Synthesis, Characterization, and Remarkable Biological Properties of Cyclodextrins Bearing Guanidinoalkylamino and Aminoalkylamino Groups on Their Primary Side

Abstract

The introduction of aminoalkylamino and guanidinoalkylamino substituents on the primary side of beta- and gamma-cyclodextrin (CDs) resulted in a series of novel compounds that were extensively characterized by NMR spectroscopy and mass spectrometry. Bromination of the primary side of beta- and gamma-CD, and reaction with neat alkylene diamines at a pressure of 7 atm afforded aminoalkylamino derivatives that were then guanylated at the primary amino group to give the corresponding guanidinoalkylamino-CDs. These compounds are water soluble and display pK(a) values that allow them to be mostly protonated at neutral pH; for example, pK(a(1)) approximately 6.4 and pK(a(2)) approximately 9.5 for the aminoethylamino-beta-CD and pK(a(1)) approximately 7.8 and pK(a(2)) approximately 11.0 for the guanidinoethylamino-beta-CD. The title CDs are rigid, cyclic alpha-D-glucopyranose oligomers (heptamers or octamers) with branches that resemble lysine and arginine side chains that enable multiple interactions with suitable substrates. Thus, they bear similarities to known cell-penetrating peptides. Indeed, the compounds were found to cross the membranes of HeLa cells and penetrate inside the cytoplasm quickly, the guadinylated ones within 15 min, as shown by fluorescence microscopy using fluorescein-labeled derivatives. The toxicity of the compounds, measured by performing MTT tests, ranged from 50 to 300 microM. Furthermore, some of the aminated CDs could facilitate the transfection of DNA expressing the green fluorescent protein (GFP) in HEK 293T cells, with effectiveness comparable to the commercial agent Lipofectamine 2000. Circular dichroism, atomic force microscopy and electrophoresis experiments confirmed the strong interaction of the compounds with DNA. Because of their carbohydrate, non-peptide nature the title compounds are not anticipated to be enzymatically labile or immunogenic, and thus they fulfill many of the criteria for non-hazardous transport vectors in biological and pharmaceutical applications.

Preorganized, Macromolecular, Gene‐Delivery Systems

“…Interestingly, the transfection efficiency of these vectors against human embryonic kidney HEK 293T cells favorably compared with that of the commercial cationic lipid formulation Lipofectamine 2000 . [61] Reineke and co-workers took advantage of the copper(I)-catalyzed Huisgen 1,3-dipolar azide-alkyne cycloaddition reaction [62] between the per-(O-2,O-3)-acetylated heptaazide 27 and acriloyl amide derivatives to synthesize a family of polycationic b-CD "click clusters" (28) bearing seven linear oligoethyleneimine branches with variable, but controlled, number of protonable amino groups (Scheme 5). [63] Agarose gel electrophoresis, DLS, and TEM experiments revealed that the click polycations complexed pDNA and protected it from nuclease degradation by forming nanoparticles with an average diameter of 80-130 nm.…”

Binding of Nucleotides and Nucleosides to Per(6‐guanidino‐6‐deoxy)cyclodextrins in Solution

“…[8] The mode of binding to DNA is not known for these CDs or for other CD-based transfection agents modified on the primary side with elaborate positively charged groups. [9][10][11] The present work is an NMR spectroscopic study of the interactions of guanidino CDs with nucleic acid components, namely, the nucleotides 2Ј-deoxyadenosine 5Ј-monophosphate (5Ј-dAMP), adenosine plexes were 1:2, except for 5Ј-dAMP/bg where oligomeric aggregates formed.…”

Insights in cellular uptake mechanisms of pDNA–polycationic amphiphilic cyclodextrin nanoparticles (CDplexes)

“…More recently, monodisperse polycationic CD conjugates with promising pDNA delivery capabilities were prepared by homogeneous functionalization of the CD primary rim (22)(23)(24). Aiming at merging the virtues of both cationic polymers and cationic lipids in a flexible, well defined framework, we have reported a modular synthetic strategy for the preparation of polycationic amphiphilic CDs (paCDs) that allows the installation of different building blocks onto either face of the truncated-cone structure in a sequential and controlled way, offering a unique opportunity for structure-activity relationship (SAR) studies (25)(26)(27) (Figure 1).…”