Orthogonal self-assembly of an organoplatinum(II) metallacycle and cucurbit[8]uril that delivers curcumin to cancer cells

Abstract

Curcumin (Cur) is a naturally occurring anticancer drug isolated from the Curcuma longa plant. It is known to exhibit anticancer properties via inhibiting the STAT3 phosphorylation process. However, its poor water solubility and low bioavailability impede its clinical application. Herein, we used organoplatinum(II) ← pyridyl coordination-driven self-assembly and a cucurbit[8]uril (CB[8])-mediated heteroternary host-guest complex formation in concert to produce an effective delivery system that transports Cur into the cancer cells. Specifically, a hexagon 1, containing hydrophilic methyl viologen (MV) units and 3,4,5-Tris[2-[2-(2-methoxyethoxy)ethoxy]ethoxy]benzoyl groups alternatively at the vertices, has been synthesized and characterized by several spectroscopic techniques. The MV units of 1 underwent noncovalent complexation with CB[8] to yield a host-guest complex 4. Cur can be encapsulated in 4, via a 1:1:1 heteroternary complex formation, resulting in a water-soluble host-guest complex 5. The host-guest complex 5 exhibited ca 100-fold improved IC₅₀ values relative to free Cur against human melanoma (C32), melanoma of rodents (B16F10), and hormone-responsive (MCF-7) and triple-negative (MDA-MB231) breast cancer cells. Moreover, strong synergisms of Cur with 1 and 4 with combinatorial indexes of <1 across all of the cell lines were observed. An induced apoptosis with fragmented DNA pattern and inhibited expression of phosphor-STAT3 supported the improved therapeutic potential of Cur in heteroternary complex 5.

Keywords: cancer; drug delivery; metallacycle; orthogonal self-assembly; supramolecular coordination complex.

Fig. 1.

(A) Synthesis of MV-functionalized discrete metallacycle 1 and cartoon representation of the formation of host–guest complex 5 from the hierarchical self-assembly of 1, CB[8], and Cur. (B) Host–guest complexation of 2′ with CB[8] and Cur.

Fig. 2.

(A and B) The ³¹P and partial (C–E) ¹H NMR spectra (CD₃OD, 25 °C) of (A and C) hexagonal metallacycle 1, (B and E) acceptor 3, and (D) donor 2'. (F) Experimental (red) and calculated (blue) ESI TOF MS spectra of discrete metallacycle [M − 5ONO₂]⁵⁺.

Fig. 3.

TEM images of self-assembled nanostructures obtained from aq. solutions of (A) 1 at a concentration of 0.14 mM and 5 at concentrations of (B–D) 0.28, 0.14, and 0.014 mM, respectively.

Fig. 4.

DLS plots of 1 and 5 at concentrations of (A) 0.14 and (B) 0.014 mM.

Fig. 5.

MTT assay results showing cell viability in (A) C32, (B) B16F10, (C) MCF-7, and (D) MDA-MB231 cells; 10,000 cells were plated in 96-well plates and treated for 48 h before performing the MTT assay. Cells were treated at concentrations of 50, 5, 0.5, and 0.05 μM aq. formulations of free Cur, 1, 4, 5, 2′, 4′, and 5′ (concentrations of 5 and 5′ are chosen such that they have the same molar equivalence of Cur). Comparison of IC₅₀ values obtained from treatment of C32, B16F10, MCF-7, and MDA-MB231 cells using samples (E) 2′, 4′, and 5′ and (F) 1, 4, and 5. Biostatistical analysis was performed using two-way ANOVA with Dunnett’s multiple comparisons test comparing cell viability from Cur treatment with other samples. Here *, **, ***, and **** represent P values < 0.05, 0.01, 0.001, and 0.0001, respectively.

Fig. 6.

(A) Histograms of C32 cells obtained after PI staining posttreatment with 5′ and 5. Cells without treatment and without PI staining were used as controls, while cells with PI staining and without treatment were secondary controls. Cur-treated cells were used as positive controls. Mechanistic studies on representative C32 cell lines. (B) DNA fragmentation assay performed on cells untreated (lane III) or treated with 5 (lane II), Cur (lane I), and 5′ (lane V); lane IV represents the DNA ladder with fragments of different polynucleotide length and molecular weight. (C) Protein bands captured from blots with pSTAT3 and β-actin expression after treatment of cells with Cur (band II), 5′ (band III), and 5 (band IV) and compared with untreated cell protein (band I) and (D) comparison of percentage protein expression from different treatments. Cells were treated with free Cur, 5, and 5′ at a concentration of 10 μM (concentrations of 5 and 5′ are chosen such that they have the same molar equivalence of Cur).