Application of PARP inhibitor combined with chemotherapy in treatment of CXorf67 high-expression tumor

Abstract

The invention provides an application of a PARP inhibitor combined with chemotherapy in treatment of CXorf67 high-expression tumors. Specifically, the invention provides an application of an active component combination, the active component combination is used for preparing a medicine or a preparation for treating or synergistically treating CXorf67 expression or high expression tumors, and the active component combination comprises (Z1) a PARP inhibitor, (Z2) a CXorf67 expression or high expression tumor inhibitor, and (Z3) a CXorf67 expression or high expression tumor inhibitor. And (Z2) a chemotherapeutic drug. It is accidentally found for the first time that a tumor cell line or tumor tissue expressing or highly expressing CXorf67 is more sensitive to PARP inhibitor combined chemotherapy (such as PARP inhibitor combined platinum drugs), the CXorf67 gene or protein thereof can be used as a marker for detecting the sensitivity of tumor cells to PARP inhibitor combined chemotherapy, and the PARP inhibitor and chemotherapy have a synergistic treatment effect.

Description

Application of PARP inhibitors combined with chemotherapy in the treatment of tumors with high CXorf67 expression Technical Field

[0001] The present invention belongs to the field of biomedicine and specifically relates to the use of a PARP inhibitor combined with chemotherapy in the treatment of tumors with high CXorf67 expression. Background Art

[0002] The homologous recombination (HR) repair pathway is an error-free repair mechanism that primarily repairs DNA breaks during the S / G2 phase of the cell cycle, utilizing homologous DNA within the cell as a template. Defects or deficiencies in HR can lead to genomic instability, which in turn contributes to the development and progression of tumors. For example, mutations in genes such as BRCA1, PLAB2, and BRCA2, which are involved in the HR pathway, have been found in many tumors. Mutations in these genes can prevent effective repair of double-strand breaks in DNA, leading to changes in gene copy number, rearrangements, and mutations, ultimately contributing to the development of tumors and diseases such as ovarian, breast, pancreatic, and prostate cancers.

[0003] In 2005, two laboratories simultaneously reported that PARP inhibitors could selectively kill tumor cells with BRCA1 / 2 mutations. Subsequent studies found that tumor cells with BRCAness (containing HR defects but not BRCA1 / 2 mutations) were also sensitive to PARP inhibitors. However, as research deepened, it was discovered that various PARP inhibitors had comparable inhibitory efficiencies on PARP catalytic activity, but their cell-killing effects varied greatly. How to further effectively kill tumors with HR repair pathway defects remains unknown.

[0004] Therefore, there is an urgent need to develop a more sensitive and effective treatment for tumors with defects in the HR repair pathway.

[0005] Summary of the Invention

[0006] The purpose of the present invention is to provide a more effective treatment method for tumors expressing or overexpressing CXorf67, especially a treatment method of combining PARP inhibitors with chemotherapy to achieve a synergistic therapeutic effect.

[0007] Another object of the present invention is to provide the use of the CXorf67 gene, mRNA, cDNA or protein or a detection reagent thereof as a marker for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0008] Another object of the present invention is to provide the use of CXorf67 gene, mRNA, cDNA or protein or its detection reagent for preparing a diagnostic reagent or kit for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0009] In a first aspect of the present invention, there is provided a use of an active ingredient combination for preparing a medicament or preparation for treating CXorf67-expressing or overexpressing tumors, or a medicament or preparation for synergistically treating CXorf67-expressing or overexpressing tumors, wherein the active ingredient combination comprises:

[0010] (Z1) first active ingredient: a PARP inhibitor; and

[0011] (Z2) Second active ingredient: chemotherapy drug.

[0012] In another preferred embodiment, the PARP inhibitor is selected from the group consisting of Talazoparib, Olaparib, Veliparib, Rucaparib, Niraparib, Fluzoparib, Pamiparib, or a combination thereof; preferably, the PARP inhibitor is selected from Niraparib.

[0013] In another preferred embodiment, the dosage of the PARP inhibitor is 10-100 mg / kg, preferably 20-80 mg / kg, more preferably 30-70 mg / kg, and most preferably 50 mg / kg.

[0014] In another preferred embodiment, the chemotherapy drugs include drugs that destroy DNA structure and / or function (such as platinum drugs).

[0015] In another preferred embodiment, the chemotherapy drug is a platinum drug.

[0016] In another preferred embodiment, the platinum drug is selected from the group consisting of Cisplatin, Carboplatin, Nedaplatin, Oxaliplatin, Lobaplatin, or a combination thereof; preferably, the platinum drug is selected from the group consisting of Carboplatin and / or Cisplatin.

[0017] In another preferred embodiment, the dosage of the platinum drug is 0.1-20 mg / kg, preferably 0.5-10 mg / kg, more preferably 1-5 mg / kg, and most preferably 2 mg / kg.

[0018] In another preferred embodiment, the drug or preparation further comprises a pharmaceutically acceptable carrier, diluent or excipient.

[0019] In another preferred embodiment, the drug or preparation further comprises other tumor therapeutic agents.

[0020] In another preferred embodiment, the CXorf67 expressing or high expressing tumor is selected from the following group: ependymoma posterior fossa group A, kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP), or a combination thereof.

[0021] In another preferred embodiment, the tumor includes a tumor with a defective HR repair pathway.

[0022] In another preferred embodiment, the tumor is a primary tumor, a metastatic tumor, a transplanted tumor, or a combination thereof.

[0023] In another preferred embodiment, the ependymoma comprises PFA.

[0024] In another preferred example, the accession number of the CXorf67 gene is Gene ID: 340602.

[0025] In another preferred embodiment, the accession number of the CXorf67 mRNA is NM_203407.3.

[0026] In another preferred embodiment, the accession number of the CXorf67 protein is NP_981952.1.

[0027] In a second aspect of the present invention, a pharmaceutical composition is provided, comprising: a PARP inhibitor, a chemotherapeutic drug, and a pharmaceutically acceptable carrier, diluent, or excipient.

[0028] In another preferred embodiment, the pharmaceutical composition further comprises other tumor therapeutic agents.

[0029] In another preferred embodiment, the PARP inhibitor is selected from the group consisting of Talazoparib, Olaparib, Veliparib, Rucaparib, Niraparib, Fluzoparib, Pamiparib, or a combination thereof; preferably, the PARP inhibitor is selected from Niraparib.

[0030] In another preferred embodiment, the dosage of the PARP inhibitor is 10-100 mg / kg, preferably 20-80 mg / kg, more preferably 30-70 mg / kg, and most preferably 50 mg / kg.

[0031] In another preferred embodiment, the chemotherapy drugs include drugs that destroy DNA structure and / or function (such as platinum drugs).

[0032] In another preferred embodiment, the chemotherapy drug is a platinum drug.

