Cutaneous Papilloma and Squamous Cell Carcinoma Therapy Utilizing Nanosecond Pulsed Electric Fields (nsPEF)

Nanosecond pulsed electric fields (nsPEF) induce apoptotic pathways in human cancer cells. The potential therapeutic effective of nsPEF has been reported in cell lines and in xenograft animal tumor model. The present study investigated the ability of nsPEF to cause cancer cell death in vivo using carcinogen-induced animal tumor model, and the pulse duration of nsPEF was only 7 and 14 nano second (ns). An nsPEF generator as a prototype medical device was used in our studies, which is capable of delivering 7-30 nanosecond pulses at various programmable amplitudes and frequencies. Seven cutaneous squamous cell carcinoma cell lines and five other types of cancer cell lines were used to detect the effect of nsPEF in vitro. Rate of cell death in these 12 different cancer cell lines was dependent on nsPEF voltage and pulse number. To examine the effect of nsPEF in vivo, carcinogen-induced cutaneous papillomas and squamous cell carcinomas in mice were exposed to nsPEF with three pulse numbers (50, 200, and 400 pulses), two nominal electric fields (40 KV/cm and 31 KV/cm), and two pulse durations (7 ns and 14 ns). Carcinogen-induced cutaneous papillomas and squamous carcinomas were eliminated efficiently using one treatment of nsPEF with 14 ns duration pulses (33/39 = 85%), and all remaining lesions were eliminated after a 2nd treatment (6/39 = 15%). 13.5% of carcinogen-induced tumors (5 of 37) were eliminated using 7 ns duration pulses after one treatment of nsPEF. Associated with tumor lysis, expression of the anti-apoptotic proteins Bcl-xl and Bcl-2 were markedly reduced and apoptosis increased (TUNEL assay) after nsPEF treatment. nsPEF efficiently causes cell death in vitro and removes papillomas and squamous cell carcinoma in vivo from skin of mice. nsPEF has the therapeutic potential to remove human squamous carcinoma

Cutaneous papilloma and squamous cell carcinoma therapy utilizing nanosecond pulsed electric fields (nsPEF).

Nanosecond pulsed electric fields (nsPEF) induce apoptotic pathways in human cancer cells. The potential therapeutic effective of nsPEF has been reported in cell lines and in xenograft animal tumor model. The present study investigated the ability of nsPEF to cause cancer cell death in vivo using carcinogen-induced animal tumor model, and the pulse duration of nsPEF was only 7 and 14 nano second (ns). An nsPEF generator as a prototype medical device was used in our studies, which is capable of delivering 7-30 nanosecond pulses at various programmable amplitudes and frequencies. Seven cutaneous squamous cell carcinoma cell lines and five other types of cancer cell lines were used to detect the effect of nsPEF in vitro. Rate of cell death in these 12 different cancer cell lines was dependent on nsPEF voltage and pulse number. To examine the effect of nsPEF in vivo, carcinogen-induced cutaneous papillomas and squamous cell carcinomas in mice were exposed to nsPEF with three pulse numbers (50, 200, and 400 pulses), two nominal electric fields (40 KV/cm and 31 KV/cm), and two pulse durations (7 ns and 14 ns). Carcinogen-induced cutaneous papillomas and squamous carcinomas were eliminated efficiently using one treatment of nsPEF with 14 ns duration pulses (33/39 = 85%), and all remaining lesions were eliminated after a 2nd treatment (6/39 = 15%). 13.5% of carcinogen-induced tumors (5 of 37) were eliminated using 7 ns duration pulses after one treatment of nsPEF. Associated with tumor lysis, expression of the anti-apoptotic proteins Bcl-xl and Bcl-2 were markedly reduced and apoptosis increased (TUNEL assay) after nsPEF treatment. nsPEF efficiently causes cell death in vitro and removes papillomas and squamous cell carcinoma in vivo from skin of mice. nsPEF has the therapeutic potential to remove human squamous carcinoma

SNP array analysis of tyrosine kinase inhibitor-resistant chronic myeloid leukemia identifies heterogeneous secondary genomic alterations

