Method and composition for delivering biotin to mitochondria

Abstract

An exemplary embodiment is directed to a biologically active composition of matter (and use thereof) configured for targeted delivery of biotin to mitochondria, the composition comprising a first D-biotin conjugated to a water-soluble, cell-permeable, peptide sequence, the peptide sequence being selected from a group of polypeptides having alternating aromatic-cationic motifs.

Description

[Technical Field] 【0001】 Cross-reference of related applications [Background technology] 【0002】 This application claims priority to U.S. Provisional Patent Application No. 63 / 075,996 (Attorney Docket 1255-003), filed on September 9, 2020, and to U.S. Provisional Patent Application No. 63 / 076,022 (Attorney Docket 1255-004), filed on September 9, 2020, which are incorporated herein by reference in their entirety. [Prior art documents] [Patent Documents] 【0003】 [Patent Document 1] U.S. Patent No. 4749742 [Patent Document 2] U.S. Patent No. 5026773 [Patent Document 3] U.S. Patent No. 7576061 [Patent Document 4] U.S. Patent No. 9388212 [Patent Document 5] U.S. Patent No. 9695214 [Patent Document 6] U.S. Patent No. 10125163 [Patent Document 7] U.S. Patent Application Publication No. 2019 / 0202861 [Patent Document 8] U.S. Patent Application Publication No. 2019 / 0015521 [Patent Document 9] U.S. Patent Application Publication No. 2012 / 0149868 [Patent Document 10] U.S. Patent No. 5,391,711 [Patent Document 11] U.S. Patent No. 5,416,016 [Patent Document 12] U.S. Patent Application Publication No. 2006 / 0149035 [Patent Document 13] U.S. Patent No. 2,720,527 [Overview of the Initiative] [Means for solving the problem] 【0004】 A wide variety of potential, feasible, and / or useful embodiments will be more readily apparent through the non-limiting, non-exclusive description of an exemplary embodiment provided herein, by referring to the accompanying illustrative drawings below. [Brief explanation of the drawing] 【0005】 [Figure 1A] Figure 1A is a chemical structure diagram showing how biotin can be conjugated to mitochondrial target peptide sequences. [Figure 1B] Figure 1B is a chemical structure diagram showing how biotin can be conjugated to mitochondrial target peptide sequences. [Figure 1C] Figure 1C is a chemical structure diagram showing how biotin can be conjugated to mitochondrial target peptide sequences. [Figure 2] Figure 2 shows fluorescence microscopy images related to the intracellular uptake of an exemplary compound. [Figure 3] Figure 3 is a graph illustrating exemplary intracellular uptake. [Figure 4] Figure 4 shows fluorescence microscopy images related to the intracellular uptake of an exemplary compound. [Figure 5] Figure 5 is a graph illustrating exemplary intracellular uptake. [Figure 6] Figure 6 shows fluorescence microscopy images related to mitochondrial targets of a particular exemplary compound. [Figure 7] Figure 7 is a graph showing exemplary cell viability. [Figure 8] Figure 8 is a microscopic image showing exemplary cell counts. [Figure 9]Figure 9 is a graph showing exemplary cell viability. [Figure 10] Figure 10 is a graph showing exemplary intracellular ATP levels. [Figure 11] Figure 11 is a graph showing exemplary intracellular ATP levels. [Figure 12] Figure 12 is a graph showing exemplary intracellular ATP levels. [Figure 13] Figure 13 is a fluorescence microscope image showing mitochondrial potential. [Figure 14] Figure 14 is a fluorescence microscope image showing exemplary mitochondrial potential. [Figure 15] Figure 15 is a microscope image of a scratch area related to an exemplary compound. [Figure 16] Figure 16 is a graph showing an exemplary scratch area. [Figure 17] Figure 17 is a graph showing an exemplary scratch area. [Figure 18] Figure 18 is a microscope image of a scratch area related to an exemplary compound. [Figure 19] Figure 19 is a graph showing an exemplary scratch area. [Figure 20] Figure 20 is a microscope image related to the cell proliferation of an exemplary compound. [Figure 21] Figure 21 is a graph showing exemplary