Synthesis, micellization and lectin binding of new glycosurfactants

doi:10.1016/j.carres.2014.07.021

Carbohydrate Research

Volume 397, 9 October 2014, Pages 31-36

https://doi.org/10.1016/j.carres.2014.07.021 Get rights and content

Highlights

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Biosourced amphiphiles were synthesized from carbohydrates, fatty acids, and PEG.
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Monodisperse micelles were obtained by self-assembly in water and characterized by DLS and SAXS.
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Biodisponibility of the surface-carbohydrates for lectin targeting was evidenced.

Abstract

Here we report the preparation and physico-chemical characterization of carbohydrate-decorated micelles and their interaction with lectins. A library of biosourced amphiphiles was prepared by copper-catalyzed azide–alkyne cycloaddition (CuAAC) between alkynyl sugars (lactose, N-acetyl-d-glucosamine) and azido-functionalized poly(ethylene glycol) esters (N₃-PEG₉₀₀-decanoate (C₁₀) and -dodecanoate (C₁₂)). In water, these glycoconjugates self-assemble into micelles of homogeneous nanometric size (11 nm) as evidenced by scattering techniques (DLS for light, and SAXS for X-ray). A comparative study with previously synthesized octadecanoate counterparts pointed out that that nature of the fatty acid has no significant influence on the particle size but only affects their compactness. These findings are in favor of a possible bulk preparation from lipid mixtures such as those encountered in renewable vegetable oils. The presence of the carbohydrate epitopes on the surface of the micelles and their bioavailability for lectin targeting were also evidenced by light scattering measurements using wheat germ agglutinin (WGA) and peanut (Arachis hypogaea) (PNA) lectins, supporting possible application as targeted drug nanocarriers.

Graphical abstract

Introduction

Nanoparticle-based biotechnology and its medical applications are rapidly heading to the forefront of drug delivery, diagnostic and other areas. Presently, several systems based on polymeric nanoparticles have been proposed as drug delivery nanocarriers. Those produced by self-assembly of amphiphilic macromolecules are very attractive candidates.1, 2, 3 Particularly, spherical micelles can be prepared using relatively simple protocols and their large surface-to-volume ratio allows high-capacity drug loading. Such nanoparticles proved attractive for passive tumor-targeting in cancer therapy. Indeed, uptake of drug-loaded nanocarriers at tumor sites can occur because of their usually leaky microvasculature. This is known as the enhanced permeation and retention (EPR) effect⁴ which is optimized when the size of the nanoparticles is below the cut-off size of the typical tumor pores (∼200 nm).⁵ Even though it is well established that nanoparticles are preferentially accumulated at tumor sites as a result of the EPR effect,6, 7 the tumor-targeting can be achieved actively through the use of specific biomarkers.⁸ Ideally, the nanocarriers should be decorated with targeting ligands to allow active drug delivery at specific receptor sites and sufficiently large (2R_H > 5 nm) to avoid fast renal clearance thus ensuring prolonged blood circulation time.9, 10

The design of functional nanoparticles possessing bio-affinity properties is a challenging endeavor in the field of nanobiotechnology and it is currently the subject of intensive research with important outcomes in medicine for cancer diagnosis and therapy. Antibodies have been extensively investigated as targeting biomolecules, however their immunogenicity limits therapeutic applications and the carbohydrate-decoration has been revealed a promising alternative.¹¹ The mechanism of sugar-mediated targeting is essentially based on specific interaction between carbohydrates and cell membrane proteins called lectins.¹² The binding of lectins typically involves the two or three terminal sugar residues of the mammalian glycans including galactose, mannose, N-acetyl-neuraminic acid, fucose, or N-acetyl-glucosamine. Therefore, the development of nanoparticles whose outer shell is decorated by glycoconjugates aiming at lectin targeting is thus considered as a promising way to improve drug delivery and cell internalization.13, 14

While carbohydrate-based amphiphiles such as sugar esters and alkyl glycosides have long been used for their surfactant properties,15, 16 a renewed interest for these molecules has raised with the desire to make a better use of the extraordinary rich saccharidic biomass and with the emergence of green chemistry. In particular, the click chemistry introduced by Sharpless¹⁷ as efficient and universal ligation reaction gave rise to exciting perspectives in many areas including biology¹⁸ and material sciences.¹⁹ Thus, a wide range of new glycosurfactants with various applications has been recently reported in the literature. One can notice maltose-based supramolecular hydrogelators,²⁰ fluorescent sucrose probes for membrane optical imaging²¹ or stabilizers for quantum dots nanoparticles.²² Glycosylated liposomes have also been largely studied for cell-selective delivery of drugs, in particular for nucleic acids.23, 24 However, the rapid clearance of circulating liposomes from the bloodstream, coupled with their high uptake by liver and spleen is often a limitation. Polymeric micelles, showing better thermodynamic stability in physiological solution, a narrow size distribution, stealth properties induced by the hydrophilic polymeric brush on the micellar surface, and great capacity to encapsulate hydrophobic drugs have thus emerged as a promising alternative to liposomes.²⁵

