A method and device for correcting homologous recombination repair defect scoring

Abstract

A method and device for correcting homologous recombination repair defect score, the method comprising: a correction step, comprising correcting the homologous recombination repair defect score according to the following formula to obtain a corrected homologous recombination repair defect score: HRD score 矫正 =LOH score+(1‑α*WGD)*LST score+(1‑β*WGD)*TAI score; HRD score 矫正 It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomere allele imbalance score, WGD refers to the whole genome duplication information value, and α and β refer to the correction coefficient. By correcting the homologous recombination repair defect score, the accuracy of homologous recombination repair defect assessment can be effectively improved, providing guidance for the use of drugs for cancer patients.

Description

Technical Field

[0001] The present invention relates to the field of bioinformatics, and in particular to a method and a device for correcting homologous recombination repair defect scores. Background Art

[0002] Ovarian cancer is a common malignant tumor of the female reproductive system, with nearly 250,000 women diagnosed with ovarian cancer each year worldwide. With surgery and platinum- and taxane-based chemotherapy, the average 5-year survival rate is about 30%. [1] In recent years, several PARP inhibitors have been approved for marketing, effectively prolonging the progression-free survival (PFS) of ovarian cancer patients. The biomarkers of ovarian cancer PARP inhibitors have been expanded from BRCA genes to homologous recombination repair deficiency (HRD) status, which means that the population of ovarian cancer patients who benefit will expand from about 20% to more than 50%, which will greatly help improve the survival prognosis of patients.

[0003] Homologous recombination repair (HRR) is the preferred repair method for DNA double-strand breaks (DSBs). HRD usually refers to the state of HRR dysfunction at the cellular level. When HRR is dysfunctional, DSBs will overly rely on low-fidelity, high-error-prone alternative DNA damage repair pathways such as non-homologous end joining (NHEJ), microhomology mediated end joining (MMEJ) and single-strand annealing (SSA), which is likely to cause genomic instability, including identifiable gene mutations, insertion / deletion patterns, chromosome structural abnormalities, gene copy number variations, etc., which is also the theoretical basis for the current construction of HRD clinical detection methods.

[0004] There is no unified standard for HRD detection. HRD detection uses the next-generation sequencing technology (NGS) method, which usually includes two parts: BRCA1 / 2 mutation status and genomic instability score (GIS), or HRD score. There are currently two major methods for scoring genomic instability. One is based on "genomic scar" analysis of single nucleotide polymorphisms (SNPs). Globally, only two HRD detection products have been verified in large-scale Phase III clinical studies and have been approved by the FDA, specifically: Myriad CDx(Myriad GeneticLaboratories,Inc.) and FoundationFocus TM CDx BRCA LOH (Foundation Medicine, Inc.). In Myriad In CDx testing, HRD positivity is defined as tumor BRCA1 / 2 mutation and / or GIS score ≥42 points. The GIS score is calculated by the comprehensive calculation of loss of heterozygosity (LOH), telomereal allele imbalance (TAI) and large-scale state transitions (LST). The threshold is set based on the HRD score of the 5th quartile of ovarian and breast cancer tumor samples with BRCA defects, and was first proven to be effective in predicting the sensitivity of breast cancer and ovarian cancer to platinum-containing chemotherapy. However, the detection product did not take into account the characteristics of the Oriental population when selecting SNP sites and setting thresholds. In FoundationFocus TM In the CDx BRCA LOH test, HRD positivity is defined as tumor BRCA1 / 2 mutation and / or genomic LOH score ≥ 16%. The threshold was initially set based on its effectiveness in distinguishing the effects of platinum-containing chemotherapy in ovarian cancer patients, and was subsequently adjusted based on its effectiveness in distinguishing the effects of rucaparib treatment in ovarian cancer patients. Studies have reported that the positive rate of BRCA1 / 2 gene mutation samples that can be identified by this test product is higher than that of Myriad Low CDx. Another major category is the detection methods based on single or multiple dimensional data features including mutation features obtained from whole exome (WES) or whole genome (WGS) data. For example, SigMA, HRDetect and ShallowHRD of mutation feature Signature 3. The disadvantages of WGS detection, such as the need for higher sequencing data volume and the inability to cover somatic mutations in the HRR signaling pathway, limit its clinical application. Summary of the invention

[0005] According to the first aspect, in one embodiment, a method for correcting homologous recombination repair deficiency score is provided, comprising:

[0006] The correction step includes correcting the homologous recombination repair deficiency score according to the following formula to obtain a corrected homologous recombination repair deficiency score:

[0007] HRD score 矫正 =LOH score+(1-α*WGD)*LST score+(1-β*WGD)*TAI score;

[0008] HRD score 矫正 It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomeric allele imbalance score, WGD refers to the whole genome copy information value (also known as WGD value), and α and β refer to the correction coefficients.

[0009] According to the second aspect, in one embodiment, a method for assessing homologous recombination repair deficiency is provided, comprising:

[0010] The prediction step includes predicting whether the patient to which the sample to be tested belongs has homologous recombination repair deficiency according to the corrected homologous recombination repair deficiency score obtained by the method of the first aspect.

[0011] According to the third aspect, in one embodiment, a device for correcting homologous recombination repair deficiency score is provided, comprising:

[0012] The correction module is used to correct the homologous recombination repair defect score according to the following formula to obtain a corrected homologous recombination repair defect score:

[0013] HRD score 矫正 =LOH score+(1-α*WGD)*LST score+(1-β*WGD)*TAI score;

[0014] HRD score 矫正It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomeric allele imbalance score, WGD refers to the whole genome copy information value (also known as WGD value), and α and β refer to the correction coefficients.

[0015] According to the fourth aspect, in one embodiment, a device for evaluating homologous recombination repair deficiency is provided, comprising:

[0016] A prediction module is used to predict whether the patient to which the sample to be tested belongs has homologous recombination repair deficiency based on the corrected homologous recombination repair deficiency score obtained by the method of the first aspect and / or the device of the third aspect.

[0017] According to the fifth aspect, in one embodiment, there is provided an apparatus, comprising:

[0018] Memory, used to store programs;

[0019] A processor is used to implement the method as described in the first aspect and / or the second aspect by executing the program stored in the memory.

