WO2017143269A1

WO2017143269A1 - Methods, systems, and apparatuses for tissue ablation using electrolysis and cryosurgical techniques

Info

Publication number: WO2017143269A1
Application number: PCT/US2017/018486
Authority: WO
Inventors: Boris Rubinsky; Franco Lugnani; Paul Mikus; Enric GÜNTHER; Liel RUBINSKY
Original assignee: Boris Rubinsky; Franco Lugnani; Paul Mikus; Günther Enric; Rubinsky Liel
Priority date: 2016-02-19
Filing date: 2017-02-17
Publication date: 2017-08-24

Abstract

Examples of cryoelectrolysis probes are described which are able to both freeze tissue and deliver electrolysis products. The probe may be metal and may serve as the anode or the cathode used to provide electrolysis products. Methods of performing cryoelectrolysis include freezing tissue before, during, or after delivering electrolysis to the same tissue.

Description

METHODS, SYSTEMS, AND APPARATUSES FO TISSUE ABLATION USING ELECTROLYSIS AM) CRYOSURGICAL TECHNIQUES

CROSS-REFERENCE TO RELATED APPLICATIONS

[061] This application claims the benefit of ^'U.S. Provisional Application No.

62/297,698 filed February 1.9, 2016, and U.S. Provisional Application No. 62/375,841 , filed August 16, 201 , each of which is hereby incorporated by reference, in its entirety, for any purpose.

TECHNICAL FIELD

[0021 The present disclosure relates to the combination of electrolysis arid cryosurgery to ablate tissue.

BACKGROUND

J063] Electrolysis has been used for minimally invasive tissue ablation since the early 1800^;s, The process of electrolysis occurs at the electrode surfaces of electrodes in an ionic conducting media. Ne chemicai species axe generated at the interface of the electrodes as a result of the electric-potexrtial-dri ven transfer between electrons and ions or atoms. The various chemical species produced near the electrodes diffuse away in a process driven by differences in electrochemical potential In physiological solutions, these chemical reactions also yield changes i H, resulting in art acidic region near the anode and a basic region near the cathode. Tissue ablation is driven by two factors; development of a cytotoxic environment due to local changes in H, as well as the presence of some of the new chemical species formed during electrolysis. Electrolysis is a chemical ablation mechanism and the extent of ablation is a function of the concentration of the chemical species and the exposure time to these chemicals. The total amount of electrolytic products generated during electrolysis is related to the charge delivered during the process, and therefore the total charge is used as a quantitative measure for the extent of electrolysis.

[004| Over the last two decades, substantial research has been conducted on tissue ablation by electrolysis, including cell and animal experiments, mathematical modeling, and clinical work, in the contemporary literature, electrolytic ablation is sometimes referred to as Electro-Chemical Therapy (EChT). Electrolytic ablation has been shown to exhibit several unique attributes. First, due to the chemical nature of the ablation process, die diffusion of chemical species in the tissue and the rate of chemical reactions dominate the time scale of the procedure. Second, the chemical products at the anode differ from those formed at the cathode, thus resulting in distinct mechanisms of ablation. Finally, electro-osmotic forces drive the migration of water from the anode to the cathode, further magnifying the contrasting physiological effects at the electrode surfaces. From an operational standpoint, electrolysis may use very low voltages and currents, providing advantages relative to other ablation techniques, e.g. reduced instrumentation complexity, it is, however, a lengthy procedure, controlled by the process of diffusion and the need for high concentrations of electrolytica!ly-produced ablative chemical species.

|00Sf Cryosurgery and cryosurgical techniques generally refer to the use of extreme cold to induce cell death, e.g. minimally invasive tissue ablation methods that employ freezing to induce cell death. Frozen tissue thaws naturally or through external warming. One of the advantages of cryosurgery over other minimally invasive tissue ablation, methods ma be the ability to image the extent of tissue freezing, with conventional medical imaging, non-irtvasivety and in real time. While medical imaging produces high quality details on the extent of freezing, the extent of cell death does not necessarily coincide with the extent of freezing, in particular on the outer rim of the frozen lesion and in the vicinity of large blood vessels.

[006) Electroporation generally harnesses an electricity-induced phenomenon; it differs from electrolysis by employing different set of biophysical principles. The bioelectric phenomenon of electroporation is characterized by the permeabilizatioo of the eel! membrane through the application of very brief, high-magnitude electric field pulses. The extent of membrane permeabilization is a function of the electric field strength. Electroporation can be used to produce reversible pores in the lipid bi!ayer, alio wins for the introduction of molecules such as aenes and druas into cells. The eiectric parameters, however, can be designed to produce irreversible defects, resulting in a cell membrane that does not reseal after the field is removed. Reversible electroporation techniques have been combined with anticancer drugs such as bleomycin to target cancerous tissues for successful clinical use in the field of electfoehemotherapv; Reversible electroporation. is also used in- other .medical and biotechnoIogfcaS applications, including bin not limited to transfection and. introduction of molecules such as siRNA or CRISPR-related compounds into cells that survive the peitneabilization process, Electroporation specifically targets the eel! membrane through the application of an electric field that develops instantaneously.

[007] The information included in this Background section of the specification, including any references cited herein and any description or discussion thereof, is included for technical reference purposes only and is not to he regarded as subject matter by which the scope of the disclosure as defined in the claims is to be bound.

SUMMARY

[008] The technology disclosed herein relates to the combination of electrolysis and cryosurgery. The combination may be used to ablate tissue. Methods of combining electrolysis and cryosurgery, as well as systems and apparatuses for combining electrolysis and cryosurgery are disclosed.

[009] In some embodiments, a method of tissue ablation includes performing electrolysis such that electrolysis products are applied to a tissue to be treated and also performing a. cryosurgical technique to a portion of the same tissue to freeze the tissue. The cryosurgery is performed during a time period that overlaps with the performance of the electrolysis.

[010] In some embodiments, the electrolysis is performed by applying an electrode to the tissue. The electrode may also be a cryosurgery probe, In some embodiments, the electrolysis products propagate outward from the electrode into the tissue.

[0.11] In some embodiments, electrolysis occurs in frozen tissiie. The electrolysis products may propagate through frozen tissue,

[012| In some embodiments, the electrolysis products include hypochlorous acid.

[013] In some embodiments, a second electrolysis is performed subsequent to performing a first electrolysis and a first cryosurgical technique.

[014] In some embodiments, the tissue is f ozen at -21. 1 °C or warmer.

[0151 In some embodiments, the cryosurgical technique is commenced after the commencement of electrolysis and terminates prior to the termination of electrolysis, lire rate of electrolysis may decrease or stop as freezing increases. (^'0I.6J In some enibc lInients, an area of tissue exposed to electrolysis products may be the same size as, smaller than, or larger than an area of frozen tissue.

[017 J In some embodiments, an apparatus for tissue ablation includes a device for producing electrolysis products as well as for freezing tissue, at least one power supply operably connected to the device, and at least one controller operably connected to the device,

|^'018| In some embodiments, the device for producing electrolysis products as well as for freezing tissue is an electrode. The device may be an anode.

|019| In some embodiments, the device for producing electrolysis products as well as for freezing tissue is a cryosurgery probe.

