WO2022022541A1 - Use of rbm10 gene - Google Patents

Abstract

Provided is a biomarker for evaluating the efficacy of Chiauranib or guiding the administration thereof or predicting the therapeutic effect on ovarian cancer. The biomarker is the RBM10 gene, an mRNA thereof, or a protein or protein fragment encoded thereby. The correlation between an RBM10 gene variation and the efficacy of Chiauranib is verified by means of using the progression-free survival (PFS) of a patient as an efficacy index. The detection of RBM10 gene mutation information can guide the clinical administration of Chiauranib and evaluate the efficacy thereof in tumor treatment, particularly for recurrent and advanced ovarian cancer.

Description

Uses of the RBM10 gene technical field

The present invention relates to the technical field of medicine, in particular to the use of RBM10 gene.

Background technique

Ovarian cancer ranks third among gynecological malignant tumors, and its mortality ranks first, and its incidence is increasing year by year. Ovarian cancer patients can achieve complete remission (CR) after ideal cytoreduction (residual tumor lesions < 1 cm) combined with platinum and paclitaxel systemic chemotherapy, but 60%-70% of patients will be Relapse occurs. Relapsed patients are less effective for re-chemotherapy: platinum-sensitive relapsed populations have an effective rate of about 30-80% for platinum-containing chemotherapy. Even platinum-sensitive patients will become platinum-resistant after 2 to 3 platinum-containing treatments; platinum-refractory The effective rate of platinum-free single-agent chemotherapy is only 5-17%, the progression-free survival time is about 3 months, and the overall survival time is less than 1 year.

In recent years, there has been more and more evidence from clinical research on targeted therapy and immunotherapy, especially the application of anti-angiogenic drugs in patients with platinum-refractory and recurrent ovarian cancer. The relevant research mainly includes:

GOG0170D study, a phase II study of bevacizumab monotherapy in patients with platinum-refractory/resistant ovarian cancer, enrolled 62 subjects, ORR was 21%, of which 2 (3.2%) CR, 11 Case (17.7%) PR. Median PFS and OS were 4.7 and 17 months, respectively.

The GOG0170F study, a phase II study of sorafenib monotherapy in ovarian cancer patients with recurrence and disease progression less than 12 months from the last platinum-containing chemotherapy, enrolled 71 patients, including platinum-sensitive 21 (29.6%) and platinum-sensitive Drug resistance was found in 50 cases (70.4%), including 59 evaluable cases. Two patients (3.4%) achieved PR, ORR was 3.4%, and median PFS and OS were 2.1 and 16.3 months, respectively.

The VEG104450 study, pazopanib monotherapy for patients with high recurrence risk ovarian cancer with elevated CA125 after initial treatment, enrolled 36 subjects, of whom 14 (39.0%) were platinum-resistant and 22 (61.0%) were platinum-resistant. %) platinum sensitive. Referring to the GCIG (Gynecological Oncology Society) standard, the response rate was 11 (31.0%), and the response rate of the evaluable cases according to the RECIST standard was 0.

In addition to anti-angiogenic drugs, other mechanism-targeted therapy or immunotherapy has also progressed in platinum-refractory/resistant recurrent ovarian cancer, such as polyadenosine diphosphate ribose polymerase (PARP) inhibitors such as olapa Niraparib and Niraparib have clear curative effect on ovarian cancer patients with BRCA mutation, but they have not obtained significant benefit for most platinum-refractory/resistant recurrent ovarian cancer patients without BRCA mutation. Ovarian cancer requires the search for more effective treatment options.

Cioroni is a small molecule anti-tumor drug targeting multiple protein kinases. It has high selective inhibitory activity on Aurora B, VEGFR/PDGFR/c-Kit, and CSF-1R targets, and has the ability to inhibit tumor cells. Multi-channel synergistic mechanisms such as mitosis, anti-tumor angiogenesis, and regulation of tumor immune microenvironment play a comprehensive anti-tumor effect. Due to the unique anti-tumor mechanism of cioronib, the research on the marker genes related to the efficacy of cioroni will help to find the potential benefit patient population and better therapeutic effect for the clinical application of cioroni .

SUMMARY OF THE INVENTION

In view of this, the purpose of the present invention is to provide the use of RBM10 gene or its mRNA or its encoded protein or protein fragment as a biomarker in evaluating the efficacy of cioronitrile or instructing the administration of cioronib or predicting the therapeutic effect of ovarian cancer .