[0033] In another preferred embodiment, the platinum drug is selected from the group consisting of Cisplatin, Carboplatin, Nedaplatin, Oxaliplatin, Lobaplatin, or a combination thereof; preferably, the platinum drug is selected from the group consisting of Carboplatin and / or Cisplatin.

[0034] In another preferred embodiment, the dosage of the platinum drug is 0.1-20 mg / kg, preferably 0.5-10 mg / kg, more preferably 1-5 mg / kg, and most preferably 2 mg / kg.

[0035] In another preferred embodiment, the PARP inhibitor is Niraparib, and the chemotherapy drug is Carboplatin or Cisplatin.

[0036] In a third aspect of the present invention, a medicine kit is provided, comprising:

[0037] (a) a first formulation comprising a PARP inhibitor and a pharmaceutically acceptable carrier;

[0038] (b) a second formulation comprising a chemotherapeutic agent and a pharmaceutically acceptable carrier; and

[0039] (c) instructions describing a method of using the first agent and the second agent in combination to treat a tumor.

[0040] In another preferred embodiment, the first preparation and the second preparation are independent of each other.

[0041] In another preferred embodiment, the first preparation and the second preparation are freeze-dried preparations or liquid preparations.

[0042] In another preferred embodiment, the dosage form of the first preparation and the second preparation is an injection or a gastrointestinal tract administration dosage form.

[0043] In another preferred embodiment, the first preparation is administered before, during or after the administration of the second preparation.

[0044] In a fourth aspect of the present invention, there is provided a use of a CXorf67 gene, mRNA, cDNA or protein or a detection reagent thereof, (i) as a marker for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy; and / or (ii) for preparing a diagnostic reagent or kit for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0045] In another preferred embodiment, the diagnostic reagent includes antibodies, primers, probes, sequencing libraries, nucleic acid chips (such as DNA chips) or protein chips.

[0046] In another preferred embodiment, the protein includes a full-length protein or a protein fragment.

[0047] In another preferred embodiment, the protein contains a PALB2-binding motif.

[0048] In another preferred embodiment, the PALB2 binding motif is located at positions 420-432 of the CXorf67 protein.

[0049] In another preferred embodiment, the CXorf67 gene, mRNA, cDNA or protein is derived from mammals, preferably from rodents (such as mice, rats), primates and humans, and more preferably from patients diagnosed with tumors with HR repair pathway defects.

[0050] In another preferred embodiment, the detection is tissue sample detection.

[0051] In another preferred embodiment, the detection includes immunohistochemistry, immunoblotting and fluorescence quantitative PCR detection.

[0052] In another preferred embodiment, the detection is the measurement of tumor tissue.

[0053] In another preferred embodiment, the detection reagent includes a CXorf67-specific antibody, a CXorf67-specific binding molecule, a specific amplification primer, a probe or a chip.

[0054] In another preferred embodiment, the CXorf67 protein or its specific antibody or specific binding molecule is coupled with or carries a detectable label.

[0055] In another preferred embodiment, the detectable label is selected from the following group: a chromophore, a chemiluminescent group, a fluorophore, an isotope or an enzyme.

[0056] In another preferred embodiment, the CXorf67-specific antibody is a monoclonal antibody or a polyclonal antibody.

[0057] In a fifth aspect of the present invention, a diagnostic kit for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy is provided, the kit comprising a container containing a detection reagent for detecting the CXorf67 gene, mRNA, cDNA or protein; and a label or instructions indicating that the kit is used for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0058] In another preferred embodiment, the detection reagent for detecting CXorf67 gene, mRNA, cDNA or protein includes:

[0059] (a) Anti-CXorf67 protein-specific antibodies; and / or

[0060] (b). Specific primers for specific amplification of CXorf67 mRNA or cDNA.

[0061] In another preferred embodiment, the detection is tissue sample detection.

[0062] In a sixth aspect of the present invention, a method for determining sensitivity to PARP inhibitors combined with chemotherapy is provided, the method comprising:

[0063] a) providing a test sample from a subject;

[0064] b) detecting the expression level of CXorf67 protein in the test sample; and

[0065] c) judging the sensitivity to PARP inhibitor combined with chemotherapy based on the expression level of CXorf67 protein determined in step b).

[0066] In another preferred embodiment, when CXorf67 protein is present in the test sample, it can be determined that the test sample is sensitive to PARP inhibitors combined with chemotherapy.

[0067] In another preferred embodiment, when the expression level of CXorf67 protein in the test sample is greater than 0.5, preferably greater than 1.5, and more preferably greater than 2, it can be determined that the patient is sensitive to PARP inhibitor combined with chemotherapy.

[0068] In another preferred embodiment, the subject is a human or non-human mammal.

[0069] In another preferred embodiment, the test sample is a tumor cell or tissue expressing or over-expressing CXorf67.

[0070] In another preferred embodiment, the test sample is a tumor cell or tissue with defective HR repair pathway.

[0071] In another preferred embodiment, the detecting step (b) comprises detecting the amount of CXorf67 mRNA, or the amount of CXorf67 cDNA; and / or detecting the amount of CXorf67 protein.

[0072] In another preferred embodiment, the expression level of CXorf67 protein in the sample is detected by fluorescent quantitative PCR or immunohistochemistry.

[0073] In another preferred embodiment, the method is non-diagnostic and non-therapeutic.

[0074] In a seventh aspect of the present invention, a method for determining a treatment regimen is provided, the method comprising:

[0075] a) providing a test sample from a subject;

[0076] b) detecting the expression level of CXorf67 protein in the test sample; and

[0077] c) determining a treatment regimen based on the expression level of CXorf67 protein in the sample.

[0078] In another preferred embodiment, the subject is a human or non-human mammal.

[0079] In another preferred embodiment, when CXorf67 protein is present in the sample (preferably the expression level of CXorf67 protein is >0.5, preferably >1.5, and more preferably >2), the treatment regimen includes: CXorf67 inhibitor therapy, PARP inhibitor combined with chemotherapy, or a combination thereof.

[0080] In another preferred embodiment, the pharmaceutical ingredients used in the CXorf67 inhibitor therapy and the PARP inhibitor combined chemotherapy are selected from the following groups:

[0081] CXorf67 inhibitor therapy: antibodies, small molecule compounds, microRNA, siRNA, shRNA, or combinations thereof that target and inhibit CXorf67;

[0082] PARP inhibitor combination chemotherapy: PARP inhibitor, platinum drug, or a combination thereof.

[0083] In another preferred example, when the subject is more sensitive to PARP inhibitor combined with chemotherapy than the general population (control group), the treatment regimen also includes: CXorf67 inhibitor therapy, PARP inhibitor combined with chemotherapy, and a combination of CXorf67 inhibitor therapy and PARP inhibitor combined with chemotherapy.