[0201 in some embodiments, the device for producing electrolysis products as well as for freezing tissue is constructed in part of stainless steel ,

[02 ij This Summary is provided to introduce a selection of concepts in a simplified form that are further described below in the Detailed Description. This Summary is not intended to identify key featores or essential features of the claimed subject matter, nor is it intended to be used to limit the scope of the claimed subject matter. A more extensive presentation of featores, derails, utilities,, and advantages of the present disclosure as defined in the claims is provided in the following written description of various embodiments and illustrated in the accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

| 22| FIG. 1 is a schematic illustration of a omttiniodalify electrolysis system according to one embodiment.

[023J FIG. 2A is a flow chart illustrating a method of cryoeiectrolysis according to one embodiment, FIG, 2B is a flow chart illustrating a method of cryoeiectrolysis according to another embodiment.

[024} FIG. 3 is a schematic diagram of a cryoeiectrolysis probe according to one embodiment,

[025) FIG. 4 shows a macroscopic comparison between lesions produced by cryosurgery (FIG, 4A) and those produced by cryoeiectrolysis (FIG. 4B).

[0261 FIG. 5 shows a macroscopic comparison between lesions produced by electrolysis (FIG. 5A) and those produce by cryoeiectrolysis (FIG. SB), (Ό27) FIG. 6 shows a microscopic comparison of the margins of the lesions of FIG. 4, {^'028 J FIGS. 7A-7F depict an experimental setup for performing freezing simultaneously wit electrolysis and FIGS. 7F-7H. depict results of the same.

029J FIGS. 8A-8B depict an experimental setup for perforaiing freezing simultaneously with electrolysis according to one implementation and FIGS. 8C-8E depict results of the same.

j^'030| FIG. 9 A depicts an experimental setup for performing freezing simultaneously with electrolysis according to another implementation and FIGS. 9B-9D depict results of the same.

{^'03 J. J FIGS. I0A-10B depict an experimental setup for performing freezing simultaneously with electrolysis according to another implementation .and FIGS. 10C- iOE depict results of the same. All images are to the same scale,

(032{ FIGS. 1 1 A-10I depict results of performing freezing simultaneously with electrolysis according to another implementation. All images are to the same scale.

{033} FIGS. 12A-.10E present data from the experi ment of FIG. 1 1.

DETAILED DESCRIPTIO

{034{ Certain details are set forth below to provide a sufficient understanding of embodiments of the disclosure. However, it will be clear to one skilled in the art that embodiments of the disclosure may be practiced without these particular details. Moreover, the particular embodiments of the present disclosure described herein are provided by way of example and should not be used to limit the scope of the disclosure to these particular embodiments. In other instances, well-known materials, components, processes, controller components, software, circuitry, timing diagrams, and/or anatomy have not been described or shown in detail in order to avoid unnecessarily obscuring the embodiments,

|0351 The technology described herein is generally understood as devices, systems, and methods fo combining cryosurgery wit electrolysis, with or without eleciroporation. hi some examples, the combination may result in a clinical effect greater than eac one alone and of a more desired clinical benefit with reduced or eliminated unwanted side effects. ) Electrolysis generally refers to process of inducing a chemical reaction that involves passing a direct current through, an ionic solution via two electrodes. Electrolysis may facilitate the removal and/or addition of electrons from atoms and/or ions, which may lead to the formation of new products. For example, by passing a DC current through a saline solution (NaCl and ¾()), hypoehSorous acid (H^'CIO) may be formed. The products formed may be based, at least in part, on the ionic composition of the solution. pH, and/or materials from which the electrodes are constructed. The amount of electrolysis products formed may be based at least in part o the magnitude of the current and/or the duration for which the current is applied. The current may be generated by coupling a power source to the electrodes. Examples of power sources may include, but are not limited to, electrical networks, batteries, a computer (e.g., coupled via a USB cable), a solar cell, external inductors like induction or gradient coils similar to ones used in MR!, for example, and combinations thereof.

) Electrolysis products may be used for sterilization and or ablation of tissue. Some electrolysis products, such as hypochiorous acid, may be toxic to cells and/or organisms. Hypochiorous acid and other electrolysis products may be introduced to a tissue by pouring a solution including the electrolysis products on. a targeted tissue or injecting the solution into tissue. Electrolysis products may degrade rapidly over time, which may reduce their effectiveness. In tissue ablation applications, it may be desirable to produce the electrolysis products at the target tissue site.

) Electrodes for performing electrolysis may be placed in contact with a tissue or a solution in contact with the tissue. When the electrodes are activated, e.g. through an electric current, electrolysis products ma form at the interface between the electrodes and the tissue or a solution in contact with the tissue. The electrolysis products may diffuse from the electrode/tissue interface outward into the tissue through diffusion, forced convection, and/or electro-osmosis. The amounts of electrolytic products generated may depend on the electric charge transferred from the electrodes to the tissue (and/or solution), the composition of the solution, and the construction of the electrodes. Cells within the target site may be ablated as a function of the concentration of the electrolytic products and or the time of exposure to the products. Some applications of tissue ablation by electrolysis may demand high concentrations of electrolytes and/or long times of exposure, which may produce undesired consequences.

[039J Electrical interactions can produce a variety of effects including the induction of

Joule beating, various modalities of electroporation, and electrolysis. Any one or more of these effects may be used for tissue ablation in som examples. The electrical parameters may be varied such that one of the effects becomes the dominant effect. These effects from the basis of the methods of tissue ablation bv thermal heatinu ablation, electroporation, or electrolysis. Because all the effects may occur simultaneously, it may be difficult to reduce or eliminate any one effect. For example, eiectrolvsis and Joule heating may also occur during electroporation.

|04O| Electrolysis may be used in combination with other treatments such as irreversible electroporation, reversible electroporation, thermal ablation, cellular peraieabilization, and/or combinations thereof. The penneabi!izatlon of cells may increase the diffusion of electrolysis products into the ceils of the target volume of tissue. Increased diffusion of electrolysis products may enhance the effectiveness of electrolysis therapy and/or reduce the amount of electrolysis products sufficient to achieve a desired effect. The combination of electrolysis and electroporalion is described in, for example, Phillips, Mary; Rubinsky, Lie!; Meir, Arie; ^'Rubinsky, Boris "Combining Electrolysis and Electroporation for Tissue Ablation," Technology i Cancer Research and Treatment, Volume 14, Issue 4, pages 395-410 (2015), which is hereby incorporated by reference in its entirety and for any purpose.

[041 j Cryosurgery (e.g., cryosurgical techniques) generally employs cooling elements inserted into or brought in contact with tissue to be treated. In an exemplary cryosurgery procedure, cooling to subzero temperatures results in freezing of tissue that begins at the tissue cryosurgery probe interface and propagates outwards from the probe into the tissue. Ice has a tight crystallographic structure and solutes in the frozen medium may be concentrated by the removal of water as ice from the solution. This increase in solute concentration may be a mechanism of cryosurgery-induced cell death. Many different imaging modalities may be employed to detect the extent of freezing in real time, which may allow significant control over the cryosurgery treatment. However, in some examples, the extent of freezing does not directly correspond to the extent of tissue ablation. The temperature of the frozen lesio produced during cryosurgery may range from cryogenic near the tissue/prob interface to. -0.56 °C on the outer edge of the frozen lesion. The .region of frozen tissue having .a temperature between -40 °C and -0.56 °C may contain living cells such that the margin of the frozen lesion does not directly coincide with the extent of complete cell ablation. This difference between frozen legion margin and dead cells may make cryosurgery less precise and may be a drawback of the procedure. The combination of electrolysis and cryosurgery may address this deficiency. Details on the field of cryosurgery may ^'be found in, for example, Rubinsky. B. "Cryosurgery" Ann. Rev. Biomed. Engr., Eds. Yarmushet al, Vol. 2, pages 157-189, {2000), which is hereby incorporated by reference in its entirety and for any purpose.

