ES2475366B1 - Methods and kits for colorectal cancer prognosis - Google Patents

Abstract

Methods and kits for the prognosis of colorectal cancer. # The invention relates to methods for predicting the risk of relapse of patients with colorectal cancer, as well as methods for providing personalized medicine to said patients, based on levels of expression of the ACSL1, ACSL4 and, optionally, GCNT3, LPL, APOC1, SCD genes, whose expression products are involved in cellular energy metabolism. The invention also relates to kits for carrying out predictive and predictive medicine methods.

Description

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DESCRIPTION

METHODS AND KITS FOR THE FORECAST OF COLORRECTAL CANCER

FIELD OF THE INVENTION

The invention relates to the field of diagnosis and, more particularly, to methods for predicting the risk of relapse of cancer patients, as well as methods for providing personalized medicine to said patients. The invention also relates to kits for carrying out the methods of diagnostic and predictive medicine.

BACKGROUND OF THE INVENTION

Colorectal cancer (CRC) is one of the most common malignancies in the western world, it is the third leading cause of death in men, after lung cancer and prostate cancer and is the second most frequent among women, after cancer of breast Colorectal cancer is the third most common cancer in men (663,000 cases, 10.0% of the total) and the second in women (571,000 cases, 9.4% of the total) worldwide. Some 608,000 deaths from colorectal cancer are estimated worldwide, representing 8% of cancer deaths, making it the fourth most common cause of cancer death. (GLOBOCAN.iarc.fr) The main treatment option for colorectal cancer is surgery, with or without adjuvant chemotherapy and / or radiotherapy, depending on the patient's individual classification and other medical factors. The selection of an appropriate treatment is essential both for the patient and for economic reasons. For patient survival, it is essential to know when to immediately use a heavy and aggressive treatment protocol to prevent the spread of malignant colorectal cancer. Otherwise, patient survival may be compromised. On the contrary, the performance of a strong and aggressive treatment when it is not necessary is highly harmful for the patient. These treatments subject patients to a degree of discomfort and inconvenience arising from adverse toxicity that can significantly affect the patient's quality of life. It should be noted that each patient incurs a possibility of 1 in 400 that the treatment results in fatal toxicity. In addition, heavy and aggressive treatments are usually very expensive, and therefore should be performed only when necessary. Currently, treatment selection is based on tumor staging, which is usually performed with the American Joint Tumor / Nodules / Metastasis (TNM) test.

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Committee on Cancer (AJCC). The TNM system assigns a number based on three categories. "T" denotes the degree of invasion of the intestinal wall, "N" the degree of lymph node involvement, and "M" the degree of metastasis. The stage of a cancer is usually cited as a number I, II, III, IV derived from the value of TNM grouped by prognosis, where a higher number indicates a more advanced cancer and a worse outcome is likely. Although the AJCC classification provides valuable information about the stage in which colorectal cancer has been diagnosed, it does not give information about the aggressiveness of the tumor and its usefulness for prognosis is limited. Considering that it is clear that stage IV patients have a poor prognosis, the diagnosis of early colorectal cancer does not exclude the possibility that the tumor may develop very quickly. In particular, the causes for which 20 to 40 percent of patients with stage II colorectal cancer (early cancer with metastasis or lymph node invasion in diagnosis) are rapidly aggravated and die are unknown. Some studies suggest that a subgroup of patients at high risk for stage II colon cancer could benefit from adjuvant therapy (Quasar et al, Lancet 2007; 370: 2020-2029). However, histopathological variables, such as high-risk characteristics in stage II disease, are only guidelines in staging therapy. When the lymph nodes are invaded by the tumor cells, the results of the TNM tests predict a poor prognosis and the patient is usually subjected to surgery followed by heavy chemotherapy. Clinical studies show that for every 25 patients identified as high-risk stage II CCR, 20 are cured, regardless of whether or not they receive treatment (Quasar et al, Lancet 2007; 370: 2020-2029). Similarly, a subgroup of patients with stage III colon cancer treated only with surgery do not relapse in 5 years, even without adjuvant treatment (Ranghammar et al, Acta Oncologica 2001; 40: 282-308). Adjuvant chemotherapy is the standard recommendation for phase III CRC, however, the prospective identification of this subgroup of patients with stage III colon cancer could cause therapy to be dispensed with. Therefore, an accurate and reliable method that identifies patients at a higher and lower risk (for example, "high risk" stage II and "low risk" stage III colon cancer) could improve selection of individualized therapy within these groups. For this reason, several methods have been described to predict the outcome of patients suffering from colorectal cancer based on the levels of molecular marker expression. US7695913 describes a method to predict the prognosis of a patient suffering from CRC that comprises determining the levels of normalized expression of genes

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INHBA, MYBL2, FAP and Ki67 in which an increase in the expression of INHBA and FAP correlates negatively with a greater probability of a positive prognosis and where the expression of the MYBL2 and Ki67 genes correlates positively with a greater possibility of a prognosis positive. The method described in this document forms the basis of the Oncotype DX kit, although the kit includes the determination of 12 genes, including the INHBA, MYBL2, FAP and Ki67 genes. WO02057787 reports the results of a study designed to determine whether survivin mRNA can be used to predict death from a recurrent colorectal carcinoma. The study was based on data obtained from biopsies of frozen tumors from 144 patients. The study shows that survivin expression is associated with a significantly higher risk of death from recurrent cancer in patients with stage II colorectal cancer. Rosati et al. (Tumor Biol. 2004, 25: 258-63) reports the results of a study designed to determine whether the expression of thymidylate synthase (TS), p53, bcl-2, Ki-67 and p27 proteins in colorectal adenocarcinoma is capable of predicting survival without relapses or overall survival. Samples of 103 patients were examined by immunohistochemistry. According to this reference, there is no statistically significant association between the expression of any of the proteins TS, p53, bcl-2, Ki-67 and p27 and a clinical outcome although a statistically significant association was observed between an unfavorable outcome and a combination of negative expression of p53 and a positive expression of Ki-67 and stage C cancers. However, despite research on this topic, today there are very few tumor markers that are useful from the point of clinical view, both for the diagnosis of CRC and for determining the status of a CRC carcinoma. A test capable of quantifying the probability of the patient benefiting from stage III chemotherapy would be very useful. A stage III patient with a low risk of recurrence similar to that of a stage II patient and a low probability of benefiting from chemotherapy may choose to give up chemotherapy. A patient with a high risk of recurrence and a low probability of benefiting from chemotherapy with 5-FU may choose an alternative treatment. Therefore, there is an unmet need in the field of markers or marker panels that allow the diagnosis of CRC and the classification of colorectal carcinomas stage with high reliability. Therefore, an accurate and reliable method that identifies patients in a situation of greater and lesser risk (for example, of "high risk" in stage II and of "low risk" in

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stage III colon cancer) could improve the selection of individualized therapy within these groups.

BRIEF DESCRIPTION OF THE INVENTION

In a first aspect, the invention relates to a method for determining the prognosis of a patient suffering from colorectal cancer which comprises determining in a sample obtained from said patient the expression levels of at least one gene selected from the group formed by GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 where an increase in the expression levels of the LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes and / or a decrease in GCNT3 gene expression levels with respect to A reference value for each gene is indicative that the patient has a poor prognosis. In another aspect the invention relates to a method for selecting a suitable treatment for patients suffering from colorectal cancer comprising the determination of the expression levels of at least one gene selected from the group consisting of GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 where an increase in the expression levels of the LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes and / or a decrease in GCNT3 gene expression levels with respect to a reference value for each gene is Indicative that the patient has to be treated is a candidate for radiotherapy or chemotherapy after surgical treatment. In another aspect, the invention relates to a kit comprising the reagents suitable for the determination of the expression levels of the GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes and, optionally, the reagents for the determination of the expression levels of one or more control genes, constitutive expression or "housekeeping". In another aspect, the invention relates to the use of a kit according to the invention to predict the outcome (prognosis) of a patient suffering from colorectal cancer or to determine if a patient with colorectal cancer is a candidate for chemotherapy or radiotherapy. after surgery.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1: Prognostic value of the LPL gene in clinical samples of patients with colon cancer. Figure 2: Prognostic value of the APOC1 gene in clinical samples of patients with colon cancer.

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Figure 3: Prognostic value of the SCD gene in clinical samples of patients with colon cancer. Figure 4: Prognostic value of the ACSL1 gene in clinical samples of patients with colon cancer. ) Prognostic value of ACSL1 in the initial series of patients. B) Predictive value of response to chemotherapy of ACSL1 in patients of the initial series. C) Predictive value of response to chemotherapy of ACSL1 in the validation series. Figure 5: A) Prognostic value of the ACSL4 gene in clinical samples of patients with colon cancer. B) Prognostic value of the ACSL4 gene in clinical samples of patients with colon cancer who have not been treated with chemotherapy. Figure 6: Prognostic value of the GCNT3 gene in clinical samples of patients with colon cancer. Figure 7: Prognostic value of a metabolic footprint based on the determination of combined gene levels. The predictive value of a genomic fingerprint based on the determination of 4 (A) and 5 genes (B) is presented, both in the initial series (upper panel) and in the validation series (central panel), as well as the sensitivity commitment and specificity of the test (lower panel).

DETAILED DESCRIPTION OF THE INVENTION

Prognostic method of the invention

The authors of the present invention have identified different genes whose expression levels allow the identification of patients with CRC with lower and higher risk of relapse (for example, of "high risk" in stage II and of "low risk" in stage III of colon cancer). For example, as shown in examples 1 to 5 of the invention, the expression levels of the LPL, APOC1, SCD, ACSL1, ACSL4 and GCNT3 genes are differentially expressed in tumor samples from patients having a worse prognosis (determined through disease-free survival) than in those patients who have a better prognosis. Therefore, in a first aspect, the invention relates to a method (hereinafter "prognostic method of the invention") for predicting the outcome of a patient suffering from colorectal cancer comprising determining the levels of expression of the GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes where an increase in the expression levels of the LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes and / or a decrease in GCNT3 gene expression levels With respect to a reference value for each gene it is indicative that the patient has a poor prognosis.

