Gene combination for risk prediction of acute graft versus host disease and donor screening before hematopoietic cell transplantation and application thereof

Abstract

The invention discloses a gene combination for risk prediction of acute graft versus host disease and donor screening before hematopoietic cell transplantation and application of the gene combination. According to the present invention, the gene combination with the post-transplantation aGVHD occurrence prediction and diagnosis value is found through the whole genome sequencing and the target gene capture sequencing method; on one hand, comprehensive judgment and evaluation are carried out by detecting gene combination mutation conditions of donors and recipients, selection of transplantation donors is optimized, and the survival rate and the survival quality of patients are improved; on the other hand, the occurrence of aGVHD after transplantation of the recipient can be predicted by detecting the gene combination mutation condition of the donor and the recipient, and a basis is provided for potential aGVHD diagnosis; and thirdly, a new genetic basis is provided for revealing the pathogenesis of the aGVHD, and a new drug target is provided for treating the aGVHD.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine, and in particular to a gene combination and application thereof for risk prediction of acute graft-versus-host disease and donor screening before hematopoietic cell transplantation. Background Art

[0002] Allogeneic hematopoietic cell transplantation is still the effective or even the only cure for hematological malignancies. Acute graft-versus-host disease (aGVHD) is a common complication after hematopoietic cell transplantation and an important cause of transplant-related death. Currently, HLA matching between donors and recipients is the main screening factor for allogeneic hematopoietic cell transplantation. Among relatives, the probability of HLA matching between siblings is only 25%, while in the Chinese population, the probability of HLA matching between two unrelated people is about 1 / 10000-1 / 100000. The lack of donors limits the widespread clinical application of hematopoietic cell transplantation.

[0003] In recent years, with the establishment of an in vitro T-cell-removed HLA haplotype-matched transplantation protocol based on GCSF and anti-thymocyte globulin-induced immune tolerance by Peking University and the widespread application of other haplotype-matched transplantation protocols at home and abroad, the patient's parents, children, siblings, and collateral relatives have become potential candidate donors. When a patient has multiple haplotype donors, how to choose the most suitable donor is of great significance for avoiding the occurrence of aGVHD and improving the prognosis of transplantation. Summary of the invention

[0004] The purpose of the present invention is to provide a gene combination and application thereof for predicting the risk of acute graft-versus-host disease and screening donors before hematopoietic cell transplantation.

[0005] In order to achieve the above-mentioned object, the present invention first provides a new use of a substance for detecting whether a gene pair in a donor and a recipient has mutated.

[0006] The present invention provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in the preparation of a product for predicting or assisting in predicting the risk of acute graft-versus-host disease in a patient after hematopoietic cell transplantation.

[0007] The present invention further provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in predicting or assisting in predicting the risk of acute graft-versus-host disease in a patient after hematopoietic cell transplantation.

[0008] The present invention also provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in the preparation of a product for diagnosing or assisting in diagnosing whether an acute graft-versus-host disease occurs in a patient after hematopoietic cell transplantation.

[0009] The present invention also provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in diagnosing or assisting in diagnosing whether an acute graft-versus-host disease occurs in a patient after hematopoietic cell transplantation.

[0010] The present invention also provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in the preparation of a product for screening or assisting in screening a donor before hematopoietic cell transplantation.

[0011] The present invention also provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in screening or assisting in screening donors before hematopoietic cell transplantation.

[0012] The present invention also provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in preparing a product for pairing a donor and a recipient before hematopoietic cell transplantation (matching a donor and a recipient before hematopoietic cell transplantation).

[0013] The present invention also provides the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in pairing a donor and a recipient before hematopoietic cell transplantation (matching a donor and a recipient before hematopoietic cell transplantation).

[0014] In order to achieve the above object, the present invention also provides a kit having any one of the following functions a1)-a4):

[0015] a1) To predict or assist in predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation;

[0016] a2) To diagnose or assist in the diagnosis of acute graft-versus-host disease in patients after hematopoietic cell transplantation;

[0017] a3) Screening or assisting in the screening of donors before hematopoietic cell transplantation;

[0018] a4) Matching between donors and recipients before hematopoietic cell transplantation (matching donors and recipients before hematopoietic cell transplantation);

[0019] The kit includes substances for detecting whether a gene pair in the donor and the recipient has mutated;

[0020] In any of the above applications or kits, the gene pair is at least one of the following gene pairs: ASCC1-ANKRD20A2, ADAMTS7-APOL4, EFCAB5-ACTN2, EFCAB5-AGAP3, EFCAB5

[0021] -ATP6V1B1, EFCAB5-CARD8, EFCAB5-CNTNAP3B, EFCAB5-DEFB126, EFCAB5-DENND3, EFCAB5-FRMD5, EFCAB5-HAPLN3, EFCAB5-HRNR, EFCAB5-KRT24, EFCAB5-KRTAP10-1, EFCAB5-LCN10, EFCAB5 -MAGEE2, EFCAB5-MUC16, EFCAB5-PABPC3, EFCAB5-SETBP1, EFCAB5-SLC35G6, EFCAB5-TLR8, EF CAB5-ZFPM1, EFCAB5-ZNF211, MYO15B-TMX3, TMEM247-CPNE1, ZNF880-ANAPC1, ZNF211-GIGYF2.

[0022] The two genes before and after the "-" in each of the above gene pairs correspond to the donor and the recipient, respectively, that is, the gene before the "-" in each of the above gene pairs is recorded as the first gene, and the gene after the "-" is recorded as the second gene. The detection of whether the gene pair in the donor and the recipient is mutated refers to detecting whether the first gene in the donor and the second gene in the recipient are mutated.

[0023] In any of the above applications or kits, the substance for detecting whether a gene pair in the donor and the recipient has mutated is a substance for detecting whether a gene pair in the peripheral blood of the donor and the recipient has mutated.

[0024] Furthermore, the substance for detecting whether a gene pair in the peripheral blood of the donor and the recipient has mutated may be a reagent and / or an instrument for detecting whether a gene pair in the peripheral blood of the donor and the recipient has mutated.

[0025] The reagents and / or instruments include but are not limited to reagents for extracting genomic DNA from human peripheral blood cells, and reagents and / or instruments required for detecting whether gene pairs in peripheral blood are mutated using various conventional methods in the prior art (such as DNA sequencing, restriction fragment length polymorphism, single-stranded conformation polymorphism, allele-specific oligonucleotide hybridization, etc.).

[0026] Furthermore, the reagents and / or instruments required for detecting whether a gene pair in peripheral blood is mutated using DNA sequencing (such as high-throughput sequencing or first-generation sequencing, etc.) include various primers, probes and sequencers used in sequencing.

