AG1

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Prof. Dr. Kerstin Borgmann, Dipl.-Biol.
Sandra Classen, MSc.MLS PhD Stud.

The aim of this project is to elucidate the molecular and biological mechanisms that lead to breast cancer at a young age. Therefore, we investigate the functional influence of mutations in genes of the DNA repair complex homologous recombination, such as PALB2, BRCA1, BRCA2 and ABRAXAS on DNA replication processes, DNA damage control and genomic instability. This question is investigated on primary cells of mutation carriers and blood samples of young compared to older breast cancer patients without family history. The focus lies on the causative factors of genomic instability through defective DNA replication as a reason for early onset of cancer (Fig. 1). The aim is to develop a possible marker for early onset cancers. Based on this, we are currently investigating whether a primary genomic instability could accelerate the occurrence of brain metastases in breast cancer patients. This issue is addressed by the project:

1. Incorrect replication processes as a driver of early onset breast cancer (Kerstin Borgmann und Lena Schuckenbrock

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Figure 1: Incorrect replication processes as markers of early onset cancer

Chromosomal instability negatively affects the survival of all subgroups of the breast cancer. This is attributed to an increased aggressiveness and poorer curability of genomically unstable tumours. An increased chromosomal instability (CIN) can be attributed to a defect in the DNA repair pathway Homologous recombination (HR). HR prevents genomic instability through DNA repair and protection of DNA replication structures. The aim is to clarify whether genetic alterations lead to a repair defect or whether the supply of metabolites and/or superordinate signaling pathways are of crucial importance and how tumor stem cells behave in this context. These issues are addressed by several projects in our group:

• Molecular mechanisms of resistance formation by defective HR using isogenic, genetically engineered cell systems (Elena Rahlf). The effects of reduced BRCA1 expression on HR capacity, tumor stem cell content and intracellular immune response are investigated in a CRISPR/Cas9 modified isogenic cell system after irradiation.
• Influence of the metabolome on S-phase DNA repair processes: identification of new targets for the further development of existing tumor therapies (Lena Schuckenbrock). The working hypothesis addressed in this project is that changes in the metabolome and extranuclear signal cascades influence the regulation of cellular DNA damage response and repair in the S-phase and thus represent an important reason for individual radiation sensitivity.

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Fig. 2: DNA Repair pathways altered in tumor stem cells are associated with DNA replication.

• Radioresistance of tumor stem cells (Felix Meyer). Tumor stem cells are resistant to conventional therapies and are responsible for recurrence and metastasis. This is attributed to upregulated DNA repair mechanisms and an increased DNA damage response (see Fig. 2). The aim of this project is the identification of druggable targets to radiosensitize tumor stem cells. For this, tumor stem cells were isolated from radioresistant breast cancer cell lines. Altered signalling cascades, DNA-replication processes and DNA-repair mechanisms are experimentally determined and important key factors inhibited to achieve radiation sensitization. Through this approach we hope to develop new, more effective therapeutic approaches to increase the efficiency of radiotherapy.

• Targeting the adaptive resistome in triple-negative breast cancer patient-derived organoids (Sandra Classen). The aim of this project is to identify druggable targets in therapy resistant triple-negative breast cancer (TNBC) associated with DNA replication and repair pathways. These pathways are known to play an important role in the adaptive resistome. For the identification of factors to resensitize the cancer cells to therapy in a clinically relevant setting patient-derived organoids from TNBC tumor samples are used. In combination with wet lab experiments, bioinformatic analysis are conducted to discover key changes within the signaling cascades of DNA repair and replication processes. We anticipate that our approach will reveal new insights into the resistance mechanisms of TNBC. This might enable the development of novel potent multi-targeting concepts in combination with conventional radiochemotherapy.

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Fig. 3: (A) Patient-derived organoids of primary tumor biopsies. (B) Immunofluorescence of the DNA damage marker yH2Ax after irradiation.

Running third party funding

DFG (Bo1868/5): Imbalance of homologous recombination and DNA replication as a driver of hereditary breast cancer. Deutsche Krebshilfe (70112960): Incorrect replication processes to identify genes of early onset breast cancer. BMBF (02NUK35B): DNA repair foci as a marker of genomic instability. BMBF (02NUK39A): Outside in: How is radiation resistance in the S-phase modulated by metabolism? Wilhelm Sander Foundation (2017.106.1): Characterization and validation of Oct4 and the Oct4-associated gene signature as potential radiation resistance biomarkers in head and neck tumors. Hamburger Krebsgesellschaft eV: The relevance of tumor stem cells for radiation resistance in triple-negative breast cancer (AM. Engel). Clinical relevance of DNA damage response in osseous metastatic breast cancer under radiotherapy (Y. Goy). Effects of mutations in the breast cancer genes PALB2, ABRAXAS, BRCA1 and BRCA2 on replication in breast cancer patients (J. Willmann).

