Electrochemotherapy in Translational Medicine: From Veterinary Experience to Human Oncology.

Electrochemotherapy (ECT) is a tumor treatment that, through the application of electric pulses with suitable amplitude and waveforms, favors the systemic or local delivery of chemotherapy agents. This procedure significantly increases the permeability of cancer cells to anticancer drugs, making them more effective and allowing their use at lower doses with less morbidity for patients. Its use in veterinary oncology is consolidated and it is currently adopted as first-line treatment for different cancers with successful results. In human oncology, ECT use is mainly in the treatment of cutaneous tumors and for the palliation of cutaneous metastases of malignant tumors. A standard operating procedure has been formulated. Currently, several preclinical and phase I and II studies are under way involving various cancers in humans to better define the efficacy and tolerability of this therapy. This review summarizes the state of the art of ECT in veterinary and human oncology, describing the most significant results achieved to date.

Adjuvant electrochemotherapy after debulking in canine bone osteosarcoma infiltration.

Osteosarcoma is a bone cancer considered rare to humans, but common in dogs. Dogs and humans share genetic homology and environmental risk factors. Improving the treatment of osteosarcoma in dogs could also be relevant to improve procedures in humans. Traditional treatments of osteosarcoma involve surgery and chemotherapy. Such treatments are commonly aggressive and not possible for many patients. Electrochemotherapy emerges as a minimally invasive, effective, and safe treatment alternative. Electrochemotherapy combines applications of high-intensity electric fields during short periods with anti-cancer drugs to improve its medicine cytotoxicity. Analyzing the electric field distribution, as well as electric current density, are essential to electrochemotherapy success. This paper brings the first case of a canine osteosarcoma treatment performed with bleomycin and electrochemotherapy. We performed in silico studies with finite element method software to observe the electric field distribution. In silico experiments help to verify possibilities and limitations of treating bone destruction and macro or micro tumor infiltrations around the primary tumor mass. Results show that both needle or plate electrodes are feasible to remove the tumor even with invasion into the bone. Plate electrodes perform well in treating micro infiltrations when associated with conductive gel and direct contact between electrode and bone (without soft tissues). Needle electrodes are effective in treating tumor infiltration on external cortical bone. Multiple applications are needed to cover all cranium layers with sufficient electric field intensity. Electrochemotherapy protocol with needle or plate electrodes does not present sufficient electric current density capable of affecting brain tissue, even in cases of bone destruction.

Plasma Bleomycin Concentrations during Electrochemotherapeutic Treatment of Fibropapillomas in Green Turtles Chelonia mydas.

Fibropapillomatosis of sea turtles is traditionally treated with surgical debulking techniques that are often associated with prolonged healing and tumor recurrence. Electrochemotherapy was recently described for green turtles Chelonia mydas and can be an alternative to surgery and even general anesthesia. The objectives of this study were to replicate an electrochemotherapy protocol from a previous report and add plasma bleomycin analysis to the treatment. After bleomycin injection into similarly sized tumors of two green turtles and immediate electroporation at two time points, plasma bleomycin reached detectable concentrations that were considerably lower than those found in human studies. At 3 months posttherapy, no healing complications or recurrences were encountered and only scar tissue remained. This study adds further support that electrochemotherapy with bleomycin has the potential to be used as an effective alternative treatment for this complex disease.

Definition of novel electrochemotherapy parameters and validation of their in vitro and in vivo effectiveness.

Electrochemotherapy (ECT) is a cancer therapy that conjugates the administration of a chemotherapy agent to the delivery of permeabilizing pulses released singularly or as bursts. This approach results in higher number of anticancer molecules delivered to their biological targets, but is also associated to undesirable side effects such as pain and muscular contractions. A new electroporator delivering train of eight biphasic pulses at the voltage of 1,300 V/cm lasting 50 + 50 µsec each, with a frequency of 1 Hz, and with 10-µsec interpulse intervals (total treatment time: 870 µsec/cm(2) of treated area) was tested in vitro on the human lung cancer cell line A549 and in vivo, both in mice xenografts and privately owned rabbits with spontaneous tumors. The tumor cell line was treated with electroporation using the new parameters, that showed improved drug efficacy in causing cell death. Mice with chemoresistant xenografts were treated as well with either the new parameters and with a previous protocol, confirming the higher tolerability and efficacy of the novel parameters. Finally, a cohort of six pet rabbits with advanced skin neoplasms were enrolled in a compassionate trial using the new parameters in adjuvant fashion. In terms of efficacy, none of the rabbits experienced tumor recurrence, showing minimal discomfort during the ECT sessions. The data described, demonstrate that the new permeabilizing protocol adopting biphasic electric pulses displays a significant higher efficacy compared to previous ECT treatments and substantial reduction of the associated morbidity.

