TWI804620B - Methods and kits for detecting dmard-induced severe cutaneous adverse drug reactions and usesof the kits - Google Patents

Abstract

The present invention provides methods of assessing the risk of a subject developing DMARD-induced severe cutaneous adverse drug reactions (SCARs), by detecting the presence of specific HLA alleles. DMARD-induced SCARs include but are not limited to Stevens-Johnson syndrome (SJS), toxic epidermal necrolysis (TEN), or drug reactions with eosinophilia and systemic symptoms (DRESS). Kits for detecting specific HLA alleles for assessing the risk of DMARD-induced SCARs and the uses thereof are also included.

Description

Evaluation of the risk of serious cutaneous adverse drug reactions to disease-modifying antirheumatic drugs Method, its detection kit and its use

The present invention provides a method for assessing the risk of cutaneous adverse drug reactions (Cutaneous Adverse Drug Reactions) caused by disease-modifying antirheumatic drugs, especially Sulfasalazine (Sulfasalazine), Mesalazine (Mesalazine), Sulfapyridine (Sulfapyridine), Olsalazine and other methods that cause the risk of adverse skin drug reactions.

Cutaneous Adverse Drug Reactions (CADRs) have always been a major clinical problem, and their manifestations are very diverse, ranging from mild papules (maculopapular eruption, MPE), fixed drug eruption (fixed drug eruption, FDE) to severe skin drug eruptions Adverse reactions (severe cutaneous adverse drug reactions, SCARs), including: drug rash with eosinophilia and systemic symptoms (DRESS), Stevens Johnson Syndrome (SJS) And toxic epidermal necrolysis (TEN), etc. Stevenson-Johnson Syndrome (SJS) and Toxic Epidermal Necrosis (TEN) In the early stage, some cold-like symptoms often appear, including fever, sore throat, and swelling of the lips, and then rapidly develop systemic erythema, blisters, inflammation and ulceration of the mucous membranes of the eyes, mouth, and genitals, and in severe cases, it is like a patient who has been scalded all over the body. The biggest difference between the two is that if the range of epidermal separation is less than 10% of the body surface area, it is called SJS, and if it exceeds 30%, it is called TEN. The main clinical features of drug rash with eosinophilia and systemic symptoms (DRESS) include fever, skin rash, increased blood eosinophilic leukocytes, enlarged lymph nodes, and invasion of internal organs. The most common and most severely violated organ is the liver, which may be complicated by fulminant hepatitis, which becomes the most common cause of death for patients. Others include nephritis, myocarditis, pneumonia, and thyroid inflammation.

Adverse drug reactions are often related to immune responses, but the immune mechanism is very complex, such as: HLA-A has more than 300 genotypes; HLA-B has more than 600 genotypes. Therefore, it is difficult to find out the immune mechanism that causes adverse drug reactions.

The disease-regulating anti-rheumatic drug Sulfasalazine (Sulfasalazine) (trade name Salazine or Salazopyrin® or Azulfidine®) is a drug that regulates the immune system and has anti-inflammatory effects. It was approved by the US Food and Drug Administration in 1950. Approved by the FDA, it can be used to treat inflammatory bowel disease and various inflammatory arthritis, such as: rheumatoid arthritis, ankylosing spondylitis, psoriatic arthritis and juvenile chronic arthritis. Sulfasalazine and its metabolites, such as Mesalazine and Sulfapyridine, and Olsalazine, a dimer of Mesalazine, have anti-inflammatory, immunosuppressive and antibacterial effects, and are used in When treating inflammatory arthritis, in addition to reducing joint pain and swelling, it can also reduce the chance of permanent joint damage and disability.

Although disease-modifying antirheumatic drugs can be used to treat a variety of inflammatory diseases, their use is limited because of their high frequency of adverse reactions in clinical practice. About 25% of patients using sulfasalazine will experience significant side effects, including: loss of appetite, nausea, headache, white blood cell Cutaneous Adverse Drug Reactions (CADRs), among which cutaneous adverse drug reactions account for the second largest proportion of side effects. Therefore, there is still a need for risk assessment of serious cutaneous adverse drug reactions (including SJS, TEN, and DRESS) caused by disease-modifying antirheumatic drugs. The present invention addresses this need.

