University of Groningen Deacetylase inhibitors & Histone inheritance Zwinderman, Martijn R. H.

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University of Groningen

Deacetylase inhibitors & Histone inheritance

Zwinderman, Martijn R. H.

DOI:

10.33612/diss.167867692

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Publication date: 2021

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Zwinderman, M. R. H. (2021). Deacetylase inhibitors & Histone inheritance. University of Groningen. https://doi.org/10.33612/diss.167867692

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Appendices

Nederlandse samenvatting

Publications

Posters

Acknowledgements

Curriculum Vitae

197

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Nederlandse samenvatting

Deacetylase remmers

Er is een dringende noodzaak voor het ontwikkelen van nieuwe therapieën voor longaandoeningen met een ontstekingscomponent. Zo sterven er wereldwijd naar schatting 3 miljoen mensen per jaar aan de gevolgen van de chronisch obstructieve longziekte COPD. Daarnaast kost longkanker het leven van ongeveer 1,5 miljoen mensen per jaar. De behandeling van deze longziektes, en ook astma, kan wellicht verbeterd worden door een therapie te ontwikkelen die verstoorde patronen van posttranslationele modificaties corrigeert. Acetylatie en methylatie zijn belangrijke voorbeelden van posttranslationele modificaties die op duizenden eiwitten gevonden worden en daarmee de eigenschappen van de gemodificeerde eiwitten beïnvloeden. De enzymen die een acetyl of methyl group aan een eiwit toevoegen of weghalen vervullen daarmee een cruciale rol in de regulatie van verschillende intracellulaire werkingsmechanismen. Toepassing van remmers van deze enzymen in preklinische modellen van astma, COPD en longkanker laten positieve resultaten zien. Er is de afgelopen jaren veel vooruitgang geboekt op dit gebied, in het bijzonder met betrekking tot de ontwikkeling en toepassing van deacetylase remmers. Toch zijn er nog verschillende stappen nodig om deacetylase remmers te kunnen toepassen als nieuwe therapie voor de genoemde longaandoeningen.

Een belangrijke stap is het ontwikkelen van remmers voor specifieke deacetylases om daarmee de rol van deze deacetylases in ontstekingsmechanismen te kunnen ontrafelen. Daarom hebben we nieuwe remmers gemaakt op basis van o-aminobenzamide gesubstitueerde heterocyclische vijfringen. De selectiviteit waarmee de moleculen histon deacetylase 1, 2 en 3 remmen hebben we gekoppeld aan de mate waarin de remmers de inflammatoire genexpressie in macrofagen beïnvloeden. Van de moleculen bleek een o-aminobenzamide gesubstitueerde thiofeen de beste remmer van histon deacetylase 1, 2 en 3. Om de selectiviteit van de

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aminobenzamide gesubstitueerde thiofeen verder te onderzoeken hebben we een nieuwe syntheseroute ontwikkelt voor 2-nitro-3-aminothiofenen die op de 5-positie met diverse groepen kunnen worden gesubstitueerd. Deze moleculen zijn een precursor voor de uiteindelijke remmers. Door alkynyl cyanides als grondstof te gebruiken hebben we een relatief eenvoudige en efficiënte methode ontwikkeld om de gewenste precursors te maken. Diverse groepen op de 5-positie zorgden ervoor dat de moleculen met een grotere selectiviteit histon deacetylase 1 en 2 remden ten opzichte van histon deacetylase 3. Vervolgens vonden we dat zowel de moleculen die histone deacetylase 1, 2 en 3 als de moleculen die voornamelijk histon deacetylase 1 en 2 remden de transcriptionele activiteit van NF-κB verhoogden en ook de genexpressie van het pro-inflammatoire cytokine IL-6. Daarentegen zorgden alleen remmers van histon deacetylase 1, 2 en 3 en niet een selectieve remmer van histon deacetylase 1 en 2 voor een verhoogde expressie van het anti-inflammatoire cytokine IL-10. Tezamen bieden deze bevindingen inzicht in de toepassing van niet-selectieve remmers van histone deacetylase 1, 2 en 3 in ziektes met een inflammatoire component zoals asthma, COPD en longkanker.

