Solution-Phase Fmoc-Based Peptide Synthesis for DNA-Encoded Chemical Libraries: Reaction Conditions, Protecting Group Strategies, and Pitfalls

Abstract

Peptide drug discovery has shown a resurgence since 2000, bringing 28 non-insulin therapeutics to the market compared to 56 since its first peptide drug, insulin, in 1923. While the main method of discovery has been biological displayphage, mRNA, and ribosomethe synthetic limitations of biological systems has restricted the depth of exploration of peptide chemical space. In contrast, DNA-encoded chemistry offers the synergy of large numbers and ribosome-independent synthetic flexibility for the fast and deeper exploration of the same space. Hence, as a bridge to building DNA-encoded chemical libraries (DECLs) of peptides, we have developed substrate-tolerant amide coupling reaction conditions for amino acid monomers, performed a coupling screen to illustrate such tolerance, developed protecting group strategies for relevant amino acids and reported the limitations thereof, developed a strategy for the coupling of α,α-disubstituted alkenyl amino acids relevant to all-hydrocarbon stapled peptide drug discovery, developed reaction conditions for the coupling of tripeptides likely to be used in DECL builds, and synthesized a fully deprotected DNA-decamer conjugate to illustrate the potency of the developed methodology for on-DNA peptide synthesis.

Metal‐Free and Open‐Air Arylation Reactions of Diaryliodonium Salts for DNA‐Encoded Library Synthesis

“…As a consequence, most of the conventional organic reactions are excluded from the toolbox of DEL chemistry. Researchers from academia and industry have since developed various DNA‐compatible reactions such as on‐DNA amide formation, [ 21 , 22 ] diazo‐transfer, [ 23 ] cross‐coupling reaction, [ 24 , 25 , 26 , 27 , 28 , 29 ] C‐H activation and functionalization, [ 30 , 31 , 32 , 33 , 34 , 35 ] ring‐closing metathesis, [ 36 , 37 ] Sulfur‐Fluoride Exchange (SuFEx) chemistry, [ 38 ] photo‐catalyzed reaction, [ 39 , 40 , 41 , 42 , 43 , 44 , 45 ] and cycloaddition reactions. [ 46 , 47 , 48 , 49 , 50 ] Recently, the strategies of micellar‐mediated on‐DNA synthesis, [ 51 , 52 ] using chemically stabilized DNA barcodes, [ 53 ] solid support mediated on‐DNA synthesis, [ 54 ] and especially the reversible adsorption of DNA on solid supports have greatly expanded the on‐DNA synthetic toolbox.…”

Quantitative Validation and Application of the Photo-Cross-Linking Selection for Double-Stranded DNA-Encoded Libraries

“…The reaction recovery rate was only 12%. We optimized the linker protecting group and deprotection conditions and found that the recoveries of the deprotection reaction of the Alloc protecting group, the Nvoc protecting group, and the Cbz protecting group could reach 99%, 88%, and 72%, respectively. − In future designs, we can use suitable linkers and protecting groups to complete the construction of photo-cross-linking libraries under milder reaction conditions (see Supporting Information for details).…”

Synthesis and Applications of Peptides and Peptidomimetics in Drug Discovery

“…In 2020, Young and coworkers reported a significant advancement in this field by constructing stapled peptides in DNA-encoded chemical libraries. [46,47] ALRN-5281 and ALRN-6924 are the two best clinical candidates synthesized by stapled peptide methodology.…”