Patent Application
20230101022
The present invention relates to a new use of ginsenoside M1 as a modulator of angiotensin regulating enzymes and relevant treatment methods using the same. In particular, the present invention provides a method for treating a disease or conditions associated with imbalance in angiotensin regulating enzymes, including those caused by viral infection and resulting damages in organs or tissues, using ginsenoside M1.
Renin-Angiotensin System (RAS) or Renin-Angiotensin-Aldosterone System (RAAS) is a hormonal system. When a large amount of blood loss or blood pressure drops, this system will be activated, which is a key hormone can regulates blood pressure and body fluid constant in the body. Both angiotensin-converting enzyme (ACE) and angiotensin-converting enzyme 2 (ACE2) play important roles in RAS. ACE2 will generate angiotensin I (Ang I) to angiotensin (Ang)-(1-9) or Ang II to Ang-(1-7) in order to achieve fluid balance. RAS plays an important role in many cardiovascular diseases. The present studies shown that ACE2 deficiency can indues acute or chronic lung injury or even pulmonary fibrosis. On the other hand, excessive activation of Ang II can lead to blood vessels and kidney damage, and progression to necrosis and fibrosis. Therefore, the lack of ACE2 can lead to excessive Ang II accumulation can cause to the lung and kidney damage. However, the latest research shows that the S protein of the new coronavirus will bind to ACE2 and invade the lung. Excessive accumulation of Ang II in the body, resulting in pulmonary fibrosis and kidney damage phenomenon. [1]
Coronaviruses pose serious health threats to humans and other animals. From 2002 to 2003, severe acute respiratory syndrome coronavirus (SARS-CoV) infected 8,000 people, with a fatality rate of ˜10%. Since 2012, Middle East respiratory syndrome coronavirus (MERS-CoV) has infected more than 1,700 people, with a fatality rate of ˜36%. Since 2013, porcine epidemic diarrhea coronavirus (PEDV) has swept throughout the United States, causing an almost 100% fatality rate in piglets and wiping out more than 10% of America's pig population in less than a year. Further, novel coronavirus (also called SARS-CoV-2) causes the COVID-19 pandemic. As of Feb. 6, 2021, more than 105 million cases have been confirmed with more than 2.29 million deaths attributed to COVID-19 according to the Wikipedia website.
In general, coronaviruses cause widespread respiratory, gastrointestinal, and central nervous system diseases in humans and other animals, threatening human health and causing economic loss. Coronaviruses are capable of adapting to new environments through mutation and recombination with relative ease and hence are programmed to alter host range and tissue tropism efficiently. Therefore, health threats from coronaviruses are constant and long-term. Understanding the virology of coronaviruses and controlling their spread have important implications for global health and economic stability.
Coronaviruses belong to the family Coronaviridae in the order Nidovirales. They can be classified into four genera: Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus. Among of them, alpha-and betacoronaviruses infect mammals, gammacoronaviruses infect avian species, and deltacoronaviruses infect both mammalian and avian species. Representative alphacoronaviruses include human coronavirus NL63 (HCoV-NL63), porcine transmissible gastroenteritis coronavirus (TGEV), PEDV, and porcine respiratory coronavirus (PRCV). Representative betacoronaviruses include SARS-CoV, MERS-CoV, bat coronavirus HKU4, mouse hepatitis coronavirus (MHV), bovine coronavirus (BCoV), and human coronavirus OC43. Representative gamma-and deltacoronaviruses include avian infectious bronchitis coronavirus (IBV) and porcine deltacoronavirus (PdCV), respectively. Coronaviruses are large, enveloped, positive-stranded RNA viruses. They have the largest genome among all RNA viruses, typically ranging from 27 to 32 kb. The genome is packed inside a helical capsid formed by the nucleocapsid protein (N protein) and further surrounded by an envelope. Associated with the viral envelope are at least three structural proteins: The membrane protein (M protein) and the envelope protein (E protein) are involved in virus assembly, whereas the S protein mediates virus entry into host cells. Some coronaviruses also encode an envelope-associated hemagglutinin-esterase protein (HE protein). Among these structural proteins, the spike forms large protrusions from the virus surface, giving coronaviruses the appearance of having crowns (hence their name; corona in Latin means crown). In addition to mediating virus entry, the spike is a critical determinant of viral host range and tissue tropism and a major inducer of host immune responses. [2]
Ginsenosides, the main active ingredients of ginseng, are known to have a variety of pharmacological activities, e.g. antitumor, antifatique, antiallergic and antioxidant activities. Ginsenosides share a basic structure, composed of gonane steroid nucleus having 17 carbon atoms arranged in four rings. Ginsenosides are metalized in the body, and a number of recent studies suggest that ginsenoside metabolites, rather than naturally occurring ginsenosides, are readily absorbed in the body and act as the active components. Among them, ginsenoside M1, also named Compound K (CK), is known as one metabolite of protopanaxadiol-type ginsenosides via the gypenoside pathway by human gut bacteria. Until now, no prior art references report the effect of ginsenoside M1 in regulating angiotensin regulating enzymes and its use for treatment for relevant diseases or conditions.