Electrochemotherapy for the treatment of fibropapillomas in Chelonia mydas.

Fibropapillomatosis continues to be an important cause of morbidity and mortality in sea turtles, particularly in Chelonia mydas. Turtles with this debilitating herpesvirus disease usually present with multiple, large, and ulcerated cutaneous masses that compromise both locomotion and feeding. There are very few available therapeutic strategies, with surgical excision being the most common. However, this surgical excision is associated with a high rate of local disease recurrence and secondary infections. Electrochemotherapy has been used for the treatment of epithelial neoplasm in several animal species. This technique is based on a combination of chemotherapy, usually with bleomycin or cisplatin, and electroporation. It consists of a series of short, high-voltage electric pulses that lead to increased membrane permeability and more efficient transport of antineoplastic drugs through the cellular membrane. Here, two C. mydas fibropapillomas were treated with a standard electrochemotherapy protocol using intralesional bleomycin sulfate injections followed by the application of electric pulses. Two sessions were performed, with a 33-day interval between sessions. Complete regression of lesions occurred without side effects or complications in each animal. There was no sign of local recurrence, even 1 yr after the end of treatment. Electrochemotherapy may be an effective therapeutic alternative for sea turtles with fibropapillomas.

A novel approach to ear pain in the horse: A case report.

Background: During electrochemotherapy (ECT), a chemotherapeutic drug is injected into the tumor and then an electroporation is provided. In horses, ear manipulation may be very painful, and combining a loco-regional technique with sedation might be a good option to avoid anesthesia-related risks. A two-injection-point block of the internal and external pinna and acoustic meatus was described in horse cadavers, and it permitted complete stain of all three branches of the great auricular nerve (GAN), internal auricular nerve branch (IAB), lateral auricular branch (LAB), and caudal auricular nerve (CAN), suggesting a lower risk of intra-parotid injection during the IAB and LAB block. Case Description: An 8-year-old Italian jumping gelding presented for ECT to treat a fibroblastic sarcoid in the left medial pinna. After intravenous sedation with acepromazine, romifidine, and butorphanol, a two-injection-point block was provided as previously described. The block of the GAN was blind, whereas an electrical nerve locator was used for the IAB, LAB, and CAN. A total of 12 ml of 0.5% ropivacaine was injected. The ECT was safely performed without any difficulties. The horse well tolerated the procedure and completely recovered 75 minutes after sedation. No complications were detected. Conclusion: The described approach seems feasible and suitable for the blockade of the sensory innervation of the equine ear in the case of ECT.

Description of outcome and adverse events in 21 cats with locally advanced nasal planum squamous cell carcinoma treated with electrochemotherapy.

OBJECTIVES: Squamous cell carcinoma (SCC) is the most common tumour in the nasal planum of cats. Surgery has traditionally been the treatment of choice but might not be feasible in locally advanced scenarios. Electrochemotherapy (ECT) has shown good control in superficial tumours, but there is a lack of robust information about efficacy in locally advanced cases. The aim of this study was to assess the safety and efficacy of ECT in the treatment of locally advanced stage nasal planum SCC in cats. METHODS: The clinical database of a veterinary referral hospital was searched retrospectively for cats diagnosed with a locally advanced nasal planum SCC (T3N0M0 or T4N0M0) that had received ECT. Local response, adverse events and outcome were documented. The data were evaluated by inferential statistics and correlations between response, recurrence, feline immunodeficiency virus/feline leukaemia virus status, number of treatments, voltage and severity of adverse events, with Kaplan-Meier curves and log-rank tests. Statistical significance was set at P <0.05. RESULTS: In total, 21 cats were enrolled over a 4-year period. Nineteen cats achieved a complete response (CR) and two cats a partial response (PR) for an overall response rate of 100%. Cats achieving a CR had a median disease-free interval of 182 days (range 128-327) and those with a PR had a median progression-free survival of 156.5 days (range 122-191). The median time to progression was not reached. The overall survival was 453 days for a median follow-up of 341 days (range 191-989). Of the cats, 62% had grade 3 or 4 toxicities, but no deaths due to the treatment were documented. Only voltage was correlated with longer survival (P = 0.001). CONCLUSIONS AND RELEVANCE: ECT appears to be an effective treatment for feline nasal planum SCC and could be considered a first-line therapy for locally advanced cases. Toxicities reported can be severe in the short term and these could be secondary to more invasive lesions and equipment used.

