Novel Multifunctional Ascorbic Triazole Derivatives for Amyloidogenic Pathway Inhibition, Anti-Inflammation, and Neuroprotection

Abstract

Alzheimer's disease (AD) is a common neurodegenerative disorder. The number of patients with AD is projected to reach 152 million by 2050. Donepezil, rivastigmine, galantamine, and memantine are the only four drugs currently approved by the United States Food and Drug Administration for AD treatment. However, these drugs can only alleviate AD symptoms. Thus, this research focuses on the discovery of novel lead compounds that possess multitarget regulation of AD etiopathology relating to amyloid cascade. The ascorbic acid structure has been designated as a core functional domain due to several characteristics, including antioxidant activities, amyloid aggregation inhibition, and the ability to be transported to the brain and neurons. Multifunctional ascorbic derivatives were synthesized by copper (I)-catalyzed azide-alkyne cycloaddition reaction (click chemistry). The in vitro and cell-based assays showed that compounds 2c and 5c exhibited prominent multifunctional activities as beta-secretase 1 inhibitors, amyloid aggregation inhibitors, and antioxidant, neuroprotectant, and anti-inflammatory agents. Significant changes in activities promoting neuroprotection and anti-inflammation were observed at a considerably low concentration at a nanomolar level. Moreover, an in silico study showed that compounds 2c and 5c were capable of being permeated across the blood-brain barrier by sodium-dependent vitamin C transporter-2.

Keywords: BACE1 inhibitor; amyloid aggregation inhibition; anti-inflammation; antioxidant; ascorbic derivatives; neuroprotective.

Figure 1

Chemical structure of ascorbic acid and its conjugation via triazole linker with tryptoline and phenolic moieties to create novel compounds with multitargeted functionality to treat Alzheimer’s disease.

Figure 2

Structures of multifunctional novel ascorbic derivatives.

Figure 3

Distance between the aromatic end group and the ascorbic furanone ring in the 3D structure of ascorbic derivatives: (a) 2c (yellow) and (b) 5c (magenta).

Figure 4

Neuroprotective activities of ascorbic derivatives at a concentration of 1 nM on cultured P19-derived neurons evaluated by serum-deprivation-induced oxidative stress. α-MEM, supplemented with 10 μM Ara-C, and 1% antibiotics–antimycotic solution without FBS were used to induce the oxidative stress condition. Each bar is presented as mean ± SD (n = 3). Letter a denotes a significant difference compared with the toxic condition (p < 0.05), and letter b denotes a significant difference compared with quercetin (1 nM) positive control (p < 0.05), based on one-way ANOVA using the Origin Pro 9.0 program, Northampton, MA, USA.

Figure 5

Effects of ascorbic acid, compound 2c, and compound 5c at 10 and 100 nM, on mRNA expression levels of COX-2 (cyclooxygenase-2) and iNOS (inducible nitric oxide synthase) genes in LPS inflammatory-activated RAW 264.7 macrophages. Each bar is presented as mean ± SD (n = 3). Different letters (a, b, c, d, and e) indicate significant differences of means between the groups (p-value < 0.05) based on Tukey’s HSD one-way ANOVA using PAST version 3.14, Oslo, Norway.

Figure 6

Ascorbic binding modes with sodium-dependent vitamin C transporter-2 (SVCT2) (pdb code: 4RP9). (a) Superimposition of ascorbic acid crystal pose (yellow) and dock pose (cyan); (b) van der Waals interaction of the residues around the active sites, with ascorbic acid dock pose.

Figure 7

Binding modes of ascorbic derivatives and vitamin C transporter 2 (pdb code: 4RP9). (a) Binding position of 5c (magenta) compared with ascorbic acid (cyan), and (b) binding position of 2c (yellow) compared with ascorbic acid (cyan).

Scheme 1

The synthesis route of ascorbic derivatives. (a) K₂CO₃, propargyl bromide, DMSO/THF (9:8), rt, 2 h. (b) N₃R, 5% mol CuSO₄, 20% mol NaAsc, t-BuOH/EtOH/H₂O (2:2:1), rt, 2 h. (c) 2,2 dimethoxypropane, p-toluenesulfonyl chloride, rt, 24 h. (d) K₂CO₃, propargyl bromide, DMSO/THF (9:8), rt, 2 h. (e) 12 N HCl, THF, rt, 2.5 h. (f) N₃R, 5% mol CuSO₄, 20% mol NaAsc, t-BuOH/H₂O (4:2) or t-BuOH/EtOH/H₂O (4:4:2), rt, 2 h.

Scheme 2

The synthesis route of (3-azidomethyl)tryptoline, c. (a) MeOH, H₂SO₄, reflux, 18 h. (b) NaBH₄, MeOH/THF (1:1), reflux, 18 h. (c) NsCl, TEA, CH₂Cl₂, rt, 4 h. (d) NaN₃, DMF, 70 °C, 6 h. (e) p-toluenethiol, DMF, 50 °C, 2 h.