Selected recent publications

• Meyer F, Becker S, Classen S, Parplys AC, Mansour WY, Riepen B, Timm S, Ruebe C, Jasin M, Wikman H, Petersen C, Rothkamm K, Borgmann K. Prevention of DNA Replication Stress by CHK1 Leads to Chemoresistance Despite a DNA Repair Defect in Homologous Recombination in Breast Cancer. Cells. 2020 Jan 17;9(1):238.
• Schulz A, Meyer F, Dubrovska A, Borgmann K. Cancer Stem Cells and Radioresistance: DNA Repair and Beyond. Cancers (Basel). 2019 Jun 21;11(6):862.
• Bose M, Sachsenweger J, Laurila N, Parplys AC, Willmann J, Jungwirth J, Groth M, Rapakko K, Nieminen P, Friedl TWP, Heiserich L, Meyer F, Tuppurainen H, Peltoketo H, Nevanlinna H, Pylkäs K,Borgmann K, Wiesmüller L, Winqvist R, Pospiech. BRCA1 mislocalization leads to aberrant DNA damage response in heterozygous ABRAXAS1 mutation carrier cells. H.Hum Mol Genet. 2019 Dec 15;28(24):4148-4160.
• Marcar L, Bardhan K, Gheorghiu L, Dinkelborg P, Pfäffle H, Liu Q, Wang M, Piotrowska Z, Sequist LV, Borgmann K, Settleman JE, Engelman JA, Hata AN, Willers H. Acquired Resistance of EGFR-Mutated Lung Cancer to Tyrosine Kinase Inhibitor Treatment Promotes PARP Inhibitor Sensitivity. Cell Rep. 2019 Cell Rep. 2019 Jun 18;27(12):3422-3432
• Dinkelborg PH, Wang M, Gheorghiu L, Gurski JM, Hong TS, Benes CH, Juric D, Jimenez RB,Borgmann K, Willers H. A common Chk1-dependent phenotype of DNA double-strand break suppression in two distinct radioresistant cancer types. Breast Cancer Res Treat. 2019 Apr;174(3):605-613.
• Verhagen CVM, Vossen DM, Borgmann K, Hageman F, Grénman R, Verwijs-Janssen M, Mout L, Kluin RJC, Nieuwland M, Severson TM, Velds A, Kerkhoven R, O'Connor MJ, van der Heijden M, van Velthuysen ML, Verheij M, Wreesmann VB, Wessels LFA, van den Brekel MWM, Vens C. Fanconi anemia and homologous recombination gene variants are associated with functional DNA repair defects in vitro and poor outcome in patients with advanced head and neck squamous cell carcinoma. Oncotarget. 2018 Apr 6;9(26):18198-18213.
• Pai GM, Zielinski A, Koalick D, Ludwig K, Wang ZQ, Borgmann K, Pospiech H, Rubio I TSC loss distorts DNA replication programme and sensitises cells to genotoxic stress. Oncotarget. 2016 Dec 20;7(51):85365-85380.
• Berte N, Piée-Staffa A, Piecha N, Wang M, Borgmann K, Kaina B, Nikolova T. Targeting Homologous Recombination by Pharmacological Inhibitors Enhances the Killing Response of Glioblastoma Cells Treated with Alkylating Drugs. Mol Cancer Ther. 2016 Nov;15(11):2665-2678.
• Nikolova T, Göder A, Parplys A, Borgmann K. DNA Fiber Spreading Assay to Test HDACi Effects on DNA and Its Replication. Methods Mol Biol. 2017;1510:103-113.
• Foertsch F, Szambowska A, Weise A, Zielinski A, Schlott B, Kraft F, Mrasek K, Borgmann K, Pospiech H, Grosse F, Melle C. S100A11 plays a role in homologous recombination and genome maintenance by influencing the persistence of RAD51 in DNA repair foci.Cell Cycle. 2016 Oct 17;15(20):2766-79.
• Borgmann K, Köcher S, Kriegs M, Mansour WY, Parplys AC, Rieckmann T, Rothkamm K. DNA Repair. Recent Results Cancer Res. 2016;198:1-24.
• Penterling C, Drexler GA, Böhland C, Stamp R, Wilke C, Braselmann H, Caldwell RB, Reindl J, Girst S, Greubel C, Siebenwirth C, Mansour WY, Borgmann K, Dollinger G, Unger K, Friedl AA. Depletion of Histone Demethylase Jarid1A Resulting in Histone Hyperacetylation and Radiation Sensitivity Does Not Affect DNA Double-Strand Break Repair. PLoS One. 2016 Jun 2;11(6):e0156599.
• Köhler C, Koalick D, Fabricius A, Parplys AC, Borgmann K, Pospiech H, Grosse F. Cdc45 is limiting for replication initiation in humans. Cell Cycle. 2016;15(7):974-85.
• Wurster S, Hennes F, Parplys AC, Seelbach JI, Mansour WY, Zielinski A, Petersen C, Clauditz TS, Münscher A, Friedl AA, Borgmann K. PARP1 inhibition radiosensitizes HNSCC cells deficient in homologous recombination by disabling the DNA replication fork elongation response. Oncotarget. 2016 Mar 1;7(9):9732-41.
• Parplys AC, Seelbach JI, Becker S, Behr M, Wrona A, Jend C, Mansour WY, Joosse SA, Stuerzbecher HW, Pospiech H, Petersen C, Dikomey E, Borgmann K. High levels of RAD51 perturb DNA replication elongation and cause unscheduled origin firing due to impaired CHK1 activation. Cell Cycle. 2015;14(19):3190-202.