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The term "determine the prognosis" is used here to refer to the probability that a patient has a specific clinical outcome, whether positive or negative. The prediction methods of the present invention can be used clinically to make decisions about choosing the most appropriate treatment for each particular patient. The prediction methods of the present invention are valuable tools for predicting whether a patient will respond favorably to a treatment regimen, such as chemotherapy. The prediction may include prognostic factors. As experts in the field will understand, the prediction, although preferred, does not have to be correct for 100% of the subjects that can be diagnosed or evaluated. The term, however, requires that a significant part of the subjects can be identified as most likely to have a certain outcome. If a subject is statistically significant it can be determined without further ado by the person skilled in the art, using different known statistical evaluation tools, for example, the determination of confidence intervals, the determination of p-value, cross-validation classification rates and details, etc., as shown in Dowdy and Wearden, Wiley Research Statistics, John & Sons, New York 1983. Recommended confidence intervals are at least 50%, at least 60%, at least 70%, at least 80%, at least 90% or at least 95%. The p-values are preferably 0.01, 0.005 or less. The term "patient", as used herein, refers to all animals classified as mammals and includes, but is not limited to, domestic and farm animals, primates and humans, for example: humans, non-primates. humans, cows, horses, pigs, sheep, goats, dogs, cats or rodents. Preferably, the patient is a human being male or female of any age or race. The term "colorectal cancer" is used in its broadest sense and refers to (1) all stages and all forms of cancer that originate from epithelial cells of the large intestine and / or rectum and / or (2) all stages and all forms of cancer that affect the lining of the large intestine and / or rectum. In staging systems for the classification of colorectal cancer, the colon and rectum are treated as a single organ. In the preferred embodiment, the patient has a stage I, II, III or IV, where stage I is defined as T1 N0 M0 or T2 N0 M0, stage II is defined as T3 N0 M0 or T4 N0 M0, the stage III is defined as any T, N1-2, M0 and stage IV corresponds to any T, any N, M1. According to the tumor / node / metastasis (TNM) staging system of the American Joint Committee on Cancer (AJCC) (Greene et al. (Eds.), AJCC cancer

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Manual Staging 6th ed. New York, NY: Springer, 2002), the different stages of colorectal cancer are defined as follows:

-

Tumor: T1: the tumor invades the submucosa, T2: the tumor invades the muscularis

propria, T3: the tumor invades the muscularis propria in the subserosa, or the tissues

pericholics

or perirectal; T4: The tumor invades directly others organs or

structures, and / or perforates.

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Node: N0: There is no metastasis to regional lymph nodes; N1: metastasis

in 1 to 3 regional lymph nodes, N2: Metastasis in 4 or more nodes

regional lymphatics

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Metastasis: M0: Distant metastasis mp; M1: distant metastasis present.

In a preferred embodiment, the patient whose outcome is predicted is a patient who has been diagnosed with colorectal cancer and who has undergone a surgical intervention of the cancer. In a preferred embodiment, the patient has had a surgical intervention of a stage I tumor, a stage II tumor, a stage III tumor or a stage IV tumor. In the present invention, the term "sample" or "biological sample" refers to the biological material isolated from a subject. The biological sample may contain any biological material suitable for detecting the desired biomarker and may comprise cells and / or non-cellular material of the subject. The sample may be isolated from any suitable biological tissue or fluid, such as, for example, prostate tissue, blood, blood plasma, serum, urine, cerebrospinal fluid (CSF) or feces. The samples used for the determination of the marker genes are samples of colon tissue preferably obtained by biopsy. On the other hand, the samples are samples of biofluids. The terms "biological fluid" and "biofluid" are used interchangeably herein and refer to aqueous fluids of biological origin. The biofluid can be obtained from any location (such as blood, plasma, serum, urine, bile, cerebrospinal fluid, vitreous or aqueous humor, or any body secretion), an exudate (such as fluid obtained from an abscess or any other site of infection or inflammation), or the fluid obtained from a joint (for example, a normal joint or a joint affected by a disease such as rheumatoid arthritis). In a first step, the first method of the invention comprises determining the expression levels of at least one gene selected from the group consisting of GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 in a sample of said patient.

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The term "GCNT3", as used herein, refers to the gene encoding mucosal glucosaminyl (Nacethyl) transferase 3, also known as C2 / 4GnT, C2GnT-M, C2GnT2 and identified in the UniProtKB / SwissProt database with Accession number O95395. The human GCNT3 gene has accession number 4205 in the HGNC database. The term "LPL", as used herein, refers to the gene encoding the lipoprotein lipase protein, P06858. The human LPL gene has accession number 6677 in the HGNC database. The term "APOC1", as used herein, refers to the gene encoding ApoC1 protein, and which is identified in the UniProtKB / SwissProt database with the access number P02654. The human APOC1 gene has accession number 607 in the HGNC database. The term "SCD", as used herein, refers to the gene encoding the stearoyl-CoA desaturase protein (also called delta-9 desaturase) and which is identified in the UniProtKB / SwissProt database with the accession number O00767. The human SCD gene has accession number 1057 in the HGNC database. The term "ACLS1", as used herein, refers to the gene encoding isoform 1 of the long chain acyl-CoA synthetase and which is identified in the UniProtKB / SwissProt database with accession number P33121. The human ACLS1 gene has accession number 3569 in the HGNC database. The term "ACLS4", as used herein, refers to the gene encoding isoform 4 of the long chain acyl-CoA synthetase and which is identified in the UniProtKB / SwissProt database with accession number O60488. The human ACLS4 gene has accession number 3571 in the HGNC database. It will be understood that the method according to the present invention may comprise the determination of naturally occurring polymorphic variants of one or more of the genes above. In a preferred embodiment, the method of the invention comprises determining the expression levels of each of the genes individually. In another preferred embodiment, the method of the invention comprises determining the expression levels of the genes grouped in pairs. Thus, the invention comprises the determination of the following GCNT3 and LPL gene pairs; GCNT3 and APOC1; GCNT3 and SCD; GCNT3 and ACSL1; GCNT3 and ACSL4; LPL and APOC1; LPL and SCD; LPL and ACSL1; LPL and ACSL4; APOC1 and SCD; APOC1 and ACSL1; APOC1 and ACSL4; SCD and ACSL1; SCD and ACSL4; ACSL1 and ACSL4. In another preferred embodiment, the method of the invention comprises determining the expression levels of the genes grouped three by three. So, the

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The invention comprises the determination of the following gene groups: GCNT3, LPL and APOC1; GCNT3, LPL and SCD; GCNT3, LPL and ACSL1; GCNT3, LPL and ACSL4; GCNT3, APOC1 and SCD; GCNT3, APOC1 and ACSL1; GCNT3, APOC1 and ACSL4; GCNT3, SCD and ACSL1; GCNT3, SCD and ACSL4; GCNT3, ACSL1 and ACSL4; LPL, APOC1 and SCD; LPL, APOC1 and ACSL1; LPL, APOC1 and ACSL4; LPL, SCD and ACSL1; LPL, SCD and ACSL4; LPL, ACSL1 and ACSL4; APOC1, SCD and ACSL1; APOC1, SCD and ACSL4; APOC1, ACSL1 and ACSL4; SCD, ACSL1 and ACSL4. In another preferred embodiment, the method of the invention comprises determining the expression levels of the genes grouped four by four. Thus, the invention comprises the determination of the following gene groups: GCNT3, LPL, APOC1 and SCD; GCNT3, LPL, APOC1 and ACSL1; GCNT3, LPL, APOC1 and ACSL4; GCNT3, LPL, SCD and ACSL1; GCNT3, LPL, SCD and ACSL4; GCNT3, LPL, ACSL1 and ACSL4; GCNT3, APOC1, SCD and ACSL1; GCNT3, APOC1, SCD and ACSL4; GCNT3, APOC1, ACSL1 and ACSL4; GCNT3, SCD, ACSL1 and ACSL4; LPL, APOC1, SCD and ACSL1; LPL, APOC1, SCD and ACSL4; LPL, APOC1, ACSL1 and ACSL4; LPL, SCD, ACSL1 and ACSL4; APOC1, SCD, ACSL1 and ACSL4. In another preferred embodiment, the method of the invention comprises determining the expression levels of the genes grouped five by five. Thus, the invention comprises the determination of the following gene groups: GCNT3, LPL, APOC1, SCD, and ACSL1; GCNT3, LPL, APOC1, SCD and ACSL4; GCNT3, LPL, APOC1, ACSL1 and ACSL4; GCNT3, LPL, SCD, ACSL1 and ACSL4; GCNT3, APOC1, SCD, ACSL1 and ACSL4; LPL, APOC1, SCD, ACSL1 and ACSL4. In another preferred embodiment, the method of the invention comprises determining the expression levels of the GCNT3, LPL, APOC1, SCD, ACSL1 and ACSL4 genes. In another preferred embodiment, the method of the invention further comprises determining the expression levels of one or more of one of the ABCA1, APOE, FADS2, CD36 genes where an increase in the expression levels of the ABCA1 genes , APOE, FADS2 and / or CD36 with respect to a reference value for each gene is indicative that the patient has a poor prognosis. The term "ABCA1", as used herein, refers to the gene encoding the ABCA1 protein (ATP-binding cassette subfmaliy A member 1), which is identified in the UniProtKB / SwissProt database with accession number O95477. The human ABCA1 gene has accession number 29 in the HGNC database.

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The term "APOE", as used herein, refers to the gene encoding the apolipoprotein E or Apo-E protein, which is identified in the UniProtKB / SwissProt database with the accession number P02649. The human APOE gene has accession number 613 in the HGNC database.

5 The term "FADS2", as used herein, refers to the gene that encodes the "delta (6) fatty acid desaturase" or "fatty acid desaturase 2" protein, which is identified in the UniProtKB / SwissProt database with the Accession number O95864. The human FADS2 gene has accession number 3575 in the HGNC database. The term "CD36", as used herein, refers to the gene that encodes the platelet protein.

