Abstract 2345: Trans-renal DNA: Urine liquid biopsies methodology to increase compliance, convenience, and yield of systemic ctDNA
Authors: Floyd E. Taub, Lindsey Ray, Charles Preston Neff, Elijah Barstis, Charles Henry, Kirsten Arnold
Journal: Cancer Research (2025)

Abstract Liquid biopsies promise monitoring residual disease, monitoring for minimal recurrence, evaluating the immediate effects of therapy, and even screening healthy subjects for cancer. Access and patient considerations should be built into the system from ground up. The success of all existing screening programs relies more on participation rates than incremental gains in sensitivity and specificity. Resistance to blood draws, especially for underserved patients is high and COVID, immunosuppression, and personal beliefs decrease participation when needle puncture is required. Venipuncture increases costs, biohazard waste and is sufficiently burdensome to lower participation and medical benefit. Urine is truly noninvasive. While long known to contain trans-renal cfDNA (tr-cfDNA), less effort has gone into specifically isolating this subset of urine DNA and using it as a substitute for blood cfDNA in cancer studies. Striking success has been seen but kits modified from blood kits typically isolate predominantly urinary DNA. Methods designed to handle shorter tr-cfDNA fragments are needed. The glomerular filtration limit of ∼10 nm permits cfDNA of ≤100 bp to transit into the urine. Nucleosomes exceed 100 nm. Thus, we have designed a system to capture short non-nucleosomal tr-cfDNA. Urine is available in many hundreds of mls, compared to blood typically providing ≤10 mls of plasma. The latter is often insufficient to have the sequence of interest. Even with urine, data suggests that the more tested, the more likely rare sequences, like those from cancer, are detected. Further detailed understanding of diurnal, biological, daily, and weekly longitudinal ctDNA dynamics, in an outpatient setting, is facilitated by urine sampling. Herein we present a patient focused device that allows self-collection of urine voids (up to 500 mls) with immediate stabilization and separation, within minutes, of trans-renal and urothelial DNA onto separate filters contained within a filter cartridge. The tightly captured DNA excludes DNase binding, the DNA is thus stable at ambient temperature. A single filter cartridge can capture multiple voids and can then be sent by standard shipping. Even from small voids ≥2 ug rather than 10-20 ng amounts of cfDNA are collected. This allows higher sensitivity, methods requiring more DNA, as well as reuse should an initial assay fail, and banking for subsequent studies. The system specifically purifies tr-cfDNA based on interaction of the anionic free DNA with cationic filter bound polymer that does not bind urothelial nucleosomal DNA with positively charged histones. In the lab DNA is washed and eluted with simple syringe-based methods resulting in a purified preparation in 10 minutes. This generates a single peak of the expected trans renal size (80-100 bp) on capillary electrophoresis. Included data demonstrate single copy PCR and high quality (95% Q30s) in NGS. Citation Format: Floyd E. Taub, Lindsey Ray, Charles Preston Neff, Elijah Barstis, Charles Henry, Kirsten Arnold. Trans-renal DNA: Urine liquid biopsies methodology to increase compliance, convenience, and yield of systemic ctDNA [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2345.