042J Systems described herein may generally include a cryosurgical system sufficient to generate cryosurgical temperatures at a targeted tissue interface, an electrolysis system sufficient to generate electrolytic species at or near a targeted tissue (with or without electroporation), and a mechanism (e.g., controller) to combine at least cryosurgery and electrolysis. Combining cryosurgical techniques and electrolysis may result in the combination working m a manner that enhances the effect of one or the other or both as a result of the combination.

043} The cryosurgical system may include a generator that regulates the cryoge (i.e., cooling agent), a control system to allow user interface and control over the application of the cryogen. and a probe for the application of the cryogen to the targeted treatment site. The electrolysis system may include a generator to produce electric current and an applicator or probe that has an electrode that conducts the current at the targeted tissue interface to produce electrolytic species. A relevant parameter in electrolysis generally is the amount of charge delivered. Generally, the higher the voltage and current the higher is the rate of electrolytic products generated. Under some conditions, the voltage may be high enough to produce electroporation as a secondary effect. The electrolysis system may be designed to produce electroporation based on the voltage used to generate the desired amount of charge (e.g., in coulombs) delivered for electrolysis.

©44J Example methods and devices described herein may improve the effectiveness of tissue ablation by cryosurgery through the addition of electrolysis to the cryosurgery procedure (with or without electroporation). The contribution of electrolysis is generally understood to be by producing chemical species that are toxic to cells at a certain concentra&on. Often electrolysis Is a lengthy process due to the high concentration of electrolytes needed. Cryosurgery (i.e. , freezing) may ablate cells in a certain low temperature range, while in the high subzero temperature range cells ma survive. In this high subzero range, freezing may cause an increase in the local concentration of the electrolytic compounds as well as permeabilize the cell membrane to those compounds. Consequently, examples of the combination cryosurgery and electrolysis may employ lower coiicentTations of toxic electrolytic compounds, such as by delivering electrolysis for shorter amounts of time, and may be effective in ablating frozen cells in the high subzero temperature range in which frozen cells survive cryosurgery. Electrolysis generally involves the delivery of electrical charges through electrodes, which may be performed with various protocols, some of which may also yield electroporation. An example of a multimodality sequence described herein includes delivery of electrolysis, followed by cryosurgery, optionally followed by another electrolysis sequence.

[045J Examples described herein include examples of the combined effect of cryosurgery with electrolysis, which may allow for more effective ablation, of tissue. Combining cryosurgery with electrolysis may produce a substantial increase in the extent of tissue ablatio as compared to the ablation produced by the same dose of electrolysis or cryosurgery separately. Without being bound by a particular theory, this phenomenon may be attributed to the electrolytically produced chemicals that, in combination with the process of cryosurgery, may cause cell damage of clinical utility under conditions in which the cryosurgery or the products of electrolysis alone will produce less damage. This damage may be direct and immediate to the affected cells, but alternatively or additionally may be delayed and indirect through, for example, an enhanced ischemic effect, which may be caused by more severe damage to the micro vasculature than in either of the two techniques when used separately.

j^'046{ Cryoabiation ma temporarily or permanently damage ceil membranes.

Electrolytic products may induce cell death more quickly if cell membranes are permeabilized than if they are intact. Combining electrolysis with cryosurgery may increase the effectiveness of cryoabiation. For example, a radius of cryoelectrolytic ablation may be increased compared to a radius of cryosurgical ablation alone. As ϊθ

.another example, cell death from eryoelectroiytic ablation may be enhanced near the edges of the frozen region compared to cryosurgical, -ablation alone. Without being, limited to any mechanism or mode of action, combination therapy by produce the highest concentration of electrolytic products at the electrode surface. Freezing may push the front of electrolytic products forward to the edges of the frozen region.

J FIG. .1 is a schematic illustration of a -multimodality electrolysis system 100 according to one embodiment. The system 100 may be capable of performing one or both of electrolysis and cryosurgery. As shown in FIG. 1, the system .100 may be employed on the siu face of a tissue 10, or the system 100 may be used inside tissue 10, proximate tissue 10. and/or in a cavity formed by tissue 10.

1 The system 100 may be generally understood as including an electrolysis device 110 and a cryosurgery device 1 15, each coupled to a controller 105 and a power supply 1.20. The power supply 120 may be a component of or integrated with any or more of the electrolysis device 110, cryosurgery device 1 15, and controller 105.

} in some embodiments, the electrolysis device 1 30 may include one or more electrodes for conducting a current through a solution. The solution may be native to the treatment site and/or it may be introduced to the treatment site. In some embodiments, the electrolysis device 1.1.0 may include an aqueous matrix in. contact with the electrodes, which may be placed proximate the treatment site. The aqueous matrix may be a gel including a saline solution, in some embodiments, the electrolysis device i 10 may be a treatment pad for surface treatments. In some embodiments, the electrolysis device 1 10 may include needle electrodes and/or a catheter for use within cavities and/or tissues.

j The cryosurgery device LI S may be configured to perforin cryosurgical ablation. The cryosurgery device 1 15 may include one or more probes through which a cryogen is internally circulated to cool the probe. Thermal conduction through the probe wall may operate to cool the target treatment site when the probe is placed at the site, such as on tissue 10. Cryosurgical abiation of the site may be affected by selection of the target temperature to be achieved at the site. The target temperature may be selected to cause the maximum amount of ablation by cryogenic freezing of the cells, or the target temperature may be selected to be below the threshold of cell death. I 1 In some embodiments, the electrolysis device 1 10 and the cryosurgery device 1 15 are separate devices. The eiecixolysis device 1 10 may be packaged with the cryosurgical device 1 15. In some embodiments, the electrolysis device 1 0 (electrode) and the cryosurgery device 115 (probe) may be placed at different locations with respect to the targeted tissue 10. For example, an electrode may be placed near the intended margin of the frozen lesion of tissue 10 while the crvosuraerv probe is placed at a site removed from the intended margin, which may promote ceil death at or near the margin without reaching low subzero temperatures in the same region, m other examples, when the devices 1 10, 1 15 are in use, they may be placed proximate to a treatment site on the tissue 10.

J in some embodiments, the electrolysis device 1.10 and the cryosurgery device 1 15 are a single device. For example, the electrodes for performing electrolysis may also perform cryosurgical ablation.

} The controller 105 may be a separate component coupled to the devices 1 10, 1 15, as shown in FIG. 1, or the controller 105 may be integrated into one or both devices 110, 1 15, or packaged together with one or both devices 1 10, 1 15. In some embodiments, the controlle 105 may include a programmabl chi coupled to the devices Ϊ 10, 115,

J The controller 105 may control the timing, strength, and/or duration of treatments provided by the devices 1 10, 1 15. The controller 105 may, for example, be programmed or programmable to provide an electronic signal to the devices 1 10, 115, The electronic signal may indicate a dose of treatment, such as a dose of electrolysis products and/or targeted cryosurgical temperature. The electronic signal may control the timing or magnitude of a current generated by the electrolysis de vice 1 10 and/or the cryosurgery device 1 15. The controller 105 may allow a user to customize treatment of the tissue 10. The controller 105 may, for example, include such a program, or include one or more processing devices (e.g. processors) coupled to a memor encoded with executable instructions for electrolysis treatment or cryosurgical treatment, in some examples, cooling of the cryosurgery system may be connected to or otherwise be able to interact with the joules heating provided by the current. Such connection or interaction may help produce a desired temperature. For example, the target tissue may be pre-cooied or cooled during ibe delivery of current to avoid carbonization, which may also avoid loss of conductivity,

J In some embodiments, die controller 105 ma be implemented using a computing device (not shown) and may be remotely coupled to the devices 1 10, 115. The computing device may be, for example, a desktop computer, laptop computer, server, handheld computing device, tablet computer, and/or a smart phone. In some examples, the computing device may be integrated with and or shared with a separate piece of medical equipment. The controller 1 5 may be coupled by a wire to the devices 1 10, 1 15 or may communicate with the devices 1 10, 1 15 wirelessly. in some embodiments, two separate controllers 105 ma be used in the system 100. Each controller 105 may be coupled to one of the devices 1 10, 115.

