Abstract 3738: Monitoring chemotherapy response status of colorectal cancer patients using liquid biopsy samples
Authors: Bartak Barbara Kinga , Fodor Tamás , Kalmár Alexandra , Nagy Zsófia Brigitta , Zsigrai Sára , Szigeti Krisztina Andrea , Kothalawala William , Valcz Gábor , Igaz Péter , Takács István , Dank Magdolna , Molnár Béla 
Published: June 2022

Abstract
Background and aims: The early detection of colorectal cancer (CRC) recurrence and the monitoring of therapeutic response are crucial steps in the determination and modification of treatment strategies. The currently used gold standard tumor markers such as CEA and CA 19-9 and the different imaging techniques have several limitations in many cases. Therefore, we aimed to establish a liquid biopsy-based approach for tracking tumor dynamics in post-operative non-metastatic (n=32) and metastatic (n=23) CRC patients.
Methods: Blood samples were collected from each patient before and during chemotherapy, and finally, patients were classified according to disease outcome. Longitudinal investigations of the total amount, global and local (SFRP2 and SDC2 genes) DNA methylation pattern of cell-free DNA (cfDNA) fraction were performed. The plasma concentration of homocysteine was also determined, as it is one of the main components of the DNA methylation process, and its level defines the methylation potential. We examined how the parameters mentioned above were affected depending on the different therapy responses.
Results: The average cfDNA amount was significantly higher (p&lt;0.05) in patients with recurrent cancer (30.4±17.6ng) and progressive disease (PD) (44.3±34.5ng) than individuals who achieved remission (REM) (13.2±10.0ng). More than 10% elevation of cfDNA from first to last sample collection was detected in 92% of PD patients, while reduced cfDNA concentration was observed in 67% of the non-metastatic CRC patients with REM. An effective differentiation was detected between patients achieving remission and showing tumor progression based on cfDNA level with 94% sensitivity and 81% specificity. The average global cfDNA methylation was determined by bisulfite pyrosequencing analysis of long interspersed nuclear element-1 (LINE-1), and it was significantly lower (p&lt;0.05) in the PD group compared to people with remission (71.0±6.7% vs. 78.9±2.0%). Methylation level changes between the study beginning and end indicated a decline (75.5±3.4% vs. 68.2±8.4%) in PD; in contrast, we found a reverse change in remission. The mean relative change of homocysteine concentration revealed an opposite trend with the global DNA methylation suggesting a linkage between these parameters, as it showed an increase (+12.5%) in the case of PD and a decrease (-15.8%) in patients with REM. Regarding local DNA methylation, SFRP2 and SDC2 genes revealed higher promoter hypermethylation in the PD set compared to patients with remission.
Conclusion: Our study offers the possibility to monitor the therapeutic response during chemotherapy with a minimally-invasive blood-based method that combines the analysis of cfDNA, its global and local DNA methylation pattern, and homocysteine level.
Citation Format: Barbara Kinga Bartak, Tamás Fodor, Alexandra Kalmár, Zsófia Brigitta Nagy, Sára Zsigrai, Krisztina Andrea Szigeti, William Kothalawala, Gábor Valcz, Péter Igaz, István Takács, Magdolna Dank, Béla Molnár. Monitoring chemotherapy response status of colorectal cancer patients using liquid biopsy samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3738.