[0084] In the eighth aspect of the present invention, a method for synergistically killing or inhibiting tumor cells for non-disease treatment purposes in vitro is provided, comprising the steps of: contacting tumor cells with a PARP inhibitor and a chemotherapy drug, thereby synergistically killing or inhibiting the tumor cells.

[0085] In another preferred embodiment, the tumor cells are tumor cells expressing or over-expressing CXorf67.

[0086] In another preferred embodiment, the chemotherapy drug is a platinum drug.

[0087] In another preferred embodiment, the method further comprises subjecting the tumor cells to radiotherapy, targeted therapy, immunotherapy, or a combination thereof.

[0088] In another preferred embodiment, the radiotherapy comprises irradiation.

[0089] In another preferred embodiment, the irradiation includes local irradiation, whole body irradiation, or a combination thereof.

[0090] In a ninth aspect of the present invention, a method for treating a tumor in a subject in need thereof is provided, comprising the step of administering a therapeutically effective amount of a PARP inhibitor and a chemotherapeutic drug to the subject.

[0091] In another preferred embodiment, the method further comprises the step of: performing other treatments on the subject selected from the group consisting of radiotherapy, surgical treatment, targeted therapy, immunotherapy, or a combination thereof.

[0092] In another preferred embodiment, the tumor is a tumor that expresses or highly expresses CXorf67.

[0093] It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features described in detail below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be listed here one by one. BRIEF DESCRIPTION OF THE DRAWINGS

[0094] Figure 1 shows that platinum-based drugs are more sensitive to tumor cells expressing or overexpressing CXorf67. CXorf67 was restored in Daoy CXorf67KO cells. The next day, different concentrations of Cisplatin or Carboplatin were added for 5 days. Cell viability was measured using the CellTiter-Glo kit, and the IC values ​​of the drugs were calculated. 50 .

[0095] Figure 2 shows that platinum-based drugs can increase the sensitivity of PARP inhibitors to CXorf67-expressing or overexpressing tumor cells. A. Daoy CXof67-KO cells were transfected with EV and CXof67 lentiviruses, or PFA-4 primary cells were transfected with sh-GFP and sh-CXorf67 viruses, respectively. The cells were plated in 96-well plates and treated the next day with cisplatin (0.1 μM) or carboplatin (1 μM). Niraparib was also added at varying concentrations. Five days later, cells were treated with CellTiter-Glo and read using a microplate reader. B. The above cell manipulations were repeated. Three days after drug treatment, cells were harvested and apoptosis was assessed using Annexin-V / PI reagent.

[0096] Figure 3 shows that PARP inhibitors can increase the sensitivity of platinum-based drugs to CXorf67-expressing or overexpressing tumor cells. A. Daoy CXof67-KO cells were transfected with EV and CXof67 lentiviruses, cultured for 72 hours, and plated in 96-well plates. The next day, cells were treated with DMSO or niraparib (1 μM) and stimulated with varying concentrations of cisplatin. Five days later, cells were treated with CellTiter-Glo and read using a microplate reader. B. PFA-4 primary cells were transfected with sh-GFP and sh-CXorf67 lentiviruses, respectively. 60 hours later, cells were plated in 96-well plates, and the above drug treatment procedures were repeated for cell viability assays.

[0097] Figure 4 shows that PARP inhibitors combined with platinum drugs can inhibit the growth of CXorf67-expressing or over-expressing tumors. A. C67-KO and C67-KO complemented with CXorf67Daoy cells that stably express Luciferase were constructed and injected into the cerebellum of nude mice. After 7 days, they were randomly divided into 4 groups and treated with Vehicle, Cisplatin, Niraparib and Cisplatin combined with Niraparib. The size of the tumor was detected every 7 days. B. The constructed PFA-4 PDX model was expanded and cultured and randomly divided into four groups of 5 mice each, and the same above-mentioned drug treatment was given. The tumor volume of the mice and the mice were measured every 7 days. When the tumor volume reached 800mm 3 The mice were euthanized and the survival curves were calculated. DETAILED DESCRIPTION

[0098] After extensive and in-depth research and extensive screening, the present inventors unexpectedly discovered for the first time that tumor cell lines or tumor tissues expressing or overexpressing CXorf67 are more sensitive to PARP inhibitors combined with chemotherapy (e.g., PARP inhibitors combined with platinum drugs). Therefore, the CXorf67 gene or its protein can be used as a marker to detect the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy. Based on this, the present inventors completed the present invention.

[0099] the term

[0100] In order to make the present invention easier to understand, certain technical and scientific terms are specifically defined below. Unless otherwise clearly defined in this article, all other technical and scientific terms used herein have the meanings generally understood by those of ordinary skill in the art to which the present invention belongs. Before describing the present invention, it should be understood that the present invention is not limited to the specific methods and experimental conditions described, because such methods and conditions can change. It should also be understood that the terms used herein are intended only to describe specific embodiments and are not intended to be restrictive, and the scope of the present invention will be limited only by the appended claims.

[0101] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. As used herein, when used in reference to a specific recited value, the term "about" means that the value may vary by no more than 1% from the recited value. For example, as used herein, the expression "about 100" includes all values ​​between 99 and 101 (e.g., 99.1, 99.2, 99.3, 99.4, etc.).

[0102] As used herein, the term "treatment" refers to administering an internal or external therapeutic agent comprising a pharmaceutical composition or active ingredient of the present invention to a patient experiencing one or more symptoms of a disease for which the therapeutic agent is known to have a therapeutic effect. Typically, the therapeutic agent is administered to a patient in an amount effective to alleviate one or more symptoms of the disease (a therapeutically effective amount).

[0103] Tumors with defects in the HR repair pathway

[0104] The homologous recombination (HR) repair pathway is an error-free repair mechanism that primarily repairs DNA breaks during the S / G2 phase of the cell cycle, utilizing homologous DNA within the cell as a template. Defects or deficiencies in HR can lead to genomic instability, which in turn contributes to the development and progression of tumors. For example, mutations in genes such as BRCA1, PLAB2, and BRCA2, which are involved in the HR pathway, have been found in many tumors. Mutations in these genes can prevent effective repair of double-strand breaks in DNA, leading to changes in gene copy number, rearrangements, and mutations, ultimately contributing to the development of tumors and diseases such as ovarian, breast, pancreatic, and prostate cancers.

[0105] In a preferred embodiment, the tumor with HR repair pathway defect is a tumor expressing or overexpressing CXorf67, selected from the following group: ependymoma (Ependymoma posterior fossa group A), kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP) ependymoma, kidney renal clear cell carcinoma (KIRC), renal papillary cell carcinoma, or a combination thereof.