In the present invention, it is unexpected found that ginsenoside M1 is effective in regulating angiotensin regulating enzymes, including upregulating ACE2 and downregulating ACE, promoting degrading angiotensin II and preventing accumulation of angiotensin II, and thus useful for treatment of a disease or condition associated with abnormal balance of ACE and ACE2 or accumulation of angiotensin II, particularly damages in organs or tissues caused by infection of virus e.g. coronavirus.
Therefore, the present invention provides a use of ginsenoside M1 for manufacturing a medicament as a modulator of an angiotensin regulating enzyme in a subject in need thereof.
In some embodiments of the invention, the angiotensin regulating enzyme is selected from the group consisting of angiotensin-converting enzyme (ACE) and angiotensin-converting enzyme 2 (ACE2).
In some embodiments, the modulator is an ACE inhibitor or an ACE2 activator or both.
In some embodiments, the modulator is effective in degrading angiotensin II and preventing accumulation of angiotensin II in the subject.
In some embodiments, the medicament is effective in treating a disease or condition associated with imbalance in ACE and ACE2 or accumulation of angiotensin II in the subject.
The present invention also provides a method for treating a disease or condition associated with abnormal balance of ACE and ACE2 or accumulation of angiotensin II in a subject in need, comprising administering to the subject an amount of ginsenoside M1 effective to treat the subject.
In some embodiments, ginsenoside M1 is administered in an amount effective in inhibiting ACE, or activating ACE2 or both in the subject.
In some embodiments, ginsenoside M1 is administered in an amount effective in degrading angiotensin II and preventing accumulation of angiotensin II in the subject.
In some embodiments, the disease or condition to be treated are associated with abnormal balance of ACE and ACE2 or accumulation of angiotensin II, including damages in organs or tissues in the subject.
In some embodiments, the damages includes damages in lung, gastrointestinal tract, spleen, lymph nodes, heart, kidney, bladder, liver, gallbladder, adrenal glands and/or testis.
In some embodiments, the damages are caused by infection of coronavirus.
In some embodiments, the coronavirus is selected from severe acute respiratory syndrome coronavirus (SARS-CoV), middle east respiratory syndrome coronavirus (MERS-CoV), and novel coronavirus (2019-nCoV).
In some embodiments, ginsenoside Ml is administered in combination with one or more additional therapeutic methods or agents useful for degrading angiotensin II and preventing accumulation of angiotensin II. In one example, such one or more additional therapeutic agents include ACE2.
In some embodiments, ginsenoside M1 is administered in combination with one or more additional therapeutic agents selected from the group consisting of corticosteroids, non-steriodal anti-inflammatory drugs (NSAIDs), cytotoxic drugs, immunosuppressants, and vasodilators.
In some embodiments, ginsenoside M1 is administered in combination with one or more additional therapeutic agents selected from the group consisting of an antibiotic, interferon and an anti-viral agent.
Specifically, the present invention provides use of ginsenoside M1 for manufacturing a medicament for treating pulmonary injuries. The present invention also provides a method for treating pulmonary injuries by administering an effective amount of ginsenoside M1 to a subject in need thereof. In particular, the pulmonary injuries are associated with abnormal balance of ACE and ACE2 or accumulation of angiotensin II in the subject.
In some embodiments, the pulmonary injuries include alveolitis, lymphocyte infiltration and/or fibrosis in the lung.
