Background/Purpose: Tumor progression locus 2 (TPL2, also known as MAP3K8) is a mitogen-activated protein kinase kinase kinase and the primary regulator of ERK-mediated gene transcription downstream of multiple proinflammatory stimuli including bacterial products (eg, LPS and bacterial peptidoglycans), damage-associated molecular patterns (DAMPs), TNFα, and IL-1β.¹ TPL2 regulates the expression of several proinflammatory cytokines, including TNFα, IL-1β, IL-6 and IL-8. As TPL2 acts on both the production of and response to TNFα and IL-1β, it acts to amplify proinflammatory signaling. Dysregulated signaling downstream of these inflammatory signals can drive uncontrolled immune cell activation and inflammation, which is associated with multiple chronic inflammatory and autoimmune diseases. As such, TPL2 inhibition represents a strategy to modulate inflammation in a variety of disease settings. We evaluated the effect of a highly selective TPL2 inhibitor (GS-4875) on inflammatory signaling and cytokine production in primary human cells and in an acute inflammation model.

Methods: GS-4875 selectivity was screened using a KINOMEscan™ selectivity assay (ScanMAX, DiscoveRx, San Diego, CA). Cells were precultured with GS-4875 prior to stimulation with LPS, TNFα, or EGF. Lewis rats were dosed orally with 3, 10, 30 or 100 mg/kg doses of GS-4875 or 1 mg/kg dexamethasone followed by IV dosing of 0.01 mg/kg LPS 2 hours later. Animals were bled for plasma at multiple time points between 0 and 5h after compound dosing and plasma concentrations of TNFα and GS-4875 were measured.

Results: GS-4875 inhibits the TPL2 kinase with an IC50 = 1.3 nM with no significant off-target binding activity. GS-4875 selectively inhibited LPS and TNFα-stimulated phosphorylation of TPL2, MEK, and ERK, with little to no inhibition of phosphorylated p38, JNK or p65 observed. Both the RNA production and secretion of TNFα, IL-1β, IL-6, and IL-8 following LPS stimulation in primary human monocytes was similarly inhibited with GS-4875. In monocyte-derived dendritic cells GS-4875 inhibited the secretion of TNFα and IL-6 following LPS stimulation. To confirm TPL2 requirement for inflammatory, but not Ras-mediated (growth factor stimulated) ERK signaling, A431 cells were stimulated with either TNFα or EGF. Although TPL2 inhibition reduced TNFα-stimulated pERK, no effect on ERK activation downstream of EGF was observed. In vivo activity and PK/PD relationship was established using a rat LPS-TNFα model of acute inflammation. GS-4875 treatment showed dose and exposure dependent inhibition of LPS-stimulated TNFα production at all time points with an estimated EC50 (±SD) of 667 ±124 nM. Inhibition of TNFα production at the highest dose tested inhibited TNFα levels at levels equivalent to that of dexamethasone.

Conclusion: This work demonstrates the selective effects of TPL2 inhibition on ERK-mediated signaling and proinflammatory cytokine production and highlights the potential for TPL2 inhibition to treat diseases associated with dysregulated inflammatory signaling and chronic inflammation.

Reference:

1. Gantke T, Sriskantharajah S, Ley S. Cell Res. 2011;21:131-145.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/gs-4875-a-first-in-class-tpl2-inhibitor-suppresses-mek-erk-inflammatory-signaling-and-proinflammatory-cytokine-production-in-primary-human-monocytes/