In the past year the Mucosal Immunity Section has been engaged in a number of research studies involving both inflammatory bowel disease and common variable immunodeficiency. In the area of IBD, we have conducted studies of IBD associated with LRRK2 risk polymorphisms, as described in detail in another annual report. In these studies we have established that the polymorphism is associated with increased levels of LRRK2 and this elevation is accompanied by increased gut inflammation in an induced colitis model driven by Dectin-1 induced NF-kappaB responses. Several observations derived from these studies have direct clinical significance. The first is that cells derived from patients who do not bear the LRRK2 risk polymorphism exhibit reduced pro-inflammatory cytokine responses in vitro when the cells are exposed to inhibitors of the kinase activity of LRRK2. The second is that induced colitis of normal mice or mice with elevated levels of LRRK2 is inhibited by administration of various LRRK2 inhibitors. These observations suggest that treatment of patients with anti-LRRK2 can have a therapeutic effect in all patients with IBD regardless of their LRRK2 status. In other IBD studies we have investigated Crohn's disease-like colitis occurring in patients with chronic granulomatous disease (CGD). These studies include the evaluation of cytokine production in the inflamed mucosa of CGD patients. In the studies conducted so far, Crohn's disease in CGD patients and conventional patients appear to be quite similar. In addition, even CGD patients without symptoms of overt Crohn's disease exhibit immunologic abnormalities suggest the presence of covert disease. This has led us to conclude that CGD is, in effect, a risk factor for the development of Crohn's disease that manifests itself in all CGD patients. Finally, we have designed and written a clinical protocol for a study of the safety and and immunologic effects of treatment of patients with moderately severe patients with Crohn's disease with vorinostat, an HDAC inhibitor. This clinical trial will be partially funded by a CRADA with Merck and will involve 26 patients who have failed other forms of Crohn's disease therapy. A wide array of studies to evaluate the immunologic effects of HDAC inhibitor therapy will be performed, including extensive studies of the drug's effect on regulatory T cells present in gut biopsy specimens. Currently, the Clinical Protocol has been written has obtained both NIAID IRB and FDA approval. Patients will be enrolled imminently. In the area of CVID we have continued to focus on gastrointestinal manifestations of this heterogeneous disease, i.e., patients with a CVID-entropathy characterized by diarrhea, malabsorption and villous atrophy. In prior studies of CVID we established that the CVID enteropathy is driven by cells producing IFN-gamma. In addition, we collaborated with Drs. Andriy Morgan and Natalia Stulzhenko in the Laboratory of Cellular and Molecular Immunology in studies of CVID that included microarray analyses of biopsy tissue of CVID patients with enteropathy showing that CVID enteropathy is characterized by up-regulation of genes related to various interferons. Based on this prior work, we have applied for and have obtained a U01 NIH cooperative research grant (PAR-13-029) with Drs. Morgan and Shulzhenko (now at Oregon State University) to expand on these findings. This Grant is entitled Anti-IL-12p40 Treatment of CVID Enteropathy: Gene Expression and Microbiota Analysis, and is centered around a clinical study of the safety and efficacy of a FDA-approved monoclonal anti-IL-12p40 in the treatment of CVID enteropathy. In addition, this Grant includes a study of the effect of this biologic agent on gene expression of inflammatory cytokines and on the microbiota in patients. In an initial phase of these studies, we have treated three patients with CVID enteropathy with a single dose of anti-IL-12p40 (270mg sc of Stelara)and have found that in all three patients experienced clinical improvement with respect to diarrhea. In one of the two patients, there was long-term and dramatic cessation of diarrhea, whereas in the other two patients diarrhea was greatly reduced but rather transient. However, improvement in objective measure of malabsorption such as stool fat assessment was observed in none of the patients. As a result of this experience we believe that a greater therapeutic effect can be obtained with a multiple dose regimen consisting of 270mg induction dose of Stelara followed by two addition 180mg doses over a six month period. Consequently, we applied to and have receive permission from the NIAID IRB to treat patients in this manner. Presently 4-5 patients have been enrolled in this more prolonged treatment protocol and we have noted initial success in treatment of the diarrhea and malabsorption. We are therefore optimistic that sustained treatment with anti-IL-12p40 can reverse both the clinical and immunologic concomitant of CVID enteropathy. Meanwhile stool and tissue specimens of CVID patients with and without enteropathy have been sent to Drs. Morgun and Shulzhenko at Oregon State University for microbiome analysis before and after anti-IL-12p40 treatment as well as identification of a possible organism causing the enteropathy. A major outcome of this collaborative study has emerged from analysis of patient immunoglobulin levels in gut specimens. We have found that those patients with enteropathy have little or no expression of IgA in intestinal biopsies whereas those patients without enteropathy have only moderately reduced levels of IgA. Moreover, this deficiency does not extend to mucosal IgG levels. This finding indicates that enteropathy occurs in the subset of CVID patients with severe mucosal B cell immunodeficiency affecting the mucosal immunoglobulin, IgA. A second major outcome from these studies is the finding the duodenal specimens from patients with enteropathy contain Acinetobacter baumanii, an organism that, acting as an pathobiont in immunodeficient patients, induces an excessive IFN-gamma synthesis and resultant enteropathy. In other studies of CVID patients we have continued to evaluate the course and characteristics of nodular regenerative hyperplasia (NRH), a complication of CVID that we extensively defined and categorized defined previously. This compilation of CVID is associated with the presence of increased concentrations of CD8+ T cells in liver specimens and is likely to be due to an abnormal immunologic response directed toward liver cells. Since some of the patients with CVID enteropathy also have NRH, we are evaluating if anti-ILp40 administration has an ameliorative effect on concomitant NRH.
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