Chronisch-entzündliche Darmerkrankungen wie Colitis ulcerosa (CU) sind Risikofaktoren für die Erkrankung an Kolorektalkarzinomen (KRK) (Eaden, Abrams et al. 2001, Lutgens, Vleggaar et al. 2008). Sowohl in CU (Hussain, Amstad et al. 2000, Takaku, Ajioka et al. 2001, Leedham, Graham et al. 2009, Kakiuchi, Yoshida et al. 2020) als auch in Colitis-assoziierten KRK (CA-KRK) (Yin, Harpaz et al. 1993, Brentnall, Crispin et al. 1994, Galandiuk, Rodriguez–Justo et al. 2012, Kobayashi, Tomita et al. 2017, Baker, Cross et al. 2019) treten vermehrt epitheliale Zellklone auf, die einen Verlust an physiologischem Tumorsupressor P53 Signaling haben. Diese Klone treiben vermutlich die Karzinogenese voran (Petitjean, Achatz et al. 2007). Bislang ist jedoch nicht verstanden, durch welchen Mechanismus diese Klone Wildtypzellen verdrängen könnten, um in der Mucosa fixiert zu werden. In der vorliegenden Studie nutze ich induzierbare Trp53 Knock-Out (KO) Mäuse, mithilfe derer ich die Auswirkungen eines Verlusts von P53 im Darmepithel im Colitis-Kontext sowohl in vivo als auch in vitro untersuchen kann. Ich zeige, dass während Trp53 für die Aufrechterhaltung der epithelialen Homöstase im Colon verzichtbar ist, es nötig ist, damit Epithelzellen nach der Schädigung durch eine chemische Colitis wieder in die Homöstase zurückkehren können. Epithel, dem P53 fehlt, verharrt nach der Schädigung in einem fetalen, regenerativen Status („locked regenerative state“), welcher durch eine andauernde Aktivierung des YAP-Signalweges charakterisiert ist. Alles in allem offenbart diese Arbeit die kontextabhängige Bedeutung des P53 Signalings im Kolonepithel im regenerativen Status nach colitis-assoziierter Schädigung und gibt erste Erklärungen für die erhöhte Häufigkeit von Zellklonen mit verringertem P53 Signaling in CU und CA-KRK. / Chronic inflammatory bowel diseases such as ulcerative colitis (UC) are risk factors for colorectal cancer (CRC) (Eaden, Abrams et al. 2001, Lutgens, Vleggaar et al. 2008). Both in UC (Hussain, Amstad et al. 2000, Takaku, Ajioka et al. 2001, Leedham, Graham et al. 2009, Kakiuchi, Yoshida et al. 2020) and UC-associated colorectal cancer (UC-CRC) (Yin, Harpaz et al. 1993, Brentnall, Crispin et al. 1994, Galandiuk, Rodriguez–Justo et al. 2012, Kobayashi, Tomita et al. 2017, Baker, Cross et al. 2019). Cell clones that lack tumor suppressor 53 (P53) signaling are observed at high frequency and are considered drivers of carcinogenesis (Petitjean, Achatz et al. 2007). The fixation of those clones could be due to a selective advantage of mutated cells specifically in the inflammatory context (Vermeulen, Morrissey et al. 2013), however mechanistic insights why and how mutant cells could outcompete wild type (WT) cells are lacking. In the present study, I have used inducible Trp53 knock-out (KO) mice through which I investigate the impact of loss of P53 signaling in colonic epithelium in the context of colitis, both in vivo and in vitro. I show that while Trp53 is dispensable for maintenance of epithelial homeostasis in the colon, it has a crucial function in epithelial cells to make them return to homeostasis after colitis-associated injury. Integrating proteome and transcriptome data, I discover that the colonic epithelium of Trp53 KO mice fails to reestablish the homeostatic crypt architecture and is locked in a “fetal-like” regenerative state that is characterized by activated Yes-associated protein 1 (YAP) signaling. This work discloses the context-dependent relevance of P53 signaling in the injury-induced regenerative state during colitis and gives reason for the higher abundance of clones lacking P53 signaling in UC and UC-CRC. It also demonstrates, how loss of P53 could contribute to chronification of the disease and a carcinogenic trajectory in affected epithelium.

No description available.

