2β-3,4-Unsaturated sialic acid derivatives: Synthesis optimization, and biological evaluation as Newcastle disease virus hemagglutinin-neuraminidase inhibitors

Abstract

The optimization of the synthetic protocol to obtain the 3,4-unsaturated sialic acid derivatives, through the fine-tuning of both the Ferrier glycosylation conditions and the subsequent hydrolysis work-up, is herein reported. The accomplishment of the desired β-anomers and some selected α-ones, in pure form, led us to evaluate their specific inhibitory activity towards NDV-HN and human sialidase NEU3. Importantly, the resulting data allowed the identification, for the first time, of three active 3,4-unsaturated sialic acid analogs, showing IC₅₀ values against NDV-HN in the micromolar range.

Introduction

Paramyxoviridae is a family of viruses that can infect humans and animals, ultimately causing also very severe diseases.¹ Interestingly, the hemagglutinin-neuraminidase (HN), a key enzyme present on the envelope of the majority of these viruses and involved in crucial steps of viral contagion (binding, fusion and spreading) has been shown to be an effective target to block viral infections.¹ In particular, several 2,3-unsaturated N-acetylneuraminic acid analogs have been developed as competitive neuraminidase inhibitors, including the natural derivative DANA 1a,^2a and some antivirals against the human parainfluenza viruses (hPIVs), as BCX-2798 1b,^2b,c and Newcastle disease virus (NDV), as compounds 1c-g^2d,e (Fig. 1).

Similarly, various 3,4-unsaturated Neu5Ac analogs, such as 2a,b or 3a,b, have been tested, as potential neuraminidase inhibitors against human influenza viruses^3a and hPIVs.^3b,c

In this regard, the synthesis of the 3,4-unsaturated sialic acid derivatives was originally accomplished by Maudrin^3a through a multistep synthetic pathway and, more recently, these molecules have been achieved using a rapid method proposed by Ikeda and coworkers^3b,c via a Ferrier glycosylation reaction on oxazoline 4, followed by hydrolysis (See Table 1).

However, despite the high yields and β-stereoselectivity reported in the Ferrier reaction (using methanol or ethanol as nucleophiles), a drastic yield reduction could be observed when alcohols or thiols with longer or branched chains were subjected to the same reaction conditions. Moreover, the hydrolytic deprotection caused an undesired variation in the anomeric ratios.^3b,c Thus, regrettably, the final compounds could be tested as hPIV-HN inhibitors^3c only as α/β diastereomeric mixtures.

Along this line, we recently developed a novel ¹³C NMR method to unequivocally assign the anomeric configuration of several 3,4-unsaturated Neu5Ac derivatives via 1,7-lactonization.4a), 14, 15, 16 Furthermore, we reported an exhaustive explanation of the rearrangement mechanism, under aqueous acidic conditions, of the oxazoline 4,^4b the starting material in the Ferrier reaction.^3b,c Thus, in our continuous effort to provide new synthetic tools in this field,2d), 4 we report here an efficient protocol to obtain the β-anomers of 3,4-unsaturated sialic acid derivatives, as they represent suitable intermediates for the synthesis of 3 or/and 4-substituted DANA derivatives, as well as new potential neuraminidase inhibitors. Indeed, we evaluated the biological inhibitory activity of the obtained derivatives on a selected Paramyxoviridae HN, the NDV-HN, as well as their selectivity for this enzyme as compared to the human sialidase NEU3.