[go: up one dir, main page]

WO2020064006A2 - Groupe de gènes utilisé pour le typage moléculaire du cancer du sein et le risque de métastase à distance, produit de diagnostic et application - Google Patents

Groupe de gènes utilisé pour le typage moléculaire du cancer du sein et le risque de métastase à distance, produit de diagnostic et application Download PDF

Info

Publication number
WO2020064006A2
WO2020064006A2 PCT/CN2019/109459 CN2019109459W WO2020064006A2 WO 2020064006 A2 WO2020064006 A2 WO 2020064006A2 CN 2019109459 W CN2019109459 W CN 2019109459W WO 2020064006 A2 WO2020064006 A2 WO 2020064006A2
Authority
WO
WIPO (PCT)
Prior art keywords
risk
breast cancer
gene
reagent
distant metastasis
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/CN2019/109459
Other languages
English (en)
Chinese (zh)
Other versions
WO2020064006A3 (fr
Inventor
周彤
胡志元
周伟庆
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Shanghai Precision Diagnostics Co Ltd
Original Assignee
Shanghai Precision Diagnostics Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Shanghai Precision Diagnostics Co Ltd filed Critical Shanghai Precision Diagnostics Co Ltd
Priority to CN201980006647.6A priority Critical patent/CN111566229B/zh
Publication of WO2020064006A2 publication Critical patent/WO2020064006A2/fr
Publication of WO2020064006A3 publication Critical patent/WO2020064006A3/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Images

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6844—Nucleic acid amplification reactions
    • C12Q1/6851—Quantitative amplification
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/112—Disease subtyping, staging or classification
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/158—Expression markers

