TW201938214A - Methods and apparatus for the delivery of hepatitis B virus (HBV) vaccines - Google Patents

Abstract

Methods and apparatus for the reproducible, consistent and efficacious delivery of an HBV vaccine to a subject. The disclosure comprises apparatus for the controlled administration of the HBV vaccine through an orifice to the subject, a plurality of penetrating electrodes arranged with a predetermined spatial relationship relative to the orifice, and an electrical signal generator operatively connected to the electrodes.

Description

Method and device for delivering hepatitis B virus (HBV) vaccine

The present invention is directed to the administration of preventive and / or therapeutic hepatitis B virus (HBV) to individuals in need, and more specifically, to provide a safe and effective treatment among heterogeneous recipient groups with minimal user training. In a consistent manner, preventive and / or therapeutic HBV vaccines, such as nucleic acids encoding HBV antigens, are reproducibly, consistently, and effectively delivered to a limited area of a selected relevant tissue site, the delivery being facilitated by a local application of an electric field.

Hepatitis B virus (HBV) is a 3.2 kb hepatotropic small DNA virus that encodes four open reading frames and seven proteins. About two billion people are infected with HBV, and about 240 million people have chronic hepatitis B infection (chronic HBV), which is characterized by the persistent presence of viruses and subviral particles in the blood for more than 6 months ( Cohen et al., J. Viral Hepat . (2011) 18 (6), 377-83). Persistent HBV infection stimulates HBV-specific T cell receptors through viral peptides and circulating antigens for a long time, resulting in depletion of circulating and intrahepatic HBV-specific CD4 + and CD8 + T cells. As a result, T cell multifunction is diminished (i.e., reduced IL-2, tumor necrosis factor (TNF) -α, IFN-γ levels, lack of proliferation, and degranulation). Therapeutic vaccination has the potential to remove HBV from chronically infected individuals (Michel et al., J. Hepatol . (2011) 54 (6), 1286-1296). An immunogenic composition containing one or more nucleic acid molecules encoding one or more HBV antigens can be used to provide therapeutic immunity to an individual in need, such as a human with a chronic HBV infection.

Although naked DNA can be delivered to cells in vivo and cause gene expression, the efficiency of gene transfer is relatively low and variable. It has been shown that the local application of electrical signals can enhance the distribution and absorption of macromolecules in living tissues. The application of such electrical signals in tissues in combination with the administration of prophylactic or therapeutic HBV vaccines can have the desired effect on tissues and / or agents to be delivered. In particular, techniques such as electroporation and iontophoresis have been used to significantly improve the distribution and / or uptake of nucleic acids including both DNA and RNA sequences. Potential clinical applications of such technologies include the delivery of nucleic acid sequences for prophylactic and therapeutic immunization, and the delivery of nucleic acid sequences encoding therapeutic proteins or peptides.

The application procedure includes administering the agent of interest to a target tissue site and applying an electric field of a magnitude and duration sufficient to induce a desired effect on the delivery, distribution, and / or efficacy of the agent. These electric fields propagate through two or more electrodes that are in conductive communication with the tissue. Suitable electrode configurations for these technologies include tissue-penetrating electrodes, surface-contact electrodes, and air-gap electrodes. Specific electrode configurations include, but are not limited to, slender needle or rod electrodes, point electrodes, meander electrodes (ie, shaped wires), planar electrodes, and combinations thereof. The specific type and arrangement of the electrodes is selected based on the type of target tissue and the purpose of the procedure. The desired result is best achieved when the electric field propagates within the target tissue in the presence of the agent of interest.

Various methods and devices have been described in which an electric field is applied to the tissue in the presence of the agent of interest to enhance the delivery of the agent in the skin and muscle tissue. These devices include the use of both surface and tissue penetrating electrode systems and combinations thereof. Despite the promise of electrically-mediated drug delivery and the potential application of these technologies to the clinic, there remains a need for reliable and consistent delivery of nucleic acids of interest to individuals. The main source of this change is caused by differences in operator skills and skill levels. Other sources of change that cannot be addressed with current systems include differences in physiological characteristics between individuals that may affect the application of the program. Other considerations for developing a suitable device include its ease of use and implementation of a design that reduces the frequency and significance of potential user errors.

Given the particular need for safe, reliable, accurate and consistent application of clinical therapies, there is a particular need to develop improved application systems. Such developments should include means for minimizing operator-related changes, while providing means to reconcile differences in individual characteristics that may be encountered during widespread clinical application of electrically mediated drug delivery. In other words, specific areas of improvement include the ability to maintain consistent performance across heterogeneous recipient groups and reduce the level of training and skills required for effective user use. In addition, the device, system or method should be designed to help avoid user or device errors and minimize their impact when errors occur.

This prior art system provided to introduce a brief contents of the embodiment and the following embodiment the invention. This prior art is not intended to help determine the scope of the claimed subject matter, nor should it be viewed as limiting the claimed subject matter to implementations that address any or all of the disadvantages or problems provided above.

The present invention provides a device, kit and method for the reproducible, consistent, and effective delivery of an HBV vaccine comprising a nucleic acid molecule encoding an HBV antigen to an individual in need thereof by using Electrically Mediated Therapeutic Agent Delivery (EMTAD) . In the examples described herein, the therapeutic agent is a HBV vaccine.

In one aspect of the present invention, there is provided a device for delivering a HBV vaccine to a predetermined site in an individual, comprising: a combination for the controlled administration of the HBV vaccine to an individual, the combination including a HBV vaccine A reservoir; at least one orifice through which the medicament is administered; and control sufficient to pass a predetermined amount of HBV vaccine from the reservoir through the orifice to a predetermined site within the individual at a predetermined rate Energy source. In addition, the device may include a plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the orifice, and an electrical signal generator operatively connected to the electrodes.

In another aspect of the present invention, there is provided a kit for delivering a HBV vaccine to a predetermined site in an individual, comprising a HBV vaccine and a device including the device for the controlled administration of the HBV vaccine to the individual. An assembly, wherein the device comprises a reservoir of the HBV vaccine; at least one orifice through which the HBV vaccine is administered; and a quantity sufficient to deliver a predetermined amount of HBV vaccine from the reservoir through the orifice at a predetermined rate A controlled sexual energy source delivered orally to a predetermined site within the individual. In addition, the device may include a plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the orifice, and an electrical signal generator operatively connected to the electrodes.

The HBV vaccine included in the device or kit of this application may include:
A first nucleic acid molecule comprising a first polynucleotide encoding a HBV polymerase antigen, the HBV polymerase antigen having an amino acid sequence at least 98% identical to SEQ ID NO: 4, wherein the HBV polymerase antigen does not have a reverse Transcriptase activity and RNAse H activity;
A second nucleic acid molecule comprising a second polynucleotide encoding a truncated HBV core antigen, the truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 14; and medicine Acceptable carrier,
The first nucleic acid molecule and the second nucleic acid molecule exist in the same nucleic acid molecule or in two different nucleic acid molecules.

Other aspects of the invention include a method for administering the HBV vaccine in a controlled manner in space and time with respect to Electric Signal Administration (ESA).

The benefits and advantages of certain embodiments according to the principles of the invention are in a variety of forms. Some embodiments allow the depth of needle and electrode insertion to be selected within heterogeneous populations with different body masses and body compositions, thereby allowing insertion into various types of desired tissue (eg, dermis, muscle, etc.). The embodiments also help adapt the methods for use in specific target groups, such as, but not limited to, human patients with chronic HBV infection and / or HBV-induced diseases. Provided herein are systems and methods that include design features that enable such systems and methods to resist accidental discharge or possible misuse, such as due to dripping, vibration, and / or shedding. In some embodiments, devices are provided that are configured for multiple injection depths. The system and method according to the principles of the present invention allow multiple safety interlocking devices to reduce the frequency and / or impact of user errors. These interlocking devices include features that facilitate the correct preparation and configuration of the dose to be administered, thereby ensuring that the device is applied to the recipient's tissue with the necessary force, that the safety cover is removed, and so on. Systems, devices, kits and methods in accordance with the principles of the present invention can achieve highly consistent delivery of therapy regardless of the type of medication or recipient. The systems, devices, kits, and methods described herein may allow for consistent force patterns to be obtained, for example, before and during delivery therapy, thereby allowing consistent administration to recipients with different skin and muscle characteristics.

This summary is provided to introduce a selection of concepts in a simplified form. These concepts are further described in the implementation section. In addition described in the Summary of the outer element or step may also be affected elements or steps, and no element or step based necessary. This summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used as an aid in determining the scope of the claimed subject matter. The claimed subject matter is not limited to implementations that solve any or all disadvantages noted in any part of this invention.

Other aspects, features, and advantages of the present invention will be readily understood from the following disclosure, including a detailed description of the present invention, its preferred embodiments, and the scope of the attached patent application.

Cross-reference to related applications

This application claims the priority of Taiwan Patent Application No. 106144667, filed on December 19, 2017, the disclosure of which is incorporated herein by reference in its entirety.
Reference to Sequence Listing Submitted Electronically

This application contains a sequence listing, which is filed electronically via EFS-Web with an ASCII format sequence listing created on December 10, 2018 with the file name "688097-405 Sequence Listing" and a size of 46.6 KB. This sequence listing submitted via EFS-Web is part of the specification and is incorporated herein by reference in its entirety.

The following description and examples illustrate embodiments of the invention in detail. It should be understood that the invention is not limited to the particular embodiments described and may therefore vary. Those skilled in the art will recognize that there are many variations and modifications of the present invention, which are encompassed within the scope of the present invention.

All terms are intended to be understood by those skilled in the art. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

The section headings used in this article are for organizational purposes only and should not be construed as limiting the subject matter described.

Although various features of the invention may be described in the context of a single embodiment, the features may also be provided separately or in any suitable combination. In contrast, although the invention may be described herein in the context of separate embodiments for clarity, the invention may also be implemented in a single embodiment.

To assist the reader of this application, the description is divided into various paragraphs or sections, or directed to various embodiments of this application. Such separations should not be considered as unrelated to the substance of one paragraph or part or embodiment and the substance of another paragraph or part or embodiment. On the contrary, those skilled in the art should understand that this description has broad applications and covers all combinations of various parts, paragraphs, and sentences that can be covered. The discussion of any embodiment is intended to be illustrative only, and is not intended to indicate the scope of the invention, including the scope of patent applications, which is limited to such examples. For example, although the embodiments of the HBV vector described herein that can be used in this application (such as plastid DNA or viral vectors) may contain specific components arranged in a particular order, including (but not limited to) certain Promoter sequences, enhancers or regulatory sequences, signal peptides, HBV antigen coding sequences, polyadenylation signal sequences, etc., but those skilled in the art should understand that the concepts disclosed herein can be equally applied to the use of Other components are arranged in other orders as applicable to the HBV vectors of this application. This application covers the use of any of the applicable components in any combination with any sequence that can be used in the HBV vector suitable for this application, whether or not a particular combination is explicitly described.

The following definitions supplement these definitions in this technology and are for this application and should not be attributed to any related or unrelated situation, such as any jointly owned patent or application. Although any methods and materials similar or equivalent to those described herein can be used in the practice of testing of the present invention, the preferred materials and methods are described herein. Accordingly, the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

In this application, the use of the singular includes the plural unless specifically stated otherwise. It must be noted that, unless the context clearly indicates otherwise, as used in this specification, the singular forms "a / an" and "the" include plural referents. In this application, the use of "or" means "and / or" unless stated otherwise. In addition, the use of the term "including" and other forms, such as "include", "includes", and "included", are not restrictive.

"Some embodiments," "one embodiment," or "other embodiments" in this specification means that a particular feature, structure, or characteristic described in connection with the embodiments is included in at least some embodiments of the invention , But not necessarily included in all embodiments.

As used in this specification and the scope of the patent application, the words "comprising" (and any form of inclusion, such as "comprise" and "comprises"), "having" (and having Any form, such as "have" and "has"), "including" (and any form of inclusion, such as "includes" and "include") or " "Containing" (and any form of inclusion, such as "contains" and "contain" are inclusive or open and do not exclude other unlisted elements or method steps. It is expected in this description Any of the embodiments discussed in this document can be implemented using any method or composition of the invention, and vice versa. In addition, the composition of the invention can be used to implement the method of the invention.

The term "about" and its grammatical equivalents as used herein with respect to a reference value may include the value itself and a series of values plus or minus 10% from the value. For example, the amount "about 10" includes 10 and any amount from 9 to 11. For example, the term "about" with respect to a reference value may also include plus or minus 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% from the value. A range of values.

The present invention provides improved systems, sets, for HBV vaccines, such as reproducible, consistent, and effective delivery of nucleic acids encoding HBV antigens, and combinations thereof with electrically mediated therapeutic agents (e.g., HBV vaccine) delivery (EMTAD), Method and device.

In one aspect, the present invention provides a device for delivering a HBV vaccine to a predetermined site within an individual, comprising: a cartridge assembly comprising an outer tube, an inner tube, and a reservoir containing the HBV vaccine , Wherein the closed system of the reservoir is contained in the outer cylinder and is configured to receive the reservoir; the applicator, which includes a cylinder assembly receiver, a needle interface, and an insertion detector, wherein the insertion detection The at least one interlocking device, wherein the interlocking device helps to correctly perform the HBV vaccine administration procedure; at least one injection orifice, the HBV vaccine is passed through the Administration of injection orifices; a plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the orifices; an electric field generator for generating electrical signals operatively connected to the electrodes; and a sufficient amount of a predetermined amount of HBV vaccine A control energy source that is delivered at a predetermined rate from the reservoir through the orifice to a predetermined location within the individual.

In another aspect, the present invention provides a kit for delivering a HBV vaccine to a predetermined site within an individual, comprising a HBV vaccine and a device, the device comprising: a cartridge assembly, the cartridge assembly comprising an outer tube , An inner cylinder, and a reservoir of the HBV vaccine, wherein the reservoir closed system is contained in the outer cylinder and is configured to receive the reservoir; an applicator, which includes a cylinder assembly receiver and a needle interface And an insertion detector, wherein the insertion detector senses the loading of the reservoir in the reservoir enclosure; at least one interlocking device, wherein the interlocking device helps to correctly perform the HBV vaccine administration procedure; at least An injection orifice through which the HBV vaccine is administered; a plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the orifice; an electric field for generating an electrical signal that is operatively connected to the electrodes A generator; and a control energy source sufficient to deliver a predetermined amount of the HBV vaccine from the reservoir through the orifice to a predetermined site within the individual at a predetermined rate.

The HBV vaccine included in the device or kit of this application may include:
A first nucleic acid molecule comprising a first polynucleotide encoding a HBV polymerase antigen, the HBV polymerase antigen having an amino acid sequence at least 98% identical to SEQ ID NO: 4, wherein the HBV polymerase antigen does not have a reverse Transcriptase activity and ribonuclease H activity;
A second nucleic acid molecule comprising a second polynucleotide encoding a truncated HBV core antigen, the truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 14; and medicine Acceptable carrier,
The first nucleic acid molecule and the second nucleic acid molecule exist in the same nucleic acid molecule or in two different nucleic acid molecules.

In certain aspects of the invention, EMTAD can be considered as the administration of HBV vaccine to the biological tissue of interest and before, at the same time or after, an electrical signal is applied to the biological tissue to achieve enhanced migration of the HBV vaccine in the tissue And / or absorption purpose. The method of EMTAD includes two elements: 1) administration of a therapeutic agent (ie, HBV vaccine) (TAA); and 2) application of an electrical signal (ESA) sufficient to induce the desired EMTAD effect. In the present invention, TAA can be implemented in a controllable manner, for example, called Controlled Therapeutic Agent Administration (CTAA). The term CTAA as used herein may refer to a method or device that provides space and / or time control over the administration of an HBV vaccine relative to the induction of the effect of EMTAD. Controlled dosing techniques can be performed using conventional syringes with needles (e.g. automatic injection devices) and / or various needle-free methods (e.g. jet syringes, transdermal / percutaneous patches, oral, gel, cream or inhaled administration )The change. The term ESA, as used herein, can refer to the application of electrical signals to promote or enhance the delivery of an agent by improving the movement and / or absorption of an active agent, such as a HBV vaccine, thereby inducing EMTAD effects. When used to facilitate or enhance the delivery of HBV vaccines, ESA methods such as electroporation, iontophoresis, electroosmosis, electropermeabilization, electrical stimulation, electromigration, and electroconvection all represent various modes of EMTAD, of which One or more may be included in the methods described herein.

Specific applications of the devices, kits and systems described herein include, but are not limited to, delivery of a HBV vaccine containing one or more nucleic acid molecules. For such applications, EMTAD has traditionally been initiated by injecting the HBV vaccine using a conventional needled syringe. After administering the medicament, a device suitable for ESA is applied to a designated location on the individual. Finally, HBV vaccine delivery is desirably promoted or enhanced with appropriate ESA protocols. However, with traditional EMTAD, the space and time relationship between the administration of the drug and the ESA may not be realized.
Spatial parameter

In some embodiments of the systems, kits, methods, and devices described herein, HBV vaccine administration is performed using a conventional syringe with a needle. The need to use EMTAD to deliver certain agents further increases the complexity of TAA implementation. As depicted in FIG. 1, any conventional injection syringe with needle, when the needle 5 is inserted into the tissue, it may be difficult to control with respect to the tissue surface 3 of the insertion depth and angle a 2. In addition, the position 4 where the needle penetrates at the tissue surface 3 may not be representative of the position of the orifice 6 and the area 7 in which the drug is administered in the target tissue. By way of illustrative example, a percutaneous intramuscular injection may not correspond to an insertion site on the skin, as two tissues usually move relative to each other.

Although this conventional method is generally applicable to the delivery of many different therapeutic agents that do not require EMTAD, these variables cause the distribution of HBV vaccines after injection to be generally inconsistent and / or uncertain and may prevent effective EMTAD. In certain embodiments described herein, the most effective use of EMTAD utilizes a predetermined relationship between HBV vaccine and ESA in an individual. Therefore, compared to devices, methods, or systems that provide space and time control, the use of conventional syringes with needles reduces the effectiveness of EMTAD applications without performing space control of the TAA in the target tissue. An illustrative example of this concept is the use of electroporation to facilitate the delivery of HBV vaccine. When TAA and ESA are co-localized within a target area of a tissue, electroporation is often the most effective in improving HBV vaccine delivery. In many cases, if the agent to be delivered and the induced electroporation are not co-localized in a target area of the tissue, the delivery of the agent is sub-optimal.

Another example that requires proper spatial control of TAA in EMTAD is iontophoresis. This EMTAD mode uses an electric field to move charged molecules. In order to achieve the required medicament movement, a proper spatial relationship between the electrodes and the HBV vaccine must be achieved. If a negatively charged agent is placed near the position of the positive electrode, then little or no movement of the agent through the tissue will be observed. In contrast, positioning the negatively charged agent near the negative electrode will cause the agent to move significantly through the tissue in the direction of the positive electrode.

As illustrated by the previous examples, it is important to control the precise position of the TAA relative to the ESA to achieve the desired effect. Thus, embodiments of the devices and methods described herein provide control over the precise location of TAA relative to the ESA, and can be used to achieve a reproducible, consistent, and well-characterized distribution of one or more HBV vaccines.
Time parameter

In the case of conventional TAA injection with a needle syringe, the injection rate may vary from operator to operator, thereby making the distribution of the agent in the tissue inconsistent. When multiple devices need to be placed to complete the EMTAD process, time variations are introduced. For example, one application of EMTAD requires the administration of plastid DNA encoding a therapeutic protein, followed by an electroporation-induced electric field. Using the traditional EMTAD method, the HBV vaccine was injected with a needle syringe, and then the electroporation device was placed and activated. Due to the need to place two separate devices (original needle syringes, followed by ESA devices), this procedure is susceptible to inter-individual changes caused by the inconsistencies in the timing of application of each device by the operator. In addition, the use of two independently placed devices results in a unavoidable time interval between the placement of the clinician and the activation of each device. This situation is more complicated where multiple application sites are needed to achieve proper delivery of the medicament to a designated area within the target tissue.

These issues are particularly important for agents, such as nucleic acids, that can degrade or inactivate in the extracellular environment. Degradation of nucleic acids in HBV vaccines can lead to reduced efficacy of the therapy and reduced consistency of therapy application. In addition, the rate of nucleic acid degradation in HBV vaccines is not constant among individuals, which leads to inconsistencies in the overall treatment of conventional needle-injector injections with ESA, and more specifically with electroporation therapy.

Due to the inherent spatial and temporal variation issues associated with the use of conventional needle-injector injection with ESA, the precise location and timing of TAA relative to ESA is often unknown. Therefore, the effective administration and dosage of HBV vaccines using EMTAD may be inconsistent and unreproducible. Although conventional syringe injections with needles are sometimes suitable for HBV vaccine administration, by controlling the spatial and temporal relationship between HBV vaccine administration and induction of the desired EMTAD effect, reproducible and consistent delivery of HBV vaccine will be significantly improved .

Therefore, although traditional EMTAD procedures may be suitable for some applications, time and space control systems are particularly desirable for clinical applications that often require a high degree of consistency and reproducibility. Compared with the conventional EMTAD method, embodiments of the method, system, and device described herein help CTAA and ESA to provide a method and device that are more beneficial to the clinical application of EMTAD. The present invention utilizes various aspects of CTAA in combination with ESA to provide reproducible, consistent, and effective HBV vaccine delivery. The method and device described in the present invention implement space and time control over the administration of HBV vaccine relative to the application of electrical signals, thereby improving the movement and / or absorption of the vaccine in the target tissue.

In some embodiments, methods and devices are provided, wherein there is a controllable spatial relationship between the administration of the HBV vaccine and the application of an electrical signal. Prior to treatment, determine the optimal position of TAA relative to ESA. This spatial relationship between TAA and ESA is determined by the treatment parameters, including the nature of the medicament administered and the characteristics of the target tissue to which the medicament is administered. In an exemplary embodiment, the electrical signal is preferentially applied to the distal end of the HBV vaccine administration site. In certain other embodiments, the spatial relationship is the application of an EMTAD-induced electrical signal to the proximal end of the medicament administration site. In some cases, co-localization between TAA and ESA is preferred. This is often the case when the desired EMTAD effect is induced using electroporation and / or iontophoresis.

In another aspect of the invention, the device described herein provides a controllable time relationship regarding the order and timing of TAAs relative to ESAs. Before treatment, determine the best order and timing of the combination of TAA and ESA. As with spatial relationships, the desired temporal relationship between TAA and ESA is determined by parameters such as the nature of the medicament administered and the characteristics of the target tissue to which the medicament is administered. In certain applications, exposure to ESA-related electric fields can adversely affect HBV vaccines. In the practice of such applications, CTAA is performed after generating such an electric field. However, the typical time relationship is ESA after CTAA.

The present invention provides improved methods and devices for reproducible, consistent, and efficient delivery of HBV vaccines containing nucleic acid-based constructs using EMTAD. This is achieved by controlling the space and time of HBV vaccine administration relative to the application of electrical signals. In one embodiment, EMTAD is initiated by injecting the HBV vaccine using a conventional needled syringe. After administering the medicament, a device suitable for ESA is applied to a designated location on the individual. Desirably promote or enhance the delivery of HBV vaccine using appropriate ESA protocols. An exemplary ESA method that has proven effective in almost all cell types is electroporation. Other exemplary methods of electrical-mediated delivery include, but are not limited to, iontophoresis, electroosmosis, electroosmosis, electrical stimulation, electromigration, and electroconvection. These terms are used for illustrative purposes only and should not be construed as limiting the invention.

Electroporation uses the application of an electric field to induce a transient increase in cell membrane permeability and move charged particles. Due to the permeability of the cell membrane in the target tissue, electroporation significantly improves the intracellular absorption of foreign substances that have been administered to the target tissue. Increased cell membrane permeability and molecular mobility caused by electroporation provides a method for overcoming cell membranes as a barrier to HBV vaccine delivery. The advantage of applying electroporation as a technique to induce EMTAD is that the physical properties of the technique allow electroporation to be applied in almost all tissue types. Therefore, various aspects and embodiments of the present invention discuss (but are not limited to) electroporation as a technique for inducing EMTAD.
HBV vaccine

The term "HBV vaccine" is used herein in its broadest sense and includes any agent capable of providing a desired or beneficial immune response, therapeutic and / or prophylactic effect against HBV to living tissues. The term therefore includes both prophylactic and therapeutic HBV vaccines, as well as any other class of agents that have such a desired effect. The scope of the invention is broad enough to include controlled delivery of any HBV vaccine, regardless of classification. HBV vaccines include, but are not limited to, medicines and vaccines, and nucleic acid sequences (such as supercoiled, loose, and linear progenitor DNA, RNA, antisense constructs, artificial chromosomes, or any other nucleic acid-based therapeutic agent) and any of their formulations . Such pharmaceutical formulations include, but are not limited to, cationic lipids, cationic and / or non-ionic polymers, liposomes, saline, nuclease inhibitors, anesthetics, poloxamers, preservatives, Sodium phosphate solution, or other compounds that can improve the administration, stability, and / or effect of HBV vaccines. Other formulations include agents and additives that impart the ability to control the viscosity and resistance of the administered agent.

