WO2023039582A1 - Sub-microsecond electrical field pulses to enhance efficacy of toxin treatments - Google Patents

Abstract

Described herein are methods (including cosmetic methods) and apparatuses for delivery of sub-microsecond electrical field pulses to enhance effects of a toxin, such as (but not limited to) botulinum toxin (BoNT).

Description

SUB-MICROSECOND ELECTRICAL FIELD PULSES TO ENHANCE EFFICACY OF

TOXIN TREATMENTS

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This patent application claims priority to U.S. provisional patent application No. 63/243,673, titled “SUB -MICRO SECOND ELECTRICAL FIELD PULSES TO ENHANCE EFFICACY OF THERAPEUTIC TOXIN TREATMENTS,” filed on September 13 2021, which is herein incorporated by reference in its entirety.

INCORPORATION BY REFERENCE

[0002] All publications and patent applications mentioned in this specification are herein incorporated by reference in their entirety to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

BACKGROUND

[0003] Botulinum toxin (BoNT) is a neurotoxic protein produced by the bacterium Clostridium botulinum which prevents the release of the neurotransmitter acetylcholine from axon endings at the neuromuscular junction, thus causing flaccid paralysis. The toxin causes the disease botulism. The toxin is also used commercially for medical and cosmetic purposes. Commercial forms are marketed under the brand names Botox™ (onabotulinumtoxinA), Dysport™/Azzalure™ (abobotulinumtoxinA), Myobloc™ (rimabotulinumtoxinB), Xeomin™/Bocouture™ (incobotulinumtoxinA), and Jeuveau™ (prabotulinumtoxinA). These commercial forms may have different dosage units.

[0004] BoNT injections are noted for the ability to treat various conditions, however, they are also used for purely cosmetic purposes, such as reducing the appearance of facial wrinkles. BoNT temporarily prevents a muscle from moving. While the primary application of neurotoxins such as Botox™, commercially sold by Allergan, Inc. (Irvine, Calif.), has been focused on cosmetic applications, such as treatment of facial wrinkles, other uses for the compound are now common. Certain applications include treatment of cervical dystonia, tremor, headache (migraine), spasticity, torticollis, hemifacial spasm, blepharospasm, meige syndrome, spastic dysphonia, writer’s cramp, hyperhydrosis, hypersalivation, bladder dysfunction, multiple sclerosis, spinal cord injury, cystic fibrosis, stroke paralysis, stuttering, and all types of pain.

[0005] Because it is a toxin, BoNT must be carefully delivered to the body, and the onset and extent of the effects should be tightly controlled. It would therefore be desirable to provide methods and apparatuses for enhancing the effects of BoNT in a subject (e.g., a patient), including reducing the onset time when BoNT takes effect and/or the amount of BoNT used in a therapy or in a purely cosmetic applications.

SUMMARY OF THE DISCLOSURE

[0006] Described herein are methods and apparatuses for treatment of tissue with submicrosecond electrical field pulses to enhance the effects of a toxin, such as (but not limited to) botulinum toxin (BoNT). In particular, described herein are methods and apparatuses (e.g., systems, devices, etc.) for delivering sub-microsecond electrical field pulses to the tissue to which a treatment with a toxin, such as neurotoxin BoNT, has been applied in order to reduce the amount of time necessary to achieve a robust effect of the BoNT, increase the effect of the BoNT on the tissue, and/or reduce the amount of BoNT that is needed to achieve the same effect as would have been by administering BoNT alone without delivering sub-microsecond electrical filed pulses. In general, any enhancement of the toxin (e.g., BoNT) effect, including as stated above, may be referred to as activating the toxin with sub-microsecond electrical field pulses. [0007] For example, described herein are various methods, including therapeutic and non- therapeutic methods. These methods may be used for enhancing treatment with a toxin, such as BoNT. In some examples, these methods may reduce the time it takes for the toxin to fully activate to treat/affect the target tissue, including reducing the time for the toxin to reduce the appearance of surface wrinkles, eliminate or reduce pain, reduce the activity of glands (sweat glands), reduce the activity of target muscles (e.g., overactive bladder, lazy eye, etc.), etc. As compared to target tissue in the same subjects when the toxin is administered alone, the methods and apparatuses described herein may reduce the time for activation, or complete activation, of the toxin, for example, by 10% or more (e.g., 35% or more, 40% or more, 45% or more, 50% or more, 55% or more, 60% or more). In some examples and implementations, any of these methods may reduce the time for activation of the injectable toxin by more than 40% (and in some cases by more than 75%) compared to untreated target area.

[0008] Thus, according to some implementations, described herein are methods comprising: applying, following an introduction of a toxin, a sub-microsecond pulsed electric field (e.g., nanosecond pulsed electric field) to a target area into which the toxin has been introduced, wherein the sub-microsecond pulsed electric filed is applied over or in proximity to the target area from one or more electrodes, thereby enhancing effect of the toxin (e.g., reducing the time for activation of the toxin), wherein the sub-microsecond pulsed electric field comprises one or more pulses each having a duration of less than 1 psec and an electric field, for example, of 5kV or less. In some examples, the toxin may be introduced by injection, in other examples the toxin may be topical. It should be understood that any appropriate method of introduction of the toxin is within the scope of the present disclosure. In some examples the voltage may be less than lOkV/cm, less than 5kV/cm, less than 2kV/cm, less than 1.5kV/cm, less than IkV/cm, less than 500 V/cm. In some examples the voltage may be between 0.75kV/cm and 1.5KV/cm. The energy applied may be, for example, between approximately 0.3 mJ to 100 mJ. While in many examples, non-penetrating electrodes (e.g., plate electrodes, surface electrodes, etc.) may be beneficial, in some examples, penetrating electrodes (e.g., very small needle electrodes) may be used as well. In any of these methods the toxin may be a botulinum toxin (BoNT).

[0009] Some of the methods may also include a step of injecting the injectable toxin into the target area. The injection may be performed by the same person or agent performing the application of the nanosecond pulsed electrical field or may be performed by a different person or agent. For example, the method may include noting (recording/storing and/or recalling/retrieving, etc.) the time of the injection and waiting for the waiting time. A timer (e.g., alarm, alert, etc.) may be set either automatically or manually to count off the waiting time period.

[0010] In some examples, the nanosecond pulses may be applied after an optional waiting time following an injection. In general, such waiting time may be, for example, at least about 5 min, about 10 min, about 15 min, about 25 minutes, at least about 30 minutes, at least about 35 minutes, at least about 40 minutes, at least about 45 minutes, at least about 50 minutes, at least about 1 hour, etc. In some examples the waiting time may be, e.g., between about 20 minutes and about 12 hours (e.g., between about 30 minutes and about 5 hours, etc.).

[0011] In general, the applied sub-microsecond (e.g., nanosecond) pulse energy field (“submicrosecond pulsed energy”, “nanosecond pulsed energy”) may be applied at an intensity that is sufficient to cause a brief muscle activation of the underlying muscle and/or nerve. The nanosecond pulsed energy may be felt by the subject (e.g., the patient being treated), but may be non-irritating, and is generally not painful. In some examples the applied nanosecond pulsed energy may be sub-sensory (e.g., may not be consciously sensed by the subject), however if higher electric fields are used, pain management may be used, as needed. In any of these methods and apparatuses, the nanosecond pulse electric field may comprise, for example, between 1 and 200 pulses (e.g., between 2-100 pulses, between 2-80 pulses, between 3-50 pulses, between 5-30 pulses, between 5-15 pulses, etc.). The nanosecond pulse electric field may have a frequency of between 1-100 Hz (e.g., between 1-90 Hz, between 1-80 Hz, between 1-50 Hz, between 1-40 Hz, between 1-30 Hz, between 5-75 Hz, between 5-50 Hz, between 5-20 Hz, between 5-10 Hz, etc.). [0012] Without being bound by a particular theory of operation, the applied pulsed energy may be sufficient to drive local stimulation of an underlying muscle or nerve within the target area (e.g., within the injection site), but may be insufficient to cause the formation of pores (e.g., poration) that may result from higher nanosecond pulsed energy. Thus, in any of these examples, the applied nanosecond pulsed energy may be limited to prevent damage to the dermis of the target region, including to prevent poration of the tissue (cells, including muscle, nerve and/or dermal cells) in the target region. In any of the methods and apparatuses described herein, the application of the nanosecond pulsed energy as described herein following the waiting time after introduction (e.g., injection) of the toxin (e.g., BoNT), may not result in any significant (and/or detectable) damage to the dermis. The skin in the treatment region may appear substantially unaffected. Typically, redness or irritation does not occur at the levels of nanosecond pulsed energy applied as described herein.

