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WO2010127369A1 - Sonoporation, ablation de tissu par ultrasons, guidées par tomographie par impédance électrique (eit), et utilisation de celles-ci - Google Patents

Sonoporation, ablation de tissu par ultrasons, guidées par tomographie par impédance électrique (eit), et utilisation de celles-ci Download PDF

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Publication number
WO2010127369A1
WO2010127369A1 PCT/US2010/033450 US2010033450W WO2010127369A1 WO 2010127369 A1 WO2010127369 A1 WO 2010127369A1 US 2010033450 W US2010033450 W US 2010033450W WO 2010127369 A1 WO2010127369 A1 WO 2010127369A1
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Prior art keywords
body mass
tissue
ultrasound
cell
eit
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English (en)
Inventor
Yong Huang
Xufeng Xi
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Rational Biotechnology Inc
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Rational Biotechnology Inc
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61N—ELECTROTHERAPY; MAGNETOTHERAPY; RADIATION THERAPY; ULTRASOUND THERAPY
    • A61N7/00—Ultrasound therapy
    • A61N7/02—Localised ultrasound hyperthermia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61N—ELECTROTHERAPY; MAGNETOTHERAPY; RADIATION THERAPY; ULTRASOUND THERAPY
    • A61N7/00—Ultrasound therapy
    • A61N7/02—Localised ultrasound hyperthermia
    • A61N2007/025—Localised ultrasound hyperthermia interstitial

Definitions

  • Sonoporation has been used as a new nonviral and non- invasive technique for gene transfection and target drug delivery.
  • Sonoporation typically utilizes ultrasound or the interaction of ultrasound with contrast agents (commonly stabilized microbubbles) to temporarily permeabilize the cell membrane.
  • Therapeutic agents mixed with ultrasound contrast agents can be injected locally or systemically, and ultrasound can be coupled and even focused into the defined area to achieve targeted delivery of the therapeutic agents.
  • the membrane permeability caused by the sonoporation is transient, leaving the agents trapped inside the cell after the ultrasound exposure.
  • Ultrasound based tissue ablation technique such as HIFU (High Intensity Focused Ultrasound) is a non-invasive therapeutic method using high- intensity ultrasound to heat and destroy malignant tissue without causing damage to overlying or surrounding health tissue.
  • HIFU High Intensity Focused Ultrasound
  • the target tissue temperature can be increased to 70-100 0 C within a couple of seconds.
  • biological effects including coagulative necrosis and structural disruption can be induced in the tissue.
  • HlFU has been used m the treatment of solid tumors such as breast cancer and other diseases.
  • EIT Electrical impedance tomography
  • EIT is an imaging technique in which an image of the conductivity or permittivity of part of the body is constructed by injecting small electrical signals (potentials or currents) through surface or embedded source electrodes to the body part, followed by measuring the electrical signals (potentials or currents) at the corresponding receiving electrodes.
  • EIT has been used to measure effectiveness of cell electroporation, under which, cell membrane impedance is compromised due to electroporation- mediated increase of cell membrane permeability. See, e.g., U.S. Patent Number 6,387,671, which is hereby incorporated by reference in its entirety.
  • electroporation requires contact for delivering the current to cause cell permeabilization. Furthermore, electroporation is limited to localized delivery of the current. EIT can be used to generate two or three-dimension impedance images. It's practice can also be reduced to electrical impedance measurement that does not necessarily generate an image.
  • An aspect of the invention is directed to a method of applying ultrasound to a section of body mass within a subject.
  • the method may comprise the steps of: sending an electric current between a first point and a second point across the section of body mass, generating an impedance profile of the section of body mass based on the electrical impedance of the section of body mass, and applying ultrasonic energy to the section of body mass subsequent to or concurrent with said sending of the electric current.
  • the body mass may be selected from the group consisting of transformed and non-transformed tissues, including muscle, vascular, endothelial, connective, retinal, hepatic, lymphatic, adipose, epithelial, neural, and hematopoietic tissues; organs and/or portions therof, including kidney, liver, lung, heart, spleen, pancreas, prostate, brain, and islets of langerhans; glands such as thyroid, parathyroid, paracrine glands, adrenal, endocrine glands, and exocrine glands; and transformed or tumor tissues, including solid tumors such as carcinomas, including cervical carcinoma, hepatocellular carcinoma, colorectal carcinoma, basal cell carcinoma, renal cell carcinoma, prostate carcinoma, small cell lung carcinoma, breast carcinoma; sarcomas, endotheliomas; mesotheliomas; including acute and chronic Hepatitis; including viral hepatitis such as Hepatitis A, Hepatitis B and He
  • the ultrasonic energy may be sufficient to cause reversible cell permeabilization in the section of body mass, or may be sufficient to cause irreversible cell permeabilization or cell death in the section of body mass.
  • the ultrasonic energy may have an energy intensity of 0.1 W/cm or greater.
  • the method may also include the step of administering to said subject a therapeutic agent, wherein application of said ultrasonic energy enhances the delivery of said therapeutic agent to said body mass.
  • the therapeutic agent may be selected from the group consisting of: peptides, polypeptides, polynucleotides, siRNA, microRNA, small molecule drugs, inorganic compounds, and organic compounds.
  • the therapeutic agent may be delivered in combination with a carrier, a contrast agent, an enhancing agent, or a combination thereof. In some instances, the therapeutic agent may be delivered across the blood brain barrier.
  • At least a portion of said body mass may be disrupted or destroyed by cavitation of microbubbles.
  • the microbubbles may be generated in the subject by the application of the ultrasonic energy.
  • the microbubbles may be administered to the patient prior to or during the application of said ultrasonic energy. Disruptions may result in a decrease in the appearance of a skin irregularity.
  • the body mass may be a blood clot, cancer tissue, fibrotic or cirrhotic tissue, viral-infected tissue, or alcoholic liver.
  • a system for applying ultrasonic energy to a section of body mass of a subject may comprise: a first electrode and a r oc e o. - . second electrode across the section of body mass, configured to provide an electric current between the first electrode and the second electrode; a sensor capable of sensing an electrical property of the section of body mass; a processor for generating an impedance profile of the section of body mass based on data correlated with the electrical property collected by the sensor; and an ultrasonic probe configured for applying ultrasonic energy to the section of body mass subsequent to or concurrent with the sensing of the electrical property.
  • the electrical property may be voltage, impedance, resistance, conductance, and/or inductance.
  • FIG. 1 shows an example of a system utilizing electrical impedance tomography (EIT) and an ultrasound transmitting system, in accordance with an embodiment of the invention.
  • EIT electrical impedance tomography
  • FIG. 2A shows an example of cells that have not undergone successful sonoporation or cell ablation.
  • FIG. 2B shows an example of cells that have undergone successful sonoporation or cell ablation.
  • FIG. 3 shows an example of an EIT system for use within the invention.
  • FIG. 4 shows an example of an ultrasound system in accordance with an embodiment of the invention, which may be used for sonoporation or ultrasound tissue ablation.
  • FIG. 5 shows an ultrasonic probe applying an ultrasonic signal to a section of body mass in accordance with an embodiment of the invention.
  • FIG. 6 shows an example of an experimental setup for ultrasound mediated siRNA delivery ex vivo. r oc e o. - .
  • FIG. 7 shows an example of how cell impedance level may vary over time under different permeabilization conditions.
  • FIG. 8A shows a conceptual illustration of electrical impedance changes due to (a) reversible and (b) irreversible membrane permeabilization of cells subjected to transient electrical or ultrasonic stimulation.
  • FIG. 8B shows an example of representative patterns of resistance changes during and after cell electroporation.
  • FIG. 9A shows examples of bright field and fluorescent images of control MDCK cell monolayer for siRNA delivery without being treated with ultrasound.
  • FIG. 9B shows examples of bright field and fluorescent images of MDCK cell monolayer for siRNA delivery that have been successfully sonoporated.
  • FIG. 1 OA shows fluorescent and bright field images of a slice from a liver tissue without siRNA and sonoporation treatment, wherein the dim fluorescence in the image is due to auto- fluorescence of liver cells.
  • FIG. 1 OB shows fluorescent and bright field images of a slice from a control liver tissue injected with siRNA but without being treated with ultrasound.
  • FIG. 1 OC shows fluorescent and bright field images of a slice from a successfully sonoporated liver tissue.
  • FIG. 1 IA shows gross images of multi-nodular HCC induced in LAP-tTA;TRE-Myc mice after 8 weeks post removal of doxycycline.
  • FIG. 1 IB shows H&E staining of liver tumor specimen.
  • polynucleotide refers to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof. Polynucleotides may have any three dimensional structure, and may perform any function, known or unknown.
  • polynucleotides coding or non-coding regions of a gene or gene fragment, loci (locus) defined from linkage analysis, exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, short interfering RNA (siRNA), short-hairpin RNA (shRNA), micro-RNA (miRNA), ribozymes, cDNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes, and primers.
  • loci locus defined from linkage analysis, exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, short interfering RNA (siRNA), short-hairpin RNA (shRNA), micro-RNA (miRNA), ribozymes, cDNA, recombinant polynucleotides, branched poly
  • a polynucleotide may comprise modified nucleotides, such as methylated nucleotides and nucleotide analogs. If present, modifications to the nucleotide structure may be imparted before or after assembly of the polymer. The sequence of nucleotides may be interrupted by non nucleotide components. A polynucleotide may be further modified after polymerization, such as by conjugation with a labeling component. r oc e o. - .
  • expression refers to the process by which a polynucleotide is transcribed into mRNA and/or the process by which the transcribed mRNA (also referred to as “transcript”) is subsequently being translated into peptides, polypeptides, or proteins.
  • the transcripts and the encoded polypeptides are collectedly referred to as "gene product.” If the polynucleotide is derived from genomic DNA, expression may include splicing of the mRNA in a eukaryotic cell.
  • polypeptide polypeptide
  • peptide and “protein” are used interchangeably herein to refer to polymers of amino acids of any length.
  • the polymer may be linear or branched, it may comprise modified amino acids, and it may be interrupted by non amino acids.
  • the terms also encompass an amino acid polymer that has been modified; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation, such as conjugation with a labeling component.
  • amino acid refers to either natural and/or unnatural or synthetic amino acids, including glycine and both the D or L optical isomers, and amino acid analogs and peptidomimetics.
  • the terms "subject,” “individual,” and “patient” are used interchangeably herein to refer to a vertebrate, preferably a mammal, more preferably a human. Mammals include, but are not limited to, murines, simians, humans, farm animals, sport animals, and pets. Tissues, cells and their progeny of a biological entity obtained in vivo or cultured in vitro are also encompassed.
  • the terms "therapeutic agent”, “therapeutic capable agent” and “treatment agent” are used interchangeably and refer to a molecule or compound that confers some beneficial effect upon administration to a subject.
  • the beneficial effect includes enablement of diagnostic determinations; amelioration of a disease, symptom, disorder, or pathological condition; reducing or preventing the onset of a disease, symptom, disorder or condition; and generally counteracting a disease, symptom, disorder or pathological condition.
  • treatment or “treating,” or “palliating” or “ameliorating” is used interchangeably. These terms refer to an approach for obtaining beneficial or desired results including but not limited to a therapeutic benefit and/or a prophylactic benefit.
  • therapeutic benefit is meant any therapeutically relevant improvement in or effect on one or more diseases, conditions, or symptoms under treatment.
  • the compositions may be administered to a subject at risk of developing a particular disease, condition, or symptom, or to a subject reporting one or more of the physiological symptoms of a disease, even though the disease, condition, or symptom may not have yet been manifested.
  • the term "effective amount” or “therapeutically effective amount” refers to the amount of an agent that is sufficient to effect beneficial or desired results.
  • the therapeutically effective amount will vary depending upon the subject and disease condition being treated, the weight and age of the subject, the severity of the disease condition, the manner of administration and the like, which can readily be determined by one of ordinary skill in the art.
  • the term also applies to a dose that will provide an image for detection by any one of the imaging methods described herein.
  • the specific dose will vary r oc e o. - . depending on the particular agent chosen, the dosing regimen to be followed, whether it is administered in combination with other compounds, timing of administration, the tissue to be imaged, and the physical delivery system in which it is carried.
  • biologically active and “bioactive,” as used herein, indicate that a composition or compound itself has a biological effect, or that it modifies, causes, promotes, enhances, blocks, or reduces a biological effect, or which limits the production or activity of, reacts with and/or binds to a second molecule that has a biological effect.
  • the second molecule can be, but need not be, endogenous.
  • a "biological effect” can be but is not limited to one that stimulates or causes an immunoreactive response; one that impacts a biological process in a cell, tissue or organism (e.g., in an animal); one that impacts a biological process in a pathogen or parasite; one that generates or causes to be generated a detectable signal; and the like.
