LP-110 development of GSTA1, CYP2C19, and CYP2B6 gene polymorphism detection methods on the response of cyclophosphamide therapy for lupus nephritis patients

Indrawijaya, Yen Yen Ari, Iwo, Maria Immaculata, Artarini, Aluicia Anita and Hamijoyo, Laniyati (2023) LP-110 development of GSTA1, CYP2C19, and CYP2B6 gene polymorphism detection methods on the response of cyclophosphamide therapy for lupus nephritis patients. Presented at Lupus KCR 2023, May 2023, Seoul, Korea.

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Abstract

Background
Cyclophosphamide, an immunosuppressant in patients with lupus nephritis, aims to prevent recurrence and is a steroid-sparing agent. The clinical response (efficacy and toxicity) of lupus nephritis patients receiving cyclophosphamide varied.1 Cyclophosphamide is a pro-drug metabolized by hydroxylation via the CYP2C19 and CYP2B6 enzymes and detoxification via the glutathione-S-transferase (GST) enzyme.2 The most common type of mutation that occurs is single nucleotide polymorphism (SNP). Several SNPs from previous studies associated with cyclophosphamide response, metabolism, and toxicity in patients with lupus nephritis, namely rs3957356 in the GSTA1, rs4244285 in the CYP2C19, rs7254579 and rs4802101 in the CYP2B6.3 Therefore, detecting and screening SNP genotypes in lupus nephritis patients can help analyze cyclophosphamide response variation. This study aims to develop a method for detecting the genotypes of the four target SNPs and several surrounding SNPs using PCR-Sanger sequencing.

Methods
The gene polymorphism analysis method was optimized before taking patient samples at the hospital. PCR and Sanger sequencing methods are used for gene polymorphism analysis because they produce chromatograms with target amplicon base lengths and several SNPs that can be analyzed simultaneously.

Results
The analysis results of the four target SNPs of the GSTA1, CYP2C19, and CYP2B6 (rs3957356, rs4244285, rs7254579, and rs4802101) showed one synonymous SNP and three SNPs (regulatory and intergenic). We have developed an SNP detection assay using four fragments from the GSTA1, CYP2C19, and CYP2B6 that can detect 15 SNPs simultaneously (figure 1). In addition, this method succeeded in distinguishing wild-type, heterozygous and homozygous genotypes. Furthermore, this method can be used to analyze GSTA1, CYP2C19, and CYP2B6 gene polymorphisms in lupus nephritis patients receiving cyclophosphamide therapy, especially in a population in West Java, Indonesia.

Conclusions
PCR-sanger sequencing is a reliable, accurate, and simple method for determining SNP genotypes from blood samples.

Item Type:	Conference (Poster)
Keywords:	SNP; PCR-Sanger sequencing; Lupus Nephritis
Subjects:	11 MEDICAL AND HEALTH SCIENCES > 1115 Pharmacology and Pharmaceutical Sciences > 111505 Pharmacogenomics
Divisions:	Faculty of Medical and Health Sciences > Department of Pharmacy
Depositing User:	Yen Yen Indrawijaya
Date Deposited:	12 Jan 2024 09:32

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