Thanh bên bài viết

PDF (English)
Đã xuất bản: 2026-06-09
Số lượt xem tóm tắt: 0
Số lượt xem PDF (English): 0
DOI: https://doi.org/10.52852/tcncyh.v202i5E18.4838
Số xuất bản
Tập 202 Số 5E18 (2026)
Chuyên mục
Các bài báo
Cách trích dẫn
Thao, T. V., Nghiep, N. T., Bao, B. T., Mai, T. T. C., & Hoa, T. K. (2026). Development and Validation of a Probe-Free Real-time SYBR Green ARMS-PCR for UGT1A1*6 genotyping in the Vietnamese Kinh Population. Tạp Chí Nghiên cứu Y học, 202(5E18), 25-34. https://doi.org/10.52852/tcncyh.v202i5E18.4838

Development and Validation of a Probe-Free Real-time SYBR Green ARMS-PCR for UGT1A1*6 genotyping in the Vietnamese Kinh Population

Ta Van Thao, Nguyen Thi Nghiep, Bui Thi Bao, Tran Thi Chi Mai, Tran Khanh Hoa

Nội dung chính của bài viết

Tóm tắt

Irinotecan-induced toxicities, such as neutropenia and diarrhea, affect up to 36% of East Asian colorectal cancer patients due to impaired UGT1A1 glucuronidation, primarily from the UGT1A1*6 variant (c.211G>A, rs4148323). We developed a real-time SYBR Green ARMS-PCR assay using three parallel reactions: internal control with outer primers (OF/OR, 353 bp amplicon), G-specific (OF/IR_G, 176 bp), and A-specific (IF_A/OR, 232 bp). The assay was validated on 30 NGS-confirmed samples (10 GG, 10 GA, 10 AA) and applied to 100 healthy Kinh individuals. It achieved 100% NGS concordance, with ΔCt thresholds (≤ -8 for GG, -3 < ΔCt < 5 for GA, ≥ 8 for AA) yielding 100% sensitivity/specificity. In the Kinh cohort, genotypes were 75 GG (75%), 23 GA (23%), and 2 AA (2%), with G allele frequency 86.5% and A allele 13.5%, conforming to Hardy-Weinberg equilibrium (p=0.877). This probe-free assay enables rapid (<2 hours), low-cost genotyping for personalized irinotecan dosing in low-resource settings.

Chi tiết bài viết

Từ khóa

Real-time SYBR Green ARMS-PCR, SNP genotyping, UGT1A1*6, irinotecan toxicity, pharmacogenetics, Kinh population

Tài liệu tham khảo

1. Marsh S, McLeod HL. Pharmacogenetics of irinotecan toxicity. Pharmacogenomics. 2004; 5(7): 835-843. doi:10.1517/14622416.5.7.835.
2. Paulík A, Nekvindová J, Filip S. Irinotecan toxicity during treatment of metastatic colorectal cancer: focus on pharmacogenomics and personalized medicine. Tumori. 2020; 106(2): 87-94. doi:10.1177/0300891618811283.
3. Cheng L, Li M, Hu J, et al. UGT1A1*6 polymorphisms are correlated with irinotecan-induced toxicity: a systematic review and meta-analysis in Asians. Cancer Chemother Pharmacol. 2014; 73(3): 551-560. doi:10.1007/s00280-013-2310-2.
4. Yang Y, Zhou M, Hu M, et al. UGT1A16 and UGT1A128 polymorphisms are correlated with irinotecan-induced toxicity: a meta-analysis. Asia Pac J Clin Oncol. 2018; 14(5): e479-e489. doi:10.1111/ajco.13028.
5. Gibson N. The use of real-time PCR methods in DNA sequence variation analysis. Clin Chim Acta. 2006; 363(1-2): 32-47. doi:10.1016/j.cca.2005.07.020.
6. Morlan JD, Baker J, Sinicropi D. Mutation detection by real-time PCR: a simple, robust and highly selective method. PLoS One. 2009; 4(2): e4584. doi:10.1371/journal.pone.0004584.
7. Karas S, Innocenti F. All you need to know about UGT1A1 genetic testing for patients treated with irinotecan: a practitioner-friendly guide. JCO Oncol Pract. 2022; 18(4): 270-277. doi:10.1200/OP.21.00624.
8. Subbarayan P, Sarkar M, Ardalan B. Isolation of genomic DNA from human whole blood. BioTechniques. 2002; 33(6): 1231-1234. doi:10.2144/02336bm10.
9. Collins A, Ke X. Primer1: primer design web service for tetra-primer ARMS-PCR. Open Bioinform J. 2012; 6: 55-58. doi:10.2174/1875036201206010055.
7. Pichler V, Sanou A, Love RR, et al. A novel tetra-primer ARMS-PCR approach for the molecular karyotyping of chromosomal inversion 2Ru in the main malaria vectors Anopheles gambiae and Anopheles coluzzii. Parasit Vectors. 2023; 16: 388. doi:10.1186/s13071-023-06014-6.
8. Bustin SA, Benes V, Garson JA, et al. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem. 2009; 55(4): 611-622. doi:10.1373/clinchem.2008.112797.
9. Yamagata H, Kobayashi A, Tsunedomi R, et al. Optimized protocol for the extraction of RNA and DNA from frozen whole blood sample stored in a single EDTA tube. Sci Rep. 2021; 11(1): 17075. doi:10.1038/s41598-021-96567-2.
10. Medrano RFV, de Oliveira CA. Guidelines for the tetra-primer ARMS-PCR technique development. Mol Biotechnol. 2014; 56(7): 599-608. doi:10.1007/s12033-014-9734-4.
11. Ye S, Dhillon S, Ke X, Collins A, Day IN. An efficient procedure for genotyping single nucleotide polymorphisms. Nucleic Acids Res. 2001; 29(17): e88. doi:10.1093/nar/29.17.e88.
12. Shin J, Jung C. Improving the accuracy of single-nucleotide variant diagnosis using on-off discriminating primers. Biosensors (Basel). 2023; 13(3): 380. doi:10.3390/bios13030380.
13. Baris I, Etlik O, Koksal V, Ocak Z, Baris S. SYBR green dye-based probe-free SNP genotyping: introduction of T-PLEX real-time PCR assay. Anal Biochem. 2013; 441(2): 225-231. doi:10.1016/j.ab.2013.06.022.