Discovery of 1,4-Naphthoquinones as a New Class of Antiproliferative Agents Targeting GPR55

Abstract

A new series of 1,4-naphthoquinones, bearing various cyclic and aliphatic amines on C2, was designed and synthesized to identify antiproliferative agents for triple-negative breast cancer, which represents a clinical challenge without targeted therapies. Among naphthoquinones, 2a and 3a inhibited the proliferation of MDA-MB-231 cells (EC₅₀ = 1.6 and 2.7 μM, respectively), compared to primary human breast cells MCF10A. Furthermore, they did not affect the viability of peripheral blood mononuclear cells (PBMC), suggesting their potential safer use for cancer treatment. Recently, correlations have emerged between the expression of G protein-coupled receptor 55 (GPR55) and both triple-negative breast cancer development and invasion, making it a promising target for the development of targeted therapies. Based on this evidence, molecular docking studies supported the hypothesis of binding to GPR55, and pharmacological tests suggested that compound 3a could exert its antiproliferative activity acting as a GPR55 inverse agonist.

Scheme 1. Synthetic Steps for Compounds 1-6a and 1-4b

Reagents and conditions: (i) 1. a: AlCl₃, NaCl, 140–150 °C; b: 170–175 °C, 5 min; 2. H₂O, HCl, rt, overnight, 99%; (ii) amine, Na₂CO₃, DCM, rt, overnight, 17–28%; (iii) CH₃I, Ag₂O, CHCl₃, rt, 22 h, 56%.

Figure 1

Dose–response curves of compound 2a (A) and 3a (B).

Figure 2

(A) Complementarity between the molecular surface of 3a (blue) and that of the GPR55 binding site (gray). (B,C) Graphical representation of the interactions between 3a and LPI and the binding site of GPR55. The GPR55–3a complex (A) is stabilized by two hydrogen bonds (blue dashed lines) involving the ligand oxygens at 4 and 5 positions and by strong hydrophobic contacts (green dashed lines) mainly involving the morpholine ring at C2. Interestingly, LPI conserved the same hydrogen bonds (blue dashed lines, panel B), while its alkyl edge is superposed to the hydrophobic portion of morpholine of 3a. (D) Rough superposition between 3a (pink and atom type) and THM (green and atom type) shows a clear correspondence between the two phenolic hydroxyls of THM and the 4-carbonyl and 5-phenolic group of 3a. Likewise, the overall size and shape of both molecules are comparable.

Figure 3

Activity of the GPR55 selective antagonist CID16020046 and 3a, alone and in combination on two TBNC cell line MDA-MB-231 and MDA-MB-468 (A,C) and on two hormone-responsive cell lines MCF-7 and T47D (B,D). Values with the same letter are not significantly different, while values with different letters are significantly different (*p < 0.05 and **p < 0.001 one-way ANOVA followed by Bonferroni’s comparison test, as posthoc; each sample was run in triplicate (n = 3)).