In a preferred aspect of the present application, the HBV vaccine used in the present invention may include:
The first nucleic acid molecule, preferably a first plastid DNA vector, comprises a first polynucleotide encoding an HBV polymerase antigen, and preferably the HBV polymerase antigen has at least 98% identity with SEQ ID NO: 4, such as Amino acids consistent with SEQ ID NO: 4 at least 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100% A sequence in which the HBV polymerase antigen does not have reverse transcriptase activity and ribonuclease H activity;
A second nucleic acid molecule, preferably a second plastid DNA vector, comprising a second polynucleotide encoding a truncated HBV core antigen, preferably the truncated HBV core antigen is represented by SEQ ID NO: 2 or SEQ ID The amino acid sequence composition of NO: 14; and a pharmaceutically acceptable carrier, and wherein the first nucleic acid molecule and the second nucleic acid molecule are present in the same nucleic acid molecule, such as the same plastid DNA vector, or It exists in two different nucleic acid molecules, such as two independent plastid DNA vectors.

As used herein, the terms "HBV core antigen", "HBcAg", and "core antigen" all refer to HBV antigens capable of eliciting an immune response against an HBV core protein, such as a humoral and / or cell-mediated response in an individual. The terms "core", "core polypeptide" and "core protein" all refer to the HBV virus core protein. The full-length core antigen is typically 183 amino acids in length and includes an assembly domain (amino acids 1 to 149) and a nucleic acid binding domain (amino acids 150 to 183). This 34-residue nucleic acid binding domain is required for capsidization of pregenomic RNA. This domain is also used as a nuclear input signal. It contains 17 arginine residues and is highly basic, consistent with its function. The HBV core protein is in the form of a dimer in solution, and the dimer self-assembles into an icosahedral capsid. Each core protein dimer has four alpha-helical bundles flanked by an alpha-helical domain on either side. A truncated HBV core protein without the nucleic acid binding domain can also form a capsid.

In one aspect of the present application, the HBV antigen used in the present invention is a truncated HBV core antigen. As used herein, "truncated HBV core antigen" refers to a HBV antigen that does not contain a full-length HBV core protein but is capable of eliciting an immune response against the HBV core protein in an individual. For example, the HBV core antigen can be modified so that the core antigen typically contains seventeen spermine (R) residues, one or Multiple amino acids are missing. The truncated HBV core antigen of the present application is preferably a C-terminal truncated HBV core protein that does not contain the HBV core nuclear input signal and / or a truncated HBV core protein that has been deleted from the C-terminal HBV core nuclear input signal. In one embodiment, the truncated HBV core antigen comprises a deletion in the C-terminal nucleic acid binding domain, such as a deletion of 1 to 34 amino acid residues in the C-terminal nucleic acid binding domain, such as 1, 2, 3, 4 , 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 , 30, 31, 32, 33 or 34 amino acid residues, preferably all 34 amino acid residues are deleted. In a preferred embodiment, the truncated HBV core antigen comprises a deletion in the C-terminal nucleic acid binding domain, preferably all 34 amino acid residues are deleted.

The HBV core antigens that can be used in this application can be common sequences derived from multiple HBV genotypes (eg, genotypes A, B, C, D, E, F, G, and H). As used herein, "common sequence" means an amino acid sequence alignment based on homologous proteins, such as an artificial amino acid sequence determined by aligning (eg, using Clustal Omega) amino acid sequences of homologous proteins. It can be the order of the most common amino acid residues found at various positions in the sequence alignment, calculated based on sequences of HBV antigens (e.g., core, pol, etc.) from at least 100 natural HBV isolates. The common sequence may be non-naturally occurring and different from the native viral sequence. The common sequence can be designed by using multiple sequence alignment tools to align multiple HBV antigen sequences from different sources, and selecting the most common amino acid at the position where the alignment is changed. Preferably, the common sequence of HBV antigens is derived from HBV genotypes B, C and D. The term "common antigen" is used to refer to antigens having a common sequence.

Exemplary truncated HBV core antigens that can be used in this application do not have a nucleic acid binding function and can elicit an immune response in mammals against at least two HBV genotypes. Preferably, the truncated HBV core antigen is capable of inducing a T-cell response in mammals against at least HBV genotypes B, C, and D. More preferably, the truncated HBV core antigen is capable of eliciting a CD8 T cell response against at least HBV genotypes A, B, C, and D in a human individual.

Preferably, the HBV core antigens used in the present application are common antigens, preferably common antigens derived from HBV genotypes B, C, and D, and more preferably truncated from HBV genotypes B, C, and D Common antigen. Exemplary truncated HBV core common antigens according to the present application are at least 90% identical to SEQ ID NO: 2 or SEQ ID NO: 14, such as at least 90%, 91 corresponding to SEQ ID NO: 2 or SEQ ID NO: 14, 91 %, 92%, 93%, 94%, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100% identical amino acid sequence composition. SEQ ID NO: 2 and SEQ ID NO: 4 are core common antigens derived from HBV genotypes B, C, and D. SEQ ID NO: 2 and SEQ ID NO: 14 delete the 34 amino acids of the nucleic acid binding domain with a large positive charge (sinine-rich) at the C-terminus of the natural core antigen.

In a specific embodiment of the present application, the HBV core antigen is a truncated HBV antigen consisting of the amino acid sequence of SEQ ID NO: 2. In another specific embodiment, the HBV core antigen is a truncated HBV antigen consisting of the amino acid sequence of SEQ ID NO: 14.

Examples of a polynucleotide sequence encoding a truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2 include, but are not limited to, being at least 90% identical to SEQ ID NO: 1, such as or in accordance with SEQ ID NO: 1 at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98% , At least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% consistent, more than A polynucleotide sequence that is approximately 98%, approximately 99%, or 100% identical to SEQ ID NO: 1.

Examples of a polynucleotide sequence encoding a truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 14 include, but are not limited to, being at least 90% identical to SEQ ID NO: 15, such as or in accordance with SEQ ID NO: 15 at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98% , At least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% consistent, more than A polynucleotide sequence that is about 98%, about 99%, or 100% identical to SEQ ID NO: 15 is preferred.

In a specific embodiment of the present application, the HBV vaccine used in the present invention comprises a non-naturally occurring nucleic acid molecule encoding a truncated HBV core antigen, and the non-naturally occurring nucleic acid molecule comprises SEQ ID NO: 1 or SEQ ID NO: 15 polynucleotide sequence.

As used herein, the terms "HBV polymerase antigen", "HBV Pol antigen" or "HBV pol antigen" refer to those capable of eliciting an immune response, such as a humoral and / or cell-mediated response, to HBV polymerase in an individual. HBV antigen. The terms "polymerase", "polymerase polypeptide", "Pol" and "pol" all refer to HBV virus DNA polymerase. HBV viral DNA polymerase has four domains, from the N-terminus to the C-terminus, including the terminal protein (TP) domain, which serves as a primer for negative-strand DNA synthesis; spacers that are not important for polymerase function; reverse for transcription Transcriptase (RT) domain; and ribonuclease H domain.

In one embodiment of the application, the HBV antigen comprises a HBV Pol antigen, or any immunogenic fragment or combination thereof. The HBV Pol antigen may contain other modifications that improve the immunogenicity of the antigen, such as by introducing mutations into the active site of the polymerase and / or ribonuclease H domains to reduce or substantially remove certain enzyme activities.

Preferably, the HBV Pol antigen that can be used in the present application does not have reverse transcriptase activity and ribonuclease H activity, and can induce an immune response in mammals against at least two HBV genotypes. Preferably, the HBV Pol antigen is capable of inducing a T-cell response in mammals against at least HBV genotypes B, C, and D. More preferably, the HBV Pol antigen is capable of eliciting a CD8 T cell response against at least HBV genotypes A, B, C, and D in a human individual.

Therefore, in some embodiments, the HBV Pol antigen line used in the present application is an inactivated Pol antigen. In one embodiment, the inactivated HBV Pol antigen comprises one or more amino acid mutations in the active site of the polymerase domain. In another embodiment, the inactivated HBV Pol antigen comprises one or more amino acid mutations in the active site of the ribonuclease H domain. In a preferred embodiment, the inactivated HBV pol antigen contains one or more amino acid mutations in the active site of both the polymerase domain and the ribonuclease H domain. For example, reverse transcription can be reduced or substantially eliminated, for example, by replacing one or more asparagine residues (D) with asparagine residues (N) to eliminate or reduce metal coordination functions. Enzyme function to mutate the "YXDD" motif required for nucleotide / metal ion binding in the polymerase domain of the HBV pol antigen. As an alternative to or in addition to the "YXDD" motif mutation, for example, by replacing one or more aspartic acid residues (D) with asparagine residues (N) and / or with gluten leucine (Q) substitutions glutamic acid residue (E), thereby reducing or substantially eliminate the RNase H function to make the desired Mg ^{2 +} ligand RNase H domain antigen of HBV pol "DEDD Primitive mutation. In a specific embodiment, the HBV pol antigen system is modified by: (1) mutating an asparagine residue (D) in the "YXDD" motif of the polymerase domain to an asparagine residue ( N); and (2) mutating the first asparagine residue (D) in the "DEDD" motif of the ribonuclease H domain to the asparagine residue (N) and making the first bran The amino acid residue (E) is mutated to the glutamate residue (N), thereby reducing or substantially eliminating the reverse transcriptase and ribonuclease H functions of the pol antigen.

In a preferred embodiment of the present application, the HBV pol antigen is a common antigen, preferably a common antigen derived from HBV genotypes B, C, and D, and more preferably a HBV genotype B, C, or D. Common antigen of inactivation. Exemplary HBV pol common antigens according to the present application contain at least 90% identity to SEQ ID NO: 4, such as at least 90%, 91%, 92%, 93%, 94%, 95%, 95.5, and SEQ ID NO: 4 %, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100% consistent, preferably at least 98% consistent with SEQ ID NO: 4, such as at least 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6 %, 99.7%, 99.8%, 99.9%, or 100% identical amino acid sequences. SEQ ID NO: 4 is a pol common antigen derived from HBV genotypes B, C, and D that contains four mutations in the active sites of the polymerase and ribonuclease H domains. Specifically, the four mutations include mutations in the asparagine residue (D) in the "YXDD" motif of the polymerase domain to asparagine residue (N); and the " The first aspartic acid residue (D) in the "DEDD" motif is mutated to asparagine residue (N) and glutamic acid residue (E) to glutamic acid residue (Q).

In a specific embodiment of the present application, the HBV pol antigen that can be used in the present application comprises the amino acid sequence of SEQ ID NO: 4. In other embodiments of this application, the HBV core antigen that can be used in this application consists of the amino acid sequence of SEQ ID NO: 4.

Examples of a polynucleotide sequence encoding a HBV pol antigen comprising the amino acid sequence of SEQ ID NO: 4 include, but are not limited to, at least 90% identity to SEQ ID NO: 3 or SEQ ID NO: 16, such as with SEQ ID NO: 3 or SEQ ID NO: 16 at least 90%, 91%, 92%, 93%, 94%, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100%, preferably 98% of SEQ ID NO: 3 or SEQ ID NO: 16 %, 99%, or 100% identical polynucleotide sequences. In a specific embodiment of the present application, the HBV vaccine useful in the present application comprises a non-naturally occurring nucleic acid molecule encoding the HBV pol antigen, and the non-naturally occurring nucleic acid molecule comprises SEQ ID NO: 3 or SEQ ID NO: 16 of the polynucleotide sequence.

As used herein, the term "fusion protein" or "fusion" refers to a single polypeptide chain having at least two polypeptide domains that are not normally present in a single natural polypeptide.

In one aspect of the present application, the HBV antigen used in the present invention may include a truncated HBV core antigen operably linked to a HBV Pol antigen, or a HBV Pol antigen operably linked to a truncated HBV core The fusion of an antigen is preferably performed via a linker. As used herein, the term "linker" refers to a compound or moiety that acts as a molecular bridge to operably link two different molecules, where a portion of the linker is operably linked to a first molecule, and wherein the linker The other part is operably linked to the second molecule. As used herein, the term "operably linked" means linked or contiguous, where the components so described are in a relationship that allows them to function in their intended manner. For example, a regulatory sequence operably linked to a nucleic acid sequence of interest can direct transcription of the nucleic acid sequence of interest, or a signal sequence operably linked to an amino acid sequence of interest can direct the amino acid sequence of interest Secreted or translocated to a component.

For example, in a fusion protein containing a first polypeptide and a second heterologous polypeptide, a linker is mainly used as a spacer between the first and second polypeptides. In one embodiment, the linker system is composed of amino acids linked together via peptide bonds, preferably from 1 to 20 amino acids linked via peptide bonds, wherein the amino acid system is selected from 20 naturally occurring Amino acids. In one embodiment, the 1 to 20 amino acids are selected from the group consisting of glycine, alanine, proline, asparagine, glutamine and lysine. Preferably, the linker system is composed of a large number of sterically hindered amino acids such as glycine and alanine. Exemplary linker systems are polyglycine, especially (Gly) 5, (Gly) 8; poly (Gly-Ala), and polyalanine. An exemplary suitable linker line (AlaGly) n, where n is an integer from 2 to 5.

Preferably, the fusion protein useful in the present application is capable of eliciting an immune response in mammals against HBV core and HBV Pol of at least two HBV genotypes. Preferably, the fusion protein is capable of inducing a T-cell response in mammals against at least HBV genotypes B, C, and D. More preferably, the fusion protein is capable of eliciting a CD8 T cell response against at least HBV genotypes A, B, C, and D in a human individual.

In one aspect of the present application, a fusion protein that can be used in the present application can include a protein having at least 90% identity to SEQ ID NO: 2 or 14, such as at least 90%, 91%, 92%, 93%, 94 %, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, Truncated HBV core antigen with 99.8%, 99.9% or 100% identical amino acid sequence; linker; and having at least 90% identity with SEQ ID NO: 4, such as at least 90%, 91%, 92%, 93 %, 94%, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, HBV Pol antigen with 99.7%, 99.8%, 99.9% or 100% identical amino acid sequences.

Preferably, the fusion protein useful in the present application comprises a truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2 or 14; a linker comprising (AlaGly) n, where n is 2 to An integer of 5; and a HBV Pol antigen having the amino acid sequence of SEQ ID NO: 4. More preferably, the fusion protein useful in the present application comprises the amino acid sequence of SEQ ID NO: 20.

In some embodiments of the present application, the fusion protein that can be used in the present application further comprises a signal sequence. Preferably, the signal sequence has an amino acid sequence of SEQ ID NO: 6 or SEQ ID NO: 19. More preferably, the fusion protein comprises the amino acid sequence of SEQ ID NO: 21.

Examples of polynucleotide sequences encoding fusion proteins useful in the present application include, but are not limited to, at least 90% identity to SEQ ID NO: 1 or SEQ ID NO: 15 such as, for example, SEQ ID NO: 1 or SEQ ID NO: 15 at least 90%, 91%, 92%, 93%, 94%, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100%, preferably 98%, 99% or 100 of SEQ ID NO: 1 or SEQ ID NO: 15 A polynucleotide sequence that is 100% identical is operably linked to be at least 90% identical to SEQ ID NO: 22, such as at least 90%, 91%, 92%, 93%, 94%, 95%, 95.5%, 96%, 96.5%, 97%, 97.5%, 98%, 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% Or 100% identical, preferably 98%, 99%, or 100% identical linker coding sequence, which is further operably linked to SEQ ID NO: 3 or SEQ ID NO: 22 16 is at least 90% consistent, such as at least 90%, 91%, 92%, 93%, 94%, 95%, 95.5%, 96%, 96.5%, 97%, SEQ ID NO: 3 or SEQ ID NO: 16 97.5%, 98% 98.5%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, or 100%, preferably the same as SEQ ID NO: 3 or SEQ ID NO: 16 has 98%, 99%, or 100% identical polynucleotide sequences. In a specific embodiment of the present application, the HBV vaccine useful in the present application comprises a non-naturally occurring nucleic acid molecule encoding a fusion protein, and the non-naturally occurring nucleic acid molecule comprises a operably linked to SEQ ID NO: 22 SEQ ID NO: 1, which is further operably linked to SEQ ID NO: 3.

In one aspect of the present application, in the HBV vaccine used in the present application, the first nucleic acid molecule and the second nucleic acid molecule are a first plastid DNA vector and a second plastid DNA vector, respectively, and the first The first and second plastid DNA vectors each contain an origin of replication, an antibiotic resistance gene, and a promoter sequence, an enhancer sequence, a signal peptide coding sequence, a first polynucleotide sequence, or a second sequence from the 5 'end to the 3' end. Polynucleotide sequences, and polyadenylation signal sequences. In a preferred embodiment of the present application, the DNA plasmonics system is suitable for a protein expression vector in mammalian host cells. Expression vectors suitable for expressing proteins in mammalian host cells include, but are not limited to, pcDNATM, pcDNA3TM, pVAX, pVAX-1, and the like. Preferably, the expression vector is based on pVAX-1, which can be further modified to optimize protein performance in mammalian cells. pVAX-1 is a common plastid in DNA vaccines and contains a strong human immediate early cytomegalovirus (CMV-IE) promoter, followed by bovine growth hormone (bGH) -derived polyadenylation sequence (pA). pVAX-1 also contains a pUC origin of replication and a kanamycin resistance gene driven by a small prokaryotic promoter that allows bacterial plastids to multiply. Preferably, the plastid DNA vector comprises an optimized kanamycin resistance having a codon having a polynucleotide sequence that is at least 90% identical to SEQ ID NO: 12, and preferably 100% identical to SEQ ID NO: 12. Sex genes.

In a specific embodiment, the HBV vaccine used in the present invention comprises:
a) The first plastid DNA vector, from the 3 'end to the 5' end, comprising a promoter sequence containing the polynucleotide sequence of SEQ ID NO: 7 and Enhancer sequence, signal peptide coding sequence containing the polynucleotide sequence of SEQ ID NO: 5, a first polynucleotide sequence containing the polynucleotide sequence of SEQ ID NO: 3, and Polyadenylation signal sequence of a polynucleotide sequence;
b) a second plastid DNA vector, from the 3 'end to the 5' end, comprising a promoter sequence containing the polynucleotide sequence of SEQ ID NO: 7 and Enhancer sequence, signal peptide coding sequence containing the polynucleotide sequence of SEQ ID NO: 5, a second polynucleotide sequence containing the polynucleotide sequence of SEQ ID NO: 1, and A polyadenylation signal sequence of a polynucleotide sequence; and
c) a pharmaceutically acceptable carrier,
Wherein the first plastid DNA vector and the second plastid DNA vector each further comprise a kanamycin resistance gene having a polynucleotide sequence of SEQ ID NO: 12 and a polynucleoside having SEQ ID NO: 10 The origin of replication of the acid sequence, and wherein the first plastid DNA vector and the second plastid DNA vector are in the same composition or two different compositions.

In these embodiments of the application in which the HBV vaccine includes a first vector (such as a first DNA plastid) and a second vector (such as a second DNA plastid), the respective amounts of the first vector and the second vector are not affected. Special restrictions. For example, the first DNA plastid and the second DNA plastid can be from 10: 1 to 1:10 by weight, such as 10: 1, 9: 1, 8: 1, 7: 1, 6 by weight : 1, 5: 1, 4: 1, 3: 1, 2: 1, 1: 1, 1: 2, 1: 3, 1: 4, 1: 5, 1: 6, 1: 7, 1: 8 , 1: 9, or 1:10 ratios exist. Preferably, the first DNA plastids and the second DNA plastids exist at a ratio of 1: 1 by weight.

The compositions and immunogenic combinations of the present application may comprise additional polynucleotides or vectors encoding additional HBV antigens and / or additional HBV antigens or immunogenic fragments thereof. However, in certain embodiments, the compositions and immunogenic combinations of this application do not include certain antigens. In a preferred embodiment, the HBV vaccine used in the present invention does not contain a nucleic acid molecule encoding a HBV antigen selected from the group consisting of hepatitis B surface antigen (HBsAg), HBV envelope (Env) antigen, and HBV L protein antigen. It also does not contain HBV antigens selected from the group consisting of hepatitis B surface antigen (HBsAg), HBV envelope (Env) antigen and HBV L protein antigen.

HBV vaccines that can be used in this application can also include a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers are non-toxic and should not interfere with the efficacy of the active ingredients. Pharmaceutically acceptable carriers may include, for example, one or more of the following: water, glycols, sugars, oils, amino acids, alcohols, preservatives, lubricants, stabilizers, colorants, and the like.

Other examples of HBV vaccines that can be used in this application are described in the file entitled `` Hepatitis B Virus (HBV) Vaccines and Uses thereof '' filed on the same day as this application The contents of the International Patent Application No. 688097-403WO1 of the Attorney are incorporated herein by reference in their entirety.
Set / System

In a general aspect, the present invention relates to a kit or system for the controlled delivery of a HBV vaccine to a predetermined tissue site in an individual in need, comprising a HBV vaccine and applying the HBV vaccine to the subject via electroporation A device for pre-determining a tissue site. For example, the kit or system can have a pre-filled container, such as a syringe, containing a pre-measured amount of the HBV vaccine, which can be loaded into the device described herein for subsequent administration of the HBV vaccine.
Methods to induce immune response

In another general aspect, the invention relates to a method for inducing an immune response against hepatitis B virus (HBV) in an individual in need, comprising using the device, kit or system of the present application to the individual Administer an immunogenicly effective amount of HBV vaccine. Any of the devices, kits, or systems of this application described herein can be used in the methods of this application. As used herein, the term "infection" refers to the invasion of a host by a pathogen. Pathogens are considered "infectious" when they can invade the host and replicate or reproduce within the host. Examples of infectious agents include viruses such as HBV and adenoviruses, prions, bacteria, fungi, protozoa and the like of certain species. "HBV infection" specifically refers to the invasion of HBV to a host organism, such as the cells and tissues of the host organism.

When used in connection with the methods described herein, "evoked immune response" as used herein encompasses eliciting a desired immune response or effect against an infection, such as an HBV infection, in an individual in need. "Induced immune response" also covers the provision of therapeutic immunity against pathogens such as HBV for treatment. As used herein, the term "therapeutic immunity" or "therapeutic immune response" means that a vaccinated individual is able to control the infection of the pathogen to which the vaccination is directed, for example by vaccination with Immunity. In one embodiment, "eliciting an immune response" means generating immunity in an individual in need, for example, to provide a therapeutic effect against a disease, such as an HBV infection. In certain embodiments, "evoked immune response" refers to eliciting or improving cellular immunity against HBV, such as a T cell response. In certain embodiments, "evoked immune response" refers to eliciting or improving a humoral immune response against HBV. In certain embodiments, "evoked immune response" refers to eliciting or ameliorating cellular and humoral immune responses against HBV.

As used herein, the term "protective immunity" or "protective immune response" means that a vaccinated individual is able to control the infection of the pathogen to which the vaccination is directed. Generally, individuals with a "protective immune response" develop only mild to moderate clinical symptoms or are completely asymptomatic. Generally, individuals with a "protective immune response" or "protective immunity" against a pathogen will not die from an infection with that pathogen.

Generally, the administration of HBV vaccines according to embodiments of the present application will have therapeutic purposes for generating an immune response against HBV after HBV infection or the development of symptoms specific to HIV infection, such as for therapeutic vaccination.

As used herein, "immunogenically effective amount" or "immunologically effective amount" means an amount of a composition, polynucleotide, carrier, or antigen sufficient to elicit a desired immune effect or immune response in an individual in need. In one embodiment, an immunogenicly effective amount means an amount sufficient to elicit an immune response in an individual in need. In another embodiment, an immunogenicly effective amount means an amount sufficient to generate immunity in an individual in need, for example, an amount that provides a therapeutic effect against a disease such as an HBV infection. The immunogenic effective amount can vary depending on a variety of factors, such as the physical condition, age, weight, health status, etc. of the individual; specific applications, such as providing protective or therapeutic immunity; and specific diseases such as viruses that need to be targeted infection. A person skilled in the art can easily determine an immunogenicly effective amount according to the present invention.

In a specific embodiment of the present application, an immunogenic effective amount refers to a composition or immunogenic combination (such as an HBV vaccine) sufficient to achieve one, two, three, four or more of the following effects: Amount: (i) reduce or improve the severity of HBV infection or its related symptoms; (ii) shorten the duration of HBV infection or its related symptoms; (iii) prevent the progression of HBV infection or its related symptoms; (iv) make HBV infection Or its related symptoms subsided; (v) preventing the development or onset of HBV infection or its related symptoms; (vi) preventing the recurrence of HBV infection or its related symptoms; (vii) reducing the hospitalization of individuals with HBV infection; (viii) shortening The length of hospitalization of individuals with HBV infection; (ix) increased survival of individuals with HBV infection; (x) elimination of HBV infection in individuals; (xi) inhibition or reduction of HBV replication in individuals; and / or (xii) enhancement Or improve the preventive or therapeutic effect of another therapy.

In other specific embodiments, the immunogenic effective amount is sufficient to reduce the HBsAg content to meet the development of clinical seroconversion; use the individual's immune system to achieve continuous elimination of HBsAg and reduce infected liver cells; induce T specific activation of HBV antigen Cell population; and / or achieving sustained impairment of HBsAg within 12 months. Examples of target indicators include lower HBsAg and / or higher CD8 counts below the HBsAg International Unit (IU) threshold of 500 copies.