[0027] In a specific embodiment of the present invention, the DNA sequencing method may specifically include the following steps:

[0028] 1. Draw venous blood from donors and recipients before hematopoietic cell transplantation and extract genomic DNA;

[0029] 2. Use the enzyme digestion method to randomly fragment 200ng genomic DNA into an average length of 150-200bp, then repair and fill the ends and add A bases at the 3' end to form sticky ends, then connect with the adapter containing the specific barcode sequence, and then use magnetic beads to screen to remove incomplete connection products and adapter self-connection products, and finally use universal primers complementary to the adapter sequence for PCR amplification to form a sequencing library;

[0030] 3. Use The pool probes were hybridized in the hybridization buffer, and then the hybridized target fragments were captured using magnetic beads and separated and purified. The captured DNA fragments were amplified by PCR and the quality of the captured library was checked (including Qubit concentration determination and 2100 Bioanalyzer fragment length determination). Finally, PE150 high-throughput sequencing was performed on the NovaSeq6000 sequencer according to Illumina's standard process.

[0031] 4. Use sequencing profiles for genotyping.

[0032] In any of the above applications or kits, the kit also includes a data processing device, and the data processing device has a built-in module, which has the following functions: predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation or diagnosing whether acute graft-versus-host disease occurs in patients after hematopoietic cell transplantation based on whether the above gene pairs in the peripheral blood of donors and recipients are mutated.

[0033] In practical applications, any one of the above gene pairs can be used to predict the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation, diagnose whether acute graft-versus-host disease occurs in patients after hematopoietic cell transplantation, or screen donors before hematopoietic cell transplantation.

[0034] Specifically,

[0035] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the ASCC1-ANKRD20A2 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation with mutations in the donor ASCC1 gene and mutations in the recipient ANKRD20A2 gene is higher than that in patients after hematopoietic cell transplantation in which at least one of the donor ASCC1 gene and the recipient ANKRD20A2 gene is not mutated.

[0036] The method for diagnosing whether acute graft-versus-host disease occurs in a patient after hematopoietic cell transplantation based on the ASCC1-ANKRD20A2 gene pair in the donor and the recipient is as follows: if the donor ASCC1 gene is mutated and the recipient ANKRD20A2 gene is mutated, the patient has developed or is a candidate to develop acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor ASCC1 gene and the recipient ANKRD20A2 gene is not mutated, the patient has not developed or is a candidate to develop acute graft-versus-host disease after hematopoietic cell transplantation.

[0037] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the ADAMTS7-APOL4 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor ADAMTS7 gene is mutated and the recipient APOL4 gene is mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor ADAMTS7 gene and the recipient APOL4 gene is not mutated.

[0038] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the ADAMTS7-APOL4 gene pair in the donor and the recipient is as follows: if the donor's ADAMTS7 gene is mutated and the recipient's APOL4 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's ADAMTS7 gene and the recipient's APOL4 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0039] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-ACTN2 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient ACTN2 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient ACTN2 gene is not mutated.

[0040] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-ACTN2 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's ACTN2 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's ACTN2 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0041] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-AGAP3 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient AGAP3 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient AGAP3 gene is not mutated.

[0042] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-AGAP3 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's AGAP3 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's AGAP3 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0043] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-ATP6V1B1 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient ATP6V1B1 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient ATP6V1B1 gene is not mutated.

[0044] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-ATP6V1B1 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's ATP6V1B1 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's ATP6V1B1 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0045] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-CARD8 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient CARD8 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient CARD8 gene is not mutated.

[0046] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-CARD8 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's CARD8 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's CARD8 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0047] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-CNTNAP3B gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient CNTNAP3B gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient CNTNAP3B gene is not mutated.

[0048] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-CNTNAP3B gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's CNTNAP3B gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's CNTNAP3B gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0049] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-DEFB126 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient DEFB126 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient DEFB126 gene is not mutated.

[0050] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-DEFB126 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's DEFB126 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's DEFB126 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0051] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-DENND3 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient DENND3 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient DENND3 gene is not mutated.

[0052] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-DENND3 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's DENND3 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's DENND3 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0053] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-FRMD5 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient FRMD5 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient FRMD5 gene is not mutated.

[0054] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-FRMD5 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's FRMD5 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's FRMD5 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0055] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-HAPLN3 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient HAPLN3 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient HAPLN3 gene is not mutated.

[0056] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-HAPLN3 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's HAPLN3 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's HAPLN3 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0057] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-HRNR gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient HRNR gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient HRNR gene is not mutated.

[0058] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-HRNR gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's HRNR gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's HRNR gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0059] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-KRT24 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient KRT24 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient KRT24 gene is not mutated.

[0060] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-KRT24 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's KRT24 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's KRT24 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0061] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-KRTAP10-1 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient KRTAP10-1 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient KRTAP10-1 gene is not mutated.

[0062] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-KRTAP10-1 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's KRTAP10-1 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's KRTAP10-1 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0063] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-LCN10 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient LCN10 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient LCN10 gene is not mutated.

[0064] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-LCN10 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's LCN10 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's LCN10 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0065] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-MAGEE2 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient MAGEE2 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient MAGEE2 gene is not mutated.

[0066] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-MAGEE2 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's MAGEE2 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's MAGEE2 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0067] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-MUC16 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation with a mutation in the donor EFCAB5 gene and a mutation in the recipient MUC16 gene is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient MUC16 gene is not mutated.

[0068] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-MUC16 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's MUC16 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's MUC16 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0069] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-PABPC3 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient PABPC3 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient PABPC3 gene is not mutated.

[0070] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-PABPC3 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's PABPC3 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's PABPC3 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0071] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-SETBP1 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient SETBP1 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient SETBP1 gene is not mutated.

[0072] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-SETBP1 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's SETBP1 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's SETBP1 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0073] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-SLC35G6 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient SLC35G6 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient SLC35G6 gene is not mutated.

[0074] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-SLC35G6 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's SLC35G6 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's SLC35G6 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0075] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-TLR8 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation with a mutation in the donor EFCAB5 gene and a mutation in the recipient TLR8 gene is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient TLR8 gene is not mutated.

[0076] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-TLR8 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's TLR8 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's TLR8 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0077] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-ZFPM1 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient ZFPM1 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient ZFPM1 gene is not mutated.