Electrochemotherapy in dogs and cats-A review.

Electrochemotherapy (ECT) is a treatment modality that combines the electroporation of cell membranes with chemotherapy to facilitate the transport of non-permeant molecules into cells. Several canine and feline studies have shown promising results, suggesting that ECT can be a valid adjuvant or alternative treatment option for some tumours. The objective of this paper is to provide a bibliographic review of the principles and applications of ECT in veterinary medicine and to compare to its use in human medicine.

Comparison of Three Chemotherapy Protocols With Electrochemotherapy for the Treatment of Feline Cutaneous Squamous Cell Carcinoma.

Electrochemotherapy (ECT) with intravenous (IV) and/or intratumoral (IT) bleomycin has shown considerable efficacy in the treatment of non-resectable feline cutaneous squamous cell carcinoma (cSCC), boasting response rates of up to 95%, but other chemotherapy protocols have not yet been investigated. The objective of this prospective multicentre study was to compare the overall response rate (ORR) and progression-free interval (PFI) between cats with cSCC treated with ECT using IT and IV carboplatin (IV + IT), IV carboplatin (IV) or IV bleomycin (IV). A total of 44 cats with unresectable cSCC across three centres were enrolled and treated with ECT using carboplatin IV + IT (n = 10), carboplatin IV (n = 11) or bleomycin IV (n = 23). Treatment response according to RECIST criteria was recorded at 2 and 4 weeks post-treatment, and patients were followed until disease progression and/or death. All three groups were comparable regarding age, sex, weight, and lesion size. Adverse events were generally mild, localised and similar between groups. ORRs were 90.0% (carboplatin IV + IT), 90.9% (carboplatin IV) and 95.6% (bleomycin IV) and were not significantly different (p = 0.79). Median PFI was not reached for carboplatin IV + IT or carboplatin IV and was 566 days for bleomycin IV, with no significant difference between the three groups (p = 0.81). This study suggests that ECT using IV or IV + IT carboplatin is a reasonable alternative therapeutic option for managing cSCC, and further studies are warranted to compare outcomes between treatment protocols.

Electrochemotherapy for the treatment of recurring aponeurotic fibromatosis in a dog.

This paper reports the clinical findings, histopathology, and clinical outcome of a rare case of aponeurotic fibromatosis in a dog. The dog was treated with 4 courses of electrochemotherapy using the drugs cisplatin and bleomycin. There was complete remission and the dog was still disease-free after 18 months.

Électrochimiothérapie pour le traitement de la fibromatose aponévrotique chez un chien. Cet article présente les résultats cliniques, l'histopathologie et le résultat clinique d'un rare cas de fibromatose aponévrotique chez un chien. Le chien a été traité avec 4 séries d'électrochimiothérapie utilisant les médicaments cisplatine et bléomycine. Il s'est produit une rémission complète et le chien était toujours exempt de maladie après 18 mois.(Traduit par Isabelle Vallières).

Treatment of spontaneous canine mast cell tumors by electrochemotherapy combined with IL-12 gene electrotransfer: Comparison of intratumoral and peritumoral application of IL-12.