[0013] The methods and apparatuses described herein may be configured to apply the nanosecond pulsed electric field to any appropriate body region. For example, the methods and apparatuses may be configured to apply the nanosecond pulsed electric field to a target area in the skin, such as, but not limited to the face, the neck, the armpits, etc. Applications to the face may be specific to, or may include: the forehead, the region between the eyebrows, the region around the eyes, the “crow’s feet” region, the nasalis, the corners of the mouth, etc. Thus, any of these methods may include applying the nanosecond pulsed electric field to a subject’s face. [0014] In general, any appropriate treatment applicator or applicator (including one or more electrodes) may be used for applying the treatments described herein. For example, any of these methods may include applying the nanosecond pulsed electric field using two or more nonpenetrating electrodes. The treatment applicator may comprise an electrode tip that may be configured to be removably attached to a handle or handpiece portion of the applicator, the applicator may in turn be operably connected to a pulse generator as described herein. The electrode tip may generally include a treatment (skin- or tissue-) contacting surface that may be configured to be applied to the target region or in a proximity to the target regions (for example, if the target region is around eyebrows, the electrode tip may be placed on the forehead, including at the top of the forehead near a hairline). In other words, the nanosecond pulsed electric field may be applied not only to the immediate area where the toxin has been introduced, but to the areas adjacent or in proximity to such area. The electrode tip may include one or more, including a plurality (e.g., two or more) electrodes. As stated above, electrodes may be nonpenetrating or penetrating. Examples of non-penetrating electrodes include surface electrodes, wire electrodes, disc-electrodes, electrode pads, etc. For example, the non-penetrating electrodes may comprise elongate electrodes extending in parallel across a contact surface of the electrode tip. Any of these electrodes may be used with a conductive gel on the surface of the target tissue. [0015] The treatment applicator and/or the electrodes may be adapted for applying energy to the target tissue region (e.g., the face, and/or regions of the face, etc.). For example, the treatment applicator may include two or more non-penetrating electrodes on the electrode tip, wherein each of the non-penetrating electrodes extend 1-20 mm along a contact surface of the electrode tip. The electrode tip may also comprise a conductive spacer positioned between two or more electrodes. In any of these examples the treatment tip may include an array of electrodes. The array of electrodes may define a treatment area of any desired shape and/or size. For example, the array of electrodes may have a longest side or a diameter of between 2.5 mm and 30 mm (e.g. a 2.5 x 2.5 mm square array, a 5x5 mm square array, a 10x10 mm square array, or other shape and sizes). The array may have a treatment area that is approximately square, rectangular, circular, semi-circular, fan-shaped, etc. It may also be configured to cover a relatively large area. [0016] For example, a method may include: applying a nanosecond pulsed electric field to a target area after waiting a waiting time following an injection of a botulinum toxin into the target area of a subject’s face, wherein the nanosecond pulsed electric filed is applied over the injected area from a non-penetrating electrode, thereby reducing the time for activation of the injectable toxin, wherein the nanosecond pulsed electric field comprises one or a series of pulses each having a duration of less than 1 psec and an electric field of 5kV/cm or less.

[0017] Also described herein are apparatuses (e.g., systems) for performing any of the methods described herein. These apparatuses may include a controller comprising control circuitry (one or more processors, one or more memories, one or more user inputs, one or more display outputs, etc.) that may include software, firmware and/or hardware for performing any of these methods. For example, an apparatus (e.g., system) may include: a pulse generator; a treatment applicator comprising one or a plurality of (e.g., non-penetrating) electrodes and configured to couple to the pulse generator; and a controller configured to control the pulse generator to apply a nanosecond pulsed electric field to or in a proximity of a target area into which the toxin has been introduced from the one or a plurality of electrodes, the nanosecond pulsed electric field may comprise one or more pulses each having a duration of less than 1 psec and an electric field of, for example, lOkV/cm or less (5kV/cm or less, 2kV/cm or less).

[0018] In some examples the controller is configured to receive confirmation that the toxin (e.g., injectable toxin) was introduced (e.g., injected) before the application of the nanosecond pulsed electric field, and in some implementations that a specified waiting time has passed (e.g., 10 minutes, 20 minutes, 30 minutes or more). For example, any of these apparatuses may include an input (including a manual input, a user interface, etc.) for the injection time and may warn/alert and/or prevent or block the user from the application of the nanosecond pulsed electric field before the expiration of the waiting time period. The waiting time period may be preset or may be user-set. In some examples the user may select from a plurality of waiting time periods (e.g., 5 min, 20 min, 25 min, 30 min, 45 min, 1 hr., etc.). In some examples the apparatus (e.g., system) includes a user interface into which the user (e.g. doctor, clinician, nurse, etc.) enters or confirms introduction time of the toxin. Alternatively, the apparatus may include an automatic detection of the injection. In some examples the system may tract the procedure, including the injection time and/or location. For example, a user may begin a treatment session with the apparatus which may track the treatment, and/or may include one or more user interfaces for recording what injections were made to what regions of the subject’s body (e.g., face, neck, etc.). The user may indicate in the user interface that an injection was made on a representation of the subject’s body. The system may then alert the user when to apply the nanosecond pulsed electrical energy to a particular region that was treated and may provide visual, audible and/or textural feedback to the user as to when and where the nanosecond pulsed electrical energy may be applied. The system may save (e.g., store) and/or transmit this information, including processing it as part of a subject’s/patient’s health record (e.g., electronic health record). [0019] Any of the applicator tips and/or electrodes (e.g., non-penetrating electrodes) described herein may be used as part of these apparatuses. For example, the plurality of electrodes may comprise elongate electrodes extending in parallel across a contact surface of the electrode tip.

[0020] According to another aspect of the present disclosure, the methods described herein include in particular, cosmetic methods for non-therapeutic applications. These cosmetic methods include methods of improving the appearance of a healthy subject (e.g., a subject’s skin), for example, by removing or reducing wrinkles. For example, the methods described herein include methods of improving the appearance of a subject having wrinkles by applying a nanosecond pulsed electric field to or near a target region of skin after introduction of a botulinum toxin (BoNT) and applying pulsed electrical energy having a pulse duration in a submicrosecond pulse range, e.g., between one or more electrodes to enhance the effect of the BoNT within the target region.

[0021] In some implementations, a cosmetic method of improving cosmetic appearance of a subject is provided. The method comprises reducing time for activation of a toxin by applying a nanosecond pulsed electric field to or in proximity to a target skin surface area of the subject into which the toxin has been introduced, wherein the nanosecond pulsed electric filed is applied over or in the proximity to the target skin surface area from one or more electrodes, and wherein the nanosecond pulsed electric field comprises one or more pulses each having a duration of less than 1 psec and an electric field of lOkV/cm or less.

[0022] In some further implementations, a cosmetic method of improving cosmetic appearance of facial wrinkles is provided. The method comprises: after waiting a waiting time following an introduction of a botulinum toxin into a target skin area of a subject’s face, applying a nanosecond pulsed electric field in the proximity and/or to the target skin area of the subject’s face to reduce the time for activation of the botulinum toxin, wherein the nanosecond pulsed electric field is applied from one or more surface or plate electrodes, and wherein the nanosecond pulsed electric field comprises a series of pulses each having a duration of less than 1 psec and a voltage of 2kV or less.

