Inflammatory T helper 17 cells in humans are distinguished by selective expression of MDR1 and are enriched in the gut of patients with Crohn’s disease.

A T-cell subset, defined as CD4+CD25hi (regulatory T-cells [Treg cells]), was recently shown to suppress T-cell activation. We demonstrate that human Treg cells isolated from healthy donors express the HIV-coreceptor CCR5 and are highly susceptible to HIV infection and replication. Because Treg cells are present in very few numbers and are difficult to expand in vitro, we genetically modified conventional human T-cells to generate Treg cells in vitro by ectopic expression of FoxP3, a transcription factor associated with reprogramming T-cells into a Treg subset. Overexpression of FoxP3 in naïve human CD4+ T-cells recapitulated the hyporesponsiveness and suppressive function of naturally occurring Treg cells. However, FoxP3 was less efficient in reprogramming memory T-cell subset into regulatory cells. In addition, FoxP3-transduced T-cells also became more susceptible to HIV infection. Remarkably, a portion of HIV-positive individuals with a low percentage of CD4+ and higher levels of activated T-cells have greatly reduced levels of FoxP3+CD4+CD25hi T-cells, suggesting disruption of the Treg cells during HIV infection. Targeting and disruption of the T-cell regulatory system by HIV may contribute to hyperactivation of conventional T-cells, a characteristic of HIV disease progression. Moreover, the ability to reprogram human T-cells into Treg cells in vitro will greatly aid in decoding their mechanism of suppression, their enhanced susceptibility to HIV infection, and the unique markers expressed by this subset.

A central challenge for improving autoimmune therapy is preventing inflammatory pathology without inducing generalized immunosuppression. T helper 17 (TH17) cells, characterized by their production of interleukin-17, have emerged as important and broad mediators of autoimmunity. Here we show that the small molecule halofuginone (HF) selectively inhibits mouse and human TH17 differentiation by activating a cytoprotective signaling pathway, the amino acid starvation response (AAR). Inhibition of TH17 differentiation by HF is rescued by the addition of excess amino acids and is mimicked by AAR activation after selective amino acid depletion. HF also induces the AAR in vivo and protects mice from TH17-associated experimental autoimmune encephalomyelitis. These results indicate that the AAR pathway is a potent and selective regulator of inflammatory T cell differentiation in vivo.

Febrifugine, one of the fifty fundamental herbs of traditional Chinese medicine, has been characterized for its therapeutic activity whilst its molecular target has remained unknown. Febrifugine derivatives have been used to treat malaria, cancer, fibrosis, and inflammatory disease. We recently demonstrated that halofuginone (HF), a widely studied derivative of febrifugine, inhibits the development of Th17-driven autoimmunity in a mouse model of multiple sclerosis by activating the amino acid response pathway (AAR). Here we show that HF binds glutamyl-prolyl-tRNA synthetase (EPRS) inhibiting prolyl-tRNA synthetase activity; this inhibition is reversed by the addition of exogenous proline or EPRS. We further show that inhibition of EPRS underlies the broad bioactivities of this family of natural products. This work both explains the molecular mechanism of a promising family of therapeutics, and highlights the AAR pathway as an important drug target for promoting inflammatory resolution.

Adoptive cell therapy (ACT) using autologous tumor-infiltrating lymphocytes (TIL) can result in complete regression of advanced cancer in some patients, but the efficacy of this potentially curative therapy might be limited by poor persistence of TIL after adoptive-transfer. Pharmacologic inhibition of the serine/threonine kinase Akt has recently been shown to promote immunologic memory in viral-specific murine models, but whether this approach may enhance features of memory (e.g. long-term persistence) in TIL which are characteristically exhausted and senescent is not established. Here we show that pharmacologic inhibition of Akt enables expansion of TIL with the transcriptional, metabolic and functional properties characteristic of memory T cells. Consequently, Akt inhibition results in enhanced persistence of TIL after adoptive transfer into an immunodeficient animal model and augments anti-tumor immunity of CD8 T cells in a mouse model of cell-based immunotherapy. Pharmacologic inhibition of Akt represents a novel immunometabolomic approach to enhance the persistence of anti-tumor T cells and improve the efficacy of cell-based immunotherapy for metastatic cancer.

We identified three RORγt-specific inhibitors that suppress T helper 17 (Th17) cell responses including Th17 cell-mediated autoimmune disease. We systemically characterized RORγt binding in the presence and absence of drug with corresponding whole-genome transcriptome sequencing. RORγt acts both as a direct activator of Th17 cell signature genes and as a direct repressor of signature genes from other T-cell lineages, with the strongest transcriptional effects on cis-regulatory sites containing the RORα binding motif. RORγt is central in a densely interconnected regulatory network that shapes the balance of T-cell differentiation. The three inhibitors identified here modulated the RORγt-dependent transcriptional network to varying extents and through distinct mechanisms. Whereas one inhibitor displaced RORγt from its target-loci, the two more potent inhibitors affected transcription predominantly without removing DNA-binding. Our work illustrates the power of a system-scale analysis of transcriptional regulation to characterize potential therapeutic compounds that inhibit pathogenic Th17 cells and suppress autoimmunity.

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