Patent Application
20120089009
This disclosure relates generally to delivering nanoparticles to tissue, and, more particularly, to electro-nanotherapy to deliver nanoparticles to tissue using electroporation.
In recent years, delivery of substances for treatment of tumors has been hampered by a greater permeability of surrounding cells than the tumor cells that are the target for treatment. This difference in permeability has resulted in a decrease in the effectiveness of treatment or a reliance on destroying the cell using a technique such as ablation.
As used in this patent, stating that any part (e.g., a component, module, subsystem, device, control, probe, injector, imager, etc.) is in any way positioned on (e.g., positioned on, located on, disposed on, or formed on, etc.) another part, means that the referenced part is either in contact with the other part, or that the referenced part is above the other part with one or more intermediate part(s) located therebetween. Stating that any part is in contact with another part means that there is no intermediate part between the two parts.
Delivery of nanoparticles (NPs) to a site of disease in a patient is a desirable modality of therapy. Nanoparticles are defined as small objects that behave as a unit with respect to its transport and properties. Nanoparticles can range in size between 1 and 100 nanometers in diameter, for example. Nanoparticles can exhibit size-dependent properties that differ from properties observed in other particles.
Nanoparticle chemistry can be used in a variety of applications including medical therapy. Superparamagnetic Iron Oxide nanoparticles (SPIOs), for example, are non-toxic and biodegradable and, when surface functionalized or coated (e.g., by covalently attaching to the surface of SPIOs) with chemotherapeutics, such as doxorubicin, are able to be absorbed or otherwise taken up by a variety of cell types. Also, chemotherapeutics attached to the SPIO nanoparticles are stable and feature a slow release profile once intracellular entry has occurred. Due to their iron content, SPIOs have a high magnetic moment and both high R2 and R2* relaxivity, and can be imaged using magnetic resonance imaging (MRI), for example, to noninvasively examine tissue uptake. Thus, SPIO nanoparticles can be used to transfect cells with chemotherapeutics and can simultaneously be used as a magnetic resonance (MR) contrast agent, permitting image-guided drug delivery.
For example, Doxorubicin is an anthracycline class antineoplastic drug used to treat a wide variety of solid and hematologic malignancies. Doxorubicin induces cytotoxicity through DNA intercalation and can be administered in chemotherapy to treat cancer in a patient.
Intravenous (IV) delivery of nanoparticles, however, is hampered by a proportionally large uptake of NPs by the reticuloendothelial system. The reticuloendothelial system (RES) is a part of a human immune system that includes phagocytic cells located in reticular connective tissue. The cells are primarily monocytes and macrophages, and they accumulate in lymph nodes and the spleen. The Kupffer cells of the liver and tissue histiocytes are also part of the RES. Cells in the RES absorb a large number of NPs injected intravenously and thereby prevent NPs from reaching desired sites in sufficient concentration.
Electroporation (abbreviated herein as EP, and also known as electropermeabilization) provides a technique to increase delivery of molecules to sites in cells and tissues that are treated. Using a series of brief electrical pulses delivered to in vivo tissues via, for example, a pair of electrodes, EP is able to make tissues more permeable to small and large molecules at the cellular level by affecting the cell plasma membrane. This permeabilization effect greatly increases an uptake or loading of NPs in the tissue that has received EP treatment and serves to “guide” NPs preferentially to the treated tissues. Using electroporation, a localized transmembrane voltage is applied to one or more points on a cell membrane. For a given electrical pulse duration and shape, a corresponding transmembrane voltage threshold is to be exceeded to manifest electroporation. Cells within areas where an electric field magnitude exceeds an electric field magnitude for electroporation provide greater permeability. Exceeding the threshold by too wide a margin can permanently damage the cells (e.g., irreversible electroporation).
Electro-nanotherapy, therefore, typically involves delivery of nanoparticles to tissues treated with electroporation. Using electro-nanotherapy (Electro-NT), the uptake of NPs in treated tissues can be greatly increased over administration of NPs alone (i.e., without electroporation treatment). Electro-nanotherapy can be used for a variety of applications including treatment of cancer, delivery of agent(s) for tissue regeneration, delivery of molecularly targeted imaging agent(s), etc. As used herein, electro-nanotherapy may also be referred to as nanoablation, for example.