  • Biologically active compositions, complexes or compounds may be used in investigative, therapeutic, prophylactic, and/or diagnostic methods and compositions.
  • Biologically active compositions, complexes or compounds act to cause or stimulate a desired effect upon a cell, tissue, organ or organism (e.g., an animal).
  • desired effects include modulating, inhibiting or enhancing gene expression in a cell, tissue, organ, or organism; preventing, treating or curing a disease or condition in an animal suffering therefrom; limiting the growth of or killing a pathogen in an animal infected thereby; augmenting the phenotype or genotype of an animal; stimulating a prophylactic immunoreactive response in an animal; and diagnosing a disease or disorder in an animal.
  • enhancing agent and "contrast agent” as used herein refer to at least one of an exogenous gas, liquid, mixture, solution, chemical, or material that enhances the disruptive cavitational bioeffects of an ultrasound wave on tissue.
  • an enhancing agent is an enhancing solution.
  • the enhancing solution contains exogenous gaseous bodies, for example, microbubbles.
  • siRNA short interfering RNA
  • siRNA siRNA induced silencing complex
  • RISC RNA induced silencing complex
  • siRNA molecule may be completely or partially complementary to the gene or genes whose expression in reduced, and silencing may be effected with or without cleavage of an mRNA transcript.
  • sonoporation is applied to facilitate delivery of siRNA into a target tissue or organ in the human body by permeabilizing cell membranes.
  • siRNA molecules There is no requirement of nuclear entry of siRNA molecules in order for them to exhibit therapeutic effects as the acting site of siRNA molecules is in cytoplasm. r oc e o. - .
  • the invention provides methods, devices and apparatuses for providing electrical impedance tomography (EIT) with ultrasound for cell sonoporation or tissue ablation.
  • EIT electrical impedance tomography
  • Various aspects of the invention described herein may be applied to any of the particular applications set forth below or for any other types of cell/tissue permeabilization or ablation.
  • the invention may be applied as a standalone tool or as part of an integrated platform used to perform and/or control cell/tissue sonoporation or ablation. It shall be understood that different aspects of the invention can be appreciated individually, collectively or in combination with each other.
  • FIG. 1 shows an example of a system utilizing EIT and an ultrasound transmitting system, in accordance with an embodiment of the invention.
  • An EIT-ultrasound system may include an EIT system 100 connected to a plurality of EIT probes 102, a control system 104 in communication with the EIT system, an ultrasound transmitting system 106 in communication with the control system, and ultrasonic transducer 108 which is communication with the ultrasound transmitting system, wherein the ultrasonic transducer may emit a focused ultrasound beam 110 and whose direction and focal depth may be controlled electronically or mechanically by a motor controlled scanning system 112.
  • EIT may be used to monitor and control ultrasonic applications, such as sonoporation and/or tissue ablation. Both EIT and such ultrasonic applications can be realized without open surgery. Thus, it may be feasible that non- invasive or minimally- invasive therapies may be provided by using a combination of EIT and ultrasonic transmissions.
  • the control system may receive information from the EIT system and/or the ultrasound system and may send instructions to the EIT system and/or ultrasound system. Such instructions may be provided based on the information received.
  • the control system may communicate with the EIT system and/or ultrasound system in real-time, periodically, or only at certain specified points in time.
  • the EIT-ultrasound system may allow the EIT system to guide the ultrasonic transmission and permit targeted delivery of the ultrasound signals.
  • One or more portions of the system may receive power from a power source.
  • a power source may be provided for the EIT-ultrasound system.
  • power sources may be provided to different components (e.g., EIT system, ultrasound transmitting system, ultrasonic transducer, control system) or sub-components of the system.
  • the power source may be from a grid utility, a local energy generator, an energy storage source (e.g., battery, capacitor, ultracapacitor, fuel cell), or may have any other characteristics or features as described elsewhere herein.
  • EIT is an imaging technique in which an image of the conductivity or permittivity of part of the body is constructed by injecting small electrical signals (potentials or currents) through surface or embedded source electrodes to the said body part, and measuring the electrical signals (potentials or currents) at the corresponding receiving electrodes.
  • EIT may be used to generate an impedance profile for a part of the body, which may or may not be used to generate an image.
  • An EIT system may be in communication with one, two, three, four, five, six, seven, eight, nine, ten or more, twelve or more, fifteen or more, twenty or more, thirty or more, or more EIT probes.
  • EIT probes may be distributed across different points of a body mass. For example, a plurality of EIT probes may be distributed across an organ, or a plurality of organs or tissues. Such EIT probes may assist with the generation of an impedance profile between the EIT probes and the organ, the section of the organ, or any bodily mass of interest. Increasing the number of EIT probes provided may increase the resolution of an impedance map.
  • the EIT probes may be relatively evenly spaced apart, while in other embodiments, they may be clustered together or have any other distribution.
  • the EIT probes may preferably attached to or be placed in close proximity to the skin of a subject. Alternatively, it may be applied directly to tissue, or other portions of body mass.
  • An EIT probe may include one, two, three, four, or more electrodes. Such electrodes may include current-providing electrodes and/or sensing electrodes.
  • each EIT probe may include at least one current-providing electrode and at least one sensing electrode. In some instances, each EIT probe may include at least one current-providing electrode pair, and at least one sensing electrode.
  • one or more four-electrode EIT probe may be provided, which may include one current-providing electrode pair and one sensing electrode pair.
  • an EIT probe may include only at least one current-providing electrode or may include only at least one sensor. Small alternating currents may be applied to some or all of the electrodes. The resulting electrical potentials or other electrical characteristics may be measured. This may be repeated for numerous different configurations of applied currents.
  • An electrode may include any conductive material, preferably a metal, most preferably a non- corrosive metal that is used to establish the flow of electrical current from that electrode to another electrode. Electrodes are made of a variety of different electrically conductive materials and may be alloys or pure metals including but not limited to copper, gold, platinum, steel, silver, silver chloride, and alloys thereof. Further, the electrode may be comprised of a non-metal that is electrically conductive such as a silicon-based material used in connection with microcircuits. Typical electrodes may include rod-shaped, flat plate-shaped or hollow needle-shaped structures. Electrodes may be used to deliver electrical current continuously or to deliver pulses.
  • the electrodes may be very application- specific and be comprised of parallel stainless steel plates, implanted wires, needle pairs and needle arrays. Those skilled in the art will design specific electrodes that are particularly useful in EIT in accordance with the present invention. r oc e o. - .
  • an electric current may be sent between a first point and a second point across a section of body mass.
  • the first point and second point may be provided at a first electrode and second electrode respectively.
  • the first and second electrodes may be provided at the same EIT probe or at different EIT probes.
  • the first electrode and the second electrode may be located with the section of body mass therebetween.
  • one, two, or more electrodes may be used by an EIT probe to provide a current across the section of the body mass.
  • a current may be provided between any number of additional points, which may include a third point and/or fourth point corresponding to a third electrode and/or fourth electrode respectively. Additionally, five, six, seven, eight, nine, ten, or more points corresponding to a fifth electrode, sixth, electrode, seventh electrode, eighth electrode, ninth electrode, or tenth electrode may be provided.
  • the electrical properties of the current provided to an electrode may be controlled.
  • the rate of current flow (current amplitude), current duration, continuous vs. pulsed currents, number of electric pulses, shape of electrical pulses, voltage, and/or any other electrical properties may be controlled or defined.
  • the electrical properties of the current provided may be slow varying or remain substantially the same, while in other embodiments, they may change.
  • the rate of current flow may fall within 0.05 mA to 100 mA.
  • a current flow of about 0.05mA, 0.1 mA, 0.2mA, 0.5 mA, 1 mA, 1.5 mA, 2 mA, 2.5 mA, 3 mA, 4 mA, 5 mA, 7 mA, 10 mA, 15 mA, 20 mA, 30 mA, 50 mA, 75 mA, or 100 mA may be provided.
  • the current flow may remain substantially constant or may vary.
  • the current may be continuous or pulsed. If the current is pulsed, the pulse lengths and/or characteristics may remain substantially the same, or may vary.
  • the current duration may vary, whether the current duration before a continuous current, a pulse, or multiple pulses. For example, current may be provided for periods of time on the order of nanoseconds, microseconds, milliseconds, seconds, or minutes. If a current is pulsed, it may have any duty cycle, which may include, but are not limited to, duty cycles of about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%. If a current is continuous, it may be a sine wave, with a frequency of 100Hz to 100KHz.
  • the electrical properties may be defined by a user, or may be automatically determined from a program based on parameters provided and/or measurements taken.
  • EIT may monitor and analyze differences in bio-electrical attributes of a sample being monitored.
  • the EIT technology can be used in connection with the present invention by creating an impedance profile and using that profile to adjust ultrasound signals to the sample to obtain desired results.
  • the profile may or may not be used to generate an image.
  • the EIT profile may be created by injecting electrical currents into the body mass and measuring the resulting voltages through an electrode array or any other sensing mechanism. This makes it possible to produce an impedance profile from the known current inputs and the measured input voltage data using a reconstruction algorithm.
  • any other electrical parameters may be directly measured, or may be derived from data measured. Inn some r oc e o. - . instances, the electrical measurements may be used to adjust electrical current provided for EIT measurement itself, which may permit feedback to create an optimized or improved impedance map and/or image.
  • EIT technology may be particularly desirable in connection with the present invention as applied to tissue in that EIT imaging provides a map of electrical impedances.
  • the map of electrical impedances essentially allows the user to determine when cell permeabilization is beginning. When permeabilization begins the user can stabilize the amount of current being applied and thereby avoid applying so much current as to result in irreversible damage to cells, if so desired. Such stabilization may also occur automatically in response to the impedance map.
  • the EIT technology makes it possible for the region of tissue undergoing ultrasonic cell permeabilization to determine changes in equivalent electrical impedance of the cells within tissue being monitored.
  • one, two, or more electrodes or other sensors may be used to sense an electrical property of the section of body mass.
  • sensed electrical properties may include voltage, impedance, resistance, conductance, or inductance.
  • one or more senor may be used to sense another property of the body mass, which may include, but is not limited to mechanical properties, thermal properties, chemical properties, magnetic properties, optical properties, acoustical properties, radiological properties, or environmental properties.
  • One example of a property that may be sensed is temperature.
  • the sensor may be provided at an EIT probe or separate from the EIT probe. In some instances, it may or may not be provided at the same EIT probe that may include a current-providing electrode. In some embodiments, a sensor may be a current-providing or current- receiving electrode.
  • a first probe and a second probe may be positioned at a first point and a second point on a body mass respectively.
  • One or more electrical parameter may be measured between the first and second points.
  • the measuring of the one or more electrical parameter may then be analyzed in order to determine a character of the body mass. This may include the characteristic of one or more cells of the body mass.
  • EIT may be used more specifically a characteristic of the membrane of the one or more cells.
  • EIT may be used to determine the characteristics of tissue within a body mass to locate a tumor, or other cells to ablate.
  • FIG. 2A shows an example of cells that have not undergone successful sonoporation or other forms of cell permeabilization.
  • cells that have not undergone permeabilization may function as barriers to electrical current, or may have a greater impedance value for electrical current.
  • the electrical current may primarily travel between and/or around cells.
  • FIG. 2B shows an example of cells that have undergone successful sonoporation or other forms of cell permeabilization.
  • Recent studies have demonstrated that during successful sonoporation, the electrical conductivity of the membranes of the treated cells undergo considerable increases due to increase in the permeability of the cell membranes. See, e.g., Cheri X. Deng, Fred Sieling, Hua Pan r oc e o. - . and Jianmin Cui. Ultrasound-induced cell membrane porosity. Ultrasound in Medicine & Biology, Volume 30, Issue 4, April 2004, Pages 519-526, which is hereby incorporated by reference in its entirety. For example, when cell membranes are permeabilized, electrical current may flow through the cells more easily, which may lead to lower impedance levels.
  • Cell permeability of targeted tissue is believed to increase during sonoporation as the result of transient and reversible membrane permeabilization. This allows the entry of foreign macromolecules, such as short interfering RNA (siRNA) or others to be discussed elsewhere herein, which otherwise can not cross normal cell membranes. Similarly, the permeabilized cell membranes also allow ions to pass through, and therefore exhibit significantly increased electrical conductivity. The increase in electrical conductivity can be measured by EIT and be used to assess the degree of membrane permeability. The effectiveness of cellular uptake of relatively small siRNA molecules (compared with DNA plasmids and proteins) may be directly related to the degree of membrane permeability. As such, electrical impedance measurement is considered to be a valid means to evaluate the effectiveness of foreign substance delivery mediated by sonoporation.
