Session Type: Abstract Submissions (ACR)
Background/Purpose: The use of specific cytokine inhibitors to treat inflammatory autoimmune conditions is common but typically utilizes monoclonal antibodies, fusion proteins, and other molecules whose large size necessitates parenteral administration. There remains a significant need for development of effective small molecule cytokine inhibitors for inflammatory conditions. Phosphoinositide 3-kinase (PI3K) is a direct upstream activator of AKT, and plays a critical role in multiple cell signaling pathways, cell cycle progression, and cell growth, and is thus a potential target for drug development. We examined the effect of a novel PI3Kδ inhibitor, GS-599220, on CD3/CD28 stimulated T-cell cytokine production.
Methods: Mouse Th17 cells were differentiated from naïve T cells in vitro, and GS-599220 was added on day 3 of differentiation prior to re-stimulation with anti-CD3/CD28 beads. On day 4, secretion of T helper signature cytokines by these cells were determined by both Luminex and ELISA, while real time PCR was used to quantify gene expression of these cytokines. The compound toxicity was determined by flow cytometry with 7AAD staining. Th17 cells were also transduced with a retrovirus encoding GFP-labeled myristoylated AKT, which renders the AKT constitutively active, and then treated with GS-599220.
Results: When added to culture after allowing naïve T-cells to polarize into different effector lineages, we observed dose-dependent suppression of multiple cytokines by Luminex and ELISA, including IL-17, IFNγ, and IL-4 from Th17, Th1, and Th2 cells, respectively. Real time PCR confirmed the suppression of cytokine gene expression suggesting the inhibition is mediated at the transcriptional level. Toxicity of this compound to Th17 cells was found to be minimal, indicating that the mechanism of cytokine inhibition is not by cell death. Experiments carried out under conditions free of antigen presenting cells suggest that this compound functions independently of antigen presenting cells and acts directly on the T cells. To investigate the downstream effects of this PI3Kδ inhibitor, constitutively active AKT was introduced into Th17 cells by retroviral transduction. Th17 cells containing constitutively active AKT appear to be resistant to PI3Kδ inhibition, as IL-17 production is not reduced even in the presence of high concentrations of GS-599220, suggesting that this inhibitor is acting through AKT signaling pathways.
Conclusion: These experiments show effectiveness of this small molecule inhibitor of PI3Kδ activity to modulate inflammation in vitro and exhibit promise as a treatment for in vivo models of inflammation.
E. E. Way,
K. D. Puri,
Gilead Sciences, Inc.,
J. K. Kolls,
« Back to 2013 ACR/ARHP Annual Meeting
ACR Meeting Abstracts - https://acrabstracts.org/abstract/dose-dependent-suppression-of-cytokine-production-from-t-cells-by-a-novel-phosphoinositide-3-kinase-delta-inhibitor/