactive labeling is used, the solid support must not be glass.

  The fluorescent marking is particularly preferred for carrying out the present invention. As an example of fluorescent markers, mention may be made of, but not limited to, fluorescein and its derivatives, such as fluorescein isothiocyanate (FITC), coumarins, cyanines 2, 3 and 5, phycocyanin derivatives, allophycocyanin (APC), phycoerythrin-cyanine (PC5) and phycoerythrin (PE), calcein (AM), tetramethyl-rhodamine red fluorescence or rhodamine and its derivatives, green fluorescence protein or GFP ( Green Fluorescent Protein), dansyl chloride, umbelliferone etc. (see also WT Mason (1999), Fluorescent and Luminescent Probes for Biological Activity, Academic Press, London, 2nd edition).

  The two markers for the test nucleic acid sample and for the reference nucleic acid sample will be chosen so as to obtain two signals characteristic of the hybridization intensity that are comparable to one another. Labeling methods are well known to those skilled in the art and are widely described in the literature (see for example "Practice in the use of nucleic probes and molecular hybridization", Gérard Lacotte, May 1992, Ed Lavoisier, Paris, and WT Mason (1999), Fluorescent and Luminescent Probes for Biological Activity, Academic Press, London, 2nd edition); kits and markers are commercially available (see, for example, the Low RNA Input Fluorescent Linear Amplification Kit, available from Agilent Technologies, or other kits / markers available from, for example, Molecular Probes, Leiden, NL, Clontech, Palo Alto, CA, Stratagene, La Jolla, CA, etc ...).

  The reference nucleic acid sample in the present application is intended to denote a pool of nucleic acid obtained from different normal and tumor cell lines. These cell lines are generally cultured on an industrial scale to produce large batches that undergo stringent quality control procedures. Such a reference nucleic acid sample serves as a control to obtain the information relating to the expression of the genes present in the test nucleic acid sample after hybridization with the representative fragments deposited on the solid support.

  As an example of a reference nucleic acid sample, mention may be made of, but not limited to, the Universal human reference RNA pool, marketed by Stratagene.

  The normalization in step e) makes it possible to overcome the differences caused by the background noise, the duration of the hybridization, the more or less important labeling of the test and reference nucleic acid samples, the variable amounts of the test and reference nucleic acid samples deposited on the solid support, etc. Normalization of the results is commonly used in molecular biology experiments and is well known to those skilled in the art. All types of methods can be used for carrying out the present invention, such as, in particular, the Lowess Global method (Yang IV et al., Genes Biol.

3, research 0062.1-0062.12, 2002).

  The analysis in step g) as a function of the expression profile obtained in step f) is carried out using the Principal Component Analysis method developed by Evry's Génome et Informatique laboratory (software GeneAnova, see point 5 analysis of the data of the example). Other appropriate multivariate analysis methods may be used to classify the mammary tumor as metastatic or localized. Examples of the following software are available on the Internet: - TIGR Multi-Experiment Viewer: www.tigr.org/software, - J-Express: www.ii.uib.no/%7Ebjarted/jexpress, and Genesis, genome.tugraz.at/Software/Genesis/ (without www).

  Preferably, the method according to the present invention is characterized in that the classification in step g) is determined by the following analysis: when the expression profile of step f) is obtained by means of the coated solid support of the set comprising and / or consisting of at least 2 fragments, preferably at least 50 fragments, and particularly preferably of the set comprising and / or consisting of 72 fragments and for detecting ganglion invasion in the case of tumors ER +, it is compared with the expression profile as defined in Tables A1 and A2; or - when the expression profile of step f) is obtained by means of the solid support coated with the set comprising and / or consisting of at least 2 fragments, preferably at least 50 or 100 fragments, and particularly preferably of the set comprising and / or consisting of 111 fragments and for detecting ganglion invasion in the case of ER- tumors, it is compared with the expression profile as defined in Tables A2 and A3; or when the expression profile of step f) is obtained by means of the solid support coated with the set comprising and / or consisting of at least 2 fragments, preferably at least 50, 100 or 150 fragments, and in particular preferred game comprising and / or consisting of 166 fragments and for detecting ganglion invasion in both ER- tumors and ER + tumors, it is compared with the expression profile as defined in Tables Al , A2 and A3.

  Preferably, when the expression profile of step f) is obtained by means of the solid support coated with the set comprising and / or consisting of 72 fragments chosen from the following fragments: a) fragments of sequence SEQ ID N 1 at 55 for Table A1, and fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, or b) fragments having at least 70% identity with the fragments as defined in a), or c) the fragments comprising the fragments as defined in a) or b), or d) the fragments whose sequences are complementary to the sequences of the fragments as defined in a), b ) or c), said support for detecting ganglion invasion in the case of ER + tumors, said profile is compared with step g) with the expression profile as defined in Tables A1 and A2 Preferably when the expression profile of step f) is obtained by means of the solid support coated with the set comprising and / or consisting of 111 fragments selected from the following fragments: a) fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, and fragments of sequence SEQ ID N 74 to 167 for Table A3, or b) fragments having at least 70% identity with the fragments as defined in a), or c) fragments comprising the fragments as defined in a) or in b), or d) the fragments whose sequences are complementary to the sequences of the fragments as defined in a), b) or c), said support for detecting ganglion invasion in the case of tumors ER-, said profile is compared with step g) with the expression profile as defined in Tables A2 and A3.

  Preferably, when the expression profile of step f) is obtained by means of the solid support coated with the set comprising and / or consisting of 166 fragments chosen from the following fragments: a) fragments of sequence SEQ ID No. 1 at 55 for Table A1, the fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, and the fragments of sequence SEQ ID N 74 to 167 for Table A3 or (b) fragments having at least 70% identity with the fragments as defined in (a), or (c) fragments comprising the fragments as defined in (a) or (b), or (d) fragments of which sequences are complementary sequences of fragments as defined in a), b) or c), said support for detecting ganglion invasion in both ER tumors and ER + tumors, said profile is compared to the step g) with the expression profile as defined in Tables A1, A2 and A3.

  As has been said previously, before determining the lymph node invasion for a given mammary tumor, it is essential to characterize the expression of estrogen receptors by the cells of said tumor. This characterization of the expression of the estrogen receptors can be carried out according to any appropriate method known to those skilled in the art such as, for example, immunohistochemistry, but also by using the method according to the present invention.

  Thus, according to a particular embodiment, the method according to the present invention also makes it possible to characterize whether or not the mammary tumor cells express the estrogen receptors, which process is characterized in that after step f), the expression profile of the set of genes of Table C obtained is compared by means of the solid support coated with the set comprising and / or consisting of at least 36, preferably at least 50, 100, 150 or 166 representative fragments of a distinct gene selected from the genes of Tables A1, A2 and A3, or the set comprising and / or consisting of at least one representative fragment of a gene chosen from the genes of Table B, with the profile of expression as defined in said Table C. The genes of Table C, namely AR, BAG1, CDH13, CEGP1, CHI3L1, COX6C, CPB1, CRIP1, CSDA, D123, SOX9, EZH2, FLJ00382, GDI2, HLA-DQB1 genes. , HSPC195, IL6, K-ALPHA-1, KRT17, KRT18, KRT7, LDB2, LIV-1, MYC, NAT1, NSEP1, PECAM1, PGR, PLAT, RRM1, S100A9, SLPI, TFF1, TPBG, TUBB and VWF, constitute a group of genes selected from the genes of Tables A1, A2, A3 and B Thus, a solid support according to the present invention, making it possible to detect ganglionic invasion in both ER tumors and in ER + tumors, and in particular comprising this group of genes, will also be able to characterize whether the cells of the tumor mammary express (ER +) or not (ER-) estrogen receptors: using this support, we will first characterize whether the tumor from which the sample was taken is an ER + tumor or an ER tumor - and in a second step, if the tumor is ER +, we will focus only on the expression profile obtained for all or part of the genes listed in Tables A1 and A2, and if the tumor is ER-, we have will interest the expression profile obtained for all or part of the genes listed in Tables A2 and A3.

  According to yet another preference, the method according to the present invention is characterized in that the comparison in step f) of the normalized hybridization intensity obtained for the labeled test nucleic acid sample with that obtained for the labeled reference nucleic acid sample comprises the following steps: a) calculating, for each representative fragment of a gene deposited on the support, the ratio of the normalized hybridization intensity obtained for the nucleic acid sample labeled test and that obtained for the labeled reference nucleic acid sample, b) calculating, for each representative fragment of a gene deposited on the support, the log2 of each ratio obtained in step a).

  Each representative fragment of the set is generally deposited once, preferably twice, more preferably three times or even four times on the solid support. Thus, according to a particular embodiment, the method according to the present invention is characterized in that, when each fragment of the game is deposited three times on the support, the log2 average of the three replicates is calculated for each of these fragments.

  According to another embodiment, the method according to the present invention is characterized in that the test and reference nucleic acid samples are RNA, preferably mRNA.

  Preferably, the method according to the present invention is characterized in that the step b) of marking the test RNA, preferably the test mRNA, of the sample with the aid of the first marker comprises the following steps i) retrotranscription of the test RNA from the sample obtained in step a), ii) incorporation of the first marker during the in vitro transcription of the test DNA obtained in step i).

  According to another preference, the method according to the present invention is characterized in that step c) of marking the reference RNA sample, preferably reference mRNA, with the aid of the second marker comprises the following steps: i) the retrotranscription of the reference RNA from step c), ii) the incorporation of the second marker during the in vitro transcription of the test DNA obtained in step i).

  More preferably, the process according to the present invention is characterized in that the first and second markers are chosen from cyanine 3 and cyanine 5.