Electroporation achieves disruption of cell membranes via application of an external electric field. The disruption causes otherwise low permeant or nonpermeant molecules to have increased permeability. A degree of cell membrane disruption at any given point on a cell membrane surface (M) is directly related to a transmembrane potential difference experienced at that point M, ΔVM. The transmembrane potential difference experienced at point M is related to an externally applied electric field and cell radius according to the following equation:
ΔVM=1.5×r×Eext×cos θ (Equation 1),
where r is a radius of a cell, Eext is an external electric field strength, and θ is a polar angle with respect to an electric field direction. Depending on a degree of electroporation, effects can either be reversible (e.g., a cell will return to normal with no deleterious effects after a certain length of time) or irreversible (e.g., the disruption of the cell membrane is permanent, thereby causing cell death). Electro-nanotherapy, or nanoablation, takes advantage of reversible EP to deliver NPs to cells without destroying them. However, in certain cases destruction of tissue may be desirable (e.g., in the case of solid tumor malignancies), and parameters of an externally applied electric field during an Electro-NT procedure can be adjusted to provide reversible or irreversible EP, as needed.
In certain examples, Electro-NT includes delivery of nanoparticles, either intravenously or intra-arterially, followed within a time period by electroporation at a tissue site where increased nanoparticle uptake is desired.
Certain examples provide a method for electro-nanotherapy of cells at a tissue site. The method includes facilitating identification of a tissue site for electro-nanotherapy. The example method includes facilitating injection of nanoparticles at the tissue site. The example method includes enabling generation of one or more electric pulses at the tissue site to create pores in a cell membrane at the tissue site such that the nanoparticles penetrate the cell membrane.
Certain examples provide a superparamagnetic iron oxide nanoparticle treated with a chemotherapeutic to be absorbed by a cell membrane in patient tissue when the tissue is stimulated using a series of electrical pulses, the nanoparticle adapted to be imaged to permit image-guided drug delivery.
Certain examples provide a system for electro-nanotherapy of cells at a tissue site in a patient. The example system includes a nanoparticle injector to facilitate injection of nanoparticles treated with a chemotherapeutic at an identified tissue site. The example system includes a controller to enable generation of one or more electrical pulses at the tissue site to create pores in a cell membrane at the tissue site such that the nanoparticles penetrate the cell membrane.
In an example, Electro-NT was applied to a rat model of hepatocellular carcinoma (HCC). First, the animal model was created by injecting a suspension of N1S1 cells (e.g., a rat hepatoma) into the extracorporeally exposed liver of two living rats. Rat livers were exposed using a surgical mini-laparotomy procedure and aseptically closed following cell injection. The rats survived for fourteen (14) days to allow for growth of the implanted tumor. At 14 days post-implantation, rats were enrolled in the Electro-NT study. Rat 1 served as the control rat and received an IV injection of 0.2 mL SPIOs at a concentration of 5 mg of iron/ml. The SPIOs in the example were 15 nm in diameter and were functionalized with doxorubicin. The control rat 1 was imaged using a T2-weighted Turbo Spin Echo (T2W-TSE) MR sequence following nanoparticle administration to confirm delivery. The control rat (Rat 1) was kept alive for 100 minutes, sacrificed and necropsied to obtain tissue samples. Rat 2 served as a treatment rat. Rat 2 received the same IV injection of nanoparticles as Rat 1, which was then followed by EP. To perform EP, the liver of Rat 2 was exposed surgically through a mini-laparotomy. EP electrodes were placed into the tumor and an electric field was applied using the following parameters: 1300 V, 8 pulses, 100 microseconds pulse time, at 100 millisecond intervals. Rat 2 was also kept alive for 100 minutes then sacrificed and necropsied for tissue harvest. Samples of tumor tissue were quantitatively analyzed for iron (Fe) content using inductively coupled plasma mass spectroscopy. The example results are shown in the following table:
As shown by the example results, the amount of SPIOs taken up or absorbed by the tumor tissue using Electro-NT was greater than the amount delivered to the tumor using IV administration alone—with increased penetration of the particles into the tumor core. These findings are also observed using MR imaging, which shown increased tissue contrast within an electroporated tumor zone compared to the control tumor.