In some embodiments, ginsenoside M1 is administered by parenteral or enteral route.
The details of one or more embodiments of the invention are set forth in the description below. Other features or advantages of the present invention will be apparent from the following detailed description of several embodiments, and also from the appending claims.
For the purpose of illustrating the invention, there are shown in the drawings embodiments. It should be understood, however, that the invention is not limited to the preferred embodiments shown. In the drawings:
Unless defined otherwise, all technical and scientific terms used herein have the meaning commonly understood by a person skilled in the art to which this invention belongs. As used herein, the following terms have the meanings ascribed to them unless specified otherwise.
The articles “a” and “an” are used herein to refer to one or more than one (i.e., at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element.
The term “comprise” or “comprising” is generally used in the sense of include/including which means permitting the presence of one or more features, ingredients or components. The term “comprise” or “comprising” encompasses the term “consists” or “consisting of.”
Ginsenoside M1, also named Compound K (CK), 20-O-β-D-glucopyranosyl-20(S)-protopanaxadiol, is one of saponin metabolites known in the art. The chemical structure of ginsenoside M1 is as follows:
Ginsenoside M1 is known as one metabolite of protopanaxadiol-type ginsenosides via the gypenoside pathway by human gut bacteria. Ginsenoside M1 can be found in blood or urine after intake. Ginsenoside M1 may be prepared from ginseng plants through fungi fermentation by methods known in the art, such as Taiwan Patent Application No. 094116005 (1280982) and U.S. Pat. No. 7,932,057, the entire content of which is incorporated herein by reference. In certain embodiments, the ginseng plants for preparing the ginsenoside M1 include Araliaceae family, Panax genus, e.g. P. ginseng and P. pseudo-ginseng (also named Sanqi). In general, the method of preparation of ginsenoside M1 includes the steps of (a) providing powder of ginseng plant materials (e.g. leaves or stems); (b) providing a fungus for fermenting the ginseng plant materials, wherein the fermentation temperature is ranged from 20-50° C., the fermentation humidity is ranged from 70-100%, the pH value is ranged from 4.0-6.0, and the fermentation period is ranged from 5-15 days; (c) extracting and collecting the fermentation products; and (d) isolating 20-O-β-D-glucopyranosyl-20(S)-protopanaxadiol from the fermentation products.
When ginsenoside M1 is described as “isolated” or “purified” in the present invention, it should be understood as not absolutely isolated or purified, but relatively isolated or purified. For example, purified ginsenoside M1 refers to one that is more purified compared to its naturally existing form. In one embodiment, a preparation comprising purified ginsenoside M1 may comprise ginsenoside M1 in an amount of more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or 100% (w/w) of the total preparation. It should be understood that when a certain number was used herein to show a ratio or dosage, said number generally includes that within the range of 10% more and less, or more specifically, the scope of 5% more and less than the number.
The term “individual” or “subject” used herein includes human and non-human animals such as companion animals (such as dogs, cats and the like), farm animals (such as cows, sheep, pigs, horses and the like), or laboratory animals (such as rats, mice, guinea pigs and the like).
The term “treating” as used herein refers to the application or administration of a composition including one or more active agents to a subject afflicted with a disorder, a symptom or conditions of the disorder, a progression of the disorder or at risk of developing the disorder, with the purpose to cure, heal, alleviate, relieve, alter, remedy, ameliorate, improve, or affect the disorder, the symptoms or conditions of the disorder, the disabilities induced by the disorder, or the onset or progression of the disorder.
The term “effective amount” used herein refers to the amount of an active ingredient to confer a desired therapeutic effect in a treated subject. In some embodiments, an effective amount as used herein can be an amount effective in downregulating ACE, upregulating ACE2, degrading angiotensin II, and/or preventing accumulation of angiotensin II. In some embodiments, an effective amount as used herein can be an amount effective in alleviating or reducing damages in organs or tissues caused by accumulation of angiotensin II, for example, pulmonary injuries e.g. alveolitis, lymphocyte infiltration and/or fibrosis in the lung.