[0023] As mentioned above, in general the methods and apparatuses described herein may provide a nanosecond pulsed electric field to or near a treatment region after the introduction (e.g., injection) of a toxin (e.g., BoNT) in order to speed up the effect of the toxin. Preliminary results show that healthy subjects who received BoNT in the forehead for cosmetic treatment showed a robust effect from the BoNT (such as smoothing of wrinkles) much more rapidly when they received nanosecond pulsed electric field energy as compared to subjects receiving introduction of toxin without the application of nanosecond pulsed electric field energy. Typically, subjects receiving nanosecond pulsed electric field treatment following BoNT injection took only two days or less to be fully active (smoothing/frozen forehead wrinkle lines), as compared to a week (5-7 days) or more with subjects receiving the same amount and location of BoNT. These methods and apparatuses may be configured and calibrated so as not to damage the dermis, including preventing irritation (redness, swelling, etc.). The atraumatic tips used (typically including two or more surface electrodes) do not cause any epidermal damage, and do not require anesthesia at the target region.

[0024] The methods and apparatuses described herein may be adapted to apply a plurality of pulses having a field strength of, for example, between 500 andlOOOO V (e.g., 800-5000V, less than 2000V, less than 1000V) and sub-microsecond pulse duration (e.g., between 1-900 ns, between 1-850 ns, between 1-800 ns, between 1-750 ns, between 1-700 ns, between 1-650 ns, between 1-600 ns, between 1-550 ns, between 1-500 ns, between 10-500 ns, between 10-450 ns, between 10-400 ns, etc.). Pulses may be applied as a train of between 1-100 Hz (e.g., between 5- 100 Hz, between 5-75 Hz, between 5-50 Hz, between 1-50 Hz, between 2-40 Hz, between 1-30 Hz, etc.). In some examples, pulses may be biphasic, or may be monophasic.

[0025] In any of the methods described herein, the electrodes may be placed into or against the subject’s skin to deliver short electrical pulses to enhance the effectiveness of the toxin. For example, any of these methods may include applying a plurality of electrodes onto or in a vicinity of a target area of a subject’s skin, for example, 15 minutes or more after a toxin was introduced (e.g., topical application, injection, etc.) into a target area. Applying the electrodes may include applying non-penetrating electrodes against the skin. In some examples the skin may be pinched or gripped between two or more electrodes on the surface of the skin.

[0026] Any of the methods and apparatuses described herein may include or be configured for applying the electrical pulses transdermally, without puncturing the skin. For example, any of these methods may include applying the set of electrodes on the surface of the subject’s skin before applying the plurality of short electrical pulses. In such variations one or more conductive or non-conductive gels or other materials may be applied to the skin, including to the electrode contact points and/or the region between them. For example, a non-conductive or lower- conductance gel may be used.

[0027] While in some variations two electrodes may be used, in other examples more than two electrodes (e.g., one or more active electrodes and one or more ground electrodes) may be used. The active electrodes may be coupled together; the ground electrodes may be coupled together.

[0028] The methods and apparatuses described herein may be configured to minimally disrupt the skin tissue. For example, applying the electrical energy may include applying a nonthermal treatment that does not disrupt the cell membrane(s) of the epidermal cells. Note that in some variations one or more additional or accessory therapies or treatments (including cryo, sonic, and/or laser therapy or treatment) may be applied in addition to the electrical energy and toxin injection (the additional treatment may include the application of thermal energy).

[0029] As mentioned, the applied electrical pulses may have any appropriate parameter values (e.g., frequency, pulse width, amplitude, etc.), so long as the energy delivered to the tissue is below the threshold for porating the cells within the tissue. In some applications, the treatment levels may be, for example, between 0.3 mJ to 100 mJ. Any appropriate dose parameter may be used for treatment. For example, the methods may include applying a single treatment dose extending for a treatment time (e.g., less than 1 second, 1 second or less, 2 seconds or less, 3 seconds or less, 4 seconds or less, 5 seconds or less, 6 second or less, 7 seconds or less, 8 seconds or less, 9 seconds or less, 10 seconds or less, 20 seconds or less, 25 seconds or less, 30 second or less, etc. For example, the method may comprise applying treatment for less than 1 minute, less than 45 seconds, less than 40 seconds, less than 30 seconds, less than 25 seconds, less than 20 seconds, etc. In some variations the number of pulses applied during treatment may be between, for example, 1 and 500 (e.g., between 5-200, between 10-150, less than 500, less than 400, less than 300, less than 200, less than 100, less than 50, etc.) For example, the method may include applying less than 100 pulses. In some examples, the devices and methods of the present disclosure may be used for stimulating nerves inside the subject’s body.

[0030] For example, applying the plurality of sub-microsecond electrical pulses may include applying after a waiting time and/or for a predetermined number of pulses (e.g., between 1 and 30 pulses, between 2 and 60 pulses, between 2 and 120 pulses, between 2 and 240 pulses, between 2 and 680 pulses, etc.). The pulses may be applied at any appropriate frequency. For example, the plurality of high-field strength, short electrical pulses may be applied between 1 Hz and 100 Hz (e.g., between 1 Hz and 150 Hz, between 1 Hz and 100 Hz, between 2 Hz and 75 Hz, between about 5 Hz and 100 Hz, between about 5 Hz and 50 Hz, between about 2 Hz and 20 Hz, etc.).

[0031] Repeated dosing may not be necessary, although in some variations additional (repeated) treatments may be applied to the same region of tissue. In some variations the same region of tissue may not be re-treated until after a waiting period of, e.g., between about 30 min- 24 hours, between 30 min- 18 hours, between 30 min- 12 hours between 30 min-8 hours, between 30 min-6 hours, between 30 min-4 hours, between 30 min-3 hours, between 30 min-2 hours, etc. [0032] In general, when applying electric fields treatments, the electrical energy applied may be in the form of one or more electrical pulses. The pulse duration may be at least 0.01 nanoseconds (ns) at the full-width-half-maximum (FWHM). The pulse duration may also be at least 1 ns at FWHM, or the pulse duration may be at least 5 ns at FWHM. The pulse duration may be less than 1000 ns or shorter.

[0033] The number of electrical pulses during a single treatment may be at least 1. The number of pulses may also be at least 10. The number of pulses may be at least 100. The number of pulses may be less than 500 (e.g., less than 450, less than 400, less than 350, less than 300, less than 250, less than 200, etc. For example, the number of pulses may be between 5 and 200, between 5 and 150, between 5 and 100, etc.

[0034] An entire tissue target region may be covered by the applicator including the plurality of electrodes. In some variations, the tissue may be treated by, e.g., dividing the treatment (e.g., treatment dose) into two or more parts, and rotating the applicator tip so that the electrodes apply the energy to the same portion of skin tissue from multiple different rotational orientations during the treatment. Adjacent and/or overlapping regions may be treated in this manner. In some variations, non-overlapping, but adjacent regions may be treated.

[0035] For example, described herein are methods of treating a skin tissue by applying pulsed electrical energy (in some examples comprising a plurality of nanosecond electrical pulses having a pulse duration of between 0.1 ns and 1000 ns) after a waiting time following injection of a toxin (e.g., BoNT) wherein the treatment is divided into different portions in which the same or adjacent regions are treated. The method may include: contacting the tissue with an applicator tip (e.g., treatment tip) having a pattern of electrodes; applying the first portion of the treatment to a region of the tissue over a treatment region that has been injected with the BoNT; and applying additional portions of the treatment to adjacent regions of tissue that have also been injected with the toxin.