[0106] Ependymomas (EPNs) are malignant neuroepithelial tumors that occur in the central nervous system (CNS) in both children and adults. EPNs primarily occur in three locations: supratentorial (ST), posterior fossa (PF), and spinal column (SP). A 2015 study further classified ependymomas in these three locations into molecular subtypes using DNA methylation analysis. ST subtypes include ST-SE, ST-EPN-YAP1, and ST-EPN-RELA; PF subtypes include PF-SE, PFA, and PFB; and SP subtypes include SP-SE, SP-MPE, and SP-EPN. PFA primarily occurs in infants and young children (mean age 3 years, range 0-51 years) and has a poor prognosis. PFB primarily occurs in adolescents (mean age 30 years, range 10-65 years) and has a better prognosis. The PFA subtype of ependymoma mainly occurs in the hindbrain of children and has a poor prognosis. Currently, surgery and radiotherapy are the main treatments, and there is a lack of effective drug treatments.

[0107] sample

[0108] As used herein, the terms "sample" or "specimen" refer to a material that is specifically associated with a subject, from which specific information about the subject can be determined, calculated, or inferred. A sample can consist entirely or in part of biological material from the subject. A sample can also be a material that has been in contact with the subject in a manner that allows testing of the sample to provide information about the subject. A sample can also be a material that has been in contact with other materials that were not from the subject, but that enable the first material to be subsequently tested to determine information about the subject, for example, a sample can be a cleaning solution from a probe or scalpel. A sample can be a source of biological material other than that in contact with the subject, as long as a person skilled in the art can still determine information about the subject from the sample.

[0109] Express

[0110] As used herein, the term "expression" includes the production of mRNA from a gene or gene portion, and includes the production of a protein encoded by RNA or a gene or gene portion, and also includes the appearance of a detection substance associated with expression. For example, cDNA, the binding of a binding ligand (such as an antibody) to a gene or other oligonucleotide, protein or protein fragment, and the chromogenic portion of the binding ligand are all included within the scope of the term "expression." Thus, an increase in half-dot density on an immunoblot such as a western blot also falls within the scope of the term "expression" based on biological molecules.

[0111] Reference value

[0112] As used herein, the term "reference value" refers to a value that is statistically correlated with a particular outcome when compared to an assay result. In a preferred embodiment, the reference value is determined based on statistical analysis of studies comparing CXorf67 expression with known clinical outcomes. Several such studies are presented in the Examples section herein. However, studies from the literature and user experience with the methods disclosed herein can also be used to generate or adjust reference values. Reference values ​​can also be determined by considering circumstances and outcomes specifically related to the patient's medical history, genetics, age, and other factors.

[0113] In the present invention, the reference value refers to a cut-off value, which refers to the expression level of CXorf67 in tumor cells or tissues with HR repair pathway defects. Preferably, the expression level of CXorf67 is >0.5, more preferably >1.5, and more preferably >2.

[0114] Non-tumor cell samples

[0115] As used herein, the term "a sample of non-tumor cells" includes, but is not limited to, a human population that does not have a tumor with a defect in the HR repair pathway.

[0116] CXorf67 protein and polynucleotide

[0117] In the present invention, the terms "protein of the present invention," "CXorf67 protein," and "CXorf67 polypeptide" are used interchangeably to refer to proteins or polypeptides having the amino acid sequence of CXorf67. These include CXorf67 proteins with or without an initiator methionine. Furthermore, the term also encompasses full-length CXorf67 and fragments thereof. The CXorf67 protein referred to herein includes its complete amino acid sequence, its secreted protein, its mutants, and its functionally active fragments.

[0118] CXorf67 (chromosome X open reading frame 67) is located at chromosome Xp11.22 and contains only one exon and no introns, encoding 503 amino acids. CXorf67 is a protein of unknown function, primarily localized in the nucleus. Website predictions indicate no known domain structure and are largely disordered.

[0119] The human CXorf67 protein is 503 amino acids in length (Accession No. NP_981952.1, ACCESSION: NP_981952XP_291352, VERSION: NP_981952.1), and the mouse CXorf67 protein is 589 amino acids in length (Accession No. NP_001159905.1, ACCESSION: NP_001159905, VERSION: NP_001159905.1).

[0120] In the present invention, the terms "CXorf67 gene" and "CXorf67 polynucleotide" are used interchangeably and both refer to nucleic acid sequences having the CXorf67 nucleotide sequence.

[0121] The full-length genome sequence of the human CXorf67 gene is 1896 bp (NCBI GenBank accession number is Gene ID: 340602), and the full-length mRNA sequence of its transcription product is 1512 bp (NCBI GenBank accession number is NM_203407.3).

[0122] The full-length of the mouse CXorf67 gene in the genome is 2203 bp (NCBI GenBank accession number: Gene ID: 102991), and the full-length of its transcription product mRNA sequence is 1770 bp (NCBI GenBank accession number: NM_001166433.1).

[0123] Human and mouse CXorf67 have a 39% similarity at the DNA level and a 39% similarity in protein sequence.

[0124] It should be understood that when encoding the same amino acid, the substitution of nucleotides in the codon is acceptable. In addition, it should be understood that when the nucleotide substitution produces a conservative amino acid substitution, the change of nucleotides is also acceptable.

[0125] Once an amino acid fragment of CXorf67 is obtained, a nucleic acid sequence encoding it can be constructed based on it, and specific probes can be designed based on the nucleotide sequence. The full-length nucleotide sequence or fragments thereof can typically be obtained using PCR amplification, recombinant methods, or synthetic methods. For PCR amplification, primers can be designed based on the CXorf67 nucleotide sequence disclosed herein, particularly the open reading frame sequence. Using a commercially available cDNA library or a cDNA library prepared by conventional methods known to those skilled in the art as a template, the relevant sequence can be amplified. Long sequences often require two or more PCR amplifications, followed by splicing the amplified fragments together in the correct order.

[0126] Once the relevant sequence is obtained, it can be obtained in large quantities by recombinant methods. This is usually done by cloning it into a vector, then transferring it into cells, and then isolating the relevant sequence from the propagated host cells by conventional methods.

[0127] In addition, the sequences can also be synthesized by artificial synthesis, especially when the fragment length is shorter. Usually, a long fragment can be obtained by synthesizing multiple small fragments and then connecting them.

[0128] Currently, DNA sequences encoding proteins of the present invention (or fragments or derivatives thereof) can be obtained entirely through chemical synthesis, which can then be introduced into various existing DNA molecules (such as vectors) and cells known in the art.

[0129] PARP inhibitors

[0130] In 2005, two laboratories simultaneously reported that PARP inhibitors could selectively kill tumor cells harboring BRCA1 / 2 mutations. Subsequent studies revealed that tumor cells with BRCAness (human retinal function defects but no BRCA1 / 2 mutations) were also sensitive to PARP inhibitors. The initial hypothesis regarding the mechanism of action of PARP inhibitors was that the addition of PARP inhibitors resulted in defects in the repair of single-strand breaks. After one round of DNA replication, single-strand breaks became double-strand breaks. Normal cells, with their intact HR repair pathway, survived, while cells with HR repair defects were unable to perform repairs and died. However, further research revealed that while various PARP inhibitors demonstrated comparable inhibition of PARP catalytic activity, their cytotoxic effects varied significantly. Why does this occur? It is currently believed that, in addition to inhibiting PARP catalysis, PARP inhibitors also bind (or trap) the PARP protein to DNA, preventing it from dissociating, blocking replication and accelerating the formation of double-strand breaks. This difference in trapping ability is responsible for the varying efficacy of PARP inhibitors.

