Forschungsprojekte aus dem DNA Reparatur Labor

Regulation of DNA repair functions in mammalian cells

The capacity of mammalian cells to cope with DNA lesions varies with cell type and differentiation. In humans also distinct inter-individual differences were observed. Analyzing normal and malignant hematopoetic cells we screen for common regulatory pattern within the “DNA damage response network” and try to identify rate-limiting steps along the major repair pathways. Functional data from leukemic cells indicate that the stringent control and fine tuning of this complex network in physiological cells is (partly) abolished during carcinogenesis. A relaxed regulation of damage response genes, in turn, may facilitates the outgrow of resistant cell clones in vivo under the selective pressure of DNA-reactive anticancer drugs.
Using lab-based immunoanalytical techniques we can visualize and measure drug-induced DNA damage at single cell resolution and thereby analyze such processes in model systems and in clinical specimens. Our research is primarily focused on the role of DNA repair mechanisms for the individual cancer risk and for the chemosensitivity of e.g. hematopoetic stem and progenitor cells.

Global genomic vs. gene-specific repair: what is important for the efficacy of anticancer drugs interacting with DNA?

Depending on their actual location within the genome of mammalian cells structural lesions can persist variably long. This “toposelectivity” of DNA repair is due to specialized mechanisms taking care only for active genes or for their transcribed strand (transcription-coupled repair; TCR). These specific features have a major impact on the mutagenic and cytotoxic potency of DNA damaging agents. The relative importance of “toposelective” versus global genomic repair for the chemosensitivity of tumor cells, however, is widely unknown. Experiments comparing wild type and TCR-defective mice have shown an altered toxicity profile for a number of anticancer drugs in “k.o.” animals. Thus, key components of these pathways may be pinpointed as potential target molecules for the pharmacological modulation of drug resistance. Our group has developed monoclonal antibodies for drug-induced DNA lesions to monitor their sequence-specific distribution and elimination in normal and in tumor tissue.

Thomale J, Hochleitner K, Rajewsky MF.
Differential formation and repair of the mutagenic DNA alkylation product O6-ethylguanine in transcribed and nontranscribed genes of the rat.
J Biol Chem. 1994 Jan 21;269(3):1681-6

Engelbergs J, Thomale J, Rajewsky MF.
Role of DNA repair in carcinogen-induced ras mutation.
Mutat Res. 2000 May 30;450(1-2):139-53.

Buschfort C, Muller MR, Seeber S, Rajewsky MF, Thomale J.
DNA excision repair profiles of normal and leukemic human lymphocytes: functional analysis at the single-cell level.
Cancer Res. 1997 Feb 15;57(4):651-8

Hochleitner K, Thomale J, Nikitin AYu, Rajewsky MF.
Monoclonal antibody-based, selective isolation of DNA fragments containing an alkylated base to be quantified in defined gene sequences.
Nucleic Acids Res. 1991 Aug 25;19(16):4467-72

Interference of signal modulating drugs with the cellular processing of DNA damage

Cancer treatment schedules frequently include molecules, which modulate critical regulatory functions in tumor cells. Some of these “signal modulators” show a pronounced increase in efficacy when combined with DNA-reactive cytostatics. The interface of this synergism is most likely located in the complex network, which coordinates the cellular response to DNA injury. In collaboration with the group of Dr Moritz (Experimental Hematology, Tumor Clinic) we investigate the interplay of signal modulation and DNA repair functions in cell lines and in primary tumor cells to provide information for rational drug combinations.

Muller MR, Seiler F, Thomale J, Buschfort C, Rajewsky MF, Seeber S.
Capacity of individual chronic lymphatic leukemia lymphocytes and leukemic blast cells for repair of O6-ethylguanine in DNA: relation to chemosensitivity in vitro and treatment outcome.
Cancer Res. 1994 Aug 15;54(16):4524-31.

Cellular and molecular parameters determining tissue-specific drug toxicity

Anticancer drugs frequently provoke typical spectra of toxic side effects and thereby limiting the applicable doses. This toxicity may be related to increased formation or prolonged persistance of drug-induced DNA lesions in critical cells of the affected tissues. As a paradigm we analyze molecular and cellular mechanisms underlying the severe cytotoxicity e.g. in the inner ear and in the peripheral nerveous system after Cisplatin chemotherapy (collaboration with collegues of the Neurology and ENT clinics). Using normal and repair-transgenic mouse models we have shown that the risk of Cisplatin-induced polyneuropathy is related to the activity of a specific DNA repair pathway in dorsal root ganglia cells. On the other hand, the onset of deafness after Cisplatin treatment coincides with a very high accumulation of drug-induced adducts in a small number of specific cells within the cochlea. We try to identify relevant pharmacodynamic parameters (like membrane transporters) and the cascade of consecutive events leading to functional loss. Furthermore, the established mouse models can be applied to develop and validate new protective strategies.



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