[0020] According to the sixth aspect, in one embodiment, a computer-readable storage medium is provided, on which a program is stored, and the program can be executed by a processor to implement the method as described in the first aspect and / or the second aspect.

[0021] According to the method and device for correcting homologous recombination repair deficiency score of the above embodiment, the accuracy of homologous recombination repair deficiency assessment is effectively improved by correcting the homologous recombination repair deficiency score, thereby providing guidance for the medication of cancer patients. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a flow chart for evaluating homologous recombination repair deficiency in Example 1;

[0023] Figure 2 This is a graph showing the consistency of HRD and WGS results in Example 2;

[0024] Figure 3 This is a graph showing the consistency of HRD and the standard product in Example 2;

[0025] Figure 4 This is a survival curve diagram of HRD and platinum efficacy in Example 3. DETAILED DESCRIPTION

[0026] The present invention is further described in detail below by specific embodiments in conjunction with the accompanying drawings. Wherein similar elements in different embodiments adopt associated similar element numbers. In the following embodiments, many detailed descriptions are for making the present application better understood. However, those skilled in the art can easily recognize that some features can be omitted in different situations, or can be replaced by other elements, materials, methods. In some cases, some operations related to the present application are not shown or described in the specification, this is to avoid the core part of the present application being overwhelmed by too much description, and for those skilled in the art, it is not necessary to describe these related operations in detail, and they can fully understand the related operations according to the description in the specification and the general technical knowledge in the art.

[0027] In addition, the features, operations or characteristics described in the specification can be combined in any appropriate manner to form various implementations. At the same time, the steps or actions in the method description can also be interchanged or adjusted in a manner that is obvious to those skilled in the art. Therefore, the various sequences in the specification and the drawings are only for the purpose of clearly describing a certain embodiment and are not meant to be a required sequence, unless otherwise specified that a certain sequence must be followed.

[0028] The serial numbers of the components in this document, such as "first", "second", etc., are only used to distinguish the objects described and do not have any order or technical meaning. The "connection" and "coupling" mentioned in this application, unless otherwise specified, include direct and indirect connections (couplings).

[0029] Explanation of terms

[0030] TAI: Telomeric Allelic Imbalance, telomeric allelic imbalance.

[0031] LST: Large-scale state Transition, large-scale migration.

[0032] WGD: whole genome duplication, also known as whole genome duplication.

[0033] CNV: Copy number variations, gene copy number variation.

[0034] SNV: Single Nucleotide Variant, single nucleotide site variation.

[0035] Reads: The nucleotide sequence generated by the sequencer is called a "read segment". The length of the sequence read segment can range from tens to thousands of nucleotides.

[0036] BAF: Ballele frequency, minor allele frequency, refers to the gene frequency of the second most common genotype (minor allele) in a given population.

[0037] HRD: homologous recombination deficiency, also known as homologous recombination deficiency, homologous repair deficiency, cancer has homologous recombination repair deficiency / homologous repair deficiency ("HRD") or is characterized by mutations or deletions in homologous recombination repair (HRR) genes.

[0038] LOH: Loss of heterozygosity, refers to the loss of one of the two alleles (or part of the nucleotide fragments) located at the same gene locus on a pair of homologous chromosomes, while it still exists on the paired chromosome.

[0039] TAI: Telomeric Allelic Imbalance, telomeric allelic imbalance.

[0040] LST: Large-scale state Transition, large-scale migration.

[0041] WGD: whole genome duplication, also known as whole genome duplication.

[0042] Reads: The nucleotide sequence generated by the sequencer is called a "read segment". The length of the sequence read segment can range from tens to thousands of nucleotides.

[0043] BRCA: Breast cancer susceptibility gene, breast cancer susceptibility gene, is an important tumor suppressor gene, including BRCA1 and BRCA2. BRCA1 / 2 genes are important biomarkers for assessing the risk of breast cancer, ovarian cancer and other related cancers, and are also biomarkers that affect the selection of individualized treatment plans for patients, so BRCA testing has important clinical significance.

[0044] PARP inhibitors: Also known as PARPi, are medical agents that affect the way cancer cells replicate themselves.

[0045] According to the first aspect, in one embodiment, a method for correcting homologous recombination repair deficiency score is provided, comprising:

[0046] The correction step includes correcting the homologous recombination repair deficiency score according to the following formula to obtain a corrected homologous recombination repair deficiency score:

[0047] HRD score 矫正 =LOH score+(1-α*WGD)*LST score+(1-β*WGD)*TAI score;

[0048] HRD score 矫正 It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomeric allele imbalance score, WGD refers to the whole genome duplication information value (also known as WGD value), α and β refer to the correction coefficient. LOH score, LST score, TAI score, and WGD can all be calculated using existing software.

[0049] In one embodiment, α may be 0.35, and β may be 0.35.

[0050] In one embodiment, the whole genome replication information value is 0, 1 or 2.

[0051] In one embodiment, the whole genome replication information value is the whole genome replication information value output by the optimal model. The screening method of the optimal model is an existing method, which can be specifically referred to the method described in paragraphs 98 to 108 of the specification of the Chinese patent "Method and device for identifying tumor purity and absolute copy number based on sequencing data" with application number 202010567812.X.

[0052] In one embodiment, the filtered system CNV and SNV mutation site set is used as the input of the open source software ABSOLUTE, and the WGD value, purity value, and ploidy value simulated by the sample to be tested under multiple models are output. The predicted models are screened, the optimal model is determined, and the WGD value, purity value, and ploidy value under the optimal model are output with reference to the method described in paragraphs 98 to 108 of the specification of the Chinese patent "Method and device for identifying tumor purity and absolute copy number based on sequencing data" with application number 202010567812.X.

[0053] In one embodiment, LOH score, LST score, and TAI score are calculated based on the single nucleotide site variation set and gene copy number variation site set of the sequencing data of the sample to be tested. Existing software can be used for calculation, including but not limited to scarHRD software.

[0054] In one embodiment, the sample to be tested includes but is not limited to a tumor tissue sample.

[0055] In one embodiment, the tumor comprises a solid tumor.

[0056] In one embodiment, the tumor includes but is not limited to ovarian cancer.

[0057] In one embodiment, the gene copy number variation site set of the sequencing data of the test sample is obtained by analyzing the sequencing data of the test sample and the sequencing data of the control sample.

