Compositions and methods for treating viral infection

Abstract

To provide nutritional supplements effective for viral infection treatment.SOLUTION: Provided herein are a composition and a method comprising a nutritional supplement for use in the treatment of viral infections (e.g., corona virus, HCV, HIV) by correcting dysregulation of apoptosis, reducing pro-inflammatory cytokine production, and increasing production of host antiviral compounds. The nutritional supplement can be used in combination with known antiviral therapies, as well as with low level radiotherapy and / or chemotherapeutic drugs for use in the treatment of neoplastic disease.SELECTED DRAWING: Figure 2

Description

[Technical Field]

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 063,542, filed August 10, 2020. These and all other referenced external materials are incorporated herein by reference in their entirety. In the event that a definition or use of a term in a document incorporated by reference contradicts or is contrary to the definition of the term provided herein, the definition of the term provided herein shall control.

[0002] (Technical field) The field of this invention is compositions and methods for treating viral infections, particularly coronavirus infections. [Background technology]

[0003] The background discussion includes information that may be useful in understanding the present invention. No admission is made that any of the information presented herein is prior art or relevant to the presently claimed invention, or that any publication specifically or implicitly referenced is prior art.

[0004] Coronaviruses are positive-sense, single-stranded, enveloped RNA viruses that have repeatedly crossed species barriers and caused disease in humans and animals. Over the past two decades, three novel human pathogenic coronaviruses have emerged, including severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003, Middle East respiratory syndrome coronavirus (MERS-CoV) since 2012, and most recently SARS-CoV-2 since December 2019, causing epidemics of severe respiratory infections in humans. SARS-CoV-2 infection can induce a significant inflammatory response, particularly in the respiratory system, which can be fatal.

[0005] Many existing drugs, including remdesivir, chloroquine, hydroxychloroquine, nafamostat, camostat, and ivermectin, have been reported to exhibit anti-SARS-CoV-2 activity in vitro. Remdesivir is a nucleotide analog with broad-spectrum antiviral activity. Chloroquine and hydroxychloroquine are mild immunosuppressants used to treat autoimmune diseases and have been proposed for the treatment of COVID-19, but recent studies have shown no clear clinical benefit and may increase morbidity. Nafamostat and camostat are serine protease inhibitors used to treat chronic pancreatitis and gastroesophageal reflux disease that may interfere with SARS-CoV-2 cell entry, but data from well-designed randomized controlled trials of these drugs are not yet available.

[0006] Therefore, there is an urgent need to identify safe and effective approaches to the treatment of COVID-19. Summary of the Invention [Problem to be solved by the invention]

[0007] The present subject matter provides compositions and methods that provide nutritional supplements effective in the treatment of viral infections (e.g., coronavirus, HCV, HIV) by, for example, correcting dysregulation of apoptosis, reducing the production of proinflammatory cytokines, and increasing the production of natural antiviral compounds (e.g., interferons). Such nutritional supplements can be used in combination with known antiviral therapies (therapeutic antibodies, small molecule antivirals), as well as low-level radiation therapy and / or chemotherapeutic agents traditionally used in the treatment of neoplastic diseases (e.g., cancer). [Means for solving the problem]

[0008] One embodiment of the inventive concept is a method of treating an individual for a viral infection by providing a dietary supplement comprising fish oil and selenium (e.g., in the form of selenium yeast) and administering the dietary supplement to the individual in an amount effective to treat the viral infection or the sequelae of the viral infection. The viral infection may be caused by infection with a virus that induces dysregulation of apoptosis in infected cells, and the dietary supplement provides fish oil and selenium in amounts sufficient to alter apoptotic events in the infected cells upon administration. In some embodiments, the amounts of fish oil and selenium are also selected to reduce serum concentrations of proinflammatory cytokines. In some embodiments, selenium is provided at 1,000 μg to 10,000 μg per day and / or fish oil is provided at 1,000 mg to 20 mg per day. The dietary supplement may also include coenzyme Q10 (e.g., 1 mg to 5,000 mg of coenzyme Q10). In some embodiments, the dietary supplement may include three or more ingredients listed in Table 1. Contemplated viral infections include those resulting from infection with coronaviruses (e.g., SARS-CoV-2), HCV, and / or HIV. Such treatment may be applied to active infections, including asymptomatic infections. In other embodiments, such treatment may be prophylactic. In some embodiments, the dietary supplement may be provided in combination with low-dose radiation therapy (e.g., doses up to 2 rem provided as single or multiple doses) and / or cancer chemotherapeutic agents (e.g., taxol, adriamycin, and avastin). Such cancer chemotherapeutic agents may be provided at lower doses (e.g., 1% to 50% or less) than those applied in cancer treatment.

[0009] Another embodiment of the inventive concept is the use of a dietary supplement comprising fish oil and selenium (e.g., in the form of selenium yeast), and administering the dietary supplement to an individual in an amount effective to treat the viral infection or the sequelae of the viral infection. The viral infection may be caused by infection with a virus that induces dysregulation of apoptosis in infected cells, and the dietary supplement provides fish oil and selenium in amounts sufficient to modify apoptotic events in the infected cells upon administration. In some embodiments, the amounts of fish oil and selenium are also selected to reduce serum concentrations of proinflammatory cytokines. In some embodiments, selenium is provided at 1,000 μg to 10,000 μg per day and / or fish oil is provided at 1,000 mg to 20 mg per day. The dietary supplement may also include coenzyme Q10 (e.g., 1 mg to 5,000 mg of coenzyme Q10). In some embodiments, the dietary supplement may include three or more ingredients listed in Table 1. Contemplated viral infections include those resulting from infection with coronaviruses (e.g., SARS-CoV-2), HCV, and / or HIV. Such treatment can be applied to active infections, including asymptomatic infections. In other embodiments, such treatment can be preventative. In some embodiments, the dietary supplement can be provided in combination with low-dose radiation therapy (e.g., doses up to 2 rem provided as single or multiple doses) and / or cancer chemotherapeutic agents (e.g., taxol, adriamycin, and avastin). Such cancer chemotherapeutic agents can be provided at lower doses (e.g., 1% to 50% or less) than those applied in cancer treatment.