J The system 100 may further include one or more sensors for measuring properties of the tissue 10. One sensor may be a pH sensor 125 for measuring pl l. One sensor may be an electric meter 130 for measuring electronic field strength. The pH sensor 125 may sense pH near the electrolysis device 1 10 and provide the pH value to the controller 105. The controller 105 may be programmed to adjust an electronic signal provided to the electrolysis device 1 .10 based on the pB near the device .1 10. A reservoir (not shown) may be provided to store and permit the addition of buffers or other solutions to the target tissue or an aqueous matrix, which may permit, adjustment of the pH. of the tissue or matrix, h another example, the pH sensor 125 may be inserted into tissue at the outer edge of the targeted volume. The pH sensor 125 may detect when the pH level at the site edge has reached a desired level, which may help ensure tissue ablation at the edge and throughout the site. Detection of a desired pH level may be a prompt fo the controller 105 to terminate the electrolysis process. In another example, the pH sensor 125 may be inserted at a selected tissue site and may detect when the pH level at the site is reaching or has reached an undesirable value. Detection of a given pH level may be a prompt for the controller 1.05 to terminate the electrolysis process, which may help avoid tissue damage.

j In some embodiments, a feedback system may be included in a communication path between the contr ller 105 and the probe, such as via a sensor positioned on the probe or positioned remotely from the probe in the treated tissue. The sensor may be, for example, a temperature sensor, a pH sensor 125, or an electric meter 130. ) FIGS. 2A and 2B are flow charts illustrating methods 200 A, 200B according to some embodiments. In some implementations, a raaltimodality electrolysis system, device, and/or apparatus may be positioned for treatment of a target site such as a tissue. The multiniodaMty electrolysis system .1 0 of FIG. 1 may be used. The treatments performed by the multimodal! ty electrolysis system may be manually controlled by a user or may be controlled by a controller such as controller 105 shown in FIG. 1 , Delivery of electrolysis products may be performed in a tissue to be treated before, during, or after cryosurgery is performed in the same tissue. Cryosurgery performed before, during, and/or after delivery of electrolysis products may improve die effectiveness of the electrolysis products in ablating the target tissue.

) In some implementations, and as shown in method 200 A, electrolysis 205 is performed and may be followed by cryosurgery 210. Electrolysis 205 may deliver electrolysis products to the target site. Cells at the target site may have increased susceptibility to cell death due to the combined delivery of the electrolysis products and cryosurgery. In some embodiments, electrolysis 205 may be repeated after cryosurger 210, In some embodiments, electrolysis 205 and cryosurgery 210 may be repeated in an alternating fashion for a desired period of time. Electrolysis 205 and cryosurgery 210 may be performed for the same or different time durations, magnitudes,, and/or other parameters. In some embodiments, electrolysis 205 and cryosurgery 210 may be separated by a period of time where no treatment i applied to die target site.

j in some implementations, and as shown in method 200B, cryosurgery 215 is performed and may be followed by electrolysis 220, Cells at the target site may have increased permeability in response to the cryosurgery 215. The electrolysis 220 may deliver electrolysis products to a target site. In some embodiments, cryosurgery 215 ma be repeated after electrolysis 220. In some embodiments, cryosurgery 215 and electrolysis 220 may be repeated i an alternating fashion for a desired period of time. Cryosurgery 215 and electrolysis 220 may be performed for the same or different, time durations., magnitudes, and/or other parameters. In some embodiments, cryosurgery 215 and electrolysis 220 may be separated by a period of time where no treatment is applied to the target site.

1 In some embodiments, electrolysis and cryosurgery may be performed at the same time or partially at the same time. For example, current to generate electrolysis products may be provided during a same period of time as cryogenic temperatures are •applied io a tissue. In sonie embodiments, electrolysis and cryosurgery may both be performed together for a continuous period of time or intermittently, in one example, electrolysis is started prior to the commencement of cryosurgery and is continued after the termination of cryosurgery. In some embodiments, one treatment may be performed continuously while the other treatment is performed intermittently. The magnitude and duration of each treatment may be modulated independently of the other treatment. For example, electrolysis may be performed for several seconds each minute, while cryosurgery may be performed continuously for several minutes. The electrolysis may be discontinued while the cryosurgery is continued. Other combinations of treatments may be possible. The time, duration, and order of the treatments ma be chosen based at least in part on the desired effect on the target site, the size of the target site, and/or local physiological conditions of the target site.

{062} FIG, 3 is a schematic diagram of a treatment probe 300 according one embodiment. The treatment probe 300 may be a needle. The treatment probe 300 may incorporate both a cryosurgery probe 305 and at least one electrolysis electrode 310. The electrode 310 for electrolysis may be electrically insulated from the cryosurgery probe 305. Electrically insulating the electrolysis electrode 310 from the cryosurgery probe 305 may permit independent optimization of both cryosurgery and electrolysis functions. For example, in the design and construction of the electrolysis electrode 310, materials may be selected for the production of speciiic electrolysis product species. As another example, the material for construction of the cryosurgery probe 305 may be selected to maximize thermal conduction. The probe 305 and electrode 310 configurations may be in any number, size and shape.

[063J in some embodiments, the treatment probe 300 ma include a plurality of electrolysis electrodes 310. hi some embodiments, the treatment probe 300 may- include plurality of cryosurgery probes 305. One or more electrodes 310 may deliver electrolysis treatment and one or more cryosurgery probes 305 may deliver cryosurgical treatment. The electrode 310 or plurality of electrodes 310 may be constructed of one or more materials. Electrode construction materials may be selected to help produce electrolysis products when a current is passed through an aqueous matrix by the electrode. Electrode materials may be selected to help produce specific electrolysis products. For example, an anode ma b constructed, at least, in pan of iridium oxide and/or rubidium oxide deposited on titanium, which may improve the production of hypochlorous acid. As another example, a cathode may include copper. The use of mixed metal oxide anode electrodes ma produce different species of eiectrolysis products that may be tailored for different clinical needs. For example, platinum may be used if inert electrodes are desired. As another example, silver electrodes or silver/silver chloride electrodes ma be selected if silver ion electrolysis products are desired, which may enhance a sterilization effect.

