N-Amino-imidazolin-2-one Peptide Mimic Synthesis and Conformational Analysis

Abstract

Base-promoted 5-exo-dig cyclizations of aza-propargylglycinamides provided N-amino-imidazolin-2-one peptide mimics, which exhibited turn geometry in X-ray crystallographic and NMR spectroscopic analyses. Sonogashira coupling prior to cyclization afforded N-amino-imidazolin-2-ones with diverse 4-position aromatic substituents with potential to serve as Phe and Trp mimics.

Identification of biologically active conformers is critical for developing therapeutics based on peptide structures, because precise folding is essential for function. Geometrically restricted analogs are thus valuable tools, because they may reduce energetic costs for folding into binding conformations and, thereby, improve potency, selectivity, and stability.¹

To constrain backbone geometry and induce turn conformations, α-amino-γ-lactams,² so-called Freidinger–Veber lactams, have been commonly introduced into peptide sequences; however, their lack of side-chain functions may translate into loss of affinity and activity. Aza analogs of amino acids possess a nitrogen atom in place of the CHα. A variety of side chains have been installed onto these semicarbazide structures, which when introduced into azapeptides restrict the backbone ϕ and ψ dihedral angles, due to the lone pair–lone pair electronic repulsion of the adjacent nitrogen and urea planarity, respectively.³ A strategy has now been devised to induce peptide turn geometry by combining the covalent constraints of α-amino-γ-lactams with the electronic restrictions and side-chain diversity of aza-amino acids through the synthesis of substituted N-amino-imidazolin-2-ones (Figure 1).

Figure 1.

N-Amino-imidazolin-2-one turn mimic conception.

Imidazolin-2-ones have utility as antitumor agents,⁴ antibacterial MurB inhibitors,⁵ dopamine D₄ and CGRP receptor antagonists,^6,7 antioxidants,⁸ and unnatural base pairs.⁹ To the best of our knowledge, however, the synthesis and biological evaluation of N-amino-imidazolin-2-ones had not been explored.

Previously, the submonomer approach for azapeptide synthesis surmounted issues of hydrazine chemistry to give access to side chains inaccessible by traditional methodology, including propargyl, allyl, and (hetero)aryl moieties.^10,11 The aza-propargylglycine side chain was later reacted in copper-catalyzed 1,3-dipolar cycloadditions to make aza-1,2,3-triazole-3-alaninyl peptide mimics.¹² Aza-propargylglycinamides have now been explored in 5-exo-dig cyclizations to access N-amino-imidazolin-2-one peptidomimetics, as well as in Sonogashira cross-coupling reactions prior to cyclization to provide their 4-arylmethyl analogues, which may mimic phenylalanine and tryptophan residues.

Imidazolin-2-ones and imidazolidin-2-ones have been respectively prepared from propargylic and allylic ureas by base-promoted 5-exo-dig cyclizations.¹³ Annulation has typically necessitated an electron-deficient urea nitrogen and activation of the π-system using transition metal salts (i.e., silver,¹⁴ palladium,¹⁵ and gold complexes).¹⁶ Moreover, cyclic amino acids, such as dehydroprolines, had been prepared respectively by Pd- and Ag-catalyzed 5-endo-dig cyclization of N-Ts- and Boc-protected propargylglycine analogs.^17,18

To study the cyclization, aza-propargylglycinyl dipeptide 1 was used and prepared by chemoselective alkylation of benzhydrylidene aza-glycinyl-d-phenylalanine tert-butyl ester (5) with propargyl bromide (Schemes 1 and 2).¹⁹ Attempted 5-exo-dig cyclization of azadipeptide 1 using homogeoneous gold catalysis [(t-Bu)₂(o-biphenyl)PAuCl (5 mol %) and AgOTf (5 mol %)] failed, likely because the urea nitrogen was insufficiently electron-deficient.

Scheme 1. N-Amino-imidazolin-2-one synthesis.

Scheme 2. Synthesis of N-(p-Methoxybenzamido)imidazolin-2-one Isopropyl Amide (10).

N-Amino-imidazolin-2-one 2 was, however, obtained in 81% yield, by adding 2.5 equiv of NaH to the mixture containing 1 and the cationic gold complex formed in situ in acetonitrile for 2 h. The impact of gold catalysis was later deemed negligible, because 2 was produced in 84% yield on reaction of 1 with 2.5 equiv of NaH in acetonitrile without a catalyst (Scheme 1). From the 5-exo-dig cyclization, an exocyclic double bond was first produced and migrated inside the ring to furnish the thermodynamically more stable N-amino-imidazolin-2-one 3. Among the solvents studied, acetonitrile proved the best (see Supporting Information (SI)). Excess NaH was necessary for high yields.

To study the effect of N-amino-imidazolin-2-one on peptide conformation, model 10 was synthesized and examined by X-ray crystallography and NMR spectroscopy (Scheme 2). Benzhydrylidene aza-glycinyl-d-phenylalanine isopropyl amide 7 was made from 5 by tert-butyl ester cleavage in a 1:1 v/v mixture of TFA/DCM and coupling to isopropylamine by way of a mixed anhydride.²⁰ Alkylation of semicarbazone 7 with propargylbromide^19,20 gave aza-propargylglycinamide 8 in 71% yield without detectable racemization; however, subsequent NaH-promoted 5-exo-dig cyclization gave imidazolin-2-one 9 in 78% yield with 10% racemization (see SI). Olefin migration occurred upon hydrazone removal, using hydroxylamine hydrochloride in pyridine,⁹ to afford N-amino-imidazolone hydrochloride which, without further purification, was treated with 4-methoxybenzoyl chloride to provide N-acyl dipeptide amide 10 in 56% overall yield.