[0078] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-ZFPM1 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's ZFPM1 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's ZFPM1 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0079] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the EFCAB5-ZNF211 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor EFCAB5 gene and the recipient ZNF211 gene are mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor EFCAB5 gene and the recipient ZNF211 gene is not mutated.

[0080] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the EFCAB5-ZNF211 gene pair in the donor and the recipient is as follows: if the donor's EFCAB5 gene is mutated and the recipient's ZNF211 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's EFCAB5 gene and the recipient's ZNF211 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0081] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the MYO15B-TMX3 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation with a mutation in the donor MYO15B gene and a mutation in the recipient TMX3 gene is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor MYO15B gene and the recipient TMX3 gene is not mutated.

[0082] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the MYO15B-TMX3 gene pair in the donor and the recipient is as follows: if the donor's MYO15B gene is mutated and the recipient's TMX3 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's MYO15B gene and the recipient's TMX3 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0083] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the TMEM247-CPNE1 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation in which both the donor TMEM247 gene is mutated and the recipient CPNE1 gene is mutated is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor TMEM247 gene and the recipient CPNE1 gene is not mutated.

[0084] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the TMEM247-CPNE1 gene pair in the donor and the recipient is as follows: if the donor's TMEM247 gene is mutated and the recipient's CPNE1 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's TMEM247 gene and the recipient's CPNE1 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0085] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the ZNF880-ANAPC1 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation with both the donor ZNF880 gene mutation and the recipient ANAPC1 gene mutation is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor ZNF880 gene and the recipient ANAPC1 gene is not mutated.

[0086] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the ZNF880-ANAPC1 gene pair in the donor and the recipient is as follows: if the donor's ZNF880 gene is mutated and the recipient's ANAPC1 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's ZNF880 gene and the recipient's ANAPC1 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0087] The method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the ZNF211-GIGYF2 gene pair is mutated in the donor and the recipient is as follows: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation with a mutation in the donor ZNF211 gene and a mutation in the recipient GIGYF2 gene is lower than that in patients after hematopoietic cell transplantation in which at least one of the donor ZNF211 gene and the recipient GIGYF2 gene is not mutated.

[0088] The method for diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation based on the ZNF211-GIGYF2 gene pair in the donor and the recipient is as follows: if the donor's ZNF211 gene is mutated and the recipient's GIGYF2 gene is mutated, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if at least one of the donor's ZNF211 gene and the recipient's GIGYF2 gene is not mutated, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation.

[0089] In practical applications, any two or three or more gene pairs (including the 27 gene pairs) among the above 27 gene pairs can also be used to predict the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation or to diagnose whether acute graft-versus-host disease occurs in patients after hematopoietic cell transplantation. When multiple gene pairs are used for joint prediction or diagnosis, each gene pair needs to meet its own judgment criteria for prediction or diagnosis, otherwise the prediction or diagnosis method is invalid.

[0090] When the 27 gene pairs are used together for prediction or diagnosis, in addition to referring to the judgment criteria of each gene pair mentioned above for prediction or diagnosis, the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation can be predicted according to the following method 1 or method 2, and whether acute graft-versus-host disease will occur in patients after hematopoietic cell transplantation can be diagnosed according to method 3 or method 4.

[0091] Method 1: When the donor ASCC1 gene is mutated and the recipient ANKRD20A2 gene is mutated, the above “method for predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation based on whether the ASCC1-ANKRD20A2 gene pair in the donor and the recipient is mutated” is used for judgment; when at least one of the donor ASCC1 gene and the recipient ANKRD20A2 gene is not mutated, the other 26 gene pairs are used for judgment: if at least one of the other 26 gene pairs in the donor and the recipient is mutated (a gene pair mutation means that both the gene corresponding to the donor (i.e., the first gene in the above text) and the gene corresponding to the recipient (i.e., the second gene in the above text) in the gene pair are mutated), the risk of acute graft-versus-host disease in the hematopoietic cell transplantation patient is lower than that in the other 26 gene pairs in the donor and the recipient where none of the mutations occurs (no mutation means that at least one of the gene corresponding to the donor and the gene corresponding to the recipient in each gene pair is not mutated).

[0092] Method 2: First, based on the gene mutation status of the above 27 gene pairs in the donor and the recipient, the total number of gene mutations is obtained, and then the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation is predicted based on the total number of gene mutations and the threshold: the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation whose total number of gene mutations is greater than or equal to the threshold is lower than that in patients after hematopoietic cell transplantation whose total number of gene mutations is less than the threshold; the total number of gene mutations is greater than or equal to 5.

[0093] Method 3: When the donor ASCC1 gene is mutated and the recipient ANKRD20A2 gene is mutated, the diagnosis is made according to the above "method for diagnosing whether acute graft-versus-host disease occurs in patients after hematopoietic cell transplantation based on whether the ASCC1-ANKRD20A2 gene pair in the donor and the recipient is mutated"; when at least one of the donor ASCC1 gene and the recipient ANKRD20A2 gene is not mutated, the diagnosis is made according to the mutation status of the other 26 gene pairs: if the donor and the recipient have at least one gene in the other 26 gene pairs If a gene pair is mutated (a gene pair mutation means that both the gene corresponding to the donor (i.e., the first gene mentioned above) and the gene corresponding to the recipient (i.e., the second gene mentioned above) in the gene pair are mutated), the patient will not develop or will not be likely to develop acute graft-versus-host disease after hematopoietic cell transplantation; if the other 26 gene pairs in the donor and the recipient are not mutated (no mutation means that at least one of the gene corresponding to the donor and the gene corresponding to the recipient in each gene pair is not mutated), the patient will develop or will be likely to develop acute graft-versus-host disease after hematopoietic cell transplantation.

[0094] Method 4: First, according to the gene mutation status of the above 27 gene pairs in the donor and the recipient, the total number of gene mutations is obtained, and then the patient after hematopoietic cell transplantation develops acute graft-versus-host disease according to the total number of gene mutations and the threshold value is diagnosed: if the total number of gene mutations is greater than or equal to the threshold value, the patient does not develop or is a candidate for not developing acute graft-versus-host disease after hematopoietic cell transplantation; if the total number of gene mutations is less than the threshold value, the patient develops or is a candidate for developing acute graft-versus-host disease after hematopoietic cell transplantation; the total number of gene mutations is greater than or equal to 5.

