Microscopy-based phenotypic monitoring of MDA-MB-231 spheroids allows the evaluation of phenotype-directed therapy

• Published in: Experimental cell research 2023-04, Vol.425 (2), p.113527-113527, Article 113527
• Main Authors: Mahmoud, Loay, Cougnoux, Antony, Bekiari, Christina, Araceli Ruiz de Castroviejo Teba, Paloma, El Marrahi, Anissa, Panneau, Guilhem, Gsell, Louise, Hausser, Jean
• Format: Article
• Language: English
• Subjects: Cell Line, Tumor; Cell phenotypes; Chemotherapies; Everolimus - therapeutic use; Female; Humans; Medicin och hälsovetenskap; Microscopy; Neoplasm Recurrence, Local; Phenotype; Spheroids; Triple negative breast cancer; Triple Negative Breast Neoplasms - drug therapy; Triple Negative Breast Neoplasms - genetics; Tumor heterogeneity
• ISSN: 0014-4827; 1090-2422; 1090-2422
• DOI: 10.1016/j.yexcr.2023.113527

Breast cancer (BC) is the most commonly diagnosed cancer among women. Prognosis has improved over the years, to a large extent, owing to personalized therapy informed by molecular profiling of hormone receptors. However, there is a need for new therapeutic approaches for a subgroup of BCs lacking molecular markers, the Triple Negative Breast Cancer (TNBC) subgroup. TNBC is the most aggressive type of BC, lacks an effective standard of care, shows high levels of resistance and relapse is often inevitable. High resistance to therapy has been hypothesized to be associated with high intratumoral phenotypic heterogeneity. To characterize and treat this phenotypic heterogeneity, we optimized a whole-mount staining and image analysis protocol for three-dimensions (3D) spheroids. Applying this protocol to TNBC spheroids located in the outer region of the spheroid the cells with selected phenotypes: dividing, migrating, and high mitochondrial mass phenotypes. To evaluate the relevance of phenotype-based targeting these cell populations were targeted with Paclitaxel, Trametinib, and Everolimus, respectively, in a dose-dependent manner. Single agents cannot specifically target all phenotypes at the same time. Therefore, we combined drugs that should target independent phenotype. With this rationale we observed that combining Trametinib and Everolimus achieves the highest cytotoxicity at lower doses from all the tested combinations. These findings suggest a rational approach to design treatments can be evaluated in spheroids prior to pre-clinical models and potentially reduce adverse effects. •Develop a whole mount staining, clearing, and scanning protocol that can image single cells and phenotypic markers.•3D single-cell spatial profiling of spheroid phenotypic heterogeneity.•Cells with dividing, migrating, high mitochondrial mass phenotypes are targeted by Paclitaxel, Trametinib, and Everolimus, respectively.•A combination of agents targeting multiple cell phenotypes simultaneously achieves higher cytotoxicity.