A research group composed of Professor Satoshi Uematsu and Associate Professor Kosuke Fujimoto of Graduate School of Medicine at Osaka Metropolitan University; and Professor Seiya Imoto of the Human Genome Center, Institute of Medical Science at the University of Tokyo jointly discovered highly virulent Enterococcus faecalis strains in the intestinal tract of hematopoietic stem cell transplant recipients. The research group revealed their involvement in the development of graft-versus-host disease (GVHD), a serious complication of hematopoietic stem cell transplantation. They also confirmed that administration of a bacteriophage-derived bacteriolytic enzyme, which was identified via metagenomic analysis, prevented the exacerbation of GVHD and significantly improved mortality in model mice. The enzyme is potentially usable as a treatment for GVHD. The results were published in the international journal Nature on July 11.
In hematopoietic stem cell transplantation for leukemia treatment, immune cells derived from the donor's hematopoietic stem cells may attack the recipient's organs as if they were foreign, leading to GVHD, which can be fatal in severe cases. Studies have shown that changes in the gut microbiome may trigger GVHD. A cohort study in 2019 reported that an increase in Enterococcus spp. was associated with a decrease in survival. Previously, the research group studied bacteriophages (phages) that infect intestinal bacteria to control intestinal bacteria.
The research group constructed a metagenomic analysis pipeline of the gut microbiome and viromes at the Institute of Medical Science at the University of Tokyo. They established a method of revealing infection relationships even when bacteria and phages cannot be cultured. Uematsu and Fujimoto also hold positions in the Division of Metagenome Medicine, Human Genome Center, Institute of Medical Science, the University of Tokyo.
In this study, the research group conducted a longitudinal fecal analysis of 46 hematopoietic stem cell transplant recipients hospitalized at Osaka Metropolitan University Hospital and observed increases in Enterococcus spp. in 30 of the 46 cases. The examination of isolated/cultured bacteria revealed the presence of highly virulent E. faecalis secreting cytolysin, which can lyse human erythrocytes and other cells. This resident bacterium is present in small amounts in healthy individuals but is significantly elevated in patients. Moreover, the incidence of GVHD was significantly higher in patients from whom highly virulent E. faecalis was isolated.
Many genes involved in biofilm formation were identified through gene analysis using DNA extracted from isolated highly virulent E. faecalis. Although the bacterium was not drug-resistant, the study explored whether it could evade antimicrobials and other agents administered during transplantation therapy by forming biofilms, thereby causing GVHD. A biofilm is a conglomerate comprising a large number of microorganisms and substances they produce. By forming biofilms, bacteria protect themselves against external factors such as drugs and environmental changes.
When bone marrow transplantation was performed in germ-free mice colonized with highly virulent E. faecalis from patients, nonvirulent E. faecalis, and Escherichia coli, only the group with highly virulent E. faecalis began to die after 5 days. No mice in other groups died even after 25 days, indicating that highly virulent E. faecalis triggered GVHD. Therefore, the research group investigated whether phages infecting E. faecalis could eradicate this bacterium and suppress GVHD.
Phage-derived bacteriolytic enzyme sequences were identified from E. faecalis metagenomic data. The phage-derived enzyme that breaks the membrane of E. faecalis was identified as a novel bacteriolytic enzyme, "endolysin." The research team also confirmed that "endolysin" lysed E. faecalis only and had no effects on other intestinal bacteria such as Enterococcus spp. and Escherichia coli. Biofilms were found to be lysed when "endolysin" was added to E. faecalis-derived biofilms in vitro. They also confirmed that administration of the enzyme significantly improved mortality rates in GVHD model mice in which germ-free mice were colonized with highly virulent E. faecalis of patient origin as well as in mice transplanted with patient feces. "Endolysin" is believed to have improved the pathophysiology of GVHD by lysing the entire biofilms derived from highly virulent E. faecalis.
Fujimoto said, "Moving forward, we would like to launch a nationwide-scale cohort study with the initial aim of understanding the target population for drug discovery. We hope this will lead to new prophylactic and therapeutic methods for GVHD."
Journal Information
Publication: Nature
Title: An enterococcal phage-derived enzyme suppresses graft-versus-host disease
DOI: 10.1038/s41586-024-07667-8
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