Patent Application
20210389297
Many diseases are difficult to be detected by a single approach or methodology. In particular, many serious diseases with high morbidity and mortality, including cancer and heart diseases, are difficult to diagnose at an early stage with high sensitively, specificity and efficiency, by using one piece of detection equipment. Current disease diagnosis devices typically detect and rely on a single type of macroscopic data and information such as body temperature, blood pressure, or scanned images of the body. For example, to detect serious diseases such as cancer, each of the diagnosis apparatus commonly used today is based on one imaging technology, such as x-ray, CT scan, or nuclear magnetic resonance (NMR). While used in combination, these diagnosis apparatuses provide various degrees of usefulness in disease diagnosis. However, each of them alone cannot provide accurate, conclusive, efficient, and cost-effective diagnosis of such serious diseases as cancer at an early stage, and is difficult to detect multiple types of cancer at the same time. Further, many of the existing diagnosis apparatus have a large size and are invasive with large footprint, such as x-ray, CT scan, or nuclear magnetic resonance (NMR).
Even the newly emerged technologies such as those deployed in DNA tests usually rely on a single diagnosis technology and cannot provide a comprehensive, reliable, accurate, conclusive, and cost-effective detection for a serious disease. In recent years, there have been some efforts in using nano technologies for various biological applications, with most of the work focused on one type of gene mapping and moderate developments in the field of disease detection. For instance, Pantel et al. discussed the use of a MicroEelectroMechanical Systems (MEMS) sensor for detecting cancer cells in blood and bone marrow in vitro (see, e.g., Klaus Pantel et al., Nature Reviews, 2008, 8, 329); Kubena et al. disclose in U.S. Pat. No. 6,922,118 the deployment of MEMS for detecting biological agents; and Weissman et al. disclose in U.S. Pat. No. 6,330,885 utilizing MEMS sensor for detecting accretion of biological matter.
In sum, to date, most of above described technologies have been limited to isolated diagnosis technology for sensing, using systems of relatively simple constructions and large dimensions but often with limited functions, and lack sensitivities and specificities. Further, the existing technologies require multiple times detection by multiple apparatus. This will increase costs and affect achieved degree of sensitivity and specificity as well.
Current cancer screening and prognosis IVD methods typically include bio-markers, circulating tumor cells (CTC), and genomics (such as circuiting tumor-DNA (ct-DNA)). While each of the above-mentioned technology offer a number of advantages, they also have a number of limitations, which include inability to simultaneously detect multiple types of cancer, inability to detect cancer early, relatively low sensitivity and specificity, and in some cases, inability to detect certain types of cancer (for example, esophageal cancer and brain tumor). Bio-markers are not effective for early stage cancer detection, but are limited for cancer types.
In the case of CTC and ct-DNA, signals occur only after solid tumor has been formed, making early stage cancer detection relatively. See, e.g., Ji et al., J Clin Oncol 33, 2015; Xuedong Du et al., J Clin Oncol 33, 2015; Jiang et al., J Clin Oncol 33, 2015; Tao et al., J Clin Oncol 33, 2015; Bettegowda et al., Science Translational Med., 2014, 6 (224):224; Phallen et al., Science Translational Med., 2017, 9 (403): 2415; Khoo et al., Science Advances, 2016, 2 (7):e1600274; Garcia-Murillas et al., Sci. Translational Med., 2015, 7 (302): 302; Abbosh et al., Nature, 2017, 545 (7655):446-451; and Herbst et al., Nature, 2018, 553 (7689):446.
To date, conventional cancer screening, detection and/or treatment methods lack markers or signals to effectively target or impact multiple types of cancer at the same time (e.g., in one test or treatment).
These drawbacks call for novel solutions that provide reliable and flexible diagnosis apparatus using multiple diverse technologies and bring improved accuracy, sensitivity, specificity, efficiency, non-invasiveness, practicality, conclusive, and speed in early-stage disease detection, particularly in screening or detection of multiple cancer types, at reduced costs.
The present invention in general relates to a novel technology for detecting disease, in which a number of different classifications of biological information are collected in a device and processed or analyzed.
It also relates to a novel technology for assessing risk levels of disease and cancer occurrence, and differentiating healthy individuals from possible disease or cancer individuals.
In traditional technology, typically only one level of biological information is collected (one dimensional), while in this novel technology, at least two levels (classifications) of information can be collected (seven dimensional, or seven factor interactions). Compared with traditional technology which typically focuses on one parameter or one level (for example, bio-marker at protein level), signal and information collected in this novel technology can be collected in a number of forms, and non-linearly amplified. There are additional 2-factor and three-factor interactions which can be collected and analyzed, which maybe missing in other technologies, since they typically only measure one type of biological information.
Existing cancer screening tests and treating methods lack the ability to effectively detect and/or impact multiple types of cancer at the same time (e.g., in one test or treatment). Compared with those traditional technologies which typically focus on one type of disease in one test, a significantly increased number of diseases - particularly cancers or pre-cancer diseases (e.g., over 20 types of cancer)—can be simultaneously detected, or even treated, by this novel technology.
Specifically, this novel technology can be used for cancer screening, assisting in diagnosis, prognosis, and follow-up tests with improved sensitivity and specificity, ability to detect cancer early, ability to detect major diseases, pre-cancer diseases and over 20 types of cancer, cost effective, and no side effects. Particularly, the present invention provides novel apparatus and methods for simultaneously detecting and/or treating the presence or monitoring the progression of multiple types of diseases, including cancer.
The novel technology offers several advantages that cannot be achieved by the traditional technology: (1) ability proven so far to detect as many as over 20 cancer types in one test, including some cancer types which cannot be detected by other in vitro tests (e.g., esophageal cancer, cerebral cancer), covering over 80% of all cancer incidences; (2) capability of early stage cancer detection; (3) high sensitivity and specificity (75%˜90% on over 20 types of cancer); (4) no side effects; (5) high speed, fully automated operations without human intervention; (6) statistical difference between cancer group and non-cancer disease group including inflammation—significantly lower false positives (higher specificity); (7) easy process, no difference between fasting blood testing and non-fasting blood testing, and (8) highly cost effective.
In addition, compared to conventional technologies, the novel cancer treatment technology of this invention has a number of major, unexpected advantages—including, e.g., lower costs, far less side efforts, easier recovery, capability of cancer prevention, improved survival rate, and ease of use and likely widespread clinical uses. Notably, the novel cancer treatment of this invention can use a low dosage and/or a weak field and/or energy for treatment.
Accordingly, one aspect of this invention relates to an apparatus for simultaneously detecting presence or monitoring progression of two or more types of cancer in a biological subject, comprising: a channel or chamber in which the biological subject passes through; at least one detector placed partially or completely alongside one or more side walls of the channel or chamber, wherein the at least one detector is configured to detect at least one bio-physical or physical property of the biological subject, and the detected bio-physical or physical property is collected for analysis to determine whether each of the two or more types of cancer is likely to be present with the biological subject or to determine the status of each cancer, thereby providing the ability to simultaneously determine or monitor progression of the two or more types of cancer.
In some embodiments, the bio-physical property comprises a mechanical property, an acoustical property, an optical property, an electrical property, an electro-magnetic property, or an electro-mechanical property.
In some further embodiments, electronic property can be surface charge, surface potential, resting potential, electrical current, electrical field distribution, surface charge distribution, cell electronic properties, cell surface electronic properties, dynamic changes in electronic properties, dynamic changes in cell electronic properties, dynamic changes in cell surface electronic properties, dynamic changes in surface electronic properties, electronic properties of cell membranes, dynamic changes in electronic properties of membrane surface, dynamic changes in electronic properties of cell membranes, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, electrical dipole, electrical quadruple, oscillation in electrical signal, electrical current, capacitance, three-dimensional electrical or charge cloud distribution, electrical properties at telomere of DNA and chromosome, DNA surface charge, DNA surrounding media electrical properties, quantum mechanical effects, capacitance, or impedance. For example, the bio-physical property may be quantum mechanical effects that affect gene replications and mutations.
In some embodiments, the bio-physical property comprises a trans-membrane potential, a membrane voltage, a membrane potential, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, a zeta potential, an impedance, an optical reflective index, an optical refractive index, potassium ions, sodium ions, chloride ions, nitride ions, calcium ions, an electro-static force, an electro-static force acting on cells, an electro-static force acting on DNA double helix, an electro-static force acting on RNA, an electrical charge on cell membrane, an electrical charge on DNA double helix, an electrical charge on RNA, quantum effects, near-field electrical properties, near-field electro-magnetic properties, membrane bilayer properties, ion types and/or concentrations, ion permeability, electrical current, electrical conductance, capacitance, or electrical resistance.
In some embodiments, the apparatus detects one or more bio-physical or physical properties each selected from the group consisting of electrolyte, potassium ion concentration, sodium ion concentration, chloride ion concentration, calcium ion concentration and distribution, net charge in extracellular region, ion concentration in extracellular, gravitational field, mechanical field, quantum mechanical field, magnetic field, electro-magnetic field, electrical field, electrical current, electrical resistance, electrical conductance, capacitance, membrane potential, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, magnetic field, electrical field, electro-magnetic field, and quantum field in the biological subject.
In some further embodiments, the apparatus directly or indirectly measures the quantum mechanical effects; and/or directly or indirectly measures ions or ion levels in a liquid sample of the biological subject; and/or directly or indirectly measures ion levels or concentrations by a bio-chemistry or electrode method; and/or directly or indirectly measures potassium ions; and/or directly or indirectly measures concentration of potassium ions; and/or directly or indirectly measures ion permeability.
In some embodiments, the apparatus directly or indirectly measures one or more ions selected from potassium ions, sodium ions, chloride ions, nitride ions and calcium ions; and/or directly or indirectly measures the concentration(s) of one or more ions selected from the group consisting of potassium ions, sodium ions, chloride ions, nitride ions and calcium ions.
In some embodiments, the biophysical physical property is related to and responsible for cell to cell interactions, cell signal, cell surface properties, cell electro-static force, cell repulsive force, DNA surface properties, DNA surface charge, DNA surrounding media electrical properties, quantum mechanical effects, gene mutation frequencies, or quantum mechanical effects.
In some embodiments, the biological subject is a liquid sample. For example, the biological subject may include a body fluid or tissue. More specifically, the body fluid may comprise whole blood, serum, plasma, sweat, tear, or urine.
In some embodiments, the bio-physical or physical property is a non-cancerous signal and exists in the biological subject from a healthy individual.
In some embodiments, the bio-physical or physical property exists and can be detected in both the biological subject from a healthy individual, the biological subject from a non-cancer disease patient, and the biological subject from a patient with each of the two or more types of cancer; and the detected properties between the healthy individual, the non-cancer disease patient, and the patient with the cancer are different.
In some embodiments, the bio-physical or physical property exits in each of the two or more types of cancer, and is able to differentiate a normal sample from abnormal samples with the cancers.
In some embodiments, the determination is by comparing the biophysical information of the detected biological subject with the same biological information of a confirmed disease-free or diseased biological subject.
In some embodiments, the status of each of the cancers comprises health stage, non-cancer disease stage, pre-cancer stage, early stage cancer stage, and mid to late stage cancer stage, with statistically significant detection or monitoring between any of the two stages.
In some other embodiments, the apparatus simultaneously can presence or monitoring progression of three or more types of cancer in the biological subject.
In some embodiments, the cancer types may comprise lung cancer, liver cancer, colorectal cancer, breast cancer, gastric cancer, esophageal cancer, brain tumor, prostate cancer, blood cancer, intestine cancer, stomach cancer, cervical cancer, ovarian cancer, rectum cancer, colon cancer, nasopharyngeal cancer, cardiac carcinoma, uterine cancer, oophoroma, pancreatic cancer, or circulating tumor cells.
In some other embodiments, an additional device is used for adjusting the physical or bio-physical property in the biological subject. For instance, the physical or bio-physical property may be first measured and then adjusted.
In some embodiments, such physical or bio-physical property comprises a mechanical property, an acoustical property, an optical property, an electrical property, an electro-magnetic property, or an electro-mechanical property. More specifically, the electrical property may comprise electrical current, electrical conductance, capacitance, electrical resistance, ion types and/or concentrations, or quantum mechanical effect.
In some embodiments, the additional device adjusts the current to a higher value, adjusts the electrical conductance to a higher value, adjusts the electrical resistance to a lower value, or alters the quantum mechanical effect.
In some embodiments, a reagent is injected into blood to adjust bio-physical properties in the blood. For example, the reagent may contain ions, oxidizers, and components to impacting electrical properties of the blood. Such electrical property may comprise electrical current, electrical conductance, capacitance, electrical resistance, ion types and/or concentrations, or quantum mechanical effect.
