Impact of SMTP Targeting Plasminogen and Soluble Epoxide Hydrolase on Thrombolysis, Inflammation, and Ischemic Stroke

Figure 1. Structures of key Stachybotrys microspora triprenyl phenol (SMTP) congeners and substituted chromanlactam core unit.

Figure 2. Putative pathway for SMTP biosynthesis. The synthetic pathway for SMTP-type and phenylspirodrimane-type triprenyl phenols may diverge, through differential cyclization mechanisms, from LL-Z1272β (ilicicolin B), a key intermediate synthesized from farnesyl diphosphate and orsellinic acid [63]. Pre-SMTP forms various SMTPs via a nonenzymatic reaction with an amine. Stachybotrydial [66] may yield a wide variety of phenylspirodrimanes via a similar mechanism, because aromatic o-dialdehydes (shaded in light blue) are highly reactive with an amine [67,68]. The precursor amine feeding method can be applied to selectively synthesize a phenylspirodrimane of interest [69] using Stachybotrys sp. F462, which forms stachybotramide, a phenylspirodrimane-type triprenyl phenol [70].

Figure 3. Plasminogen species. Schematic structures of the four plasminogen species and plasmin, and the effect of SMTP-7 on the activation of the four plasminogen species [97] are presented. A key disulfide bond connects the two chains of plasmin (green line). PAN, plasminogen–apple–nematode domain; K, kringle domain; P, protease domain.

Figure 4. Proposed mechanism for the action of SMTP on plasminogen. (A) Schematic structure of Glu-plasminogen with closed spiral conformation, which is resistant to fibrin binding and plasminogen activator (PA)-catalyzed activation (see Figure 3 legend for designation of the symbol). The image incorporates essential structural information derived from crystallographic studies [94,95]. (B) An SMTP cofactor (phospholipid or unsaturated fatty acid) possibly binds to a site in K5 partly overlapping the aminohexyl site (see [98] for oleic acid–K5 binding) to modify the PAN–K5 interaction (image represents PAN–K5-disrupted molecule). (C) Cofactor-modified conformation may allow SMTP binding (possibly to K5, because SMTP does not affect micro-plasminogen, which lacks intact K5 [97]). SMTP binding leads to large-scale conformational change in plasminogen, enabling (D) robust fibrin binding and PA-catalyzed activation to plasmin (not shown). The resulting plasmin cleaves fibrin to yield C-terminal lysine, to which K1 (as well as K2 and K4) binds strongly, recruiting more plasminogen to accelerate fibrinolysis [99]. Thus, the promotion of plasminogen–fibrin binding and activation of the bound…

Figure 5. Structure–activity relationships of SMTP congeners with respect to plasminogen modulation and soluble epoxide hydrolase (sEH) inhibition. Plots are based on data in [20,60]. Plot number represents SMTP number (see Table S1 for structure). Details of the dimensions in the three axes are described in [20]. Briefly, the larger the value for each parameter, the stronger the activity (plasminogen modulation or sEH ihibition). C-EH, C-terminal epoxide hydrolase of soluble epoxide hydrolase; N-Phos, N-terminal phosphatase of soluble epoxide hydrolase.

Figure 6. Summary of pharmacological activity of SMTP in comparison with tissue-type plasminogen activator (t-PA). (A) Pharmacological activity of SMTP is derived from a triad of activities: (i) modulation of plasminogen (Plg) conformation, promoting fibrin binding and conversion to plasmin (Pm), and subsequent fibrinolysis; (ii) inhibition of sEH, suppressing inflammation; and (iii) antioxidative activity, preventing oxidative damage. Process (i) is dependent on fibrin and endogenous PAs, allowing the SMTP action to promote physiological fibrinolysis. Therefore, pharmacological doses of SMTP do not cause bleeding. In process (ii), inhibition of C-EH-catalyzed hydrolysis of anti-inflammatory epoxy fatty acids such as epoxyeicosatrienoic acid (EET) may play a role; inhibition of N-phos may play an additional role (not shown). In process (iii), radical-scavenging activity inherent in the SMTP structure plays a role in the suppression of oxidative modifications of lipids, proteins, nucleic acids, and sugars [27,30,133,158,159], which are implicated in cytotoxicity, tissue damage, and inflammation. Anti-inflammatory/antioxidative effects contribute to the suppression of neuronal damage and…