Browsing by Author "Coutermarsh-Ott, Sheryl Lynn"

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    Investigating the ablative and immunomodulatory effects of high frequency irreversible electroporation on osteosarcoma in-vitro
    Patwardhan, Manali Nitin (Virginia Tech, 2024-05-23)
    Osteosarcoma (OS) is the most common primary bone tumor with an annual incidence rate of 3-4 individuals per million particularly affecting children and young adults. The 5-year survival rate stands at 60-80% with the current standard of care for human OS patients who do not have metastatic disease at presentation, but this drops to 20% for patients with metastatic disease which frequently occurs in the lungs. OS is much more common in canines, with metastasis being the major contributor to mortality, the same as in humans. Metastatic OS warrants novel treatment strategies to improve prognosis and survival. High-frequency irreversible electroporation (H-FIRE) is a promising, non-thermal, minimally invasive technique that induces cell death by applying pulsed electric fields in targeted regions, potentially triggering an anti-tumor immune response that could also target and prevent metastases. Such a dual functionality of H-FIRE is uniquely suited to treat pulmonary metastatic OS. The goal of this thesis was to study the ablative and immunomodulatory effects of H-FIRE on OS in-vitro with the overall hypothesis that H-FIRE completely ablates OS cells, induces the release of damage-associated molecular patterns (DAMPs), and promotes pro-inflammatory immune activating signatures in macrophages and T cells. Using an in-vitro model, my master's thesis focused on 1) Determining the electric field strength that completely ablates OS cells 2) Evaluating the immunomodulatory effects of H-FIRE by co-culturing H-FIRE treated OS cells with macrophages and T cells separately. Our study has utilized murine, canine, and human OS and immune cells, thus demonstrating a unique cross-species approach, 3) Evaluating DAMPs (ATP, calreticulin, and HMGB1) post-H-FIRE ablation of human OS cells. Overall, our study showed that H-FIRE successfully ablated OS cells in-vitro, induced the release of DAMPs from treated cells, and promoted activation signatures in immune cells. This thesis provides foundational data for future investigations developing H-FIRE as an immunomodulatory strategy for treating metastatic OS.
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    Mechanical High-Intensity Focused Ultrasound (Histotripsy) in Dogs with Spontaneously Occurring Soft Tissue Sarcomas
    Yang, Ester (Virginia Tech, 2023-09-06)
    Background: Histotripsy is a non-thermal, non-invasive high-intensity focused ultrasound (HIFU) ablative technique that causes mechanical fragmentation of tissue, resulting in liquefied acellular debris with histologically clear demarcated boundaries between treated and non-treated tissues. The acellular debris may include tumor antigens with preserved immunogenicity and the potential to generate systemic immune response against tumor cells. Soft tissue sarcomas (STS) are a common form of cancer in dogs with biological behavior similar to STS in humans. Long-term tumor control requires complete removal with extensive surgical resection, which in many cases is not feasible. As a result, there is need for alternative therapies. Objectives: The primary objective of this study was to demonstrate safety and feasibility of histotripsy in a small animal model of spontaneous STS. The secondary objective was to characterize the impact of histotripsy on the immunologic response. Materials and methods: Pet dogs diagnosed with spontaneous STS were recruited. CT scan of the chest, abdomen, and the tumor was performed for staging and treatment planning. Pretreatment biopsies were obtained. Safety was monitored with physical examinations, owner reports, and CBC/serum biochemistry. Partial tumor ablation was performed using a 500 kHz prototype histotripsy system. A spherical treatment zone of up to 3 cm diameter in each tumor was treated with histotripsy according to the patient-specific treatment plan using 1-2 cycle pulses applied at a pulse repetition frequency (PRF) of 500 Hz. Anatomical ablation zones were evaluated with contrast CT at 1- and 4-days post-treatment, with tumor resection at 4-6 days post-treatment. Tumor microenvironment (TME) gene expression was evaluated with the Nanostring Canine IO panel, and the systemic immune response was evaluated using multiplex serum cytokine levels. Results: Ten dogs were recruited and treated. Tumor histologies included 3 grade III STS, 4 grade II STS, 2 grade I STS, and 1 malignant mesenchymoma. Six dogs were alive, three dogs were euthanized due to disease progression, and one dog was lost to follow up. Histotripsy-related complications were generally self-limiting, with only one patient having increased cutaneous injury score from 1 to 2 (scale 1-5) post-treatment, likely due to prefocal cavitation at the skin. No significant adverse events impacting patient outcome were noted in any of the patients. Visible histotripsy cavitation bubble clouds were seen on real-time ultrasound imaging in nine of ten treatments. Post-treatment histopathology indicated sharply defined regions of ablation that were clearly identifiable grossly and histologically in all samples. Treatment zones were characterized by loss of cell viability, hyalinization, and acute hemorrhage. Post-treatment contrast-enhanced CT images revealed clear, demarcated regions of histotripsy ablated tissue in seven of ten patients. Differential gene expression analysis identified 79 genes with at least 2-fold change following treatment. Genes associated with inflammation, immune cell migration, and immune cell interactions were the highest upregulated. Amongst the gene set analyses, the myeloid compartment gene sets obtained the highest significance score. There were no statistically significant differences between pre- and post-treatment cytokine concentrations for any of the analytes. Conclusions: Histotripsy can achieve safe and effective tumor ablation in dogs diagnosed with STS. Histotripsy induced pro-inflammatory changes within the tumor microenvironment. Histotripsy as an immunotherapeutic treatment option needs to be further investigated. Histotripsy has a potential to be a precise, non-invasive treatment for canine STS.