In situ Click chemistry

Mgr. Juraj DobiašKOCH, PRIF UK

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:

What is Click Chemistry ?

O

N N N

NN

N

O

Synthesis of 1,2,3-triazoles

R1R2 N

NN

14

R1N

NN

R2

4 15

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

2002, Fokin, Sharpless Melda

1950-70 Huisgen

2005, FokinShrapless

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

HO

DIBO

MMps inhibitors

12

8

MMP7 selective, low mcM inhibitors

Org. Lett. 8 2006 3821-24.

Cu(I)

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 preclickcomponents 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 clickO

N3

ON

NH

NH

NH O

NH

71 randomly selecteddiverse alkynes

ONN

N

ON

N

580 nM

22 nM

ON

NH

NH

NH O

NH

Triazole komplex

NH

NH

NH O

NH

ON

NH

ONNN

ON

Asp142Glu144

Asp215Arg294

Trp403

Arg338

N

Trp220

Trp973WD1

Captured transition state

NH

NH

NH O

NH

ON

NH

ONN

ON

Asp142Glu144

Asp215Arg294

Trp403

N

Trp220

Trp97

N

• Alkene doesn’t cocrystalyze without azide even at high concentration – induced fit.• Different pose of azide with and without alkene – double induced fit.

NH

NH

NH O

NH

Asp142Glu144

Asp215Trp403

Arg338

NHO

NNN

O

N

Arg294

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.