Systems, devices, and methods for sensing, detecting, and implementing in a micro-nano environment

Abstract

Describes a device that is on the order of μm or nm in size and can include integrated circuit logic, a sensing subsystem, a decision-making subsystem, an implementation subsystem, and a power harvesting system based on sub-10nm SIA transistor nodes. The sensing subsystem can identify pathogenic entities, which include disease-related cells (such as cancer cells, autoimmune cells, or pathogenic microorganisms) or viruses. The sensing subsystem can include at least one pad made of a conductive material and at least one linker attached to the at least one pad. Each linker can also be attached to a targeting agent (such as an antibody fragment) or a reference agent. After the targeting agent binds to the entity of interest, information is transmitted to the logic circuit, which processes the binding event information and determines whether the entity is disease-related (e.g., a disease-related cell or virus) and requires the delivery of a therapeutic agent or other treatment.

Description

[0001] Cross - reference to related applications

[0002] This application claims the benefit of co-pending U.S. Provisional Patent Application No. 62 / 830,762, filed on April 8, 2019, which is incorporated herein by reference in its entirety. Technical Field

[0003] The present invention relates to therapeutic or medical nanorobots, including nano-devices or micro-devices that sense and identify chemical and biological materials and can dispense targeted therapeutic agents and other chemicals. Background Art

[0004] Disease-causing entities, including disease-associated cells (“DACs”) and pathogenic viruses, are a widespread and major cause of morbidity and mortality in diseases that are at the root of pathological mechanisms. Such diseases include, but are not limited to, cancer, autoimmune diseases, and infectious diseases (including those caused by viruses, parasites, bacteria, etc.). In 2019, cancer alone caused approximately 600,000 deaths in the United States, with an estimated 1.8 million new cancer diagnoses. Similar trends are observed in autoimmune and infectious diseases, as well as in highly prevalent cardiovascular, neurological, and metabolism-related diseases (such as metabolic syndrome, obesity, diabetes) in terms of high economic costs and disease prevalence and mortality.

[0005] The primary reason for the lack of progress in developing effective and definitive therapeutic agents targeting disease-causing entities, including DACs, parasites, viruses, and others, is the inherent ability of these disease-causing entities to adapt and develop resistance mechanisms that render many therapeutic agents ineffective. There is now increasing concern about infectious diseases that are even resistant to the most advanced antibiotics. The ability of pathogens to adapt antibiotic resistance mechanisms creates a continuous need to develop new antibiotic strategies. In addition, there is an ever-present threat that we will encounter new microbial or viral resistant strains that are untreatable and will effectively force medicine back to the pre-penicillin era before treatment was possible. Despite many efforts made in the past century to develop new small molecule-, biologic-, and nanoparticle-based therapeutic agents to neutralize disease-causing entities, including against DACs, parasites, and viruses, viable technologies that can act on cells resistant to treatment have emerged. In almost all attempts in all treatment modalities developed to date, some biological process has been utilized—the process by which disease-causing entities, including DACs, parasites, and viruses, evolve and develop while counteracting resistance mechanisms. The present disclosure describes logic-based and non-bionic effector systems, devices, and methods for neutralizing disease-causing entities against which corresponding resistance cannot be generated.

[0006] With new and growing concerns in the ability to safely treat specific cell types, avoiding off-target effects and pathological activation of the immune system in patients, as in the case of viral vector-related gene therapies and CAR-T therapies, for example, chimeric antigen receptor T cell (CAR-T) therapies, clustered regularly interspaced short palindromic repeats (“CRISPR”), siRNA, and other gene therapies. New solutions are needed to not only improve the precision of these drugs, but also to be able to deliver these drugs to specific pathogenic entities, including DACs and viruses, which will reduce the potential toxicity (off-target effects) of these drugs and significantly increase their efficacy at high concentrations released locally at the pathogenic entity by delivering the therapeutic agent or therapeutic action directly to the pathogenic entity. This disclosure describes how to deliver any therapeutic agent or imaging agent only to the pathogenic entity of interest (e.g., only to DACs, such as cancer cells), facilitate entry of the reagent into specific DACs regardless of the size or chemical composition of the therapeutic agent or imaging agent, and reduce pathological immune activation and other toxic off-target effects.

[0007] This disclosure describes systems, devices, and methods that can achieve such highly specific therapeutic delivery within biological systems including, but not limited to, humans. Finally, this disclosure describes embodiments that include uses beyond traditional therapeutic and imaging applications. As described herein, specific embodiments of the subject matter described herein can be used as sensors, chemical synthesis tools, and delivery agents, and they can be used to transmit collected information to an external sensor or within a specific embodiment, with or without the need to introduce it within a biological or in vivo system. SUMMARY OF THE INVENTION

[0008] In one embodiment, the MNSDED includes a non-conductive housing having a surface and containing support logic circuitry. At least one conductive needle extending from the housing is provided, the needle being connected to the logic circuitry and a needle drive circuit. At least one conductive pad is located on the housing surface and is connected to the logic circuitry and at least one targeting agent capable of interacting with a target. In response to target interaction, the logic circuitry allows a voltage from the needle drive circuit to be applied to at least one needle.

[0009] In one embodiment, the non-conductive housing of the MNSDED has a major axis length from 100 nanometers to 500 micrometers and the logic circuitry is fabricated using 10 nm or smaller SIA transistor nodes.

[0010] In one embodiment, a biocompatible protective agent is provided around at least a portion of the surface of the non-conductive housing.

[0011] In one embodiment, each targeting agent is connected to a conductive linker that is attached to the conductive pad.

[0012] In one embodiment, at least one needle is capable of electrically interacting with the bilayer of a target cell attached to a targeting agent, and a sufficient voltage from a needle drive circuit can be used to facilitate at least one of ablation and electroporation of the target cell.

[0013] In one embodiment, the MNSDED includes a housing having a surface that contains support logic circuitry and a biocompatible protective agent surrounding at least a portion of the surface. At least one conductive pad is located on the housing surface and is connected to the logic circuitry and a targeting agent capable of interacting with a target. A reservoir is configured in the housing to accommodate at least one of a therapeutic agent and a diagnostic agent. In response to a target interaction with the targeting agent, the logic circuitry can initiate the release of at least one of the therapeutic agent and the diagnostic agent.

[0014] In one embodiment, a release barrier is provided that prevents the release of at least one of a therapeutic agent and a diagnostic agent until electrically activated by the logic circuitry. The release barrier can include a thermoresponsive material that dissolves when electrically activated by the logic circuitry. Alternatively or additionally, the release barrier can include a material that dissolves when placed near a target cell.

[0015] In one embodiment, the MNSDED includes a housing having a surface and containing support logic circuitry. A first conductive pad is located on the housing surface and is connected to the logic circuitry and a first conductive junction that is also attached to a targeting agent capable of interacting with a target. A second conductive pad is located on the housing surface and is connected to the logic circuitry and a second conductive junction that is attached to a reference agent that cannot interact with the target. In response to a target interaction with the targeting agent, the logic circuitry can initiate an effector mechanism.

[0016] In one embodiment, in response to a target interaction with the targeting agent, a readout amplifier connected to the first pad and the second pad is triggered, and a decision logic circuitry can facilitate at least one of ablation and electroporation of the target or initiate the release of at least one of a therapeutic agent and a diagnostic agent from the reservoir.

[0017] In one embodiment, the MNSDED includes a housing having a surface and containing support logic circuitry. A biocompatible protective agent is provided surrounding at least a portion of the surface. At least one conductive pad is located on the housing surface and is connected to the logic circuitry and a targeting agent capable of interacting with a target. A gasket is also provided surrounding the at least one conductive pad.

[0018] In one embodiment, when the targeting agent interacts with a target, the gasket facilitates the binding of the targeting agent. The gasket can be formed of molecules that extend longer than the molecules forming the biocompatible protective agent.

[0019] In one embodiment, the gasket boundary is printed on the MNSDED surface using biosafe lithography surrounding the at least one conductive pad.

[0020] In one embodiment, the MNSDED includes a non-conductive housing having a surface and containing support logic circuitry. The antenna is supported by the non-conductive housing and is connected to an energy harvesting circuit to power the logic circuitry and a voltage amplifier circuit. At least one conductive pad is located on the housing surface and is connected to the logic circuitry and a targeting agent capable of interacting with a target.

[0021] In one embodiment, the antenna further comprises multiple turns and multiple layers of coils. In addition to the energy harvesting circuit, one or more antennas may also be connected to a signaling circuit.

[0022] In one embodiment, the MNSDED includes a non-conductive housing having a surface and containing support logic circuitry. The antenna is supported by the non-conductive housing and is connected to a signaling circuit connected to the logic circuitry. At least one conductive pad is located on the housing surface and is connected to the logic circuitry and a targeting agent capable of interacting with a target.

[0023] In one embodiment, the signaling circuit is capable of enabling communication with an external transceiver. Alternatively or additionally, the signaling circuit may enable communication with another MNSDED.

[0024] In one embodiment, a method of treating disease-associated cells (DACs) includes introducing a plurality of MNSDEDs into a patient, each MNSDED having a non-conductive housing and at least one conductive needle extending from the housing, the housing having a surface and containing support logic circuitry. The needle is connected to the logic circuitry and a needle drive circuit, and further includes at least one conductive pad located on the housing surface and connected to the logic circuitry and a targeting agent capable of interacting with the DAC. In response to the interaction between the DAC and the MNSDED via the targeting agent, a voltage is applied from the needle drive circuit to at least one needle to ablate the DAC.

[0025] In one embodiment, a method of treating a pathogenic entity including disease-associated cells (DACs) and a pathogenic virus includes introducing a plurality of MNSDEDs into a patient, each MNSDED having a non-conductive housing having a surface and containing support logic circuitry. A reservoir is configured in the housing to contain at least one of a therapeutic agent and a diagnostic agent, the reservoir being connected to the logic circuitry. At least one conductive pad is located on the housing surface and is connected to the logic circuitry and a targeting agent capable of interacting with the pathogenic entity. In response to the interaction between the pathogenic entity and the MNSDED via the targeting agent, at least one of the therapeutic agent and the diagnostic agent can be released using the logic circuitry. Alternatively or additionally, applying a voltage from the needle drive circuit to at least one needle can perforate the DAC and allow at least one of the therapeutic agent and the diagnostic agent to enter the DAC.

[0026] In one embodiment, the testable MNSDED system includes a wafer that includes a plurality of casings configured to have a biocompatible coating attached thereto, each casing having a surface and including a substrate that supports sub-10 nm transistor logic circuitry. At least one conductive test pad is located on the surface and is connected to the logic circuitry. A plurality of individual test logic circuits may be positioned adjacent to each of the plurality of casings and are separately connected to respective conductive test pads.

[0027] In one embodiment, a therapeutic composition includes a pharmaceutically acceptable carrier and a plurality of MNSDEDs contained within the carrier. The MNSDEDs include non-conductive casings having surfaces and including support for logic circuitry. At least one conductive needle extends from the casing, the needle being connected to the logic circuitry and a needle driver circuit. At least one conductive pad is located on the surface of the casing and is connected to the logic circuitry and a targeting agent capable of interacting with a target.

[0028] In one embodiment, a therapeutic composition includes a pharmaceutically acceptable carrier and a plurality of MNSDEDs contained within the carrier. The MNSDEDs include casings having surfaces that include support for logic circuitry and having a biocompatible protective agent surrounding at least a portion of the surface. At least one conductive pad is located on the surface of the casing and is connected to the logic circuitry and a targeting agent capable of interacting with a target. A reservoir is configured within the casing to accommodate at least one of a therapeutic agent and a diagnostic agent.

[0029] In one embodiment, a method of treating disease-associated cells (DACs) includes introducing a plurality of MNSDEDs into a patient, each MNSDED having a non-conductive casing that has a surface and includes support for logic circuitry and further includes at least one conductive pad located on the surface of the casing that is connected to the logic circuitry and at least one targeting agent capable of interacting with a targeted DAC. After at least one targeting agent associates with a DAC, power is supplied to the MNSDED using an antenna connected to the logic circuitry. Using a sensing subsystem circuit within each MNSDED, a signal may be generated to indicate binding of at least one targeting agent to the targeted DAC. Then, based on one or more inputs or signals received from the sensing subsystem, a decision subsystem within each MNSDED may determine whether at least one targeting agent has bound to the targeted DAC. If the decision subsystem logic circuitry determines that the MNSDED is bound to a predetermined DAC, an actuator subsystem may be activated to direct current to at least one of the nano-needle and reservoir associated circuitry of each bound MNSDED.

[0030] In one embodiment, the MNSDED may include a non-conductive housing having a surface and containing a supporting logic circuit. At least one conductive pad is located on the housing surface and connected to the logic circuit. At least one targeting agent is capable of interacting with a target, and at least one conductive connector is provided to connect each targeting agent to the conductive pad. The MNSDED has a biocompatible protective agent surrounding at least a portion of the surface of the non-conductive housing, and the zeta potential of the MNSDED is between 0 mV and -20 mV. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] The embodiments will be easily understood by the following detailed description in conjunction with the accompanying drawings. For ease of description, the same reference numerals represent the same structural elements. The embodiments are shown in the accompanying drawings by way of example and not limitation. According to convention, the various features shown in the drawings are not drawn to scale, but are intended to emphasize specific features associated with the present invention. In addition, the size of the features and the thickness of the layers may be very different from the scales showing these features and layers.

[0032] Figure 1 An exemplary geometry of a MNSDED is shown.

[0033] Figure 2 An exemplary architecture and layout of MNSDED is shown.

[0034] Figure 3 An exemplary architecture and layout of a MNSDED sensing subsystem is shown.

[0035] Figure 4 An exemplary design and resistor-capacitor ("RC") equivalent circuit of an EEBI using DNA as the charge-carrying polymer is shown.

[0036] Figure 5 A general exemplary architecture and layout of an EEBI molecular fine-tuning pair including one reference EEBIP is shown.

[0037] Figure 6 A general exemplary architecture and layout of an EEBIMTP coupled to two reference EEBIs is shown.

[0038] Figure 7 An embodiment of an EEBI shim associated with a MNSDED sensing subsystem is shown.

[0039] Figure 8 A-8C shows the general exemplary architecture, layout, and equivalent circuits of one embodiment of a MNSDED implementation subsystem including electroporation / electroporation nanoneedles.

[0040] Figure 9Shows a general exemplary architecture and layout of an embodiment of an MNSDED implementation subsystem including a drug / imaging agent delivery component.

[0041] Figure 10 Is a flowchart of an exemplary process for fabricating one or more MNSDEDs.

[0042] Figure 11 Is a flowchart of an exemplary biocompatible lithography process for fabricating MNSDEDs.

[0043] Figure 12 A-12D shows the general exemplary steps of a biocompatible lithography and etch lift-off process for fabricating MNSDEDs.

[0044] Figure 13 Is a flowchart describing an exemplary treatment method using an MNSDED system.

[0045] Figure 14 A-14B shows an embodiment of a sense amplifier with an associated cross-coupled latch.

[0046] Figure 15 A-15B shows an alternative binding detection circuit scheme.

[0047] Figure 16 A-16B shows a schematic diagram of the MNSDED binding detection process.

[0048] Figure 17 Shows a representative diagram of a 3-EEBI pairing system.

[0049] Figure 18 Shows an example of binding mode determination using a simple logic circuit.

[0050] Figure 19 Shows an exploded view embodiment of a multi-layer coil radio frequency energy harvesting circuit (“RFEHC”) embedded in an MNSDED.

[0051] Figure 20 A-20C shows details of an embodiment of the RFEHC coil design.

[0052] Figure 21 A-21B shows an embodiment of a voltage multiplier circuit connected to an RFEHC LC (coil capacitor) circuit.

[0053] Figure 22 Shows an embodiment of a test method for MNSDEDs for high-volume production on a silicon wafer.

[0054] Figure 23A-23B shows a mode for testing and activating only those MNSDEDs that pass a set of logic tests. Detailed Description

[0055] Methods for fabricating and utilizing one or more micro / nano sensing-determining-actuating-devices (“MNSDEDs”) are described herein, as well as descriptions of the components that make up the MNSDED assembly, including: barcode targeting recognition (e.g., multiple unique targeting ligands that, upon binding to a DAC receptor, generate an electronic signal that activates or inactivates an effector mechanism), integrated circuits (e.g., NAND, current mirror, differential amplifier), effector mechanisms (e.g., electroporation nanoneedles, therapeutic release modules), power sources (e.g., inductive power transfer via an RF antenna), and other components that will be described in detail herein. The structures and manners in which these component MNSDEDs are interconnected and interoperate are also described herein.

[0056] Examples of specific MNSDED designs, related compositions, and delivery methods are also described herein, which can be used together for specific therapeutic and non-therapeutic applications. These designs, compositions, and methods will describe how an MNSDED (or a compound of interest contained within the MNSDED) can detect, neutralize, eliminate, or treat a pathogenic entity (including DAC and viruses) after administration to a subject. In addition, methods of using the MNSDED as a chemical sensor, chemical synthesis tool, and chemical delivery agent without introducing it into a biological or in vivo system will also be described.

[0057] Definitions:

[0058] Unless otherwise provided herein, the following terms and phrases have the meanings indicated below. The present disclosure may employ other terms and phrases that are not explicitly defined herein. These other terms and phrases shall have the meanings that would be ascribed to them by a person of ordinary skill in the art in the context of the present disclosure. In some cases, a term or phrase may be defined as singular or plural. In such cases, it should be understood that any term in the singular form may include its plural counterpart and vice versa, unless explicitly indicated to the contrary.