In some embodiments, the reagent is a drug capable of adjusting the biological properties in the blood. In some further embodiments, the drug is capable of releasing, upon intake, ions and charged components and capable of adjusting electrical properties of the blood. Such property may comprise electrical current, electrical conductance, capacitance, electrical resistance, ion types and/or concentrations, or quantum mechanical effect.
In some other embodiments, at least one bio-marker is added to the liquid sample for physical or bio-physical property and related properties to be measured. In some embodiments, the bio-marker provides at least some indicative information of risks of two or more types of cancer occurrence at a given organ and location.
In still some other embodiments, the detected property is analyzed in conjunction with information and data obtained from test(s) comprising of bio-marker tests, genomics tests, circulating tumor DNA, circulating free tumor DNA, and circulating tumor cell tests, and overall cancer risks and location(s) of possible cancer occurrence are obtained.
In some embodiments, the apparatus comprises an ion injector, configured to add a desired amount of ions to the biological subject. For instance, the ions may comprise potassium ions, sodium ions, chloride ions, nitride ions, or calcium ions. Particularly, the ions may include potassium ions.
In some embodiments, the apparatus comprises one or more channels, and the one or more channels comprise one or more detectors; wherein the one or more detectors are configured to detect one or more bio-physical or physical properties of the biological subject.
In some embodiments, the apparatus further comprises one or more ion injectors on sidewalls, and each of the one or more ion injectors is configured to add a desired amount of ions to the biological subject. The ions added by ion injectors can be the same or different; and the bio-physical or physical properties detected by different detectors can be the same or different.
In some embodiments, the detector comprises one or more sensors placed partially or completely alongside one or more side walls of the channel or chamber. Each sensor may be independently a thermal sensor, optical sensor, acoustical sensor, biological sensor, chemical sensor, electro-mechanical sensor, electro-chemical sensor, electro-optical sensor, electro-thermal sensor, electro-chemical-mechanical sensor, bio-chemical sensor, bio-mechanical sensor, bio-optical sensor, electro-optical sensor, bio-electro-optical sensor, bio-thermal optical sensor, electro-chemical optical sensor, bio-thermal sensor, bio-physical sensor, bio-electro-mechanical sensor, bio-electro-chemical sensor, bio-electro-optical sensor, bio-electro-thermal sensor, bio-mechanical-optical sensor, bio-mechanical thermal sensor, bio-thermal-optical sensor, bio-electro-chemical-optical sensor, bio-electro-mechanical optical sensor, bio-electro-thermal-optical sensor, bio-electro-chemical-mechanical sensor, physical sensor, mechanical sensor, piezo-electrical sensor, piezo-electro photronic sensor, piezo-photronic sensor, piezo-electro optical sensor, bio-electrical sensor, bio-marker sensor, electrical sensor, magnetic sensor, electromagnetic sensor, image sensor, or radiation sensor.
More specifically, the thermal sensor may include a resistive temperature micro-sensor, a micro-thermocouple, a thermo-diode and thermo-transistor, and a surface acoustic wave (SAW) temperature sensor; the image sensor may include a charge coupled device (CCD) or a CMOS image sensor (CIS); the radiation sensor may include a photoconductive device, a photovoltaic device, a pyro-electrical device, or a micro-antenna; the mechanical sensor may include a pressure micro-sensor, micro-accelerometer, flow meter, viscosity measurement tool, micro-gyrometer, or micro flow-sensor; the magnetic sensor may include a magneto-galvanic micro-sensor, a magneto-resistive sensor, a magneto diode, or magneto-transistor; the biochemical sensor may include a conductimetric device, a bio-marker, a bio-marker attached to a probe structure, or a potentiometric device.
In some embodiments, at least one sensor is a probing sensor and applies a probing or disturbing signal to the biological subject. In some further embodiments, at least another sensor, different from the probing sensor, is a detection sensor and detects a response from the biological subject upon which the probing or disturbing signal is applied.
In some other embodiments, a read-out circuitry is connected to at least one sensor and transfers data from the sensor to a recording device. The connection between the read-out circuit and the sensor may be digital, analog, optical, thermal, piezo-electrical, piezo-photronic, piezo-electrical photronic, opto-electrical, electro-thermal, opto-thermal, electric, electromagnetic, electromechanical, or mechanical.
In some embodiments, the chamber or channel has a length ranging from 1 micron to 50,000 microns, from 1 micron to 15,000 micron, from 1 micron to 10,000 microns, from 1.5 microns to 5,000 microns, or from 3 microns to 1,000 microns.
In some embodiments, the chamber or channel has a width or height ranging from 0.1 micron to 100 microns; from 0.1 micron to 25 microns, from 1 micron to 15 microns, or from 1.2 microns to 10 microns.
In some embodiments, at least four sensors are located on one side, two opposite sides, or four sides of the interior surface of the chamber or channel.
In some embodiments, an application specific integrated circuit chip is internally bonded to or integrated into the detector.
In some embodiments, the apparatus is fabricated by integrated circuit technologies.
Another aspect of this invention relates to a method for screening or detecting the presence or progression of two or more types of cancer in a biological subject, comprising measuring a physical or biophysical property at a microscopic level of cells in the biological subject with an apparatus of this invention, wherein information related to the measured property of the cells in the biological subject is detected by the detector and collected for analysis to determine whether each of the cancers is likely to be present with the biological subject or to determine the status of each cancer, thereby providing the ability to simultaneously determine or monitor progression of the two or more types of cancer.
In some embodiments, the measured property is collected for analysis to simultaneously determine or monitor progression of three or more types of cancer.
Still a further aspect of this invention provides an apparatus for treating a biological subject, comprising: a channel or chamber in which the biological subject passes through; and at least one transducer placed partially or completely in the channel or chamber; wherein the transducer is configured to transmit at least one bio-physical property, bio-physical energy, material or element onto the biological subject, thereby providing a simultaneous treatment of two or more types of cancer in the biological subject.
In some embodiments, the biological subject is a liquid sample of a mammal. The biological subject may be a blood sample, a urine sample, or a sweat sample of the mammal. More specifically, the biological subject may comprise blood, proteins, red blood cells, while blood cells, T cells, other cells, gene mutations, quantum mechanical effects, DNA, RNA, or other biological entities.
In some embodiments, the cancers comprise lung cancer, liver cancer, colorectal cancer, breast cancer, gastric cancer, esophageal cancer, brain tumor, prostate cancer, blood cancer, intestine cancer, stomach cancer, cervical cancer, ovarian cancer, rectum cancer, colon cancer, nasopharyngeal cancer, cardiac carcinoma, uterine cancer, oophoroma, pancreatic cancer, or circulating tumor cells.
In some embodiments, the at least one bio-physical property, bio-physical energy, material or element comprises a mechanical property or energy, an acoustical property or energy, an optical property or energy, an electrical property or energy, an electro-magnetic property or energy, or an electro-mechanical property or energy.
In some embodiments, the at least one electrical property or energy comprises electrical current, electrical voltage, electrical field, electro-magnetic field, electrical conductance, capacitance, electrical resistance, net electrical charge in extracellular region, membrane potential, membrane polarization, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, ion concentrations, electro-static force and charge on DNA double helix and RNA double helix, or quantum mechanical effect.
In some embodiments, the at least one bio-physical property, bio-physical energy, material or element comprises a trans-membrane potential, a membrane voltage, a membrane potential, a membrane ion channel property, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, a zeta potential, an impedance, an optical reflective index, an optical refractive index, potassium ions, sodium ions, chloride ions, nitride ions, calcium ions, an electro-static force, an electro-static force acting on cells, an electro-static force acting on DNA double helix, an electro-static force acting on RNA, an electrical charge on cell membrane, an electrical charge on DNA double helix, an electrical charge on RNA, quantum effects, near-field electrical properties, near-field electro-magnetic properties, membrane bilayer properties, ion types and/or concentrations, ion permeability, electrical current, electrical conductance, capacitance, or electrical resistance.
In some embodiments, the transmitted bio-physical property or energy adjusts the current of the biological subject to a higher value, adjusts the electrical conductance of the biological subject to a higher value, adjusts the electrical resistance of the biological subject to a lower value, or alters the quantum mechanical effect of the biological subject.
In some embodiments, the at least one transducer is placed alongside a side wall of the channel or chamber, and is configured to apply a pulsed electrical voltage to the biological subject passing through the channel.
In some further embodiments, the biological subject is the blood sample, and the applied voltage is configured to impact an electrical field, charge distribution, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, or membrane potential of the blood sample.
In some other embodiments, the transducer is a generator configured to apply at least one type of energy or field onto the biological subject.
In some further embodiment, the generator is looped around the channel in which the biological subject flows through or stays statically for a desired period of time. The energy may comprise physical energy, bio-physical energy, bio-chemical energy, electrical energy, electro-magnetic energy, magnetic energy, optical energy, acoustic energy, thermal energy, mechanical energy, gravitational field energy, quantum mechanical energy, or radiation energy. Also, the energy can be applied in constant, alternating, or pulsed form.
In some embodiments, the channel applying the energy is a coil surrounding channel.
In some embodiments, the apparatus comprises at least one optical energy generator, acoustic energy generator, mechanical force generator, gravitational field generator, quantum mechanical field generator, electrical field generator, electro-magnetic field generator, electrical voltage generator, thermal energy generator, or radiation energy generator mounted along the channel.
In some embodiments, the field comprises electrical field, magnetic field, electro-magnetic field, quantum field, mechanical force field, or gravitational field.
In still some other embodiments, the apparatus includes at least one ion injector connecting to the channel, wherein the ion injector is configured to add a desired amount of ions to the biological subject.
In some embodiments, the apparatus comprises one or more channels, and the one or more channels comprise one or more transducers on sidewalls, and one or more ion injectors connecting to the one or more channels, optionally through one or more small openings; wherein at least one transducer is configured to transmit a bio-physical energy to the biological subject, and the at least one ion injector is configured to add a desired amount of ions to the biological subject. In some further embodiments, the biological subject can be the blood sample. Moreover, the bio-physical energy can be an electrical pulse; and the added ions may comprise potassium ions. Such an apparatus can enhance an electrical conductivity of the blood sample, a net ion concentration, an electrolyte concentration, a net cell surface charge, a net DNA surface charge, a net RNA surface charge, a net protein surface charge, a net electrical charge in the blood sample, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, a membrane potential, a membrane voltage, or a polarization of membrane potential.
In some embodiments, the apparatus further includes at least one detector placed partially or completely alongside one or more side walls of the channel or chamber, wherein the at least one detector is configured to detect at least one bio-physical or physical property of the biological subject.
In some embodiments, the bio-physical or physical signal exist and can be detected in both the biological subject from a healthy individual, the biological subject from a non-cancer disease patient, and the biological subject from a patient with any of the cancers to be treated; and the detected signals between the healthy individual, the non-cancer disease patient, and the patient with the cancers are different.
In some embodiments, the apparatus comprises one or more channels, and one or more detectors, one or more ion injectors, and one or more generators in or around the channels; wherein the one or more generators are configured to apply at least one type of energy or field onto the biological subject; the one or more detectors are configured to detect one or more bio-physical or physical properties of the biological subject, and each of the one or more ion injectors is configured to add a desired amount of ions to the biological subject.
In some embodiments, the apparatus is capable of simultaneously impacting two or more cancer statuses, each involving converting at least a property of the biological subject from a cancer state to an earlier cancer stage, a pre-cancer state, a non-cancer disease state or a healthy state; converting at least one property of the biological subject from a pre-cancer state to a non-cancer disease state or healthy state; or converting at least one property of the biological subject from a non-cancer disease state to a healthy state.
In some embodiments, the property comprises a physical property, bio-physical property, bio-chemical property, protein property, cellular property, molecular property, genomic property, or immunology property. In some other embodiments, the property comprises an optical property, acoustical property, thermal property, quantum property, gravitational property, mechanical property, electrical property, magnetic property, or electro-magnetic property.
In some embodiments, the apparatus is configured to impact at least one of the following properties in the biological subject: electrolyte concentration and distribution, potassium ion concentration and distribution, sodium ion concentration and distribution, chloride ion concentration and distribution, calcium ion concentration and distribution, net charge in extracellular region, ion concentration in extracellular, gravitational field, mechanical field, quantum mechanical field, magnetic field, electro-magnetic field, electrical field, electrical current, electrical resistance, electrical conductance, capacitance, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, membrane voltage, and membrane potential.
In some embodiments, the apparatus is fabricated by integrated circuit technologies. The integrated circuit technologies may comprise thin film deposition, lithography, etch, diffusion, ion implantation, annealing, cleaning, or polishing processes.