In certain examples, an NP therapeutic can be scaled-up and produced in bulk for sale. An NP therapeutic material can be adapted or configured to treat a variety of solid tumor malignancy(ies), cause tissue growth in a solid organ requiring it (e.g., liver regeneration, heart regeneration after a myocardial infarction, etc.), deliver molecular imaging agent(s) to tissue(s) of interest, etc. Coupled with EP, NPs can be guided specifically to target tissue(s) with high resulting uptake or absorption in the tissue(s). Electro-nanotherapy can be developed to treat and/or diagnose a variety of disease processes including cancers, solid organ disease, cardiac disease, etc., using efficient delivery of NPs to desired site(s) in a body.
As discussed above, electroporation (EP) can be utilized to modulate influx of chemotherapeutics into tumor cells, both in vitro and in vivo. During EP, when cells are exposed to brief direct current, the electric field induces transient cell membrane channels, which form temporary pores. The temporary pores allow passage of extracellular macromolecules into the cytosol of the cell. The electric pulses also induce transient vascular hypoperfusion within the treated zone. A resultant restriction of flow diminishes washout of therapeutics from the treated zone. In certain examples, the therapeutic agent has already been administered to the patient and is already within the target EP zone at the time of treatment. In other examples, the therapeutic agent is administered to the patient at the time of EP treatment and/or shortly following EP treatment with the treated zone still exhibits greater permeability.
To explore the relationship between the timing of therapeutic delivery and tumor EP, therapeutic superparamagnetic iron oxide nanoparticles (SPIOs) were utilized in an example to serve as a dual MR imaging agent and drug delivery vehicle. Eight VX2 tumors were surgically implanted in rabbit hind limbs. SPIO-NPs were obtained and functionalized with doxorubicin (e.g., mean diameter=10 nm). In the example, all animals underwent anatomic T2 turbo spin echo (TSE) imaging to confirm tumor growth and location, and T2 weighted (T2*W) imaging to determine baseline tumor signal intensity. Following scans, animals were transferred to the angiography suite for carotid artery catheterization and femoral artery angiography under X-Ray Digital Subtraction Angiography guidance to confirm ideal catheter placement prior to therapeutic delivery (e.g., SPIO+ethiodol). Each tumor was then electroporated at different times relative to SPIO embolization (e.g., range: −5 minutes to +3 minutes). T2*W images were then obtained post-procedurally to confirm NP delivery and evaluate tumor signal changes. The rabbits were then euthanized and tissues were harvested to determine SPIO uptake using inductively coupled plasma mass spectroscopy (ICP-MS). Mean SPIO-NP concentration within all tumors were compared between timing groups using ANOVA with post-hoc Tukey analysis. A p<0.05 was considered significant.
In the example, ICP-MS analysis of iron content demonstrated that tumors that underwent EP for 1.5 to 2.25 minutes following injection showed a 2.9 fold increase in SPIO concentration compared to all other time points (p<0.05). These findings were confirmed by a noted decrease in MR signal intensity on T2*W imaging. Groups that underwent EP outside this window did not demonstrate appreciable T2*W signal changes within their tumors.
The timing of EP, relative to intra-arterial (IA) therapeutic embolization, for example, can affect tumor uptake of the NPs. In the example, a therapeutic window was observed to occur between 1.5 to 2.25 minutes following therapeutic delivery, for example. A decrease in uptake was noted when EP occurred outside this window. Furthermore, these findings can be observed non-invasively using T2*W imaging in vivo. EP can be shown to provide efficacy in tumor therapy using these timing parameters.