Crystals were grown by slow diffusion of hexanes into an ethyl acetate/chloroform solution of 10. X-ray diffraction revealed two turn conformations in the solid state (Figure 2): 10a exhibiting a type II′ β-turn with an intramolecular ten-membered hydrogen bond between residues i and i + 3, and 10b showing a seven-membered hydrogen bonded conformer in an inverse γ turn. The X-ray structures for 10 deviate primarily by rotation of the ψ_i+2 dihedral angle, as shown by comparison of their φ and ψ dihedral angles with an ideal turn geometry and crystal structures of azapeptide and α-amino-γ-lactams, which adopted a turn geometry (Table 1).²¹⁻²³ In contrast to amino lactams, the planar geometry of the N-amino-imidazalone causes the ψ_i+1 dihedral angle to deviate by 33°–46° from that of an ideal type II′ β-turn, the geometry of which is contingent on the stereochemistry of the C-terminal residue (i.e., Phe).

Figure 2.

X-ray structures of N-(amido)imidazolin-2-one amide 10. Broken lines represent inferred hydrogen bonds.

Table 1. Structures 10–13 and Their ϕ and ψ Dihedral Angles (in degrees) from Crystal Analyses Compared with Ideal Turns.

type of turn	φi+1	ψi+1	φi+2	ψi+2
β-II′	60	–120	–80	0
inverse γ	n/a	n/a	–70	60
10a	58.9	–153.3	–69.1	–4.6
10b	62.1	–166.1	–71.7	65.7
β-II	–60	120	80	0
11	–55.4	120.9	89.3	17.8
12	–42	133	89	–6.9
13	–40	116	96	–97

Measurement of the amide chemical shift values of 10 as a function of DMSO-d₆ % (1 to 100%) in CDCl₃ indicated relatively little variation (0.45 ppm) for the isopropylamide NH signal compared to the benzamide chemical shift (1.21 ppm; see SI), consistent with solvent-shielded (hydrogen-bonded) and exposed hydrogens,²⁴ as found in the X-ray structure.

To access constrained Phe, Trp, and His mimics, Sonogashira couplings were performed on dipeptide 1, using various aryl iodides, Pd(PPh₃)₂Cl₂, and CuI in a 1:1 DMF/Et₂NH mixture (Scheme 3, Table 2). Electron-rich and -poor aryl iodides as well as N-protected indole and imidazole iodides all reacted in the coupling reaction to furnish aza-arylpropargylglycines 14 in 50–93% yields.

Scheme 3. Sonogashira/Cyclization Reaction Sequence for the Synthesis of 4-Substituted N-Amino-imidazolin-2-ones.

Table 2. Sonogashira/Cyclization Reaction Sequence Yields.

^aStarting material was recovered.

Exposure of 14 to the NaH-promoted 5-exo-dig cyclization produced mixtures possessing endo- and exocyclic double bonds. For example, imidazolin-2-ones 15a and 16a were isolated as isomeric mixtures in 69% yield. Although either Z or E geometry were possible for 15, a through-space interaction between the vinyl proton and the methylene of imidazolidinone 15c in a 2D NOESY experiment revealed an exclusive exocyclic Z double bond geometry. Acid cleavage of the tert-butyl ester promoted double bond migration inside the five-membered ring to furnish 17a (Scheme 3).²⁵

In the NaH-promoted 5-exo-dig cyclization, the fluorine p-substituent was well tolerated and gave 15c in 64% yield (Table 2). Substrates 14 with electron-withdrawing substituents (i.e., trifluoromethyl) reacted rapidly giving complete consumption of the starting material, albeit with lower yields due to decomposition. In contrast, electron-rich aza-p-methoxyphenylpropargylglycinamide 14b afforded N-amino-imidazolin-2-one 15b in only 10% yield with recovered starting material. Imidazolyl alkyne 14g failed to react and was exclusively recouped. In contrast, N-Boc-3-indolyl alkyne 14f underwent base-promoted cyclization to afford constrained tryptophan mimic imidazolin-2-one 15f in 40% yield with recovered starting material.

4-Substituted N-amino-imidazolin-2-ones have been prepared as hybrids of the covalent and electronic constraints of α-amino-γ-lactams and aza-amino acids. Opportunity for adding side-chain functionality was demonstrated by using a Sonogashira arylation prior to NaH-promoted 5-exo-dig cyclization of aza-propargylglycinamide to afford 4-substituted N-amino-imidazolin-2-one mimics. The propensity of the N-amino-imidazolin-2-one subunit to induce turn conformations was confirmed using X-ray crystallography and NMR spectroscopy of model peptide 10. Considering their conformational preferences and potential for their diversification, N-amino-imidazolinones represent a promising class of geometrically restrained mimics for studying peptide structure. Incorporation of N-amino-imidazolinones into a biologically active peptide sequence is currently under investigation and will be reported in due time

The Table 1 graphic contained an error in the version published ASAP August 14, 2012. The correct version reposted August 22, 2012.

Supporting Information Available

Experimental procedures, compound characterization data, and NMR spectra for new compounds. Crystallographic information files (CIF). This material is available free of charge via the Internet at http://pubs.acs.org.

The authors declare no competing financial interest.