Browsing by Author "Hay, Alayna N."
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- Ablative and Immunostimulatory Effects of Histotripsy Ablation in a Murine Osteosarcoma ModelHay, Alayna N.; Imran, Khan Mohammad; Hendricks-Wenger, Alissa; Gannon, Jessica M.; Sereno, Jacqueline; Simon, Alex; Lopez, Victor A.; Coutermarsh-Ott, Sheryl; Vlaisavljevich, Eli; Allen, Irving C.; Tuohy, Joanne L. (MDPI, 2023-10-09)Background: Osteosarcoma (OS) is the most frequently occurring malignant bone tumor in humans, primarily affecting children and adolescents. Significant advancements in treatment options for OS have not occurred in the last several decades, and the prognosis remains grim with only a 70% rate of 5-year survival. The objective of this study was to investigate the focused ultrasound technique of histotripsy as a novel, noninvasive treatment option for OS. Methods: We utilized a heterotopic OS murine model to establish the feasibility of ablating OS tumors with histotripsy in a preclinical setting. We investigated the local immune response within the tumor microenvironment (TME) via immune cell phenotyping and gene expression analysis. Findings: We established the feasibility of ablating heterotopic OS tumors with ablation characterized microscopically by loss of cellular architecture in targeted regions of tumors. We observed greater populations of macrophages and dendritic cells within treated tumors and the upregulation of immune activating genes 72 h after histotripsy ablation. Interpretation: This study was the first to investigate histotripsy ablation for OS in a preclinical murine model, with results suggesting local immunomodulation within the TME. Our results support the continued investigation of histotripsy as a novel noninvasive treatment option for OS patients to improve clinical outcomes and patient prognosis.
- Characterizing the Ablative Effects of Histotripsy for Osteosarcoma: In Vivo Study in DogsRuger, Lauren N.; Hay, Alayna N.; Vickers, Elliana R.; Coutermarsh-Ott, Sheryl; Gannon, Jessica M.; Covell, Hannah S.; Daniel, Gregory B.; Laeseke, Paul F.; Ziemlewicz, Timothy J.; Kierski, Katharine R.; Ciepluch, Brittany J.; Vlaisavljevich, Eli; Tuohy, Joanne L. (MDPI, 2023-01-25)Osteosarcoma (OS) is a malignant bone tumor treated by limb amputation or limb salvage surgeries and chemotherapy. Histotripsy is a non-thermal, non-invasive focused ultrasound therapy using controlled acoustic cavitation to mechanically disintegrate tissue. Recent ex vivo and in vivo pilot studies have demonstrated the ability of histotripsy for ablating OS but were limited in scope. This study expands on these initial findings to more fully characterize the effects of histotripsy for bone tumors, particularly in tumors with different compositions. A prototype 500 kHz histotripsy system was used to treat ten dogs with suspected OS at an intermediate treatment dose of 1000 pulses per location. One day after histotripsy, treated tumors were resected via limb amputation, and radiologic and histopathologic analyses were conducted to determine the effects of histotripsy for each patient. The results of this study demonstrated that histotripsy ablation is safe and feasible in canine patients with spontaneous OS, while offering new insights into the characteristics of the achieved ablation zone. More extensive tissue destruction was observed after histotripsy compared to that in previous reports, and radiographic changes in tumor size and contrast uptake following histotripsy were reported for the first time. Overall, this study significantly expands our understanding of histotripsy bone tumor ablation and informs future studies for this application.
- Engrafting Horse Immune Cells into Mouse Hosts for the Study of the Acute Equine Immune ResponsesLeeth, Caroline M.; Adkins, Janie; Hay, Alayna N.; Bogers, Sophie Helen; Potter, Ashley; Witonsky, Sharon G.; Zhu, Jing (MDPI, 2021-10-14)Immunological studies in the horse are frequently hampered by lack of environmental control, complicated study design, and ethical concerns when performing high risk studies. The purpose of the current study was to investigate the utility of a xenograft model for studying acute equine immune responses. Immunocompromised non obese diabetic (NOD). sudden combined immunodeficiency (scid).gamma-/- (NSG) mice were engrafted with either equine peripheral blood lymphocytes (PBLs) or equine bone marrow to determine an optimal protocol for equine lymphocyte engraftment. We found that both PBL and bone marrow grafts populated the host mice successfully. Bone marrow transplants were technically more challenging and required further processing to retard graft versus host disease. Graft vs host disease was apparent at 28 days post-PBL transfer and 56 days post-bone marrow transfer. The results of these studies support the use of mouse hosts to study acute equine immune responses and that different engraftment techniques can be used depending on the experimental design.
