Discovery of Anti-Inflammatory Alkaloids from Sponge Stylissa massa Suggests New Biosynthetic Pathways for Pyrrole-Imidazole Alkaloids

Abstract

Pyrrole-imidazole alkaloids (PIAs) are a class of marine sponge derived natural products which have complex carbon frameworks and broad bioactivities. In this study, four new alkaloids, stylimassalins A-B (1-2), 3, and 5, together with two known compounds (4 and 6), were isolated from Stylissa massa. Compounds 2, 4, and 6 are the C-2 brominated analogues of 1, 3, and 5, respectively. Their structures display three different scaffolds, of which scaffold 1 (compounds 1,2) is new. A new biosynthetic pathway from oroidin, through spongiacidin, to latonduine and scaffold 1 was proposed by our group, in which the C12-N13-cleavaged compounds of spongiacidin (scaffold 2), dubbed seco-spongiacidins (3 and 4), are recognized as a key bridged scaffold, to afford PIA analogues (1,2 and 5,6). An anti-inflammatory evaluation in a zebrafish inflammation model induced by copper sulphate (CuSO₄) demonstrated that stylimassalins A and B (1 and 2) could serve as a promising lead scaffold for treating inflammation.

Keywords: Stylissa massa; anti-inflammatory; biosynthetic pathway; latonduine; pyrrole–imidazole alkaloids; seco-spongiacidin.

Figure 1

Chemical structures of compounds 1–6. Stylimassalins A and B (1 and 2) (with a 5/6/6 scaffold) and compounds 3 and 4 are 12,13-secospongiacidin PIAs (with a 5/7 scaffold), and compounds 5 and 6 are latonduine PIAs (with a 5/7/6 scaffold).

Figure 2

Structural elucidation of compounds 1, 2, 3, and 5. HMBC correlations are shown using red (acquired via DMSO-d₆) and pink (acquired via methanol-d₄) arrow lines and COSY correlations are shown using blue bold lines.

Figure 3

X-ray structures of compounds 2, 4, and 6, which have three different scaffolds. The data on compound 4 have been published [12].

Figure 4

The absolute configurations of 1 and 2 were determined by time-dependent density functional theory ECD (TDDFT-ECD) calculations (9S for 1a and 2a; 9R for 1b and 2b).

Scheme 1

A new biosynthetic hypothesis of compounds 1–6.

Figure 5

Compounds 1 and 2 inhibit CuSO₄-induced inflammatory responses in zebrafish. (A) Representative fluorometric images of macrophage migration in the CuSO₄-induced inflammation model. The red arrows indicate macrophages that migrated to the lateral line neural crest. (B) Statistical analysis of the number of macrophages in the lateral line neural crest. The data are presented as the means ± SEMs (n = 6). ^# p < 0.05, ^## p < 0.01, and ^### p < 0.001 compared with the control group; **** p < 0.0001 compared with the model group.