[0010] That is, the present invention provides the following.

[0011] 1. A method of treating an individual for a viral infection, comprising: Obtaining a dietary supplement containing fish oil and selenium; and administering to the individual the dietary supplement in an amount effective to treat the viral infection or the sequelae of the viral infection; The method, wherein the viral infection is caused by infection with a virus that induces dysregulation of apoptosis in cells infected with the virus, and the dietary supplement provides fish oil and selenium in amounts sufficient to alter apoptotic events in cells infected with the virus upon administration. Preferably, the amounts of fish oil and selenium are selected to further reduce serum concentrations of pro-inflammatory cytokines in the individual. Preferably, the selenium is provided in the form of selenium yeast. Preferably, the selenium is provided in an amount of 1,000 μg to 10,000 μg per day. Preferably, the dietary supplement further comprises coenzyme Q10. Preferably, the dietary supplement contains 1 mg to 5,000 mg of coenzyme Q10. Preferably, the fish oil is provided at 1,000 mg to 20 mg per day. Preferably, the dietary supplement comprises three or more of the ingredients listed in Table 1. Preferably, the virus is a coronavirus. Preferably, the coronavirus is SARS-CoV-2. Preferably, the virus is HIV. Preferably, the virus is HCV. Preferably, the treatment includes treatment of an active infection. Preferably, the active infection is asymptomatic. Preferably, the treatment is prophylactic. Preferably, it involves the administration of radiation therapy. Preferably, said radiation therapy comprises a non-zero radiation dose of up to 2 rem. Preferably, said radiation therapy comprises a single application of radiation. Preferably, the radiation therapy comprises multiple applications of radiation. Preferably, it includes combination therapy with a cancer chemotherapy drug. Preferably, the cancer chemotherapeutic agent is selected from the group consisting of taxol, adriamycin, and avastin. Preferably, when the cancer chemotherapeutic drug is administered for the treatment of cancer, it is applied at a dose equivalent to 1% to 50% of the dose of the cancer chemotherapeutic drug.

[0012] 1. Use of a nutritional supplement comprising fish oil and selenium for the treatment of a viral infection in an individual, wherein the nutritional composition is provided in an amount effective to treat the viral infection or the sequelae of the viral infection, wherein the viral infection is caused by infection with a virus that induces dysregulation of apoptosis in virus-infected cells, and the nutritional supplement provides fish oil and selenium in amounts sufficient to modify apoptotic events in the virus-infected cells upon administration. Preferably, the amounts of fish oil and selenium are selected to further reduce serum concentrations of pro-inflammatory cytokines in the individual. Preferably, the selenium is provided in the form of selenium yeast. Preferably, the selenium is provided at up to 10,000 μg per day. Preferably, the fish oil is provided in up to 20 grams per day. Preferably, the dietary supplement further comprises coenzyme Q10. Preferably, the dietary supplement contains 1 mg to 5,000 mg of coenzyme Q10. Preferably, the dietary supplement comprises three or more of the ingredients listed in Table 1. Preferably, the virus is a coronavirus. Preferably, the coronavirus is SARS-CoV-2. Preferably, the virus is HIV. Preferably, the virus is HCV. Preferably, said treatment includes treatment of an active infection. Preferably, the active infection is asymptomatic. Preferably, the treatment is prophylactic. Preferably, said use is in combination with radiotherapy. Preferably, said radiation therapy comprises a non-zero radiation dose of up to 2 rem. Preferably, said radiation therapy comprises a single application of radiation. Preferably, the radiation therapy comprises multiple applications of radiation. Preferably, said use is in combination with a combination therapy with a cancer chemotherapy drug. Preferably, the cancer chemotherapeutic agent is selected from the group consisting of taxol, adriamycin, and avastin. Preferably, when the cancer chemotherapeutic drug is administered for the treatment of cancer, it is applied at a dose equivalent to 1% to 50% of the dose of the cancer chemotherapeutic drug.

[0013] Various objects, features, aspects and advantages of the present subject matter will become apparent from the following detailed description of preferred embodiments, along with the accompanying drawings. [Brief explanation of the drawings]