) In some embodiments, the cryosurgery probe 305 may be constructed of one material (e.g., stainless steel), and the electrolysis electrode 31.0 may be constructed of a different material (e.g., titanium, iridium oxide, or platinum). The different physical properties of the materials may aid in electrolytic production in a discrete area of the treatment probe 300 while avoiding electrolytic production in other areas of the probe 300.

| in some embodiments, a cryosurgery probe includes an. electrolysis sheath. The eiectrolysis sheath may permit the combination of eiectrolysis electrodes with cryosurgery probes. The eiectrolysis sheath may include electrodes, which, may generate electrolytic species at the interface between the sheath and the target tissue. The sheath may be a fixed length or a variable length, which ma permit placement over the therapeutically active area of the cryosurgery probe, or retraction away from the therapeutically active area of the cryosurgery probe. The electrolysis sheath may aid in removal of the cryosurgery probe from the tissue. The sheath may be constructed at least in part from a non-electrieally conductive material, which may restrict the electrical activity to only the conductive areas of the sheath. The thermal insulation of the non-conductive regio may be enhanced, as ma be the non-cryo-active portion of the cryosurgery probe.

1 Example methods and systems combining electrolysis and cryosurgery may overcome drawbacks of each of electrolysis and cryosurgery used alone. The combination cryosurgery and electrolysis may occur in. various combinations suc as electrolysis first, cryosurgery next; cryosurgery first, electrolysis next; electrolysis and freezing simultaneously; or electrolysis and thawing simultaneously. The combined modality of freezing and electrolysis or thawing and electrolysis may shorten the time of a cryo lectrolysis procedure, ^'which may be beneficial in time-sensitive clinical ■applications.;

[067J An exemplary combination method may include the following steps. First, an electrolysis protocol may be implemented. During electrolysis, an electric current may be delivered to electrodes in contact with a target tissue to generate electrolysis products in the tissue. Compared to electrolysis alone, the current delivered in the exemplary combined method may be low; such that the electrolysis products alone may not be sufficient t cause cell death by electrolysis.

[068) Second, freezing may be performed by cryosurgery. The effects of freezing may include some or all of the following: concentrate the products of electrolysis, which may be due to removal of water from the solution as ice; force the flow of electrolytic products to the exterior of the frozen lesion, which may be a result of forced convection (e.g., ice has a lower density than water and freezing may act to generate a flow of the unfrozen solution in front of the freezing interface); and induce phase transformation in the cell membrane, which may permeabili e the membrane and open the cell interior to highly concentrated products of electrolysis. In examples described herein, any or ail of the effects of f ezing may lead to cell death throughout the frozen lesion at a temperature range in which cells can survive freezing.

[069) In some methods, a second deployment of electrolysis follows cryosurgery.

Electric currents also produce heat as result of a Joule heating effect, and the heat may melt the region of frozen tissue adjacent to the probe. The availability of fluid near the probe may further produce electrolytic products in the vicinity of the probe. The localized production of electrolytic products .may facilitate the removal of the probe from the frozen lesion while ablating cells electrolyiically along the tract of the probe. When used to ablate cancerous tissue, the concurrent probe removal and cell ablation may help reduce or eliminate of the opportunity for cancer cells to translocate and seed.

[0?0) i some methods, the frozen lesion may be thawed by body heat and the dead cells may be removed by the body's immune system. In some examples, active thawing is employed, such as to remove cryoneedles or to enhance the destructive effects of the combination therapy.

[071) Freezing of tissue by a probe may result in adherence of the probe to the tissue, in cryosurgery, when several probes are used simultaneously, it may be beneficial to use a sticking (adherence) mode to fix the probes in place, ^'cryoelectrolysis probes ma adhere during the simultaneous applications of freezing and electrolysis or thawing and electrolysis. Probe adhesion during cryoelectrolysis may he used such as in tumor treatment or tissue ablation.

|0?2| In some methods, electrolysis is performed first, then cryosurgery, then electrolysis again. The first application of each of electrolysis and cryosurgery is performed either sequentially or simultaneously, which may .maximize the treatment effect The subsequent application of electrolysis may be performed to aid in removal of the treatment probes. The removal may be effected by applying the DC current for electrolysis, which may result in thermal heating and thawing around the cryosurgery probe shaft. The removal may be effected b the production of electrolytic species, which may result in pH-mduced melting of ice at the surface of the cryosurgery probe.

(073) In some methods, electrolysis is performed to aid in removal of the treatment probes after cryosurgery alone. The removal may be effected by any method described above for the combination of electrolysis and cryosurgery.

[074J In some examples, an electric modality may be implemented at the beginning of electrolysis phase to produce an acute microvascu!ature stop, which may help create a partially or completely sealed area where electrolysis products may be retained longer than in the absence of the stop. In some examples, the electric modality may be implemented prior to the commencement of cryosurgical techniques in order to enhance permeabihzation of the cell membrane.

[075) Combination cryosurgery and electrolysis devices and methods may be used for any application for which cryosurgery alone or electrolysis alone may be employed. Applications for cryoelectrolysis devices and methods include treatment of tumors; cardiovascular treatments such as treatment of atrial fibrillation, arrhythmias, deep vein thrombosis, percutaneous transvascular applications, congenital heart disease, peripheral artery disease, restenosis, and other lumen-based treatments; nerve ablation; renal denervation; and ablation of fat, such as for dermatological or cosmetic purposes. Each combination treatment may provide enhanced results compared to either treatment alone.

(076) Tumors treated by cryoelectrolysis may be malignant or benign. ^'Tumors may include, but are not limited to, prostate, bladder, breast lung, liver, brain, colon. esophagus, kidn y, rectal, skin, stomach, pancreas, eye, and uterine tumors, and benign prostatic hypertroph , fibroids and niyonias.

J Other dermatologies! or cosmetic applications of cryoelectrolysis include treating benign skin tumors, actinic keratosis, basal cell carcinoma, dysptastie nevi, melanoma, and squamous cell carcinoma; skin resurfacing; skin tightening; skin lesion removal; hair removal; wrinkle removal or reduction; and acne removal, reduction, or prevention.

} In some embodiments, the electrolysis and cryosurgery may be combined with other modalities for tissue treatment such as thermal ablation, radiation, chemical ablation, irreversible electroporation, reversible etectroporaiion, and/or gene therapy.} it is to be understood that the examples provided of both the delivery systems and the clinical applications are not the limit of the uses of the combination of cryosurgery and electrolysis. Many configurations of delivery systems exist, as well as applications that would benefit from the use of the delivery systems and applications disclosed herein. It is to be appreciated that any one of the above embodiments or processes may be combined with one or more other embodiments and or processes or be separated and/or performed amongst separate devices or device portions in accordance with the present systems, devices and methods,

} Finally, the discussion herein is intended to be merely illustrative of the present devices, apparatuses, systems, and methods and should not be construed as limiting to any particular embodiment or group of embodiments. Thus, while the present disclosure has been described in particular detail with reference to exemplary embodiments, it should also be appreciated that numerous modifications and alternative embodiments may be devised by those having ordinary skill in the art without departing fro the broader and intended spirit and scope of the present disclosure. Accordingly, the specification and drawings are to be regarded in an illustrative manner and are not intended to li mit the scope of the appended claims.

EXAMPLES

Example 1 : Modality comparison in pig liver

} A study was conducted on in vivo pig liver using two female pigs, weighing 30 kg to 4⁽3 kg, treated in accordance with Good Laboratory Practice regulations as set forth in 21 CFR 58. Each procedure began with anesthetization of the animal under general anesthesia per SOP #33156. The treatment, was delivered us.bg two Endoeare (Austin, TX) 2 mm cryosurgery probes, which served, for both cryosurgery and to deli er electrolysis.

