Universität zu Köln

Orschel, Charlotte ORCID: 0000-0002-5323-9475 (2026). Deciphering metastatic signaling cascades in small cell lung cancer. PhD thesis, Universität zu Köln.

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Abstract

Down to the present-day small cell lung cancer (SCLC) remains one of the deadliest cancer entities. Due to its early metastasis formation and wide therapy resistance, SCLC occupies a special place within cancer research. Given the limited treatment options, particularly in advanced stages, it is vital to examine how certain signaling cascades contribute to the aggressive phenotype, immune evasion and high migration potential of SCLC. My dissertation mainly focuses on the CD29-dependent signaling pathway and MHC-I dependent regulation of the tumor micro-milieu and discusses their involvement in processes of metastasis formation and immune evasion mechanisms of SCLC. In our initial screening, we observed elevated CD29 expression in liver metastases in both human and murine samples. Therefore, CD29 Knockout (KO) clones were generated using CRISPR to test the characteristics on functional and cellular level in comparison to the original liver metastasis-derived cell line. To evaluate migration behavior, the different clones and the wild type were subjected to a series of cell migration and cell–cell interaction tests (Scratch Assay, Boyden Chamber). To elucidate the functional role of CD29 in metastasis formation, the CD29 KO clones were also tested in an in vivo setting. In vivo, tumors were induced via orthotopic and intravenous injection of the respective tumor cells into immunocompetent and immune deficient mouse models. A fluorescence-activated cell sorting (FACS) readout of tumor and metastasis tissue provided insights into immune cell involvement and tumor expression markers. Additionally, immunohistochemistry (IHC) was applied to assess tumor cells and desmoplastic reactions ex vivo. In vitro migration assays demonstrated that CD29 KO abrogates fibronectin- and angiopoietin-2 (ANG-2) stimulated migration. In vivo experiments further revealed that, in contrast to the wild type SCLC cells, CD29 KO clones did not form liver metastasis, which was confirmed by flow cytometry and IHC. Altogether, CD29 is a potent driver of metastasis in SCLC, which can be abrogated by knockout. Furthermore, my dissertation explores MHC-I and its role in SCLC immune evasion and metastasis formation. In a patient sample screen, we found a downregulation of MHC-I in SCLC liver metastases. To investigate the impact of MHC-I loss on tumor-immune interactions, MHC-I-deficient clones (MHC-I KO) were generated by targeted knockout of the β2 microglobulin (B2M) gene. The MHC-I KO was tested in vivo by orthotopic and intravenous injection and presented a more aggressive, fast-growing phenotype. In MHC-I KO tumors, we found an enhanced reaction of the innate immune system in the liver metastases and a decreased T cell answer in the pleural effusion. A comparison between primary tumors and metastases revealed increased phosphorylation of ERBB2 in SCLC metastases. Initial in vitro experiments with an ERBB2 inhibitor demonstrated suppression of key downstream signaling pathways and induction of immune activating signals via TBK1. Notably, in our autochthonous SCLC mouse model, combined blockade of PD-1 and ERBB2 resulted in a significant extension of both overall and progression-free survival. Overall, this study states the importance of the CD29-ANG-2 signaling axis in SCLC metastasis formation and portrays the immune evading effects caused by MHC-I loss. Based on these findings, potential therapeutic approaches are discussed and tested in preclinical setting.