As a general guideline, an effective amount of immunogenicity when applied to DNA plastids can range from about 0.1 mg / mL to 10 mg / mL total DNA plastids, such as 0.1 mg / mL, 0.25 mg / mL, 0.5 mg / mL, 0.75 mg / mL, 1 mg / mL, 1.5 mg / mL, 2 mg / mL, 3 mg / mL, 4 mg / mL, 5 mg / mL, 6 mg / mL, 7 mg / mL, 8 mg / mL, 9 mg / mL, or 10 mg / mL. Preferably, the immunogenic effective amount of DNA plastids is less than 8 mg / mL, more preferably less than 6 mg / mL, even more preferably 3-4 mg / mL. An immunogenicly effective amount can be from one vector or plastid, or from multiple vectors or plastids. An immunogenicly effective amount can be administered in a single composition or in multiple compositions, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 compositions (e.g., lozenges, capsules, or injectables ), Wherein the administration of the plurality of capsules or multiple injections generally provides an immunogenicly effective amount to the individual. It is also possible to administer an immunogenicly effective amount to an individual and then to administer another dose of the immunogenicly effective amount to the individual in a so-called initial-plus treatment regimen. The general concept of this initiation-plus treatment course is well known to those skilled in the vaccine field. As needed, additional supplements can be administered during the course of treatment as appropriate.

According to the embodiment of the present application, an immunogenic combination comprising two DNA plastids (for example, a first DNA plastid encoding a HBV core antigen and a second DNA plastid encoding a HBV pol antigen) can be obtained by mixing the two The plastids and the mixture is delivered to a single anatomical site for administration to the individual. Alternatively, two separate immunizations can be performed, each delivering a single plastid. In such embodiments, the first DNA plastids and the second DNA plastids can be weighted by 10 regardless of whether the two plastid systems are administered as a mixture in a single immunization administration or in two separate immunization administrations. : 1 to 1:10, such as 10: 1, 9: 1, 8: 1, 7: 1, 6: 1, 5: 1, 4: 1, 3: 1, 2: 1, 1: 1 by weight: Dosing at a ratio of 1, 1: 2, 1: 3, 1: 4, 1: 5, 1: 6, 1: 7, 1: 8, 1: 9 or 1:10. Preferably, the first DNA plastid and the second DNA plastid system are administered at a ratio of 1: 1 by weight.

In some embodiments, the individuals treated by the method of the present application are infected with HBV, particularly individuals with chronic HBV infection. Acute HBV infection is characterized by efficient activation of the innate immune system coupled with subsequent extensive acquired responses (eg, HBV-specific T cells, neutralizing antibodies), which often results in successful inhibition of replication or removal of infected liver cells. On the contrary, such reactions will be weakened or reduced due to high virus and antigen load, for example, a large amount of HBV envelope protein is produced, and it can release 1,000 times more infectious virus in subviral particles.

Chronic HBV infection is described in stages based on characteristics such as viral load, liver enzyme content (necrotic inflammatory activity), HBeAg or HBsAg load, or the presence of antibodies against these antigens. The cccDNA content remains relatively constant, with about 10 to 50 replicas per cell, even though viremia can vary greatly. The persistence of cccDNA species causes chronicity. More specifically, each stage of chronic HBV infection includes: (i) an immune tolerance period, characterized by a high viral load and normal or minimal elevation of liver enzymes; (ii) an immune-activated HBeAg-positive period, observed at this stage To low or decreased viral replication and significantly elevated liver enzymes; (iii) inactive HBsAg carrying period, which is a low replication state with low viral load and normal in serum that can follow HBeAg seroconversion Liver enzyme content; and (iv) HBeAg-negative phase, during which virus replication (reactivation) occurs periodically and accompanied by fluctuations in liver enzyme content, mutations in the precore and / or basal core promoters often cause infected cells Unable to produce HBeAg.

As used herein, "chronic HBV infection" means that the presence of HBV in an individual can be detected for more than 6 months. Individuals with chronic HBV infection can be at any stage of chronic HBV infection. Chronic HBV infection is understood in terms of its general meaning in the field. Chronic HBV infection can be characterized, for example, by HBsAg persistence for 6 months or more after acute HBV infection. For example, the chronic HBV infection mentioned in this article follows the definition published by the Centers for Disease Control and Prevention (CDC). According to this definition, chronic HBV infection can be characterized by laboratory standards, such as: (i) Negative IgM antibodies against hepatitis B core antigen (IgM anti-HBc) and positive nucleic acid tests against hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg) or DNA related to hepatitis B virus ; Or (ii) a positive nucleic acid test against HBsAg or related HBV DNA, or a positive test against HBeAg at least twice every 6 months.

According to a particular embodiment, an immunogenicly effective amount refers to an amount of a composition or immunogenic combination sufficient to treat chronic HBV infection.

In some embodiments, individuals with chronic HBV infection are undergoing nucleoside analog (NUC) treatment and are inhibited by NUC. As used herein, "inhibited by NUC" means that an individual has an undetectable HBV virus content and a stable alanine aminotransferase (ALT) content for at least six months. Examples of nucleoside / nucleotide analog treatments include HBV polymerase inhibitors, such as entacavir and tenofovir. Preferably, individuals with chronic HBV infection do not develop advanced liver fibrosis or cirrhosis. Such individuals will typically have a METAVIR score for fibrosis of less than 3 points and a fibroscan result of less than 9 kPa. METAVIR score is a commonly used scoring system to evaluate the degree of inflammation and fibrosis by the histopathological evaluation of liver sections of patients with hepatitis B. The scoring system specifies two standardized values: one reflecting the degree of inflammation and one reflecting the degree of fibrosis.

Eliminating or alleviating chronic HBV can allow early disease interception of severe liver diseases, including virus-induced cirrhosis and hepatocellular carcinoma. Therefore, the method of the present application can also be used as a treatment for HBV-induced diseases. Examples of HBV-induced diseases include, but are not limited to, liver cirrhosis, cancer (eg, hepatocellular carcinoma), and fibrosis, particularly advanced fibrosis characterized by a METAVIR score for fibrosis of 3 or higher. In such embodiments, an immunogenicly effective amount is an amount sufficient to achieve a permanent loss of HBsAg and significantly reduce clinical disease (eg, cirrhosis, hepatocellular carcinoma, etc.) within 12 months.

The method according to embodiments of the present application further comprises administering to the individual in need of another immunogenic agent (such as another HBV antigen or other antigen) or another anti-HBV agent (such as a nucleoside analog or other anti-HBV agent) Medicament) in combination with the composition of the present application.

The ability to elicit or stimulate an anti-HBV immune response when administered to an animal or human organism can be evaluated in vitro or in vivo using a variety of standard analytical methods in this technology. For a general description of techniques that can be used to assess the initiation and activation of an immune response, see, for example, Coligan et al. (1992 and 1994, Current Protocols in Immunology; ed. J Wiley & Sons Inc., National Institute of Health). The measurement of cellular immunity can be achieved by measuring the cytokine profile secreted by activated effector cells, including those derived from CD4 + and CD8 + T cells (e.g., cells that produce IL-10 or IFN γ by ELISPOT) By determining the activation status of immune effector cells (e.g., T cell proliferation analysis by classical [3H] thymidine uptake), by analyzing antigen-specific T lymphocytes in sensitized individuals (e.g., in cytotoxicity analysis Peptide-specific dissolution, etc.).

The ability to stimulate cellular and / or humoral responses can be determined by antibody binding and / or competitive binding (see, eg, Harlow, 1989, Antibodies, Cold Spring Harbor Press). For example, the potency of antibodies produced in response to the administration of a composition that provides an immunogen can be measured by an enzyme-linked immunosorbent assay (ELISA). The immune response can also be measured by neutralizing antibody analysis, where virus neutralization is defined as the loss of infectivity caused by the reaction / inhibition / neutralization of the virus via specific antibodies. Immune response can also be measured by antibody-dependent cellular phagocytosis (ADCP) analysis.
Target organization

Target tissues that are particularly suitable for EMTAD using the methods, devices, and systems described herein include both healthy and diseased cells located in, for example, the epidermis, dermis, subcutaneous tissue, connective tissue, and muscle tissue. The technology can also be used in healthy or diseased organs that must be accessed via minimally invasive or other surgical methods. Such target tissues include liver, lung, heart, blood vessels, lymph, brain, kidney, pancreas, stomach, intestine, colon, bladder, and reproductive organs. In some embodiments, a desired therapeutic effect can be achieved by using a method or device described herein to deliver an amount of a medicament to a cell type normally located in the target tissue and other cell types abnormally found in the tissue (e.g., chemotherapy) Agents to treat tumors).

As previously discussed, and as depicted in FIG. 1, a conventional EMTAD lack of precision and reproducibility in the space between the HBV vaccine administered and the time relations with the electrical signal. Compared to traditional EMTAD methods, the present invention describes methods and devices that combine CTAA and ESA to make EMTAD more advantageous for clinical applications. The present invention utilizes various aspects of CTAA in combination with ESA to provide reproducible, consistent, and effective HBV vaccine delivery. The methods and devices provided herein provide space and time control over the administration of HBV vaccine relative to the application of electrical signals, thereby improving the movement and / or absorption of the agent in the target tissue.
method

In one aspect, the invention described herein provides systems, kits and devices for use in the controlled administration of HBV vaccine to individuals in need, followed by ESA. As used herein, "individual" means any animal, preferably a mammal, and most preferably a human, that will be treated or has been treated using the method according to one embodiment of the present application. As used herein, the term "mammal" encompasses any mammal. Examples of mammals include, but are not limited to, cattle, horses, sheep, pigs, cats, dogs, mice, rats, rabbits, guinea pigs, non-human primates (NHP) such as monkeys or apes, humans, etc., and more Better for humans.

In another aspect, the invention described herein provides systems, kits and devices for use in a method for performing ESA followed by the controlled administration of a HBV vaccine. In another aspect, the invention described herein provides systems and devices for a method for the controlled administration of HBV vaccine while performing ESA. These methods include determination of treatment parameters, individual preparation procedures, CTAA, ESA, and other measures, but the scope or order relationship is not limited to this.
Determination of treatment parameters

In some embodiments, the treatment parameters are based on the amount of HBV vaccine required and / or the duration of the dose. HBV vaccine doses may depend, for example, on the specific indication or therapeutic application (such as the type and location of the target tissue), as well as various individual parameters (such as age and body mass). The dose of HBV vaccine can be controlled by parameters related to the administration of HBV vaccine and ESA. Exemplary controllable parameters related to CTAA include, but are not limited to, agent volume, agent viscosity, and injection rate. Exemplary controllable parameters related to ESA include, but are not limited to, characteristics of electrical signals, tissue volume exposed to the electrical signals, and electrode array form. The relative timing and location of CTAA and ESA are parameters that provide further control of HBV vaccine administration.
Individual preparation

In the embodiments described herein, the methods described herein may include individual preparation steps. Individual preparation may include, but is not limited to, disinfectant cleaning and anaesthetic administration, including local or regional, nerve block, spinal block, epidural block, or general anesthesia. In an exemplary case of intramuscular (IM) ESA, a course of treatment that minimizes the effects of electrical stimulation of the muscle can be included in the methods described herein, including thermal control (e.g., cooling the muscle), administration of anesthetic, and / Or an alternative stimulation pattern sufficient to ease discomfort. It should be understood that the choice of individual preparation techniques should not adversely affect the efficacy of treatment, whether or not there are acceptable alternatives. For example, it has been shown in some cases that intramuscular administration of amidines may have an undesirable effect on intramuscular delivery of plastid DNA-based therapies, presumably because these anesthetics have mild muscle toxicity and inhibit Muscle cells exhibit the ability to encode proteins encoded by the DNA sequence to which they are administered.
CTAA and ESA

In some embodiments, a method is described herein in which CTAA and ESA are combined to achieve consistent and reproducible HBV vaccine delivery. In some cases, a device or kit suitable for CTAA is provided, including, for example, a device containing at least one of an automatic injection device and a jet injector.

The present invention provides the ability to percutaneously deploy a plurality of elongated electrodes to a target depth in a safe and consistent manner on a target site of a drug distribution in a recipient with uneven skin thickness and composition to support the application of an electric field in a tissue, thereby enhancing the therapeutic agent or Prophylactic agents, methods, kits and devices for intramuscular, intradermal and / or subcutaneous administration of nucleic acids, pharmaceutical agents, antibodies, peptides, proteins or combinations thereof.

The system and method according to the principles of the present invention can consistently percutaneously deploy a plurality of slender, tissue-penetrating electrodes to a predetermined target tissue site so that an electric field can propagate through the skin, subcutaneous tissue, and / or skeletal muscle. The invention provided herein is designed to enable a user to consistently deploy the electrodes to a target depth with minimal training, while maintaining a proper spatial relationship between the plurality of electrodes, even when applied at sites with different skin characteristics The procedure. Such changes can be attributed to changes in skin characteristics at different locations within the individual or changes in skin characteristics among heterogeneous recipient groups. In other words, the system and method according to the principles of the present invention should allow consistent patterns regardless of the donor or recipient. In some embodiments, the deployment of the electrodes is accompanied by the insertion of one or more injection needles that are configured to administer the HBV vaccine to a target area of the tissue and regarding the ESA The equal electrodes are arranged in a predetermined spatial relationship. In an exemplary embodiment, the electrodes are arranged such that any electrical signals from the electrodes are preferentially applied to the distal end of the site where the HBV vaccine is administered by insertion of one or more injection needles. In another embodiment, the electrodes are arranged such that any electrical signal is preferentially applied to the proximal end of the site where the HBV vaccine is administered by insertion of one or more injection needles.

Aspects of the invention can be used alone or in combination to support percutaneous insertion of electrodes for application of an electric field in vivo, thereby enhancing intramuscular, intradermal and / or subcutaneous application of nucleic acids, small molecule drugs, antibodies, peptides, proteins, and combinations thereof Vote for. In some embodiments, electrode deployment and subsequent electric field propagation are performed in a manner that is coordinated with the distribution of the agent of interest in the target tissue site. In an exemplary embodiment, the administration of the medicament and the application of one or more electric fields are performed in a controlled and monitored manner, thereby maximizing the probability that the distribution of the medicament and the electric field application site are co-located in space and time.

In general, the present invention provides for the percutaneous deployment of electrodes to a predetermined site within the skin, subcutaneous tissue, and / or skeletal muscle of a recipient in combination with the administration of the agent of interest and the application of a localized electric field to improve the delivery and absorption of the agent And / or biological methods and devices. In some embodiments, the implementation of the present invention allows the user to effectively and reliably set up and use the device with minimal training. In another embodiment, the present invention also includes the implementation of various interlocking devices, sensors, and feedback loops to reduce the frequency and / or potential impact of user errors that may occur during the device setup and use. Referring to FIG. 2 , one embodiment of the device described herein includes a "cylinder assembly" 100 removably interfaced to an "applicator" 400 configured to be connected to a controller 700 that acts Electrode activation and subsequent electric field generation, as well as diagnostic and other control routine electrical energy sources. The controller 700 further provides a user interface, a tray, a rack for the applicator 400 , and various other features described. As seen in FIG. 2, with a suitable uniform size and general shape of the reservoir 101 may be inserted into the cylinder assembly 100 in the method of use.

Details of the cartridge assembly 100 presented in some embodiments of the devices described herein are described, for example, in FIGS. 3A - 12 , and the cooperative part of the corresponding applicator 400 is described in FIGS. 13A - 18C . In related cases, then each section controller 700 described in FIG. 19 - 20D are. The rest of the applicator 400 is described next, followed by the rest of the controller 700 .

Referring again to FIGS. 3A - 4 , in some embodiments described herein, the cartridge assembly 100 may include a support structure configured to interface with the applicator 400 and accommodate two or two mounted on the structure to form an array. More than elongate electrodes 122 . In order to avoid the unnecessary propagation of current in the device, the design and materials of the electrode mounting structure should be specified so that appropriate dielectric barriers exist between electrodes of opposite polarity in the device. The distal region 137 of the elongated electrode is bonded to the mounting structure using standard mechanical features and / or adhesives suitable for the electrode mounting structure and the material composition of the electrodes.

In an exemplary embodiment, the configuration of the cartridge outer shell structure 102 interfaces with a fluid reservoir 101 containing a HBV vaccine, wherein the configuration of the reservoir 101 and the cartridge shell structure 102 is operatively connected to at least An injection orifice (needle 105 ) through which the HBV vaccine is administered into the target tissue. In some embodiments, this configuration facilitates the co-location of the distribution of the agent of interest and the site of application of the electric field. In another embodiment, this configuration facilitates a predetermined spatial relationship between the devices used for ESA and CTAA. In another exemplary embodiment, the syringe 101 is inserted into the barrel 100 , wherein when the barrel is loaded into the applicator 400 , the syringe 101 moves forward to mate with the needle interface 152 and connect the barrel to the needle.

Certain embodiments of the invention may include the use of a syringe, vial, ampoule, cartridge or equivalent structure to store one or more HBV vaccines. In some embodiments, the reservoir may include at least one of glass and plastic, where the selection of the material is compatible with the agent of interest. Coatings can be applied to the reservoir to provide the required lubrication or protection characteristics. As disclosed above, the electrodes 122 may be hollow and in some cases configured with injection orifices that are operatively connected to a fluid reservoir. Alternatively, the injection orifice may include one or more hypodermic needles and / or needleless injection ports positioned relative to the electrodes. The choice of the type and size of the injection orifice can depend on the desired route of administration, tissue distribution, and physical characteristics of the agent of interest. In one embodiment, the cartridge structure is designed to ensure a predetermined spatial relationship between the injection orifice and the electrode in the deployed state, so that the distribution of the agent of interest is basically defined by the conductive areas of the plurality of electrodes Happen in the organization. To minimize the need for the user to handle sharps, in one embodiment, the configuration of the barrel 100 designed for a hypodermic needle allows the hypodermic needle to fit into the barrel at the time of manufacture, rather than More common practice of fitting a needle to a syringe during use. In certain embodiments, one aspect of the present invention may be incorporated in the barrel and / or needle to ensure the retention of the needle during manufacture, distribution, handling and use, and to ensure that the reservoir is in use before use Features that fit properly with the needle. In some embodiments, such features can minimize the risk of the agent leaking from the reservoir or the reservoir orifice interface due to rupture or improper fit.

In some embodiments, the cartridge may include a tissue contact interface at the distal end of the sub-assembly. In some embodiments, the tissue contact interface includes a substantially planar structure that is oriented perpendicular to the direction of elongation of the electrodes and has one or more pores, the configuration of the one or more pores allows the electrodes to pass through Contact interface through tissue. For embodiments incorporating an integrated reservoir and injection orifice, the interface also has an aperture to accommodate the injection orifice or needleless injection. In order to minimize the risk of contamination of these electrodes and injection needles and the occurrence of unintended exposure of sharp objects to users or recipients, in an embodiment, the pores that accommodate these electrodes and injection needles have Suitable for preventing accidental contact with these electrodes and needles. Most commonly, the tissue contact interface consists of one or more plastics suitable for at least short-term tissue contact.

In order to avoid cross-contamination of biological materials that may occur between recipients, the cartridge assembly 100 is configured for single use. In some cases, the cartridge includes one or more mechanical, electrical, and / or identification elements that define the cartridge for a single administration. Examples of mechanical elements having this property include, for example, but not limited to, latches and / or stoppers that fasten electrode mounting structures and / or puncture shields (see below) that are in a deployed state after use. Examples of electrical components include, for example, fuses or connections that are configured in series with one or more electrodes, which are deactivated under the effect of electrical energy at the end of the first use of the cartridge. Examples of identification elements include, for example, serialized radio frequency identification devices, bar codes or quick response codes, the configuration of which is read by an applicator and / or an electrical energy source. Identification information about a particular cartridge can then be used to prevent the cartridge from being accidentally or intentionally reused by the applicator and / or energy source. In one embodiment, one or more redundant features are incorporated to minimize the potential for reuse of the cartridge.

In one embodiment of the arrangement described herein, as shown in FIG. 2, the applicator 400 comprises a configuration wherein the support structure which contact the dielectric tube assembly 100, UI 410--418 (FIG. 13B), the conductive The electrical connection is configured to operatively connect the conductive contact area located on the distal region of the electrode with the electrical energy source when the elongated electrode is deployed in the recipient's target tissue. In some cases, the user interface includes a handle, one or more display features designed to communicate information to the user, and one or more features capable of accepting user input. In some embodiments, the configuration of the display features communicates the operating status of the device during setup and use, and related warning / error messages. Such displays may include mechanical features, lights, alphanumeric displays, and / or electronic display screens. In some cases, the configuration of features that can accept user input allows the user to disable the safety features in the device to prevent accidental ejection at appropriate stages of the procedure, selecting specific parameters of the procedure (such as a predetermined injection depth), And initial program administration, and may include buttons, triggers, mechanical sliders and / or levers.

In some embodiments, the applicator 400 also includes an actuating mechanism that interfaces with the barrel assembly and is configured for percutaneous deployment of the electrodes, positioning the injection orifice relative to the target tissue, and paying attention to The medicament is discharged from the reservoir through the orifice and sent to the target tissue site, and / or an electrical signal is transmitted from an electric field generator such as the controller 700 to the cartridge 100 . The applicator 400 is configured so that the user can supply energy to actuate the mechanism, or more preferably, the device can incorporate one or more non-biological energy sources operatively connected to the actuation mechanism within the applicator . Such non-biological energy sources include, for example, electromechanical devices (solenoids, motors, lead screws), mechanical components (springs and related devices), and compressed gases.

One exemplary embodiment of the cartridge assembly 100 as described in FIG. 3A, the cartridge assembly 100 includes a reservoir 140 and a load port for receiving and containing a medicament reservoir 101. The reservoir 142 is closed. Because a device that will generate an electric field and be used as part of a therapy requires an electric field generator, such as the controller 700 to electrically interface with a device containing electrodes configured to contact the target tissue, a cartridge assembly 100 is needed. Since the configuration of the controller is for multiple use, and the reservoir 101 can be scheduled for a single use, there can be a cylinder assembly 100 to hold the electrode 122 and the reservoir 101 , and reusable devices Interfaced and used to configure the cartridge assembly 100 for single use. Therefore, the applicator 400 may be a reusable component and the configuration of the cartridge assembly 100 is for a single use. The cartridge assembly 100 can also be placed in a certain state to prevent subsequent use in the event of an error or interference when inserting the reservoir 101 , or in the event of a defect.

The term reservoir 101 may refer to a syringe, a vial, or a device with an orifice, such as any other device that interfaces with a needle, that can hold a drug or HBV vaccine and that can be shown as needle 105 with needle interface 152 in FIG. 4 . For a given type of cartridge assembly 100 , the reservoir 101 generally has a common shape and size. Each component within the cartridge assembly 100 allows for certain errors in precise dimensional design and / or manufacturing tolerances, but generally requires a common shape and size to reduce the erroneous delivery of drugs not labeled for the cartridge 100 with the device risks of. If the user does not provide an appropriately sized reservoir 101 , one or more interlocking devices in the cartridge assembly 100 may not be deactivated and the system may be unavailable until the appropriately sized reservoir 101 is inserted.

As seen in Figure 3B, the reservoir 101 may be equipped with a port generally for drug release and the plunger 156. A removable cover 158 may also be provided to maintain the sterility and integrity of the medicament prior to insertion and use of the reservoir. The port for drug release may be proximal to the needle interface 152 used, and the plunger may be opposite this port for drug release. As an alternative to an open port for drug release, in some embodiments, the reservoir is configured with a baffle assembly to cover and seal the opposite end of the container from the plunger. The separator may be made of an elastic compound, such as silicone or butyl rubber, and the specific formulation and coating of the material are selected for stability and compatibility with the medicament contained in the reservoir. The baffle assembly is usually held in place by a crimp seal or other securing mechanism. This partition seal configuration avoids the need for a removable cap, but requires the needle 105 to be equipped with a suitable piercing member such as a needle, spike or other access to the fluid contained in the reservoir feature. Specific implementations of this configuration include a bilateral needle configuration and a vial adapter with a pointed tip.

The cartridge assembly 100 is not only configured to receive the reservoir 101 , but also configured to be received in the applicator cartridge assembly receiving port 401 of the applicator 400 ( FIG. 2 ). As such, the cartridge assembly 100 includes a device that allows the applicator 400 to at least partially pull and hold it within the internal volume. In some embodiments, the device is one or more racks on the surface of the barrel assembly 100 that engage a corresponding motorized pinion assembly in the applicator 400 . In other embodiments, the applicator 400 can interface with the cartridge assembly 100 without the need to pull it into the internal volume. In still other embodiments, other techniques can be used to engage the cartridge assembly 100 with the applicator 400 , such as a motorized track or bracket and the like, to which the cartridge assembly 100 can interface.

The applicator 400 further has an interface element that enables it to control certain actions within the cartridge assembly 100 . In detail, the configuration of the applicator 400 can use various subsystems to control needle insertion, drug delivery, electrode insertion, and electrode activation. In some cases, the steps are connected such that a single action of the applicator 400 starts multiple of the steps. In some embodiments, the steps other than drug delivery and electrode activation are caused by a single action, as described in the following exemplary embodiments.

After the electrical or optical signals, such as the use of mechanical, electrical, or optical components transmitted to the cartridge assembly 100 are properly activated, the applicator 400 can be activated to test the subsystems within the cartridge assembly 100 to ensure that it operates correctly and is properly assembled State for drug delivery and electric field application. By way of example, such subsystems including applicator 400 can be tested to ensure that the cartridge assembly 100 has not been previously used; the reservoir has been properly placed in the cartridge assembly; when applied via the alignment guide / tilt shield 108 Appropriate force is applied to the body of the individual; the test user has positively selected the depth; and the outer canister cover 110 has been removed. In addition, the configuration of the applicator 400 can monitor the functional status of the cartridge during program execution. For example, such subsystems including the applicator 400 can be tested to ensure that the electrodes 122 are properly deployed in the individual before drug administration begins; before the electric field is applied, the plunger in the reservoir has been properly activated. The user maintains an appropriate force on the individual's body during the administration process, and so on.

