Center for Molecular Medicine Cologne

von Karstedt, Silvia - A 06

Elucidation of ferroptosis-inducing therapy (FIT) for the treatment of pancreatic cancer

Introduction

Pancreatic ductal adenocarcinoma (PDAC) is amongst the deadliest cancers with a 5-year survival rate of below 5%. This poor prognosis is caused by absence of specific early symptoms and primary resistance to chemotherapy highlighting an urgent need to address these two problems. Molecularly, PDAC is characterised by the presence of activating point mutation in the small GTPase KRAS in 95% of the cases. This point mutation leads to constitutive activation of KRAS, promoting proliferation, invasion and cell death resistance. We recently contributed to a study identifying that the cystine/glutamate antiporter xCT (SLC7A11) promotes Ras-mediated transformation by protecting cells from reactive oxygen species (ROS) overload (Lim et al., 2019). Interestingly, xCT has been implicated in the protection against a recently discovered form of regulated necrosis, termed ferroptosis. Sensitivity to ferroptosis, in turn, has emerged as a selective vulnerability in cancer cells which escape targeted treatment. Based on these findings, we have experimentally explored whether the KRASG12D mutation which is prevalent in PDAC might be promoting malignancy through evading ferroptosis. The aim of this project is to establish i) whether ferroptosis evasion might determine KRASG12D fitness in PDAC development ii) and how breaking ferroptosis evasion might affect the development of PDAC precursor lesions in a genetically-engineered mouse model. We thereby anticipate to identify cell death vulnerabilities in conjunction with potential markers during early PDAC progression for the development of novel treatment strategies. 

Figure 1

Clinical relevance

The poor 5-year survival rate of below 5% of PDAC patients has not significantly improved since the 1970ies. This has marked PDAC as one of the cancers with unmet needs warranting the urgent requirement to explore novel therapeutic strategies. We propose to test ferroptosis-inducing therapy (FIT) in in-vitro and in-vivo PDAC model systems in order to address this need. Findings made in these model systems will be corroborated in a PDAC patient tissue bank.

Our Aims

  1. Elucidate mechanisms of ferroptosis response in KRAS-mutated cells
    • Define proteomics of KRAS mutated and wild type cells upon induction of ferroptosis
    • Define expression changes of KRAS mutated and wild type cells upon induction of ferroptosis
    • Identify differential adaptation mechanisms in KRAS-mutated PDAC upon ferroptosis
    • Validation of role of differentially regulated factors
  2. Elucidate the role of genetic ferroptosis induction in the development and progression of PDAC
    • Determine the role of ferroptosis in PDAC development in vivo
    • Test FIT in KRAS-driven genetically engineered mouse models
    • Validation of expression of ferroptosis pathway components and potential prognostic value in a patient biobank

Previous and ongoing Work

Most tumours are known to undergo constant cycles of selection via immune effector cells. One pathway via which immune effector cells kill target cells is extrinsic apoptosis induction via ligand/receptor binding of Tumor necrosis factor (TNF)- and TNF-receptor (TNFR) superfamily (SF) members (Karstedt et al., 2017). Yet, given the fast kinetics and efficiency by which extrinsic apoptosis can eliminate targeted cells, it is not surprising that tumours frequently develop escape mechanisms against extrinsic apoptosis. In KRAS-driven PDAC, we previously found that extrinsic apoptosis is disabled (Karstedt et al., 2015). Interestingly, Ferroptosis is a recently discovered type of regulated necrosis which, unlike apoptosis, is independent of caspase. Instead, ferroptotic cells die following iron-dependent lipid peroxidation, a process which is antagonised by glutathione peroxidase 4 (GPX4), by the cystine/glutamate antiporter xCT (SLC7A11) and ferroptosis suppressor protein 1 (FSP1) (reviewed in Bebber et al. 2020). Importantly, tumour cells escaping other forms of regulated cell death maintain or acquire sensitivity to ferroptosis (Bebber et al. 2021). Moreover, ferroptosis is a lytic type of cell death during which cellular plasmamembranes are permeabilised (Pedrera et al. 2020) making it potentially immunogenic and thereby particularly interesting for cancer therapy. As part of this project, we tested whether KRAS-mutated PDAC can respond to ferroptosis-inducing therapy (FIT).