In the present invention, Cioroni capsules are used to conduct Phase Ib clinical trials for relapsed and refractory advanced ovarian cancer. Through the detection and analysis of plasma cell-free tumor DNA (ctDNA), 548 tumor-related genes are used to conduct accompanying research on therapeutic efficacy-related biomarkers. . Blood samples were drawn from all patients before enrollment, and gene sequences were detected for tumor-related genes, including gene mutations and copy number abnormalities. The detection results selected all genes with a mutation rate of more than 0.4%, and used the patient's progression-free survival (PFS) as the efficacy index to analyze the correlation between tumor-related gene abnormalities and the efficacy of cioronib. The results showed that among the evaluated tumor-related genes, the RBM10 gene was significantly associated with the efficacy of cioronib.

The specific test results showed that the mutation of RBM10 gene was significantly associated with the PFS of cioroni in ovarian cancer patients. Among the 23 evaluable subjects, the median PFS was 232 days in RBM10-mutated patients and 106 days in wild-type patients, with a P value of 0.02. It is indicated that this gene can be a biomarker related to the efficacy of ciorone.

According to the above technical effect, the present invention accordingly relates to the preparation of a reagent for detecting the RBM10 gene or its mRNA or its encoded protein or protein fragment for evaluating the curative effect of cioronib or guiding the administration of cioronib or predicting the treatment of ovarian cancer Use in kits or microarrays for effect. In one embodiment, if there is variation in the RBM10 gene or its mRNA or the protein encoded by the subject compared with the wild type, it indicates that for the subject, cioronib has better curative effect and can The use of cioroni therapy or ovarian cancer treatment is better. In another embodiment, the reagent for detecting the RBM10 gene or its mRNA or its encoded protein or protein fragment is a binding agent that binds to the RBM10 gene encoded protein or protein fragment, or hybridizes with the RBM10 gene or its mRNA Or a substance that amplifies the RBM10 gene or its mRNA. In another embodiment, the binding agent that binds to the protein or protein fragment encoded by the RBM10 gene is an anti-RBM10 antibody. In another embodiment, the substance that hybridizes to or amplifies the RBM10 gene or its mRNA is an oligonucleotide primer or probe.

In another aspect, the present invention relates to a kit or microarray for evaluating the curative effect of cioronib or instructing the administration of cioronib or predicting the therapeutic effect of ovarian cancer, comprising a kit or microarray for detecting the RBM10 gene or its mRNA or its Reagents for encoded proteins or protein fragments. In one embodiment, if there is variation in the RBM10 gene or its mRNA or the protein encoded by the subject compared with the wild type, it indicates that for the subject, cioronib has better curative effect and can The use of cioroni therapy or ovarian cancer treatment is better. In another embodiment, the reagent for detecting the RBM10 gene or its mRNA or its encoded protein or protein fragment is a binding agent that binds to the RBM10 gene encoded protein or protein fragment, or hybridizes with the RBM10 gene or its mRNA Or a substance that amplifies the RBM10 gene or its mRNA. In another embodiment, the binding agent that binds to the protein or protein fragment encoded by the RBM10 gene is an anti-RBM10 antibody, and the substance that hybridizes with the RBM10 gene or its mRNA or amplifies the RBM10 gene or its mRNA is an oligonucleotide Nucleotide primers or probes.

In another aspect, the present invention relates to the use of cioronib in the preparation of a medicament for the treatment of ovarian cancer, especially ovarian cancer with RBM10 gene mutation.

In yet another aspect, the present invention relates to a biomarker for evaluating the efficacy of cioronib or guiding the administration of cioronib or predicting the treatment effect of ovarian cancer, the biomarker is the RBM10 gene or its mRNA or its encoding protein or protein fragment.

In yet another aspect, the present invention relates to the use of the RBM10 gene or its mRNA or its encoded protein or protein fragment as a biomarker in evaluating the efficacy of cioronib or guiding the administration of cioronib or predicting the therapeutic effect of ovarian cancer.

In yet another aspect, the present invention relates to the use of the RBM10 gene or its mRNA or its encoded protein or protein fragment in the preparation of biomarkers for evaluating the efficacy of cioronib or guiding the administration of cioronib or predicting the therapeutic effect of ovarian cancer.

In yet another aspect, the present invention relates to a method for evaluating the curative effect of cioronitrile or instructing the administration of cioronib or predicting the therapeutic effect of ovarian cancer, comprising the following steps:

(1) Obtain a biological sample from the subject; and

(2) detecting whether there is a mutation in the RBM10 gene or its mRNA or its encoded protein or protein fragment in the sample;

Among them, if there is a mutation in the RBM10 gene or its mRNA or its encoded protein or protein fragment in the subject, for the subject, the curative effect of cioronib is better, and the cioronib can be used for treatment or ovarian cancer. Cancer treatment is better.