Objective: Two chromosomal instability (CIN) pathways are described in glioblastoma multiforme (GBM), type 1 and type 2, which can be observed in up to 70% of the cases. Microsatellite instability (MSI) plays a pathogenic role in sporadic cancers such as colon, gastric and endometrial carcinomas with deficient mismatch repair (MMR). We aimed to perform a comprehensive analysis of the relationship between CIN and MSI mechanisms in sporadic glioblastomas. Methods: 129 GBMs were examined (109 newly diagnosed and 20 relapses) investigating MSI, immunohistochemical expression of MMR proteins as well as sequencing and promoter methylation of hMLH1. We characterized the molecular changes frequently correlated with CIN in MSI+ GBMs and compared them with 26 microsatellite-stable tumors. Results: Low-level MSI was observed in 11 of 129 (8.5%) cases and was higher in relapses than in primary GBMs (25 vs. 5.5%, p = 0.027). High-level MSI was not found in any case. A deficient expression of MLH1 and PMS2 without hMLH1 inactivation was observed only in one giant cell GBM. 55% of the MSI+ GBMs showed a profile which did not correspond to one of the known CIN pathways. An inverse association was observed between MSI and mutations of both p53 and PTEN. Conclusions: Our data suggest that CIN and MSI contribute to the genomic instability in GBMs via independent pathways. Since MSI was significantly more frequent in relapses, it might play a role in the malignant progression of GBM. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Objective: Two chromosomal instability (CIN) pathways are described in glioblastoma multiforme (GBM), type 1 and type 2, which can be observed in up to 70% of the cases. Microsatellite instability (MSI) plays a pathogenic role in sporadic cancers such as colon, gastric and endometrial carcinomas with deficient mismatch repair (MMR). We aimed to perform a comprehensive analysis of the relationship between CIN and MSI mechanisms in sporadic glioblastomas. Methods: 129 GBMs were examined (109 newly diagnosed and 20 relapses) investigating MSI, immunohistochemical expression of MMR proteins as well as sequencing and promoter methylation of hMLH1. We characterized the molecular changes frequently correlated with CIN in MSI+ GBMs and compared them with 26 microsatellite-stable tumors. Results: Low-level MSI was observed in 11 of 129 (8.5%) cases and was higher in relapses than in primary GBMs (25 vs. 5.5%, p = 0.027). High-level MSI was not found in any case. A deficient expression of MLH1 and PMS2 without hMLH1 inactivation was observed only in one giant cell GBM. 55% of the MSI+ GBMs showed a profile which did not correspond to one of the known CIN pathways. An inverse association was observed between MSI and mutations of both p53 and PTEN. Conclusions: Our data suggest that CIN and MSI contribute to the genomic instability in GBMs via independent pathways. Since MSI was significantly more frequent in relapses, it might play a role in the malignant progression of GBM. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Da vorliegende Studien unserer Arbeitsgruppe auf eine Beteiligung von Connexinen an der oralen Plattenepithelkarzinogenese hindeuten, war es Ziel dieser Arbeit in-vivo am Hamstermodell die Expression der Connexine 26, 43 und 45 im Verlauf der Karzinogenese zu untersuchen. Die durch 9,10-Dimethyl-1,2-Benzanthrazen (DMBA) induzierten Karzinome wurden makroskopisch, histologisch, sowie auf ihren Inflammationsgrad hin untersucht, um die Effektivität des Tiermodells zu prüfen. Weiterhin erfolgte sie Genexpressionsanalyse mittels RNA-Isolation, PCR-Analyse und statistischer Auswertung. Es konnte insgesamt gezeigt werden, dass die Applikation von DMBA in Abhängigkeit von Behandlungsdauer makroskopisch, histologisch und inflammatorisch einen deutlichen karzinogenen Effekt hatte und, dass das histopathologische Grading signifikant mit der Länge der Behandlungsdauer korrelierte. Im Rahmen der Genexpressionsanalyse konnte im Verlauf der DMBA-Behandlung für Connexin 26 und 45 eine signifikante Überexpression beschrieben werden. Connexin 43 zeigte sich als nicht differentiell exprimiert. Ein Zusammenhang zwischen histologischem Grading und Genexpression war nicht nachvollziehbar.

T-cell receptor (TCR) polyclonal mature T cells are surprisingly resistant to oncogenic transformation after retroviral insertion of T-cell oncogenes. In a mouse model, it has been shown that mature T-cell lymphoma/leukemia (MTCLL) is not induced upon transplantation of mature, TCR polyclonal wild-type (WT) T cells, transduced with gammaretroviral vectors encoding potent T-cell oncogenes, into RAG1-deficient recipients. However, further studies demonstrated that quasi-monoclonal T cells treated with the same protocol readily induced MTCLL in the recipient mice. It has been hypothesized that in the TCR polyclonal situation, outgrowth of preleukemic cells and subsequent conversion to overt malignancy is suppressed through regulation of clonal abundances on a per-clone basis due to interactions between TCRs and self-peptide-MHC-complexes (spMHCs), while these mechanisms fail in the quasi-monoclonal situation. To quantitatively study this hypothesis, we applied a mathematical modeling approach. In particular, we developed a novel ordinary differential equation model of T-cell homeostasis, in which T-cell fate depends on spMHC-TCR-interaction-triggered stimulatory signals from antigen-presenting cells (APCs). Based on our mathematical modeling approach, we identified parameter configurations of our model, which consistently explain the observed phenomena. Our results suggest that the preleukemic cells are less competent than healthy competitor cells in acquiring survival stimuli from APCs, but that proliferation of these preleukemic cells is less dependent on survival stimuli from APCs. These predictions now call for experimental validation.

No description available.

T-cell receptor (TCR) polyclonal mature T cells are surprisingly resistant to oncogenic transformation after retroviral insertion of T-cell oncogenes. In a mouse model, it has been shown that mature T-cell lymphoma/leukemia (MTCLL) is not induced upon transplantation of mature, TCR polyclonal wild-type (WT) T cells, transduced with gammaretroviral vectors encoding potent T-cell oncogenes, into RAG1-deficient recipients. However, further studies demonstrated that quasi-monoclonal T cells treated with the same protocol readily induced MTCLL in the recipient mice. It has been hypothesized that in the TCR polyclonal situation, outgrowth of preleukemic cells and subsequent conversion to overt malignancy is suppressed through regulation of clonal abundances on a per-clone basis due to interactions between TCRs and self-peptide-MHC-complexes (spMHCs), while these mechanisms fail in the quasi-monoclonal situation. To quantitatively study this hypothesis, we applied a mathematical modeling approach. In particular, we developed a novel ordinary differential equation model of T-cell homeostasis, in which T-cell fate depends on spMHC-TCR-interaction-triggered stimulatory signals from antigen-presenting cells (APCs). Based on our mathematical modeling approach, we identified parameter configurations of our model, which consistently explain the observed phenomena. Our results suggest that the preleukemic cells are less competent than healthy competitor cells in acquiring survival stimuli from APCs, but that proliferation of these preleukemic cells is less dependent on survival stimuli from APCs. These predictions now call for experimental validation.