Robust Suzuki–Miyaura Cross-Coupling on DNA-Linked Substrates

Ding, Yun; Clark, Matthew

doi:10.1021/co5001037

Cited by 115 publications

(111 citation statements)

References 30 publications

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“…9,20,33–35 Following a test reaction, sensor beads are analyzed as before, and the solution is analyzed by HPLC and MALDI-TOF MS to determine the conversion of 5′-amino oligonucleotide starting material to DNA-conjugated product (Table 2). Condensation of 5′-amino oligonucleotide substrate with Fmoc-Lys(Alloc)-OH in the presence of the activator DMT-MM, a staple amide bond forming reaction in DNA-encoded library synthesis, 20,41 proceeds with 65% yield and 69% DNA remaining.…”

Section: Resultsmentioning

confidence: 99%

“…Subsequent Alloc deprotection 34 yields a mix of Alloc-deprotected (70%) and fully deprotected product (30%), and 50% DNA remaining. Published Suzuki-Miyaura cross-coupling conditions 33 leave 30% of amplifiable DNA remaining. Conventional solution-phase library preparation includes HPLC purification, likely eliminating synthesis products attached to Pd-catalyzed DNA cleavage products (see Supporting Information).…”

Section: Resultsmentioning

confidence: 99%

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What is a “DNA-Compatible” Reaction?

2016

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DNA-encoded synthesis can generate vastly diverse screening libraries of arbitrarily complex molecules as long as chemical reaction conditions do not compromise DNA’s informational integrity, a fundamental constraint that “DNA-compatible” reaction development does not presently address. We devised DNA-encoded reaction rehearsal, an integrated analysis of reaction yield and impact on DNA, to acquire these key missing data. Magnetic DNA-functionalized sensor beads quantitatively report the % DNA template molecules remaining viable for PCR amplification after exposure to test reaction conditions. Analysis of solid-phase bond forming (e.g., Suzuki-Miyaura cross-coupling, reductive amination) and deprotection reactions (e.g., allyl esters, silyl ethers) guided the definition and optimization of DNA-compatible reaction conditions (> 90% yield, > 30% viable DNA molecules), most notably in cases that involved known (H+, Pd) and more obscure (Δ, DMF) hazards to DNA integrity. The data provide an empirical yet mechanistically consistent and predictive framework for designing successful DNA-encoded reaction sequences for combinatorial library synthesis.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

What is a “DNA-Compatible” Reaction?

2016

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“…In general terms, we have found that the usual amount of DNA that we append to the beads 118 will tolerate 3–4 acylations of secondary amines, or 5–6 acylations of primary amines under our standard conditions before the amplifiable level of DNA drops too low to be of use as an encoding tag. Some organometallic reactions, such as Suzuki couplings 119 , also inactivate about 50% of the DNA templates on the bead. Therefore, in the future it will be important to develop new library designs of different types of molecules that can be made with gentler reactions.…”

Section: Dna-encoded Oboc Librariesmentioning

confidence: 99%

Towards vast libraries of scaffold-diverse, conformationally constrained oligomers

Kodadek

McEnaney

2016

Chem. Commun.

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There is great interest in the development of probe molecules and drug leads that would bind tightly and selectively to protein surfaces that are difficult to target with traditional molecules, such as those involved in protein-protein interactions. The currently available evidence suggests that this will require molecules that are larger and have quite different chemical properties than typical Lipinski-compliant molecules that target enzyme active sites. We describe here efforts to develop vast libraries of conformationally constrained oligomers as a potentially rich source of these molecules.

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“…6−8 DELs, however, bridge a significant chemotype divide by moving library content away from the space of naturally occurring biopolymers found in display libraries and into the more drug-like space of high-throughput screening (HTS) compound collections. 4,9,10 The plunging costs of both DNA synthesis and next-generation sequencing (NGS) have positioned DEL technology as a promising companion to industrial HTS platforms 11 because it offers access to larger compound collections and lead identification is more economical and expeditious.…”

Section: Introductionmentioning

confidence: 99%

An Integrated Microfluidic Processor for DNA-Encoded Combinatorial Library Functional Screening

2017

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DNA-encoded synthesis is rekindling interest in combinatorial compound libraries for drug discovery and in technology for automated and quantitative library screening. Here, we disclose a microfluidic circuit that enables functional screens of DNA-encoded compound beads. The device carries out library bead distribution into picoliter-scale assay reagent droplets, photochemical cleavage of compound from the bead, assay incubation, laser-induced fluorescence-based assay detection, and fluorescence-activated droplet sorting to isolate hits. DNA-encoded compound beads (10-μm diameter) displaying a photocleavable positive control inhibitor pepstatin A were mixed (1920 beads, 729 encoding sequences) with negative control beads (58 000 beads, 1728 encoding sequences) and screened for cathepsin D inhibition using a biochemical enzyme activity assay. The circuit sorted 1518 hit droplets for collection following 18 min incubation over a 240 min analysis. Visual inspection of a subset of droplets (1188 droplets) yielded a 24% false discovery rate (1166 pepstatin A beads; 366 negative control beads). Using template barcoding strategies, it was possible to count hit collection beads (1863) using next-generation sequencing data. Bead-specific barcodes enabled replicate counting, and the false discovery rate was reduced to 2.6% by only considering hit-encoding sequences that were observed on >2 beads. This work represents a complete distributable small molecule discovery platform, from microfluidic miniaturized automation to ultrahigh-throughput hit deconvolution by sequencing.

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Robust Suzuki–Miyaura Cross-Coupling on DNA-Linked Substrates

Cited by 115 publications

References 30 publications

What is a “DNA-Compatible” Reaction?

What is a “DNA-Compatible” Reaction?

Towards vast libraries of scaffold-diverse, conformationally constrained oligomers

An Integrated Microfluidic Processor for DNA-Encoded Combinatorial Library Functional Screening

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