In one embodiment, the biological sample is ctDNA, tumor tissue, circulating tumor cells, or tissue from other sources of the human body. In one embodiment, the detection method is gene sequencing, PCR, FISH, immunohistochemistry, ELISA, Western or flow cytometry. In some embodiments, the ovarian cancer tumor tissue ctDNA is used as the detection sample, and the next-generation sequencing technology is used for detection.

In yet another aspect, the present invention relates to the use of cioronib in the treatment of ovarian cancer, especially ovarian cancer with mutations in the RBM10 gene.

Description of drawings

Figure 1 shows the progression-free survival of RBM10 mutant and wild-type.

detailed description

Several aspects of the invention are described below with reference to example applications for illustration. It should be understood that numerous specific details, relationships and methods are set forth to provide a thorough understanding of the present invention. However, one of ordinary skill in the relevant art will readily recognize that the invention may be practiced without one or more of the specific details or in other ways.

The present invention relates to the relationship between the mutation and/or expression of a newly discovered biomarker (ie, RBM10) and the efficacy of cioronib, the administration of cioronib, and/or the therapeutic effect of ovarian cancer. The biomarkers described herein provide methods for evaluating the efficacy of cioronib, guiding the administration of cioronib, and/or predicting the effect of treatment in ovarian cancer. Thus, one embodiment of the present invention represents an improvement in biomarkers suitable for evaluating the efficacy of cioronib, guiding the administration of cioronib and/or predicting the effect of ovarian cancer treatment. In yet another embodiment, the newly discovered biomarkers of the present invention (ie, RBM10) may be combined with one or more other cancer markers known in the art (eg, CEA, NSE, CA 19-9, CA 125, CA 72-4, PSA, proGRP, SCC, NNMT, VEGFR2, HER2, MISIIR, VEGFA, CD24) in combination, for example, for evaluating the efficacy of cioronib, guiding the administration of cioronib and/or predicting the treatment effect of ovarian cancer or For the preparation of kits for this purpose.

The invention verifies the correlation between the RBM10 gene variation and the curative effect of cioroni by taking the patient's progression-free survival (PFS) as the curative effect index, and the detection of the RBM10 gene mutation information can guide the clinical medication and treatment of cioroni. To evaluate its curative effect on tumor treatment, especially for ovarian cancer, especially for recurrent and advanced ovarian cancer.

In the present invention, "the curative effect of cioronib is better, the treatment effect of cioronib can be better, or the effect of ovarian cancer treatment is better" especially refers to that cioronib can significantly prolong the progression-free survival period of patients such as ovarian cancer patients (PFS). In certain embodiments, the kit of the present invention is used to evaluate the efficacy of cioronib in the treatment of ovarian cancer and/or to guide the administration of cioronib for the treatment of ovarian cancer.

The term "sample" means a material known or suspected to express or contain a biomarker (i.e. RBM10) or a binding agent, such as an antibody specific for the biomarker (i.e. RBM10). Samples may be derived from biological sources ("biological samples"), such as tissues (eg, biopsy samples), extracts or cell cultures including cells (eg, tumor cells), cell lysates, and biological or physiological fluids, such as Whole blood, plasma, serum, saliva, cerebrospinal fluid, sweat, urine, milk, peritoneal fluid, etc. Samples obtained from sources or after pretreatment to improve sample characteristics (eg, preparation of plasma from blood, dilution of mucus, etc.) can be used directly. In certain aspects of the invention, the sample is a human physiological fluid, such as human serum. In certain aspects of the invention, the sample is a biopsy sample such as tumor tissue or cells obtained by tissue examination. In certain aspects of the invention, the sample is a malignant or normal tissue sample such as a paracancerous normal tissue sample.

Samples that can be analyzed in accordance with the present invention include polynucleotides of clinical origin. As will be understood by those of skill in the art, target polynucleotides can include RNA, including but not limited to total cellular RNA, poly(A)+ messenger RNA (mRNA) or a portion thereof, cytoplasmic mRNA, or RNA transcribed from cDNA ( i.e. cRNA).

The target polynucleotide can be detectably labeled on one or more nucleotides using methods known in the art. Detectable labels can be, without limitation, luminescent, fluorescent, bioluminescent, chemiluminescent, radioactive, and colorimetric labels.