The present invention provides a method for assessing the risk of severe skin adverse drug reactions caused by disease-modulating antirheumatic drugs in patients, wherein serious skin adverse drug reactions include: Stevens Johnson Syndrome (Stevens Johnson Syndrome, SJS), toxic epidermal necrosis ( toxic epidermal necrolysis (TEN) or drug rash with eosinophilia and systemic symptoms (DRESS). HLA-B*1502, HLA-B*3802 or their allogene combinations, HLA-B*1301 and HLA-B*3901 allogene combinations are associated with severe skin adverse drug reactions induced by disease-modifying antirheumatic drugs.

Specifically, the present invention provides a method of assessing the risk of a patient to develop a serious cutaneous adverse drug reaction to a disease-modifying antirheumatic drug, comprising determining the presence of at least one allele selected from the group consisting of: HLA-B*1502, HLA- B*3802, or a combination of HLA-B*1301 and HLA-B*3901, where the presence of at least one allele is an indicator of risk for serious cutaneous adverse drug reactions. In a specific example, the drugs are disease-modifying anti-rheumatic drugs (Disease-modifying anti-rheumatic drugs, DMARDs). Disease modifying antirheumatic drugs include, but are not limited to, Sulfasalazine, Mesalazine, Sulfapyridine, or Olsalazine. Serious cutaneous adverse drug reactions include at least one adverse reaction selected from the group consisting of: Stevens Johnson Syndrome (SJS), toxic epidermal necrosis (toxic epidermal necrolysis (TEN) or drug rash with eosinophilia and systemic symptoms (DRESS). In a specific example, the patient carries the HLA-B*1502 allele. In a specific example, the patient carries the HLA-B*3802 allele. In one embodiment, the patient carries both HLA-B*1502 and HLA-B*3802 alleles. In one embodiment, the patient carries alleles of HLA-B*1301 and HLA-B*3802 and HLA-B*3901. In one embodiment, the patient has alleles of HLA-B*1301 and HLA-B*1502 and HLA-B*3802 and HLA-B*3901.

The present invention provides a set for detecting the alleles of HLA-B*1502, HLA-B*3802 or the combination of HLA-B*3901 and HLA-B*1301, which is used in the preparation of evaluation of disease-modulating anti-rheumatic drugs causing serious skin drug adverse effects Use of the risk of reaction, the kit comprising reagents for detecting at least one allele selected from the group consisting of: HLA-B*1502, HLA-B*3802, or HLA-B*1301 combined with HLA-B*3901 .

HLA-B*1502, HLA-B*3802, HLA-B*1301 and HLA-B*3901, HLA-B*1502 and HLA-B*3802, HLA-B*1301 and HLA-B*3802 and HLA- The presence of B*3901 or HLA-B*1301 and HLA-B*1502 and HLA-B*3802 and HLA-B*3901 alleles means that the patient is more likely than HLA-B*1502, HLA-B*3802, HLA-B *1301 and HLA-B*3901, HLA-B*1502 and HLA-B*3802, HLA-B*1301 and HLA-B*3802 and HLA-B*3901 or HLA-B*1301 and HLA-B*1502 Patients with the absence of alleles of HLA-B*3802 and HLA-B*3901 have more than one time, more than two times, more than three times, more than four times, more than five times, high More than six times, more than seven times, more than eight times, more than nine times, more than ten times, more than eleven times, more than twelve times, more than thirteen times, Higher than fourteen times, higher than fifteen times, higher than sixteen times, higher than seventeen times, higher than eighteen times, higher than nineteen times More than, more than 20 times, more than 30 times, more than 40 times, more than 50 times, more than one time to more than fourteen times the risk of drug allergic reaction.