- Equine Protozoal Myeloencephalitis: investigating immunopathogenesis and treatment efficacy in mouse models and clinically affected horsesHay, Alayna N. (Virginia Tech, 2020-01-09)Equine protozoal myeloencephalitis (EPM), predominantly caused by the protozoa Saracocystis neurona, is a common neurologic disease in horses from North America. Equine exposure to the parasite occurs frequently as the protozoa is excreted in opossum (Didelphis virginiana) feces and contaminates the horse's environment. However, clinical neurologic disease only emerges in a small fraction of exposed horses. The seemingly protective immune response that develops in some exposed horses but not all is not fully defined. Previous reports utilizing horse EPM models and immune compromised mouse models, which develop disease simulating EPM after infection with S. neurona, have reported a role of T-lymphocytes and the cytokine interferon gamma, in disease protection. As part of this dissertation, the role of T-lymphocytes and IFNγ was further elucidated. It was determined that IFNγ production is essential for T-lymphocytes to offer protection against S. neurona induced encephalitis, in immune compromised mice. Another factor hindering prognosis of EPM affected horses is treatment failure. The efficacy of the antiprotozoal decoquinate, was tested and found to be ineffective at preventing S. neurona encephalitis, in immune compromised mice. However, the antiprotozoal, diclazuril, was found to be effective at preventing S. neurona encephalitis in immunocompromised mice but once treatment was terminated, infection persisted, and neurologic disease developed. In-situ methods were employed to extensively evaluate the immunopathology of spinal cord tissue samples collected from EPM affected horses. A novel in-situ hybridization technique was successfully utilized to identify S. neurona in tissue samples collected from horses with EPM. This technique will create new opportunities for investigating the immunopathology of EPM. Overall results from the studies conducted in this dissertation suggest that IFNγ production from T lymphocytes is essential for them to offer protection against S. neurona encephalitis. Additionally, further insight on FDA approved and non-FDA approved treatment options for S. neurona infection was gained through the use of the B6Ifnγ -/- mouse model. Collectively, these studies expanded on the knowledge of an understudied equine neurologic disease.
- High intensity focused ultrasound for the treatment of solid tumors: a pilot study in canine cancer patientsCarroll, Jennifer; Coutermarsh-Ott, Sheryl; Klahn, Shawna L.; Tuohy, Joanne L.; Barry, Sabrina L.; Allen, Irving C.; Hay, Alayna N.; Ruth, Jeffrey; Dervisis, Nikolaos G. (Taylor & Francis, 2022-01)Purpose: To investigate the safety, feasibility, and outcomes of High-Intensity Focused Ultrasound (HIFU) for the treatment of solid tumors in a spontaneous canine cancer model. Methods: Dogs diagnosed with subcutaneous solid tumors were recruited, staged and pretreatment biopsies were obtained. A single HIFU treatment was delivered to result in partial tumor ablation using a commercially available HIFU unit. Tumors were resected 3-6 days post HIFU and samples obtained for histopathology and immunohistochemistry. Total RNA was isolated from paired pre and post treated FFPE tumor samples, and quantitative gene expression analysis was performed using the nCounter Canine IO Panel. Results: A total of 20 dogs diagnosed with solid tumors were recruited and treated in the study. Tumors treated included Soft Tissue Sarcoma (n = 15), Mast Cell Tumor (n = 3), Osteosarcoma (n = 1), and Thyroid Carcinoma (n = 1). HIFU was well tolerated with only 1 dog experiencing a clinically significant adverse event. Pathology confirmed the presence of complete tissue ablation at the HIFU targeted site and immunohistochemistry indicated immune cell infiltration at the treated/untreated tumor border. Quantitative gene expression analysis indicated that 28 genes associated with T-cell activation were differentially expressed post-HIFU. Conclusions: HIFU appears to be safe and feasible for the treatment of subcutaneous canine solid tumors, resulting in ablation of the targeted tissue. HIFU induced immunostimulatory changes, highlighting the canine cancer patient as an attractive model for studying the effects of focal ablation therapies on the tumor microenvironment.
- Histotripsy Ablation of Bone Tumors: Feasibility Study in Excised Canine Osteosarcoma TumorsArnold, Lauren; Hendricks-Wenger, Alissa; Coutermarsh-Ott, Sheryl; Gannon, Jessica; Hay, Alayna N.; Dervisis, Nikolaos G.; Klahn, Shawna L.; Allen, Irving C.; Tuohy, Joanne L.; Vlaisavljevich, Eli (Elsevier, 2021-12)Osteosarcoma (OS) is a primary bone tumor affecting both dogs and humans. Histotripsy is a non-thermal, non-invasive focused ultrasound method using controlled acoustic cavitation to mechanically disintegrate tissue. In this study, we investigated the feasibility of treating primary OS tumors with histotripsy using a 500-kHz transducer on excised canine OS samples harvested after surgery at the Veterinary Teaching Hospital at Virginia Tech. Samples were embedded in gelatin tissue phantoms and treated with the 500-kHz histotripsy system using one- or two-cycle pulses at a pulse repetition frequency of 250 Hz and a dosage of 4000 pulses/point. Separate experiments also assessed histotripsy effects on normal canine bone and nerve using the same pulsing parameters. After treatment, histopathological evaluation of the samples was completed. To determine the feasibility of treating OS through intact skin/soft tissue, additional histotripsy experiments assessed OS with overlying tissues. Generation of bubble clouds was achieved at the focus in all tumor samples at peak negative pressures of 26.2 ± 4.5 MPa. Histopathology revealed effective cell ablation in treated areas for OS tumors, with no evidence of cell death or tissue damage in normal tissues. Treatment through tissue/skin resulted in generation of well-confined bubble clouds and ablation zones inside OS tumors. Results illustrate the feasibility of treating OS tumors with histotripsy. CORRIGENDUM: The authors regret that errors were present in the above article. The legend for Figure 5 on page 3441 should read “Fig. 5. Normal, healthy, non-neoplastic bone was excised from amputated canine limbs and subjected to histology. No histological differences were noted between untreated (a: magnification 4 x, b: magnification 40 x) and treated samples (c: magnification 4 x, d: magnification 40 x).” Also, the final section heading on page 3439 should read “Histotripsy ablation of ex vivo bone and nerve specimens.” Finally, the reference after the last complete sentence on page 3437 is incomplete and should read “Focal pressure waveforms for the 500-kHz transducer were measured using a custom-built fiberoptic hydrophone (FOPH) in degassed water at the focal point of each transducer (Parsons et al. 2006).” The authors would like to apologise for any inconvenience caused.