  A final aspect of the present invention is a kit comprising the solid support of the present invention. Kits comprising a solid hybridization support are sufficiently described in the literature, and generally comprise, in addition to the solid support, reagents necessary for hybridization, such as, in particular, markers or hybridization and washing buffers.

  The legends of the figures and examples which follow are intended to illustrate the invention without in any way limiting its scope.

FIGURES

  Figure lA: Classification obtained by Analysis in Multiple Components for ER + tumors (PCR, GeneAnova software, Evry genome and computer laboratory).

  Figure 1B: Classification obtained by Multiple Component Analysis for ER- tumors.

  Figure 2: Classification obtained by Multiple Component Analysis for the study of estrogen receptor (ER + / ER-) expression by mammary tumor cells.

EXAMPLES

  Example 1 Study of Ganglion Involvement 1. Patients Surgical fragments of mammary tumors in patients operated at the Jean Perrin Center (Clermont-Ferrand) were taken. The criteria for tumor selection were the histological type (invasive ductal carcinomas) and the occurrence of these tumors in a sporadic context. A total of 69 tumors are studied.

  2. Biological samples The extraction of RNA is carried out from the frozen tumors in liquid nitrogen with the Trizol protocol (Invitrogen, Cergy-Pontoise, France). After verifying the quality of the RNAs obtained by microelectrophoresis on the bioanalyzer (Agilent Technologies, Massy, France), 10 g of total RNA are used for the on-chip study.

  A pool of RNA from cell lines is used as the reference RNA (Universal human reference RNA, Stratagene, La Jolla, CA).

  This reference pool of reference RNA is commercially available and includes a collection of RNAs obtained from a number of cell lines to cover an optimal range of genes.

  3. Preparation of biochips (or solid supports) Molecular probes (or representative fragments) are oligonucleotides of 50 bases synthesized by MWG. The sequence of some oligonucleotides is available in the MWG banks, 28,475,512 others have been sought at the request of the inventors. These oligonucleotides are diluted to a concentration of 25 M in a solution of 50% DMSO (dimethylsulfoxide) and then deposited on Ultragaps slides (Corning, New York, US) using a robot (Microgrid II, Biorobotics, Genomic Solutions, Cambridge, UK). ). Each molecular probe is deposited in triplicate on the slide. After deposition, the DNA molecules are fixed by incubation on slides at 80 C for 2 hours. A chemical treatment as defined by the company Corning is carried out. The slides are then stored at room temperature in the dark.

  4. Marking and Hybridization For each sample, 10 g RNA is retranscribed in the presence of amino-allyl dUTP (Fairplay Microarray labeling kit, Stratagene, La Jolla, US). The cDNAs obtained from the RNAs of the tumor and the reference RNA are respectively incubated with cyanine and cyanine 3 (Amersham Biosciences, Quebec, CA) which will bind to the amino-allyl group. The fluorescent cDNAs are then purified and dried under vacuum. The pellet is taken up in 160 of hybridization buffer (Corning, New York, US) and then the denaturation is carried out at 100 C. Hybridization is done with a coverslip which makes it possible to maintain the contact between the target and the surface of the blade, at 37 C and 50% humidity, during the night. The slides are then washed in solutions of SDS (sodium dodecyl sulphate) and SSC (saline sodium citrate) at different concentrations. Fluorescence is detected by passing the chip through a scanner (GMS 418, Affymetrix).

  5. Data analysis The images obtained are analyzed with image analysis software (Jaguar, Affymetrix, UK). The normalization of the data is performed according to the global lowess method because there is a systematic dependence between the value of the log2 (ratio) and the intensity, which usually appears with high log2 (ratio) values for the values of the log2 (ratio). low intensity. This method makes it possible to remove such effects. Then, the ratio of tumor / reference intensities is calculated, then the ratio box. For each gene, the average log2 (ratio) of the three replicates is calculated, each molecular probe being deposited three times on the chip.

  The Student test is then used. This is a significance test that can be used when comparing two averages. The t-test is calculated by performing the ratio of the difference of the averages over the standard error. A value p <0.05 is considered significant. This test is involved to evaluate the level of significance between the means of expression of the different genes between the group of sensitive samples and the group of resistant samples. The expression profiles of the discriminant genes (p <0.05) are then studied by a principal component analysis (PCA).

  Principal Component Analysis (GeneAnova software, Genome and Computer Science Laboratory of Evry, FR) is a so-called descriptive method, it aims at structuring and simplifying data from several variables, without favoring one of them. The PCA allows to visualize the geometry of the cloud of observations when the space has more than 3 dimensions. It proceeds in 2 steps, the determination of the axes of inertia of the cloud then its projection on the planes determined by the axes of inertia taken 2 to 2. The analysis of the sphere of the correlations makes it possible to visualize the similarities between different profiles of 'expression. The linear correlation between 2 profiles is equal to the cosine of the angle between the vectors corresponding to these experimental conditions (an angle of 90 shows a total absence of correlation).

  6. Results and discussion Two groups of patients are studied (see Table Clinical data of patients). The first corresponds to patients whose tumors express the estrogen receptor (estrogen receptor positive, ER +), the second to those who do not express it (estrogen receptor negative, ER -).

  111 genes (see Tables A2 and A3) have an expression profile that makes it possible to discriminate ER- / N- tumors of ER- / N + tumors against 72 genes (see Tables A1 and A2) for ER + tumors. The classifications (FIGS. 1A for ER + tumors and IB for ER tumors) obtained by multiple component analysis reveal two groups, one corresponds to N tumors, the other corresponds to N + tumors.

  The biochip used in this study can therefore be used to determine the lymph node status of a tumor, thus avoiding axillary dissection to patients.

  Table Clinical Data of Patients N Patient Invaded Ganglia / No. of Status N ER Ganglion Group Taken 9 0/5 N- + Group 1 177 0/14 N- + 178 0/13 N- + 186 0/13 N- + 187 0 / 11 N- + 188 0/9 N- + 193 0/15 N- + 194 0/9 N- + 2 6/21 N + + 8 1/16 N + + 23 1/13 N + + 28 6/18 N + + 33 2/10 N + + 38 '1/12 N + + 1/22 N + + 48 2/15 N + + 49 2/9 N + + 53 1/3 N + + 58 3/10 N + + 10/10 N + + 61 3 / 13 N + + 63 5/12 N + + 69 7/12 N + + 73 5/13 N + + 76 9/13 N + + 91 2/6 N + + 96 1/12 N + + I35c 127 2/13 N + + 128 6/8 N + + 129 3/5 N + + 157 2/11 N + + 159 4/19 N + + 2/12 N + + 181 3/15 N + + 189 8/19 N + + 191 1/13 N + + 192 1/6 N + + 196 2/9 N + + 13/25 N + + 201 3/10 N + + 32 ND + 56 ND + 64 ND + 68 ND + 78 ND + 168 ND + 176 ND + Nb invaded ganglia / number of lymph nodes removed 0/7 N patient Status N ER Group Group 2

NOT-

  31 0/21 N-59 0/8 N-92 0/11 N-174 0/8 N-0/5 N-0/25 N-0/7 N-198 0/10 N-1 6/12 N + 4 1/18 N + 24 1/11 N + 41 25/28 N + 2/24 N + 57 3/12 N + 81 2/13 N + 163 1/10 N + 2/20 N + - 166 1/8 N + - 203 3/14 N + - 153 ND -

ND -

  Group 1 corresponds to patients whose tumors express the estrogen receptor (ER +).

  Group 2 corresponds to patients whose tumors do not express the estrogen receptor (ER-).

  ND: Ganglionic status not determined.

  N: N + status, ganglionic invasion; N-, no ganglionic invasion.

  Example 2: Study of the Expression of the Estrogen Receptors Breast cancer is hormonodependent. Tumor cells express estrogen receptors (ERs). ER + tumors have a better prognosis than ER tumors.

  The data used are those of the patients studied for the characterization of ganglionic invasion.

  The technical approach is identical (see Example 1). Only ER-tumor-discriminating genes of ER-tumors change and are shown in Table C. The classification obtained is that of Figure 2.

  The patient n 68 initially characterized ER + is classified in the group ER-.

  Further studies are under way to know the biological significance of this result, a present but non-functional receptor being an explanation.