Taking into account the above considerations, the present work reports the synthesis, self-assembly, and the interaction with lectins of a library of new glycosylated polymeric amphiphiles. The main objective is to move forward in the development of novel carbohydrate-decorated micelles whose surface allows the site-directed vectorization of active molecules through receptor–ligand binding. A series of new amphiphilic glycoconjugates was prepared from mono-azido end-functionalized poly(ethylene glycol) following a recently reported procedure.²⁶ PEG is commonly used to produce stealth nanocarriers and a molecular weight of 900 g mol⁻¹ was selected to provide a final hydrophobic volume fraction close to ∼20% which is required for the formation of spherical micelles.²⁷ The target macromolecular amphiphiles were obtained after esterification of N₃-PEG₉₀₀-OH with fatty acids (decanoic and dodecanoic acids) and subsequent ‘click-glycosylation’ with lactose and N-acetyl-β-d-glucosamine through copper-catalyzed azide–alkyne cycloaddition.17, 28 Self-assembly and lectin adhesion properties of the glycoconjugates were characterized by means of dynamic light scattering (DLS), fluorescence spectroscopy, small-angle X-ray scattering (SAXS).

Section snippets

Materials

All reagents were of commercial grade and were used as received unless otherwise noticed. The solvents were dried and distilled prior to use.

Synthesis of the amphiphiles

C₁₀PEG₉₀₀GlcNAc, C₁₂PEG₉₀₀GlcNAc, C₁₀PEG₉₀₀Lac and C₁₂PEG₉₀₀Lac were synthesized according to the previously reported procedure.²⁶ Experimental details and characterization are reported in Supplementary information file.

Nanoparticles preparation

The aqueous micellar solutions were prepared by direct dissolution of the amphiphiles in Milli-Q water at typically 0.5 mg mL⁻¹, followed by

Synthesis and characterization of the amphiphiles

In a previous work, we have reported the synthesis and self assembly of amphiphilic stearate-based glycoconjugates into homogeneous nanometric micelles.²⁶ In the present study, we aimed to assess the influence of the fatty acid chain on the micellization and synthesized new glycoconjugates incorporating C₁₀ and C₁₂ fatty acids. Esterification of N₃-PEG₉₀₀-OH 1²⁶ with decanoic and dodecanoic acids in the presence of dicyclohexylcarbodiimide afforded 2 and 3 in 81% and 78%, respectively.

Conclusion

A library of sugar-functionalized amphiphiles containing saturated lipids of different molecular weights was developed. The carbohydrate groups (GlcNAc and lactose) were efficiently introduced onto azido-terminated poly(ethylene glycol) fatty acid esters by CuAAC click chemistry. All the amphiphiles, whatever the size of the fatty acid, self-assembled into highly monodisperse micelles upon simple dissolution in water. Interestingly, all the particles displayed a similar size with a mean

Acknowledgments

The authors acknowledge the support from CNRS, Carnot Polynat, Labex Arcane (ANR-11-LABX-0003-01), CAPES-COFECUB (Project 620/08) and from the Mass spectrometry and NMR platforms of ICMG (FR 2607). The ESRF and LNLS are acknowledged for supplying the beam time (proposals 02 01 784 and 12356, respectively). The technical assistance of Dr. C. Rochas and Dr. B. Jean during the experiments at ESRF is greatly acknowledged. F.C.G. thanks FAPESP (Grant 2010/06348-0). A.G.D.B. acknowledges the

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Synthesis, micellization and lectin binding of new glycosurfactants

Highlights

Abstract

Graphical abstract

Introduction

Section snippets

Materials

Synthesis of the amphiphiles

Nanoparticles preparation

Synthesis and characterization of the amphiphiles

Conclusion

Acknowledgments

J. Controlled Release

Adv. Drug Delivery Rev.

Biochim. Biophys. Acta

J. Controlled Release

Carbohydr. Res.

Prog. Polym. Sci.

Nat. Nanotechnol.

Pharm. Res.

Proc. Natl. Acad. Sci. U.S.A.

Cancer Res.