[0095] The method for obtaining the total number of gene mutations in any of the above genes is as follows: if ASCC1-ANKRD20A2, ADAMTS7-APOL4, EFCAB5-ACTN2, EFCAB5-AGAP3, EFCAB5-ATP6V1B1, EFCAB5-CARD8, EFCAB5-CNTNAP3B, EFCAB5-DEFB126, EFCAB5-DENND3, EFCAB5-FRMD5, EFCAB5-HAPLN3, EFCAB5-HRNR, EFCAB5-KRT24, EFCAB5-KRTAP10-1, EFCAB5-LCN10, EFCAB5-MA Mutation of the GEE2, EFCAB5-MUC16, EFCAB5-PABPC3, EFCAB5-SETBP1, EFCAB5-SLC35G6, EFCAB5-TLR8, EFCAB5-ZFPM1, EFCAB5-ZNF211, MYO15B-TMX3, TMEM247-CPNE1, ZNF880-ANAPC1 or ZNF211-GIGYF2 gene pair (mutation of a gene pair means that both the gene corresponding to the donor (i.e., the first gene mentioned above) and the gene corresponding to the recipient (i.e., the second gene mentioned above) in the gene pair are mutated) was recorded as 1, and the mutation scores of the 27 gene pairs were accumulated to obtain the total number of gene mutations.

[0096] Mutation in any of the above gene pairs means that both the gene corresponding to the donor (i.e. the first gene before the “-” in the above text) and the gene corresponding to the recipient (i.e. the second gene after the “-” in the above text) in the gene pair are mutated.

[0097] Mutations in any of the above genes include any base mutations that can change the amino acid sequence of the protein encoded by the gene. The base mutations include base insertions, base deletions, base substitutions, etc. The number of base mutations includes 1 or 2 or more base mutations.

[0098] The fact that any of the above genes have not undergone mutations includes the fact that there are no base mutations (completely identical to the wild-type sequence of the gene) or any base mutations that do not change the amino acid sequence of the protein encoded by the gene.

[0099] Any of the above thresholds may specifically be 17.5.

[0100] In practical applications, the risk of acute graft-versus-host disease after hematopoietic cell transplantation can be first predicted according to the above method, and then the donors before hematopoietic cell transplantation can be screened according to the risk of acute graft-versus-host disease, or other clinical indicators can be combined to assist in the screening of donors before hematopoietic cell transplantation, so as to optimize the selection of donors before hematopoietic cell transplantation and improve the survival rate and quality of life of patients before hematopoietic cell transplantation.

[0101] In any of the above applications or kits, the donor is a healthy individual who provides hematopoietic cells before hematopoietic cell transplantation, and the recipient is a patient before hematopoietic cell transplantation.

[0102] In any of the above uses or kits, the hematopoietic cell transplantation is allogeneic hematopoietic cell transplantation.

[0103] The allogeneic hematopoietic cell transplantation is HLA haplotype-matched allogeneic hematopoietic cell transplantation.

[0104] The patient before hematopoietic cell transplantation may be an acute myeloid leukemia patient, an acute lymphoid leukemia patient, a chronic myeloid leukemia patient, a chronic lymphoid leukemia patient, a multiple myeloma patient, a myelodysplastic syndrome patient or a primary systemic amyloidosis patient.

[0105] The use of ASCC1, ANKRD20A2, ADAMTS7, APOL4, ACTN2, AGAP3, ATP6V1B1, CARD8, CNTNAP3B, DEFB126, DENND3, FRMD5, HAPLN3, HRNR, KRT24; KRTAP10-1, LCN10, MAGEE2, MUC16, PABPC3, SETBP1, SLC35G6, EFCAB5, TLR8, ZFPM1, EFCAB5, ZNF211, MYO15B, TMX3, TMEM247, CPNE1, ZNF880, ANAPC1, ZNF211 and / or GIGYF2 as targets in the preparation of products for preventing and / or treating acute graft-versus-host disease also falls within the scope of protection of the present invention.

[0106] The NCBI IDs of any of the above ASCC1, ANKRD20A2, ADAMTS7, APOL4, ACTN2, AGAP3, ATP6V1B1, CARD8, CNTNAP3B, DEFB126, DENND3, FRMD5, HAPLN3, HRNR, KRT24; KRTAP10-1, LCN10, MAGEE2, MUC16, PABPC3, SETBP1, SLC35G6, EFCAB5, TLR8, ZFPM1, EFCAB5, ZNF211, MYO15B, TMX3, TMEM247, CPNE1, ZNF880, ANAPC1, ZNF211 and GIGYF2 are shown in Table 1 below.

[0107] The present invention has discovered gene combinations with predictive and diagnostic value for the occurrence of aGVHD after transplantation through whole genome sequencing and target gene capture sequencing. On the one hand, by detecting the gene combination mutations of donors and recipients, comprehensive judgment and evaluation can be made to optimize the selection of transplant donors and improve the survival rate and quality of life of patients; on the other hand, by detecting the gene combination mutations of donors and recipients, the risk of aGVHD after transplantation in recipients can be predicted, providing a basis for potential aGVHD diagnosis; thirdly, the present invention provides a new genetic basis for revealing the pathogenesis of aGVHD and a new drug target for the treatment of aGVHD. BRIEF DESCRIPTION OF THE DRAWINGS

[0108] Figure 1 This is the result of analyzing the significant genes together with HLA-A, HLA-B, HLA-DRB1, and the total number of HLA-matching loci.

[0109] Figure 2 Figure 3 shows the distribution of the total number of LoF gene pair mutations in the aGVHD-positive group and the aGVHD-negative group.

[0110] Figure 3 Figure 3 is the ROC curve of the total number of LoF gene pair mutations for diagnosing aGVHD. DETAILED DESCRIPTION

[0111] The present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.

[0112] The experimental methods in the following examples, unless otherwise specified, are all conventional methods, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples, unless otherwise specified, can all be obtained from commercial channels.

[0113] The HLA haplotype matching involved in the present invention means that the HLA-A, HLA-B, and HLA-DRB1 alleles between the related donor and recipient are the same on the same chromosome, but at least one of the above alleles on another chromosome is different (i.e. 3 / 6, 4 / 6, or 5 / 6 matching).

[0114] Example 1. Obtaining LoF mutation gene pairs associated with aGVHD

[0115] 1. Research subjects

[0116] The research subjects of this example are as follows: peripheral blood samples of 106 patients with aGVHD and their donors (defined as the aGVHD positive group), and peripheral blood samples of 106 patients without aGVHD and their donors (defined as the aGVHD negative group).