[0036] As stated above, any appropriate type of penetrating or non-penetrating electrode may be used, in particular non-penetrating electrodes (e.g., surface electrodes). In some variations the treatment tip includes an array of electrodes. The electrodes may be fixed relative to the distal face of the treatment tip, and may be flush, recessed or may extend proud of the skin-contacting surface of the applicator tip. For example, the electrodes may extend proud by between 0.1 mm and 1 mm (e.g., between 0.1 mm and 0.5 mm, between 0.1 mm and 0.4 mm, between 0.1 mm and 0.3 mm, etc.). In some variations the electrodes may be configured to retract relative to the treatment tip (e.g., retract into the treatment tip).

[0037] In any of the methods described herein, the movement of the electrodes to different regions may be performed manually, semi-manually, or automatically. The movement of the electrodes (e.g., movement of the applicator and/or applicator tip) between different target regions of the skin and/or over the same target region of the skin for treatment may be performed robotically. In addition, any or all of the steps of the methods disclosed herein, including moving, removing and/or reapplying the electrodes, as well as coordinating application of the pulsed electrical energy, may be performed by a robotic system, for example, under computer control.

[0038] Thus, any of these systems may be robotic systems wherein the applicator comprising a treatment tip with one or more (e.g., an array of) electrodes is coupled to a moveable arm. For example, the robotic system may receive instructions from the controller and move the treatment tip to change position relative to the target skin. For example, a system for applying pulsed electrical treatment to tissue may include: a movable arm (e.g., robotic arm); an applicator operatively coupled to the movable arm, the applicator configured to apply pulsed electrical energy from the one or more electrodes of the applicator; and a processor comprising a set of instructions for executing operations, the set of instructions including instructions for: moving the movable arm to contact a region of tissue with the applicator; directing application of the pulsed electrical treatment to the first region of the tissue; directing moving an applicator to a different region within a target area after a waiting time or after a certain number of pulses were performed at the first region of the tissue; and applying a subsequent pulsed electrical treatments to the different region of tissue. The instructions may also comprise an instruction for directing application of the pulsed electrical treatment to the first region of the tissue after waiting a waiting time since a toxin was injected or otherwise introduced into a target area.

[0039] In some examples, the robotic system may include a navigation interface comprising, for example, an image acquisition device and the navigation interface may be configured to receive imaging data and/or determine a path for application of treatment energy) based on the previous injection site. The navigation interface may determine the distance between the tissue (as well as the location of the target treatment site on the tissue) and the plurality of electrodes/treatment tip, and/or the orientation of the plurality of electrodes/treatment tip and the target tissue, to allow control and guidance of the treatment tip relative to the target tissue.

[0040] The applicator may be operably connected to the movable arm, such as held by the movable (e.g., robotic) arm. Alternatively, the applicator may be integrated into the movable arm.

[0041] As mentioned above, described herein are apparatuses (e.g., systems and devices) configured to perform any of these methods. According to some examples, a system may include: an applicator comprising at least one electrode; a pulse generator, wherein the applicator is configured to couple to the pulse generator; and a controller configured to cause the pulse generator to apply a nanosecond pulsed electric field to or in proximity to a target area into which a toxin has been introduced, the nanosecond pulsed electric field comprising a series of pulses each having a duration of less than 1 psec and a voltage of lOkV or less. In some examples, a system may include a pulse generator; an applicator having a plurality of electrodes at a treatment tip of the applicator, the applicator tip configured to apply energy from the pulse generator to the plurality of electrodes; and a controller configured to control, at least partially, operation of the pulse generator and the applicator tip. The controller may comprise a processor having a set of instructions, wherein the set of instructions, when executed by the processor causes the controller to apply the pulsed electrical treatment.

[0042] For example, a system for treating tissue may include: a pulse generator; an applicator configured to apply energy from the pulse generator to a plurality of electrodes at a treatment tip of the applicator, and a controller configured to control, at least partially, operation of the pulse generator and the applicator as described herein. The controller may include a processor having a set of instructions, wherein the set of instructions, when executed by the processor, causes the controller to apply the pulsed electrical treatment. In some variations multiple controllers may be included as part of the system. The controller may be integrated with the other parts of the system (e.g., the applicator and/or pulse generator, etc.). For example, in some variations the controller (including one or more processors) may be housed within a housing to which all or a portion of the pulse generator is also enclosed. Alternatively, in some variations the controller and/or one or more processors may be remote to the pulse generator. [0043] Although the majority of the embodiments descried here are specific to nonpenetrating electrodes, in some variations the electrode may be tissue-penetrating (e.g., needle electrodes, knife electrodes, etc.).

[0044] All of the methods and apparatuses described herein, in any combination, are herein contemplated and can be used to achieve the benefits as described herein.

BRIEF DESCRIPTION OF THE DRAWINGS

[0045] A better understanding of the features and advantages of the methods and apparatuses described herein will be obtained by reference to the following detailed description that sets forth illustrative embodiments, and the accompanying drawings of which:

[0046] FIG. 1 is an example of a system for generation and delivering sub-microsecond electrical field pulses, e.g., pulses in the nanosecond range.

[0047] FIG. 2A is an example of a system for generation and delivering sub-microsecond electrical field pulses as described herein. FIG. 2B shows an example of a handpiece for a system such as the one shown in FIG. 2A. FIG. 2C is an example of one variation of an applicator tip or electrode tip as described herein.

[0048] FIG. 3 A shows an example of an applicator tip as described herein. FIG. 3B shows an enlarged view of the applicator tip, including the non-penetrating electrodes on the tissuecontacting surface shown in FIG. 3 A.

[0049] FIGS. 4A-4B show schematic overviews of some examples of methods as described herein.

[0050] FIG. 5 is a histological section through a region of treated tissue as described herein; the dermal tissue is not modified by the application of the sub-microsecond electrical field pulses applied as described herein.

DETAILED DESCRIPTION

[0051] Described herein are methods and apparatuses (including devices, systems, etc.) for treating a target region (or regions) that has been previously injected with a toxin, such as a botulinum toxin, with a sub-microsecond pulsed electrical field in order to enhance the effect of the toxin on the target tissue. Some examples of the toxins include but are not limited to the commercial forms that are marketed under the brand names Botox™ (onabotulinumtoxinA), Dysport™/Azzalure™ (abobotulinumtoxinA), Myobloc™ (rimabotulinumtoxinB), Xeomin™/Bocouture™ (incobotulinumtoxinA), and Jeuveau™ (prabotulinumtoxinA) just to name a few. In particular, the therapeutic and/or cosmetic effects of a toxin, such as botulinum toxin (BoNT) may be enhanced by the methods and apparatuses described herein. For example, enhancing the effects of a BoNT introduction (e.g., injection) may include enhancing a reduction (including elimination) in wrinkles in the skin of the target region, and/or the reduction in the tonic muscle activity and/or reduction in gland activity (e.g., sweating).

[0052] The methods and apparatuses for performing these methods may include or be used in conjunction with an application (e.g., an injection) of a toxin such as BoNT. Any of methods may include, for example, following injection of a target region of tissue with an injectable toxin, and after waiting for a waiting time period, non-invasively applying sub-microsecond (e.g., nanosecond), pulsed electrical field energy to all or a portion of the treatment region of the tissue injected with the toxin. The sub-microsecond pulsed electrical energy may be applied for a certain period of time (that may be referred to as an activation period) so that energy delivered activates the underlying muscle and/or nerve but does not create any epidural damage or effect and does not result in pain. Specifically, the applied energy may be below the level that would result in forming pores in the cells within the treated region. In some examples, the applied energy may be sensed by the subject (e.g., patient) receiving the treatment, and may be felt as a contraction of the muscle, a tingling sensation, etc. In some examples the applied energy may not be detected by the subject and may be below the level for sensory detection.

[0053] The methods described herein have been shown to reduce the time required for an injected BoNT toxin to achieve robust, full effect by more than half. Without any applied submicrosecond pulses to a region of skin (such as the forehead or other facial region) treated by injection of BoNT to reduce wrinkles may require more than 5-7 days to achieve full effect. The application of sub-microsecond pulsed electrical energy after waiting for a wait period (e.g., 30 minutes or more) to a region of the forehead injected with BoNT has been found to result in accelerating the time for the injected region achieving robust effect of the toxin, for example, reducing wrinkles within 2-3 days, or faster.

