Background: The aim of this study was to present the first domestic animal trial of nanosecond electroporation with chemotherapy and CO₂ laser surgery. Methods: sixteen-year-old domestic cat with diagnosed melanoma on oral cavity was the case used in the study. Firstly, CO₂ laser surgery was used for the removal of most of tumor mass. Then nanoelectrochemotherapy with bleomycin was applied including appropriate margin of healthy tissue. A 15 ± 4 kV/cm × 15 ns pulsing protocol was employed with a total of 800 ± 100 pulses. Only one session of nanoelectrochemotherapy (nanoECT) was performed. Results and conclusions: during the next two weeks areas of focal necrosis were replaced by the granulation tissue. Complete wound healing was observed four weeks after initial treatment. After 15 weeks post treatment, no local recurrence was apparent.

Telomeres are repetitive DNA sequences at the ends of chromosomes that protect against genomic instability and prevent unwanted DNA damage responses. In most somatic cells, telomeres progressively shorten with each division, limiting cellular lifespan. However, cancer cells bypass this limitation by activating telomerase or the alternative lengthening of telomeres, enabling unchecked proliferation and tumor progression. This review examines the molecular mechanisms underlying telomere maintenance and their intricate relationship with DNA repair pathways. We discuss how telomere-associated proteins regulate genomic stability and explore therapeutic strategies targeting telomerase and alternative lengthening of telomeres. Challenges such as resistance mechanisms and off-target effects are also considered, highlighting the need for precision approaches in telomere-based cancer therapies.

The phenomenon known as bipolar cancellation is observed when biphasic nanosecond electric field pulses are used, which results in reduced electroporation efficiency when compared to unipolar pulses of the same parameters. Basically, the negative phase of the bipolar pulse diminishes the effect of the positive phase. Our study aimed to investigate how bipolar cancellation affects Ca²⁺ electrochemotherapy and cellular response under varying electric field intensities and pulse durations (3–7 kV/cm, 100, 300, and 500 ns bipolar 1 MHz repetition frequency pulse bursts, n = 100). As a reference, standard microsecond range parametric protocols were used (100 µs × 8 pulses). We have shown that the cancellation effect is extremely strong when the pulses are closely spaced (1 MHz frequency), which results in a lack of cell membrane permeabilization and consequent failure of electrochemotherapy in vitro. To validate the observations, we have performed a pilot in vivo study where we compared the efficacy of monophasic (5 kV/cm × ↑500 ns × 100) and biphasic sequences (5 kV/cm × ↑500 ns + ↓500 ns × 100) delivered at 1 MHz frequency in the context of Ca²⁺ electrochemotherapy (B16-F10 cell line, C57BL/6 mice, n = 24). Mice treated with bipolar pulses did not exhibit prolonged survival when compared to the untreated control (tumor-bearing mice); therefore, the bipolar cancellation phenomenon was also occurrent in vivo, significantly impairing electrochemotherapy. At the same time, the efficacy of monophasic nanosecond pulses was comparable to 1.4 kV/cm × 100 µs × 8 pulses sequence, resulting in tumor reduction following the treatment and prolonged survival of the animals.

Electrochemotherapy (ECT) with bleomycin has been effectively used in recent years to treat various skin tumors. Microsecond electric pulses significantly improve bleomycin (BLM) delivery and its anticancer potential. Up to now, we can determine electric field distribution in the targeted tissue, however, the distribution of the injected drug is still not well known. In this study, we propose the combination of indocyanine green (ICG) with bleomycin as a practical approach for ECT, enabling drug distribution control and detection. Normal skeletal muscle (L6) and fibrosarcoma (WEHI-164) cells were used for the viability evaluation by MTT assay after 24 and 72 h. Cells were exposed to the ESOPE protocol alone and in combination with drugs. Additionally, visualization of the uptake of ICG and ICG + BLM supported by electroporation was performed by confocal microscopy. The mast cell tumor (MCTs) was diagnosed in the feline case. The mixture of ICG + BLM was injected into the tumor, and ECT was performed under near-infrared fluorescence imaging (NIRF). The obtained results indicate the safety of the used procedure in vitro and in vivo. ICG does not affect ECT protocols in vitro. No significant cell viability decrease was noted only in the case of WEHI-164 cells post-ECT. Moreover, it does not adversely affect the procedure; in the case of in vivo surgery, it helps to control the drug distribution before and after ECT and identify the sentinel lymph node.