ARF6 Inhibition Stabilizes the Vasculature and Enhances Survival during Endotoxic Shock

Abstract

The vascular endothelium responds to infection by destabilizing endothelial cell-cell junctions to allow fluid and cells to pass into peripheral tissues facilitating clearance of infection and tissue repair. During sepsis, endotoxin and other pro-inflammatory molecules induce excessive vascular leak, which can cause organ dysfunction, shock, and death. Current therapies for sepsis are limited to antibiotics and supportive care, which are often insufficient to reduce morbidity and prevent mortality. Previous attempts at blocking inflammatory cytokine responses in humans have proven ineffective at reducing the pathologies associated with sepsis, thus highlighting the need for a new therapeutic strategy. The small GTPase ARF6 is activated by a MYD88-ARNO interaction to induce vascular leak through disruption of endothelial adherens junctions. Here we show that the MYD88-ARNO-ARF6 signaling axis is responsible for lipopolysaccharide (LPS)-induced endothelial permeability and is a destabilizing convergence point utilized by multiple inflammatory cues. We also show that blocking ARF6 with a peptide construct of its N-terminus is sufficient to reduce vascular leak and enhance survival during endotoxic shock without inhibiting host cytokine response. Our data highlight the therapeutic potential of blocking ARF6 and reducing vascular leak for the treatment of inflammatory conditions such as endotoxemia.

Endothelial Robo4 suppresses breast cancer growth and metastasis through regulation of tumor angiogenesis

“…To confirm the role of Robo4, we investigated the effect of enhancing Robo4 signaling with the small molecule drug (SecinH3) which targets downstream of Robo4. Similar to previous studies of sepsis and arthritis (Davis et al., 2014; Zhu et al., 2012), SecinH3 showed robust effects on enhancing Robo4 signaling. It rescued tight junction protein ZO‐1 expression and inhibited breast tumor angiogenesis, tumor growth and metastasis.…”

“…Deactivation of ARF6 could be achieved by pharmacologically inhibiting ARF6 activating proteins, guanine nucleotide exchange factors, with the small molecule drug SecinH3 (Grossmann et al., 2013; Zhu et al., 2012). This drug has been shown to be effective in enhancing Robo4 function in treating arthritis and endotoxemia (Davis et al., 2014; Zhu et al., 2012). Hence, we hypothesize that SecinH3 also enhances Robo4 signaling in regulating breast cancer angiogenesis.…”

Small GTPase ARF6 controls VEGFR2 trafficking and signaling in diabetic retinopathy

“…We hypothesized that the small GTPase ARF6 may be involved in the enhanced vascular permeability seen in diabetic retinopathy, as ARF6 has been shown to mediate permeability in other disease states (14,15). We recently discovered the essential role of ARF6 in uveal melanoma and found upregulated protein levels of ARF6 in human tumor samples (16).…”

Preserving Vascular Integrity Protects Mice Against Multidrug-Resistant Gram-Negative Bacterial Infection

“…Stabilizing the vasculature to prevent fluid leak, without compromising the beneficial innate immune system response, may allow the host ample time to clear the infection without causing organ failure. The activation state of ARF6, which is a convergence point in the signaling pathways of several inflammatory mediators and cytokines, plays a major role in controlling vascular permeability (13–16). When ARF6 is in its activated GTP-bound state, permeability is increased, and when it is in its inactive GDP-bound state, the vasculature is stabilized leading to decreased permeability.…”

“…Proinflammatory agonists such as IL-1β, GNB lipopolysaccharides (LPS), and vascular endothelial growth factor (VEGF) activate ARF6, which in turn promotes the internalization of the adherens junction protein VE-cadherin and thereby increases endothelial permeability (13, 15, 16). We have shown that stabilizing the vasculature by reducing ARF6 activity is an effective strategy for reversing pathology and increasing survival rates in several preclinical models of inflammatory and vascular disease, including arthritis (15), LPS-induced endotoxemia (13), and diabetic retinopathy (16). Here, we show that stabilizing the vasculature through pharmacological inhibition of ARF6 activation by small molecules can increase survival rates and reduce pathology in preclinical models of MDR GNB infections.…”