[0054] For example, described herein are methods (e.g., methods of enhancing the effect of a toxin such as BoNT) by applying sub-microsecond, e.g., nanosecond, electrical field pulses after introduction/inj ection of toxin to a treatment area. The applied sub-microsecond pulses may be configured so as to not result in any significant epidermal effect on its own (e.g., in the absence of a previously injected toxin), including avoiding irritation (redness, swelling, pain, etc.).

[0055] The applied sub-microsecond pulsed electrical field energy may be applied so that the pulses (which maybe monophasic, positive or negative, or biphasic) have a pulse width of less than 1000 ns (e.g., between 1-999 ns, between 1-950 ns, between 1-900 ns, between 1-850 ns, between 1-800 ns, between 1-750 ns, between 1-700 ns, between 1-650 ns, between 1-600 ns, between 1-550 ns, between 1-500 ns, between 1-450 ns, between 1-400 ns, less than 1000 ns, less than 900 ns, less than 800 ns, etc.). Each of the pulses typically have a voltage of less than 2000V (e.g., between 500 V-2000V, between 700 V and 2000V, etc.). The pulses may be delivered at a frequency of between 1-100 Hz, such as between 1-50 Hz, between 1-40 Hz, between 1-30 Hz, between 1-20 Hz, between 5-100 Hz, between 5-50 Hz, between 5-30 Hz, between 5-20 Hz, etc.). Between 2-100 pulses may be applied (e.g., between 5-75 pulses, between 5-50 pulses, between 5-30 pulses, between 5-20 pulses, between 1-15 pulses, etc.). [0056] In general, the applied sub-microsecond pulses may be applied following injection of the toxin into the target tissue (e.g., skin, such as the skin of the forehead or around the eyes, nose and/or mouth, neck, armpits, etc.). In some implementations, the method or apparatus may be configured to include a waiting time period from injection of or application of a toxin to a treatment site before applying the enhancing sub-microsecond pulsed electrical energy. For example, the waiting time period may be at least about 5 minutes, at last about 20 minutes, at least about 30 minutes, at least about 40 minutes, at least about 45 minutes, at least about 50 minutes, at least about 55 minutes, at least about 1 hour, at least about 1.25 hours, at least about 1.5 hours, at least about 1.75 hours, at least about 2 hours, etc. In any of the methods and apparatuses described herein the wait period may be between, e.g., 10 min and 3 hours (e.g., 30 min-2 hours, 30 min - 1.5 hours, etc.). The waiting time period may be predetermined (e.g., may correspond to a predetermined time period). The waiting time period may be set by the user, e.g., doctor, clinician, nurse, technician, etc., or it may be automatically selected by the system (for example, based on some historical data or available tables). The waiting time period may be determined by one or more factors, such as one or more of: the amount of toxin applied or injected, the location of the injection region, the age, weight and/or gender of the subject, and/or the type of toxin.

[0057] As will be described in detail below, any size or shape of applicator and/or electrodes may be used. For example, the applicator may include an array of 2 or more electrodes that are arranged on the tissue-contacting surface of the applicator. These electrodes may be nonpenetrating, e.g., surface, electrodes. Non-penetrating electrodes are configured so as not to penetrate into the tissue, but to apply energy from the surface of the tissue. Examples of nonpenetrating electrodes include wire electrodes, coil electrodes, plate electrodes, disc electrodes, etc. The electrodes may be arranged on the surface of the tissue-contacting surface of the applicator tip, recessed slightly into the tissue-contacting surface (and/or surrounded by an annular lip or rim) and/or extending or extendable proud of the tissue-contacting surface. The electrodes may be flat. The electrodes may be formed of a conductive material, such as a metal or electrically conductive polymer. The tissue contacting surface of the electrode tip may be formed of an electrically insulating material, which may be soft or compliant (e.g., such as silicone). In some implementations the tissue contacting surface may include one or more passages for applying a vacuum to form a seal against the tissue (e.g., skin) so that electrical contact may be sustained against the tissue during the application of energy.

[0058] In general, described herein are methods and apparatuses for treating tissue (e.g., skin) to enhance the effect of a toxin, including BoNT. In particular, the methods and apparatuses (e.g., devices, systems, etc.) described herein may apply pulsed (e.g., submicrosecond, such as nanosecond) electric pulses to the target region of the tissue to which toxin has previously been introduced. The methods described herein may enhance the effect of the toxin without increasing dermal turnover, or otherwise irritating the skin.

[0059] The methods and apparatuses described herein may be used to treat, for example, skin tissue by generally applying a treatment, e.g., a pulsed electrical treatment, to the skin. Without being bound by a particular theory of operation, the sub-microsecond pulsing following treatment may drive activity in the target muscles and/or nerves into which the toxin has already been (or begun) being taken up. The non-thermal treatment employed in the methodology described herein is typically an electrical treatment (e.g., very short, relatively high-field strength electric pulses, typically in the sub-microsecond range) adapted to enhance toxin activity. These pulses may affect the target tissue without provoking an inflammatory response (e.g., without increasing the density of leukocytes and/or melanocytes above a threshold percentage compared to untreated skin) and without leading to poration and/or turnover of dermal cells overlying or surrounding the nerves, glands, and/or muscles within the skin.

[0060] Illustrative embodiments are now discussed. Other embodiments may be used in addition or instead. Details which may be apparent or unnecessary may be omitted to save space or for a more effective presentation. Conversely, some embodiments may be practiced without all of the details which are disclosed, or various features shown in one example may be combined with the features of a different example, as appropriate.

[0061] In any of the methods described herein, the pulsed electrical treatment may be nanosecond electric pulsed treatment, which may include the application of electrical pulses with duration of less than 1,000 nanoseconds (ns). Although the examples described herein focus primarily on pulses having a width (e.g., pulse width) within the nanosecond range, other pulse widths may be used. For example, in some variations, pulses may have pulse widths in picosecond ranges.

[0062] The pulsed electrical treatment may be achieved by providing electrical energy to the target skin region in a form of one or more electrical pulses. Further, these methods may be generally non-thermal, and may be configured to prevent a substantial inflammatory response. [0063] The treatment may comprise at least one treatment session. For example, the treatment session may comprise an administration of the electrical energy to the skin region of a subject by physician at an office visit. The treatment of a region with the sub-microsecond electric pulses may follow shortly (e.g., within a wait period) injection of toxin to the region. The applied sub-microsecond pulsed electrical field energy may be applied to the same (overlapping region) of the tissue that was injected. The region of the toxin introduction (e.g., toxin injection region) and the region to which the sub-microsecond pulses are applied may overlap, but they need not perfectly overlap. In some examples the region of the tissue that the sub-microsecond pulses of electrical energy are applied is only approximately coextensive with the injected region of the toxin. The injected region may be larger or smaller than the target region to which the submicrosecond pulses of electrical energy are applied. For example, in practice at least 40% (e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, etc.) of the region of the skin injected with the toxin may be treated by applying sub-microsecond pulses of energy. In some examples, all or a portion (e.g., 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, etc.) of the injection sites (the regions in which the toxin was injected into the skin) overlap with the region of the tissue to which the sub-microsecond pulsed electrical energy is applied.

[0064] Any system suitable for delivery of sub-microsecond pulsed electrical energy within the target energy parameters may be used. A pulse generator may be any pulse generator that is capable of generating pulses, for example, with a duration of less than 1,000 ns. The pulse delivery device may be any device that can deliver electrical pulses to the skin. This device may have an applicator tip that may comprise at least one or a plurality of electrodes for both bi-polar and monopolar applications. In some embodiments, additional electrodes may be electrically floating and may be switched to become active, as desired. This applicator may comprise at least one ground electrode.