Item Type:	Thesis (PhD thesis)
Translated title:	Title Language Entschlüsselung metastatischer Signalkaskaden beim kleinzelligen Lungenkarzinom German
Translated abstract:	Abstract Language Das kleinzellige Lungenkarzinom (SCLC) zählt nach wie vor zu den tödlichsten Krebserkrankungen. Die frühe Metastasierung und die schnelle Entwicklung von Resistenzen gegenüber klassischen Therapien stellen eine besondere Herausforderung dar. Da es vor allem im fortgeschrittenen Stadium kaum wirksame Behandlungsoptionen gibt, ist es von besonderer Bedeutung, die Mechanismen zu ergründen, die für die aggressive Natur, die hohe Migrationsfähigkeit und das Umgehen des Immunsystems des SCLCs verantwortlich sind. In meiner Dissertation fokussiere ich mich auf den CD29-abhängigen Signalweg und die MHC I-abhängige Regulation im Tumormikromilieu und erörtere deren Beteiligung an Prozessen der Metastasenbildung und Immunresistenz. Zunächst zeigte ein breites Screening von SCLC-Probenpaaren aus Primärtumor und Lebermetastasen, menschlichen und murinen Ursprungs, eine erhöhte CD29 Expression im Metastasengewebe. Um die genaue Funktion von CD29 zu verstehen, wurden Knockout-(KO) SCLC Klone mithilfe von CRISPR erzeugt. Diese Klone wurden auf zellulärer und funktioneller Ebene mit der ursprünglichen, aus Lebermetastasen gewonnenen Zelllinie verglichen. Dazu wurden die Verfahren des Scratch Assays und der Boyden Chamber angewandt, um das Migrations- bzw. Invasionsverhalten des CD29 KO zu analysieren. Zusätzlich wurden in vivo Experimente durchgeführt, bei denen CD29 KO und Wildtyp SCLC Tumorzellen in immunkompetente und immundefiziente Mäuse injiziert wurden, um die Metastasenbildung zu untersuchen. Die Ergebnisse zeigten, dass der CD29 KO das Migrationsvermögen der Tumorzellen hemmte, insbesondere unter Stimulation mit Fibronektin und Angiopoietin-2 (ANG-2). In vivo konnten die CD29 KO Tumorzellen im Gegensatz zum Wildtyp keine Lebermetastasen bilden, was durch immunhistochemische Analysen und Durchflusszytometrie bestätigt wurde. Insgesamt zeigt sich, dass CD29 eine entscheidende Rolle bei der Metastasenbildung des SCLCs spielt, was durch einen Knockout unterdrückt werden kann. Im weiteren Verlauf, konzentriert sich meine Dissertation auf MHC-I und dessen Rolle bei der Umgehung der Immunabwehr und Metastasenbildung. Bei der Analyse der gepaarten SCLC Patientenproben zeigte sich eine reduzierte MHC-I Expression in den Lebermetastasen. Zur Untersuchung der Auswirkungen eines MHC-I Verlusts auf die Wechselwirkungen zwischen Tumor und Immunsystem wurden MHC-I defiziente Klone (MHC-I KO) durch gezielten Knockout des β2-Mikroglobulin-Gens (B2M) erzeugt. In vivo zeigte sich, dass die MHC-I KO Tumore deutlich aggressiver wuchsen. Dabei wurde eine verstärkte Aktivierung des angeborenen Immunsystems in den Lebermetastasen und eine reduzierte T-Zell-Antwort im Pleuraerguss beobachtet. Allerdings zeigte sich keine ausgeprägte Lebermetastasierung. Ein Vergleich zwischen Primärtumoren und Metastasen ergab eine verstärkte Phosphorylierung von ERBB2 in den SCLC Metastasen. Erste in vitro Experimente mit einem ERBB2 Inhibitor 10 zeigten eine Hemmung zentraler nachgeschalteter Signalwege und die Aktivierung immunstimulierender Signale durch TBK1. In unserem autochthonen SCLC Mausmodell erzielte die kombinierte Blockade von PD-1 und ERBB2 eine signifikante Verlängerung des Gesamtüberlebens, sowie des progressionsfreien Überlebens. Zusammenfassend stellt diese Dissertation die entscheidende Bedeutung der CD29-ANG-2-Signalkaskade für die Metastasenbildung von SCLC sowie die immunologischen Folgen des MHC-I Verlusts dar, und erörtert darauf basierend mögliche Therapieansätze. German
Creators:	Creators Email ORCID ORCID Put Code Orschel, Charlotte orschelcharlotte@gmail.de https://orcid.org/0000-0002-5323-9475 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-795987
Date:	2026
Language:	English
Faculty:	Faculty of Medicine
Divisions:	Faculty of Medicine > Innere Medizin > Klinik I für Innere Medizin - Hämatologie und Onkologie
Subjects:	Medical sciences Medicine
Uncontrolled Keywords:	Keywords Language Metastasis German SCLC German Targeted theraoy German
Date of oral exam:	4 December 2025
Referee:	Name Academic Title Meder, Lydia Professorin Dr. nat. med. Stripecke, Renata Universitätsprofessorin Dr. rer. nat.
Funders:	Mildred Scheel School of Oncology
Projects:	Dissertation
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Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/79598