Triazoles as amide bond isosters

Pursuing the idea of replacing amide bonds by proteolytically stable and planar chemical groups, the triazole, which shares electronic and topologic characteristics with natural peptide bonds, emerged in the focus of peptidomimetic chemists.³⁰ The 1,2,3-triazoles represent a class of aromatic heterocycles, whose characteristics make them promising candidates for the development of bioactive peptidomimetics with improved pharmacologic properties.³¹ Like amide bonds, 1,2,3-triazoles are planar, have a strong dipole moment and are capable of accepting and donating hydrogen bonds. While the distance between the α-carbons of 1,5-disubstituted triazoles matches quite well for the cis amide bond (3.0 compared to 3.2 Å), the distances for the trans amide bond does not comply well (3.7 compared to 4.9 Å) with the 1,4-disubstituted triazole (Figure 13 ).³¹

Figure 13. Comparison of trans- and cis amide bonds and their resembling 1,4- and 1,5-disubstituted triazole surrogates.

The synthesis of triazoles by a cycloaddition between mono-substituted alkynes (as dipolarophiles) and azides (as 1,3-dipolar components) was first described by Michael et al.

in 1893 and reviewed by Huisgen.³² The thermal reaction between phenylazide and phenylacetylide gave a nearly 1:1 regioisomer-mixture of the 1,4- and 1,5-disubstituted diphenyltriazole.

Since the independent discovery of the copper(I) catalysed azide alkyne cycloaddition (CuAAC) by the groups of Meldal and Sharpless in 2002 (Scheme 7),^{33, 34} which leads exclusively to 1,4-disubstituted 1,2,3-triazoles, the CuAAC has found wide spread use by the chemical community.³⁵

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Scheme 7. CuAAC between Propargyl phenyl ether and benzylazide in aqueous media without the exclusion of oxygen. The reductive “CuSO4/sodium ascorbate”-system serves as a regenerative Cu^I source; a) CuSO4.5H2O (1 mol%), sodium ascorbate (5 mol%), H2O/^tBu (2:1), RT, 8 h.³⁴

Several key features account for the popularity of this versatile reaction. It proceeds very selectively and without side products also in aqueous media and is orthogonal towards biologically relevant functional groups. Due to its perfect atom economy and straightforward workup, Sharpless and co-workers defined the CuAAC as a prototype of a “click-reaction”.³⁶ Because the reaction mechanism is initiated by a formation of a copper acetylide derivative, the CuAAC can be only performed with terminal alkynes.³⁴

In 2005, the groups of Fokin and Jia published its counterpart, the ruthenium(II) catalysed azide alkyne cycloaddition (RuAAC) based on the complex Cp*RuCl(L2), which enabled the selective synthesis of the 1,5-disubstituted 1,2,3-triazole regioisomer (Scheme 8).³⁷

Scheme 8. RuAAC between 2-ethynylnaphthalene and benzylazide under elevated temperatures; a) Cp*RuCl(PPh3)2 (1 mol%), benzene, 80 °C, 4 h.³⁷

Although the reaction does not tolerate water or protic solvents, it is quite tolerable towards functional groups like alcohols, aldehydes, alkenes, amides, amines, boronic esters, ketones and halides.³⁸ Since the reaction is initiated by a π-complex between the acetylide and the Ru(II) center, the reaction also tolerates internal alkynes as substrates, which would lead to trisubstituted triazoles with another possible regioisomeric combination.

A recent publication from Kim et al. in 2017 describes the nickel(II) catalysed [3+2] alkyne azide cycloaddition to obtain 1,5-disubstitued triazoles in aqueous media under oxygen atmosphere, thereby addressing the disadvantages of the RuAAC (Scheme 9).³⁹

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Scheme 9. Optimized reaction condition example of a NiAAC between benzylazide and phenylacetylide. The regioisomers 1,5-disubstitued and 1,4-disubstituted triazole were isolated in a ratio of 94% to 6%; a) Cp2Ni (10 mol%), Xantphos (10 mol%), Cs2CO3 (1 eq), toluene, rt, air, 12 h.