  • siRNA short interfering RNA
  • Excessive membrane permeabilization can be irreversible, and may lead to rapid cell and tissue damage. Irreversible permeabilization may be desirable in certain situations targeting cell/tissue ablation, which may be achieved using high intensity ultrasound beam, such as HIFU. Fortunately, an undesirable damage mode can also be detected by electrical impedance measurement. This is because irreversible membrane permeabilization or cell damage due to physical stimulation (e.g., electroporation, sonoporation, etc.) is not transient therefore results in permanent increase of electrical conductivity in the damaged area of the treated tissue/organ (see, e.g., FIG. 8A). In contrast, tissue that undergoes reversible membrane permeabilization normally can recover after withdraw of stimulation. This is evidenced by the gradual increase of cell electrical impedance due to membrane recovery after withdrawal of stimulation (see, e.g., FIG. 8A).
  • physical stimulation e.g., electroporation, sonoporation, etc.
  • EIT can determine whether cell membranes have been successfully sonoporated or not. EIT can also determine the degree to which the cell membranes for cells in a tissue have been sonoporated. Moreover, in addition to the degree of cell membrane permeability changes caused by sonoporation, EIT can also be able to map the regions where cells are successfully sonoporated, and where cells are not.
  • EIT effectiveness of sonoporation on specific tissues or organs of the human body
  • Such control may be implemented by using EIT to measure the locations and degrees of cell sonoporation in a body mass, and further to provide feedback to control the dosage and targeting area of ultrasonic energy, thereby achieving a desirable level of sonoporation at desirable locations.
  • EIT may provide r oc e o. - . feedback that allows the ultrasonic energy location to be adjusted accordingly, thereby permitting the sonoporation of cells at various locations and/or scanning over a wide area.
  • the body mass or section of body mass that can be used in connection with the subject methods includes but is not limited to, transformed and non-transformed tissues, including muscle, vascular, endothelial, connective, retinal, lymphatic, adipose, epithelial, neural, and hematopoietic tissues; organs and/or portions thereof, including kidney, liver, lung, heart, spleen, pancreas, prostate, brain, and islets of langerhans; glands such as thyroid, parathyroid, paracrine glands, adrenal, endocrine glands, exocrine glands, and the like; and transformed or tumor tissues, e.g.
  • transformed and non-transformed tissues including muscle, vascular, endothelial, connective, retinal, lymphatic, adipose, epithelial, neural, and hematopoietic tissues
  • organs and/or portions thereof including kidney, liver, lung, heart, spleen, pancreas, prostate, brain, and islets of
  • solid tumors such as carcinomas, including cervical carcinoma, hepatocellular carcinoma, colorectal carcinoma, basal cell carcinoma, renal cell carcinoma, prostate carcinoma, small cell lung carcinoma, breast carcinoma; sarcomas, endotheliomas; mesotheliomas; including acute and chronic Hepatitis; including viral hepatitis such as Hepatitis A, Hepatitis B and Hepatitis C; including alcoholic liver, liver fibrosis and liver cirrhosis; including fibrotic and cirrhotic organs/tissues; and other joined or discreet groups of cells and cell types.
  • Cells in a body mass may be of the same or of a number of different types. These cells may preferably be organized to carry out a specific function.
  • a body mass may be present within a living organism as well as removed and may refer to in vivo or in vitro situations. Further, the body mass may be from any organism including plants and animals or a tissue developed using genetic engineering and thus be from an artificial source. In one embodiment a body mass includes a plurality of cells present within a distinct area of a human.
  • the section of body mass may be provided within a living subject, a partial or dead subject, or may be in a controlled environment. For example, the section of body mass may be in vivo, ex vivo, or in vitro.
  • a subject may be a patient, or may be a clinical or pre-clinical test subject, or any other human, mammal, or any other animal that may be interacting with the EIT-ultrasound system.
  • EIT may be proposed as a medical imaging means for applications including lung function monitoring, skin and breast cancer detection, etc.
  • EIT may be used to measure effectiveness of cell membrane permeabilization, under which, cell membrane impedance is compromised due to sonoporation- mediated reversible increase of cell membrane permeability.
  • EIT- ultrasound system EIT may be used with ultrasound.
  • the EIT may be used to assist with guiding the ultrasound.
  • the ultrasound may be used for sonoporation (cell permeabilization using ultrasound signals) or tissue ablation (i.e., leading to irreversible cell permeabilization, cell rupture).
  • cell permeability of damaged cell/tissue increases substantially, conceptually such change can also be detected with EIT; the cell/tissue impedance measurement can then indicate whether a cell, part of the whole tissue is successfully ablated, furthermore, such measurement can be used to guide adjustment of an ultrasound beam property (such as intensity, cycle, r oc e o. - . treatment time, treatment location, etc.) to achieve successful cell/tissue ablation at a desirable location of a tissue/organ.
  • an ultrasound beam property such as intensity, cycle, r oc e o. - . treatment time, treatment location, etc.
  • FIG. 3 shows an example of an EIT system for use within the invention.
  • the EIT system may be used to carry out a process on a body mass (e.g., tissue 71).
  • a current source 72 may be controlled by a signal generator 73 and may be used to drive an electrical current into the body mass sample 71 through a pair of computer controlled multiplexers 74 and 75 which may lead to a differential amplifier 76 and demodulator 77.
  • the measured signals may be compared to an original signal in order to record amplitude and phase data for later profile construction.
  • the controlling computer 78 may typically choose which pair of electrodes will inject current while reading the remaining electrode voltages. There may be a number of different hardware configurations which can be utilized in connection with the present invention.
  • the EIT system as shown in FIG. 3 may be generally referred to as a serial system because of its single current source and measurement amplifier. Varying degrees of parallelism (multiple current sources and voltage measuring amplifiers) may be utilized in other embodiments of the invention, thereby increasing the flexibility and speed of the current injection system.
  • any other type of power source may be used instead of the current source.
  • a voltage source may be used.
  • a current source, voltage source, or any other type of power source may be used interchangeably herein to describe any means for providing electrical power, current or voltage thereby creating a flow of electrical current between the electrodes.
  • the device may preferably be capable of providing for a controlled mode and amplitude and may provide constant DC current or AC current, provide pulse voltage or continuous voltage.
  • Preferred devices are capable of exponentially decaying voltage, ramp voltage, ramped current, or any other combination.
  • a power supply may be used in combination with a chip of the type used in connection with microprocessors and provide for high-speed power amplification in connection with a conventional wall circuit providing alternating current at 110 volts.
  • the pulse shape may be generated by a microprocessor device such as a Toshiba laptop running on a Lab View program with output fed into a power amplifier.
  • a processor device such as any network device described elsewhere herein, may be used. Any processor or network device may be specially programmed to perform one or more function or step as described herein.
  • a wide range of different commercially-available power supplies can provide the desired function. However, the range may be amplification-specific and can be extended outside the range for any desired application.
  • any number of multiplexers or pairs of multiplexers may be used (e.g., one, two, three, four, five, six, seven, eight, or more multiplexers).
  • the multiplexers may be designed to multiplex signals from various locations on the body mass. For example, if EIT probes are positioned at six different locations on a section of body mass, one or more multiplexers may be used to accommodate six different signals. Any additional signal processing components may be used to adjust and/or maintain the signal being provided to the data acquisition system 78. Any communications r oc e o. - . within the EIT system or between the EIT system and any other system may be provided between a wired connection or a wireless connection.
  • communications may be provided over a network, which may be a local area network, or a wide area network, such as the Internet.
  • a network which may be a local area network, or a wide area network, such as the Internet.
  • the data acquisition system 78 may be provided over a network and need not be in close physical proximity to the rest of the EIT system. Alternatively, the data acquisition system may be in close physical proximity to the rest of the EIT system.
  • EIT systems may be used. Some examples of commercially available EIT systems may include Sheffield Mark 3.5 by Maltron International, Goe MF II by Drager Medical, Viasys Health Care, or a system by Sim-Tecknika. Alternatively, EIT systems may be developed for use within the EIT-ultrasound system. Components, features, characteristics of EIT systems known or later developed in the art may be used in the EIT-ultrasound system.
  • a control system may be in communication with a data acquisition system of the EIT system. Alternatively, the data acquisition of the EIT system may be part of the control system. The control system will be discussed in greater detail elsewhere herein.
  • Reconstruction algorithms may be used in order to take the voltage measured on an outer surface of a region of interest in the body (the injected current data) and information relating to the electrode geometry, and produce an image which represents spatial tissue impedance tissue distribution inside the region of the tissue 71.
  • Static imaging is the production of an absolute impedance distribution. See, e.g., Cook, R. D. et al. ACT3: a high speed, high precision electrical impedance tomography, IEEE, Trans. Biomed. Eng. 41, 713-22 (1994), which is hereby incorporated by reference in its entirety.
  • Differential imaging methods produced distributions based on differences between two data sets. See, Barber, D.
  • an image may or may not be generated.
  • a map of an electrical property such as an impedance map may be generated.
  • the impedance map may include information about electrical properties of the body mass at and/or between various locations.
  • Various algorithms may be used in order to generate an impedance map.
  • an impedance map may include impedance values for body mass at and/or between locations of EIT probes. r oc e o. - .
  • a control system can be placed in communication with an EIT system and/or an ultrasound transmitting system.
  • the control system may separately communicate with the EIT system and the ultrasound transmitting system.
  • the EIT system and the ultrasound system may or may not be in communication with one another.
  • the control system may communicate with only one of the EIT system or the ultrasound transmitting system.
  • the control system may communicate with the EIT system and/or the ultrasound transmitting system via a wired connection or wireless connection. Examples of wireless communications may include, but are not limited to, radio frequency communications, microwave communication, or infrared (IR) communication.
  • IR infrared
  • communications may be provided over a network, which may be a local area network, or a wide area network, such as the Internet.
  • two-way communications may be provided between the control system and the EIT system and/or ultrasound transmitting system.
  • communications may be one way from the control system or to the control system.
  • the control system may include a memory and a processor.
  • the memory may store tangible computer readable medium as previously described.
  • the processor may access the memory and implement steps based on the computer readable medium.
  • the control system may include a user interactive display or device.
  • a user may be able to view data and/or provide data or instructions to the system.
  • a video display screen may be presented to the user.
  • Video displays may include devices upon which information may be displayed in a manner perceptible to a user, such as, for example, a computer monitor, cathode ray tube, liquid crystal display, light emitting diode display, touchpad or touchscreen display, and/or other means known in the art for emitting a visually perceptible output.
  • a display page shown on the video display may comprise well known features of graphical user interface technology, such as, for example, frames, windows, scroll bars, buttons, icons, and hyperlinks, and well known features such as a "point and click" interface. Pointing to and clicking on a graphical user interface button, icon, menu option, or hyperlink also is known as "selecting" the button, option, or hyperlink.
  • Other user interactive components may be included that may permit a user to interact with the display, such as a mouse, keyboard, touchscreen, remote controller, trackball, or stick.
  • a display page according to the invention also may incorporate multimedia features.
  • a user may be anyone interacting with the EIT-ultrasound system. For example, a user may be a lab technician, a medical practitioner, or the subject.
  • the control system may include a network device, such as a computer. Any discussion of a network device, or any specific type of network device may include, but is not limited to, a personal computer, server computer, or laptop computer; personal digital assistants (PDAs) such as a Palm-based device or Windows CE device; phones such as cellular phones or location-aware portable phones (such as GPS); a roaming device, such as a network-connected roaming device; a wireless device such as a wireless email device or other device capable of communicating wireless with a computer network; or any other type of network device that may communicate over a network and handle electronic transactions.
  • the control system may include multiple devices.
  • the control system may include a client-server architecture.
  • network devices may be specially programmed to perform one or more step or calculation or perform any algorithm, as described herein.
  • the control system may receive data from the EIT system and the ultrasound transmitting system, and based on the data provide instructions to the EIT system and the ultrasound transmitting system. This may occur with user interaction and/or automatically without user interaction.
  • the EIT system may provide an impedance map or profile to the control system.
  • An ultrasound transmitting system may provide information about a characteristic of energy being transmitted and/or the positioning of an ultrasonic transducer. Based on this information, the control system may instruct the ultrasound transmitting system to vary and/or maintain one or more characteristic of energy being transmitted and/or the position of the ultrasonic transducer. This will be discussed in greater detail elsewhere herein.
  • the control system may determine that a desired degree of cell permeabilization has been achieved in one section of a body mass, but not in another section of the body mass.
  • the control system may provide instructions to the ultrasound transmitting system to actuate one or more motor to adjust the position of an ultrasonic transducer to oc e o. - . target the section of the body mass where the desired degree of cell permeabilization has not yet occurred.