TABLE Al

  Sequence id Symbol Unigene ER-tumors ER-tumors N-N + NN + gene name Seq id n 1 AP3S2 Hs.154782 X adapor-related protein complex 3, sigma 2 subunit Seq id n 2 APOA1 Hs.93194 X apolipoprotein AI Seq id n 3 BYSL Hs.106880 X bystin-like Seq id 4 CBX3 Hs.381189 X chromobox homolog 3 Seq id n 5 CCND1 Hs.371468 X cyclin Dl Seq id n 6 CDC20 Hs. 82906 X cell division cycle 20, S.cerevisiae homolog Seq id n 7 CDH15 Hs. 148090 X cadherin 15, M-cadherin (myotubule) Seq id n 8 CDKNIA Hs. 370771 X cyclin-dependent kinase inhibitor 1A (p21, Cipl) Seq id n 9 CHK1 Hs.24529 X chkl checkpoint s.pombe homolog chekl; hchkl kinase Seq id 10 COL4A2 Hs.407912 X alpha 2 type iv collagen Seq id n 11 CRABP2 Hs.183650 X cellular retinoic acid-binding protein 2 Seq id n 12 CRMP 1 Hs.155392 X collapsin response mediator protein 1 Seq id n 13 CUL4A Hs.270788 X cullin 4A Seq id 14 DDB2 Hs.446564 X damage-specific DNA binding protein 2 (48kD) Seq id 15 DTR Hs.799 X diphtheria toxin receptor (heparin-binding epidermal growth factor-like growth factor ) Seq id n 16 EDNRB Hs.82002 endothelin receptor type B Seq id 17 EXT1 Hs.184161 X exostoses (multiple) 1 Seq id n 18 FGF2 Hs. 284244 X fibroblast growth factor 2 (basic) Seq id n 19 GAPD Hs. 80206 X-glyceraldehyde-3-phosphate dehydrogenase Seq id 20 GSS Hs. 82327 X glutathione synthase seq id 21 IGF1 Hs.85112 _ insulin-like growth factor 1

X

  Seq id n 22 IGF1R Hs.239176 X _ insulin-like growth factor receptor 1 Seq id 23 IGF2R Hs.76473 X insulin-like growth factor 2 receptor Seq id n 24 IGFBP1 Hs.401316 X _ insulin-like growth factor binding protein 1 Seq id 25 KRT18 Hs.406013 X keratin 18 Seq id n 26 LDB2 Hs.4980 X LIM binding domain 2 Seq id n 27 LRP1 Hs.162757 X low density liEo.rotein- related protein alpha-2-macroglobulin receptor Seq id n 28 MAPK1 Hs. 324473 X mtogen-activated protein kinase 1 Seq id 29 MFNG Hs. 371768 _ manie fringe (Drosophila) homolog

X

  Seq id n 30 MLH1 Hs.433618 X mutL (E. coli) homolog 1 (colon cancer, nonpolyposis type 2) Seq id n 31 MS4A7 Hs.188011 X ms4a7 Seq id n 32 MTMR4 Hs.141727 - X myotubularin related protein 4 Seq id n 33 MYC Hs.202453 X v-myc avian myelocytomatosis viral oncogene homolog Seq id n 34 NME1 Hs.118638 X non-metastatic cells 1, protein (NM23A) expressed in (NME1) Seq id n 35 NONO Hs.355861 X no -POU-domain- containing, octamer-binding Seq id 36 ORC6L Hs.49760 X origin recognition complex subunit 6 yeast homolog-like Seq id n 37 PAICS Hs. 444439 _ _ X multifunctional polypeptide similar to SAICAR synthetase and AIR carboxylase Seq id n 38 PAI-RBP1 Hs.356427 X pal-1 mrna-binding protein Seq id n 39 PRAME Hs.30743 X Preferentially expressed antigen in melanoma Seq id n 40 PRKARIA Hs.183037 X protein kinase, cAMP-dependent, regulatory, type I, alpha (tissue specific extinguisher 1) Seq id n 41 PSMD12 Hs.4295 X proteasome (prosome, macropain) 26S subunit, non-ATPase, 12 Seq id n 42 RB 1 Hs.408528 X retinoblastoma 1 (including osteosarcoma) Seq id n 43 RBL2 Hs.283604 X retinoblastoma-like 2 (p130) Seq id n 44 RELA Hs.132594 X v-rel avian reticuloendotheliosis viral oncogene homolog A (nuclear factor of kappa light polypeptide gene enhancer in B-cells 3 (p65)) Seq id n 45 S 100A9 Hs.112405 X S100 calcium-binding protein A9 (calgranulin B) Seq id 46 SFRS5 Hs.166975 X splicing factor srp40-1 srp40 member of the family sr protein factor-factors, arginine / serine-rich 5 Seq id n 47 STAT3 Hs. 421342 X signal transducer and activator'of transcription 3 (acute-phase response factor) Seq id n 48 TAP2 Hs.502 X ATP-binding cassette, sub- family B (MDR / TAP), member 3 (ABCB3) Seq id n 49 TRAG3 Hs.458484 X TXL resistance associated gene 3 Seq id n 50 TRIP13 Hs.436187 Homo sapiens HPV16 El protein binding protein mRNA, complete cds Seq id n 51 UMPS Hs. 2057 X uridine monophosphate synthetase (orotate phosphoribosyl transferase and orotidine-5'-decarboxylase) Seq id n 52 WARS Hs. 82030 X tryptophanyl-tRNA synthetase Seq id n 53 WISP1 Hs.194680 X wntl inducible signaling pathway protein 1 Seq id n 54 WISP2 Hs.194679 X WNT1 inducible signaling pathway protein 2 Seq id 55 XBP 1 Hs.437638 X X-box binding protein 1

TABLE A2

  Sequence id Unigenic Tumor ER-Neoplasms ER + N-N + NN + Seq id n 56 ABL1 Hs.446504 XX c-abl (v-abl Murine Abelson leuken - viral oncogene homolog 1) Seq id n 57 AP2B1 Hs. 370123 XX adaptor-related protein complex 2, beta 1 subunit Seq id 58 BIRC5 Hs.1578 XX apoptosis inhibitor 4 (survivin) Seq id 59 BRCA1 Hs.194143 XX breast cancer 1, early onset Seq id n 60 FLT1 Hs.433670 XX fms-related tyrosine kinase 1 (vascular endothelial growth factor / vascular permeability factor receptor) Seq id n 61 GCLC Hs.414985 XX glutamate-cysteine ligase, catalytic subunit Seq id 62 HSPD1 Hs.79037 _ X heat shock 60kD protein 1 ( chaperonin)

X

  Seq id 63 KIAA0788 Hs.246112 X X KIAA0788 protein Seq id 64 NRAS Hs. 260523 X X neuroblastoma viral RAS (v-ras) oncogene homolog Seq id 65 ODCI Hs.443409 X X ornithine decarboxylase 1 Seq id n 66 ZNF607 Hs. 75323 X _ X zinc finger protein Seq id n 67 PHB Hs.75323 X _ X prohibitin Seq id n 68 PIK3R2 Hs.211586 X X phosphoinositide-3-kinase, regulatory subunit, polypeptide 2 (p85 beta) Seq id n 69 STC2 Hs. 155223 X _ X stanniocalcin 2 Seq id n 70 SYT1 Hs.154679 XX synaptotagmin I Seq id n 71 TUBB Hs.512712 XX homo sapiens clone 24703 beta-tubulin mRNA, complete cds Seq id n 72 TUBB Hs.512712 X _ tubulin, beta polypeptide

