1. Andreas Brunschweiger
TU Dortmund
Accessing genetically tagged
heterocycle libraries via a
chemoresistant DNA sequence

2. Some DNA sequences are more equal than others

3. Identification of bioactive molecules
for chemical biology and medicinal chemistry?
CHEMISTRY BIOLOGY
HYPOTHESIS
?
Peptides
Macrocycles
Oligonucleotides
Small Molecules
PROBE
Natural products
CombiChem
Signaling cascade
Protein-Protein Interaction
Splicing
Cell fate
Differentiation
Autophagy
The gap between chemistry and biology

4. HYPOTHESIS
Peptides
Macrocy
Oligo
Small Molecules
PROBE
Natural pro
CombiChem
plicing
Cell fate
Differentiation
cles S
ducts Aut
nucleotides
Protein-
ophagy
High-throughput screening
CHEMISTRY BIOLOGY
Signaling cascade
Protein Interaction

5. HYPOTHESIS
Peptides
Macrocycles
Oligonucleotides
Small Molecules
PROBE
Natural products
CombiChem
Splicing
Cell fate
Differentiation
Autophagy
Bridging the gap with encoded libraries
CHEMISTRY BIOLOGY
DNA-tagging of drug-like compounds
Signaling cascade
Protein-Protein Interaction

6. Detection of DNA-small molecule conjugates
PCR:
40 μL volume
25 cycles
Taq-polymerase
genetic information

7. DNA-encoded library screen: Selection
ATG GAC
bioactive
compounds
 efficient
 generic
 no dedicated infrastructure
validationpool
Salamon, ACS Chem. Biol.2016

8. A validated technology
NUEVOLUTION
Philochem
Franzini, Angew Chem Int Ed. 2015; Podolin, Prostaglandins 2013; Fernandez-Montalvan, ACS Chem Biol. 2017
Kollmann, Bioorg Med Chem. 2014; Harris, J. Med. Chem. 2017; Lewis, Nat Chem Biol.2015

9. DNA-recorded libraries
 split-and-pool synthesis
 alternated synthesis and encoding steps
 exponential growth of libraries

10. The chemical space of DELs: appendage diversity
5 x 108 compounds
(Harris, J Med Chem.2016)
1 x 105 compounds
(Franzini, Angew Chem Int Ed. 2015)
3 x 106 compounds
(Young, ACS Med Chem Lett. 2015)
4 x 108 compounds
(Kollmann, Bioorg Med Chem.2014)
DNA-compatible, robust, high-yielding reactions with broad scope
 large numbers, high appendage diversity
 few heterocycle-forming reactions
 few reactions confering three-dimensionality:
low structural diversity
Salamon, ACS Chem. Biol. 2016
Klika Skopic, Med. Chem. Commun.2016

11. encoded library
synthesis
Klika-Skopic, MedChemComm, 2016
DELs based on privileged scaffolds
workflow established

12. DEL screen: proof of concept with streptavidin
POI: Streptavidin
sequence Building block1
TCGAACGT 10a

13. DEL screen: results
rank scaffold BB1 BB2
1 amino acid B10a C11
2 amino acid B10a F11
3 amino acid B10a E11
4 amino acid B10a F06
5 amino acid B10a E10
6 amino acid B10a F09
7 amino acid B10a G07
8 amino acid B10a G04
9 amino acid B10a F04
10 amino acid B10a F12
rank scaffold BB1 BB2
1 pyrazolopyr B10a C11
2 pyrazolopyr B10a F04
3 pyrazolopyr B10a E11
4 pyrazolopyr B10a F04
5 pyrazolopyr B10a G04
6 pyrazolopyr B10a F06
7 pyrazolopyr B10a B04a
8 pyrazolopyr B10a A05a
9 pyrazolopyr B06 A02a
10 pyrazolopyr B10a C09
rank scaffold BB1 BB2
1 benzodiazepine B10a F11
2 benzodiazepine B10a F02
3 benzodiazepine B10a E11
4 benzodiazepine B10a F08
5 benzodiazepine B10a F06
6 benzodiazepine B10a E10
7 benzodiazepine B10a F03
8 benzodiazepine B10a F09
9 benzodiazepine B10a F05
10 benzodiazepine B10a E08
selected DEL selected DEL selected DEL
Massive enrichment of building block B10a connected to each scaffold!
POI: Streptavidin

14. DEL screen: results
Desthiobiotin, a streptavidin binder!
Library, selection, and sequencing validated!
POI: Streptavidin

15. DEL screen: results
POI: Streptavidin
The 2nd ranking streptavidin binder

16. DEL screen: results
POI: Streptavidin
The 2nd ranking streptavidin binder
Identified in a second DEL

17. • adapting the scaffold to the needs of DNA
• only appendage diversity
• methodology restricted: two reactions
Coverage of chemical space? Perspective??
encoded library
synthesis

18. appendage diversity
condensation-
macrocycle-
Diels-Alderreaction-
Hantzschreaction-
!!!
scaffold diversity!
carbonyl
reactions
urea-
thiourea-
tert. amine-
amide-
sulfonamide-
sec. amine -
triazole-
biaryl-
alkyne-
hetarylamine -
Expand the chemical space of DNA-encoded libraries!
Our vision for DNA-encoded chemistry

19. Our vision for DNA-encoded chemistry

20. HPLC trace of a DNA prior
Au(I)-catalyzed reaction
… the day after:
armageddon
Running against a massive wall…

21. HPLC trace of a DNA prior
treatment with 1% TFA
… and the day after
a massacre
Running against a massive wall…

22. HPLC trace of a DNA prior
treatment with 1% TFA
… and the day after
a massacre
Running against a massive wall…

23. =
DNA-encoded libraries do not benefit from progress
in many fields of organic chemistry.