[0059] As used herein, unless the context clearly indicates otherwise, the singular forms “a,” “an,” and “the” include plural referents. For example, reference to “a substituent” includes a single substituent as well as two or more substituents, etc.

[0060] As used herein, the term "about", when used in reference to a range of values, a cutoff value, or a specific value, is used to indicate that the value may vary by up to 10% from the listed value. Since many of the values used herein are determined experimentally, those skilled in the art will appreciate that such determinations can and often will vary in different experiments. The values used herein should not be construed as being unduly limited by such inherent variations. The term "about" is used to encompass such variations up to or equal to 10%.

[0061] As used herein, the term "attach / attached / attachment" means to connect or join by chemical bonds, linkages, or forces to hold two or more components together.

[0062] As used herein, "for example", "such as", or "including" are intended to introduce examples that further illustrate a more general subject. Unless otherwise expressly indicated, such examples are provided only as an aid to understanding the embodiments illustrated in the present disclosure and are not meant to be limiting in any way. These phrases also do not denote any kind of preference for the embodiments disclosed.

[0063] As used herein, "polymer" means a macromolecule composed of repeating structural units ("-mers") that are typically connected by covalent chemical bonds.

[0064] Although the present invention admits of many different forms of embodiments, several specific embodiments are shown in the drawings and will be described in detail herein. It is to be understood that the present disclosure is to be considered as illustrative of the principles of the invention and is not intended to limit the invention to the embodiments shown.

[0065] Abbreviations

[0066] ARC - Anti-reflection coating.

[0067] BBB - Blood-brain barrier.

[0068] BMBC - Brain metastases of breast cancer.

[0069] BLM - Bilayer lipid membrane.

[0070] bps - Base pairs.

[0071] cDNA - Complementary DNA.

[0072] CDC - Clock distribution circuit.

[0073] CK - Clock signal.

[0074] CMOS - Complementary metal oxide semiconductor.

[0075] DAC - Disease-associated cells.

[0076] DNA - Deoxyribonucleic Acid.

[0077] EEBI - Environment - Electron - Binding - Interface.

[0078] EEBITA - EEBI Targeting Agent

[0079] EEBIG - EEBI Gasket.

[0080] EEBIL - EEBI Linker.

[0081] EEBIP - EEBI Pad.

[0082] ER - Estrogen Receptor.

[0083] FAB - Fragment Antigen Binding.

[0084] Gl - Gastrointestinal.

[0085] HER2 - Human Epidermal Growth Factor Receptor 2.

[0086] IC - Integrated Circuit.

[0087] kDa - Kilodalton.

[0088] MNSDED - Micro / Nano Sensing - Decision - Execution Device.

[0089] nm - Nanometer.

[0090] PEG - Polyethylene Glycol.

[0091] PEI - Polyethyleneimine.

[0092] PR - Photoresist.

[0093] RF - Radio Frequency.

[0094] RFEHC - Radio Frequency Energy Harvesting Circuit.

[0095] RNA - Ribonucleic Acid.

[0096] SDE - Sensing - Decision - Execution.

[0097] SED - Standard Spin - Exposure - Development.

[0098] siRNA - Small Interfering RNA.

[0099] ssDNA - Single - Stranded DNA.

[0100] UV / vis - Ultraviolet / Visible Light

[0101] μm - micrometer.

[0102] MNSDED Overview

[0103] The term "micro-nano-sensing-determining-enabling-device" ("MNSDED") as used herein refers to a composite nanorobotic structure of micrometer- to nanometer-scale dimensions. The portion of the MNSDED that contacts the external environment of the MNSDED is generally identified as the surface of the MNSDED. A typical MNSDED will be ovoid to cubic, although different morphologies and deviations from this basic geometry are possible and may be necessary depending on the specific requirements of the MNSDED. The MNSDED may have a major axis length that is typically in the range of about 100 nm to about 500 μm. Although typically ovoid to cubic, the embodiments of the MNSDED described herein will be described by spherical approximations having diameters from about 100 nm to about 500 μm. Factors driving these size scales and dimensions include, but are not limited to, size limitations imposed by factors such as in vivo mobility through tissues, target DAC size, and circuit functional requirements. The specific MNSDED diameter will depend at least in part on the composition of the MNSDED and the functional design that enables the MNSDED to be used for its intended use. For example, a MNSDED with limited functional capabilities (eg, comprising a few functional components as described herein) may have a diameter of about 1 μm or less, and a MNSDED with broad functional capabilities (eg, comprising a variety of functional components as described herein) may have a diameter of about 1 μm to about 500 μm.

[0104] More specifically, the MNSDED consists of transistors and associated integrated circuits ("ICs") contained within a "housing" of a dielectric or any non-conductive material, which may be coated with one or more biocompatible surface coatings or protective agents and one or more targeting ligands. The one or more biocompatible surface coatings or protective agents may include, but are not limited to, polyethylene glycol ("PEG"). The one or more targeting ligands may include, but are not limited to, antibodies. The MNSDED may include targeting ligands and conductive connectors capable of sensing a chemical or biological environment, a conductive surface component capable of ingesting information sensed from the environment and converting it into an electrical signal, an IC component capable of making a logical decision on an expected action based on the sensed environment or processing a logical decision, and a component capable of implementing one or more appropriate actions based on the decision.

[0105] MNSDED Dimensions: In some embodiments, the MNSDED may be small enough to enable relevant cell-MNSDED therapeutic interactions. Staphylococcus aureusThe cells have a diameter of ~1 μm, and thus the related MNSDEDs designed for therapy will be of a corresponding size of 1 μm in diameter. Accordingly, the MNSDEDs can incorporate full - function ICs with a standard Semiconductor Industry Association (“SIA”) node of no more than 10 nm, which is the size to achieve a sufficient number of components of related sense - decide - effectuate (“SDE”) transistors on an MNSDED effective for diagnosis or therapy. These ICs are designed and arranged to perform multiple functions, including supporting radio - frequency (“RF”) inductive power, employing differential amplifiers for signal detection in combination, applying input / output (“I / O”) logic for ligand - binding to process specific barcode - type patterns, and activating effector mechanisms, etc.

[0106] The systems, devices, and methods disclosed herein can rely in part on the cellular transport of the MNSDEDs to allow for cellular internalization and sub - cellular localization of the MNSDEDs and transport through various tissues by diffusion or convection. In some cases, the described systems, devices, and methods can utilize some form of endosome - or phagosome - facilitated transport through target cells. In designing MNSDEDs for therapeutic use, this and other practical aspects, such as affinity, can cause size to be a factor. For example, the affinity of the MSNDEDs can be adjusted such that they can irreversibly associate (bind) with DAC, but reversibly associate (bind and release) with healthy cells. Some MNSDED embodiments can be designed to be administered by stereotactic injection into dense tissues (e.g., some solid tumors, brain parenchyma), and the MNSDED size can be adjusted such that it can be transported through those tissues at a relatively high diffusion rate (e.g., with a diameter of 500 nm or less). In other embodiments, for example, the MNSDEDs can be designed to be orally administered and targeted to eliminate the Helicobacter pyloriinfected, and the size of the MNSDED can be adjusted such that it can bind maximally to the bacterium having a size diameter of about 1 to 4 μm and employ effector mechanisms against the bacterium, regardless of transport through dense tissue. In other embodiments, for example, the MNSDED can be intravenously injected and can be designed to freely flow through small blood vessels, including capillaries, requiring the MNSDED size to be no more than 4 to 8 μm in diameter. In other embodiments, for example, the MNSDED can be designed to bind to selected neurons in neural tissue to modulate neuronal activity by an electrically based effector mechanism, and the size of the MNSDED can be adjusted to be at the same size diameter level of about 4 to 100 μm of the neuronal cell body. In other embodiments, for example, the MNSDED can be designed to bind to and eliminate adipocytes after direct administration into adipose tissue, and the size of the MNSDED can be adjusted to be at the same size diameter level of adipocytes of about 100 - 500 μm.

[0107] MNSDED Zeta Potential: The systems, devices, and methods described herein can rely at least in part on the diffusion or convection of the MNSDED through various tissues such as plasma and whole blood. When designing the MNSDED to specifically interact with pathogenic entities including DACs and viruses, it is useful to consider aspects of the MNSDED that can inhibit non - specific interactions of the MNSDED with interstitial fluid components, plasma components, whole blood components, and bulk components of the tissue to which the MNSDED is administered. One such property is the ζ - potential of the MNSDED. Accordingly, the systems, devices, and methods described herein can be carried out using a targeted MNSDED comprising an MNSDED core as described herein, conjugated to any of the targeting agents described herein, and further conjugated to a biocompatible surface - protecting coating, wherein the ζ - potential of the MNSDED is slightly negative to near neutral. The ζ - potential of the MNSDED can vary depending on the material used to fabricate the MNSDED core, the linker used, the targeting agent, and the biocompatible surface - protecting coating or protecting agent (all described herein). In most cases, the ζ - potential of the targeted MNSDED will fall within the negative range. To reduce the likelihood of immunogenicity and reduce the likelihood of biofouling, the ζ - potential of the MNSDED used with the systems, devices, and methods described herein can be in the range of about 0 mV to about - 20 mV. Other desired properties of the MNSDED (including but not limited to surface charge and steric stability) can also vary depending on the specific application of interest.

[0108] MNSDED Detection Method: The size and properties of MNSDED can be detected or measured by techniques known in the art. Exemplary techniques for detecting particle size include, but are not limited to, dynamic light scattering (“DLS”) and nanoparticle tracking analysis (“NTA”), as well as various microscopes including transmission electron microscopy (“TEM”), scanning electron microscopy (“SEM”), and atomic force microscopy (“AFM”). Exemplary techniques for detecting particle morphology include, but are not limited to, TEM and AFM. Exemplary techniques for detecting the surface charge of MNSDED include, but are not limited to, the zeta potential method and NTA. Other techniques applicable to detecting other chemical properties include 1 H, 13 C, and other supported isotope nuclear magnetic resonance (“NMR”), UV / Vis and infrared / Raman spectroscopy, fluorescence spectroscopy and microscopy (when MNSDED is used in combination with a fluorescent label, or when the MNSDED transistor is activated and emits infrared light), and other techniques that will be apparent to one of ordinary skill in the art.

[0109] MNSDED Administration Mode: The MNSDED described herein can be administered orally in any acceptable dosage form, such as capsules, tablets, aqueous suspensions, solutions, etc. MNSDED can also be administered parenterally, including but not limited to subcutaneous, dermal, intranasal, intravenous, intraarterial, intramuscular, intraarticular, intrasynovial, intrasternal, intraocular, intrathecal, intracerebral, intravaginal, intraurethral, intrarectal, intrapulmonary, and intracranial injection, or by stereotactic assistance or infusion techniques. Alternatively, MNSDED can be administered directly into the tissue of interest, such as by direct injection into adipose tissue, or, for example, by intravenous or intraperitoneal injection, or, for example, by injection through a J-tube or G-J tube into the gastrointestinal (“GI”) system.

[0110] MNSDED Use Cases: Some embodiments can be designed for specific modes of administration for the corresponding target diseases, including but not limited to oral administration — for treating GI-related pathologies, including GI-related cancers (esophageal cancer, colon adenocarcinoma, etc.), GI-related infections (e.g., Clostridium difficile ( Clostridium difficile ) or Helicobacter pylori, and for systemic treatment of diseases where the MNSDED is designed to cross the blood - gut barrier; administration via a J - tube, G - J tube, or any other administration requiring a GI procedure - for treating GI tract disorders related to dysphagia or other oral ("PO") difficulties and for treating lower GI tract - related diseases, where it is advantageous to prevent MNSDED exposure to the upper GI tract (e.g., acidity problems); intravenous, intra - arterial, intracardiac, intravascular administration - for systemic treatment of various pathologies (especially for treating blood cancers and sepsis); intraperitoneal ("IP") administration - for treating IP - related conditions, including endometriosis, peritonitis, etc.; subcutaneous administration - for treating local pathologies, such as skin cancer "melanoma", and for treating regional - targeted therapeutic concentrations in local lymph for treating pathologies, including lymphoma, breast cancer, autoimmune diseases, and any other lymph - related pathologies; intramuscular administration - for musculoskeletal - related diseases (e.g., gene therapy for sarcoma, muscular dystrophy); intraocular administration - for retinoblastoma, or for reducing inflammatory responses in other ophthalmic diseases - e.g., retinitis pigmentosa; intrathecal or intracerebral administration - for treating central nervous system - related pathologies, including glioma and glioblastoma multiforme, reducing central nervous system ("CNS") - related inflammation and other diseases (e.g., multiple sclerosis, lysosomal storage diseases, including Batten disease), and directly treating neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, and Huntington's disease; intravaginal and intraurethral administration - for treating sexually transmitted diseases and gynecological infectious diseases and cancers (cervical cancer, vaginal cancer, bladder cancer, etc.), and for treating urinary tract infections; rectal administration - for treating lower GI and rectal - related diseases, including Clostridium difficile (Clostridium difficile) infections, Crohn's disease, and other inflammatory bowel diseases, and lower GI cancers, etc.; pulmonary administration - for treating pulmonary pathologies, including lung cancer, restrictive lung diseases, and infectious diseases; intranasal administration - for targeted CNS treatment and ear, nose, and throat ("ENT") - related pathologies; local administration (e.g., stereotactic injection into solid tumors for related treatment, or direct injection into adipose tissue for targeted reduction of adipocytes to treat obesity).

[0111] The MNSDED is not limited to in - vivo use cases and can be designed for any use case involving chemical or biological environmental sensing. The MNSDED can further be used for any purpose, system, device, and method described herein that employs logic - based decision - making and subsequent effector mechanisms.

[0112] MNSDED Structure and Method of Operation

[0113] The following will outline the main subsystems and components of MNSDED, associated circuitry and non-circuit-related components, the composition of the components of MNSDED, and descriptions of the ways in which these MNSDEDs can be synthesized, organized, specialized, managed, and used. Any examples described herein are not intended to limit the scope of the systems, devices, methods, or compositions, but rather to provide clarification of broader concepts through specific descriptions.

[0114] A typical MNSDED can be divided into multiple subsystems that interoperate with each other to achieve desired therapeutic, imaging, or chemical sensing results. Each of the subsystems described below can be modified with specific design features to achieve the desired end-use case or result. Generally, an MNSDED consists of at least the following subsystems: (1) a sensing subsystem; (2) a decision-making subsystem; (3) an implementation subsystem, and (4) an associated power and signaling subsystem.

[0115] MNSDED Sensing Subsystem:

[0116] The MNSDED sensing subsystem consists of at least the following components and materials: (1) an environment-electron-binding-interface targeting agent; (2) an environment-electron-binding-interface pad; (3) an environment-electron-binding-interface junction; (4) an environment-electron-binding-interface molecular tuning pair; (5) an environment-electron-binding-interface spacer; and (6) a biocompatible surface protection coating.

[0117] Environment - Electron - Binding - Interface ("EEBI") is a term that will be used herein to describe the assembly of the components and materials of MNSDED that are associated with, interact with, and sense the surrounding environment, and that, in some embodiments, transfer chemical or biochemical information from that environment to other components of the MNSDED by converting chemical or biochemical information into an electrical signal that can be processed by the MNSDED's electronic components. The term "sensing" refers to the specific process by which the MNSDED associates with the environment through specific binding interactions and converts chemical and biological information into electrical information that is propagated into the MNSDED circuitry.

[0118] EEBI Targeting Agent ("EEBITA") (also described herein as "target-binding molecule", "targeting ligand", "ligand", "targeting agent", or "capture agent") can act as an interface that specifically binds the MNSDED to any virus or cell type of interest (e.g., DAC), or to a specific chemical or biological entity of interest in the environment (the latter may not be cell-associated). The term "target" as used herein refers to any chemical entity or any biochemical entity that includes any biological system of interest, including organs, tissues, cells, or any part thereof, and can include In vitro or In vivoA biological system or any part thereof. The term "target" as used herein can also denote a non-biological entity of interest, such as a molecule or chemical not derived from a natural source. The terms "EEBITA" or "target-binding molecule" or "targeting agent" or "ligand" or "targeting ligand" or "capture agent" as used in the present disclosure denote any molecule that can be present on the surface of the MNSDED, the purpose of which is to engage a specific target, for example, by enabling the MNSDED itself to attach to a specific DAC receptor, and in particular a specific cell or chemical recognition. Examples of suitable EEBITA include, but are not limited to, vitamins (such as folic acid), proteins (such as transferrin and monoclonal antibodies), monosaccharides (such as galactose), peptides, oligomers, small molecules, amino acid-based biomolecules (such as DNA, cDNA, RNA), and polysaccharides. Specifically, the EEBITA can be an antibody directed against certain surface cell receptors (e.g., CD19 involved in non-Hodgkins lymphoma). The term "antibody" includes immunoglobulin molecules that react with a specific antigen. The term also includes genetically engineered forms, such as chimeric antibodies (e.g., humanized murine antibodies), heteroconjugate antibodies (e.g., bispecific antibodies), and recombinant single-chain variable fragments ("scFv"), disulfide-stabilized ("dsFv") Fv fragments, or pFv fragments. The term "antibody" also includes antigen-binding forms of antibodies known to those skilled in the art (e.g., Fab', F(ab')2, Fab, Fv, rlgG, and others) (see, for example, as disclosed in U.S. Patent No. 7,081,518, which is incorporated herein by reference in its entirety). An antibody that immunoreacts with or is "specific for" a specific antigen is a relative term. As provided in the "Definitions" herein, the term "connected, attached to" as used herein means to connect or join by a chemical bond, link, or force to hold two or more components together. This encompasses direct or indirect attachment, such as embodiments where the first compound binds directly to the second compound and embodiments where one or more intermediate compounds are disposed between the first compound and the second compound. The interaction of the EEBITA with a predetermined chemical or biomolecular target will initiate a series of events described herein, which will send an electrical signal to the MNSDED, the electrical signal having information specifying that a specific environmental, chemical, or biomolecular EEBITA binding event has occurred.