The therapeutically effective amount may change depending on various reasons, such as administration route and frequency, body weight and species of the individual receiving said pharmaceutical, and purpose of administration. Persons skilled in the art may determine the dosage in each case based on the disclosure herein, established methods, and their own experience. For example, in certain embodiments, the oral dosage of ginsenoside M1 used in the present invention is 10 to 1,000 mg/kg daily. In some examples, the oral dosage of ginsenoside M1 used in the present invention is 100 to 300 mg/kg daily, 50 to 150 mg/kg daily, 25 to 100 mg/kg daily, 10 to 50 mg/kg daily, or 5 to 30 mg/kg daily. In addition, in some embodiments of the invention, ginsenoside M1 is administered periodically for a certain period of time, for example, daily administration for at least 15 days, one month or two months or longer.
In one embodiment, a therapeutically effective amount of the active ingredient may be formulated with a pharmaceutically acceptable carrier into a pharmaceutical composition of an appropriate form for the purpose of delivery and absorption. Depending on the mode of administration, the pharmaceutical composition of the present invention preferably comprises about 0.1% by weight to about 100% by weight of the active ingredient, wherein the percentage by weight is calculated based on the weight of the whole composition.
As used herein, “pharmaceutically acceptable” means that the carrier is compatible with the active ingredient in the composition, and preferably can stabilize said active ingredient and is safe to the individual receiving the treatment. Said carrier may be a diluent, vehicle, excipient, or matrix to the active ingredient. Some examples of appropriate excipients include lactose, dextrose, sucrose, sorbose, mannose, starch, Arabic gum, calcium phosphate, alginates, tragacanth gum, gelatin, calcium silicate, microcrystalline cellulose, polyvinyl pyrrolidone, cellulose, sterilized water, syrup, and methylcellulose. The composition may additionally comprise lubricants, such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifying and suspending agents; preservatives, such as methyl and propyl hydroxybenzoates; sweeteners; and flavoring agents. The composition of the present invention can provide the effect of rapid, continued, or delayed release of the active ingredient after administration to the patient.
According to the present invention, the form of said composition may be tablets, pills, powder, lozenges, packets, troches, elixers, suspensions, lotions, solutions, syrups, soft and hard gelatin capsules, suppositories, sterilized injection fluid, and packaged powder.
The composition of the present invention may be delivered via any physiologically acceptable route, such as oral, parenteral (such as intramuscular, intravenous, subcutaneous, and intraperitoneal), transdermal, suppository, and intranasal methods. Regarding parenteral administration, it is preferably used in the form of a sterile water solution, which may comprise other substances, such as salts or glucose sufficient to make the solution isotonic to blood. The water solution may be appropriately buffered (preferably with a pH value of 3 to 9) as needed. Preparation of an appropriate parenteral composition under sterile conditions may be accomplished with standard pharmacological techniques well known to persons skilled in the art, and no extra creative labor is required.
According to the present invention, ginsenoside M1 may be used as an active ingredient for treating a disease or condition associated with abnormal balance of ACE and ACE2 or accumulation of angiotensin II in the subject. In some embodiments, such disease or condition includes damages in organs or tissues in the subject such as those in lung, gastrointestinal tract, spleen, lymph nodes, heart, kidney, bladder, liver, gallbladder, adrenal glands and/or testis. In certain examples, the damages are caused by infection of coronavirus, including but not limited to SARS-CoV, MERS-CoV and 2019-nCoV.
In some embodiments, ginsenoside M1 or compositions comprising ginsenoside M1 as the active ingredient may be used in combination with existing therapeutic methods or medicaments, for example, those for treating a disease or condition associated with abnormal balance of ACE and ACE2 or accumulation of angiotensin II as described herein. In one example, ginsenoside M1 is administrated with ACE2. Accordingly, a particular combination of ginsenoside M1 with ACE2 is administrated to a subject in need as described herein.
In some embodiments, ginsenoside M1 or compositions comprising ginsenoside M1 as the active ingredient may be used in combination with corticosteroids (such as prednisolone), non-steriodal anti-inflammatory drugs (NSAIDs), cytotoxic drugs (such as cyclophosphamide, chlorambucil, and azathioprine), immunosuppressants (such as cyclosporine and Mycophenolate Mofetil), and vasodilators (such as angiotensin-converting-enzyme inhibitors (ACE inhibitors)).