Definitions

  • the invention belongs to the field of biotechnology, and particularly relates to breast cancer subtypes and distant metastasis risk gene groups, and in vitro diagnostic products and applications thereof.
  • Breast cancer accounts for the first incidence of malignant tumors in women in China, and it is increasing year by year at a rate of about 4%.
  • Breast cancer is a highly heterogeneous tumor, and its pathological molecular mechanism is very complicated, often involving mutations of multiple genes in many types of somatic cells.
  • Breast cancer classification according to the pathological characteristics of the tumor, improving the accuracy of clinical diagnosis and prognosis prediction of breast cancer, is the basis of accurate treatment after breast cancer surgery, and also the key to improving the survival rate of breast cancer patients.
  • the accurate treatment of breast cancer after surgery at home and abroad still faces great challenges.
  • the internationally recognized breast cancer molecular typing product based on tumor gene expression is the PAM50 (Prosigna) jointly developed by Professor Perou of the University of North Carolina and NanoString Corporation, which has been approved by the US FDA.
  • PAM50 divides breast cancer into four subtypes, lumenal A (Luminal A), lumenal B (Luminal B), and HER2 enriched type ( HER2-enriched (HER2 for short) and Basal-like (Basal), and the risk of distant metastasis in breast cancer within 10 years was evaluated based on the subtype and tumor proliferation index.
  • the present invention relates to a group of gene groups, including 66 molecular typing and genes related to distant metastasis risk assessment, and the use of the gene group in molecular diagnosis of breast cancer.
  • the gene population can be used for molecular typing of breast cancer and / or assessing its risk of distant metastasis.
  • the 66 molecular typing and distant metastasis risk assessment related genes include:
  • the gene population further includes a housekeeping gene.
  • the housekeeping genes include at least one of the following (eg, 1, 2, 3, 4, 5, or 6), preferably at least 3, and most preferably all 6: GAPDH, GUSB, MRPL19, PSMC4, SF3A1, and TFRC.
  • the present invention relates to a gene group (also referred to herein as Precitype) for molecular typing of breast cancer and / or assessing its risk of distant metastasis, which includes 72 Genes, that is, 66 molecular typing and distant metastasis risk assessment related genes and 6 housekeeping genes, of which
  • the 66 molecular typing and distant metastasis risk assessment related genes include:
  • the six housekeeping genes include: GAPDH, GUSB, MRPL19, PSMC4, SF3A1, and TFRC.
  • the present invention relates to the use of the genetic population in performing breast cancer molecular typing and / or assessing the risk of distant metastases.
  • the invention also relates to the use of the gene population in the preparation of a diagnostic product for molecularly typing breast cancer and / or assessing the risk of distant metastases.
  • the invention also relates to a method of molecularly typing breast cancer and / or assessing the risk of distant metastases, said method being performed by using a diagnostic product of the invention.
  • the present invention also relates to a diagnostic product for molecularly typing breast cancer and / or assessing the risk of distant metastases, which comprises a reagent for detecting the expression level of a gene in the gene group of the present invention.
  • the diagnostic product is in the form of an in vitro diagnostic product, preferably in the form of a diagnostic kit.
  • the diagnostic product further comprises a total RNA extraction reagent, a reverse transcription reagent, and / or a second-generation sequencing reagent.
  • the reagent is a probe or a primer.
  • the breast cancer includes luminal type A, luminal type B, HER2-enriched type, basal cell type, and immune-enhanced type (Immune-enhanced, which may also be referred to herein as Immuno ).
  • Figure 1 shows that according to the expression level of 66 genes in breast cancer tissue, breast cancer is divided into luminal A (Lum A), luminal B (Lum B), basal cell type (Basal), and HER2 enriched type (HER2 ) And immune-enhanced (Imm).
  • Lum A luminal A
  • Lum B luminal B
  • Basal basal cell type
  • HER2 HER2
  • Imm immune-enhanced
  • A 1951 European and American cases
  • B 824 Chinese cases.
  • Invasion-related genes ie, CTSL2 and MMP11
  • CTSL2 and MMP11 have fewer genes and have fewer gene expression regions in the heat map, so they are not shown in Figure 1.
  • Figure 2 shows that the risk of distant metastasis is different for each subtype of breast cancer.
  • the risk of distant metastasis of luminal type A was significantly lower than the other four subtypes, and the risk of distant metastasis of immune-enhanced type was significantly lower than that of luminal type B, basal cell type and HER2 enriched type.
  • Lumen B is less than basal cell type (Basal), while basal cell type (Basal) is less than HER2 enriched type (HER2); 10 years after surgery There is no significant difference between the three.
  • the lumen B type was smaller than the basal cell type and the HER2 enriched type within 3 years of surgery. After 10 years of surgery, the HER2 enriched type had the worst prognosis.
  • A 1951 European and American cases
  • B 824 Chinese cases.
  • Figure 3 shows that the immune index has an important effect on the prognosis of breast cancer.
  • strong immune function can significantly reduce the risk of distant metastasis of breast cancer in patients ;
  • low-risk breast cancer subtype lumen A the strength of immune function has no significant effect on the prognosis.
  • A 1951 European and American cases; B, 824 Chinese cases.
  • Figure 4 shows that according to the calculated recurrence risk index, the risk of distant tumor metastasis is divided into three groups, low risk (0-32), medium risk (33-49), and high risk (50-100).
  • A 1951 European and American cases; B, 824 Chinese cases.
  • FIG. 5 shows the second-generation sequencing (NGS) -based gene expression detection of the present invention, and its sensitivity and detection throughput are better than the existing gene chip, quantitative PCR, and Nanostring technologies.
  • NGS second-generation sequencing
  • Figure 6 shows the results of reproducible experiments, which show that the RNA (left) extracted from paraffin sections and the RNA (right) extracted from fresh tissue have excellent correlation.
  • the expression “at least one (species)” or similar expression “one (species) or multiple (species)” means 1 (species), 2 (species), 3 (species), 4 ( Species), 5 (species), 6 (species), 7 (species), 8 (species), 9 (species) or more (species).
  • breast cancer originates from ducts and acinar epithelium at all levels of the breast, and gradually develops from glandular epithelium to atypical hyperplasia. It is divided into carcinoma in situ (non-invasive cancer), early invasive cancer to invasive cancer. In a preferred embodiment, the breast cancer is invasive breast cancer.
  • risk of recurrence refers to the likelihood of a breast cancer patient's tumor recurrence within a specified number of years, including, but not limited to, localized breast cancer recurrence, regional breast cancer recurrence, and distant metastasis. As used herein, "risk of relapse” preferably indicates the possibility of distant metastasis. Therefore, in this article, the assessment of "Risk of Recurrence Score” (RRS) can be expressed as “distant recurrence risk score” or "distant metastasis risk score”.
  • breast cancer recurrence risk assessment can be used to assess the possibility of local recurrence or distant metastases in breast cancer patients.