[0119] EEBITA can include ligands specific to particular chemical functional groups, chemical bonds, or combinations of chemical functional groups, spatial regions, or other chemically relevant entities where interactions may occur. EEBITA can also include ligands specific to receptors expressed on cancer cells, autoimmunity-related cells, infectious disease-related cells, or any other DACs. These receptors can be specific to membrane-associated receptors, internalized receptors, transcytotic receptors, receptors associated with lysosomal trafficking, or receptors associated with trafficking to any other subcellular compartment. For therapeutic applications, for example, EEBITA associated with a particular receptor can define the overall MNSDED design in terms of both the integrated circuit and the therapeutic mechanism. For example, EEBITA targeting receptors associated with cancer cells in solid tumors can be used in MNSDEDs having an activating component in an acidic environment. This can include an acid-soluble membrane for releasing therapeutic agents from the MNSDED drug compartment, which will be described in further detail herein.

[0120] Ideally, a targeted therapeutic agent will be able to reach multiple tissue targets to treat a wide range of diseases. For example, to enable an MNSDED to cross the blood-brain barrier ("BBB"), receptors that undergo transcytosis at the blood-brain barrier ("BBB") may be targeted. Chemical changes experienced during receptor-mediated transcytosis can also be exploited to increase the accumulation of MNSDEDs within anatomically distinct organs. Methods are disclosed herein for delivering an MNSDED to anatomically distinct and privileged regions (e.g., brain, retina, testis, prostate, etc.) of a subject that are separated from the circulation by most vascular barriers by administering to the subject an MNSDED having an EEBITA that is designed to target receptors that facilitate transport across such barriers. A variety of EEBITAs can be used to facilitate the delivery of the disclosed MNSDEDs. Brain-specific therapeutic strategies can include treating multiple sclerosis by eliminating disease-associated CD4 T cells or any other specific neuroinflammatory mechanism, or treating meningitis / encephalitis by eliminating bacterial DACs or viruses, or treating primary or secondary brain cancers such as gliomas, glioblastomas, or secondary breast cancers.

[0121] To perform the method, it may be advantageous to control the number and type of EEBITA attached to the MNSDED. The MNSDED can contain as few as one EEBITA, and they can contain as many as the physical space available on the surface of the MNSDED can accommodate. The number and type of EEBITA can vary depending on the type of MNSDED being delivered, the targeted pathogenic entity, or many other factors. The number and type of EEBITA can be altered to maximize the electronic signal transmitted to the IC of the MNSDED while ensuring that the MNSDED maintains biocompatible characteristics (e.g., biocompatible ζ potential, minimized immunogenic potential, minimized potential to bind to unintended targets with a corresponding maximized therapeutic effect and minimized off-target effects, etc.). The number and type of EEBITA can be adjusted such that the MNSDED binds irreversibly to pathogenic entities (including DAC and viruses) with a small binding dissociation constant (K d ) and binds reversibly or transiently to healthy cells with a large binding dissociation constant.

[0122] EEBI Pad ("EEBIP") Located on the surface of the MNSDED, and composed of a conductive material, and its surface can be chemically modified as described herein. These materials can include, but are not limited to, metals (such as gold, silver, copper, titanium, or other metal materials known to those skilled in the field of microelectronics manufacturing) and semiconductor materials (such as silicon), which are sufficiently doped by methods known to those skilled in the art to become conductive materials. These EEBIPs will be direct physical and electrical connections from the MNSDED to the surface of the MNSDED integrated circuit, which will be described in further detail herein. The EEBIP itself can be of any size limited by the size of the MNSDED surface and can be oval or linear with a critical dimension (such as diameter or major axis length) large enough to accommodate the hydrodynamic radius of the relevant EEBITA. For example, in some embodiments, the critical dimension length of the EEBIP can be not less than about 10 nm, which is sufficient to accommodate the EEBITA, which is an antibody with a hydrodynamic radius of about 10 nm. In other embodiments, the critical dimension length of the EEBIP can be not less than about 30 nm, which is sufficient to accommodate two EEBITAs, both of which are antibodies, each with a hydrodynamic radius of about 10 nm, and each with a nominal amount of spatial freedom allowing for spatial degrees of freedom. Each EEBIP will be large enough to contain or accommodate at least one EEBITA or targeting ligand, and the EEBIP size can vary to be large enough to contain or allow the utilization of as many EEBITAs as possible for the specified purpose of the MNSDED. The design of the EEBIP critical dimension will take into account the size of the relevant EEBITA and the linker. Ideally, the EEBIP critical dimension can be no more than about 10% larger than the hydrodynamic radius of the EEBITA and is typically smaller. In embodiments where the linker has a larger hydrodynamic radius than the EEBITA, the EEBIP critical dimension can ideally be no more than about 10% larger than the hydrodynamic radius of the linker and is typically smaller. The EEBIP will be designed to be as small as possible on the MNSDED surface to ensure that the electronic conduction signal through the linker is higher than the electronic signal-to-noise ratio, where the signal is related to the specific binding event of the EEBITA to its target, and the noise is related to non-specific binding events and background electrochemical potential and thermal fluctuations.

[0123] EEBITA can be directly conjugated to EEBIP, as described in further detail in the "EEBI Linker" section herein. For each unique EEBITA, there can be at least one EEBIP, and there can be at least one EEBITA included within or associated with each MNSDED, having as many EEBITAs as required in the functional design of the MNSDED, and which can be spatially placed on the surface of the MNSDED. For example, one embodiment can be designed to target CD19 and CD71, and thus this embodiment can include an EEBIP having a first unique EEBITA (which is an antibody targeting CD19) and a second EEBIP having a second unique EEBITA (which is an antibody targeting CD71), where there can be a number of CD19-related EEBIPs and CD71-related EEBIPs that can actually fit on the surface of the MNSDED.

[0124] Once all of the EEBITAs have been attached to the EEBIPs, a protecting agent small molecule (such as methoxy-(n-alkyl)-thiol or other molecules) can be linked to the remaining exposed regions of the EEBIPs for shielding against non-specific contamination by environmental molecules.

[0125] EEBI Adapter ("EEBIL") Can be used to attach the EEBITA to the EEBIP and carry any electron transfer or charge transfer caused by the specific binding event of the EEBITA to the EEBIP. The EEBIL can be composed of any combination of small molecules, biomolecules, compounds, polymers, moieties, or other entities that are bioconjugated or chemically linked, and they can be designed to be as small as possible (e.g., in embodiments using polymers as the EEBIL, a low "oligomer" length) to minimize the attenuation of charge transfer across the EEBIL and to increase the ability of the EEBIL to cause a voltage change at the EEBIP through charge transfer facilitated by the EEBIL when there is a specific EEBITA-target binding event.

[0126] As used herein, the term "polymer" refers to a macromolecule composed of repeating structural units ("-mers") typically connected by covalent chemical bonds. Suitable polymers can be linear and / or branched, and they can take the form of homopolymers or copolymers. If copolymers are used, the copolymers can be random copolymers or branched copolymers. Exemplary polymers are water-dispersible, particularly water-soluble polymers. For example, suitable polymers include, but are not limited to, polysaccharides, polyesters, polyamides, polyethers, polycarbonates, polyacrylates, and the like. Ideally, the polymer will maintain high stability under the conditions seen in a biological environment (e.g., a temperature of 37 °C, a pH in the range of 2 to 7.4) and under MNSDED manufacturing conditions (e.g., a temperature in the range of 25 °C to 100 °C, and other conditions as will be further described herein). For therapeutic and / or pharmaceutical uses and applications, the polymer ideally has low toxicity and cytotoxicity characteristics. Suitable polymers include those having a molecular weight of about 10,000 Da and below.

[0127] Ideally, the EEBIL selected to be used as the medium can transfer electrons or alter the electric field (i.e., electrically bind information) along the entire length of the EEBIL, independent of the Debye screening length and independent of the solvent-related bilayer capacitance. In some physiological and biochemical environments where MNSDED may be expected to function, the Debye screening induced by high (physiological) salt concentrations is widely accepted as overly attenuating the electric field effects, otherwise the EEBIP would experience electric field effects when a specific binding event causes an electric field change that would induce a voltage change in the EEBIP. Due to the Debye screening problem under physiological conditions, some embodiments may require the EEBIL to carry a charge to overcome this Debye screening barrier, which would render the field effect modulation of the potential at the EEBIP ineffective in many in vivo applications. The EEBIL molecules carrying a charge can be polymers (e.g., polyethyleneimine ("PEI") or the conductive polymer polypyrrole) or biomolecules (e.g., DNA wires), or any other chemical or biological composition that can facilitate charge transfer or electron transfer. In such cases, the length of the EEBIL can be adjusted to balance the stability of the linker with the ability of the linker to transfer charge to the EEBIP or to effect a local change in the potential at the EEBIP. For example, in some embodiments, the EEBIL will be composed of DNA, and the size of the EEBIL will be determined by the total number of DNA base pairs ("bps"). In this embodiment, the total number of bps will be adjusted to ensure sufficient binding strength between complementary DNA strands (e.g., the annealing temperature (T 退火), a physiological temperature of at least greater than 37°C), while ensuring that there are few enough base pairs to ensure sufficient conductivity to cause charge transfer to the EEBIP without significant attenuation of the charge transfer, which charge transfer can occur entropy - related or over large distances by other mechanisms (e.g., according to Slinker, typically a DNA wire can carry a current with less than 100 complementary bps over up to 34 nm, with an inverse relationship between conductivity and the total number of base pairs - doi: 10.1038 / nchem.982). Ideally, in this embodiment, the length of the EEBIL will be defined by the minimum number of bps to achieve (T 退火 >T 生理 ). The condition is that the number of bps is low enough to achieve an electrical signal that results in a reference - to - target EEBIP potential difference of at least in the picovolt range, as will be further described herein. T 退火 Under additional constraints such that it must be less than any temperature that would denature, degrade, or otherwise impair the EEBITA (e.g., many antibodies degrade at greater than 70°C and many proteins denature at greater than 41°C). Under these constraints, in some embodiments, the length of the EEBIL will not be greater than about 10 nm (or about 30 bps) and may be smaller. Additionally, under these constraints, the selection of the base pair sequence will be optimized such that T 退火 is able to achieve the lowest number of bps and possibly the shortest EEBIL.

[0128] Attachment of EEBIL to EEBIP can be facilitated by a functional handle placed on the surface of EEBIP. The functional handle can be any chemical functional group described herein and desirably has a small length (e.g., a small molecule of less than 400 Da and a chemical length of no more than about 2 nm) and specific chemical functionality that promotes the different conjugate reactions described herein. An exemplary functional handle can be achieved by reacting thio-(n-alkyl)-amine with the metal EEBIP surface to produce a metal-thiolate ligand bond that will promote a primary amine functional handle near the metal EEBIP surface. This primary amine functional handle can then undergo further conjugation chemistry capable of binding a larger EEBIL molecule (e.g., a complementary functionalized DNA strand). For example, in some embodiments, the above-described EEBIP-related primary amine functional handle can react with an N-hydroxysuccinimide-ssDNA molecule (or other nucleobase-binding molecule) to form an amide covalent bond between the cDNA molecule and EEBIP. Additionally, as needed, a functional handle can be placed on the EEBIL itself or on EEBITA to facilitate the binding of EEBIL to EEBIP or EEBIL to EEBITA. For example, a protein EEBITA can be thiol-functionalized by reacting the primary amine of a protein lysine group with N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP) and then treating with dithiothreitol (DTT) for reduction. The resulting thiol group on the protein can be used as a functional handle to react with maleimide-ssDNA to form ssDNA-protein (or EEBIL-EEBITA).

[0129] Various functional handles can facilitate the linking of EEBIL to EEBIP and EEBIL to EEBITA, and these functional handles can be used to perform the disclosed methods. In some embodiments, the functional handle can individually or combinatorially include one or more of the following: amine, thiol, carboxylate, hydroxyl, aldehyde, ketone, diazo, aryl azide, haloaryl azide, benzophenone, anthraquinone, alkyne, biotin, polypeptide, or any other chemically reactive group. Those skilled in the art will understand that other functional handles can be employed in a similar manner to effect the association between EEBITA and EEBIP. These functional handles can be covalently bound to EEBIP through relevant chemical reactions guided by photolithographic patterning, as will be described in further detail herein. Desirably, the functional handle will be as small as possible, and in applications where MNSDED is used in a physiological environment, they are typically less than about 2 nm in length to overcome Debye shielding for charge transfer from the charge-carrying polymer, through the functional handle, and to EEBIP. In most applications, the length of the functional handle can be no more than about 1 nm. The functional handle can be used in conjugation reactions with chemical moieties that can chemically interact with the functional handle with high specificity and selectivity and facilitate the binding of EEBIP to EEBIL or EEBIL to EEBITA. For example, an amine functional handle can react with isothiocyanates, isocyanates, acyl azides, NHS esters, sulfonyl chlorides, aldehydes, glyoxals, epoxides, ethylene oxides, carbonates, arylating agents, imidates, carbodiimides, acid anhydrides, fluorophenyl esters, hydroxymethylphosphine derivatives, and guanidyl formates. Thiols can react with haloacetyl derivatives, haloalkane derivatives, maleimides, aziridines, propiolyl derivatives, arylating agents, thio-disulfide exchangers (pyridyl disulfide, TNB-thiol), vinyl sulfone derivatives, metal-thiol coordination bonds, and cisplatin derivatives. Carboxylates can react with diazoalkanes, diazoacetyl compounds, carbonyldiimides, and carbodiimides. Hydroxyls can react with epoxides, ethylene oxides, carbonyldiimides, N,N'-disuccinimidyl carbonate, N-hydroxysuccinimidyl chloroformate, periodates, alkyl halogens, and isocyanates. Aldehydes and ketones can react with hydrazine derivatives or can undergo Schiff base formation, reductive amination, or Mannich condensation. Diazo can react with active hydrogen sites on aromatic rings. Aryl azides and haloaryl azides can react with active sites associated with N-H and C-H. Benzophenone can react with C-H active sites. Anthraquinone can react with specific radical pairs. Alkynes can react with azide functional groups. Biotin can react with avidin.

[0130] Based on the foregoing disclosure, one of ordinary skill in the art will understand that a variety of functional handle molecules can be used with the EEBIP, EEBIL, and EEBITA such that a particular EEBITA can be associated with a corresponding EEBIP. Such alternatives, which are obvious to one of ordinary skill in the art in view of the present disclosure, are considered to be within the scope of the present disclosure. In addition, a functional handle described as being associated with an EEBIP can be associated with an EEBITA and vice versa.

[0131] EEBIL itself can be a capture agent, such as an oligonucleotide stem-loop receptor, which undergoes a conformational change upon binding to a target and which juxtaposes the phosphate backbone charge of the EEBIP to less than the Debye length (<1 nm), which can create a specific voltage change within the EEBIP (as in Nakatsuka, N. et al. (2018) "Aptamer-field-effect transistors overcome Debye length limitations for small-molecule sensing" Science 362, 319 - 324, which is incorporated herein by reference in its entirety).

[0132] EEBI Molecular Tuning Pair ("EEBIMTP") A term for the entire assembly that describes a reference molecule (described herein) conjugated to a reference-associated EEBIP ("reference EEBIP") by the same linker and functional handle assembly (same EEBIL assembly) used to conjugate an EEBITA to a target-associated EEBIP ("target EEBIP"). The voltage of the reference EEBIP is compared to the voltage of the target EEBIP by a differential amplifier. In the case of a non-specific binding event occurring on the target EEBIP, the EEBIMTP can function as a voltage and capacitance comparator, with the voltage and capacitance differences determined by a differential amplifier, as further described herein. These EEBIMTPs can ensure that, upon non-specific binding of the target EEBIP, there will be a negligible voltage difference (enabled by voltage comparison by the differential amplifier) between the target EEBIP and the corresponding reference EEBIP. Thus, the EEBIMTP can ensure that a latch (described herein) will not trigger a non-specific interaction event associated with the target EEBIP, such as a false positive. The EEBIMTP can be calibrated by identifying a "reference" molecule or biomolecule and conjugating it to the reference EEBIP. The "reference" molecule or biomolecule can be adjusted or selected such that when it non-specifically interacts with the environment (including non-specific cellular interactions), it will affect a nearly equal voltage change in the reference EEBIP as will occur in the target EEBIP when the EEBITA participates in non-specific interactions. These EEBIMTPs can be configured and designed based on the specific application of the MNSDED and can be configured and designed through experimental development, where the configuration and design of the EEBIMTP are further informed by the specific environment in which the MNSDED is to be administered. For example, in some embodiments, if the MNSDED is administered intravenously to a human, the EEBIMTP "reference" molecule can potentially be based on an albumin molecule (to mimic the immediate surrounding environment of the MNSDED), or in the case where an antibody is the MNSDED targeting ligand, the EEBIMTP "reference" molecule can be based on an antibody specific for an antigen not found in humans. The "reference" molecule or biomolecule to be conjugated to the reference EEBIP can include, but is not limited to, polymers (e.g., PEG, PEI, etc.), glycoproteins, proteins (e.g., antibodies, antibody fragments, albumin), nucleic acids, lipids (e.g., palmitic acid), small molecules, and any other molecule or biomolecule that achieves the purpose and function of the EEBIMTP.