In some embodiments, the apparatus comprises semiconductor, electrically insulating, and electrically conductive materials. More specifically, the apparatus may comprise a material selected from group consisting of silicon, germanium, glass, silicon dioxide, silicon nitride, poly-silicon, tungsten, aluminum, copper, gold, and silicon carbide.
Also within the scope of this invention is a method for treating or slowing progression of two or more types of cancer in a patient, comprising administering to the patient thereof a therapeutic agent that enhances or increase the level of a biophysical property at the microscopic level of the patient, wherein the level of the biophysical property simultaneously impacts the status of each of the two or more type cancers.
In some embodiments, the therapeutic agent is administered orally or by intravenous injection.
In some embodiments, the biophysical property is an electronic property. For instance, the electronic property is surface charge, surface potential, resting potential, electrical current, electrical field distribution, surface charge distribution, cell electronic properties, cell surface electronic properties, dynamic changes in electronic properties, dynamic changes in cell electronic properties, dynamic changes in cell surface electronic properties, dynamic changes in surface electronic properties, membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, electronic properties of cell membranes, dynamic changes in electronic properties of membrane surface, dynamic changes in electronic properties of cell membranes, electrical dipole, electrical quadruple, oscillation in electrical signal, electrical current, capacitance, three-dimensional electrical or charge cloud distribution, electrical properties at telomere of DNA and chromosome, DNA surface charge, DNA surrounding media electrical properties, quantum mechanical effects, capacitance, or impedance.
In some embodiments, physical or bio-physical detection method may be combined with other detection methods to achieve unique and much more comprehensive detection performance results. Examples of other detection methods to be combined include but not limited to bio-chemistry, immunological, genomic, circulating tumor cell, and imaging detection technologies. Physical or bio-physical detection tends to be simpler and cost effective tests, which gives more fundamental, earlier and overall (more global in terms of covering common properties of more types of cancer/tumor, such as brain tumor and esophageal cancer) information about cancer occurrence, while other tests tend to be more complex and provide additional, more specific information - such as more specific and/or detailed information about a given type of cancer and its location. The combination tests can yield non-linearly more information. For instance, physical or bio-physical cancer detection method can be combined with bio-marker tests to obtain both (a) earlier stage cancer or pre-cancer information (with physical or bio-physical detection method), as well as more specific cancer type information including location or organ information (with bio-marker tests). In an immunological measurement, bio-markers may include alpha-fetoprotein, prostate specific antigen, arcinoembryonic antigen, carbohydrate antigen 50, carbohydrate antigen 242, carbohydrate antigen 125, carbohydrate antigen 153, carbohydrate antigen 199, and carbohydrate antigen 724. The above combination test approach can be called platform approach. In this work, it is called CDA (cancer differentiation analysis) platform, while physical or bio-physical detection approach alone is called CDA method.
In some embodiments, the cancers may include lung cancer, liver cancer, colorectal cancer, breast cancer, gastric cancer, esophageal cancer, brain tumor, prostate cancer, blood cancer, intestine cancer, stomach cancer, cervical cancer, ovarian cancer, rectum cancer, colon cancer, nasopharyngeal cancer, cardiac carcinoma, uterine cancer, oophoroma, pancreatic cancer, or circulating tumor cells.
In still another aspect, this invention provides a therapeutic agent for treating or slowing progression of two or more types of cancer in a patient, comprising a component that alters or enhances an electronic property of the patient.
Examples of the component include electrolytes or components which release electrolytes. The component may enhance electrical current, electro-static charges on DNA surfaces, and/or electrical conductance, reduces electrical resistance, and/or alters quantum mechanical effect.
The present invention can detect the presence of multiple types of diseases, including multiple types of cancer, with a single test. Examples of cancer comprise breast cancer, lung cancer, esophageal cancer, intestine cancer, cancer related to blood, liver cancer, stomach cancer, cervical cancer, ovarian cancer, rectum cancer, colon cancer, nasopharyngeal cancer, cardiac carcinoma, uterine cancer, oophoroma, pancreatic cancer, prostate cancer, brain tumor, and circulating tumor cells; examples of the inflammatory disease include acne vulgaris, asthma, autoimmune diseases, autoinflammatory diseases, celiac disease, chronic prostatitis, diverticulitis, glomerulonephritis, hidradenitis suppurativa, hypersensitivities, inflammatory bowel diseases, interstitial cystitis, otitis, pelvic inflammatory disease, reperfusion injury, rheumatic fever, rheumatoid arthritis, sarcoidosis, transplant rejection, and tasculitis; examples of the lung disease include asthma, chronic obstructive pulmonary disease, chronic bronchitis, emphysema, acute bronchitis, cystic fibrosis, pneumonia, tuberculosis, pulmonary edema, acute respiratory distress syndrome, pneumoconiosis, interstitial lung disease, pulmonary embolism, and pulmonary hypertension; examples of the diabetes include Type 1 diabetes, Type 2 diabetes, and gestational diabetes; examples of the heart disease include coronary artery disease, enlarged heart (cardiomegaly), heart attack, irregular heart rhythm, atrial fibrillation, heart rhythm disorders, heart valve disease, sudden cardiac death, congenital heart disease, heart muscle disease (cardiomyopathy), dilated cardiomyopathy, hypertrophic cardiomyopathy, restrictive cardiomyopathy, pericarditis, pericardial effusion, marfan syndrome, and heart murmurs; examples of the liver disease include fascioliasis, hepatitis, alcoholic liver disease, fatty liver disease (hepatic steatosis), hereditary diseases, Gilbert's syndrome, cirrhosis, primary biliary cirrhosis, primary sclerosing cholangitis, and Budd-Chiari syndrome; examples of the gastric disease include gastritis, gastric polyp, gastric ulcer, benign tumor of stomach, acute gastric mucosa lesion, antral gastritis, and gastric stromal tumors; examples of the biliary disease include calculus of bile duct, cholecystolithiasis, cholecystitis, cholangiectasis, cholangitis, and gallbladder polyps; the cardiovascular disease comprises coronary artery disease, peripheral arterial disease, cerebrovascular disease, renal artery stenosis, aortic aneurysm, cardiomyopathy, hypertensive heart disease, heart failure, pulmonary heart disease, cardiac dysrhythmias, endocarditis, inflammatory cardiomegaly, myocarditis, valvular heart disease, congenital heart disease, rheumatic heart disease, coronary artery disease, peripheral arterial disease, cerebrovascular disease, and renal artery stenosis.
A further aspect of this invention provides an apparatus for treating a disease of the biological subject, comprising a coil surrounding channel, in which the biological subject passes through, wherein the coil surrounding channel is configured to apply at least one type of energy or field onto the biological subject.
Ins some embodiments, the coil surrounding channel comprises a multiple-layer structure. For instance, the coil may comprise: a top conductive layer, a middle layer further comprising conductive plugs, and a bottom conductive layer; wherein the top conductive layer, the conductive plugs, and the bottom conductive layer are surrounded by one or more insulating materials; and the conductive plugs connects the top conductive layer and the bottom conductive layer, thereby forming the coil surrounding the channel.
In some embodiments, the apparatus simultaneously treats two or more types of cancer. Examples of the cancers include, but are not limited to, lung cancer, liver cancer, colorectal cancer, breast cancer, gastric cancer, esophageal cancer, brain tumor, prostate cancer, blood cancer, intestine cancer, stomach cancer, cervical cancer, ovarian cancer, rectum cancer, colon cancer, nasopharyngeal cancer, cardiac carcinoma, uterine cancer, oophoroma, pancreatic cancer, or circulating tumor cells.
In some embodiments, the energy comprises physical energy, bio-physical energy, bio-chemical energy, electrical energy, electro-magnetic energy, magnetic energy, optical energy, acoustic energy, thermal energy, mechanical energy, gravitational field energy, quantum mechanical energy, or radiation energy. The energy can be applied in constant, alternating, or pulsed form.
In some embodiments, the field comprises electrical field, magnetic field, electro-magnetic field, quantum field, mechanical force field, or gravitational field.
In some embodiments, the apparatus further includes at least one detector placed partially or completely in the channel, wherein the at least one detector is configured to detect at least one bio-physical or physical signal of the biological subject; and an ion injector connecting to the channel, wherein the ion injector is configured to add a desired amount of ions to the biological subject.
In some embodiments, the apparatus includes one or more coil surrounding channels, one or more detectors, and one or more ion injectors; wherein the one or more coil surrounding channels are configured to apply at least one type of energy or field onto the biological subject; the one or more detectors are configured to detect one or more bio-physical or physical properties of the biological subject, and each of the one or more ion injectors is configured to add a desired amount of ions to the biological subject.
In some embodiments, the apparatus is capable of simultaneously impacting two or more cancer statuses, each involving converting at least a property of the biological subject from a cancer state to an earlier cancer stage, a pre-cancer state, a non-cancer disease state or a healthy state; converting at least one property of the biological subject from a pre-cancer state to a non-cancer disease state or healthy state; or converting at least one property of the biological subject from a non-cancer disease state to a healthy state.
For instance, the property comprises a physical property, bio-physical property, bio-chemical property, protein property, cellular property, molecular property, genomic property, or immunology property. The property may also comprise an optical property, acoustical property, thermal property, gravitational property, mechanical property, quantum property, electrical property, magnetic property, or electro-magnetic property.
In some embodiments, the apparatus is configured to impact at least one of the following properties in the biological subject: electrolyte concentration and distribution, potassium ion concentration and distribution, sodium ion concentration and distribution, chloride ion concentration and distribution, calcium ion concentration and distribution, net charge in extracellular region, ion concentration in extracellular, gravitational field, mechanical field, quantum mechanical field, magnetic field, electro-magnetic field, electrical field, electrical current, electrical resistance, electrical conductance, capacitance, and membrane ion channel properties, resting membrane potential, trans-membrane potential, depolarized trans-membrane potential, membrane voltage, membrane potential.
The biological subject can be a liquid sample of a mammal, such as a blood sample, a urine sample, or a sweat sample of the mammal. Such a biological subject may include blood, proteins, red blood cells, while blood cells, T cells, other cells, gene mutations, quantum mechanical effects, DNA, RNA, or other biological entities.
In some embodiments, the apparatus is fabricated by integrated circuit technologies, including, e.g., thin film deposition, lithography, etch, diffusion, ion implantation, annealing, cleaning, or polishing processes. The apparatus may be made of semiconductor, electrically insulating, and/or electrically conductive materials.
Still within the scope of this invention is a method of fabricating a micro-device for treating a disease, wherein the method uses microelectronics process and comprises the following step: fabricating a coil surrounding channel by semiconductor or integrated circuit process, wherein the coil surrounding channel is configured to apply at least one type of energy or field onto a biological subject passing through or staying in the channel.
For instance, the method may comprise the following steps: providing a substrate; depositing material A onto the surface region of the substrate, and patterning or etching material A to form first recessed areas; depositing material B onto the surface region of material A and filing into the first recessed areas; etching or polishing material B to remove material B from the top surface of material A and leaving sufficient amount of material B in the recessed areas coplanar with the top surface of material A; depositing material C onto the surfaces of materials A and B to form a thin layer; depositing layer material D onto the surface of material C, and patterning material D to form second recessed areas; depositing sacrificial material E, and etching or polishing material E to remove material E from the top surface of material D and leaving sufficient amount of material E in the second recessed areas coplanar with the top surface of material D; depositing material F onto the surfaces of materials D and E, and using an etching process to etch a small hole through material F; using a further etching process to etch off sacrificial material E, thereby forming micro-channels in material D, depositing material G onto the surface of material F to form a thin etch stop layer; depositing material H onto the surface of material G; using pattering and etching processes to form trench areas in material H, and deep hole areas through materials G, F, D, and C stopping on the top surface of material B; and depositing material I into the trench areas and the deep hole areas, thereby contacting material B, wherein material I and material B together form a coil, which surrounds the micro-channels formed in material D.
In some embodiments, the substrate is a semiconductor or insulating substrate.
In some embodiments, material A is an insulating material. Material B can be a conductive material, and can be polished via chemical mechanical polishing (CMP). Material C may comprise silicon nitride or poly-silicon.
In some embodiments, material D may be an insulating material, optionally being the same as material A. In some embodiments, material E is polished via chemical mechanical polishing (CMP).
Still in some embodiments, material F is an insulating material, optionally same as material A. Materials A, D, and F may be the same insulating material. Material G may also comprise silicon nitride or poly-silicon, and optionally being the same as material C.