In addition to the subsystems operated by the applicator 400 , the cartridge assembly 100 may also incorporate appropriate subsystems, including subsystems that interact with the applicator 400 and subsystems that do not so interact, to achieve the goals of drug delivery and electric field application therapy. These subsystems include subsystems used to cause needle and electrode insertion; subsystems used to protect users from sharp objects after therapy administration; subsystems used to provide different needle / electrode insertion depths; Subsystems that have exerted an appropriate force on the recipient's organization before the process was initiated and subsequently during the application of the administration process, and so on. Although generally described in the context of deployable needles and electrodes, it should be noted here that this situation is not absolutely necessary and that systems with needles and electrodes that are not deployable or fixed also benefit from Systems and methods, including the described subsystems.

In an exemplary embodiment, as shown in FIG. 4 , the cartridge assembly 100 includes an outer cartridge 102 , referred to as a housing in some cases. The outer cylinder 102 is terminated at the distal end by an outer cylinder cover 106 . The outer cylinder 102 includes an inner cylinder closed body 150 for receiving the inner cylinder 103 , the inner cylinder being received and movable in a slidable manner relative to the outer cylinder 102 . The inner cylinder 103 includes a reservoir enclosure 142 in which the reservoir 101 can be placed. The inner cylinder 103 is engaged with the inner cylinder cover 104 at a distal end. The inner cylinder cover 104 has multiple functions, including locking the electrode 122 in place (the inner cylinder 103 itself has a seam for the electrode 122 to be placed therein) and provides a support surface for the puncture shield 134 . Locking the cover to the inner tube 104 on the inner cylinder 103.

The barrel end 112 is received in a portion of the reservoir enclosure 142 in the inner barrel 103 , in a portion opposite to a portion of the reservoir cap 104 in the reservoir closed body. The reservoir detection cover 118 engages the barrel end 112 via the reservoir detection spring 116 . The barrel lock ring 114 locks the system in place, including locking the barrel end 112 to the inner barrel 103 . The reservoir detection spring 116 is also used to push the reservoir 101 into engagement with the needle interface 152 , and is also used to adjust the tolerance of the size of the reservoir 101 .

The reservoir interlock 120 provides a mechanical interlock to prevent inadvertent or unnecessary actuation of the cartridge function. In detail, the reservoir interlocking device 120 , also referred to as the first reservoir insertion trigger, is a finger-like member placed below the inner cylinder 103 and having a slot or hole extending through the inner cylinder 103 ( (See Figure 5B ). These fingers prevent the barrel end 112 from slidingly moving relative to the inner barrel 103 , and in particular, prevent the barrel end from facing the inner barrel 103 in the inner barrel 103 before inserting the reservoir into the reservoir enclosure 142 . The cover 104 moves.

When the reservoir 101 is correctly inserted into the reservoir enclosure 142 , the reservoir interlocking device 120 is pushed down and the fingers are pushed downward so that it no longer extends into the reservoir enclosure 142 . This downwardly pushed or lowered configuration of the reservoir interlocking device 120 can also provide an auditory, tactile or tactile "click" that can inform the user of the correct insertion. Once depressed, the barrel end 112 is no longer blocked by the fingers of the reservoir interlock 120 , and then allowed to move, and in particular, allowed to move in the direction toward the inner barrel cap 104 .

When the barrel assembly 100 is inserted into the applicator 400 in the manner described below, the barrel end 112 is caused to move in this manner under the action of the spring cover / cylinder interface 470 . When the barrel end 112 moves far enough forward, it locks in place, securing the reservoir 101 in the reservoir enclosure 142 and ensuring that it is properly positioned relative to the needle interface 152 to ensure self-reservoir 101 Complete fluid path to the orifice of needle 105 .

For an embodiment of a device that incorporates an injection needle into the barrel 100, a standard "off-the-shelf" single-use hypodermic needle can be used within the device. However, the operation and reliability characteristics of the device can be improved by incorporating custom-designed elements that are not present in hypodermic needles intended for use in conventional parenteral administration procedures. Particular aspect of the needle hub 152 may include a sample of the material constituting the interface is prevented from holding the needle hub 152 wherein the inner tube 103 is incorporated into the shift during dispensing and use, and the orientation of the needle with respect to any feature of the bevel of the interface.

Conventional single-use disposable injection needles typically consist of injection molded polypropylene thermoplastics. However, for many applications, the impact strength, tensile strength, and flexural strength of polypropylene may not be suitable for ensuring the integrity of the interface when subjected to the forces unique to needle deployment and injection with this device. Specific faults of interest include damage to the interface wall caused by impact or injection force, and damage to the interface needle joint caused by the same cause. Although the design of the interface, including its geometry and wall thickness, can be used to prevent such failures, the design is changed to fully prevent the interface from being damaged, while ensuring that the interface remains as the International Standards Organization (ISO) ISO 80369-7: 2016 Small-bore connectors for liquids and gases in healthcare applications-Part 7: Connectors for intravascular The dimensional characteristics required for proper mating with a conical male luer slip connector, as described in the relevant standards disclosed in the or hypodermic application, are not always feasible. Specifically, in view of the forces experienced by the syringe needle and interface during deployment, in some embodiments, materials with improved impact strength, tensile strength, and flexural strength are used. One example is the use of injection molded polycarbonate plastics (such as ZELUX® GS, Makrolon, or Lexan) or copolyesters (such as Eastman Tritan ™ Copolyester MX731, MX711, and MX 730). When evaluated according to ISO 180: 2000 Plastics-Determination of Izod impact strength, a notched impact strength of at least 70 kJ / m2 is considered suitable for this application. When evaluated in accordance with ISO 527-1: 2012 Plastics-Determination of tensile properties-Part 1: General principles (Plastics--Determination of tensile properties--Part 1: General principles), a tensile strength of at least 30 MPa is considered appropriate. In this application. When evaluated in accordance with ISO 178: 2010 Plastics--Determination of flexural properties, a flexural strength of at least 50 MPa is considered suitable for this application. In some embodiments, the particular resin selected exhibits compatibility with a predetermined sterilization method, such as gamma radiation, while not exhibiting adverse changes in physical properties that may impair its function.

For embodiments utilizing a custom injection needle, one or more mechanical features that are not normally present on a conventional syringe interface to enable the device to be inserted into the inner barrel 103 are included. Such features may include tabs, latches, or ridges with corresponding mechanical features positioned on the inner barrel 103 . In some cases, the implementation of these features makes the interface cooperate with the inner cylinder 103 in a consistent orientation. Combined with a needle manufacturing method capable of orienting the bevel or other needle orifice features uniformly, thereby ensuring that deviations in the injection position or drug distribution caused by the orifice position and design can be considered in the device design. For example, a needle with an asymmetrically penetrating tip feature, such as a beveled incision, may exhibit a directional deviation during deployment into the tissue due to the interaction between the tissue and the asymmetrically penetrating feature on the needle. If the electrodes have symmetrical penetrating tip features (such as a trocar tip), the electrodes will not exhibit corresponding deviations in their deployment features. Therefore, the mounting feature for mounting the needle interface 152 on the inner cylinder 103 may include a positional offset of the injection orifice on the needle 105 relative to the electrode 122 before deployment, thereby causing the expected deployment of the asymmetrical bevel of the needle feature. The exact size of the offset may depend on the nature of the target tissue and the range of expected penetration depths, and in some embodiments, for each 10 mm penetration depth, the needle is offset by 0.5-1 mm. Such features advantageously ensure co-localization of the drug distribution and the application of the electric field when using electrodes and injection needles with different tip shapes or when the tip shapes must be aligned with each other consistently.

The incorporation of the syringe detection cap 118 mounted to the syringe detection spring 116 ensures that the cartridge assembly 100 can receive the syringe 101 and position the syringe correctly relative to the needle interface 152 within the manufacturing tolerances expected for the syringe 101 . When the spring cover / canister interface 470 within the applicator 400 moves distally relative to the canister assembly 100 , the applicator 400 moves the cannula end 112 forward during the loading procedure. This action occurs when the cartridge assembly 100 is loaded into the applicator 400 and the cartridge assembly 100 is pushed into the applicator 400 under the action of a loading mechanism such as a rack and pinion mechanism described below, for example.

The movement of the barrel end 112 can be used as a second interlocking device. Certain words, in one embodiment, as shown in Figure 5C - 5D seen, the receiver in the cartridge assembly within the sensor 403 appropriately configured locking hole 144 via a set of reservoir '(FIG. 5D) visible and Detect sight. This line of sight is visible when the cartridge assembly 100 is loaded into the applicator 400 . The visible line of sight or obstruction of the line of sight can serve as part of the second interlocking device, which must be disabled to allow the controller 700 to allow activation and triggering of the device, including needle and electrode insertion, drug delivery, and electrode activation.

For example, in one embodiment, the reservoir locking hole 144 ' ( FIG. 5D ) must be closed for the device to operate. If, for example, the presence of visible line of sight is detected by a pair of IR or visible light transmitters and detectors within the applicator barrel assembly receiver 403 , the device can be rendered inoperable and displayed on the applicator display 404 and / Or the controller 700 , including the controller display 712 , generates and displays an error message to inform the user of the status of the device and the recommended steps to be taken to resolve the error.

Therefore, in this embodiment, an error condition may be that no reservoir 101 is loaded into the cartridge 100 or that the syringe is not properly seated in the inner cartridge 103 . In this case, since there is no reservoir 101 that performs this action, the reservoir interlocking device 120 cannot be depressed. In this case, the barrel end 112 cannot move forward toward the inner barrel cover 104 in the distal direction. The construction of these components can cause the open tail state to produce an open line of sight 146 through the first reservoir lock hole 144 ' ( Fig. 5D ). The tail end 112 of the auxiliary inspection system can not be closed, and this itself can be manifested as that the cylinder cannot move backward or distal to the required distance to enter the cylinder assembly receiver 403 . Since the system defines these conditions as error states, they can be identified and used to generate error messages, such as on the user interface on the applicator display 404 and / or the controller 700 , including the controller display 712 . Present appropriate information. A similar error condition may occur if the reservoir 101 is improperly loaded, or if the reservoir interlock 120 is damaged. Generally, in this case, the appropriate error message may be accompanied by instructions that require the user to remove the cartridge, reinstall a new reservoir, and attempt to reintroduce the cartridge assembly 100 into the applicator 400 .

Another error state may be that the user manually moves the barrel end 112 forward without the reservoir 101. This situation occurs when the user manually ejects the reservoir interlocking device 120 from the reservoir enclosure 142 . . This situation can also be defined as an error state, and since another (second) group of reservoir lock holes 144 ( FIG. 5C ) is placed on a part of the outer cylinder 102 , this situation can be detected. If the reservoir is not in place, but the barrel end 112 is moved forward by the depressed reservoir interlocking device 120 , the reservoir locking hole 144 ' ( Fig. 5D ) and the reservoir locking hole 144 ( Fig. 5C ) Alignment, regenerating the open line of sight 146 and then presenting an error state. This error condition may also occur when the reservoir interlock 120 is damaged and its fingers are no longer inside the reservoir enclosure 142 . In one embodiment, this error condition may not be remedied by attempting to reinstall the reservoir, as the reservoir may not fit in the reservoir enclosure 142 locked at the end 112 of the barrel. In one embodiment, a new cartridge assembly 100 is required .

In contrast, if the reservoir 101 of the appropriate size is correctly positioned in the proper position, it pushes the reservoir detection cover 118 backward against the reservoir detection spring 116 , and the movement of the reservoir detection cover 118 The reservoir locking hole 144 ( FIG. 5C ) and the reservoir locking hole 144 ' ( FIG. 5D ) are closed. In this case, there is no error state, thereby allowing the device to operate. The closure and the detection of the closure occur in the main body of the applicator 400 after the cartridge assembly 100 is inserted, and therefore, are not easily affected by a user's attempt to resist the interlocking device, whether intentionally, accidentally, or caused by a defect of. It should be noted that even if the user intentionally or unintentionally closes the tail end 112 itself during the processing of the cartridge 100 , this "error-free" state will still occur.

Depending on what error condition occurs, the cartridge assembly 100 may remain usable or unusable. If the barrel end 112 is locked in place, the barrel assembly 100 is rendered unusable. However, if the barrel end 112 is not locked in place, the barrel assembly 100 can be removed from the applicator 400 and a new reservoir 101 can be inserted.

Although it has been found that the set of two interlocking devices described above (one is a mechanical interlocking device using a reservoir interlocking device 120 and one using a light emitter and collector and a reservoir latching hole 144 and 144 ' ) is particularly special in some embodiments Useful, but it should be understood that other types of interlocking devices can also be used ( Figures 5C - 5D ). For example, the configuration of the error state occurs when the reservoir lock hole 144 is closed (and the clear line of sight 146 corresponds to a non-error state) (via changes in the position of the reservoir lock hole and program logic), rather than An error condition occurs when the reservoir lock hole 144 is not closed. The reservoir interlocking device 120 may incorporate additional mechanical marking features that are recessed into the barrel when the interlocking device is active, but change with time when the syringe 101 is properly inserted and the reservoir interlocking device 120 is pressed into its unlocked position. It must be visible to a properly configured sensor. In other variations, other ways may be used to determine whether there is a line of sight, such as optical, acoustic, electrical, or similar, as long as it is suitable for the emitter and the collector to be positioned within the applicator 400 . In view of this teaching, those skilled in the art should understand that other methods can also be used to determine whether the reservoir 101 is properly loaded, such as mechanical technology. Depending on the implementation, if an error condition is detected, the applicator 400 may prevent operation with the cartridge assembly 100 in place, or the applicator 400 may even prevent receiving the cartridge assembly 100 first . For example, the cartridge may be designed such that the reservoir interlock 120 includes a mechanical tab or locking feature that extends from one or more cartridge surfaces before the syringe 101 is properly inserted into the cartridge 100 . The mechanical tabs are designed to interact with corresponding lock pin features located in the applicator 400 , thereby physically blocking the loading of the barrel 100 in the applicator 400 unless the reservoir interlock 120 is unlocked. This mechanical interaction will provide feedback to the user or the system that the error condition must be resolved before loading the cartridge 100 into the applicator 400 . In other variations, more or less than two interlocking devices may be provided, but the interlocking devices may be associated with different security types accordingly.

In certain embodiments, other features may also be used in the above determinations, or to facilitate the above determinations. For example, when the cartridge assembly 100 is pulled into the applicator 400 using a motor, a sensor may be used to detect the spatial position of the cartridge assembly 100 during the insertion procedure as described below. In other words, the applicator 400 can detect that the cartridge assembly 100 is inside the cartridge assembly receiver 403 . In some cases, this may allow for additional error states to be determined, either directly or by prompting another sensor to activate to evaluate the state of the device. For example, in a used canister, the cannula end 112 is locked into place. If it is intended to reuse the used cartridge assembly 100 , the optical detector detects that the cartridge assembly 100 is at a different position from the unused cartridge assembly 100 . Using an electric motor to implement one or more system functions also provides an opportunity to monitor its operating status, including the amount of voltage and current supplied to the motor and the number of revolutions the motor has performed during a particular operation. Measurements of these quantities during system operation can be used as the primary or secondary method for detecting possible or actual fault conditions. For example, a mechanical interlocking device designed to prevent the loading procedure when the cartridge 100 is not properly configured may be coupled with sensors and logic circuits that monitor the motor to ensure that the cartridge 100 is properly loaded into the applicator 400 . For example, the interaction of the mechanical features described above designed to prevent loading of the cartridge 100 when the reservoir 102 is not properly inserted will increase the load on the motor drive mechanism, resulting in higher current draw. Detecting an increase in the current draw of the motor will prompt the loading procedure to stop and display the fault condition to the user, such as on the stimulator display 712 and / or the applicator display 404 .

As it may happen that a drug that is not intended to be administered using this method is contained in a reservoir having a size and configuration similar to that intended for the delivery method. Therefore, another aspect of the system incorporates one or more methods to ensure that the reservoir 101 inserted by the user into the cartridge 100 is explicitly intended for the device. The implementation of such features will reduce the risk of administering the wrong drug to a given individual. Conventionally, specific information on user instructions and drug labeling include the route and method of administration. However, to further reduce the possibility of user error, it may be necessary to incorporate mechanical, optical, and / or electrical features within the reservoir and device. In one embodiment, the syringe may be designed to incorporate one or more unique mechanical features not present in other reservoirs, and the syringe may be similar to the reservoirs designed for this method of delivery. For example, the reservoir may be designated to incorporate ribs or other elongated features on the flange or barrel of the reservoir. In this embodiment, a corresponding mating feature will be included on the reservoir interlocking device 120 so that the reservoir interlocking device will be deactivated only if a reservoir with appropriate mating characteristics is properly inserted into the device. Where it is not feasible to implement this feature directly in the design of the reservoir, an alternative embodiment would include placing a second mechanical component on the reservoir, which would be unique to the reservoir intended for the device. For example, a ring or other suitably configured feature designed to slide on the barrel of the reservoir can be used, with the outer cylinder 102 , the inner cylinder 103 , the reservoir interlock 120 , the reservoir detection Corresponding features in the cover 118 or other suitable features in the cartridge 100 cooperate to "key" the reservoir for the device. Further embodiments are custom indicia of suitable size, color and / or conductivity applied to predetermined locations on the outer surface of the reservoir intended for the device. The configuration of the corresponding optical or inductive sensor in the applicator 400 is used to evaluate the presence or absence of the mark on the surface of the reservoir in order to verify that the drug inserted into the cartridge is intended for the device. Detection methods will include the use of optical or electrical signals applied to the surface of the marker in order to assess its presence or absence. In this way, a reservoir containing a drug that is not intended for the device (and therefore missing associated labels) can be detected and possible misuse prevented.

The configuration of the sensor for performing the cartridge loading and syringe detection determination described above will now be described. Such sensors further form part of the cartridge loading sub-assembly within the applicator 400 . In more detail, and referring again to FIGS. 6 , 17B, and 18C , the exemplary way of detecting the position of the cartridge assembly 100 is by using a cartridge loading sensor 436 and a cartridge loading sensor 438 , which form a loading drive sub-assembly 454 of a portion of the sub-assembly 454 further comprises a cylindrical guide 442 and the loading motor 444, the loading motor has a connecting member connected to the pinion gear assembly 448, the upper pinion assembly via the base of the outer cylinder 102 of the The rack 154 pulls the cartridge assembly 100 into the cartridge assembly receiver 403 . In more detail, when the cartridge loading sensor 436 detects the start mark 172 (see FIG. 6 ) on the cartridge assembly 100 , the motor can start loading. When the cartridge loading sensor 438 detects the same flag, the loading can be stopped. A continuation flag 174 may be used, the presence of which is required for loading to proceed.

The configuration of the first "tooth" of the rack 154 is to provide the user with tactile sensation (or hearing or tactile sensation) when the user inserts the cartridge assembly 100 into the cartridge assembly receiver 403 . Such configurations may include the shape and / or size of the rack teeth 154 and the amount of deflection allowed to position them in the outer cylinder. By using a rack and pinion implementation, the required degree of tactile feedback can be achieved while ensuring that it does not provide a large force to the loading motor 444 to prevent receiving and loading the cartridge assembly 100 .

Referring again to FIG. 17A , and as indicated above, the cartridge assembly 100 is inserted into the cartridge assembly receiver 403 within the applicator 400 . Although this insertion can be performed in various ways, a particularly useful way found is by means of a pinion assembly 448 that engages a rack 154 on the outer cylinder 102 . Using more than one rack provides additional stability, particularly torsional stability during the loading phase. Referring also to the insertion / injection driving assembly 456 of FIG. 18A , in addition to pulling the cartridge assembly 100 in the cartridge assembly receiver 403 , the insertion action also compresses the electrode / needle insertion spring 472 via the spring cover / cylinder interface 470 . The electrode / needle insertion spring 472 is used as the main driving force for needle and electrode insertion during drug delivery.

This hybrid motor / spring effect provides multiple benefits. The motor drive is advantageous in that it is highly controllable and allows the cartridge 100 to be loaded into the applicator 400 in a semi-automatic manner with minimal user input of mechanical force. As described above, the implementation of the motor-based mechanism provides monitoring of the operating status of the system. For example, the logic and control circuits that pass current draw and revolution counts to the system provide a complementary method for detecting and diagnosing possible fault conditions. Notwithstanding these advantages, in some cases, electric motors may not be suitable for effective transdermal deployment within a short enough time scale that includes the plurality of elongated electrodes and is ideal for the sequence of hypodermic needles in selected embodiments. Apply the required linear force. In detail, penetration of skin tissue can be achieved most consistently by applying large linear forces within a short time scale. In some embodiments, the most advantageous insertion feature is achieved when the electrode is penetrated, and if present, the injection needle contacts the skin at a higher speed. This is because sharp objects that travel to the skin contact point at a higher speed cause less tissue deformation when cutting or penetrating the tissue. Therefore, in some embodiments, it is desirable for the sharp object to accelerate rapidly before contacting the skin. In some embodiments, multiple electrodes and injection needles are often utilized. In another embodiment, the speed of the electrode is at least 50 mm / sec before contacting the skin. In yet another embodiment, the speed of the electrode is at least 500 mm / sec before contacting the skin. This deployment method minimizes the discomfort experienced by the individual during electrode penetration and is most beneficial to maintaining a consistent spatial relationship between the plurality of electrodes. Compared with the electromechanical motor, the spring drive mechanism exhibits a more favorable ejection pattern, which can give the electrodes and injection needles the rapid impulse required for percutaneous electrode implantation. In detail, the force exerted by the compression spring reaches its peak when initially discharged. It is advantageous to achieve high speed at the skin contact point, and due to the viscoelasticity of the skin tissue, the maximum force is required to penetrate the skin, especially when the skin is in contact with multiple electrodes and / or injection needles. Institutions are good for percutaneous deployment. In addition, spring-based mechanisms can generate this force from a simple, durable, and compact form factor that can be easily integrated into the form of a handheld device. However, the disadvantage of the spring-based mechanism is that it usually requires the user to input a large amount of mechanical force to trigger its operation, especially for springs with higher force constants and / or larger displacements. As described in the present invention, a hybrid motor and spring mechanism is used to achieve the required deployment force characteristics, while the user is simple to operate. Although the hybrid motor and spring mechanism is a preferred embodiment, depending on the implementation, other hybrid mechanisms incorporating two or more drive mechanisms, one of which can generate a rapid impulse and the other can trigger the impulse mechanism, For example, a pump capable of compressing the gas into the chamber and then discharging the compressed gas in order to apply an impulse to deploy the electrode, and a hypodermic needle may also be used when applicable.

In any event, once loaded, the depth of electrode deployment and / or medicament administration required by the physician or other drug administrator is positively selected and transferred to the applicator 400 . Referring again to FIG. 13B , the depth can be selected by the depth selection button 409 (or other equivalent interface such as a jump switch or slide switch) and the result can be displayed on the injection depth selection indicator 408 (or, similarly, other equivalent interfaces) on. The available injection depth is communicated to the user by appropriate markings of the applicator 400 and / or the cartridge 100 . In some embodiments, any markings regarding the depth of injection are located on the cartridge 100 and remain visible to the user after installation in the applicator 400 . For example, the available injection depth may be marked on the upper surface of the alignment guide / tilt shield 108 . In order to avoid the user from forgetting or ignoring the choice of injection depth, preferably, the device does not allow the user to continue the administration process until such a positive choice is made. This can be achieved by implementing appropriate logic controls within the system so that subsequent device settings or usage of the device are not available until the user types in a valid depth selection. In certain embodiments, when the user is prompted to positively select the injection depth, the controller display can convey to the user information about an appropriate method for assessing the individual and determining the injection depth suitable for the selected administration site.

Referring again to FIGS. 18A - 18B , when the cartridge assembly is properly inserted, the spring cover / cylinder interface 470 engages and presses the spring cover hole 471 and the tab 491 . Although a large spring force presses the inner cylinder 103 , the inner cylinder is prevented from moving forward by engaging a set of holding posts 488 that abut against the side wall 494 of the outer cylinder 102 . However, the force applied by 470 tilts apart the tab 491 (for preventing the lock ring from inadvertently rotating during processing), thereby rotating the cartridge lock ring 114 under the action of the motor driving mechanism. Rotation of the cartridge lock ring 114 causes rotation of the holding post 488 . The holding post 488 can be rotated into a channel of a first depth 490 or a channel of a second depth 492 . The length of the channel of the first depth 490 corresponds to one depth selection, and the length of the channel of the second depth 492 corresponds to another depth selection, where one depth or the other depth is selected by the user using the button 409 . For example, the length of the channel 490 can be in the range of 20-30 mm and the length of the channel 492 can be in the range of 12-20 mm. The user's affirmative choice of depth requirements provides yet another chain. Without a positive selection, the applicator may not allow activation / needle insertion. Therefore, according to the regulations of the user, the clockwise or counterclockwise rotation guided by the applicator 400 causes the barrel lock ring 114 to rotate. The need to rotate a set of struts into such channels to enable the deployment of electrodes and (if present) injection needles greatly reduces the chance of accidental discharge, even during severe vibrations or drops.