[0117] Inclusion criteria: 1) Patients underwent HLA haplotype-matched allogeneic hematopoietic cell transplantation at Peking University People's Hospital from 2018 to 2019; 2) Patients had malignant blood diseases before transplantation, including acute myeloid leukemia, acute lymphoid leukemia, chronic myeloid leukemia, chronic lymphoid leukemia, multiple myeloma, myelodysplastic syndrome, and primary systemic amyloidosis; 3) Patients and donors were aged between 14 and 65 years old; 4) The aGVHD-positive group included patients diagnosed with aGVHD 2-4 within 100 days after transplantation; the aGVHD-negative group included patients who did not develop aGVHD symptoms within 100 days after transplantation and were paired 1:1 with the aGVHD-positive group. There was no statistical difference in age and gender between the two groups.

[0118] Exclusion criteria: 1) The patient's disease before transplantation was not acute promyelocytic leukemia or aplastic anemia; 2) This was not the first hematopoietic cell transplantation for the patient; 3) The patient died within three months after transplantation and the cause of death was not diagnosed as aGVHD.

[0119] 2. Sample preparation

[0120] The genomic DNA was extracted from the collected peripheral blood samples using the phenol-chloroform method to test the purity, concentration, integrity and contamination of the DNA. The qualified samples were sequenced for genome sequencing, and the experimental process was carried out according to the library preparation steps and sequencing steps provided by Illumina.

[0121] 3. Next-generation sequencing

[0122] The library was constructed using the human whole exome sequence capture kit Truseq Nano, and hybridized with probes for capture and enrichment. The samples were then sequenced using the Illumina Novaseq S4 PE150 high-throughput sequencing platform, with an average sequencing depth of 35x for each sample.

[0123] 4. Analysis of sequencing results

[0124] FastQC software was used to evaluate the quality of the output raw data, and BWA (parameter-M) software was used to align the measured data with the reference genome. All analyses were performed based on the human reference genome hg38. The mutations that affect protein structure annotated by ANNOVAR were defined as protein loss-of-function mutations (LOF mutations) in this study. The ANNOVAR annotations included "stopgain", "startloss", "frameshift insertion", "frameshift deletion" and "frameshift block substitution". When LoF mutations occur in two genes in a pair of donors and recipients (such as mutations in gene A in the donor and mutations in gene B in the recipient), the donor A gene and the recipient B gene are defined as a LoF mutation gene pair.

[0125] 5. Analysis results

[0126] Through analysis and screening, 1150 LoF mutation gene pairs related to aGVHD were obtained. To further exclude the influence of HLA, the significant genes were analyzed together with HLA-A, HLA-B, HLA-DRB1, and the total number of HLA-matched loci, and it was found that all 1150 LoF mutation gene pairs were significantly associated with aGVHD ( Figure 1 ).

[0127] Example 2: Screening of LoF mutation gene pairs associated with aGVHD

[0128] 1. Research subjects

[0129] The research subjects of this example are as follows: peripheral blood samples of 247 patients with aGVHD and their donors (defined as the aGVHD positive group), and peripheral blood samples of 224 patients without aGVHD and their donors (defined as the aGVHD negative group).

[0130] Inclusion criteria: 1) Patients underwent HLA haplotype-matched allogeneic hematopoietic cell transplantation at Peking University People's Hospital from 2019 to June 2020; 2) Patients had malignant blood diseases before transplantation, including acute myeloid leukemia, acute lymphoid leukemia, chronic myeloid leukemia, chronic lymphoid leukemia, multiple myeloma, myelodysplastic syndrome, and primary systemic amyloidosis; 3) Both patients and donors were between 14 and 65 years old.

[0131] Exclusion criteria: 1) The patient's disease before transplantation was not acute promyelocytic leukemia or aplastic anemia; 2) This was not the first hematopoietic cell transplantation for the patient; 3) The patient died within three months after transplantation and the cause of death was not diagnosed as aGVHD.

[0132] 2. Sample preparation

[0133] The peripheral blood samples collected were used to extract genomic DNA and test the purity, concentration, integrity and contamination of the DNA. The qualified samples were then tested for the next step.

[0134] 3. Gene Selection

[0135] The top 100 LOF mutation gene pairs in the whole genome sequencing were selected for probe verification.

[0136] 4. Probe Sequencing

[0137] 200 ng of genomic DNA was randomly fragmented into 150-200 bp average length using enzyme digestion, and then end-repaired and flattened with an A base added to the 3' end to form a sticky end. Then, it was connected to a connector containing a specific barcode sequence. Magnetic bead screening was used to remove incomplete connection products and connector self-connection products. PCR amplification was performed using universal primers complementary to the connector sequence to form a sequencing library. Then, the library was used to generate a sequencing library. The pool probes were hybridized in the hybridization buffer, and the hybridized target fragments were captured using magnetic beads and separated and purified. The captured DNA fragments were amplified by PCR and the PCR products were purified. The quality control of the captured library was then performed, including Qubit concentration determination and 2100 Bioanalyzer fragment length determination. Finally, PE150 high-throughput sequencing was performed on the NovaSeq6000 sequencer according to Illumina's standard process.

[0138] 5. Results Analysis

[0139] FastQC software was used to evaluate the quality of the raw data, and BWA (parameter-M) software was used for genome alignment. All analyses were performed based on the human reference genome hg38. Univariate analysis was performed on the donor-recipient LOF mutation gene pairs to obtain the donor-recipient LOF mutation gene pairs that were still associated with aGVHD.

[0140] The results are shown in Table 1. There were 27 donor-recipient LOF mutation gene pairs that were still significantly different between the two aGVHD groups.

[0141] Table 1

[0142]

[0143]

[0144] The specific mutation status of each LoF mutation gene pair in 471 donor-recipient pairs is as follows:

[0145] There were 17 cases of ASCC1-ANKRD20A2 gene mutation in 1,471 donor-recipient pairs, including 13 cases in the aGVHD-positive group and 4 cases in the aGVHD-negative group. The aGVHD grouping and gene pair mutation are shown in Table 2. After chi-square test analysis, P=0.049, there was a statistically significant difference between the two groups.

[0146] Table 2

[0147]

[0148] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0149] 2. Among the 471 donor-recipient pairs, 114 cases had ADAMTS7-APOL4 gene mutations, including 49 cases in the aGVHD-positive group and 65 cases in the aGVHD-negative group. The aGVHD grouping and gene pair mutations are shown in Table 3. After chi-square test analysis, P=0.021, there was a statistically significant difference between the two groups.

[0150] Table 3

[0151]

[0152] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0153] There were 27 cases of EFCAB5-ACTN2 gene mutation in 3.471 donor-recipient pairs, including 9 cases in the aGVHD-positive group and 18 cases in the aGVHD-negative group. The aGVHD grouping and gene pair mutation are shown in Table 4. After chi-square test analysis, P=0.042, there was a statistically significant difference between the two groups.