[0014] [Figure 1] FIG. 1 shows the effect of selenium and fish oil application on BAX expression in cells with reduced apoptosis (in this case primary tumor cells). [Figure 2] FIG. 2 shows the effect of selenium and fish oil application on BAX expression in cells with reduced apoptosis (in this case, metastatic tumor cells). [Figure 3] FIG. 3 shows the effect of selenium and fish oil application on Bcl-2 expression in cells with reduced apoptosis (in this case primary tumor cells). [Figure 4] FIG. 4 shows the effect of selenium and fish oil application on Bcl-2 expression in cells with reduced apoptosis (in this case, metastatic tumor cells). [Figure 5] FIG. 5 shows the effect of selenium and fish oil application on caspase 3 expression in cells with reduced apoptosis (tumor cells in this case). [Figure 6] FIG. 6 shows the effect of selenium and fish oil application on the expression of protein markers associated with apoptosis (VEGF, p53, and HIF-α). [Figure 7] FIG. 7 shows the effect of selenium and fish oil application on the expression of caspase 8, which is associated with apoptosis. [Figure 8] FIG. 8 shows the effect of selenium and fish oil application on the expression of cytochrome C, which is associated with apoptosis. [Figure 9] FIG. 9 shows the effect of application of selenium, fish oil, and a combination of selenium and fish oil on the growth of tumor cells in culture, which may act as analogs of certain virus-infected cells. [Figure 10] FIG. 10 shows the effect of application of selenium, fish oil, and a combination of selenium and fish oil on the survival of tumor cells in culture, which may act as analogs of certain virus-infected cells, as shown by staining with trypan blue. [Figure 11] FIG. 11 shows the effect of administering a dietary supplement containing selenium and fish oil on serum TNF-α concentrations in an animal model of inflammation. [Figure 12] FIG. 12 shows the effect of administering a dietary supplement containing selenium and fish oil on serum IL-6 concentrations in an animal model of inflammation. [Figure 13] FIG. 13 shows the effect of applying a dietary supplement containing selenium and fish oil on the expression of muscle-related proteins (UCP3, UCP2) and inflammation-related cytokines (IL6) found in mitochondria and associated with inflammation. [Figure 14] FIG. 14 shows the effect of applying a dietary supplement containing selenium and fish oil on the expression of VEGF. [Figure 15] FIG. 15 shows the effect of applying a dietary supplement containing selenium and fish oil on the expression of VEGF. [Figure 16] FIG. 16 shows the effect of applying a dietary supplement containing selenium and fish oil on the expression of TNF-α. [Figure 17]FIG. 17 shows the effect of applying a dietary supplement containing selenium and fish oil on the expression of TNF-α. [Figure 18] FIG. 18 shows the effect of applying a dietary supplement containing selenium and fish oil on the expression of IL-1β and IL-10. [Figure 19] FIG. 19 shows the effect of applying a dietary supplement containing selenium and fish oil on the expression of IL-1β. [Figure 20] FIG. 20 shows the effect of applying a dietary supplement containing selenium and fish oil (N) on the expression of IL2. [Figure 21] FIG. 21 shows the effect of applying a dietary supplement containing selenium and fish oil (N) on the expression of IL2 and IFN-γ. DETAILED DESCRIPTION OF THE INVENTION

[0015] (Detailed Description of the Invention) The present subject matter provides compositions and methods that provide dietary supplements effective in reducing the inflammatory response that is dysregulated in COVID-19 and modulating apoptotic pathways that are disrupted by SARS-CoV-2 infection of cells. The combined effect is believed to be highly beneficial in treating COVID-19 by (1) reducing SARS-CoV-2 viral load by restoring apoptotic pathways in infected cells and (2) reducing the inflammatory response in COVID-19. The inventors believe that this approach to treating viral infections may also be applicable to other viruses in which apoptosis is disrupted in infected cells and / or there is a clinically significant inflammatory component in active disease (e.g., influenza, HIV, etc.).

[0016] It should be understood that such treatment of viral infections can be treatment of active viral infections. Such active viral infections can be symptomatic, or in some embodiments, asymptomatic infections. Alternatively, in some embodiments, treatment of viral infections can be preventative or prophylactic. In such embodiments, treatment with a dietary supplement of the inventive concept can be effective in preventing active viral infections (e.g., by enhancing effective apoptosis of cells during initial exposure to pathogenic viruses).

[0017] The following discussion provides many exemplary embodiments of the inventive subject matter. While each embodiment represents a single combination of inventive elements, it is understood that the inventive subject matter includes all possible combinations of the disclosed elements. Thus, if one embodiment includes elements A, B, and C and a second embodiment includes elements B and D, it is understood that the inventive subject matter also includes any other remaining combinations of A, B, C, or D, even if not explicitly disclosed.

[0018] In some embodiments, numerical values ​​expressing properties such as amounts and concentrations of ingredients, reaction conditions, and the like, used to describe and claim certain embodiments of the present invention should be understood to be optionally modified by the term "about." Accordingly, in some embodiments, the numerical parameters set forth in the specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained in a particular embodiment. In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodiments of the present invention are approximations, the numerical values ​​set forth in the specific examples are reported as precisely as possible. The numerical values ​​presented in some embodiments of the present invention may contain certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0019] As used throughout this specification and the claims that follow, the meaning of "a," "an," and "the" includes plural referents unless the context clearly dictates otherwise. Also, as used herein, the meaning of "in" includes "in" and "on," unless the context clearly dictates otherwise.

[0020] The recitation of ranges of values ​​herein is merely intended as a shorthand method of referring individually to each individual value falling within the range. Unless otherwise indicated herein, each individual value is incorporated herein by reference as if set forth individually herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or clearly contradicted by context. The use of any and all examples presented with respect to certain embodiments herein, or exemplary language (e.g., "etc."), is intended merely to better illustrate the invention and does not impose limitations on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.

[0021] Groupings of other elements or embodiments of the invention disclosed herein are not to be construed as limitations. Each group member may be referred to and claimed individually or in combination with other members of the group or other elements found herein. One or more members of a group may be included in, or deleted from, a group for reasons of convenience and / or patentability. When such inclusions or deletions are made, the specification herein is deemed to include the group as modified so as to satisfy all Markush group descriptions used in the appended claims.