- Histotripsy Ablation of Spontaneously Occurring Canine Bone TumorsRuger, Lauren N.; Hay, Alayna N.; Gannon, Jessica M.; Sheppard, Hannah O.; Coutermarsh-Ott, Sheryl L.; Daniel, Gregory B.; Kierski, Katharine R.; Ciepluch, Brittany J.; Vlaisavljevich, Eli; Tuohy, Joanne L. (IEEE, 2023-01)Objective: Osteosarcoma (OS) is a devastating primary bone tumor in dogs and humans with limited non-surgical treatment options. As the first completely non-invasive and non-thermal ablation technique, histotripsy has the potential to significantly improve the standard of care for patients with primary bone tumors. Introduction: Standard of care treatment for primary appendicular OS involves surgical resection via either limb amputation or limb-salvage surgery for suitable candidates. Biological similarities between canine and human OS make the dog an informative comparative oncology research model to advance treatment options for primary OS. Evaluating histotripsy for ablating spontaneous canine primary OS will build a foundation upon which histotripsy can be translated clinically into a standard of care therapy for canine and human OS. Methods: Five dogs with suspected spontaneous OS were treated with a 500 kHz histotripsy system guided by real-time ultrasound image guidance. Spherical ablation volumes within each tumor (1.25-3 cm in diameter) were treated with single cycle histotripsy pulses applied at a pulse repetition frequency of 500 Hz and a dose of 500 pulses/point. Results: Tumor ablation was successfully identified grossly and histologically within the targeted treatment regions of all subjects. Histotripsy treatments were well-tolerated amongst all patients with no significant clinical adverse effects. Conclusion & Significance: Histotripsy safely and effectively ablated the targeted treatment volumes in all subjects, demonstrating its potential to serve as a non-invasive treatment modality for primary bone tumors.
- Investigation of High Frequency Irreversible Electroporation for Canine Spontaneous Primary Lung Tumor AblationHay, Alayna N.; Aycock, Kenneth N.; Lorenzo, Melvin F.; David, Kailee; Coutermarsh-Ott, Sheryl; Salameh, Zaid; Campelo, Sabrina N.; Arroyo, Julio P.; Ciepluch, Brittany; Daniel, Gregory; Davalos, Rafael V.; Tuohy, Joanne (MDPI, 2024-09-07)In this study, the feasibility of treating canine primary lung tumors with high-frequency irreversible electroporation (H-FIRE) was investigated as a novel lung cancer treatment option. H-FIRE is a minimally invasive tissue ablation modality that delivers bipolar pulsed electric fields to targeted cells, generating nanopores in cell membranes and rendering targeted cells nonviable. In the current study, canine patients (n = 5) with primary lung tumors underwent H-FIRE treatment with an applied voltage of 2250 V using a 2-5-2 µs H-FIRE waveform to achieve partial tumor ablation prior to the surgical resection of the primary tumor. Surgically resected tumor samples were evaluated histologically for tumor ablation, and with immunohistochemical (IHC) staining to identify cell death (activated caspase-3) and macrophages (IBA-1, CD206, and iNOS). Changes in immunity and inflammatory gene signatures were also evaluated in tumor samples. H-FIRE ablation was evident by the microscopic observation of discrete foci of acute hemorrhage and necrosis, and in a subset of tumors (n = 2), we observed a greater intensity of cleaved caspase-3 staining in tumor cells within treated tumor regions compared to adjacent untreated tumor tissue. At the study evaluation timepoint of 2 h post H-FIRE, we observed differential gene expression changes in the genes IDO1, IL6, TNF, CD209, and FOXP3 in treated tumor regions relative to paired untreated tumor regions. Additionally, we preliminarily evaluated the technical feasibility of delivering H-FIRE percutaneously under CT guidance to canine lung tumor patients (n = 2). Overall, H-FIRE treatment was well tolerated with no adverse clinical events, and our results suggest H-FIRE potentially altered the tumor immune microenvironment.