However, the reaction conditions seem to be critical for the regioselectivity. A deviation from the standard conditions, considering temperature, amount of base and ligand, leads to varying formation of undesired 1,4-disubstituted triazole, which might be challenging to separate from its regioisomer. Even the optimized protocol leads to the formation of 6%

1,4-disubstituted triazole, and to 94% of the 1,5-disubstituted in toluene (for the specific reaction example between benzylazide and phenylacetylide), whereas a change of solvent using water as a reaction solvent leads to a 91% to 6% ratio in isolated yield. Although the authors claimed a broad substrate scope similar to the CuAAC, biocompatibility could not be proven, since the NiAAC failed with unprotected sugars as substrates, further experiments with amino acids were done with fully protected starting materials.

An interesting, alternative approach for the synthesis of 1,5-disubstituted triazoles, without the use of alkynes, was published in 2011 by Dey et al.⁴⁰ They described a metal-free and regioselective formation of 1,5-disubstituted triazoles in water under reflux conditions, utilizing vinyl sulfones and azides (Scheme 10).

Scheme 10. Metal-free formation of 1,5-disubstituted triazoles between vinylsulfones and azides in water, both aromatic or aliphatic substrates are tolerated; a) Ph-N3, water or toluene, reflux, 12 h.

This approach works for either aromatic or aliphatic substrates. Furthermore, the substrate scope included several ethers, free alcohols, mesylated amines and ketals. Although, vinyl sulfones are readily available from alkenes, 1,2-diols, epoxides and aldehydes,⁴¹ to the best of our knowledge, a synthesis of 3-tosyl-prop-2-en-1-amines, starting from chiral amino aldehydes under preservation of the chiral integrity, has not been described in the

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literature yet (which would be suitable starting materials for peptide-based building blocks).

If “click chemistry” is supposed to be used in living organisms and in vivo applications, the toxic nature of transition metals may pose a problem, this could be circumvented employing electron withdrawing substituents and by ring-strain activated alkynes, which readily react selectively with azides to form the desired disubstitued triazoles within minutes, without transition metal catalysts. The group of Bertozzi published a bioorthogonal in vivo ligation employing difluorinated cyclooctyne as an activated alkyne for the copper-free click chemistry,⁴² therefore combining the rate-enhancing features of ring-strain and electron-withdrawing substituents (Scheme 11).

Scheme 11. Difluorinated strained alkynes react readily with azides in a biological environment to form trisubstituted triazoles as “click”-product.

However, the formation of both regioisomers might impose a problem for the synthesis of smaller ligation products but is generally not considered an issue in the labelling or immobilization of larger biomolecules.

With these tools in hand, chemists started to come up with applications for both, 1,4- and 1,5-disubstitued triazoles.⁴³ A successful replacment of a backbone amide bond by a 1,4-disubstituted triazole was demonstrated by Nahrwold et al. in Cryptophycin-52, while preserving its bioactivity (Figure 14).⁴⁴

Figure 14. Schematic depiction of the cryptophycin-52 and its peptidomimetic, which contains a 1,4-disubstituted triazole between the 3-chloro-4-methoxy-phenylalanine and β-Aib moieties.

Cryptophycin-52 is a cyclic depsipeptide, which shows high in vitro cytotoxicity against multidrug resistant human cancer cell lines (KB-V1), after the incorporation of the

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disubstituted triazole, reproducting the trans configured amide bond (in this position), the bioactivity was only slightly reduced (IC50= 3.2 nM compared to 0.7 nM).⁴⁴

The 1,5-disubstituted triazole was demonstrated to be a cis amide bond surrogate by Tam et al. in 2007.⁴⁵ In this work, the dipeptide surrogate -Xaa[5Tz]Ala- was shown to be a general substitute for -Xaa-cis-Pro- (Figure 15), built into the turn region of RNase A by an expressed protein ligation (Asn113-Pro114 were replaced). The semisynthetic folded enzymes where compared to their wild-types, resulting in thoroughly retained catalytic activity and similar CD-spectra.⁴⁵

Figure 15. Comparison of the turn region -Gly-Asn-cis-Pro-Tyr- and its mimic -Gly-Asn[5Tz]Ala-Tyr-.⁴⁵