  • breast cancer can be classified into low risk, medium risk, and high risk according to the recurrence risk score.
  • Chemotherapy is generally considered to be low risk, while chemotherapy at high risk reduces the risk of recurrence or distant metastases.
  • the medium risk requires the clinician to make judgments based on the actual clinical situation. Such a "medium risk" has less guiding significance in clinical treatment. Therefore, the fewer cases of medium risk, the more efficient the assessment will be.
  • the risk of breast cancer recurrence is preferably assessed for the possibility of distant metastases.
  • RNA transcript refers to total RNA, that is, coding or non-coding RNA, including RNA directly from tissue or peripheral blood samples, and also indirectly from tissue or blood samples after cell lysis.
  • Total RNA includes tRNA, mRNA, and rRNA, where mRNA includes mRNA transcribed from the target gene, and mRNA from other non-target genes.
  • polypeptide refers to a compound consisting of amino acids connected by peptide bonds, including the full length or amino acid fragments of a polypeptide.
  • the amount of the polypeptide encoded by the gene can be standardized with respect to the amount of total protein in the sample or the amount of the polypeptide encoded by the housekeeping gene.
  • RNA transcripts can be detected and quantified, for example, by methods of hybridization, amplification or sequencing. For example, hybridize RNA transcripts with probes or primers.
  • hybridization refers to the process under which the two nucleic acid fragments are combined by stable and specific hydrogen bonds to form a double helix complex under appropriate conditions.
  • amplification primer or “primer” refers to a nucleic acid fragment containing 5 to 100 nucleotides, preferably, 15 to 30 cores capable of initiating an enzymatic reaction (eg, an enzymatic amplification reaction). Glycylic acid.
  • hybridization probe refers to a nucleic acid sequence including at least 5 nucleotides, for example, containing 5 to 100 nucleotides, which can interact with the expression product of the target gene or the expansion of the expression product under specified conditions. The gain products hybridize to form a complex.
  • the hybridization probe may further include a label for detection.
  • the label includes, but is not limited to, a label for fluorescent quantitative PCR or fluorescent in situ hybridization.
  • the label may be FAM, HEX, VIC, Cy5, or the like.
  • the label may be biotin, digoxin and the like.
  • the detection of the expression level of a gene described herein may employ assay methods known in the art, including, but not limited to, a method of detecting the amount of an RNA transcript of the gene or the amount of a polypeptide encoded by the gene.
  • RNA transcript of a gene can be converted into cDNA complementary thereto by methods known in the art, and the amount of RNA transcript can be obtained by measuring the amount of complementary cDNA.
  • the amount of a gene's RNA transcript or its complementary cDNA can be normalized to the amount of total RNA or total cDNA in a sample or to a set of housekeeping genes' RNA transcripts or its complementary cDNA.
  • RNA transcripts can be detected and quantified by methods such as hybridization, amplification or sequencing, including but not limited to methods for hybridizing RNA transcripts with probes or primers, by polymerase chain reaction (PCR) -based A method of detecting the amount of RNA transcript or its corresponding cDNA product by various quantitative PCR techniques or sequencing techniques.
  • the quantitative PCR technology includes, but is not limited to, fluorescent quantitative PCR, real-time PCR, or semi-quantitative PCR technology.
  • the sequencing technology includes, but is not limited to, Sanger sequencing, second-generation sequencing, third-generation sequencing, and single-cell sequencing.
  • the sequencing technology is second-generation sequencing, more preferably targeted RNA-seq technology.
  • the amount of polypeptide can be detected by, for example, proteomics or reagents.
  • the reagent is an antibody, an antibody fragment or an affinity protein.
  • the invention relates to a group of gene groups, which includes 66 genes for molecular typing and distant metastasis risk assessment.
  • the 66 molecular typing and distant metastasis risk assessment related genes include:
  • the gene population of the present invention further includes a housekeeping gene.
  • the housekeeping gene includes at least one of the following (eg, 1, 2, 3, 4, 5, or 6), preferably at least 3, and most preferably 6: GAPDH, GUSB, MRPL19, PSMC4, SF3A1, and TFRC.
  • a set of gene groups is provided, which includes 72 genes, that is, 66 molecular typing and distant metastasis risk assessment related genes, and 6 housekeeping genes.
  • This gene group is also referred to herein as Precitype.
  • the 66 molecular typing and distant metastasis risk assessment related genes and 6 housekeeping genes are as described above.
  • the gene group of the present invention is shown in Table 1.
  • Housekeeping genes are used to normalize and correct target gene expression levels. Inclusion criteria for housekeeping genes that can be considered are: 1. Stable expression in tissues, whose expression level is not affected or less affected by pathological conditions or drug treatment; 2. Expression levels should not be too high to avoid expression data ( (For example, obtained through second-generation sequencing), the proportion of data obtained is too high, which affects the accuracy of data detection and interpretation of other genes. Therefore, housekeeping genes may be included in the gene group of the present invention.
  • the housekeeping genes considered include at least one of GAPDH, GUSB, MRPL19, PSMC4, SF3A1, and TFRC, such as 1, 2, 3, 4, 5, or 6, preferably at least 3, and most preferably all 6 .
  • the housekeeping genes considered include GAPDH, GUSB, MRPL19, PSMC4, SF3A1, TFRC, or any combination thereof.
  • the present invention also relates to a group of immune-related genes comprising 17 genes: APOBEC3G, CCL5, CCR2, CD2, CD3D, CD52, CD53, CORO1A, CXCL9, GZMA, GZMK, HLA-DMA, HLA-DQA1, IL2RG, LCK, LYZ, and PTPRC.
  • the genes in the immune-related gene group are also shown in the related information in Table 1.
  • the gene population of the present invention can be used for molecular typing (subtyping) of breast cancer and / or assessing its risk of distant metastasis.
  • the subtypes of breast cancer include luminal A, luminal B, HER2-enriched, basal cell-like, and immune-enhanced.
  • the invention relates to a diagnostic product for molecularly typing breast cancer and / or assessing its risk of distant metastasis, which comprises an agent for detecting the expression level of genes in the gene group of the invention.
  • the gene population is as described above.
  • the gene group of the present invention may include 66 genes related to molecular typing and distant metastasis risk assessment.
  • the 66 molecular typing and distant metastasis risk assessment related genes include:
  • the gene population of the present invention further includes a housekeeping gene.
  • the housekeeping gene includes at least one of the following (eg, 1, 2, 3, 4, 5, or 6), preferably at least 3, and most preferably 6: GAPDH, GUSB, MRPL19, PSMC4, SF3A1 and TFRC.
  • the gene group is shown in Table 1.
  • the diagnostic product is an in vitro diagnostic product.
  • the diagnostic product is a diagnostic kit.
  • the diagnostic product is used for breast cancer subtype and / or distant metastasis risk assessment.
  • the reagent is a reagent that detects the amount of RNA, particularly mRNA, transcribed by the gene. In yet another embodiment, the reagent is a reagent that detects the amount of cDNA complementary to the mRNA.