[0154] Table 4

[0155]

[0156] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0157] Among the 471 donor-recipient pairs, 31 cases had EFCAB5-AGAP3 gene mutation, including 11 cases in the aGVHD-positive group and 20 cases in the aGVHD-negative group. The aGVHD grouping and gene pair mutation status are shown in Table 5. After chi-square test analysis, P=0.050, there was a statistically significant difference between the two groups.

[0158] Table 5

[0159]

[0160] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0161] There were 21 cases of EFCAB5-ATP6V1B1 gene mutation in 5.471 donor-recipient pairs, including 6 cases in the aGVHD-positive group and 15 cases in the aGVHD-negative group. The aGVHD grouping and gene pair mutation are shown in Table 6. After chi-square test analysis, P=0.025, there was a statistically significant difference between the two groups.

[0162] Table 6

[0163]

[0164] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0165] There were 28 cases of EFCAB5-CARD8 gene mutation in 6.471 donor-recipient pairs, including 9 cases in the aGVHD-positive group and 19 cases in the aGVHD-negative group. The aGVHD grouping and gene pair mutation are shown in Table 7. After chi-square test analysis, P=0.027, there was a statistically significant difference between the two groups.

[0166] Table 7

[0167]

[0168] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0169] There were 13 cases of EFCAB5-CNTNAP3B gene mutation in 7.471 donor-recipient pairs, including 2 cases in the aGVHD-positive group and 11 cases in the aGVHD-negative group. The aGVHD grouping and gene pair mutation are shown in Table 8. After chi-square test analysis, P=0.007, indicating a statistically significant difference between the two groups.

[0170] Table 8

[0171]

[0172] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0173] There were 28 cases of EFCAB5-DEFB126 gene mutation in 8,471 donor-recipient pairs, including 9 cases in the aGVHD-positive group and 19 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 9. After chi-square test analysis, P=0.027, there was a statistically significant difference between the two groups.

[0174] Table 9

[0175]

[0176] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0177] Among the 9,471 donor-recipient pairs, 34 cases had EFCAB5-DENND3 gene mutations, including 12 cases in the aGVHD-positive group and 22 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutations are shown in Table 10. After chi-square test analysis, P = 0.038, indicating a statistically significant difference between the two groups.

[0178] Table 10

[0179]

[0180] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0181] There were 17 cases of EFCAB5-FRMD5 gene mutation in 10,471 donor-recipient pairs, including 4 cases in the aGVHD-positive group and 13 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 11. After chi-square test analysis, P = 0.015, there was a statistically significant difference between the two groups.

[0182] Table 11

[0183]

[0184] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0185] There were 23 cases of EFCAB5-HAPLN3 gene mutation in 11,471 donor-recipient pairs, including 6 cases in the aGVHD-positive group and 17 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 12. After chi-square test analysis, P = 0.009, there was a statistically significant difference between the two groups.

[0186] Table 12

[0187]

[0188] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0189] There were 34 cases of EFCAB5-HRNR gene mutation in 12,471 donor-recipient pairs, including 12 cases in the aGVHD-positive group and 22 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 13. After chi-square test analysis, P = 0.038, indicating a statistically significant difference between the two groups.

[0190] Table 13

[0191]

[0192] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0193] Among the 13,471 donor-recipient pairs, 31 cases had EFCAB5-KRT24 gene mutation, including 11 cases in the aGVHD-positive group and 20 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation status are shown in Table 14. After chi-square test analysis, P = 0.050, there was a statistically significant difference between the two groups.

[0194] Table 14

[0195]

[0196] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0197] There were 23 cases of EFCAB5-KRTAP10-1 gene mutation in 14,471 donor-recipient pairs, including 7 cases in the aGVHD-positive group and 16 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 15. After chi-square test analysis, P = 0.030, indicating a statistically significant difference between the two groups.

[0198] Table 15

[0199]

[0200] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0201] There were 25 cases of EFCAB5-LCN10 gene mutation in 15,471 donor-recipient pairs, including 8 cases in the aGVHD-positive group and 17 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation status are shown in Table 16. After chi-square test analysis, P=0.035, there was a statistically significant difference between the two groups.

[0202] Table 16

[0203]

[0204] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0205] Among the 16,471 donor-recipient pairs, 30 cases had EFCAB5-MAGEE2 gene mutations, including 9 cases in the aGVHD-positive group and 21 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation status are shown in Table 17. After chi-square test analysis, P = 0.011, and there was a statistically significant difference between the two groups.

[0206] Table 17

[0207]

[0208] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0209] There were 34 cases of EFCAB5-MUC16 gene mutation in 17,471 donor-recipient pairs, including 22 cases in the aGVHD-positive group and 12 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 18. After chi-square test analysis, P = 0.038, there was a statistically significant difference between the two groups.

[0210] Table 18

[0211]

[0212] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0213] Among the 18,471 donor-recipient pairs, 31 cases had EFCAB5-PABPC3 gene mutations, including 11 cases in the aGVHD-positive group and 20 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation status are shown in Table 19. After chi-square test analysis, P = 0.050, there was a statistically significant difference between the two groups.

[0214] Table 19

[0215]

[0216] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0217] There were 28 cases of EFCAB5-SETBP1 gene mutation in 19,471 donor-recipient pairs, including 9 cases in the aGVHD-positive group and 19 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 20. After chi-square test analysis, P = 0.027, and there was a statistically significant difference between the two groups.

[0218] Table 20

[0219]

[0220] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0221] There were 34 cases of EFCAB5-SLC35G6 gene mutation in 20,471 donor-recipient pairs, including 12 cases in the aGVHD-positive group and 22 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 21. After chi-square test analysis, P = 0.038, there was a statistically significant difference between the two groups.

[0222] Table 21

[0223]

[0224] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0225] There were 34 cases of EFCAB5-TLR8 gene mutation in 21,471 donor-recipient pairs, including 12 cases in the aGVHD-positive group and 22 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 22. After chi-square test analysis, P = 0.038, there was a statistically significant difference between the two groups.

[0226] Table 22

[0227]

[0228] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0229] There were 34 cases of EFCAB5-ZFPM1 gene mutation in 22,471 donor-recipient pairs, including 12 cases in the aGVHD-positive group and 22 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation status are shown in Table 23. After chi-square test analysis, P=0.038, there was a statistically significant difference between the two groups.