[0022] The inventors have developed a dietary supplement based on high-dose fish oil and selenium, in addition to other supporting and nutritional ingredients. An example of such a formulation is shown in Table 1. The selenium is preferably provided as selenium yeast or its components (e.g., peptides and / or amino acids prepared from selenium yeast). Thus, the benefits found in studies of fish oil and selenium yeast can be extended to the use of this dietary supplement. This formulation has been found to be well tolerated and has a high level of tolerance. [Table 1] TIFF2025138818000003.tif248168TIFF2025138818000004.tif130168

[0023] In some embodiments, one or more minerals (e.g., selenium, molybdenum, chromium) can be provided in the form of a yeast preparation and / or metal-substituted amino acids. In a preferred embodiment, selenium, molybdenum, and / or chromium are provided in the form of selenium yeast, molybdenum yeast, and chromium yeast, respectively. In a preferred embodiment, the dietary supplement includes at least three ingredients as shown in Table 1, e.g., fish oil, selenium (e.g., in the form of selenium yeast), and coenzyme Q (e.g., 10-5,000 mg of coenzyme Q10, 50 mg-3,000 mg of coenzyme Q10, or 200 mg-1,500 mg of coenzyme Q10). Such additional ingredients provide high levels of essential vitamins, minerals, and amino acids. Other ingredients (e.g., enzymes, lecithin) function to aid in the digestion and absorption of the components of the composition when ingested. The combination of these complementary activities provides a synergistic effect that exceeds the simple additive effects of the individual ingredients. It should be understood that the compositions shown in Table 1 also include certain flavorants (e.g., brown sugar, honey, vanilla flavor, and masking agents) that serve to improve palatability and acceptability. Certain ingredients (e.g., honey, brown sugar, milk, rice protein, casein) can provide both flavor and calories. The inventors have found that combinations of the above flavors are effective in providing compliance with the consumption of the dietary supplement in effective amounts. In some embodiments, such flavors can be omitted without negatively impacting the efficacy of the dietary supplement.

[0024] The ingredients shown in Table 1 can be provided as a single formulation (e.g., as a pill, tablet, capsule, powder, liquid, suspension, etc.) or can be separated into different formulations (e.g., as a pill, tablet, capsule, powder, liquid, suspension, or combinations thereof). The amounts shown in Table 1 are exemplary and represent typical daily dosages provided to an otherwise healthy adult of normal height. These amounts can be adjusted to account for differences in weight, gender, medical condition, etc. For example, a relatively small patient weighing 40 kilograms or less may benefit from a dosage provided at or below the lower end of the range provided, while a relatively large patient weighing 100 kilograms or more may require a dosage provided at the upper end (or higher) of the range described. In some embodiments, such a daily dose can be distributed as multiple doses throughout the day. In some such embodiments, the composition of each such distributed dose can be identical. In other embodiments, the composition of such distributed doses can be different, provided that the sum of such doses provides the required supplementation.

[0025] As shown in a co-pending patent application, the inventors have demonstrated that such nutritional compositions can be effectively used in the treatment of cancer, particularly in combination with conventional therapies. In some embodiments, synergistic effects are observed in such combination therapies. In the context of this specification, synergistic effects are those in which the effect of a combination therapy exceeds the sum of the individual effects of the monotherapies that make up the combination therapy. In investigating the molecular basis of this phenomenon, the inventors have found that the administration of a dietary supplement containing fish oil and selenium is effective in modulating markers related to apoptosis and alleviating the inflammatory effects of these disease processes.

[0026] The inventors have noted that infection with viruses, particularly coronaviruses, can result in the disruption of the apoptotic process in infected cells, and that the proinflammatory "cytokine storm" that can occur in infected individuals refers to a dysregulation of the inflammatory process. Based on this, the inventors have concluded that tumor cells, in certain embodiments, serve as an appropriate model for elements of the viral infection process.

[0027] Without being bound by theory, the inventors believe that dietary supplements containing fish oil and selenium (e.g., as described above) have a similar effect for correcting dysregulation of apoptosis in virus-infected cells and / or reducing inflammation that may accompany viral infections, which cause dysregulation of apoptosis and / or inflammation (e.g., through proinflammatory cytokine expression or release). Examples of such viruses include coronaviruses (e.g., SARS-CoV-2), HIV, HCV, etc. It should be understood that the induction of apoptosis in virus-infected cells is a line of defense against viral infections. For example, Fung and Liu (Frontiers in Microbiology, vol. 5, June 2014) showed that coronaviruses disrupt several cellular pathways that drive coronavirus-infected cells toward programmed cell death, thereby extending cell survival and allowing the cells to produce and release more of the virus. All publications herein are incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference. If the definition or use of a term in an incorporated document contradicts or is contrary to the definition of the term provided herein, the definition of the term provided herein shall apply and the definition of the term in the document shall not apply. Similarly, Hanauske-Abel et al. (PLOS ONE, vol. 8, 2013) have shown that infection with HIV blocks apoptotic pathways, and that their reversal leads to apoptosis of HIV-infected cells.

[0028] Without wishing to be bound by theory, the inventors believe that the application of a dietary supplement containing fish oil and selenium, which have been shown to be effective in regulating apoptosis markers in cancer cells, may be effective in correcting the dysregulation of apoptosis associated with infection with certain viruses. Such viruses include, but are not limited to, SARS-CoV-2 (e.g., in COVID-19), HIV, and HCV. In addition, because such supplements have been shown to be effective in reducing inflammation, the inventors believe that they may also be effective in treating inflammation caused by such viral infections. [Example]

[0029] Figures 1 and 2 show the effect of treatment with a dietary supplement containing fish oil and selenium on BAX-expressing cells exhibiting reduced apoptotic activity (T), in this case primary tumor BAX expression and metastatic (lung) tumor BAX expression (respectively). BAX is considered a marker of apoptosis. Because apoptosis is also suppressed in cells infected with certain viruses (e.g., coronaviruses), the inventors believe this system is highly relevant in the context of viral infections. As shown, BAX expression is low in cells with low apoptotic activity (i.e., untreated tumor cells, T). Treatment with dietary supplements containing fish oil and selenium (PTN, TN) resulted in a dramatic increase in BAX expression, resulting in elevated BAX expression. Therefore, the inventors believe that administration of a composition containing fish oil and selenium (e.g., as shown in Table 1) may be effective in increasing BAX expression in cells infected with certain viruses (e.g., coronaviruses, HIV, HCV, etc.) and may act to correct the suppression of apoptosis in such infected cells. Such modification of apoptosis serves to reduce cell-to-cell spread, reducing viral load and may prevent or reduce the severity of viral disease.