In one particular embodiment, ginsenoside M1 is administrated in combination with ACE2 for use treatment of pulmonary injuries, for example, alveolitis, lymphocyte infiltration and/or fibrosis in the lung.
In some embodiments, ginsenoside M1 is administered in combination with an antibiotic, interferon and an anti-viral agent.
In one embodiment, the medicament or therapeutic method used in combination may be used simultaneously (parallel) or sequentially. When medicaments are used in combination, the medicaments may be mixed in the same formula or put in different formulas separately, such as separate capsules, pills, tablets, and injections.
The present invention is further illustrated by the following examples, which are provided for the purpose of demonstration rather than limitation. Those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments which are disclosed and still obtain a like or similar result without departing from the spirit and scope of the invention.
Ginsenoside M1, also named Compound K (CK), 20-O-β-D-glucopyranosyl-20(S)-protopanaxadiol (named LCHK168 below), was prepared by methods known in the art, such as those described in Taiwan Patent Application No. 094116005 (I280982) and U.S. Pat. No. 7,932,057.
According to previous research, the spike protein (S protein) on coronavirus will bind to ACE2, and the amount of ACE2 will affect the health of the lungs and kidneys (ACE2 Expression in Kidney and Testis May Cause Kidney and Testis Damage After 2019-nCoV Infection https://doi.org/10.1101/2020.02. 12.20022418, Posted Feb. 13, 2020; and Angiotensin-Converting Enzyme 2 Attenuates Bleomycin-Induced Lung Fibrosis in Mice, Cellular Physiology and Biochemistry, 2015; 36: 697-711). This study uses ginsenoside M1 to regulate the content of ACE2 to maintain a healthy body.
Angiotensin II (Ang II)-mediated renal injury represents a major pathogenetic mechanism in most chronic kidney diseases. We found that ginsenoside M1 attenuated Ang II expression by upregulating angiotensin-converting enzyme 2 (ACE2) in the renal tissue. We confirmed this finding in an exogenous Ang II-infused mice model of renal injury, and the results showed consistent in two models. Local renin-angiotensin system (RAS) activation has been shown to play an important role in the pathogenesis of idiopathic pulmonary fibrosis (IPF). It has been reported that ACE2 could inhibit RAS-mediated epithelial injury and fibrogenesis and ACE2 deficiency could aggravate acute and chronic lung injury.
1. Material and Methods
1.1 Animals and Treatments.
1.1.1 Rat (Renal Damages)
A total of 20 Wistar-Kyoto rats (WKY) and 20 SHR (male, 16˜17 weeks old) animals were grouped into four: WKY group (8 WKY, orally administered 0.5% CMC-Na); SHR group (8 SHR, orally administered 0.5% CMC-Na); WKY+Ginsenoside M1 group (8 WKY, orally administered 20 mg/kgGinsenoside M1); SHR +Ginsenoside M1 group (8 SHR, orally administered 20 mg/kg Ginsenoside M1). Ginsenoside M1 or placebo administration was carried out once daily for 42 days is was followed by animal sacrifice and blood and renal tissue sample collection. The renal tissue specimens underwent fixation with 4% formalin (histopathology) or were snap-frozen with liquid nitrogen and kept at −80° C. for reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA).
1.1.2 Mice (Infusion Model) (Renal Damages)
A total of twenty C57BL/6 mice (male, 10 weeks old) were maintained with rodent chow and water at will. Subcutaneous implantation of a 1002 osmotic minipump was performed at the dorsum of the neck for Ang II (1.5 mg/kg) or normal saline infusion animals were assigned to four groups: Saline group (5 mice, infused with normal saline and orally administered 0.5% CMC-Na); Ang II group (5 mice, infused with Ang II and orally administered 0.5% CMC-Na); Saline+Ginsenoside M1 group (5 mice, infused with normal saline and orally administered 20 mg·kg−1·d−1 Ginsenoside M1); Ang II +Ginsenoside M1 group (5 mice, infused with Ang II and orally administered 20 mg/kg Ginsenoside M1). Ginsenoside M1 or placebo administration was performed daily for 14 days, followed by animal sacrifice and blood and renal tissue sample collection. The renal tissue specimens underwent fixation with 4% formalin (histopathology) or were snap-frozen with liquid nitrogen and kept at −80° C. (RT-qPCR and ELISA).