[0058] In one embodiment, the control sample and the test sample are from the same patient.

[0059] In one embodiment, the control sample includes but is not limited to at least one of blood cells and paracancerous tissues. When the control sample is blood cells, the effective component to be detected is the white blood cells in the blood cells.

[0060] In one embodiment, the organism is a human.

[0061] In one embodiment, both the test sample and the control sample contain DNA.

[0062] In one embodiment, the sequencing data is sequencing data generated based on next generation sequencing (NGS).

[0063] In one embodiment, the sequencing data is sequencing data aligned to a reference genome sequence.

[0064] In one embodiment, the reference genome sequence is derived from a reference cohort.

[0065] In one embodiment, the reference genome sequence comprises a consensus sequence from a reference population.

[0066] In one embodiment, the reference genome sequence comprises at least a portion of a hs37d5 genome, a GRCh37 genome, a b37 genome, a hgl8 genome, a hgl7 genome, a hgl6 genome, or a hg38 genome.

[0067] In one embodiment, the reference genome sequence is GRCh37 (also known as hg19).

[0068] In an embodiment, the sequencing data comprises capture sequencing data.

[0069] In one embodiment, the capture sequencing data includes at least one of whole exome sequencing (WES) data and regional capture sequencing data.

[0070] According to the second aspect, in one embodiment, a method for assessing homologous recombination repair deficiency is provided, comprising:

[0071] The prediction step includes predicting whether the patient to which the sample belongs has homologous recombination repair deficiency according to the corrected homologous recombination repair deficiency score obtained by the method of the first aspect. The result obtained by this method is only an intermediate result, not a final diagnosis result, and therefore does not belong to the diagnosis and treatment method of the disease.

[0072] In one embodiment, the clinical benefit of a patient receiving drug treatment can be predicted based on whether the patient has homologous recombination repair deficiency.

[0073] In one embodiment, the drug includes but is not limited to a PARP inhibitor.

[0074] In one embodiment, based on the relationship between the corrected homologous recombination repair deficiency score and the threshold, it is predicted whether the patient to which the sample to be tested belongs has homologous recombination repair deficiency.

[0075] In one embodiment, if the corrected homologous recombination repair defect score is ≥ the threshold, the patient to which the sample to be tested belongs is predicted to be a homologous recombination repair defect-positive patient; if the corrected homologous recombination repair defect score is < the threshold, the patient to which the sample to be tested belongs is predicted to be a homologous recombination repair defect-negative patient.

[0076] In one embodiment, the homologous recombination repair deficiency-positive patients include patients with functional loss of breast cancer susceptibility genes.

[0077] In one embodiment, a patient with a breast cancer susceptibility gene loss of function refers to a patient with a breast cancer susceptibility gene with biallelic mutations, and a patient with a breast cancer susceptibility gene with intact function refers to a patient with a breast cancer susceptibility gene with a monoallelic mutation or a wild-type breast cancer susceptibility gene.

[0078] In one embodiment, the threshold is 39.

[0079] According to the third aspect, in one embodiment, a device for correcting homologous recombination repair deficiency score is provided, comprising:

[0080] The correction module is used to correct the homologous recombination repair defect score according to the following formula to obtain a corrected homologous recombination repair defect score:

[0081] HRD score 矫正 =LOH score+(1-α*WGD)*LST score+(1-β*WGD)*TAI score;

[0082] HRD score 矫正It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomeric allele imbalance score, WGD refers to the whole genome copy information value (also known as WGD value), and α and β refer to the correction coefficients.

[0083] According to the fourth aspect, in one embodiment, a device for evaluating homologous recombination repair deficiency is provided, comprising:

[0084] The prediction module is used to predict whether the patient to which the sample to be tested belongs has homologous recombination repair deficiency based on the corrected homologous recombination repair deficiency score obtained by the method of the first aspect and / or the device of the third aspect.

[0085] According to the fifth aspect, in one embodiment, there is provided an apparatus, comprising:

[0086] Memory, used to store programs;

[0087] A processor is used to implement the method of the first aspect and / or the second aspect by executing a program stored in a memory.

[0088] According to the sixth aspect, in one embodiment, a computer-readable storage medium is provided, on which a program is stored, and the program can be executed by a processor to implement the method of the first aspect and / or the second aspect.

[0089] Expert consensus on clinical detection and application of homologous recombination repair deficiency (2021 edition) [2] It is suggested that the sample size, data volume, and somatic mutations required for comprehensive detection can be combined to use a high-depth WES plus high-density SNPs method to comprehensively detect genetic mutations, somatic mutations, and genomic instability in the HRR signaling pathway in tumor samples. In one embodiment, the present invention uses a high-depth WES plus high-density SNPs method to evaluate HRD.

[0090] In one embodiment, the present invention uses a high-depth WES plus high-density SNPs method to simultaneously and comprehensively detect inherited mutations, somatic mutations, and genomic instability states in the HRR signaling pathway in tumor samples, thereby providing more comprehensive medication guidance for ovarian cancer patients.

[0091] Example 1

[0092] The evaluation method of this embodiment mainly includes the following steps:

[0093] (1) For tumor samples and paired blood cell samples, the sequencing data is used to generate sequencing data files (bam format) after alignment, sorting, filtering, and duplicate marking;

[0094] (2) Use Mutect2 software to perform SNV analysis on the tumor samples to obtain the mutation set of the system SNV, including information such as mutation frequency and mutation site depth;

[0095] (3) Annotate the obtained system SNVs;

[0096] (4) Using the cnvkit software, the bam of the tumor sample to be tested and the paired sample are used as input to analyze and obtain the segment where the CNV mutation occurs in the sample, and output information such as the size of the segment, the number of probes contained in the segment, and the BAF value of the segment;

[0097] (5) Using ABSOLUTE software to analyze the purity and ploidy of the tumor sample to be tested, and obtain the purity, ploidy, and whole genome duplication WGD value of the tumor sample to be tested;

[0098] (6) The obtained CNV and SNV information of the tumor sample to be tested is used as the input of the scarHRD software to calculate the LOH score, TAI score, and LST score of the sample to be tested. The HRD score of the homologous recombination repair defect score is the sum of the LST score, TAI score, and LOH score;

[0099] (7) Use the WGD value of the tumor sample to correct the LST score and TAI score, and output the corrected HRD score.