[0030] Bcl-2 is associated with reduced apoptosis. As shown in Figures 3 and 4, Bcl-2 expression was found to be elevated in cells (T) with low apoptotic activity, in this case, tumor cells and metastatic (lung) tumor cells (respectively). Bcl-2 expression was reduced by treatment with either a dietary supplement containing fish oil and selenium (PTN.TN). Therefore, the inventors believe that administration of a composition containing fish oil and selenium (e.g., as shown in Table 1) may be effective in increasing BAX expression in cells infected with certain viruses (e.g., coronavirus, HIV, HCV, etc.) and may act to correct the suppression of apoptosis in such infected cells. Such correction of apoptosis may serve to reduce cell-to-cell spread, reduce viral load, and prevent or reduce the severity of viral disease.

[0031] Figure 5 shows the effect of treatment with a dietary supplement containing selenium and fish oil on caspase 3 expression in cells with reduced apoptosis, in this case, tumor cells (T). As shown in Figure 5, caspase 3 expression is reduced in such cells (T). Caspase 3 expression in such cells is increased by treatment with dietary supplements containing fish oil and selenium (TN, PTN). Therefore, the inventors believe that administration of a composition containing fish oil and selenium (e.g., as shown in Table 1) may be effective in increasing caspase 3 expression in cells infected with certain viruses (e.g., coronavirus, HIV, HCV, etc.) and may act to correct the suppression of apoptosis in such infected cells. Such correction of apoptosis may serve to reduce cell-to-cell spread, reduce viral load, and prevent or reduce the severity of viral disease.

[0032] Table 2 shows typical results of a qPCR study on the expression of apoptotic markers (Bax, Bcl-2, and caspase 3) in cells with low apoptotic activity, in this case tumor cells.

[0033] [Table 2]

[0034] As shown, the Bax / Bcl-2 expression ratio is low in cells with low apoptotic activity (T). Surprisingly, treatment with dietary supplements containing fish oil and selenium (TN, PTN) significantly increased this ratio, indicating increased apoptosis. Caspase 3 was dramatically increased by treatment with dietary supplements containing fish oil and selenium. The inventors believe that dietary supplements of the present invention may induce similar changes in virus-infected cells, such as cells infected with coronavirus, HIV, and / or HCV, thereby increasing apoptosis in infected cells.

[0035] Apoptosis and / or apoptotic events are associated with certain cell cycle phases. Studies of cell cycle phase distribution suggest that treatment with a supplement containing fish oil and selenium modulates cell cycle phase distribution, which in turn can induce apoptosis and / or apoptotic events in treated cells and / or cells of treated individuals. The inventors have found that a dietary supplement containing fish oil and selenium is effective in enhancing apoptosis in tumors in an in vivo model of breast cancer, which the inventors believe can act as an analog of virus-infected cells. For example, Figure 6 shows the effects of using such a dietary supplement (N), certain therapeutic small molecules (Avastin, Taxol, Adriamycin), and combination therapy on the expression of certain apoptotic markers (p53, VEGF, HIF-α) on tumor cells and on the expression of apoptotic markers (specifically, VEGF, p53, and HIF-α) in advanced tumors in mice injected with breast cancer cells. As shown, use of the dietary supplement increases p53 expression while decreasing VEGF and HIF-α expression. The results of a similar study on caspase 8 are shown in Figure 7. Figure 8 shows the effect of treatment with a dietary supplement comprising fish oil and selenium ("N"), certain small molecule drugs (Taxol, Adriamycin, Avastin), and combination treatments on cytochrome C expression in tumor cells, which the inventors believe can serve as analogs or models for virally infected cells. The inventors believe that a dietary supplement of the inventive concept can induce similar changes in virally infected cells, e.g., cells infected with coronavirus, HIV, and / or HCV, thereby promoting apoptosis of such infected cells.

[0036] In addition to biochemical markers of apoptosis, apoptotic effects can be observed directly in tumor cells, which the inventors believe can serve as analogs of virus-infected cells. For example, as shown in Figure 9, the growth of A549 cells in culture is slightly affected by exposure to a selenium-containing supplement or a fish oil-containing supplement, but dramatically affected by a supplement containing both fish oil and selenium. Similarly, as shown in Figure 10, trypan blue staining of A549 cells in culture shows only a slight effect on cell death induction when exposed to a selenium-containing supplement or a fish oil-containing supplement, but a dramatic increase in cell death when exposed to a supplement containing both fish oil and selenium. The inventors believe that a dietary supplement according to the present inventive concept can induce similar effects on growth and cell death induction in virus-infected cells, such as cells infected with coronavirus, HIV, and / or HCV.

[0037] Viral infections can also result in inflammation, which can be characterized by the presence of proinflammatory cytokines in serum. Some viruses (especially coronaviruses) are known to induce a so-called "cytokine storm," which can lead to severe shock and, in some cases, death. Alleviating the body's defensive inflammatory response is an important factor in treating such viral infections. Inflammatory responses, such as those mediated by proinflammatory cytokines, are also observed in tumor-bearing animals. Therefore, the applicant believes that such animals can serve as models of inflammatory conditions, including those induced by viral infections.

[0038] Inflammation is mediated, at least in part, by the elaboration of proinflammatory cytokines. The inventors have noted that elevated levels of such cytokines are also found in some tumors. Without wishing to be bound by theory, the inventors believe that therapies effective in modulating the inflammatory response in such conditions (e.g., decreasing proinflammatory cytokines and / or increasing anti-inflammatory cytokines) may be useful in alleviating the inflammatory response in individuals with certain viral infections (e.g., coronavirus, HCV, HIV, etc.).