X

  Seq id n 73 ZNF161 Hs.414455 X _ X zinc fnger protein 161

TABLE A3

  Sequence Symbol Tumors ER-Tumors Gene name id ER + N- N + N- N + Seq id n 74 ABCC1 Hs.21330 X P-glycoprotein 1, MDR1 (ATP-binding cassette, sub-family B, member 1) Seq id n 75 ABCC6 Hs.107911 X ATP-binding cassette, sub-family C (CFTR / MRP), member 6 Seq id n 76 ABCG2 Hs. 194720 X ATP-binding cassette, sub-family G (WHITE), member 2 Seq id n 77 ACTRIA Hs.153961 X _ ARP1 (actin-related protein 1, yeast) homolog A (centractin alpha) Seq id n 78 ADPRT Hs. 177766 X ADP-ribosyltransferase (NAD +; poly (ADP-ribose) polymerase) Seq id n 79 AKT1 Hs.368861 X v-akt murine thymoma viral oncogene homolog 1 Seq id n 80 ARHB Hs.406064 X ras homolog gene family, member B Seq id n 81 B2M Hs.48516 X beta-2-microglobulin Seq id n 82 BBC3 Hs.35052 X bd-2 binding component 3 Seq id 83 BCAR3 Hs.201993 X breast cancer anti-estrogen resistance 3 Seq id n 84 BCL2 Hs.79241 X B-cell CLL / lymphoma 2 Seq id 85 BCL2L1 Hs. 305890 X BCL2-like 1 Seq id 86 BLMH Hs.387183 X bleomycin hydrolase Seq id 87 BNC Hs.64025 X basonuclin Seq id 88 CBP Hs.136227 X CREB-binding protein Seq id 89 CD68 Hs.246381 X CD68 antigen Seq id n 90 CDC27 Hs.406631 X cell cycle division 27 Seq id n 91 CHIT1 Hs. 201688 X chitinase 1 (chitotriosidase) Seq id n 92 CRADD Hs. 155566 X CASP2 and RIPK1 domain containing adaptator with death domain Seq id n 93 CSF1 Hs.173894 X colony stimulating factor 1 (macrophage) Seq id n 94 CTNND1 Hs.166011 X catenin, delta 1 Seq id n 95 CTSC Hs. 128065 X cathepsin C Seq id n 96 CYP19 Hs.187471 X cytochrome P450, subfamily XIX (aromatization of androgens) Seq id n 97 DENR Hs. 22393 X density regulated protein Seq id 98 DLX4 Hs.172648 X DLX4 Seq id 99 E2F3 Hs.1189 X E2F transcription factor 3 Seq id EDR2 Hs. 165263 Early development regulator 2 n 100 Seq id EEF1D Hs. 334798 X eukaryotic translation elongation factor 1 delta (guanine nucleotide exchange protein) n 101 Seq id EGFR Hs.77432 X epidermal growth factor receptor (avian erythroblastic leukemia viral (v-erb-b) oncogene n 102 homolog) Seq id EPAS 1 Hs. 8136 X endothelial NO domain protein 1 n 103 Seq id ERCCI Hs.435981 X _ excision repair cross-complementing rodent repair deficiency, complementation group 1 n 104 Seq id FABP7 Hs.26770 X _ fatty acid binding protein 7, brain n 105 Seq id FGF8 Hs.57710 X _ fibroblast growth factor 8 (androgen-induced) n 106 Seq id FLJ00382 Hs.439852 X Homo sapiens mRNA for FLJ00382 protein n 107 Seq id FOXM1 Hs.239176 X forkhead (Drosophila) -like 16 n 108 Seq id G22P 1 Hs.169744 X thyroid autoantigen 70kd antigen n 109 Seq id GATA3 Hs. 169946 X GATA-binding protein 3 n 110 Seq id GDI2 Hs.56845 X gdp dissociation inhibitor 2 n 111 Seq id GPD1 Hs.350554 X glycerol-3 phosphate dehydrogenase 1 (soluble) n 112 Seq id GPX4 Hs. 433951 X glutathione peroxidase 4 (phospholipid hydroperoxidase) n 113 Seq id GSTA3 Hs.102484 X _ glutathione S-transferase A3 n 114 Seq id GSTM1 Hs.301961 X glutathione s-transferase; gstmlb glutathione S-transferase M1 n 115 Seq id H2AFX Hs.147097 X H2A histone family, member X n 116 Seq id IFI27 Hs.278613 X interferon alpha inducible protein 27 n 117 Seq id IFITM1 Hs.458414 X interferon-induced protein 17 n 118 Seq id IGH Hs.381568 X Human Ig J Chain Bene n 119 _ Seq id IL1B Hs. 126256 X interleukin 1, beta n 120 Seq id ITGA6 Hs.212296 X integrin, alpha 6 n 121 Seq id ITGB3 Hs.87149 X integrin, beta 3 (platelet glycoprotein IIIa, antigen CD61) n 122 Seq id JUN Hs.78465 X v on avian sarcoma virus 17 oncogene homolog 123 Seq id K Hs.446608 X Alpha tubulin n 124 ALPHA-1 Seq id KIAA0430 Hs.432741 X KIAA0430 gene product 125 Seq id KIAA1067 Hs.511946 X kiaalO67 n 126 Seq id KLK3 Hs. 171995 X _ _ kallikrein 3, (prostate specific antigen) n 127 Seq id LMNB1 Hs.89497 X lamin B1 n 128 Seq id LMNB2 Hs.76084 X lamin B2 n 129 Seq id LMO4 Hs.3844 X lim domain only 4 n130 Seq id LSP1 Hs. 56729 X lymphocyte-specific protein 1 n 131 Seq id LTBP1 Hs. 241257 X _ latent transforming growth factor beta binding protein 1 n 132 Seq id MAPK3 Hs.861 X ERK1, mitogen-activated protein kinase 3 n 133 Seq id MGB1 Hs.46452 X mainmaglobin 1 n 134 Seq id MMP17 Hs.159581 X matrix metalloproteinase 17 n 135 Seq id MTA1 Hs.101448 X metastasis-associated gene 1 n 136 Seq id NFX1 Hs.413074 X nuclear transcription factor, X-box binding 1 n 137 Seq id OXTR Hs.2820 X oxytocin receptor n 138 Seq id PA2G4 Hs .374491 X proliferation-associated 2G4, 38kD n 139 Seq id PDGFB Hs.1976 X platelet-derived growth factor beta polypeptide (simian sarcoma viral (v-sis) oncogene n 140 homolog) Seq id PMS2 Hs. 177548 X postmeiotic segregation increased (S. cerevisiae) 2 n 141 _ Seq id PRGI Hs.1908 X proteoglycan 1, secretory granule n 142 Seq id PRKCE Hs. 155281 X protein kinase C, epsilon n 143 Seq id PRKCG Hs.2890 X protein kinase C, gamma n 144 Seq id PRKCZ Hs.407181 X protein kinase C, zeta n 145 Seq id PSME1 Hs.75348 X proteasome (prosome, macropain) activator subunit 1 (PA28 alpha) n 146 Seq id PTGES Hs.146688 X _ prostaglandin E synthase n 147 Seq id PTK2 Hs.434281 X PTK2 protein tyrosine kinase 2 n 148 Seq id RAF1 Hs.257266 X v-raf-1 murine leukemia viral oncogene homolog raft v-raf-1 murine leukemia viral oncogene n 149 homolog 1 Seq id RPS28 Hs.153177 ribosomal protein S28 n 150 Seq id SLC7A5 Hs. 184601 X _ solute carrier family 7 (cationic amino acid transporter, y + system), member 5 n 151 Seq id SLPI 'X secretory leukocyte protease inhibitor (antileukoproteinase) n 152 Hs.251754 Seq id SNL Hs. 118400 _ _ singed (Drosophila) -like (sea urchin fascin homolog like) n 153 X Seq id SPS Hs.124097 X Selenophosphate synthetase; human selenium donor protein n 154 Seq id STAB1 Hs301989 X _ kiaa0246 protein n 155 Seq id TERT Hs.439911 X _ telomerase reverse transcriptase n 156 Seq id TGFBR2 Hs.82028 X _ _ transforming growth factor, beta receptor II (70-80kD) n 157 Seq id TGFBR3 Hs342874 X _ _ transforming growth factor, beta receptor III (betaglycan, 300kD) n 158 Seq id TOB1 Hs.178137 X transducer of erbb2 1; tob 1 n 159 Seq id TRAF4 Hs.8375 X TNF associated factor 4 n 160 Seq id TRAF6 Hs.444172 X TNF receptor-associated factor 6 n 161 Seq id TRB X63456 X T-cell receptor, beta cluster n 162 Seq id UCH37 Hs . 25223 X ubiquitin c-terminal hydrolase uch37 n 163 Seq id UP Hs. 314828 X uridine phosphorylase n 164 Seq id VHL Hs.421597 X von Hippel-Lindau syndrome n 165 Seq id XPA Hs.288867 X xeroderma pigmentosum, complementation group A n 166 Seq id ZNF217 Hs.155040 X zinc finger protein 217 n 167

TABLE B

  Sequence id Symbol Unigene Gene name Seq id n 168 TFAP2A Hs. 210911 transcription factor AP-2 alpha (activating enhancer-binding protein 2 alpha) Seq id n 169 TFAP2C Hs.440411 transcription factor ap-2 gamma (activating enhancer binding protein 2 gamma) Seq id n 170 A2M Hs. 74561 alpha-2-macroglobulin Seq id n 171 ABCC5 Hs.310735 ATP-binding cassette, sub-family C (CFTR / MRP), member 5 Seq id n 172 ACP5 Hs. 1211 acid phosphatase 5, tartrate resistant Seq. Id 173 ACTA2 Hs. 208641 actin, alpha 2, smooth muscle, aorta Seq id 174 ADH1B Hs. 4 alcohol dehydrogenase 2 (class I), beta polypeptide Seq id 175 ADRBK1 Hs.83636 adrenergic, beta, receptor kinase 1 Seq id 176 AKAP2 Hs.42322 a kinase (prka) anchor protein 2; akap2 Seq id n 177 ALDHIAI Hs.76392 aldehyde dehydrogenase 1 family, member Al Seq id n 178 ALDH4A1 Hs.77448 aldehyde dehydrogenase family member al Seq id n 179 ALDOA. Hs.273415 aldolase A, fructose-bisphosphate Seq id n 180 ANXA1 Hs.287558 annexin Al Seq id n 181 ANXAB Hs.87268 annexin A8 Seq id n 182 APEX Hs.73722 apex nuclease (multifunctional dna repair enzyme) Seq id n 183 APOD Hs.75736 apolipoprotein D Seq id n 184 AQP1 Hs. 76152 aquaporin 1 (channel-forming integral protein, 28kD) Seq id n 185 AQP7 Hs.455323 aquaporin 7 Seq id n 186 AR Hs.99915 androgen receptor (dihydrotestosterone receptor; testicular feminization; spinal and bulbar muscular atrophy; Kennedy disease) Seq id n 187 ARVCF Hs. 326730 armadillo repeat gene deletes in velocardiofacial syndrome Seq id n 188 ATDC Hs.82237 ataxia-telangiectasia group D-associated protein Seq id 189 ATF3 Hs.460 activating transcription factor 3 Seq id 190 BAG1 Hs.377484 BCL2-associated athanogene Seq id n 191 BARD1 Hs. 54089 BRCA1 associated RING domain 1 Seq id 192 BAX Hs.159428 BCL2- associated X protein Seq id 193 BCAR1 Hs.273219 breast cancer anti-estrogen resistance 1 Seq id n 194 BCAR1 Hs.273219 breast cancer anti-estrogen resistance 1 Seq id n 195 BPAG1 Hs.443518 bullous pemphigoid antigen 1 (230 / 240kD) Seq id 196 BRCA2 Hs.34012 breast cancer 2, early onset Seq id 197 BRF1 Hs.424484 butyrate response factor 1 (EGF- response factor 1) Seq id n 198 BUB1B Hs.36708 budding uninhibited by benzimidazoles 1 (yeast homolog), beta Seq id n 199 CAD Hs. 377010 carbamoylphosphate synthetase 2aspartate transcarbamylasedihydroorotase Seq id n 200 CAV1 Hs.74034 caveolin 1, caveolae protein, 22kD Seq id n 201 CCNB 1 Hs.23960 cyclin BI Seq id 202 CCNE1 Hs.244723 cyclin El seq id 203 CCNE1 Hs.244723 cyclin el Seq id 204 CCNE2 Hs.408658 cyclin e2, isoform 1 Seq id 205 CCNG1 Hs. 79101 cyclin G1 Seq id 206 CD2 Hs.89476 CD2 antigen (p50), sheep red blond cell receptor Seq id 207 CD34 Hs.374990 CD34 antigen Seq id 208 CD36 Hs.443120 CD36 antigen (collagen type I receptor, thrombospondin receptor CD3D Hs.95327 CD3D antigen, delta polypeptide (TiT3 complex) Seq id 210 CD3G H5.2259 CD3G antigen, gamma polypeptide (TiT3 complex) Seq id 211 CDC25C Hs.656 cell division cycle 25C Seq id n 212 CDC42BPA Hs.18586 CDC42-binding protein kinase alpha isoform A Seq id 213 CDC6 Hs.405958 CDC6 (cell division cycle 6, S. cerevisiae) homolog Seq id 214 CDC7L1 Hs.28853 CDC7 (cell division cycle 7, S. cerevisiae, homolog) -like 1 Seq id n 215 CDHI Hs. 194657 cadherin 1, type 1, E-cadherin (epithelial) Seq id n 216 CDH13 Hs. 63984 cadherin 13, H-cadherin (heart) Seq id n 217 CDH3 Hs. 191842 cadherin 3, type 1, P-cadherin (placental) Seq id n 218 CDK4 Hs. 95577 Cyclin-dependent kinase 4 Seq id 219 CDKNIB Hs.238990 cyclin-dependent kinase inhibitor 1B (p27, Kipl) Seq id n 220 CDKN 1 C Hs. 106070 cyclin-dependent kinase inhibitor 1C (p57, Kip2) Seq id n 221 CDKN2A Hs.421349 cyclin-dependent kinase inhibitor 2A (melanoma, p16, CDK4 inhibitors) Seq id n 222 CEACAM3 Hs.220529 carcinoembryonic antigen Seq id n 223 CEGP1 Hs .435861 cegpl protein Seq id n 224 CENPA Hs. 1594 centromere protein a Seq id 225 CENPE Hs.75573 centromere protein E (3121 (D) Seq id n 226 _ Hs.77204 centromere protein F (350 / 400kD, mitosin)