[0133] In some embodiments, for each target EEBIP, there will be at least one reference EEBIP. The reference EEBIP corresponding to the target EEBIP can be located adjacent to the associated target EEBIP and can be coupled to the target EEBIP through a differential amplifier described in further detail herein. In some embodiments, each target EEBIP can have two or more reference EEBIPs. In the case where there is more than one reference EEBIP associated with a target EEBIP, in addition to each reference EEBIP being coupled to the target EEBIP through an additional dedicated differential amplifier, these reference EEBIPs can also be coupled to each other through a differential amplifier (one differential amplifier between each pair of reference EEBIPs). In such an embodiment, these reference EEBIPs will be used to minimize or possibly eliminate any false positive identification events of the target EEBIP by comparing the voltage difference between the reference EEBIPs with the voltage difference between the reference EEBIP and the target EEBIP. By way of illustration, for example, in the case where there is a difference of less than a few percent between the voltage differences of these reference EEBIPs and the target EEBIP, the latch (described herein) will not be triggered and the state of the MNSDED effector subsystem will remain unchanged.

[0134] A readout amplifier will be incorporated into an integrated circuit to amplify the voltage difference between any two EEBIPs (e.g., the voltage difference between a target EEBIP and a reference EEBIP, or the voltage difference between two reference EEBIPs). When the EEBITA on an EEBIP attaches to its target (e.g., CD19-specific IgG attached to CD19 on a lymphoma cell), the electric field distribution around the target EEBIP changes with the conductance of the associated electrical signal through its corresponding EEBIL, which is different from the electric field distribution around the reference EEBIP and the conductance of the associated electrical signal through the associated EEBIL with its non-specific background voltage. The resulting difference in target-to-reference EEBIP voltage can be detected and amplified from the initial voltage difference and converted to complementary metal oxide semiconductor ("CMOS") logic levels for use by the MNSDED control circuit. The readout amplifier output can undergo further signal conditioning (e.g., amplification, filtering) and then be latched into a storage circuit. A regenerative readout amplifier incorporating clocked cross-coupled pairs with precharge, measurement, and latch phases can be used to boost the small voltage difference between the reference and target EEBIPs to CMOS logic levels. These clocked readout amplifiers will output multiple measurements based on one or more clock frequencies. If these kinds of readout amplifiers are employed, the threshold number of "positive" measurements per time interval can be counted by a counting / threshold circuit that triggers downstream logic when counting a predetermined number of "binding events" or multiple measurements of the same binding event. The counting circuit can then send a logic signal to a decision circuit. This data or logic signal constitutes an input for determining whether the EEBITA has bound to its specific target and becomes an input to the MNSDED control logic circuit.

[0135] EEBI Spacer ("EEBIG") will be used to enhance the barrier created by cell surface - associated glycocalyx / biomolecules and MNSDED - related surface protectants (e.g., PEG). When the relevant EEBITA of the target EEBIP has bound to its specific target, the spacer can help isolate each target EEBIP from the surrounding environment. This can help buffer the ion concentration near the EEBITA - target binding site, thereby reducing / stabilizing the Debye screening length and increasing / stabilizing the capacitive (field - effect) sensitivity of the EEBIP to target - specific binding events (through field - effect charge transfer through the non - conductive component of the EEBIL, e.g., in some embodiments, the EEBIL can be conjugated to the functional handle of the EEBIP, and in some embodiments, the EEBIL can be conjugated to a portion of the EEBITA). The EEBIG can be a barrier around the EEBIP and can be composed of any molecule, including but not limited to polymers, lipids, proteins, oligomers, nucleic acids, or small molecules. Using the biocompatible lithography techniques further described herein, these molecules can be specifically positioned around the EEBIP. The molecules and the MNSDED - related linkers can be designed such that they are long enough compared to the biocompatible surface - protecting coating to create a robust seal but not so long as to unduly prevent the diffusion of cell - associated receptors to the targeted ligand through the fluid - mosaic diffusion process. For example, in one embodiment, the molecules of the EEBIG and the MNSDED - related linkers can be not less than about 10 nm but not more than about 20 nm in length to exhibit the above - described sealing and diffusion characteristics. The molecules can be tuned to be "ionic" sponges and contain molecular structures that act as ionic sponges (e.g., 1,8 - bis(dimethylamino) - naphthalene, and others known to those skilled in the art), which can additionally reduce the ionic strength of the internal environment of the EEBIG itself. The molecules can be adjusted to be hydrophobic to repel any aqueous associations or hydrophilic entities, which can also reduce the ionic strength of the internal environment of the EEBIG. This can also significantly reduce the Debye screening and increase the capacitive and field - effect sensitivity of the EEBIP.

[0136] Biocompatible Surface Protection Coating: A protective coating containing any molecule (including but not limited to biomolecules, lipids, components typically constituting vesicles, microsomes or biological membranes (such as red blood cell-mimicking vesicles), and polymers such as PEG, PEI, etc.) can be added to the surface of MNSDED to ensure that the molecule is biocompatible, non-immunogenic, and stable in physiological solutions. Ideally, the protective coating will be thin enough to ensure that the targeting ligand is not masked (e.g., 500 Da to 2 kDa), but will be large enough (e.g., 2 kDa to 10 kDa) to ensure sufficient protection of MNSDED from physiological solutions with high solute concentrations, which are known to reduce the Leonard-Jones potential between individual MNSDEDs and allow Van der Waals interactions between surfaces, and the Van der Waals interactions can cause MNSDED aggregation and biofouling. The protective layer can additionally contribute to any field effects generated on the EEBIP by the EEBITA-target interaction, which can increase charge transfer, electron transfer, or field effects, and the transfer of potential in the MNSDED resulting from the specific binding of EEBITA to the target receptor. The EEBIP field effect can be assisted by increasing the Debye length, which is achieved by reducing the Debye shielding by adding such a biocompatible surface protective coating adjacent to the EEBIP, which can reduce the effective solute concentration near the EEBIP or further inhibit the penetration of ionic entities into the EEBIG.

[0137] MNSDED decision-making subsystem

[0138] The MNSDED decision-making subsystem consists of at least the following components and materials: (1) a differential amplifier / comparator / latch circuit facilitating the detection of the initial binding of EEBITA to the target, and (2) a CMOS logic circuit that converts the binding pattern into a logical decision to exert or not exert an effect on the bound cell.

[0139] Can be employed within MNSDED Decision Logic Circuit To facilitate the logical processing of combinatorial states, such as when a single or a group of different EEBITAs have bound (or not bound) their respective targets. The term "barcode type identification" as used herein refers to a logic-based decision made by the MNSDED integrated circuit that one or more EEBITAs have bound (or not bound) their respective targets, and further to initiate effector mechanisms based on a predefined combination of triggered (and in some cases non-triggered) latches that are specific to a set of EEBITA binding events (e.g., for positive feedback) and the possible lack of EEBITA binding events (for negative feedback control).

[0140] These decisions to activate the implementation subsystem can be facilitated by a logic circuit acting as a control component that will dictate the output response for each bound or unbound EEBIP pair (target and reference EEBIP pair) or bound or unbound EEBIP grouping (target EEBIP and its associated one or more reference EEBIPs). If required by the MNSDED design, the logic circuit (in the example described herein) can meet the conditions of Boolean function integrity and can include any combination of logic gates (e.g., a combination of CMOS logic gates that can enable any Boolean function), and the final logic circuit design can be dictated by the MNSDED application decision requirements. These logic circuits can obtain data derived from each EEBIP pair or grouping (i.e., target EEBIP and its associated one or more reference EEBIPs) and combine all these signals in a suitable logic / judgment circuit to determine whether the MNSDED should activate the implementation subsystem. Logic circuits for implementing "AND", "OR", "NOT" Boolean logic are well known in the art, as is the practice of combining them into more complex logic decision "logic blocks" to determine whether a data pattern matches one decision criterion or multiple decision criteria.

[0141] For example, for a therapeutic indication, the decision subsystem can be designed such that an EEBITA binding event will contribute to an "on" or "therapeutic active state" through an "AND" or "OR" circuit or logic, in which the therapeutic implementation subsystem circuit described below is activated. The logic circuit can also be designed such that an EEBITA that binds to a self-antigen or "safe-disarmed antigen" can be associated with a "NOT" gate (or a circuit equivalent to a "NOT" gate) and can effectively disable the therapeutic agency circuit. Since as many unique EEBITAs as possible will be placed on the MNSDED surface (limited by the size of the MNSDED and the size of the target), the MNSDED has the ability to effectively sense its unique binding to the DAC by barcode type identification, with both positive (disease-related) and negative (non-disease-related) inputs.

[0142] MNSDED Implementation Subsystem

[0143] The MNSDED implementation subsystem can consist of at least one or more of the following components and materials: (1) electroporation / electroshock nanoneedles; and / or (2) targeted drug / imaging agent delivery components.

[0144] Electroporation / Electropuncture Needle It can be coupled to a voltage amplifier circuit, which will increase the voltage generated by the radio frequency energy harvesting circuit (described further herein), enabling the MNSDED to perform electroshock or electroporation on the combined DAC. The electroshock or electroporation action can be carried out through a series of at least two nanoneedles - one serving as the anode needle and the second serving as the cathode needle (or ground potential). These nanoneedles can protrude from the surface of the MNSDED, and they can be geographically placed on the MNSDED surface as close as possible to the EEBIP to allow the target EEBIP and their associated reference EEBIP to be as close as possible to the nanoneedles, and also to ensure that the nanoneedles are as close as possible to the DAC bilayer lipid membrane ("BLM"), while minimizing the likelihood of the nanoneedles ablating or perforating adjacent healthy cells of the MNSDED. The position of the nanoneedle pair on the MNSDED surface can be far enough from the EEBIP to reduce the electric field and local Joule heat associated with the current between the nanoneedles, which may cause the EEBITA to separate from the target or damage the EEBITA and the EEBIMTP itself. Ideally, the nanoneedles are typically no less than about 100 nm from the nearest EEBIP, although the exact distance of the nanoneedles to the nearest EEBIP will be specified by the functional MNSDED design and the total MNSDED size, as well as the total area available for accommodating all the EEBIPs, nanoneedles, and other MNSDED surface-related components described herein. The nanoneedles can be spaced apart such that the distance between their bases is sufficient to prevent electrical short circuits between the bases, which is specified by the optical proximity correction and etching profile limitations associated with the manufacturing method described further herein. In one embodiment, for example, the edge of the base of one nanoneedle is no less than about 10 nm from the edge of the base of the second nanoneedle.

[0145] These nanoneedles can be designed to penetrate the BLM when the MNSDED attaches to the cell of interest through specific EEBITA binding. These nanoneedles can protrude through the biocompatible surface protective coating of the MNSDED by a length sufficient to ensure that the needles penetrate the DAC BLM, after which they can generate an electric field of sufficient strength to perforate or ablate (destroy) the BLM when specifically binding to the DAC. Additionally, in some embodiments, the nanoneedles can be designed such that they are not overly long to minimize the likelihood of their protruding through the BLM when the MNSDED is non-specifically and transiently associated with non-target cells through any non-specific (e.g., electrostatic) interactions. For example, in one embodiment, the length of the nanoneedles should be no less than about 10 nm and no greater than about 500 nm. These nanoneedles can be designed to include a biocompatible protective agent conjugated to the surface while leaving the tips of the needles exposed to achieve optimal reduced immunogenicity and retained electrical conductivity, especially when the MNSDED effector is activated, leaving a free path for electrons to transfer from the anode to the cathode (ground potential). In other embodiments, the nanoneedles can be designed such that they do not penetrate the BLM when the MNSDED binds to the DAC through one or more EEBITAs, while they can still generate an electric field of sufficient strength to ablate or perforate the BLM.

[0146] When the MNSDED effector is not activated, the ideal nanoneedles will have a high aspect ratio with nanoscale dimensions to facilitate penetration of the BLM by the nanoneedles without damaging the attached cells, which would indicate non-specific interactions with non-DACs. For example, the nanoneedles can have a diameter of less than about 100 nm and a length of about 100 nm - 1 μm. Ideally, the nanoneedles are mechanically robust in an aqueous environment, and the surface of the nanoneedles can be functionalized similar to the biocompatible surface protective coating described above. See Yum, K., Wang, N., and Yu, M-F. (2010) "Nanoneedle: A multifunctional tool for biological studies in living cells". Nanoscale ", 2, 363 - 372, which is hereby incorporated by reference in its entirety.

[0147] When the decision - making subsystem is activated, a voltage sufficient to effect electroporation (transient perforation of the DAC BLM) or ablation (i.e., large - scale structural breakdown of the BLM and associated irreversible membrane rupture) can be applied via the nanopin. As described further herein, the voltage sufficient for ablation will be used in applications intended to eliminate a pathogenic entity (e.g., DAC), while the voltage sufficient for perforation will be used in embodiments intended to treat, image, detect, or modify a pathogenic entity by introducing a therapeutic or imaging agent into the pathogenic entity via transient pores generated in the BLM (see Teissie, J. and Tsong, T. Y. (1981) "Electric field induced transient pores in phospholipid bilayer vesicles" Biochemistry Biophys. J. 20, 1548 - 1554, the entire content of which is incorporated herein by reference). In one embodiment intended for irreversible membrane breakdown, a voltage of not less than about 80 mV and up to about 5 V will be applied (irreversible membrane breakdown is expected at 1 V to 5 V), and this voltage will be applied for not less than about 1 millisecond and up to several minutes. In embodiments for electroporation only, a voltage less than about 1 V will be applied for a shorter time, which is suitable for perforation or reversible membrane rupture (e.g., it can be less than about 80 mV). Such electroporation can be coupled with small molecules (e.g., L - DOPA) or macromolecules (e.g., siRNA, CRISPR / CAS9), or with the release of a therapeutic or imaging agent that can facilitate the rapid entry of these entities into only the targeted cells. This delivery is described in further detail herein.

[0148] These nanoneedles can be fabricated from any materials known to those skilled in the field of microelectronic manufacturing, including but not limited to silicon, polysilicon, doped polysilicon, metals (such as gold, copper, titanium), hydrogels, polymers, sugars, and carbon nanotubes. The nanoneedles can be fabricated by a process of material growth or deposition (e.g., depositing silicon on the MNSDED surface), followed by appropriate doping to produce a conductive material, followed by lithography of the area where the nanoneedles reside, and then etching away all materials leaving only the nanoneedles as surface features on the MNSDED. The etching process can be adjusted such that the nanoneedles will be confined within the above dimensions and such that it will have a high aspect ratio with a reasonable tip (e.g., tapering down to <10 nm tip from about 100 nm). The sharper the nanoneedle tip, the greater the voltage gradient at the tip, thus maximizing the electroablation and electroporation capabilities of the nanoneedles. The nanoneedles can similarly be made of metals (such as copper, gold, aluminum, titanium), or polysilicon-based nanoneedles can be coated with gold or other conductive thin films (such as TiN) that can reduce potential immunogenicity. The metal nanoneedles can be produced by any of the following techniques, including but not limited to (1) metal coating of non-metal nanoneedles (e.g., via atomic layer deposition (“ALD”)), (2) metal filling within lithographically printed holes or vias, (3) lithography-assisted etching of a metal layer (e.g., reactive ion etching of Ti). The metals used in these techniques can include but are not limited to gold, silver, aluminum and its oxides, copper, tungsten, platinum, ruthenium, nickel, titanium, and titanium nitride. Other nanoneedle fabrication techniques can be used as long as the resulting nanoneedles have the structural and performance characteristics described herein. Ideally, the generation of the nanoneedles should be completed prior to any lithography-assisted chemical or biological conjugation with the EEBI as described herein.

[0149] Targeted Drug / Imaging Agent Delivery Component: The disclosed systems, devices, and methods can be used to deliver one or more therapeutic agents and / or imaging agents to pathogenic entities (including DACs and viruses) by loading the therapeutic agent or imaging agent into one or more compartments or “reservoirs” of the MNSDED prior to administration of the MNSDED to a subject. After delivery of the loaded MNSDED, the EEBITA can facilitate the binding of the MNSDED to the target DAC (e.g., tumor cells). After the EEBITA binds to the target on the DAC, the MNSDED can decide whether to activate the therapeutic agent or imaging agent delivery component through the above-described decision subsystem logic circuit. In the case where the implementation subsystem is activated by the decision subsystem, the implementation subsystem can release the therapeutic agent or imaging agent contained within the reservoir through the devices and methods described herein.

[0150] Each reservoir can be a compartment within the MSNDED, sealed from the external environment by a barrier made of a metal or polymer material or a barrier made of any other chemical or biological material. The barrier can be opened, dissolved, or degraded (when determined by the logic circuit) to release a therapeutic or imaging agent through any barrier removal mechanism appropriately corresponding to the barrier material itself, which can include but is not limited to pH-catalyzed chemical removal, electric field or current-assisted removal, or thermoelectric-based temperature-assisted removal.