Histon overerving

Tijdens DNA replicatie worden gelijke hoeveelheden histonen verdeeld over het gedupliceerd DNA door hergebruik van oude histonen en toevoeging van nieuwe histonen. Moleculaire mechanismen die oude en nieuwe histonen gebalanceerd verdelen over het gedupliceerde DNA zijn grotendeels onbekend. Daarom hebben wij de verdeling van nieuwe histonen over gedupliceerd DNA onderzocht middels een door onszelf ontwikkelde methode genaamd double-click-seq. In de controle situatie vonden we dat nieuwe histonen meer voorkwamen op DNA gedupliceerd door de lagging strand polymerase ten opzichte van de leading strand en dit werd versterkt door behandeling met een PARP remmer, met name in regio’s met veel adenosine en thymine. Het effect van de PARP remmer kan verklaard worden door de noodzakelijkheid van PARP in lagging strand synthese. Opvallend genoeg zorgde replicatie stress als gevolg van behandeling met hydroxyurea,

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curaxin of een ATR remmer, of uitschakeling van p53, ervoor dat nieuwe histonen meer voorkwamen op de DNA streng gedupliceerd door de leading strand polymerase. Om deze observaties te verklaren hebben wij een model opgesteld waarin de asymmetrische verdeling van nieuwe histonen volgt uit ontkoppeling van de helicase van de leading danwel de lagging strand polymerase. Tenslotte speculeren we over mogelijke gevolgen van de asymmetrische verdeling van nieuwe histonen tijdens DNA replicatie en in het bijzonder tijdens replicatie stress.

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Publications

2021

Martijn R. H. Zwinderman*, Thamar Jessurun Lobo*,

Petra E. van der Wouden, Diana C. J. Spierings, Marcel A. T. M. van Vugt, Peter M. Lansdorp, Victor Guryev, Frank J. Dekker. *Equal contribution.

Inversion of asymmetric histone deposition upon replication stress. Submitted.

Martijn R. H. Zwinderman, Fangyuan Cao, Petra E. van der Wouden, Ronald van Merkerk, Wim J. Quax, Frank J. Dekker.

Effect of Class I selective HDAC inhibitors with an

o-aminobenzamide substituted thiophene scaffold on inflammatory gene expression in macrophages.

European Journal of Medicinal Chemistry, under revision. 2020

Bin Liu, Siwei Chen, Anouk La Rose, Deng Chen, Fangyuan Cao, Martijn R. H. Zwinderman, Dominik Kiemel, Manon Aïssi,

Frank J. Dekker, Hidde J. Haisma.

Inhibition of histone deacetylase 1 (HDAC1) and HDAC2 enhances CRISPR/Cas9 genome editing.

Nucleic Acids Research. 2019

Martijn R. H. Zwinderman, Fangyuan Cao, Frank J. Dekker. Acetylation and Methylation in Asthma, COPD, and Lung Cancer.

Chemical Epigenetics.

Martijn R. H. Zwinderman, Sander de Weerd, Frank J. Dekker. Targeting HDAC Complexes in Asthma and COPD.

Epigenomes.

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Fangyuan Cao, Martijn R. H. Zwinderman, Ronald van Merkerk, Petra E. van der Wouden, Wim J. Quax, Frank J. Dekker. Inhibitory selectivity among class I HDACs has a major impact on

inflammatory gene expression in macrophages. European Journal of Medicinal Chemistry.

Magdalena Wójcik, Nikolaos Eleftheriadis, Martijn R. H. Zwinderman, Alexander S. S. Dömling, Frank J. Dekker, Ykelien L. Boersma. Identification of potential antivirulence agents by substitution-oriented

screening for inhibitors of Streptococcus pyogenes sortase A. European Journal of Medicinal Chemistry.

2018

Fangyuan Cao, Martijn R. H. Zwinderman, Frank J. Dekker. The Process and Strategy for Developing Selective Histone Deacetylase 3

Inhibitors. Molecules.

2017

Viktoria Krieger, Alexandra Hamacher, Christoph G. W. Gertzen, Johanna Senger, Martijn R. H. Zwinderman, Martin Marek, Christophe Romier, Frank J. Dekker, Thomas Kurz, Manfred Jung,

Holger Gohlke, Matthias U. Kassack, Finn K. Hansen. Design, multicomponent synthesis and anticancer activity of a

focused histone deacetylase (HDAC) inhibitor library with peptoid-based cap groups.

Journal of Medicinal Chemistry. 2016

Nikolaos Eleftheriadis, Stephanie A. Thee, Martijn R. H. Zwinderman, Niek G. J. Leus, Frank J. Dekker.

