Sphaerococcenol A Derivatives: Design, Synthesis, and Cytotoxicity

Abstract

Sphaerococcenol A is a cytotoxic bromoditerpene biosynthesized by the red alga Sphaerococcus coronopifolius. A series of its analogues (1-6) was designed and semi-synthesized using thiol-Michael additions and enone reduction, and the structures of these analogues were characterized by spectroscopic methods. Cytotoxic analyses (1-100 µM; 24 h) were accomplished on A549, DU-145, and MCF-7 cells. The six novel sphaerococcenol A analogues displayed an IC₅₀ range between 14.31 and 70.11 µM on A549, DU-145, and MCF-7 malignant cells. Compound 1, resulting from the chemical addition of 4-methoxybenzenethiol, exhibited the smallest IC₅₀ values on the A549 (18.70 µM) and DU-145 (15.82 µM) cell lines, and compound 3, resulting from the chemical addition of propanethiol, exhibited the smallest IC₅₀ value (14.31 µM) on MCF-7 cells. The highest IC₅₀ values were exhibited by compound 4, suggesting that the chemical addition of benzylthiol led to a loss of cytotoxic activity. The remaining chemical modifications were not able to potentiate the cytotoxicity of the original compounds. Regarding A549 cell viability, analogue 1 exhibited a marked effect on mitochondrial function, which was accompanied by an increase in ROS levels, Caspase-3 activation, and DNA fragmentation and condensation. This study opens new avenues for research by exploring sphaerococcenol A as a scaffold for the synthesis of novel bioactive molecules.

Keywords: algae; anticancer; apoptosis; hemi-synthesis; marine natural products; reactive oxygen species.

Scheme 1

Work on sphaerococcenol A reactions performed by Cafieri and collaborators [19] (A). Chemical modifications were performed in this study using sphaerococcenol A as scaffold (B).

Scheme 2

Hemi-synthetic compounds of sphaerococcenol A obtained by thio-Michael addition and reduction.

Figure 1

Partial stacking ¹H NMR spectra of diastereosiomers 5 and 6.

Figure 2

Effects of compound 1 (1–100 µM; 24 h) in A549 cell viability estimated by MTT (A), calcein-AM (B), and LDH (C) assays. Symbols represent significant differences (ANOVA, Dunnett’s test, p  <  0.05) when compared to vehicle. At least three independent experiments were carried out in triplicate.

Figure 3

Reactive oxygen species levels produced by A549 cells following treatment with compound 1 (10, 30, and 100 µM) for 3 and 6 h. Symbols represent significant differences (two-way ANOVA, Bonferroni’s test, p  <  0.05) when compared to vehicle. At least three independent experiments were carried out in triplicate.

Figure 4

Effects of compound 1 treatment (10, 30, and 100 µM) on A549 cells regarding alterations in mitochondrial membrane potential after 3 and 6 h (A), Caspase-3 activity after 6 and 12 h (B), and alterations in DNA morphology after 12 and 24 h (C). Symbols represent significant differences (two-way ANOVA, Bonferroni’s test, p  <  0.05) when compared to vehicle. At least three independent experiments were carried out in triplicate. Images are representative of one well of each treatment condition and were obtained using an inverted fluorescence microscope at ×400. Red arrows indicate alterations in DNA compared to vehicle.

Figure 4

Effects of compound 1 treatment (10, 30, and 100 µM) on A549 cells regarding alterations in mitochondrial membrane potential after 3 and 6 h (A), Caspase-3 activity after 6 and 12 h (B), and alterations in DNA morphology after 12 and 24 h (C). Symbols represent significant differences (two-way ANOVA, Bonferroni’s test, p  <  0.05) when compared to vehicle. At least three independent experiments were carried out in triplicate. Images are representative of one well of each treatment condition and were obtained using an inverted fluorescence microscope at ×400. Red arrows indicate alterations in DNA compared to vehicle.