Novel Semi-Nested Real-Time PCR Assay Leveraging Extendable Blocking Probes for Improved SHOX2 Methylation Analysis in Lung Cancer

Autor: Ngoc Anh Phuong, Trang Thuy Dao, Phuong Bich Pham, Ung Dinh Nguyen, Ba Van Nguyen, Tho Huu Ho
Jazyk: angličtina
Rok vydání: 2024
Předmět:
Zdroj: Biomolecules, Vol 14, Iss 6, p 729 (2024)
Druh dokumentu: article
ISSN: 2218-273X
DOI: 10.3390/biom14060729
Popis: Lung cancer is the leading cause of cancer deaths globally, necessitating effective early detection methods. Traditional diagnostics like low-dose computed tomography (LDCT) often yield high false positive rates. SHOX2 gene methylation has emerged as a promising biomarker. This study aimed to develop and validate a novel semi-nested real-time PCR assay enhancing sensitivity and specificity for detecting SHOX2 methylation using extendable blocking probes (ExBPs). The assay integrates a semi-nested PCR approach with ExBPs, enhancing the detection of low-abundance methylated SHOX2 DNA amidst unmethylated sequences. It was tested on spiked samples with varied methylation levels and on clinical samples from lung cancer patients and individuals with benign lung conditions. The assay detected methylated SHOX2 DNA down to 0.01%. Clinical evaluations confirmed its ability to effectively differentiate between lung cancer patients and those with benign conditions, demonstrating enhanced sensitivity and specificity. The use of ExBPs minimized non-target sequence amplification, crucial for reducing false positives. The novel semi-nested real-time PCR assay offers a cost-effective, highly sensitive, and specific method for detecting SHOX2 methylation, enhancing early lung cancer detection and monitoring, particularly valuable in resource-limited settings.
Databáze: Directory of Open Access Journals
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