[0100] Figure 1 The flowchart of the present embodiment for evaluating homologous recombination repair deficiency is shown. The evaluation method is specifically divided into the following steps:

[0101] (1) The steps of obtaining the system SNV mutation site set include: obtaining the bam by comparing the data of the tumor sample to be tested and its paired blood cell sample, which is used as the input of the SNV mutation analysis software Mutect2 to analyze and obtain the SNV mutation sites of the tumor sample to be tested and the mutation frequency and mutation site depth of each SNV mutation site; using annotation software to annotate the SNV mutation sites at the gene level to obtain the gene sites where SNV mutations occur.

[0102] (2) The step of obtaining the system CNV mutation site set includes analyzing the segment segments of the tumor sample to be tested where CNV mutations occur, the size of each segment, the number of probes contained in each segment, and the BAF value of each segment based on the comparison bam result file of the tumor sample to be tested, using cnvkit software to annotate the CNV mutation sites and obtain the system CNV mutation site set;

[0103] (3) Purity and ploidy value acquisition step: Use the filtered system CNV and SNV mutation site set as the input of the open source software ABSOLUTE, and output the WGD value, purity value (purity value), and ploidy value (ploidy value) simulated under multiple models of the sample to be tested. Refer to the method described in paragraphs 98 to 108 of the specification of the Chinese patent application number 202010567812.X "Method and device for identifying tumor purity and absolute copy number based on sequencing data", the predicted model is screened to determine the optimal model, and the WGD value, purity value, and ploidy value under the optimal model are output.

[0104] (4) The homologous recombination repair defect scoring step includes using the obtained system SNV mutation site set and system CNV mutation site set, and using scarHRD software to analyze and calculate the LOH score, TAI score, and LST score of the tumor sample to be tested; the homologous recombination repair defect score HRD score is the sum of the LST score, TAI score, and LOH score.

[0105] (5) Correction of homologous recombination repair defect score (HRD score) using WGD: Using the exhaustive method, the correction coefficients of LST index and TAI index were determined to be 0.35 respectively. When the threshold was 39, the HRD positive ratio of BRCA-positive was 90.8% (59 / 65), which was greater than 90%; the HRD positive ratio of BRCA-negative was 47% (40 / 85), which was less than 50%. The HRD score was the sum of the corrected LST score, the corrected TAI score, and the original LOH score. The specific formula is as follows:

[0106] HRD score 矫正 =LOH score+(1-0.35*WGD)*LST score+(1-0.35*WGD)*TAI score;

[0107] Among them, HRD score 矫正 It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomeric allele imbalance score, WGD refers to the whole genome duplication information value (also known as WGD value), and 0.35 is the correction coefficient.

[0108] The reasons for correction mainly include: the HRD scores of samples with WGD are significantly higher than those of samples without WGD. If no correction is made, there will be more false positive results.

[0109] The exhaustive method is as follows: exhaustive α and β coefficients (0, 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45), exhaustive thresholds (integers between 20 and 49), a total of 3000 combinations, from which BRCA-positive HRD-positive ratios greater than 90% and BRCA-negative HRD-positive ratios less than 50% were screened out, and the HRD values ​​of samples with WGD (genome duplication) can be corrected back to normal (no significant difference from samples without WGD).

[0110] The HRD score threshold is determined as follows:

[0111] 1. Whole-genome DNA Extraction

[0112] After adding proteinase K and incubation buffer to paraffin-embedded tissue (FFPE) for incubation outside the machine, the whole genome DNA was extracted by magnetic beads using a Promega Maxwell 16 instrument. The specific steps are as follows:

[0113] (1) Scraping of paraffin-embedded tissue. The order is from top to bottom and from right to left.

[0114] (2) The samples were centrifuged briefly and incubated overnight at 70°C and 1220 rpm.

[0115] (3) Transfer the incubated paraffin into a new 1.5 mL EP tube and add 400 μL of lysis buffer. Centrifuge the sample mixture with lysis buffer.

[0116] (4) Open the FFPE automated extractor and set program 01, load the reagent strip into the support plate, insert the resolubility tube into the corresponding hole of the support plate, and add 75 μl of TE buffer into each resolubility tube.

[0117] (5) Remove the seal of the reagent strip, add the sample mixture into the sample well of the reagent strip, and mix by pipetting. After the transfer, insert the LEV tube sleeve into the reagent strip, remove the re-dissolution tube, and centrifuge the re-dissolution tube instantly.

[0118] (6) Transfer the DNA and magnetic bead mixture in the centrifuged resolubilization tube into a new 1.5 mL EP tube and place the tube on the magnetic rack. After all the magnetic beads are adsorbed on the tube wall, aspirate 60 μL of DNA into the labeled 1.5 mL EP tube.

[0119] 2. Library Construction

[0120] The whole genome DNA was fragmented using an AFA ultrasonic disruptor. The main peak size of the fragmented DNA was 200-250 bp. The experimental steps included:

[0121] (1) Add sample and start interruption procedure.

[0122] (2) After fragmentation, 60 ng of the fragment was taken for electrophoresis detection to determine the fragmentation result. Samples with qualified fragmentation results were used to build the library.

[0123] The fragmented samples are purified by magnetic beads to recover the fragmented DNA, and then undergo end repair and "A" addition, adapter ligation, purification after adapter ligation, pre-capture PCR (Non-C-PCR), and post-PCR purification to obtain the pre-capture intermediate library. The specific operations are as follows:

[0124] (21) Terminal repair and adding "A"

[0125] Prepare the end repair and "A" reaction Premix (Mix 1) according to Table 1, vortex to mix and centrifuge. Premix refers to the premixed solution.

[0126] Table 1 EP Mix preparation table

[0127] Components Single reaction volume (μL) NEBNext Ultra II End Prep Reaction Buffer 7 NEBNext Ultra II End Prep Enzyme Mix 3 Total volume 10

[0128] Premix (Mix 1) was dispensed into 10 μL per reaction, added to 50 μL of the DNA sample after interruption and purification, vortexed to mix, and centrifuged. Incubated on a thermomixer or PCR instrument according to the reaction conditions in Table 2. After incubation, centrifuged briefly and collected the evaporated droplets into a tube.