Interestingly, we identified that expression of oncogenic as compared to WT KRAS in isogenic cellular systems renders cells more resistant to ferroptosis. Mechanistically, we find that cells with mutant KRAS show a specific lack of ferroptosis-induced lipid peroxidation along with upregulated expression of ferroptosis suppressor protein 1 (FSP1). Indeed, elevated levels of FSP1 in KRAS mutant cells are responsible for mediating ferroptosis resistance. Consequently, we find that pharmacological induction of ferroptosis in pancreatic organoids derived from the LsL-KRASG12D expressing mouse model is effective at killing organoids only in combination with FSP1 inhibition. Lastly, FSP1 is upregulated in non-small cell lung cancer (NSCLC), colorectal cancer (CRC) and pancreatic ductal adenocarcinoma (PDAC) as compared to the respective normal tissue of origin. Based on these data, we propose that ferroptosis-inducing therapy should be combined with FSP1 inhibitors for efficient therapy of KRAS-mutant cancers (Müller et al. under review).

  • Christina M. Bebber, Emily S. Thomas, Jenny Stroh, Zhiyi Chen, Ariadne Androulidaki, Anna Schmitt, Michaela N. Höhne, Lukas Stüker, Cleidson de Pádua Alves, Armin Khonsari, Marcel A. Dammert, Fatma Parmaksiz, Hannah L. Tumbrink, Filippo Beleggia, Martin L. Sos, Jan Riemer, Julie George, Susanne Brodesser, Roman K. Thomas, H. Christian Reinhardt and Silvia von Karstedt. Ferroptosis response segregates small cell lung cancer (SCLC) neuroendocrine subtypes. Nature Communications 12, 2048 (2021).
  • Pedrera L, Espiritu RA, Ros U, Weber J, Schmitt A, Stroh J, Hailfinger S, Karstedt von S, García-Sáez AJ. Ferroptotic pores induce Ca 2+ fluxes and ESCRT-III activation to modulate cell death kinetics. Cell Death and Differentiation. Nature Publishing Group; 2021 Dec 17;149:1–14.
  • Karstedt, von, S. & Walczak, H. An unexpected turn of fortune: targeting TRAIL KRAS-driven cancer. Cell Death Discovery 6, 14–8 (2020).
  • Clemente LP, Rabenau M, Tang S, Stanka J, Cors E, Stroh J, Culmsee C, Karstedt von S. Dynasore Blocks Ferroptosis through Combined Modulation of Iron Uptake and Inhibition of Mitochondrial Respiration. Cells 2020, Vol. 9, Page 2259. Multidisciplinary Digital Publishing Institute; 2020 Oct 1;9(10):2259
  • Bebber CM, Müller F, Prieto Clemente L, Weber J, von Karstedt S. Ferroptosis in Cancer Cell Biology. Cancers (Basel). 2020 Jan 9;12(1). pii: E164. doi: 10.3390/cancers12010164. Review. PubMed PMID: 31936571.
  • Karstedt, von, S., Conti, A., Nobis, M., Montinaro, A., Hartwig, T., Lemke, J., Legler, K., Annewanter, F., Campbell, A.D., Taraborrelli, L., et al. (2015). Cancer cell-autonomous TRAIL-R signaling promotes KRAS-driven cancer progression, invasion, and metastasis. Cancer Cell 27, 561–573.
  • Karstedt, von, S., Montinaro, A., and Walczak, H. (2017). Exploring the TRAILs less travelled: TRAIL in cancer biology and therapy. Nature Publishing Group 17, 352–366.
  • Lim, J.K.M., Delaidelli, A., Minaker, S.W., Zhang, H.-F., Colovic, M., Yang, H., Negri, G.L., Karstedt, von, S., Lockwood, W.W., Schaffer, P., et al. (2019). Cystine/glutamate antiporter xCT (SLC7A11) facilitates oncogenic RAS transformation by preserving intracellular redox balance. Proc. Natl. Acad. Sci. U.S.a. 1092, 201821323.
  • Montinaro A, Areso Zubiaur I, Saggau J, Kretz AL, Ferreira RMM, Hassan O, Kitzig E, Muller I, El-Bahrawy MA, von Karstedt S, Kulms D, Liccardi G, Lemke J, and Walczak H (2022). Potent pro-apoptotic combination therapy is highly effective in a broad range of cancers. Cell Death Differ29, 492-503. doi:10.1038/s41418-021-00869-x.
  • Bebber CM, Thomas ES, Stroh J, Chen Z, Androulidaki A, Schmitt A, Hohne MN, Stuker L, de Padua Alves C, Khonsari A, Dammert MA, Parmaksiz F, Tumbrink HL, Beleggia F, Sos ML, Riemer J, George J, Brodesser S, Thomas RK, Reinhardt HC, and von Karstedt S (2021). Ferroptosis response segregates small cell lung cancer (SCLC) neuroendocrine subtypes. Nat Commun12, 2048. doi:10.1038/s41467-021-22336-4.
  • Bragelmann J, Lorenz C, Borchmann S, Nishii K, Wegner J, Meder L, Ostendorp J, Ast DF, Heimsoeth A, Nakasuka T, Hirabae A, Okawa S, Dammert MA, Plenker D, Klein S, Lohneis P, Gu J, Godfrey LK, Forster J, Trajkovic-Arsic M, Zillinger T, Haarmann M, Quaas A, Lennartz S, Schmiel M, D'Rozario J, Thomas ES, Li H, Schmitt CA, George J, Thomas RK, von Karstedt S, Hartmann G, Buttner R, Ullrich RT, Siveke JT, Ohashi K, Schlee M, and Sos ML (2021). MAPK-pathway inhibition mediates inflammatory reprogramming and sensitizes tumors to targeted activation of innate immunity sensor RIG-I. Nat Commun12, 5505. doi:10.1038/s41467-021-25728-8.
  • Pedrera L, Espiritu RA, Ros U, Weber J, Schmitt A, Stroh J, Hailfinger S, von Karstedt S, and Garcia-Saez AJ (2021). Ferroptotic pores induce Ca(2+) fluxes and ESCRT-III activation to modulate cell death kinetics. Cell Death Differ28, 1644-1657. doi:10.1038/s41418-020-00691-x.
  • Willms A, Schupp H, Poelker M, Adawy A, Debus JF, Hartwig T, Krichel T, Fritsch J, Singh S, Walczak H, von Karstedt S, Schafer H, and Trauzold A (2021). TRAIL-receptor 2-a novel negative regulator of p53. Cell Death Dis12, 757. doi:10.1038/s41419-021-04048-1.