Hepatocellular carcinoma (HCC) is the sixth most common cancer worldwide and the third most common cause of cancer death. New therapies can help interventional radiologists to improve patient outcomes. As discussed above, nanoparticles represent a promising new drug delivery platform. However, systemic administration results in sequestration by the healthy liver tissue, and minimal tumor uptake. Using an N1-S1 rat model, reversible electroporation (EP) was tested in an example to determine an increase intratumoral uptake of therapeutic superparamagnetic iron oxide (SPIO) nanoparticles loaded with doxorubicin (DOX).
In the example, using the N1-S1 rodent model, hepatomas were grown in twelve Sprague-Dawley rats that were divided into treatment and control groups. Magnetic resonance (MR) imaging was performed at 10-14 days to confirm tumor growth. For both groups, 0.56 mg/kg body weight SPIO-DOX nanoparticles were injected via the femoral vein. For the treatment group, EP electrodes were inserted and 8 pulses (e.g., 100-ns pulse duration, 1,300-V/cm field strength) were applied to liver tumors 1.5 minutes post-SPIO-DOX injection. T2*-weighted imaging was performed on both groups to visualize nanoparticle delivery and uptake. Both groups were sacrificed and tumors were harvested for evaluation by ICP-MS for iron concentration. Prussian Blue staining was done to visualize iron content. Iron concentrations between the groups were compared with paired t-tests, with p<0.5 considered significant.
In the example, N1S1 tumors were grown in all twelve rats. Electroporation resulted in increased uptake within the tumor tissue over IV delivery alone. Within the tumor core a 5.57 times or 6.3 times, for example, increase in iron content over IV delivery was observed (p<0.05). In an example, these findings were also confined on T2*W MRI in vivo and Prussian Blue staining of tumor specimens.
Thus, EP enhances tumor uptake of SPIO-DOX nanoparticles and can serve to improve tumor uptake of other therapeutic nanoparticles.
In the illustrated example, the controller 210, such as an AngioDynamics HPV01 Generator, supplies a series of one or more electrical pulses to the probe 220. A user and/or program setting can specify one or more characteristics of the series of pulses using the controller 210. The controller 210 can be used to set a voltage, number of pulses, pulse duration, and/or pulse interval, etc. The configured series of pulses is triggered by the controller 210 for generation through the probe 220 (e.g., an AngioDynamics NanoKnife™ needle and/or other single or multiple electrical probes).
In the illustrated example, the controller 210 includes a function generator. One or more electrodes forming the probe 220 are attached to the function generator to apply a voltage at a target site with respect to the patient 250. Parameters used to configure the probe 220 for electroporation can include, for example, a voltage between 250-1500 Volts per centimeters (V/cm); a number of pulses between 4 and 10; a pulse duration between 99 microseconds (μs) to 100 milliseconds (ms); a pulse interval between 100 ms and 1 Hertz (Hz); and a number of electrodes between 1 and 8.
The probe 220 of the illustrated example is positioned with respect to a cell site of interest on and/or in the patient 250. For example, the probe 220 can be positioned on the skin of the patient 250 against and/or over the cell site of interest (e.g., cutaneously). Alternatively or in addition, the probe 220 can be inserted into the patient 250 to be adjacent to the cell site of interest within the patient 250 (e.g., inserting a needle probe 220 near an organ and/or other tumor site of interest), for example. Thus, the electroporation device 220 can be placed through the skin (percutaneously) or via surgical laparoscopy to access the tissue site. In some examples, a combination of surgical, percutaneous, and/or cutaneous probe(s) 220 can be used to facilitate electroporation.