[0065] For example, a nanosecond pulse generator system such as those shown and described in US2017/0245928 Al titled “HIGH-VOLTAGE ANALOG CIRCUIT PULSER WITH FEEDBACK CONTROL”, which is incorporated herein by reference in its entirety, may be adapted for use as described herein. The pulse generator system may provide pulses having a duration of 1,000 ns or less to the skin. The system may comprise a power supply, a controller, a pulse generator, and a pulse delivery device (e.g., a wand, or treatment applicator). An example of this system is schematically shown in FIG. 1. FIG. 1 illustrates one example of a nanosecond pulse generator system (NsPEF system) 100. The pulse generator system 100 includes a treatment applicator 102, footswitch 103, and interface 104. Footswitch 103 is connected to housing 105 and the electronic components therein through connector 106. Electrodes on the treatment applicator 102 in this example are electrically connected to the pulse generator within the housing 105 and the corresponding electronic components therein through high voltage connector 112. NsPEF system 100 also includes a handle 110 and storage drawer 108. As shown in DETAIL A portion of FIG. 1, nsPEF system 100 may also include a holster 116, which is configured to hold an exemplary treatment applicator 102 at its handle portion 114.

[0066] A human operator may interact with a user interface (e.g., shown on monitor 104) and may input control commands. The user may select a treatment paradigm including or specifying (or the user may optionally specify) a number of pulses, amplitude, pulse duration, and frequency information, for example, into a numeric keypad or a touch screen of interface 104, and/or some or all of these parameters may be automatically determined based on a target treatment protocol. In some embodiments, the pulse width can be varied. A controller may send signals to pulse control elements within nsPEF system 100. In some embodiments, fiber optic cables allow control signaling while also electrically isolating the contents of the metal cabinet with nsPEF generation system 100, the high voltage circuit, from the outside. In order to further isolate the system, system 100 may be battery powered instead of from a wall outlet.

[0067] The applicator may include or be coupled to a treatment applicator 102 having one or more (e.g., a plurality) of electrodes. The system may generally include a controller 121. The controller may control operation of the system, and may include one or more processors, one or more memories, and the like. In some examples, one or more processors may be separate from the controller. The controller and/or one or more processors may include logic (e.g., hardware, software, firmware) including instructions that, when executed by the one or more processor(s), may control the system to apply the electrical treatment as described herein. For example, the set of instructions may operate a robotic actuator (e.g., robotic arm) to move the treatment electrodes to the target tissue region and/or control the application of pulsed electrical energy treatment to the tissue. The set of instructions may include instructions controlling the application of the pulses, movement of electrodes applying the energy, and/or placement of the applicator on/off of the tissue. In some variations, the applicator may control the application of pulsed electrical energy to enhance toxin previously injected into the target skin tissue.

[0068] In some examples the apparatus may be adapted to allow the user or a person assisting the user, to enter (manually and/or automatically) the sites of injection or introduction of toxin into the treatment area. The apparatus may therefore monitor the time the injection/introduction occurred and track the waiting time period to indicate when the pulsed electrical energy should be applied and/or the locations it should be applied.

[0069] The electrical energy may be applied to the skin in the form of at least one electrical pulse. For example, between 1 and 500 pulses may be applied (e.g., between 5 and 100, between 5-50, etc.). In one embodiment, at least 10 pulses, at least 100 pulses, etc., may be applied to treat the skin during a single treatment. The treatment may be triggered by activation of a control on the handle portion or handpiece and/or input(s) of the system. The duration of one or more of the pulses may be set to be in the target range, e.g., between 1 ns to 1,000 ns. For example, the pulse width may be between 10 and 900 ns (e.g., between 200 and 800 ns, etc.). The duration of one or more of the pulses may generally be in the sub-microsecond range.

[0070] FIG. 2A shows another example of a system as described herein, similar to the system shown in FIG. 1. In this example, the system includes a console 200 including a monitor (204; e.g., touchscreen) and a handle or handpiece 216 coupled to the electronics (e.g., pulse generator) housed within the housing 221. The handpiece/handle 216 in FIG. 2A is shown coupled to a treatment tip 202. FIG. 2B shows an enlarged view of the handpiece 216 and FIG. 2C shows an example of a treatment tip 202. In this example, the treatment tip is adapted to engage with and releasably lock onto the handpiece so that electrical and mechanical connection is made between the two. In other examples (not shown) the handle portion and the tip may be configured to be a one-piece unitary treatment applicator.

[0071] FIG. 3 A shows an enlarged view of one example of an applicator tip 300. In this example the applicator tip includes two non-penetrating electrodes 355, 356 that extend slightly proud of a tissue-contacting side 359 of an electrode housing 357. The tissue-contacting side between the electrodes 355, 356 may comprise a conductive spacer as described in the PCT application PCT/US2022/024283 entitled “Conductive Spacer in an Electrode Assembly of an Electrical Treatment Apparatus,” which is also owned by the Applicant of the present disclosure and incorporated herein by reference in its entirety. In some embodiments, the conductive spacer may be a hydrogel, a conductive adhesive, a conductive gel, a conductive silicone, a urethane rubber, conductive thermoset, thermoplastic resins, any other biocompatible material with the desired conductivity, any semi-conductor material, or any combination thereof. In some embodiments, a conductivity of the conductive spacer may be substantially equal to ten times (lOx) a conductivity of a tissue of the treatment area, however, in various examples, it may be less, equal or greater than a conductivity of a tissue of the treatment area. In some embodiments, a height of the conductive spacer may be based on a distance between the electrodes. In some embodiments, a height of the conductive spacer may be greater than twenty percent (20%) or more of a distance between the electrodes. The electrodes 355, 356 may be further separated by additional spacers (not shown) that may be either insulators (in some examples) or may be also conductive. It has been discovered by the inventors of the present disclosure that in some examples it may be beneficial for the efficacy of the electric field to have two conductive spacers separated by an insulative (or less conductive) spacer. If the additional spacers are also conductive spacers, they may have conductivity that is different (e.g., lower) from the conductivity of the first conductive spacers. In the example of FIG. 3 A, the electrode housing 357 extends from a tip housing 360. In some examples, the electrode housing may extend or retract into the tip housing and may be biased to extend but may be pushed against the tissue when engaging with the tissue. In some examples, there tip housing and electrode housing may be a single unitary housing. The system may detect pressure against the electrode housing. In some examples, activation of the system may be controlled in part by detecting pressure against the tissue.

[0072] In the example of the applicator tip 300 shown in FIGS. 3 A and 3B, the nonpenetrating electrodes are each approximately 10 mm long and about 1.5 mm wide. The applicator tip may form an array of electrodes (e.g., two electrodes) that has an area, for example, of approximately 10 mm x 10 mm. As shown, the electrodes are separated (center line to center line) by approximately 10 mm, and approximately 11.5 mm wide (outside edge to outside edge). All of the dimensions shown are illustrative only; other dimensions may be used. [0073] In FIGS. 3A and 3B the electrodes extend slightly proud of the tissue-contacting surface 359, e.g., by about 0.25 mm. The edges of the electrode are smoothed to prevent damage to the tissue.

[0074] Any of the applicator tips described herein, including the tip 300 shown in FIGS. 3 A and 3B may include a mechanical attachment 365 that may reliably attach (e.g., clip, snap, etc.) onto a wand or handpiece (as shown in FIGS. 1 and 2A), and may additionally include one or more electrical attachment(s) 370 for coupling the electrodes to the pulse generator.

[0075] The tip and associated electrodes shown in FIGS. 3A-3B illustrate just one example of electrodes that may be used with the methods and apparatuses described herein. Other electrodes may be integrated into similar applicator tips, or different applicator tips may be used. For example, in some variations the same applicator tip may be integrated and reusable with the handpiece. Alternatively, the applicator tip may be single-use or single-subject use. It should be understood that the terms “handle portion” or “handpiece” as used herein is intended to describe a proximal portion of the treatment applicator and is not limiting. It refers to any structure to support, hold or attach to the electrode portion of the applicator, whether it is intended to be hand-held, or attached to the robotic arm. In some examples the handpiece may be configured to be hand-held and may include a manual grip. In some examples, the handpiece may be configured to be held by a robotic manipulator (e.g., arm, etc.).