  • the diagnostic product further includes a total RNA extraction reagent, a reverse transcription reagent, and / or a second-generation sequencing reagent.
  • the total RNA extraction reagent may be a conventional total RNA extraction reagent in the art. Examples include, but are not limited to, Qiagen 73504, RNA Storm CD201, Invitrogen and ABI AM1975.
  • the reverse transcription reagent may be a conventional reverse transcription reagent in the art, and preferably includes a dNTP solution and / or an RNA reverse transcriptase.
  • Examples of reverse transcription reagents include, but are not limited to, NEB M0368L, Thermo K1622, ABI 4366596.
  • the second-generation sequencing reagent may be a reagent commonly used in the art, as long as it can meet the requirements for performing second-generation sequencing on the obtained sequence.
  • Second-generation sequencing reagents can be commercially available products, examples of which include, but are not limited to, Illumina Reagent Kit v3 (150cycle) (MS-102-3001), Targeted RNA Index Kit A-96 Indicators (384 Samples) (RT-402-1001).
  • Next-generation sequencing is conventional second-generation sequencing in the art, such as targeted RNA-seq technology. Therefore, next-generation sequencing reagents can also include Illumina custom-made reagents that can be used to build libraries that target RNA-seq, such as Targeted RNA Custom Panel Kit (96 Samples) (RT-102-1001).
  • the reagent is a probe or a primer.
  • the reagent is a reagent that detects the amount of the polypeptide encoded by the gene, and preferably, the reagent is an antibody, an antibody fragment, or an affinity protein.
  • the reagent is a primer.
  • the primer is a synthetic oligonucleotide fragment (preferably having high specificity) as long as it is complementary to a partial sequence of a gene in the gene group of the present invention, and the gene therein is amplified.
  • Primers can be synthetic. More preferably, the sequences of the primers are as shown in SEQ ID No. 1-SEQ ID No. 132 and SEQ ID NO. 133-SEQ ID NO. 144. The specific protocol of the primers can be seen in Table 2.
  • an embodiment of the present invention also relates to a diagnostic product (preferably an in vitro diagnostic product, especially a kit), which comprises a primer whose sequence is as described in SEQ ID NO.1-SEQ ID NO.132.
  • the diagnostic product further comprises a primer, and the sequence of the primer is as described in SEQ ID NO. 133-SEQ ID NO. 144.
  • the diagnostic product of the present invention (preferably an in vitro diagnostic product, especially a kit) comprises a primer whose sequence is shown in SEQ ID NO.1-SEQ ID NO.144 (see also Table 2 ).
  • the diagnostic product (preferably in the form of a kit) of the present invention also preferably includes a device for extracting a test sample from a subject; for example, a device for extracting tissue or blood from a subject, preferably any blood that can be used for blood collection Needles, syringes, etc.
  • the subject is a mammal, preferably a human, especially a female with breast cancer.
  • the primers can be used to detect the expression level of the gene population.
  • the present invention also relates to a method for determining a breast cancer molecular typing and / or distant metastatic risk in a subject, the method comprising
  • the method of the invention can be used for diagnostic or non-diagnostic purposes.
  • the subject for use in the method of the invention is a mammal, preferably a human, especially a female with breast cancer.
  • the method of providing a sample in step (1) and the sample used are not particularly limited, as long as the expression level of genes in a gene group can be obtained therefrom, for example, total RNA of a subject can be extracted from the sample.
  • the sample is preferably a sample of tissue, blood, plasma, body fluid or a combination thereof, preferably a tissue sample, especially a paraffin tissue sample.
  • the sample is a tumor tissue sample or a tissue sample containing tumor cells, particularly a breast tumor tissue sample or a tissue sample containing breast tumor cells.
  • step (2) the expression level of a gene in the gene group of the present invention is determined.
  • This gene group is as described above.
  • the gene group can also be described in Table 1.
  • the method of the present invention can be performed by a reagent for detecting a gene expression level in the gene group.
  • the reagent is a reagent that detects the amount of RNA, particularly mRNA, transcribed by the gene.
  • the reagent is a reagent that detects the amount of cDNA complementary to the mRNA.
  • the reagent is a reagent that detects the amount of the polypeptide encoded by the gene, and preferably, the reagent is an antibody, an antibody fragment, or an affinity protein.
  • the reagent is a probe or a primer, especially a primer. More preferably, the sequence of the primer is as described in SEQ ID NO.1-SEQ ID NO.132. In one embodiment, a primer is also used, and the sequence of the primer is as described in SEQ ID NO. 133-SEQ ID NO. 144. The specific protocol of the primers can also be shown in Table 2.
  • the method of the present invention can be performed by a diagnostic product (especially a diagnostic kit) of the present invention.
  • step (2) may include
  • the extraction in step (2-1) can be performed by a conventional method in the art, and preferably, a total RNA of fresh frozen tissue or paraffin-embedded tissue of a subject is detected by using an RNA extraction kit.
  • extraction can be performed using Roche's RNA extraction kit (product number: RocheCatalogNumber # 3270289001) or Qiagen's RNA extraction kit (Qiagen, Rnease, FFPE, Kitalog ##, 504). .
  • a method for constructing a library may include the following steps:
  • the extracted total RNA was reverse transcribed to generate cDNAs of the genes described in Table 1.
  • the ends were filled and phosphorylated at the 5 ′ end.
  • 30 ⁇ l of DNA, 45 ⁇ l of purified water, 10 ⁇ l of T4 DNA ligase buffer with 10 mM ATP, 4 ⁇ l of 10 mM dNTP Mix, 5 ⁇ l of T4 DNA polymerase, 1 ⁇ l of Klenow enzyme, and 5 ⁇ l of T4 ligase were mixed. Incubate at 20 ° C for 30 minutes (reagent is Illumina sample preparation kit PE-102-1001). After incubation, use QIAGEN QIAquick PCR purification kit (part # 28104) to purify DNA.
  • End suspension A Dissolve the product from the previous step in 32 ⁇ l of buffer, add 5 ⁇ l of Klenow buffer, 10 ⁇ l of 1 mM dATP, and 3 ⁇ l of Klenow ⁇ , and keep at 37 ° C for 30 minutes (reagent is Illumina sample preparation kit).
  • the products were ligated by QIAGEN MinElute PCR purification kit (part # 28004): DNA was dissolved in 10 ⁇ l buffer, DNA ligase buffer 2 ⁇ 25 ⁇ l, PE adapter Oligo Mix 10 ⁇ l, DNA ligase 5 ⁇ l, and kept at 20 ° C for 15 minutes ( The reagent was Illumina sample preparation kit PE-102-1001). After warming, the DNA was purified using QIAGEN QIAquick PCR purification kit (part # 28104) to obtain a library.
  • Step (2-3) can be completed by RNA sequencing.
  • the sequencing method may be a conventional RNA-seq sequencing method for determining a gene expression level in the art.
  • Illumina NextSeq / MiSeq / MiniSeq / iSeq series sequencers are preferably used for next-generation sequencing.
  • the primers in the kit were used to amplify the genes shown in Table 1.
  • the obtained gene sequence could be sequenced twice.
  • the secondary sequencing is a targeted RNA-seq technology, and double-ended sequencing is performed with an Illumina NextSeq / MiSeq / MiniSeq / iSeq sequencer. Such a process can be done automatically by the instrument itself.
  • step (3) may be completed by performing statistical analysis on the obtained sequencing results.