1.1.3 Mice (Pulmonary Damages)
The animals were randomly divided into 5 groups as: (1) Saline group: 0.9% saline (200 μL) injection via the tail vein; (2) ACE2 group: a single intraperitoneal injection of ACE2 (0.2 mg/kg); (3) BLM group: a single injection of bleomycin at 150 mg/kg via the tail vein; and (4) BLM+ACE2 group: an injection of bleomycin at 150 mg/kg via the tail vein, followed by an intraperitoneal injection of ACE2 at 0.2 mg/kg. (5) BLM+ACE2+M1 group: an injection of bleomycin at 150 mg/kg via the tail vein, followed by an intraperitoneal injection of 0.2 mg/kg ACE2 and 60mg/kg M1. After administration, ten animals randomly selected from each group were euthanized on days 7, 14 and 28 by cervical dislocation.
1.2 Histopathological Assessment.
Renal tissue specimens underwent fixation with 4% formalin, paraffin embedding, sectioning at 4 μm, and staining with hematoxylin and eosin (H&E) and Masson trichrome stain, respectively. Then, a Nikon E100 light microscope was employed for analysis.
1.3 Immunohistochemistry (IHC)
ACE and ACE2 primary antibodies were produced by Bioss Antibodies. Peroxidase-linked goat anti-rabbit secondary antibodies and the DAB and two-step rabbit IHC kits were provided by ZSGB-BIO. IHC was carried out as proposed by ZSGB-BIO.
1.4 Ang II and Ang-(1-7) Level Assessment in the Renal Tissue
The supernatants of renal specimens were prepared as follows. A total of 100 mg of the renal tissue was homogenized in 900 μL of ice-cold normal saline and submitted to centrifugation (1000 g, 4° C. for 15 min). The resulting supernatants were kept at −80° C. until analysis with Ang II and Ang-(1-7) ELISA kits, respectively, in accordance with the manufacturer's protocols.
2. Results
2.1 Ginsenoside M1 Reduces Ang II Levels in the Renal Tissue in SHR
There are two main ACE which regulate the levels of Ang II in vivo, including ACE and ACE2. ACE transforms angiotensin I into Ang II, and ACE2 transforms Ang II into Ang-(1-7). So upregulation of ACE induces Ang II increase, while ACE2 upregulation attenuates Ang II increase. Spontaneously hypertensive rat (SHR) is an animal model of genetic hypertension. According to ELISA data (
2.2 Ginsenoside M1 Reduces Ang II Levels in the Renal Tissue in Mice Infused Ang II
According to ELISA data (
2.3 Treatment of Alveolitis and Fibrosis by M1 and ACE2
As shown in
1. Journal of Molecular Medicine/October 2006, Volume 84, Issue 10, pp 814-820| Cite as/Lessons from SARS: control of acute lung failure by the SARS receptor ACE2. Pharmacology & Therapeutics/Volume 128, Issue 1, October 2010, Pages 119-128/Trilogy of ACE2: A peptidase in the renin—angiotensin system, a SARS receptor, and a partner for amino acid transporters.
2. Published in final edited form as: Annu Rev Virol. 2016 Sep. 29; 3(1): 237-261. doi:10.1146/annurev-virology-110615-042301./Structure, Function, and Evolution of Coronavirus Spike Proteins/Fang Li Department of Pharmacology, University of Minnesota Medical School, Minneapolis, Minn. 55455
3. Antioxidant Treatment and Alcoholism/Camila S. Silva PhD, . . . Helio Vannucchi MD, PhD, in Molecular Aspects of Alcohol and Nutrition, 2016
4. Beta-Glucosidase From Penicillium/Gustavo Molina, . . . Glàucia M. Pastore, in New and Future Developments in Microbial Biotechnology and Bioengineering, 2018
5. Ginseng and Gastrointestinal Protection*/Min-Hyun Kim, Hyeyoung Kim, in Gastrointestinal Tissue, 2017
This application claims the benefit of U.S. provisional application No. 62/978,995, filed Feb. 20, 2020, the entire content of which is incorporated herein by reference.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/CN2021/076808 | 2/19/2021 | WO |
| Number | Date | Country | |
|---|---|---|---|
| 62978995 | Feb 2020 | US |