[0129] Table 2 EP reaction conditions

[0130]

[0131]

[0132] (22) Connector connection

[0133] The dissolved NEBNext Μltra II Ligation Master Mix, NEBNext Ligation Enhancer and adapter were mixed by vortexing and centrifuged. According to Table 3, the adapter ligation reaction Premix (Mix 2) was prepared, mixed by vortexing and centrifuged.

[0134] Table 3 Connection Mix configuration table

[0135] Components Single reaction volume (μL) NEBNext Ultra II Ligation Master Mix 30 NEBNext Ligation Enhancer 1 Total volume 31

[0136] Please refer to Table 4 for the corresponding relationship of adapter addition amount.

[0137] Table 4 Adapter addition amount comparison table

[0138] Starting amount for library construction (ng) 15μM adapter volume (μL) 100～800 4 20～100 2

[0139] The amount of the linker added in this example is 4 μL.

[0140] Premix (Mix 2) is dispensed. According to the amount of each reaction, add Premix (Mix 2) and the corresponding volume of connectors to the reaction tube in turn, shake to mix and centrifuge. Place the reaction tube on a constant temperature mixer and incubate at 20°C for 15 minutes. During the incubation, dispense the molecular label (also known as index, the molecular label of the sample, used to distinguish different samples) according to the task list. After the incubation is completed, centrifuge briefly to centrifuge the liquid on the tube wall to the bottom of the tube.

[0141] (23) Purification after adapter ligation

[0142] Add 87 μL of Axygen magnetic beads to each reaction tube, mix by pipetting or lightly shaking, and incubate at room temperature for 10 minutes to allow the magnetic beads to fully bind to the DNA fragments. Prepare 80% ethanol during the incubation period. After slight centrifugation, place the centrifuge tube on the magnetic rack until it is clear. Discard the supernatant, open all the centrifuge tubes on the magnetic rack, and add 500 μL of 80% ethanol in sequence.

[0143] (24) Pre-capture PCR (Non-C-PCR)

[0144] Dissolve the Index and KAPA HiFi HotStart ReadyMix at room temperature in advance, shake and mix the dissolved reagents, and centrifuge.

[0145] Arrange PCR tubes according to the sample quantity on the task sheet, and mark the corresponding serial numbers on the tubes. Sort the dissolved index and samples according to the order on the task sheet, add the corresponding reaction components to the PCR tubes, ensure that the index, sample, and PCR tube correspond to the task sheet one by one, mix well, and centrifuge. As shown below, Table 5-1 is the NC-PCR Mix preparation table for single-end primers, and Table 5-2 is the NC-PCR Mix preparation table for double-end primers. Add the primers correctly according to the primer types on the task sheet.

[0146] Table 5-1 Single-end primer NC-PCR Mix preparation table

[0147]

[0148]

[0149] Table 5-2 Preparation of double-ended primer NC-PCR Mix

[0150] Components Single reaction volume (μL) index-GF(10p) 2.5 index-GR(10p) 2.5 KAPA HiFi HotStart ReadyMix 25 Adapter-Ligated library 20 Total volume 50

[0151] Set up negative and positive references: Each PCR reaction needs to set up positive and negative references. TE is used instead of samples in the negative reference wells. 1 μL of positive reference is added to the positive reference wells and the volume is filled up with TE. Specific indexes are used for PCR for positive and negative samples. Place the samples in the PCR instrument and check the number and PCR reaction program. The PCR program is shown in Table 6, with a hot cover temperature of 105°C and a 50 μL system.

[0152] Table 6 NC-PCR reaction procedure

[0153]

[0154] (25) NC-PCR product purification

[0155] Prepare a set of 1.5mL EP tubes according to the task, mark the serial number on the tube cap, print the label, and stick it on the corresponding EP tube; use 45μL Axygen magnetic beads to purify the PCR product, and finally dissolve it in 31μL TE (pH 8.0) buffer. Transfer the purified product to the prepared EP tube.

[0156] 3. Hybridization capture

[0157] The intermediate library with qualified concentration quality control is mixed (pooling refers to mixing libraries of different samples together), evaporated, probe hybridized, eluted, PCR of eluted products, and purified after PCR to obtain the common library after hybridization. The specific operation steps are as follows:

[0158] (301) Prepare the plate: According to the number of hybridization groups in the hybridization table, add 5 μL of cot1 solution to the wells of the hybridization plate, and start the pooling automated PCR plate operation.

[0159] (302) Place the pooled hybridization plate in a vacuum concentrator and prepare the plate for evaporation. Seal the hybridization plate concentrated into dry powder and store it for later use.

[0160] (303) Start the PCR instrument hybridization program at least 30 min in advance, set the cover temperature to 100 °C, and start the thermostatic mixer in advance, set the temperature to 25 °C, 4 min, and 2000 rpm;

[0161] (304) Denature before hybridization, prepare denaturation buffer, shake and mix, and then dispense into the evaporated hybridization plate, add the prepared probe into the hybridization plate, 4 μL / case; shake and mix thoroughly and centrifuge, place on a constant temperature mixer for 5 min at room temperature, 4 min at 25 °C, 2000 rpm;

[0162] (305) Place the hybridization plate in a PCR instrument and follow the procedure as shown in the table below. Start hybridization and record the hybridization start time. The hybridization time should be no less than 4 h.

[0163] Table 7

[0164] step Temperature(℃) time 1 95 30s 2 65 hold

[0165] (306) Take out the M-270 magnetic beads from 4 °C 30 min in advance and equilibrate to room temperature. The amount of magnetic beads used is 20 μL / case. The magnetic beads are divided into eight rows and labeled with serial numbers.

[0166] (307) Adjust the volume of the pump gun to 200 μL, add Bead Wash Buffer to the M-270 magnetic beads, pipette to mix, place on a magnetic stand, let stand until clear, and discard the supernatant. Repeat once.

[0167] (308) Adjust the volume of the pump gun to 100 μL, add Bead Wash Buffer to the M-270 magnetic beads, pipette to mix, place on a magnetic rack, let stand until clear, discard the supernatant, and use a 10 μL pump gun to remove any remaining liquid.

[0168] (309) Place the hybridization PCR plate in a 65°C constant temperature mixer, transfer the hybridization product to the magnetic beads, and shake to mix.

[0169] (310) The eight-well plate was placed in a 65 °C PCR instrument and incubated for 45 min, with shaking every 15 min.

