Detection of mutations in liquid biopsy of patients with breast cancer and its response to personalized treatment in organoid culture

Abstract

Breast cancer (BC) is the most prevalent type of tumor among women and has high mortality rate attributed to the failure in decteting this disease early and the absence of effective markers to estimate the risk of cancer progression and to predict the tumor response to chemotherapy and radiation therapy. In dogs, about 50% of breast tumors are diagnosed as malignant. In addition, canine BC shares many similarities with these tumors in humans, which makes them excellent models for the study of cancer biology and tests with therapeutic agents and for comparative purposes of mutational profiles. BC is characterized by high intratumoral heterogeneity, that is the coexistence of different clones in the same tumor. This process of clonal evolution occurs due to the sequential acquisition of mutations throughout the tumor establishment and that can culminate in drug resistance throughout the treatment, which demonstrates the need for a personalized therapeutic approach for each patient. Currently, studies are beginning to investigate the ways of early cancer diagnosis and to monitor treatment response by detecting circulating tumor cells (CTCs) and DNA fragments (ctDNAs) released by the primary tumor or its metastases in the blood. This innovative procedure is known as a liquid biopsy and is so called because it only requires the collection of blood or other body fluids, thus avoiding invasive procedures such as conventional tissue biopsies. The liquid biopsy allows to access tumor information that guides the patient prognosis and monitoring during the follow-up period. Thus, the objective of this study is to verify the presence of mutations by liquid biopsy as well as to qualify them in the primary tumor and to test the sensitivity to specific drugs in organoid culture. Liquid biopsies will be collected for ctDNA extraction and mutation panel analysis by Next Generation Sequencing (NGS), with a total of 98 mutations for humans and 88 for dogs related to the acquisition of chemoresistance. Primary culture cells from tumor fragments obtained by surgical excision will be cultured in organoids treated with different chemotherapeutics (initial tests will be performed with doxorubicin, trastuzumab, everolimus, cetuximab and erlotinib) to evaluate the specific response. In the end, the results of the liquid biopsy will be analyzed and its ability to detect mutations verified, and the relationship between the different mutations and the responsiveness of the tumor cells to the chemotherapeutic will be defined as an attempt to determine a personalized therapy for patients with BC.