Fecal microbiota transplantation for refractory immune checkpoint inhibitor-associated colitis

Abstract

We report the first case series of immune checkpoint inhibitors (ICI)-associated colitis successfully treated with fecal microbiota transplantation, with reconstitution of the gut microbiome and a relative increase in the proportion of regulatory T-cells within the colonic mucosa. These preliminary data provide evidence that modulation of the gut microbiome may abrogate ICI-associated colitis.

Figure 1:

Endoscopic changes and characterization of colonic mucosal infiltrate throughout clinical course for Patient 1 (a-c) and Patient 2 (d-f). (a) Changes in colonic mucosa as assessed by full colonoscopy. Near the time of diagnosis (row 1), multiple large ulcers and diffuse inflammatory exudate are present (in the distal 40cm of the colon only, with the normal appearing proximal colon) and remain despite months of treatment with steroids and biologic immunosuppressive agents (steroid + 2 doses infliximab + 1 dose vedolizumab) (row 2). Approximately one month after FMT (row 3), colonic mucosa exhibits grossly normal vasculature, minimal patchy erythema, and near-complete healing of prior ulcers. Yellow arrows point to ulcerative lesions. This patient had full colonoscopic evaluations that examined every segment of the colon (ascending, transverse, descending, sigmoid and rectum). Endoscopy was performed once at each time point. Given the qualitative nature of endoscopic data collection and inability to provide true statistical analyses, we chose to include multiple other representative photos from the same colonoscopic evaluations. Additional representative photos are presented in Supplemental Figure 4a. (b) Immunohistochemical analysis of mucosal biopsies of the colon/rectum prior to, and following, FMT. A single slide representative of the endoscopic biopsy specimen as a whole was stained for each patient for each time point. Representative slides from additional time points are included in Supplemental Figure 5a. (c) Analysis of changes in the density immune cell subsets (CD8 red squares, CD4 blue circles, FOXP3 black triangles) over time, expressed as a fold change from baseline based on total densities of cells expressing these markers (absolute densities are presented in Supplemental Figure 6a). Timepoints include time of diagnosis, prior to FMT, following steroids and biologic immunosuppression and following FMT. Date of FMT is represented by dotted vertical line and is designated Day 0. These data represent the average cell density from 4 regions of interest (ROIs) per sample (single slide per patient at each time point) with each ROI measuring 500 μm x 500 μm for a total of 0.25 mm². We report the mean # IHC-positive cells/mm² divided by the mean # IHC-positive cells/mm² at baseline. (d) Changes in colonic mucosa as assessed by full colonoscopy. Near the time of diagnosis (row 1), multiple large ulcers and inflammatory exudate is present (throughout the entire colon) and remains after unsuccessful treatment with steroids and biologic immunosuppressive agents (steroid + 2 doses infliximab + 4 doses vedolizumab) (row 2). There is notable improvement following first FMT (row 3) but residual ulcers remain. Following second FMT (row 4) we note near complete resolution of all ulcerative lesions. Again full endoscopic examinations were performed, once for each time point. Additional representative photos from the colonoscopic evaluation are shown in Supplemental Figure 4b. (e) Immunohistochemical analysis of mucosal biopsies of the colon/rectum taken prior to first FMT and following first FMT. A single slide representative of the endoscopic biopsy specimen was stained for each patient for each time point. Representative slides from additional time points are included in Supplemental Figure 5b. (f) Analysis of changes in the density immune cell subsets (CD8 red squares, CD4 blue circles, FOXP3 black triangles) over time, expressed as a fold change from baseline based on total densities of cells expressing these markers (absolute densities are presented in Supplemental Figure 6b). These data represent the average cell density from 4 regions of interest (ROIs) per sample (single slide per patient at each time point) with each ROI measuring 500 μm x 500 μm for a total of approximately 1 mm². We report the mean # IHC-positive cells/mm² divided by the mean # IHC-positive cells/mm² at baseline. Date of first (Day 0) and second FMT (Day 67) are represented by dotted vertical lines.

Figure 2:

Microbiome analysis of patient and donor intestinal bacteria by 16S deep sequencing. The patients’ stool microbiomes were longitudinally sampled at indicated time points before and after FMT, along with samples from the FMT donor. Between 3380 and 42,776 sequences were obtained for each sample (average 10,003). (a) Alpha diversity, quantified by the inverse Simpson index after rarefying to 3000 sequences, as well as total observed OTU numbers, was evaluated for patient and FMT donor samples. (b) Using principal coordinate analysis of unweighted UniFrac distances, microbiome samples from (a) are depicted in space with more similar samples located closer together. (c) Bacterial 16S sequences from samples in (a) were classified by origin (unique to patient baseline, unique to donor, present in both patient baseline and donor, or absent in both patient baseline and donor). (d) Sequences were classified by taxonomy at the Class level. (e) Changes in the abundances of top varying 10 bacterial genera over time.