Optimising intratumoural treatment of head and neck squamous cell carcinoma mouse models with EBC-46

Barnett, Catherine (2015). Optimising intratumoural treatment of head and neck squamous cell carcinoma mouse models with EBC-46 MPhil Thesis, School of Medicine, The University of Queensland. doi:10.14264/uql.2015.895

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Author Barnett, Catherine
Thesis Title Optimising intratumoural treatment of head and neck squamous cell carcinoma mouse models with EBC-46
School, Centre or Institute School of Medicine
Institution The University of Queensland
DOI 10.14264/uql.2015.895
Publication date 2015-09-28
Thesis type MPhil Thesis
Supervisor Benedict Panizza
Total pages 116
Language eng
Subjects 1112 Oncology and Carcinogenesis
Formatted abstract
The five-year survival rate for patients with head and neck squamous cell carcinoma (HNSCC) has remained at ~50% for the past 30 years despite advances in treatment. EBC-46 is a novel diterpene ester developed by QBiotics Pty Ltd that induces HNSCC cell death in vitro.

The aims of this study were to identify a human HNSCC xenograft that responds poorly to intratumoural injection of EBC-46, improve efficacy of EBC-46 treatment by altering different administration parameters, and confirm the molecular mechanism of HNSCC cell death in vivo following intratumoural treatment with EBC-46.

Subcutaneous xenografts of HNSCC cell lines were grown in BALB/c Foxn1nu and NOD/SCID mice and treated with intratumoural injection of 30 μg EBC-46 or a control solution. A difficult cell line was identified and administration parameters were adjusted in an attempt to overcome treatment resistance. Treated tumours were stained for endothelial cell, macrophage and neutrophil markers. In vitro cytotoxic assays were employed to further investigate the mechanism by which EBC-46 works.

A tongue SCC cell line (SCC-15) was identified as the most resistant cell line. A single 30 μg bolus in 40% propylene glycol was the most efficacious administration. Immunohistological staining of treated tumours in BALB/c Foxn1nu mice identified tumour vessel disruption, red cell influx, and recruitment of macrophages and neutrophils following treatment with EBC-46. At 24 hr after treatment most of the cells present at the treated site were neutrophils. In contrast, no neutrophil infiltration was present in treated tumours at 24 hr in NOD/SCID mice. EBC-46 at doses comparable to the concentration administered in vivo rapidly killed tumour cells by necrosis in vitro. Simultaneous treatment with the pan- PKC inhibitor BIS-1 did not completely prevent EBC-46’s action. Dual fluorescence labelling observed under the confocal microscope revealed that mitochondrial potential was lost before uptake of propidium iodide.

In conclusion, high dose EBC-46 killed tumour cells in vitro by necrosis, associated with loss of mitochondrial potential. The picture is more complex in vivo. Disruption of tumour vasculature seen in tissue sections indicated a process of hemorrhagic necrosis. The difference in efficacy between BALB/c Foxn1nu and NOD/SCID mouse models indicated a requirement for neutrophils in the host and merits further investigation such as ablation of neutrophils in BALB/c Foxn1nu. These results provide new insight into the mechanism of action of EBC-46 and extend the number of HNSCC models further supporting the use of EBC-46 is a suitable agent for progression to human clinical trials in HNSCC.
Keyword Head and neck squamous cell carcinoma (HNSCC)
Protein kinase C
Diterpene ester
Intratumoural injection

Document type: Thesis
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Created: Sat, 12 Sep 2015, 10:53:05 EST by Ms Catherine Barnett on behalf of Scholarly Communication and Digitisation Service