Abstract 5893: Feasibility of DNA analysis in peritoneal fluid from patients with peritoneal carcinomatosis
Authors: Muhammad Talha Nawaz, Muhammad Talha Waheed, Kirsten Dennison, Bradon McDonald, Stephanie McGregor, Syed Nabeel Zafar, Mustafa Raoof, Muhammed Murtaza
Journal: Cancer Research (2025)

Abstract Introduction: Gastrointestinal (GI) cancers often metastasize to the peritoneum. Current diagnostic tools for peritoneal carcinomatosis (PC) show limited accuracy. Reliable biomarkers to detect and monitor treatment response for PC are urgently needed. While circulating tumor DNA in plasma is a promising cancer biomarker, contribution of tumor DNA from PC into blood is minimal. We hypothesize that peritoneal fluid tumor DNA (ptDNA) may be useful for PC detection and response monitoring. Currently, there are no established laboratory methods for ptDNA analysis. In this study, we evaluate the feasibility of two DNA extraction methods using peritoneal fluid (PF) samples. Methods: We evaluated the performance of two cell-free DNA extraction kits based on magnetic beads (MagMax) or silica columns (QIAamp) using commercially obtained PF from two individuals, at 1 mL and 4 mL starting volumes. Based on these results, we evaluated the optimal method further using PF samples from 24 patients with PC from GI cancers (biliary tract, appendiceal, and colorectal primary) undergoing surgery or paracentesis at City of Hope National Medical Center. PF samples were collected immediately before the start of the procedure. We extracted DNA using 4 mL inputs, eluted in 30 µL buffer and assessed using electrophoresis (TapeStation) and fluorometry (Qubit). Results: In commercially obtained ascites samples, adequate DNA yield and quality was obtained using both kits. Compared to QIAamp, mean DNA concentration was slightly lower using MagMax (24.9 ng/mL [range 13.9 - 35.9] vs. 25.7 ng/mL [range: 16.1-35.3; p=0.91). Similarly, mean fragment length was also shorter using MagMax compared to QIAamp (287.5 bp [range 202-350] vs. 294.3 bp [range: 239-348; p=0.80). In contrast to our prior experience with plasma, peritoneal fluid DNA was observed at higher concentrations (3.7-fold higher) and longer fragment lengths. Based on these results and processing considerations, we selected MagMax kit to evaluate patient PF samples. We observed similar fragment lengths (mean: 281.5 bp, range: 136-369), with concentrations ranging from 0.80 ng/mL to 105 ng/mL in PF (mean: 20.6 ng/mL). Conclusions: Our results demonstrate the feasibility of DNA analysis using PF samples in patients with PC Furthermore, the yield and quality of DNA extracted is adequate for downstream molecular analyses. We were able to detect differences in PF vs. plasma DNA and observe inter-patient variability in concentration and fragmentation. On-going work will evaluate detection and quantification of peritoneal tumor-specific DNA as a biomarker for PC detection and monitoring. Citation Format: Muhammad Talha Nawaz, Muhammad Talha Waheed, Kirsten Dennison, Bradon McDonald, Stephanie McGregor, Syed Nabeel Zafar, Mustafa Raoof, Muhammed Murtaza. Feasibility of DNA analysis in peritoneal fluid from patients with peritoneal carcinomatosis [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 5893.