Peripheral blood cell-free DNA testing as a predictor for relapse post-allogeneic stem cell transplant for AML.

6555 Background: Allogeneic stem cell transplantation (allo-SCT) is a curative option for acute myelogenous leukemia (AML), but relapse is a challenge. Monitoring minimal residual disease post-transplant through tumor-derived circulating cell-free DNA (cfDNA) in peripheral blood (PB) and bone marrow is an emerging strategy. Persistent mutations in cfDNA may be prognostic indicators of relapse. Methods: This single-center retrospective study included 120 AML patients who received allo-SCT from 2018 to 2022, with PB cfDNA collected between Days 30-200. Samples were analyzed by commercial assays (Liquid Trace or Hematology Profile Plus), that use next generation sequencing, Sanger Sequencing, and fragment length analysis to identify molecular abnormalities in DNA of 179 genes associated with hematologic neoplasms. cfDNA positivity was determined by identifying gene amplifications, deletions, single nucleotide variations and indels, including reported variant allele frequency (VAF) above 0. cfDNA negativity was defined by absence of genomic alterations. The primary endpoints were the association of cfDNA presence with overall survival (OS) and relapse-free survival (RFS). A secondary endpoint was the association of mutation risk (adverse/intermediate) with OS and RFS. Results: Patients were grouped by cfDNA presence at Day 45±15 (n=30, median survival time 1.173 years) and Day 150±50 (n=90, median survival time 1.4822 years). Kaplan-Meier analysis revealed that patients positive for PB cfDNA at Day 150±50 had significantly worse OS (p<0.0001) and RFS (p<0.0001) compared to cfDNA negative. Similarly, cfDNA positivity at Day 45±15 also correlated with worse OS (p<0.01) and RFS (p<0.0007). Regarding mutation risk, adverse mutations at Day 150±50 were linked to worse OS (p<0.0001) and RFS (p<0.0001). Multivariate analysis revealed that adverse-risk mutations were significantly associated with relapse (odds ratio [OR] 29.48, 95% CI 4.306-350.4, p<0.002), RFS (hazard ratio [HR] 12.62, 95% CI 3.541-44.35, p<0.0001), and OS (HR 19.24, 95% CI 5.242-75.73, p<0.0001). Intermediate-risk mutations also correlated with relapse (OR 13.27, 95% CI 2.818-89.91, p<0.002), RFS (HR 9.325, 95% CI 2.748-35.25, p<0.0005), and OS (HR 11.48, 95% CI 3.279-45.61, p<0.0002). Transplant age, donor type, CMV status, and GvHD regimen were not statistically significant. Conclusions: This study demonstrates that cfDNA detection in PB post-allo-HSCT is strongly associated with increased relapse and mortality in AML patients. Persistent high-risk mutations correlate with increased risk of relapse and poor survival outcomes. These findings highlight the potential of PB cfDNA as a predictive marker, potentially enabling earlier intervention to alter post-transplant treatment strategies.