[0131] Currently, a variety of PARP inhibitors have been approved for clinical treatment, such as Olaparib, Rucaparib, Niraparib and Talazoparib from abroad, and Fluzoparib and Pamiparib from China, which have been approved for the treatment of ovarian cancer and breast cancer with BRCA mutations.

[0132] It has been found that Niraparib and Pamiparib can pass through the blood-brain barrier (BBB) ​​well, and Olaparib has also been reported to pass through the BBB in brain tumor patients. However, some people believe that the integrity of the BBB in brain tumor patients has been destroyed, so the BBB does not matter. The current progress in neuro-oncology is mostly concentrated in preclinical and clinical research, and has not yet been approved. In addition to the development of new PARP inhibitors, there are many studies to expand their indications, such as identifying new markers and combination therapies. In fact, in many CNS tumors, there are few HR gene mutations, but some markers that indirectly cause BRCAness have been found, such as the expression or high expression of CXorf67 previously discovered by the inventors can inhibit HR repair and thus increase the sensitivity of PARP inhibitors.

[0133] Typically, the PARP inhibitor of the present invention is selected from the group consisting of Talazoparib, Olaparib, Veliparib, Rucaparib, Niraparib, Fluzoparib, Pamiparib, or a combination thereof.

[0134] In the present invention, the inventors, after extensive screening, unexpectedly discovered for the first time that tumor cell lines or tumor tissues expressing or overexpressing CXorf67 are more sensitive to PARP inhibitors combined with chemotherapy (e.g., PARP inhibitors combined with platinum drugs). Therefore, the CXorf67 gene or its protein can be used as a marker to detect the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy. In addition, PARP inhibitors combined with platinum drugs also have a synergistic therapeutic effect on tumor cell lines or tumor tissues expressing or overexpressing CXorf67. On this basis, the inventors completed the present invention.

[0135] Platinum drugs

[0136] The mechanism of action of platinum-based drugs is to halt DNA replication in tumor cells and prevent cell division. After entering tumor cells, platinum-based drugs hydrolyze into hydrates, which further deprotonate to form hydroxylated coordination ions. These ions are highly reactive and, in vivo, complex with the two guanine bases at the N27 position of DNA to form a closed five-membered chelate ring. This disrupts the hydrogen bonds between the purine and cytosine groups on the two polynucleotide chains, disrupting the normal double helix structure of DNA and causing its partial denaturation and inactivation, resulting in a loss of replication capacity, thus achieving its anti-cancer effect.

[0137] Typically, the platinum drug of the present invention is selected from the group consisting of Cisplatin, Carboplatin, Nedaplatin, Oxaliplatin, lobaplatin, or a combination thereof.

[0138] In the present invention, the inventors unexpectedly discovered that platinum drugs and PARP inhibitors have a synergistic effect.

[0139] Pharmaceutical compositions or active ingredient combinations and uses thereof

[0140] The present invention also provides a pharmaceutical composition for synergistically treating CXorf67-expressing or overexpressing tumors, wherein the pharmaceutical composition comprises: a PARP inhibitor, a chemotherapy drug, and a pharmaceutically acceptable carrier, diluent, or excipient.

[0141] The present invention also provides a use of an active ingredient combination for preparing a medicament or formulation for treating tumors expressing or overexpressing CXorf67, or for synergistically treating tumors expressing or overexpressing CXorf67, wherein the active ingredient combination comprises: (Z1) a first active ingredient: a PARP inhibitor; and (Z2) a second active ingredient: a chemotherapeutic agent. Preferably, the medicament or formulation further comprises another tumor therapeutic agent, or a pharmaceutically acceptable carrier, diluent, or excipient.

[0142] Preferably, the drug, preparation or pharmaceutical composition may also be used in combination with other treatment methods, and the other treatment methods are selected from the following group: radiotherapy, surgical treatment, targeted therapy, immunotherapy, or a combination thereof.

[0143] Typically, the PARP inhibitor is selected from the following group: Talazoparib, Olaparib, Veliparib, Rucaparib, Niraparib, Fluzoparib, Pamiparib, or a combination thereof; the chemotherapy drug is a platinum drug selected from the following group: Cisplatin, Carboplatin, Nedaplatin, Oxaliplatin, lobaplatin, or a combination thereof.

[0144] Typically, the CXorf67-expressing or high-expressing tumor includes a tumor with a defective HR repair pathway, which can be a primary tumor and / or a metastatic tumor selected from the following group: ependymoma (Ependymoma posterior fossa group A), kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP), or a combination thereof; wherein the ependymoma includes PFA type.

[0145] Detection method

[0146] Taking advantage of the fact that CXorf67 is present in tumor cells or tissues (such as tumors with HR repair pathway defects, preferably ependymomas) and is closely related to the sensitivity to PARP inhibitors combined with chemotherapy, the present invention also provides a method for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0147] In a preferred embodiment of the present invention, the present invention provides a high-throughput second-generation sequencing method for detecting CXorf67 as well as Sanger sequencing, fluorescence quantitative PCR (qPCR), in situ immunofluorescence (FISH) and immunohistochemistry.

[0148] Detection kit

[0149] Based on the correlation between CXorf67-expressing or high-expressing tumor cells and the sensitivity to PARP inhibitors combined with chemotherapy, tumor cells expressing or high-expressing CXorf67 are more sensitive to PARP inhibitors combined with chemotherapy. Therefore, CXorf67 can be used as a biomarker to guide the use of PARP inhibitors combined with chemotherapy.

[0150] The present invention also provides a diagnostic kit for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy, which contains a detection reagent for detecting the CXorf67 gene, mRNA, cDNA, or protein; and a label or instructions, wherein the label or instructions indicate that the kit is used to detect the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0151] Detection methods and kits

[0152] The present invention relates to diagnostic assays for quantitative and localized detection of human CXorf67 protein or mRNA levels. These assays are well known in the art. The human CXorf67 protein levels detected in these assays can be used to assess the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0153] A method for detecting whether CXorf67 protein is present in a sample is to use a specific antibody against CXorf67 protein for detection, which comprises: contacting the sample with the specific antibody against CXorf67 protein; and observing whether an antibody complex is formed. The formation of the antibody complex indicates the presence of CXorf67 protein in the sample.

