Autotaxin exacerbates tumor progression by enhancing MEK1 and overriding the function of miR-489-3p.

Upregulated expression of autotaxin, a secreted phospholipase and phosphodiesterase enzyme, appears in malignant disease. The identification of a circulating miRNA signature should distinguish autotaxin-mediated disease and also elucidate unknown molecular mechanisms that rationalize its malignant potential. Using female transgenic 'AT-ATX' mice, whereby human wild-type autotaxin is expressed in liver under the control of the alpha-1 antitrypsin promoter, transgenic animals express augmented autotaxin in circulation and a percentage develop tumors. Serum collected at necropsy had circulating miRNAs analyzed for statistical significance. The ensuing autotaxin-mediated miRNome differentiated between groups: healthy FVB/N mice versus AT-ATX mice with and without tumors. Intriguingly, miR-489-3p was sharply increased in AT-ATX tumor-bearing mice. Tissue analysis showed a correlation between miR-489-3p expression in tumors and surrounding milieu with autotaxin concentration in circulation. Sequence alignment suggested miR-489-3p targets MEK1, which was confirmed through in vitro studies. Exogenously added miR-489-3p, which decreases MEK1 in normal cells, dramatically increased MEK1 expression in cells stably expressing autotaxin. Taken together, this suggests that autotaxin overrides the normal regulatory function of miR-489-3p to inhibit MEK1 via coordinately increased miR-489-3p appearing in serum.

Vinyl sulfone analogs of lysophosphatidylcholine irreversibly inhibit autotaxin and prevent angiogenesis in melanoma.

Autotaxin (ATX) is an enzyme discovered in the conditioned medium of cultured melanoma cells and identified as a protein that strongly stimulates motility. This unique ectonucleotide pyrophosphatase and phosphodiesterase facilitates the removal of a choline headgroup from lysophosphatidylcholine (LPC) to yield lysophosphatidic acid (LPA), which is a potent lipid stimulator of tumorigenesis. Thus, ATX has received renewed attention because it has a prominent role in malignant progression with significant translational potential. Specifically, we sought to develop active site-targeted irreversible inhibitors as anti-cancer agents. Herein we describe the synthesis and biological activity of an LPC-mimetic electrophilic affinity label that targets the active site of ATX, which has a critical threonine residue that acts as a nucleophile in the lysophospholipase D reaction to liberate choline. We synthesized a set of quaternary ammonium derivative-containing vinyl sulfone analogs of LPC that function as irreversible inhibitors of ATX and inactivate the enzyme. The analogs were tested in cell viability assays using multiple cancer cell lines. The IC50 values ranged from 6.74 to 0.39 µM, consistent with a Ki of 3.50 µM for inhibition of ATX by the C16H33 vinyl sulfone analog CVS-16 (10b). A phenyl vinyl sulfone control compound, PVS-16, lacking the choline-like quaternary ammonium mimicking head group moiety, had little effect on cell viability and did not inhibit ATX. Most importantly, CVS-16 (10b) significantly inhibited melanoma progression in an in vivo tumor model by preventing angiogenesis. Taken together, this suggests that CVS-16 (10b) is a potent and irreversible ATX inhibitor with significant biological activity both in vitro and in vivo.

Regulator of G-Protein Signaling 5 Reduces HeyA8 Ovarian Cancer Cell Proliferation and Extends Survival in a Murine Tumor Model.

The regulator of G-protein signaling 5 (RGS5) belongs to a family of GTPase activators that terminate signaling cascades initiated by extracellular mediators and G-protein-coupled receptors. RGS5 has an interesting dual biological role. One functional RGS5 role is as a pericyte biomarker influencing the switch to angiogenesis during malignant progression. Its other functional role is to promote apoptosis in hypoxic environments. We set out to clarify the extent to which RGS5 expression regulates tumor progression-whether it plays a pathogenic or protective role in ovarian tumor biology. We thus constructed an inducible gene expression system to achieve RGS5 expression in HeyA8-MDR ovarian cancer cells. Through this we observed that inducible RGS5 expression significantly reduces in vitro BrdU-positive HeyA8-MDR cells, although this did not correlate with a reduction in tumor volume observed using an in vivo mouse model of ovarian cancer. Interestingly, mice bearing RGS5-expressing tumors demonstrated an increase in survival compared with controls, which might be attributed to the vast regions of necrosis observed by pathological examination. Additionally, mice bearing RGS5-expressing tumors were less likely to have ulcerated tumors. Taken together, this data supports the idea that temporal expression and stabilization of RGS5 could be a valuable tactic within the context of a multicomponent approach for modulating tumor progression.