Eleni Zivla

Background/Objectives: Esophageal squamous cell carcinoma (ESCC) is an aggressive malignancy with poor prognosis and limited treatment options, highlighting the need for reliable preclinical models for translational research. Although xenograft models are widely used in cancer biology, systematic characterization of subcutaneous ESCC models under defined implantation conditions remains limited. The objective of this study was to systematically compare implantation conditions and define histopathological and growth-related parameters that enable reproducible tumor establishment while maintaining a practical experimental window for therapeutic studies. Methods: We established and characterized a subcutaneous ESCC xenograft model using human ESCC cell line KYSE-30 implanted into immunodeficient J:NU mice. Tumor growth dynamics were compared under three implantation conditions: cells suspended in phosphate-buffered saline (PBS), Matrigel (Corning, basement membrane matrix), or Matrigel diluted with PBS. Results: Tumors generated from Matrigel-embedded cells exhibited rapid growth, reaching substantial size within approximately one week after implantation, which limited their utility for controlled therapeutic studies. In contrast, tumors generated from cells suspended in Matrigel + PBS demonstrated slower but consistent expansion, providing a more suitable and reproducible experimental window for therapeutic intervention. Cells injected in PBS alone showed inefficient tumor establishment. Histopathologic and immunohistochemical analyses revealed matrix-dependent differences in necrosis, stromal organization, and proliferative activity, while confirming epithelial tumor identity and baseline vascularization. Conclusions: Collectively, these findings define a reproducible KYSE-30 subcutaneous xenograft model and identify Matrigel dilution with PBS as an optimal implantation strategy that balances reliable tumor engraftment with controlled tumor growth suitable for therapeutic studies.

Gelonin is a ribosome-inactivating protein with extreme intracellular toxicity but poor permeation into cells. Targeted disruption of cell membranes to facilitate gelonin entry is explored for cancer and tissue ablation. We demonstrate a hundreds- to thousands-fold enhancement of gelonin cytotoxicity by pulsed electric fields in the T24, U-87, and CT26 cell lines. The effective gelonin concentration to kill 50% of cells (EC₅₀) after electroporation ranged from 50 was unattainable even at the highest gelonin concentration of 1000 nM, which reduced cell survival by only 5–15%. For isoeffective electroporation treatments using 300 ns, 9 µs, and 100 µs pulses, longer pulses were more efficient at lowering gelonin EC₅₀. Increasing the electric field strength of 8, 100 µs pulses from 0.65 to 1.25 kV/cm reduced gelonin EC₅₀ from 128 nM to 0.72 nM. Conversely, the presence of 100 nM gelonin enabled a more than 20-fold reduction in the number of pulses required for equivalent cell killing. Pulsed electric field-mediated delivery of gelonin shows promise for hyperplasia ablation at concentrations sufficiently low to minimize or avoid systemic toxicity.