[0154] CXorf67 protein or its polynucleotide can be used for the diagnosis and treatment of diseases associated with CXorf67 protein. Part or all of the polynucleotide of the present invention can be immobilized as a probe on a microarray or DNA chip for analysis of differential gene expression in tissues and genetic diagnosis. Antibodies against CXorf67 can be immobilized on a protein chip for detection of CXorf67 protein in a sample.

[0155] The main advantages of the present invention include:

[0156] 1) The present inventors have discovered for the first time that the CXorf67 gene, mRNA, cDNA or protein or its detection reagent can be used as a marker for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0157] 2) The inventors have discovered for the first time that the CXorf67 gene, mRNA, cDNA or protein or its detection reagent can also be used to prepare a diagnostic reagent or kit for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

[0158] 3) The present inventors discovered for the first time that platinum drugs can increase the sensitivity of PARP inhibitors to CXorf67-expressing or overexpressing tumor cells.

[0159] 4) The present inventors discovered for the first time that PARP inhibitors can increase the sensitivity of platinum drugs to CXorf67-expressing or overexpressing tumor cells.

[0160] 5) The present inventors discovered for the first time that PARP inhibitors combined with platinum drugs can inhibit the growth of CXorf67-expressing or overexpressing tumors.

[0161] 6) The present inventors discovered for the first time that PARP inhibitors combined with chemotherapy (such as platinum drugs) have a synergistic killing effect on tumor cells expressing or overexpressing CXorf67.

[0162] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are intended to illustrate the present invention only and are not intended to limit the scope of the present invention. The experimental methods in the following examples, for which specific conditions are not specified, are generally performed under conventional conditions, such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or according to the conditions recommended by the manufacturer. Unless otherwise stated, percentages and parts are by weight.

[0163] Experimental Materials

[0164] Daoy cell line was obtained from the Stem Cell Bank of the Chinese Academy of Sciences and cultured in MEM (supplemented with 10% FBS, non-essential amino acids, GlutaMAX, and Sodium pyruvate).

[0165] The CXorf67-KO Daoy cell line was constructed using CRISPR-Cas9 technology.

[0166] PFA-type ependymoma primary cells were isolated and cultured from PDX tumors in this experiment and cultured in Neurobasal medium (supplemented with B2, N2, GLutaMAX, EGF, FGF, and heparin). All cells were cultured in an incubator at 37°C with 5% CO2.

[0167] Niraparib (HY-10619), cisplatin (HY-17394), and carboplatin (HY-17393) were purchased from MCE. CXorf67-shRNA and GFP-shRNA plasmids were constructed using the pLKO.1 vector. CXorf67 (HPA004003) antibody was purchased from Sigma-Aldrich. GAPDH (10494-1-AP) antibody was purchased from Proteintech. FITC Annexin V apoptosis detection kit (#556547) was purchased from BD Biosciences.

[0168] General Methods

[0169] 1. Cell Survival Assay

[0170] (1) To test the effect of CXorf67 on the sensitivity of platinum drugs (cisplatin and carboplatin), CXorf67-KO and CXorf67-KO Daoy cells were plated in 96-well plates. The next day, different concentrations of cisplatin and carboplatin were added, and the cells were cultured for another five days, with additional culture medium and drugs added as needed. On the fifth day, 50 μl of CellTiter-Glo reagent was added, and the cells were lysed at room temperature for 20 minutes, and then the fluorescence values ​​were read using a microplate reader.

[0171] (2) In the experiment to test the sensitivity of platinum drugs and PARP inhibitors (Niraparib) to CXorf67-expressing or overexpressing tumor cells, CXorf67-KO and KO-complemented CXorf67 Daoy cells were also plated in 96-well plates. On the second day, different concentrations of Niraparib were added and combined with Cisplatin (0.1μM) or Carboplatin (1μM); or different concentrations of Cisplatin were added and combined with Niraparib (1μM). After 5 days of culture, the viability was measured as described above. For PDX-derived primary cells, the cells were knocked down for CXorf67 (sh-CXorf67), and sh-GFP was used as a control, and then plated in 96-well plates. Subsequently, the cells were treated and viability was measured as described above with different concentrations of Niraparib combined with platinum drugs, or different concentrations of Cisplatin combined with Niraparib.

[0172] 2. Cell Apoptosis Experiment

[0173] The Daoy CXorf67-KO cell line was cultured and then infected with a CXorf67 overexpressing lentivirus. After screening, the cells were stably cultured for 2-3 generations, and then the KO and CXorf67-replenished cells were plated into 96-well plates. The next day, Niraparib (0.5μM), Cisplatin (0.1μM), and Carboplatin (1μM) were treated alone, or Niraparib combined with Cisplatin and Niraparib combined with Carboplatin were treated. After the cells were cultured for another 3 days, they were digested and collected. The cells were washed twice with the Binding buffer in the Annexin-V / PI kit, and then incubated with FITC-Annexin and PI at room temperature for 15 minutes. The cells were then analyzed using a flow cytometer.

[0174] 3. Mouse Brain Orthotopic Tumor Transplantation Experiment

[0175] All mouse experiments and procedures were approved by the Animal Ethics Committee of the Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences. Six-week-old female nude mice were purchased from Shanghai Slack and housed in an SPF environment.

[0176] The constructed CXorf67-KO and CXorf67-KO Daoy cells were transfected with Luciferase lentivirus, and 2×10 5 The cells were injected into the cerebellum of nude mice (2mm below the lambda line, 1mm to the right of the midline, and 3mm in depth). The tumor formation was then determined by in vivo imaging, and the mice were then randomly divided into four groups: control group, Niraparib (50mg / kg) group, Cisplatin (2mg / kg) group, and Niraparib combined with Cisplatin group. Cisplatin was dissolved in saline and administered by intraperitoneal injection once a week. Niraparib was dissolved in 0.5% methylcellulose and administered orally 5 times a week. Subsequently, the tumor size was detected once every seven days by IVIS SpectrumCT imaging. Each mouse was injected with 200μl of D-luciferin substrate (15mg / ml) and detected ten minutes later. The signal intensity was then calculated by software.

[0177] 4. Mouse subcutaneous transplant tumor experiment

[0178] The PFA-4 ependymoma-derived PDX model established in the laboratory was expanded and passaged subcutaneously in nude mice. The obtained tumor was digested into single cells and counted. The cells (3×10 6 After mixing with Matrigel at a ratio of 1:1, the mixture was inoculated subcutaneously into 7-week-old female nude mice. 3 Around 60 days, mice were randomly divided into four groups: a control group, a niraparib (50 mg / kg) group, a cisplatin (2 mg / kg) group, and a niraparib combined with cisplatin group. Cisplatin was dissolved in saline and administered via intraperitoneal injection once a week. Niraparib was dissolved in 0.5% methylcellulose and administered orally five times a week. Mouse body weight and tumor volume were measured weekly, calculated using the formula (length * width * width) / 2.

