Genomic analysis of high grade serous ovarian cancer (HGSC) has revealed that up to 50% of HGSCs harbor a genomic alteration in a DNA damage repair (DDR) gene, mostly in the BRCA repair pathway. Functional profiling of DDR capacity of patient derived HGSC organoids has revealed that over 60% of HGSCs harbor defects in the stalled replication fork protection DDR pathway. Further analysis of these HGSC organoids reveals that in patients who receive neoadjuvant chemotherapy, a tumor which initially had a replication fork protection defect and was carboplatin sensitive can be induced by the neoadjuvant chemotherapy to undergo replication fork stabilization and become carboplatin resistant. Transcriptional analysis of the post-neoadjuvant fork stable organoids reveals that these cultures have undergone an epithelial-mesenchymal transition. Based on these results, the hypothesis is that defects in the stalled replication fork protection DDR pathway are a fundamental molecular defect in HGSC that when perturbed by treatment with neoadjuvant carboplatin can lead to loss of the defect at the molecular level, alterations in the overall state of the tumor cells, and changes in the way the tumor cells interact with the surrounding stroma and immune cells, all of which combine to create a broadly therapy resistant tumor microenvironment. The research challenge I will pursue is to dissect the evolving mechanisms by which tumor cells which originally harbor fork protection defects and are induced to lose the defect interact with the surrounding stroma and immune cells and how these changing interactions might be manipulated to prevent a deeply resistant tumor microenvironment. The work will utilize a novel HGSC organoid co-culture system which will be highly innovative for the field because it will allow real time assessment of the interactions between DDR defective or altered tumor cells and the surrounding normal cells over time and treatment. Approach: The goal of Aim 1 is to generate growth conditions for co-cultures of fork unstable platinum sensitive organoids with patient matched intra-tumoral stromal cells and all immune cells (T cells, B cells, NK cells, dendritic cells, macrophages, and neutrophils). The goal of Aim 2 will be to generate isogenic pairs of the organoids from Aim 1 that are fork stable and unstable and then study how the interaction of the tumor cells with the surrounding stromal cells changes in the organoid co-culture system as the tumor cells undergo selective fork stabilization. The goal of Aim 3 is to utilize the isogenic pairs of fork unstable and stable organoids from Aim 2 in the co-culture system and determine how the interaction of the tumor cells with the surrounding immune cells changes as the tumor cells undergo fork stabilization and how this may alter response to immuno-oncologic agents. This work will have major impact in HGSC because it will help understand the evolution of the tumor-normal cell interaction as the tumor cell fork protection defects are altered and stressed. This may allow for the development of rational combination therapies that simultaneously target the tumor cell defects and also prevent problematic tumor-normal cell interactions.
Up to 80% of high grade serous ovarian cancer (HGSC) patients succumb to their disease due to both lack of early detection and limited therapeutic options. Genomic analysis of these tumors has suggested that up to 50% may have difficulty in repairing damage to their DNA which might make them therapeutically sensitive to a variety of available drugs; and functional analyses of HGSC tumor cells has revealed that many tumor cells are defective in repairing specific damage generated when the cells duplicate their DNA. The work in this proposal will help to better understand how HGSC tumor cells with defects in repairing this type of damage interact with the surrounding normal cells, and how these interactions lead to sensitivity and resistance to certain therapies.