In more detail, the rotation of the cylinder lock ring 114 is transmitted to the cylinder assembly 100 through the retaining column 488. When the cylinder assembly 100 is correctly inserted, the retaining columns are arranged in the grooves on the insertion mechanism gear drive ring 478 . . In FIG. 18A , the grooves on the insert mechanism gear drive ring 478 are disposed at the 3 o'clock and 9 o'clock positions. The insertion mechanism gear drive ring 478 is mounted to a part of the insertion gear ring 479 which is driven by the insertion mechanism drive motor 482 . This part of the insertion gear ring 479 is driven to rotate the insertion mechanism gear drive ring 478 clockwise or counterclockwise. The mark 481 on the ring 480 and the accompanying insertion mechanism position sensor 483 are used to determine the position of the insertion mechanism gear drive ring 478 , and further used to drive the insertion mechanism gear when the applicator 400 is reused in another cylinder The ring returns exactly to the 3 and 9 o'clock positions.

The above embodiments provide various advantages. For example, the user must effectively perform the deep selection step before proceeding with the dosing process. In doing so, the user must evaluate the injection depth appropriate for the selected injection site and prepare the site as described in the instructions or the instruction manual record. As mentioned, the insertion mechanism that requires rotational movement for deployment obviously withstands accidental deployment caused by shedding, dripping, vibration, and the like.

Those skilled in the art will understand the changes. For example, although two channels and two retaining columns are used for each depth, one channel and one retaining column may be used. Various types of motors and mechanisms can be used to transmit the rotary motion required to rotate the pillar into the channel. Other variations should also be understood, including the use of solenoids and the like. As an alternative to using channels, motor-driven deployments can be used to provide varying depths, which can be controlled only by controlling the motor to drive the distance of the deployment. Preferably, in this case, the described configuration of the hybrid drive mechanism uses an impact discharge mechanism (such as a spring) for initial deployment through the dermis and then uses a motor drive mechanism to advance the electrodes to their desired depth.

One or more interlocking devices may be in place, which must be deactivated before inserting the mechanism gear drive ring 478 , thereby rotating the retaining post 488 into the channel.

First, the force detection interlocking device subsystem may be in place, which requires a greater than a predetermined amount of force to apply the device to the individual before allowing the initiation of the administration procedure. This force can be measured by an appropriate mechanical or electromechanical system and the result is fed back into the controller 700 and used as an interlock to prevent the device from being activated when the provided force is insufficient. In some embodiments, the controller 700 can communicate the state of the force detection interlocking device to the user via a visual, tactile or auditory signal, so that the state of insufficient force can be corrected and the user can perform administration. In the case where the user attempts to administer under a state of insufficient force contact (for example, by pressing the trigger 407 or other activation button), the applicator 400 or the controller 700 may provide additional visual, tactile or auditory signals To inform users that the interlocking device must be disabled before proceeding.

The force applied to an individual can be detected in a variety of ways. And with reference to FIGS. 8A 4 - 8B of the particular embodiment, the alignment guide / protective cover 108 is inclined contactless reader 128 powerful equipment. You may be used or a plurality of contact springs 126 force mechanically the alignment guide / protective cover 108 is inclined toward the distal direction (toward the subject), FIG. 4 shows four contact spring force. The force contact reader 128 changes its position by means of a force applied to the alignment guide / tilt shield 108 . In doing so, it also changes the state of the circuit formed by the force contact reader 128 , a set of first pads 162 , a set of second pads 164 and the flexible circuit 160 . In detail, by testing the continuity between one or more shims 162 and one or more corresponding shims 164 , it is possible to determine the force applied to bring the alignment guide / tilt shield 108 back or near The distance that the end moves, and from this determine if there is sufficient force for proper delivery. The applicator 400 uses the sensor contact 434 to read the state of the circuit (see FIG. 16 ). If sufficient force is indicated, the force contact interlock is disabled, thereby allowing the user to operate the device.

In one embodiment, at the first point of contact, the system may not record that any particular force has been applied. At the second point of contact, the system can record that it is under partial (but insufficient) pressure. At the third point of contact, the system can record that the specified pressure level required for program administration has been reached, and the interlocking device can be deactivated. Preferably, the state of the force contact circuit is provided to the user via the applicator display 404 .

Those skilled in the art will understand the changes. For example, a force contact interlock may form an electrical lock that is deactivated within the controller 700 or within the applicator 400 itself. In another variation, the configuration of the force contact circuit provides information about the status of the device during the entire process. In detail, the system may include a feedback loop between the force contact circuit and the controller 700 , wherein a reduction in the force applied by the user causes the controller 700 and / or the applicator 400 to generate a visual, tactile or auditory signal, This can correct a state where the force is reduced. In one embodiment, there is a feedback loop between the force contact circuit and the controller 700 , and thus detecting a change in the applied force prompts the system to initially check whether the electrodes are still properly deployed in the tissue of the individual, For example via impedance or resistance check between electrodes. If the check is passed, the procedure usually continues. When the electrode position is no longer desirable, the process may be suspended and the user may be notified of the status of the device via the controller 700 and / or the applicator 400 to generate a visual, tactile or auditory signal. This feedback loop is of particular significance during drug injection. By monitoring the position of the device and the state of the electrodes, the feedback loop between the force contact circuit and the electrode resistance / impedance monitor, the system can detect whether the electrodes (and therefore the injection needle) are no longer in the individual. This allows the system to stop the operation of the injection drive mechanism 456 to stop depressing the reservoir plunger 484 and terminate the injection of the drug. Although this embodiment is relatively easy to implement using a motor-driven injection drive 456 , other changes can be implemented in the case of manual operation or a spring-driven mechanism, where a mechanical interlock device can be used to start after a fault condition is detected. Stop the actuation of the reservoir plunger. This feature is particularly useful in preventing the HBV vaccine from being unintentionally ejected into the environment when the applicator 400 is removed from the tissue of an individual before drug delivery is completed. Such a design avoids unintentional excretion / exposure for drugs or therapeutic agents that may be harmful to the user or the environment.

Because the number of doses delivered to an individual in a partial dose situation is critical to inform the clinician of a decision about further treatment, it is preferred that the injection drive mechanism 456 includes appropriate sensors and control features to The position of the injection driving plunger 484 is determined when a failure state is detected or other conditions need to be stopped to stop the injection stroke. Preferably, this is achieved by monitoring the number of revolutions of the motor used to drive the injection plunger 484 , but other methods can be used to monitor the position of the injection driven plunger 484 , including optical or inductive sensors. Based on the known dimensions, insertion depth, and reservoir 101 of the injection-driven plunger 484 , the position of the injection-driven plunger can be converted to an estimate of the volume of drug remaining in the syringe at the end of the injection stroke using appropriate logic and control circuits value. Such information may be communicated to the user via the display 712 .

In addition to providing a termination feature to terminate the injection stroke if a fault condition is detected, using the motor injection driver 456 can also provide a supplemental detector for detecting fault conditions or other operational problems. Specifically, the injection can be determined to be within a defined standard by incorporating appropriate measurement and logic circuits to monitor the circuits caused by the motor during the injection stroke. Expected range may be determined at various stages of the injection stroke caused by the motor current, comprising an injection plunger drive the plunger 484 contacts the stop member 159 prior to the initial high-speed operation, the injection drive the plunger 484 and the plunger 159 stop member The initial interface between them, the actuation of the plunger stop 159 in the barrel of the reservoir 101 , and the injection stroke ends when the plunger stop 159 contacts the end of the barrel of the reservoir 101 . By correlating the position of the injection-driven plunger 484 and the measured value of the motor-induced current during each stage of the medicament injection with the expected value, the system is able to identify possible fault conditions and communicate them to the user. For example, if the user inadvertently inserts the reservoir 101 of the plunger stop 159 partially (and therefore cannot contain the full predetermined dose of medication) into the cartridge 100 , the system can detect that the injection drive When the plunger 484 reaches an external tolerance regarding plunger positioning, the expected increase in current caused by the motor injection driver 456 does not occur. In this case, the system can terminate the dosing process and notify the user that a failure has been detected. Additional fault conditions may be detected by this method, including (but not limited to) faulty components in applicator 400 , rupture of reservoir 101 , and / or defective cartridge 100 .

The alignment guide / tilt shield 108 shown in FIG. 9C provides another "chaining device" that facilitates proper execution of the administration procedure. In this figure, the alignment guide / tilt shield 108 is shown as having a tilt feature 168 for slidably moving the puncture shield 134 and a hole 170 defined therein. The puncture protection cover 134 is shown in FIG. 9B , and the electrode hole 167 is also shown. For percutaneous insertion of electrodes and injection needles where relevant, having a consistent skin interface helps deploy the electrodes into the target tissue while maintaining the required spatial relationship between the multiple components. In particular, the most consistent and appropriate deployment is achieved when the skin is positioned perpendicular to the direction of deployment. In addition, when the skin is under tension in an orientation perpendicular to the deployment direction, the misalignment of the electrodes and the injection needle is reduced. It can be seen that the tilt shield 108 includes mechanical features, including ribs and edges, to engage the skin and place it in a tension state perpendicular to the deployment direction. Combined with the force contact circuit reading system described above to ensure a uniform force applied to the skin, the alignment guide / tilt shield 108 ensures that the skin orientation and tension direction are perpendicular to the electrode deployment direction. Although this embodiment uses the mechanical rib feature to engage the skin at the device interface, a variety of other designs and features can also be used, including the use of alternative materials with high coefficients of friction when placed in contact with the skin (such as rubber inserts, adhesive patches) Tablets and the like) or alternative mechanical features, including molded structures, cut pieces, and jagged features that can keep the skin in tension.

It can be seen that the alignment guide / tilt shield 108 has a better orientation limitation. As described above, the spatial and temporal co-localization of the drug distribution and the application of the electric field is desirable. It is worth noting that the inherent structural characteristics of skeletal muscles cause a characteristic oval distribution pattern of intramuscular injection, in which the major axis of the oval is aligned with the stripes of the muscle fibers. Therefore, for applications involving intramuscular injection, it is advantageous to arrange the electrodes to produce an elliptical electric field pattern. In order to ensure that the electrode array and the resulting electric field pattern are properly oriented relative to the stripes of the target skeletal muscle, the use of alignment guidance features has a special effect for intramuscular administration. The purpose of the alignment guide feature is to assist the placement of the device so that the orientation of the electrode array relative to the muscle stripes and the resulting post-injection drug distribution are most favorable. For an arm injection, the alignment guide / tilt shield 108 will ideally surround the arm horizontally like an armband. The legs will need to be similarly oriented and the sloping shield covers the legs horizontally. It has been found that having an alignment guide / tilt shield 108 configured in this manner allows the user to achieve> 98% accurate drug delivery even in the absence of oral instructions. The preferred orientation shown and the resulting placement of the skin are particularly suitable for intramuscular injection, as the diamond-shaped electrode array (see the array formed by the end of the electrode 137 in FIG. 9A roughly matches the puncture shield 134 and its associated hole 167 Shape) is then appropriately oriented to deliver the drug and cause electroporation of the drug along the preferred direction of muscle streaks. The main feature is that, when properly aligned, the skin should be positioned flush with the orifice, such as the skin interface where needle ejection occurs. In this way, an interface consistent with the skin is obtained.

In the case of improper alignment relative to the target muscle, an arc appears between the alignment guide / tilt shield 108 and the skin, generally causing a noticeable gap, such as 2-5 mm. The clearly visible void can be used as a reminder for the user to reorient the applicator 400 . The applicator 400 may be less stable against an individual's skin. The alignment guide feature is better for facilitating proper placement, where the distance between the edges of the "wings" of the feature is at least 1.3 times the vertical height of the feature. In addition, in some cases, the feature is designed so that the tissue interface can be placed flush with the skin in the desired orientation, while exhibiting an air gap of at least 2 mm when placed at a 90-degree angle with respect to the desired orientation.

Those skilled in the art should understand the changes in the design of the alignment guide. For example, a "V" shaped alignment guide / tilt shield 108 may be used instead of the arc. Other variations should also be understood, the key feature of which is that the device can be placed directly against the skin in the desired orientation, and when the device is misaligned with the stripes of the target muscle, there is 2 mm between the tissue interface and the recipient's skin Or larger clearly visible voids.

As noted above, in some embodiments, the orientation of the alignment guide / tilt shield 108 is related to the shape of the electrode array because the diamond symmetry of the electrode array has some distribution related to it, and this distribution should be related to the The shape of the quasi-guide / tilt shield 108 has some predetermined relationship. Referring to Figures 9A - 9B , electrodes are shown, and more specifically the distal portion 137 , the number of which is four. The electrodes are positioned in a diamond array, which means that they have a secondary symmetry, that is, double symmetry, because they can rotate and look the same at two different positions. The electrodes extend from the holes 167 defined in the puncture shield 134 in this array. The use of a diamond array is particularly useful because this array is generally required for intramuscular injection as discussed above. Again, this secondary symmetrical array causes a secondary symmetrically applied electric field. If the alignment guide / tilt shield 108 is properly oriented, this type of applied electric field may have a better direction along the muscle stripes. Therefore, the alignment guide / tilt shield 108 and the secondary electric field orientation work together to achieve drug propagation along the muscle streak direction.

Broadly speaking, in a rhombus shape or other secondary symmetrical shapes, there are generally a major (long) axis and a minor (short) axis. The axes may have a predetermined relationship with a preferred axis of the alignment guide / tilt shield 108 . For example, if the alignment guide / tilt shield 108 is considered to have wings and the arcuate shape of the wings surrounds the arm, the line segment connecting the centers of the wings may be perpendicular to the The main axis.

Those skilled in the art will understand the changes. For example, one possible shape of a diamond-shaped electrode array, and another exemplary shape is rectangular, which is also second-order symmetrical. The electrodes can be placed in place, but can be moved within a predetermined or required tolerance, such as within 5%, 10%, or the like. The electrode can take various other shapes as long as it generates a second-order rotationally symmetric electric field.

In another variation, for non-intramuscular injection, other levels of symmetry can be used. For example, for the skin, there is generally no better direction of drug transmission, and therefore, a circular electrode array can even be used for intradermal injection.

Other considerations regarding electrode sequences are as follows. The simplest array configuration consists of two electrodes connected to opposite poles of an electrical energy source. As disclosed in U.S. Patent Nos. 5,873,549 and 6,041,252, which are incorporated herein by reference in their entirety, three or more electrodes that function simultaneously in a multi-element array can be used to increase the target volume of the tissue and improve the target tissue. Uniformity of electric field propagation. For the application of electric fields in tissues, a variety of geometric electrode arrangements and activation modes have been developed. These include electrodes arranged in a linear, rectangular, circular, or triangular configuration capable of propagating an electric field in a large mass of tissue that is generally elliptical, rectangular, cylindrical, or spheroidal. Most commonly, the electrodes are arranged parallel to each other and their configuration is inserted through the skin in an orientation substantially perpendicular to the surface of the skin. In order to ensure that the target volume and shape of the tissue are affected by the application of the electric field, it is necessary to achieve a predetermined spatial relationship between each electrode in the array after percutaneous insertion. In particular, unexpected changes in electrode-to-electrode spacing should be avoided, as these changes can cause changes in the magnitude of the electric field propagating within the target tissue, which can lead to negative consequences for the safety and / or efficacy of the procedure.

Another interlocking device involves the use of an external canister cover 110 . In detail, the outer cover 110 is used to cover the alignment guide / tilt shield 108 during storage. The outer canister cover 110 is configured not only to protect the distal end of the device generally before use, but also to be a feature of the interlocking device itself. Although the distal end of the protective barrel assembly 100 is for the purpose of protecting the needle and the electrode from environmental influences, a problem often encountered is that the user often forgets to remove such a cover. One solution is to make the cover a bright color different from the other components in the cartridge 100 in order to inform the user of its presence and therefore remind the user to remove it. Another part of this solution includes an extension or reminder tab 190 as shown by an arcuate section of an adjacent rectangular section, which can be seen even when the tube assembly 100 is placed on an individual. Even if the rest of the outer canister 110 is not seen, when the reminder tab 190 is seen, it can still be reminded to remove the outer canister 110 .

Referring in more detail to FIGS. 10A - 10B, the outer cover 110 comprises a cylindrical distal end and a facing surface facing the outside of the individual alignment guide 108 of the inner surface of the member / inclined shields. A plurality of hooks 176 are provided on the inner surface, which engage respective sidewalls 180 defined on the alignment guide / tilt shield 108 . The hooks hold the outer canister cover 110 in place.

However, if the external cylinder cover 110 inadvertently left in place, and the pressing force applied to the insertion portion 400, the alignment guide member / protective cover 108 is applied obliquely to the cylindrical outer cover 110 tends to force the alignment guide by The cover / tilt shield 108 pushes the outer canister cover 110 away from the hooked position and causes it to "explode". However, a chamfered surface 178 is provided on the hook 176. When pressure is applied, these chamfered surfaces often act on the puncture protection cover 134 , tilting the hook 176 outwards, increasing its abutment against the alignment guide / tilt The retaining force of the protective cover 108 prevents the outer cylinder cover 110 from bursting open. In addition, the chamfered surface further acts on the puncture shield, preventing the alignment guide / tilt shield 108 from moving relative to the puncture shield 134 . If the alignment guide / tilt shield 108 cannot move relative to the puncture shield 134 , the force detection interlock cannot be disabled because the force contact reader 128 cannot move to the point where the full pressure is detected as discussed above. Third force contact point (or even for a second force contact point). When this happens to the controller, the controller can provide a report. For example, a user interface indicator such as "The outer cylinder cover needs to be removed." May be displayed, which also causes the same display when a pull trigger is detected in the absence of an appropriate force.

Although certain interlocking devices have been described above, more or fewer interlocking devices may be provided in any given embodiment. For example, other methods can be used for force detection and as a trigger signal for interlocking devices. Other types of interlocking devices can also be used, including trigger locks, safety switches, and the like. Based on this teaching, those skilled in the art will understand yet other types.

After the user positively selects the insertion depth, and the outer cylinder cover 110 is removed, and the force interlocking device detects an appropriate force, the activation of the trigger causes the cylinder lock ring 114 to rotate clockwise or counterclockwise, in which direction of rotation Depends on the insertion depth selected by the user. The rotation moves the holding post 488 into the channel of the selected depth, which in turn causes the needle and electrode to be deployed. In detail, the inner cylinder 103 , the tail end 112 , the reservoir 101 , the inner cylinder cover 104 , the needle interface 152 , the needle 105 , the electrode 122, and related components move forward under the influence of the electrode / needle insertion spring 472 . In this embodiment, the elements are rigidly connected to each other and therefore move forward as a whole.

To further minimize the risk of sharp objects damaging the user, preferably, the cartridge assembly also incorporates a mechanism for sheathing the electrodes and needles after the electrodes and any injection needles have been removed from the recipient's tissue . This can be achieved by incorporating the stab wound shield feature, which is housed before the electrode and injection needle (if present) are used and then extended after the electrode and injection needle (if present) are deployed and removed from the individual's tissue Cross the electrodes and injection needle (if present). Typically, the tissue contact interface includes a distal surface that is a feature of the shield. Although the use of a manually operated shield feature may be considered, preferably, the device incorporates a mechanism that once the electrode is removed from the recipient's tissue, the mechanism automatically extends the shield feature across the electrode and further engages the locking feature to maintain Features of the shield in the extended state. Examples include slidably engaging the outer casing of the cartridge and operatively connected to a stored energy source, such as a shield feature of a spring, which causes the shield to slide forward as the electrode is withdrawn from the recipient's tissue. Alternatively, the structure may be contained within the applicator and its configuration reverses the electrode deployment step, thereby retracting the electrode and any associated injection needle after the tissue contacts the interface. This can be achieved using simple electromechanical mechanisms such as motors or solenoids that move linearly.

In a specific embodiment, and referring back to Figure 4 and Figures 11A and 11C , the configuration of the puncture shield 134 is held in place when drug delivery occurs, but the forward direction of the inner barrel 103 and other related components during drug delivery Move the compression stab shield spring 138 . This compressed stab wound shield spring 138 then relaxes when the applicator is removed from the subject, as the stab wound guard 134 is caused to move forward when the applicator 400 is removed from the subject, covering the needle and electrodes, and protecting the subject and others Protected from uncovered sharps. It is worth noting that this feature also prevents sharp object features from being observed during extraction, and thus may assist individuals who experience needle-related anxiety.

Although the slack spring will be able to be recompressed additionally, and in particular if the user moves the puncture shield 134 proximally, the spring is prevented from doing so by the puncture shield support arm 132 , which punctures the shield support arm It is installed inside the outer cylinder 102 and acts as a ratchet when the puncture protection cover 134 moves forward. In detail, and referring to FIGS. 11A and 11C , the stab protection cover support arm 132 moves across each continuous holding wall, and this operation can be easily performed when the stab protection cover 134 is moved distally. However, in the proximal direction, since the puncture shield support arm 132 is ratcheted against the holding wall in this direction and is not allowed to pass, the puncture shield 134 is prevented from moving.

An initial set of retaining walls 188 prevents the puncture protection shield 134 from moving proximally before the cartridge assembly 100 is used. A set of first depth retaining walls 184 prevents the proximal end of the puncture shield 134 from moving after being expelled at the first selected depth. A set of second depth retaining walls 186 prevents the proximal end of the puncture shield 134 from moving after being expelled at the second selected depth. Under the action of the puncture protection cover retaining hook 182 acting on the inner cylinder 103 , the puncture protection cover 134 is completely removed from the cylinder assembly 100 .

Those skilled in the art will understand the changes. For example, in one embodiment, the puncture shield support arm 132 may be provided by a stamped metal support arm that interfaces with the outer cylinder 102 . In some embodiments, the support arm feature is directly integrated with the injection molded plastic feature in the outer cylinder cover 106 and / or the alignment guide / tilt shield 108 . The specific material selection should achieve sufficient rigidity to provide support for the elongate electrode while maintaining sufficient elasticity to prevent the component from breaking under load. Materials should also take into account predetermined sterilization methods for electrode arrays (such as gamma radiation, steam sterilization, ethylene oxide, or electron beam) to ensure that these characteristics maintain proper material properties after sterilization. In an exemplary embodiment, the features implemented should be able to prevent the proximal movement of the puncture shield 134 when a force of at least 5 N, but more preferably at least 15 N is applied. Other means of providing co-operation to prevent movement of the proximal end of the puncture shield can also be employed, including a rack and pinion implemented on the puncture shield 134 and a corresponding ratchet feature implemented in the outer cylinder cover 106 .

Referring to FIG. 4 , each of the elongated electrodes 122 in the array includes a distal end portion 137 and a proximal end portion 135 that are in conductive communication. The configuration of the distal portion is used for tissue penetration and electric field propagation in the tissue, and the configuration of the proximal portion is capable of reliably achieving conductive communication with the electrical connection contained in the applicator having a suitable electrical energy source. Conductive contact area. The electric energy source can cause an electric field in space and time in the individual's body by the time change of the power applied to individual electrodes. The electric field is generally limited to the volume near the drug distribution area, and it can be used to facilitate electricity Perforation therapy.

Relevant characteristics of the electrode include shape, diameter, tip shape, length, material composition, and electrical conductivity, the details of which are selected based on the intended application. Most commonly, the electrode contains a conductive elongated rod having a curved cross section with a diameter of 0.1-1.5 mm. The electrodes can be solid core or hollow. The most commonly used hollow electrode incorporates one or more orifices and an operative connection to a fluid reservoir, thereby allowing administration of a medicament of interest or other related medicament, including anesthetics, surfactants, proteins, adjuvants or Enhancer from the electrode itself. Depending on the application, suitable metal electrode materials include, but are not limited to, titanium, gold, silver, aluminum, copper, tantalum, tungsten, molybdenum, tungsten, stainless steel, MP35N, and alloys thereof. The electrodes can also be made of conductive ceramic or plastic. To minimize unnecessary electrochemical reactions at the tissue electrode interface, it is often necessary to coat the one or more electrodes with a conductive material, thereby providing improved electrochemical stability than the electrode material itself. Such coating materials include, but are not limited to, platinum, iridium, palladium, osmium, gold, silver, titanium, and alloys thereof.

As described above, the tip of the distal portion is configured to penetrate tissue. Common tip types include, but are not limited to, trocars, bevels, cones, blades, spears, and wedges. For deployment of percutaneous electrodes, a tip with one or more blades is preferred. For some applications that are not suitable for generating the required electric field in the form of a penetrating tip, the tip may be composed of a non-conductive material fixed to the distal tip of the electrode, or the penetrating tip may use a biocompatible electrically insulating material (Such as poly (p-xylene) polymer, polyolefin, polyvinyl chloride, polyurethane, polyester, polyimide, polysiloxane, thermoplastic elastomer / rubber, ethylene tetrafluoroethylene, fluorine Adhesive coating or tube covering consisting of ethylene propylene and / or perfluoroalkoxy plastic).

The proximal end of the penetrating tip is one or more conductive regions that are configured to propagate an electric field in the tissue. In order to limit the propagation of the electric field to the target area of the tissue, usually, especially for subcutaneous and intramuscular administration, the configuration is to penetrate at least a portion of the electrode length into the tissue using a biocompatible electrically insulating material such as Poly (p-xylene) polymer, polyolefin, polyvinyl chloride, polyurethane, polyester, polyimide, silicone rubber, thermoplastic elastomer / rubber, ethylene tetrafluoroethylene, fluorinated ethylene Adhesive coating or tube covering made of acrylic and / or perfluoroalkoxy plastic. In this way, the configuration of the electrodes can be initiated only at specific depths within the tissue.