  • Instructions from the control system may be general (e.g., the desired new orientation of the transducer), and the EIT system and/or ultrasound transmitting system may interpret the instructions from the control system and implement them specifically (e.g., which motors need to be actuated to move the transducer into the desired orientation).
  • specific instructions may be provided from the control system itself. Any distribution or arrangement may be provided between the control system and the EIT and/or ultrasound transmitting system that may allow instructions to be implemented.
  • control system may determine that the electrical signals provided to the EIT probes in electrical communication with a section of body mass are not sufficient to generate a very accurate impedance profile of the body mass.
  • the control system may provide instructions to the EIT system to adjust the current provided to the EIT probes in order to provide an improved impedance profile.
  • An EIT-ultrasound system may include an ultrasound transmitting system.
  • the ultrasound transmitting system may be in communication with an ultrasonic transducer/probe that may provide an ultrasound beam to a section of body mass.
  • an ultrasound beam may be transmitting at an energy intensity sufficient for sonoporation or tissue ablation.
  • Sonoporation may utilize ultrasound or the interaction of ultrasound with contrast agents (commonly stabilized microbubbles) to temporarily permeabilize the cell membrane. Sonoporation may be used as a new nonviral and non-invasive technique for gene transfection and target drug delivery.
  • therapeutic agents mixed with ultrasound contrast agents could be injected locally or systemically, and ultrasound could be coupled and even focused into the defined area to achieve targeted delivery of the therapeutic agents.
  • Therapeutic ultrasound tissue ablation is a non-invasive therapeutic method using high- intensity ultrasound to heat and destroy malignant tissue without causing damage to overlying or surrounding health tissue. With greater than 1000 W/cm 2 of intensity level, the target tissue temperature could be increased to 70-100 0 C within a couple of seconds. As a result, biological effects including coagulative necrosis and structural disruption can be induced in the tissue. Therapeutic ultrasound tissue ablation has been widely used m the treatment of solid tumors such as breast cancer and other diseases. High intensity therapeutic ultrasound may cause cell ablation and/or irreversible cell permeabilization.
  • the same ultrasound transmitting system may be used for both sonoporation and tissue ablation.
  • different ultrasound transmitting systems may be used for sonoporation and tissue ablation.
  • the energy intensity delivered for sonoporation may range from less than 1 W/cm' to about 5 W/cm 2 .
  • the energy delivered for tissue ablation may be range from about 10 W/cm 2 to 10,000 W/cm 2 . r oc e o. - .
  • FIG. 7 shows an example of how cell impedance level may vary over time under different permeabilization conditions.
  • the time ti indicates when an ultrasonic signal is provided to a body mass
  • t 2 indicates when the ultrasound signal is no longer provided
  • t 3 indicates roughly when an recovery may occur (which may vary for different degrees of signal).
  • cell impedance level may drop a little, but then return to normal impedance level or close to the initial impedance level when the ultrasonic energy is no longer provided.
  • FIG. 8A shows a conceptual illustration of electrical impedance changes due to (a) reversible and (b) irreversible membrane permeabilization of cells subjected to transient electrical or ultrasonic stimulation, much like FIG. 7.
  • ⁇ Z P represents the maxima impedance
  • ⁇ Z R indicates degree of membrane recovery from permeabilization
  • Z 0 is the initial impedance.
  • the external stimulation (for electroporation or sonoporation) is shown as being provided as either on or off. In some implementations, the degree of external stimulation may vary.
  • FIG. 8B shows an example of representative patterns of resistance changes during and after cell electroporation.
  • the upper curve (blue) is from primary human mammary epithelial cells (HMEC) underwent reversible electroporation, which routinely produces 90%+ siRNA delivery and 95%+ viability in the cells.
  • HMEC primary human mammary epithelial cells
  • EIT should also be able to map the regions where cells are successfully sonoporated, and where cells are not.
  • effectiveness of sonoporation on specific tissues or organs of the human body could be monitored and controlled as illustrated in FIG. 1 , by using EIT to measure the locations and degrees of cell sonoporation in a tissue or organ, and further to provide feedback to control the dosage and targeting area of ultrasonic energy, so to achieve desirable level of sonoporation at desirable locations.
  • therapeutic ultrasound affects electrical properties of cells, such as cell impedance.
  • Therapeutic ultrasound may cause irreversible cell permeabilization or cell death.
  • Current commercial available ultrasound tissue ablation system may use either magnetic resonance imaging (MRI) or ultrasound imaging system to guide the high intensity focused ultrasound beam to the targeted area.
  • MRI magnetic resonance imaging
  • Imaging techniques such as MRI, may also not provide real-time feedback.
  • MRI typically does not operate in real-time with therapeutic ultrasound because typically image acquisition occurs every six seconds or so using a full scan of k-space.
  • an imaging system such as MRI, may be in used in conjunction with EIT.
  • EIT has been introduced as an inexpensive medical imaging means in breast cancer detection.
  • the electrical impedance of the treated tissue will change significantly because cells in tumor tissue will go through irreversible permeabilization or membrane rupture.
  • the temperature increment in the tissue under tissue ablation treatment will also affect the electrical impedance.
  • the electrical impedance change could be detected by using EIT.
  • EIT may be able to detect such changes in real-time (e.g., without having to wait for a scan of an image). Allowing real-time monitoring of delivery status may advantageously optimize the target state (e.g., drug delivery or cell ablation) without causing or reducing unwanted damage.
  • EIT could be used to guide therapeutic ultrasound beam to the targeted area and the electrical impedance change detected by EIT could be used as the feedback to control the ultrasound output to achieve optimized treatment in the targeted area by avoiding the surrounding health tissue to be overheated.
  • therapeutic ultrasound may allow targeted cell ablation with little or no damage to surrounding cells.
  • EIT should also be able to map the regions where tissues are successfully ablated, which will help to accomplish a complete treatment of the interested area.
  • the EIT guided tissue ablation may be especially suitable in treating breast cancer because it is not necessary to insert EIT probes inside a patient's body, which keeps the EIT controlled tissue ablation treatment non- invasive. This application and other applications are discussed in greater detail elsewhere herein. r oc e o. - .
  • the ultrasound transmitting system could also be combined with EIT which can guide the transmitted ultrasound beam focused to the targeted location or scan through the whole issue or organ.
  • EIT monitoring and ultrasound emission e.g. for sonoporation or tissue ablation
  • the EIT monitoring may advantageously provide feedback on the type of tissue provided, which may assist with determining a setting for the ultrasound emission.
  • the EIT monitoring may or may not provide real-time feedback on the degree of cell permeabilization or ablation achieved.
  • EIT measurements may be taken before or while ultrasonic signals are transmitted.
  • EIT measurements may be taken after ultrasonic signals have been transmitted. EIT measurements may be taken periodically or continuously while ultrasonic signals are being transmitted. The ultrasonic emission may advantageously cause targeted cell permeabilization or ablation without having to contact the cells. Thus, this may provide a controlled, less invasive procedure.
  • the ultrasound transmitting system may be in communication with an ultrasonic transducer/probe, which may direct a focused ultrasonic beam to a desired section of body mass.
  • the transducer may be mounted on a scanning system structure that may vary and/or maintain the position of the ultrasonic transducer.
  • the body mass may be provided within a subject. In other embodiments, the body mass may be provided outside the subject.
  • one ultrasonic transducer/probe may be provided for the ultrasound transmitting system. Alternatively, two, three, four, or more probes may be provided. Each probe may operate with similar ultrasonic emission characteristics, or each probe may be individually controllable and may have different ultrasonic emission characteristics.
  • the scanning system may include one or more actuators that may allow the position of the ultrasonic transducer to be varied.
  • the ultrasonic beam position may be adjusted by varying the height of the transducer, the lateral displacement of the transducer, the forward/backward displacement of the transducer, the distance of the transducer from the body mass, or the orientation of the transducer with respect to any angle or degree of freedom.
  • the actuators may include, but are not limited to, motors, solenoids, linear actuators, pneumatic actuators, hydraulic actuators, electric actuators, piezoelectric actuators, or magnets.
  • the actuators may be in communication with the ultrasound transmitting system and/or the control system.
  • the ultrasound transmitting system and/or the control system may provide instructions to the actuators to control the position of the ultrasonic transducer.
  • one or more actuator may provide information to the ultrasonic transmitting system and/or the control system on the actuator state or position, which may assist with determining the current position of the ultrasonic transducer.
  • any distance may be provided between the ultrasonic transducer and a target area.
  • the ultrasonic transducer may contact the skin of a subject in proximity to the target area.
  • a gap may be provided between the ultrasonic transducer and the subject using acoustic r oc e o. - . coupling media.
  • the gap may be about 1 mm, 2 mm, 3 mm, 4 mm, 5 mm, 7 mm,
  • the ultrasonic transducer may emit an ultrasonic beam.
  • the degree of focus of the ultrasonic beam may be altered.
  • the ultrasonic beam may be focused over a relatively small area. Alternatively, it may be focused to cover a wider area.
  • the ultrasonic beam may focus to an area on the order of about 1 nm , 1 ⁇ m , 0.1 mm , 1 mm , 10 mm , or 1 cm 2 .
  • the focus of the beam may be varied.
  • the beam may penetrate different depths of a body mass, or different locations of the body mass.
  • an ultrasonic transducer may be reoriented to scan through a body mass, and cause cell permeabilization or ablation at desired locations.
  • using ultrasonic emissions may allow a greater area or volume to be covered at a more rapid rate than requiring physical connections, because the ultrasonic emissions may remotely target cells and then move on.
  • the ultrasonic beam may also include varying energy intensities, which may include but are not limited to 0.1 W/cm 2 to 1 MW/cm 2 .
  • the energy intensity delivered for sonoporation or tissue ablation may be greater than 1 W/cm 2 .
  • the energy intensity delivered may fall below 10,000 W/cm 2 .
  • the energy intensity delivered may be about 0.1 W/cm 2 , 0.5 W/cm 2 , 1 W/cm 2 , 1.5 W/cm 2 ,
  • Ultrasonic energy may also be delivered at varying frequencies.
  • the ultrasonic frequency may fall within 100 kHz to 300 MHz.
  • ultrasound emissions may be provided at about 100 kHz, 500 kHz, 750 kHz, 1 MHz, 1.2 MHz, 1.5 MHz, 2MHz, 3 MHz, 5 MHz, 7 MHz, 10 MHz, 12 MHz, 15 MHz, 18 MHz, 20 MHz, 25 MHz, 30 MHz, 40 MHz, 50 MHz, 75 MHz, 100 MHz, 200 MHz, or 300 MHz.
  • the ultrasonic signals may be provided continuously or may be pulsed. If pulsed, they may be provided on any duty cycle. For example, they may be pulsed on about 10%, 20%, 30%, 40%, 50%, 60% 70%, 80%, or 90% duty cycle.
  • the ultrasonic signals may be provided for any duration (e.g., continuous duration or pulse duration). For example, such durations may be provided on the order of nanoseconds, microseconds, milliseconds, seconds, or minutes.
  • the ultrasonic signal may be substantially constant, or properties of the ultrasonic signal may vary over time.
  • the actuators may be in communication with a power source.
  • the power source may be integral to the device or may be external to the device.
  • power sources may include a battery, generator, grid utility, renewable power sources, or any combination thereof.
  • Any commercially available ultrasonic device may be used by the system.
  • a Sonitron 2000 may be used for sonoporation.
  • Ablatherm Robotic HIFU, or Sonoblate 500 may be examples of tissue ablation devices that may be used or incorporated. Characteristics, features, or r oc e o. - . components of commercially available ultrasonic devices may be incorporated within an EIT- ultrasound system.
  • FIG. 4 shows an example of an ultrasound system in accordance with an embodiment of the invention, which may be used for sonoporation or tissue ablation.
  • an ultrasound transmitting system 400 may be in communication with an ultrasound probe 402.
  • the ultrasound transmitting system may control the properties of the ultrasonic signals transmitted by the ultrasound probe.
  • properties include energy intensity, frequency, duration, duty cycle, or other ultrasound properties.
  • the ultrasonic signals from the ultrasound probe 402 may be provided to a sample body mass, such as a tissue sample 404.
  • the ultrasound probe may or may not contact the sample body mass directly (e.g., tissue).
  • the ultrasound probe may or may not contact the skin of the subject in proximity to the sample body mass.
  • the tissue sample may be provided within a tissue holding chamber 406.
  • the tissue sample may be provided within a subject and need not be within a holding chamber.
  • a tissue holding chamber may include an acoustically transparent membrane 408 and an acoustic absorber 410. Such features may assist with avoiding standing wave formation in tissue, which normally not happens in in vivo treatment.
  • One or more electrode pairs 412 may be provided to the sample body mass 404.
  • the electrodes may be used for EIT.
  • the electrodes may be in communication with a current and/or voltage source.