10 glycoprotein 4 (thrombospondine receptor) ”, which is identified in the UniProtKB / SwissProt database with the UniProtKB / Swiss-Prot accession number: P16671. The human CD36 gene has the access number in the HGNC database. In a preferred embodiment, the method of the invention comprises determining the expression levels of one of the following gene groups:

15 -ACSL4, SCD, APOE and FADS2; -ACSL4, SCD, ABCA1 and FADS2; -ACSL4, SCD, ABCA1 and APOC1; -ACSL4, ACSL1, SCD and ABCA1; -ACSL4, ACSL1, SCD, FADS2 and ABCA1

20 -ACSL4, APOE, SCD, FADS2 and ABCA1 -ACSL4, APOE, SCD, FADS2 and CD36 -ACSL4, SCD, FADS2, ABCA1 and CD36

Virtually any conventional method can be used within the framework of the

Invention for detecting and quantifying the levels of said genetic markers. By way of non-limiting example, expression levels are determined by quantifying the levels of mRNA encoded by the genes involved or by quantifying the levels of the corresponding proteins. Methods for determining the amount of mRNA are well known in the art. By

For example, the nucleic acid containing the sample (for example, cells or tissues prepared from the patient) is first extracted according to standard methods, for example, the use of lithic enzymes or chemical solutions or extracted with nucleic acids fixing resins following the manufacturer's instructions. The extracted mRNA is detected by hybridization (for example, Northern blot analysis or DNA microarrays after

35 mRNA conversion into a labeled cDNA) and / or amplification (eg, RT-PCR). The

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Determination can be carried out qualitatively, quantitatively, or semi-quantitatively. The quantitative real-time or semi-quantitative determination of RT-PCR is particularly advantageous. Preferably, the primer pairs are designed to include a region that contains an intron, so that cDNA amplification can be distinguished from putative genomic contamination. Suitable primers can be easily designed by the expert. Other amplification methods include ligase chain reaction (LCR), amplification mediated transcription (TMA), chain shift amplification (SDA) and nucleic acid based sequence amplification (NASBA). Preferably, the amount of mRNA is measured quantitatively or semi-quantitatively from RT-PCR or quantitative in real time or semi-quantitative from RT-PCR. The term "specific hybridization", as used herein, refers to the conditions that allow the hybridization of two polynucleotide sequences under high stringent conditions or moderately severe conditions. The terms "highly strict conditions" and "moderately strict conditions" are defined below in relation to the kit of the invention and are equally applicable in the context of the present method. On the other hand, it is also possible to determine the expression levels of the marker genes by determining the expression levels of the proteins encoded by said genes, an increase in the amount of corresponding protein must take place. The expression levels of different proteins can be determined using any conventional method. By way of non-limiting example, said determination can be made using antibodies capable of specifically binding the protein to be determined (or fragments thereof with the antigenic determinants) and subsequent quantification of the antigen-antibody complex derivatives. The antibodies to be used in this type of analysis can be, for example polyclonal sera, hybridoma supernatants or monoclonal antibodies, antibody fragments, Fv, Fab, Fab 'and F (ab') 2, scFv, diabodies, triabodies , humanized tetrabodies and antibodies. At the same time, the antibodies may or may not be labeled. Illustrative, but not exclusive, examples of markers that can be used include radioactive isotopes, enzymes, fluorophores, chemiluminescent reagents, enzyme cofactors or substrates, enzyme inhibitors, particles, dyes, etc. There are a wide variety of tests well known that can be used in the present invention, labeled antibodies (primary antibody) and labeled antibodies (secondary antibodies), which include Western-blot or immunoblot techniques, ELISA (Enzyme-linked immunosorbent assay), RIA ( radioimmunoassay), competition EIA (enzyme immunoassay), DAS-ELISA (double sandwich antibodies

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ELISA), immunocytochemical and immunohistochemical techniques, techniques based on the use of biochips or microarrays of proteins as specific antibodies or assays based on colloidal precipitation in formats such as test strips. Other forms of protein detection and quantification include affinity chromatography, ligand binding assays, etc. Once the expression levels of the genes above a sample of a patient have been determined, the levels are compared with reference values for each of said genes. Normally, the reference values are the level of expression of the genes that are compared in a reference sample. A "reference sample", as used herein, means a sample obtained from a group of healthy subjects that does not have a particular disease state or phenotype. For example, the reference sample may include samples of the colon mucosa of patients who do not suffer from colon cancer or who have no history of colon cancer. On the other hand, the reference sample could be a sample or a set of colon cancer samples with a low risk of recurrence. This sample or set of samples can be obtained from patients who have had a surgical resection of the tumor and who have not suffered a relapse, preferably in the absence of adjuvant chemotherapy. In another embodiment, the reference sample is a sample of a type that CCR or a group of type I CCR. The appropriate reference levels of gene expression can be determined by measuring the expression levels of such genes on various suitable topics, and those reference levels can be adjusted to specific populations (for example, a reference level may be age-related, so comparisons can be made between levels of expression in samples of subjects of a certain age and reference levels for a particular disease, phenotype, or lack thereof in a given age group ). In a preferred embodiment, the reference sample is obtained from several healthy subjects or patients without a previous history of colorectal cancer. On the other hand, the reference sample is a sample or a set of colon cancer samples from patients who have had a surgical resection of the tumor and who have not suffered a relapse, preferably in the absence of adjuvant chemotherapy. The person skilled in the art will appreciate that the type of reference sample may vary depending on the specific method to be performed. Thus, in the event that a diagnosis or prognosis of the disease is carried out, the sample referred to may be a set of non-tumor tissue of colorectal tissue, that of people who do not have a history of colorectal cancer or of a distal group of non-tumor tissues with respect to

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the respective tumor tissues, or a sample or a set of colon cancer samples from patients who have had a surgical resection of the tumor and who have not suffered a relapse, preferably in the absence of adjuvant chemotherapy. In the event that the method of the invention is directed to determine the effect of a therapy on a patient, the reference sample is preferable a sample obtained from said patient before starting treatment. The expression profile of the genes in the preference reference sample can be generated from a population of two or more people. The population, for example, can contain 3, 4, 5, 10, 15, 20, 30, 40, 50 or more people. In addition, the expression profile of the genes in the reference sample and in the sample of the person to be diagnosed according to the methods of the present invention can be generated from the same person, provided the profiles are analyzed and the reference profile are generated from the biological samples taken at different times and compared to each other. For example, a sample of an individual can be obtained at the beginning of a study period. A reference biological marker profile of this sample can be compared with the biomarker profiles generated from subsequent samples of the same person. In a preferred embodiment, the reference sample is a set of samples from several individuals, and corresponds to portions of colon tissue that are far from the tumor area and that have been preferably obtained in the same biopsy, but which do not have No pathological characteristics of the tumor tissues. Once the expression levels of the marker genes in relation to the reference values for said genes have been determined, it is necessary to identify whether there are alterations in the expression of said genes (increase or decrease in expression). The expression of a gene is considered increased in a sample of the subject matter when the levels of increase with respect to the reference sample are at least 5%, at least 10%, at least 15%, by at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55 %, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90 %, at least 95%, at least 100%, at least 110%, at least 120%, at least 130%, at least 140%, at least 150%, or more. Similarly, the expression of a gene is considered decreased when its levels decrease with respect to the reference sample by at least 5%, at least 10%, at least 15%, at least 20%, by at least 25%, at least 30%, at least

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35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 100% (i.e. absent). Finally, the patient is classified as having a high risk of negative result, if the marker genes show an increase in expression levels with respect to a reference sample and that have a low risk of negative results, if the marker genes show a decrease in expression levels with respect to a reference sample. In a preferred embodiment, the patient is classified as having a high risk of negative result, if the expression levels of the gene is higher than the expression level of the same gene in a sample or set of colon cancer samples from patients who have had a surgical resection of the tumor and have not suffered a relapse, preferably in the absence of adjuvant chemotherapy. The term "good prognosis", as used in the present invention, is used to indicate an improvement in some measure of the patient's condition, including those normally used in the art, such as an increase in the duration of the free interval of recurrence (RFI), an increase in overall survival time (OS), an increase in disease-free survival time (DFS), an increase in the duration of recurrence-free interval (DRFI), and the like . An increase in the probability of positive clinical results corresponds to a decrease in the probability of cancer recurrence. The term "poor prognosis", as used in the present invention, is used to indicate a worsening of any measure of the patient's condition, even normally used in the art, such as a decrease in the duration of the recurrence free interval. (RFI), a decrease in overall survival time (OS), a decrease in disease-free survival time (DFS), a decrease in the duration of recurrence-free interval (DRFI), and the like. An increase in the probability that the negative clinical outcome corresponds to an increase in the probability of cancer recurrence In a preferred modality, the outcome in a given patient is measured as disease-free survival. The term "disease-free survival ", as used herein, refers to the time elapsed between primary cancer treatment and the time in which the patient survives without the appearance of signs or symptoms Ace of disease.

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The prognostic factors are those variables related to the natural history of colorectal cancer, which influence the recurrence and prognosis rates of patients once they have developed colorectal cancer. Clinical parameters that have been associated with a worse prognosis, for example, lymph node involvement and high-grade tumors. Prognostic factors are frequently used to categorize patients into subgroups with different baseline relapse risks. In a preferred embodiment, the clinical prognostic factor used in the method of the invention is the stage of the tumor, a stage in which the increase in the tumor is indicative of an increased risk of recurrence or in which the tumor stage decreases is indicative that the patient shows a low risk of recurrence. Patients analyzed according to the present invention may have been treated with one or more therapies aimed at decreasing the size of the tumor. Therefore, in a preferred embodiment, the patients are treated before the determination of the expression levels of the different genes according to the invention with a therapy selected from the group consisting of chemotherapy, radiotherapy or surgery. The terms "chemotherapy", "Radiotherapy" and "surgery" are defined below in detail and used with the same meaning in the context of the present invention. In a preferred embodiment, patients have been treated with a fluoropyridine-based therapy. The term "fluoropyridine-based therapy", as used in the present invention, refers to a therapy comprising a fluoropyridine as an active ingredient, and includes both therapies where fluoropyridine is the only active ingredient and compositions comprising a fluoropyridine and one or more active ingredients. The term "fluoropyridine", as used in the present invention, refers to a chemotherapy agent comprising a pyrimidine ring wherein said ring is substituted by a fluorine atom. Examples of fluorpyridines include, without limitation, 5-fluorouracil, floxuridine, tegafur, 5fluoro-2'-deoxyuridine, ftorafur, emitefur, eniluracil and capecitabine. Compositions comprising a fluoropyridine and one or more active ingredients include, without limitation, FOLFOX (a composition comprising folinic acid, 5-FU and oxaliplatin), XELOX (a composition comprising capecitabine and oxaliplatin), FOLFIRI (5-FU, leucovirin and irinotecan), S-1 (a combination of ftorafur (a prodrug of 5-FU) and 2 5-FU modulators called 5-chloro-2,4-dihydroxyproidine and oxonic acid at a molar ratio of 1: 0.4: 1 ). UFT (a combination of ftorafur and uracil in a 1: 4 molar ratio).