The combined treatment of electrochemotherapy (ECT) and interleukin-12 (IL-12) gene electrotransfer (GET) has already been used in clinical studies in dogs to treat various histological types of spontaneous tumors. The results of these studies show that the treatment is safe and effective. However, in these clinical studies, the routes of administration of IL-12 GET were either intratumoral (i.t.) or peritumoral (peri.t.). Therefore, the objective of this clinical trial was to compare the two IL-12 GET routes of administration in combination with ECT and their contribution to the enhanced ECT response. Seventy-seven dogs with spontaneous mast cell tumors (MCTs) were divided into three groups: one treated with a combination of ECT + GET peri. t. (29 dogs), the second with the combination of ECT + GET i.t. (30 dogs), and the third with ECT alone (18 dogs). In addition, immunohistochemical studies of tumor samples before treatment and flow cytometry of peripheral blood mononuclear cells (PBMCs) before and after treatment were performed to determine any immunological aspects of the treatment. The results showed that local tumor control was significantly better in the ECT + GET i.t. group (p < 0.050) than in the ECT + GET peri.t. or ECT groups. In addition, disease-free interval (DFI) and progression-free survival (PFS) were significantly longer in the ECT + GET i.t. group than in the other two groups (p < 0.050). The data on local tumor response, DFI, and PFS were consistent with immunological tests, as we detected an increased percentage of antitumor immune cells in the blood after treatment in the ECT + GET i.t. group, which also indicated the induction of a systemic immune response. In addition, we did not observe any unwanted severe or long-lasting side effects. Finally, due to the more pronounced local response after ECT + GET i.t., we suggest that treatment response assessment should be performed at least two months after treatment, which meets the iRECIST criteria.

Canine squamous cell carcinoma: Electrochemotherapy association with surgery and correlation with overall survival.

Squamous cell carcinoma (SCC) is an important malignancy in dogs, due to its incidence and clinical presentation, which can be of locally aggressive single or multiple lesions with a metastatic potential. The objective of this investigation was to evaluate SCC response to treatment, anatomopathological and immunohistochemical characteristics, disease-free interval and overall survival time. 54 dogs with histopathologically diagnosed SCC were included in this study. Their mean age was 9.16 years with a range of 1-14 years. Of the 54 animals in the study, 34 (65.4%) had white skin and white fur coats. There was a significant correlation between fur coat colour and the development of tumours in areas of sun exposure (p = .001). Animals with tumours in areas of the body exposed to the sun had longer overall survival time than animals with tumours in areas not associated with sun exposure (p = .001). Surgery combined with electrochemotherapy (ECT) yielded a survival rate 32% higher than using a surgical approach alone (HR = 0.32, p = .038, IC = 0.11-0.94). ECT, with or without surgery, had an objective response rate of 90.9%. Local lymph node and/or distant site metastasis at diagnosis, or at some point during follow-up, occurred in 34.6% (18/52) of animals. Animals with tumours in sun exposed locations had more aggressive histopathological characteristics but had longer overall survival time. This is probably due to individualised therapeutic treatment with both surgery and ECT.

Preclinical models in electrochemotherapy: the role of veterinary patients.

Electrochemotherapy is a tumor treatment that adapts the systemic or local delivery of anticancer drugs by the application of permeabilizing electric pulses with appropriate amplitude and waveforms. This allows the use of lipophobic drugs, which frequently have a narrow therapeutic index, with a decreased morbidity for the patient, while maintaining appropriate anticancer efficacy. Electrochemotherapy is used in humans for the treatment of cutaneous neoplasms or the palliation of skin tumor metastases, and a standard operating procedure has been devised. In veterinary oncology, the electrochemotherapy approach is gaining popularity, becoming a first-line treatment in consideration of its high efficacy and low toxicity. This review summarizes the state of the art in veterinary oncology as a preclinical model.

Electrochemotherapy for the treatment of cutaneous solid tumors in equids: A retrospective study.

