MALBAC-based chromosomal imbalance analysis: a novel technique enabling effective non-invasive diagnosis and monitoring of bladder cancer

Hao Liu, Wang He, Bo Wang, Kewei Xu, Jinli Han, Junjiong Zheng, Jun Ren, Lin Shao, Shiping Bo, Sijia Lu, Tianxin Lin, Jian Huang, Hao Liu, Wang He, Bo Wang, Kewei Xu, Jinli Han, Junjiong Zheng, Jun Ren, Lin Shao, Shiping Bo, Sijia Lu, Tianxin Lin, Jian Huang

Abstract

Background: The gold standard for bladder cancer detection is cystoscopy, which is an invasive procedure that causes discomfort in patients. The currently available non-invasive approaches either show limited sensitivity in low-grade tumours or possess unsatisfying specificity. The aim of the present study is to develop a new non-invasive strategy based on chromosomal imbalance levels to detect bladder cancer effectively.

Methods: We enrolled 74 patients diagnosed with bladder cancer (BC), 51 healthy participants and 27 patients who were diagnosed with non-malignant urinary disease (UD). The Chromosomal Imbalance Analysis (CIA) was conducted in the tumours and urine of participants via the multiple annealing and looping-based amplification cycles-next-generation sequencing (MALBAC-NGS) strategy. The threshold of the CIA was determined with the receiver operating characteristic (ROC) curve. The comparison of the CIA with voided urine cytology was also performed in a subgroup of 55 BC patients. The consistency and discrepancy of the different assays were studied with the Kappa analysis and the McNemar test, respectively. The performance of the urinary CIA was also validated in an additional group of 120 BC patients, 15 UD and 45 healthy participants.

Results: Good concordance (87.0%) in the assessments of patient tumour tissues and urine was observed. The urine-based evaluation also demonstrated a good performance (accuracy = 89.0%, sensitivity = 83.1%, specificity = 94.5%, NPV = 85.4% and PPV = 93.7%; AUC = 0.917, 95%CI =0.868-0.966, P < 0.001) in the training group, particularly in the patients with CIA-positive tumours (accuracy = 92.7%, sensitivity = 89.8%). The sensitivity and specificity in the validation group were 89.2 and 90.0%, respectively. Even in Ta/T1 and low-grade tumour patients, the sensitivity was 85-90%. The CIA also exhibited a significantly improved sensitivity compared to voided urine cytology.

Conclusions: This is the first study employing the concept of whole genome imbalance combined with the MALBAC technique to detect bladder cancer in urine. MALBAC-CIA yielded significant diagnostic power, even in early-stage/low-grade tumour patients, and it may be used as a non-invasive approach for diagnosis and recurrence surveillance in bladder cancer prior to the use of cystoscopy, which would largely reduce the burden on patients.

Keywords: Bladder Cancer; CNV; Chromosomal imbalance analysis; MALBAC; NGS.

Conflict of interest statement

Ethics approval and consent to participate

Written informed consent was obtained from all participants, and this study was approved by the Medical Ethics Committee of Sun Yat-sen Memorial Hospital, Sun Yat-sen University.

Competing interests

S. Lu is the co-founder of Yikon Genomics.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1
Fig. 1
ROC curve analysis for urine CIA scores. To determine the best cut-off value that discriminated between malignant BC patients and control groups, urine CIA scores from 71 BC, 23 UD and 51 healthy participants were included. The cut-off was defined as 24 [Accuracy = 89.0%, sensitivity = 83.1%, specificity = 94.5%, NPV = 85.4% and PPV = 93.7%]. Area under the curve (AUC) =0.917, 95%CI =0.868–0.966, P < 0.001
Fig. 2
Fig. 2
The distribution of CIA scores. a CIA scores in tumour tissues from bladder cancer patients (BC-t), urine of BC patients (BC-u), urine of BC patients with paired CIA positive tumour tissues (BC-u (t+)), urine of non-malignant urinary disease patients (UD-u) and healthy controls (NC-u); b Tumour tissue CIA scores in different stages and grades; c Urine CIA scores in different stages and grades. The cut-off for positive CIA definition was set to 24. The P value was calculated from Fisher’s Exact Test
Fig. 3
Fig. 3
The demonstration of chromosomal CNV patterns in BC patients. The CNV profiles in tumour tissue and paired urine samples for patients No. 22, No. 27 and No. 28 (See Additional file 2: Table S1) are shown

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