The presence of an allele can be detected by any method known in the relevant art, such as, but not limited to, assays using oligonucleotides that specifically hybridize to the nucleic acid encoding the allele, serotyping, or microscopic cytotoxicity. To measure cDNA, RNA or protein products of alleles [Kenneth D. McClatkey. Clinical Laboratory Medicine. 2002]. In one embodiment, nucleic acid-specific hybridization oligonucleotide assays are performed using DNA prepared from peripheral blood of a patient. Among them, the specific oligonucleotides can be directed against the most variable sequences in the alleles of HLA-B*1301 and/or HLA-B*1502 and/or HLA-B*3802 and/or HLA-B*3901 design. In a specific example, the forward primer oligonucleotide sequence used to detect the presence of HLA-B*1502 is 5'-ATGGCGCCCCGGG-3' (Sequence 1), and the reverse primer sequence (reverse primer) is 5'-TAGTAGCCGCGCAGGTTCC-3' (sequence 2), probe 1 (probe 1) sequence is 5'-AACACACAGATCTACAAGG-3' (sequence 3) and probe 2 (probe 2) sequence is 5'-AACACACAGATCTCCAAGA-3' ( Sequence 4). In a specific example, the forward primer oligonucleotide sequence used to detect the presence of HLA-B*3802 is 5'-GCCGCGAGTCCGAGAGA-3' (SEQ ID NO: 5), and the reverse primer sequence (reverse primer) is 5'-GTGCGCAGGTTCTCTCGGTA-3' (SEQ ID NO: 6), probe 1 (probe 1) sequence is 5'-CCGGAGTATTGGGAC-3' (SEQ ID NO: 7) and probe 2 (probe 2) sequence is 5'-CCGGAATATTGGGAC-3' ( Sequence 8). In another specific example, the forward primer oligonucleotide sequence used to detect the presence of HLA-B*1301 is 5'-AGCCCCGCTTCATCACC-3' (SEQ ID NO: 9), and the reverse primer sequence (reverse primer) It is 5'-TCCTTGCCGTCGTAGGCTAA-3' (SEQ ID NO: 10), the sequence of probe 1 (probe 1) is 5'-CACATCATCCAGAGGAT-3' (SEQ ID NO: 11) and probe 2 (probe2) sequence is 5'-ACACTTGGCAGACGAT-3' (SEQ ID NO: 12). In another specific example, the forward primer oligonucleotide sequence used to detect the presence of HLA-B*3901 is 5'-GCGAGTCCGAGAGAGGAGC-3' (SEQ ID NO: 13), and the reverse primer sequence (reverse primer) It is 5'-TAGTAGCCGCGCAGGTTCC-3' (SEQ ID NO: 14), the sequence of probe 1 (probe 1) is 5'-TCCAATTCACAGACTGA-3' (SEQ ID NO: 15) and the sequence of probe 2 (probe 2) is 5'-CAACACACAGACTGA-3' (Sequence 16).

The present invention provides a detection kit for assessing the risk of serious adverse skin drug reactions caused by disease-modifying anti-rheumatic drugs. The detection kit includes a reagent capable of detecting at least one allele selected from the following: HLA-B*1502; HLA-B*3802 or HLA-B*1301 combined with HLA-B*3901, wherein the presence of at least one allele represents a higher disease-modifying antirheumatic drug in the patient than in patients without the at least one allele Risk of serious adverse skin drug reactions. In a specific example, the serious skin adverse drug reaction includes at least one adverse reaction selected from the group consisting of Stevenson-Johnson syndrome, toxic epidermal necrosis or drug eruption combined with eosinophilia and systemic symptoms.

The present invention provides methods for reducing the incidence of, or treating, severe cutaneous adverse drug reactions induced by disease-modifying antirheumatic drugs.

The present invention also provides a method for assessing the risk of adverse drug reactions caused by disease-modifying antirheumatic drugs and treating the adverse drug reactions, comprising the following steps: (a) detecting at least one allele selected from the following in a patient sample: HLA-B* 1502, HLA-B*3802, or HLA-B*1301 combined with HLA-B*3901, (b) if at least one of the following alleles is present in the sample: HLA-B*1502, HLA-B*3802, or HLA-B The combination of *1301 and HLA-B* 3901 can identify that the patient has an adverse drug reaction caused by a disease-modifying antirheumatic drug and (c) administer the drug to treat the adverse drug reaction.

In one embodiment, an adverse drug reaction is treated by administering a drug including, but not limited to, fluids, steroids, immunoglobulins, cyclosporine, anti-TNF-α agents, or plasmapheresis .

The present invention also relates to a method for evaluating the risk of adverse drug reactions caused by disease-modulating antirheumatic drugs and reducing the incidence of adverse drug reactions, comprising the following steps: (a) detecting at least one allele selected from the following in a patient sample: HLA- Presence of B*1502, HLA-B*3802, or HLA-B*1301 in combination with HLA-B*3901, (b) if at least one of the following alleles is present in the sample: HLA-B*1502, HLA-B*3802 or the combination of HLA-B*1301 and HLA-B*3901, the patient can be identified as having an increased risk of developing an adverse drug reaction and (c) the patient is not given a disease-modifying antirheumatic drug.

The terms "invention" and "the present invention" as used herein are intended to refer broadly to all claimed objects of the present invention, as well as the claims. Statements containing these terms should be understood not to limit the object of the application described herein or to limit the meaning or scope of the claims of the invention. Embodiments of the invention covered by this invention are defined by the claims, not this summary. This Summary is a high-level overview of various aspects of the invention and introduces some concepts that are further described below in the Detailed Description section. This summary is not intended to identify key or essential features of the claimed subject matter, nor is it intended to be used in isolation to determine the scope of the claimed subject matter. The subject matter of the application should be understood by reference to any or all of the drawings and each claim, as appropriate, throughout the specification.