Background: Electrochemotherapy (ECT) promotes the increased uptake of antitumor agents through the administration of permeabilizing electric pulses, thus enhancing chemotherapy effectiveness. Aim: Our study aimed to describe the tolerability and efficacy of ECT alone or in association with surgery to manage solid neoplasms in equids. Methods: Medical records of equids with a diagnosis of malignant tumors treated with ECT alone or in combination with surgery were retrospectively evaluated. Each equid received local treatment within the tumors or the tumors' bed with cisplatin at the dose of 0.5 mg/cm2. Trains of permeabilizing biphasic electric pulses were then applied under spinal or general anesthesia. Results: Sixteen equids were enrolled in this study. There were nine melanoma cases, four fibrosarcoma, and three squamous cell carcinoma. Of those 16 equids, 7 received ECT for treatment of intraoperative local disease, while in 9 cases, ECT was the only treatment modality. The seven equids treated with the combination of ECT and surgery still have no evidence of disease at different times ranging from 9 to 60 months. The remaining nine had the following responses: two complete remissions, five partial responses, one stable disease, and one progressive disease. The treatment was well-tolerated, and local side effects were minimal. No systemic effects were documented. Conclusion: This retrospective study suggests that ECT may be beneficial for equids with solid neoplasms and could be a useful addition to the current therapeutic options considering its low cost, limited toxicity, and ease of administration.

Evaluation of electrochemotherapy in the management of apocrine gland anal sac adenocarcinomas in dogs: A retrospective study.

Background: Electrochemotherapy (ECT) combines the administration of anticancer drugs with the delivery of electric pulses, thus increasing the drug uptake through the cell membranes, resulting in increased efficacy. Aim: The aim of our study was to describe the tolerability and efficacy of ECT alone or in association with other treatment modalities for the management of apocrine gland anal sac adenocarcinoma (AGASAC). Methods: Medical records of dogs with a diagnosis of AGASAC that were treated with ECT alone or in combination with surgery/chemotherapy were retrospectively evaluated. Each dog received 20 mg/m2 of bleomycin intravenously. Based on the clinician's decision, the primary tumor or tumor bed was also infiltrated with cisplatin at the dose of 0.5 mg/cm2. Trains of permeabilizing biphasic electric pulses were then applied under general anesthesia. Results: Ten dogs were enrolled in the study. Of those 10 dogs, only one received ECT for treatment of microscopic local disease, while in six cases ECT was the only treatment modality. In three dogs, ECT was followed by systemic medical treatment. Six dogs (60%) had a partial response (PR), three dogs (30%) had stable disease, and one dog treated for microscopic disease did not show any sign of local relapse for 305 days after treatment, being still alive and in complete remission at the time of writing this article. The median time to progression was 303 days and the median survival time was 365 days. The treatment was well tolerated and local side effects were minimal. No systemic effects were documented. Conclusion: This preliminary study suggests that ECT may be beneficial for dogs with AGASAC and could be a useful addition to the current therapeutic options in consideration of its low cost, limited toxicity, and ease of administration.

Health-related quality of life in dogs treated with electrochemotherapy and/or interleukin-12 gene electrotransfer.

The aim of this study was to evaluate the owners' perception of health-related quality of life (HRQoL) of dogs after treatment with electrochemotherapy (ECT) alone or combined with interleukin-12 gene electrotransfer (IL-12 GET) and/or surgery. The owners of 44 dogs with histologically different tumours were offered the ¼Cancer Treatment Form« at least one month after treatment. The owners assessed their dogs' quality of life (QoL) after treatment as good (mean 7.4) (from 1-very poor to 10-excellent) and the general health compared with the initial diagnosis of cancer as improving (mean 3.9) (from 1-worse to 5-better). The assessment of the current QoL was better within the group of dogs treated with non-invasive treatment (ECT and/or IL-12 GET only), compared with those that received invasive treatment, where, in addition to ECT and/or IL-12 GET, surgery was performed (p < .05). The owners of dogs that achieved an objective response (OR) to the treatment assessed the QoL as significantly better compared with those whose dogs did not respond to the treatment (p < .05). The majority of the owners (86.4%) would opt for the therapy again, regardless of the financial costs. In conclusion, the results of this study demonstrate that the majority of the owners of dogs assessed their dogs' QoL as good and felt that it improved after the treatment, especially in dogs, treated with non-invasive treatment and in those that responded to the treatment. This supports further use of ECT and IL-12 GET as suitable methods for the treatment of selected tumours in veterinary medicine.

Outcome Following Curative-Intent Electrochemotherapy for Extramedullary Plasmocytoma in Dogs - Case Reports.

