L.O. Oliveira; M. Telló; R.T. Oliveira; C.S. Cardoso; R. Zanella; H.R. Bittencourt; A. Cardona; O. Parise Jr.; A.C. Buzaid
Electrochemical Therapy (EChT) is a local treatment for cancer. EChT implies treatment of tumors with direct current, which flowing through electrodes produces an electrolytic process. Electrochemical products of electrolysis are formed. The toxicity of these products has destructive effects on cancer cells. Hitherto more than one hundred animals with different types of cancer were treated by our research group at the Veterinary Hospital of the Federal University of Rio Grande do Sul. In 2004 our group published a book (in Portuguese) titled: The Use of Electric Current in Cancer Treatment (Telló et al. 2004). EChT is delivered to tumors that present poor response to surgery and other conventional therapeutic methods, or when the owners are not willing to submit the animal to surgery or chemotherapy. Figure 1 shows an application of EChT in a melanoma of oral cavity in a dog.
In veterinary medicine, very few clinical trials with EChT were done worldwide. Concerning EChT in human medicine, an EChT project was initiated in China in 1987. More than 20,000 patients have been treated with EChT in China. Despite the Chinese clinical data, essential preclinical studies and clinical trials are still missing (von Euler et al. 2004). Additional studies on EChT are needed, especially related to mechanisms of anti-tumor effectiveness. Our research group in October of 2001 started EChT studies at the Veterinary Hospital of the Federal University of Rio Grande do Sul. EChT experiments were conducted in animals that acquired the malignant tumor during their life. The aim of our study was to standardize experiments in animals, to establish an adequate dose-response relationship and to determine the response that each different type of tumor presents after EChT. We started EChT evaluations treating canine mammary carcinomas. Nowadays, we are treating different skin types of cancer in pet dogs and cats.
Materials and Methods
EChT experiments using animals were approved by the Veterinary Hospital of the Federal University of Rio Grande do Sul. The animals accepted for EChT had recurrent or progressive cancer, had previously undergone another treatment, or the owner refused other standard treatment, such as surgery or systemic chemotherapy. Initially, mongrel dogs were used in our experiments. The dogs were provided by the Municipal Kennel and were housed individually at the Veterinary Hospital of Federal University of Rio Grande do Sul. Eight mammary glands of six mongrel dogs were treated by EChT. After treatment, three mongrel dogs were submitted to euthanasia. Necropsies were done in these dogs to evaluate the possibility of systemic effects caused by EChT.
Considering our results with mongrel dogs, we extended the new therapy to pet dogs with different types of skin and mammary tumors. Before treatment, the owners signed an informed consent. Equipment: An electronic device was used to apply variable levels of dc current into the target volume (tumor area). The equipment developed was designed to deliver a direct current of a pre-determined dose of "Coulombs" [Dose in Coulomb = Current (Ampere) x time (second)]. The device is galvanically isolated to uncouple the animal under treatment from the power supply. EChT procedure: All animals presenting cancer were clinically examined; the location and size of malignant tumors recorded, and the masses were submitted to fine needle aspiration biopsy for diagnosis. Thoracic radiographs were taken to verify the presence of metastasis. One or more sessions of EChT were designed, depending on the response of the animal. Animals were placed under general anesthesia for application of the direct current. For each treatment continuous current was applied in the range of 60 to 90 minutes, through two or more platinum electrodes (90% Platinum + 10% Rhodium) inserted 1 to 3 cm apart in the cancerous region. The positive electrodes (anode) were introduced at the center of the lesion and negative electrodes (cathode) 1 to 3 cm away. After treatment, the animals were evaluated clinically by visualization, palpation and measurement of the nodules, blood tests, cytological exams, and measurement of body temperature. Follow-up physical examinations were conducted before each session of treatment. Any event occurring during or after the treatment was recorded. Aspirative fine needle biopsies were done at 15 days interval after the treatment, for as long as necessary. The treatment was interrupted when cytological exams of the treated region showed absence of neoplastic cells. The evaluation included the study of the animal's response to therapy; comparing the initial volume of the nodules and tumor regression in relation to the current used (dose applied per cubic volume).