  • an electric current may be provided, and in communication with a voltage-current (V/I) converter 414, which may be in communication with a digital-analog (D/A) converter 416.
  • a voltage sensing electrode may be in communication with a differential amplifier 418 which may be connected to an analog-digital (AIO) converter 419.
  • the D/A converter and the A/D converter may be in communication with an input-output (I/O) interface 420.
  • the I/O interface may also be in communication with the ultrasound transmitting system 400 and a control device 422, such as a computer.
  • FIG. 5 shows an ultrasonic probe 500 applying an ultrasonic signal 502 to a section of body mass 504 via acoustic coupling media 507 in accordance with an embodiment of the invention.
  • One or more electrode 506 may be provided.
  • the electrode may be part of an EIT probe, that may be in communication with an EIT system.
  • a current may be provided between a first point and a second point.
  • the first point and second point may be provided by a first electrode and second electrode.
  • An electrical property between the first point and second point such as voltage, impedance, resistance, conductance, or any other electrical property mentioned elsewhere herein may be measured.
  • the electrical property properties may be measured by a third and fourth electrode, or any other sensors.
  • the section of body mass may be the portion of body mass between the first point and second point.
  • the body mass may or may not be within a subject while said ultrasonic signals are provided. r oc e o. - .
  • FIG. 6 shows an example of an experimental setup for ultrasound mediated siRNA delivery in vitro.
  • the ultrasound probe 600 may be inserted into the siRNA solution 602 and the distance between the tip of the probes and the cells monolayer may be controlled. In one example, the distance may be about 2 ⁇ 3mm. Care was taken to avoid air bubbles being trapped between the ultrasound probe and the cell layer.
  • the siRNA delivery may be performed within a well plate 604. A well plate of any dimensions, and with any number of wells may be used.
  • the well plate may be supported by a supporting frame 606.
  • An acoustic absorber 608 may be placed at the bottom of a water tank 610 to minimize the standing wave effect. The water temperature within the tank may be kept at a desired temperature.
  • the water temperature may be kept at 37°C.
  • therapeutic agents such as small molecule drugs, siRNA and DNA
  • small molecule drugs, siRNA and DNA which are not able or have poor ability to cross normal cell membranes, can enter the cells through the sonoporated cell membranes. See, e.g., K Iwanaga, K Tominaga, K Yamamoto, M Habu, H Maeda, S Akifusa, T Tsujisawa, T Okinaga, J Fukuda and T Nishihara. Local delivery system of cytotoxic agents to tumors by focused sonoporation.
  • a therapeutic agent is mixed with ultrasound contrast agents (such as OPTISON) in 0.85% NaCl based on the optimized ratio.
  • the mixture is then injected locally or systemically into a specific human tissue or organ.
  • the tissue or organ is then sonoporated at optimized setting (such as 1 MHz input frequency with a 50% duty cycle and 0.5-2.0 W/cm 2 output intensity) using an ultrasound transmitting system.
  • this process can be controlled by an EIT system, which measures cell membrane permeability to provide feedback on the effectiveness of sonoporation, and guidance to optimize application of ultrasonic energies to achieve optimal degree of sonoporation at desirable site of the tissue or organ.
  • a drug, compound, or other substance of interest can be administered prior to, during, or following exposure to ultrasonic energy. Administration may be achieved by any technique known in the art, including but not limited to inhalation, topical, oral, buccal, rectal, parenteral, intraperitoneal, intradermal, transdermal, intratracheal, intravascular, intravenous, intra-arterial, intraduodenal, via the jejunum (or ileum or colon), subcutaneously, intramuscular, intraparenteral, via direct injection into a tissue, organ, or cavity of interest, etc.
  • Injection in the targeted region can also include intra-arterial or intravenous injection into blood vessels that feed into the targeted area, intra-muscular, or intra- tissue injection.
  • delivery can be at, near, adjacent, in, or distant from the target body mass.
  • a drug or other substance may be injected into a body mass in close proximity to the desired delivery site. Delivery may also occur via local circulation (e.g., via IV or injection to a vein or artery). Sonoporation may advantageously cause cell permeabilization which may increase uptake of a substance within the cell, thus allowing for lower dosage for effective delivery. Having a lower dosage of drugs may prevent or reduce unwanted side effects, while still allowing the drug to function effectively.
  • pharmaceutical agent or “drug”, as employed herein, refer to any therapeutic or prophylactic agent which may be used in the treatment (including the prevention, diagnosis, alleviation, or cure) of a malady, affliction, disease or injury in a patient.
  • therapeutically useful peptides, polypeptides and polynucleotides may be included within the meaning of the term pharmaceutical or drug, as are various other therapeutically useful organic or inorganic compounds.
  • bacterial endotoxin such as lipopolysaccharide and macrophage activation factor); subunits of bacteria (such as Mycobacteria and Cornebacteria); the synthetic dipeptide N-acetyl-muramyl-L-alanyl-D-isoglutamine; antifungal agents such as ketoconazole, nystatin, griseofulvin, flucytosine (5-fc), miconazole, and amphotericin B; toxins such as ricin; immunosuppressants such as cyclosporins; and antibiotics such as ⁇ -lactam and sulfaphonequeln; hormones such as growth hormone, melanocyte stimulating hormone, estradiol, beclomethasone dipropionate, betamethasone, betamethasone acetate, betamethasone sodium phosphate, betamethasone disodium phosphate, betamethasone sodium phosphate, cortisone acetate, dexamethasone, dexamethas
  • alkaline phosphatase alkaline phosphatase; anti- allergens such as amelexanox; anticoagulation agents such as phenprocoumon and heparin; tissue plasminogen activators (TPA), streptokinase, and urokinase; circulatory drugs such as propranolol; metabolic potentiators such as glutathione; antibiotics such as p- aminosalicyclic acid, isoniazid, capreomycin sulfate cycloserine, ethambutol hydrochloride ethionamide, pyrazinamide, rifampin, streptomycin sulfate dapsone, chloramphenicol, neomycin, ceflacor, cefadroxil, cephalexin, cephadrine erythromycin, clindamycin, lincomycin, amoxicillin, ampicillin, bacampicillin, carbenicillin, dicloxicillin, cyclacillin,
  • a composition comprising a therapeutic agent may further comprise a carrier.
  • the carrier employed may comprise a wide variety of materials.
  • Carriers may include, for example, lipids, polymers, proteins, surfactants, inorganic compounds, metal ions, and the like, alone or in combination r oc e o. - . with water and/or a solvent, or the carrier may simply comprise water and/or a solvent.
  • the lipids, proteins, and polymers may be in liquid form or solid form (such as, for example, the form of particles, fibers, sheets, layers, etc.), or may take the form of a vesicle or other stable, organized form, which may include but is not limited to, such forms commonly referred to as, for example, liposomes, micelles, bubbles, microbubbles, microspheres, lipid-, polymer-, and/or protein-coated bubbles, microbubbles and/or microspheres, microballoons, aerogels, hydrogels, clathrates, hexagonal HII phase structures, and the like.
  • the internal void of the vesicle or other stable form may, for example, be filled with a liquid (including, for example, a gaseous precursor), a gas, a solid, or solute material, or any combination thereof, including, for example, the compound to be delivered, the organic halide, and/or any targeting ligand, as desired.
  • the carrier is provided as an aqueous milieu, such as water, saline (such as phosphate buffered saline), and the like, with or without other carrier components, although other non-aqueous solvents may also be employed, if desired.
  • the carrier may comprise a mixture in the form of an emulsion, suspension, dispersion, solution, and the like.
  • a therapeutic agent as a part of a vesicle to enhance delivery, for example as a contrast agent or enhancing agent.
  • a vesicle refers to an entity which is generally characterized by the presence of one or more walls or membranes which form one or more internal voids. Vesicles may be formulated, for example, from stabilizing compounds, such as a lipid, including the various lipids described herein, a polymer, including the various polymers described herein, or a protein, including the various proteins described herein, as well as using other materials that will be readily apparent to one skilled in the art.
  • lipids, polymers, proteins, surfactants, inorganic compounds, and/or other compounds may be natural, synthetic or semi-synthetic.
  • Preferred vesicles are those which comprise walls or membranes formulated from lipids. The walls or membranes may be concentric or otherwise.
  • the stabilizing compounds may be in the form of a monolayer or bilayer, and the mono- or bilayer stabilizing compounds may be used to form one or more mono- or bilayers. In the case of more than one mono- or bilayer, the mono- or bilayers may be concentric, if desired.
  • Stabilizing compounds may be used to form unilamellar vesicles (comprised of one monolayer or bilayer), oligolamellar vesicles (comprised of about two or about three monolayers or bilayers) or multilamellar vesicles (comprised of more than about three monolayers or bilayers).
  • the walls or membranes of vesicles prepared from lipids, polymers or proteins may be substantially solid (uniform), or they may be porous or semi-porous.
  • the vesicles described herein include such entities commonly referred to as, for example, liposomes, micelles, bubbles, microbubbles, microspheres, lipid-, protein- and/or polymer-coated bubbles, microbubbles and/or microspheres, microballoons, microcapsules, aerogels, clathrate bound vesicles, hexagonal H II phase structures, and the like.
  • the vesicles may also comprise a targeting ligand, if desired. r oc e o. - .
  • tissues of interest include, but are not limited to, transformed and non-transformed tissues, including muscle, vascular, endothelial, connective, retinal, lymphatic, adipose, epithelial, neural, and hematopoietic tissues; organs and/or portions therof, including kidney, liver, lung, heart, spleen, pancreas, prostate, brain, and islets of langerhans; glands such as thyroid, parathyroid, paracrine glands, adrenal, endocrine glands, exocrine glands, and the like; and transformed or tumor tissues, e.g. solid tumors such as carcinomas, including cervical carcinoma, colorectal carcinoma, basal cell carcinoma, renal cell carcinoma, prostate carcinoma, small cell lung carcinoma, breast carcinoma; sarcomas, endotheliomas; mesotheliomas; etc.
  • solid tumors such as carcinomas, including cervical carcinoma, colorectal carcinoma, basal cell carcinoma, renal cell carcinoma, prostate carcinoma, small cell lung carcinoma, breast carcinoma
  • the compounds delivered to the targeted tissue may be any biologically compatible exogenous agent, particularly agents that are not freely diffused into cells or tissues under normal physiological conditions due to size, hydrophobicity, etc. Included are imaging agents, pharmacologically active drugs, genetically active molecules, etc. Larger compounds, ranging from about 10 nm to 200 nm or larger, include liposomes, e.g. anionic, cationic or neutral liposomes, which may encapsulate a variety of therapeutic agents; proteins, e.g. antibodies, cytokines, hormones, growth factors, etc.; nucleic acids, e.g.
  • treatment agents may include macromolecules that decrease the cell's ability to repair itself or that cause cell death in the tissue that is treated. Examples of such types of treatment agents include lidocaine, and non-ablative heating of cells to be treated.
  • treatment agents include, local anesthetics such as marcaine, vasoconstrictive agents such as epinephrine, hypotonic saline, potassium, agitated saline, microbubbles, commercially available ultrasound contrast agents, microspheres, adipocytes, fat, autologous tissues (e.g. lysed fat cells to produce clean adipocytes to form a tissue graft to minimize hostile response from the body), PLLA, hydroxyappetite. Treatment agents may be delivered prior to, during or following the treatment of the present invention.
  • Compounds of interest include chemotherapeutic agents for neoplastic tissues, anti- inflammatory agents for ischemic or inflamed tissues, hormones or hormone antagonists for endocrine tissues, ion channel modifiers for cardiovascular or other tissues, and neuroactive agents for the central nervous system.
  • Exemplary of pharmaceutical agents suitable for this invention are those described in The Pharmacological Basis of Therapeutics, Goodman and Gilman, McGraw-Hill, New York, New York, (1993) under the sections: Drugs Acting at Synaptic and Neuroeffector Junctional Sites; Drugs Acting on the Central Nervous System; Autacoids: Drug Therapy of Inflammation; Water, Salts and Ions; Drugs Affecting Renal Function and Electrolyte Metabolism; Cardiovascular Drugs; Drugs Affecting Gastrointestinal Function; Drugs Affecting Uterine Motility; Chemotherapy of Parasitic Infections; Chemotherapy of Microbial Diseases; Chemotherapy of Neoplastic Diseases; Drugs Used r oc e o.
  • nucleic acids that correct genetic deficiencies can be introduced into a targeted tissue, usually a solid tissue, e.g. pancreatic cells for the treatment of diabetes, liver cells to treat hepatic deficiencies, etc. Also of interest is the delivery of nucleic acids to accomplish genetic immunization. Genetic immunization involves delivery of a nucleic acid to cells for expression of the encoded immunogen within the target tissue.
