Journal of Molecular Diagnostics最新文献

Epidermal growth factor receptor variant (EGFRv)-III, a common oncogenic variant in glioblastoma and other solid tumors, results from an in-frame deletion of exons 2 to 7 from the EGFR gene. Detection of EGFRvIII is crucial for understanding tumor biology, guiding targeted therapies, and developing personalized treatment strategies. In this study, two detection approaches based on DNA next-generation sequencing-read depth (RD)-based and split read (SR)-based detection-were compared to evaluate their sensitivity and accuracy in identifying EGFRvIII. Thirty-one tumor samples, including glioblastoma and pancreatic adenocarcinoma, were analyzed using both methods. The SR-based method detected EGFRvIII in 20 of 31 samples, while the RD-based method identified it in only 12 samples (P < 0.001), demonstrating that the SR-based method had significantly higher sensitivity. RNA next-generation sequencing confirmed EGFRvIII expression in most SR-positive cases. Additionally, the SR-based method identified multiple breakpoints in several tumors, revealing intratumor heterogeneity and the subclonal origins of EGFRvIII. The RD-based method was prone to false negatives, particularly in cases with high EGFR amplification or low tumor cell percentage, where copy number variations could be masked by background noise. The findings highlight the sensitivity and accuracy of SR-based detection in identifying EGFRvIII and capturing intratumor heterogeneity. SR-based analysis is recommended as the method for EGFRvIII detection in both clinical and research settings.

Breast cancer is the most common cancer among women, and metastasis to the lung is associated with poor prognosis. Reliable biomarkers for predicting lung metastasis are urgently needed to improve early detection and clinical decision-making. This study used microarray data sets comprising gene expression profiles and clinical data from primary breast cancer patients who were followed up for lung metastasis outcomes. High-throughput screening combined with Venn diagram analysis was used to identify common candidate probes, and the least absolute shrinkage and selection operator method were used to select 11 genes for model development. Logistic regression was used to construct predictive models, and the final risk signature consisted of 10 candidate genes (CDK19, GLUD1, GTPBP4, HLCS, HYI, KCND3, MAP2K1, NMUR1, PRKD3, and SLC16A3). The model achieved strong performance in training and validation cohorts (areas under the curve >0.87) and generalized to the independent METABRIC data set (area under the curve = 0.706). Subset analyses restricted to patients with early-stage disease confirmed that the signature retained predictive value. Kaplan-Meier analyses showed that patients with high-risk scores had shorter lung metastasis-free survival, recurrence-free survival, and overall survival. Multivariate Cox analysis confirmed that the risk signature provided independent predictive information from clinical variables. In conclusion, the risk signature accurately identifies patients with breast cancer at risk of lung metastasis, enabling clinicians to better assess risk and tailor effective treatment strategies.

Cameroon is an epicenter of diverse HIV-1 strains, with diagnostic and management challenges. The objective herein was to update HIV-1 non-M prevalence and compare diagnostic performance of two- versus three-test algorithms. A facility-based study was conducted in February 2024 on 2207 HIV-1 clinical samples at the Chantal Biya International Reference Centre (Yaoundé, Cameroon). HIV-1 non-M were identified by molecular phylogeny. Rapid diagnostic tests (RDTs) used in the two-test (Determine and KHB colloidal gold) versus three-test (First Response, One Step, and KHB) algorithms were evaluated on non-M, with ACRO (HIV1/2 and p24) as independent RDT. No group N (0%) nor P (0%) was found, whereas nine group O were identified (0.4%; 95% CI, 0.2%-0.8%). For individuals harboring group O (mean age, 43 ± 12 years; 50% female), median (IQR): duration since HIV diagnosis was 627 (423 to 775) weeks; viremia, 12,385 (5340 to 72,682) copies/mL; and CD4 count, 52 (39 to 228) cells/mm³. One Step, KHB, and ACRO detected eight of eight group O (100%); First Response HIV1-2.0, seven of eight (87.5%); and Determine HIV1/2, six of eight (75%), P = 1.00. In this Cameroonian setting, HIV-1 group N and P are scarce, whereas group O remains low (<1%). Transitioning from the two-test (75% performance) to the three-test algorithm (87.5% performance) could lead to improved diagnostic performance on currently circulating HIV-1 group O, calling for updates in RDTs to adapt to viral dynamics.

Conventional methods for spinal muscular atrophy (SMA) screening have been challenging in detecting SMN1/2 single-nucleotide variants (SNVs) and small insertions and deletions, SMN1 2 + 0 silent carrier, and the copy number (CN) of SMN2. To address these limitations, a long-read sequencing (LRS)-based approach termed comprehensive analysis of SMA 2 (CASMA2) was developed. CASMA2 was used to perform CN analysis by integrating Poisson distribution with an endogenous reference gene, the first such method developed for LRS platforms. The performance and clinical feasibility of CASMA2 were evaluated by using 414 retrospective peripheral blood samples and 303 prospective dried blood spot samples. CASMA2 displayed 100% accuracy in SMN1/2 CN analysis and identified the SNVs/insertions and deletions in SMN1/2. CASMA2 also showed the capability of screening for the SMN1 2 + 0 silent carrier with family-trio haplotype analysis. It achieved a 99.0% (410 of 414) first-attempt success rate for long-term peripheral blood samples and a 98.7% (299 of 303) rate for dried blood spot samples. CASMA2 offers a clinically feasible, precise, and efficient method for SMA carrier and newborn screening.

Splice-site variants within the ABO gene have the potential to impair ABO glycosyltransferase biosynthesis, leading to decreased expression of A or B antigens on the surface of red blood cells. This study characterized how four intron 6 splice-site variants-three in the A1 allele (c.374+4A>T, c.374+4A>G, and c.374+5G>A) and one in the ABO∗B.01 allele (c.374+2_374+3insT)-affect ABO pre-mRNA splicing. A combination of serologic, molecular genetic, bioinformatic, and minigene assays established a genotype-splicing-phenotype association model. Bioinformatics predictions showed that all variants impaired the recognition of the 5' splice site. Specifically, the A allele variant c.374+5G>A induced approximately 2.8% exon 6 retention, whereas the c.374+4A>G (A_finn/A_weak) and c.374+4A>T (A_weak) variants preserved approximately 6.9% and 10.2% functional transcripts, respectively. The novel B allele insertion variant c.374+2_374+3insT (B_el subtype) retained approximately 4.7% exon 6-containing transcripts, sufficient for trace B glycosyltransferase expression. Quantitative real-time PCR analysis confirmed that the transcriptional levels of the ABO gene in the AB_el subtype individual carrying the c.374+2_374+3insT variant were approximately 51.7% of those in the ABO∗A1.02/ABO∗B.01 control. This study demonstrated that splice-site variants in the ABO gene reduce the abundance of functional transcripts via altered transcriptional regulation, which, in turn, leads to decreased ABO glycosyltransferase expression, ultimately resulting in weak antigen phenotypes of varying intensity.