Activity-Based Probes for 15-Lipoxygenase-1. Angewandte Chemie - International Edition.

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Niek G. J. Leus, Martijn R. H. Zwinderman, Frank J. Dekker. Histone deacetylase 3 (HDAC 3) as emerging drug target in

NF-kappa B-mediated inflammation. Current Opinion in Chemical Biology.

Maria E. Ourailidou, Martijn R. H. Zwinderman, Frank J. Dekker. Bioorthogonal metabolic labelling with acyl-CoA reporters:

targeting protein acylation. MedChemCommun.

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Poster DECHEMA

Frankfurt, Germany, 2019 (1st_Prize)

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Poster CHAINS

Veldhoven, The Netherlands, 2017

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Acknowledgements

The research presented in this thesis was performed at the Groningen Research Institute of Pharmacy of the University of Groningen. This research has been financially supported with a VIDI grant from the Dutch Research Council (NWO) and a starting grant from the European Research Council (ERC), both awarded to Frank Dekker.

First and foremost, I would like to thank Frank Dekker. We happen to have approximately the same age difference as Watson and Crick, both featured prominently on the cover of this thesis. I could have featured each of our portraits, but that may be frowned upon. However, it would have shown how much I am indebted to you as my mentor in science, history, politics and several other aspects of life. Our lively discussions about the model of inversion of histone deposition asymmetry during replication stress is how I imagine Watson and Crick discussed their model of the double helix. Furthermore, you always gave me the freedom to make my own mistakes and helped me stay focused. You not only provided great guidance on establishing research topics, but also contributed significantly to the dissemination of research output through academic publications and presentations at key conferences. The only regret that I have of my PhD is that I have never nominated you for the university’s PhD supervisor of the year award, a title you most definitely deserve. Our history together goes back to my bachelor and master research projects and I hope it will continue to expand.

I would also like to express my sincere thanks to Petra Ettema, who has helped me a great deal with the experiments. I truly enjoyed our conversations on just about anything and have relied heavily on your organizational and experimental skills in the lab. I was always welcome to pitch any idea to you and you were always ready to give it a serious thought. Furthermore, I would like to thank Hidde Haisma as my co-promotor for guidance, positive feedback and confidence in me during the evaluation meetings. Partly because

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of the group trip with your boat I have taken up sailing myself! Also a big thanks to Wim Quax, Ykelien Boersma and Gerrit Poelarends for the fruitful discussions about my work during the group meetings. Similarly, Diana Spierings, Peter Lansdorp and Marcel van Vugt have provided helpful suggestions that have guided my work at key moments. Marianne Rots has showed me how to organize a successful international symposium and Martina Schmidt helped me to get the PhD position by providing an incredibly kind motivation letter. Thamar Jessurun Lobo and Victor Guryev were instrumental in the discussions regarding the asymmetry in histone deposition. I would also like to thank them for always being available for a nice conversation and providing the crucial analysis of the datasets.

The members of our group when it was still called pharmaceutical gene modulation, Nikolaos Eleftheriadis, Maria-Eleni Ourailidou, Thea van den Bosch, Niek Leus and Hannah Wapenaar were important for my first years in the lab. They have helped me get started, especially Nikolaos Eleftheriadis and Maria-Eleni Ourailidou, whom I got to know on a personal level. I am happy to know that you’re both doing well in your careers. I’ve also enjoyed the discussions on chemistry with people from the drug design group, Pravin Patil, Tryfonas Zizigas-Zarganis, Dinos Neochoritis, Edwin Kroon, André Boltjes and Robin van der Straat. They have also let me use several important chemicals of their lab to test my ideas. Similarly, Eduard Post and Catharina Reker-Smit have kindly let me use materials from their inventory.

To Robbert Cool, Ronald van Merkerk, Pieter Tepper and Rita Setroikromo I owe properly working labs and a tremendous amount of experience that I was allowed to make use of. Special mention goes to Ronald for our discussions on molecular modeling and Pieter for showing me how to set up a completely new lab. I am sure that a future return to the lab will surprise me with how many new regulations and signs you have put up. Jennefer Beenen, Karina Hoekstra-Wakker and Nancy Halsema have given me important advice on experimental procedures required for obtaining a suitable DNA library for

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sequencing. Similarly, Marcel de Vries has given me good tips on how to prepare protein samples for mass spectrometry.