In some embodiments, material H is an insulating material, optionally the same material as material A. Materials A, D, F, and H may be the same material.
In some embodiments, the formation of the trench areas and the deep hole areas further comprises: patterning and etching material H to form the trench areas, and then further patterning and etching to form the deep hole areas through materials G, F, D, and C, stopping on the top surface of material B. For instance, the hole area is etched by different etch chemistries, e.g., with one chemistry for materials G and C, and another chemistry for materials F and D.
Alternatively, the trench areas and the deep hole areas are formed by using a double lithography exposure, for trench area and deep hole area respectively, and doing a double-etch, thereby etching the trench area first and then the hole area.
In some embodiments, material I is a conductive material, optionally being the same as material B.
Still in another aspect, the present application provides a method of fabricating a micro device with a coil surrounding channel, comprising the following steps: providing a substrate; depositing material A1 onto the surface region of the substrate, and patterning or etching material A1 to form first recessed areas; depositing material B1 onto the surface region of material A1 and filing into the first recessed areas; etching or polishing material B1 to remove material B1 from the top surface of material A1 and leaving sufficient amount of material B1 in the recessed areas coplanar with the top surface of material A1 ; depositing material C1 onto the surfaces of materials A1 and B1 to form a thin layer; depositing layer material D1 onto the surface of material C1 , and patterning material D1 to form second recessed areas; depositing sacrificial material E1 , and etching or polishing material E1 to remove material E from the top surface of material D1 and leaving sufficient amount of material E1 in the second recessed areas coplanar with the top surface of material D1 ; depositing material F onto the surfaces of materials D1 and E1, and using an etching process to etch a small hole through material F1; using a further etching process to etch off sacrificial material E1 , thereby forming micro-channels in material D1, depositing material G1 onto the surface of material F1 to form a thin etch stop layer; depositing material H1 onto the surface of material G1 ; using pattering and etching processes to form trench areas in material H1, and deep hole areas through materials G1, F1, D1, and C1 stopping on the top surface of material B1; and depositing material I1 into the trench areas and the deep hole areas, thereby contacting material B, wherein material I1 and material B1 together form a coil, which surrounds the micro-channels formed in material D1.
In some embodiments, the coil surrounding channel is configured to apply at least one type of energy or field onto a biological subject passing through or staying in the channel. In some embodiments, the substrate is a semiconductor or insulating substrate.
In some embodiments, each of materials A1 , D1, F1, and H1 is an insulating material (optionally the same material). Each of materials B1 and I1 can be a conductive material (optionally the same material). At least one of materials C1 and G1 may comprise nitride or poly-silicon. Materials C1 and G1 may be the same.
In some embodiments, at least one of materials B1 and El is polished via chemical mechanical polishing (CMP).
In some embodiments, the formation of the trench areas and the deep hole areas further comprises: patterning and etching material H1 to form the trench areas, and then further patterning and etching to form the deep hole areas through materials G1, F1, D1, and C1, stopping on the top surface of material B 1. The hole area may be etched by different etch chemistries, e.g., with one chemistry for materials G1 and C1, and another chemistry for materials F1 and D1.
Alternatively, the trench areas and the deep hole areas are formed by using a double lithography exposure, for trench area and deep hole area respectively, and doing a double-etch, thereby etching the trench area first and then the hole area.
As used herein, the term “biomarker” means a measurable indicator of the severity or presence of some disease state, but more generally a biomarker is anything that can be used as an indicator of a particular disease state or some other physiological state of an organism. A biomarker can be a substance that is introduced into an organism as a means to examine organ function or other aspects of health. For example, rubidium chloride is used in isotopic labeling to evaluate perfusion of heart muscle. It can also be a substance whose detection indicates a particular disease state, for example, the presence of an antibody may indicate an infection. More specifically, a biomarker indicates a change in expression or state of a protein that correlates with the risk or progression of a disease, or with the susceptibility of the disease to a given treatment. Biomarkers can be specific cells, molecules, or genes, gene products, enzymes, or hormones.
As used herein, the term “or” is meant to include both “and” and “or”. It may be interchanged with “and/or.”
As used herein, a singular noun is meant to include its plural meaning. For instance, a micro device can mean either a single micro device or multiple micro-devices.
As used herein, the term “patterning” means shaping a material into a certain physical form or pattern, including a plane (in which case “patterning” would also mean “planarization”).
As used herein, the term “a biocompatible material” refers to a material that is intended to interface with a living organism or a living tissue and can function in intimate contact therewith. When used as a coating, it reduces the adverse reaction a living organism or a living tissue has against the material to be coated, e.g., reducing the severity or even eliminating the rejection reaction by the living organism or living tissue. As used herein, it encompasses both synthetic materials and naturally occurring materials. Synthetic materials usually include biocompatible polymers, made either from synthetic or natural starting materials, whereas naturally occurring biocompatible materials include, e.g., proteins or tissues.
As used herein, the term “a biological subject” or “a biological sample” for analysis or test or diagnosis refers to the subject to be analyzed by a disease detection apparatus. It can be a single cell, a single biological molecular (e.g., DNA, RNA, or protein), a single biological subject (e.g., a single cell or virus), any other sufficiently small unit or fundamental biological composition, a sample of a subject's or patient's organ or tissue that may having a disease or disorder, or a patient.
As used herein, the term “disease” is interchangeable with the term “disorder” and generally refers to any abnormal microscopic property or condition (e.g., a physical condition) of a biological subject (e.g., a mammal or biological species).
As used herein, the term “subject” generally refers to a mammal, e.g., a human person.
As used herein, the term “microscopic level” refers to the subject being analyzed by the disease detection apparatus of this invention is of a microscopic nature and can be a single cell, a single biological molecular (e.g., DNA, RNA, or protein), a single biological subject (e.g., a single cell or virus), and other sufficiently small unit or fundamental biological composition.
As used herein, an “apparatus” or a “micro-device” or “micro device” can be any of a wide range of materials, properties, shapes, and degree of complexity and integration. The term has a general meaning for an application from a single material to a very complex device comprising multiple materials with multiple sub units and multiple functions. The complexity contemplated in the present invention ranges from a very small, single particle with a set of desired properties to a fairly complicated, integrated unit with various functional units contained therein. For example, a simple micro-device could be a single spherical article of manufacture of a diameter as small as 100 angstroms with a desired hardness, a desired surface charge, or a desired organic chemistry absorbed on its surface. A more complex micro device could be a 1 millimeter device with a sensor, a simple calculator, a memory unit, a logic unit, and a cutter all integrated onto it. In the former case, the particle can be formed via a fumed or colloidal precipitation process, while the device with various components integrated onto it can be fabricated using various integrated circuit manufacturing processes. In some places, a micro-device or micro device represents a sub-equipment unit.
As used herein, the term “parameter” refers to a particular detection target (e.g., a property of microscopic level, physical property such as hardness, viscosity, current, or voltage, or chemical property such as pH value) of the biological subject to be detected, and can include micro-level property.
As used herein, the term “level” refers to chemical composition (including biochemical composition such as protein, genetic materials, e.g., DNA and RNA), cellular classification, or molecular classification of the biological subject to be detected.
As used herein, the term “component” refers a lower division or building block of a level described above. For instance, a protein level can include such components as alpha-feto protein or sugar protein; and the level of a cellular classification can include such components as surface voltage and membrane composition.
As used herein, if not specifically defined, a “channel” or “chamber” can be either an inter-unit channel or an intra-unit channel.
Biological subjects that can be detected by the apparatus include, e.g., blood, urine, saliva, tear, and sweat. The detection results can indicate the possible occurrence or presence of a disease (e.g., one in its early stage) in the biological subject.
As used herein, the term “absorption” typically means a physical bonding between the surface and the material attached to it (absorbed onto it, in this case). On the other hand, the word “adsorption” generally means a stronger, chemical bonding between the two. These properties are very important for the present invention as they can be effectively used for targeted attachment by desired micro devices for measurement at the microscopic level.
As used herein, the term “contact” (as in “the first micro-device contacts a biologic entity”) is meant to include both “direct” (or physical) contact and “non-direct” (or indirect or non-physical) contact. When two subjects are in “direct” contact, there is generally no measurable space or distance between the contact points of these two subjects; whereas when they are in “indirect” contact, there is a measurable space or distance between the contact points of these two subjects.
As used herein, the term “probe” or “probing,” in addition to its dictionary meaning, could mean applying a signal (e.g., an acoustic, optical, magnetic, chemical, electrical, electro-magnetic, bio-chemical, bio-physical, or thermal signal) to a subject and thereby stimulating the subject and causing it to have some kind of intrinsic response.
As used herein, the term “thermal property” refers to temperature, freezing point, melting point, evaporation temperature, glass transition temperature, or thermal conductivity.
As used herein, the term “optical property” refers to reflection, optical absorption, optical scattering, wave length dependent properties, color, luster, brilliance, scintillation, or dispersion.
As used herein, the term “electrical property” refers to surface charge, surface potential, electrical field, charge distribution, electrical field distribution, resting potential, action potential, or impedance of a biological subject to be analyzed.
As used herein, the term “magnetic property” refers to diamagnetic, paramagnetic, or ferromagnetic.
As used herein, the term “electromagnetic property” refers to property that has both electrical and magnetic dimensions.
As used herein, the term “acoustical property” refers to the characteristics found within a structure that determine the quality of sound in its relevance to hearing. It can generally be measured by the acoustic absorption coefficient. See, e.g., U.S. Pat. No. 3,915,016, for means and methods for determining an acoustical property of a material; T. J. Cox et al., Acoustic Absorbers and Diffusers, 2004, Spon Press.
As used herein, the term “biological property” is meant to generally include chemical and physical properties of a biological subject.
As used herein, the term “chemical property” refers to pH value, ionic strength, or bonding strength within the biological sample.
As used herein, the term “physical property” refers to any measurable property the value of which describes a physical system's state at any given moment in time. The physical properties of a biological sample may include, but are not limited to absorption, albedo, area, brittleness, boiling point, capacitance, color, concentration, density, dielectrical, electrical charge, electrical conductivity, electrical impedance, electrical field, electrical potential, emission, flow rate, fluidity, frequency, inductance, intrinsic impedance, intensity, irradiance, luminance, luster, malleability, magnetic field, magnetic flux, mass, melting point, momentum, permeability, permittivity, pressure, radiance, solubility, specific heat, strength, temperature, tension, thermal conductivity, flow rate, velocity, viscosity, volume, surface area, shape, and wave impedance.
As used herein, the term “mechanical property” refers to strength, hardness, flow rate, viscosity, toughness, elasticity, plasticity, brittleness, ductility, shear strength, elongation strength, fracture stress, or adhesion of the biological sample.
As used herein, the term “disturbing signal” has the same meaning as “probing signal” and “stimulating signal.”
As used herein, the term “disturbing unit” has the same meaning as “probing unit” and “stimulating unit.”
As used herein, the term “conductive material” (or its equivalent “electrical conductor”) is a material which contains movable electrical charges. A conductive material can be a metal (e.g., copper, silver, or gold) or non-metallic (e.g., graphite, solutions of salts, plasmas, or conductive polymers). In metallic conductors, such as copper or aluminum, the movable charged particles are electrons (see electrical conduction). Positive charges may also be mobile in the form of atoms in a lattice that are missing electrons (known as holes), or in the form of ions, such as in the electrolyte of a battery.
As used herein, the term “electrically insulating material” (also known as “insulator” or “dielectric”) refers to a material that resists the flow of electrical current. An insulating material has atoms with tightly bonded valence electrons. Examples of electrically insulating materials include glass or organic polymers (e.g., rubber, plastics, or Teflon).
As used herein, the term “semiconductor” (also known as “semiconducting material”) refers to a material with electrical conductivity due to electron flow (as opposed to ionic conductivity) intermediate in magnitude between that of a conductor and an insulator. Examples of inorganic semiconductors include silicon, silicon-based materials, and germanium. Examples of organic semiconductors include such aromatic hydrocarbons as the polycyclic aromatic compounds pentacene, anthracene, and rubrene; and polymeric organic semiconductors such as poly(3-hexylthiophene), poly(p-phenylene vinylene), polyacetylene and its derivatives. Semiconducting materials can be crystalline solids (e.g., silicon), amorphous (e.g., hydrogenated amorphous silicon and mixtures of arsenic, selenium and tellurium in a variety of proportions), or even liquid.