In the following example, we collected 32 patients (including 11 SJS/TEN and 21 DRESS patients) who were treated with the disease-modifying antirheumatic drug sulfasalazine (Sulfasalazine) and had severe skin adverse drug reactions for HLA typing and compared with 941 A comparative analysis was performed with a control group of normal healthy people. The results showed that HLA-B*1301, HLA-B*1502, HLA-B*3802, HLA-B*3901, HLA-B*1301 and HLA-B*3901, HLA-B*1502 and HLA-B*3802, HLA-B*1301 and HLA-B*3802 and HLA-B*3901 or HLA-B*1301 and HLA-B*1502 and HLA-B*3802 and HLA-B*3901 alleles and sulfasalazine Serious cutaneous adverse drug reactions were associated (Table 1). In the case of HLA-B*1301 allele distribution, 8 of the 21 DRESS patients had this genotype (38.10%), and only 114 of the 941 normal healthy controls had this genotype (12.11%) , showing that HLA-B*1301 is associated with DRESS caused by sulfasalazine (DRESS vs. healthy control group: P=2.59x10 ^-3 , odds ratio (Odds Ratio or OR)=4.5 (1.8-11.0), Sensitivity: 38.10%, Specificity: 87.89%). In the case of HLA-B*3802 allele distribution, 6 of 11 SJS/TEN patients had this genotype (54.54%), and only 71 of 941 general healthy controls (General population) had this genotype. Genotype (7.55%), showing that HLA-B*3802 is associated with SJS/TEN caused by sulfasalazine (SJS/TEN vs. healthy control group: P=7.72x10 ^-5 , odds ratio (Odds Ratio or OR)=14.7 (4.4-49.4), sensitivity: 54.54%, specificity: 92.45%). Further combined analysis of HLA-B*1301 and HLA-B*3901 showed that the correlation and sensitivity of sulfasalazine-induced DRESS were significantly improved after the combination (DRESS vs. healthy control group: P=4.27x10 ^-8 , odds ratio = 13.1 (5.0-34.2), sensitivity: 71.43%, specificity: 83.95%). In the case of HLA-B*1502 allele distribution, 5 of the 11 SJS/TEN patients had this genotype (45.45%), and only 87 of the 941 normal healthy controls had this genotype (9.25%). %), showing that HLA-B*1502 is associated with SJS/TEN induced by sulfasalazine (SJS/TEN vs. healthy control group: P=2.19x10 ^-3 , odds ratio=8.2(2.4-27.4), Sensitivity: 45.45%, Specificity: 90.75%). In the case of HLA-B*3901 allele distribution, 8 of the 21 DRESS patients had this genotype (38.10%), and only 43 of the 941 normal healthy controls had this genotype (4.57%) , showing that HLA-B*3901 is associated with DRESS caused by sulfasalazine (DRESS vs. healthy control group: P=4.30x10 ^-6 , odds ratio (Odds Ratio or OR)=12.2 (4.6-32.5), Sensitivity: 38.10%, Specificity: 95.43%). Further combined analysis of HLA-B*1502 and HLA-B*3802, the results showed that the correlation and sensitivity of SJS/TEN caused by sulfasalazine were also significantly improved (SJS/TEN vs. healthy control group: P =5.98x10 ^-5 , odds ratio=13.7 (3.6-52.4), sensitivity: 72.72%, specificity: 83.74%). If HLA-B*1301, HLA-B*3802 and HLA-B*3901 were further combined and analyzed, the results showed that the correlation and sensitivity to severe cutaneous adverse drug reactions (SCAR) caused by sulfasalazine were significantly improved ( SCAR vs. healthy control group: P=3.21x10 ^-8 , odds ratio=7.6 (3.4-17.1), sensitivity: 68.75%, specificity: 78.32%). Further combined analysis of HLA-B*1301, HLA-B*1502, HLA-B*3802 and HLA-B*3901, the results showed the correlation with the serious skin adverse reaction (SCAR) caused by sulfasalazine And the sensitivity was significantly improved (SCAR vs. healthy control group: P=2.67x10 ^-7 , odds ratio=7.1 (3.0-16.9), sensitivity: 75.00%, specificity: 60.35%). From the above results, it is known that the detection of HLA-B*1301, HLA-B*1502, HLA-B*3802, HLA-B*3901, HLA-B*1301 and HLA-B*3901, HLA-B*1502 and HLA- B*3802, HLA-B*1301 and HLA-B*3802 and HLA-B*3901 or HLA-B*1301 and HLA-B*1502 and HLA-B*3802 and HLA-B*3901 alleles exist or not Can be used to assess the risk of cutaneous adverse drug reactions to the antiepileptic drug lamotrigine.