[0179] Example 1. Tumor cells expressing CXorf67 are more sensitive to platinum drugs

[0180] First, CXorf67 expression was restored in Daoy CXorf67KO cells. The next day, different concentrations of Cisplatin or Carboplatin were added for treatment for 5 days, and the cell viability was detected using the CellTiter-Glo kit.

[0181] As shown in FIG1 , tumor cells expressing CXorf67 were found to be more sensitive to cisplatin and carboplatin.

[0182] Example 2. Platinum drugs can increase the sensitivity of PARP inhibitors to tumor cells expressing CXorf67

[0183] As shown by the data in Example 1, platinum alone can increase the sensitivity of CXof67-expressing cells to drugs. In this example, we will further explore whether platinum drugs can increase the sensitivity of PARP inhibitors.

[0184] The inventors first restored CXof67 in Daoy-CXof67KO cells, then treated them with Cisplatin and Carboplatin respectively, and simultaneously added different concentrations of Niraparib for 5 days, and tested the cell activity using Celltiter-Glo.

[0185] As shown in FIG2A , it was found that Cisplatin or Carboplatin indeed significantly increased the inhibitory effect of PARP inhibitors on cells expressing CXof67.

[0186] Secondly, the inventors also conducted a CXof67 knockdown experiment in PFA-type ependymoma-derived tumor cells.

[0187] As shown in Figure 2A , in contrast to the overexpression experiments, knockdown of CXof67 significantly inhibited the sensitization effect of cisplatin or carboplatin to PARP inhibitors.

[0188] In addition, the inventors used the Annexin-V / PI kit to test the killing effects of Cisplatin and Carboplatin combined with PARP inhibitors on cells with and without CXorf67.

[0189] As shown in FIG2B , it was also found that the apoptotic ratio of cells in the combination treatment groups (ie, Niraparib+Carboplatin; Niraparib+Cisplatin) was significantly increased compared with the single treatment groups (ie, Niraparib, Carboplatin, Cisplatin alone).

[0190] The above results confirm that platinum drugs can increase the sensitivity of PARP inhibitors (such as Niraparib) to tumor cells expressing CXorf67, and the effect of the combination of the two is significantly better than that of single use.

[0191] Example 3. PARP inhibitors can increase the sensitivity of platinum drugs to tumor cells expressing CXorf67

[0192] In Example 2, it was demonstrated that platinum drugs can increase the sensitivity of PARP inhibitors to CXorf67-expressing or overexpressing tumor cells. In this example, we will further explore whether PARP inhibitors can increase the sensitivity of platinum drugs.

[0193] The inventors also restored CXof67 in CXof67KO Daoy cells, treated them with different concentrations of Cisplatin, and simultaneously treated them with 1 μM Niraparib or DMSO for 5 days, and tested the cell activity using Celltiter-Glo.

[0194] As shown in Figure 3A, Niraparib reduced the IC of cisplatin in cells replenished with CXorf67. 50 The decrease from 451nM to 62.52nM did significantly increase the sensitivity of Cisplatin.

[0195] In addition, the inventors tested the effect of Niraparib on cislatin sensitivity in CXorf67 knockdown PFA-4 PDX-derived primary tumor cells.

[0196] As shown in Figure 3B, consistent with the previous results, the IC of Cislatin can be significantly reduced in the presence of CXorf67. 50 ; However, when CXorf67 was knocked down, the synergistic effect was significantly weakened.

[0197] The above results confirmed that PARP inhibitors can increase the sensitivity of platinum drugs to CXorf67-expressing or high-expressing tumor cells, and the effect of combining the two is significantly better than that of using them alone.

[0198] Example 4. PARP inhibitors combined with platinum drugs can inhibit the growth of tumors expressing CXorf67

[0199] To verify whether platinum drugs and PARP inhibitors also have synergistic effects in vivo, the inventors first constructed Daoy C67-KO and KO cells stably transfected with Luciferase, in which CXorf67 was complemented. The cells were injected into the cerebellum of immunodeficient mice, and tumor formation was monitored approximately 7 days later. The mice were then randomly divided into four groups: ① Vehicle (empty vector); ② Cisplatin (2 mg / kg, intraperitoneal injection, once a week); ③ Niraparib (50 mg / kg, gavage, five times a week); and ④ Niraparib (50 mg / kg, gavage, five times a week) + Cisplatin (2 mg / kg, intraperitoneal injection, once a week). After dosing, tumor size was assessed weekly using in vivo imaging.

[0200] As shown in FIG4A , the results showed that both Niraparib and Cisplatin could reduce the volume of tumors (especially CXorf67-expressing or high-expressing tumors), and the combination of Niraparib and Cisplatin could significantly inhibit tumor formation.

[0201] In addition, the same assay was performed on a PFA-type ependymoma patient-derived PDX model. PFA-4 cells were mixed with Matrigel 1:1 and injected subcutaneously into BALB / c nude mice (3×10 6 cells / mouse), 5 mice per group, when the tumor volume reached 100 mm 3 Around 6:00 p.m., mice were administered with ① Vehicle; ② Cisplatin (2 mg / kg, intraperitoneal injection, once a week); ③ Niraparib (50 mg / kg, oral gavage, five times a week); and ④ Niraparib (50 mg / kg, oral gavage, five times a week) + Cisplatin (2 mg / kg, intraperitoneal injection, once a week). Tumor volume and mouse body weight were measured weekly.

[0202] As shown in FIG4B , the results showed that both Niraparib and Cisplatin can reduce the volume of tumors (especially CXorf67-expressing or high-expressing tumors), and the combination of the two can significantly inhibit tumor growth, showing a synergistic effect.

[0203] The specific experimental data of the above results are shown in Tables 1 to 4:

[0204] Table 1

[0205] Table 2

[0206] Table 3

[0207] Table 4

[0208] The data in Tables 1 to 4 are averaged and summarized in Table 5:

[0209] Table 5

[0210] According to the data in Table 5, it can be seen that 36 days after treatment, Z3>Z1+Z2 (i.e., 739.3354>426.0953+262.4895), indicating that the degree of tumor volume reduction achieved by the combination of Niraparib and Cisplatin exceeded the sum of the tumor volume reductions achieved by Niraparib alone and Cisplatin alone.

[0211] From the above analysis, it can be seen that PARP inhibitors and chemotherapy (including platinum drugs) have a synergistic killing effect on tumor cells expressing or overexpressing CXorf67.

[0212] All documents mentioned in this application are incorporated herein by reference, just as if each document were incorporated herein by reference individually. It should also be understood that after reading the above teachings of the present invention, those skilled in the art may make various changes or modifications to the present invention, and that such equivalents also fall within the scope of the claims appended hereto.

[0213] discuss

[0214] CXorf67 is located at position p11.22 on chromosome X and contains only one exon and no introns. Its CDS (Coding Sequence) is 1512 base pairs, encoding 503 amino acids. Work in the inventor's laboratory in 2020 discovered another function of CXorf67: it can influence the efficiency of DNA homologous recombination (HR) repair by regulating the binding of PALB2 and BRCA2.