  • An immune response against the immunogen is mounted in the animal, resulting in development of humoral and/or cellular immunity.
  • Administration of nucleic acids according to the method of the invention which results in genetic alteration of cells and induction of an immune response can be used to elicit protective immunity.
  • the method of the invention is also useful in a variety of other settings in which genetic alteration of tissue is desirable.
  • the method of the invention can be used to introduce exogenous coding sequences into solid tumors, where the encoded gene product provides for recruitment of immune cells, induces apoptosis, inhibits angiogenesis, etc. in the tumor.
  • transgenic animals can be created by transfection of targeted tissues with a nucleic acid of interest.
  • the nucleic acid materials for delivery to targeted tissue can comprise a nucleic acid of interest that encodes a gene product for which expression is desired, and a promoter for expression of the gene product.
  • Nucleic acids of interest include, but are not limited to, any DNA, RNA or analog thereof that encodes a polypeptide or other gene product that is desirable for expression in tissue of a subject.
  • the gene product can include a polypeptide, an anti-sense mRNA, structural RNA, snoRNA, snRNA, shRNA, siRNA, non-coding RNA, or other gene product that is desirably expressed.
  • the nucleic acid delivered to the tissue in vivo can take any number of forms.
  • the nucleic acid can be introduced as a linear or circular molecule (e.g., a circular plasmid or other construct).
  • a linear or circular molecule e.g., a circular plasmid or other construct.
  • the optimal quantity and spacing of individual dosages of nucleic acids will be determined by the precise form and components of the nucleic acid formulation to be delivered, the site of administration, the use to which the method is applied (e.g., immunization, treatment of a condition, production of transgenic animals, etc.), and the particular subject to which the nucleic acid formulation is to be delivered, and that such optimums can be determined by conventional techniques.
  • the optimal dosing regimen i.e., the number of doses of nucleic acids
  • a dosing regimen will involve administration of the selected nucleic acid formulation at least once, and may be performed multiple times over a period of days or weeks.
  • the amount of DNA to accomplish expression of a desired gene product at an effective level e.g. a level effective to elicit an immune response, to alleviate a symptom of a condition or disease, etc.
  • an effective level e.g. a level effective to elicit an immune response, to alleviate a symptom of a condition or disease, etc.
  • the amount of DNA administered is an amount sufficient to provide for transformation of a number of cells that in turn provides for a level of gene product expression from the introduced DNA to provide for a desired effect.
  • Dosages are routinely determined in the art, and can be extrapolated from the amounts of DNA effective in an animal mode (e.g., a rodent (mouse or rat) or other mammalian animal model), in which factors such as the efficiency of transformation and the levels of gene product expression achieved can be readily assessed and extrapolated to other vertebrate subjects.
  • an animal mode e.g., a rodent (mouse or rat) or other mammalian animal model
  • the nucleic acids of the invention encode a biologically active polypeptide, such as an immunity-conferring polypeptide, e.g. for genetic immunization, or a therapeutic polypeptide, e.g. for amelioration of a symptom associated with a polypeptide deficiency, or for reduction of a tumor.
  • a polypeptide is understood to be any translation product of a nucleic acid regardless of size and glycosylation.
  • the gene product can be any gene product that exhibits a desired biological activity, e.g.
  • Immunity-conferring polypeptides include those polypeptides that comprise an epitope that upon exposure to the immune system of a vertebrate (generally, a mammal), either alone or in the presence of a molecule that facilitates immune response induction (known in the immunology art as a carrier molecule), can act as an endogenous immunogen to provoke a humoral immune response, a cellular immune response, or both.
  • nucleic acid construct having a eukaryotic promoter operably linked to a DNA of interest can be used in the invention.
  • a bacterial plasmid, viral construct, or other DNA construct can be genetically engineered to provide a recombinant DNA molecule having a sequence encoding the desired gene product.
  • the present invention provides a method and apparatus for treating subcutaneous tissue.
  • the present invention includes an apparatus for treating soft tissue.
  • the present invention includes a method for treating tissue.
  • the present invention further includes a method and apparatus for treating a subcutaneous fat layer including fat cells and septae.
  • the present invention further includes a method and apparatus for treating cellulite.
  • the present invention may be useful for a temporary reduction in the appearance of cellulite or the permanent reduction of cellulite.
  • the invention may also be used as an adjunct to liposuction.
  • the invention further provides for a subcutaneous infusion and ultrasonic dispersion of fluid to temporarily improve the appearance of cellulite.
  • the present invention is directed to methods and apparatus for targeting and disrupting subcutaneous structures, such as collagen, connective tissue, adipose tissue (fat r oc e o. - . cells) and the like (collectively included in "target tissue" or "subcutaneous structures") in order to improve the aesthetic appearance of the targeted region and/or remove, disrupt, decrease the appearance of, or otherwise ameliorate a skin irregularity.
  • subcutaneous structures such as collagen, connective tissue, adipose tissue (fat r oc e o. - . cells) and the like
  • Targeted regions may consist of any surface or contour of the human form that it is desirable to enhance, including the face, chin, neck, chest, breasts, arms, torso, abdominal region (including pelvic region), thighs, buttocks, knees and legs.
  • the target tissue may include the connective tissue or septae of the region, or the underlying tissues that may exacerbate the unwanted body contour, such as subdermal and deeper fat deposits or layers.
  • Skin irregularities refer to conditions that decrease a person's satisfaction with their outward appearance, such as cellulite, scarring, or fat deposits or excess fat in certain regions, such as neck, chin, breasts, hips, buttocks, abdomen, arms and the like.
  • Microbubbles (either endogenous or exogenous), being compressible, alternately contract and expand in an ultrasound field. These expansions and contractions may be generally equal and symmetrical at lower ultrasound pressures. This behavior is referred to by some skilled in the art as moderately oscillating. As the ultrasound driving pressure increases, more complex phenomenon occurs, for example, with bubble expansion larger than contraction. Furthermore, there may be relatively slow expansion followed by rapid collapse. This behavior is referred to by some as strongly collapsing. It is associated with the production of harmonic signals. The transition from the moderately oscillating to the strongly collapsing state may be abrupt, wherein the microbubble implodes and releases energy to tissue in the proximity of the microbubble. The energy released by bubble implosion, when bubbles are exposed to ultrasound, is one factor producing observed subcutaneous cavitational bioeffects.
  • Microbubbles may be encapsulated (in a gel type coating) or free (created by agitating a solution such as saline). Microbubbles are also sold commercially as echocardiographic contrast agents, several types of which are described in more detail below.
  • the outer shell and gas may be chosen to prolong the life of the microbubbles in blood or tissue. Another way to prolong the life of a gaseous body in tissue is to increase the viscosity of the carrier solution. For example, a solution of sodium hyaluronate will maintain gaseous bodies in solution much longer than a solution of saline.
  • microbubbles may be generated in solution in the apparatus of the present invention or in other ways known in the art.
  • microbubbles may be generated as disclosed in U.S. Ser. No. 10/798,876 filed Mar. 11, 2004 and entitled "APPARATUS, SYSTEM AND METHOD FOR GENERATING BUBBLES ON DEMAND,” the entirety of which is included herein by reference.
  • Gas bodies may be created in a solution by agitating the solution in the presence of a gas.
  • the simplest microbubbles may be formed by agitating a solution and room air back and forth between two syringes connected together, for example by a tube or stopcock.
  • the solution to be injected in the present invention may include microbubbles.
  • the solution to be injected in the present invention may not include microbubbles.
  • the solution may be a tumescent solution.
  • the solution may be a saline solution.
  • the solution may be a hypotonic solution.
  • the solution may be a hypotonic tumescent solution. In yet another embodiment, the solution may be a hypotonic saline solution.
  • gases other than room air may be used to create the microbubbles.
  • the gas bodies or microbubbles may include various gases, for example, oxygen, carbon dioxide, nitrogen, and/or other suitable gases.
  • Free gas bubbles represent the simplest form of ultrasound contrast media.
  • the bubbles may pre-exist in solution, or they may be created via cavitation during or following injection.
  • Intravascular injection of physiological saline including gas bubbles for example, room air gas bubbles, has been used as a contrast medium in echocardiography since the late nineteen sixties.
  • the utility of free air gas bubbles is highly limited in intravascular diagnostic imaging due to the rapid absorption of room air or oxygen in blood. These bubbles are also too large to pass the pulmonary vasculature.
  • gas bubbles larger than 10 pm may transiently obstruct the capillaries and act as gas emboli.
  • Commercial ultrasound contrast media having various stabilizing coatings or shells have been developed to produce encapsulated gas microbubble contrast media.
  • room air bubbles may be used in the solution because they may be absorbed less rapidly in fat than in blood.
  • the present invention it is also possible to exploit cavitational bioeffects for the purpose of disrupting tissue and tissue ablation without directly heating tissue with ultrasound.
  • the present invention may advantageously introduce exogenous microbubbles to the target tissue, and then use low power ultrasonic energy to cavitate the microbubbles and destroy the target tissue by subcutaneous cavitational bioeffects without the generation of enough heat for direct thermal injury to the tissue being treated.
  • Microbubbles infiltrated into the tissue by way of direct injection will also serve as a nidus for cavitation and tissue disruption.
  • the present invention makes use of the microbubble cavitational bioeffects to destroy subcutaneous tissues without significant thermal effects.
  • the subcutaneous cavitational bioeffects produced by the present invention are advantageous for the disruption of superficial and/or deep fat and/or septae, for example, for the treatment of cellulite and focal fat deposits.
  • Cavitational effects in a more superficial tissue layer may shield a deeper tissue layer from the full power and intensity of an ultrasound wave applied first to the skin.
  • the shielding effect of superficial cavitation may result in insufficient acoustic wave power to simultaneously cavitate the deeper tissue layer or may result in inconsistent tissue disruption in the deeper layers.
  • treatment at various subcutaneous tissue depths is performed in stages. Each injection may be followed by an application of acoustic waves to the tissue to be treated.
  • the acoustic waves applied are low acoustic pressure ultrasound waves.
  • the acoustic waves applied are in the power range of diagnostic ultrasound. For example, r oc e o. - .
  • each subsequent stage of injection is performed at a depth about 0.5 mm to 2.0 cm more superficial than the previous stage of injection. In one embodiment, each subsequent stage of injection is performed at a depth about 0.5 mm more superficial than the previous stage of injection. In another embodiment, each subsequent stage of injection is performed at a depth about 1.0 mm more superficial than the previous stage of injection.
  • each subsequent stage of injection is performed at a depth about 2 mm more superficial than the previous stage of injection. In another embodiment, each subsequent stage of injection is performed at a depth about 5 mm more superficial than the previous stage of injection. In yet another embodiment, each subsequent stage of injection is performed at a depth about 1.0 cm more superficial than the previous stage of injection. In yet one further embodiment, each subsequent stage of injection is performed at a depth about 1.5 cm more superficial than the previous stage of injection. In one further embodiment, each subsequent stage of injection is performed at a depth about 2.0 cm more superficial than the previous stage of injection. In yet one other embodiment, infiltrating the subcutaneous tissue is performed in stages at depths of about 30 mm, about 25 mm, and about 20 mm.
  • infiltrating the subcutaneous tissue is performed in stages at depths of about 15 mm, about 10 mm, about 5 mm and about 2 mm.
  • one series of ultrasound waves may be applied to the tissue after all depths have been injected, rather than the ultrasound waves being applied between injections.
  • the tissue to be treated may be injected between the dermal layer and the deep fat layer. In another embodiment, the tissue to be treated may be injected between the superficial fat layer and the muscle layer. In yet one other embodiment, the tissue to be treated may be injected between the dermal layer and the muscle layer. In one embodiment, the tissue to be treated may be injected at depths of about 2 mm to 4.0 cm. In one embodiment, the tissue to be treated may be injected at depths of about 0.5 mm. In at least one embodiment, the tissue to be treated may be injected at depths of about 1.0 mm. In yet one additional embodiment, the tissue to be treated may be injected at depths of about 1.5 mm.
  • the tissue is injected and treated at a depth of about 2 mm. In another embodiment, the tissue is injected and treated at a depth of about 5 mm. In yet another embodiment, the tissue is injected and treated at a depth of about 1.0 cm. In yet one further embodiment, the tissue is injected and treated at a depth of about 1.5 cm. In one further embodiment, the tissue is injected and treated at a depth of about 2.0 cm. In one further embodiment, the tissue is injected and treated at a depth of about 2.5 cm. In one further embodiment, the tissue is injected and treated at a depth of about 3.0 cm. In one further embodiment, the tissue is injected and treated at a depth of about 3.5 cm. In one further embodiment, the tissue is injected and treated at a depth of about r oc e o. - .
