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Session Item

Monday
November 30
08:00 - 08:40
Physics Stream 2
4D imaging for radiation therapy using MRI and PET
3025
Teaching Lecture
Physics
17:01 - 17:09
The Adaptive Resistome in TNBC: Functional Targeting in Patient-derived Organoids and Cell Lines
PH-0234

Abstract

The Adaptive Resistome in TNBC: Functional Targeting in Patient-derived Organoids and Cell Lines
Authors: CLASSEN|, Sandra(1)*[s.classen@uke.de];Rahlf|, Elena(1);Görte|, Josephine(2);Parplys|, Ann Christin(1);Schumacher|, Udo(3);Petersen|, Cordula(4);Rothkamm|, Kai(1);Toulany|, Mahmoud(5);Cordes|, Nils(2);Borgmann|, Kerstin(1);
(1)University Medical Center Hamburg-Eppendorf, Laboratory for Radiobiology and Experimental Radiooncology, Hamburg, Germany;(2)OncoRay – National Center for Radiation Research in Oncology- Technische Universität Dresden, Molecular and Cellular Radiobiology, Dresden, Germany;(3)University Medical Center Hamburg-Eppendorf, Department of Anatomy and Experimental Morphology, Hamburg, Germany;(4)University Medical Center Hamburg-Eppendorf, Department of Radiotherapy and Radiation Oncology, Hamburg, Germany;(5)University of Tuebingen, Division of Radiobiology & Molecular Environmental Research Department of Radiation Oncology, Tuebingen, Germany;
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Purpose or Objective

Despite a good initial response to radio- and chemotherapy, triple-negative breast cancer (TNBC) remains the subtype with the worst overall prognosis and survival. One major challenge in TNBC treatment is the acquired resistance against radio- and chemotherapy of recurrent tumors. Resensitizing the tumor cells by targeted inhibition of resistome-related factors seems to be a promising therapeutic approach. Therefore, this project aims to elicit druggable factors associated with DNA replication and repair pathways, which are known to play an important role in the adaptive resistome.

Material and Methods

Patient-derived organoids and isogenic MCF7 clones with different BRCA1 expression levels were grown in 3D cell culture. For validation of the 3D architecture, the organoids were stained with hematoxylin and eosin. As the BRCA1 level has a direct impact on homologous recombination (HR) and radioresistance, the viability of the clones after irradiation up to 12 Gy was monitored using an ATP-luminescence assay. Additionally, the DNA fiber assay was employed to investigate the replication stress level after irradiation with 6 Gy. The number of stem cells in the cultures was detected by analysis of the stem cell marker aldehyde dehydrogenase 1 (ALDH1) using the AldefluorTM assay.

Results

The histological analysis revealed a successful growth of the patient-derived organoids and 3D organized spheroids of the isogenic MCF7 clones. As expected, the radiosensitivity was dependent on the BRCA1 expression level. Likewise, sensitization to irradiation correlated with an increase in the replication stress level represented by fork stalling after 6 Gy irradiation (R2 = 0.96), whereas resistance to irradiation was accompanied by an increase up to 25 % in the number of ALDH1-positive stem cells. Dependent on the results of the viability analysis and replication stress level after irradiation the clones were classified as radio-resistant or sensitive. Since this classification could be successfully employed, we are currently conducting the PAMgene technology to characterize the serine/threonine kinase profiles of radio-resistant and –sensitive organoids to identify possible druggable kinases. The radioresistant organoids will be treated with a combination of clinically relevant inhibitors against the identified kinases to determine the resensitizing effects.

Conclusion

Collectively, these results indicate that we can identify radio-resistant or -sensitive tumors by monitoring the survival and replication stress level after irradiation. This classification enables the targeted analysis of factors (e.g. kinases) in radioresistant cells, thus, providing possible new targets to resensitize the cells to radiotherapy. Clinically, this would offer a new approach for the treatment of radioresistant TNBCs.