As used herein, the term “biological material” has the same meaning of “biomaterial” as understood by a person skilled in the art. Without limiting its meaning, biological materials or biomaterials can generally be produced either in nature or synthesized in the laboratory using a variety of chemical approaches utilizing organic compounds (e.g., small organic molecules or polymers) or inorganic compounds (e.g., metallic components or ceramics). They generally can be used or adapted for a medical application, and thus comprise whole or part of a living structure or biomedical device which performs, augments, or replaces a natural function. Such functions may be benign, like being used for a heart valve, or may be bioactive with a more interactive functionality such as hydroxyl-apatite coated hip implants. Biomaterials can also be used every day in dental applications, surgery, and drug delivery. For instance, a construct with impregnated pharmaceutical products can be placed into the body, which permits the prolonged release of a drug over an extended period of time. A biomaterial may also be an autograft, allograft, or xenograft which can be used as a transplant material. All these materials that have found applications in other medical or biomedical fields can also be used in the present invention.
As used herein, the term “microelectronic technology or process” generally encompasses the technologies or processes used for fabricating micro-electronic and optical-electronic components. Examples include lithography, etching (e.g., wet etching, dry etching, or vapor etching), oxidation, diffusion, implantation, annealing, film deposition, cleaning, direct-writing, polishing, planarization (e.g., by chemical mechanical polishing), epitaxial growth, metallization, process integration, simulation, or any combinations thereof. Additional descriptions on microelectronic technologies or processes can be found in, e.g., Jaeger, Introduction to Microelectronic Fabrication, 2nd Ed., Prentice Hall, 2002; Ralph E. Williams, Modern GaAs Processing Methods, 2nd Ed., Artech House, 1990; Robert F. Pierret, Advanced Semiconductor Fundamentals, 2nd Ed., Prentice Hall, 2002; S. Campbell, The Science and Engineering of Microelectronic Fabrication, 2nd Ed., Oxford University Press, 2001, the contents of all of which are incorporated herein by reference in their entireties.
As used herein, the term “selective” as included in, e.g., “patterning material B using a microelectronics process selective to material A”, means that the microelectronics process is effective on material B but not on material A, or is substantially more effective on material B than on material B (e.g., resulting in a much higher removal rate on material B than on material A and thus removing much more material B than material A).
As used herein, the term “carbon nano-tube” generally refers to as allotropes of carbon with a cylindrical nanostructure. See, e.g., Carbon Nanotube Science, by P. J. F. Harris, Cambridge University Press, 2009, for more details about carbon nano-tubes.
Through the use of a single micro-device or a combination of micro-devices integrated into a disease detection apparatus, the disease detection capabilities can be significantly improved in terms of sensitivity, specificity, speed, cost, apparatus size, functionality, and ease of use, along with reduced invasiveness and side-effects. A large number of micro-device types capable of measuring a wide range of microscopic properties of biological sample for disease detection can be integrated and fabricated into a single detection apparatus using micro-fabrication technologies and novel process flows disclosed herein. While for the purposes of demonstration and illustration, a few novel, detailed examples have been shown herein on how microelectronics or nano-fabrication techniques and associated process flows can be utilized to fabricate highly sensitive, multi-functional, and miniaturized detection devices, the principle and general approaches of employing microelectronics and nano-fabrication technologies in the design and fabrication of high performance detection devices have been contemplated and taught, which can and should be expanded to various combination of fabrication processes including but not limited to thin film deposition, patterning (lithography and etch), planarization (including chemical mechanical polishing), ion implantation, diffusion, cleaning, various materials, and various process sequences and flows and combinations thereof.
While existing cancer screening tests and treating methods lack the ability to effectively detect and/or impact multiple types of cancer at the same time (e.g., in one test), the present invention provides a novel technology that can simultaneously detect (or even treat) a significantly increased number of diseases—including cancers or pre-cancer diseases (e.g., over 20 types of cancer)—with higher speed, higher sensitivity and specificity (75%˜90% on over 20 types of cancer), simpler process, reduced costs, and no side effects. Compared to conventional technologies, the novel cancer treatment technology of this invention has a number of major, unexpected advantages—including, e.g., lower costs, far less side efforts, easier recovery, capability of cancer prevention, improved survival rate and ease of use. The cancer treatment of this invention can use a low dosage and/or a weak field and/or energy for treatment.
One aspect of the present invention relates to apparatus for detecting a disease in a biological subject in vivo or in vitro (e.g., human being, an organ, a tissue, or cells in a culture). Each apparatus comprises a delivery system, at least two sub-equipment units, and optionally a central control system. Each sub-equipment is capable of measuring at least a microscopic property of a biological sample. Accordingly, the apparatus of this invention can detect different parameters of the biological subject and provide accuracy, sensitivity, specificity, efficiency, non-invasiveness, practicality, conclusive, and speed in early-stage disease detection at reduced costs. In addition, the apparatus of this invention has some major advantages, such as reducing effective foot print (e.g., defined as function per unit space), reducing space for the medical devices, reducing overall cost, and providing conclusive and effective diagnosis by one device.
The delivery system can be a fluid delivery system. By the constant pressure fluid delivery system, microscopic biological subjects can be delivered onto or into one or more desired sub-equipment units of the apparatus.
As a key component of the apparatus, the micro-device should include means to perform at least the function of addressing, controlling, forcing, receiving, amplifying, or storing information from each probing address. As an example, the apparatus can further include a central control system for controlling the biological subject matter to be transported to one or more desired sub-equipment units and reading and analyzing a detected data from each sub-equipment unit. The central control system includes a controlling circuitry, an addressing unit, an amplifier circuitry, a logic processing circuitry, a memory unit, an application specific chip, a signal transmitter, a signal receiver, or a sensor.
In some embodiments, the fluid delivering system comprises a pressure generator, a pressure regulator, a throttle valve, a pressure gauge, and distributing kits. As examples of these embodiments, the pressure generator can include a motor piston system and a bin containing compressed gas; the pressure regulator (which can consist of multiple regulators) can down-regulate or up-regulate the pressure to a desired value; the pressure gauge feeds back the measured value to the throttle valve which then regulates the pressure to approach the target value.
The biological fluid to be delivered can be a sample of a biological entity to be detected for disease or something not necessarily to be detected for disease. In some embodiments, the fluid to be delivered is liquid (e.g., a blood sample or a lymph sample). The pressure regulator can be a single pressure regulator or multiple pressure regulators which are placed in succession to either down-regulate or up-regulate the pressure to a desired level, particularly when the initial pressure is either too high or too low for a single regulator to adjust to the desired level or a level that is acceptable for an end device or target.
Optionally, the apparatus includes additional features and structures to deliver a second liquid solution containing at least an enzyme, protein, oxidant, reducing agent, catalyst, radio-active component, optical emitting component, or ionic component. This second liquid solution can be added to the sample to be measured before or during sorting of the biological subject sample to be measured, or before or during the measurement (i.e., detection) of the biological subject sample, for the purposes of further enhancing the apparatus' measurement sensitivity.
In some other embodiments, the system controller includes a pre-amplifier, a lock-in amplifier, an electrical meter, a thermal meter, a switching matrix, a system bus, a nonvolatile storage device, a random access memory, a processor, or a user interface. The interface can include a sensor which can be a thermal sensor, a flow meter, an optical sensor, an acoustic detector, a current meter, an electrical sensor, a magnetic sensor, an electro-magnetic sensor, a pH meter, a hardness measurement sensor, an imaging device, a camera, a piezo-electrical sensor, a piezo-photronic sensor, a piezo-electro photronic sensor, an electro-optical sensor, an electro-thermal sensor, a bio-electrical sensor, a bio-marker sensor, a bio-chemical sensor, a chemical sensor, an ion emission sensor, a photo-detector, an x-ray sensor, a radiation material sensor, an electrical sensor, a voltage meter, a thermal sensor, a flow meter, or a piezo- meter.
In still some other embodiments, apparatus of this invention further includes a biological interface, a system controller, a system for reclaiming or treatment medical waste. The reclaiming and treatment of medical waste can be performed by the same system or two different systems.
Another aspect of this invention provides apparatus for interacting with a cell, which include a device for sending a signal to the cell and optionally receiving a response to the signal from the cell.
In some embodiments, the interaction with the cell can be probing, detecting, sorting, communicating with, treating, or modifying with a coded signal that can be a thermal, optical, acoustical, biological, chemical, electro-mechanical, electro-chemical, electro-optical, bio-electro-optical, bio-thermal optical, electro-chemical optical, electro-chemical-mechanical, bio-chemical, bio-mechanical, bio-electro-mechanical, bio-electro-chemical, bio-electro-chemical-mechanical, electric, magnetic, electro-magnetic, physical, or mechanical signal, or a combination thereof.
In some other embodiments, the device or the sub-equipment unit contained in the apparatus can include multiple surfaces coated with one or more elements or combinations of elements, and a control system for releasing the elements. In some instances, the control system can cause release of the elements from the device surface via an energy including but not limited to thermal energy, mechanical energy, gravitational field energy, quantum mechanical energy, optical energy, acoustic energy, electrical energy, electro-magnetic energy, magnetic energy, radiation energy, or mechanical energy in a controlled manner. The energy can be in the pulsed form at desired frequencies.
In some other embodiments, the device or the sub-equipment unit contained in the apparatus includes a first component for storing or releasing one element or a combination of elements onto the surface of the cell or into the cell; and a second component for controlling the release of the elements (e.g., a circuitry for controlling the release of the elements). The elements can be a biological component, a chemical compound, ions, catalysts, Ca, C, Cl, Co, Cu, H, I, Fe, Mg, Mn, N, O, P, F, K, Na, S, Zn, or a combination thereof. The signal, pulsed or constant, can be in the form of a released element or combination of elements, and it can be carried in a liquid solution, gas, or a combination thereof. In some instances, the signal can be at a frequency ranging from about 1×10−4 Hz to about 100 MHz or ranging from about 1×10−4 Hz to about 10 Hz, or at an oscillation concentration ranging from about 1.0 nmol/L to about 10.0 mmol/L. Also, the signal comprises the oscillation of a biological component, a chemical compound, Ca, C, Cl, Co, Cu, H, I, Fe, Mg, Mn, N, O, P, F, K, Na, S, Zn, or a combination thereof, e.g., at desired oscillating frequencies.
In some embodiments, the signal to be sent to the cell can be in the form of oscillating element, compound, or an oscillating density of a biological component, and a response to the signal from the cell is in the form of oscillating element, compound, or an oscillating density of a biological component.
In some embodiments, the device or the sub-equipment unit can be coated with a biological film, e.g., to enhance compatibility between the device and the cell.
In some other embodiments, the device or the sub-equipment unit can include components for generating a signal to be sent to the cell, receiving a response to the signal from the cell, analyzing the response, processing the response, and interfacing between the device and the cell.
Still another aspect of this invention provides devices or sub-equipment units each including a micro-filter, a shutter, a cell counter, a selector, a micro-surgical kit, a timer, and a data processing circuitry. The micro-filter can discriminate abnormal cells by a physical property (e.g., dimension, shape, or velocity), mechanical property, electric property, magnetic property, electro-magnetic, thermal property (e.g., temperature), optical property, acoustical property, biological property, chemical property, electro-chemical property, bio-chemical property, bio-electro-chemical property, bio-electro-mechanical property, or electro-mechanical property. The devices each can also include one or more micro-filters. Each of these micro-filters can be integrated with two cell counters, one of which is installed at the entrance of each filter well, while the other is installed at the exit of each filter well. The shape of the micro-filter's well is rectangle, ellipse, circle, or polygon; and the micro-filter's dimension ranges from about 0.1 μm to about 500 μm or from about 5 um to about 200 um. As used herein, the term “dimension” means the physical or feature size of the filter opening, e.g., diameter, length, width, or height. The filter can be coated with a biological or bio-compatible film, e.g., to enhance compatibility between the device and the cell.
In addition to separation of biological entity by its size and other physical features, the filter can also contain additional features and functions to perform biological entity separation via other properties, which comprise of mechanical property, electric property, magnetic property, electro-magnetic, thermal property (e.g., temperature), optical property, acoustical property, biological property, chemical property, electro-chemical property, bio-chemical property, bio-electro-chemical property, bio-electro-mechanical property, and electro-mechanical property.
In some embodiments of these devices, the shutter sandwiched by two filter membranes can be controlled by a timer (thus time shutter). The timer can be triggered by the cell counter. For instance, when a cell passes through the cell counter of the filter entrance, the clock is triggered to reset the shutter to default position, and moves at a preset speed towards the cell pathway, and the timer records the time as the cell passes through the cell counter at the exit.
