Cytotoxic and Anti-HSV-1 Effects of Caulerpin Derivatives

Abstract

Marine organisms represent a potential source of secondary metabolites with various therapeutic properties. However, the pharmaceutical industry still needs to explore the algological resource. The species Caulerpa lamouroux Forssk presents confirmed biological activities associated with its major compound caulerpin, such as antinociceptive, spasmolytic, antiviral, antimicrobial, insecticidal, and cytotoxic. Considering that caulerpin is still limited, such as low solubility or chemical instability, it was subjected to a structural modifications test to establish which molecular regions could accept structural modification and to elucidate the cytotoxic bioactive structure in Vero cells (African green monkey kidney cells, Cercopithecus aethiops; ATCC, Manassas, VA, USA) and antiviral to Herpes simplex virus type 1. Substitution reactions in the N-indolic position with mono- and di-substituted alkyl, benzyl, allyl, propargyl, and ethyl acetate groups were performed, in addition to conversion to their acidic derivatives. The obtained analogs were submitted to cytotoxicity and antiviral activity screening against Herpes simplex virus type 1 by the tetrazolium microculture method. From the semi-synthesis, 14 analogs were obtained, and 12 are new. The cytotoxicity assay showed that caulerpin acid and N-ethyl-substituted acid presented cytotoxic concentrations referring to 50% of the maximum effect of 1035.0 µM and 1004.0 µM, respectively, values significantly higher than caulerpin. The antiviral screening of the analogs revealed that the N-substituted acids with methyl and ethyl groups inhibited Herpes simplex virus type 1-induced cytotoxicity by levels similar to the positive control acyclovir.

Keywords: Caulerpa racemosa; HSV-1; Vero cells; caulerpin; indolic derivatives.

Figure 1

Structure of the bis-indolic alkaloid, caulerpin (1).

Scheme 1

2–6 derivatives of caulerpin (1). Reagents and conditions: (a) 2: KOH, Me₂SO₄; MeOH, Acetone/room temperature, magnetic stirring; (b) 3–6: KOH, RX (X = Cl or Br); DMF/room temperature, magnetic stirring.

Scheme 2

Derivatives 7 and 8 of caulerpin (1). Reagents and conditions: (a) 7: KOH, 3,4,5-trihydroxy-benzyl chloride; DMF, 50 °C, magnetic stirring; (b) 8: KOH, ethyl bromoacetate; DMF/50 °C, magnetic stirring.

Scheme 3

Acidic derivatives 9–13 from hydrolysis of 1–6. Reagents and conditions: (a) KOH, Acetonitrile: water, 60 °C, magnetic stirring.

Scheme 4

Transesterified derivatives 14 and 15. Reagents and conditions: (a) KOH, EtOH; CH₂Cl₂, room temperature, magnetic stirring.

Figure 2

Graphs of the effect of 1 and its analogs on cell viability.

Figure 3

Percentage of HSV-1 inhibition by caulerpin analogs in Vero cells infected with HSV-1 (MOI 0.2). The cells were treated with the compound’s CC₂₀ and further incubated at 37 °C in 5% CO₂ for 72 h. Cell viability was assessed using the MTT method. Statistical analyses were performed with ANOVA and Tukey’s posttest (**** p < 0.0001 versus control). ACV—Acyclovir.

Figure 4

Inhibition of HSV-1-induced cytotoxicity by compound 1 and its analogs. Cells were infected with HSV-1 (MOI 0.2) for 1 h, washed, and then treated with the compound’s CC20. The plates were incubated at 37 °C in 5% CO₂ for 72 h and cell viability was determined using the MTT method. Significant increase in the percentage of cell inhibition (ANOVA, with Dunnet’s posttest, * p < 0.05; ** p < 0.005; *** p< 0.0005; **** p < 0.0001): experimental groups versus control (infected and untreated).