) The ct opro es were inserted into the liver in a roughly axial parallel configuration, normal to the liver surface, under ultrasound monitoring. The cryoprobes were connected to a 180 bar argon gas pressure supply and also connected to an Arbitrary Function Generator (AFG 3102, Tektronix, Beaverton. OR) to produce a constant current for a fixed period of time, for deli vering electrolysis.

) Up to six lesions, separated by 3 cm each, were produced in the liver of each animal by one of the following three procedures: (a) cryosurgery in which the two probes were connected to the argon pressure bottle and the tissue was frozen for 10 minutes at the lowest, temperature achievable by the apparatus (i.e., -180 °C); (b) electrolytic ablation in which an electric current of 00mA was delivered from a 48 V power supply for 10 minutes to produce a charge of 42 C; and (c) cryoelectrolysis in which electrolysis (b) was followed by cryosurgery (a).

') Animals were sacrificed at 4 to 5 hours post procedure, and the treated tissues were examined tor macroscopic histology and for histology using H&E and trichrome blue staining.

) FIG. 4 shows a macroscopic comparison between lesions, produced by cryosurgery (procedure a) (FIG. 4A) and those produced by cryoelectrolysis- (procedure c) (FIG, 4B). FIG. 5 shows a macroscopic comparison between lesions produced by electrolysis (procedure b) (FIG. 5A) and those produce by cryoelectrolysis (procedure c) (FIG. SB). FIG. 6 shows a microscopic comparison of the margins 600 of the lesions of FIG. 4. Cryosurgery alone resulted in. a. diffuse margin between live cells 602 and dead cells 604. (See FIG. 6 A.) In contrast, cryoelectrolysis resulted in a sharp margin 600 between live cells 602 and dead cells 604. (See FIG. 6B.)

Example 2: Electrolysis through a freezing gel

} Petri dishes 700 (IQ-cni diameter) were filled with about ½ inch of a simulated tissue made from solidified agar gel 702 comprising physiological saline and 5% agar. Methyl red dye was added to the gel at 1 ral/L as an electrolysis (acidification) indicator. (Ό87) Temperature was measured using a digital temperature meter 706 and a thermocouple 708, as shown in FIG. IB.

[088J An anode 710 was constructed of a flat, rectangular Ι25-μη* titanium foil coated with a 2~μπι layer of iridium oxide, which facilitates production of chloride species at the anode. The cathode 712, a J -mm titanium rod, was inserted in the gel as shown in

FIG. 7C\

|089J A freezing probe 714 was constructed of a metal cylinder 70 having an inner chamber 716, flat bottom 718, 2-inch outer diameter, and .1.5-inch inner diameter. The chamber 71 was filled with a salt solution that froze at - -10 °C and was pre-frozen in a freezer at -20 °C (FIG. 7D). As shown in FIG. 7E, the bottom 718 of the .freezing probe 71.4 was covered with a thin electrically insulating layer of a Parafi!m® plastic paraffin film 720.

[090^'| As shown in FIG. 7F, the anode 7.10^' was placed on the gel 702 and the flat bottom 718 surface of the freezing probe 714 was placed on the anode 710. Electrolysis (25 V, 194 mA from power supply 730; FIG. 7A) was performed simultaneously with freezing and until the temperature of the gel 702 was about -1 1 °C (FIG. 7B).

[0911 FIG, 7G shows the appearance of the gel 702 after a 2-minute application, of the freezing probe71.4. The anode 710 is in the center surrounded by stained gel 736, which indicates the extent of electrolysis. A round, frozen lens 722 indicates the location of the outer edge of the freezing probe 714.

[092| FIG. 7FS demonstrates that electrolysis may be performed simultaneously with freezing on tissue-simulating gel to produce a region affected by both freezing and electrolysis. Accordingly, tissue ablation may be achieved by simultaneously freezing tissue and subjecting it to electrolysis, which may enhance the effectiveness of both ablation methods.

Example 3: Adherence of freezing probe to frozen region during cryoe!ectrolysis

[093} Simulated tissue, an electrode, and a freezing probe were prepared as described in Example 2 except as noted. Corresponding reference numerals reflect corresponding components of the experimental setup.

[094} The anode 810 was constructed of a 1 -mm diameter titanium needle coated with i ridi um oxide, as sho wn in the center of the Petri dish.800 of FIG. 8A. (^'095} As showrs m FIGS. SB & 8C, the anode 810 was placed horizontally on the gel 802 and the freezing probe 814 was placed on. the anode 810. Electrolysis (about 150- 250 niA) was performed simultaneously with freezing for about 2 minutes.

[096] During electrolysis and freezing, the freezing probe 81.4 was lifted upwards, thereby lifting the Petri dish SOO due to adherence of the probe 814 to the gel 802. FIG. SB shows the dish 800 and probe 814 before lifting and FIG. 8C shows the dish 800 and probe 814 after lifting.

[G97J FIG. 8D shows the appearance of the gel 802 immediately after the freezing probe 814 was detached from the gel 802, The stained gel 836 indicates the extent of electrolysis. An ice lens 722 surrounded the anode 810 and an arrow points to the margin of the frozen region. FIG. BE, a cross section of the treated gel 802, demonstrates that the stained region 836— reflecting the electrolysis products— extends through the entire 1.2 nun cross section.

[098J Results demonstrate that a freezing probe may bind tightly to a frozen region.

The pH front produced by electrolysis products may propagate through the frozen region and beyond. During cryoelectrolysis, freezing may be used to bring the electrode and other structures into tight contact with the tissue to be frozen.

Example 4: Electrolysis perfo med in completely f ozen tissue simulating gel

|099j Simulated tissue, an electrode, and a freezing probe were prepared as described in Example 3 except as noted. Corresponding reference numerals reflect corresponding components of the experimental setup.

{01001 The gel-filled Petri dish 900 was f ozen overnight in a freezer at -20 ¾, During the experiment, the temperature of the dish 900 was maintained in a constant temperature bath at from -2 °C to -20 °C, The anode 910 was constructed as in Example 3 but was placed perpendicular to the surface of the gel 902, as shown in FIG. 9A. Electrolysis (25V, 200 raA) was performed simultaneously with f eezing for 2 minutes.

fOIOIJ FIGS, 9B-D show the propagation of the pH front over time (i.e.. after 30 seconds, FIG. 9B; after 1 minute, FIG, 9C; after 2 minutes, FIG, 9D) in the gel 702 held at. -10 °C. For reference, a ruler (in cm) is shown adjacent the propagating electrolysed zone. 0102] Results demonstrate that electrolysis may take place in a completely frozen tissue-simulating gel., in general, the higher the subfieeztng temperature, the faster the propagation of the pH front produced by electrolysis products.

Example S: Full cryosurgery protocol In combination with electrolysis protocol

01 ⁾31 A simulated tissue was prepared by mixing 1 liter of water with 9 grams aCI and 7 grains of agarose. The solution was stirred and heated for 10 minutes and then removed from the heat. Two pH indicator dyes were added after five minutes of cooling. For analysis of electrolysis near the anode 1010, methyl red at 1. ml. per 100 h agar solution was added. For analysis of electrolysis near the cathode 1012, 0.5 wt % phenolphthalein in ethano! at 1 mL per 100 ml, agar solution was used. The agar gel 1002 was cast to a height of 4 era in a 20-cm diameter cylindrical glass vessel 1026 having radial walls coated with a 200-μηι thick copper foil (t serve as an electrode 1028).

