Unraveling cytotoxic effects of jacaric acid using breast cancer cell lines as a model

Details

Supervisors

Larondelle, Yvan ; Rezsohazy, René

Faculty

Faculté des bioingénieurs

Degree label

Master [120] : bioingénieur en sciences agronomiques, à finalité spécialisée

Abstract

Cancer is a term used to define a broad group of diseases in which abnormal cells divide uncontrollably and can spread to other organs in the body. It stands as the second leading cause of death worldwide, with approximately ten million cancer-related deaths in 2020. Of these, breast cancer accounted for 685,000 deaths in women, making it the deadliest cancer in women. Breast cancer is divided into distinct subtypes based on their molecular phenotype, which is determined by the expression of specific receptors by the cancer cells. Among these subtypes, triple-negative breast cancer (TNBC) is the most aggressive, with a poor prognosis and high recurrence rates. Unfortunately, effective therapeutic options for TNBC patients are currently limited. However, recent investigations have revealed a potential vulnerability of TNBC cells to ferroptosis, a recently discovered form of non-apoptotic cell death characterized by the accumulation of lipid hydroperoxides. Interestingly, ferroptosis can be induced by supplementing cancer cells with highly peroxidable conjugated linolenic acids (CLnAs). Of particular interest, jacaric acid (JA), a CLnA, has demonstrated notable cytotoxicity against cancer cells, including TNBC cells. Therefore, investigating the underlying mechanisms of JA-induced ferroptosis holds great promise for the discovery of novel treatment strategies against cancer cells, particularly TNBC. To evaluate the cytotoxic effects of JA, three cell lines were studied: MCF7, which belongs to the luminal A subtype, and Hs578T and MDA-MB-468, which belong to the TNBC subtype. The experiments were performed using JA either alone or in combination with a second ferroptotic inducer, RAS-selective-lethal-3 (RSL3), known to induce ferroptotic cell death through enzymatic inhibition of the major antioxidant defense system, glutathione peroxidase 4 (GPX4). Cell viability assays clearly confirmed the dose-dependent cytotoxicity of JA and RSL3 in all three cell lines, although to different degrees. TNBC cells showed increased sensitivity. Furthermore, the cytotoxicity of JA was found to be synergistically enhanced when combined with RSL3. In addition, the effect of JA, either alone or in combination with RSL3, was evaluated in three-dimensional spheroids, which represent a more complex in vitro model with increased intercellular interactions. Although spheroids showed increased resistance compared to monolayer cells, the higher sensitivity of TNBC cells to ferroptosis was consistently observed. To elucidate the underlying causes of JA cytotoxicity, both alone and in combination with RSL3, and the differential sensitivity between cell lines, fatty acid profiles were analyzed in the three tested cell lines. JA was found to be incorporated into various lipid fractions, including phospholipids, which are known to be the lipid fraction subject to peroxidation, resulting in ferroptotic cell death. However, the cause of the difference in susceptibility between the cell lines remained elusive by the analysis of the fatty acid profile, suggesting that it may be due to factors such as the differential expression of some enzymes. This master's thesis highlights the potential of inducing ferroptosis by supplementing cancer cells with JA, either alone or in combination with RSL3, as a promising avenue for the development of novel therapeutic approaches against malignant tumors. However, at the end of the present work, numerous questions remain unanswered, requiring further investigation to understand the mechanisms governing JA-induced ferroptosis and the specific sensitivity exhibited by TNBC cells.