Abstract Microsatellite instability (MSI) is associated with high immunogenicity in tumors due to the abundance of neoantigens, making MSI cancers particularly responsive to immune checkpoint blockade (ICB) therapy. However, a substantial proportion of patients with MSI gastrointestinal (GI) adenocarcinomas do not benefit from ICB, and non‐invasive biomarkers for monitoring treatment response are lacking. This study investigated the utility of MSI detection in extracellular vesicle (EV) DNA as a liquid biopsy‐based approach for therapy monitoring in patients with advanced MSI GI cancers undergoing ICB therapy and compared it with the analysis of cell‐free (cf) DNA. Blood ( n = 52) and tumor samples ( n = 16) were collected from 19 patients before and during therapy. Plasma from 18 patients and 30 healthy controls was analyzed for MSI using four diagnostic mononucleotide markers in both EV DNA and cell‐free DNA (cfDNA). MSI was detected in EV DNA in 8 out of 18 patients and in cfDNA in 9 out of 18 patients, with high concordance (93%) between the two approaches. None of the healthy controls showed MSI signals. MSI detection was more frequent before therapy initiation compared to during therapy time points, and a transition from MSI to non‐MSI status during treatment was associated with clinical benefit or objective response. This switch often occurred before the first staging at 3 months. Thus, MSI analysis in EV DNA is a promising minimally invasive tool for real‐time monitoring of ICB therapy response in MSI GI cancer patients, yielding results comparable to cfDNA while providing handling advantages, and warrants further validation in larger cohorts.
Ahadova et al. (2026) studied this question.