CENPF

  Seq id n 227 CHI3L1 Hs.382202 chitinase 3-like 1 (cartilage glycoprotein39) Seq id n 228 CKS2 Hs.83758 CDC28 protein kinase 2 Seq id n 229 COL1A1 Hs.172928 collagen, type I, alpha 1 Seq id n 230 COL3A1 Hs . 443625 collagen, type III, alpha 1 (Ehlers-Danlos syndrome type IV, autosomal dominant) Seq id n 231 COL6A1 Hs.415997 collagen, type VI, alpha 1 Seq id 232 COMP H5.1584 cartilage oligomeric matrix protein (pseudoachondroplasia, epiphyseal dysplasia 1, multiple) Seq id n 233 COX6C Hs.351875 cytochrome c oxidase subunit vic cox6c nuclear encoding mitochondrial protein cytochrome c oxidase subunit Vic Seq id n 234 COX7A2L Hs.423404 cytochrome c oxidase subunit VIIa polypeptide 2 like Seq id n 235 CP Hs 282557 ceruloplasmin (ferroxidase) Seq id 236 CPB1 Hs.437028 carboxypeptidase B1 (tissue) Seq id n 237 CRIP1 Hs. 70327 cysteine-rich protein 1 (intestinal) Seq id n 238 CSDA Hs. 221889 cold shock domain protein A Seq id n 239 CSFIR Hs.174142 colony stimulating factor 1 receptor, formally McDonough feline sarcoma viral (v-fms) oncogene homolog Seq id 240 CST6 Hs.139389 cystatin M precursor Seq id n 241 CTPS Hs. 251871 ctp synthase Seq id n 242 CTSD Hs. 343475 cathepsin D (lysosomal aspartyl protease) Seq id n 243 CTSZ Hs. 252549 cathepsin Z Seq id 244 CX3CL1 _Hs.80420 chemokine (C-X3-C motif) ligand 1 Seq id 245 D123 Hs.412842 D123 gene product Seq id n 246 DCK H5.709 deoxycytidine kinase Seq id n 247 DDR1 Hs. 423573 discoidin domain receptor family, member 1 Seq id n 248 DDX11 Hs. 443960 DEAD / H (Asp-Glu-Ala-Asp / His) box polypeptide 11 (S.cerevisiae CHL1-like helicase) Seq id n 249 Hs.279604 desmin Seq id n 250 DHFR Hs. 83765 dihydrofolate reductase Seq id 251 DNMT1 Hs.202672 DNA (cytosine5 -) - methyltransferase 1 Seq id 252 E2-EPF Hs.396393 ubiquitin carrier protein Seq id 253 EBAG9 Hs.9222 estrogen receptor binding site associated, antigen, 9 Seq id n 254 ECT2 Hs.293257 strong similarity to mouse transforming protein ect2 Seq id n 255 EEFlE1 Hs.88977 eukaryotic translation elongation factor 1 epsilon 1 Seq id n 256 EGF Hs. 419815 epidermal growth factor (beta-urogastrone) Seq id 257 EIF3S4 Hs. 28081 eukaryotic translation initiation factor 3, subunit 4 (delta, 44kD) Seq id 258 ELF3 Hs.67928 E74-like factor 3 Seq id 259 ENDOG Hs. 420106 endonuclease G Seq id n 260 ENG H5.7675 endoglin (Osier-Rendu-Weber syndrome 1) Seq id n 261 SOX9 H5.2316 SRY (sex-determining region Y) -box 9 Seq id n 262 DC13 H5.6879 DC13 protein Seq id n 263 ERBB2 Hs. 446352 v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 2 (neuro / glioblastoma derived oncogene homolog) Seq id n 264 ESM1 Hs. 410668 endothelial cell-specific molecule 1 precursor Seq id 265 ESR1 H5.1657 estrogen receptor 1 Seq id n 266 ESR2 Hs. 443150 estrogen receptor 2 (ER beta) Seq id n 267 EZH2 Hs.444082 enhancer of zest (Drosophila) homolog 2 Seq id 268 FABP4 Hs.391561 fatty acid binding protein 4, adipocyte Seq id n 269 FBP 1 Hs.360509 fructose- bisphosphatase 1 Seq id n 270 FEN 1 Hs.409065 flap structure-specific endonuclease 1 Seq id n 271 FGFRI Hs.284244 fibroblast growth factor receptor 1 (fms-related tyrosine kinase 2, Pfeiffer syndrome) Seq id n 272 FHIT 506366 fragile histidine triad gene Seq id n 273 FHL1 Hs. 421383 four domains LIM 1 Seq id n 274 FOS Hs.25647 human cellular oncogene c-fos Seq id n 275 FXYD1 Hs.442498 FXYD domain- containing ion transport regulator 1 (phospholemman) Seq id n 276 GALNT3 Hs.278611 UDP -N-acetyl-alpha-D-galactosamine: polypeptide Nacetylgalactosaminyltransferase 3 (GaINAc-T3) Seq id n 277 GGH Hs. 78619 gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) Seq id n 278 GJA1 Hs.74471 gap junction protein alpha 1, 43 kD or connexin 43 Seq id n 279 GMPS Hs.144057 guanine monophosphate synthetase Seq id n 280 GNAI3 _Hs.73799 guanine nucleotide binding protein (g protein), alpha inhibiting activity polypeptide 3 Seq id n 281 GNAZ Hs.437081 guanine nucleotide binding protein, alpha z polypeptide; gnaz Seq id 282 GRIN2A Hs.125338 glutamate receptor, ionotropic, N-methyl D-aspartate 2A Seq id n 283 GRIN2C Hs. 436980 glutamate receptor, ionotropic, N-methyl D-aspartate 2C Seq id n 284 GSTM3 Hs.2006 glutathione s-transferase m3 (brain)