  • Breast cancer typing and risk prediction can optionally be performed according to the single sample prediction method (SSP (Single Sample Predictor) and Parker) pioneered by Hu et al.
  • SSP Single Sample Predictor
  • the gene expression data of the obtained sequencing results are analyzed to obtain a subtype of a single sample, and the risk of distant metastasis can be calculated.
  • the self-developed analysis software EPIG was used to calculate the close relationship with breast cancer metastasis through unsupervised clustering and paired correlation analysis. Relevant and statistically significant gene expression profiles, and based on this to obtain meaningful genes in the expression profile. Among them, the two most relevant genes are the cell cycle and immune response genes, respectively.
  • the specific method is to randomly extract 1017 chip data (50%) from 2034 gene chip expression data, calculate expression profiles and related genes, and repeat the extraction and calculation 1000 times.
  • the detection method of the present invention can be used for diagnostic or non-diagnostic purposes.
  • the present invention also provides the application of the gene population of the present invention to molecular typing of breast cancer and / or assessing the risk of distant metastasis.
  • the invention also provides the application of the gene group of the invention in the preparation of a product for molecularly typing breast cancer and / or assessing its risk of distant metastasis.
  • the product is in the form of a detection kit.
  • the invention further provides the application of the reagent for detecting the gene expression level in the gene group of the invention in preparing an in vitro diagnostic product for molecularly typing breast cancer and / or assessing the risk of distant metastasis.
  • the product is in the form of a detection kit.
  • the reagent is a reagent that detects the amount of RNA, particularly mRNA, transcribed by the gene.
  • the reagent is a reagent that detects the amount of cDNA complementary to the mRNA.
  • the reagent is a reagent that detects the amount of the polypeptide encoded by the gene, and preferably, the reagent is an antibody, an antibody fragment, or an affinity protein.
  • the reagent is a probe or a primer, especially a primer. More preferably, the sequences of the primers are as shown in SEQ ID No. 1-SEQ ID No. 132 and SEQ ID NO. 133-SEQ ID NO. 144. The specific protocol of the primers can also be shown in Table 2.
  • the subtypes of breast cancer include lumen A, lumen B, HER2 enriched, basal cell, and immune-enhanced.
  • the detection sample used in the present invention is preferably a tissue from a test object (subject object), as long as the total RNA of the test object can be extracted from the test sample.
  • the test sample is preferably one or more of a tissue sample, blood, plasma, and body fluid, and more preferably a tissue sample, such as a paraffin tissue sample.
  • the test sample is a tissue with a high content of tumor cells.
  • the present invention also relates to a group of proliferation-related genes including ASPM, AURKA, BIRC5, CCNB1, CDC20, CDK1, CENPU, CEP55, MELK, MKI67, NEK2, PRC1, PTTG1, RRM2, TOP2A, TPX2, TYMS, UBE2C, and ZWINT ( (See also the relevant information in Table 1).
  • the present invention also relates to a group of immune-related genes, including APOBEC3G, CCL5, CCR2, CD2, CD3D, CD52, CD53, CORO1A, CXCL9, GZMA, GZMK, HLA-DMA, HLA-DQA1, IL2RG, LCK, LYZ and PTPRC ( (See also the relevant information in Table 1).
  • the present invention also relates to a combination of a proliferation-related gene and an immune-related gene, wherein the proliferation-related gene and the immune-related gene are as described above.
  • the present invention also relates to the immune-related genes (APOBEC3G, CCL5, CCR2, CD2, CD3D, CD52, CD53, CORO1A, CXCL9, GZMA, GZMK, HLA-DMA, HLA-DQA1, IL2RG, LCK, LYZ and PTPRC) or all The proliferation-related genes (ASPM, AURKA, BIRC5, CCNB1, CDC20, CDK1, CENPU, CEP55, MELK, MKI67, NEK2, PRC1, PTTG1, RRM2, TOP2A, TPX2, TYMS, UBE2C, and ZWINT) or a combination of the two, Or the use of related reagents for detecting the immune-related genes, the proliferation-related genes, or a combination thereof in the preparation of in vitro diagnostic products for molecular typing of breast cancer and / or assessment of the risk of distant metastasis of breast cancer.
  • the proliferation-related genes (ASPM, AURKA, BI
  • the reagent is a reagent that detects the amount of RNA, particularly mRNA, transcribed by the gene.
  • the reagent is a reagent that detects the amount of cDNA complementary to the mRNA.
  • the reagent is a reagent that detects the amount of the polypeptide encoded by the gene, and preferably, the reagent is an antibody, an antibody fragment, or an affinity protein.
  • the reagent is a probe or a primer, especially a primer. More preferably, the sequences of the primers can be shown in Table 2.
  • the present invention also relates to a method for determining a breast cancer molecular typing and / or distant metastatic risk in a subject.
  • the steps of the method are described above, wherein the corresponding gene group is the immune-related gene, the proliferation-related gene, or a combination thereof.
  • a new breast cancer subtype (immunoenhancement type) can be determined, and different effects and effects on distant metastasis of breast cancer can be shown in different subtypes.
  • a gene group for molecular typing of breast cancer and / or assessing the risk of distant metastasis characterized in that the gene group comprises 66 genes related to molecular typing and risk assessment for distant metastasis, wherein:
  • the 66 molecular typing and distant metastasis risk assessment related genes include:
  • the gene group also includes a housekeeping gene
  • the housekeeping gene includes at least one of the following (such as 1, 2, 3, 4, 5, 6), preferably at least 3, and most preferably 6: GAPDH, GUSB, MRPL19, PSMC4, SF3A1, and TFRC.
  • a gene group for molecular typing of breast cancer and / or assessing its risk of distant metastasis characterized by:
  • the gene group includes 66 genes related to molecular typing and distant metastasis risk assessment, and 6 housekeeping genes, of which,
  • the 66 molecular typing and distant metastasis risk assessment related genes include:
  • the six housekeeping genes include: GAPDH, GUSB, MRPL19, PSMC4, SF3A1, and TFRC.
  • a diagnostic product for molecularly typing breast cancer and / or assessing its risk of distant metastasis comprising a reagent for detecting the expression level of a gene in the gene group according to any one of items 1-3.
  • reagent is a reagent that detects the amount of RNA, particularly mRNA, transcribed by the gene.
  • reagent is a reagent that detects an amount of cDNA complementary to the mRNA.
  • the diagnostic product further comprises a total RNA extraction reagent, a reverse transcription reagent, and / or a second-generation sequencing reagent.
  • the reagent is a reagent for detecting the amount of the polypeptide encoded by the gene, and preferably, the reagent is an antibody, an antibody fragment, or an affinity protein.
  • a set of primers for molecular typing of breast cancer and / or assessing the risk of distant metastasis wherein the sequence of said primers is shown in SEQ ID NO.1-SEQ ID NO.132.
  • the primer set of item 15 further comprising a primer having a sequence shown in SEQ ID NO. 133-SEQ ID NO. 144.
  • a set of primers for molecular typing of breast cancer and / or assessing the risk of distant metastasis wherein the sequence of said primers is shown in SEQ ID NO.1-SEQ ID NO.144.
