In situ Click chemistry Mgr. Juraj Dobiaš KOCH, PRIF UK

What is Click Chemistry ? joining molecules by an „ideal chemical reaction“ fast, irreversible reaction, simple conditions starting materials are readily available, stable (biocompatible) high yielding, high atom economy, wide application large thermodynamic driving force to give predictable outcome easy work-up and product isolation preferably proceeding in water, insensitive to oxygen the best by some fusion reaction:

Synthesis of 1,2,3-triazoles Thermal Huisgen [3+2] cycloaddition 80-120°C, 12-24h, both regioisomers ca 1/1 E#A= 24-26 kcal/mol Cu(I) catalyzed (CuSO4 / sodium L-ascorbate) only 1,4-regioisomer, high yield, rt, t-BuOH / water environment E#A= 15 kcal/mol (106 times faster than Huisgen r.) Ru catalyzed (Cp*RuCl(PPh3)2) mainly 1,5-regioisomer 1950-70 Huisgen 2002, Fokin, Sharpless Melda 2005, Fokin Shrapless

Click Chemistry Exploitation Material sciences (copolymers, functionalized surfaces, adhesives, dendrimers, large macrocycles, ....) Bioorganic chemistry (biosensors, bioconjugates: tagging of proteins, nucleotides or in situ whole organisms, SPAAC – no metal) Drug development – Medicinal Chemistry

MMps inhibitors 8 Cu(I) 12 MMP7 selective, low mcM inhibitors Org. Lett. 8 2006 3821-24.

In Situ Click Chemistry (TDS) target driven synthesis TDS reduces the number of inactive compounds Compensate the lack of precision in the predictive ability of in Silico chemistry In situ AA Click chemistry is completely biocompatible, uses irreversible reaction to unite reagents inside the protein´s binding pocket Target will pick best fitting ligands from diverse sets of chemical building blocks Significant portion of the activation barrier is entropic (pieces have to approach each other in precisely the right orientation), pre-assembly of building blocks on the target active site can accelerate cycloaddition. DDT 9 2004 348.

Observation of the controlled assembly of preclick components in the in situ click chemistry generation of a chitinase inhibitor chitinase inhibitors fight against infectious and inflammatory diseases – onchocerciasis. screening of over 10,000 extracts from soil microorganisms 2013 PNAS 110 15892-97

In situ click

Triazole komplex 3WD1

Captured transition state Alkene doesn’t cocrystalyze without azide even at high concentration – induced fit. Different pose of azide with and without alkene – double induced fit. 3WD2 3WD4

Summary Identified triazole inhibitor of chinase B by in situ click chemistry approach. Solved crystal structure of transition state with alkene analog. Performed DFT calculations, but did not observe any enthaplic barrier decrease. Confirmed in situ click chemistry principle that increase in reaction rate is caused by entropic factors and greater effective concentration. Solved crystal structures that demonstrate protein flexibility and lack of in silico methods accuracy.