[0039] Figures 11 and 12 show the effect of administering a dietary supplement containing fish oil and selenium on the levels of proinflammatory cytokines in tumor-bearing mice. Figure 11 shows the levels of the proinflammatory cytokine serum TNF-α. It is clear that untreated tumor-bearing animals (T) exhibit very high levels of the proinflammatory cytokine TNF-α. Treatment with a dietary supplement containing fish oil and selenium (TN, TPN) resulted in a decrease in serum TNF-α, indicating a reduction in inflammation. The inventors believe that treatment with a dietary supplement containing selenium and fish oil can similarly reduce inflammation in individuals with viral infections (e.g., coronavirus, HCV, HIV, etc.) and can also reduce the occurrence or severity of inflammation, particularly inflammation that leads to the development of a cytokine storm.

[0040] Figure 12 shows the results of a similar study characterizing serum concentrations of IL-6, which has pro-inflammatory activity. As shown, untreated animals (T) exhibit elevated levels of IL-6. Treatment of animals with dietary supplements containing selenium and fish oil (TN, PTN) exhibited a decrease in serum IL-6, indicating reduced inflammation. The inventors believe that dietary supplements of the present invention can induce similar changes in viral infections, such as with coronavirus, HIV, and / or HCV. The inventors believe that treatment with dietary supplements containing selenium and fish oil can similarly reduce inflammation in individuals with viral infections (e.g., with coronavirus, HCV, HIV, etc.) and can reduce the occurrence or severity of inflammation—particularly inflammation that leads to the development of a cytokine storm.

[0041] The reduction in inflammation-related markers by treatment with a dietary supplement containing fish oil and selenium is also evident at the molecular level. The inventors have observed that the use of such a dietary supplement is effective in preventing or reversing changes in the expression of certain biochemical markers associated with inflammation. This is also evident at the molecular level, as shown in Figure 13, which shows the expression of muscle-related proteins (UCP3, UCP2) found in mitochondria and associated with inflammation, and an inflammation-related cytokine (IL6) in the cytosol. The inventors believe that a dietary supplement according to the inventive concept may induce similar changes in viral infections, such as infections with coronavirus, HIV, and / or HCV.

[0042] VEGF is associated with increased angiogenesis during inflammatory processes and contributes to COVID-19-related inflammation. Surprisingly, the inventors discovered that a dietary supplement containing fish oil and selenium can modulate VEGF levels. As shown in Figure 14, in an in vivo model of breast cancer (which the inventors believe acts as an analog of virally infected cells), the use of such a dietary supplement (N), alone or in combination with certain small molecule drugs (in this example, taxol, or "tax," and adriamycin, or "adria"), was found to reduce plasma VEGF concentrations, demonstrating its ability to provide anti-inflammatory and anti-angiogenic effects in viral infections (e.g., coronavirus, HCV, HIV, etc.). Similar results were observed with Avastin (see Figure 15). While such effects were observed with the supplement alone, combination therapy with such a supplement enhanced the VEGF-lowering effects of chemotherapy drugs.

[0043] As shown in Figures 16 and 17, a similar effect is seen in the reduction of the pro-inflammatory cytokine TNF-α in treated animals (TN) versus untreated animals (T), an in vivo breast cancer model that the inventors believe may function as an analog of viral infection. A similar effect is seen in the reduction of the pro-inflammatory cytokines IL1-β and IL-10 (see Figure 18) in treated animals (TN) versus untreated animals (T). Similarly, the effect of a dietary supplement (N) containing a combination of fish oil and selenium, alone or in combination with certain small molecule drugs (taxol and adriamycin), on plasma IL-1β in an in vivo model of breast cancer is shown in Figure 19. The inventors believe that a dietary supplement of the inventive concept may induce similar changes in viral infections, such as coronavirus, HIV, and / or HCV infections, thereby reducing inflammation and improving patient outcomes in the setting of such viral infections.

[0044] Conversely, the inventors have found that the use of a dietary supplement containing fish oil and selenium can increase the plasma concentrations of certain cytokines, both individually and in combination with chemotherapy drugs. For example, as shown in Figures 20 and 21 (right panel), plasma concentrations of IL2 are increased in a treated in vivo model of breast cancer. It should be understood that IL2 is believed to be useful in promoting T cell proliferation and the generation of effector and memory T cells, which are important in responding to viral infections (e.g., COVID-19). IL2 has also been recognized as important in the remission and recovery of patients with COVID-19. Plasma concentrations of IFN-γ, another immune-activating cytokine, have also been observed in studies such as those shown in Figure 21 (left panel). In particular, interferon therapy has been found to be useful in treating various viral infections. The inventors believe that a dietary supplement according to the inventive concept can induce similar changes in viral infections, such as coronavirus, HIV, and / or HCV infections.

[0045] As described above, the present inventors have discovered that treatment with fish oil and selenium (preferably in the form of selenium yeast) can effectively reduce inflammation and improve apoptosis dysregulation, both of which are associated with infection with certain viruses (e.g., SARS-CoV, MERS-CoV, HCV, and HIV). A suitable treatment protocol may include treatment with a composition containing fish oil that provides at least 1 g, 2 g, 3 g, 4 g, 5 g, 6 g, 7 g, 8 g, 9 g, 10 g, 12 g, 14 g, 16 g, 18 g, 20 g, 30 g, or 50 g of fish oil per day for an average-sized individual. Such fish oil may have an EPA:DHA ratio of approximately 3:2. Similarly, such a treatment protocol may involve treatment with a composition containing selenium (preferably in the form of selenium yeast) that provides at least 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 15 mg, 20 mg, 30 mg, 40 mg, or 50 mg per day for an average-sized person. These amounts can be adjusted to accommodate individuals of above- or below-average size. Similarly, such amounts can be adjusted (e.g., adjusted over the course of treatment) to accommodate the severity of the viral infection.