[0170] (311) The 65 °C incubation reagent was transferred to a 65 °C thermomixer. The sample was removed from the PCR instrument, 100 μL of 65 °C Wash Buffer I was added, the sample was mixed by pipetting, and the sample was placed on a magnetic rack. The sample was allowed to stand until clear, the supernatant was discarded, and the sample was removed from the magnetic rack.

[0171] (312) Add 170 μL of Stringent Wash Buffer taken from the magnetic rack, mix well by pipetting, and incubate in a 65°C PCR instrument for 5 min. This operation is repeated twice.

[0172] (313) The sample was placed on a magnetic rack and allowed to stand until it was clear. The supernatant was discarded and the sample was removed from the magnetic rack. 170 μL of Wash Buffer I, Wash Buffer II, and Wash Buffer III were added in sequence and mixed by pipetting. Wash Buffer I, Wash Buffer II, and Wash Buffer III are reagents in the 96rxn xGEN-lockdown-reagents kit (purchased from IDT).

[0173] (314) Place the sample on a magnetic rack, let it stand until it is clear, discard the supernatant, remove it from the magnetic rack, and add 20 μL NF-H 2 O and mix by oscillation.

[0174] (315) Set up negative and positive references: Use NF-H for the negative reference well. 2 O replaces the sample, and 1 μL positive reference + 19 μL NF-H is added to the positive reference well. 2 O.

[0175] (316) Take 20 μL of Yang Shen sample and add it to the eight-row plate containing Mix. Set up the PCR program shown in the table below.

[0176] Table 8

[0177]

[0178]

[0179] (317) After the program is finished, centrifuge the sample and place it on a magnetic rack. Take a shallow well plate and transfer the sample to the plate in order. Add 60 μL of Axygen magnetic beads to each well and mix by pipetting 30 times. After incubating at room temperature for 10 min, place the shallow well plate on a magnetic rack and let it stand until it is clear. Discard the supernatant.

[0180] (318) Add 170 μL of 80% ethanol to each well, pipette gently, let stand for a while, and discard the supernatant. Repeat once.

[0181] (319) The residual liquid was sucked out with a 10 μL spray gun and the shallow well plate was placed on a 38 °C dryer to heat and dry until cracks appeared on the surface of the magnetic beads. The shallow well plate was then removed from the dryer.

[0182] (320) 31 μL of TE (pH 8.0) buffer was added to the shallow well plate, pipetted to mix, and incubated at room temperature for 5 min to allow the DNA fragments to fully dissolve in the TE buffer.

[0183] (321) The shallow well plate was placed on a magnetic stand until completely clarified, and the supernatant purified product was transferred to a 1.5 mL EP tube.

[0184] (322) The purified product was quantified using an ELISA reader, and the eluted library was sent to the quality control unit for Labchip quality control.

[0185] 4. Sequencing and Information Analysis

[0186] Paired-end (PE100) sequencing was performed using the Gene+Seq2000 sequencer, and the sequencing process was subjected to multiple quality control checks.

[0187] 4.1 Data Management System

[0188] Genenjia's data management system matches sample type, sampling site, sample number, library number, index number, and computer information one by one, and binds this information to each link of data analysis. After the data analysis is completed, it records the process name and version information, database name and version information used in this data analysis, and saves the key file information of the fastq, BAM and VCF files corresponding to the sample.

[0189] 4.2 FASTQ Data Output

[0190] The software splitBarcode (version: 0.1.3) was used to extract the full-length reads information corresponding to each sample from the offline file in combination with the index sequence information corresponding to the sample, and the reads of the double-end sequencing were stored in two fastq files in a fixed naming format. The sequencing data volume of the tumor sample was 60G, with an effective depth of 500×; the sequencing data volume of the control sample was 18G, with an effective depth of 200×.

[0191] 4.3Index matching exception check

[0192] In the mutation detection stage, index matching abnormalities or cross-contamination between samples are detected by identifying abnormalities and matches of homozygous sites in tumor samples and control samples.

[0193] 4.4 Data comparison and bam file generation

[0194] Before data alignment, fastp (version: 0.20.0) software was used to filter low-quality reads in the input fastq file. The filtered high-quality reads were aligned to the human genome (version: hs37d5) by BWA (version: 0.7.15-r1140) software to generate the bam file of the initial alignment result. Then, sentieon software (version: 201808) was used to mark the PCR duplicate reads in the initial alignment result file in turn, and the indel regions within the 50 bp range of both ends of the detection chip were re-aligned in combination with the common indel mutations in the 1000 database and dbSNP (version: 138) database. The base quality values ​​within the 50 bp range of both ends of the detection chip were re-corrected in combination with the information in the 1000 database, dbSNP (version: 138) database, and COSMIC database.

[0195] 4.5 Sample Quality Control (QC)

[0196] (1) Sample pairing error

[0197] The bioinformatics process is as follows: the consistency between the homozygous sites in the control sample and the tumor sample within the range of 50bp at both ends of the chip interval is calculated to determine whether the sample pairing is abnormal. If the consistency is lower than 90%, it is considered that the control and tumor samples tested are at risk of pairing errors.

[0198] (2) Sample contamination

[0199] The Calculate Contamination module in the GATK (version: 4.1.4) software was used to combine the bam file information of the control sample and the tumor sample. The reads information supporting the reference base in the homozygous site in the test sample was read and counted to evaluate the cross-contamination of the sample. If the contamination rate was higher than 3%, it was considered that there was a risk of cross-contamination between the samples, and the sample was discarded without proceeding to the subsequent steps.

[0200] 4.6 SNV and CNV calling

[0201] The bam obtained by comparing the data of the tumor sample to be tested and its paired blood cell sample is used as the input of the SNV mutation analysis software Mutect2. The SNV analysis of the tumor sample to be tested is performed using Mutect2 software to obtain the mutation set of the system SNV, including information such as mutation frequency and mutation site depth, and the obtained system SNV is annotated. The mutations obtained in this step are filtered, and the mutations that meet the following conditions are retained: (1) mutations located in the hotspot set and the number of reads supporting the mutation (AD) ≥ 4 and the mutation frequency (AF) ≥ 0.01; (2) mutations located in the hotspot area and the mutation type is not SNV and AD ≥ 4 and A ≥ 0.01; (3) mutations not located in the hotspot area and the hotspot set and AF ≥ 0.012 and AD ≥ 8 and the mutation label is PASS. Mutations that meet any of the above three conditions are retained.