[0076] FIG. 4A illustrates one example of a method as described herein. In practice the method may include the optional step of introducing a toxin, for example, by injection (e.g., BoNT) into a target area (step 401). The toxin may be injected in one or multiple overlapping or non-overlapping regions (injection regions), for example, in the subject’s skin. For example, BoNT injections may be made on the skin of the forehead, around the eyes (crow’s feet), around the mouth, around the nose (nasalis), between the eyebrows, etc. In some examples the injection location and/or time may be recorded, including recording (manually and/or automatically) on the apparatus.

[0077] Following injection, optionally, the user may wait for a wait time (optional step 403). As discussed above, the wait time may be, for example, several minutes or more (e.g., between 30 min-4 hours, between 30 min-2hours, etc.). The wait time may be calculated by the system. The wait time may be based on the location of the injection(s) and/or the concentration (e.g., amount) of the toxin injected, and/or one or more subject characteristics (e.g., age, weight, skin type, etc.).

[0078] The applicator tip may then be applied to the treatment site to be treated (step 405). As mentioned, the treatment site may be overlapping with the toxin introduction (e.g., injection) site(s) or it may be adjacent or in proximity to the toxin introduction site. The electrodes of the applicator tip may be applied against the treatment site. In any of these examples a conductive gel may be used to make the electrical contact with the subject skin and the electrodes. In some examples suction may be applied to make and/or seal the electrode in contact with the skin at the site(s) to be treated.

[0079] The sub-microsecond pulsed electrical energy may be applied to the skin from the electrodes (step 407), as described above. The user may trigger activation of the energy using the handpiece (or wand), or other input (e.g., foot petal, touchscreen, etc.). The system may calculate and deliver the dose once the trigger is made. The system may provide feedback on the application of the dose (e.g., indicating successful delivery of energy, etc.), location of current or next dose, etc.).

[0080] These steps may be repeated as necessary (optional step 409). As mentioned, in some examples the injection sites may be fully overlapped by the sub-microsecond pulsed energy delivery sites (treatment sites), or a subset of the injection sites may be treated.

[0081] FIG. 4B schematically illustrates another example of a method of applying submicrosecond pulses in conjunction with the application of a botulinum toxin into a subject’s skin, which may be performed as part of a cosmetic procedure. For example, in FIG. 4B, the method may include waiting for a wait time following the introduction or application of a botulinum toxin to a target region of a subject’s body (e.g., face) 411. Any appropriate wait time may be used, for example the wait time may be several minutes or more (e.g., between 30 min-4 hours, between 30 min-2hours, etc.). The wait time may be calculated by the system. The wait time may be based on the location of the application(s) and/or the concentration (e.g., amount) of the toxin, and/or one or more subject characteristics (e.g., age, weight, skin type, etc.).

[0082] Immediately following the wait time, a nanosecond pulsed electrical field may be applied to the subject in the proximity of and/or to the target region of the subject’s body (e.g., face), to reduce the time for activation of the botulinum toxin 413. The step of waiting and applying sub-microsecond (e.g., nanosecond) pulsing may be repeated to treat all of the desired areas of the subject’s skin (e.g., the subject’s face) 415.

[0083] Any of the methods (including user interfaces) described herein may be implemented as software, hardware or firmware, and may be described as a non-transitory computer-readable storage medium storing a set of instructions capable of being executed by a processor (e.g., computer, tablet, smartphone, etc.), that when executed by the processor causes the processor to control perform any of the steps, including but not limited to: displaying, communicating with the user, analyzing, modifying parameters (including timing, frequency, intensity, etc.), determining, alerting, or the like.

[0084] FIG. 5 illustrates a section through tissue treated as described herein. The image shows epidermis on the top, which appears as a healthy-looking epidermal cell layer, and underlying dermis. The application of sub-microsecond pulses as described above, did not damage the epidermal or dermal layers.

[0085] It should be appreciated that all combinations of the foregoing concepts and additional concepts discussed in greater detail below (provided such concepts are not mutually inconsistent) are contemplated as being part of the inventive subject matter disclosed herein and may be used to achieve the benefits described herein.

[0086] When a feature or element is herein referred to as being "on" another feature or element, it can be directly on the other feature or element or intervening features and/or elements may also be present. In contrast, when a feature or element is referred to as being "directly on" another feature or element, there are no intervening features or elements present. It will also be understood that, when a feature or element is referred to as being "connected", "attached" or "coupled" to another feature or element, it can be directly connected, attached or coupled to the other feature or element or intervening features or elements may be present. In contrast, when a feature or element is referred to as being "directly connected", "directly attached" or "directly coupled" to another feature or element, there are no intervening features or elements present. Although described or shown with respect to one embodiment, the features and elements so described or shown can apply to other embodiments. It will also be appreciated by those of skill in the art that references to a structure or feature that is disposed "adjacent" another feature may have portions that overlap or underlie the adjacent feature. [0087] Terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. For example, as used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise. It will be further understood that the terms "comprises" and/or "comprising," when used in this specification, specify the presence of stated features, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, steps, operations, elements, components, and/or groups thereof. As used herein, the term "and/or" includes any and all combinations of one or more of the associated listed items and may be abbreviated as "/".

[0088] Spatially relative terms, such as "under", "below", "lower", "over", "upper" and the like, may be used herein for ease of description to describe one element or feature's relationship to another element(s) or feature(s) as illustrated in the figures. It will be understood that the spatially relative terms are intended to encompass different orientations of the device in use or operation in addition to the orientation depicted in the figures. For example, if a device in the figures is inverted, elements described as "under" or "beneath" other elements or features would then be oriented "over" the other elements or features. Thus, the exemplary term "under" can encompass both an orientation of over and under. The device may be otherwise oriented (rotated 90 degrees or at other orientations) and the spatially relative descriptors used herein interpreted accordingly. Similarly, the terms "upwardly", "downwardly", "vertical", "horizontal" and the like are used herein for the purpose of explanation only unless specifically indicated otherwise.

[0089] Although the terms “first” and “second” may be used herein to describe various features/elements (including steps), these features/elements should not be limited by these terms, unless the context indicates otherwise. These terms may be used to distinguish one feature/element from another feature/element. Thus, a first feature/element discussed below could be termed a second feature/element, and similarly, a second feature/element discussed below could be termed a first feature/element without departing from the teachings of the present invention.

[0090] Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise”, and variations such as “comprises” and “comprising” means various components can be co-jointly employed in the methods and articles (e.g., compositions and apparatuses including device and methods). For example, the term “comprising” will be understood to imply the inclusion of any stated elements or steps but not the exclusion of any other elements or steps.

[0091] In general, any of the apparatuses and methods described herein should be understood to be inclusive, but all or a sub-set of the components and/or steps may alternatively be exclusive and may be expressed as “consisting of’ or alternatively “consisting essentially of’ the various components, steps, sub-components or sub-steps.

[0092] As used herein in the specification and claims, including as used in the examples and unless otherwise expressly specified, all numbers may be read as if prefaced by the word "about" or “approximately,” even if the term does not expressly appear. The phrase “about” or “approximately” may be used when describing magnitude and/or position to indicate that the value and/or position described is within a reasonable expected range of values and/or positions. For example, a numeric value may have a value that is +/- 0.1% of the stated value (or range of values), +/- 1% of the stated value (or range of values), +/- 2% of the stated value (or range of values), +/- 5% of the stated value (or range of values), +/- 10% of the stated value (or range of values), etc. Any numerical values given herein should also be understood to include about or approximately that value unless the context indicates otherwise. For example, if the value " 10" is disclosed, then "about 10" is also disclosed. Any numerical range recited herein is intended to include all sub-ranges subsumed therein. It is also understood that when a value is disclosed that "less than or equal to" the value, "greater than or equal to the value" and possible ranges between values are also disclosed, as appropriately understood by the skilled artisan. For example, if the value "X" is disclosed the "less than or equal to X" as well as "greater than or equal to X" (e.g., where X is a numerical value) is also disclosed. It is also understood that the throughout the application, data is provided in a number of different formats, and that this data, represents endpoints and starting points, and ranges for any combination of the data points. For example, if a particular data point “10” and a particular data point “15” are disclosed, it is understood that greater than, greater than or equal to, less than, less than or equal to, and equal to 10 and 15 are considered disclosed as well as between 10 and 15. It is also understood that each unit between two particular units are also disclosed. For example, if 10 and 15 are disclosed, then 11, 12, 13, and 14 are also disclosed.

