CAP TODAY Pathology/Laboratory Medicine/Laboratory Management
With the second approach, tumor and germline tissue are sequenced first to identify somatic mutations, and then the cfDNA sample is classified as positive if one of those somatic mutations is identified in the urine as well.
Since both approaches have advantages and disadvantages—for example, tumor-informed profiling requires obtaining tumor and germline tissue but offers the ability to identify mutations that can then be used to detect cancer with better specificity—the group employed both methods and compared them using their workflow.
They assessed the assay technically using urine from 33 young, healthy controls. They then applied the thresholds set based on that group to two separate clinical groups, the first consisting of 54 patients with early-stage, biopsy-proven bladder cancer (urine obtained before treatment). Thirty-four controls without bladder cancer were approximately matched according to age and smoking status.
“In this context, with the tumor-informed approach we achieved a sensitivity of 93 percent and a specificity of 96 percent,” while the tumor-naive approach yielded a similar specificity (97 percent) but a sensitivity that was slightly lower, at 83 percent, Dr. Dudley said. The sensitivity of cytology was 14 percent, or about sixfold lower, though it had 100 percent specificity.
“Within patients who had tumor DNA detected, the concentration of tumor DNA in the urine was very strongly correlated with high-risk clinical parameters, including T stage, lesion morphology, lesion grade, as well as cytology status,” he said.
To establish the utility of the panel for bladder cancer surveillance, the researchers tested stored specimens from 67 patients with a history of bladder cancer, including 37 patients who ultimately developed recurrent disease. They stored the urine samples with EDTA at 4°C and used or froze them within one week.
“In this context, as a gold standard we used biopsy-proven follow-up for patients and at least a year of negative clinical follow-up for true negatives,” Dr. Dudley said.
The panel’s performance was similar in the surveillance context as it had been for early bladder cancer detection, with a sensitivity of 91 percent and 100 percent specificity using a tumor-informed approach and 84 percent sensitivity and 96 percent specificity using a tumor-naive approach.
Using statistical analyses, they determined that detection of utDNA was “very predictive of recurrence,” with a hazard ratio of 8.8, and detectable utDNA preceded clinical recurrence in 92 percent of patients by a median of 2.7 months.
Cytology was only 38 percent sensitive in identifying recurrence, performing slightly better than it had for early-stage detection, Dr. Dudley said, noting that cytology was more likely to detect recurrent high-grade cancers.
He also reported that across both cohorts, uCAPP-Seq detected tumor DNA in 100 percent of bladder cancers detected by cytology and 82 percent of cases that cytology missed.
Cystoscopy and cytology together, the current standard of care in surveillance, detected 53 percent of recurrent cancers.
When they used UroVysion in a subset of seven patients who developed recurrent disease, it was positive for three and thus its sensitivity was 43 percent.
Other studies have explored the use of cellular or cfDNA as a biomarker for bladder cancer. “Our approach differs in using hybrid-capture target enrichment, interrogating a significantly larger genomic territory and number of mutations per case, and detecting somatic variants and copy-number alterations in one assay,” Dr. Dudley and coauthors write in Cancer Discovery.
Although their method will need to be tested prospectively and in larger studies, Dr. Dudley tells CAP TODAY, “these initial results suggest it could offer a significant improvement over the sensitivity of cytology without compromising cytology’s high specificity.” And, in a surveillance context, it could reduce the frequency of expensive and invasive cystoscopy procedures.
“Given the long lead time observed in some cases,” he and coauthors say, “it is tempting to speculate that intervention at the time of utDNA positivity might increase the likelihood of success of bladder-sparing interventions, though this will need to be tested in prospective trials.”
David Wild is a writer in Toronto. Dr. Dudley received the AMP’s Young Investigator Award for this work.