[0202] Using cnvkit software, the bam files of the tumor samples to be tested and the paired samples were used as input to analyze the segments where CNV mutations occurred in the samples, and output information such as the size of the segment, the number of probes contained in the segment, and the BAF value of the segment.

[0203] 5. Determination of HRD score threshold

[0204] The filtered CNV and SNV mutation site sets were used as inputs to the open source software ABSOLUTE, and the WGD values, purity values, and ploidy values ​​of the samples to be tested were output under multiple models. The predicted models were screened to determine the optimal model (the optimal model for each sample was different) with reference to the method described in paragraphs 98 to 108 of the specification of the Chinese patent "Method and device for identifying tumor purity and absolute copy number based on sequencing data" with application number 202010567812.X. The WGD value, purity value, and ploidy value under the optimal model were output. The scarHRD software was used to analyze and calculate the LOH score, TAI score, and LST score of the tumor samples to be tested; the homologous recombination repair defect score HRD score was the sum of the LST score, TAIscore value, and LOH score value, and finally the homologous recombination repair defect score (HRD score) was corrected using WGD.

[0205] The reasons for correction are as follows: The HRD scores of samples with WGD are significantly higher than those without WGD. If not corrected, there will be more false positive results. When BRCA-negative samples were not corrected, the HRD score p value between the sample set with WGD and the sample set without WGD was 0.0023, which was significantly different. The p value after correction was 0.82, which was not significantly different. The HRD positive proportion of the sample set without WGD was 49.3% (33 / 67); the HRD positive proportion of the sample set with WGD (uncorrected) was 74.3% (29 / 39), which was high; the HRD positive proportion of the sample set with WGD (corrected) was 43.6% (17 / 39).

[0206] When WGD does not exist, WGD=0; when WGD exists, WGD=1 or 2.

[0207] In order to determine the threshold of HRD score, this example selected 151 samples of high-grade serous ovarian cancer patients, divided into two groups of samples: BRCA-deficient (65 cases) and BRCA-functional (86 cases). Specifically, the BRCA-deficient tumors were selected to have (a) one deleterious mutation in BRCA1 or BRCA2, LOH in the wild-type copy; or (b) two deleterious mutations in the same gene, and the BRCA-functional tumors were selected to have (a) monoallelic mutations in BRCA1 or BRCA2; or (b) no BRCA1 or BRCA2 mutations. The My Choice CDx product approved by the U.S. Food and Drug Administration (FDA) for companion diagnostics defines HRD positivity as the ability to screen out 95% of patients with BRCA functional loss, and the cutoff value that meets the conditions is set at 42.[3] . This embodiment corrects the HRD score and sets the threshold to 39, which ensures that 90.8% (59 / 65) of patients with BRCA dysfunction are screened out. Although the percentage of accurately screened patients is reduced, these patients are theoretically more genomically unstable and are expected to achieve better drug treatment effects. The higher the HRD value, the more unstable the genome, and it is speculated that the use of PARP inhibitors (PARPi) will have a better therapeutic effect.

[0208] This embodiment can detect that more than 90% of BRCA1 / 2 mutation samples are HRD-positive samples, and about 50% of BRCA1 / 2-negative samples are HRD-positive samples.

[0209] The test results of 151 samples are shown in the following table.

[0210] Table 9

[0211]

[0212]

[0213]

[0214]

[0215] This cohort was used to define HRD score thresholds designed to reflect HR-deficient versus HR-nondeficient states.

[0216] BRCA deficiency can be defined as meeting one of the following two conditions: (1) BRCA1 / 2 biallelic mutations; (2) LOH of the BRCA1 / 2 wild-type copies in BRCA1 / 2 mutant samples. The above-mentioned mutation information can be obtained by the detection method of the present invention.

[0217] BRCA is a gene associated with hereditary breast cancer, and the wild type refers to a genotype without mutations.

[0218] Example 2 HRD Accuracy Assessment

[0219] This example uses the method of Example 1 (abbreviated as WES detection method) for evaluation.

[0220] (1) Comparison of consistency with WGS testing

[0221] In this example, 50 ovarian cancer FFPE samples (Formalin-Fixed and Paraffin-Embedded, FFPE samples for short) were used to compare the results of the WES detection method in Example 1 with the HRD values ​​obtained by WGS detection. The results are as follows: Figure 2 As shown, Spearmanρ=0.88, showing good consistency.

[0222] (2) Comparison of consistency with standard products

[0223] This example tested 12 pairs of HRD standards and compared them with reference values ​​to investigate the accuracy of the test method. The results are as follows: Figure 3 It was shown that the HRD detection result of the detection method of this embodiment had a high consistency with the standard, Spearmanρ=0.87.

[0224] Example 3 HRD score and clinical efficacy

[0225] This example adopts the method of Example 1 to test 59 samples of high-grade serous ovarian cancer patients who were treated with platinum chemotherapy drugs for the first time, and statistically analyzes the relationship between the first platinum efficacy data and HRD, including 45 cases of platinum sensitivity data and 14 cases of platinum resistance data. The statistical results are shown in Figure 4 .from Figure 4 It can be seen that HRD-positive samples have higher PFS than HRD-negative samples (Log rank p=0.04, HR=0.55). PFS refers to progression-free survival, which specifically refers to the time from randomization to the first occurrence of disease progression or death from any cause.

[0226] Those skilled in the art will appreciate that all or part of the functions of the various methods in the above-mentioned embodiments can be implemented by hardware or by computer programs. When all or part of the functions in the above-mentioned embodiments are implemented by computer programs, the program can be stored in a computer-readable storage medium, and the storage medium can include: read-only memory, random access memory, disk, optical disk, hard disk, etc., and the program is executed by a computer to implement the above-mentioned functions. For example, the program is stored in the memory of the device, and when the program in the memory is executed by the processor, all or part of the above-mentioned functions can be implemented. In addition, when all or part of the functions in the above-mentioned embodiments are implemented by computer programs, the program can also be stored in a storage medium such as a server, another computer, disk, optical disk, flash disk or mobile hard disk, and can be downloaded or copied and saved in the memory of the local device, or the system of the local device is updated, and when the program in the memory is executed by the processor, all or part of the functions in the above-mentioned embodiments can be implemented.

