Abstract
PurposeThere is a need to supplement or supplant the conventional diagnostic tools, namely, cystoscopy and B-type ultrasound, for bladder cancer (BC). We aimed to identify novel DNA methylation markers for BC through genome-wide profiling of BC cell lines and subsequent methylation-specific PCR (MSP) screening of clinical urine samples.Experimental DesignThe methyl-DNA binding domain (MBD) capture technique, methylCap/seq, was performed to screen for specific hypermethylated CpG islands in two BC cell lines (5637 and T24). The top one hundred hypermethylated targets were sequentially screened by MSP in urine samples to gradually narrow the target number and optimize the composition of the diagnostic panel. The diagnostic performance of the obtained panel was evaluated in different clinical scenarios.ResultsA total of 1,627 hypermethylated promoter targets in the BC cell lines was identified by Illumina sequencing. The top 104 hypermethylated targets were reduced to eight genes (VAX1, KCNV1, ECEL1, TMEM26, TAL1, PROX1, SLC6A20, and LMX1A) after the urine DNA screening in a small sample size of 8 normal control and 18 BC subjects. Validation in an independent sample of 212 BC patients enabled the optimization of five methylation targets, including VAX1, KCNV1, TAL1, PPOX1, and CFTR, which was obtained in our previous study, for BC diagnosis with a sensitivity and specificity of 88.68% and 87.25%, respectively. In addition, the methylation of VAX1 and LMX1A was found to be associated with BC recurrence.ConclusionsWe identified a promising diagnostic marker panel for early non-invasive detection and subsequent BC surveillance.
Highlights
Bladder cancer (BC) is one of the leading causes of cancer-related morbidity and mortality and the sixth most common cancer in the world [1]
A total of 1,627 hypermethylated promoter targets in the BC cell lines was identified by Illumina sequencing
Validation in an independent sample of 212 BC patients enabled the optimization of five methylation targets, including VAX1, KCNV1, TAL1, PPOX1, and CFTR, which was obtained in our previous study, for BC diagnosis with a sensitivity and specificity of 88.68% and 87.25%, respectively
Summary
Bladder cancer (BC) is one of the leading causes of cancer-related morbidity and mortality and the sixth most common cancer in the world [1]. BC incidence increases with age; the average age at the time of diagnosis is approximately 60 years, and it is 3 times more common in men than in women [3]. 75–80% of new BC cases occur as superficial or carcinoma in situ lesions, whereas the remaining 20–25% present as a more advanced disease, with a poor prognosis. Even in the superficial tumors, only 20% are curable. 60–70% of patients will relapse within 5 years, and 10–20% of tumors will progress to a more aggressive disease [5], which necessitates frequent monitoring for disease recurrence [6]. Cystoscopy is the most common diagnostic BC procedure, and it shows high sensitivity (SN) and specificity (SP). Cystoscopy requires high operator proficiency, and the invasive nature of cystoscopy reduces its value as a screening tool. Other optimal methods are needed for the early, non-invasive detection and surveillance of BC
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