0 041 With reference to the experimental setup shown in FIG. Ί0Α, an Endocare® R2.4 stainless steel cryoprobe 1024 (or "cryoelectrotysis probe") with a diameter of 2.4 mm was connected to an Endocare® single port control console .1038 to regulate flow duration and monitor feed-hack temperatures ( Endocare Inc., Austin, XX). The cryoelectrolysis probe 1 24 was supplied by a pressurized argon gas contai ner through the control console 1038 at a constant pressure of 3000 psi. The cryoelectrolysis probe 1024 was cooled through a Joule-Thomson internal valve. The probe temperature is capable of reaching -180 °C at a rate of cooling governed in part by the thermal environment in which the cryoelectrolysis probe 1024 is inserted, A 30 μηι gold foil was wrapped several times around the cryoelectrolysis probe 1024 to minimize the participation of the cryoprobe metal in the electrolysis process. The metal body of the cryoelectrolysis probe 1024 was connected to a DC power supply 1030 (Agilent E3631 A, Santa Clara, CA). The electrical circuit, included the power supply 1030, the cryoelectrolysis probe .1024 electrode, the gel 1002, and the copper electrode 1028 around the vessel 1026.

01051 The cryoelectrolysis probe 1024 was inserted vertically into th center of the gel 1002. A 1 mm T-type thermocouple 1008 was inserted into the gel 1 02 less than 5 mm from the outer surface of the cryoelectrolysis probe 1024, as shown in FIG. 10B, and the gel temperature was recorded continuousl throughout the experiment. A camera 1032 was focused o» the experimental setu to continuously record the position of the change-oiVphase interface, position of the pH .from (produced by electrolysis products)., voltage, current, and time (via a timer 1034).

01 6J The electrical circuit was connected to the power supply 1030 first and remained connected during both freezing and thawing. The flow of cryogen (argon gas) began one minute after the circuit was connected and continued for 10 minutes. A constant pressure of 3000 psi was used to generate the argon gas flow in a manner typical for clinical cryosurgical treatment with the selected cryoelectrolysis probe 1024, Application of the cryogen produced a frozen lesion, which was left to thaw in situ. The electrical circuit remained connected to the power suppl 1030 for an additional 15 minutes after the cryogen .flow was terminated. The current was set to 50 niA to 400 raA and the voltage was allowed to change to provide the desired current. If changes in resistance demanded a voltage higher than the saturation voltage of 25 V, the current dropped accordingly.

01O7J FIGS. I OC 8c 10D show the progressio of the pli front during application of electrolysis without cooling. The cryoelectrolysis probe 1024 served as the anode and delivered a 400 niA current. FIG. IOC shows the radially symmetric pB front (stained gel 1036) around the cryoelectrolysis probe 1.024 anode and FIG.. l OD shows the same gel 1.002 and vessel 1026 after electrolysis was continued for several minutes. The pH front advanced over time while remaining radially symmetric. The insert in FIG. l OD (as well as FIG. 10F & 10H) is a magnified view of the region near the cryoelectroiysis probe 1 24 anode. A dark gap 1040 formed between the cryoelectrolysis probe 1 24 anode and the gel 1.002. The gap 1040 may have been caused by the electro-osmotic- driven flow of solution away from the cryoelectrolysis probe 1024 anode, towards the copper foil cathode electrode 1028. Diffusion- and iontophoresis-driven electro- osmosis may cause the propagation of the pH front from the cryoelectrolysis probe 1024 anode outward.

^'01.081 FIGS, lO^'E & I OF show the progression of the pH front and the ice front during cryoelectrolysis. The cryoelectrolysis probe 1024 served as the anode and delivered a 400 niA current. FIG. I E shows the frozen lesion .1042 at the end of the cooling stage and FIG. 10.F shows the same gel 1002 and vessel 1026 taken several minutes after the cooling was stopped but while the power supply 1 30 continued to deliver current to the electrical circuit. The extent of the frozen lesion 1042 in FIG, 10F did not change over time from that in FIG. I0E. However, the pH front (stained gel 1036} in FIG. 10F advanced beyond the frozen lesion 1042. These results demonstrate that electrolysis may occur through a frozen region. A gap 1040 formed between the cryoelectrolysis probe 1024 anode and the gel 1002. The gap 1040 may have been generated by electro-osmotic flow. With reference to FIG. 913. the gap 1040 formed during conventional electrolysis may also occur during cryoelectrolysis (FIG. 9F). This result further demonstrates that electrolysis ma occur through a frozen region.

0109| FIGS. I 0G &10H show the progression of the pH front and the ice front during cryoelectrolysis. The cryoelectrolysis probe 1024 served as the cathode instead of an anode as in FIGS. IOC & l OD and delivered a 50 niA current. Each of the pH front (stained gel 1036) and ice front propagated asymmetrically. The difference, as seen by comparing FIGS. 10G & 10H with 10F & IGF, respectively, may be caused by the direction of the electro-osmotic flow, which in FIGS. 10G & 10H is towards the cryoelectrolysis probe 1024 cathode. The electro-osmotic flow may generate a high solution flow rate at the interface between the cryoelectrolysis probe 1024 cathode and the gel 1002. A gushing flow of water has been observed, at the interface between the cryoelectrolysis probe 1024 cathode and the gel 1002, independent of the current level used. The water contained hydrogen gas from the reduction reaction nea the cathode. Splashed droplets of acidic fluid created a splattering of stained dots 1044 on the right- hand side of the gel 1022.

Oil Of When the cryoelectrolysis probe 1 24 is the cathode, th electro-osmotic pressure may cause various random and detrimental effects. For example, at higher currents of 200 mA and 400 mA. the electro-osmotic pressure-driven flow may have caused f actures or cracks in the gel 1002 (not shown). At lower currents, such as 50 mA, the electro-osmotic pressure-driven flow may have produced the lack of symmetry seen in FIGS. 10G & I 0B.

0111 J FIG. FOB shows the same gel 1002 and vessel 1026 as in FIG. 10G but was taken after cooling had stopped and only electrolysis was occurring (i.e., a similar stage of the cryoelectrolysis protocol as thai of FIG. 10F). The pH front (stained gel 1036) propagated irregularly both within and beyond the frozen lesion 1042. These results demonstrates that electrolysis may occur through a frozen region when the cryoelectrolysis probe 1024 is either an anode (FIG. 10F) or a cathode (FIG. I OH). With reference to FIG. 1QE, the magnified insert of the region near the cryoelectrolysis probe 1024 cathode shows a bulging volume of ice 1046 and an ice-filled crack in the gel 1002. These physical features are in contrast to the dark gap 1040 between the cryoelectrolysis probe 1 24 anode and the gel 1002 in FIG. 10F.

01121 I summary of FIG, 1 , electrolysis may occur throug a frozen milieu at both the anode and cathode, and electro-osmotic flows may play a role .in the physical events that, occur during cryoelectrolysis and the flows may be different between a cryoelectrolysis cathode probe and a cryoelectrolysis anode probe. Also, the sequence and timing of electrolysis and cryosurgical technique (i.e., freezing) may be selected and adjusted to control the absol ute size of the area through which electrolysis products propagate (shown here as pH-dependent stained gel) and the area of the frozen lesion, as well as the size of each area relati ve to the other.

Example 6: Full cryosurgery protocol in combination with electrolysis protocol O^'l 1.31 Freezing and electrolysis were performed as in Example 5 except where noted.

Corresponding reference numerals reflect corresponding components of the experimental setup. The cryoelectrolysis probe 1124 was the anode and the current was set at a constant 200 niA.