[0215] The PARP (poly(ADP-ribose) polymerase) family currently consists of 17 members, primarily located in the nucleus. They share a common ART (ADP-ribosyltransferase) catalytic motif, which catalyzes PARylation. PARP1 is the most intensively studied member. As a DNA damage-sensing protein, it uses NAD+ to link the negatively charged PAR to DNA damage repair proteins, transmitting repair signals. PARP1 was initially found to play a role in single-strand break and base excision repair, and more recently, it has been shown to play a role in nucleic acid excision, mismatch repair, and homologous recombination repair.

[0216] Double-strand breaks (DSBs) in cellular DNA are the most serious type of DNA damage faced by cells. DSB repair is generally categorized into two pathways, depending on whether homologous sequences are involved. One is classical non-homologous end joining (cNHEJ), which is independent of homologous sequences and primarily operates during the G0 / G1 phase of the cell cycle. The other is the homologous recombination (HR) repair pathway, an error-free repair pathway that primarily repairs DNA breaks during the S / G2 phase of the cell cycle using homologous DNA within the cell as a template. This homologous DNA can be sequences on sister chromatids, homologous chromosomes, or ectopic chromosomes.

[0217] Tumors with defects in the HR repair pathway are sensitive to PARP inhibitors. When PARP inhibitors block the repair of single-strand breaks in DNA, the single-strand breaks in DNA will turn into double-strand breaks after one round of replication. The HR repair defects in tumor cells will lead to cell death; while normal cells have complete HR repair, so they will not be killed. The inventors' preliminary results found that the expression or high expression of CXorf67 can inhibit DNA homologous recombination repair by competing with BRCA2 for binding to PALB2, and for the first time proposed that CXorf67 can be used as a biomarker for the use of PARP inhibitors in tumor cells. In addition, the inventors also found that PARP inhibitors combined with radiotherapy can more effectively kill tumor cells that express or overexpress CXorf67. However, the effectiveness of the combination of PARP inhibitors with other treatment methods remains unclear, and how to further efficiently kill tumors with defects in the HR repair pathway remains unknown.

[0218] Based on the above needs, in the present invention, the inventors unexpectedly discovered for the first time after a large number of screenings that tumor cell lines or tumor tissues expressing or overexpressing CXorf67 are more sensitive to PARP inhibitors combined with chemotherapy (such as PARP inhibitors combined with platinum drugs). Therefore, the CXorf67 gene or its protein can be used as a marker to detect the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy. In addition, PARP inhibitors combined with platinum drugs also have a synergistic killing effect on tumor cell lines or tumor tissues expressing or overexpressing CXorf67. On this basis, the present invention provides a more sensitive and effective treatment method for tumors with defects in the HR repair pathway.

Claims

1. Use of an active ingredient combination, characterized in that: The active ingredient combination is used to prepare a drug or preparation for treating a tumor expressing or over-expressing CXorf67, or a drug or preparation for synergistically treating a tumor expressing or over-expressing CXorf67, wherein the active ingredient combination comprises: (Z1) a first active ingredient: a PARP inhibitor; and (Z2) Second active ingredient: chemotherapy drug.

2. The use according to claim 1, characterized in that The PARP inhibitor is selected from the group consisting of Talazoparib, Olaparib, Veliparib, Rucaparib, Niraparib, Fluzoparib, Pamiparib, or a combination thereof; preferably, the PARP inhibitor is selected from the group consisting of Niraparib.

3. The use according to claim 1, characterized in that The chemotherapy drug is a platinum drug, and the platinum drug is selected from the following group: Cisplatin, Carboplatin, Nedaplatin, Oxaliplatin, lobaplatin, or a combination thereof; preferably, the platinum drug is selected from: Carboplatin and / or Cisplatin.

4. The use according to claim 1, characterized in that The medicine or preparation also includes other tumor therapeutic agents.

5. The use according to claim 1, characterized in that The CXorf67 expressing or highly expressing tumor is selected from the following group: ependymoma posterior fossa group A, kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP), or a combination thereof.

6. The use according to claim 1, characterized in that The tumors include tumors with defects in the HR repair pathway.

7. A pharmaceutical composition, characterized in that The pharmaceutical composition comprises: a PARP inhibitor, a chemotherapeutic drug and a pharmaceutically acceptable carrier, diluent or excipient.

8. The pharmaceutical composition according to claim 7, characterized in that The dosage of the PARP inhibitor is 10-100 mg / kg, preferably 20-80 mg / kg, more preferably 30-70 mg / kg, and most preferably 50 mg / kg.

9. The pharmaceutical composition according to claim 7, characterized in that The chemotherapy drug is a platinum drug, and the dosage of the platinum drug is 0.1-20 mg / kg, preferably 0.5-10 mg / kg, more preferably 1-5 mg / kg, and most preferably 2 mg / kg.

10. A medicine box, characterized in that: The medicine kit comprises: (a) a first preparation, which contains a PARP inhibitor and a pharmaceutically acceptable carrier; (b) a second formulation comprising a chemotherapeutic drug and a pharmaceutically acceptable carrier; and (c) instructions describing a method of using the first agent and the second agent in combination to treat a tumor.

11. A use of CXorf67 gene, mRNA, cDNA or protein or a detection reagent thereof, characterized in that: (i) used as a marker for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy; and / or (ii) used for preparing a diagnostic reagent or kit for detecting the sensitivity of tumor cells to PARP inhibitors combined with chemotherapy.

12. A diagnostic kit for detecting the sensitivity of tumor cells to PARP inhibitor combined with chemotherapy, characterized in that: The kit comprises a container containing a detection reagent for detecting CXorf67 gene, mRNA, cDNA or protein; and a label or instructions, wherein the label or instructions indicate that the kit is used to detect the sensitivity of tumor cells to PARP inhibitor combined with chemotherapy.

13. A method for determining sensitivity to PARP inhibitor combined with chemotherapy, characterized in that: The method comprises: a) providing a test sample from a subject; b) detecting the expression level of CXorf67 protein in the test sample; and c) based on the expression level of CXorf67 protein determined in step b), determining the sensitivity to PARP inhibitor combined with chemotherapy.

14. A method for determining a treatment regimen, characterized in that: The method comprises: a) providing a test sample from a subject; b) detecting the expression level of CXorf67 protein in the test sample; and c) determining a treatment regimen based on the expression level of CXorf67 protein in the sample.

15. A method for synergistically killing or inhibiting tumor cells in vitro for non-disease treatment purposes, characterized in that: The method comprises the steps of: contacting tumor cells with a PARP inhibitor and a chemotherapy drug, thereby synergistically killing or inhibiting the tumor cells, wherein the tumor cells are tumor cells expressing or highly expressing CXorf67.

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