24. Adapt the encoding strategy
to the chemistry!
„HexT“

25. TIDEC
Klika Skopic, Chem. Sci. Chem. Sci., 2017, 8, 3356-3361
Brunschweiger, Krause, Antonchick, et al. PCT/EP2016/081845
HexT
Initiating library synthesis with the hexT

26. hexT: recognition by T4 ligase
fluorescence scan
detection of
DNA
stain 5,5% agarose gel; 75 min; 150V;
gel stain: Midori Green
grey: primer sequence

27. β-carbolines by Pictet-Spengler reaction

28. aq. NH3
Mass (calc): 2375.7
Mass (found): 2374.2
aq. NH3
Access to hexT-β-carboline conjugates?

29. aq. NH3
Mass (calc): 2375.7
Mass (found): 2374.2
aq. NH3
Access to hexT-β-carboline conjugates.
Mass (calc): 2467.8
Mass (found): 2468.6

30. aq. NH3
catalysts concentration time
diphenyl phosphate 1 %, CH2Cl2 overnight
TCA 10 % (1-3 %), CH2Cl2 4 h (overnight)
TFA 10 % (1-3 %), CH2Cl2 4 h (overnight)
formic acid 50 %, CH2Cl2 4 h
Solvent DCM DMF ACN THF DCE DMSO MeOH 1,4-Dioxane Toluene
Result
x x x x x

31.  broad scope: 80 conjugates synthesized with diverse aldehydes
 only 15/80 purified
efficient entry into alkaloid-type DELs

32. Suzuki, Org. Lett.2011
Au(I)-catalyzed access to pyrazolines

33. Access to hexT-pyrazoline conjugates?
aq. NH3

34. aq. NH3
HPLC of the crude hexT-conjugate
Ms (calc.): 2391.8
Ms (found):2392.3
aq. NH3
Access to hexT-pyrazoline conjugates.

35. Towards pyrazol(in)e libraries
71 amino-substituted hexT-pyrazol(in)es synthesized
selection of reaction conditions: access to two core structures
scope: aromatic and branched aliphatic aldehydes, diverse hydrazides,
aliphatic, and aromatic alkynes

36. Library synthesis
Klika Skopic, Chem. Sci., 2017, 8, 3356-3361
15.000 DNA-encoded compounds

37. Library synthesis
Klika Skopic, Chem. Sci., 2017, 8, 3356-3361
15.000 DNA-encoded compounds
Perspective?

38. Bode J. W.et. al. Org. Lett., 2014, 16,1236-1239.
mass calc.= 2260.6
mass found= 2261.6
SnAP reagent
MALDI-MS of the purified
hexT-small molecule conjugate
Expanding chemistry: Cu(II)-mediated cyclization
HPLC trace
of the crude

39. Expanding chemistry: Ugi reaction
mass calc.= 2256.7
mass found= 2257.5
MALDI-MS of the purified
hexT-small molecule conjugate
HPLC trace
of the crude

40. From sequence to sequences, mixed T,C-DNA

41. From sequence to sequences, mixed T,C-DNA
Towards alternative tagging strategies.
M. Klika Skopic, Org. Biomol. Chem., 2017,15, 8648-8654

42. Conclusion
sequence Building block1
TCGAACGT 10a

43. Conclusion
appendage diversity
structural diversity
DNA-tagging
strategies

44. Outlook
micellar catalysis
DEL chemistryscreening
lab-on-a-chip techniques
with Dr. Dirk Janasek (ISAS Dortmund)with Prof. Ralf Weberskirch, TU Dortmund

45. The DNA-Team
Mateja Klika Skopic
Hazem Salamon
Kathrin Jung
Olivia Bugain
Chemistry
Prof. Ralf Weberskirch, TU Dortmund
Dr. Andrea Ernst
Hanne Petersen
Prof. Norbert Krause, TU Dortmund
Dr. Bernd Wagner
Justin Schieven
Financial support from
is most gratefully acknowledged!
Cheminformatics
Dr. Tuomo Kalliokoski, LDC Dortmund
Dr. Oliver Koch, TU Dortmund
Faculty
Prof. Daniel Rauh
Prof. Herbert Waldmann
Analysis of sequencing
data
Dr. Michael Blank
Dr. Carsten Gröber
Screening
Prof. Günter Mayer, Universität Bonn
Franziska Pfeiffer, Universität Bonn

46. Presented at the Global Medicinal
Chemistry and GPCR Summit
To find out more, visit:
www.global-engage.com