To elucidate whether tyrosine kinase inhibitor (TKI) resistance in chronic myeloid leukemia is associated with characteristic genomic alterations, we analyzed DNA samples from 45 TKI-resistant chronic myeloid leukemia patients with 250K single nucleotide polymorphism arrays. From 20 patients, matched serial samples of pretreatment and TKI resistance time points were available. Eleven of the 45 TKI-resistant patients had mutations of BCR-ABL1, including 2 T315I mutations. Besides known TKI resistance-associated genomic lesions, such as duplication of the BCR-ABL1 gene (n = 8) and trisomy 8 (n = 3), recurrent submicroscopic alterations, including acquired uniparental disomy, were detectable on chromosomes 1, 8, 9, 17, 19, and 22. On chromosome 22, newly acquired and recurrent deletions of the IGLC1 locus were detected in 3 patients, who had previously presented with lymphoid or myeloid blast crisis. This may support a hypothesis of TKI-induced selection of subclones differentiating into immature B-cell progenitors as a mechanism of disease progression and evasion of TKI sensitivity

Treatment of cell lines <i>in vitro</i> with nsPEF.

<p>nsPEF exposure of Jurkat cells (Human T cell leukemia, 2×10⁶/mL) to 50, 100 or 200 pulses of 30 ns duration, at 50 Hz and a varying peak voltage (Panel<b> A,</b> left), and at a fixed peak voltage resulting in a field of 30 kV/cm and a varying number of pulses (Panel <b>A,</b> right). nsPEF exposure of 11 solid tumor cell lines (2×10⁶/mL in 1 mm cuvette): glioblastoma multiforme (GBM) cells (U118, T98G, U373); colon cancer cells ( HCT116); skin cancer cells (SRB-1, SRB-12, SCC-13, Colo-16, HaCaT); as well as early transformed cells [AK (actinic keratosis), KA (keratoacanthoma)]. Cells were exposed to 100 or 200 pulses of 30 ns duration, at 50 Hz and a varying peak voltage, and at a fixed peak voltage resulting in a field of 30 kV/cm and a varying number of pulses (Panels B. C. D. E). The effect of nsPEF exposure on cell viability is represented by the percentage of viable cells remaining after exposure calculated as a fraction of viable control cells, not exposed but handled similarly. Trypan blue was used to measure viable cells after nsPEF exposure at one hour. The dashed lines indicate either the pulse number or peak voltage associated with a 50% (ED50) reduction in cell viability. Results are values obtained from three experiments under identical conditions (mean + SD).</p

Effect of nanosecond pulse electric fields on squamous carcinomas <i>in vivo</i>.

<p>Effect of nanosecond pulse electric fields on squamous carcinomas <i>in vivo</i>.</p

Sensitivity of different cell types to cell death (ED50) caused by nsPEF (30k V/cm).

<p>Sensitivity of different cell types to cell death (ED50) caused by nsPEF (30k V/cm).</p

Required pulse number at 50, 100, or 200 pulses to kill 50% cells (ED50) of various cancer types.

<p>Required pulse number at 50, 100, or 200 pulses to kill 50% cells (ED50) of various cancer types.</p

Expression of Bcl-xl <i>in vitro</i> and <i>in vivo</i>.

<p>Western blot analyzed the Bcl-xl expression post nsPEF exposure. Glioblastoma multiforme cell line (U118) was exposed to varying numbers of pulses <i>in vitro</i> [20 ns duration, 50 Hz, and 30 kV/cm) in 1 mm cuvette]. 1 hour post nsPEF exposure, Bcl-xl expression was measured (Panel <b>A</b>). Squamous carcinoma cell line (SRB-12) was injected subcutaneously into immunocompromised mice. After one week, established tumors were exposed to nsPEF 200 pulses of 14 ns duration, 50 Hz and 40 kV/cm nominal electric field. One hour later, cells were harvested and western blot was performed to measure Bcl-xl expression (Panel <b>B</b>). Three induced tumors were either untreated or treated with nsPEF (40 kV/cm nominal electric field, 50 Hz, 1.75 mm tip 200p). Protein was extracted from these tumors after 1 and 3 hours, followed by Western blotted and probed with antibody to Bcl-xl (Panel <b>C</b>). GAPDH was used as loading control.</p

Visual changes over time following nsPEF exposure of induced papillomas and squamous cell carcinomas.

<p>Solid circle surround squamous cell carcinomas and dash lines surround normal skin. Appearance at 24 hours (<b>d1</b>) post-exposure is shown following nsPEF at 200 and 400 pulses of 14 ns duration, 50 Hz and 40 kV/cm peak nominal electric field and at 200 pulses of 14ns duration, 50 Hz and 31 kV/cm peak nominal electric field. Additional images are shown on alternate days up to one week (<b>d3</b>, <b>d5</b> and <b>d7</b>) for nsPEF exposure of 200 pulses of 14ns duration, 50 Hz and 40 kV/cm nominal electric field.</p