Development of Oral Immunotherapy against Clostridioides difficile Transmission

Clostridioides difficile is a gram-positive bacterial pathogen responsible for severe gastrointestinal infections, often leading to life-threatening complications, particularly in healthcare settings. C. difficile produces environment resistant spores that are the major source of transmission. Both symptomatic patients and asymptomatic carriers in hospitals and nursery homes serve as reservoirs for C. difficile spores. Current preventive strategies for C. difficile infections (CDI) are limited and there is no treatment available to reduce spore production and prevent the spread of this pathogen. We have recently identified a panel of single-domain variable fragments of heavy-chain only antibodies (VHHs) against conserved Cwp84, a surface-associated cysteine protease important for bacterial colonization and sporulation. We found that a VHH heterodimer could inhibit the sporulation in vitro. Most importantly, oral administration of mice with the engineered yeast secreting the bi-specific antibody during the CDI not only reduced disease severity, but also significantly diminished spore shedding. In this project, we propose to develop a novel probiotic yeast-based immunotherapy that utilizes the engineered probiotic yeast as a vehicle to deliver the neutralizing antibodies against Cwp84 in gut to reduce C. difficile colonization and spores shedding subsequently preventing transmission. This study will further screen for lead bi-specific VHH antibodies against Cwp84 and generate the lead yeast strains expressing the anti-Cwp84 antibodies. Several CDI animal models will be employed to assess the efficacy of the lead strains on reducing disease symptoms, bacterial colonization, spore shedding and preventing disease transmission. The successful completion of this project will result in a novel probiotic yeast-based immunotherapy that can effectively prevent the transmission of C. difficile. This project addresses a critical gap in the prevention of C. difficile transmission. The development of a probiotic yeast-based immunotherapy represents a novel and potentially transformative strategy that could significantly reduce the incidence of CDI, improve patient outcomes, and decrease healthcare costs associated with C. difficile infections. Project Number: 5R01AI190438-02 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of Allergy and Infectious Diseases (NIAID) | Principal Investigator: Yongrong Zhang (+1 co-PI) | Institution: UNIVERSITY OF MARYLAND BALTIMORE, BALTIMORE, MD | Award Amount: $719,022 | Activity Code: R01 | Study Section: Drug Discovery and Molecular Pharmacology A Study Section [DMPA] View on NIH RePORTER: https://reporter.nih.gov/project-details/5R01AI19043802