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Combined effects of Olaparib and DNA damage inducing therapies in a zebrafish xenograft model of triple negative breast cancer
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BRCA1/2 genes were identified as determinant risk-factors in breast cancer and are particularly relevant for triple negative breast cancer where approximately one in five patients has BRCA1/2 inactivation. Triple negative breast cancer is the molecular subtype associated with a worst prognosis due to unavailability of targeted therapies. In the treatment options for advanced triple negative breast cancer PARP inhibitor Olaparib is given as a third-line treatment to patients with a BRCA1/2 mutation. Unfortunately, many patients with BRCA1/2 mutations show resistance to the drug whereas wild type patients can have a striking sensitivity, making BRCA status a poor biomarker for treatment choice. Therefore, a test that determines the sensitivity to PARP inhibitors is an unmet need for the treatment management of these patients. Both BRCA1/2 and PARP are components of the genetic machinery responsible for the maintenance of genomic stability and considered as caretaker genes due to their role in DNA damage repair pathways. DNA damage induces the activation of a complex network of pathways that have the main goal of DNA repair or, in cases where this damage is unrepairable leads to the induction of cellular fates such as apoptosis, autophagy, mitotic catastrophe or senescence. In this sense, the therapeutic strategy behind PARP inhibition has as a rationale the concept of synthetic lethality – cells that have deficiencies in one or more DNA repair pathways can become dependent on PARP for repair and by inhibiting the PARP dependent pathways the cell has no more alternative options to repair DNA damage and undergoes cell death. Recently, Fior et al. showed that zebrafish xenografts have resolution to reveal intra and inter-tumor heterogeneity and differential response to therapies in just 4 days. This approach unveils not only a feasible clinical platform for screening for personalized treatments, but also a valuable model to study the interlink between tumor microenvironment, cancer development and therapeutic response. Previous work from the Lab, tested whether the zebrafish xenograft model could be used to test Olaparib sensitivity. As a proof of concept, isogenic Chinese Hamster cell lines varying only on their BRCA2 status were xenotransplated to zebrafish larvae. After 4 days of Olaparib treatment different effects could be identified depending on BRCA status. Furthermore, when using Olaparib in combination with ionizing radiation, the levels of activated caspase 3 increased in both mutant and wild-type cell lines. These results show that zebrafish larvae xenografts constitute a model capable of discerning the tumor response to treatment with a PARP inhibitor in monotherapy or in combination with ionizing radiation, in a rapid and highly sensitive manner. However, this study was performed in hamster cell lines, therefore in order to have a more translational application, in the present study our goal is to evaluate the differential tumor responses to PARP inhibition, with human cell lines, both as a single agent or combined with DNA damage inducing therapies. Zebrafish xenografts were developed with human triple negative breast cancer cell lines with different BRCA status. When challenged with the PARP inhibitor treatment as a single agent, different cell lines showed different responses, revealing a sensitivity phenotype independent of BRCA status. The combination with DNA damage inducing therapies increased the cytotoxic effects in both sensitive and resistant cell lines and led to a reduction in both metastatic potential and angiogenesis. In conclusion, this work reveals not only the potential of the zebrafish xenograft model as a sensitivity profiling platform for Olaparib, but also as a screening platform capable of determining the effects of different therapeutic combinations. Also, given the translational aspect of this study we can also report clinical considerations in the reinforcement of the need for a better selection method for the patients that will benefit from treatment with Olaparib but also that the combination with other therapies has the potential to increase the cytotoxic effects in both sensitive and resistant patients.
Descrição
Tese de mestrado, Biologia Molecular e Genética, Universidade de Lisboa, Faculdade de Ciências, 2019
Palavras-chave
Cancro da mama BRCA1/2 PARP Olaparib Xenotransplantes Peixe-zebra Teses de mestrado - 2019