[0230] Table 23

[0231]

[0232] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0233] There were 29 cases of EFCAB5-ZNF211 gene mutation in 23,471 donor-recipient pairs, including 9 cases in the aGVHD-positive group and 20 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation status are shown in Table 24. After chi-square test analysis, P=0.017, there was a statistically significant difference between the two groups.

[0234] Table 24

[0235]

[0236] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0237] There were 25 cases of MYO15B-TMX3 gene mutation in 24,471 donor-recipient pairs, including 8 cases in the aGVHD-positive group and 17 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 25. After chi-square test analysis, P = 0.035, there was a statistically significant difference between the two groups.

[0238] Table 25

[0239]

[0240] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0241] There were 4 cases of TMEM247-CPNE1 gene mutation in 25.471 donor-recipient pairs, all of which were in the aGVHD negative group (17 cases); the aGVHD grouping and gene pair mutation status are shown in Table 26. After chi-square test analysis, P = 0.035, there was a statistically significant difference between the two groups.

[0242] Table 26

[0243]

[0244] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0245] There were 157 cases of ZNF880-ANAPC1 gene mutation in 26,471 donor-recipient pairs, including 71 cases in the aGVHD-positive group and 86 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation status are shown in Table 27. After chi-square test analysis, P=0.027, there was a statistically significant difference between the two groups.

[0246] Table 27

[0247]

[0248] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0249] There were 20 cases of ZNF211-GIGYF2 gene mutation in 27,471 donor-recipient pairs, including 6 cases in the aGVHD-positive group and 14 cases in the aGVHD-negative group; the aGVHD grouping and gene pair mutation are shown in Table 28. After chi-square test analysis, P=0.040, there was a statistically significant difference between the two groups.

[0250] Table 28

[0251]

[0252] Note: No mutation in the table means that at least one of the two genes in the gene pair has not mutated, and mutation means that both genes in the gene pair have mutated.

[0253] Example 3: Application of 27 LOF mutation gene pairs in predicting the risk of aGVHD in hematopoietic cell transplantation patients

[0254] 1. Application of mutations in 27 LOF mutation gene pairs in predicting the risk of aGVHD in hematopoietic cell transplantation patients

[0255] The 417 patients and their donors in Example 2 were used as research subjects, and the correlation between the occurrence of 27 LOF mutation gene pairs and the occurrence of aGVHD was statistically analyzed.

[0256] The results showed that among the 471 patients, 281 had any mutation in the 27 LOF mutation gene pairs, and 149 of the 281 patients with mutations in the LOF mutation gene pairs developed aGVHD after hematopoietic cell transplantation (Table 29). After chi-square analysis, P = 0.004. Therefore, in practical applications, the risk of aGVHD in patients after hematopoietic cell transplantation can be predicted based on whether the 27 LOF mutation gene pairs of the donor and recipient are mutated.

[0257] Table 29

[0258]

[0259] Note: "Present" in the table means that at least one of the 27 LOF mutation gene pairs is mutated (a gene pair mutation means that both the donor's corresponding gene and the recipient's corresponding gene in the gene pair are mutated), and "absent" means that none of the 27 LOF mutation gene pairs are mutated (no mutation means that at least one of the donor's corresponding gene and the recipient's corresponding gene in each LOF mutation gene pair is not mutated).

[0260] II. Application of the number of 27 LOF mutation gene pairs in predicting the risk of aGVHD in hematopoietic cell transplantation patients

[0261] The 417 patients and their donors in Example 2 were used as research objects, and the total number of gene mutations for each patient was obtained according to the following method: if ASCC1-ANKRD20A2, ADAMTS7-APOL4, EFCAB5-ACTN2, EFCAB5-AGAP3, EFCAB5-ATP6V1B1, EFCAB5-CARD8, EFCAB5-CNTNAP3B, EFCAB5-DEFB126, EFCAB5-DENND3, EFCAB5-FRMD5, EFCAB5-HAPLN3, EFCAB5-HRNR, EFCAB5-KRT 24. Mutations in EFCAB5-KRTAP10-1, EFCAB5-LCN10, EFCAB5-MAGEE2, EFCAB5-MUC16, EFCAB5-PABPC3, EFCAB5-SETBP1, EFCAB5-SLC35G6, EFCAB5-TLR8, EFCAB5-ZFPM1, EFCAB5-ZNF211, MYO15B-TMX3, TMEM247-CPNE1, ZNF880-ANAPC1, or ZNF211-GIGYF2 were recorded as 1, and the cumulative number of mutations in the 27 LOF mutation gene pairs was calculated as the total number of gene mutations.

[0262] The number of LOF mutation gene pairs in the peripheral blood of donors and recipients was compared with the occurrence of aGVHD by Mann-Whitney U test, and the statistical difference between the two groups of aGVHD was compared. The receiver operating characteristic (ROC) curve was used to evaluate the specificity and sensitivity of diagnosis. The Yoden Index was used to calculate the diagnostic cut-off value.

[0263] After analysis, it was found that when patients had 5 or more LOF mutation gene pairs, the total number of gene mutations in the aGVHD negative group was significantly higher than that in the aGVHD positive group ( Figure 2 , P<0.001). ROC curve analysis showed that the area under the curve of the total number of gene mutations for diagnosing aGVHD was 0.841 ( Figure 3 , 95% confidence interval is 0.696-0.986), the total number of gene mutations corresponding to the maximum Youden index is 17.5, and 17.5 is used as the diagnostic threshold for aGVHD, and the sensitivity of the test is 81.8%. Therefore, in practical applications, the risk of aGVHD in patients can be predicted based on the number of 27 LOF mutation gene pairs in donors and recipients.

[0264] The present invention has been described in detail above. It will be apparent to those skilled in the art that the present invention may be implemented in a wide range under equivalent parameters, concentrations and conditions without departing from the spirit and scope of the present invention and without the need for unnecessary experimentation. Although the present invention provides specific embodiments, it should be understood that further improvements may be made to the present invention. In short, according to the principles of the present invention, this application intends to include any changes, uses or improvements to the present invention, including changes made by conventional techniques known in the art that depart from the scope disclosed in this application. Applications of some of the basic features may be made within the scope of the following appended claims.

