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Imprint
The role of endocrine active substances and hormone receptors for centrosome amplification-induced chromosomal instability in colorectal cancer
Project
Project code: BfR-ZEBET-08-1322-733
Contract period: 01.12.2018
- 31.12.2019
Purpose of research: Experimental development
Endocrine-active substances (EAS) refer to a group of substances with hormone or hormone-like actions, which may be of natural or synthetic origin (xenoestrogens) and encounter us as biocides, pesticides or as components in many plastic packages. In addition to effects on the hormone system, they are also closely related to the development of colorectal cancer. Specifically, they can promote the development and progression of cancer cells by affecting cell division processes or by stimulating metastasis. However, the underlying cellular and molecular mechanisms of this hormone-induced risk potential are largely unclear. As the third most common cancer type in the world, colorectal cancer belongs to the most prevalent diseases with mortality rates of up to 21%. In order to uncover the underlying cellular and molecular mechanisms of this hormone-induced risk potential, many animal experiments are currently performed. Given important concerns, not only from an ethical but also from an economic point of view, and considering difficulties regarding the transferability to humans, the development of alternative methods for animal testing is indispensable. In order to develop alternative methods according to the 3R principles of Russel and Burch (1959), the identification of the underlying mechanisms that lead to colon cancer represents a necessary prerequisite. Hence, we plan to investigate a potential link between natural and synthetic hormones, the generation of supernumerary centrosomes and chromosomal instability (CIN) using molecular and cellular biology-based methods which take receptor-mediated signaling pathways into account. Thereby, the identification and characterization of the mode-of-action of EAS regarding cell proliferation, does not only provide important implications for human health risks(e.g. EAS-induced colorectal cancer), but does also allow the derivation of endpoints important for the development of in vitro alternative methods with respect to EAS-identification (e.g. an EAS-induced centrosome amplification). We are therefore planning the following experiments: In a first step, microscopy-based methods will be used to investigate the karyotypic stability of various chromosomally stable intestinal (cancer) cell lines after ongoing EAS treatment. Since our first investigations point to an association between EAS and chromosomal maldistribution, we plan to verify numerical chromosome aberrations using karyotype analysis upon EAS treatment. To uncover the underlying mechanisms, we refer to our preliminary data, which suggest a link between EAS and the generation of supernumerary centrosomes. Numerical centrosome aberrations are closely associated with the formation of CIN and may arise from different causes. The aim of this project is to use high-resolution microscopy analysis to investigate whether hormones can cause structural changes of the centrosomes. Since centrosome amplification (CA) can often be triggered by altered expression levels of important centrosomal structural proteins and kinases, the expression level of selected centrosome-associated components should also be investigated after EAS treatment on mRNA and protein level. It is known that CA can lead to defective and hyper-stable attachments of the spindle fibers to the contact points of the chromosomes (kinetochores), which subsequently promote chromosome maldistribution and CIN. For this reason, the half-lives of microtubule binding to the kinetochores should be further investigated within the scope of a collaboration. Incorrect attachments due to EAS treatment should be directly detected using super resolution microscopy. Since EAS-mediated adverse effects may depend on classical (ERa/b) and/or alternative hormone receptors(GPER1/GPR30), we plan to determine the endogenous expression levels of the respective receptors in the presence and absence of EAS in different intestinal (cancer) cell lines. Our preliminary work indicates an important role for GPER1/GPR30 in EAS-mediated centrosome amplification. Therefore, the next step is to investigate whether other EAS-dependent lesions, such as chromosome maldistribution and centrosomal aberrations (structural and expression levels), depend on GPER1/GPR30. Last but not least, we want to investigate whether natural and xenoestrogens induce cancer-associated adverse effects, such as increased cellular migration and invasion or cancer-like tumor growth, and whether this depends on GPER1/GPR30 and the downstream signaling cascade.
Section overview
Subjects
- Biotechnology
- Toxicology
Framework programme
Funding programme
Excutive institution
BfR - Centre for Documentation and Evaluation of Alternatives to Animal Experiments (BfR - ZEBET)
