Abstract 245: Development of a novel urine cell-free DNA preservation method for molecular profiling in genitourinary tumors
Authors: zhang jie , liu fang , wang min , deng cuishan , mo wei , tao seng , wang amy , jia shidong , zhao zhixin , du pan , xiang binggang , luo shujun 
Published: April 2023

Abstract
Introduction: Due to the benefits of its non-invasive testing, liquid biopsy technology has been applied therapeutically as an adjuvant to disease detection and monitoring in areas such as prenatal cell-free DNA (cfDNA) screening and cancer. Although the majority of current liquid biopsy techniques focus on DNA from blood, DNA from urine also conveys important genetic information from the human body. Studies have revealed that urine cfDNA can potentially be utilized for bladder cancer detection and monitoring. Urine samples can be quickly and painlessly collected; however, because urine DNA is readily damaged, maintaining high-quality cfDNA in urine is challenging. In this study, we developed a robust and user-friendly urine preservation method to maintain integrity of cfDNA in urine and to facilitate the transportation of urinary samples to clinical laboratories.
Methods: Urine samples from 10 donors were collected with Predicine’s proprietary urine preservation method. Two urine samples were spiked with BL21(DE3) E.coli competent cells to simulate the urinary tract infection. All samples were incubated at 4°C, room temperature, and 37°C to mimic extreme storage or transporting environments. cfDNA was extracted after incubation of 0, 4, and 7 days and measured by Qubit and Fragment analyzer. Extracted DNA was also quantified by ddPCR with two markers: EGFR as an endogenous DNA marker and ARv7 (synthetic oligos containing intron-deleted sequence, post-incubation spike-in) as an inner control for the extraction and detection assays. Whole-genome sequencing was conducted to measure the quality of the cfDNA and bacterial contamination. Other commercial urine preserve buffers are used as control.
Results: cfDNA derived from urine stored in Predicine’s proprietary urine preservation buffer remained stable till day 7 at different temperatures. There was no significant change in normalized copies of EGFR in the treated urine samples for up to 7 days. cfDNA yield was considerably higher in the novel preservation buffer than that using commercial buffers. Furthermore, the mapping rates in bacteria-contaminated samples were significantly improved.
Conclusion: Predicine’s proprietary urine preservation method demonstrated excellent urine cfDNA preserving capacity at various temperatures for at least 7 days, outperforming commercially available preserve buffers in terms of DNA integrity and yield. The innovative technique offers an easy-to-use solution for obtaining high-quality urine cfDNA for clinical use.
Citation Format: jie zhang, fang liu, min wang, cuishan deng, wei mo, seng tao, amy wang, shidong jia, zhixin zhao, pan du, binggang xiang, shujun luo. Development of a novel urine cell-free DNA preservation method for molecular profiling in genitourinary tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 245.