The term "marker" as used herein refers to a molecule to be used as a target for analyzing a patient's experimental sample. Examples of such molecular targets are genes, proteins or polypeptides. The genes, proteins or polypeptides used as markers in the present invention are intended to include naturally occurring variants of said genes or proteins as well as fragments of said genes or proteins or said variants, particularly immunologically detectable fragments . Immunologically detectable fragments preferably comprise at least 6, 7, 8, 10, 12, 15 or 20 contiguous amino acids of the marker polypeptide. One skilled in the art will recognize that proteins released by cells or present in the extracellular matrix may be damaged (eg, during inflammation) and may be degraded or cleaved into such fragments. Certain markers are synthesized in an inactive form, which can then be activated by proteolysis. As the skilled artisan will appreciate, proteins or fragments thereof can also be present as part of a complex. Such complexes can also be used as markers in the sense of the present invention. Variants of marker polypeptides are encoded by the same gene but may differ in their isoelectric point (=PI) or molecular weight (=MW) or both, e.g. as a result of alternative mRNA or pre-mRNA processing . The amino acid sequence of the variant is 95%, 96%, 97%, 98%, 99% or more identical to the corresponding marker sequence. Additionally, or in the alternative, marker polypeptides or variants thereof may carry post-translational modifications. Non-limiting examples of post-translational modifications are glycosylation, acylation and/or phosphorylation.

Expression of the marker can also be identified by detecting translation of the marker (ie, detection of the marker protein in the sample). Methods suitable for detecting marker proteins include any suitable method for detecting and/or measuring proteins from cells or cell extracts. Such methods include, but are not limited to, immunoblotting (eg, Western blotting), enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), immunoprecipitation, immunohistochemistry, and immunofluorescence. Particularly preferred methods for detecting proteins include any cell-based assay, including immunohistochemistry and immunofluorescence assays. Such methods are well known in the art.

The terms "subject", "patient" and "individual" are used interchangeably herein to refer to a warm-blooded animal, such as a mammal. The term includes, but is not limited to, livestock, rodents (eg, rats and mice), primates, and humans. Preferably the term refers to humans.

RBM10 (NG_012548.1) belongs to a family of RNA motif-containing binding proteins, located on chromosome Xp11.23, encoding a 930 amino acid nuclear protein, and contains two RNA recognition domains (RRM), two zinc finger domains and A G-patch domain. This gene is also known as DXS8237E, GPATC9, GPATCH9, S1-1, TARPS and ZRANB5.

The term "wild-type" is to be understood according to the general understanding of those skilled in the art and refers to a nucleic acid or amino acid sequence in its naturally occurring form without any artificial mutation, nucleotide change or amino acid modification.

The term "mutant" should be understood according to the general understanding of those skilled in the art. A nucleic acid sequence is referred to as "mutated" if it contains at least one nucleotide addition, deletion or substitution in its nucleic acid sequence compared to its native or native nucleic acid sequence, ie if it contains a nucleic acid mutation. An amino acid sequence is referred to as "mutated" if it contains at least one added, deleted or substituted amino acid in its amino acid sequence compared to its native or native amino acid sequence, ie if it contains an amino acid mutation.

In the present invention, the variation of the RBM10 gene or its mRNA refers to the mutation in the wild-type RBM10 gene or its mRNA, which includes addition, deletion and substitution of one or more nucleotides and changes in copy number (including amplification and missing). The nucleotides may be nucleotides of a coding region or a non-coding region. The coding region may be a nucleotide sequence encoding an RNA recognition domain (RRM), a zinc finger domain and/or a G-patch domain. The variation of the RBM10 gene or its mRNA can lead to the increase or decrease of the expression level of the RBM10 gene and/or the amplification or deletion of the copy number. For example, the level or copy number is at least about 1.1, 1.25, 1.5, 2, 3, 4, 5, 6, 7, 8, 9 or 10 times or more or at most about 1 times the control or standard, respectively /1.1, 1/1.25, 1/1.5, 1/2, 1/3, 1/4, 1/5, 1/6, 1/7, 1/8, 1/9 or 1/10 or less. Copy number amplifications or deletions can be detected by techniques well known in the art, such as whole gene sequencing.

In the present invention, the variation of the protein or protein fragment encoded by the RBM10 gene includes: 1) mutations in the protein or protein fragment encoded by the wild-type RBM10 gene, which include addition, deletion or substitution of one or more amino acids; and 2 ) Changes in the expression level of the protein encoded by the wild-type RBM10 gene. The amino acid may be an amino acid of an RNA recognition domain (RRM), a zinc finger domain and/or a G-patch domain. The protein fragments refer to polypeptides having amino-terminal deletions, carboxy-terminal deletions and/or intermediate deletions compared to the full-length native protein. The fragments may also contain modified amino acids compared to the native protein. In certain embodiments, fragments are about 5-500 amino acids in length. For example, fragments can be at least 5, 6, 8, 10, 14, 20, 50, 70, 100, 110, 150, 200, 250, 300, 350, 400, or 450 amino acids in length. In one embodiment, the fragment is an immunologically detectable fragment preferably comprising at least 6, 7, 8, 10, 12, 15 or 20 contiguous amino acids of the marker polypeptide. The change in the protein expression level refers to at least about 1.1, 1.25, 1.5, 2, 3, 4, 5, 6, 7, 8, 5, 5, 6, 7, 8, 9 or 10 times or more or at most about 1/1.1, 1/1.25, 1/1.5, 1/2, 1/3, 1/4, 1/5, 1/6 of the expression level of a control or standard , 1/7, 1/8, 1/9 or 1/10 or less.

