Administration of Panobinostat Is Associated with Increased IL-17A mRNA in the Intestinal Epithelium of HIV-1 Patients

Intestinal CD4+ T cell depletion is rapid and profound during early HIV-1 infection.This leads to a compromised mucosal barrier that prompts chronic systemic inflammation.The preferential loss of intestinal T helper 17 (Th17) cells in HIV-1 disease is a driver of the damage within the mucosal barrier and of disease progression.Thus, understanding the effects of new therapeutic strategies in the intestines has high priority. Histone deacetylase (HDAC) inhibitors (e.g., panobinostat) are actively under investigation as potential latency reversing agents in HIV eradication studies. These drugs have broad effects that go beyond reactivating virus, including modulation of immune pathways. We examined colonic biopsies from ART suppressed HIV-1 infected individuals (clinicaltrials.gov: NCT01680094) for the effects of panobinostat on intestinal T cell activation and on inflammatory cytokine production. We compared biopsy samples that were collected before and during oral panobinostat treatment and observed that panobinostat had a clear biological impact in this anatomical compartment. Specifically, we observed a decrease in CD69+ intestinal lamina propria T cell frequency and increased IL-17A mRNA expression in the intestinal epithelium. These results suggest that panobinostat therapy may influence the restoration of mucosal barrier function in these patients

Administration of Panobinostat Is Associated with Increased IL-17A mRNA in the Intestinal Epithelium of HIV-1 Patients

Intestinal CD4+ T cell depletion is rapid and profound during early HIV-1 infection. This leads to a compromised mucosal barrier that prompts chronic systemic inflammation. The preferential loss of intestinal T helper 17 (Th17) cells in HIV-1 disease is a driver of the damage within the mucosal barrier and of disease progression. Thus, understanding the effects of new therapeutic strategies in the intestines has high priority. Histone deacetylase (HDAC) inhibitors (e.g., panobinostat) are actively under investigation as potential latency reversing agents in HIV eradication studies. These drugs have broad effects that go beyond reactivating virus, including modulation of immune pathways. We examined colonic biopsies from ART suppressed HIV-1 infected individuals (clinicaltrials.gov: NCT01680094) for the effects of panobinostat on intestinal T cell activation and on inflammatory cytokine production. We compared biopsy samples that were collected before and during oral panobinostat treatment and observed that panobinostat had a clear biological impact in this anatomical compartment. Specifically, we observed a decrease in CD69+ intestinal lamina propria T cell frequency and increased IL-17A mRNA expression in the intestinal epithelium. These results suggest that panobinostat therapy may influence the restoration of mucosal barrier function in these patients

Infliximab Induces Clonal Expansion of γδ-T Cells in Crohn's Disease: A Predictor of Lymphoma Risk?

BACKGROUND: Concominant with the widespread use of combined immunotherapy in the management of Crohn's disease (CD), the incidence of hepato-splenic gamma-delta (γδ)-T cell lymphoma has increased sharply in CD patients. Malignant transformation of lymphocytes is believed to be a multistep process resulting in the selection of malignant γδ-T cell clones. We hypothesised that repeated infusion of anti-TNF-α agents may induce clonal selection and that concurrent treatment with immunomodulators further predisposes patients to γδ-T cell expansion. METHODOLOGY/PRINCIPAL FINDINGS: We investigated dynamic changes in the γδ-T cells of patient with CD following treatment with infliximab (Remicade®; n=20) or adalimumab (Humira®; n=26) using flow cytometry. In patients with a high γδ-T cell level, the γδ-T cells were assessed for clonality. Of these 46 CD patients, 35 had a γδ-T cells level (mean 1.6%) comparable to healthy individuals (mean 2.2%), and 11 CD patients (24%) exhibited an increased level of γδ-T cells (5-15%). In the 18 patients also receiving thiopurines or methotrexate, the average baseline γδ-T cell level was 4.4%. In three male CD patients with a high baseline value, the γδ-T cell population increased dramatically following infliximab therapy. A fourth male patient also on infliximab monotherapy presented with 20% γδ-T cells, which increased to 25% shortly after treatment and was 36% between infusions. Clonality studies revealed an oligoclonal γδ-T cell pattern with dominant γδ-T cell clones. In support of our clinical findings, in vitro experiments showed a dose-dependent proliferative effect of anti-TNF-α agents on γδ-T cells. CONCLUSION/SIGNIFICANCE: CD patients treated with immunomodulators had constitutively high levels of γδ-T cells. Infliximab exacerbated clonal γδ-T cell expansion in vivo and induced γδ-T cell proliferation in vitro. Overall, young, male CD patients with high baseline γδ-T cell levels may be at an increased risk of developing malignant γδ-T cell lymphomas following treatment with anti-TNF-α agents

Anti-TNF-α agents promote the expansion of γδ-T cells in IL-2 stimulated PBMNC cultures.

<p>PBMCs were labelled with CFSE and were analysed by flow cytometry after 5 days of culture. γδ-T cells were stained with APC. The anti-TNF-α agents (infliximab, adalimumab, and etanercept) induced proliferation of γδ-T cells in vitro (middle panel), whereas the proliferation on non-γδ-T cells was negligible (right panel). In IL-2 -supplemented (200 IU/ml) PBMC cultures, infliximab (0.1 µg/ml or 1.0 µg/ml) induced a dose-dependent proliferation of γδ-T cells (middle panel). Etanercept induced proliferation of γδ-T cells in the same magnitude.The anti-IL-12p40 antibody ustekinumab had no effect on γδ-T cell proliferation in vitro. PBMCs from three CD patients and two healthy controls were analyzed.</p

Analysis of <i>TCRD</i> rearrangement and heteroduplex analysis confirms the clonal expansion of γδ-T cells.

<p>For the fragment and heteroduplex analyses two representative examples of three CD patients are shown. A) Genescan analysis of patient number 47 shows a single monoclonal peak at 199 bp with a fluorescence intensity of 3574. B) Heteroduplex analysis of the same sample demonstrates a dominant band of approximately 200 bp. C) Genescan of patient number 8 shows a single monoclonal peak at 184 bp with a fluorescence intensity of 7120. D) Heteroduplex analysis of the same sample shows a very prominent band in the range of 180–190 bp. E) Genescan of a healthy control shows various polyclonal bands with different sizes and generally low fluorescence intensity. F) Heteroduplex analysis of the same sample resulted in a DNA smear with weak fluorescence intensity.</p

The γδ-T cell level of three CD patients who were treated with infliximab increased shortly after infliximab infusion.

<p>A) The results from two representative young male CD patients are shown. B) Illustration of the gating strategy. Using multicolour flow cytometry, γδ-T cells were identified by positive gating on live lymphocytes based on forward/side scatter (left panel), the selection of both CD4+ and CD4- cells among the CD3+ T cells (middle panel), and the separation of γδ-T cells from αβ-T cells (right panel).</p

Summarised baseline characteristics.

<p>Summarised baseline characteristics.</p

Baseline γδ-T cell characteristics in 46 CD patients.

<p>A) Of the 46 CD patients, 35 (76%) had a γδ-T cell level comparable to healthy volunteers, while 11 (24%) had a level ranging from 5 to 15% (left). The γδ-T cell level correlated with the expression of the Vδ2 subunit (middle), and the γδ-T cell levels correlated negatively with age (right). B) There were no statistically significant associations between γδ-T cell levels and markers of mucosal inflammation (faecal calprotectin), clinical disease activity (Harvey-Bradshaw Index), or systemic inflammation (C-reactive protein).</p