 

  • von Karstedt S, Walczak H.: An unexpected turn of fortune: targeting TRAIL-Rs in KRAS-driven cancer.Cell Death Discov. 2020 Mar 17;6:14. doi: 10.1038/s41420-020-0249-4. eCollection 2020. PMID: 32194994 Free PMC article. Review.
  • Allgöwer C, Kretz AL, von Karstedt S, Wittau M, Henne-Bruns D, Lemke J. Cancers (Basel).: Friend or Foe: S100 Proteins in Cancer. 2020 Jul 24;12(8):2037. doi: 10.3390/cancers12082037. PMID: 32722137 Free PMC article. Review.
  • Clemente LP, Rabenau M, Tang S, Stanka J, Cors E, Stroh J, Culmsee C, and von Karstedt S (2020). Dynasore Blocks Ferroptosis through Combined Modulation of Iron Uptake and Inhibition of Mitochondrial Respiration. Cells 9.
  • Bebber CM, Müller F, Prieto Clemente L, Weber J, von Karstedt S. Ferroptosis in Cancer Cell Biology. Cancers (Basel). 2020 Jan 9;12(1). pii: E164. doi: 10.3390/cancers12010164. Review. PubMed PMID: 31936571.
Prof. Dr. Silvia von Karstedt CMMC Cologne
Prof. Dr. Silvia von Karstedt

Department of Translational Genomics - CECAD Research Center

CMMC - PI - A 06

+49 221 478 84340

Department of Translational Genomics - CECAD Research Center

Joseph-Stelzmann-Str. 26

50931 Cologne

http://vonkarstedt.cecad-labs.de/Home.824.0.html

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Publications - Silvia von Karstedt

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Group Members

Ariadne Androulidaki, PostDoc
Eric Seidel, PostDoc
Christina Bebber, PhD student
Fabienne Müller, PhD student
Sofya Tishina, PhD student
Fatma Isil Yapici, PhD student
Fanyu Liu, PhD student
Emmanuel Sarfo Gyamfi, M.Sc student
Ioanna Kotouza, M.Sc student
Jenny Stroh, Technician
Alina Dahlhaus, Technician
Julia Beck, SHK