_

  Seq id n 285 GSTP1 Hs. 411509 glutathione S-transferase pi Seq id n 286 GZMA Hs.90708 granzyme A (granzyme 1, cytotoxic T-lymphocyte-associated serine esterase 3) Seq id n 287 HADHA Hs.75860 hydroxyacyl-Coenzyme A dehydrogenase / 3-ketoacyl-Coenzyme A thiolase / enoyl-Coenzyme A hydratase (trifunctional protein), subunit alpha Seq id n 288 HEC Hs. 414407 highly expressed in cancer, rich in leucine heptad repeats Seq id n 289 HGF Hs.396530 hepatocyte growth factor Seq id n 290 HLA-DQB1 Hs. 409934 major histocompatibility complex, clans II, DQ beta 1 Seq id n 291 HMG2 Hs.434953 high-mobility group (chromosomal nonhistone) protein 2 Seq id n 292 HMGIY Hs.57301 high mobility group (chromosomal nonhistone) protein isoforms I and Y Seq id n 293 HMMR Hs.72550 hyaluronan-mediated motility receptor (RHAMM) Seq id n 294 HNF3A Hs.163484 hepatocyte nuclear factor 3, alpha Seq id n 295 HPN Hs.432750 hepsin (transmembrane protease, serine 1) Seq id n 296 HRAS Hs.37003 v-Ha-ras Harvey rat sarcoma viral oncogene homolog Seq id n 297 HRB Hs.528387 HIV-1 Rev binding protein Seq id n 298 HSPC195 Hs.356509 hypothetical protein; ## EQU1 ## 302 IGFBP5 Hs.512226 insulin-like growth factor binding protein 5 Seq id 303 IGFBP6 Hs.274313 insulin-like growth factor binding protein 6; igfbp6 Seq id 304 IL10RA Hs.327 interleukin 10 receptor, alpha Seq id 305 IL 1A Hs.1722 interleukin 1, alpha Seq id 306 IL2RB Hs. 75596 interleukin 2 receptor, beta Seq id n 307 IL3RA Hs. 130058 interleukin 3 receptor, alpha (low affinity) Seq id n 308 IL6 Hs. 512234 interleukin 6 (interferon, beta 2) Seq id n 309 IL8 Hs. 624 interleukin 8 Seq id 310 ILF2 Hs.75117 interleukin enhancer binding factor 2 Seq id n 311 ING1 Hs.46700 inhibitor of growth 1 family, member 1 Seq id n 312 ING2 Hs.1179 inhibitor of growth family, member 2 Seq id n 313 INPP4B Hs.153687 inositol polyphosphate-4-phosphatase, type II, 105kD Seq id n 314 ITGA3 Hs.265829 integrin, alpha 3 (antigen CD49C, alpha3 subunit of VLA-3 receptor) Seq id n 315 ITGA7 Hs.74369 integrin , alpha 7 Seq id 316 ITGAL Hs.174103 integrin, alpha L (antigen CD11A (p180), lymphocyte function-associated antigen 1; alpha polypeptide) Seq id n 317 ITGB 1 Hs.287797 integrin, beta 1 (fibronectin receptor, beta polypeptide, antigen CD29 includes MDF2, MSK12) Seq id n 318 ITGB2 Hs. 375957 integrin, beta 2 (antigen CD18 (p95), lymphocyte function-associated antigen 1, macrophage antigen 1 (mac-1) beta subunit) Seq id n 319 ITGB4 Hs.85266 integrin, beta 4 Seq id n 320 ITGB8 Hs.355722 integrin, beta 8 Seq id n 321 KAI1 Hs. 323949 kangai 1 (suppression of tumorigenicity 6, prostate; CD82 antigen (R2 leukocyte antigen, antigen detected by monoclonal and antibody IA4)) Seq id n 322 KCNN2 Hs.98280 potassium intermediate / small conductance calcium-activated channel, subfamily N, member 2 Seq id n 323 KIAA0008 Hs. 77695 KIAA0008 gene product Seq id n 324 KIAA0042 Hs.3104 KIAA0042 gene product Seq id n 325 KIAA0074 Hs.308045 KIAA0074 protein Seq id n 326 KIAA0101 Hs.81892 KIAA0101 gene product Seq id n 327 KIAA0125 Hs. 38365 KIAA0125 gene product Seq id n 328 KIAA0159 Hs.5719 chromosome condensation-related SMC-associated protein 1 Seq id n 329 KIAA0758 Hs. 362806 KIAA0758 protein Seq id n 330 KIAA1442 Hs.274307 kiaal442 Seq id n 331 KIAAO601 Hs.348515 kiaa0601 Seq id n 332 KISS1 Hs.95008 kiss-1 metastasis suppressor Seq id n 333 KLK10 Hs.275464 kallikrein 10 Seq id n 334 KLK13 Hs .165296 kallikrein 13 Seq id n 335 KLK6 Hs.79361 kallikrein 6 (neurosin, zyme) Seq id n 336 KNTC1 Hs.300559 KIAA0166 gene product (kinetochore associated 1) Seq id n 337 KRT17 H5.2785 keratin 17 Seq id n 338 KRT19 Hs.309517 keratin 19 Seq id n 339 KRT5 Hs.433845 keratin 5 (epidermolysis bullosa simplex, Dowling-Meara / Kobner / Weber-Cockayne types) Seq id n 340 KRT7 Hs.23881 keratin 7 Seq id n 341 KRT8 Hs.356123 keratin 8 Seq id n 342 LAD 1 Hs.18141 Ladinin 1 Seq id n 343 LAMA3 Hs. 83450 laminin, alpha 3 (nicein (150kD), kalinin (165kD), BM600 (150kD), epilegrin) Seq id n 344 LAMC2 Hs.54451 laminin, gamma 2 (nicein (100kD), kalinin (105kD), BM600 (100kD) , Herlitz junctional epidermolysis bullosa)) Seq id n 345 LASP1 Hs.334851 LIM and HS3 protein 1 Seq id n 346 LCK H5. 1765 lymphocyte-specific protein tyrosine kinase Seq id n 347 LCN1 H5. 2099 lipocalin 1 (protein migrating faster than albumin, tear prealbumin) Seq id n 348 LCN2 Hs.204238 lipocalin 2 Seq id n 349 LDHB Hs. 234489 lactate dehydrogenase B Seq id n 350 LENG4 Hs.78768 breast and bladder overexpressed gene 1 Seq id n 351 LIV-1 Hs.79136 LIV-1 protein, estrogen regulated Seq id n 352 LPL Hs.180878 lipoprotein lipase Seq id n 353 LUM Hs.406475 lumican Seq id n 354 MAD2L1 Hs.79072 MAD2 (mitotic arrest deficient, yeast, homolog) -like 1 Seq id n 355 MAP2KIIP1 Hs. 433332 mitogen-activated protein kinase kinase 1 interacting protein 1 Seq id n 356 MCM2 Hs.57101 minichromosome maintenance deficient (S. cerevisiae) 2 (mitotin) Seq id n 357 MCM6 Hs.444118 mcm6 minichromosome maintenance deficient 6 mis5 homologs. pombe cerevisiae Seq id 358 MCM7 Hs.438720 mcm7 minichromosome maintenance deficient 7 s. Cerevisiae Seq id n 359 MGST1 Hs.389700 microsomal glutathione S-transferase 1 Seq id n 360 MKI67 Hs.80976 antigen identified by monoclonal antibody Ki-67 Seq id 361 MMP11 Hs.143751 matrix metalloproteinase 11 (stromelysin 3) Seq id n 362 MMP13 Hs.2936 matrix metalloproteinase 13 (collagenase 3) Seq id n 363 MMP14 Hs.2399 matrix metalloproteinase 14 (membrane-inserted) Seq id n 364 MMP9 Hs. 151738 matrix metalloproteinase 9 (gelatinase B, 92kD gelatinase, 92kD type IV collagenase) Seq id n 365 MSE55 Hs.148101 serum constitute protein Seq id n 366 MSH3 Hs.42674 mut S (E.coli) homolog 3 Seq id n 367 MTAP44 Hs .82316 interferon-induced, hepatitis C-associated microtubular aggregate protein (44kD) Seq id n 368 MUC1 Hs.89603 secreted epithelial tumor mucin antigen Seq id n 369 MX2 H5.926 myxovirus (influenza) resistance 2, homolog of murine Seq id n 370 MXI1 Hs. 118630 MAX-interacting protein 1 Seq id n 371 MYB Hs.407830 v-myb avian myeloblastosis viral oncogene homolog Seq id n 372 MYBL2 Hs.179718 v-myb avian myeloblastosis viral oncogene homolog-like 2 Seq id n 373 NAT1 Hs. 155956 N-acetyltransferase 1 (arylamine N-acetyltransferase) Seq id n 374 NCF1 H5.1583 neutrophil cytosolic factor 1 (47kD, chronic granulomatous disease, autosomal 1) Seq id n 375 NCOA3 Hs.382168 nuclear receptor coactivator 3 Seq id n 376 NDRG1 Hs.318567 N-myc downstream regulated Seq id n 377 NIFU Hs.350702 nitrogen fixation cluster-like Seq id n 378 NMU Hs.418367 neuromedin u Seq id n 379 NPM1 Hs. 411098 nucleophosmin Seq id n 380 NSEP1 Hs.74497 nuclease sensitive element binding protein 1 Seq id n 381 OSF2 Hs.122116 osteoblast specific factor 2 (fasciclin I-like) Seq id n 382 OXCT Hs.177584 3-oxoacid coa transferase Seq id n 383 P28 Hs.406050 dynein, axonemal, light intermediate polypeptide Seq id 384 P2RX4 Hs.321709 purinergic receptor P2X, ligand-gated ion channel, 4 Seq id n 385 PCNA Hs.78996 proliferating cell nuclear antigen Seq id n 386 PDGFRB Hs. 307783 platelet-derived growth factor receptor, beta polypeptide Seq id 387 PECAM1 Hs. 78146 platelet / endothelial cell adhesion molecule (CD31 antigen) Seq id n 388 PES1 Hs.132037 pescadillo (zebrafish) homolog 1, containing BRCT domain Seq id n 389 PFKP Hs.26010 phosphofructokinase, platelet Seq id n 390 PGK1 Hs.78771 phosphoglycerate kinase 1 Seq id No. 391 PGR H5. 2905 progesterone receptor Seq id n 392 PHYH Hs.172887 phytanoyl-coa hydroxylase precursor Seq id n 393 PIB5PA Hs.21492 Human phosphatidylinositol (4,5) bisphosphate 5-phosphatase mRNA homolog, partial cds Seq id n 394 PIG7 Hs.76507 LPS-induced TNF-alpha factor Seq id No. 395 PIP Hs.99949 prolactin-induced protein Seq id No. 396 PLA2G2A Hs. 76422 phospholipase A2, group IIA (platelets, synovial fluid) Seq id n 397 PLAK H5.2340 plakoglobin Seq id n 398 PLAT Hs.274404 plasminogen activator, tissue type isoform 1 preproprotein; dish Seq id n 399 PLG Hs. 143436 plasminogen Seq id 400 PLK Hs.329989 polo (Drosophia) -like kinase Seq id 401 PLSCR1 Hs.348478 phospholipid scramblase 1 Seq id 402 PMS2L3 Hs.278467 postmeiotic segregation increased 2-like 3 Seq id n 403 PMSCL1 Hs. 91728 polymyositis / scleroderma autoantigen 1 (75kD) Seq id n 404 POU2F2 Hs.1101 POU domain, class 2, transcription factor 2 Seq id n 405 PPPICB Hs.21537 proteinphosphatase 1, catalytic subunit, beta isoform Seq id n 406 PRC1 Hs.344037 protein regulator of cytokinesis 1 Seq id n 407 PRIM1 Hs.32916 primase, polypeptide 1 (49kD) Seq id n 408 PRL H5.1905 prolactin Seq id n 409 PRLR Hs.212892 prolactin receptor Seq id 410 PRSS8 Hs.75799 protease, serine , 8 (prostasin) Seq id n 411 PSMC1 Hs.356654 proteasome (prosome, macropain) 26S subunit, ATPase, 1 Seq id n 412 PTK6 Hs.51133 PTK6 protein tyrosine kinase 6 Seq id n 413 PTTG1 Hs.350966 pituitary tumor-transforming 1 Seq id n 414 PVALB Hs.295449 parvalbumin Seq id n 415 RAB6B Hs.352530 rab6b, member ras oncogene fami ly Seq id n 416 RAD51 Hs.446554 RAD51 (S. cerevisiae) homolog (E coli RecA homolog) Seq id n 417 RAD54L Hs. 66718 RAD54 (S.cerevisiae) -like Seq id n 418 RAP2A Hs.48554 RAP2A, member of RAS oncogene family Seq id n 419 RARRES 1 Hs.82547 retinoic acid receptor responder (tazarotene induced) 1 Seq id n 420 RBP4 Hs. 418083 retinol-binding protein 4, interstitial Seq id n 421 RERG Hs. 416854 RAS-like, estrogen-regulated, growth-inhibitor Seq id n 422 RFC4 Hs.35120 replication factor C (activator 1) 4 (37kD) Seq id n 423 RGS5 Hs.24950 regulator of G-protein signaling 5 Seq id n 424 RPL13 Hs.410817 ribosomal protein L13 Seq id n 425 RPL19 Hs. 381061 ribosomal protein L19 Seq id n 426 RRM1 Hs.383396 ribonucleotide reductase M1 polypeptide Seq id n 427 S100A2 Hs.515713 5100 calcium-binding protein A2 Seq id n 428 S100A4 Hs.81256 5100 calcium-binding protein A4 (calcium protein, calvasculin, metastasin, murine placental homolog) Seq id n 429 S 100A8 Hs.416073 S100 calcium binding protein A8 Seq id 430 S 100B Hs.422181 S100 calcium-binding protein, beta (neural) Seq id n 431 S 100P Hs.2962 S100 calcium binding protein P Seq id n 432 SCYA18 Hs.16530 small inducible cytokine subfamily A (Cys-Cys), member 18, pulmonary and activation-regulated Seq id n 433 SCYD1 Hs. 80420 small inducible cytokine subfamily D (Cys-X3-Cys), member 1 (fractalkine, neurotactin) Seq id n 434 SELENBP1 Hs.334841 selenium binding protein 1 Seq id n 435 SERFIA Hs.32567 small edrk-rich factor la, telomeric Seq id n 436 SERPINB5 Hs.55279 serine proteinase inhibitor, clade B, member 5 (maspin) Seq id n 437 SERPINC1 Hs.75599 serine (or cysteine) proteinase inhibitor, clade C (antithrombin), member 1 Seq id n 438 SERPINE1 Hs. 414795 serine (gold cysteine) proteinase inhibitor, clade E (nexin, plasminogen activator inhibitor type 1), member 1 Seq id n 439 SERPINI2 Hs.158308 serine (gold cysteine) proteinase inhibitor, clade I (neuroserpin), member 2 Seq id n 440 SFRP1 H5.7306 frizzled related protein 1 _secreted Seq id n 441 SLC2A3 Hs.419240 solute carrier family 2 (facilitated glucose transporter), member 3 Seq id n 442 SLC9A3R1 Hs. 396783 solute carrier family 9 (sodiurn / hydrogen exchanger), isoform 3 regulatory factor 1 Seq id n 443 SNCG Hs.349470 synuclein, gamma (breast cancer-specific protein 1) Seq id n 444 SNRPB2 Hs.280378 small nuclear ribonucleoprotein polypeptide B Seq id n 445 SOD3 H5.2420 superoxide dismutase 3, extracellular Seq id n 446 SPHAR Hs.296169 s-phase response (cyclin-related); sphar Seq id n 447 ST13 Hs.377199 progesterone receptorassociated p48 protein Seq id n 448 ST14 Hs.56937 suppression of tumorogenicity 14 Seq id n 449 ST7 Hs.203557 suppression of tumorogenicity 7 Seq id n 450 STAT1 Hs.21486 signal transducer and activator of transcription 1, 9lkD Seq id n 451 STK15 Hs. 250822 serine / threonine kinase 15 Seq id 452 TEK Hs.89640 TEK tyrosine kinase, endothelial (venous malformations, multiple cutaneous and mucosal) Seq id n 453 TFF1 Hs.350470 trefoil factor 1 (breast cancer, estrogen-inducible sequence expressed in) Seq id n 454 TFPI Hs.102301 tissue factor pathway inhibitor (lipoprotein-associated coagulation inhibitor) Seq id n 455 TGFA Hs.170009 transforming growth factor, alpha Seq id n 456 TGFB3 H5.2025 transforniing growth factor beta 3 Seq id n 457 THBS1 Hs.164226 thrombospondin 1 Seq id n 458 THY1 Hs.134643 Thy-1 cell antigen surface Seq id n 459 TIAF1 Hs.354085 TGFB1-induced anti-apoptotic factor 1 Seq id n 460 TIE Hs.78824 tyrosine kinase with inununoglobulin and epidermal growth factor homology domains Seq id n 461 TIMP1 Hs.446641 tissue inhibitor of metalloproteinase 1 (erythroid potentiating activity, collagenase inhibitor) Seq id n 462 TIMP2 Hs. 6441 tissue inhibitor of metalloproteinase 2 Seq id n 463 TIMP3 Hs. 245188 tissue inhibitor of metalloproteinase 3 (Sorsby fundus dystrophy, pseudoinflarnmatory) Seq id n 464 TNNI2 Hs.83760 troponin I, skeletal, fast Seq id n 465 TOP2A Hs.156346 topoisomerase (DNA) II alpha (1701d)) Seq id n 466 TOP2B Hs.282346 topoisomerase (DNA) II beta (180kD) Seq id n 467 TP53 Hs.408312 tumor protein p53 (Li-Fraumeni syndrome) Seq id n 468 TP53BP2 Hs.44585 tumor protein p53 binding 2 Seq id n 469 TPBG Hs. 82128 oncofetal trophoblast glycoprotein (5T4) Seq id n 470 TRD Hs.2014 T-cell receptor, delta (V, D, J, C) Seq id n 471 TROAP Hs.171955 trophinin associated protein (tastin) Seq id n 472 TSN Hs .75066 translin Seq id n 473 TYMS Hs.87491 thymidylate synthetase Seq id n 474 UGTREL1 Hs. 154073 UDP-galactose transporter related Seq id n 475 UNG Hs.78853 uracilDNA glycosylase Seq id n 476 VEGF Hs.73793 vascular endothelial growth factor Seq id n 477 VIM Hs.435800 vimentin Seq id n 478 VLDLR Hs. 370422 very low density lipoprotein receptor Seq id n 479 VWF Hs. 440848 von Willebrand factor Seq id n 480 XPOT Hs.85951 exportin, tRNA (nuclear export receptor for tRNAs) Seq id n 481 YARS Hs.322735 tyrosyltRNA synthetase Seq id n 482 ZNF 147 Hs.88914 zinc finger protein 147 (estrogen responsive finger protein) Seq id 483 ZNF22 Hs.108642 zinc finger protein 22 (kox 15)