Custom therapy methods

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The prognostic methods defined above also offer personalized therapies for patients suffering from colorectal cancer. In particular, patients considered to have a poor prognosis are candidates for adjuvant therapy (after tumor removal). In contrast, patients who show a low risk of relapse can give up additional therapeutic treatment after surgery and avoid the unwanted effects of adjuvant therapy. Therefore, in another aspect, the invention relates to a method (hereinafter, the first personalized therapeutic method of the invention) for the selection of a suitable treatment for colorectal cancer in a patient comprising the determination of the levels of expression of at least one gene selected from the group consisting of GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 in a sample of said patient, where there is an increase in the expression levels of the LPL, APOC1, SCD, ACSL1 genes and / or ACSL4 and / or a decrease in GCNT3 gene expression levels with respect to a reference value for each gene is indicative that the patient has to be treated with adjuvant therapy. As used herein, "treatment" refers to clinical intervention in an attempt to prevent, cure, delay, reduce the severity of, or improve one or more symptoms of the disease or disorder or recurrent disease or disorder, or in order to prolong the survival of a patient beyond what is expected in the absence of such treatment. The term "colorectal cancer" has been described in detail in the context of the prognostic method of the invention and is used with the same meaning in the context of custom methods according to the invention. In a first step, the personalized therapy method according to the invention comprises determining the level of expression of GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 in a sample of said patient. The terms "colorectal cancer", "patient", "GCNT3 gene", "LPL gene", "APOC1 gene", "SCD gene", "ACSL1" gene, "ACSL4" gene, "expression levels", "sample" they have been described in detail above, and apply equally to the methods according to the current method. In a preferred embodiment, the personalized therapy method according to the invention comprises the determination of the expression levels of each of the genes individually. In another preferred embodiment, the personalized therapy method according to the invention comprises determining the expression levels of the genes grouped in pairs. Thus, the invention comprises the determination of the following GCNT3 and LPL gene pairs; GCNT3 and APOC1; GCNT3 and SCD; GCNT3 and ACSL1; GCNT3 and

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ACSL4; LPL and APOC1; LPL and SCD; LPL and ACSL1; LPL and ACSL4; APOC1 and SCD; APOC1 and ACSL1; APOC1 and ACSL4; SCD and ACSL1; SCD and ACSL4; ACSL1 and ACSL4. In another preferred embodiment, the personalized therapy method according to the invention comprises determining the expression levels of the genes grouped three by three. Thus, the invention comprises the determination of the following gene groups: GCNT3, LPL and APOC1; GCNT3, LPL and SCD; GCNT3, LPL and ACSL1; GCNT3, LPL and ACSL4; GCNT3, APOC1 and SCD; GCNT3, APOC1 and ACSL1; GCNT3, APOC1 and ACSL4; GCNT3, SCD and ACSL1; GCNT3, SCD and ACSL4; GCNT3, ACSL1 and ACSL4; LPL, APOC1 and SCD; LPL, APOC1 and ACSL1; LPL, APOC1 and ACSL4; LPL, SCD and ACSL1; LPL, SCD and ACSL4; LPL, ACSL1 and ACSL4; APOC1, SCD and ACSL1; APOC1, SCD and ACSL4; APOC1, ACSL1 and ACSL4; SCD, ACSL1 and ACSL4. In another preferred embodiment, the personalized therapy method according to the invention comprises the determination of expression levels of genes grouped four by four. Thus, the invention comprises the determination of the following gene groups: GCNT3, LPL, APOC1 and SCD; GCNT3, LPL, APOC1 and ACSL1; GCNT3, LPL, APOC1 and ACSL4; GCNT3, LPL, SCD and ACSL1; GCNT3, LPL, SCD and ACSL4; GCNT3, LPL, ACSL1 and ACSL4; GCNT3, APOC1, SCD and ACSL1; GCNT3, APOC1, SCD and ACSL4; GCNT3, APOC1, ACSL1 and ACSL4; GCNT3, SCD, ACSL1 and ACSL4; LPL, APOC1, SCD and ACSL1; LPL, APOC1, SCD and ACSL4; LPL, APOC1, ACSL1 and ACSL4; LPL, SCD, ACSL1 and ACSL4; APOC1, SCD, ACSL1 and ACSL4. In another preferred embodiment, the personalized therapy method according to the invention comprises determining the expression levels of the genes grouped five by five. Thus, the invention comprises the determination of the following gene groups: GCNT3, LPL, APOC1, SCD, and ACSL1; GCNT3, LPL, APOC1, SCD and ACSL4; GCNT3, LPL, APOC1, ACSL1 and ACSL4; GCNT3, LPL, SCD, ACSL1 and ACSL4; GCNT3, APOC1, SCD, ACSL1 and ACSL4; LPL, APOC1, SCD, ACSL1 and ACSL4. In another preferred embodiment, the personalized therapy method according to the invention comprises the determination of the expression levels of the GCNT3, LPL, APOC1, SCD, ACSL1 and ACSL4 genes. In another preferred embodiment, the method of the invention further comprises determining the expression levels of one or more of one of the ABCA1, APOE, FADS2, CD36 genes where an increase in the expression levels of the ABCA1 genes , APOE, FADS2 and / or CD36 with respect to a reference value for each gene is indicative that the patient has to be treated with adjuvant chemotherapy.

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The terms "ABCA1", "APOE", "FADS2" and "CD36" have been defined in the context of the first method of the invention. In a preferred embodiment, the method of the invention comprises determining the expression levels of one of the following gene groups:

-ACSL4, SCD, APOE and FADS2; -ACSL4, SCD, ABCA1 and FADS2; -ACSL4, SCD, ABCA1 and APOC1; -ACSL4, ACSL1, SCD and ABCA1. -ACSL4, ACSL1, SCD, FADS2 and ABCA1 -ACSL4, APOE, SCD, FADS2 and ABCA1 -ACSL4, APOE, SCD, FADS2 and CD36 -ACSL4, SCD, FADS2, ABCA1 and CD36

The expression levels of the different genes used in the first personalized therapeutic method of the invention can be determined by determining the levels of mRNA encoded by the genes involved or by determining the levels of the polypeptide encoded by the genes involved. In a second step, the method of assigning a personalized therapy according to the invention comprises the identification of those patients who present an increase in the expression levels of the LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes, alone or combined with an increase in the expression of ABCA1, APOE, FADS2, CD36 and / or a decrease in GCNT3 gene expression levels with respect to a reference value for such genes as candidates for adjuvant radiotherapy or chemotherapy after treatment Surgical or patient who shows decreased levels of gene expression with respect to a reference value as a patient who is not a candidate for radiotherapy or chemotherapy after surgery. The term "surgery", as used herein, means any therapeutic procedure that involves a methodical action of the hand or hand with an instrument, in the body of a human or other mammal, to produce healing or recovery. As the term "chemotherapy" is used herein, "chemotherapy drug" broadly refers to the use of a chemical drug or a combination of both for the treatment of cancer, tumors or malignant neoplasms, including both cytotoxic or cytostatic drugs. Examples of chemotherapy agents that may be in accordance with the present invention include:

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-

Alkylating agents (for example, mechlorethamine, chlorambucil, cyclophosphamide,

ifosfamide,

streptozocin, carmustine, lomustine, melphalan, busulfan,

dacarbazine, temozolomide, thiotepa or altretamine);

-

Platinum drugs (for example, cisplatin, carboplatin and oxaliplatin);

-

Antimetabolite drugs (for example, 5-fluorouracil, capecitabine, 6

mercaptopurine, methotrexate, gemcitabine, cytarabine, fludarabine or pemetrexed);

-

Anti-tumor antibiotics (for example, daunorubicin, doxorubicin, epirubicin,

idarubicin, actinomycin D, bleomycin, mitomycin C, or mitoxantrone);

-

Inhibitors from the mitosis (for example, paclitaxel, docetaxel, ixabepilone,

vinblastine, vincristine, vinorelbine, vindesine or estramustine), and

-

Topoisomerase inhibitors (e.g., etoposide, teniposide, topotecan,

irinotecan, or diflomotecan elomotecan).

The term "radiotherapy" is a term commonly used in the art to refer to various types of radiation therapy, including internal and external radiation therapies, or radioimmunotherapy, and the use of various types of radiation such as X-rays, gamma rays. , alpha particles, beta particles, photons, electrons, neutrons, radioisotopes, and other forms of ionizing radiation. In a preferred embodiment, the treatment is adjuvant or neoadjuvant chemotherapy. The term "neoadjuvant therapy", as used herein, refers to any type of cancer treatment given before surgical resection of the primary tumor, in a patient with a cancer. The most common reason for neoadjuvant therapy is to reduce the size of the tumor in order to facilitate more effective surgery. Neoadjuvant therapies include radiation therapy and, preferably, systemic therapy, such as hormonal therapy, chemotherapy, immunotherapy and monoclonal antibody therapy. The term "adjuvant therapy," as used herein, refers to any type of cancer treatment (for example, chemotherapy or radiotherapy) as an additional treatment, usually after surgical resection of the primary tumor, in a patient. with a cancer that is at risk of metastasis and / or likely to recur. The objective of this type of adjuvant treatment to improve the prognosis. Adjuvant therapies include radiation therapy and, preferably, systemic therapy, such as hormonal therapy, chemotherapy, immunotherapy and monoclonal antibody therapy. In a preferred embodiment, the therapy is a chemotherapy other than a fluoropyridine-based therapy. The terms "fluoropyridine-based therapy" and "fluoropyridine" have been described in detail in relation to the prognostic method of the invention and are used in the context of the present method with the same meaning.