The above is a specific description of the technical characteristics of the present invention for the preferred embodiments of the present invention; however, those who are familiar with this technology should be able to change and modify the present invention without departing from the spirit and principles of the present invention. Such changes and modifications shall all be covered in the scope defined by the scope of the patent application as follows.

<110> Chang Gung Medical Foundation Linkou Chang Gung Memorial Hospital

<120> Method for assessing the risk of severe cutaneous adverse drug reactions caused by disease-modifying antirheumatic drugs, its detection kit and its use

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Claims (11)

A method for assessing the risk of a patient developing Stevens Johnson Syndrome (SJS) and/or toxic epidermal necrosis (TEN) caused by disease-modifying antirheumatic drugs, the method comprising determining the patient's HLA-B*3802 allele, wherein the presence of the allele represents that the patient has higher disease-modulating antirheumatic drug-induced Stevens Johnson Syndrome (SJS) and/or the presence of the allele Or the risk of toxic epidermal necrolysis (TEN), where the disease-modifying antirheumatic drug is Sulfasalazine. The method according to claim 1, further comprising measuring the HLA-B*1502 allele. A method for assessing the risk of developing severe cutaneous adverse drug reactions (SCARs) caused by disease-modifying antirheumatic drugs in a patient, the method comprising determining any of the following alleles in the patient: HLA-B* 1301 and HLA-B*3901; HLA-B*3802, HLA-B*1301 and HLA-B*3901; or HLA-B*3802, HLA-B*1301, HLA-B*1502 and HLA-B*3901 , wherein the presence of at least any one group of alleles represents that the patient has a higher risk of severe cutaneous adverse drug reactions caused by disease-modifying anti-rheumatic drugs than patients without at least one group of alleles, wherein the disease-modulating anti-rheumatic drugs The rheumatic drug is Sulfasalazine. The method as described in any one of claim 1 to claim 3, wherein the determination of HLA-B*1502, HLA-B*3802, HLA-B*1301 or HLA-B*3901 alleles is performed by peripheral blood from the patient Determination of samples prepared from DNA, RNA, protein, cells or serum. A test set for assessing the risk of developing disease-modulating antirheumatic drugs in patients with Stevens Johnson Syndrome (SJS) and/or toxic epidermal necrosis (TEN), the test set includes A reagent for detecting HLA-B*3802 and HLA-B*1502 in a test sample of a patient, wherein the disease-modifying antirheumatic drug is Sulfasalazine. A detection kit for assessing the risk of a patient developing a serious cutaneous adverse drug reaction caused by a disease-modifying antirheumatic drug, the detection kit includes any one of the following groups of reagents in the detection sample used to detect the patient: HLA-B*1301 with HLA-B*3901; HLA-B*3802, HLA-B*1301 with HLA-B*3901; or HLA-B*3802, HLA-B*1301, HLA-B*1502 with HLA-B*3901, The disease-modifying antirheumatic drug is Sulfasalazine. The detection kit according to any one of claim 5 to claim 6, wherein the detection kit comprises oligonucleotides specifically hybridized with the nucleic acid of the allele. A kit for detecting HLA-B*3802 alleles is being prepared for the evaluation of disease-modulating antirheumatic drugs causing Stevens Johnson Syndrome (SJS) and/or toxic epidermal necrosis (TEN) use of risk, where The disease modifying antirheumatic drug is Sulfasalazine. The use as described in Claim 8, further comprising a kit for detecting HLA-B*1502 alleles. A use of a kit for detecting alleles in preparation for assessing the risk of severe adverse skin drug reactions caused by disease-modulating antirheumatic drugs, wherein the alleles are selected from any group of alleles described below: HLA-B*1301 and HLA -B*3901; HLA-B*3802, HLA-B*1301 and HLA-B*3901; or HLA-B*3802, HLA-B*1301, HLA-B*1502 and HLA-B*3901, where the The disease-modifying antirheumatic drug was Sulfasalazine. The use according to any one of claim 8 to claim 10, wherein the set comprises oligonucleotides specifically hybridized with the nucleic acid of the allele.