  • the scheme of the present invention introduces immune regulation genes into the molecular typing of breast cancer for the first time, and can further strengthen the rationality of molecular typing of breast cancer by considering the expression levels of other genes And improve the ability to guide the clinical treatment of breast cancer, and molecularly type breast cancer into lumen A, lumen B, HER2 enriched, basal cell, and immune-enhanced.
  • a more accurate distinction can also be made between these types of transfer risks.
  • the risk of distant metastasis of luminal type A is significantly lower than the other four subtypes, while the risk of distant metastasis of immune-enhanced type is significantly lower than that of luminal type B, basal cell type and HER2 enriched type.
  • the prognosis of breast cancer with different types can be judged more accurately.
  • strong immune function can significantly reduce the risk of distant breast cancer metastasis.
  • the immune genome can be further subdivided.
  • the second-generation sequencing is used to detect gene expression and is used for molecular typing.
  • the gene group and corresponding products of the present invention can increase detection sensitivity, improve detection capability and efficiency, and greatly reduce detection costs.
  • the present invention is more accurate for molecular typing of breast cancer, can calculate the risk of distant metastasis, predict the risk of distant metastasis, and can guide clinical treatment more accurately.
  • the products and methods of the present invention have high reliability, high sensitivity, and high repeatability.
  • the products and methods of the present invention can significantly reduce the proportion of types that cannot be typed, thereby providing more accurate and efficient typing methods and products.
  • the products and methods of the present invention can also significantly reduce the so-called "medium risk” ratio. And this type of "medium risk” is difficult to judge in clinical treatment, and the guidance is poor.
  • Example 1 Screening of genes associated with breast cancer subtypes and distant metastasis risk
  • Results A total of 66 genes and 6 housekeeping genes related to breast cancer subtype and distant metastasis risk were screened out, that is, 72 gene test combinations. The gene list is shown in Table 1.
  • Example 2 Using the screened breast cancer subtypes and distant metastasis risk-related gene groups to perform molecular typing and distant metastasis risk assessment of breast cancer
  • the 66 breast cancer molecular genotyping groups shown in Table 1 were used to molecularly type the above breast cancer cases to divide breast cancer tumors into 5 subtypes (Figure 1), and their treatment can be given Different suggestions:
  • Patients with luminal A tumors have a low mutation rate of p53 gene and a good prognosis. They are not sensitive to chemotherapy and are suitable for endocrine therapy. Therefore, it has guiding significance for clinical endocrine therapy.
  • Lumen B is a tumor that is sensitive to endocrine therapy, but for HER2-positive patients, tamoxifen treatment is less effective than lumen A, and aromatase inhibitors are better. For patients with HER2-positive lumen type B tumors, molecular targeted therapy can be performed.
  • HER2-enriched tumors have a high mutation rate of p53 gene and relatively poor tumor differentiation. This type is relatively sensitive to targeted molecular therapy but has a poor prognosis.
  • HER2-enriched tumors are widely treated with Herceptin combined with systemic chemotherapy.
  • Basal cell tumors are the most invasive tumors, namely triple negative tumors (ER-, HER2-, PR-). Basal cell tumor patients are not sensitive to current breast cancer treatment options and generally have a poor clinical prognosis; however, if the tumor tissues have strong immune-related gene expression, the prognosis is relatively good.
  • the Kaplan-Meier survival curve can be drawn to obtain the corresponding metastatic risk.
  • the distant transfer risks of the five subtypes are different.
  • Luminous type A has the lowest risk of distant metastasis, which is a low-risk subtype.
  • luminal type B is lower than the HER2 enriched type and basal cell type. It is said that the lumen B type, HER2 enriched type and basal cell type are not significantly different, and they are all high-risk subtypes, while the immune-enhancing type is between the two groups and belongs to intermediate-risk subtypes.
  • Each subtype can be further divided into two groups based on the immune index, that is, the strong immune group and the weak immune group, and the difference in metastatic risk between the two groups is observed.
  • the strong immune group and the weak immune group the difference in metastatic risk between the two groups is observed.
  • Lumenal B Basal-like and HER2-enriched subtypes, the risk of distant metastasis was significantly lower in patients with strong immunity (P ⁇ 0.05). Immunely weak cases; Lumenal A, immunologically weak cases and distantly metastatic cases showed no significant difference in distant metastatic risk (Figure 3).
  • the immune index can be obtained by averaging the expression levels of the 17 immune genes:
  • the Cox model is used to calculate the risk of distant metastasis of the tumor.
  • the occurrence and time of distant metastasis are used as the observation endpoint.
  • Corresponding coefficients are determined based on the relative risk of the tumor subtype, immune index, and proliferation index for distant metastasis. Methods as below:
  • RRS 0.2x Basal + 0.4x HER2-0.2x Immune-0.1x LumA + 0.2x LumB + 0.3x proliferation index-0.1x immune index
  • Basal represents the pearson correlation coefficient between the tumor and the basal cell tumor
  • HER2 represents the pearson correlation coefficient between the tumor and the HER2-enriched tumor
  • “LumA” represents the pearson correlation coefficient between this tumor and lumen type A tumor
  • “LumB” represents the pearson correlation coefficient between the tumor and luminal type B tumor
  • Immuno represents the pearson correlation coefficient between the tumor and the immune-enhanced tumor.
  • the risk of distant metastasis of the tumor can be divided into three groups, low risk (0-32), medium risk (33-49), and high risk ( 50-100).
  • RNA-seq technology based on Illumina mid-throughput second-generation sequencing platform MiSeq was used to detect fresh tumor tissue or paraffin-embedded tumor tissue of 300 Chinese breast cancer patients.
  • the expression levels of 72 genes including genes were used as the basis for further breast cancer typing.
  • Second-generation sequencing database The raw data of all next-generation sequencing of fresh tumor tissue or paraffin-embedded tumor tissue of 300 Chinese breast cancer patients described in Example 3 will be uploaded to a web-based data storage and analysis database.
  • This method uses Java software to develop and apply many J2EE (Java Enterprise Edition) components and modes. It can 1) directly input data from Illumina NextSeq / MiSeq / MiniSeq / iSeq instruments; 2) it can display the entered data in a flexible manner.
  • J2EE Java Enterprise Edition
  • Data can be indexed according to different needs, such as gene correlation, samples or experimental groups; 3) calculation of gene expression profiles after normalization of housekeeping genes; 4) analysis of specific element details; 5) , Output data in different formats, such as XML, excel and text formats; 6), can be safely managed to ensure data privacy protection.
  • RNA sequencing Obtain postoperative paraffin samples from Chinese breast cancer patients through partner hospitals, extract RNA in accordance with the previous steps, perform quality inspection, build a library by reverse transcription, use the Illumina NextSeq / MiSeq / MiniSeq / iSeq platform for RNA sequencing, and detect 72 gene expression levels
  • PRS Precitype Risk Score
