Characterization of novel murine mammary cell lines with different aggressive phenotype and identification of potential targets by mass spectrometry.

JOHANNA ELENA SIDABRA; GOTTARDO, MARÍA FLORENCIA; CAPOBIANCO, CARLA SABRINA; ALONSO DANIEL F.; FARINA HERNÁN G

Mar del Plata

Congreso; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica; 2019

Sociedad Argentina de Investigación Clínica

Breast cancer is the first cause of death from female cancer. After treatments against the primary tumor, a few cells can prevail in a dormant state for a long period, becoming minimal residual disease. These cells can resume growth and establish as metastasis, which are responsible for 90% of deaths from cancer. Patients with breast cancer diagnosed at early stage with small tumors have a 25-30% chance of recurrence after 10-15 years after treatment. These new growth often entails chemo and radio resistance and there is not response to conventional treatment. For all these reasons, there is a clear need for the development of novel therapies against these cells in a dormant state. The main limitation on the study of these dormant cells is the lack of relevant in vitro an in vivo models. In this work, we isolated and characterized two novel cell lines F3II-TP and F3II-NM. F3II-TP represents tumors with high proliferation rate whereas F3II-NM represents a dormant cell phenotype. The comparison between these cell lines is expected to yield new molecular targets for the design of new drugs that point dormant cellsThe study began with the isolation and establishment of new tumor cell lines. After inoculation of the F3II murine mammary carcinoma cell line, we established cell populations in vitro, one from the primary tumor and another from a spontaneous metastatic nodule, F3II TP and F3II NM cell lines respectively. First, we analyzed the expression of EMT markers such as E-cadherin, N-cadherin, Vimentin, Pan-Cytoqueratin and β-catenin by immunofluorescence. In in vitro assays, F3II NM presented major doubling time, major adhesion capacity and lower clonogenic potential. In addition, the expression of the dormancy related genes such as NR2F1 and BHLHE41 was analyzed by qPCR, and we found that F3II-NM showed higher levels of transcript than F3II-TP. In addition, F3II NM showed lower chemo sensibility to cisplatin and in an orthotopic model of BALB/c mice, presented a longer latency time and lower tumor growth rate in comparison to F3II-TP. Taking all these together, F3II NM present a dormant phenotype and F3II TP a proliferative one. Finally, analysis and comparison of the cell lines using LFQ-Mass spectrometry revealed 256 differentially expressed proteins between the lines studied. Overexpressed genes of survival paths were identified in F3II-NM, such as DDX42, COX-2, ATGB4 involved in the inhibition of apoptosis, promotion of tumor recurrence via SOX-2 and therapy resistance, respectivelyTo sum up, we established a new interesting dormant tumor cell model, F3II NM that could help to a better understanding of the behavior and survival of dormant cells. Moreover, we identified interesting protein targets that encourage the possibility of design new drugs that point and eliminate dormant cells