[0151] For example, in some embodiments, the barrier can be composed of a thermoresponsive material (e.g., poly(N-isopropylacrylamide), PNIPAM), which is stable at physiological temperature but can dissolve when heated to its melting temperature, and the melting temperature is facilitated by passing an electric current through a corresponding resistive heating element contained within the MNSDED or by Joule heating of the environment immediately surrounding the barrier (e.g., interstitial fluid, plasma, etc.). In such embodiments, the thermoresponsive material is ideally stable at physiological temperature (about 37 °C) and will have a melting transition temperature slightly above this physiological temperature (e.g., for poly(γ-2-(2-(2-methoxyethoxy)-ethoxy)ethoxy-ε-caprolactone)- b -poly(γ-octyloxy-ε-caprolactone), the transition temperature is 40 °C, or for poly(lactic-co-glycolic acid) (“PLGA”) the glass transition temperature is 45 °C). In other embodiments, the barrier can be composed of the polymer polypyrrole, which can be removed by electrochemical redox and current. In other embodiments, the barrier can be composed of phospholipids or PEG, which are perforated when stimulated by an electric current or potential promoted by the MNSDED IC. In other embodiments, the barrier can be composed of a polymer or other chemical or biological entity that is stable at physiological pH (7.4) but whose stability decreases at acidic pH, such that the membrane breaks down when the MNSDED is exposed to acidic pH (e.g., if the MNSDED is intended to enter an acidic compartment after administration, such as a lysosomal compartment, tumor stroma, or gastric mucosa). In these embodiments, the barrier material will ideally be stable at physiological pH (7.4) and will decompose at a slightly lower (or higher) pH (e.g., PEG-poly(β-amino-ester) is stable at pH 7.4 but decomposes at pH 6.4 - 6.8).

[0152] The MNSDED reservoir can be constructed by lithography and etching methods and techniques known to those skilled in the field of microelectronic manufacturing. Specifically, before functionalizing the EEBIP with the above-mentioned functional handle and EEBIL, the final layer of the MNSDED can be modified with trenches, holes, or any voids of any shape achieved by pre-designed lithographic patterning and post-lithographic etching. In some embodiments, the final layer of the MNSDED can consist of an interlayer dielectric material (not related to EEBIP), which is any dielectric material (such as an oxide, typically an oxide of silicon or a non-conductive oxide of another material). This oxide can be etched with a corresponding etching technique that has high selectivity for the dielectric material, which is well-known to those skilled in the field of microelectronic manufacturing. Ideally, this etching will be facilitated by lithography, which protects the EEBIP from the etching process and enables geometries and dimensions that maximize the etched surface area and etching depth. In some embodiments, the MSNDED surface will be designed such that the reservoir can have sufficient dimensions to encapsulate large chemical or biological molecules (such as CAS9 and large genetic material sequences such as DNA, for delivery to and incorporation into the DAC genome), which can be, for example, an ellipsoidal inlet to the reservoir with a major axis diameter of no less than about 20 nm and a depth of no less than about 40 nm. Once the reservoir has been etched, biocompatible lithographic functionalization of the EEBIP (as described herein) can be performed, followed by an etch stripping process (as described herein). In some embodiments, before adding a linker that utilizes a conjugation chemical between the linker and the functional handle associated with the EEBIP, the MNSDED device can be filled with a therapeutic agent or imaging agent by immersing it in a solution of the therapeutic agent or imaging agent having a predetermined concentration (e.g., a concentration close to the solubility limit of the therapeutic agent or imaging agent), which also contains a material that forms a membrane, the concentration of which is such that the membrane can self-assemble on the outer opening of the reservoir (at an appropriate concentration, pH, and temperature). Once the MNSDED has had sufficient time to encapsulate the therapeutic agent or imaging agent in the reservoir, the MNSDED can be separated from the solution (which contains the therapeutic agent or imaging agent and the membrane material) by continuous centrifugation and removal of the supernatant, or by another filtration method known to those skilled in the field of nanoparticle synthesis. Then, the MNSDED can be treated with a conjugation chemical that further couples any linker and EEBITA, as further described herein.

[0153] After delivering the MNSDED with a filled reservoir to a subject, EEBITA can facilitate the association of the MNSDED with the DAC. In some embodiments, EEBITA can be designed to target the DAC receptor such that after the MNSDED binds to the DAC, the MNSDED can be internalized by the DAC through the process of intracellular phagocytosis or endocytosis. The membrane of the loaded compartment that internalizes the MNSDED can be destabilized and the loaded agent secreted in a phagocytic or endosomal / lysosomal environment (e.g., by pH-catalyzed lysis) by the methods described above, thereby delivering the agent to the target DAC. Alternatively, after the EEBITA binds to a specific pattern of DAC receptors, the MNSDED decision-making subsystem can provide a corresponding output that activates the effector subsystem, and after power-up, the membrane of the loaded compartment can be removed by the aforementioned methods and the loaded agent can be locally secreted to the DAC, near the surface of the DAC. In a variant of this embodiment, upon activation of the effector subsystem, consistent with the lysis of the compartment membrane, the nanoneedles can have an applied voltage such that the DAC BLM is transiently perforated and the released therapeutic or imaging agent can diffuse through the pores in the BLM.

[0154] In some embodiments, the method can be implemented to deliver chemotherapeutic agents that can be used to treat cancer. For example, in some embodiments, the described systems, devices, and methods can be implemented to deliver chemotherapeutic agents such as irinotecan or 5-fluorouracil (5-FU), which can be used to treat cancer such as gastric adenocarcinoma. Other agents for treating cancer can also be delivered to cancer cells via the systems, devices, and methods. In other embodiments, radiotherapy can be specifically delivered to tumor cells by this method. In some embodiments, the method can be implemented to deliver antibiotics that can be used to treat infectious diseases. In some embodiments, the method can be implemented to deliver immunosuppressants, which can be used to treat autoimmune diseases. Imaging agents that may not easily reach the DAC alone can also be delivered using the method. For example, in some embodiments, the method can be used to deliver MNSDED carrying the imaging agent 64 Cu to the DAC tissue of a subject to allow imaging, i.e., by positron emission tomography ("PET"). Additionally, the method can be used to deliver one or more therapeutic agents, imaging agents, or a combination of both therapeutic and imaging agents to the pathogenic entities of a subject, including DACs and viruses.

[0155] MNSDED Power and Signaling Subsystem

[0156] The MNSDED can include a power source and can incorporate a signaling subsystem that consists of at least the following components and materials: (1) a radio frequency energy harvesting circuit, (2) a power regulation and distribution circuit; and (3) a signaling circuit (depending on the use case).

[0157] Radio Frequency Energy Harvesting Circuit ("RFEHC"): The MNSDED sensing, decision-making, and implementation subsystem will require a power supply voltage suitable for the CMOS technology employed in its construction. These voltages can be provided by collecting radio frequency ("RF") energy received through an antenna, which will then be rectified and multiplied to a voltage sufficient to enable stable analog and logic circuit biasing and operation. The RF energy harvesting circuit ("RFEHC") can include an inductor or one or more antennas, matching / tuning circuits, rectifying and voltage multiplying circuits, and voltage regulation circuits. An example of such an approximate schematic and layout is shown in Figures 19 to 21 below.

[0158] Depending on one or more RF frequencies employed by the RFEHC, the antenna structure can include a simple planar or multi-layer planar spiral loop with multiple turns, or more complex dipoles or other structures. Antenna efficiency (radiation resistance and internal losses) can be characterized by the number of turns, geometry, and layout, as well as the antenna impedance constrained by the geometry.

[0159] Depending on the antenna impedance (Z ant ), the RF voltage from the antenna can be passed through an impedance transformation network to ensure optimal recovery of RF energy from the antenna to the rectifying / multiplying circuit. Depending on the impedance requirements of the subsequent rectifying / multiplying circuit, an impedance transformation of a larger apparent real Z may be required (see Figure 22 ).

[0160] Rectifying and voltage multiplying circuits are well known in the field of RF energy harvesting. A simple example is a low-pass T-network, which, as is known to those skilled in the art, boosts the impedance of a planar spiral from a few mOhm to a larger value.

[0161] Figure 21 A simple 8-stage rectifying and multiplying (voltage multiplier) circuit is shown. The required DC voltage will depend on the required CMOS operating characteristics, process technology, and total circuit power requirements. For example, an MNSDED that does not need to transmit energy can be done with a smaller power budget. The number of multiplier stages can be determined by the available ambient HF incident power, CMOS transistor and capacitor characteristics, and connecting parasitics.

[0162] Power Regulation and Distribution Circuit: The rectified output of the RFEHC can be fed "as is" (without regulation) to the logic and communication circuits of the MNSDED, as described herein, or it can be further regulated using active or passive components, such as additional filter capacitors, diodes, and / or series-pass regulator circuits. Given the small circuit load of the MNSDED at semiconductor nodes not exceeding 7 nm, additional regulation may not be necessary, but can be easily implemented by those familiar with conventional CMOS power delivery systems using passive components (e.g., filter capacitors) or actively (e.g., mirror circuits, series-pass regulation).

[0163] MNSDED Signaling from External Circuit: In addition to powering the MNSDED inductively from an external RF power source, the RFEHC inductor can also be used to receive information from an external signaling source such as an RF transmitter.

[0164] Antenna Structure and Matching Circuit: The MNSDED can also employ the RFEHC antenna or a separate antenna structure to receive RF signals from an external circuit. The receiving circuit can include an antenna structure, a matching circuit, and a receiver circuit. For high-frequency (“HF”) frequencies (30 MHz and below), the small size of the MNSDED and its immersion in a weakly conductive medium (such as blood, lymph, or extracellular or intracellular fluid or cytosol) precludes more elaborate antenna structures other than a simple loop, since the MNSDED is much smaller than most radio-frequency wavelengths of interest. For THz to near-infrared (“NIR”) frequencies, it is possible to fabricate more complex antennas, including but not limited to microstrip patch antennas, micro-bowtie antennas, or similar structures. Both the simple loop and the more complex structures can use standard T, Pi, or other multi-element matching structures for impedance transformation.

[0165] Signaling Circuit: The MNSDED can also employ the same antenna, a portion of the RFEHC antenna, or a separate antenna structure to transmit signals to an external circuit. The external signaling circuit can include an antenna structure, a matching circuit such as for a receiver, and can in fact employ the same antenna structure incorporating a duplexer / receiver protection circuit. The MNSDEDs can communicate with each other or with an external transceiver using far-field or near-field (e.g., inductive coupling) methods. For both methods, the required transmitter circuits can be similar in nature. Signals encoded by phase / frequency modulation (e.g., quadrature phase shift keying (“QPSK”)) techniques or simple pulse code modulation (“PCM”) can be demodulated from the carrier frequency using standard FM or AM techniques. Using code division multiple access (“CDMA”) techniques, more complex channel sharing is also possible, along with an increase in receiver complexity and thus DC power requirements. These techniques are well established in wireless communication and are familiar to those skilled in the art of radio-frequency communication.

[0166] Via the signaling circuit, communication can occur between each of the MNSDEDs and from the MNSDED to an external circuit. The external device can compile information from the MNSDEDs and use that information to perform calculations to generate clinically relevant insights or guide treatment actions, either by a physician or by real-time feedback to the MNSDED via the signaling circuit. In some applications, the MNSDEDs can be designed to be sub-specialized. For example, some MNSDEDs can only electro-ablate the DAC, while other MNSDEDs can electroporate at the DAC and locally release a therapeutic agent, and when the DAC is conjugated, yet another MNSDED can release an imaging agent, and yet another MNSDED can call upon other MNSDEDs, attract other MNSDEDs or act as a signal relay path from an external or internal node to one MNSDED. Each of these sub-specialized MNSDEDs can communicate with each other via the signaling circuit or with the external circuits described herein.

[0167] MNSDED Fabrication, Testing, Activation, and Etch Release

[0168] The methods disclosed herein also outline key steps in the MNSDED fabrication and testing processes. The methods outlined herein include: an etch "release" technique that involves etching the polysilicon layer under the MNSDEDs on the wafer in a biocompatible manner and releasing the MNSDEDs from the wafer into solution; electrically testing the MNSDEDs on the wafer with a voltage sufficient to render all MNSDEDs with malfunctioning logic circuits inoperative ("e-testing"), leaving all properly functioning MNSDEDs intact; functionalizing each EEBIP with a unique EEBITA via a biocompatible lithography technique that allows for EEBIP-specific functional handles and EEBIP-specific EEBILs, specifically by using EEBITA and linkers that are not interfered with by the biocompatible lithography process (e.g., pH-appropriate and chemically benign trilayers, resists, topcoats, developers, and other solvents, and appropriate materials to maintain a biologically relevant temperature during the spin exposure development ("SED") process). The fabrication method also includes selective polymer attachment (by methods including directed self-assembly).

[0169] Lithography Technology for Biocompatible Lithography to Produce EEBI: The methods provided herein include biocompatible lithography techniques. One such technique utilizes ssDNA attached to EEBIP, which can then bind to a complementary ssDNA sequence that itself binds to a targeting ligand. For example, a thiol-R-thiol bifunctional linker can be added to the wafer surface in an aqueous solution to functionalize the surface of the metallic EEBIP with thiol groups. Once the EEBIP has been functionalized with a thiol-functional handle, a selected EEBIP can be covalently linked to a selected DNA sequence, which can be achieved by conjugating the ssDNA to the thiol groups associated with the EEBIP, the conjugation being via a complementary functional handle to a thiol (such as ssDNA-maleimide) that is itself covalently coupled to the ssDNA.

[0170] To achieve this EEBIP-ssDNA sequence specificity, during the process of lithographic patterning by mask-based patterning, a pre-designed set of EEBIPs can be kept exposed while all other EEBIPs are covered with three layers (described herein), which will remain non-reactive with the specific ssDNA sequence. The lithographic patterning can employ three layers (a bioprotective layer (such as SU-8, PEG, etc.), an anti-reflection coating ("ARC"), and a photoresist ("PR") layer (such as ideally but not limited to known biocompatible resists, such as SU-8), which may contain a top coat (such as if an immersion lithography process is indicated)), materials well-known to those skilled in the lithography art, and the lithographic patterning can be accomplished by standard spin-exposure development ("SED") processes well-known to those skilled in the lithography art themselves.

[0171] The subsequently exposed EEBIPs can be treated with ssDNA oligomers (such as cDNA as described herein) containing a maleimide group at one end, which will react with the thiol-functional handles previously conjugated to the EEBIP surface to create a thioester bond between the EEBIP and the ssDNA oligomer. Once the ssDNA oligomers are linked to the EEBIP in a valence and covalent manner, the three layers can be removed by conventional methods (including but not limited to methods using biocompatible solvents or other known solvents that do not disrupt the ssDNA, such as, for example, N-methyl-2-pyrrolidone (NMP)), and then by heating to 80 °C to remove the solvent. The biocompatible protective layer deposition, ARC deposition, PR deposition, SED, ssDNA conjugation, and PR / ARC removal processes can be repeated as needed multiple times in order to add ssDNA oligomers of different and potentially unique sequences to strategically and lithographically exposed EEBIPs. Once all of the ssDNA oligomers have been added to the appropriate EEBIPs, the MNSDED can be released from the wafer surface by an etch-based lift-off process, using methods that will be described herein.

[0172] Some embodiments may require ssDNA as the EEBIL because, for SED-related short time scales on the order of minutes, ssDNA is thermally stable at SED-related temperatures up to but not exceeding approximately 90 °C (as demonstrated by conventional PCR protocols that take ssDNA to 90 °C for several minutes), and ssDNA is acid-stable with respect to the pH and time exposure conditions experienced during the SED process. A bioprotective layer that can be added around the ssDNA and prior to ARC and SED will ideally maintain a pH-neutral environment and, in addition to standard bioprotective materials such as PEG, it can include materials that act as proton sponges (such as, including but not limited to amine-containing polymers such as poly-L-lysine, poly(β-amino esters), poly-(γ-benzyl-L-glutamate), or polyhistidine).

[0173] "Lift-off" after etching; the resulting colloidal suspension of MNSDED can be reacted with EEBITA and a reference ligand that has itself been bio-conjugated to the complementary sequence of DNA via a bifunctional linker (as described previously herein). For example, in one embodiment, IgG with reduced thiols can be used as the EEBITA and the reference ligand and can react with DNA-maleimide to form a thioester bond between the Fc portion of the antibody and the cDNA fragment. The pre-prepared EEBITA-cDNA and reference-ligand-cDNA can be stirred together in solution with MNSDED. The subsequent conjugation of the reference ligand and EEBITA to their expected EEBIPs on the MNSDED will occur via the complementary ssDNA (cDNA) sequences, where the annealing of the complementary sequences of the ssDNA can be thermally controlled such that the annealing occurs at no less than 5 °C below the melting temperature (T m ) of the complementary strand of the ssDNA. For example, in some embodiments, the cDNA sequence can be designed such that T m is 55 °C, and the reaction mixture can be incubated between approximately 50 °C and 55 °C, where 50 °C does not exceed T mBelow 5 °C, another limitation is that the reaction temperature does not exceed the temperature at which the ligands (EEBITA and reference ligands) start to degrade or denature (e.g., an antibody can start to denature at about 55 °C). The EEBITA-MNSDED coupling reaction can be carried out in an aqueous environment with a mild buffer solution (e.g., < 50 mM sodium phosphate, pH 7.4). Once all the complementary EEBITA and reference ligand-bound ssDNA anneals with the EEBIP-bound ssDNA, the remaining surface of the metal can be backfilled (e.g., with a methoxy-(n-alkyl)-thiol monolayer), which will further protect the EEBIP from non-specific binding by environmental components (e.g., plasma proteins, interstitial proteins, etc.). Through this process, "EEBIP X" will be associated with "ssDNA sequence X", which will anneal with "complementary ssDNA associated with sequence X", which binds to "EEBITA X", where "X" refers to the expected association with a specific "receptor X" on "DAC X", such that binding to "receptor X" will cause "EEBIP X" to trigger its associated latch, and the associated latch is associated with the logic output of the IC to appropriately respond to the binding of receptor X.