[0046] Such a dose of the dietary supplement comprising selenium and fish oil can be provided in any suitable schedule. For example, the daily dose can be distributed throughout the day (e.g., from once an hour to once every 12 hours) or can be provided as a single dose. In some embodiments, such a dose of the dietary supplement can be administered daily, every two days, every three days, weekly, every two weeks, or at intervals longer than that. In some embodiments, the treatment period can be from one day to continuous, and can be limited to one week, two weeks, three weeks, one month, two months, three months, six months, one year, or more than one year.

[0047] In some embodiments of the present invention, one or more supplementary drugs useful in the treatment of viral infections can be administered in combination with treatment with a dietary supplement comprising selenium and fish oil. For example, monoclonal antibodies against SARS-CoV-2, remdesivir, corticosteroids (e.g., dexamethasone), IL-6 inhibitors, barictinib, and / or tocilizumab have been used to treat COVID-19. Protease inhibitors and reverse transcriptase inhibitors have been used to treat HIV infection. Interferons, ribavirin, daclatasvir, elbasvir, grazoprevir, glecaprevir, pibrentasvir, ombitasvir, paritaprevir, ritonavir, dasabuvir, simeprevir, sofosbuvir, velpatasvir, and voxilaprevir have been used to treat HCV infection. In some embodiments, the one or more additional drugs can be provided on the same schedule as the dietary supplement comprising selenium and fish oil. In other embodiments, one or more additional drugs can be provided on a schedule that is different from the schedule of the dietary supplement containing selenium and fish oil, and such combination therapy can provide a synergistic effect (i.e., a greater than additive effect observed from treatment with the individual components of the combination therapy) in treating viral diseases.

[0048] The inventors noted that the developing data from these studies indicate that the administration of radiation therapy can enhance the observed effects of treatment with dietary supplements fish oil and / or selenium. These in vivo studies used low doses of radiation, on the order of 2 rem or less. As shown in Figures 1-5 and Figures 11 and 12, combination therapy using such low levels of radiation in combination with a dietary supplement containing selenium and fish oil (TRN, PTRN) can enhance the effects seen with treatment with the dietary supplement alone and provide a synergistic effect (i.e., greater than the additive effect of treatment with radiation and the dietary supplement individually). Thus, in some embodiments of the inventive concept, fish oil and selenium are provided in combination with low-dose (e.g., 2 rem or less) radiation therapy to treat viral infections. Based on the data obtained on the use of such dietary supplements in combination with radiation therapy, the inventors believe that such radiation therapy can enhance the effects of selenium and fish oil therapy in reducing inflammation and correcting dysregulated apoptosis in individuals infected with the viruses detailed above. Such radiation can be administered before, during, or after administration of a suitable nutritional supplement. Such radiation therapy can be administered once or as a series of repeated applications. The frequency of repeated applications of radiation therapy can be daily, every other day, every three days, twice a week, weekly, or every two weeks.

[0049] The inventors have noted that the developing data from these studies indicate that the administration of chemotherapy with small molecule drugs used to treat cancer (e.g., taxol or "tax," adriamycin or "adria," and avastin) can enhance the observed effects of treatment with dietary supplements fish oil and / or selenium. As shown in Figures 6-8 and 13-18, combination therapy using such chemotherapy drugs in combination with a dietary supplement containing selenium and fish oil can enhance the effects seen with treatment using the dietary supplement alone and can provide a synergistic effect (i.e., greater than the additive effect of treatment with the chemotherapy drug and the dietary supplement individually). The inventors believe that such enhanced effects of a dietary supplement containing selenium and fish oil can be observed at lower doses of chemotherapy drugs than those used in cancer chemotherapy (e.g., 1%, 3%, 5%, 10%, 20%, 30%, 40%, or 50% of conventional doses), allowing them to be used with minimal or no side effects associated with cancer chemotherapy. Thus, in some embodiments of the present invention, fish oil and selenium are provided in combination with a cancer chemotherapy drug to treat viral infections. Based on data obtained from the use of such dietary supplements in combination with chemotherapy, the inventors believe that combination therapy with one or more such cancer chemotherapy drugs can enhance the effectiveness of selenium and fish oil therapy in reducing inflammation and correcting the dysregulation of inflammation and / or apoptosis in individuals infected with the viruses detailed above. In such embodiments, one or more chemotherapy drugs used in cancer treatment can be provided before, during, or after the administration of a suitable dietary supplement. Such combination therapy with one or more chemotherapy drugs used in cancer treatment can be administered once or as a series of repeated applications. The frequency of repeated combination therapy with one or more chemotherapy drugs used in cancer treatment can be daily, every other day, every three days, twice a week, weekly, or every two weeks.

[0050] It will be apparent to those skilled in the art that many modifications beyond those already described are possible without departing from the inventive concepts herein. Accordingly, the inventive subject matter is not to be limited except insofar as the spirit of the appended claims is apparent. Moreover, in interpreting both this specification and the claims, all terms should be interpreted in the broadest possible manner consistent with the context. In particular, the terms "comprises" and "comprising" should be interpreted to refer to elements, components, or steps in a non-exclusive manner, indicating that the referenced element, component, or step may be present, utilized, or combined with other elements, components, or steps not specifically mentioned. When this specification and claims refer to at least one of something selected from the group consisting of A, B, C, and N, such context should be interpreted as requiring only one element from the group, and not A+N, or B+N, etc.

