Inhibition of GTPase Rac1 in Endothelium by 6-Mercaptopurine Results in Immunosuppression in Nonimmune Cells: New Target for an Old Drug

Abstract

Azathioprine and its metabolite 6-mercaptopurine (6-MP) are well established immunosuppressive drugs. Common understanding of their immunosuppressive properties is largely limited to immune cells. However, in this study, the mechanism underlying the protective role of 6-MP in endothelial cell activation is investigated. Because 6-MP and its derivative 6-thioguanosine-5′-triphosphate (6-T-GTP) were shown to block activation of GTPase Rac1 in T lymphocytes, we focused on Rac1-mediated processes in endothelial cells. Indeed, 6-MP and 6-T-GTP decreased Rac1 activation in endothelial cells. As a result, the compounds inhibited TNF-α–induced downstream signaling via JNK and reduced activation of transcription factors c-Jun, activating transcription factor-2 and, in addition, NF κ-light-chain-enhancer of activated B cells (NF-κB), which led to decreased transcription of proinflammatory cytokines. Moreover, 6-MP and 6-T-GTP selectively decreased TNF-α–induced VCAM-1 but not ICAM-1 protein levels. Rac1-mediated generation of cell membrane protrusions, which form docking structures to capture leukocytes, also was reduced by 6-MP/6-T-GTP. Consequently, leukocyte transmigration was inhibited after 6-MP/6-T-GTP treatment. These data underscore the anti-inflammatory effect of 6-MP and 6-T-GTP on endothelial cells by blocking Rac1 activation. Our data provide mechanistic insight that supports development of novel Rac1-specific therapeutic approaches against chronic inflammatory diseases.

Targeting Pancreatic Cancer Metastasis by Inhibition of Vav1, a Driver of Tumor Cell Invasion

“…Though azathioprine has been used clinically for decades, it was only recently discovered that a metabolite of azathioprine is a potent inhibitor of Vav1/Rac (11–13). Azathioprine, which is converted to 6-mercaptopurine (6-MP), is metabolized to 6-thio-GTP, which can be loaded onto Rac1 in place of GTP.…”

Azathioprine with Allopurinol

“…Inhibition of GTP signaling, specifically the RAC1 GTPase which is elevated in expression in activated T-cells, 40 may be one mechanism, at least in vitro. 23,41,42 However, evidence that this underlies clinical responses is lacking 43 and interference in RAC1 signalling was not apparent from our RNA-seq data. We found a clear response with respect to the downregulation of genes involved in host-defense mechanisms against microorganisms in LDAA patients.…”

6-Mercaptopurine modifies cerebrospinal fluid T cell abnormalities in paediatric opsoclonus–myoclonus as steroid sparer

“…In murine microglial cultures, 6-MP attenuated TNF-a production, and potential therapeutic use to down-regulate inflammation mediated by microglia was proposed [30]. 6-MP also inhibits atherosclerosis in mice by reducing local CCL2 and macrophages [31]; affords neuroprotection to focal cerebral occlusion in rats [32]; and regulates the Bcl-2/Bax ratio [33], transcriptional activity at the nuclear receptor NR4A level [34,35] and Rac1-mediated signalling [36,37]. In OMS, we did not find a reduction in CSF T cell activation, but our patients did not manifest the very high percentages we often encounter [2].…”