[0174] Additionally, in other embodiments, a preselected group of EEBIPs can be kept exposed during a lithography process by mask-based patterning as described above. The exposed EEBIPs can be functionalized with a chemical functional handle (e.g., thiol) as described above, and EEBIL (ideally a conductive polymer as described above) will be coupled to the functional handle (e.g., through a maleimide-linker bioconjugation reaction), where EEBIL itself is bifunctional, having separate and distinct functional handles (e.g., NHS-ester, primary amine, or other chemically reactive moieties) at its opposite ends. This separate functional handle (e.g., NHS-ester) is unique to the complementary reactive group located on EEBITA (e.g., a primary amine on EEBITA, which can couple with the NHS-ester on EEBIL). In a manner similar to the complementary ssDNA strategy, the biocompatibility protectant layer deposition, ARC deposition, PR deposition, SED, and PR / ARC removal processes can be repeated as many times as needed to add unique functional handles to the lithographically exposed EEBIPs. Then, each unique functional handle can be coupled to its corresponding EEBITA through its complementary reactive group as described herein. Combinations of functional handles and their complementary reactive groups are described herein.

[0175] Each MNSDED can be manufactured in large quantities on a silicon wafer (e.g., a 300 mm wafer) (e.g., 100 million or more MNSDEDs can be manufactured on a 300 mm wafer). To ensure that all MNSDEDs that are peeled off are fully functional or disabled, the following electrical test ("e-test") and enable / disable circuits and methods can be employed. In the case of employing such an e-test, these MNSDEDs will be non-functional until the device is tested and confirmed to be functional. On the wafer, multiple MNSDEDs can be placed in a constellation around a signal classification test chip. The test chip can be probed with a standard classification probe card. The probe card can power the test chip, and the test chip can then perform a set of pre-programmed tests on each MNSDED (e.g., excite only for the combined EEBIP pattern of 011101), and can report back the success or failure of each MNSDED attached to it. The test chip can input a test signal into a test communication circuit (described below) within the MNSDED, and if the MNSDED logic circuit passes the test mode set, the test chip can blow the internal fuse of the MNSDED, thereby enabling the energy harvesting circuit of the MNSDED. In this embodiment, the MNSDED will be non-functional unless the internal fuse has been disabled.

[0176] This test can help identify faulty MNSDEDs. For example, some EEBIPs can be designed to identify tissues that the MNSDED is not intended to affect. The test can ensure that only those patterns associated with the target tissue rather than off-target tissues will activate and trigger the MNSDED logic circuit. For example, due to cardiotoxicity, HER2 / Neu-positive gastric adenocarcinoma cannot be treated with trastuzumab and anthraquinone simultaneously because HER2 is present on cardiomyocytes. The MNSDED strategy will allow the treatment of HER2-positive gastric adenocarcinoma when the MNSDED is programmed to be inactive when identifying cardiomyocyte-specific antigens. During the MNSDED test and activation process, those MNSDEDs with faulty circuits associated with identifying cardiomyocyte-related antigens will fail the test, so the internal fuse will not be disabled, and the MNSDED will be non-functional or unavailable for its intended purpose. This will increase the safety of the MNSDED by enabling MNSDEDs that do not have off-target (i.e., unintended) effects.

[0177] The MNSDED wafer-level test circuit within each MNSDED can be attached to the test chip via wire bonds (providing at least power, ground, transmit, receive, and fuse functions). Transmit and receive can be at least two-wire communication buses to each chip, much like a serial bus. Each MNSDED can have an RFEHC as part of the power supply. The RFEHC can feed a power regulation circuit, which can then provide DC power to the rest of the MNSDED. Each MNSDED can be fabricated to have a fusible link between the output of the power regulation circuit and ground potential, effectively shorting the power supply to the chip until the fuse is blown during testing. This helps prevent the production of chips with faulty logic.

[0178] MNSDED wafers can be fabricated as an array of test interface chips ("TICs") that have multiple MNSDEDs placed in a second array around the TIC. Each wafer can have thousands of these TICs. Each TIC can have approximately hundreds to thousands of MNSDEDs attached via wire bonds.

[0179] Each MNSDED can contain logic circuitry associated with a fusible connection. The TIC can send a set of patterns to the logic circuitry. If the logic circuitry sends back an appropriate response via the serial bus, the fusible link will be blown, enabling the MNSDED power subsystem.

[0180] Once the fabrication and testing of the device are complete, each individual device can be protected with a biocompatible protective surface coating material (e.g., high molecular weight PEG coupled via a functional handle, parylene coupled to the MNSDED surface via CVD, etc.), and the MNSDED can be released from the substrate via standard etch and lift-off techniques. One such exemplary technique includes the following steps: (1) forming trenches around the device via an etching process during fabrication and before adding EEBITA to EEBIP; (2) then lithographically applying the protective biocompatible material to the surface of the wafer such that the material only covers the surface of the MNSDED; (3) then subjecting the wafer surface to an etchant material (e.g., tetramethylammonium hydroxide ("TMAH")), which will then be selective for the underlying substrate beneath the MNSDED such that the etchant will undercut and release each MNSDED while not reacting with the protective layer; (4) washing the etchant material from the released MNSDED via successive centrifugation, supernatant removal, and aqueous buffer addition steps; (5) storing the MNSDED under inert conditions after freeze-drying for further processing.

[0181] Exemplary MNSDED Usage and Operation

[0182] The therapeutically effective amounts of MNSDED will be determined experimentally through preclinical and clinical trials, and these amounts may vary depending on the specific disease targeted, the degree of disease progression, and other characteristics specific to the subject, where the subject can be any animal, including but not limited to humans, mice, rats, hamsters, guinea pigs, rabbits, cats, dogs, monkeys, donkeys, cows, horses, pigs, birds, reptiles, etc. In some embodiments, at least one of the MNSDED may be administered to the subject at a daily dose, where the dose range is proportional to the body weight or surface area of the subject. The dose administered to the subject can also be measured based on the total amount of at least one of the MNSDED administered per day. For example, in some embodiments, at least one type or class of the MNSDED may be administered to the subject at multiple doses per day, as may be required to achieve a therapeutically relevant dose (e.g., multiple administrations may be required to achieve a clinically relevant outcome, such as multiple rectal administrations to eliminate specific pathogenic bacteria in the gut microbiota). In other embodiments, for example, a dose may be administered to the subject at least once a year, which may be required to ensure that the active and stable MNSDED is located in the body space to detect recurrence of cancer cells and also signal to an extracorporeal device that the cancer has been detected. In other embodiments, for example, the method may be performed to administer the MNSDED described herein to the subject weekly, bi-weekly, monthly, bi-monthly, semi-annually, or annually, as needed according to a clinical or preclinical development plan. Treatment may be started with a smaller dose than the optimal dose and the dose may then be increased during the course of treatment until the optimal effect in a medical setting is achieved.

[0183] The methods provided herein can be carried out by administering MNSDED suspended in a pharmaceutically acceptable carrier to a subject. Such carrier compositions can be used, for example, to administer to a patient to treat cancer, autoimmune diseases, and infectious diseases. The carrier compositions can be formulated into any of the various formulations known and suitable in the art. In some embodiments, the carrier composition is an aqueous formulation. An aqueous solution can be prepared by mixing MNSDED in water or a suitable physiological buffer and optionally adding suitable colorants, preservatives, stabilizers, and thickening agents as needed. An aqueous suspension can also be prepared by dispersing MNSDED in water or a physiological buffer with a viscous substance, such as natural or synthetic gums, resins, methylcellulose, sodium carboxymethylcellulose, and other well-known suspending agents.

[0184] Also included are liquid formulations and formulations in solid form, which can be converted to liquid formulations shortly before use. Such liquids include solutions, suspensions, syrups, slurries, and emulsions. The liquid formulations can be prepared with pharmaceutically acceptable additives by conventional methods, such as suspending agents (e.g., sorbitol syrup, cellulose derivatives, or hydrogenated edible fats or oils); emulsifying agents (e.g., lecithin or gum arabic); non-aqueous vehicles (e.g., almond oil, oily esters, or fractionated vegetable oils); and preservatives (e.g., methyl paraben or propyl paraben or sorbic acid). In addition to the active agent, these formulations can also contain colorants, flavoring agents, stabilizers, buffers, artificial and natural sweeteners, dispersing agents, thickening agents, solubilizing agents, etc. The carrier composition can be in powder or lyophilized form for combination with a suitable vehicle such as sterile water, physiological buffer, saline solution, or alcohol before use.

[0185] The carrier composition can be formulated for injection into a subject. For injection, the carrier composition can be formulated as an aqueous solution, such as water or alcohol, or a physiologically compatible buffer, such as Hanks' solution, Ringer's solution, or saline buffer solution. The solution can contain one or more formulations, such as suspending agents, stabilizers, or dispersing agents. Injectable formulations can also be prepared as solid-form formulations, which are intended to be converted to a liquid-form formulation suitable for injection shortly before use, such as by formulating with a suitable vehicle (e.g., sterile water, saline solution, or alcohol) before use.

[0186] The carrier composition can be formulated into a final oral solution, capsule, pill, spray, syrup, or any other standard oral formulation known to those skilled in the art. Additionally, the carrier composition can be formulated for inhalation via a nebulizer or atomizer.

[0187] Examples of Usage Methods:

[0188] The above examples of specific embodiments are illustrative only and are not intended to be limiting in any way, and are described herein to demonstrate methods of administering MNSDED and the manner in which MNSDED operates when administered to a subject. This example will describe how to use MNSDED to treat a subject diagnosed with solitary brain metastases of well-defined primary breast cancer ("BMBC").

[0189] Step 1: One or more MNSDEDs can be designed and formulated such that in a reconstituted sterile solution of cerebrospinal fluid, e.g., calcium chloride dihydrate (1.0 mg); magnesium chloride hexahydrate (0.8 mg); potassium chloride (1.1 mg); sodium chloride (44.0 mg); disodium hydrogen phosphate, heptahydrate (0.55 mg); sodium dihydrogen phosphate, monohydrate (0.4 mg); and 5 mL of water for injection. For safe intracerebral or intraventricular administration, the pH of the solution can be between about 6.0 and about 7.0. The MNSDED itself can be designed with an EEBITA that recognizes receptors identified as specific for BMBC cells (e.g., estrogen receptor (“ER”), human epidermal growth factor receptor (“HER2”), and other receptors), and the MNSDED can be designed such that it electroablates any cell identified (by a pre-determined EEBITA binding event) as having said specific BMBC cell surface receptor expressed on its membrane surface.

[0190] Step 2: One or more MNSDEDs can be administered by direct, stereotactic injection into the intraventricular, intracerebral, or intratumoral space. Such administration requires aseptic technique and should be administered by a physician in the field of intracerebral or intraventricular administration. The MNSDED can be administered directly into the cerebrospinal fluid, brain parenchyma, or the tumor itself through a surgically implanted reservoir and catheter (intraventricular device) by single injection or infusion. The recommended dose will be determined by one or more clinical safety, toxicity, and efficacy trials, which in turn will be guided by non-clinical good laboratory practice safety and toxicity studies. The MNSDED can be administered by infusing the above intraventricular electrolytes with MNSDED and pre-treating with any one of an antihistamine, an antipyretic, or a corticosteroid. Each infusion will be formulated in a vial sufficient for one administration, and a filtration set with a filter not less than 0.5 μm larger than the characteristic size of the MNSDED will be used for complete infusion (e.g., for a MNSDED of size 1.5 μm, a 2 μm filter may be required). After completion of the vial infusion, the infusion line, the infusion needle, and the intraventricular access device can be flushed with intraventricular electrolytes in order to administer the dose completely.

[0191] Step 3 (Sensing) ：In the case where one or more MNSDEDs are directly administered into the intratumoral space, the MNSDEDs will be allowed to traverse the intratumoral space (e.g., by diffusion), or their transport through the tumor can be assisted by additional infusion of intraventricular electrolytes (i.e., assisted convective transport). To allow the MNSDEDs sufficient time to interact with tumor cells and bind to the target BMBC cell surface receptors, the MNSDEDs can be allowed to traverse the tumor space for a minimum time span determined to be therapeutically effective or longer. During this period, one or more EEBITAs on each MNSDED will attach to the associated BMBC cell surface receptors. For example, EEBITA targeting ER will bind to the BMBC cell surface ER. In this case, the EEBITA-ER association can promote the binding of HER2 to the HER2-EEBITA associated with the MNSDED, which is achieved by immobilizing the MNSDED on the BMBC surface, followed by diffusion of HER2 towards the associated EEBITA on the MNSDED through the fluid mosaic diffusion process.

[0192] Step 4 (Power Supply) ： Once the MNSDEDs have had sufficient time to associate with the BMBCs, the subject can be placed in an RF field (e.g., an MRI machine, where the MRI itself is a source of RF), and under the supervision of a physician with knowledge of chemotherapy, the subject can be exposed to electromagnetic ("EM") radiation in the radiofrequency range of the EM spectrum, which has sufficient intensity to induce power in each MNSDED. The above-described one or more RFEHCs within each MNSDED "collect" or mediate the EM radiation. In this case, the RF power and / or frequency can be tuned or modulated such that each MNSDED has sufficient power to electroablate the BMBC cells, as will be described in subsequent steps.

[0193] Step 5 (Decision) ：In this embodiment, the MNSDED will be designed such that it will activate the electro-ablation mechanism if and only if the MNSDED EEBITA associated with ER, HER2, and any other predetermined BMBC-specific surface receptor is bound. In any of these binding events, the reference-target EEBIP voltage difference will be amplified by a differential amplifier circuit, which is itself powered by the above-described RF inductive power mechanism. If the readout amplifier detects a voltage difference above a predetermined threshold, it will trigger an integrator latch, which will send an electronic signal to the MNSDED logic circuit that a DAC binding event has been achieved. This logic circuit will gate the power supply to the implementation subsystem, and in this case, the subsystem will initiate the electro-ablation event that will be described in subsequent steps. If the logic circuit has a predefined combination of inputs from the latches, the predefined combination of which is associated with the recognition of the binding of all required BMBC-specific receptors, the logic circuit will trigger the implementation subsystem.

[0194] Step 6 (Implementation): Once the implementation subsystem is activated, it will be powered by the above-described RF inductive power mechanism, which is assisted by a voltage amplifier and a capacitor that will release a conventional current through the MNSDED nanoneedle cathode. This cathode will be designed to deliver current across the BMBC bilayer lipid membrane, and the MNSDED circuit and RF power will be designed such that the current will be sufficient in terms of voltage and duration to irreversibly ablate the bilayer lipid membrane. The MNSDED nanoneedle anode will be located on the MNSDED such that it is as far as possible from the cathode needle on the MNSDED and such that it can complete the electronic circuit initiated by the cathode nanoneedle and conducted through the BMBC bilayer lipid membrane and the cytosol.

[0195] Step 7 (End of Treatment) ： Once sufficient RF power has been administered to the subject to ablate all BMBC cells that have been bound by the MNSDED, the patient can be removed from the MRI machine, and the intraventricular device can be left in place for additional infusion of the MNSDED, or the device can be removed if a physician in the oncology field determines that the intraventricular device is no longer applicable. Multiple doses of RF may be required within a treatment course to optimally ablate all MNSDED-bound BMBC, and additionally, multiple infusions followed by multiple doses of RF may be required to ablate the entire tumor and eliminate all associated BMBC cells.

[0196] Step 8 (Monitoring): Once the MNSDED administration process is completed, the patient can be monitored for a predetermined period (possibly several months to a year) to determine if there is a recurrence of BMBC in the CNS or any adverse reactions associated with MNSDED administration. The MNSDED can remain in the space where the tumor has been ablated to serve as a bound "sentinel" for detecting other cancer cells if they recur. In some designs, information about the "bound" state of the MNSDED can be probed via RF signaling from the MNSDED, and the oncologist can monitor for any additional recurrence of BMBC cells, even before tumor formation that may be large enough to be observed via standard CT or MRI imaging. In the event additional BMBC cells are identified, an additional RF dose can be applied, repeating steps 4 through 8.

[0197] Referring now to the drawings, Figure 1 an exemplary geometry and surface features of the MNSDED are shown. In this embodiment, the MNSDED has a spherical or cubic geometry associated with an etch-stripping process, and the MNSDED surface 101 is associated with the wafer surface used to fabricate the MNSDED. The MNSDED surface 101 is substantially flat. Generally, the MNSDED is characterized by a major axis length 103 and a minor axis length 104 of the MNSDED and has a MNSDED thickness 102 (z-height). The MNSDED surface 101 contains a plurality of EEBI pads 105a - h, which are conductive regions; a plurality of MNSDED communication bus pads 106a - b, which are conductive regions that allow the MNSDED to communicate electrically with the test process; and a plurality of nanopin pairs 107a - b composed of nanopins 107a and 107b, which project significantly upward from the MNSDED surface 101. The MNSDED bottom 108 of the MNSDED is composed of a material that is not susceptible to the effects of the etch-stripping chemicals, and the MNSDED sides 109 can be made of a different material that is also resistant to the stripping chemicals and provides a substantially airtight seal for the sides of the device. The MNSDED top surface 101 also contains solution-to-bulk silicon connection pads 110, which serve as a potential "ground" reference for making electrical measurements relative to the cell surface or other surface or object in the biological solution having the MNSDED. The solution-to-bulk silicon connection pads 110 can be coated with a conductive polymer or polyelectrolyte layer to alter their electrical properties in solution. The solution-to-bulk silicon connection pads 110 can be electrically connected to the bulk silicon of the MNSDED.