The terms "polypeptide" and "protein" are used interchangeably herein to refer to at least one molecular chain of amino acids linked by covalent and/or non-covalent bonds. The term includes post-translational modifications of peptides, oligopeptides, and proteins and polypeptides, such as glycosylation, acetylation, phosphorylation, and the like. Protein fragments, analogs, muteins or variant proteins, fusion proteins, etc. are also included within the meaning of the term.

In certain embodiments, determining "protein expression level", "gene expression" or "gene expression level" as used herein includes, but is not limited to, determining corresponding RNA, protein or peptide levels (or combinations thereof). The present invention is not limited to specific methods and reagents for measuring protein, peptide or RNA levels, all of which are well known in the art.

Methods for determining the amount or concentration of protein in a sample are known to the skilled artisan. Such methods include radioimmunoassays, competitive binding assays, Western blot analysis and ELISA assays. For methods using antibodies, both monoclonal and polyclonal antibodies are suitable. The antibody may be immunologically specific for a protein, protein epitope or protein fragment.

The term "oligonucleotide" refers to a polymeric form of nucleotides of any length, either ribonucleotides or deoxyribonucleotides. The term includes double- and single-stranded DNA and RNA, modified and unmodified forms such as methylation or capping of polynucleotides. The terms "polynucleotide" and "oligonucleotide" are used interchangeably herein. The oligonucleotide may, but need not, include other coding or non-coding sequences, or it may, but need not, be linked to other molecules and/or vectors or support materials. The oligonucleotides used in the methods or kits of the invention can be of any length suitable for the particular method. In certain applications, the term refers to an antisense nucleic acid molecule (eg, an mRNA or DNA strand in the opposite orientation to a sense polynucleotide encoding a cancer marker of the invention (eg, RBM10)).

Oligonucleotides for use in the present invention include complementary nucleic acid sequences and nucleic acids substantially identical to those sequences, and also include sequences that differ from nucleic acid sequences due to the degeneracy of the genetic code. Oligonucleotides useful in the present invention also include nucleic acids that hybridize to oligonucleotide cancer marker nucleic acid sequences under stringent conditions, preferably high stringency conditions.

Nucleotide hybridization assays are well known in the art. Hybridization assay procedures and conditions will vary depending on the application and are chosen according to known general binding methods, see eg, J. Sambrook et al., Molecular Cloning: A Laboratory Guide (Third Edition. Science Press, 2002); and Young and Davis , PNAS, 80:1194 (1983). Methods and apparatus for performing repetitive and controlled hybridization reactions have been described in US Pat. Nos. 5,871,928, 5,874,219, 6,045,996, 6,386,749, and 6,391,623, each of which is incorporated herein by reference.

In some cases, it may be necessary to amplify the sample. Genomic samples can be amplified by various mechanisms, some of which employ PCR. Samples can be amplified on the array. See, eg, US Patent No. 6,300,070 and US Patent Application Serial No. 09/513,300.

Other suitable amplification methods include ligase chain reaction (LCR) (eg Wu and Wallace, Genomics 4, 560 (1989), Landegren et al. Science 241, 1077 (1988) and Barringer et al. Gene 89:117 (1990)), transcription Amplification (Kwoh et al., Proc. Natl. Acad. Sci. USA 86, 1173 (1989) and WO 88/10315), self-sustaining sequence replication (Guatelli et al., Proc. Nat. Acad. Sci. USA, 87, 1874 (1990) ) and WO90/06995), selective amplification of target polynucleotide sequences (US Pat. No. 6,410,276), consensus-primed polymerase chain reaction (CP-PCR) (US Pat. No. 4,437,975), arbitrarily primed polymerase chain reaction reaction (AP-PCR) (US Pat. Nos. 5,413,909, 5,861,245) and Nucleic Acid-Based Sequence Amplification (NABSA) (see US Pat. Nos. 5,409,818, 5,554,517 and 6,063,603, each of which is incorporated herein by reference).

Reagents that can be used to detect RBM10 expression levels and/or copy number are well known in the art. Such reagents suitable for use in the present invention are commercially available or routinely prepared by methods well known to those skilled in the art.