Claims

1. 50mg-20,000mg fish oil, 30µg-10,000µg selenium yeast, 10,000mg-50,000mg maltodextrin, 5,000mg-60,000mg whey protein isolate, 1,000mg-50,000mg whey protein concentrate, 40mg-15,000mg fructooligosaccharides / inulin, 1,000mg-9,000mg granulated honey, 500mg-15,000mg oat fiber, 500mg-20,000mg natural French vanilla flavor, 500mg-50,000mg soy protein Protein, 500mg-10,000mg brown powdered sugar, 500mg-5,000mg natural vanilla masking flavor, 200mg-10,000mg lecithin, 50mg-5,000mg nonfat milk, 50mg-5,000mg rice protein powder, 50mg-2,000mg calcium caseinate, 100mg-7,000mg flax oil, 100mg-7,000mg canola oil, 100mg-7,000mg borage oil, 100mg-7,000mg olive oil, 100mg-1,000mg pure lemon oil, 50mg-1, 000mg pure orange oil, 0.5mg-200mg mixed tocopherols, 200mg-1,500mg potassium phosphate, 100mg-5,000mg calcium carbonate, 150mg-2,500mg choline bitartrate, 100mg-2,000mg sodium chloride, 100mg-2,000mg tribasic calcium phosphate, 50mg-3,000mg ascorbic acid, 50mg-2,000mg potassium chloride, 50mg-500mg magnesium oxide, 30µg-3,000µg chromium yeast, 30µg-2,000µg Molybdenum yeast, 10 mg to 5,000 mg inositol, 5 mg to 200 mg zinc sulfate monohydrate, 5 IU to 2,000 IU dried vitamin E acetate, 5 mg to 500 mg niacinamide, 3 mg to 100 mg ferric orthophosphate, 3 mg to 200 mg calcium pantothenate, 3 mg to 100 mg manganese sulfate monohydrate, 1 mg to 100 mg beta-carotene, 1 mg to 15 mg copper gluconate, 25 IU to 5,000 IU vitamin D3, 2 μg to 1,000 μg vitamin K2, 0.5 mg to 200 mg pyridoxine HCl, 0.5mg to 1,500mg potassium iodide, 0.5mg to 1,000mg riboflavin, 0.5mg to 2,500mg thiamine hydrochloride, 1μg to 500μg dried vitamin K1, 500IU to 100,000IU vitamin A acetate, 100μg to 10,000μg folic acid, 10μg to 10,000μg d-biotin, 1μg to 3,000μg vitamin B12, 300mg to 30,000mg L-carnitine, 500m g to 60,000 mg of L-glutamine, 500 mg to 30,000 mg of L-arginine base, 50 mg to 2,000 mg of taurine, 50 mg to 2,000 mg of L-lysine, 10 mg to 1,000 mg of alpha lipoic acid, 15 mg to 1,500 mg of resveratrol, 10 mg to 5,000 mg of coenzyme Q10, 5 mg to 1,000 mg of glycine, 5 mg to 1,000 mg of proline, 2 mg to 500 mg of Lact. Acidophilus (total of approximately 10 billion), 2 mg to 500 mg of Bifidobacterium bifidum (total of approximately 10 billion), 2 mg to 500 mg of Lac. Bulgaricus (approximately 10 billion in total), 2mg-500mg Bifidobacterium Longum (approximately 10 billion in total), 2mg-500mg Strep. thermophilus (approximately 10 billion in total), 5mg-100mg papain, 5mg-100mg pepsin, 5mg-100mg lipase, 5mg-100mg bromelain, 0.5mg-100mg pancreatin 4X, 1mg-100mg lactase, 3mg-100mg betaine HCl, 2mg-500mg pineapple juice powder, 2mg-500mg papaya fruit powder, 30mg-3,000mg quercetin, 25mg-600mg EGCG, 15mg-500mg 1. A dietary supplement for treating a viral infection or the sequelae of said viral infection, comprising: 15 mg of OPC, 15 mg to 5,000 mg of anthocyanins, 10 mg to 300 mg of ellagic acid, 2 mg to 90 mg of astaxanthin, 20 mg to 1,500 mg of fucoidan, 5 mg to 6,000 mg of cordyceps, 15 mg to 10,000 mg of Ganoderma lucidum, 40 mg to 15,000 mg of shiitake mushroom, 30 mg to 15,000 mg of maitake mushroom, and 30 mg to 15,000 mg of Coriolus versicolor. wherein the nutritional ingredients are provided in an amount effective to treat the viral infection or sequelae of the viral infection, the viral infection being due to infection with the virus that induces dysregulation of apoptosis in virus-infected cells, the nutritional supplement providing fish oil and selenium in amounts sufficient to modify apoptotic events in the virus-infected cells upon administration, and the nutritional supplement is formulated for use in combination with radiation therapy or a cancer chemotherapy drug.

2. 10. The dietary supplement of claim 1, wherein the amount of said dietary supplement is effective to reduce serum concentrations of pro-inflammatory cytokines.

3. 2. The dietary supplement of claim 1, wherein the virus is a coronavirus.

4. 4. The dietary supplement of claim 3, wherein the coronavirus is SARS-CoV-2.

5. 2. The dietary supplement of claim 1, wherein the virus is HIV.

6. 2. The dietary supplement of claim 1, wherein the virus is HCV.

7. 10. The dietary supplement of claim 1, wherein said treatment comprises treatment of an active infection.

8. 8. The dietary supplement of claim 7, wherein the active infection is asymptomatic.

9. 10. The dietary supplement of claim 1, wherein the treatment is prophylactic.

10. 10. The dietary supplement of claim 1, wherein said radiation therapy comprises a non-zero radiation dose of up to 2 rem.

11. 10. The dietary supplement of claim 9, wherein said radiation therapy comprises a single application of radiation.

12. 10. The dietary supplement of claim 9, wherein said radiation therapy comprises multiple applications of radiation.

13. 10. The dietary supplement of claim 1, wherein the cancer chemotherapy drug is selected from the group consisting of taxol, adriamycin, and avastin.

14. 10. The dietary supplement of claim 1, wherein said cancer chemotherapy drug is provided in a dose equivalent to 1% to 50% of the dose of said cancer chemotherapy drug provided for the treatment of cancer.

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