Graduation Date

Summer 8-14-2020

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Cancer Research

First Advisor

Joyce C Solheim

Second Advisor

Jennifer Black

Third Advisor

Tatiyana Bronich

Fourth Advisor

Michael A Hollingsworth

Abstract

Solid tumors, such as pancreatic cancer, often result in dismally low survival outcomes for patients due to insufficient understanding of disease development and progression. Pancreatic cancer is the fourth leading cause of cancer-related deaths in the United States and although oncogenic drivers (such as KRAS mutation or loss of tumor suppressor p53) and stages of disease development have been studied, further understanding of pancreatic cancer development is greatly needed. Studies from our laboratory have identified novel and varied functions of amyloid precursor-like protein 2 (APLP2) in the development and progression of pancreatic cancer. These functions include promoting cancer cell migration, proliferation, and invasion. APLP2 has also been found to bind to MHC class I molecules, thus leading to their internalization, loss of surface expression on cancer cells, and possible functions in cancer cell immune evasion. We furthered our investigation into elucidating the role of APLP2 in pancreatic cancer by employing the genetically engineered KPC/KC mouse models and incorporated a conditional APLP2 knockout (KPCA). These genetically engineered mouse strains enabled us to investigate how APLP2 loss, in a pancreas-specific manner, modulates pancreatic cancer development and progression. Loss of APLP2 in the pancreas delayed tumor development, inhibited metastasis, and prolonged survival in KPCA mice. We derived cell lines from primary tumors collected from mice bearing wild type APLP2, heterozygous knockout of APLP2, or homozygous knockout of APLP2 and confirmed modulation in APLP2 expression at the protein at mRNA levels.

An additional hurdle in successfully treating solid tumors, such as neuroblastoma or pancreatic cancer, is the lack of efficacious, durable treatment agents that are capable of inducing a robust antitumor immune response. Neuroblastoma is the most common extracranial solid tumor diagnosed in children and patients with aggressive metastatic disease or refractory/relapsed neuroblastoma face dauntingly low survival prognoses. It has been reported, by our laboratory and others, that chemoattractant C-C motif chemokine ligand 21 (CCL21) is effective as an intratumoral therapy, able to deter tumor growth and induce an antitumor immune response. We hypothesized that utilizing CCL21 in a novel, slow-release alginate nanoformulation would provide prolonged release of CCL21, leading to a steady influx of immune cells into the tumor mass, delaying tumor growth, and halting disease progression in a manner superior to CCL21 alone. Initial studies resulted in the characterization of alginate-nanoformulated CCL21 in vitro, including rates of release. When injected intratumorally into mice bearing subcutaneous neuroblastoma lesions, alginate-nanoformulated CCL21 significantly prolonged survival and reduced the tumor growth rate compared to CCL21 alone, empty nanoparticles, or buffer control. Notably, complete tumor clearance and subsequent protection against tumor re-challenge was observed in 33% of nanoformulated CCL21-treated mice. Further analysis revealed superior intratumoral retention of nanoformulated CCL21 compared to free CCL21 at days 1 and 2 post treatment as determined via fluorescent labeling and tracking of CCL21. A general pattern of prolonged increases in antitumor cytokines and relatively lower levels of protumor cytokines was observed in nanoformulated CCL21-treated mice when compared to CCL21-treated or buffer-treated tumors. Overall, we report that nanoformulated CCL21 is an effective and novel treatment for neuroblastoma. In addition, this novel nanoformulation has potential for future development as a slow-release modality for other immunotherapies, as well as for the delivery of CCL21 when used in a combinatorial therapeutic approach.

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