  • the therapeutic compound delivered to the treatment site includes a plurality of microbubbles having, for example, a gas formed therein.
  • a therapeutic compound containing microbubbles is referred to herein as a "microbubble therapeutic compound".
  • the microbubbles are formed by entrapping micro spheres of gas into the therapeutic compound. In one embodiment, this is accomplished by agitating the therapeutic compound while blowing a gas into the therapeutic compound.
  • this is accomplished by exposing the therapeutic compound to ultrasonic energy with a sonicator under a gaseous atmosphere while vibrating the therapeutic compound.
  • gases that are usable to form the microbubbles include, but are not limited to, air, oxygen, carbon dioxide, and inert gases.
  • the therapeutic compound includes approximately 4 ⁇ 10 7 microbubbles per milliliter of liquid.
  • the therapeutic compound includes between approximately 4 ⁇ 10 6 and approximately 4 ⁇ 10 8 microbubbles per milliliter of liquid.
  • the microbubbles have a diameter of between approximately 0.1 ⁇ m and approximately 100 ⁇ m. Other parameters can be used in other embodiments.
  • the efficacy of the therapeutic compound is enhanced by the presence of the microbubbles contained therein.
  • the microbubbles act as a nucleus for cavitation, and thus allow cavitation to be induced at lower levels of ultrasonic energy. Therefore, a reduced amount of ultrasonic energy can be delivered to the treatment site without reducing the efficacy of the treatment. Reducing the amount of ultrasonic energy delivered to the treatment site reduces risks associated with overheating the treatment site, and, in certain embodiments, also reduces the time required to treat a vascular occlusion. In certain embodiments, cavitation also promotes more effective diffusion and penetration of the therapeutic compound into surrounding tissues, such as the vessel wall and/or the clot material.
  • the mechanical agitation caused motion of the microbubbles is effective in mechanically breaking up clot material.
  • a microbubble therapeutic compound is infused intra-arterially or intravenously to the treatment site before the ultrasound radiating members are activated. Therefore, once the ultrasound radiating members begin to generate ultrasonic energy, the microbubble therapeutic compound is already at the treatment site.
  • the microbubble therapeutic compound is delivered using the same catheter that is used to the deliver the ultrasonic energy.
  • the microbubble therapeutic compound is delivered using a separate catheter from the member used to deliver the ultrasonic energy, and delivery of the microbubble therapeutic compound to the treatment site is optionally via the general vascular circulation.
  • the microbubble therapeutic compound is delivered from an ultrasonic catheter intermittently with ultrasonic energy.
  • the microbubble therapeutic compound is delivered without ultrasonic energy.
  • the first and second treatment phases are alternately repeated several times.
  • the duration of the first and second phases are each on the order of approximately a few minutes. This configuration reduces the amount of cavitation occurring within the fluid delivery lumen of the ultrasonic catheter.
  • the therapeutic compound delivered to the treatment site is alternated between a therapeutic compound that contains microbubbles and a therapeutic compound that does not contain microbubbles.
  • the phases with ultrasonic energy correspond to the periods during which the therapeutic compound that does not contain microbubbles is applied.
  • the microbubble therapeutic compound is injected directly into a vascular obstruction — such as a clot — at the treatment site.
  • a vascular obstruction such as a clot — at the treatment site.
  • one or more ultrasound radiating members mounted within a catheter can be energized, thereby delivering ultrasonic energy to the infused microbubble therapeutic compound.
  • the ultrasonic energy delivering member can be optionally repositioned to direct additional ultrasonic energy into the infused microbubble therapeutic compound.
  • This technique allows microbubbles to be suspended within the obstruction.
  • ultrasonic energy is applied to the obstruction, thereby causing mechanical agitation of the microbubbles.
  • the mechanical agitation of the microbubbles is effective in mechanically breaking up clot material.
  • the invention provides methods and apparatus for the treatment of cancer.
  • This includes cancer cells from the bladder, blood, bone, bone marrow, brain, breast, colon, esophagus, gastrointestine, gum, head, kidney, liver, lung, nasopharynx, neck, ovary, pancreas, prostate, skin, stomach, testis, tongue, or uterus.
  • the cancer may specifically be of the following histological type, though it is not limited to these: neoplasm, malignant; carcinoma; carcinoma, undifferentiated; giant and spindle cell carcinoma; small cell carcinoma; papillary carcinoma; squamous cell carcinoma; lymphoepithelial carcinoma; basal cell carcinoma; pilomatrix carcinoma; transitional cell carcinoma; papillary transitional cell carcinoma; adenocarcinoma; gastrinoma, malignant; cholangiocarcinoma; hepatocellular carcinoma; combined hepatocellular carcinoma and cholangiocarcinoma; trabecular adenocarcinoma; adenoid cystic carcinoma; adenocarcinoma in adenomatous polyp; adenocarcinoma, familial polyposis coli; solid carcinoma; carcinoid tumor, malignant; branchiolo-alveolar adenocarcinoma; papillary adenocarcinoma; chromophobe carcinoma; acid
  • inflammatory carcinoma paget's disease, mammary; acinar cell carcinoma; adenosquamous carcinoma; adenocarcinoma w/squamous metaplasia; thymoma, malignant; ovarian stromal tumor, malignant; thecoma, malignant; granulosa cell tumor, malignant; androblastoma, malignant; Sertoli cell carcinoma; leydig cell tumor, malignant; lipid cell tumor, malignant; paraganglioma, malignant; extra-mammary paraganglioma, malignant; pheochromocytoma; glomangiosarcoma; malignant melanoma; amelanotic melanoma; superficial spreading melanoma; malig melanoma in giant pigmented nevus; epithelioid cell melanoma; blue nevus, malignant; sarcoma; fibrosarcoma; fibrous histi
  • ultrasonic energy is used to directly provide the desired therapeutic effect, for example irreversible membrane permeabilization, thermal ablation, and other ultrasonic energy induced effects discussed herein.
  • ultrasonic energy is used to enhance the delivery of anti-cancer therapeutic agents into a target cancer cell, tissue, mass, growth, etc.
  • Anticancer therapeutic agents can be any known in the art.
  • Non- limiting examples of anti-cancer therapeutic agents for delivery to a cancer cell in accordance with the methods and systems of the present invention include, cytotoxic agents, and non-peptide small molecules such as Gleevec® (Imatinib Mesylate), Velcade® (bortezomib), Casodex (bicalutamide), Iressa® (gefitinib), and Adriamycin; alkylating agents such as thiotepa and cyclosphosphamide (CYTOXANTM); alkyl r oc e o. - .
  • cytotoxic agents and non-peptide small molecules
  • non-peptide small molecules such as Gleevec® (Imatinib Mesylate), Velcade® (bortezomib), Casodex (bicalutamide), Iressa® (gefitinib), and Adriamycin
  • alkylating agents such as thiotepa and cyclosphospham
  • sulfonates such as busulfan, improsulfan and piposulfan; aziridines such as benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelamines including altretamine, triethylenemelamine, trietylenephosphoramide, triethylenethiophosphaoramide and trimethylolomelamine; nitrogen mustards such as chlorambucil, chlornaphazine, cholophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard; nitrosureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, ranimustine; antibiotics such as aclacinomysins, actinomycin
  • paclitaxel TAXOLTM, Bristol-Myers Squibb Oncology, Princeton, NJ.
  • docetaxel T AXOTERETM, Rhone-Poulenc Rorer, Antony, France
  • retinoic acid esperamicins
  • capecitabine ecitabine
  • chemotherapeutic cell conditioners are anti-hormonal agents that act to regulate or inhibit hormone action on tumors such as anti- estrogens including for example tamoxifen, (NolvadexTM), raloxifene, aromatase inhibiting 4(5)-imidazoles, 4- hydroxytamoxifen, trioxifene, keoxifene, LY 117018, onapristone, and toremifene (Fareston); and anti- androgens such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogs such as cisplatin and carbop latin; vinblastine; platinum; etoposide (VP- 16); ifosfamide; mitomycin C; mitoxantrone; vincristine; vinorelbine; navelbine
  • the compounds or pharmaceutical composition of the present invention can r oc e o. - . be used in combination with commonly prescribed anti-cancer drugs such as Herceptin®, Avastin®, Erbitux®, Rituxan®, Taxol®, Arimidex®, Taxotere®, ABVD, AVICINE, Abagovomab, Acridine carboxamide, Adecatumumab, ⁇ -N-Allylamino- ⁇ -demethoxygeldanamycin, Alpharadin, Alvocidib, 3-Aminopyridine-2-carboxaldehyde thiosemicarbazone, Amonafide, Anthracenedione, Anti-CD22 immunotoxins, Antineoplastic, Antitumorigenic herbs, Apaziquone, Atiprimod, Azathioprine, Belotecan, Bendamustine, BIBW 2992, Biricodar, Brostallicin, Bryostatin, Buthionine s
  • a direct non-destructive cavitation-based detectable change is induced at a selected location in the brain by applying ultrasound to the selected location of the brain at frequencies ranging from 20 kHz to 5 MHz, and with sonication duration ranging from 100 nanoseconds to 1 minute.
  • a phased array or other ultrasound energy emitter can deliver ultrasound to the selected location in the brain (or a location based thereon) to heat or to cause cavitation sufficient to open the blood-brain barrier, thereby, effecting uptake of neuropharmaceuticals, potential neuropharmaceuticals or other compounds in the blood into that location of the brain.
  • nondestructive heat-based opening of the blood-brain barrier is induced at the selected location in the brain applying ultrasound to the selected location of the brain at frequencies ranging from 250 kHz to 10 MHz, and with sonication duration ranging from 0.10 microseconds to 30 minutes.
  • nondestructive cavitation-based opening (sonoporation) of the blood-brain barrier is induced at the selected location in the brain applying ultrasound to the selected location of the brain at frequencies ranging from 20 kHz to 10 MHz, sonication duration ranging from 100 nanoseconds to 30 minutes, with continuous wave or burst mode operation, where the burst mode repetition varies from 0.01 Hz to 1 MHz.
  • Compounds for delivery across the blood-brain barrier can include, by way of non- limiting example, neuropharmacologic agents, neuroactive peptides (e.g., hormones, gastrointestinal peptides, angiotensin, sleep peptides, etc.), proteins (e.g, calcium binding proteins), enzymes (e.g., cholineacetyltransferase, glutamic acid decarboxylase, etc.), gene therapy, neuroprotective or growth factors, biogenic amines (e.g., dopamine, GABA), trophic factors to brain or spinal transplants, immunoreactive proteins (e.g, antibodies to neurons, myelin, antireceptor antibodies), receptor binding proteins (e.g., opiate receptors), radioactive agents (e.g., radioactive isotopes), antibodies, and cytotoxins, among others.
  • neuroactive peptides e.g., hormones, gastrointestinal peptides, angiotensin, sleep peptides, etc.
  • compounds to be administered into the bloodstream can include high molecular weight complexes formed by combining relatively inert substances, such as EDTA, with neuropharmaceuticals or other substances currently known to pass through the blood-brain barrier. Due to their sizes and/or molecular configurations, such complexes are prevented from crossing the barrier, except at selected locations in the brain opened via ultrasound as described herein. Use of such complexes in connection with the invention, therefore, permits localized application of compounds that might otherwise produce unwanted effects in other parts of the brain or body. [00146] It will be appreciated that administration of the compound need not necessarily precede application of the ultrasound.
  • the ultrasonically-opened blood-brain barrier typically permits uptake of administered compounds for at least a short period of time
  • the compound can be introduced into the blood stream after the barrier-opening ultrasound dose is applied.
  • the methods and apparatus described herein can be employed for treating neurological disorders by image-guided or EIT-guided ultrasonic deliver of compounds through the blood-brain barrier.
  • Such disorders include tumors, cancer, degenerative disorders, sensory and motor abnormalities, seizure, infection, immunologic disorder, mental disorder, behavioral disorder, and localized CNS disease, among others.
  • the foregoing apparatus and methods can be used to introduce selective cytotoxins into selected locations of the brain to destroy all or selected cell types there.
  • these apparatus and methods can be employed to introduce immunologic agents at those selected locations. Still further, they can be employed in neural pathway tracing studies using retrograde or anteretrograde axonal transport, or in neurophysiological testing using localized delivery of activation or inhibition. In still further related aspects, the invention provides methods for modification of neurologic and neurologically-related activity (e.g., behavioral activity, memory-related activity, and sexual activity, among others) by such methods.
  • neurologic and neurologically-related activity e.g., behavioral activity, memory-related activity, and sexual activity, among others
  • the invention provides methods and apparatus for the delivery of short- interfering RNA (siRNA) into an organ, tissue, cells, or other target body mass, and specifically liver or a portion of liver or liver tissue.
