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

Abstract

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 – 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.

MicroRNA Regulation of the Autotaxin-Lysophosphatidic Acid Signaling Axis

“…Additional studies suggest that serum measurements of autotaxin concentration may be useful as a biomarker for follicular lymphoma [41]. Within the tumor microenvironment, autotaxin is found where tumor cells are undergoing angiogenesis [42], due to its normal role in vasculogenesis [43,44]. In general, inhibiting autotaxin through chemical means or siRNA delivery will kill cells and prevent metastatic progression, in vitro and in vivo [34,42,45,46,47,48].…”

Development and therapeutic potential of autotaxin small molecule inhibitors: From bench to advanced clinical trials

“…The conduct of further in vivo studies in a xenograft melanoma experimental animal model demonstrated that 18 significantly inhibited tumor progression at 40 to 50 mg/kg dose after 65 days, exerting also a superior efficacy compared with other known and potent ATX inhibitors. Of interest, the mechanism through which 18 exhibited its antitumor effect was by preventing angiogenesis and not by inhibiting mitogenesis as initially thought based on the in vitro data results . However, health deterioration of the subjects and unexpected death of one of them during the study cast doubts about the safety profile of this class of inhibitors and its therapeutic potential against melanoma progression.…”

“…In an interesting example of inhibitor design, a small series of vinyl sulfone derivatives was recently prepared by Georgia University, as part of a broader effort to explore the possibility of identifying irreversible ATX inhibitors (Figure ) . The concept of developing irreversible ATX inhibitors is supported by the presence of a critical threonine residue in the catalytic site of the enzyme (Thr 210 ) acting as a nucleophile in the hydrolysis of LPC to LPA and choline.…”

“…Consequently, the incorporation of an electrophilic center into the inhibitor structure could behave as an activation site, which upon reacting covalently with the nucleophilic group of Thr 210 of ATX catalytic site would irreversibly deactivate enzyme function (compiled in a simplistic function in Figure ). With this in mind, four vinyl sulfone analogs were made containing a quaternary ammonium head group on one side, mimicking the choline part of LPC, and different lengths of saturated and unsaturated alkyl chains on the other side ( 17‐20 ; Figure ) . All analogs were tested in a recombinant ATX enzyme showing good inhibitory activity with the exception of 20 having the shortest alkyl chain (Figure ).…”

“…Structures of vinyl sulfone analogs synthesized and studied, accompanied by their in vitro autotaxin inhibitory activity and their mode of binding [Color figure can be viewed at wileyonlinelibrary.com]…”