|Figure 1. EChT Session in a Dog, Showing the Electrodes Inserted in the Tumor.|
All dogs tolerated well the sessions of anesthesia and treatment, recoveries from anesthesia were uneventful. Treated regions showed an initial increase of the mass after treatment, due to edema. Subsequently foci of necrosis and ulceration of the neoplasm were developed and a gradual decrease of the tumor mass occurred. Tissue response to EChT includes inflammation and necrosis, with abundant macrophages in early response. After destruction of the neoplastic mass, second intention healing was initialized at the treated tumors. Destruction of tumor cells was confirmed by cytological exams at the end of the treatment. Tumors with different volumes were submitted to different values of charge (dose) imparted in it. A total of 27 dogs with mammary cancer were entered into a noncomparative study with the objective to evaluate the response of mammary tumors considering the rate Charge/ tumor volume. Figure 2 shows the obtained results as complete response (CR), partial response (PR), no responsive (NR) and in evaluation of treatment (ET). According to Figure 2, 20 C/cm3 is the smaller value of current that must be applied into a neoplastic mass to achieve CR of canine mammary tumors to EChT. Dogs with mammary tumor with positive lymph node had poor response to therapy and shorter survival time. EChT was successfully associated with surgery in two cases. One dog with positive lymph node had PR to EChT, treatment was associated with surgery but the dog developed metastasis 5 months after surgery. A variety of other types of neoplasia in 36 dogs and 20 cats were treated with EChT. This group was formed by a heterogeneous group of patients. Feline mammary cancer (3 patients), skin and subcutaneous tumors (7 basal cell carcinomas, 16 squamous cell carcinomas, 3 melanomas, 1 fibrosarcoma, 2 histiocytomas, 2 fibromas and 7 mast cell tumors), oral cavity tumors (1 melanoma and 2 carcinomas), 1 nasal mucinous carcinoma, 7 venereal transmissible tumors, 2 osteosarcoma and 2 mammary lymph node metastasis were treated. Many of these dogs and cats had late stage tumors (large tumors, metastasis, lymph node evolvement or recidivant) and treatment was palliative. Some dogs had more than one nodule treated. For this miscellaneous group of dogs, the type of the tumors, initial volume, dose of current used and response were evaluated through statistical analysis.
|Figure 2. EChT Mammary Tumors Response Considering the Rate Charge / Tumor Volume.|
EChT Statistical Analysis: EChT statistical analyses were done. The SPSS software version 11.5 was used at the statistical analysis. The total number of animals statistically evaluated (animals submitted to the EChT) was 79 (147 tumors), having different types of neoplastic disease. Basically, our interest is related with: a) the treatment response (CR) as function of applied dose (C/cm3) and b) the treatment response (CR) as function of tumor volume (cm3). For each indicated case of statistical interest [items a) and b)] we must specify the meaning of the terms Sensitivity and Specificity. So, in general, Sensitivity is the proportion of actual positives which are correctly identified as such. Specificity is the proportion of negatives which are correctly identified. The graph showing the Sensitivity and the Specificity results is called Receiver Operating Curve, or simply ROC curve. ROC curve allows identifying the parameter that maximizes the estimated probability of cure.
Statistical Analysis of EChT Response x Dose: Initially, we chose 10 C/cm3 as reference dose (software exigency). This value of reference dose is arbitrary. So, the Sensibility for the dose 10 C/cm3 is defined as the proportion of cases with dose equal or higher than 10 C/cm3 among all animals cured. The Specificity is the proportion of cases with dose smaller than 10 C/cm3 among all patients not cured. Figure 3 shows the ROC Curve. The Sensitivity is 97.6% and the Specificity is approximately 49.3%. We wish both "Sensitivity and Specificity" to be highest as possible. According ROC Curve the "ideal/optimum" dose is situated among 20.8 and 23.9 C/ cm3. Our research group evaluate statistically the EChT Response x Volume. The obtained results showed that for volumes smaller than 6.35 cm3 the Sensibility is approximately 73%. That means, among all cured tumors 73% shown volume smaller or equal than 6.35 cm3. About Specificity, our evaluation indicated that among all non-cured tumors, 70% had volume greater than cm3.
|Figure 3. ROC Curve (EChT Response x Dose).|
Discussion and Conclusions
Cytological exams of the tumors after treatment revealed development of inflammation and necrosis, with abundant macrophages in early response. Infiltration of macrophages and lymphocytes increased after treatment; this may be due to antigens released from damaged tumor cells following EChT. Antitumor effects of EChT are mediated partly via alterations in ion concentrations, pH state and cell cycle control leading to apoptosis (Tang et al. 2005). Secondary cell destruction was caused by necrosis in the cathodic region and apoptosis in the anodic region. Low pH and hypoxia seems to be important factors to the development of apoptosis in the tumors (von Euler et al. 2004). As EChT is a local treatment for cancer, it should be used in selected initial cases. In dogs with lymph node involvement, the responses were poorer. In cases of palliative treatment of large or metastatic tumors, the relief of pain due to local necrosis was considered a good result. According to the statistical analysis, the initial volume of the tumor and the dose of current used for treatment were important prognostic factors for cure. Our results indicates that EChT is clinically effective, had few local side effects, has low cost, improves the quality of life for the patients, has the ability to administer repeated treatments, is compatible with other cancer therapies and has high benefit/cost ratio for the patient.
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