β-Silyl alkynoates: Versatile reagents for biocompatible and selective amide bond formation

Abstract

The study introduces a previously unidentified method for amide bond formation that addresses several limitations of conventional approaches. It uses the β-silyl alkynoate molecule, where the alkynyl group activates the ester for efficient amide formation, while the bulky TIPS (triisopropylsilane) group prevents unwanted 1,4-addition reactions. This approach exhibits high chemoselectivity for amines, making the method compatible with a wide range of substrates, including secondary amines, and targets the specific ε-amino group of lysine among the native amino ester's derivatives. It maintains stereochemistry during amide bond formation and TIPS group removal, allowing a versatile platform for postsynthesis modifications such as click reactions and peptide-drug conjugations. These advancements hold substantial promise for pharmaceutical development and peptide engineering, opening avenues for research applications.

Fig. 1.. Background and reaction design.

(A) Amide bonds in bioactive molecules. (B) Classical methods for amide bond formation. (C) This work: Biocompatible amide bond formation using β-silyl alkynoates via 1,2 additions. (D) Chemical reactivity of amino esters. (E) Application of the reaction in protein, drug, and peptide modification.

Fig. 2.. Control experiments.

(A) Effect of β-substitution on chemoselective 1,2- versus 1,4-addition. (B) Comparison of ester reactivity. (C) Nucleophilic competitive experiment. Reaction yield was determined from the crude reaction mixture by ¹H NMR using CH₂Br₂ as an internal standard. N.R, no reaction; PBS, phosphate-buffered saline (pH 7.0).

Fig. 3.. Substrate scope of amine nucleophiles to β-silyl alkynoates.

1a (50 mg, 0.196 mmol), 2 (0.295 mmol, 1.5 equiv.), and solvent (0.5 ml) for 48 hours. Isolated yield. ^aThe remaining starting materials were recovered (for details, see Supplementary Materials). ^bPBS buffer (pH = 7.0)/CH₃CN (4:1) was used.

Fig. 4.. Scope of the amine-containing bioactive molecule and drug modification.

1a (50 mg, 0.196 mmol), 2 (0.295 mmol, 1.5 equiv.), and solvent (0.5 ml) for 48 hours. Isolated yield. ^aPBS buffer (pH = 7.0)/CH₃CN (4:1) solvent was used. ^bPBS buffer (pH = 7.0) was used as a solvent. ^cThe remaining starting materials were recovered (for details, see Supplementary Materials).

Fig. 5.. Stereoselectivity assessment in peptides using ¹³C NMR spectrum for epimer of amine-conjugated product.

Fig. 6.. Application of N-substituted propiolamide.

(A) Gram-scale synthesis. (B) Product derivatization of N-benzylpropiolamide (C) Application in linker chemistry for peptide-drug conjugation (PDCs).

Fig. 7.. Selectivity comparison between activated esters and ethyl alkynoates.

Fig. 8.. Amino ester selectivity and polypeptide modification with 1a.

(A) Specific amino ester selectivity. (B) Polypeptide modification with 1a. (C) Modified lanreotide conjugates.

Fig. 9.. Modification of bovine serum albumin, myoglobin, lysozyme, and cytochrome c protein using β-silyl alkynoates.