[0198] Figure 2 an exemplary architecture and layout of the MNSDED are shown, including features and components associated with the MNSDED surface. Figure 2Depicts the following design components: the MNSDED bottom 108, which consists of: a silicon oxide or other insulating layer on which the MNSDED is fabricated; a plurality of biocompatible protective molecules 201; and an integrated circuit layer 202 constructed on a silicon-on-insulator ("SOI") wafer by a microelectronic manufacturing process. The MNSDED surface 101, which consists mainly of a non-conductive material such as silicon oxide or any other dielectric, includes a plurality of EEBI pads 105a-h and a plurality of MNSDED communication bus pads 106a-b, each EEBI pad 105a-h being coupled to the integrated circuit and each MNSDED communication bus pad 106a-b being coupled to the integrated circuit. The functional handles 203a-b are respectively bound to exemplary EEBI pads 105a-b and are used to bind conductive connectors (EEBILs) 205a-b to the EEBI pads 105a-b. For EEBI pad 105a, a target binding molecule (EEBITA) 204 is bound to the EEBIL 205a. For EEBI pad 105b, a reference molecule 206 is bound to the EEBIL 205b. Conductive nanopin pairs 107a-b are associated with the integrated circuit and consist of a nanopin 107a as the anode and a nanopin 107b as the cathode. An optional loading compartment 207 may contain a therapeutic agent or an imaging agent. The MNSDED side 109 consists of a material such as silicon nitride, which provides a substantially airtight seal for the device.

[0199] Figure 3Shows an exemplary architecture and layout of the MNSDED sensing subsystem and its features and components. Two environmental electronic binding interfaces ("EEBI") 301 and 302 each contain a combination of features and components of the MNSDED that are associated with, interact with, and sense the surrounding environment. These combinations transfer chemical or biochemical information from the environment to other components of the MNSDED by converting chemical or biochemical information into electrical signals that can be processed by the MNSDED logic circuitry. EEBI 301 is a targeted EEBI and has the following features and components: at least one conductive EEBIP 105a; a plurality of biocompatible protective molecules 201 (only one is labeled for clarity) surrounding the perimeter of EEBIP 105a to prevent biofouling or corrosion; a metal-linking compound 303 that is either homofunctional or heterofunctional in nature and capable of binding valence to the gold or other metal surface of EEBIP 105a. The metal-linking chemical moiety 303 is used to covalently or valence-attach a second chemical moiety 304 to EEBIP 105a (where 303 and 304 together are components of a homofunctional or heterofunctional handle 203). The chemical moiety 304 itself is a functional group that can be bound to a complementary reactive moiety 305 by conjugation chemistry, and the complementary reactive moiety 305 is itself covalently attached to EEBIL. This EEBIL can be any conductive polymer or material, and for clarity, in this exemplary figure, it is single-stranded DNA (ssDNA) 306. A second strand of ssDNA 307 is strictly complementary to ssDNA 306, and this ssDNA 307 binds to ssDNA 306 in the finished device. At the distal end of ssDNA 307 is a chemical moiety 308 that facilitates chemical reactions, chemical or bioconjugation chemistry, as known to those skilled in the art. Moiety 308 binds ssDNA 307 to a targeting agent (EEBITA) 204. On any given EEBIP (e.g., 105a), there can be multiple instances of the metal-linking chemical moiety 303 that are bound to the gold or other metal surface of the EEBIP, whereby each instance of the metal-linking chemical moiety 303 on the EEBIP is used to attach a chemical moiety 304 to the EEBIP. Each instance of the chemical moiety 304 linked to the EEBIP is capable of attaching a single EEBIL (e.g., ssDNA 306 and complementary ssDNA 307), and subsequently binding the targeting agent binding moiety 308 to EEBITA 204. In this way, multiple EEBITAs can be attached or linked to the same single EEBIP. EEBI 302 is a reference EEBI and is almost identical to the targeted EEBI 301, except that instead of the EEBITA molecule 204, the reference EEBI 302 includes a non-binding reference agent 206 that has electrochemical properties similar to 204 described herein.

[0200] Figure 4 An RC equivalent circuit showing an exemplary EEBI design using DNA as a charge-carrying polymer is shown. The components include: an EEBIP 105a located on an MNSDED surface 101 having a conductive region that is electrically connected to an integrated circuit below the surface; a metal linker chemical moiety 303; a chemical moiety 304 having a complementary reactive moiety 305; a conductive EEBI L, which in this exemplary figure consists of a first ssDNA 306 and a second ssDNA 307 containing complementary sequences. The first ssDNA strand 306 is coupled to the linker moiety 305, while the second ssDNA strand 307 is coupled to a target-binding molecule (EEBITA) 204 via a binding moiety 308. The EEBITA 204 is capable of specifically binding to a target antigen 401.

[0201] The double-stranded combination of ssDNA 306 and ssDNA 307 provides a path for the flow of charge generated by the binding of the EEBITA 204 to the target antigen 401. The EEBIP 105a provides a connection to the integrated circuit and an associated readout amplifier circuit 402, which is capable of detecting a change in capacitance or voltage upon binding of the target antigen 401. The equivalent circuit model of the arrangement of the EEBI components (105a, 303, 304, 305, 306, 307, 308, 204) is given by a circuit comprising a capacitor 403 in series with a resistor 404 and a parallel RC circuit 405. The combination of the binding moiety 308 and the EEBITA 204 is represented by the capacitor 403. The resistor 404 accounts for the conductivity of the DNA or other conductive polymer "wire" formed by ssDNA 306 and ssDNA 307. The parallel RC circuit 405 accounts for the metal linker moiety 303, as well as the chemical linker moiety 304 and its complementary reactive chemical moiety 305. The conductive EEBIP 105a is connected to the readout amplifier circuit 402 using standard integrated circuit technology. Another input to the readout amplifier circuit 402 comes from a conductive EEBI pad 105b, which is part of a reference EEBI 302. The first voltage input to the readout amplifier circuit 402 generated by the binding of the EEBITA 204 to the target antigen 401 (i.e., associated with the target EEBI 301) is the target voltage 406, while the second voltage input to the readout amplifier circuit 402 generated from a reference molecule 206 (i.e., associated with the reference EEBI 302) is the reference voltage 407.

[0202] Figure 5Shows the general exemplary architecture and layout of the EEBI molecule fine-tuning pair (“EEBIMTP”). Components include: a targeted EEBI 301 including a target EEBIP 105a and an associated EEBITA 204; a reference EEBI 302 including a reference EEBIP 105b and a reference molecule 206; a readout amplifier circuit 402 having a first input target voltage 406 from the target EEBIP 105a and a second input reference voltage 407 from the reference EEBIP 105b. The readout amplifier circuit 402 has an output voltage 501.

[0203] Figure 6 Shows the general exemplary architecture and layout of an alternative EEBIMTP coupled to two reference EEBIs 302a and 302b, including its features and components. Features and components include: a first reference EEBI 302a associated with a first reference molecule 206a and a reference voltage 407a; a second reference EEBI 302b associated with a second reference molecule 206b and a reference voltage 407b; a targeted EEBI 301 associated with an EEBITA 204 and a target voltage 406; a first readout amplifier circuit 402a having a voltage input 407a and 406 and a first readout amplifier circuit output voltage 502a; a second readout amplifier circuit 402b having a voltage input 407b and 406 and a second readout amplifier circuit output voltage 502b. Output voltages 502a and 502b representing binding events are fed to a logic gate circuit 601, which can be any 2-input logic gate.

[0204] Figure 7 Shows the EEBI gasket associated with the MNSDED sensing subsystem of certain embodiments. Components include: a targeted EEBI 301 (including its corresponding EEBIP 105a coupled to an EEBITA 204) and a reference EEBI 302 (including its corresponding EEBIP 105b coupled to a reference molecule 206); a plurality of biocompatible protective molecules 201 on the MNSDED surface 101 to prevent biofouling or corrosion; and a plurality of molecular gasket molecules 701 deposited on the periphery of the EEBIPs 105a and 105b, which create a privileged environment to modify and assist the EEBITA in binding and sensing.

[0205] Decision subsystem / power supply / signaling subsystem

[0206] Figure 8 A shows the general exemplary architecture and layout of an embodiment of the MNSDED implementation subsystem, including the electroporation / electroablation nanoneedles of the implementation subsystem and other features and components. As Figure 8As shown in A, the nanoneedles 107a and 107b protrude from the MNSDED surface 101, having a characteristic height 806. The nanoneedle bases are no more than the characteristic pitch 807 apart, and each nanoneedle base is at least the offset distance 808 from the EEBIP arrays 105a-h. A simple implementation of the nanoneedle drive circuit is shown in Figure 8 C, and its components and features include: one or more nanoneedle pairs, each pair consisting of a nanoneedle anode 107a and a nanoneedle cathode 107b, and each nanoneedle pair 107a and 107b is connected to a nanoneedle drive circuit 801 controlled by one or more logic inputs 802. In Figure 8 the simple example shown in B, Figure 8 the nanoneedle drive circuit 801 of C is represented by a field effect transistor (FET) switch 804 and a FET activation circuit 805 (surrounded by a dashed box). One needle is connected to the nanoneedle drive circuit power supply 803 through the FET switch 804. The FET switch 804 is referenced to the substrate 202 and activated by the FET activation circuit 805, and the FET activation circuit 805 is controlled by the nanoneedle drive circuit logic input 802. Figure 8 The combination of the FET switch 804 and the FET activation circuit 805 in B is used to apply a voltage between the nanoneedle 107a and the nanoneedle 107b under the control of the MNSDED logic (through the logic input 802). Returning to Figure 8 the implementation shown in C, the nanoneedle drive circuit 801 can optionally incorporate a more complex gate driver circuit capable of generating more complex waveforms.

[0207] Figure 9 The general exemplary architecture and layout of an embodiment of the MNSDED implementation subsystem are shown, including the drug / imaging agent delivery components and other features and components of the implementation subsystem. The components and features include: one or more loading compartments or reservoirs 207, each of the reservoirs being a cavity with characteristic dimensions of a reservoir length 901, a reservoir width 902, and a reservoir height 903, and each of the loading compartments or reservoirs 207 is located near other MNSDED surface features including the nanoneedles 107a and 107b, the EEBIP arrays 105a-h, and the MNSDED communication bus pads 106a and 106b. Each reservoir 207 includes a reservoir barrier 904, which can be controllably removed by the MNSDED logic circuit (through a dissolution or electrochemical conversion process), and the reservoir barrier 904 seals or holds a volume of drug or imaging agent 905 within the reservoir 207. The MNSDED surface is covered by a plurality of biocompatible protecting molecules 201, and in this example, it has an oxide MNSDED bottom surface 108 and a nitride MNSDED side surface 109.

[0208] Figure 10FIG. 0 is a flow chart of an exemplary process for fabricating one or more MNSDEDs and preparing the MNSDEDs for delivery to a patient by electrical testing. Depending on the end use and application of the particular MNSDED, variations and deviations from this exemplary process for fabricating and preparing the MNSDED are obvious to those skilled in the art. In step 1001, the physical components and structures associated with the functional MNSDED design (including field effect transistors (FETs), contacts, metals, vias, capacitors, loading compartments, associated interlayer dielectric (ILD) layers, and nanoneedles) are fabricated using a <10 nm technology node or process technology (such as may be characterized by the International Technology Roadmap for Semiconductors (ITRS)). In step 1002, after the nanoneedles are synthesized, each MNSDED undergoes electrical testing, as described in more detail herein, such that defect-free MNSDEDs can be used for future administration and use. Conversely, defective MNSDEDs are not enabled. In step 1003, a charge-carrying linker (e.g., ssDNA) is coupled to the EEBIP using biosafe lithography techniques. In the particular case where the MNSDED uses a spacer, biosafe lithography can be used to print the associated spacer boundary region while coupling the spacer-associated functional handle. In step 1004, the MNSDED is then released from the silicon wafer on which the MNSDED is fabricated using the biosafe wet etch lift-off techniques described in more detail herein. If the specific MNSDED design or use case requires delivery of a drug, imaging agent, and / or other therapeutic agent, then in step 1005, the MNSDED loading compartment or reservoir is filled. In this step, the MNSDED is immersed in a solution containing the drug, imaging agent, and / or other therapeutic agent, and a "cap" (e.g., a membrane) is added by self-assembly to contain the drug, imaging agent, and / or other therapeutic agent within the loading compartment or reservoir. Regardless of whether the specific MNSDED design or use case requires delivery of a drug, imaging agent, and / or other therapeutic agent, in step 1006, the MNSDED is immersed in a solution containing complementary ssDNA that attaches to the previously added ssDNA that binds to the EEBIP. In step 1007, the MNSDED is then immersed in a solution containing a surface biocompatibility protectant that provides biocompatibility-protecting molecules to the MNSDED surface. In the particular case where the MNSDED contains a spacer, the associated spacer molecules are also placed in solution at this time in step 1007. In step 1008, the MNSDED is then washed or rinsed in a wash suspension. Finally, in step 1009, the MNSDED is prepared for delivery to a patient by suspending the MNSDED in a final formulation medium, the composition of which may vary with the specific end use or application of these particular MNSDEDs.

[0209] Figure 11A flowchart of an exemplary biocompatible lithography process for fabricating one or more MNSDEDs. Figure 11 Also depicted are exemplary chemical structures and reactions corresponding to the steps of the biocompatible lithography process. Depending on the end use and application of the specific MNSDED, variations and deviations from this exemplary process for fabricating MNSDEDs will be apparent to those skilled in the art. In step 1101, a thiol-R-thiol molecule is attached to the surface of all EEBIPs fabricated on the MNSDED. In step 1102, a multilayer protective film stack (MPL) consisting of a photoresist, an anti-reflective coating, and a bioprotective layer covers all EEBIPs except one, leaving an EEBIP reactive functional handle (thiol) exposed. In step 1103, ssDNA is coupled to the exposed thiol (using, for example, maleimide-thiol linkage chemistry). In step 1104, the MPL is removed from the EEBIPs previously covered in step 1102 using techniques well known in the art. In step 1105, those EEBIPs that have been coupled to ssDNA are covered in the MPL by standard lithography techniques, leaving only the EEBIPs to be coupled to a pre-designed ssDNA sequence uncovered. Steps 1103 through 1105 are repeated until all of the desired EEBIPs are coupled to ssDNA. In step 1106, after all EEBIPs are coupled to ssDNA, all surfaces are covered with a protective layer consisting of biocompatible protective molecules. In step 1107, a lift-off trench patterning resist is applied to prepare for separating the MNSDED from the silicon wafer using an etch lift-off process.

[0210] Figure 12 Figure A-12D is an illustration of an exemplary etch lift-off process used in the fabrication of one or more MNSDEDs. Variations and deviations from this exemplary etch lift-off process for fabricating MNSDEDs will be apparent to those skilled in the art. In Figure 12 Figure A, each MNSDED region 1201 is surrounded by wells or trenches on all four sides by an anti-lift-off material 1202, a region 1203 filled with a different material reserved for trench etching, and the underlying layer of each MNSDED 1204. The entire assembly of 1201 - 1204 is built on a layer of material 1205 vulnerable to wet etching. The assembly is located on top of a silicon substrate 1206. At this stage of the manufacturing process, the nanoneedles 107 have been fabricated. The MNSDEDs are fabricated and tested on the silicon wafer, and in Figure 12 Figure B, etch lift-off access trenches 1207 are etched around each MNSDED by removing the material 1203 using conventional semiconductor processing (lithography and etching). This exposes the trenches on all four sides of each MNSDED. In Figure 12In C, the silicon wafer is then immersed or subjected to wet chemical treatment, and the wet chemical enters the now-open trench 1207, selectively dissolving the underlying layer 1205 to create a gap 1208 until the MNSDED underlying layer 1205 is completely dissolved from the underlying layer 1206, as Figure 12 shown in D, and each individual MNSDED 1209 can float. Alternatively, in a simpler embodiment of the lift-off process, layers 1204, 1205, and 1206 can all be part of a silicon substrate. Then, trenches 1207 are fabricated with sufficient depth to allow the formation of gaps 1208 by wet etching to release the MNSDED 1209, while leaving sufficient material at the bottom of each MNSDED for use by the functional device.

[0211] Figure 13 is a flowchart depicting an exemplary treatment method using the MNSDED system proposed. Variations and departures from this exemplary treatment method will depend on the specific MNSDED end use and application. In Figure 13 the embodiment shown, one or more MNSDEDs can be used to treat solitary brain metastases of primary breast cancer ("BMBC"). This flowchart depicts the general steps of formulating one or more MNSDEDs, administering one or more MNSDEDs to a patient, activating the MNSDEDs such that the MNSDED sensing, decision-making, and actuation subsystems operate as described herein, and monitoring the patient's condition after treatment.

