Peptoids and polyamines going sweet: Modular synthesis of glycosylated peptoids and polyamines using click chemistry

Sugar moieties are present in a wide range of bioactive molecules. Thus, having versatile and fast methods for the decoration of biomimetic molecules with sugars is of fundamental importance. The glycosylation of peptoids and polyamines as examples of such biomimetic molecules is reported here. The method uses Cu-catalyzed azide alkyne cycloaddition to promote the reaction of azidosugars with either polyamines or peptoids. In addition, functionalized nucleic acids were attached to polyamines via the same route. Based on a modular solid-phase synthesis of peralkynylated peptoids with up to six alkyne groups, the latter were modified with azidosugar building blocks by using copper-catalyzed azide alkyne cycloadditions. In addition, the up-scaling of some particular azide-modified sugars is described.

Multiplicities of signals are described as follows: s = singlet, bs = broad singlet, d = doublet, t = triplet, q = quartet, m = multiplet. Coupling constants (J) are given in hertz (Hz). Multiplicities in the 13 C NMR spectra were determined by DEPT (distortionless enhancement by polarization transfer) measurements. Mass spectra (ESI) were obtained by using an Agilent 6230 TOF LC/MS. QToF electrospray-ionisation (ESI) MS was performed with a Micromass QTof2 spectrometer. MALDI-TOF mass spectra were obtained by using either a Bruker Biflex IV spectrometer with a pulsed ultraviolet nitrogen laser, 200 µJ at 337 nm and a time-of-flight mass analyzer with a 125 cm linear flight path, or a Micromass TOFSpecE spectrometer in reflectron mode. 2,5-Dihydroxybenzoic acid and -cyano-4-hydroxy cinnamic acid were used as the matrix. HPLC was performed on a Jasco HPLC system, with a C18 column (30 × 190 mm). Flow rate: 15 mL/min; solvent A: 0.1% TFA in water; solvent B: 0.1% S3 TFA in MeCN. Analytical TLC was performed on MERCK ready-to-use plates with silica gel 60 (F254). Column chromatography: MERCK silica gel 60, 0.04-0.063 mm.
The analytical data of resin-bound substrates were taken from the unpurified free product after test cleavage from the resin.

General procedures for solid-phase synthesis
General washing procedure for resin. In the steps with two solvents they were used alternatingly. Per 1 g of the resin, 50 mL solvent was used.

Nosyl (Ns) deprotection.
For cleavage of the Ns protecting group the resin was swelled in DMF for 15 min. Then, 20.0 equiv of -mercaptoethanol and 20.0 equiv of DBU were added. After agitation overnight the reaction mixture was removed and the resin was washed according to procedure A.
Cleavage from the resin. The product was cleaved from the resin by adding 1% TFA in CH 2 Cl 2 . Meanwhile the color of the resin turned to red. After 5-10 min the product was washed off with CH 2 Cl 2 and MeOH, followed by evaporation of the solvent in high vacuum.

Synthesis of azidosugars 1-3:
To a solution of D-mannosamine/ D-glucosamine/ Dgalactosamine hydrochloride (1.00 g; 4.64 mmol) in methanol (50 mL), sodium methanolate (30% w/w NaOMe in MeOH, 1.66 mL, 4.64 mmol, 1.00 equiv) was added and the mixture was stirred at room temperature for 30 min until complete dissolution. NEt 3 (0.47 g; 4.64 mmol, 1.00 equiv) and chloroacetic anhydride (871 mg; 5.10 mmol, 1.10 equiv) were added to the solution, and stirred at room S4 temperature overnight. The solvent was evaporated and the crude product was used in subsequent reaction without further purification. Comment: If necessary, sodium bicarbonate was added to neutralize the solution.
Subsequently, the reaction mixture was concentrated and dried in vacuo. The residue was then suspended in pyridine (20 mL) and acetic anhydride (20 mL) was added to the solution. The reaction was stirred at room temperature overnight. After concentration in vacuo, the residue was dissolved in EtOAc (50 mL) and washed with 1N HCl, NaHCO 3 and brine (each 50 mL) (CAUTION: extraction with sodium bicarbonate causes gas formation and excess pressure in the separating funnel).

Sperminyl-
The resin was agitated for 16 h, then washed according to procedure A and dried under reduced pressure. After drying, 1.94 g (1.00 equiv) resin and 1.45 g (15.0 equiv) K 2 CO 3 were suspended together in 20 mL DMF. Then, 1.0 mL (10.0 equiv) of 5-chloropent-1-yne was added and the mixture was agitated at 60 °C for 16 h. After cooling down, the solvent was removed and the resin was washed following procedure A until all K 2 CO 3 was removed. An orange resin was obtained after drying under reduced pressure. NMR data and MS were obtained after cleavage from the resin according to the general procedures for solid-phase synthesis.  1H-1,2,3-triazole) 13. A suspension of 200 mg (1.00 equiv) resin 9 in 2 mL DMF reacted with 12.3 mg (0.500 equiv) CuSO 4 ·5H 2 O in 1 mL DMF, 97.72 mg (5.00 equiv) sodium ascorbate in 0.5 mL aqua dest. and 40.08 mg (2.00 equiv) 1-azido-1-deoxy--D-glucopyranosid or 71.75 mg (2.00 equiv). After agitating for 2 d the reaction mixture was removed and the resin was washed according to procedure B. After deprotection of the nosyl group and washing of the resin according to procedure A the products were cleaved from the resin according to general procedures.  N-(N'''-1-β-D-lactopyranosyl-4-propyl-1H-1,2,3-triazole)
After agitating for 2 d the reaction mixture was removed and the resin was washed according to procedure B. After deprotection of the nosyl group and washing of the resin according to procedure A the products were cleaved from the resin according to general procedures.

S8
The suspension was agitated for 2.5 d, then the reaction mixture was removed from the resin. After that, the orange resin was washed according to procedure B, deprotected from the Nosyl group and dried under vacuum.

Synthesis of hexaalkyne 27 on 2-chloro tritylchloride resin
For the synthesis, 50 mg (0.103 mmol, 1.00 equiv) of 2-chlorotrityl chloride resin and washed in a fritted plastic syringe (Multisyntech) with 1 mL of dichloromethane, followed by swelling in 1 mL of dichloromethane for 5 min. The first submonomer was added by reacting 77.2 mg of bromoacetic acid (0.555 mmol, 5.40 equiv) and 99 µL of DIPEA (71.8 mg, 0.555 mmol, 5.00 equiv) in 1 mL of dichloromethane on a shaker S9 platform for 40 min at room temperature, followed by washing with dichloromethane (three times with 2 mL) and DMF (three times with 2 mL). The bromoacetylated resin

Cu-catalyzed alkyne azide cycloaddition on resin 27 and cleavage from the resin
For the reaction with the sugar, the resin 27 was swollen in THF, and 70 mg Cu(CH 3 CN) 4 PF 6 (0.188 mmol, 1.83 equiv) was directly weighed into the plastic syringe containing the resin. In 2 mL dry THF, 310 mg of Ac 4 GalNAz (2, 0.721 mmol, 7.00 equiv) and 100 µL 2,6-lutidine (92.0 mg, 0.859 mmol, 8.34 equiv) were dissolved and added to the resin. The resin was shaken for 18 h at rt.

S10
After the reaction, the resin was washed with saturated aqueous sodium ascorbate solution, water, THF and dichloromethane (three times 2 mL each).
The peptoid was cleaved from the resin using 3 mL 33% HFIP in dichloromethane (v/v)