Human fallopian tube organoid models of high grade serous ovarian cancer initiation

Ovarian cancer is the sixth most lethal cancer in women. Out of all ovarian cancers – the high grade serous (HGSC) subtype is the most common and deadliest. This cancer originates from fallopian tube secretory cells and has several putative precursor lesions that have been described in the fallopian tube. These precursors show a spectrum of morphology from normal to abnormal and have been identified in patients with and without cancer. Genetically, HGSC is characterized by widespread genomic instability and ubiquitous TP53 mutation. A variety of model systems have attempted to interrogate how carcinogenesis occurs in the fallopian tube, but few examine the effect of TP53 mutation alone, and virtually all use other species to model human disease. In this thesis I describe the development, validation, and characterization of normal human fallopian tube derived organoids that have had exogenous mutations in TP53 introduced by the targeted cutting of the Cas9 nuclease. Analysis of these organoids was undertaken by live-cell imaging, histochemistry, and single-cell genomic sequencing. Relative to controls, mutant organoids showed normal morphology and growth but abnormal p53 immunohistochemistry. Single-cell sequencing further validated the strength of this model system and revealed comparable genomic stability between TP53 mutants and controls. These findings make sense in light of p53 signature precursor lesions’ normal morphology and low proliferative index; and support the idea that TP53 mutation alone is not sufficient to cause carcinogenesis in the fallopian tube. Reagents and theoretical approaches to interrogating the effects of TP53 missense mutations and homologous recombination deficiency pathway members have also been designed. This research supports genetic perturbation of patient-derived normal organoids as a robust and promising avenue of research.Medicine, Faculty ofMedical Genetics, Department ofGraduat