[0227] References:

[0228] [1] Reid BM, Permuth JB, Sellers TA. Epidemiology of ovarian cancer:areview. Cancer Biol Med. 2017; 14:9-32.

[0229] [2] Chen Rui, Ji Yuan, Shen Peng, Yu Jinpu, Zhang Haiwei, Zhao Weipeng. Expert consensus on clinical detection and application of homologous recombination repair deficiency (2021 edition)[J]. Chinese Journal of Cancer Prevention and Treatment, 2021, 13(04): 329-338.

[0230] [3]Telli ML, Timms KM, Reid J, et al. Homologous recombinationdeficiency(HRD)score predicts response to platinum-containing neoadjuvantchemotherapy in patients with triple-negative breast cancer[J]. Clinicalcancer research, 2016, 22(15):3764-3773.

[0231] The above specific examples are used to illustrate the present invention, which is only used to help understand the present invention and is not intended to limit the present invention. For those skilled in the art, according to the concept of the present invention, some simple deductions, modifications or substitutions can be made.

Claims

1. A method for correcting homologous recombination repair defect scores, characterized in that: include: The correction step includes correcting the homologous recombination repair deficiency score according to the following formula to obtain a corrected homologous recombination repair deficiency score: HRD score 矫正 =LOH score+(1-α*WGD)*LST score+(1-β*WGD)*TAI scor e； HRD score 矫正 It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomere allele imbalance score, WGD refers to the whole genome duplication information value, α and β refer to the correction coefficients, wherein LOH score, LST score and TAI score are obtained according to the single nucleotide site variation set and gene copy number variation site set of the sequencing data of the sample to be tested and analyzed by scarHRD software; the sequencing data is the sequencing data generated based on the next generation sequencing, the copy number variation site set is obtained according to the sequencing data of the sample to be tested and the sequencing data of the control sample, the sample to be tested is a tumor tissue sample, and the control sample includes at least one of blood cells and adjacent cancer tissues.

2. The method according to claim 1, characterized in that α is 0.35 and β is 0.

35.

3. The method according to claim 1, characterized in that The whole genome replication information value is the whole genome replication information value output by the optimal model; The whole genome duplication information value is 0, 1, or 2.

4. The method according to claim 1, characterized in that The LOH score, LST score, and TAI score are calculated based on the single nucleotide site variation set and the gene copy number variation site set of the sequencing data of the sample to be tested.

5. The method according to claim 4, characterized in that The sample to be tested includes a human tumor tissue sample; The tumor includes a solid tumor; The tumors include ovarian cancer; The gene copy number variation site set of the sequencing data of the sample to be tested is obtained by analyzing the sequencing data of the sample to be tested and the sequencing data of the control sample; The control sample and the sample to be tested are derived from the same patient; The control sample includes at least one of blood cells and paracancerous tissues; The test sample and the control sample both contain DNA; The sequencing data is sequencing data generated based on next generation sequencing; The sequencing data is sequencing data aligned to a human reference genome sequence; The reference genome sequence is derived from a reference group; The reference genome sequence comprises a consensus sequence from a reference group; The reference genome sequence includes the hg19 human genome; The sequencing data includes capture sequencing data; The capture sequencing data includes at least one of whole exome sequencing data and regional capture sequencing data.

6. A method for evaluating homologous recombination repair deficiency, characterized in that include: The prediction step comprises predicting whether the patient to which the sample to be tested belongs has homologous recombination repair deficiency according to the corrected homologous recombination repair deficiency score obtained by the method according to any one of claims 1 to 5.

7. The method according to claim 6, characterized in that Predicting whether the patient to which the sample to be tested belongs has homologous recombination repair deficiency based on the relationship between the corrected homologous recombination repair deficiency score and the threshold; If the corrected homologous recombination repair defect score is ≥ the threshold, it is predicted that the patient to which the sample to be tested belongs is a homologous recombination repair defect-positive patient; If the corrected homologous recombination repair defect score is less than the threshold, it is predicted that the patient to which the sample to be tested belongs is a homologous recombination repair defect-negative patient; The homologous recombination repair deficiency-positive patients include patients with functional loss of breast cancer susceptibility genes; The patient with breast cancer susceptibility gene functional loss refers to a patient with breast cancer susceptibility gene containing biallelic mutations; The threshold is 39; In the prediction step, the clinical benefit of the patient receiving drug treatment is predicted based on whether the patient has homologous recombination repair deficiency; The drugs include PARP inhibitors.

8. A device for correcting homologous recombination repair defect scores, characterized in that: include: The correction module is used to correct the homologous recombination repair defect score according to the following formula to obtain a corrected homologous recombination repair defect score: HRD score 矫正 =LOH score+(1-α*WGD)*LST score+(1-β*WGD)*TAI scor e； HRD score 矫正 It refers to the corrected homologous recombination repair defect score, LOH score refers to the loss of heterozygosity score, LST score refers to the large fragment migration score, TAI score refers to the telomere allele imbalance score, WGD refers to the whole genome duplication information value, α and β refer to the correction coefficients, wherein LOH score, LST score and TAI score are obtained according to the single nucleotide site variation set and gene copy number variation site set of the sequencing data of the sample to be tested and analyzed by scarHRD software; the sequencing data is the sequencing data generated based on the next generation sequencing, the copy number variation site set is obtained according to the sequencing data of the sample to be tested and the sequencing data of the control sample, the sample to be tested is a tumor tissue sample, and the control sample includes at least one of blood cells and adjacent cancer tissues.

9. A device for evaluating homologous recombination repair deficiency, characterized in that include: A prediction module is used to predict whether the patient to which the sample to be tested belongs has homologous recombination repair deficiency according to the corrected homologous recombination repair deficiency score obtained by the method according to any one of claims 1 to 5 and / or the device according to claim 8.

10. A device for evaluating homologous recombination repair deficiency, characterized in that include: Memory, used to store programs; A processor, configured to implement the method according to any one of claims 1 to 7 by executing the program stored in the memory.

11. A computer-readable storage medium, characterized in that: The medium stores a program, and the program can be executed by a processor to implement the method according to any one of claims 1 to 7.

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