Role of the heparan sulfate proteoglycan Syndecan-1 in radiation resistance of breast cancer

• Applicants: Professor Burkhard Greve, Ph.D.; Professor Dr. Martin Götte
• Subject Area: Gynaecology and Obstetrics ; Biochemistry ; Nuclear Medicine, Radiotherapy, Radiobiology
• Term: from 2020 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 438042087

Although radiotherapy plays an important role in the treatment of breast cancer, the molecular mechanisms underlying recurrence are only poorly understood. Joint work of the applicants has indicated a previously unknown link between the cell surface proteoglycan Sdc-1 and resistance of breast cancer cells to radiation. Along with increased integrin-dependent cell motility and invasiveness, we could demonstrate an increased activation of FAK in Sdc-1-depleted breast cancer cells. Notably, recent work by others has provided a link between activation of integrins / FAK and resistance of cancer cells to radiation. We hypothesize that downregulation of Sdc-1 in breast cancer cells induces resistance to radiation and increased cell motility and invasiveness via increased activation of beta-integrins and their downstream signaling kinase FAK. To test our hypothesis, we will address the following objectives: (i) We will characterize the Sdc-1-dependent radiation phenotype in breast cancer cells investigating apoptotic/necrotic cell death, cell proliferation and DNA double strand breaks and repair capacity. (ii) We will perform video microscopy analysis of Sdc-1 depleted cells +/- radiation to investigate a potential role of Sdc-1 in radiation. Induced cell motility and invasiveness. (iii) We will confirm the results of a transcriptomic analysis of MDA-MB-231 cells subjected to Sdc-1 siRNA knockdown and radiation in independent biological replicates by qPCR and Western blotting to identify the molecular players relevant to the radiation phenotype in an unbiased approach. Relevant molecules will be further studied in functional complementation assays. (iv) We will study the role of Sdc-1 dependent signal transduction pathways in the altered response to radiation using an unbiased phosphokinase array screening approach and a more detailed analysis of beta-integrin / FAK-related signal transduction pathways. (v) We will determine the functional impact of Sdc-1 pathway inhibition on the radiation response of breast cancer cells using (a) a combination of Sdc-1 siRNA depletion and an antiintegrin antibody known to modulate the radiation response of tumor cells, (b) heparitinase degradation of the heparan sulfate chains of Sdc-1, (c) HPLC-purified Sdc-1 ectodomain, (d) an anti-human Sdc-1 antibody and (e) pharmacological inhibitors of signaling pathways identified in the preceding work packages, including a FAK inhibitor. Readouts will be colony formation, cell motility (video microscopy) and matrigel invasiveness. Overall, these objectives will help to identify and validate the molecular players involved in the Sdc-1-dependent modulation of the response of breast cancer cells to radiation therapy. If successfully implemented, this work program will provide mechanistic clues as a groundwork for developing targeted pharmacological approaches for improving the response to radiation therapy, and minimizing side effects such as radiation-induced metastasis.

• DFG Programme: Research Grants