Development of Stable Amino-Pyrimidine-Curcumin Analogs: Synthesis, Equilibria in Solution, and Potential Anti-Proliferative Activity

Abstract

With the clear need for better cancer treatment, naturally occurring molecules represent a powerful inspiration. Recently, curcumin has attracted attention for its pleiotropic anticancer activity in vitro, especially against colorectal and prostate cancer cells. Unfortunately, these encouraging results were disappointing in vivo due to curcumin's low stability and poor bioavailability. To overcome these issues, herein, the synthesis of eight new pyrimidine-curcumin derivatives is reported. The compounds were fully characterized (¹H/¹³C NMR (Nuclear Magnetic Resonance), LC-MS (Liquid Chromatography-Mass Spectrometri), UV-Vis spectroscopy), particularly their acid/base behavior; overall protonation constants were estimated, and species distribution, as a function of pH, was predicted, suggesting that all the compounds are in their neutral form at pH 7.4. All the compounds were extremely stable in simulated physiological media (phosphate-buffered saline and simulated plasma). The compounds were tested in vitro (48 h incubation treatment) to assess their effect on cell viability in prostate cancer (LNCaP and PC3) and colorectal cancer (HT29 and HCT116) cell lines. Two compounds showed the same anti-proliferative activity as curcumin against HCT116 cells and improved cytotoxicity against PC3 cells.

Keywords: cancer cells; cell proliferation; curcumin; pyrimidine derivatives; solution equilibria.

Figure 1

General scheme for the synthesis of amino-pyrimidine–curcumin derivatives, together with atom numbering used for NMR assignments. Red numbers refer to atom numbering used for NMR assignment.

Figure 2

¹H-NMR spectrum of MPY1 in DMSO-d₆ at 600 MHz (298 K). Highlighting boxes show the resonances of olefinic protons in the E configuration.

Figure 3

pH-metric spectrophotometric titration of MPY1 at 25 °C in aqueous solution ((MPY1) = 25 µM; (NaNO₃) = 1 mM)) in the 250–650 nm spectral range. Red spectrum, pH = 2; green spectrum, pH = 7; blue spectrum, pH = 11. The inset shows the absorbance vs. pH at 355 nm (black) and 406 nm (red).

Figure 4

Species distribution curves for MPY1, MPY3, and PY1 (from top to bottom); [L]_tot = 1 × 10⁻⁴ mol L⁻¹.

Figure 5

Absorption spectra of MPY1 (left) and PY1 (right) in PBS (phosphate-buffered saline) at 298 K ((MPY1) = (PY1) = 20 µM).

Figure 6

The histogram represents the distribution of LNCaP cells in the different phases of the cell cycle following the administration of DMSO, PY1, PY3, and curcumin for 48 h. The values represent means ± SEM of three independent experiments (two-way ANOVA with Fisher’s LSD test: * p < 0.05, *** p < 0.001, **** p < 0.0001, n = 3).