Abstract Introduction: The gut microbiome has emerged as a biomarker of response to immune checkpoint inhibitor (ICI). Fecal microbiota transplantation (FMT) in combination with ICI has shown therapeutic promise with 6 positive trials. Polyethylene glycol (PEG)–based bowel preparation is routinely performed prior to FMT. However, its impact on gut microbiome and potential influence on ICI outcomes are unknown, creating uncertainty on whether placebo arms should receive bowel preparation. Methods: We conducted a prospective trial in 20 healthy volunteers (HV) (NCT06831539) who consumed 4L of PEG, and fecal samples were collected at baseline and multiple timepoints post-PEG. Shotgun metagenomics was analyzed using the MetaPhlAn4 and findings were validated by qPCR. To model PEG-induced gut microbiome disruption, mice were transiently exposed to 50% or 75% PEG for 12 days with longitudinal stool collection. To assess the effect of PEG-induced dysbiosis on ICI efficacy, tumor-bearing mice (MCA-205 or HKP1) were treated with anti-PD-1 or isotype under water, PEG, or antibiotics exposure. Tumor-infiltrating lymphocytes (TILs) were profiled by flow cytometry, and colonic tight-junction integrity was assessed using RT-qPCR and immunofluorescence. Results: In the HV cohort, median age was 30 years and 5 (25%) were male. Metagenomics revealed a transient decrease in α diversity (Shannon index, p=0.07) at the time of bowel preparation and most patients recovered α and β diversity at 3 days post-PEG, confirmed by qPCR. At 3 days post-PEG, 4 HV (20%) experienced a decrease in α diversity and change in β diversity from baseline, with complete recovery at 1 week except for one HV. All participants fully recovered at the 2 week timepoint. In mice, transient exposure to PEG led to recovery of α and β diversity at 3 days, and did not interfere with anti-PD-1 response. Conversely, continuous exposure to PEG significantly and persistently altered the gut microbiome composition, and impaired anti-PD-1 response in MCA-205 and HKP1 tumor-bearing mice, with reduced memory CD8+ and ICOS+CD4+ T-cell populations in TILs of PEG-treated mice compared to water controls. PEG significantly decreased colonic tight junction protein expression, though unlike antibiotics the detrimental effect of PEG on ICI response was independent of MAdCAM-1. To examine the relevance of prolonged PEG exposure in patients, clinical data from 197 advanced NSCLC patients showed that 17g PEG was associated with shorter progression-free survival to ICI (5.5 vs 11 months, HR 1.5, p=0.03), confirmed in multivariable analysis (HR 1.8, p=0.005). Conclusion: PEG bowel preparation causes transient disruption of the gut microbiome, with most healthy volunteers showing rapid recovery. However, a subset did not return to baseline by day 3, highlighting the need for caution when using PEG in placebo arms of microbiome-focused RCTs, specifically, monitoring the gut microbiome post-PEG, and allowing for adequate recovery prior to ICI start. Citation Format: Yongjia Hu, Alyse Filin, Diana Rusu, Sebastien Hunter, Diogjena Prifti, Xiaojing Dong, Meriem Messaoudene, Mayra Ponce, Jade Maillou, Wiam Belkaid, Julie Malo, Bertrand Routy, Arielle Elkrief. Impact of polyethylene glycol bowel preparation on the gut microbiome composition and response to immune checkpoint inhibition abstract. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr C065.
Hu et al. (2026) studied this question.