[0212] More specifically, in step 1301, a sterile solution formulation is prepared and MNSDED is added to the sterile solution to obtain a final formulation. In step 1302, the final solution containing MNSDED is then directly injected into the intratumoral space of the patient. In step 1303, a period of time is allowed sufficient for the MNSDED to traverse the tumor and specifically engage the desired target binding molecule. In step 1304, an RF field is applied to the patient. The RF energy is collected by the RFEHC and powered within each MNSDED. In step 1305, if a specific predetermined antigen binds to the EEBI of the MNSDED, a signal is generated by the sensor subsystem circuitry within each MNSDED. In step 1306, the logic circuitry of the decision subsystem within each MNSDED determines whether there is a predetermined (pre-programmed) "correct" pattern of inputs or signals received from the sensor subsystem, corresponding to the determination that a particular MNSDED is bound to at least one BMBC cell. In step 1307, if the decision subsystem logic circuitry determines that the MNSDED is bound to at least one BMBC cell, the effector subsystem is activated to direct current to the nanoneedle and / or reservoir associated circuitry. In step 1308, this current is applied to electroablate or electroporate the BMBC cell membrane and / or release the contents of the loading compartment or reservoir into the BMBC cell by the nanoneedles. In step 1309, the patient is monitored for tumor disruption-mediated toxicity or other effects. In step 1310, depending on the prescribed treatment protocol, the observed effects on the targeted intratumoral space or other factors, the RF field can be removed to end the treatment, reapplied for subsequent treatment cycles, and / or additional doses of MNSDED can be administered to the patient in a manner similar to repeating steps 1302 to 1309.

[0213] Figure 14 A shows a simple example of a sense amplifier (SA) circuit. The SA differentially amplifies the voltages from a pair of EEBIs, targeting EEBI 301 (V eebi靶 ), and reference EEBI 302 (V eebi参考 (corresponding to the targeting EEBI 301 and reference EEBI 302 in Figure 3 ), and incorporates a cross-coupled latch (1401) with output 1406. The SA acts as a comparator that changes the logic state of its output when a large enough difference between EEBI301 and EEBI 302 is detected. A change in capacitance when the target antigen binds at the targeting EEBI 301 will cause a voltage difference, V eebi靶 - V eebi参考, which is amplified and detected by an SA (1410) that provides a clock signal 1408. The clock signal 1408 (node CK) enables the NMOS transistor current source 1404 and the PMOS current source 1402 during the linear amplification and the regenerative amplification phases. It is powered at 1405, and the output voltage V out . V out 's complement V outbar (measured at 1407) is 180 degrees out of phase with V out . According to the final detection scheme, a latch circuit or other logic element will be connected to 1406 (V out ) and 1407; the variations of this latch circuit will be obvious to those skilled in the art of CMOS comparator design. The output of the latch circuit 1406 represents the combined detection signal. The schematic diagram of the detection circuit including the sense amplifier is represented by the sense amplifier 1410 in Figure 14 B, where the representative capacitors 1412 and 1411 represent the capacitances at the targeted EEBI and the reference EEBI respectively. It is powered at 1405, and the SA combined signal is output at 1403. The sense amplifier 1410 is clocked by the clock circuit 1409 that outputs the clock signal 1408. All the sensing circuits are referenced to the solution-to-bulk silicon connection pad 110.

[0214] Figure 15 A-15B shows the variations of the combined detection circuit. Figure 15 A shows the linear differential detection scheme, where the EEBIs 301 and 302 with capacitances 1412 and 1411 are driven by the AC signals 1502 and 1503 with opposite phases respectively, and the resulting currents are subtracted and amplified through a chain of amplifiers 1506 and 1507 having capacitances and / or other impedances 1504, 1507 and 1511 in their feedback paths to create an integrating, differentiating, linear or non-linear amplifier. Then, the output of the amplifier 1507 at the end of the chain is demodulated using the drive voltage source 1502 and its 90-degree phase-shifted copy to create a pair of DC voltages representing the impedance difference between the target EEBI 301 and its reference EEBI 302. Then these voltages are compared with a reference threshold using the sense amplifiers 1501a and 1501b. The output from the comparator is fed to the sense amplifier output 1406 and used as part of the MNSDED combined detection logic. Figure 15B shows a clocked readout amplifier 1410 used in conjunction with pulsed voltage sources 1508 and 1509 having a common mode (bias) voltage source 1510 to apply DC and time-varying voltages to EEBIs capacitors 1412 and 1411. When a target molecule binds to the EEBI 301, the capacitance change between the target and reference EEBIs 301 and 302 will result in different discharge rates of 1412 and 1411, thus resulting in a time-varying voltage difference that varies with the clock signal 1408 output from the follower clock circuit 1409, and this voltage difference will then be detected by the readout amplifier 1410. Variations and departures from these exemplary impedance measurement circuits will be apparent to those of ordinary skill in the art.

[0215] Figure 16 A-16B shows an exemplary schematic diagram of the entire MNSDED binding detection process. Variations and departures from such an exemplary schematic diagram of the entire MNSDED binding detection process are possible. In Figure 16 A, the EEBITA or target binding molecule 204 and the reference molecule 206 are bound to the conductive EEBI pads 105a and 105b via ssDNA 306 and complementary ssDNA 307 in the manner described previously herein (see, for example Figure 3 ). Each of the EEBI pads 105a and 105b is electrically connected to a readout amplifier ("SA") 1410 (see, for example Figure 14 A-14B or Figure 15 A-15B), where the SA can include Figure 14 any detection method described in A-14B and Figure 15 A-15B, such as a clocked comparator, a linear current or voltage amplifier followed by a clocked comparator, a linear amplifier chain followed by demodulation and a clocked comparator, or other methods for converting the capacitance change between EEBIs 301 and 302 upon a target binding event. The SA 1410 is clocked by a clock circuit 1409 that outputs a clock signal 1408. The voltage output V out 1406 of the SA is a logic voltage connected to a count threshold circuit 1601. In Figure 16 B, if the count threshold circuit 1601 detects a predetermined number of positive binding events with the target antigen 401 within a predetermined time interval starting from the SA 1410, corresponding to a predetermined number of signals 1406, it sends a binding signal 1603 to the logic circuit 1602. The logic "decision" circuit 1602 receives the input from the logic circuit 1601 and any other logic inputs closely related to the binding decision, and via the nanopin drive circuit input signal 1604, activates the nanopin drive circuit 801, provides a voltage to the nanopins 107a-b, and produces a predetermined effect (such as electroablation) on the target cell that is the source of the binding event. Return referenceFigure 16 A, if the counting threshold circuit 1601 fails to detect a predetermined number of binding events or signal 1406 (e.g., corresponding to absence or insufficient target binding) within a predetermined time interval starting from SA 1410, then the signal 1603 to the logic circuit 1602 remains unasserted, the corresponding signal to the logic "decision" circuit 1602 remains quiescent, no signal activates the nanoneedle drive circuit 801, and the nanoneedles 107a-b remain de-energized.

[0216] Figure 17 A representative diagram of the 3-EEBI pairing system is shown. Variations and departures from this exemplary schematic of the 3-EEBI pairing system are possible. The EEBI pairs (301a, 302a), (301b, 302b), and (301c, 302c) include specific EEBITA and reference molecules. The differential voltages from each EEBI pair are measured by or input to the clocked readout amplifiers 1410a, 1410b, and 1410c, respectively. The clock signal 1408 in this embodiment is generated by the same RF that powers the MNSDED; the clock signal is recovered from the RF energy harvesting circuit ("RFEHC") 1701 (including the RFEHC coil 1704) and sent to the clock distribution circuit ("CDC") 1409 for distribution throughout the MNSDED on the clock line 1703. The CDC 1409 also provides the counting threshold circuit 1601 via the clock line 1703. The readout amplifiers 1410a, 1410b, and 1410c send their output signals 1406a, 1406b, and 1406c to the counting threshold circuit 1601 via lines 1705, 1706, and 1707, respectively. The counting threshold circuit 1601 sends each of the received signals 1406a, 1406b, and 1406c (representing binding or non-binding events) to the logic / control circuit 1702, which determines whether one or more predetermined patterns, combinations, or threshold numbers of EEBITA binding events have occurred (based on a circuit implementation such as Boolean logic). If the logic / control circuit 1702 determines that one or more predetermined patterns, combinations, or threshold numbers of EEBITA binding events have occurred, then it sends an activation signal to the nanoneedle drive circuit 801 via the control circuit 1708, and the activation signal activates or triggers the nanoneedles 107a-b. If the logic / control circuit 1702 determines that no predetermined pattern, combination, or threshold number of EEBITA binding events have occurred, then it inhibits sending an activation signal to the nanoneedle drive circuit 801, and the nanoneedles 107a-b are not activated or triggered.

[0217] Figure 18An example of a binding mode determination process using a logic circuit is shown. Variations and departures from this exemplary binding mode determination process are possible. The EEBI pairs (301a, 302a), (301b, 302b), and (301c, 302c) include specific EEBITA and reference molecules. The differential voltages from each EEBI pair are measured by or input to the clocked readout amplifiers 1410a, 1410b, and 1410c, respectively. Each clocked readout amplifier 1410a, 1410b, and 1410c outputs to the corresponding clocked count threshold circuits 1601a, 1601b, and 1601c, which are provided with the clock signal 1408 from the clock distribution circuit 1409. Each count threshold circuit outputs signals 1603a, 1603b, and 1603c, which are input to the 3-input logic circuit 1801, which implements a logical determination output as the voltage level or signal 1802. If the count threshold circuit output signals 1603a and 1603c indicate bound antigen, but the count threshold circuit output signal 1603b does not indicate bound antigen, this specific 3-input logic circuit embodiment will trigger only the nanoneedle drive circuit 801 ( Figure 18 not shown in) via the 3-input logic circuit output signal 1802, the status is indicated by 1803 and circled in the Figure 18 logic table in the lower right. In this example, the count threshold circuit output signal 1603b can represent an inhibitory binding event. This particular combination of the count threshold circuit output signals 1603a, 1603b, and 1603c, along with all other combinations of the count threshold circuit output signals that do not trigger the nanoneedle drive circuit 801, are illustrated in the Figure 18 mapping table in the lower right. In this particular mapping table, input A corresponds to the EEBI pair 301a and 302a (and its corresponding count threshold circuit output signal 1603a), input B corresponds to the EEBI pair 301b and 302b (and its corresponding count threshold circuit output signal 1603b), and input C corresponds to the EEBI pair 301c and 302c (and its corresponding count threshold circuit output signal 1603c).

[0218] Figure 19is an exploded perspective view of a multi-layer coil radio frequency energy harvesting circuit ("RFEHC") 1701 embedded in MNSDED. Variations and departures from this exemplary RFECH design and structure are possible. The first layer of coil 1901 is shown at the top layer 101, but it can be buried in a lower layer or implemented as a multi-turn multi-layer coil with more layers / turns in the lower metal / ILD stack 1902. The coil winding start 1903 and the coil winding end 1904 are connected to other components of the RFEHC in the lower integrated circuit layer of the MNSDED. Each layer of the RFEHC coil is connected to the layers above and below by a plurality of via structures 1905. The MNSDED is bounded by the MNSDED side 109 that forms an isolation well. The EEBI pad 105, the communication bus pad 106, the solution-to-bulk silicon connection pad 110, and the nanopins 107a-b are shown, and the RFEHC coil 1901 is shown wound around the outside of other surface components. A buried layer 1906 of an additional high magnetic susceptibility (high μ) film can be placed inside the inner layer of the RFEHC coil.

[0219] Figure 20 A-20C shows details of an exemplary RFEHC coil design. Variations and departures from this exemplary RFEHC coil design and structure are possible. The single-layer multi-turn planar RFEHC coil is shown as 1901 in Figure 20 A, with most of its turns at the outer radius because the inner radius turns do not cross as much magnetic flux as the outer radius turns and contribute mostly to losses. The RFEHC coil 1901 can be part of a series or parallel resonant circuit that has a capacitor associated with a voltage multiplier circuit or connected to a more complex impedance transformation circuit, depending on the intended frequency. Figure 20 B shows the circuit topology of a multi-layer RFECHC coil 1902, with a starting point 1903 of the winding, an ending point 1904 of the winding, and the layers connected in series by via structures 1905. By placing high permeability films (magnetic susceptibility μ >> μ0, the magnetic susceptibility of free space) 1906a, 1906b, 1906c, etc. inside one or more planar coils, the flux collection efficiency of the coil can be improved, as Figure 20 shown in C. If more than one permeable layer is used, the permeable layers can be connected using via structures made of a high permeability material 2001.

[0220] Figure 21 A-21B is an example of a voltage multiplier circuit connected to an RFEHC LC (coil-capacitor) circuit. Variations and departures from this example of a voltage multiplier circuit connected to an RFEHC LC (coil-capacitor) circuit are possible. In these examples, the RFEHC coil 1901 and the capacitor 2101 are connected in series, but more complex energy storage circuits can also be used. InFigure 21 In the circuit of A, the RFEHC coil is connected to a Cockcroft-Walton voltage multiplier composed of diode 2103 and capacitor 2102. In Figure 21 In the circuit of B, the RFEHC coil is connected to diode-connected MOSFETS 2104. The rectified and multiplied output voltage for both the upper and lower circuits appears at output node 2105.

[0221] Figure 22 A possible test configuration of mass-produced MNSDED 1209 fabricated on silicon wafer 1206 is shown. Variations and deviations from the design and structure of this exemplary test configuration are possible. Each MNSDED 1209 is connected to a much larger logic test chip 2201 via a communication bus having cross-connections 2202 and 2203.

[0222] Figure 23 A-23B is a mode for testing and activating only those MNSDEDs that pass a set of logic tests. Variations and deviations from this exemplary mode for testing and implementing defect-free MNSDEDs are possible. In Figure 23 In A, the conductive fuse 2301 shorts the power delivery circuit 2105 by default in each MNDSED during manufacturing. The pin driver 801 of the MNSDED circuit is represented by block 2304. If the MNSDED passes the logic test applied by the Figure 22 logic test chip 2201, a current pulse transmitted through line 2303 is used to fuse the fuse 2301 (shown as fuse 2302 being fused in Figure 23 B), enabling the MNSDED to be powered by RFECHC. Otherwise, the MNSDED remains inert, as Figure 23 shown in A.

[0223] Although specific embodiments have been illustrated and described above, it should be understood that the disclosure provided is not limited to the exact configurations, steps, and components disclosed. With the aid of this disclosure, various modifications, changes, and variations to the arrangements, operations, and details of the disclosed methods and systems will be apparent to those skilled in the art.

[0224] Without further elaboration, it is believed that those skilled in the art can utilize the foregoing description to the fullest extent of the disclosure. The examples and embodiments disclosed herein should be construed as merely illustrative and exemplary, and in no way limiting the scope of the disclosure. It will be apparent to those skilled in the art that changes can be made to the details of the above embodiments without departing from the basic principles of the disclosure.

Claims

1. A micro / nano sensing-determining-actuating device, comprising: A non-conductive housing having a surface and containing support logic circuitry; At least one conductive needle extending from the housing, the needle being electrically connected to the logic circuitry and a needle driving circuit; and At least one conductive pad positioned on the housing surface and electrically connected to the logic circuitry and at least one targeting agent capable of binding to a target; Among them, In response to a target binding event, the logic circuitry sends an activation signal to the needle driving circuit to trigger application of a voltage from the needle driving circuit to the at least one needle.

2. The micro / nano sensing-determining-actuating device according to claim 1, wherein the non-conductive housing has a major axis length from 100 nanometers to 500 micrometers, and the logic circuitry is fabricated using a 10 nm or smaller SIA transistor node.

3. The micro / nano sensing-determining-actuating device according to claim 1, further comprising a biocompatible protective agent surrounding at least a portion of the surface of the non-conductive housing.

4. The micro / nano sensing-determining-actuating device according to claim 1, wherein each targeting agent is connected to a conductive linker that attaches to the conductive pad.

5. The micro / nano sensing-determining-actuating device according to claim 1, wherein the at least one needle is capable of electrically interacting with a bilayer lipid membrane of a target cell attached to the targeting agent, and a sufficient voltage from the needle driving circuit is capable of being used to promote at least one of ablation and electroporation of the target cell.

6. A method for using a micro / nano sensing-determining-actuating device, comprising: Introducing a plurality of micro / nano sensing-determining-actuating devices into a system having a target, each micro / nano sensing-determining-actuating device having a non-conductive housing and at least one conductive needle extending from the housing, the housing having a surface and containing support logic circuitry, the needle being electrically connected to the logic circuitry and a needle driving circuit, and the micro / nano sensing-determining-actuating device further comprising at least one conductive pad positioned on the housing surface and electrically connected to the logic circuitry and a targeting agent capable of binding to the target; and In response to a target binding event, sending an activation signal to the needle driving circuit to trigger application of a voltage from the needle driving circuit to the at least one needle.

7. A therapeutic composition, comprising: A pharmaceutically acceptable carrier; A plurality of micro / nano sensing-determining-actuating devices contained within the carrier, the micro / nano sensing-determining-actuating devices comprising: A non-conductive housing having a surface and containing support logic circuitry; At least one conductive needle extending from the housing, the needle being electrically connected to the logic circuitry and a needle driving circuit; and At least one conductive pad located on the housing surface and electrically connected to the logic circuitry and a targeting agent capable of binding to a target, wherein, in response to a target binding event, the logic circuitry sends an activation signal to the needle driving circuit to trigger application of a voltage from the needle driving circuit to the at least one needle.

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