The tremendously informative meetings with Bernard van Vliet and Jack den Hartog have given me a much broader scope of my PhD, enlightening the path towards clinical development of drug candidates. I hope that they continue to mentor me on this path towards success in getting the developed HDAC inhibitors to the clinic to treat COPD. In this respect I am also indebted to Reinoud Gosens and Loes Kistemaker from Aquilo. I hope we will receive funding to test the HDAC inhibitors that have been left out of the scope of this thesis. I would also like to thank Reinoud for being my mentor in managing my PhD and for showing me the interesting research going on in his lab.

I’ve had great fun guiding several bachelor and master students in their research projects. Francesca Serratore and Faisal Benmhammed, thank you for helping me on several projects and making sure I knew every detail of them. Furthermore, big thanks to Sander de Weerd, who was willing to write a review with me. You’ve read countless of papers and distilled only the most relevant information and did a great job. I wish you good luck with your future creative and entrepreneurial projects.

To my research family, Hao Guo, Fangyuan Cao, Zhangping Xiao, Bin Liu, Olivia Diaz, Siwei Chen and Deng Chen, thank you for making my PhD a wonderful experience. I have received so much kindness and support from all of you it is difficult to put in words. Special thanks to Bin for keeping me up-to-date with the latest, highly relevant papers and interesting discussions about Cas9. Also, thank you for letting me try a thousand-year-old-egg and asking me to become your paranymph for your defense. You’ve done well and we had a great party. Hao for letting me play basketball with his fellow Chinese friends and Hao and Yu for welcoming me for dumpling parties. Thanks Fangyuan for being my partner in crime in the lab. I think we made a wonderful team. Together we solved some complex med chem problems.

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Zhangping, Siwei and Deng you’ve been really helpful through your kindness and interest in my work. I hope I’ve returned the favor and wish you luck finishing your PhD, especially now during the corona crisis. Finally, thanks Olivia for the orchid you gave me. I never managed to let it become so beautiful as yours, but it was a constant reminder of your kindness.

The people that have helped me to set-up the Honours Review foundation, Michael Lerch, Alexander Pietrus-Rajman, Sébastien Volker, Hanny Elzinga and Kees de Vey Mestdagh. I’ve gained several useful skills from this endeavor, for instance how to do graphic design, which let me win a best poster prize in a conference in Frankfurt (see poster on page 204). I have also learned how to set-up an organization and ensure continuity by networking.

Support from Winant and Nadie and my parents was always there and cannot be put in words. Thanks to Groningse nuchterheid I am sure that you know how much I rely on you. Winant, I am glad you are willing to start the spin-off Griphingo Pharmaceuticals with me and hope that together we will make an impact on people’s quality of life in the future.

I’d like to thank my roommate Ferdi Koetje for allowing me to use his room as a writing room. Furthermore, my other roommate Koen ten Hertog and all other non-lab friends from Pekela and Winschoten have ensured my business as usual approach to a PhD. In the end a PhD project is no different than any other project-based job and I’m very much looking forward to working on new projects at Avebe under the guidance of Johan Hopman.

Finally, the single most important event during my PhD was my chance encounter with Marjolein van der Putten. All of the chemistry that I’ve done in the lab does not even closely match the chemistry between the two of us. We realized our unique connection and quickly brought our love start-up outside of academia. At the moment we have left the start-up phase and entered the scale-up phase with a beautiful apartment in Zwolle. I have already had countless of wonderful experiences with you and hope that continues for years to come!

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Curriculum vitae

Martijn (Reint Harm) Zwinderman was born on November 17, 1990 in Winschoten, The Netherlands. He graduated from Dollard College, Winschoten in 2009, and received a Bachelor’s degree in Pharmacy at the University of Groningen in 2012. During the Bachelor program he also completed the Honours College track for excellent students. At the same university, he graduated from the Master in Pharmacy (cum laude) in 2015. During the Master program he also co-founded and chaired Honours Review, a biannual university-wide student journal with artwork from the Minerva Art Academy in Groningen.

Since then, Martijn has been a PhD candidate at the University of Groningen under the guidance of Professor Frank J. Dekker. During his PhD trajectory he focused on the synthesis and effects of novel regulators of epigenetic processes in inflammatory diseases. Additionally, he spearheaded a multidisciplinary project team to unravel the fundamental dynamics of histone deposition during DNA replication by developing a new research method, the results of which are described in this thesis. He also had the honour to present his research outcomes at the FIGON Dutch Medicine Days student competition to represent the University of Groningen.

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