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Kits of the invention

In another embodiment, the invention relates to a kit comprising suitable reagents for the determination of expression levels of at least one gene selected from the group consisting of GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4. In the context of the present invention, "kit" is understood as a product containing the different reagents necessary to carry out the methods of the invention adapted to allow transport and storage. Suitable materials for packing kit components include (polyethylene, polypropylene, polycarbonate and the like) of glass, plastic, bottles, jars, paper, envelopes, etc. In addition, the kits of the invention may contain instructions for the simultaneous, sequential or separate use of the different components found in the kit. Said instructions may be in the form of printed material or in the form of an electronic medium capable of storing instructions so that they can be read by a subject, such as electronic storage media (magnetic discs, tapes and the like), optical media (CD -ROM, DVD) and the like. Additionally or alternatively, the media may contain Internet addresses that offer such instructions. The term "reagent that allows to determine the level of expression of a gene" means a compound or group of compounds that allows to determine the level of expression of a gene, both by determining the level of mRNA and by determining the level of protein level. Therefore, reagents of the first type include probes capable of hybridizing specifically with the mRNA encoded by the genes involved. Reagents of the second type are compounds that specifically bind to proteins encoded by the marker genes and, preferably, antibodies are included, although they may be specific aptamers. In a preferred embodiment, the reagents suitable for the determination of expression levels of one or more genes comprise at least 10%, at least 20%, at least 30%, less 40%, at least 50%, less 60 %, at least 70%, at least 80%, at least 90% or less than 100% of the total amount of reagents suitable for determining the expression levels of the genes that make up the kit. Thus, in the particular case of kits comprising the reagents for the determination of the expression levels of the GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes, the specific reagents for said gene (for example, the probes that are able to hybridize under strict conditions with the mRNAs encoded by the GCNT3, LPL, APOC1, SCD, genes

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ACSL1 and / or ACSL4) comprise at least 10%, at least 20%, at least 30%, less 40%, at least 50%, less 60%, at least 70%, at least 80%, at least 90% or less 100% of the probes present in the kit. In other embodiments, suitable reagents for the determination of expression levels of one or more genes comprise at least 55% at least 60%, at least 65%, at least 70%, less 75%, 80 At least%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or less 99% of the total amount of reagents that make up the kit . In another preferred embodiment, the kit of the invention contains reagents suitable for the determination of at least two of the marker genes. Thus, the invention comprises kits comprising reagents suitable for the determination of the following GCNT3 and LPL gene pairs; GCNT3 and APOC1; GCNT3 and SCD; GCNT3 and ACSL1; GCNT3 and ACSL4; LPL and APOC1; LPL and SCD; LPL and ACSL1; LPL and ACSL4; APOC1 and SCD; APOC1 and ACSL1; APOC1 and ACSL4; SCD and ACSL1; SCD and ACSL4; ACSL1 and ACSL4. In another preferred embodiment, the kit of the invention contains reagents suitable for the determination of at least three of the marker genes. Thus, the invention comprises kits comprising suitable reagents for the determination of the following gene groups: GCNT3, LPL and APOC1; GCNT3, LPL and SCD; GCNT3, LPL and ACSL1; GCNT3, LPL and ACSL4; GCNT3, APOC1 and SCD; GCNT3, APOC1 and ACSL1; GCNT3, APOC1 and ACSL4; GCNT3, SCD and ACSL1; GCNT3, SCD and ACSL4; GCNT3, ACSL1 and ACSL4; LPL, APOC1 and SCD; LPL, APOC1 and ACSL1; LPL, APOC1 and ACSL4; LPL, SCD and ACSL1; LPL, SCD and ACSL4; LPL, ACSL1 and ACSL4; APOC1, SCD and ACSL1; APOC1, SCD and ACSL4; APOC1, ACSL1 and ACSL4; SCD, ACSL1 and ACSL4. In another preferred embodiment, the kit of the invention contains reagents suitable for the determination of at least four of the marker genes. Thus, the invention comprises kits comprising reagents suitable for the determination of the following gene groups: GCNT3, LPL, APOC1 and SCD; GCNT3, LPL, APOC1 and ACSL1; GCNT3, LPL, APOC1 and ACSL4; GCNT3, LPL, SCD and ACSL1; GCNT3, LPL, SCD and ACSL4; GCNT3, LPL, ACSL1 and ACSL4; GCNT3, APOC1, SCD and ACSL1; GCNT3, APOC1, SCD and ACSL4; GCNT3, APOC1, ACSL1 and ACSL4; GCNT3, SCD, ACSL1 and ACSL4; LPL, APOC1, SCD and ACSL1; LPL, APOC1, SCD and ACSL4; LPL, APOC1, ACSL1 and ACSL4; LPL, SCD, ACSL1 and ACSL4; APOC1, SCD, ACSL1 and ACSL4. In another preferred embodiment, the kit of the invention contains reagents suitable for the determination of at least five of the marker genes. Thus, the invention

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comprises kits comprising suitable reagents for the determination of the following gene groups: GCNT3, LPL, APOC1, SCD, and ACSL1; GCNT3, LPL, APOC1, SCD and ACSL4; GCNT3, LPL, APOC1, ACSL1 and ACSL4; GCNT3, LPL, SCD, ACSL1 and ACSL4; GCNT3, APOC1, SCD, ACSL1 and ACSL4; LPL, APOC1, SCD, ACSL1 and ACSL4. In another preferred embodiment, the kit of the invention contains reagents suitable for the determination of at least five of the marker genes. Thus, the invention comprises kits comprising suitable reagents for the determination of the following gene groups: GCNT3, LPL, APOC1, SCD, and ACSL1; GCNT3, LPL, APOC1, SCD and ACSL4; GCNT3, LPL, APOC1, ACSL1 and ACSL4; GCNT3, LPL, SCD, ACSL1 and ACSL4; GCNT3, APOC1, SCD, ACSL1 and ACSL4; LPL, APOC1, SCD, ACSL1 and ACSL4; In another preferred embodiment, the kit of the invention contains reagents suitable for the determination of the GCNT3, LPL, APOC1, SCD, ACSL1 and ACSL4 genes. In another preferred embodiment, the kit of the invention additionally contains reagents suitable for the determination of the expression levels of one or more genes selected from the group ABCA1, APOE, FADS2 and CD36. In preferred embodiments, the kit of the invention comprises reagents suitable for determining the expression levels of the following gene groups:

-

ACSL4, SCD, APOE and FADS2;

-

ACSL4, SCD, ABCA1 and FADS2;

-

ACSL4, SCD, ABCA1 and APOC1;

-

ACSL4, ACSL1, SCD and ABCA1.

-

ACSL4, ACSL1, SCD, FADS2 and ABCA1

-

ACSL4, APOE, SCD, FADS2 and ABCA1

-

ACSL4, APOE, SCD, FADS2 and CD36

-

ACSL4, SCD, FADS2, ABCA1 and CD36

In a particular embodiment of the kits of the invention, the kit reagents are nucleic acids that are capable of specifically detecting the mRNA level of the aforementioned genes and / or the level of proteins encoded by one or more of the aforementioned genes. previously. Such detection is achieved thanks to the ability of nucleic acids to hybridize specifically with the aforementioned genes. The nucleic acids that are part of the kit of the invention can be probes that specifically recognize the target genes or can be one or more pairs of oligonucleotide primers for the specific amplification of fragments of mRNAs (or their corresponding cDNAs) of said genes

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In a preferred embodiment, the first component of the kit of the invention comprises a probe that can specifically hybridize to the genes mentioned above. The term "specific hybridization", as used herein, refers to the conditions that allow the hybridization of two polynucleotides under highly stringent conditions or moderately stringent conditions. "Rigor" of hybridization reactions is easily determined by one skilled in the art, and is usually an empirical calculation depending on the length of the probe, the washing temperature, and the salt concentration. In general, longer probes require higher temperatures for proper annealing, while shorter probes need lower temperatures. Hybridization generally depends on the ability of denatured DNA to resume when complementary strands are present in an environment below their melting temperature. The higher the degree of homology desired between the probe and the hybridizable sequence, the higher the relative temperature, which can be used. As a result, it follows that higher temperatures in relation would tend to make the reaction conditions more astringent, while lower temperatures not so much. For more details and explanations of the astringency of hybridization reactions, see Ausubel et al., Current Protocols in Molecular Biology, Wiley Interscience Publishers, (1995). "High stringency conditions", as defined in the present invention, generally involve: (1) low ionic strength and high temperature for washing, for example sodium chloride 0.015 / sodium citrate 0.0015 M /0.1 % sodium dodecyl sulfate at 50 ° C., (2) used during the hybridization of a denaturing agent, such as formamide, for example, 50% (v / v) formamide with 0.1% bovine serum albumin /0.1% Ficoll / 0.1% polyvinylpyrrolidone / 50 mM sodium phosphate buffer at pH 6.5 with 750 mM sodium chloride, 75 mM sodium citrate at 42 º C, or (3) use 50% formamide, 5xSSC ( 0.75 M NaCl, 0.075 M sodium citrate), 50 mM sodium phosphate (pH 6.8), 0.1% sodium pyrophosphate, 5 times Denhardt solution, sonic salmon sperm DNA (50 mg / ml ), 0.1% SDS, and 10% dextran sulfate at 42 ° C, with washings at 42 ° C in 0.2xSSC (sodium chloride / sodium citrate) and 50% formamide, followed by a wash of high rigor that Consists of 0.1xSSC containing EDTA at 55 ° C. "Moderately stringent conditions" can be identified as described by Sambrook et al, Molecular Cloning :. A Laboratory Manual, New York: Cold Spring Harbor Press, 1989, and include the use of wash solution and hybridization conditions (e.g. temperature, ionic strength and% SDS) less stringent than those described above. An example of moderately stringent conditions is incubation

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overnight at 37 ° C in a solution comprising: 20% formamide, 5xSSC (150 mM NaCl, 15 mM sodium citrate), 50 mM sodium phosphate (pH 7.6), 5x Denhardt solution, 10% sulfate of dextran, and 20 mg / ml denatured DNA sheared from salmon sperm, followed by washing the filters in 1xSSC at about 37-50º C. The person skilled in the art recognizes how to adjust the temperature, ionic strength, etc. . as necessary to accommodate factors such as probe length and the like. In the event that the expression levels of several of the genes identified in the present invention are determined at the same time, it is useful to include the probes for all the genes whose expression is to be determined in a microarray hybridization. The microarrays comprise a plurality of nucleic acids that are spatially distributed and stably associated with a support (for example, a biochip). Nucleic acids have a sequence complementary to particular sub-sequences of genes whose expression must be detected, so that they are capable of hybridizing with said nucleic acids. In the methods of the invention, a microarray comprising a set of nucleic acids is contacted with a preparation of the isolated nucleic acids of the patient under study. The incubation of the microarrays in the preparation of the nucleic acids is carried out under the conditions suitable for hybridization. Subsequently, after the elimination of nucleic acids that have not bound to the support, the hybridization pattern is detected, which provides information on the genetic profile of the analyzed sample. Although the microarrays are capable of providing both qualitative and quantitative information of the nucleic acids present in a sample, the invention requires the use of matrices and methodologies capable of providing quantitative information. The invention contemplates a variety of matrices in relation to the type of probes and in relation to the type of support used. The probes included in the matrices that are capable of hybridizing with the nucleic acids may be nucleic acids or analogs thereof that maintain the ability to hybridize, such as, for example, the nucleic acids in which the phosphodiester bond has been substituted with a phosphorothioate. , methylimine, methylphosphonate, phosphoramidate, nucleic acids in which the nucleotide ribose is replaced by another hexose, nucleic peptide (APN). The length of the probes can be from 5 to 50 nucleotides and, preferably, from 7, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, nucleotides 100 and they vary in the range of 10 to 1000 nucleotides, preferably in the range of 15 to 150 nucleotides, more preferably in the range of 15 to 100 nucleotides and nucleic acids may be single stranded or double stranded. The matrix can contain all the specific probes of a given mRNA of a certain length or can