Claims

1. The use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in the preparation of a product for predicting or assisting in predicting the risk of acute graft-versus-host disease in a patient after hematopoietic cell transplantation; or, the use of a substance that detects whether a gene pair in a donor and a recipient is mutated in predicting or assisting in predicting the risk of acute graft-versus-host disease in a patient after hematopoietic cell transplantation; The gene pair is at least one of the following gene pairs: ASCC1-ANKRD20A2, ADAMTS7-APOL4, EFCAB5-ACTN2, EFCAB5-AGAP3, EFCAB5-ATP6V1B1, EFCAB5-CARD8, EFCAB5-CNTNAP3B, EFCAB5-DEFB126, EFCAB5-DENND3, EFCAB5-FRMD5, EFCAB5-HAPLN3, EFCAB5-HRNR, EFCAB5-KR T24, EFCAB5-KRTAP10-1, EFCAB5-LCN10, EFCAB5-MAGEE2, EFCAB5-MUC16, EFCAB5-PABPC3, EFCAB5-SETBP1, EFCAB5-SL C35G6, EFCAB5-TLR8, EFCAB5-ZFPM1, EFCAB5-ZNF211, MYO15B-TMX3, TMEM247-CPNE1, ZNF880-ANAPC1, ZNF211-GIGYF2.

2. The use of substances for detecting whether gene pairs in donors and recipients are mutated in the preparation of products for diagnosing or assisting in diagnosing whether patients develop acute graft-versus-host disease after hematopoietic cell transplantation; Or, the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in diagnosing or assisting in diagnosing whether a patient develops acute graft-versus-host disease after hematopoietic cell transplantation; The gene pair is at least one of the following gene pairs: ASCC1-ANKRD20A2, ADAMTS7-APOL4, EFCAB5-ACTN2, EFCAB5-AGAP3, EFCAB5-ATP6V1B1, EFCAB5-CARD8, EFCAB5-CNTNAP3B, EFCAB5-DEFB126, EFCAB5-DENND3, EFCAB5-FRMD5, EFCAB5-HAPLN3, EFCAB5-HRNR, EFCAB5-KR T24, EFCAB5-KRTAP10-1, EFCAB5-LCN10, EFCAB5-MAGEE2, EFCAB5-MUC16, EFCAB5-PABPC3, EFCAB5-SETBP1, EFCAB5-SL C35G6, EFCAB5-TLR8, EFCAB5-ZFPM1, EFCAB5-ZNF211, MYO15B-TMX3, TMEM247-CPNE1, ZNF880-ANAPC1, ZNF211-GIGYF2.

3. The use of substances for detecting whether gene pairs in donors and recipients are mutated in the preparation of products for screening or assisting in the screening of donors before hematopoietic cell transplantation; or, the use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in screening or assisting in screening donors before hematopoietic cell transplantation; or, use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in the preparation of a product for matching a donor and a recipient before hematopoietic cell transplantation; or, use of a substance for detecting whether a gene pair in a donor and a recipient is mutated in matching a donor and a recipient before hematopoietic cell transplantation; The gene pair is at least one of the following gene pairs: ASCC1-ANKRD20A2, ADAMTS7-APOL4, EFCAB5-ACTN2, EFCAB5-AGAP3, EFCAB5-ATP6V1B1, EFCAB5-CARD8, EFCAB5-CNTNAP3B, EFCAB5-DEFB126, EFCAB5-DENND3, EFCAB5-FRMD5, EFCAB5-HAPLN3, EFCAB5-HRNR, EFCAB5-KR T24, EFCAB5-KRTAP10-1, EFCAB5-LCN10, EFCAB5-MAGEE2, EFCAB5-MUC16, EFCAB5-PABPC3, EFCAB5-SETBP1, EFCAB5-SL C35G6, EFCAB5-TLR8, EFCAB5-ZFPM1, EFCAB5-ZNF211, MYO15B-TMX3, TMEM247-CPNE1, ZNF880-ANAPC1, ZNF211-GIGYF2.

4. The use according to any one of claims 1 to 3, Features: The hematopoietic cell transplantation is allogeneic hematopoietic cell transplantation.

5. The use according to any one of claims 1 to 4, Features: The substance for detecting whether a gene pair in the donor and the recipient has mutated is a substance for detecting whether a gene pair in the peripheral blood of the donor and the recipient has mutated.

6. A kit having any one of the functions a1) to a4) below, comprising a substance for detecting whether a gene pair in a donor and a recipient has mutated; a1) To predict or assist in predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation; a2) To diagnose or assist in the diagnosis of acute graft-versus-host disease in patients after hematopoietic cell transplantation; a3) Screening or assisting in the screening of donors before hematopoietic cell transplantation; a4) Matching between donor and recipient before hematopoietic cell transplantation; The gene pair is at least one of the following gene pairs: ASCC1-ANKRD20A2, ADAMTS7-APOL4, EFCAB5-ACTN2, EFCAB5-AGAP3, EFCAB5-ATP6V1B1, EFCAB5-CARD8, EFCAB5-CNTNAP3B, EFCAB5-DEFB126, EFCAB5-DENND3, EFCAB5-FRMD5, EFCAB5-HAPLN3, EFCAB5-HRNR, EFCAB5-KR T24, EFCAB5-KRTAP10-1, EFCAB5-LCN10, EFCAB5-MAGEE2, EFCAB5-MUC16, EFCAB5-PABPC3, EFCAB5-SETBP1, EFCAB5-SL C35G6, EFCAB5-TLR8, EFCAB5-ZFPM1, EFCAB5-ZNF211, MYO15B-TMX3, TMEM247-CPNE1, ZNF880-ANAPC1, ZNF211-GIGYF2.

7. The kit according to claim 6, Features: The hematopoietic cell transplantation is allogeneic hematopoietic cell transplantation.

8. The kit according to claim 6 or 7, Features: The substance for detecting whether a gene pair in the donor and the recipient has mutated is a substance for detecting whether a gene pair in the peripheral blood of the donor and the recipient has mutated.

9. A kit according to any one of claims 6 to 8, Features: The kit also includes a data processing device, which has a module inside. The module has the following functions: predicting the risk of acute graft-versus-host disease in patients after hematopoietic cell transplantation or diagnosing whether acute graft-versus-host disease occurs in patients after hematopoietic cell transplantation or screening donors before hematopoietic cell transplantation based on whether gene pairs in the peripheral blood of donors and recipients are mutated.

10. Use of ASCC1, ANKRD20A2, ADAMTS7, APOL4, ACTN2, AGAP3, ATP6V1B1, CARD8, CNTNAP3B, DEFB126, DENND3, FRMD5, HAPLN3, HRNR, KRT24; KRTAP10-1, LCN10, MAGEE2, MUC16, PABPC3, SETBP1, SLC35G6, EFCAB5, TLR8, ZFPM1, EFCAB5, ZNF211, MYO15B, TMX3, TMEM247, CPNE1, ZNF880, ANAPC1, ZNF211 and / or GIGYF2 as targets in the preparation of products for the prevention and / or treatment of acute graft-versus-host disease.

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