Uncovering Novel Drivers of Mammary Tumorigenesis and Osteosarcoma Chemotherapy Resistance
2022-12
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Uncovering Novel Drivers of Mammary Tumorigenesis and Osteosarcoma Chemotherapy Resistance
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2022-12
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Breast cancer is one of the most common cancers globally, with nearly a quarter of a million new cases diagnosed every year. Although extensive study identified molecular and clinical subsets of the disease, the enormous heterogeneity in gene expression, genome arrangement, clinical behavior, and phenotype makes identifying driver genes and the cells of origin challenging. Known breast cancer driver gene mutations that have been identified fail to show absolute associations with these molecular subtypes, and it has been suggested that phenotype is influenced by factors like epigenetic alterations, gene copy number changes, and the cell of origin. The study presented here explored the contributions of these factors to breast cancer development. A Sleeping Beauty forward mutagenesis screen was performed using a known breast cancer driver gene mutation, Pik3ca^H1047R as a background initiating mutation. Mobilization of SB transposons accelerated mammary tumor development in both mouse mammary tumor virus (MMTV)-Cre and whey acidic protein (Wap1)-Cre lineage cells. Histopathology and RNA sequencing (RNA-seq) revealed estrogen receptor (ER) positive and negative tumor subsets with corresponding ER-dependent gene expression increases or decreases. Large-scale DNA and RNA sequencing of tumors and matched normal tissues identified common insertion sites of the SB transposon and distinguished early, truncal mutations from later-occurring progression mutations. The SB screen identified known breast cancer driver genes, such as Pten, along with novel driver genes. γ-catenin (Jup) was a top common insertion site in both DNA and RNA sequencing, and was functionally validated as a mammary oncogene. Osteosarcoma (OS) is the most common primary bone tumor and has a propensity to metastasize early and aggressively, leading to frequent treatment failure, particularly for patients with metastatic disease at diagnosis. Successful treatment of the metastatic OS disease state has eluded clinicians for decades and a better understanding of how OS metastases evade chemotherapy and immune cells is sorely needed to improve patient survival. The study presented here explores the mechanisms by which a component of the extracellular matrix, hyaluronan (HA), contributes to chemotherapy resistance in OS lung metastases.
A pilot study of induced OS lung metastases in an immunodeficient mouse model revealed that OS lung metastases are rich in HA, as are primary tumors in an SB-driven OS model developed previously in our laboratory and primary human tumors. Primary tumors are significantly enriched for HA compared to normal bone. We evaluated human OS cell lines and determined that they produce their own HA, with the highest HA production from highly metastatic OS or from immortalized osteoblasts. Moreover, treatment with chemotherapy induces expression of the HA synthase genes in OS cell lines. Systemic administration of a PEGylated hyaluronidase (PEGPH20) improved chemotherapy penetrance into lung metastases, decreased the number of actively dividing tumor cells, and trended toward inducing apoptosis. Critically, the combination of PEGPH20 with standard of care chemotherapy halved the number of metastases in the lungs compared to chemotherapy alone. This pilot study supports the hypothesis that targeting HA improves chemotherapy efficacy and highlights the need for further investigations in an immunocompetent mouse model to investigate whether immune cell infiltration into OS lung metastases improves with HA breakdown.
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University of Minnesota Ph.D. dissertation. December 2022. Major: Comparative and Molecular Biosciences. Advisor: David Largaespada. 1 computer file (PDF); xii, 344 pages.
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Pope, Emily. (2022). Uncovering Novel Drivers of Mammary Tumorigenesis and Osteosarcoma Chemotherapy Resistance. Retrieved from the University Digital Conservancy, https://hdl.handle.net/11299/259634.
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