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contain probes selected from different regions of an mRNA. Each probe is analyzed in parallel with a probe with a change base, preferably in a central area of the probe. The matrix is contacted with a sample containing nucleic acids with sequences complementary to the probes of the matrix and the hybridization signal with each of the probes and the corresponding hybridization controls are determined. The probes in which a greater difference is observed between the hybridization signal with the probe and its hybridization control are selected. The optimization process may include a second optimization round in which the hybridization matrix is hybridized with a sample that does not contain sequences complementary to the matrix probes. After the second round of selection, probes with hybridization signals below a threshold will be selected. Therefore, the probes that pass the controls, that is, showing a minimum level of non-specific hybridization and a maximum level of specific hybridization with the target nucleic acid are selected. The microarrays of the invention not only contain specific probes for polynucleotides that indicate a certain pathophysiological situation, but also contain a series of control probes, which can be of three types: normalization controls, expression level control and Hybridization controls. In the particular case of expression controls, it is useful to use probes that specifically recognize genes that encode proteins that perform essential functions of the cell and, therefore, show constant expression throughout the cell cycle. As a guideline, genes such as β-2-microglobulin 18S ubiquitin, ribosomal protein, cyclophilin A, transferrin receptor, actin, GAPDH, tyrosine 3 monooxygenase / tryptophan 5-monooxygenase protein activation (YWHAZ) can be used and betaactin. The microarrays can be high density matrices, obtained by fixing thousands of oligonucleotides using photolithography (Fodor et al., 1991, Science, 767 to 773). This type of matrix is usually redundant, that is, several probes are included for each mRNA to be detected. In a preferred embodiment, the matrices are low density matrices or LDA contain less than 10,000 probes per square centimeter. In said low density matrices, the different probes applied manually with the help of a pipette in different places of a solid support (for example, a glass surface, a membrane). The supports used to fix the probes can be obtained from a wide variety of materials, such as plastic, ceramics, metals, gels, membranes, crystals, etc. Microarrays can be obtained with any methodology known to the person skilled in the art.

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After hybridization, in cases where the non-hybridized nucleic acid is capable of emitting a signal in the detection step, a previous washing step is used to remove said non-hybridized nucleic acids. The washing step is carried out using methods and solutions known to the person skilled in the art. In the event that the expression levels of the genes according to the present invention are determined by measuring the levels of the polypeptide or polypeptides encoded by said gene or genes, the kits according to the present invention comprise reagents that are capable of specifically binding to said polypeptide or polypeptides. Thus, in one embodiment, the invention relates to a kit comprising antibodies specific for the polypeptide encoded by the GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes. For this, the use of antibody matrices such as those described by De Wildt et al. (2000) Nat. Biotechnol. 18: 989-994; Lueking et al. (1999) Anal. Biochem 270: 103-111; Ge et al. (2000) Nucleic Acids Res. 28, e3, I-VII; MacBeath and Schreiber (2000) Science 289: 1760-1763, WO01 / 40803 and WO 99 / 51773A1 are useful. Matrix antibodies include any immune agent capable of binding a high affinity ligand, including IgG, IgM, IgA, IgD and IgE, as well as antibody-like molecules that have an antigen binding site, such as Fab ' , Fab, F (ab ') 2, single domain antibodies or DABS, Fv, scFv and the like. The techniques for the preparation of said antibodies are well known to those skilled in the art and include the methods described by Ausubel et al. (Current Protocols in Molecular Biology, eds. Ausubel et al, John Wiley & Sons (1992)). In another aspect, the invention relates to the use of a kit of the invention to determine the prognosis of a patient suffering from colorectal cancer or to determine whether a patient with colorectal cancer is the candidate for adjuvant therapy. In a preferred embodiment, the use of the kits according to the invention is carried out in patients with stage II or stage III CCR.

Therapeutic methods In another aspect, the invention relates to the use of an inhibitor of a gene selected from the group consisting of LPL, APOC1, SCD, ACSL1 and ACSL4 for the preparation of a medicament for the treatment of cancer. In another aspect, the invention relates to an inhibitor of a gene selected from the group consisting of LPL, APOC1, SCD, ACSL1 and ACSL4 for use in the treatment of cancer.

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In another aspect, the invention relates to a method for the treatment of cancer in a subject comprising administering to said subject an inhibitor agent of a gene selected from the group consisting of LPL, APOC1, SCD, ACSL1 and ACSL4. The term "gene inhibitor", as used in the present invention, refers both to a

5 compounds capable of causing a decrease in the activity of the expression product of the genes LPL, APOC1, SCD, ACSL1 and ACSL4 as well as compounds capable of causing a decrease in the expression of said genes, either inhibiting the transcription of the corresponding gene, either by inhibiting the translation of the corresponding mRNA or by causing a decrease in the levels of the corresponding mRNA or mRNA.

In a preferred embodiment, the ACSL1 and ACSL4 gene inhibitor is selected from the group formed by the inhibitors in Table 1.

Triacsin C

Valproate or valproic acid.

Compounds of the thiazolidinediones family (for example troglitazone, rosiglitazone and pioglitazone).

Enoximone

Nonsteroidal anti-inflammatories (for example 2-arylpropionic acid, ibuprofen, fenoprofen, ketoprofen and naproxen).

Table 1: ACSL1 and ACSL4 inhibitors. Valproate or valproic acid: it is a specific ACSL4 inhibitor, thiazolidinediones are specific ACSL4 inhibitors.

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In a preferred embodiment, the APOC1 inhibitor is Vorinostat (suberoylanilide hydroxamic acid or SAHA). In a preferred embodiment, the SCD gene inhibitor is selected from the group consisting of the inhibitors of Table 2

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Rapamycin

Piperazine

3- (2-hydroxyethoxy) -N- (5-benzylthiazol-2-yl) -benzamides

3- [4- (2-Chloro-5-fluorophenoxy) -1 -piperidinyl] -6- (5-methyl-1,3,4-oxadiazol2-yl) -pyridazine

5- (tetradecyloxy) -2-furancarboxylic acid or TOFA

(N (2 (6 (3,4-dichlorobenzylamino) -2 - (4-methoxyphenyl) -3oxopyrido [2,3-b] pyrazine-4 (3H) -yl) ethyl) acetamide or CVT-11127

1- (4-phenoxypiperidin-1-yl) -2-arylaminoethanone

4- (2-Chlorophenoxy) -N- (3- (3-methylcarbamoyl) phenyl) piperidine-1carboxamide

2- (5- (3- (4- (2-Bromo-5-fluorophenoxy) piperidin-1-yl) isoxazol-5-yl) 2H-tetrazol-2-yl) acetic acid MK-8245

Table 2: SCD inhibitors

In a preferred embodiment, the LPL gene inhibitor is selected from the group consisting of the inhibitors of Table 3

TNF-α

Poloxamer 407 (P-407 or Pluronic F-127)

Tetrahydrolipstatin (THL)

Tyloxapol

Angiopoietin-like protein 3 (ANGPTL3)

Angiopoietin-like protein 4 (ANGPTL4)

Melanoma-derived LPL inhibitor (MLPLI)

APOC1

Orlistat or S) - ((S) -1 - ((2S, 3S) -3-hexyl-4-oxooxetan-2-yl) tridecan-2-yl) 2formamido-4-methylpentanoate

GSK 264220A or N- [2-Methyl-5- (1-piperidinylsulfonyl) -3-furanyl] -N'phenylurea

Table 3: LPL inhibitors

In a preferred embodiment, the inhibitor agent is an inhibitor antibody specific to the product encoded by said gene. In another preferred embodiment, the inhibitor is selected from the group consisting of a specific antisense olignucleotide.

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for said gene, an interference RNA specific for said gene and a ribozyme specific for said gene. In a preferred embodiment, the inhibitor agent for use in the present invention is a specific inhibitor antibody for the product encoded by a gene selected from the group consisting of LPL, APOC1, SCD, ACSL1 and ACSL4. Antibodies against an epitope of proteins encoded by LPL, APOC1, SCD, ACSL1 and ACSL4 genes can effectively block the function of these proteins and, therefore, can be used as inhibitors in the compositions of the present invention. "Inhibitor antibody" as used herein, refers to antibodies that are capable of at least partially inhibiting the biological activities of the polypeptides encoded by the LPL, APOC1, SCD, ACSL1 and ACSL4 genes. The determination of the inhibitory capacity on the biological activity of the LPL-encoded product can be carried out, for example, using the method described by Mayes and Felts (Biochem. J., 1968, 108,483-487) based on the determination of the ability of the antibody to inhibit the degradation of triglycerides in the presence of LPL. The determination of the inhibitory capacity on the biological activity of the product encoded by SCD can be carried out, for example, using the method described by Strittmatter et al. (J. Biol. Chem. 1988, 263; 2532-2535) based on the determination of the ability of the antibody to inhibit phosphatidylcholine degradation in the presence of SCD. The determination of the inhibitory capacity on the biological activity of the product encoded by ACSL1 or ACSL4 can be carried out, for example, using the method described by Gassler et al. (Gastroenterology, 2007, 133: 587-98) based on the determination of the ability of the antibody to inhibit the synthesis of palmitoyl-CoA from palmitic acid and CoA in the presence of ACSL1 or ASCL4. Antibodies or inhibitor fragments specific for the polypeptides encoded by the LPL, APOC1, SCD, ACSL1 and ACSL4 genes can be purchased commercially or easily produced using conventional molecular biology techniques. Suitable inhibitory antibodies include polyclonal antibodies, monoclonal antibodies, Fab, F (ab ’) 2, Fab’ and scFv fragments thereof, bispecific, heteroconjugate antibodies, diabodies, human and humanized antibodies. In another preferred embodiment, the inhibitor agent is an interference RNA (RNAi) specific for silencing a gene selected from the group consisting of GCNT3, LPL, APOC1, SCD, ACSL1 and ACSL4. Interference RNAs suitable for silencing the LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes include, without limitation,

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Small interference RNA (siRNA), micro RNA (miRNA) and short bracketed RNA (hRNA). In another embodiment, the inhibition of the expression of the genes LPL, APOC1, SCD, ACSL1 and / or ACSL4 is carried out by the use of ribozymes designed to cut

5 catalytic form transcripts of mRNA targets generated by the transrciption of different genes. Types of ribozymes suitable for inhibition of gene expression include, without limitation, hammerhead ribozymes and endoribonuclease RNA (Cech type ribozymes) In another preferred embodiment, the inhibitory agent is an antisense nucleic acid.