[0093] Although various illustrative embodiments are described above, any of a number of changes may be made to various embodiments without departing from the scope of the invention as described by the claims. For example, the order in which various described method steps are performed may often be changed in alternative embodiments, and in other alternative embodiments one or more method steps may be skipped altogether. Optional features of various device and system embodiments may be included in some embodiments and not in others, and some feature(s) described in reference to one example may be incorporated in the other provided examples. Therefore, the foregoing description is provided primarily for exemplary purposes and should not be interpreted to limit the scope of the invention as it is set forth in the claims. [0094] The examples and illustrations included herein show, by way of illustration and not of limitation, specific embodiments in which the subject matter may be practiced. As mentioned, other embodiments may be utilized and derived there from, such that structural and logical substitutions and changes may be made without departing from the scope of this disclosure. Such embodiments of the inventive subject matter may be referred to herein individually or collectively by the term “invention” merely for convenience and without intending to voluntarily limit the scope of this application to any single invention or inventive concept, if more than one is, in fact, disclosed. Thus, although specific embodiments have been illustrated and described herein, any arrangement calculated to achieve the same purpose may be substituted for the specific embodiments shown. This disclosure is intended to cover any and all adaptations or variations of various embodiments. Combinations of the above embodiments, and other embodiments not specifically described herein, will be apparent to those of skill in the art upon reviewing the above description.

Claims

CLAIMS What is claimed is:

1. A cosmetic method of improving cosmetic appearance of a subj ect, the method comprising: reducing time for activation of a toxin by applying a nanosecond pulsed electric field to or in proximity to a target skin surface area of the subject into which the toxin has been introduced, wherein the nanosecond pulsed electric filed is applied over or in the proximity to the target skin surface area from one or more electrodes, and wherein the nanosecond pulsed electric field comprises one or more pulses each having a duration of less than 1 psec and an electric field of lOkV/cm or less.

2. The method of claim 1, wherein the toxin is botulinum toxin.

3. The method of claims 1 or 2, wherein the one or more electrodes are surface or plate electrodes.

4. The method of any one of claims 1 to 3, the method comprising applying the nanosecond pulsed electric field after waiting a waiting time following an introduction of the toxin.

5. The method of claim 4, wherein the waiting time is 10 minutes or more.

6. The method of any one of claims 1 to 5, wherein the nanosecond pulse electric field comprises between 1 and 200 pulses.

7. The method of any one of claims 1 to 6, wherein the nanosecond pulse electric field has a frequency of between 1-100 Hz.

8. The method of any one of claims 1 to 7, wherein reducing the time for activation of the toxin comprises reducing the time by more than 40% compared to the target skin surface area into which the toxin has been introduced without a follow-up application of the nanosecond pulsed electric field.

9. The method of any one of claims 1 to 8, wherein applying the nanosecond pulsed electric field comprises applying to a skin surface area of the subject’s face.

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10. The method of any one of claims 1 to 9, wherein the nanosecond pulsed electric field is applied using two or more surface or plate electrodes on an electrode tip, and wherein each of the non-penetrating electrodes extends from 1 to 20 mm along a contact surface of the electrode tip.

11. The method of any one of claims 1 to 10, the method comprising applying the electrical pulses transdermally.

12. A method of improving cosmetic appearance of facial wrinkles, the method comprising: after waiting a waiting time following an introduction of a botulinum toxin into a target skin area of a subject’s face, applying a nanosecond pulsed electric field in the proximity and/or to the target skin area of the subject’s face to reduce time for activation of the botulinum toxin, wherein the nanosecond pulsed electric field is applied from one or more surface or plate electrodes, and wherein the nanosecond pulsed electric field comprises a series of pulses each having a duration of less than 1 psec and a voltage of 2kV or less.

13. A method, the method comprising: applying, a nanosecond pulsed electric field to or in proximity to a target area into which a toxin has been introduced, wherein the nanosecond pulsed electric filed is applied over or in the proximity to the target area from one or more electrodes, thereby enhancing effect of the toxin, wherein the nanosecond pulsed electric field comprises one or more pulses each having a duration of less than 1 psec and an electric field of lOkV/cm or less.

14. The method of claim 13, wherein the toxin is botulinum toxin.

15. The method of claims 13 or 14, further comprising injecting the toxin into the target area.

16. The method of any one of claims 13 to 15, wherein enhancing the effect of the toxin comprises reducing time for activation of the toxin.

17. The method of any one of claims 13 to 16, wherein the one or more electrodes are nonpenetrating electrodes.

18. The method of any one of claims 13 to 17, the method comprising applying the nanosecond pulsed electric field after waiting a waiting time following an introduction of the toxin.

19. The method of any one of claims 13 to 18, wherein the nanosecond pulse electric field comprises between 1 and 200 pulses.

20. The method of claim 18, the method comprising setting a timer for the waiting time.

21. The method of any one of claims 1 to 20, wherein the nanosecond pulse electric field had a frequency of between 1-100 Hz.

22. The method of claims 18 or 20, wherein the waiting time is 10 minutes or more.

23. The method of claim 22, wherein the waiting time is between 30 minutes and 10 hours.

24. The method of claim 16, wherein reducing the time for activation of the toxin comprises reducing the time by more than 40% compared to the target area into which the toxin has been introduced without a follow-up application of the nanosecond pulsed electric field.

25. The method of any one of claims 13 to 24, wherein applying the nanosecond pulsed electric field comprises applying to a subject’s face.

26. The method of any one of claims 13 to 25, wherein applying the nanosecond pulsed electric field comprises applying using two or more non-penetrating electrodes on an electrode tip, and wherein each of the non-penetrating electrodes extend 1-20 mm along a contact surface of the electrode tip.

27. The method of any one of claim 1 to 26, wherein the method is performed by a computer- controlled system.

28. A system, the system comprising: an applicator comprising at least one electrode; a pulse generator, wherein the applicator is configured to couple to the pulse generator; and a controller configured to cause the pulse generator to apply a nanosecond pulsed electric field to or in proximity to a target area into which a toxin has been introduced, the nanosecond pulsed electric field comprising a series of pulses each having a duration of less than 1 psec and a voltage of lOkV or less.

29. The system of claim 28, wherein the controller is configured to receive a confirmation that the toxin was introduced at a specified time.

30. The system of claim 29, wherein the controller is further configured to prevent application of the nanosecond pulsed electric field before expiration of a waiting time after introduction of the toxin.

31. The system of any one of claims 28 to 30, wherein the system is configured to provide visual, audible and/or textural feedback for a timing and a location of an application of the nanosecond pulsed electric field.

32. The system of any one of claims 28 to 31, wherein the at least one electrode comprises a plurality of non-penetrating electrodes extending in parallel across a contact surface of an electrode tip, and the applicator comprises a conducting spacer between at least two of the plurality of non-penetrating electrodes.

33. The system of any one of claims 28-32, wherein each of the at least one electrode extends between 1 and 20 mm along a contact surface of the applicator.

34. The system of any one of claims 28-33, wherein the at least one electrode comprises an array of non-penetrating electrodes, and further wherein the array of non-penetrating electrodes has a longest side or a diameter of between 2.5 mm and 30 mm.

35. The system of any one of claims 28-34, wherein the system is a robotic system.

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