Still a further aspect of this invention provides methods for fabricating a micro-device with micro-trench and probe embedded in the micro-trench's sidewalls. A micro-trench is an unclosed tunnel (see, e.g.,
In some embodiments, the method further includes coupling two devices or sub-equipment units that are thus fabricated and symmetric (i.e., a flipped mirror) to form a detecting device with channels. The entrance of each channel can be optionally bell-mouthed, e.g., such that the size of channel's opening end (the entrance) is larger than the channel's body, thereby making it easier for a cell to enter the channel. The shape of each channel's cross-section can be rectangle, ellipse, circle, or polygon. The micro-trenches of the coupled two micro-devices can be aligned by the module of alignment marks designed on the layout of the micro-device. The dimension of the micro-trench can range from about 0.1 um to about 500 um.
Alternatively, the method can also include covering the micro-trench of the micro-device with a flat panel. Such a panel can comprise or be made with silicon, SiGe, SiO2, Al2O3, quartz, low optical loss glasses, or other optical materials. Examples of other potentially suitable optical materials include acrylate polymer, AgInSbTe, synthetic alexandrite, arsenic triselenide, arsenic trisulfide, barium fluoride, CR-39, cadmium selenide, caesium cadmium chloride, calcite, calcium fluoride, chalcogenide glass, gallium phosphide, GeSbTe, germanium, germanium dioxide, glass code, hydrogen silsesquioxane, Iceland spar, liquid crystal, lithium fluoride, lumicera, METATOY, magnesium fluoride, agnesium oxide, negative index metamaterials, neutron super-mirror, phosphor, picarin, poly(methyl methacrylate), polycarbonate, potassium bromide, sapphire, scotophor, spectralon, speculum metal, split-ring resonator, strontium fluoride, yttrium aluminum garnet, yttrium lithium fluoride, yttrium orthovanadate, ZBLAN, zinc selenide, and zinc sulfide.
In other embodiments, the method can further include integrating three or more sub-equipment units or devices thus fabricated to yield an enhanced device with an array of the channels.
Another aspect of this invention relates to a set of novel process flows for fabricating micro-devices (including micro-probes and micro-indentation probes) for their applications in disease detection by measuring microscopic properties of a biological sample. The micro-devices can be integrated into detection apparatus of this invention as sub-equipment units to measure one or more properties at microscopic levels. For example, a cancerous cell may have a different hardness (harder), density (denser), and elasticity than a normal cell.
Another aspect of this invention is to involve in cellular communications and regulate cellular decision or response (such as differentiation, dedifferentiation, cell division and cell death) with fabricated signals generated by the micro-devices disclosed herein. This could be further employed to detect and treat diseases.
Another aspect of the current application is that the inventive method or measured parameter in the method is a function of at least two levels F (level 1, level 2), where level 1 can be a biological entity such as protein and level 2 can be another biological entity such as genetics, where the measured signal strength of F (level 1, level 2) is greater than the sum of the signal containing only level 1 information f (level 1) and the signal containing only level 2 information f (level 2):
Signal strength of F (level 1, level 2)>signal strength of f (level 1)+signal strength of f (level 2)
The above novel feature and property can be extended to a measured parameter which is a function containing many levels F (level 1, level 2, level 3 level n). One novel and unobvious feature of this innovation is that the measured signal in a parameter containing multiple biological levels is synergistically enhanced over the measured signals with each signal containing a single biological level only. With this approach, the typically weak detection signal in disease detection such as cancer detection (especially in early stage cancer detection) can be effectively enhanced or magnified, making early disease detection possible and more effective.
To further enhance measurement capabilities, multiple micro-devices can be implemented into a piece of detection apparatus as sub-equipment units employing the time of flight technique, in which at least one probing micro-device and one sensing micro-device placed at a preset, known distance. The probing micro-device can apply a signal (e.g., a voltage, a charge, an electrical field, a laser beam, a thermal pulse, a train of ions, or an acoustic wave) to the biological sample to be measured, and the detection (sensing) micro-device can measure response from or of the biological sample after the sample has traveled a known distance and a desired period of time. For instance, a probing micro-device can apply an electrical charge to a cell first, and then a detection (sensing) micro-device subsequently measures the surface charge after a desired period of time (T) has lapsed and the cell has traveled a certain distance (L).
The micro-devices or the sub-equipment units contained in the apparatus of this invention can have a wide range of designs, structures, functionalities, flexibilities, and applications due to their diverse properties, high degree of flexibilities, and ability of integration, miniaturization, and manufacturing scalability. They include, e.g., a voltage comparator, a four point probe, a calculator, a logic circuitry, a memory unit, a micro cutter, a micro hammer, a micro shield, a micro dye, a micro pin, a micro knife, a micro needle, a micro thread holder, micro tweezers, a micro laser, a micro optical absorber, a micro mirror, a micro wheeler, a micro filter, a micro chopper, a micro shredder, micro pumps, a micro absorber, a micro signal detector, a micro driller, a micro sucker, a micro tester, a micro container, a signal transmitter, a signal generator, a friction sensor, an electrical charge sensor, a temperature sensor, a hardness detector, an acoustic wave generator, an optical wave generator, a heat generator, a micro refrigerator and a charge generator.
Further, it should be noted that advancements in manufacturing technologies have now made fabrications of a wide range of micro-devices and integration of various functions onto the same device highly feasible and cost effective. The typical human cell size is about 10 microns. Using state-of-the-art integrated circuit fabrication techniques, the minimum feature size defined on a micro-device can be as small as 0.1 micron or below. Thus, it is ideal to utilize the disclosed micro-devices for biological applications.
In terms of materials for the micro-devices in the apparatus of this invention, the general principle or consideration is the material's compatibility with a biological entity. Since the time in which a micro-device is in contact with a biological sample (e.g., a cell) may vary, depending on its intended application, a different material or a different combination of materials may be used to make the micro-device. In some special cases, the materials may dissolve in a given pH in a controlled manner and thus may be selected as an appropriate material. Other considerations include cost, simplicity, ease of use and practicality. With the significant advancements in micro fabrication technologies such as integrated circuit manufacturing technology, highly integrated devices with minimum feature size as small as 0.1 micron can now be made cost-effectively and commercially. One good example is the design and fabrication of micro electro mechanical devices (MEMS), which now are being used in a wide variety of applications in the electronics industry and beyond.
Good disease (cancer and non-cancer) detection results in terms of measurement sensitivity and specificity have been obtained on multiple types of cancer tested, demonstrating validity of the apparatus of this invention for improved ability to detect diseases (e.g., cancers), particularly in their early stages. The present invention provides novel “Cancer Differentiation Analysis” (CDA) liquid biopsy technology. The experimental results have also shown that multiple cancer types can be detected using the disclosed apparatus, which itself is an improvement over many existing detection apparatuses.
Specifically, studies utilizing the apparatus of this invention have been carried out on multiple types of cancer and non-cancer diseases (including an inflammatory disease, diabetes, a lung disease, a heart disease, a liver disease, a gastric disease, a biliary disease, or a cardiovascular disease). In these studies, whole blood samples were used within 5 days after being obtained and/or properly transported/stored in a 0.5-20° C. refrigerated environment. The samples of the control group were obtained from healthy people confirmed by physical examinations with normal AFP and CEA values (in normal ranges).
CDA value is obtained from an algorithm using calculation based on tested values from the studies. CDA value increases with risks of diseases. In other words, the higher the CDA values, the higher the risks of diseases.
As the above tables show, the CDA values are higher for various diseases (mid 40s) than those of control (healthy) group (around 36). Statistical analysis of CDA values for those two groups shows that there was a statistically significant difference in CDA values between those two groups. Accordingly, the studies above show that the apparatus and methods of this invention were able to distinguish some major diseases from control group, with sensitivity and specificity likely higher than existing technologies.
Set forth below are several illustrations or examples of apparatus of this invention containing a class of innovative micro-devices that are integrated as sub-equipment units.
To enhance detection speed and sensitivity, a large number of micro-devices can be integrated into a single apparatus of this invention. Each micro-device can be an independent sub-equipment unit in the apparatus. To achieve the above requirements, the detection apparatus should be optimized with its surface area maximized to contact the biological sample and with large number of micro-devices integrated on the maximized surface.
Instead of measuring a single property of a biological subject for disease diagnosis, various micro-devices can be integrated into a detection apparatus to detect multiple properties. Various micro-devices can constitute different sub-equipment units.
As illustrated herein, it is desirable to optimize the detection apparatus design to maximize measurement surface area, since the greater the surface area, the greater number of micro-devices that can be placed on the detection apparatus to simultaneously measure the sample, thereby increasing detection speed and also minimizing the amount of sample needed for the test.
Yet another aspect of this invention relates to a set of novel fabrication process flows for making micro-devices or sub-equipment units for disease detection purposes. Thus, a micro-device with two probes capable of measuring a range of properties (including mechanical and electrical properties) of biological samples is fabricated, using the above novel fabrication process flow.
Detection apparatus integrated with micro-devices disclosed in this application is fully capable of detecting pre-chosen properties on a single cell, a single DNA, a single RNA, or an individual, small sized biological matter level. In another further aspect, the invention provides the design, integration, and fabrication process flow of micro-devices capable of making highly sensitive and advanced measurements on very weak signals in biological systems for disease detection under complicated environment with very weak signal and relatively high noise background. Those novel capabilities using the class of micro-devices disclosed in this invention for disease detection include but not limited to making dynamic measurements, real time measurements (such as time of flight measurements, and combination of using probe signal and detecting response signal), phase lock-in technique to reduce background noise, and 4-point probe techniques to measure very weak signals, and unique and novel probes to measure various electronic, electromagnetic and magnetic properties of biological samples at the single cell (e.g., a telomere of DNA or chromosome), single molecule (e.g., DNA, RNA, or protein), single biological subject (e.g., virus) level.
For example, in a time of flight approach to obtain dynamic information on the biological sample (e.g., a cell, a substructure of a cell, a DNA, a RNA, or a virus), a first micro-device is first used to send a signal to perturb the biological subject to be diagnosed, and then a second micro-device is employed to accurately measure the response from the biological subject. In one embodiment, the first micro-device and the second micro-device are positioned with a desired or pre-determined distance L apart, with a biological subject to be measured flowing from the first micro-device towards the second micro-device. When the biological subject passes the first micro-device, the first micro-device sends a signal to the passing biological subject, and then the second micro-device detects the response or retention of the perturbation signal on the biological subject. From the distance between the two micro-devices, time interval, the nature of perturbation by the first micro-device, and measured changes on the biological subject during the time of flight, microscopic and dynamic properties of the biological subject can be obtained. In another embodiment, a first micro-device is used to probe the biological subject by applying a signal (e.g., an electronic charge) and the response from the biological subject is detected by a second micro-device as a function of time.
To further increase detection sensitivity, a novel detection process for disease detection is used, in which time of flight technique is employed.
The utilization of micro-devices (e.g., made by using the fabrication process flows of this invention) as discussed above and illustrated in
In addition to the above examples in measuring electrical properties (e.g., charge, electronic states, electronic charge, electronic cloud distribution, electrical field, current, and electrical potential, and impedance), mechanical properties (e.g., hardness, density, shear strength, and fracture strength) and chemical properties (e.g., pH) in a single cell, and in
One of the key aspects of this invention is the design and fabrication process flows of micro-devices and methods of use the micro-devices for catching and/or measuring biological subjects (e.g., cells, cell substructures, DNA, and RNA) at microscopic levels and in three dimensional space, in which the micro-devices have micro-probes arranged in three dimensional manner with feature sizes as small as a cell, DNA, or RNA, and capable of trapping, sorting, probing, measuring, and modifying biological subjects. Such micro-devices can be fabricated using state-of-the-art microelectronics processing techniques such as those used in fabricating integrated circuits. Using thin film deposition technologies such as molecular epitaxial beam (MEB) and atomic layer deposition (ALD), film thickness as thin as a few monolayers can be achieved (e.g., 4 A to 10 A). Further, using electron beam or x-ray lithography, device feature size on the order of nanometers can be obtained, making micro-device capable of trapping, probing, measuring, and modifying a biological subject (e.g., a single cell, a single DNA or RNA molecule) possible.
Another aspect of this invention relates to micro-indentation probes and micro-probes for measuring a range of physical properties (such as mechanical properties) of biological subjects. Examples of the mechanical properties include hardness, shear strength, elongation strength, fracture stress, and other properties related to cell membrane which is believed to be a critical component in disease diagnosis.