01141 FIG. 1 is sequence of images showing the pH front and the ice front at different time points during the cryoelectrolysis protocol Captured time points (in minutes) are as follows; A, 1 minute; B, 2; C 3.5; IX 1 1; E, 12.5; F, 16; G„ 18.5; H, 21; and L 26 minutes. The data in FIG, 12 was gathered during the same experiment as the images of FIG. 1 1 and displays the diameter of the pfl front (solid line) and the ice front (dashed line) (FIG: 12 A); the measured current (FIG. 12B); the measured voltage (FIG. I 2C); the calculated resistance (FIG. 12D); and the temperature as measured by a thermocouple (FIG, 12E)₅ each as a fraction of time.

0115| FIG. 1 1A shows the appearance of the pH front (stained gel 1 136) one minute after the start of the experiment, just prior to the start of the freezing process. FIG. Γ1Β shows the appearance of the frozen lesion (ice front 1 142) and the pH front 1 136 one minute after the start of freezing and two minutes after the start of the experiment. A comparison of FIGS. 1 LA and 1 IB reveals that during the first one minute of freezing, each of the ice front 1 142 and pB front 1 136 advanced. 0116] With reference to the same time period in FIG. 12, as the temperature near the cryoeJectrolysis probe 1 1.24 dropped below freezing (FIG. 12 E), the voltages increased and reached the 25 V saturation level (FIG. 12C) after which the current began to decrease (FIG. 12 B). The current decrease may be caused by an increase in resistance in the freezing domain. FIG. 1 I B and the data in FIG. 12 demonstrate that electrolysis mav occur in the earlv. hiuh subzero temperature staue of the freezins process.

M il] FIGS, 1 1C₅ 1 iC, and 12 show that beyond one minute of freezing the pH front

1136 stopped advancing (i.e.. no electrolysis) while the ice front 1 142 propagated fUrther, FIG. 12 shows that, during the same time period, the temperature dropped further (FIG. I2E), the electrical resistance of the frozen lesion increased (FIG. 12D), and the flow of electrical current stopped (FIG. 1.2B).

0118] FIG. 12D shows that the gel 1 122 temperature began to rise as soon as the cooling stopped (i.e., 1.1 minutes after the start of the experiment). Surprisingly, the temperature remained close to and below the phase transformation temperature for most of the remainder of the cryoelectrolysis protocol. Without being limited to a mechanism or mode of action, the observed temperature changes may be related to the change in. enthalpy during phase transition of ice into water, which is very large relati e to the change in enthalpy due to change in temperature of ice. During melting, heat may be extracted from the frozen domain, through the change-of-phase interface, by the environment surrounding the interface. The temperature of the interface may be fixed by equilibrium thermodynamics of a two-phase system at constant pressure. As long as an ice and water mixture is present in domain, the temperature of that domain m y not exceed the thermodynamic phase transition temperature of the solution. The phase transformation process (melting) may occur on the change-of-phase interface, which ma propagate very slowly because of the large change in enthalpy involved. Because the enthalpy associated with changes of temperature in the frozen domain ma be very small relative to the change in enthalpy of phase transformation, the temperature of the frozen region may become elevated and reach the phase transition temperature quickly throughout the frozen region, while the region is still frozen. Consequently, while the extent of the frozen region may remain essentially unchanged at the end of cooling (FIGS. 1 lE-l lG) the temperature of the frozen region ma rise to become close to and below the change-of-phase temperature (FIG, 12E). The increase in the temperature of the frozen region may cause a. gradual increase in voltage (FIG. 12C) and a decrease in resistance (FIG. 12D). Electrolysis may continue and the pH front 1 .136 may expand beyond the margin of the froze region (ice front 1 142), while the region is still frozen (FIG, I I P- 1 11).

(0119} Further, but without being limited to a mechanism or mode of action, electrolysis in a high subzero frozen media may be associated with the process of freezing in solutions and tissues. Ice may have a tight crystallographic structure and may not. contain any solutes. Constitutional supercooling may dictate that during freezing of a solution, fmger-!ike ice crystals may form and the salt may be rejected along the ice crystals. High concentration salt solutions may form along the ice crystals. This process may occur during freezing of any aqueous medium, in solutions, gels, and tissues. While ice electrical conductivity may be essentially zero, electrical currents may flow through the high concentration brine channels until the temperature reaches the eiitectic -21.1 ^CC. Performing cryosurgery techniques at temperatures of -21.1 °C or above may be used in some examples. FIG. 12 B shows that current may flow through the high subzero temperature region of a frozen gel Flow of ionic current may be associated with electrolysis and may explain why the pH front may .advance while the tissue is still frozen. The flow of current through the brine channels may elevate the local temperature of the channels and may cause local melting and expansion or collapse of the brine channels, it is possible that this phenomenon is responsible for the jumps in voltage measured occasionally after freezing has stopped (Fig. 12C),

(0120} The above specification, examples, and data provide a complete description of the structure and use of exemplary embodiments of the disclosure as defined in the claims. Although various embodiments have been described above with a certain degree of particularity, o with reference to one or more individual embodiments, those skilled in the art could make numerous alterations to the disclosed embodiments without departing from the spirit or scope of the disclosure. Other embodiments are therefore contemplated. It is intended that all matter contained in the above description and shown in the accompanying drawings shall be interpreted as illustrative only of particular embodiments and not limiting. Changes in detail or structure may be made without departing from the basic elements of the invention as defined in the following claims.

Claims

CLAIMS What is claimed is:

1. A method of tissue ablation,_, the method comprising:

performing a first electrolysis such that electrolysis products are applied to a tissue; and

performing, during a time period that overlaps with the performance of the first electrolysis, a first cryosurgical technique such that at least a portion of the tissue is frozen.

2. The method of claim 1, wherein electrolysis is performed fa applying an electrode to the tissue.

3. The method of claim 2, wherein the electrode is also a cryosurgery probe.

4. The method of claim 2 or claim 3, wherein the electrolysis, products propagate outward from the electrode into the tissue.

5. The method of claim ! , wherein the electrolysis occurs in frozen tissue.

6. The method of claim 5, wherei the electrolysis products propagate through frozen tissue.

7. The method of any one of claims 1. to 6, wherein the electrolysis products include hypochloroiis acid,

8. The method of claim 1. further comprising -performing a second electrolysis subsequent to performing the first electrolysis and the first cryosurgical technique.

9. The method of any one of claims ί to 8 , wherein the tissu is frozen ai - 2.1.1°C or wanner.

!ø, The method of claim 1, wherein the cryosurgical technique is ^•commenced after die commencement of electrolysis aad terminates prior to the term mati on of e ! ectro I y sis.

1 1. The method of cl ai m 10, wherei n the rate of electrolysis may decrease or stop as freezing increases.

12. The .method of any one of claims I to 11 , wherein fin area of tissue exposed to electrolysis products may be the same size as, smaller than, or larger than an area of frozen tissue.

13. An apparatus for tissue ablation, the apparatus comprising:

a device for producing electrolysis products as well as for freezing tissue;

at least one powe simply operably connected to the. device for producing electrolysis products and freezin tissue; and

at least one controller operably connected to the device for producing electrolysis products and freezing tissue,

1.4 The apparatus of claim .13, wherein the device is an electrode.

Ϊ . The apparatus of claim 14, wherein the device is an anode.

16. The apparatus of claim. 13, wherein the device is a. cryosurgery probe.

17. The apparatus of any one of claim 13 to 16, wherein the device is constructed in part of stainless steel.