TABLE C

  Sequence id Symbol Unigene ER + ER- Gene name Seq id n 186 AR H5.

99915 X androgen receptor (dihydrotestosterone receptor; testicular feminization; spinal and bulbar muscular atrophy; Kennedy disease) Seq id 190 BAG1 Hs.377484 X BCL2-associated athanogene Seq id 216 CDH13 H5.

63984 X cadherin 13, H-cadherin (heart) Seq idn 223 CEGP1 Hs.

435861 X cegpl protein Seq id n 227 CHI3L1 H5.382202 X chitinase 3-like 1 (cartilage glycoprotein-39) Seq id n 233 COX6C Hs.351875 X cytochrome c oxidase subunit vic cox6c nuclear encoding mitochondrial protein cytochrome c oxidase subunit Vic Seq id 236 CPB 1 Hs.

437028 X carboxypeptidase B1 (tissue) Seg id No. 237 CRIP1 H5.

70327 X cysteine-rich protein 1 (intestinal) Seq id n 238 CSDA H5.

221889 X cold shock domain protein A Segid n 245 D123 H5.412842 X D123 gene product Seq id n 261 SOX9 H5.2316 X SRY (sex determining region Y) - box 9 Seq id n 267 EZH2 Hs.444082 X enhancer of zest ( Drosophila) homolog 2 Seq id n 107 FLJ00382 Hs.439852 X Homo sapiens mRNA for FLJ00382 protein Seq id n 111 GDI2 H5.56845 X gdp dissociation inhibitor 2 Seq id n 290 HLA-DQB1 Hs.409934 X major histocompatibility complex, class II, DQ beta 1 Seq id 298 HSPC195 Hs.356509 X hypothetical protein; hspc195 Seq id 308 IL6 H5.512234 X Interleukin 6 (interferon, beta 2) Seq id n 124 K-ALPHA-1 _Hs.446608 X Alpha tubulin Seq id n 337 KRT17 H5.

2785 X keratin 17 Seq id 25 KRT18 Hs.406013 X keratin 18 Seq id n 340 KRT7 H5.23881 X keratin 7 _Seq id n 26 LDB2 Hs.4980 _ X LIM binding domain 2 Seq id n 351 LIV-1 H5.