Another novel approach provided by this invention is the use of phase lock-in measurement for disease detection, which reduces background noise and effectively enhances signal to noise ratio. Generally, in this measurement approach, a periodic signal is used to probe the biological sample and response coherent to the frequency of this periodic probe signal is detected and amplified, while other signals not coherent to the frequency of the probe signal is filtered out, which thereby effectively reduces background noise. In one of the embodiments in this invention, a probing micro-device can send a periodic probe signal (e.g., a pulsed laser team, a pulsed thermal wave, or an alternating electrical field) to a biological subject, response to the probe signal by the biological subject can be detected by a detecting micro-device. The phase lock-in technique can be used to filter out unwanted noise and enhance the response signal which is synchronized to the frequency of the probe signal. The following two examples illustrate the novel features of time of flight detection arrangement in combination with phase lock-in detection technique to enhance weak signal and therefore detection sensitivity in disease detection measurements.
To illustrate how a micro-device can be used to simulate an intracellular signal, calcium oscillation is taken as an example mechanism. First, a Ca2+-release-activated channel (CRAC) has to be opened to its maximal extent, which could be achieved by various approaches. In an example of the applicable approaches, a biochemical material (e.g., thapsigargin) stored in the cartridge 924 is released by an injector 925 to the cell, and the CRAC will open at the stimulus of the biological subject. In another example of the applicable approaches, the injector 924 forces a specific voltage on cell membrane, which causes the CRAC to open as well.
The Ca2+ concentration of a solution in the injector 928 can be regulated as it is a desirable combination of a Ca2+-containing solution 926, and a Ca2+ free solution 927. While the injector 930 contains a Ca2+ free solution, then injectors 928 and 930 are alternately switched on and off at a desired frequency. As such, the Ca2+ oscillation is achieved and the content inside the cell membrane are then exposed to a Ca2+ oscillation. Consequently, the cell's activities or fate is being manipulated by the regulated signal generated by the apparatus.
Meanwhile, the cell's response (e.g., in the form of a thermal, optical, acoustical, mechanical, electrical, magnetic, electromagnetic property, or a combination thereof) can be monitored and recorded by the probes integrated in this apparatus.
As surface charge will affect the shape of a biological subject, by using novel and multiple plates, information on the shape and charge distribution of biological subjects can be obtained. The general principle and design of the micro-device can be extended to a broader scope, thereby making it possible to obtain other information on the biological subject via separation by applying other parameters such as ion gradient, thermal gradient, optical beam, or another form of energy.
Alternatively, a probe 1020 can be designed to trigger optical emission such as florescence light emission 1043 in the targeted biological subject such as diseased cells, which can then be detected by an optical probe 1032 as illustrated in
The channel included in the apparatus of this invention can have a width of, e.g., from 1 nm to 1 mm. The apparatus should have at least one inlet channel and at least two outlet channels.
The biological subject 2501 flows in the x direction from the entrance channel 2510 to the accelerating chamber 2530. A bio-compatible fluid 2502 flows from disturbing fluid channel 2520 to the accelerating chamber 2530, it then accelerates the biological subject 2501 in the y-direction. The acceleration correlates with the radius of the biological subject and the larger ones are less accelerated than the smaller ones. Then, the larger and smaller subjects are separated into different selecting channels. Meanwhile, probes can be optionally assembled on the sidewalls of the channels 2510, 2520, 2530, 2540, and 2550. The probes could detect, at the microscopic level, electrical, magnetic, electromagnetic, thermal, optical, acoustical, biological, chemical, biochemical, electro-mechanical, electro-chemical, electro-chemical-mechanical, physical, mechanical properties, or combinations thereof.
Probe 3112 is a fine probing device which is coated by a piezo-electrical material. There is a distance ΔL between probe 3111 and probe 3112.
When the biological subjects are tested when getting through 3111, if the entity is identified to be a suspected abnormal one, the system would trigger the piezo-electrical probe 3112 to stretch into the channel and probe particular properties after a time delay of Δt. And probe 3112 retracts after the suspected entity passed through.
The probing device is capable of measuring at the microscopic level an electrical, magnetic, electromagnetic, thermal, optical, acoustical, biological, chemical, electro-mechanical, electro-chemical, electro-chemical-mechanical, bio-chemical, bio-mechanical, bio-electro-mechanical, bio-electro-chemical, bio-electro-chemical-mechanical, physical or mechanical property, or a combination thereof, of the biological subject.
The width of the micro-channel can range from about 1 nm to about 1 mm.
When a biological subject is tested while getting through 3211, if it is normal, the valve 3221 of the flush channel is open, while the detection channel valve 3222 is closed, the biological subject is flushed out without a time-consuming fine detection.
When the biological subject is tested while getting through 3211, if it is suspected to be abnormal or diseased, the valve 3221 of the flush channel is closed, while the detection channel valve 3222 is open, the biological subject is conducted to the detection channel for a more particular probing.
The width of the micro-channel can range from about 1 nm to about 1 mm.
The probing device is capable of measuring at the microscopic level an electrical, magnetic, electromagnetic, thermal, optical, acoustical, biological, chemical, electro-mechanical, electro-chemical, electro-chemical-mechanical, bio-chemical, bio-mechanical, bio-electro-mechanical, bio-electro-chemical, bio-electro-chemical-mechanical, physical or mechanical property, or a combination thereof, of the biological subject.
The probing device is capable of measuring at the microscopic level an electrical, magnetic, electromagnetic, thermal, optical, acoustical, biological, chemical, electro-mechanical, electro-chemical, electro-chemical-mechanical, bio-chemical, bio-mechanical, bio-electro-mechanical, bio-electro-chemical, bio-electro-chemical-mechanical, physical or mechanical property, or a combination thereof, of the biological subject.
As illustrated in
In
Like
To effectively sorting, separating, screening, probing, or detecting of diseased biological entities, a chamber (or chambers) integrated with various channels can be deployed as shown
To significantly speed up the sorting, screening, probing and detection operations using the disclosed device and process, a high number of desired structures such as those discussed in
One of the key aspects of the present invention relates to a novel technology for detecting disease, in which a number of different classifications of biological information are collected in a device and processed or analyzed. For instance,
Tests were carried out in the laboratory with the apparatus of this invention on certain cancerous tissue samples (with multiple samples for each type of cancer) although the apparatus of this invention can be used for detection of other types of cancer or other types of treatment. In the tests, healthy control samples were obtained from animals with no known cancer disease at the time of collection and no history of malignant disease. Both cancerous samples and healthy control samples were collected and cultured in the same type of culture solution. The cultured samples were then mixed with a dilution buffer and diluted to the same concentration. The diluted samples were maintained at the room temperature for different time intervals and processed within a maximum of 6 hours after being recovered. The diluted samples were tested at the room temperature (20˜23 oC) and in the humidity of 30%˜40%. The samples were tested with an apparatus of this invention under the same conditions and stimulated by the same pulse signal.
The tests show that, in general, the control groups' tested (measured) values (i.e., measured values in relative units for the testing parameter) were lower than the cancerous or diseased groups. Under the same stimulation (in terms of stimulation type and level) with a stimulating or probing signal applied by a probing unit of the tested apparatus of this invention, the difference shown in the measured values between the control groups and the cancerous groups became much more significant, e.g., ranging from 1.5 times to almost 8 times in terms of level of increase in such difference, compared with that without simulation. In other words, the cancerous groups' response to the stimulating signal was much higher than that of the control groups. Thus, the apparatus of this invention has been proven to be able to significantly enhance the relative sensitivity and specificity in the detection and measurement of diseased cells, in comparison to the control or healthy cells.
Further, the test results show that in terms of the novel parameter utilized by the apparatus of this invention, the cancerous group and the control group showed significantly different response. Such difference is significantly greater than the measurement noise. There was a large window to separate the control groups from the cancerous groups, showing a high degree of sensitivity of the novel measurement method and apparatus.
Studies were also undertaken to examine the effect of adding molecular level reaction triggering agent on the efficacy of the apparatus and methods for detecting disease of this invention. The results provided in
The apparatus and methods of this invention has been used in test of more than 20 different types of cancer in all stages of development and showed expectedly high sensitivity and specificity.
Another major novel aspect of this application relates to an effective method to probe and track ability (including immune system) to detect and prevent potential diseases, ability to flight diseases, and the state of a life body, including but not limited to healthy state, non-cancer disease state, pre-cancer state, and cancer state.
Using a novel microfluidic device equipped with sensitive sensors and a fully automated testing machine developed in this work, the method of this invention has been demonstrated on about 100,000 samples which included control (healthy group), disease group, pre-cancer disease group, and cancer group individuals. The test results showed statistically significant blood micro-electrical current level decreasing from healthy group to disease group, and further decreasing to cancer group, signifying potential importance of this new detection technology for early stage cancer detection. In early stage non-small cell lung cancer (NSCLC) tests, sensitivity and specificity reached ˜85% and 93%, respectively. It has also shown that it is capable to detect over 20 types of cancer, including esophageal cancer and brain tumor which do not have other effective screening methods. As the class of electrical properties is a fundamental bio-physical sub-field and impacting many aspects of human blood, it has multi-level effects at cellular, protein, and even molecular levels. Data appear to reveal that this novel technology provides a potentially powerful insight into how cancer evolves and can be highly valuable for pre-cancer and early stage cancer detection. Its mechanism, potential significance, and ramifications will be presented.
Since the liquid media (for example, blood) is interfacing, connecting and communicating with both cells, proteins, and genetic components (DNAs, RNAs, etc.), it plays a critical role in the interfacing, interactions, and communications (for example, cell signaling) between cells, proteins, and genetic components (DNAs, RNAs, etc.) and other biological entities, and the occurrence and progression of diseases including but not limited to non-cancer diseases, pre-cancer diseases and cancer. On the other hand, in the transition from a healthy individual to a disease state, immune system is degraded and disease detection and killing agents such as T cell lost function. In this invention, it is believed that immune system degradation (decrease) and loss in disease detection and disease fighting will and action is caused by changes in properties in the said liquid media surrounding cells, proteins, genetic components (DNAs, RNAs, etc.) and other biological entities. Specifically, those properties can be biological properties (protein concentration, protein types, DNA sequence, DNA static electrical force, DNA surface charge, DNA surrounding media electrical properties, quantum mechanical effects, etc.), bio-chemistry properties, physical properties (thermal, mechanical, electrical, and electro-magnetic properties), bio-physical properties, properties. For example, the shift in the above property (for example, reduction in the above said physical properties) may affect (for example, reduction in effectiveness and efficiency, and transduction degradation) cell signaling and communications by cells and between cells and other biological entities, resulting in the compromise of immune system, loss of detection capability of cells such as T cells to detect cancer cells and ability to kill cancer cells. Therefore, by measuring the above properties including physical and bio-physical properties, one is able to detect the onset of disease and track disease from one stage to the next stage, making early detection and prevention of disease possible.
Mechanism
A micro fluidic device was fabricated by an integrated circuit method in which micro-channels were formed along which sample fluid can be passed, and on whose sides detection transducers (i.e., sensors) were formed to probe the fluid. During dada collection, a voltage meter with automated data recording capabilities was used. When fluid sample arrives at a micro-channel, sensors in the channel can probe the sample via applying a constant voltage while recording micro- electrical current response as a function of time dependent behavior (time sweep) as shown
Cell Line Characteristics
Four cell lines were utilized in the preliminary research. Human non-small cell lung cancer cell line A-549 (Cat. No. TCHu150), human embryonic lung cell line MRC-5 (Cat. No. GNHu41), human hepatoma cell line QGY (Cat. No. TCHu 42) and human hepatocyte cell line HL-7702 (Cat. No. GNHu 6), which were purchased from Cell Bank of Typical Culture Preservation Committee of Chinese Academy of Sciences/Cell Resource Center of Shanghai Academy of Life Sciences, Chinese Academy of Sciences, were cultured in complete growth medium of RPMI-1640 medium which contain 10% FBS (fetal bovine serum) and 1% penicillin-streptomycin in atmosphere of 95% air and 5% carbon dioxide in 37° C. Cell suspension solutions were prepared for testing.
Blood Sample Characteristics
Samples used in a CDA test were whole blood or serum samples, with whole blood typically used.
Whole blood was drawn into an EDTA tube with anticoagulant agent. In addition, cell lines for both control (healthy) and cancer samples were also used in initial development phase of the work to test and validate signals of the technology.
Algorithm
With a large data base from retrospective studies, an algorithm has been built with a CVD test numbers along with cut-off values as a test outcome which is correlated to cancer risk, which (CDA value) is proportional to cancer risk. Based on CDA values, three regions were divided, healthy, medium risk, and high risk.
Results
Both retrospective studies and population screenings were carried out. Follow-up was carried out on a group of individuals with low, medium, and high risk values, among which feedback on 5809 individuals was obtained.
