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Background: The tumor necrosis factor-alpha (TNF-α) -308GA polymorphism (rs1800629) may affect TNF-α expression and the effectiveness of TNF-α inhibitors in autoimmune conditions such as ankylosing spondylitis (AS). For clinical use, it is very important to have fast and accurate genotyping methods. This study aimed to comprehensively evaluate a fluorescence quantitative polymerase chain reaction (PCR) method based on TaqMan minor groove binder (MGB) probe technology in detecting the TNF-α rs1800629 polymorphism and investigate the correlation between this SNP and the efficacy of TNF-α inhibitor therapy in patients with AS.
Methods: A TaqMan-MGB FQ-PCR detection system was constructed for the detection of the TNF-α single nucleotide polymorphism (SNP) (-308G>A). The limit of detection, accuracy, precision, specificity, sensitivity, and stability were assessed. A multicenter clinical trial was conducted to assess the detection system using a double-blind method involving 1,045 patients with immune diseases. The genotypes of TNF-α -308G/A from 20 patients with hyperlipidemia or jaundice were tested using the detection systems to evaluate the influence of interference. The correlation between the clinical response to TNF-α inhibitors and TNF-α -308G/A SNP was investigated using the 32 patients with ankylosing spondylitis.
Results: The detection range limit was 151.4 ng/μL - 0.5 ng/μL. The genotype detected using the research detection system and Sanger sequencing was 100%, with a Kappa 1.00 > 0.75, p = 0 < 0.001. The coefficient of variation of precision was less than 8%. Interfering substances did not affect the detection system. The follow-up trial indicated that the patients with the GA genotype, three out of the four cases, had no response for TNF-α inhibitors. In contrast, all patients with the GG genotypes experienced better treatment efficacy with TNF-α inhibitors.
Conclusions: The detection system utilizing the TaqMan-MGB probe for TNF-α polymorphism - 308G/A demonstrated high reliability. TNF-α -308G/A polymorphisms are potential biomarkers for predicting the treatment efficacy of TNF-α inhibitors for patients with ankylosing spondylitis.
DOI: 10.7754/Clin.Lab.2025.250918
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