In summary, the number of electrodes and the distance between the electrodes, the penetration depth of the electrodes, and the length of the conductive area determine the volume of the tissue of the applied electric field. These parameters are selected based on the specific purpose of the dosing procedure, including the volume, dose, and viscosity of the target tissue site and the agent to be delivered, as well as changes in skin and subcutaneous tissue thickness in the intended recipient population. In general, when intradermal administration is performed on human recipients, the array contains 2-16 electrodes with a diameter of 0.2-0.7 mm, the distance between the electrodes is 2-8 mm, the electrode penetration depth is 0.5-4 mm, and the conductive length It is 0.5-4 mm. In general, when subcutaneous administration is performed on human recipients, the array contains 2-8 electrodes with a diameter of 0.3-0.8 mm, the distance between the electrodes is 4-10 mm, the electrode penetration depth is 5-15 mm, and the conductive length is 2-8 mm. In general, when intramuscular administration is performed on human recipients, the array contains 2-8 electrodes with a diameter of 0.3-1.2 mm, among which the distance between the electrodes is 4-12 mm, the electrode penetration depth is 10-60 mm, and it is conductive The length is 2-20 mm.

To avoid undesired observation and exposure to the electrodes during use of the device, the preferred configuration of the cartridge assembly 100 is such that the electrode 122 is recessed into the device before being inserted into the recipient's tissue. Most commonly, this is accomplished by providing an outer housing structure 102 that slidably engage an electrode mounting support. In one embodiment, the electrode mounting support includes an inner barrel 103 that appears to be formed therein, and the electrode system is positioned In the inner tube. Prior to use, the electrodes are recessed into the outer housing 102 . During use, the electrode mounting structure, for example, the inner tube 103 slidably engages the outside of the housing, allowing the electrode 122 with respect to the forward sliding of the distal tip of the outer housing 102, whereby the outer housing 102 from electrode 122 to deploy. The length of the sliding joint between the electrode mounting structure and the outer housing corresponds to the maximum electrode penetration depth required in a given application.

As can be seen in Figure 4 , the electrode 122 has a proximal portion 135 and a distal portion 137 . The proximal portion and the distal portion are separated by a shoulder or bend 139 . The shoulder portion or the bent portion 139 is fastened by the inner cylinder cover 104 and the inner cylinder 103 and is between the inner cylinder cover and the inner cylinder. This shoulder or bend 139 provides multiple functions. First, the electrode at the distal end of the cartridge assembly 100 needs to form an array having a certain size and shape as described above, and the electrode 122 is placed at the desired array size and shape at the proximal end of the cartridge assembly 100 due to the reservoir. The existence of 101 is impractical. In other words, the electrode 122 must be bent "out-of-the-way" to make room for the reservoir 101 . In addition, the curved portion provides a surface for the electrode 122 to abut when the force of needle and electrode insertion recoils in the distal direction due to the electrode 122 interacting with the tissue during its deployment, especially when the curved portion rubs against When the mode is locked in place between the inner cylinder 103 and the inner cylinder cover 104 . The bend can resist axial rotation, which is beneficial so that the electrode does not exhibit significant torsional movement and rotates away from its electrical contact pad 130 (described below). In addition, compared with the prior art in which multiple electrodes are separated from the coaxial ring axially, which requires different configurations of the electrodes, the system of the present invention allows all four electrodes to use a common electrode type.

Systems and methods according to the principles of the present invention provide for the transmission of an electric field in the recipient's skin, subcutaneous tissue, and / or skeletal muscle, thereby facilitating intracellular delivery of the HBV vaccine within the recipient's tissue. In this aspect, the device includes two or more elongated electrodes 122 arranged in a predetermined spatial relationship, the configuration of which is intended for deployment to a target tissue that has or will distribute the agent of interest. The required enhancement of the pharmacological activity depends on the co-localization of the distribution site of the medicament and electric field propagation with a sufficient amplitude to induce the desired physiological effect. Therefore, the device is required to achieve an electrode deployment at a target depth with a specified inter-electrode pitch. In particular, this ensures that the electric field propagates in the appropriate tissue site, and since the amplitude of the electric field propagating in the tissue varies with the inter-electrode spacing, this ensures a safe and effective electric field strength. However, changes in the thickness, density, and composition of skin and subcutaneous tissue between various sites in a given recipient and among recipient groups may result in significant changes in electrode deployment characteristics. For example, recipients with increased skin thickness have an increased chance of electrode deformation and / or insufficient penetration depth, which may affect co-localization with drug distribution sites.

The system and method according to the principles of the present invention also facilitate consistent percutaneous insertion of electrode arrays containing 2-16 electrodes with a diameter of 0.2-1.3 mm in recipients with uneven skin thickness, composition, and state. These electrode arrays The distance between the electrodes is 2-12 mm and the electrode penetration depth is 0.5-60 mm. To allow percutaneous insertion, the tissue penetrating electrodes are most often elongated and configured to have a tissue penetrating tip and tissue contact area in the distal portion and an electrical contact area in the proximal portion. Due to its elongated structure, it is prone to tortuosity, bending, and / or buckling (collectively referred to as deformation) when subjected to the compressive forces generated during percutaneous insertion into a recipient. Such deformations during electrode insertion are undesirable because such deformations can lead to improper electrode insertions characterized by insufficient penetration depth and / or excessive changes in the inter-electrode spacing. It is worth noting that humans and animals exhibit significant intra- and inter-species differences in skin thickness, composition, and state, and any of them can have a significant effect on the force experienced by the electrode during percutaneous insertion. In some embodiments, the device for percutaneous insertion of an elongated electrode is designed to withstand the forces that exist under the most stringent conditions of electrode insertion to be encountered in a target recipient group. The occurrence of electrode deformation can be partially alleviated by selecting electrode materials, reducing the length of the electrodes in the array, and increasing their diameter. However, material selection may be constrained by issues of efficiency, cost, manufacturability and biocompatibility. In addition, reducing electrode length may prevent adequate coverage of target groups with uneven tissue thickness, while larger diameter electrodes increase discomfort and tissue trauma. Therefore, the present invention is designed to provide methods and devices that facilitate percutaneous insertion of electrodes without these variables.

In a first embodiment of the present invention, the secondary assembly barrel that houses the electrode array incorporates one or more dynamic support members that are in physical contact with one of the electrodes during tissue insertion, and its configuration is constrained perpendicular to The electrodes in the insertion direction move and maintain the required spatial relationship between the electrodes. For example, and referring to FIG. 12 , an electrode support feature 124 is shown, in which a hole 192 is provided for an electrode and a hole 194 is provided for a needle to pass through. The tendency of the elongated electrode to deform or buckle under load can be exacerbated by any bending, eccentricity, or twisting of the electrode that may have been introduced during manufacturing or assembly. Therefore, a device such as electrode support feature 124 is advantageously employed to constrain the electrode to prevent, or correct, any bending, eccentricity, or tortuosity that it may exhibit when stationary, and to prevent electrode instability caused by loading that occurs during deployment into tissue Necessary vertical movement.

In the context of the present invention, a dynamic support component is defined as a structural element that slides into engagement with an elongated electrode during insertion into a recipient's tissue, and is configured to undergo changes in position, size, and / or configuration during electrode insertion in order to Maintain the required spatial relationship of the electrode as it is deployed to the target tissue depth. Since the electrodes experience the maximum loading force during initial contact with the skin, the engagement of the dynamic support members with the electrodes constituting the array is preferably achieved before the initial contact with the recipient's tissue and continues to provide support when the electrodes are deployed to a full penetration depth. It is also advantageous that the dynamic support member is designed to provide support for the electrode and the injection needle while minimizing the loss of electrode penetration due to friction.

Although the main function of the dynamic support component is to maintain the required spatial relationship of the plurality of electrodes in the array relative to each other and to the injection needle, the design of the dynamic support component also needs to include features that can stabilize the entire array. This is preferably achieved by providing additional structural support features that restrict the lateral movement of the support member. In one embodiment, the support features are integrated into a sharp object shield, which is also used to prevent accidental exposure of the user to electrodes and / or injection needles. Various embodiments of the dynamic support member according to the present invention are disclosed below.

One embodiment of the invention described above as the electrode support feature 124 relates to the use of a planar support structure 124 positioned vertically with respect to the elongated orientation of the electrode. The planar support structure is configured with one or more apertures 192 , which correspond to the positions of the electrodes in a specified spatial relationship. The configuration of the size, shape and location of the pores allows the support structure to slide smoothly along the length of the electrode elongation within the array, despite restraining unnecessary movement perpendicular to the electrode deployment direction. Generally, the pores are configured as holes or grooves 192 in the planar structure having gaps suitable for the electrodes (at least 10% larger than the maximum cross section of the electrode including any coating or other adhesive material). However, if a particular electrode requires greater support, one or more of these apertures may constitute a tubular structure 196 arranged perpendicular to the planar support structure. The pores constituting such a tubular structure increase the surface area of the contact electrode and thus the support provided to the elongated electrode. The planar support structure may be made of any material including metal, polymer, ceramic, or composite materials with appropriate structural characteristics, and may be formed, machined, molded, or made by other methods. To avoid unnecessary electrical interaction with the electrodes, preferably, the interface between the electrodes and the planar support structure is non-conductive. The choice of materials and manufacturing methods should also minimize the amount of friction at the interface between the electrode and the dynamic support member. Due to a number of factors including low cost, ease of manufacture, and favorable electrical characteristics, electrode support structures are typically made of thermoplastics such as polycarbonate, polystyrene, polypropylene, acrylic, or polyethylene. The specific material selection should achieve sufficient rigidity to provide support for the elongate electrode while maintaining sufficient elasticity to prevent the component from breaking under load. Material selection should also consider the intended sterilization method of the electrode array (such as gamma radiation, steam sterilization, ethylene oxide, or electron beam) to ensure that the dynamic support components are compatible. The specific size and design of the support structure depends on the characteristics of the selected material. However, it is desirable to minimize the size of the support structure so that it does not unduly limit the distance at which electrodes can be deployed or interfere with other functional characteristics of the device. A rigid planar structure with a thickness of 0.5 mm-2 mm is usually sufficient.

Another embodiment of a dynamic support member uses a compression spring whose aperture accommodates the electrode and restricts its lateral movement. Compression springs can be made of metal, polymer, or elastic materials and can be formed, machined, molded, or made by other methods. When stationary, the spring is uncompressed or minimally compressed, and the electrode is inserted into the aperture along the length of the spring. When the electrode is deployed, the spring is compressed in the deployment direction, and the aperture accommodates the sliding movement of the electrode perpendicular to the spring coil. This embodiment has a special effect when combined with a protective cover or sheath for accommodating a penetrating electrode / needle after removal from a tissue site. In such embodiments, the force applied to the spring by the forward deployment of the electrodes can be used to deploy a sheath or shield over the electrodes when the device is removed from the tissue.

In the embodiment of FIG. 4 , the puncture protection cover spring 138 may be used to partially support the electrode support 124 , and in particular, the configuration of the spring radius matches (or is only slightly larger than) the side wall 198 of the electrode support 124 . Radius . In this manner, the electrode support 124 can be inserted in the middle of the puncture shield spring 138 during use. The electrode support 124 may then slide inside the puncture shield 134 . Because it is placed in the center of the spring, the electrode support 124 naturally maintains the position in the center of the spring, thereby providing a "halfway point" required to support the electrode, that is, approximately at the inner cylinder cover 104 Between its support point and its penetration point at the tissue interface.

Covers other spring-based electrode supports. For example, the compression spring formed is positioned in the region of the electrode to provide adaptive electrode support. The resulting compression spring can be shaped and scaled to closely follow the relative position of the electrodes in order to limit its lateral movement. An optional biasing element can be positioned in conjunction with the compression spring formed, and can be used to bias the electrode outward against the compression spring formed. The resulting compression spring may be made of metal, polymer, or elastic material, and may be formed, machined, molded, or made by other methods. The biasing element may be made of metal, polymer, or elastic material and may be formed, machined, molded, or made by other methods.

It also covers electrode supports based on other mechanisms, such as telescoping sleeves. A telescoping sleeve is used to support the electrode during its insertion into the individual. The telescoping sleeves can be sized to move freely relative to each other, or they can be sized to move only when an axial force is applied to them.

Other support structures are covered, such as those based on movable, flexible or pivoting support components. The lateral support members can be attached to the electrodes at optional hinge features. The lateral support member may be integrally formed with the structure for accommodating the electrodes, or may be a separate component attached by a conventional method (lock, welding, adhesive, fixing member, etc.).

Another embodiment uses a compressible matrix material into which the electrodes are embedded. When the electrodes are deployed, the material is compressed in the deployment direction, thereby providing lateral support in the direction of travel. Examples of compressible matrix materials include cellulose, foamed plastic or rubber polymers such as microcellular plastics, foamed silicone or foamed polychloroprene, or carbon foamed matrix. Since these materials are designed to contact the electrodes and / or injection needles (if applicable), the choice of these materials should be compatible with indirect tissue contact.

The above structure thus supports the percutaneous deployment of a plurality of elongated electrodes at a tissue depth of up to 60 mm, while maintaining the spatial relationship required between the plurality of electrodes and, in specific embodiments, the orifices of the hypodermic injection needle. Such support members engage the plurality of elongated electrodes during percutaneous insertion and constrain the deflection of the electrodes in one or more directions perpendicular to their elongated orientation. Another aspect of the present invention provides the use of the force required to apply a biocompatible lubricating compound to the surfaces of the plurality of electrodes in order to reduce the need to achieve consistent percutaneous deployment of the plurality of elongated electrodes at a depth of up to 60 mm. Method and device. In an exemplary embodiment, a biocompatible polysiloxane such as Dow Corning 360 Medical Fluid or Dow Corning MDX4-4159 can be applied to a plurality of electrodes constituting the array by a conventional spray or dip coating method to improve Electrode insertion features. The specific choice of coatings and application conditions, such as coating method and thickness, depends on the number, size, composition, and tip configuration of the electrodes and the target tissue where the electrodes are deployed.

Referring to FIG. 4, and 7A - 7B, the proximal portion 122 of electrode 135 may be positioned on the exterior of the inner tube 100 of cartridge assembly 103 (FIG. 7B). In this way, when the inner cylinder 103 is slidably disposed inside the outer cylinder 102 ( FIG. 7A ), the proximal portion 135 contacts the corresponding electrode contact 130 , and specifically the inner portion 133 of the outer cylinder of the electrode contact 130 , the The internal portion 133 is configured for electrical communication with the proximal portion 135 . Due to the length of the proximal portion 135 and the length of the inner portion 133 of the outer tube, electrical communication can be made at multiple locations along its continuous interface, regardless of the longitudinal position of the inner tube with respect to the outer tube. The electrode contact 130 further includes an outer cylinder external contact 131 configured to be in electrical communication with a corresponding connector 496 on the applicator 400 (see FIG. 18C ). The electrode contacts 130 are made of a conductive material sufficient to transmit electrical signals. In certain embodiments, the design and material selection ensure that the contact provides proper engagement to ensure a corresponding conductive interface within the range of manufacturing variations of the intended electrode and contact location, while not inhibiting or interfering with mounting on the inner barrel 103 The electrode 122 travels forward. The design must also allow the joint to retain the shelf life of the product label when exposed to expected storage conditions. In one embodiment, the electrical contacts 130 are made of stamped or formed metal with appropriate hardness to facilitate bonding with the electrodes and may include a coating such as gold or copper to ensure the integrity of the electrical contacts and avoid corrosion . In addition to ensuring that electrical contact with the electrode can be maintained at multiple locations along its continuous interface, incorporating the electrode contact into the outer cylinder 102 in this configuration also ensures that the electrode 122 and the electrode contact 130 are caused by a sliding interaction Any wear occurs within the cartridge 100 designed for single use, thereby allowing a static interface to be formed between the outer barrel contact 131 and the applicator electrical contact 496 , which allows the use of an applicator designed for multiple uses. Possible mechanical wear on the electrode connection 496 is minimized. This configuration is beneficial to extend the functional life of the multi-use applicator.

The rest of the applicator 400 will now be described. These sections are generally independent of the operation of the cartridge assembly 100 . Referring first to FIG. 13A , the applicator 400 includes a handle 402 and a multi-wire cable 406 designed to carry power and control signals and electrical signals to be applied to the tissue. The cable 406 is typically terminated with a connector and the corresponding connector interface is in the controller 700 . The applicator 400 also includes a user interface 404 , in which the user can view the appearance of the program, and in which the user can instruct the applicator to perform various functions, in particular, depth selection. The applicator 400 also includes a program start trigger 407 that the individual uses to initiate the program.

Referring again to FIG. 13B , the applicator 400 includes a program countdown timer 410 , which informs the user of the remaining duration of the program, and also implicitly instructs the user that the countdown trigger should not reset itself before the countdown trigger count reaches zero time. Remove applicator 400 . Power indicator 418 is arranged for the supply and satisfactory connection instruction to the controller 700. The program fault indicator 414 is configured to indicate whether the user has a fault, for example, one of the interlocking devices described above has not been deactivated. The application status indicator 412 is provided to inform the user if appropriate pressure has been obtained for the individual tissue, thereby allowing the procedure to begin.

FIG. 14 indicates various structural components of the applicator 400 , including a connector 426 (not to scale) for connection to the controller 700 , a top case 420 , side cases 422 and 424 , and an inner protective case 432 . A front cover 430 and an end cover 428 are provided . Various electromechanical sub-assemblies 450 are also provided, some of which have been described above.

FIG. 15 shows a more detailed view of the applicator 400 , indicating the cartridge pressure sensor contact 434 and the secondary assembly 452 corresponding to the cartridge loading, electrode insertion, and injection function and related sensors.

FIG. 16 shows a more detailed view of the group 452 of sub-assemblies, including a load-driven sub-assembly 454 and a cartridge-loaded sub-assembly 456 . The operations of these sub-assemblies have been described above.

FIG. 17A shows a more detailed view of the load-driven subassembly 454 , an overview of which has been described above. It should be noted here that the loading is triggered by a mark on the cartridge assembly 100 , and the detection of the mark causes the system to be triggered at the switch 464 . The load drive sub-assembly is mounted to the applicator housing using brackets 466 and 468 . The motion of the motor 444 is transmitted to the pinion assembly 448 through the motor transmission shaft 462 .

FIG. 17B shows a more detailed view of the insertion / injection drive assembly 456. FIG. Many of these components have been described above. It should be noted here that the operational assembly is mounted to the applicator housing by a mounting bracket 476 . After inserting the electrode 122 and the needle 105 , the injection driving plunger 484 depresses the plunger of the needle 105 , and its action pushes the medicine out of the orifice of the needle. The injection driving plunger is driven by an injection driving motor and a gear assembly 486 .

Generally speaking, the injection-driven plunger driven by the injection-driven assembly 486 moves forward until it can no longer move forward, that is, the position moved distally, indicating that it has reached the end of each stroke. This indication is generally provided by the generally increased current used in the injection drive motor 486 , indicating that the reservoir plunger has reached the end of the reservoir and the injection has been completed. However, in some cases, the number of motor rotations can be used to determine the amount of drug delivered. Such features can be used to support a metered dose of a drug or to deliver a volume that is not within the expected total volume, such as the entire volume of the reservoir 101 intended to be injected, but based on the plunger rod, which cannot be emptied In case, inform the user. Such situations may occur before the procedure is completed, such as when the user fails to hold the applicator 400 against the individual's body before the countdown timer counts down to zero. As noted above, within the intermediate program time range, when the force to the individual is no longer detected, an impedance check can be performed to determine if the electrode is still within the individual. If it is not present, it can be speculated that the applicator is removed prematurely, and a signal can be sent to the injection drive motor 486 to stop the injection, thereby limiting the amount of drug that is pushed out of the individual.

Referring to FIG. 19 , it can be seen that the controller system / combination 700 includes an electric field controller / generator 750 and a handle 702 and an applicator bracket 706 . The controller is configured for use on a table or on a cart. In a cart-mounted configuration, it is advantageous to include a storage bin 704 . Details of the controller in a cart-mounted configuration are shown in Figures 20A - 20D , including a wheel lock for fastening the controller assembly to prevent movement in the operating environment, for placement including personal preparation , Tray / 710 for reservoirs / vials / syringes, and the like. The applicator connection port 708 is shown for connecting the applicator 400 to the controller assembly 700 . The display 712 displays the status of the administration procedure, specifically, the status regarding the attachment of the IFU.

The cartridge eject button 714 is provided to eject the cartridge assembly 100 from the applicator 400 . The menu navigation button 716 allows navigation and manipulation of components as seen on the display 712 . The mute button 718 is provided to mute an alert or other audible indicator when necessary. The power button 722 is configured to power the unit, and it is activated when the main power switch 726 ( FIG. 20D ) is turned on.

The display also includes a battery indicator 720 , which provides an indication of battery power, and a battery backup system is provided therein. Such a battery backup system may be included in the controller / generator 750 for reconciling situations where power loss prevents the main power source from powering the unit. It can also be used as a backup in case the program starts under main power but has encountered a loss of main power. In this case, the logic and control circuits are implemented to provide a substantially seamless conversion of autonomous power to battery power, whereby the battery backup can be used to complete the process. The favorable situation is. The controller includes a battery monitoring circuit that can monitor whether the battery has enough power to complete the procedure after the main power loss. In some embodiments, the controller also includes a display for notifying the user when the current power state of the battery is insufficient to complete the procedure when the main power is lost.

Referring to FIG. 20D , a rear view of the controller 750 is shown. It can be seen that the controller includes a USB port 724 , a main power switch 726, and a main power input 728 .

Referring to FIG. 21 , in a method of use, as illustrated by flowchart 800 , the controller / generator 750 is powered and its program is automatically started (step 802). The applicator is connected (step 804), and if the applicator is not yet connected, the user may be shown an instruction or instruction on performing this action on the display screen 712 . The system may perform a self-test (step 806), which not only ensures proper operation of the controller / generator 750 , but also ensures proper connection of the applicator 400 with the controller / generator 750 .

The program can cause the display screen to provide the user with instructions on the preparation site (step 808). This step may include ensuring that the correct medication is delivered, that the medication has not expired, that contraindications have been checked, and that warnings / precautions have been followed.

The user then removes the reservoir cap and inserts the reservoir 101 into the cartridge assembly 100 (step 810). In some embodiments, the user experiences an auditory, tactile, or tactile click (step 811), indicating that the reservoir is properly placed in the cartridge assembly.

The user then inserts the cartridge assembly 100 into the applicator 400 (step 816). It then tests, for example, an error condition regarding proper reservoir placement (step 818), and if an error condition is detected, the process is stopped, an error message is displayed, and the user is instructed to take remedial action. If the error condition can be corrected, for example, the user inserts the reservoir improperly and cannot engage or close the end of the canister, the user can be instructed to remove the canister and reinsert the reservoir correctly (state 820). In some cases, the canister is automatically ejected, and in other cases, the user must press the "canister eject" button to achieve the canister ejection. In other error states, such as when the end of the cartridge is closed, the user can be instructed to use a new cartridge.

In any case, once the device is installed and has reached the "error-free" state, the user can administer medication and electroporation therapy (step 822).

The main function of the controller is to generate the electric field required to achieve the required drug delivery, control the operation of the system during installation and use, monitor the state of the system during installation and use, and monitor the system during installation and use. The status is communicated to the user. In some embodiments, the controller can provide recommendations and instructions for using the device in user training situations and during resolution of fault conditions during normal use.

The controller system and controller operation method can be fully implemented using a variety of computing devices, including a microprocessor, a microcontroller, and a programmable logic controller. Generally, the instructions are arranged on a computer-readable medium that is generally non-transitory, and such instructions are sufficient to allow a processor in a computing device to implement the methods of the present invention. The computer-readable medium may be a hard drive or solid-state memory with instructions loaded into the random access memory when running. For example, a plurality of users or any user may input an application program through a plurality of suitable computer input devices. For example, a user can use a keypad, keyboard, mouse, touch screen, joystick, track pad, other pointing device, or any other such computer input device to enter calculation-related data. Data can also be entered by means of inserted memory chips, hard drives, flash drives, flash memory, optical media, magnetic media or any other type of file storage media. The output can be delivered to the user by means of a video graphics card or integrated graphics chipset coupled to a display visible to the user. Alternatively, the system can output electronic files in one or more forms or can use a printer to print out the results. In light of this teaching, it should be understood that the present invention encompasses a variety of other tangible outputs. For example, the output can be stored on a memory chip, a hard drive, a flash drive, flash memory, optical media, magnetic media, or any other type of output. It should also be noted that the present invention can be implemented on many different types of computing devices, such as personal computers, laptop notebook computers, mini notebook computers, handheld computers, personal digital assistants, mobile phones, smart phones, tablets Computers and devices specifically designed for these purposes. In one embodiment, a user of a smart phone or wi-fi connected device uses a wireless internet connection to download a copy of the application from the server to his device. Proper verification procedures and secure transaction laws can make payments to sellers. Apps can be downloaded via a mobile connection or WiFi or other wireless network connection. The application can then be run by the user. Such networked systems can provide a suitable computing environment for implementations where multiple users provide independent input to the system and method. In the following system covering the control of the applicator, the plurality of inputs may allow a plurality of users to input related data at the same time.
Component list
Examples

The following examples of the invention further illustrate the nature of the invention. It should be understood that the following examples do not limit the present invention and the scope of the present invention is determined by the scope of the attached patent application.

Example 1 : Production of HBV core and Pol antigen sequences and plastid optimization Can be used to improve the breadth of the response. Therefore, the hepatitis B core and polymerase protein were selected as the antigens for the design of a therapeutic hepatitis B virus (HBV) vaccine.