Furthermore, there is noticeable difference between control, disease and liver cancer samples (
Data for a typical control whole blood sample and a liver cancer whole blood sample are shown in
Having initially confirmed feasibility of this new technology for disease detection, multiple retrospective clinical studies have been carried out. Data on over 20 types of cancer have been collected, and an algorithm has been built based upon a large data base. A set of test parameters have been built around the above-mentioned algorithm. The key parameter calculated from this algorithm based on raw data is CDA indicator, whose value is proportional to the cancer risk, and inversely proportional micro- electrical current value of the sample tested.
Table 8 shows significance test of difference—non-parametric test of various types of cancer. In Table 8, the distribution of CDA is the same across the categories of Group. Asymptotic significances are displayed. The significance level is 0.05. Table 8 shows that the difference in CDA values between control group and various cancer types are of statistical significance.
A summary of cancer screening sensitivity and specificity for control group and a number of cancer types from retrospective study is given in Table 9. Table 9 showed that overall, both sensitivity and specificity of CDA technology of various cancer types are relatively high, demonstrating CDA technology is potentially suited for a large number of cancer types. In addition, statistical analysis of the data Table 8 showed that P values for each two groups (each cancer group and control group) are all less than 0.001, also meaning that the difference in CDA values between control group and various cancer types listed in Table 8 are of statistical significance.
Table 10 shows CDA values of non-small lung cancer samples at various stages and control sample, and corresponding sensitivity and specificity, which are higher than traditional methods, particularly at stage I.
Esophageal cancer is a cancer which still does not have a bio-marker and IVD screening method. In this investigation, CDA technology has been evaluated for esophageal cancer screening. Esophageal cancer results are summarized in Table 11. Results showed even at stage I, sensitivity and specificity are above 80%, far better than those by other technologies, which will have significant clinical meaning in catching esophageal cancer early.
CDA technology was utilized to screen ˜100,000 general populations. Based on CDA values, screened individuals were divided into three groups: high risk (beyond the standard, CDA value: ≥50.0), medium risk (need attention, CDA value: 42.0≤CDA<50.0), and low risk (in the normal range, CDA value: <42.0). Follow-up was conducted, and was able to make contact and obtain response from over 5809 individuals through the phone by professionally trained staff. For the period of follow-up (over one year to date), the high risk group was first contacted phone within 15 days after the report; the medium risk group was first contacted within 3 months after the report; and the low risk group was first contacted within 6 months after the report. For patients who are willing to accept follow-up visits, follow-up calls were or will be made every six months after the first phone visit.
Table 12A shows the initial follow-up data statistics. To date, follow-up was carried out on a group of individuals with low, medium, and high risk values, out of which 5809 individuals were able to have made contact and willing to share results from follow-up tests and diagnosis. Based on follow-up on 5809 individuals initially tested with low, medium and high CDA values and later confirmed by oncologists, Table 12B shows cancer cases screened out by CDA technology; Table 12C shows pre-cancer and benign tumor cases screened out by CDA technology; and Table 12D shows other disease cases screed out by CDA technology. As shown in Tables 12A-12D, at the time of the follow-up contact, 129 individuals were diagnosed by oncologists having cancer, 284 individuals were confirmed with pre-cancer and benign tumor, and 297 individuals were confirmed with other diseases. Follow-up is still on-going with remaining individuals.
In addition, in initial retrospective studies, CDA test results on Caucasian group showed comparable sensitivity and specificity as those on Chinese Han ethnic group.
While the functions and properties of bio-physics have played a critical role in physiology, they have not been extensively utilized in the field of IVD of cancer, which has traditionally been more heavily replied upon bio-chemistry, immunology, and genomics. Prior work has not elucidated how electrical properties evolve from a normal state to cancerous state, nor it has been developed into a practical cancer detection tool. This work represents a novel approach and breakthrough in the field cancer detection. Results demonstrated that this technology has unique advantage to detect cancer early, and can be an effective approach to track disease progression, as it showed statistical difference between healthy group and disease group, and between disease group and cancer group. Compared with traditional approaches, the current approach detects a signal which is much more foundational and it is in existence in all human being including healthy individuals. Therefore, its signal is much earlier in nature in detecting occurrence of cancer. Further, micro- electrical current has shown to decrease significantly from healthy group to disease group and from disease to cancer group, making it ideal for early stage cancer detection and tracking diseases leading to cancer.
Results from tests (a) using samples with increasing amount of cancer cells, (b) using samples with increasing amount of bio-marker concentration CEA, and (c) with samples with and without an assay which is known to cause a molecular level reaction showed that CDA values are proportional to increasing amount of cancer cells and bio-marker CEA concentrations. In addition, CDA values are dependent on with and without molecular level reactions. Based on the above observations, it can be stated that CDA values are a function of cellular, protein, and molecular levels (as shown in
Initial data indicates that this novel technology is correlated to protein level (bio-chemistry), cellular level and molecular level expressions, and it has multi-level, and multi-parameter characteristics. Having demonstrated viability of this new technology for pre-cancer and early stage cancer detection, possible mechanism can be further proposed. A scheme of cells, proteins, and genetic components (DNA, RNA, etc.) and their surrounding liquid media (e.g., blood) is described above and provided in
Compared with other traditional cancer detection technologies, CDA technology has many unique features and clear advantages. First, many existing technologies detect cancer signals after cancer has already formed which make those technologies ineffective for early stage cancer detection, while CDA technology detects a bio-physical parameter which exists in healthy individuals and rises as the risk of cancer increases (as shown in
In addition, based on CDA value dependent disease progression behavior (disease progresses with decreasing micro- electrical current of the blood sample); based on the above proposed hypothesis, new model for cancer occurrence is proposed as follows. In this new model, as a major bio-physical parameter, the shift in electrical properties of blood, specifically, decreasing in micro- electrical current and/or changing quantum mechanical effects (which affect gene replications and mutations) is causing negative effects at multi-levels which include (1) reduced surface charge, cell repulsion, and cell signaling efficiency at cellular level, and (2) reduced electrostatic force, DNA surface charge, and possibly increased mutation at DNA level. Further, it is hypothesized that reduced micro-electrical current (and conductance) also causes reduced surveillance capability of T cells for cancer cell detection and reduced immunity which increase occurrence of cancer. The above hypothesis is supported by data collected in this work showing that decreasing (increasing CDA values) in micro- electrical current is correlated with disease progress from healthy group to disease group, from disease group to pre-cancer group, and from pre-cancer group to cancer group.
In this invention, changes in electrical properties in blood and DNA level can be used as a tool for disease detection. As electrical current and conductance decrease, a number of molecular level (DNA surface charge decreases, quantum mechanical effect change, and DNA mutation increases) properties degrade, resulting in increased disease and cancer occurrence. As shown in
Furthermore, the new technology according to this invention can also be used in assisting in diagnosis, such as assisting in diagnosis of lung cancer. As shown in
As also shown in
Initial clinical study results show that the novel technology according to this invention is capable of evaluating effectiveness of drug treatment of cancer. In this case (e.g., as shown in
One of key aspects of this invention is that the bio-physical properties and its associated behaviors disclosed in this novel work are of common to a large number of cancer types, and can be used for detection of a large number of cancer types, making the disclosed method a viable technology for cancer screening, assisting in diagnosis, prognosis, therapy selection and reoccurrence detection.
As shown in
In one embodiment, utilizing a micro-fluidic device with micro-channels and sensitive sensors, electrical properties of blood samples at near field of cells illustrated in above figure (schematic of cellular membranes) can be measured, and related electrical properties including electrical current across the region, trans-membrane potential, and ion levels (potassium ions, sodium ions, chloride ions, calcium ions, and nitride ions) can be directly and indirectly measured. Since disease state of mammals is related to the above-mentioned cellular bio-physical properties (and DNA, RNA and other biological entities in the cells), the above inventive measurement technology can be used to detect diseases including pre-cancer and cancer diseases. The membrane potential can regulate the balance between normal cellular activities including normal growth and replications, and carcinogenesis. As such, both ion level and concentration (potassium ions, sodium ions, chloride ions and calcium ions) and membrane potential could be used as a new, novel bio-marker for cancer prevention and early stage cancer detection.
The present invention provides a new cancer detection technology using a bio-physical approach based on electrical properties of liquid samples for IVD applications. In this new technology, a micro- electrical current is detected which has shown to be very effective in detecting pre-cancer and early stage cancer. This technology has the advantages of screening/detecting or even treating multiple types of diseases (e.g., multiple cancer types) at the same time, detecting cancer early, high sensitivity and specificity, covering a wide range of cancer types, and relatively simple and cost effective. Based on how CDA values are correlated to control, disease and cancer groups in this work, and possible effects of electrical properties in blood on disease progression, a new hypothesis on cancer occurrence model is proposed in which a reduction in blood micro electrical current (and conductance) and/or a change of quantum mechanical effects is proposed to cause a number of negative effects at cellular and molecular levels, resulting in reduced cell to cell signaling, cell to cell repulsion, and immunity, and increased gene mutation frequency, and hence increased occurrence of cancer.
While for the purposes of demonstration and illustration, the above cited novel, detailed examples show how microelectronics and/or nano-fabrication techniques and associated process flows can be utilized to fabricate highly sensitive, multi-functional, powerful, and miniaturized detection devices, the principle and general approaches of employing microelectronics and nano-fabrication technologies in the design and fabrication of high performance detection devices have been contemplated and taught, which can and should be expanded to various combination of fabrication processes including but not limited to thin film deposition, patterning (lithography and etch), planarization (including chemical mechanical polishing), ion implantation, diffusion, cleaning, various materials, combination of processes and steps, and various process sequences and flows. For example, in alternative detection device design and fabrication process flows, the number of materials involved can be fewer than or exceed four materials (which have been utilized in the above example), and the number of process steps can be fewer or more than those demonstrated process sequences, depending on specific needs and performance targets. For example, in some disease detection applications, a fifth material such as a biomaterial-based thin film can be used to coat a metal detection tip to enhance contact between the detection tip and a biological subject being measured, thereby improving measurement sensitivity.
Applications for the detection apparatus and methods of this invention include detection of diseases (e.g., in their early stage), particularly for serious diseases like cancer. Since cancer cell and normal cell differ in a number of ways including differences in possible microscopic properties such as electrical potential, surface charge, density, adhesion, and pH, novel micro-devices disclosed herein are capable of detecting these differences and therefore applicable for enhanced capability to detect diseases (e.g., for cancer), particularly in their early stage. In addition to micro-devices for measuring electrical potential and electrical charge parameters, micro-devices capable of carrying out mechanical property measurements (e.g., density) can also be fabricated and used as disclosed herein. In mechanical property measurement for early stage disease detection, the focus will be on the mechanical properties that likely differentiate disease or cancerous cells from normal cell. As an example, one can differentiate cancerous cells from normal cells by using a detection apparatus of this invention that is integrated with micro-devices capable of carrying out micro-indentation measurements.
Another aspect of this invention provides treatment of diseases.
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Finally, as shown in
Although specific embodiments of this invention have been illustrated herein, it will be appreciated by those skilled in the art that any modifications and variations can be made without departing from the spirit of the invention. The examples and illustrations above are not intended to limit the scope of this invention. Any combination of detection apparatus, micro-devices, fabrication processes, and applications of this invention, along with any obvious their extension or analogs, are within the scope of this invention. Further, it is intended that this invention encompass any arrangement, which is calculated to achieve that same purpose, and all such variations and modifications as fall within the scope of the appended claims.
All publications or patent applications referred to above are incorporated herein by reference in their entireties. All the features disclosed in this specification (including any accompanying claims, abstract and drawings) may be replaced by alternative features serving the same, equivalent or similar purpose, unless expressly stated otherwise. Thus, unless expressly stated otherwise, each feature disclosed is one example of a generic series of equivalent or similar features.
This application claims priority to U.S. Application No. 62/741,843, filed Oct. 5, 2018, U.S. Application No. 62/776,605, filed Dec. 7, 2018, U.S. Application No. 62/818,909, filed Mar. 15, 2019, U.S. Application No. 62/830,354, filed Apr. 5, 2019, PCT Application No. PCT/US2019/028785, filed Apr. 23, 2019, and U.S. Application No. 62/865,816, filed Jun. 24, 2019, the contents of all of which are incorporated herein by reference in their entirety.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/US2019/055060 | 10/7/2019 | WO | 00 |
| Number | Date | Country | |
|---|---|---|---|
| 62865816 | Jun 2019 | US | |
| 62830354 | Apr 2019 | US | |
| 62818909 | Mar 2019 | US | |
| 62776605 | Dec 2018 | US | |
| 62741843 | Oct 2018 | US |
| Number | Date | Country | |
|---|---|---|---|
| Parent | PCT/US2019/028785 | Apr 2019 | US |
| Child | 17282770 | US |