The nanoparticle injector 230 of the illustrated example provides nanoparticles to the patient 250 (e.g., to a particular tissue site of interest in the patient 250) via systemic and/or local delivery. In the example of
Some examples of needle electrode probes are shown in
After nanoparticles have been introduced in the patient 250 via the injector 230 and a series of pulses have been applied at a patient cell site of interest by the probe 220 to increase permeability for the nanoparticles at the cell site, results can be evaluating using the example imager 240 of
While an example manner of implementing an electro-nanotherapy has been illustrated in
A flowchart including blocks representative of example machine readable instructions for implementing some or all of the system 200 of
As mentioned above, the example processes of
At block 420, positioning of an electric field source (e.g., a needle or probe electrode) is performed and/or enabled to be performed with respect to the tissue site. For example, a user, such as a surgeon and/or other clinician, can position the electric field source on and/or in the patient with respect to the tissue site the user wishes to have increased permeability. The electric field source can include and/or be connected to a controller including a user interface and a power unit. The user interface accepts user input and calculates treatment parameters based on the input and possibly other stored information. The power unit generates electrical pulses based on the treatment parameters. Using the controller, timing delays and triggering signals can be configured and provided to the electric field source.
At block 430, nanoparticles are injected and/or injection is facilitated into the patient. Injection can be performed using one or more devices include IV injection, IA injection, surgical introduction of nanoparticles, bathing of a tumor and/or other cell site in NPs, etc.
At block 440, a series of one or more electric pulses is generated and applied at the tissue site. Electroporation can employ micro to millisecond electric pulses to create pores in the cell membrane, thus allowing molecules that, due to their physical and/or chemical properties, would normally not be able to cross the cell membrane, to enter the cell. Using a control, electroporation can be triggering using a series of electrical pulses generated at a needle and/or other probe using a power supply in the control. In electrochemotherapy, a combination of chemotherapy and electroporation of tumors, the effects of nanoparticles or drugs are increased. The opening of pores in the cell membrane allows a chemotherapeutic agent to enter the cell at greater, more effective concentration. An electroporator device can use one or more electrodes to apply an electric field with a desired appropriate shape and intensity to homogenously cover the target tissue. A switching unit can be used with the controller to route treatment to the electrodes in sequenced fashion.
In some examples, nanoparticles are injected prior to generation and application of electric pulses. In some examples, nanoparticles are injected while and/or after electric pulses are applied to the tissue site.
At block 450, electro-nanotherapy at the tissue site is monitored. For example, one or more images, such as MRI images, can be obtained to monitor and review results of the electro-nanotherapy. Alternatively or in addition, results can be verified using a mass spectrometry analysis of a tissue sample to determine absorption of the nanoparticles. Monitoring can also include images taken to visualize electrode placement and determine other positioning information for image-guided surgery, image-guided delivery of nanoparticles, etc., before, during, and/or after nanoparticle insertion and/or electrical pulse generation, for example.
The processor 512 of
The system memory 524 can include any desired type of volatile and/or non-volatile memory such as, for example, static random access memory (SRAM), dynamic random access memory (DRAM), flash memory, read-only memory (ROM), etc. The mass storage memory 525 can include any desired type of mass storage device including hard disk drives, optical drives, tape storage devices, etc.
The I/O controller 522 performs functions that enable the processor 512 to communicate with peripheral input/output (“I/O”) devices 526 and 528 and a network interface 530 via an I/O bus 532. The I/O devices 526 and 528 can be any desired type of I/O device such as, for example, a keyboard, a video display or monitor, a mouse, etc. The network interface 530 can be, for example, an Ethernet device, an asynchronous transfer mode (“ATM”) device, an 802.11 device, a DSL modem, a cable modem, a cellular modem, etc. that enables the processor system 500 to communicate with another processor system.
While the memory controller 520 and the I/O controller 522 are depicted in
From the foregoing, it will appreciate that methods, apparatus, and articles of manufacture have been described which improve cell permeability and/or delivery of nanoparticles to a target tissue site for cell treatment.
Although certain example methods, apparatus and articles of manufacture have been described herein, the scope of coverage of this patent is not limited thereto. On the contrary, this patent covers all methods, apparatus and articles of manufacture fairly falling within the scope of the claims of this patent.
The present application claims the benefit of priority to U.S. Provisional Application No. 61/391,855 filed on Oct. 11, 2010, entitled “Methods and Apparatus to Deliver Nanoparticles to Tissue Using Electro-nanotherapy”, which is herein incorporated by reference in its entirety.
| Number | Date | Country | |
|---|---|---|---|
| 61391855 | Oct 2010 | US |