  • Example 4 Analysis method of second-generation sequencing test kit for assessing breast cancer molecular typing and distant metastasis risk gene groups
  • Step 1 Take the tumor or paraffin-embedded tissue of the test object, and use the method in the test kit to obtain the region of the test object that contains high tumor cells as the original material.
  • Step 2 Extract total RNA from the tissue.
  • Step 3 The obtained RNA is made into a library for sequencing.
  • the RNA of the obtained tissue is made into a library that can be used for targeted RNA-seq second-generation sequencing.
  • the method for preparing the library includes the following steps:
  • RNA of the extracted tissue was generated under the guidance of a specific primer using reverse transcriptase to generate cDNAs of a variety of genes of interest (72 genes described in Table 1). The ends were filled and phosphorylated at the 5 ′ end.
  • 30 ⁇ l of DNA 45 ⁇ l of purified water, 10 ⁇ l of T4 DNA ligase buffer with 10 mM ATP, 4 ⁇ l of 10 mM dNTP Mix, 5 ⁇ l of T4 DNA polymerase, 1 ⁇ l of Klenow enzyme, 5 ⁇ l of ⁇ 4 After mixing, incubate at 20 ° C for 30 minutes (reagents from Illumina sample preparation kit PE-102-1001).
  • End suspension A Dissolve the product from the previous step in 32 ⁇ l of buffer, add 5 ⁇ l of Klenow buffer, 10 ⁇ l of 1 mM dATP, and 3 ⁇ l of Klenow ⁇ -3 ⁇ l, and keep at 37 ° C for 30 minutes (reagent from Illumina sample preparation kit).
  • MinElute PCR purification kit (part # 28004) ligation: DNA was dissolved in 10 ⁇ l buffer, DNA ligase buffer 2 ⁇ 25 ⁇ l, PE adapter Adapter Oligo Mix 10 ⁇ l, DNA ligase 5 ⁇ l, and kept at 20 ° C for 15 minutes (reagent is Illumina sample Prepare the kit PE-102-1001). After warming, use the QIAGEN QIAquick PCR purification kit (part # 28104) to purify the DNA to obtain the library.
  • Step 4 Use the primer sequences in Table 2 of Example 3 to perform the next-generation sequencing on the resulting DNA library using NextSeq / MiSeq / MiniSeq / iSeq. Double-ended sequencing was performed with an Illumina NextSeq / MiSeq / MiniSeq / iSeq sequencer. This process is done automatically by the instrument itself (Illumina).
  • Step 5 Statistical analysis of results.
  • the obtained sequencing results were analyzed statistically, and breast cancer typing and risk prediction were performed according to the single sample prediction method (SSP (Single Sample Predictor) or Parker) proposed by Hu et al.
  • SSP Single Sample Predictor
  • the gene expression data of the obtained sequencing results were analyzed.
  • the detection method of the present invention is the best in both sensitivity and copy number detection capabilities.
  • the experimental research of the present invention shows that the sensitivity of the second-generation sequencing to determine the gene expression profile is much higher than that of the gene chip method, and at the same time, it is better than the quantitative PCR and Nanostring method in the detection throughput (Figure 5).
  • the detection method of the present invention has high repeatability. Correlation coefficients were higher than 0.97 in 7 repeated experiments with paraffin tissue RNA. With 15 replicates of fresh frozen tissue, the correlation coefficient was higher than 0.99 ( Figure 6).
  • CN 104293910A discloses a group of gene groups, which are composed of 60 genes (Comparative I). It can be seen from the comparison between the scheme of the gene group of the present invention and the scheme of the present invention that a new immune-enhancing type is introduced, so that the subtype of breast cancer can be classified more accurately. Moreover, these types of transfer risks can be more accurately distinguished. As mentioned above, the risk of distant metastasis of luminal type A is significantly lower than the other four subtypes, while the risk of distant metastasis of immune-enhanced type is significantly lower than that of luminal type B, basal cell type and HER2 enriched type. Furthermore, the prognosis of breast cancer with different types can be judged more accurately. For lumen B, basal cell, and HER2-enriched types in high-risk breast cancer subtypes, strong immune function can significantly reduce the risk of distant metastasis of breast cancer.
  • Example 2 of the present invention 1951 breast cancer tumors with complete clinical information in 2034 cases were subjected to typing analysis and distant metastasis risk assessment.
  • the scheme of the present invention (such as 66 in Table 1) Molecular typing and risk of distant metastasis and 6 housekeeping genes) were compared with comparison I. The results are shown in Tables 4 and 5. For a detailed description of the results, please refer to Comparative Example 2.
  • the scheme of the present invention can significantly reduce the proportion of Mixed cases.
  • the solution of the present invention provides a more complete analysis system, for example, the types of typing are increased. At the same time, more accurate typing can be achieved. It also significantly reduced the number of Mixed cases.
  • the effect of the scheme of the present invention is obviously better than that of the prior art, and the proportion of medium risk is significantly reduced.
  • Comparison II (59 genes): The 20 proliferation-related genes in the above comparison I were replaced with 19 proliferation-related genes used in the scheme of the present invention.
  • Comparison III (77 genes): The above comparison I was added to the 17 immune-related genes used in the present invention.
  • Contrast I 60 genes
  • Contrast II 59 genes
  • 12.70% Contrast III 77 genes
  • the scheme of the present invention can significantly reduce the proportion of Mixed cases. From the differences between the protocol of the present invention and Comparative II or the difference between Comparative I and Comparative III shown in the table above, it can be known that the protocol of the present invention introduces immune regulatory genes into the molecular typing of breast cancer for the first time, making the mixed case The proportion is significantly reduced.
  • the protocol of the present invention (66 molecular typing and distant metastatic risk and 6 housekeeping genes described in Table 1) is compared with Comparative I, Comparative II, and Comparative III below.
  • the distant metastasis risk score was calculated based on the gene expression level in the 1951 breast cancers described above, and the risk of distant metastasis was divided into three groups: low risk, medium risk, and high risk. The results are shown in Table 7, where Shows the proportion of "medium risk" types that are less instructive in clinical treatment in the total sample.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Genetics & Genomics (AREA)
  • Zoology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Wood Science & Technology (AREA)
  • Immunology (AREA)
  • Analytical Chemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • Molecular Biology (AREA)
  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Microbiology (AREA)
  • Biophysics (AREA)
  • Pathology (AREA)
  • Biomedical Technology (AREA)
  • Hospice & Palliative Care (AREA)
  • Oncology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Plant Pathology (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention a trait au domaine de la biotechnologie, et concerne spécifiquement un groupe de gènes utilisé pour le sous-typage du cancer du sein et l'évaluation de risque de métastase à distance, un produit de diagnostic in vitro, et une application.
PCT/CN2019/109459 2018-09-30 2019-09-30 Groupe de gènes utilisé pour le typage moléculaire du cancer du sein et le risque de métastase à distance, produit de diagnostic et application Ceased WO2020064006A2 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201980006647.6A CN111566229B (zh) 2018-09-30 2019-09-30 乳腺癌分子分型及远处转移风险基因群及诊断产品和应用