10 specific for a gene selected from the group consisting of LPL, APOC1, SCD, ACSL1 and ACSL4. Antisense nucleic acids can be linked to the gene that is desired to be inhibited by conventional base complementarity, or, for example, in the case of binding to double stranded DNA, through specific interactions in the major groove of the double helix. In general, these methods refer to the range of techniques generally employed in the

Technique, and include any method that relies on specific binding to oligonucleotide sequences. Exemplary nucleic acid molecules for use as antisense oligonucleotides include:

-Oligonucleotides comprising modified bases such as 5-fluorouracil, 5-bromouracil, 5-chlorouracil, 5-iodouracil, hypoxanthine, xanthine, 420 acetylcytosine, 5- (carboxyhydroxythiethyl) uracil, 5-carboxymethylaminomethyl-2-thioxymethyl, 2-carboxymethyl dihydrouracil, beta-D-galactosylkeosine, inosine, N6-isopentenyladenine, 1-methylguanine, 1-methylinosine, 2,2-dimethylguanine, 2-methylladenine, 2-methylguanine, 3-methylcytosine, 5-methylcytosine, N6-adenine, 7-methylguanine 5-Methylaminomethyluracil, 5-methoxyaminomethyl-225 thiouracil, beta-D-mannosylkeosine, 5'-methoxycarboxymethyluracil, 5-methoxyuracil, 2-methylthio-N6-isopentenyladenine, uracil-5-oxyacetic acid (v), wybutoxosylocyclosine, pseudoxyracilosine, p 2-Thiocytosine, 5-methyl-2-thiouracil, 2-thiouracil, 4-thiouracil, 5-methyluracil, uracil-5-oxyacetic acid methyl ester, uracil5-oxyacetic acid (v), 5-methyl-2-thiouracil, 3- (3-amino-3-N-2-carboxypropyl) uracil,

30 (acp3) w, and 2,6-diaminopurine. -Oligonucléotides comprising modified sugars such as arabinose, 2fluoroarabinous, xylulose and hexose. -Oligonucleotides comprising a modified phosphate skeleton selected from the group consisting of a phosphorothioate, a phosphorodithioate, a

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phosphoramidothioate, a phosphoramidate, a phosfordiamidate, a methylphosphonate, an alkyl phosphotriester, and a formacetal or a peptide nucleic acid (ANP) -

5 Proc. Natl. Acad. Sci. U.S.A. 86: 6553-6556, 1989; Lemaitre et al., Proc. Natl. Acad. Sci. 84: 648-652, 1987; PCT Publication No. WO88 / 09810) or the blood brain barrier (see, for example, PCT Publication No. WO89 / 10134), hybridization triggers (see, for example, Krol et al., BioTechniques 6: 958 -976, 1988) intercalating agents (see, for example, Zon,

10 Pharm. Res. 5: 539-549, 1988). - Alpha-anomeric oligonucleotides - Morpholino oligonucleotides In another preferred embodiment, the expression inhibitor of the LPL, APOC1, SCD, ACSL1 and / or ACSL4 genes is a DNA enzyme.

15 The terms "cancer" and "tumor" refer to the physiological condition in mammals characterized by deregulated cell growth. Types of cancer that can be treated according to the methods of the present invention include, without limitation, cancer of the esophagus, stomach, liver, pancreas, gallbladder, small intestine, rectum, colon, colorectal, prostate, lung, ovary and mom. In a preferred embodiment of the invention the

20 tumor / cancer to be treated with said compositions is colon cancer. In another preferred embodiment of the invention the tumor / cancer to be treated with said compositions is breast cancer.

*** The invention detailed below by way of the following examples are merely illustrative and in no way limit the scope of the invention.

EXAMPLES

Materials and methods

30 Tissue tissue samples embedded in paraffin from patients with stage B colon cancer from the La Paz University Hospital in Madrid were used. The study consists of a retrospective analysis of the prognostic value of the expression of GCNT3, LPL, APOC1, SCD, ACSL1 and / or ACSL4 in these patients. The time until

35 progression was used for disease-free survival analysis. ROC (receiver operating characteristic) curves were used to determine the expression cut-off values of these genes for disease-free survival according to the relationship between sensitivity and false positive ratio (1-specificity), establishing the best combination marked by the ROC curves . The Kaplan-Meier method was used to estimate the

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5 disease-free survival. All values of p referred are two-tailed. Statistical significance was defined as p <0.05. Statistical analysis was performed using SPSS software (version 19.0). 83 patients with stage B colon cancer have been included in this study, with a median cancer survival of 69 months. The median free survival of

10 disease in these patients is 60 months, with 22 patients (26.5%) with relapse at the time of analysis. The clinical and pathological characteristics of this series of patients are included in Table 4.

Tumor stage:

B 83 (100%)

Sex

Female Male 34 (41%) 49 (59%)

Diff grade:

Poor Moderate Good 5 (6%) 71 (8%) 6 (7%)

Desmoplasia:

No Yes 37 (44%) 29 (35%)

Vascular invasion

No Yes 57 (69%) 25 (30%)

Chemotherapy

Do not 34 (41%)

(UFT-LV, xeloda)

Yes 49 (59%)

Age at diagnosis

Medium: Minimum: Maximum: 69 32 86

Table 4. Characteristics of the patients.

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Example 1. Prognostic value of the LPL gene in clinical samples of patients with colon cancer.

As can be seen in Figure 1, patients with higher levels of LPL have significantly lower disease-free survival (p = 0.006) and 20 lower overall survival (cancer death) (p = 0.04) than those with levels of lower. The increased risk of relapse and death from cancer in these patients is three times higher than patients with low levels of LPL (HR = 3.3 and 3.2 respectively). The stratification of patients according to whether they have received chemotherapy or not has no influence.

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in the data maintaining the HR in both cases. These results indicate that LPL overexpression has both a prognostic and predictive value of the response to treatment in these patients.

Example 2: Prognostic value of the APOC1 gene in clinical samples of patients with colon cancer.

As can be seen in Figure 2, patients with higher levels of APOC1 have significantly lower disease-free survival (p = 0.02), and lower overall survival (p = 0.009) than those with lower levels. The increased risk of relapse and death from cancer is more than twice higher for patients with high levels of APOC1 (HR = 2.6 and 4.7 respectively). The stratification of patients according to whether they have received chemotherapy or not has no influence on the data, maintaining HR in both cases. These results indicate that overexpression of APOC1 has both a prognostic and predictive value of the response to treatment in these patients.

Example 3: Prognostic value of the SCD gene in clinical samples of patients with colon cancer.

As can be seen in Figure 3, patients with higher levels of SCD have significantly lower disease-free survival (p = 0.02), and lower overall survival (p = 0.03) than those with lower levels. The increased risk of relapse and death from cancer is more than twice higher for patients with high levels of SCD (HR = 2.6 and 3.3 respectively). The stratification of patients according to whether they have received chemotherapy or not has no influence on the data, maintaining HR in both cases. These results indicate that the overexpression of SCD has both a prognostic and predictive value of the response to treatment in these patients.

Example 4: Prognostic value of ACSL1 and ACSL4 genes in clinical samples of patients with colon cancer. As can be seen in Figure 4A, patients with higher levels of ACSL1 have significantly lower disease-free survival (p = 0.02), and lower overall survival (p = 0.03) than those with lower levels. The increased risk of relapse and death from cancer is more than twice higher for patients with high levels of ACSL1 (HR = 2.6 and 3.1 respectively). The stratification of patients according to whether they have received chemotherapy or not followed by the corresponding multivariate analysis shows a significant interaction (p = 0.032), indicating that while ACSL1 overexpression has no influence on relapse or survival of patients who have not received chemotherapy. , there is an almost 10-fold increase in the risk factor for relapse in those patients who have received chemotherapy and have overexpressed ACSL1, with a HR = 9.8 (p = 0.009) (Figure 4B). These results indicate that the

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5 Overexpression of ACSL1 has a marked predictive value of a failure in response to treatment with oral fluoropyrimidines (UFT, xeloda) of patients with colon cancer. In order to validate these data, tumor samples from a second series of 58 patients with stage B colon cancer (validation series) from La Paz University Hospital were used, all treated with treatments also including

10 oral fluoropyridines (FOLFOX or XELOX). The clinical and pathological characteristics of this series of patients are included in Table 5.

Tumor stage:

B 58 (100%)

Sex:

Female Male 22 (38%) 36 (62%)

Diff grade:

Poor Moderate Good 5 (8%) 43 (74%) 10 (17%)

Vascular invasion

No Yes 46 (79%) 12 (21%)

Chemotherapy: (Xelox, Folfox)

Yes 58 (100%)

Age at diagnosis

Medium Minimum Maximum 60 23 77

Table 5. Characteristics of the patients in the validation series.

