Design, synthesis and anticancer activity of constrained sphingolipid-phenoxazine/phenothiazine hybrid constructs targeting protein phosphatase 2A

Abstract

Inspired by the cytotoxicity of perphenazine toward cancer cells and its ability to activate the serine/threonine protein phosphatase 2A (PP2A), we prepared series of ether-carbon linked analogs of a constrained synthetic sphingolipid analog 3, known for its cytotoxicity, nutrient transporter down-regulation and vacuolation properties, incorporating the tricyclic neuroleptics phenoxazine and phenothiazine to represent hybrid structures with possible synergistic cytotoxic activity. While the original activity of the lead compound 3 was diminished by fusion with the phenoxazine or phenothiazine tethered moieties, the corresponding 3-pyridyltetryl ether analog 10 showed cytotoxicity and nutrient transporter down-regulation similar to the lead compound 3, although it separated these PP2A-dependent phenotypes from that of vacuolation.

Keywords: Cytotoxicity FL5.12; Hybrid structures; Nutrient transport down-regulation; Sphingolipid; Vacuolation.

Figure 1.

Structures of natural and synthetic anti-proliferative compounds.

Figure 2.

Vacuolation in FL5.12 cells. (A) Two representative vacuolation images of FL5.12 cells with vacuolation scores of: (i) 0 (no vacuoles); (ii) 1 (a few small vacuoles; (iii); 2 (multiple vacuoles); (iv) 3 (multiple large vacuoles). (B) Calyculin A protects from vacuolation caused by 3.

Figure 3.

Structures of known PP2A activators.

Figure 4.

Structures of synthetic hybrid compounds 6-9.

Figure 5.

Perphenazine (4) and SMAP (5) are less potent than 3 but share PP2A dependence. (A) IC₅₀, nutrient transporter loss, and vacuolation of PP2A activators 3-5. ^‡Denotes a score of 0. (B) Nutrient transporter loss at 2x IC₅₀ of PP2A activators shown in (A) ± pretreatment with 5 nM Calyculin A. Data in (A) are means ± S.D. and data in (B) are means ± range.

Figure 6.

IC₅₀, nutrient transporter loss and vacuolation of synthetic hybrid compounds with increasing linker length. Data shown are means ± S.D.

Figure 7.

Structures of heteroaromatic linked analogs of 3.

Figure 8.

IC₅₀, nutrient transporter loss, and vacuolation of heteroaromatic linker analogs of 3. Data shown are means ± S.D.