Derivation of common sequence of HBV core and polymerase antigen <br/> Based on HBV genotypes B, C and D, a common sequence of HBV pol and core antigen was generated. Different HBV sequences were obtained from different sources and aligned for core and polymerase proteins, respectively. The original sequence alignment of all subtypes (A to H) was then restricted to HBV genotypes B, C, and D. Based on the alignment of each protein, common sequences were determined in each individual isoform and in all conjugating BCD sequences. In changing alignment positions, the most common amino acids in the common sequence are used.

Optimization of HBV core antigen
The HBV core antigen common sequence is optimized by creating deletions in native viral proteins. In detail, thirty-four amino acids required for capsidization of pregenomic RNA corresponding to a large positively charged segment at the C-terminus were deleted.

Optimization of HBV Pol Antigen <br/> Specifically, the asparagine residue (D) in the "YXDD" motif of the reverse transcriptase domain was changed to asparagine residue (N) to remove any ligand Site function, and thereby eliminate nucleotide / metal ion binding. In addition, the first aspartate residue (D) in the "DEDD" motif of the ribonuclease H domain was changed to asparagine residue (N) and the first glutamic acid residue ( E) Change to glutamate residue (A) to remove Mg2 + coordination. In addition, the HBV pol antigen sequence is codon-optimized to disrupt envelope proteins, including S proteins and the internal open reading frames of pre-S1 and pre-S2 in the form of S-proteins with N-terminal extensions. Therefore, the open reading frames of the envelope proteins (pre-S1, pre-S2, and S protein) and X protein were removed.

Optimization of HBV core and Pol antigen performance strategies AGAG fuses in-frame to produce a single core-Pol fusion protein (Figure 25A); (2) Represents the core and pol from a plastid by means of a ribosome slippage point, especially the FA2 slippage point from foot and mouth disease (FMDV) The antigen is a bicistronic expression vector (Fig. 25B) that expresses individual core and pol proteins from a single mRNA; and (3) two independent plastids encode the HBV core and pol antigen, respectively (Fig. 25C).

In vitro performance analysis <br/> The common HBV core and pol antigen coding sequences were cloned into the commercially available performance plastid pcDNA3.1 based on each of the above three performance strategies. HEK-293T cells were transfected with these vectors and protein expression was evaluated by HBV core-specific antigen by Western blotting method.

Optimization of post-transcriptional regulatory elements <br/> For improving protein performance by stabilizing major transcripts, promoting their nuclear export and / or improving transcription-translation coupling, evaluate four different post-transcriptional regulatory elements: (1) soil Woodchuck HBV post-transcriptional regulatory element (WPRE) (GenBank: J04514.1); (2) Intron / exon sequence derived from the human apolipoprotein A1 precursor (GenBank: X01038.1); (3) Non-translated R-U5 domain (GenBank: KM023768.1) of human T cell leukemia virus (HTLV-1) long terminal repeat (LTR); and (4) HTLV-1 LTR, synthetic rabbit β -A complex sequence (triple complex sequence) composed of a hemoglobin intron (GenBank: V00882.1) and a splicing enhancer. Enhancer sequences were introduced between the CMV promoter and the HBV antigen coding sequence in these plastids. No significant difference in core antigen expression was observed using Western blot methods when expressed in the presence or absence of WPRE elements (Figure 25D). However, when the core antigen performance in HEK293T cells transfected with plastids with three other post-transcriptional regulatory sequences was evaluated by Western blotting method, the triple enhancer sequence caused the strongest core antigen performance (Figure 25E).

Select signal peptides for efficient protein secretion <br/> Evaluate three different signal peptides introduced in the frame at the N-terminus of the HBV core antigen: (1) Ig heavy chain γ signal peptide SPIgG (BAA75024.1); (2) Ig heavy chain ε signal peptide SPigE (AAB59424.1); and (3) cystatin S precursor signal peptide SPCS (NP_0018901.1). The Signal P prediction program was used to optimize the signal peptide cleavage site of the core fusion by computer simulation. Secretion efficiency was determined by analyzing the core protein content in the supernatant. Using Western blot analysis, the core antigen secretion display using three different signal peptides fused at the N-terminus, the cystatin S signal peptide caused the most efficient protein secretion (Figure 25F).

DNA vaccine vector optimization <br/> The optimized expression cassette containing the triple complex enhancer sequence upstream of the HBV antigen coding sequence with the N-terminal cystatin S signal peptide sequence was cloned into the DNA vaccine vector pVax-1 (Life Technologies, Thermo Fisher Scientific, Waltham, MA). The expression cassette in pVax-1 contains the human CMV-IE promoter followed by a bovine growth hormone (BGH) -derived polyadenylation sequence (pA). Bacterial reproduction lines are driven by the origin of pUC replication and the kanamycin resistance gene (Kan R) is driven by a small eukaryotic promoter. The origin of pUC replication, the KanR performance cassette, and the performance cassette driven by the CMV-IE promoter all showed the same orientation in the plastid backbone. However, the expression of the core antigen observed in the pVax-1 vector was significantly lower than that observed in the pcDNA3.1 vector.

Several strategies were used to improve protein performance: (1) reverse the entire pUCori-KanR cassette in a counterclockwise orientation (pVD-core); and (2) change the codon usage of the KanR gene and use Amp from the pcDNA3.1 vector The promoter replaces the Kan promoter (pDK-core). Both strategies restored core antigen performance, but the core antigen performance was highest with the pDK vector, which contains the codon-adjusted Kan R gene, the AmpR promoter (instead of the KanR promoter), and the reverse-oriented pUCori-KanR card cassette.

Four different HBV core / pol antigen-optimized performance cassettes as shown in Figure 25G were introduced into the pDK plastid backbone to test the three performance strategies shown in Figures 25A-25C, respectively. By Western blot analysis, core and pol specific antibodies were used to test these plastids in vitro for core and pol antigen performance. When the core and pol antigens are encoded by independent vectors, that is, individual DNA vectors pDK-core and pDK-pol, the most consistent representation of cells and secreted core and pol antigens is achieved (Figure 25H). Schematic representations of pDK-pol and pDK-core vectors are shown in Figures 26A and 26B, respectively.

TriGrid Example 2 using the delivery system. (TDS - IM) based on an electroporation-mediated nucleic acid of the biological agent with muscle <br/> administered using the device for the exemplary device, such as type II TriGrid delivery system provided herein ( TDS-IM) can enhance intracellular delivery of nucleic acid sequences in the skeletal muscles of the upper or lower limbs of an individual. In some embodiments, the TDS-IM device is used in combination with an agent approved for research use of the TDS-IM device. In an exemplary embodiment, the approved agent is a nucleic acid, that is, DNA or RNA. In some embodiments, the use of TDS-IM devices is limited to individuals in need. In some embodiments, the use of TDS-IM devices is limited to individuals recruited in open clinical trials of electroporation-mediated intramuscular nucleic acid delivery.

In order to start the system for administration, the main power source of the device is connected to the stimulator and the battery of the system is fully charged for use. Turn on the main power switch and press the front panel power button. Connect the applicator to the applicator connector. The applicator power indicator lights to determine the proper connection of the applicator. Once the system has completed all self-tests, the start screen appears. Press the OK button for program administration.

To insert the syringe into the TDS barrel, remove the syringe cap from the syringe and align the syringe flange with the TDS barrel syringe loading port. The syringe should snap into place and be fully seated in the TDS barrel. After loading the syringe to it, press the OK button on the pulse stimulator to continue. The cartridge cap should remain attached to the cartridge until the cartridge is loaded into the applicator.

To insert the syringe-loaded cartridge into the applicator, align the cartridge with the applicator with the cartridge syringe loading port facing up. When the cartridge is inserted into the applicator, the cartridge is automatically pulled into its fully loaded position in the applicator. Successful loading of the canister is indicated in the stimulator. Immediately after loading the cartridge, the applicator is returned to its stent and the appropriate injection site is selected on the individual. In some embodiments, the injection site for intramuscular nucleic acid delivery is in the middle of the deltoid muscle at about three finger widths below the edge of the acromion (scapula). In an exemplary embodiment, the injection depth at the middle of the deltoid muscle is about 0.75 "-1.25" (19-30 mm). In some embodiments, the injection site for intramuscular nucleic acid delivery is the lateral femoral muscle (outer thigh) at approximately the midpoint between the hip and knee. In an exemplary embodiment, the injection depth at the lateral femoral muscle is about 1.0 "-1.5" (25-38mm). After the injection site is selected, the applicator depth selection button is pressed, followed by the injection depth selection button corresponding to the injection site / depth. In some embodiments, the injection depth selection indicator becomes solidly lit to determine the selected injection depth. In some embodiments, the depth selection button on the right-hand side corresponds to a deeper injection depth. In one embodiment where the injection site to be initially selected is changed, a selection button corresponding to another injection depth is pressed, and when returning to the selection injection depth screen, another injection depth is selected.

To begin administering the approved medicament to the individual via the TDS-IM device, the canister cap is removed and discarded. Align the device with the target injection site and press firmly against the target injection site. When the device is firmly pressed against the target injection site, all four rows of applicator placement indicators light up, and the program countdown timer lights up and displays "8" seconds, indicating the remaining time of the administration program. The applicator trigger is depressed to administer the medicament while maintaining consistent pressure. When the program countdown timer reaches "0", electrical stimulation is delivered. After the administration procedure is completed, the procedure completion indicator lights up and the device can be withdrawn from the injection site. The device will not exit from the injection site until the program is complete or the program fault indicator is lit. In some embodiments where the device detects a problem during the administration process, the device aborts the administration process and lights up a process failure indicator. In an exemplary embodiment in which the device suspends the dosing procedure and the device is removed from the individual immediately, the device's stimulator display provides further instructions.

In order to eject the cartridge from the applicator after administering the medicament to the individual in need, the eject button on the stimulator is pressed down. The applicator automatically advances the cartridge to a position where it can be manually removed from the applicator. After the cartridge stops moving, the sides of the cartridge, as indicated by the arrows, can be gripped to pull the cartridge out of the applicator. After the cartridge is removed, the complete injection can be verified by checking the position of the syringe plunger. To shut down the device, place the applicator in a rack and press the front panel power button for 5 seconds.

Example 3 TDS to mice and non-human primates of -. IM electroporation using TriGrid <br/> delivery system - intramuscular (TDS-IM) electroporation techniques enhance muscle tissue of DNA-based delivery construct . The TriGrid electrode array for intramuscular (IM) delivery consists of four electrodes arranged in two equilateral triangles around the center injection needle to form a diamond shape ( Figure 22 ). The drug delivery and electric field propagation are integrated in a single device to ensure the induction of electroporation at the drug distribution site and allow the injection of plastid structures and the application of electrical pulses in a single step. In this way, IM delivery of plastid DNA is achieved in an efficient and reproducible manner.

The TriGrid electrode array used for each animal model was scaled to efficiently deliver DNA plastids, such as those prepared in Example 1, to selected muscles, regardless of size and overlying tissue characteristics. The major electroporation device parameters used for each animal model are scaled to TriGrid size (i.e., electrode spacing), applied voltage (250 V / cm, where cm is the size of TriGrid), electrode diameter, and electrode penetration depth . Secondary adjustments for this device include changes in syringe volume, plastid DNA volume, and hypodermic needle size / length.

The mouse research was performed using a TDS-IM v1.0 device using an electrode array with an electrode spacing of 2.5 mm. This adjustment of electrode array size should be adapted to the smaller size of the mouse muscle. Figure 23A depicts a TDS-IM v1.0 TriGrid version suitable for use in a mouse model. Non-GLP monkey studies were also performed using TDS-IM v1.0 devices, but using an electrode array pitch of 6 mm. Figure 23B depicts a TDS-IM v1.0 TriGrid version suitable for use in a non-human primate (NHP) model.

The device used in GLP monkey research is the TDS-IM v2.0 device, which is also intended for human clinical research. This device has an electrode pitch of 6 mm and the same startup conditions as the TDS-IM v1.0 device. The electrode material of the TDS-IM v2.0 device is the same as the TDS-IM v1.0 device used in the NHP study described above, but the cartridge support structure and the syringe used to administer it are different. In addition, humans and non-human primates have different insertion depths. Therefore, the TDS-IM v2.0 device is equipped with a spacer in order to make the TDS-IM v2.0 device suitable for much smaller NHP muscles. Figure 24 depicts a TDS-IM v2.0 TriGrid version suitable for use in a non-human primate (NHP) model.

In each animal study, the animals were anesthetized and fixed prior to the administration of the electroporation procedure. The administration site was located using the local anatomical landmark as a reference point and an electric hair clipper was used, followed by a sterile swab to remove hair from the selected site. The TDS-IM array was inserted percutaneously into selected muscles, and the major axis of the diamond configuration was oriented parallel to the muscle fibers. After electrode insertion, an injection is initiated to distribute the DNA into the muscle. After the IM injection was completed, a 250 V / cm electric field was locally applied at a 10% duty cycle for a total duration of 400 ms (ie, effectively applied a total of 40 ms for a duration of 400 ms). After completing the electroporation procedure, the TriGrid ™ array is removed and the animals are allowed to recover. The component changes of each TDS-IM version and / or animal model are shown in Tables 1 and 2 below.
Table 1 : TDS-IM Version and TriGrid Details
Table 2 : Details of TDS-IM version and pulse stimulator

Example 4 for the human TDS -. IM <br/> electroporation using TDS-IM v2.0 devices in clinical studies. This device is basically the same as that used for GLP monkeys. Human and non-human primates have different insertion depths. Therefore, the TDS-IM v2.0 device is equipped with a spacer in order to make the TDS-IM v2.0 device suitable for much smaller NHP muscles. The stimulation conditions were essentially the same as those used for GLP monkeys, except that the penetration depth was about 19 mm, not 9 mm in GLP monkeys.

Although preferred embodiments of the invention have been shown and described herein, it should be apparent to those skilled in the art that these embodiments are provided by way of example only. Those skilled in the art will now think of many variations, changes, and substitutions without departing from the invention. It should be understood that various alternatives to the embodiments described herein, or a combination of one or more of the embodiments or aspects described therein can be used to practice the invention. The following patentable scope is expected to define the scope of the invention, and therefore methods and structures within the scope of these patented scopes and their equivalents.

1‧‧‧ insertion depth

2‧‧‧ insertion angle

3‧‧‧ Tissue surface

4‧‧‧ Needle penetration position

5‧‧‧ needle

6‧‧‧ orifice

7‧‧‧ Area where pharmacy is administered

100‧‧‧ tube assembly

101‧‧‧Reservoir

102‧‧‧ Outer tube, also known as shell

103‧‧‧Inner tube

104‧‧‧Inner cylinder cap

105‧‧‧ needle

106‧‧‧ Outer cylinder cap

108‧‧‧ Alignment Guide / Tilt Feature

110‧‧‧External (safety) canister cap

112‧‧‧Tail end

114‧‧‧ cylinder lock ring

116‧‧‧Reservoir detection spring

118‧‧‧Reservoir detection cover

120‧‧‧Reservoir interlock device, also known as reservoir insert trigger

122‧‧‧electrode

124‧‧‧electrode support

126‧‧‧ Force contact spring

128‧‧‧ Force contact reader

130‧‧‧electrode contact

131‧‧‧ External outer barrel electrode part for coupling to applicator

132‧‧‧Stab wound shield support

133‧‧‧Internal outer tube electrode part for coupling to inner tube

134‧‧‧Stab wound protection cover

135‧‧‧ Proximal inner tube electrode part for coupling to outer tube

137‧‧‧Distal tube electrode part for tissue insertion

138‧‧‧ Force contact with flexible circuit

139‧‧‧ electrode shoulder or bend

140‧‧‧ reservoir loading port

142‧‧‧Reservoir enclosure

144‧‧‧Reservoir locking hole

144'‧‧‧ Reservoir locking hole (Group 1 or Group 2)

146‧‧‧Optical sight

148‧‧‧Insert detectors, such as emitter / collector IR sensor in applicator

150‧‧‧Inner tube closed body

152‧‧‧ Needle connector

154‧‧‧ rack

156‧‧‧Release port

158‧‧‧reservoir cover

159‧‧‧Plunger stop

160‧‧‧ flexible circuit

162‧‧‧The first group of gaskets

164‧‧‧Second set of gaskets

166‧‧‧Stab wound shield interface

167‧‧‧ puncture protection cover hole

168‧‧‧Tilt feature

170‧‧‧Alignment guide hole for puncture protection cover

172‧‧‧Starting sign

174‧‧‧Continue to sign

176‧‧‧External tube cover hook

178‧‧‧Chamfered surface of outer can

180‧‧‧ Hook to engage the side wall of the alignment guide / tilt shield 108

182‧‧‧Stab wound protection cover holding hook

184‧‧‧ first depth retaining wall

186‧‧‧Second depth retaining wall

188‧‧‧ initial or standing retaining wall

190‧‧‧ reminder tab

192‧‧‧electrode support feature electrode hole

194‧‧‧ electrode support feature pinhole

196‧‧‧electrode support feature electrode hole support structure

198‧‧‧ Electrode support feature sidewall

400‧‧‧ Applicator

401‧‧‧Applicator tube assembly receiving port

402‧‧‧Handle

403‧‧‧Cylinder assembly receiver defined by the shell

404‧‧‧user interface

406‧‧‧multi-wire cable

407‧‧‧Trigger

408‧‧‧ Injection depth selection indicator

409‧‧‧ Injection depth selection button (in another embodiment, it can be a jump switch or other form)

410‧‧‧Program countdown timer

412‧‧‧Application status indicator

414‧‧‧Procedure fault indicator

416‧‧‧Program completion indicator

418‧‧‧Power Indicator

420‧‧‧Top case

422‧‧‧First side housing

424‧‧‧Second side shell

426‧‧‧electrical connector

428‧‧‧end cap

430‧‧‧Front cover

432‧‧‧Inner protective shell

433‧‧‧for electrical contacts of motor driver 444

434‧‧‧Cylinder Force Sensor Contact

435‧‧‧Switch connector

436‧‧‧ Cartridge Load Sensor

438‧‧‧Cylinder Load Sensor

440‧‧‧Guide or track

442‧‧‧Drum guide

444‧‧‧Load drive motor

446‧‧‧Motor trigger connector

448‧‧‧ pinion assembly

450‧‧‧Electro-mechanical sub-assembly

452‧‧‧ Cartridge loading, electrode insertion and injection subassembly

454‧‧‧Load drive sub-assembly

456‧‧‧Cylinder loading sub-assembly

462‧‧‧Motor drive shaft

464‧‧‧System trigger switch

466‧‧‧Gear cover bracket

468‧‧‧Mounting bracket

470‧‧‧Spring cover / tube interface

471‧‧‧Spring cover hole

472‧‧‧electrode / needle insertion spring

474‧‧‧Insert Gear Bushing

476‧‧‧Mounting bracket

478‧‧‧Insertion gear drive ring

479‧‧‧Insert gear ring

480‧‧‧ Logo holder

481‧‧‧Insertion mechanism flag

482‧‧‧Insertion drive motor

483‧‧‧ Insertion mechanism position sensor

484‧‧‧Injection driven plunger

486‧‧‧ Injection drive motor and transmission

488‧‧‧holding column (holding feature)

490‧‧‧First Depth Channel

491‧‧‧locking tab

492‧‧‧Second Depth Channel

494‧‧‧butt side wall

496‧‧‧Applicator Electroporation Electrode Contact

700‧‧‧controller assembly

702‧‧‧Handle

704‧‧‧Storage silo

706‧‧‧Applicator holder

708‧‧‧Applicator connection port

710‧‧‧tray

712‧‧‧display

714‧‧‧Eject button

716‧‧‧Menu navigation buttons

718‧‧‧ mute button

720‧‧‧ Battery indicator

722‧‧‧Power button

724‧‧‧USB port

726‧‧‧Main power switch

728‧‧‧Main power port

750‧‧‧ Electric field generator

800‧‧‧flow chart

802‧‧‧step

804‧‧‧step

806‧‧‧step

808‧‧‧step

810‧‧‧step

811‧‧‧step

816‧‧‧step

818‧‧‧step

820‧‧‧step

822‧‧‧step

The features of the invention are elaborated in the scope of the attached patent application. A better understanding of the features and advantages of the present invention will be obtained with reference to the following detailed description, which illustrates exemplary embodiments utilizing the principles of the present invention and its accompanying drawings:

Figure 1 illustrates possible sources of spatial variation related to conventional syringe injections with needles.

FIG. 2 is an overview of a system according to the principles of the present invention. The system includes a cartridge assembly 100 , an applicator 400, and a controller system 700 .

FIG 3A - 3B show views of various aspects of the device described herein. FIG. 3A shows a side view of a cartridge assembly 100 according to the principles of the present invention. FIG. 3B shows a side view of the HBV vaccine reservoir 101 embodied as a syringe in accordance with the principles of the present invention.

FIG. 4 shows various exemplary components of a cartridge assembly 100 according to the principles of the present invention.

5A - 5E show views of the inner barrel and the cartridge breech in the device described herein. FIG. 5A illustrates a top view of the inner cylinder 103 according to the principles of the present invention. FIG. 5B shows a bottom view of the inner cylinder 103 and the tail end of the cylinder according to the principles of the present invention. FIG. 5C illustrates a detailed side perspective view of the inner cylinder 103 according to the principles of the present invention. FIG. 5D shows a side view of the barrel end 112 according to the principles of the present invention. FIG. 5E illustrates a reservoir interlock 120 having an improved locking feature to prevent the barrel end 112 from inadvertently moving forward.

FIG. 6 illustrates details of the barrel assembly according to the principles of the present invention, showing the rack 154 , the start mark 172 and the continue mark 174 .

FIG. 7A - 7B show a view of the device of the aspects described herein, the electrode and / or the electrode contact. FIG. 7A shows details of the electrode contacts 130 and various electrode contact portions according to the principles of the present invention. FIG. 7B shows details of the electrode 122 and various electrode contact portions according to the principles of the present invention.

FIG 8A - 8B show the device described herein, the contact force aspect of the view of the chain systems. FIG. 8A shows a top view of a force contact interlocking system according to the principles of the present invention. FIG. 8B illustrates details of a force contact interlocking system according to the principles of the present invention.

FIG. 9A - 9D show various aspects of view of the device described in this document. FIG. 9A shows a needle 105 and a distal inner barrel electrode 137 for tissue insertion according to the principles of the present invention. FIG. 9B illustrates details of a stick shield 134 according to the principles of the present invention. FIG. 9C illustrates an alignment guide 108 and a splay shield 168 according to the principles of the present invention. FIG. 9D illustrates the puncture protection cover support member integrally formed with the outer cylinder cover 106 .

10A - 10D are views showing the appearance of the outer can cover in the device described herein. FIG. 10A shows an external canister cover 110 according to the principles of the present invention. Figure 10B shows a side view of the outer cover 110 according to the principles of the present invention is cylindrical. FIG. 10C shows the outer canister cover 110 used in the alignment guide 108 and the tilt shield according to the principles of the present invention. FIG. 10D shows the outer canister cover 110 with an extension member designed to hold the inner canister 103 in place during handling and loading.

FIGS 11A - 11C show details of the shield puncture device of the aspect of the described herein. FIG. 11A shows the puncture protection cover retaining hook 182 of the puncture protection cover 134 according to the principles of the present invention. FIG. 11B shows details of the puncture protection cover 134 according to the principles of the present invention. FIG. 11C shows the puncture shield support 132 holding the puncture shield 134 in place in accordance with the principles of the present invention.

FIG. 12 shows details of an electrode support 124 according to the principles of the present invention.

FIGS 13A - 13B show the device as described herein in view of the applicator. FIG. 13A shows a side view of an applicator 400 according to the principles of the present invention. FIG. 13B shows a top view of an applicator 400 according to the principles of the present invention.

FIG. 14 is an exploded view of an applicator 400 according to the principles of the present invention.

FIG. 15 shows details of the side housing and the electroporation electrode connector 496 of the applicator 400 according to the principles of the present invention.

Figure 16 is an exploded view of the applicator according to the principles of the present invention, showing the cartridge loading sub-assembly 456 .

Figures 17A - 17B show views of the applicator in the device described herein. FIG. 17A is an exploded view of an applicator according to the principles of the present invention, showing a load-driven subassembly 454. FIG. FIG. 17B shows a rack 154 in a load driving sub-assembly 454 according to the principles of the present invention.

FIG. 18A - 18C is a view of a display mode of the device described herein of applying. FIG. 18A shows the details of the cartridge loading sub-assembly 456 according to the principles of the present invention, showing the insertion / injection driving assembly of the applicator and the cartridge assembly. FIG. 18B shows a cross-sectional view of a cartridge assembly according to the principles of the present invention, showing the insertion / injection driving assembly of the applicator and the cartridge assembly. FIG. 18C shows details of the cartridge loading, electrode insertion, and injection sub-assembly 452 of the applicator 400 according to the principles of the present invention.

FIG. 19 shows various components of a controller system according to the principles of the present invention.

FIG 20A - 20D show views of the device described in this document. FIG. 20A shows various components of a controller system according to the principles of the present invention. FIG. 20B shows details of the applicator connection port 708 and the tray 710 of the controller system according to the principles of the present invention. FIG. 20C shows details of a stimulator display screen of a controller system according to the principles of the present invention. FIG. 20D shows a detail of a rear view of a controller system according to the principles of the present invention.

FIG. 21 is a flowchart showing a method of operation according to the principles of the present invention.

FIG. 22 illustrates a TriGrid electrode array (section) for intramuscular (IM) delivery, which includes four electrodes arranged in two equilateral triangles around a central injection needle to form a diamond shape.