Plasma cell tumors can occur as solitary collections referred as extramedullary plasmocytoma (EMP). The present report describes four cases of EMP treated with a local nonthermal ablative approach. Four dogs were diagnosed with extramedullary plasmocytomas (EMP) in different body regions (oral cavity, digits, and lip). Since surgical excision was declined by the owners (maxillectomy; amputation or lip reconstruction), a curative-intent approach was indicated as solely treatment- electrochemotherapy (ECT). All the patients received ECT under general anesthesia using bleomycin intravenously (15,000 UI/m²) or cisplatin intratumorally (1mg/cm³). All dogs developed transitory ulceration and swelling one-week after procedure that completely healed within 30 days post-ECT. Complete remission was achieved in all cases and lasted for 515 (oral case), 695 (one digit), 90 (another digit case) and 240 (lip) days. These results suggested that ECT promoted remission in EMP cases being a possibility for local control in dogs affected by this disease.

Combination of bleomycin and cisplatin as adjuvant electrochemotherapy protocol for the treatment of incompletely excised feline injection-site sarcomas: A retrospective study.

Background: fFeline injection-site sarcomas (FISSs) are mesenchymal tumors that can occur in cats after injections of different medical agents and are easily prone to recurrence. Aim: The aims of this study were to report treatment outcomes for cats with feline injection-site sarcomas (FISSs) treated with both bleomycin and cisplatin, per adjuvant electrochemotherapy (ECT) protocol. Methods: The medical records of cats with a diagnosis of FISS that were treated with ECT using both bleomycin and cisplatin were retrospectively evaluated. A total of 27 cats were available for statistical evaluation of their response. The cats received intravenous 20 mg/m2 bleomycin, and the tumor bed and margins were infiltrated with cisplatin at the dose of 0.5 mg/cm2. Then, the trains of permeabilizing biphasic electric pulses lasting 50 + 50 µseconds each were delivered in bursts of 1,300 V/cm using caliper electrodes under sedation. A second session was performed 2 weeks later. Results: Side effects were limited to local inflammation in three cats. Three cats developed local tumor recurrence at days 180, 180, and 545 after surgery, two cats developed recurrence and metastases at 100 and 505 days after surgery, and two cats experienced distant metastases. A median time to recurrence could not be calculated as over 80% of the study population remained disease free or were censored due to death from other causes. Mean survival time was 985 days, and median cumulative survival for all cases was 1,000 days. Conclusion: When compared to historical controls, the results of this study demonstrate the superior rates of tumor-free survival and disease-free interval. This adjuvant therapy could be a useful addition to the current options for FISS in consideration of its efficacy, limited toxicity, and ease of administration.

Electrochemotherapy in treatment of canine oral non-tonsillar squamous cell carcinoma. A case series report.

Non-tonsillar squamous cell carcinoma (ntSCC) is a common and locally aggressive oral tumour in dogs. The treatments of choice are currently surgery and radiotherapy. Electrochemotherapy (ECT) is a local ablative anti-tumour technique using electric pulses to enhance the intracellular diffusion of cytotoxic drugs. The aim was to retrospectively evaluate the outcome of patients with oral ntSCC treated with ECT. Twelve dogs with ntSCC were retrospectively enrolled. ECT was combined with IV bleomycin (15 000 UI/m2 ) alone in 11 cases and post-surgery in 1. Parameters considered were: tumour site and size, electroporation parameters, response rate (complete remission [CR], partial remission [PR]), median survival time (MST), recurrence rate (RR), median disease-free interval (DFI) and treatment toxicity (6-point scale). Median tumour size was 1.65 cm (range 0.3-8.0 cm) and the response rate was 90.9% (10/11; 8 CR and 2 PR). Two dogs underwent a second ECT. MST for dogs dead with tumour (n = 2) was 110 days and for dogs dead without tumour (n = 3) was 831 days. Among five surviving dogs, one experienced tumour recurrence and four were in CR. Results from two dogs were analysed separately. Overall RR was 27.3%. DFI and MST for dogs with recurrence were 50 and 115 days, respectively. Treatment toxicity was very low. We noticed that all dogs with tumours smaller than 1-2 cm achieved CR without recurrence suggesting a favourable prognosis when using ECT. ECT for canine ntSCC could be considered a valid treatment option especially for smaller tumours, but a larger caseload would be needed to confirm this statement.