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Treatments based on electroporation (EP) induce the formation of pores in cell membranes due to the application of pulsed electric fields. We present experimental evidence of the existence of pH fronts emerging from both electrodes during treatments based on tissue EP, for conditions found in many studies, and that these fronts are immediate and substantial. pH fronts are indirectly measured through the evanescence time (ET), defined as the time required for the tissue buffer to neutralize them. The ET was measured through a pH indicator imaged at a series of time intervals using a four-cluster hard fuzzy-c-means algorithm to segment pixels corresponding to the pH indicator at every frame. The ET was calculated as the time during which the number of pixels was 10% of those in the initial frame. While in EP-based treatments such as reversible (ECT) and irreversible electroporation (IRE) the ET is very short (though enough to cause minor injuries) due to electric pulse characteristics and biological buffers present in the tissue, in gene electrotransfer (GET), ET is much longer, enough to denaturate plasmids and produce cell damage. When any of the electric pulse parameters is doubled or tripled the ET grows and, remarkably, when any of the pulse parameters in GET is halved, the ET drops significantly. Reducing pH fronts has relevant implications for GET treatment efficiency, due to a substantial reduction of plasmid damage and cell loss.

Gene electrotransfer (GET) protocols, based on the introduction into the cells of genes encoding immunomodulatory molecules, constitute a safe and powerful strategy for inducing an immune response against cancer. But GET efficiency can be significantly affected by damage due to the products of electrolysis, in particular, pH fronts. To elucidate the role of pH fronts and damage in GET efficiency we present an analysis of the pH fronts-tissue natural buffer interaction through a theoretical model using the Nernst-Planck equations for ion transport assuming a tissue with a bicarbonate buffering system and its validation with experimental measurements. pH front-buffer interaction measurements unveil a remarkable behavior tuned by pulse length and frequency: during the ON pulse critical pH front trajectories (pH=8.5 or 5.5) jump forward, during the OFF pulse, they recede due to tissue natural buffer attenuation. Theory shows that they are intimately related to ion transport mode: during the ON pulse, ion transport is mainly governed by migration and trajectories jump forward in time; during the OFF pulse, migration ceases, ion transport is governed solely by diffusion and trajectories recede due to buffer attenuation. Experiments and theory show that regardless of the presence of buffer attenuation, pH fronts remain during several minutes in a non-physiological state after the treatment. These results suggest that regions enclosed by pH fronts trajectories (thus subjected to non-physiological pH values during a sufficiently long time) may be subjected to plasmid damage during a GET treatment. Ways to minimize this effect, thus optimizing GET efficiency are suggested. [ABSTRACT FROM AUTHOR]

Abstract: We have recently shown that in the classical electrochemical treatment (EChT) of tumors, i.e., the passage of a direct electric current through two electrodes inserted locally in the tumor tissue, from an initial uniform condition two pH fronts evolve, expanding towards each other, inducing extreme pH changes and tumor destruction, mainly by necrosis. Here we extend these results introducing a one-probe two-electrode device (OPTED) containing the cathode and the anode very close to each other (1mm). Experiments show that upon application of the OPTED-EChT, two half-spherical pH fronts, one basic and the other acid (from cathode and anode, respectively), expand towards the periphery configuring a distorted full sphere. Tracking of the pH front advance reveals a time scaling close to t 1/2, signature of a diffusion-controlled process. An analytic model presented allows the estimation of the time needed for total tumor destruction with a minimum compromise of healthy tissue. Main advantages of the OPTED are the insertion of one applicator rather than two or more (thus minimizing tissue intrusion, for instance, in the nervous system), the ability to reach tumors beyond capabilities of conventional surgery and the minimization of electric current circulation through the treated organ. We propose here this new design which could have significant implications in EChT optimal operative conditions, in particular, in the way in which the evolving pH spherical fronts can cover and destroy a cancer cell spherical casket. [Copyright &y& Elsevier]

Nasal cavity tumors are usually diagnosed late, when they already have infiltrated adjacent tissues thus requiring very aggressive treatments with serious side effects. Here we use electrochemotherapy (ECT), a well demonstrated treatment modality for superficial tumors.

Electrochemotherapy (ECT), a medical treatment widely used in human patients for tumor treatment, increases bleomycin toxicity by 1000 fold in the treated area with an objective response rate of around 80%. Despite its high response rate, there are still 20% of cases in which the patients are not responding. This could be ascribed to the fact that bleomycin, when administered systemically, is not reaching the whole tumor mass properly because of the characteristics of tumor vascularization, in which case local administration could cover areas that are unreachable by systemic administration.