The term "binding agent" refers to substances such as polypeptides, antibodies, ribosomes or aptamers that specifically bind to the biomarkers of the invention (RBM10). A substance "specifically binds" to a biomarker of the invention if it reacts at a detectable level with the biomarker of the invention but not detectably reacts with a peptide containing an unrelated sequence or a sequence of a different polypeptide. Binding properties can be assessed using an ELISA that can be readily performed by those skilled in the art.

The binding agent may be a ribosome, RNA or DNA molecule or polypeptide with or without a peptide component. The binding agent may be a polypeptide comprising a polypeptide biomarker sequence, a peptide variant thereof, or a non-peptide mimetic of such a sequence.

Aptamers include DNA or RNA molecules that bind nucleic acids and proteins. Aptamers that bind the markers of the present invention can be generated using conventional techniques without undue experimentation. [See, for example, the following publications describing in vitro selection of aptamers: Klug et al., Mol. Biol. Reports 20:97-107 (1994); Wallis et al., Chem. Biol. 2:543-552 (1995); Ellington, Curr. Biol. 4:427-429 (1994); Lato et al., Chem. Biol. 2:291-303 (1995); Conrad et al., Mol. Div. 1:69-78 (1995); and Uphoff et al., Curr. Opin . Struct. Biol. 6:281-287 (1996)].

Antibodies useful in the present invention include, but are not limited to, synthetic antibodies, monoclonal antibodies, polyclonal antibodies, recombinant antibodies, antibody fragments (eg, Fab, Fab', F(ab')2), dAbs (domain antibodies; see Ward et al. , 1989, Nature, 341:544-546), antibody heavy chain, intrabody, humanized antibody, human antibody, antibody light chain, single-chain Fv (scFv) (eg, including monospecific, bispecific, etc.) , anti-idiotypic (ant-Id) antibodies, proteins comprising antibody moieties, chimeric antibodies (eg, antibodies containing the binding specificity of a murine antibody but where the remainder is of human origin), derivatives such as enzyme conjugates or labeled Derivatives, diabodies, linear antibodies, disulfide-linked Fvs (sdFv), multispecific antibodies (eg, bispecific antibodies), epitope-binding fragments of any of the foregoing, and antigen recognition comprising the desired specificity Any other modified configuration of the immunoglobulin molecule at the site. Antibodies include antibodies of any class (e.g., IgA, IgD, IgE, IgG, IgM, and IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or any subclass (e.g., IgG2a and IgG2b), and antibodies do not have to be is any particular type, class or subclass. In certain embodiments of the invention, the antibody is an IgG antibody or a class or subclass thereof. Antibodies can be from any animal source, including birds and mammals (eg, human, murine, donkey, sheep, rabbit, goat, guinea pig, camel, horse, or chicken).

For example, antibodies for use in the present invention are commercially available from, for example, Invitrogen (Cat. Nos. PA5-83253, PA5-40331, PA5-40330, etc.), Abcam (eg, antibody under Cat. No. ab224149), and the like. Alternatively, the antibodies can be prepared by recombinant methods well known in the art. In some embodiments, the antibody is a monoclonal antibody. For the preparation of monoclonal antibodies see, eg, Kohler et al. (1975) Nature 256:495-497; Kozbor et al. (1985) J. Immunol Methods 81:31-42; Cote et al. (1983) Proc Natl Acad Sci 80:2026-2030 and Cole et al. (1984) Mol Cell Biol 62:109-120.

The kit of the present invention can be prepared by conventional methods in the art. The kits may contain materials or reagents (including reagents for detecting the RBM10 gene or its mRNA or its encoded protein or protein fragment) for use in carrying out the methods of the present invention. Kits may include storage reaction reagents (eg, primers, dNTPs, enzymes, etc. in suitable containers) and/or support materials (eg, buffers, instructions for performing assays, etc.). For example, a kit may include one or more containers (eg, cassettes) containing the respective reaction reagents and/or support materials. Such contents may be delivered to the intended recipient together or separately. As an example, the kit may contain reagents, buffers, and instructions for use in detecting the RBM10 gene or its mRNA or its encoded protein or protein fragment. The kit may also contain polymerase and dTNP, among others. The kit may also contain internal standards, positive and negative controls, etc. for quality control. The kit may also contain reagents for preparing nucleic acid, eg, DNA, from the sample. The above examples should not be construed as limiting the kits and their contents suitable for use in the present invention.