79136 X LIV-1 protein, estrogen regulated Seq id n 33 MYC Hs.202453 X v-myc avian viral myelocytomatosis oncogene homolog Seq id n 373 NAT1 Hs.

155956 X N-acetyltransferase 1 (arylamine N-acetyltransferase) Seq id n 380 NSEP1 H5.74497 X nuclease sensitive element binding protein 1 Seq id n 387 PECAM1 Hs.78146 _ X platelet / endothelial cell adhesion molecule (CD31 antigen) Seq id n 391 PGR H5.2905 X progesterone receptor Seq id n 398 PLAT Hs.274404 X plasminogen activator, tissue type isoform 1 preproprotein; plate Seq id n 426 RRM1 Hs.383396 X ribonucleotide reductase M1 polypeptide Seq id 45 S 100A9 H5.112405 X S100 calcium-binding protein A9 (calanulin B) _Seq id n 152 SLPI Hs.251754 X secretory leukocyte protease inhibitor (antileukoproteinase) Seq id n 453 TFF1 Hs.350470 X trefoil factor 1 (breast cancer, estrogen-inducible sequence expressed in) _Seq id n 469 TPBG Hs.82128 X oncofetal trophoblast glycoprotein (5T4) Seq id n 72 TUBB Hs.512712 X tubulin, beta polypeptide Seq id 479 VWF Hs.440848 X von Willebrand factor

Claims (1)

53 Claims   1. Solid support coated with a set of representative fragments of genes whose expression profile for all of these genes in a subject with a mammary tumor allows the diagnosis or prognosis of the metastatic or localized nature of said tumor, characterized in that said set comprises at least 2 fragments, each of the 2 fragments being representative of a distinct gene chosen from the genes of Tables A1, A2 and A3.   2. Solid support according to claim 1, characterized in that the set comprises at least 50 fragments, each of the 50 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3.   3. Solid support according to claim 2, characterized in that the set comprises at least 100 fragments, each of the 100 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3.   4. Solid support according to claim 3, characterized in that the set comprises at least 150 fragments, each of the 150 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3.   5. Solid support according to claim 2, characterized in that the set comprises 72 fragments, each of the 72 fragments being representative of a distinct gene selected from the genes of Tables A1 and A2.   6. Solid support according to claim 5, characterized in that the 72 fragments are chosen from the following fragments: a) fragments of sequence SEQ ID N 1 to 55 for Table A1, and fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, or b) fragments having at least 70% identity with fragments as defined in a), or c) fragments comprising fragments as defined in a) or b), or d) fragments whose sequences are complementary to the sequences of the fragments as defined in a), b) or c).   7. Solid support according to claim 3, characterized in that the set comprises 111 fragments, each of the 111 fragments being representative of a distinct gene selected from the genes of Tables A2 and A3.   8. solid support according to claim 7, characterized in that the 111 fragments are chosen from the following fragments: a) fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for the Table A2, and the fragments of sequence SEQ ID N 74 to 167 for Table A3, or b) the fragments having at least 70% identity with the fragments as defined in a), or c) the fragments comprising the fragments as defined in a) or b), or d) fragments whose sequences are complementary to the sequences of the fragments as defined in a), b) or c).   9. Solid support according to claim 4, characterized in that the set comprises 166 fragments, each of the 166 fragments being representative of a distinct gene selected from the genes of Tables A1, A2 and A3.   10. Solid support according to claim 9, characterized in that the 166 fragments are chosen from the following fragments: a) the fragments of sequence SEQ ID N 1 to 55 for Table A1, the fragments of sequence SEQ ID N 56 to 70 , SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, and fragments of sequence SEQ ID N 74 to 167 for the Table A3, or   b) fragments having at least 70% identity with the fragments as defined in a), or e) fragments comprising fragments as defined in a) or b), or d) fragments whose sequences are complementary sequences fragments as defined in a), b) or c).   11. Solid support according to claim 9, characterized in that the set further comprises at least one representative fragment of a gene selected from the genes of Table B. 12. Solid support according to claim 11, characterized in that the set further comprises at least one representative gene fragment for all the genes of Table B. 13. Solid support according to one of claims 1 to 12, characterized in that at least one representative fragment of a gene selected from the genes of Table A1, A2, A3 or B are chosen from the following fragments: a) fragments of sequence SEQ ID N 1 to 55 for Table A1, fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, the fragments of sequence SEQ ID N 74 to 167 for Table A3, and the fragments of sequence SEQ ID N 168 to 192, SEQ ID N 193 or 194, SEQ ID N 195 to 201, SEQ ID N 202 or 203, and SEQ ID N 204 to 483 for Table B, or b) the fragments ay at least 70% identity with fragments as defined in a), or c) fragments comprising fragments as defined in a) or b), or d) fragments whose sequences are complementary to sequences of fragments as defined in a), b) or c).   14. solid support according to one of claims 1 to 12, characterized in that at least one representative gene fragment for all of the genes of Table A1, A2, A3 or B is chosen from the following fragments: the fragments of sequence SEQ ID N 1 to 55 for Table A1, the fragments of sequence SEQ ID N 56 to 70, SEQ ID N 71 or 72, and SEQ ID N 73 for Table A2, the fragments of sequence SEQ ID N 74 to 167 for Table A3, and the fragments of sequence SEQ ID N 168 to 192, SEQ ID N 193 or 194, SEQ ID N 195 to 201, SEQ ID N 202 or 203, and SEQ ID N 204 to 483 for the Table B, or b) fragments having at least 70% identity with fragments as defined in a), or c) fragments comprising fragments as defined in a) or b), or d) fragments whose sequences are complementary to the sequences of the fragments as defined in a), b) or c).   15. A method for diagnosing or prognosing in vitro the metastatic or localized character of a mammary tumor from a sample of said tumor in a subject, characterized in that said method comprises a step in which a sample of a nucleic acid extract, optionally after retro-transcription, from said tumor sample, on a solid support according to one of claims 1 to 14, and in that the amount of nucleic acid having hybridized with the set of representative fragments deposited on said support.   16. The method of claim 15, characterized in that it comprises the following steps: a) the extraction of a test nucleic acid sample from the tumor sample, b) the labeling of the nucleic acid test of the sample obtained in step a) using a first marker, c) in parallel, the marking of a reference nucleic acid sample with a second marker different from the first marker of step b), d) contacting the labeled test nucleic acid sample obtained in step b) and the labeled reference nucleic acid sample obtained in step c) with the carrier according to any one of claims 1 to 14 under conditions suitable for hybridization, each representative fragment of a solid support gene being capable of hybridizing with a fragment of the gene present in the sample of Nucleic acid labeled test and a fragment of the gene present in the acid sample reference nuclei; e) normalization, for each representative gene fragment deposited on the support, of the hybridization intensity obtained for the labeled test nucleic acid sample and that obtained for the nucleic acid sample of labeled reference, f) comparing, for each representative fragment of gene deposited on the support, the normalized hybridization intensity obtained for the labeled test nucleic acid sample with that obtained for the nucleic acid sample. labeled reference, so as to obtain a gene expression profile whose representative fragments have been deposited on the solid support, g) the classification of the mammary tumor as being metastatic or localized after analysis of the expression profile obtained at the step f).   17. Method according to claim 16, characterized in that the classification in step g) is determined by the following analysis: when the expression profile of step f) is obtained by means of the solid support according to one of claims 1, 2, 5 and 6, it is compared with the expression profile as defined in Tables A1 and A2; or - when the expression profile of step f) is obtained by means of the solid support according to one of claims 1 to 3, 7 and 8, it is compared with the expression profile as defined in Tables A2 and A3; or when the expression profile of step f) is obtained by means of the solid support according to one of claims 1 to 4, 9 and 10, it is compared with the expression profile as defined in Tables Al , A2 and A3.   18. The method of claim 16 also for characterizing whether or not the mammary tumor cells express the estrogen receptors, characterized in that after step f), the expression profile of the set of Table C genes obtained by means of the solid support according to one of claims 12 to 14, with the expression profile as defined in said Table C. 19. Process according to one of claims 15 to 18, characterized in that that the comparison in step f) of the normalized hybridization intensity obtained for the labeled test nucleic acid sample with that obtained for the labeled reference nucleic acid sample comprises the following steps: a) the calculating, for each representative fragment of a gene deposited on the support, the ratio between the normalized hybridization intensity obtained for the labeled test nucleic acid sample and that obtained for the nucleic acid sample ic labeled reference, b) calculating, for each representative fragment of a gene deposited on the support, the log2 of each report obtained in step a).   20. Method according to one of claims 15 to 19 characterized in that, when each fragment of the game is deposited three times on the support, the log2 average of the three replicates is calculated for each of these fragments.   21. Method according to one of claims 15 to 20, characterized in that the test and reference nucleic acid samples are RNA.   22. The method according to claim 21, characterized in that step b) of marking the test RNA of the sample with the aid of the first marker comprises the following steps: i) retrotranscription of the test RNA the sample obtained in step a), ii) the incorporation of the first marker during the in vitro transcription of the test DNA obtained in step i).   23. The method according to claim 21, characterized in that step c) of marking the reference RNA sample with the aid of the second marker comprises the following steps: i) the retrotranscription of the reference RNA of step c), ii) the incorporation of the second marker during the in vitro transcription of the test DNA obtained in step i).   24. Method according to one of claims 15 to 23, characterized in that the first and second markers are chosen from cyanine 3 and cyanine 5.   25. Kit comprising the solid support according to one of claims 1 to 14.