Development of Methods to Validate Synthetic Lethalitty In vitro and In vivo

CASTELLARO, AM; NIGRA, AD; CARBAJOSA, S; RODRIGUEZ-BAILI, MC.; RACCA, AC; PANSA, MF; GARCÍA, LA; GIL, GA; SORIA, G

Buenos Aires

Congreso; LIII REUNIÓN ANUAL DE LA SOCIEDAD ARGENTINA DE INVESTIGACIÓN BIOQUÍMICA Y BIOLOGÍA MOLECULAR (SAIB); 2017

One of the current challenges in the development of anticancer therapies lies in the identification of drugs that selectively kill cancer cells with minor impact on normal cells. A strategy of selective cytotoxicity that has yielded promising results is the induction of synthetic lethality (SL). SL induction involves Tumor-acquired mutations that sensitize cells to a given treatment when they are in combination. We are focused in the study of drugs that cause SL in BRCA1/2-deficient cancer cells (Bd-cells). Herein we show the validation of potential drugs which have been previously identified as SL-inducers using a cell-based high-throughput screening platform based on flow cytometry. Three validation methods were developed to assess the performance of these drugs on a Bd-cell model in comparison to the wild type cell line (Bw-cells). The first method evaluates the clonogenic potential of Bd-cells and Bw-cells after the treatmentwith the drugs, using an adaptation of the colony formation assay.The second method involves the use of 3D chimeric spheroids constituted by an equal proportion of Bd-cells and Bw-cells. Both cell populations are tagged with different fluorescent proteins and can therefore be quantified by flow cytometry after the treatments with the drugs. The third model is a double tumor xenograft approach in Nude mice. This model consists in the generation of flanking tumorsin a single mouse, one from Bd-cells and the other from Bw-cells.Mice are then treated with potential SL-inducers or vehicle and the progression of each tumor-type is assessed in order to expose the SL phenomena in vivo