A microarray refers to a solid support with a flat surface that has an array of nucleic acids, each member of the array comprising identical copies of oligonucleotides or polynucleotides immobilized on spatially defined regions or sites, so The regions or sites do not overlap with regions or sites of other members of the array; that is, the regions or sites are spatially discrete. Furthermore, a spatially defined hybridization site may be "addressable" in that its location and the identity of its immobilized oligonucleotide are known or predetermined (eg, known or predetermined prior to its use). definite). Typically the oligonucleotide or polynucleotide is single stranded and is usually covalently attached to the solid support from the 5'- or 3'-end. The density of nucleic acids containing non-overlapping regions in the microarray is typically greater than 100/cm ² , more preferably greater than 1000/cm ² . Microarray technology is disclosed, for example, in the following references: Microarrays: A Practical Approach, edited by Schena (IRL Press, Oxford, 2000); Southern, Current Opin. Chem. Biol., 2:404-410, 1998, the entire contents of which are via Reference is incorporated herein.

The invention discloses the use of the RBM10 gene, and those skilled in the art can learn from the content of this article and appropriately improve the process parameters to achieve. It should be particularly pointed out that all similar substitutions and modifications are obvious to those skilled in the art, and they are deemed to be included in the present invention. The uses described in the present invention have been described through the preferred embodiments, and it is obvious that relevant persons can make changes or appropriate changes and combinations of the uses described herein without departing from the content, spirit and scope of the present invention, so as to realize and apply the technology of the present invention .

The use of the RBM10 gene provided by the present invention will be further described below.

Example 1: Phase Ib clinical trial of cioronib monotherapy in the treatment of relapsed and refractory ovarian cancer

Test drug: Cioroni capsules, specifications: 5mg, 25mg. Produced by Shenzhen Microchip Biotechnology Co., Ltd.

Dosing regimen: Cioroni capsules were administered at 50 mg/day, QD (no adjustment for body weight or body surface area). Take each morning on an empty stomach, with water, and swallow the capsule whole. Continuous administration for 28 days is one treatment cycle, and there is no interval between each treatment cycle.

Number of cases: 25 patients were enrolled, of which 23 patients were evaluated for efficacy.

standard constrain:

1. Age ≥18 years old, ≤70 years old, female;

2. Histologically diagnosed epithelial ovarian cancer, fallopian tube cancer or primary peritoneal cancer;

3. Subjects need to have received platinum-based chemotherapy regimens, i.e.

a) Platinum-resistant patients (disease progression or recurrence within 6 months after the completion of platinum-containing chemotherapy regimens), should have received at least 2 different chemotherapy regimens for disease progression or recurrence;

b) If platinum-sensitive patients (disease progression or recurrence >6 months after the completion of platinum-containing chemotherapy regimens), should have received at least 2 chemotherapy regimens for disease progression or recurrence, or the subject refuses to receive further chemotherapy;

4. According to RECIST1.1 criteria, there is at least one measurable target lesion;

5. ECOG physical fitness score 0-1 points;

6. The interval between the previous chemotherapy, radiotherapy, targeted therapy, immunotherapy or study drug treatment should be more than 4 weeks. If the chemotherapy regimen includes mitomycin, the interval should be more than 6 weeks;

7. Major organ functions meet the following criteria:

Blood routine: absolute value of neutrophils ≥ 1.5×10 ⁹ /L, platelets ≥ 90×10 ⁹ /L, hemoglobin ≥ 90g/L;

Blood biochemistry: total bilirubin ≤ 1.5 times the upper limit of the reference range (ULN), AST, ALT ≤ 2 times ULN (liver metastases: ≤ 5 times ULN); serum creatinine ≤ 1.5 times ULN;

Coagulation function: prothrombin time-international normalized ratio (PT-INR) ≤ 1.5 times ULN.

8. Expected survival time ≥ 3 months;

9. Voluntarily signed a written informed consent.

Treatment plan:

The test subjects took cioroni capsules 50 mg orally once a day, every 28 days as a treatment cycle, and there was no discontinuation interval during the treatment cycle. Throughout the trial, all subjects continued treatment until any of the following occurred (whichever occurred first): disease progression, intolerable toxicity, death, withdrawal of informed consent, or loss to follow-up.

Efficacy evaluation: According to RECIST1.1 criteria, the evaluation was performed at the baseline and the 4th week after treatment, and repeated every 8 weeks until disease progression. Tumor imaging examinations include CT or MRI of the neck, chest, whole abdomen, and pelvis. Examinations of other parts are performed according to clinical indications and when necessary. The same techniques and methods should be used for baseline and follow-up assessment of lesions.

Safety assessment: including physical examination, vital signs, ECOG performance score, blood routine, urine routine, 12-lead ECG, blood biochemistry, electrolytes, coagulation function, cardiac enzymes, troponin, TSH, FT3, FT4, amylase, Echocardiography, 24-hour urine protein quantification (if necessary), adverse events.

Biomarker Research:

10 ml of peripheral blood was taken before the subjects took cioroni for the first time and when the disease progressed, and the gene sequences of plasma cell-free tumor DNA (ctDNA) and leukocyte extracted DNA (control) were detected, including a total of 548 tumor-related genes, Test results include genetic mutations and copy number abnormalities.