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN201811166148.7A CN110964815A (zh) 2018-09-30 2018-09-30 乳腺癌分子分型及远处转移风险基因群及诊断产品和应用
CN201811166148.7 2018-09-30

Publications (2)

Publication Number Publication Date
WO2020064006A2 true WO2020064006A2 (fr) 2020-04-02
WO2020064006A3 WO2020064006A3 (fr) 2020-04-30

Family

ID=69950303

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2019/109459 Ceased WO2020064006A2 (fr) 2018-09-30 2019-09-30 Groupe de gènes utilisé pour le typage moléculaire du cancer du sein et le risque de métastase à distance, produit de diagnostic et application

Country Status (2)

Country Link
CN (2) CN110964815A (fr)
WO (1) WO2020064006A2 (fr)

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114672556A (zh) * 2020-12-25 2022-06-28 上海善准生物科技有限公司 结直肠癌分子分型及生存风险基因群及诊断产品和应用
CN114480652A (zh) * 2022-02-21 2022-05-13 深圳市陆为生物技术有限公司 评价乳腺癌患者对于辅助内分泌治疗的响应性的产品

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP3722810A3 (fr) * 2009-02-11 2021-01-13 Caris MPI, Inc. Profilage moléculaire de tumeurs
CN102586410A (zh) * 2011-01-18 2012-07-18 苏州科贝生物技术有限公司 一种定量评估乳腺癌远期复发风险的试剂盒
US20160115551A1 (en) * 2013-05-13 2016-04-28 Nanostring Technologies, Inc. Methods to predict risk of recurrence in node-positive early breast cancer
WO2015103743A1 (fr) * 2014-01-08 2015-07-16 李铮 Puce de leptine pour identifier une série de maladies hépatiques sur base de la chaîne de sucre de glycoprotéine salivaire et application correspondante
CN104293910B (zh) * 2014-07-23 2016-07-06 周彤 一组评估乳腺癌分子分型基因群及其检测试剂盒

Also Published As

Publication number Publication date
WO2020064006A3 (fr) 2020-04-30
CN111566229A (zh) 2020-08-21
CN111566229B (zh) 2021-03-26
CN110964815A (zh) 2020-04-07

Similar Documents

Publication Publication Date Title
CA3081061C (fr) Methode d'utilisation de l'expression de klk2 pour determiner le pronostic du cancer de la prostate
US20230366034A1 (en) Compositions and methods for diagnosing lung cancers using gene expression profiles
EP2504451B1 (fr) Procédés destinés à prédire l'issue clinique d'un cancer
JP5420247B2 (ja) 肝細胞癌腫分類および予後判定のための方法
CN102575295B (zh) 可用于检测braf突变的方法、引物、探针和试剂盒
Vazquez-Mena et al. Amplified genes may be overexpressed, unchanged, or downregulated in cervical cancer cell lines
Colacino et al. Comprehensive analysis of DNA methylation in head and neck squamous cell carcinoma indicates differences by survival and clinicopathologic characteristics
CN110964814B (zh) 用于核酸序列变异检测的引物、组合物及方法
CN116761899B (zh) 结直肠癌分子分型及生存风险基因群及诊断产品和应用
WO2021036620A1 (fr) Application d'un groupe de gènes liés au pronostic du cancer de l'ovaire
CN105683391A (zh) 神经胶质瘤和肿瘤亚群中tert启动子突变
JP2016515800A (ja) 肺癌の予後および治療選択のための遺伝子サイン
WO2017112738A1 (fr) Procédés pour mesurer l'instabilité microsatellitaire
US20180044736A1 (en) Biomarker based prognostic model for predicting overall survival in patients with metastatic clear cell kidney cancer
WO2021175236A1 (fr) Groupe de gènes liés à la voie de signalisation de l'interféron, produit de diagnostic et application
WO2020064006A2 (fr) Groupe de gènes utilisé pour le typage moléculaire du cancer du sein et le risque de métastase à distance, produit de diagnostic et application
CN106498028B (zh) Egfr的t790m突变的诊断方法及试剂盒
CN108456730B (zh) 一种复发风险基因群作为标志物在制备评估乳腺癌分子分型内远处复发风险的产品中的应用
US9708666B2 (en) Prognostic molecular signature of sarcomas, and uses thereof
CN110295218B (zh) 量化靶基因的突变型等位基因负担的方法
WO2015127103A1 (fr) Méthodes de traitement du carcinome hépatocellulaire
WO2012045019A2 (fr) Déficit en gène brca et méthodes d'utilisation associées
CN110964833B (zh) 一种一管检测血浆游离dna中kras和braf基因突变的试剂盒
US20140024028A1 (en) Brca deficiency and methods of use
HK40107615A (en) Methods to predict clinical outcome of cancer

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 19865959

Country of ref document: EP

Kind code of ref document: A2

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 19865959

Country of ref document: EP

Kind code of ref document: A2