Lithium Attenuates Graft-Versus-Host Disease Via Effects on the Intestinal Stem Cell Niche

Abstract

Allogeneic hematopoietic stem cell transplantation (HCT) is a curative therapy for the majority of hematological malignancies but is limited by graft-versus-host disease (GVHD) and opportunistic infections. Severe acute GVHD in the gastrointestinal (GI) tract is a major determinant of mortality and causes progressive epithelial and crypt loss leading to denudation of intestinal mucosa. Since the current approaches to prevent severe GVHD to date focus on the immune suppression of T cells with calcineurin inhibitors, anti-metabolites or alkylating agents, therapeutic strategies that protect intestinal epithelium from immune injury represents an attractive alternative approach. The renewal of intestinal stem cells (ISC) and their progeny, intestinal epithelial cells (IEC), is critical for the maintenance of gut integrity after transplantation. Multiple signaling pathways including WNT-β-Catenin signaling regulate this process. Lithium inhibits glycogen synthase kinase-3 (GSK-3), a pluripotent kinase that antagonizes WNT signaling, resulting in cellular proliferation and epithelial regeneration. In a clinical pilot study conducted at Fred Hutchinson Cancer Research Center (FHCRC) and the University of Washington, twenty patients with steroid-refractory gut GVHD and mucosal denudation at endoscopy were treated with oral lithium carbonate in addition to second line therapy. Ten of the 20 patients (50 %) had a complete remission, and 7 (35 %) survived more than 2 years 1. These results were encouraging since only 10% of the patients with steroid-refractory stage 3-4 intestinal GVHD at FHCRC survived for more than 2-years historically1. We thus investigated the activity of lithium in gastrointestinal GVHD in preclinical models. The administration of oral lithium chloride (LiCl) from 2 days before HCT to day 7 after T cell-replete HCT (B6 donors → B6D2F1 recipients) significantly decreased the GVHD-related mucosal injury in the ileum (pathology score: 7.4 ± 0.6 in control vs 4.6 ± 0.6 in LiCl (mean ± sem), p = 0.0049) but not the colon (13.4 ± 0.5 in control vs 12.4 ± 0.9 in LiCl, p > 0.05). We undertook multispectral immunofluorescence analysis in a second BALB/c → B6 system where recipient ISC were marked by Lgr5-driven GFP. LiCl significantly increased the frequency of Paneth cells (6.14 ± 0.4 % in control vs 15.0 ± 1.5 % in LiCl (mean ± sem), p = 0.0031) and their antimicrobial lysozyme content (% cytoplasm occupancy: 61.1 ± 0.7 % in control vs 65.0 ± 0.8 % in LiCl (mean ± sem), p = 0.0055). Lgr5+ ISCs in the ileum were not quantitatively different at this early timepoint. Given that Paneth cells are primarily distributed in the small intestine rather than the colon, these data explain the protection from GVHD afforded by LiCl in the ileum but not the colon. Since ISC regeneration is ultimately essential to repair severe ulceration of the epithelium, we examined the renewal of ISC at later timepoints. Compared to the vehicle control, LiCl treatment significantly increased the percentage of Lgr5+ ISCs (2.33 ± 0.2 % in control vs 4.80 ± 0.6 % in LiCl (mean ± sem), p = 0.0129) in the ileum on day 10. LiCl did not alter the frequencies of CD4+ and CD8+ T cells infiltrating the ileum. These findings suggest that LiCl exerts a protective effect in GVHD by promoting Paneth cell function and subsequent ISC regeneration. Our findings provide further rationale for studies of GSK inhibition, direct WNT activation, and other agents such as IL-22 and IL-28 that provide support of the ISC niche during GVHD, as a complement to immune suppression. Reference:Steinbach G, Hockenbery DM, Huls G, et al. Pilot study of lithium to restore intestinal barrier function in severe graft-versus-host disease. PloS one. 2017;12(8):e0183284.