Clinical trial results:

Of the 25 subjects, 2 had no post-baseline efficacy assessment, resulting in 23 evaluable cases. Among the 23 evaluable subjects, the best response during the trial was: 2 (8.7%) PR (of which 1 was confirmed at follow-up assessment), 14 (60.9%) SD, 7 (30.4%) PD , the confirmed ORR was 4.3% and the unconfirmed ORR was 8.7%. Of the 23 evaluable subjects, 3 were platinum-sensitive and 20 were platinum-resistant. Both PR subjects were platinum-resistant subjects, and the confirmed response was 1 (5.0%). The results show that cioronib monotherapy is effective in ovarian cancer.

A concomitant study of efficacy-related biomarkers was performed on 548 tumor-related genes for the detection and analysis of cell-free tumor DNA (ctDNA) in the plasma of the evaluated patients. The detection results selected all genes with a mutation rate of more than 0.4%, and used the patient's progression-free survival (PFS) as the efficacy index to analyze the correlation between tumor-related gene abnormalities and the efficacy of cioronib. The results showed that among 548 tumor-related genes, a gene mutation that was significantly correlated with the efficacy of Cioroni was found, which was the RBM10 gene. The median PFS was 232 days in 7 of the 23 evaluable subjects with RBM10 mutations (30.4%, these mutations included nucleotide additions, deletions and substitutions, and copy number changes) and 16 with RBM10 wild-type (69.6%). The median PFS of ) was 106 days, and there was a significant difference between the two groups, suggesting that the RBM10 gene is associated with the efficacy of cioronib in the treatment of ovarian cancer and can be a potential biomarker for evaluating the efficacy. See Table 1 and Figure 1 for the results.

Table 1: RBM10 gene mutation and median PFS correlation analysis

Note: P value = 0.0242, HR = 0.133

The above are only the preferred embodiments of the present invention. It should be pointed out that for those skilled in the art, without departing from the principles of the present invention, several improvements and modifications can be made. It should be regarded as the protection scope of the present invention.

Claims (10)

1. Use of a reagent for detecting RBM10 gene or its mRNA or its encoded protein or protein fragment in the preparation of a kit or microarray for evaluating the efficacy of cioronib or guiding the administration of cioronib or predicting the therapeutic effect of ovarian cancer .
2. The use according to claim 1, wherein if compared with the wild type, there is variation in the RBM10 gene or its mRNA or its encoded protein in the subject, indicating that for the subject, cioroni The curative effect is better, can use cioroni therapy or ovarian cancer treatment effect is better.
3. The use according to claim 1, wherein the reagent for detecting the RBM10 gene or its mRNA or its encoded protein or protein fragment is a binding agent that binds to the protein or protein fragment encoded by the RBM10 gene, or binds to the RBM10 gene. A substance that hybridizes or amplifies the RBM10 gene or its mRNA by a gene or its mRNA.
4. The use according to claim 3, wherein the binding agent that binds to the protein or protein fragment encoded by the RBM10 gene is an anti-RBM10 antibody.
5. The use according to claim 3, wherein the substance that hybridizes with the RBM10 gene or its mRNA or amplifies the RBM10 gene or its mRNA is an oligonucleotide primer or probe.
6. A kit or microarray for evaluating the curative effect of cioronib or guiding the administration of cioronib or predicting the therapeutic effect of ovarian cancer, characterized in that it comprises a kit or microarray for detecting the RBM10 gene or its mRNA or its encoded protein or protein Fragment reagents.
7. The kit or microarray according to claim 6, wherein if compared with the wild type, there is variation in the RBM10 gene or its mRNA or the protein encoded by the subject, it indicates that for the subject, Cioroni has better efficacy, can be treated with cioroni, or has better results in ovarian cancer treatment.
8. The kit or microarray according to claim 6, wherein the reagent for detecting the RBM10 gene or its mRNA or the protein or protein fragment encoded by it is a binding agent that binds to the protein or protein fragment encoded by the RBM10 gene , or a substance that hybridizes with the RBM10 gene or its mRNA or amplifies the RBM10 gene or its mRNA.
9. The kit or microarray according to claim 8, wherein the binding agent that binds to the protein or protein fragment encoded by the RBM10 gene is an anti-RBM10 antibody, and the binding agent to the RBM10 gene or its mRNA is hybridized or amplified The substance of the RBM10 gene or its mRNA is an oligonucleotide primer or probe.
10. Use of cioronib in the preparation of a medicament for treating ovarian cancer, especially ovarian cancer with RBM10 gene mutation.

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