Figures 23A - 23B depict TDS-IM v1.0 TriGrid devices suitable for use in a mouse model ( Figure 23A ) and a non-human primate (NHP) model ( Figure 23B ).

Figure 24 depicts a TDS-IM v2.0 TriGrid device suitable for use in a non-human primate (NHP) model.

Figures 25A - 25H show the design and optimization of performance cassettes and DNA plastids encoding HBV polymerase and core antigen as described in Example 1; Figure 25A is a schematic representation of a performance strategy in which HBV core and polymerase antigen The coding sequence is in-frame fusion; Figure 25B is a schematic representation of a performance strategy, in which the core and polymerase antigen coding sequences are expressed from a single plastid by means of the ribosome FA2 slippage point; Figure 25C is a performance strategy Schematic representation, in which the core and polymerase antigens are expressed by two independent plastids; Figure 25D shows the expression of core antigens in HEK293T cells transfected with plastids that express core antigens in the presence and absence of post-transcriptional regulatory elements WPRE Western blot (Western blot); performance is tested using α-core antibodies in cell lysates (left) and supernatant (sup; right); Figure 25E shows western blot analysis, showing the use of cores to express plastid transformation Comparison of core performance in stained HEK293T cells. The core performance plastids include intron / exon sequences derived from the human apolipoprotein A1 precursor ("AI intron"), type 1 human T cell white blood Disease virus (HTLV-1) long terminal repeat (LTR) non-translated R-U5 domain ("HTLV R"), or HTLV-1 LTR, synthetic rabbit β-hemoglobin intron and splice enhancer Tertiary enhancer complex sequence ("triple"); unlabeled lanes are purified core proteins as size markers; performance is tested in lysates (left) and supernatants (sup; right); using three The core antigen expression of the enhancer complex sequence is the highest; Figure 25F is a western blot analysis of core antigen secretion using fusions of different signal peptides with the N-terminus of the HBV core antigen; the most efficient observed with cystatin S signal peptide Protein secretion; Figure 25G is a schematic representation of the optimized HBV core / pol antigen performance cassette for each of the three performance strategies shown in Figures 25A - 25C ; CMVpr: human CMV-IE promoter; TRE: triple enhancer sequence; SP : Cystatin S signal peptide; FA2: FMDV ribosomal slippage point; pA: BGH polyadenylation signal; Figure 25H is about HBV core of pDK vector containing each of the performance cassettes shown in Figure 25G Western blot analysis of polymer and antigen expression; lanes 1 and 2 pDK-core; lanes 3 and 4: pDK-polymerase; lanes 5 and 6: pDK-core FA2 polymerase; lanes 7 and 8: pDK-core-polymerase fusion: when the antigenic lines are encoded by independent vectors , Observed the most consistent performance map of cells and secreted core and polymerase antigen;

Figures 26A - 26B show schematic representations of DNA plastids according to an embodiment of the present application; Figure 26A shows DNA plastids encoding HBV polymerase (pol) antigens according to an embodiment of the present application; Figure 26B shows The DNA plastid encoding the HBV core antigen in one embodiment of the application; the HBV core and pol antigen are expressed under the control of the CMV promoter and carry the cystatin S signal peptide at the N-terminus. Cleavage from the expressed antigen; the plastid's transcriptional regulatory elements include an enhancer sequence located between the CMV promoter and the polynucleotide sequence encoding the HBV antigen and a bGH polymer located downstream of the polynucleotide sequence encoding the HBV antigen Adenylation sequence; a second expression cassette containing a kanamycin resistance gene under the control of the Amp ^r (bla) promoter in a reverse orientation in this plastid; also includes a reverse orientation Origin of replication (pUC);

Similar reference numerals refer to similar elements throughout. Elements are not drawn to scale unless indicated otherwise.

Claims (92)

A device for the controlled delivery of a HBV vaccine to a predetermined tissue site in an individual in need, comprising: A canister assembly comprising an outer canister, an inner canister, and a reservoir containing the HBV vaccine, wherein a reservoir closure system is contained within the outer canister and is configured to receive the reservoir; An applicator comprising a cartridge assembly receiver, a needle interface, and an insertion detector, wherein the insertion detector senses the loading of the reservoir in the reservoir enclosure; At least one interlocking device, wherein the interlocking device facilitates proper implementation of the HBV vaccine administration procedure; At least one injection orifice through which the HBV vaccine is administered; A plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the aperture; An electric field generator for generating electrical signals operatively connected to the electrodes; and A control energy source sufficient to deliver a predetermined amount of the HBV vaccine from the reservoir through the orifice to a predetermined site within the individual at a predetermined rate, The HBV vaccine contains: A first nucleic acid molecule comprising a first polynucleotide encoding a HBV polymerase antigen, wherein the HBV polymerase antigen comprises an amino acid sequence at least 98% identical to SEQ ID NO: 4 and wherein the HBV polymerase antigen does not have Reverse transcriptase activity and RNAse H activity; A second nucleic acid molecule comprising a second polynucleotide encoding a truncated HBV core antigen, the truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2; and A pharmaceutically acceptable carrier, The first nucleic acid molecule and the second nucleic acid molecule exist in the same nucleic acid molecule or in two different nucleic acid molecules. A kit for the controlled delivery of a HBV vaccine to a predetermined tissue site in an individual in need, comprising: (i) the HBV vaccine, which comprises: A first nucleic acid molecule comprising a first polynucleotide encoding a HBV polymerase antigen, wherein the HBV polymerase antigen comprises an amino acid sequence at least 98% identical to SEQ ID NO: 4 and wherein the HBV polymerase antigen does not have Reverse transcriptase activity and ribonuclease H activity; and A second nucleic acid molecule comprising a second polynucleotide encoding a truncated HBV core antigen, the truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2; and A pharmaceutically acceptable carrier, The first nucleic acid molecule and the second nucleic acid molecule exist in the same nucleic acid molecule or in two different nucleic acid molecules; (ii) a device comprising: A canister assembly comprising an outer canister, an inner canister, and a reservoir of the HBV vaccine, wherein a reservoir closure system is contained within the outer canister and is configured to receive the reservoir; An applicator comprising a cartridge assembly receiver, a needle interface, and an insertion detector, wherein the insertion detector senses the loading of the reservoir in the reservoir enclosure; At least one interlocking device, wherein the interlocking device facilitates proper implementation of the HBV vaccine administration procedure; At least one injection orifice through which the HBV vaccine is administered; A plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the aperture; An electric field generator for generating electrical signals operatively connected to the electrodes; and A control energy source sufficient to deliver a predetermined amount of the HBV vaccine from the reservoir via the orifice to a predetermined site within the individual at a predetermined rate. The device of claim 1 or the set of claim 2, wherein the electrodes are a plurality of elongated electrodes. A device as claimed in claim 1 or a set as claimed in claim 2, wherein the interlocking device prevents inadvertent activation of the cartridge function. The device of any one of 3 and 4, or the set of any one of claims 2 to 4, wherein at least one interlocking device is selected from the group consisting of a mechanical interlocking device, a light emitter / collector, a tube tail End, force interlocking device, alignment guide and tilt protective cover, trigger lock and safety switch. If the device of any one of claims 1 and 3 to 5, or the set of any one of claims 2 to 5, wherein the device includes at least two sets of interlocking devices, wherein the first interlocking device is a mechanical interlocking device, And the second interlocking device is selected from the group consisting of: a light emitter / collector, a barrel end, a force interlocking device, an alignment guide and a tilting protective cover, a trigger lock and a safety switch. The device or kit according to any one of claims 5 to 6, wherein the mechanical interlocking device is a reservoir interlocking device. The device or kit of claim 7, wherein the reservoir interlocking device further comprises at least one reservoir lock hole. The device or kit of any one of claims 5 to 8, wherein the end of the barrel provides optical sight through at least one reservoir latching hole. The device or kit of any one of claims 7 to 9, wherein the reservoir interlocking device further comprises a tab extending from the surface of one or more cartridges, wherein the tab ensures that the reservoir is properly inserted into the cartridge. The device or kit of claim 6, wherein the device further comprises a third interlocking device. The device or kit of claim 11, wherein the third interlocking device is a force interlocking device. The device or kit of claim 12, wherein the force interlocking device senses a force applied to a predetermined tissue site of the individual and prevents the HBV vaccine from being administered to the individual when the provided force is insufficient. The device or kit of any one of claims 12 to 13, wherein the force interlocking device further forms an electrical lock in the applicator. If the device of any one of claims 1 and 3 to 14, or the set of any one of claims 2 to 14, the device further includes a button for the reservoir, wherein the button is in the reservoir The barrel of the device is slid to ensure proper fit in the barrel assembly. The device or kit of any one of claims 5 to 6, wherein the inclined protective cover comprises at least one rib and at least one edge for engaging with a predetermined tissue site of the individual and keeping the device in contact with the deployment for The needle administered with the HBV vaccine is under vertical tension. The device of claim 1 or the kit of claim 2, wherein the barrel assembly further comprises a puncture protection cover. The device or kit of any one of 6 and 16, wherein the inclined protective cover includes at least one hole for slidably moving the puncture protective cover. The device of any one of claims 1 and 3 to 18, or the kit of any one of claims 2 to 18, wherein after the cartridge assembly is loaded into the applicator, the reservoir is directed to Move forward to fit the needle interface and bring the barrel into contact with the injection needle when the HBV vaccine is administered. The device according to any one of claims 1 and 3 to 19, or the kit according to any one of claims 2 to 19, wherein the inner cylinder is slidably moved relative to the outer cylinder. If the device of any one of claims 1 and 3 to 20, or the kit of any one of claims 2 to 20, wherein the inner cylinder is engaged with the inner cylinder cap at the distal end, wherein the inner cylinder cap will be The electrodes are locked in place and provide a support surface for the puncture shield. A device as claimed in claim 1 or a set as claimed in claim 2, wherein the device further comprises at least one sensor. The device or kit according to claim 22, wherein the sensor is selected from the group consisting of a cartridge loading sensor, a cartridge loading sensor, a cartridge force sensor, an insertion mechanism position sensor, and an insertion detection device. Detector, optical detector and inductive detector. The device or kit of claim 23, wherein the cartridge loading sensor and the cartridge loading sensor form part of a load driving sub-assembly. The device or kit of claim 24, wherein the loading driving sub-assembly further comprises at least one barrel guide and a loading motor. The device or kit according to any one of claims 24 to 25, wherein the load driving sub-assembly further has a connecting member connected to the pinion assembly, and the at least one rack on the base of the outer cylinder The cartridge assembly is pulled into the cartridge assembly receiver. The device or kit of claim 26, wherein the pinion assembly is engaged with a rack on the outer cylinder. The device or kit of any one of claims 26 to 27, wherein the rack includes at least one rack tooth. The device or kit of any one of claims 26 to 28, wherein the first rack teeth provide tactile sensation when the barrel assembly is inserted into the barrel assembly receiver. The device or kit of any one of claims 26 to 29, wherein the rack teeth provide torsional stability. The device or kit of any one of claims 23 to 26, wherein the cartridge loading sensor detects a start mark on the cartridge assembly to initiate loading. The device or kit according to any one of claims 23 to 26, wherein the cartridge loading sensor detects a start mark on the cartridge assembly to stop loading. The device of any one of claims 1 and 3 to 32, or the kit of any one of claims 2 to 32, wherein the device further includes a continue flag for continuing the loading of the cartridge. The device or kit of claim 23, wherein the insertion detector is a light emitter / collector IR sensor. The device or kit of any one of claims 22 to 34, wherein the sensor detects a reservoir marker and validates the HBV vaccine. A device for the controlled delivery of an HBV vaccine to a predetermined tissue site in an individual, comprising: A cartridge assembly comprising a housing and a reservoir containing the HBV vaccine, wherein the reservoir enclosure is contained within the housing and is configured to receive the reservoir; An applicator comprising a cartridge assembly receiver, a needle interface, and an insertion detector, wherein the insertion detector senses the loading of the reservoir in the reservoir enclosure; at least one injection orifice, the HBV vaccine Administered through the orifice; A plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the aperture; An electrode support structure including a hole and a side wall, wherein the electrode support structure prevents the electrodes from inadvertently moving vertically; An electric field generator for generating electrical signals operatively connected to the electrodes; and A control energy source sufficient to deliver a predetermined amount of the HBV vaccine from the reservoir through the orifice to a predetermined site within the individual at a predetermined rate, The HBV vaccine contains: A first nucleic acid molecule comprising a first polynucleotide encoding a HBV polymerase antigen, wherein the HBV polymerase antigen comprises an amino acid sequence at least 98% identical to SEQ ID NO: 4 and wherein the HBV polymerase antigen does not have Reverse transcriptase activity and ribonuclease H activity; A second nucleic acid molecule comprising a second polynucleotide encoding a truncated HBV core antigen, the truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2; and A pharmaceutically acceptable carrier, and The first nucleic acid molecule and the second nucleic acid molecule exist in the same nucleic acid molecule or in two different nucleic acid molecules. A kit comprising: (i) HBV vaccine, which contains A first nucleic acid molecule comprising a first polynucleotide encoding a HBV polymerase antigen, wherein the HBV polymerase antigen comprises an amino acid sequence at least 98% identical to SEQ ID NO: 4 and wherein the HBV polymerase antigen does not have Reverse transcriptase activity and ribonuclease H activity; A second nucleic acid molecule comprising a second polynucleotide encoding a truncated HBV core antigen, the truncated HBV core antigen consisting of the amino acid sequence of SEQ ID NO: 2; and A pharmaceutically acceptable carrier, and Wherein the first nucleic acid molecule and the second nucleic acid molecule exist in the same nucleic acid molecule or in two different nucleic acid molecules; and (ii) a device comprising: A cartridge assembly comprising a housing and a reservoir of the HBV vaccine, wherein a reservoir enclosure is contained within the housing and is configured to receive the reservoir; An applicator comprising a cartridge assembly receiver, a needle interface, and an insertion detector, wherein the insertion detector senses the loading of the reservoir in the reservoir enclosure; At least one injection orifice through which the HBV vaccine is administered; A plurality of penetrating electrodes arranged in a predetermined spatial relationship with respect to the aperture; An electrode support structure including a hole and a side wall, wherein the electrode support structure prevents the electrodes from inadvertently moving vertically; An electric field generator for generating electrical signals operatively connected to the electrodes; and A control energy source sufficient to deliver a predetermined amount of the HBV vaccine from the reservoir via the orifice to a predetermined site within the individual at a predetermined rate. The device of claim 36 or the set of claim 37, wherein when the electrodes are deployed in a predetermined tissue site within the individual, the electrode support structure is on a conductive contact area located on a distal region of the electrodes An operational connection is provided to this control energy source. The device of claim 36 or 38, or the kit of claim 37 or 38, wherein the electrode support structure includes at least one hole through which the electrodes pass and a hole through which the injection needle passes. If the device of any one of claims 36 and 38 to 39, or the set of any one of claims 37 to 39, wherein the electrode support structure is a flat support positioned vertically with respect to the elongation direction of the electrodes structure. The device or kit of claim 40, wherein the planar support structure includes at least one aperture configured to allow the electrode to pass through the predetermined tissue site. The device or kit of any one of claims 40 to 41, wherein the aperture comprises at least one tubular structure arranged perpendicular to the planar support structure. The device or kit of any one of claims 40 to 42, wherein the planar support structure is oriented perpendicular to the electrodes. The device according to any one of claims 36 and 38 to 43, or the kit according to any one of claims 37 to 43, wherein the electrode supporting structure is an adaptable electrode support. The device or kit of claim 44 wherein the adjustable electrode support is a compression spring. The device or kit of claim 45, wherein the compression spring is made of metal, polymer or elastic material. The device of any one of claims 36 and 38 to 46, or the kit of any one of claims 37 to 46, wherein the electrode support structure includes at least one telescoping sleeve. The device according to any one of claims 36 and 38 to 47, or the kit according to any one of claims 37 to 47, wherein the electrode supporting structure further comprises a puncture shield spring. The device of claim 36 or the set of claim 37, wherein the electrode support structure further comprises at least one lateral support member attached to the electrodes with at least one optional hinge feature. The device of any one of claims 36 and 38 to 49, or the kit of any one of claims 37 to 49, wherein the electrode support structure is made of metal, polymer, ceramic, composite or compressible matrix Material manufacturing. The device or kit of claim 50, wherein the compressible matrix material is selected from the group consisting of cellulose, foamed plastic, rubber polymer, microporous plastic, foamed silicon, and foamed polychloroprene And carbon foam matrix. The device according to any one of claims 36 and 38 to 51, or the kit according to any one of claims 37 to 51, wherein the electrode supporting structure is made of a non-conductive material. The device of any one of claims 36 and 38 to 52, or the kit of any one of claims 37 to 52, wherein the electrode support structure is made of a thermoplastic material. The device or kit of claim 53, wherein the thermoplastic material is selected from the group consisting of polycarbonate, polystyrene, polypropylene, acrylic, and polyethylene. A device as claimed in any of claims 36 and 38 to 54 or a kit as claimed in any of claims 37 to 54 wherein the electrode support structure supports percutaneous deployment of the electrode and is maintained at up to 60 mm Organizational depth. The device according to any one of claims 36 and 38 to 55, or the set according to any one of claims 37 to 55, wherein the proximal portion of the electrode is an electrode contact and is positioned in the inner tube of the tube assembly On the outside, the configuration of the electrode contact is in electrical communication with the corresponding connector on the applicator. The device or kit of claim 56, wherein the electrode contact further includes an external contact of the outer cylinder. The device or kit of any one of claims 56 to 57, wherein the electrode contact provides a conductive interface with the corresponding electrode without interfering with the forward travel of the electrodes mounted on the inner cylinder. The device of any one of 3 to 36 and 38 to 58 or the kit of any one of claims 2 to 35 and 37 to 58, wherein the applicator further comprises an injection-driven assembly, wherein the injection-driven assembly Cooperate with the tube assembly. The device of any of 3 to 36 and 38 to 58 or the set of any one of claims 2 to 35 and 37 to 58, wherein the device further includes a depth selection button. The device or kit of claim 60, wherein the depth selection button is selected from the group consisting of a jump switch, a switch and a slide switch. The device of any one of 3 to 36 and 38 to 58 or the kit of any one of claims 2 to 35 and 37 to 58, wherein the device further comprises a plurality of channels and a plurality of holding columns. The device of any one of 3 to 36 and 38 to 58 or the kit of any one of claims 2 to 35 and 37 to 58, wherein the device further includes an insert mechanism gear drive ring. The device or kit of claim 63, wherein the rotation of the insertion mechanism gear ring rotates the holding post into the channel. The device of any one of 3 to 36 and 38 to 64, or the set of any one of claims 2 to 35 and 37 to 64, wherein the cartridge combination system is for a single use. The device of any of 3 to 36 and 38 to 64, or the kit of any one of claims 2 to 35 and 37 to 64, wherein the applicator is for multiple use. The device of any one of 3 to 36 and 38 to 66, or the kit of any one of claims 2 to 35 and 37 to 66, wherein the applicator further comprises a top case, a side case, an inner protective case, Front cover and end cover. The device of any of 3 to 36 and 38 to 67, or the set of any one of claims 2 to 35 and 37 to 67, wherein the applicator further includes a user interface, a program start trigger, and a program countdown Timer, program fault indicator or application status indicator. The device of any one of 3 to 36 and 38 to 68, or the kit of any one of claims 2 to 35 and 37 to 68, wherein the device further includes a controller. The device of any one of 3 to 36 and 38 to 69, or the kit of any one of claims 2 to 35 and 37 to 69, wherein the applicator further includes a connector for connecting to the controller. The device or kit of any one of claims 69 to 70, wherein the controller further comprises an electric field controller. The device of any one of 3 to 36 and 38 to 71, or the kit of any one of claims 2 to 35 and 37 to 71, wherein the penetrating electrodes and / or the injection orifices are at least 50 mm The speed per second is in contact with the predetermined tissue site. The device of any one of 3 to 36 and 38 to 71, or the kit of any one of claims 2 to 35 and 37 to 71, wherein the penetrating electrodes and / or the injection orifices are at least 500 mm The speed per second is in contact with the predetermined tissue site. The device of any of 3 to 36 and 38 to 73, or the kit of any one of claims 2 to 35 and 37 to 73, wherein the HBV vaccine is capable of eliciting in mammals against at least two HBV genotypes The immune response is preferably that the HBV vaccine is capable of inducing a T cell response against at least HBV genotypes B, C, and D in a mammal, and more preferably, the HBV vaccine is capable of inducing a human at least HBV genotype A , B, C, and D CD8 T cell responses. The device or kit of claim 74, wherein the first non-naturally occurring nucleic acid molecule is present in a first plastid DNA vector, and the second non-naturally occurring nucleic acid molecule is present in a second plastid DNA vector in. The device or kit of claim 75, wherein the first plastid DNA vector and the second plastid DNA vector each include an origin of replication, an antibiotic resistance gene, and a promoter sequence from the 5 'end to the 3' end, An enhancer sequence, a signal peptide coding sequence, the first polynucleotide sequence or the second polynucleotide sequence, and a polyadenylation signal sequence. The device or kit of claim 76, wherein the antibiotic resistance gene has a kana that is at least 90% identical to SEQ ID NO: 12, preferably 100% identical to SEQ ID NO: 12 Mycin resistance gene. The device or kit of claim 77, wherein the HBV vaccine comprises: a) A first plastid DNA vector, from the 3 ′ end to the 5 ′ end, comprising the promoter sequence containing the polynucleotide sequence of SEQ ID NO: 7 and the polynucleotide sequence containing SEQ ID NO: 8 The enhancer sequence, the signal peptide coding sequence containing the polynucleotide sequence of SEQ ID NO: 5, the first polynucleotide sequence containing the polynucleotide sequence of SEQ ID NO: 3, and the sequence containing SEQ ID The polyadenylation signal sequence of the polynucleotide sequence of NO: 11; b) a second plastid DNA vector, from the 3 ′ end to the 5 ′ end, comprising the promoter sequence containing the polynucleotide sequence of SEQ ID NO: 7 and the polynucleotide sequence containing SEQ ID NO: 8 Control sequence, the signal peptide coding sequence containing the polynucleotide sequence of SEQ ID NO: 5, the second polynucleotide sequence containing the polynucleotide sequence of SEQ ID NO: 1, and the second polynucleotide sequence containing SEQ ID NO: The polyadenylation signal sequence of the polynucleotide sequence of 11; and c) a pharmaceutically acceptable carrier, Wherein the first plastid DNA vector and the second plastid DNA vector each further comprise a kanamycin resistance gene having a polynucleotide sequence of SEQ ID NO: 12 and a polynucleoside having SEQ ID NO: 10 Origin of replication of the acid sequence, and The first plastid DNA vector and the second plastid DNA vector are in the same composition or two different compositions. The device or kit of claim 78, wherein the HBV vaccine does not contain a nucleic acid molecule encoding a HBV antigen selected from the group consisting of hepatitis B surface antigen (HBsAg), HBV envelope (Env) antigen, and HBV L protein antigen, It also does not contain HBV antigens selected from the group consisting of hepatitis B surface antigen (HBsAg), HBV envelope (Env) antigen, and HBV L protein antigen. A method for inducing an immune response against HBV infection in an individual in need, comprising using a device as in any of claims 1, 3 to 36 and 38 to 79, or as in claims 2 to 35 and 37 to 79 The kit of any one delivers a HBV vaccine to a predetermined tissue site within the individual, wherein the individual is preferably a human with a chronic HBV infection. A method for treating an HBV-induced disease in a subject in need, comprising using a device such as any of claims 1, 3 to 36, and 38 to 79, or any one of claims 2 to 35, and 37 to 79 The set of items delivers the HBV vaccine to a predetermined tissue site in the individual, preferably wherein the system is a human individual and wherein the HBV-induced disease is selected from the group consisting of advanced fibrosis, cirrhosis and hepatocellular carcinoma (HCC) group. The method of claim 80 or 81, wherein the predetermined tissue site is located in a skeletal muscle of the individual. The method of claim 82, wherein the individual's skeletal muscle is the middle of the deltoid muscle. The method of claim 83, wherein the injection depth at the middle of the deltoid muscle is about 3-30 mm. The method of claim 82, wherein the individual's skeletal muscles are lateral femoral muscles. The method of claim 85, wherein the injection depth at the lateral femoral muscle is about 3-38 mm. The method of any one of claims 80 to 86, wherein the electric signal operatively connected to the electrodes comprises an electric field strength of 100 V / cm to 400 V / cm, preferably 250 V / cm. The method of claim 87, wherein the electrical signal operatively connected to the electrodes has a voltage of 50 to 200 V, preferably about 150 V. The method of claim 88, wherein the electric signal operatively connected to the electrodes has a current of 0.5 to 5 A / sec, preferably 0.6 to 4 A / sec, and more preferably 0.16 A / sec. The method of claim 89, wherein the electrical signal operatively connected to the electrodes has about 1 to 10 electrical pulses, preferably 6 pulses. The method of claim 90, wherein the electrical signal operatively connected to the electrodes has an effective duration of 30 to 50 milliseconds, preferably 40.8 milliseconds, and the total duration of the effective duration applied is 200 to 500 milliseconds , Preferably about 370 milliseconds. The device of any one of 3 to 36 and 38 to 79, or the set of any one of claims 2 to 35 and 37 to 79, is used to elicit an immune response against at least two HBV genotypes, and more In particular, for treating HBV-induced diseases, the device is suitable for performing electroporation of the HBV vaccine in individuals in need, preferably humans.