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As can be seen in Figure 4C, patients with over-expression of ACSL1, despite having received the same treatment, relapse significantly before (p = 0.025), and, consequently, have a lower survival (p = 0.04), which those patients with low levels of ACSL1. These results confirm that ACSL1 is a predictive factor of

20 response to conventional treatments applied in patients with colon cancer in non-advanced stages. ACSL4 is an isozyme of the ACSL1 family. Within these two family members, we have observed a significant correlation between ACSL1 and ACSL4 (Spearman's correlation 0.5, p = 0.002), indicating that both genes have a tendency to "coexpress" in the

25 same patients. However, being a correlation value not close to 1, these results indicate the interest in analyzing both genes to have a global view of the value

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marker of this route. When determining the prognostic value of ACSL4, we observe that patients with higher levels of ACSL4 have significantly lower disease-free survival (p = 0.0001), and lower overall survival (p = 0.001) than those with higher levels. low. The increased risk of relapse and death from cancer is more than four times higher for patients with high levels of ACSL4 (HR = 4.39 and 5.7 respectively) (Figure 5A). The stratification of the patients according to whether they have received chemotherapy or not does not present a significant interaction, but it is of greater magnitude in those patients who have not received chemotherapy (Figure 5B) (also maintaining a qualitatively similar behavior in the treated patients). These results indicate that over-expression of ACSL4 has both a prognostic value (with greater magnitude) and a predictive response to treatment in these patients. Therefore, these results indicate that the determination of ACSL1 and ACSL4 (simultaneous, or not) in patients with colon cancer in advanced stages provides important information both additive and complementary on the prognosis of patients (more marked by ACSL4) and its response to conventional therapy (more marked by ACSL1).

Example 5. Prognostic value of the GCNT3 gene in clinical samples of patients with colon cancer. As can be seen in Figure 6, patients with lower levels of GCNT3 have significantly lower disease-free survival than those with higher levels. The median disease-free survival of patients with low levels of the GCNT3 gene is 25 months, while it is not reached in patients with GCNT3 levels above the cut-off point established by the ROC curve (p = 0.001). These results indicate that the low expression of GCNT3 has a poor prognostic marker value in patients with non-advanced colon cancer.

Example 6. Prognostic value of combinations of different genes

An analysis using the “survival” package of the R software in which survival was analyzed by a multivariate analysis of COX after discriminating the variables and making K-means comparisons of gene pairs indicates that the combination of some of these genes increase the predictive power of the test (table 6).

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Gene1

Gene2 pValue.gen pValue.gen pValue

APOC1.Hs00155790_m ACSL1.Hs00253457_m1 ACSL4.Hs01547083_m1 ACSL1.Hs00253457_m1

SCD.Hs01682761_m SCD.Hs01682761_m SCD.Hs01682761_m APOC1.Hs00155790 0.033053 0.006501 0.080951 0.006501 0.001048 0.001048 0.001048 0.033053 0.000019 0.000123 0.000831 0.001247

Table 6. Multivariate COX analysis of the value as a prognostic marker of the combination of gene pairs.

Example 7. Prognostic value of the combinations of these genes with four additional genes involved in nearby metabolic pathways.

Subsequently, a multivariate analysis was carried out including 31 other genes of related metabolic pathways, which individually, although they marked a certain tendency to have a predictive value, did not present a significant and validated marker value. This analysis sought to find a “metabolic footprint” prognostic marker in patients 10 with colon cancer according to a prognostic index derived from a Cox regression model. For this, a Cross-Validation analysis (CV, Cross-Validation) was performed, using 5fold CV, in which it is observed as combinations of ACSL1, ACSL4, SCD, LPL and / or APOC1 with APOE, FADS2, ABCA1 and / or CD36, significantly increase the predictive value of the test. While the individual analysis of one of the marker genes indicates

15 a maximum risk of relapse of 7 times in a positive patient versus another that is not, the fact of also determining the expression of these additional genes that by themselves do not mark response, increases the prognostic value identifying patients who have 19 times more risk of relapse than those without this footprint (Table 7).

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Gene1

Gene4 Gene3 Gene4 Gene5 C.INDEX HR signature p (test) validation p (val)

ACSL4

SCD ABCA1 FADS2 0.79 12.96 0 4.91 0.001

ACSL4

SCD APOE FADS2 0.796 12.79 0 3.96 0.0093

ACSL4

SCD ABCA1 APOC1 0.77 7.71 0 5.05 0.0009

ACSL4

SCD ABCA1 ACSL1 0.775 9.54 0 2.83 0.05

ACSL4

SCD ABCA1 FADS2 ACSL1 0.795 10.29 0 6.16 0.0002

ACSL4

SCD ABCA1 APOE FADS2 0.8 12.96 0 6.16 0.0002

ACSL4

SCD FADS2 APOE CD36 0.811 19.22 0 3.53 0.02

ACSL4

SCD ABCA1 CD36 FADS2 0.791 13.36 0 4.25 0.0033

Table 7. Genomic footprint based on genes involved in metabolism. Predictive value in Hazard Ratio (HR) of the genes in combination forming a metabolic footprint constituted by the joint determination of the following genes. These models,

5 identified in the initial series, have been validated in the validation series, and ratified in a meta-analysis with the global patient.

Thus, patients with higher values of this metabolic footprint have a significantly higher risk of relapse than patients with intermediate and low levels (Figure 7).

10

Example 8. Antitumor activity of agents that enhance GCNT3 expression.

The fact that a low expression of GNCT3 is indicative of a poor prognosis of the disease suggests that it is a factor of tumor aggressiveness. Therefore, agents that potentiate the expression of this gene could have a good antitumor activity. The GCNT3 enhancer activity of a series of bioactive compounds has been studied, finding that rosemary extract significantly enhances the expression of this gene in different human colon and breast cancer cells both in vitro and in vivo. These results suggest that agents capable of increasing GCNT3 levels may be promising antitumor agents. Likewise, agents capable of

Inhibiting the expression or activity of LPL, APOC1, SCD, ACSL1 or ACSL4 may also result in efficient agents with antitumor activity, especially for patients in whom these markers are altered.

Claims (15)

image 1 1. Method for determining the prognosis of a patient affected by colorectal cancer that includes the determination of the levels of expression of the ACSL1 and / or genes 5 ACSL4 in a sample of said patient, where an increase in the expression levels of the ACSL1 and / or ACSL4 genes with respect to a reference value for each gene is indicative that the patient has a poor prognosis. 2. Method according to claim 1 further comprising determining 10 the expression levels of at least one gene selected from the group consisting of GCNT3, LPL, APOC1 and SCD, where an increase in the expression levels of the LPL, APOC1 and / or SCD genes and / or a decrease in levels GCNT3 gene expression with respect to a reference value for each gene is indicative that the patient has a poor prognosis. fifteen 3. Method according to claims 1 or 2 further comprising determining the expression levels of at least one gene selected from the group consisting of ABCA1, APOE, FADS2 and CD36, wherein an increase in the expression levels of ABCA1 genes , APOE, FADS2 and / or CD36 with respect to a value of 20 reference for each gene is indicative that the patient has a poor prognosis. 4. Method according to any of claims 1 to 3 wherein the patient has been treated with a fluoropyridine-based therapy. 25 5. Method for selecting a suitable treatment for a patient affected by colorectal cancer comprising the determination of the expression levels of the ACSL1 and / or ACSL4 genes in a sample of said patient, wherein an increase in the expression levels of ACSL1 and / or ACSL4 genes with respect to a reference value for each gene is indicative that the patient has to be treated with 30 adjuvant chemotherapy. Method according to claim 5, further comprising determining the expression levels of at least one gene selected from the group consisting of GCNT3, LPL, APOC1 and SCD, wherein an increase in the expression levels of the 35 LPL, APOC1 and / or SCD genes and / or a decrease in gene expression levels 39 image2 GCNT3 with respect to a reference value for each gene is indicative that the patient has to be treated with adjuvant chemotherapy.

7.

Method according to claims 5 or 6, further comprising determining the expression levels of at least one gene selected from the group consisting of ABCA1, APOE, FADS2 and CD36, wherein an increase in the expression levels of the ABCA1, APOE genes , FADS2 and / or AGPAT2 with respect to a reference value for each gene is indicative that the patient has to be treated with adjuvant chemotherapy.

8.

Method according to any one of claims 5 to 7 wherein said adjuvant chemotherapy is a therapy other than fluoropyridine-based therapy.

9.

Method according to any of claims 1 to 8 wherein the expression levels of the SCD and ACSL1 genes are determined; SCD and ACSL4; APOC1 and ACSL1; ACSL4, SCD, APOE and FADS2; ACSL4, SCD, ABCA1 and FADS2; ACSL4, SCD, ABCA1 and APOC1; ACSL4, ACSL1, SCD and ABCA1 or LPL, ACSL4, FADS2 and AGPAT2.

10.

Method according to any one of claims 1 to 9 wherein the patient suffers from stage B colorectal cancer.

eleven.

Method according to any one of claims 1 to 10 wherein the sample is selected from the group consisting of a tumor biopsy or a biofluid.

12.

Method according to claim 11 wherein the biofluid is selected from the group consisting of blood, plasma and serum.

13.

A kit comprising the appropriate reagents for the determination of the expression levels of the ACSL1 and / or ACSL4 genes and, optionally, the reagents for the determination of the expression levels of one or more constitutive expression genes, wherein said reagents they are selected from the group consisting of probes that specifically hybridize with said genes and antibodies capable of specifically binding to the polypeptides encoded by said genes.

14.

Kit according to claim 13 further comprising suitable reagents for the determination of the expression levels of at least one gene selected from the group consisting of GCNT3, LPL, APOC1 and SCD.

40 image3 15. Kit according to claims 13 or 14 further comprising reagents suitable for the determination of the expression levels of at least one gene selected from the group consisting of ABCA1, APOE, FADS2 and CD36. 16. Kit according to any of claims 13 to 15 comprising reagents 10 suitable for the determination of the expression levels of the SCD and ACSL1 genes; SCD and ACSL4; APOC1 and ACSL1; ACSL4, SCD, APOE and FADS2; ACSL4, SCD, ABCA1 and FADS2; ACSL4, SCD, ABCA1 and APOC1; ACSL4, ACSL1, SCD and ABCA1; ACSL4, ACSL1, SCD, FADS2 and ABCA1; ACSL4, APOE, SCD, FADS2 and ABCA1; ACSL4, APOE, SCD, FADS2 and CD36; or ACSL4, SCD, FADS2, ABCA1 and  15 CD36. 17. Use of an inhibitor agent of the CSL1 and / or ACSL4 genes for the preparation of a medicament for the treatment of cancer in which the ACSL1 and / or ACSL4 inhibitor is selected from the group consisting of an inhibitor antibody specific for the 20 product encoded by said genes, an antisense oligonucleotide specific to said genes, an interference RNA specific to said genes, a ribozyme specific to said genes, a DNA enzyme specific to said genes and a compound as indicated in Table 1. 41