  • siRNA short- interfering RNA
  • the invention provides methods and apparatus for the treatment of liver diseases, including but not limited to alcoholic liver, liver fibrosis and cirrhosis.
  • liver diseases including but not limited to alcoholic liver, liver fibrosis and cirrhosis.
  • TNF-alpha activation plays crucial roles in alcoholic liver injuries, and activation of RSK protein is believed to trigger excessive production of collagen and resulting in fibrotic or cirrhotic liver.
  • intervention of these pathological pathways by inhibiting or knocking-down expression of certain proteins, such as RSK and TNF-alpha, on the pathways is utilized to prevent progression or reverse liver diseases such as liver cirrhosis and liver tumor.
  • siRNA molecules that target TNF-alpha can be delivered by sonoporation to prevent further liver damages in patients with alcoholic liver.
  • delivery of siRNA molecules that target key proteins on the pathway of RSK activation, such as C/EBP-beta-Ala217 can be used to treat patients with fibrotic or cirrhotic livers.
  • delivery of therapeutic siRNA moleucles to the liver is facilitated by sonoporation, optionally guided by EIT.
  • siRNA solution is introduced into liver via hepatic artery injection or portal vein injection, and electrical impedance of liver tissue and EIT image of the whole liver is measured and generated before, during and after ultrasound exposure.
  • the electrical signal collected serves as a real-time feedback used for real time sonoporation optimization so as to achieve the best siRNA delivery efficiency while minimizing induced damage in the treated liver area.
  • Ultrasound can be focused to deep tissue and scanned through the area of interest, and can provide controlled sonoporation across the entire organ for whole-liver siRNA delivery.
  • Example 1 Sample ultrasonic setup
  • an ultrasonic setup may include a commercially available system Sonitron 2000 (Rich Mar, Chattanooga, Tennessee) with IMHz 6 mm probe. It is appreciated that other sonoporation devices and probes may be used. In one example, this system may be sufficient for experiments with small liver samples. Localized sonoporation targeting specific regions of a large liver can be achieved by using focused ultrasound beams generated by phase array sonoprobes or curved transducers with a long focal length controlled by using a more comprehensive transmitting control system. The Sonitron system may be calibrated to provide reliable intensity output level from 0 ⁇ 20W/cm2.
  • Pulse length, pulse repetition frequency and treatment duration may also be adjustable.
  • a 6 mm probe may be chosen because of its even ultrasound energy r oc e o. - . distribution and a beamwidth of 5-6 mm in the range of 3-7 mm from the tip of the probes, which is sufficient to induce sonoporation in small tissue samples.
  • an ultrasound transmission gel which has acoustic transmission properties similar to human tissues, will be applied between the sonoprobe and the liver tissue, in order to avoid formation of air bubbles that can substantially impede ultrasound transmission.
  • Such experimental procedures are often not necessary for in vivo studies.
  • sonoprobe(s) are placed outside the patient's body, ultrasound energy is coupled into the body through acoustically transparent media and focused to the target area, but not in direct contact with the liver.
  • MDCK Mesh-Darby canine kidney cells in vitro.
  • MDCK cells were used because they are epithelial cells, the same cell type of liver hepatocytes, which are notoriously hard to transfect using commercially available transfection techniques such as lipofection.
  • Alexa Fluro 488 a bright and stable green dye from QIAGEN was used to evaluate the effectiveness of the siRNA delivery.
  • a high concentration (1 ⁇ M) of the labeled siRNA was used in order to detect cellular siRNA uptake using an inverted fluorescent microscope (1X71 Olympics).
  • MDCK cells were seeded in the wells of a 24-well culture plate with a density of 1 x 10 5 cells/well and cultured in an incubator (37°C, 5% CO 2 ) for 24 hours.
  • the culture medium was replaced with the siRNA solution with a final concentration of 1 ⁇ M after mixing with ultrasound contrast agent solution (Artison, Inola, OK).
  • ultrasound contrast agent solution Artison, Inola, OK.
  • the ultrasound was irradiated using a commercially available sonoporation system Sonitron 2000 (Rich Mar, Chattanooga, Tennessee) with a 6mm IMHz probe. As shown in FIG. 6, the ultrasound probe was inserted into the siRNA solution and the distance between the tip of the probes and the cell monolayer was controlled at 2 ⁇ 3mm.
  • FIG. 9 shows bright field and corresponding fluorescent images of negative control cells (column a) and sonoporated cells (column b).
  • the images were taken using an Olympus DP 1200 r oc e o. - . digital camera with a 1Ox PC objective.
  • the fluorescent image of the control shows virtually no uptake of the siRNA (the two bright dots are most- likely dead cells).
  • significant siRNA uptake was observed in more than 80% (estimated using an imaging analysis program) of the cells receiving sonoporation treatment.
  • Minimal cell damage was also achieved, as shown in the bright field images. This result clearly demonstrates that the siRNA molecules can be delivered into MDCK cells with a high efficiency while keeping most of the cells intact.
  • Example 3 Ex vivo siRNA delivery to mouse liver tissue via sonoporation
  • siRNA was delivered ex vivo by intrahepatic injection of siRNA during sonoporation treatment.
  • Harvested mouse liver tissues were obtained as lab waste from Antagene Inc. (Mountain View, CA), and used in the experiments.
  • a final concentration of 4 ⁇ M Alexa- 488 labeled siRNA (QIAGEN) solution was prepared after mixed with 15% (v/v) ultrasound contrast agent (Artison, Inola, OK).
  • Mouse liver lobes were submerged in DPBS (GIBCO) solution and the ultrasound probe was inserted into the solution.
  • DPBS DPBS
  • the location of the probe tip was adjusted to the position of lmm above the liver lobes.
  • 100 ⁇ l of the mixed siRNA solution were injected directly into the liver lobes over a period of 2 minutes, and in the mean time, the liver lobes were treated with I MHz ultrasound transmitted at 2 W/cm 2 output level with a duty cycle of 20%.
  • the treated liver lobes were embedded in O. CT compound (Sakura Finetek USA, Inc) and then immediately immersed in liquid nitrogen.
  • the frozen samples were cryosectioned. The thickness of each section was 15 ⁇ m.
  • the control liver lobes were handled with the same procedure except for treatment with ultrasound.
  • the control and treated sections were examined under the fluorescent microscope and images were taken using an Olympus DP 1200 digital camera with a 2Ox objective.
  • FIG. 10 shows example images of tissue slices from both control and sonoporated liver tissues.
  • FIG. 1 Oa shows auto-fluorescence of liver tissue that was not treated with fluorescently labeled siRNA and sonoporation.
  • FIG. 10b shows that the dye-labeled siRNA molecules have difficulty passing the cell membrane without facilitation.
  • FIG. 10c shows that substantial amount of fluorescence above auto- fluorescence background.
  • Example 4 In vivo delivery of siRNA to mouse liver
  • the efficacy of siRNA as a therapeutic delivered using sonoporation guided by EIT can be evaluated using double transgenic mice (LAP-tTA;TRE-Myc) that overexpress human c-Myc in hepatocytes under the control of doxycycline as a model for the development of liver cancer.
  • the double transgenic mice develop multiple liver tumor nodules within 1 to 2 months post removal of doxycycline (FIG. 1 Ia). Histological examination reveals that the tumor resembles poorly differentiated hepatocellular carcinoma (HCC) in which tumor cells show prominent nuclei and little cytoplasm stains (FIG. 1 Ib).
  • HCC poorly differentiated hepatocellular carcinoma
  • siRNA targeted to c-Myc can be delivered to livers of the mice via tail vein injection.
  • a scrambled siRNA that does not target c-Myc can be used in some transgenic mice to provide a negative control group.
  • Ultrasound can then be directed to target the liver using EIT in accordance with the methods of the invention.
  • a second control group can receive the siRNA targeting c-Myc, but without receiving ultrasound treatment. This process can be repeated, for example, once every two weeks for three months. After three months, the livers can be examined for the number and size of tumors, with a decrease in one or both in the experimental group compared to the control groups indicating the therapeutic effectiveness of siRNA delivered by EIT guided sonoporation.

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Abstract

L'invention porte sur des procédés, sur des dispositifs et sur des appareils pour fournir une tomographie par impédance électrique (EIT) avec des ultrasons. Les ultrasons peuvent être utilisés pour une sonoporation de cellules/tissu ou une ablation de tissu. La EIT peut aider à la détermination du niveau et/ou de l'emplacement d'ultrasons devant être appliqués. La EIT peut également assurer une surveillance et une commande pour un ciblage guidé des ultrasons pour une perméabilisation de cellules/tissu ou une ablation de tissu.
PCT/US2010/033450 2009-05-01 2010-05-03 Sonoporation, ablation de tissu par ultrasons, guidées par tomographie par impédance électrique (eit), et utilisation de celles-ci Ceased WO2010127369A1 (fr)

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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012012750A1 (fr) 2010-07-23 2012-01-26 Trustees Of Boston University Inhibiteurs anti-despr comme agents thérapeutiques pour l'inhibition de l'angiogenèse pathologique et de l'invasivité des cellules tumorales et pour l'imagerie moléculaire et l'administration ciblée
FR3012042A1 (fr) * 2013-10-23 2015-04-24 Edap Tms France Appareil de generation d'ondes ultrasonores focalisees a temps de traitement reduit
WO2016037357A1 (fr) * 2014-09-12 2016-03-17 重庆海扶医疗科技股份有限公司 Dispositif de rééducation post-partum
WO2020197400A1 (fr) 2019-03-27 2020-10-01 Umc Utrecht Holding B.V. Anticorps iga modifiés et procédés d'utilisation
WO2023180811A3 (fr) * 2022-03-22 2023-11-23 Insightec Ltd. Surveillance de la perméabilité d'un tissu pendant des procédures ultrasonores

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20010031243A1 (en) * 1997-09-15 2001-10-18 Imarx Pharmaceutical Corp. Novel methods of ultrasound treatment using gas or gaseous precursor-filled compositions
US20020077627A1 (en) * 2000-07-25 2002-06-20 Johnson Theodore C. Method for detecting and treating tumors using localized impedance measurement
US20030130711A1 (en) * 2001-09-28 2003-07-10 Pearson Robert M. Impedance controlled tissue ablation apparatus and method
US20070055179A1 (en) * 2005-09-07 2007-03-08 Deem Mark E Method for treating subcutaneous tissues
US20070083120A1 (en) * 2005-09-22 2007-04-12 Cain Charles A Pulsed cavitational ultrasound therapy

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20010031243A1 (en) * 1997-09-15 2001-10-18 Imarx Pharmaceutical Corp. Novel methods of ultrasound treatment using gas or gaseous precursor-filled compositions
US20020077627A1 (en) * 2000-07-25 2002-06-20 Johnson Theodore C. Method for detecting and treating tumors using localized impedance measurement
US20030130711A1 (en) * 2001-09-28 2003-07-10 Pearson Robert M. Impedance controlled tissue ablation apparatus and method
US20070055179A1 (en) * 2005-09-07 2007-03-08 Deem Mark E Method for treating subcutaneous tissues
US20070083120A1 (en) * 2005-09-22 2007-04-12 Cain Charles A Pulsed cavitational ultrasound therapy

Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012012750A1 (fr) 2010-07-23 2012-01-26 Trustees Of Boston University Inhibiteurs anti-despr comme agents thérapeutiques pour l'inhibition de l'angiogenèse pathologique et de l'invasivité des cellules tumorales et pour l'imagerie moléculaire et l'administration ciblée
EP3696195A1 (fr) 2010-07-23 2020-08-19 Trustees of Boston University Inhibiteurs anti-despr comme agents thérapeutiques pour l'inhibition de l'angiogenèse pathologique et de l'invasivité des cellules tumorales et pour l'imagerie moléculaire et l'administration ciblée
FR3012042A1 (fr) * 2013-10-23 2015-04-24 Edap Tms France Appareil de generation d'ondes ultrasonores focalisees a temps de traitement reduit
EP2865420A1 (fr) 2013-10-23 2015-04-29 EDAP TMS France Appareil de génération d'ondes ultrasonores focalisées à temps de traitement réduit
US10004922B2 (en) 2013-10-23 2018-06-26 Edap Tms France Apparatus for generating focused ultrasound waves with reduced treatment time
WO2016037357A1 (fr) * 2014-09-12 2016-03-17 重庆海扶医疗科技股份有限公司 Dispositif de rééducation post-partum
WO2020197400A1 (fr) 2019-03-27 2020-10-01 Umc Utrecht Holding B.V. Anticorps iga modifiés et procédés d'utilisation
WO2023180811A3 (fr) * 2022-03-22 2023-11-23 Insightec Ltd. Surveillance de la perméabilité d'un tissu pendant des procédures ultrasonores

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