cell proliferation. [Figure 22] Figure 22 is a microscope image related to the cell proliferation of an exemplary compound. [Figure 23] Figure 23 is a graph showing exemplary cell proliferation. [Figure 24] Figure 24 is a microscope image related to the mitochondrial potential of an exemplary compound. [Figure 25] [[ID=**48**]]Figure 25 is a graph showing exemplary mitochondrial potential. [Figure 26] Figure 26 is a fluorescence microscope image related to the retinal uptake of an exemplary compound. [Figure 27]Figure 27 shows fluorescence microscopy images related to retinal uptake of an exemplary compound. [Modes for carrying out the invention] 【0006】 Some exemplary embodiments described herein may relate to methods and compositions for enhancing the intracellular and / or mitochondrial uptake of biotin. Targeted delivery of biotin to mitochondria can improve the efficacy of biotin and avoid the use of high doses of biotin. 【0007】 One exemplary embodiment described herein can provide a method for using a water-soluble, cell-permeable, mitochondrial-targeting peptide sequence to deliver biotin to mitochondria. 【0008】 Some exemplary embodiments described herein relate to short water-soluble peptide sequences selected from a group of polypeptides having a common aromatic-cationic motif, meaning that the amino acids of the polypeptide group can be arranged, for example, as [aromatic-cationic-aromatic-cationic] or [cationic-aromatic-cationic-aromatic]. 【0009】 Some exemplary embodiments described herein may include polypeptides composed of naturally occurring amino acids. 【0010】 Some exemplary embodiments described herein may include D-amino acids, which can help make the peptide more resistant to hydrolysis by peptidase enzymes. Some exemplary embodiments described herein may include polypeptides composed of one or more amino acids that do not exist naturally. Amino acids that do not exist naturally are typically those that are not synthesized in the normal metabolic processes of living organisms and are not naturally present in proteins. Amino acids that do not exist naturally may include derivatives of naturally occurring amino acids. 【0011】 Peptides useful in this invention may contain one or more amino acids that do not exist in nature. These amino acids may be L-, dextrorotatory (D), or a mixture thereof. Optimally, the peptides do not contain any naturally occurring amino acids. 【0012】 Amino acids that do not exist naturally are amino acids that do not exist naturally in proteins. Furthermore, amino acids that do not exist naturally and are useful in this invention are preferably generally not recognized by proteases. 【0013】 Amino acids that do not exist in nature can be present at any position in a peptide. For example, amino acids that do not exist in nature can be present at the N-terminus, the C-terminus, or any position between the N-terminus and the C-terminus. 【0014】 Unnatural amino acids may contain, for example, alkyl, aryl, or alkylaryl groups. Some examples of alkyl amino acids include α-aminobutyric acid, β-aminobutyric acid, γ-aminobutyric acid, δ-aminovaleric acid, and ε-aminocaproic acid. Some examples of aryl amino acids include ortho-, meta-, and para-aminobenzoic acid. Some examples of alkylaryl amino acids include ortho-, meta-, and para-aminophenylacetic acid, and γ-phenyl-β-aminobutyric acid. 【0015】 Amino acids that do not exist in nature include derivatives of naturally occurring amino acids. Derivatives of naturally occurring amino acids may include, for example, the addition of one or more chemical groups to naturally occurring amino acids. 【0016】 For example, one or more chemical groups can be attached to one or more of the 2', 3', 4', 5', or 6' positions of the aromatic ring of a phenylalanine or tyrosine residue, or to one or more of the 4', 5', 6', or 7' positions of the benzo ring of a tryptophan residue. The group can be any chemical group that can be attached to an aromatic ring. Some examples of such groups include branched or unbranched C1-C4 alkyl groups such as methyl, ethyl, n-propyl, isopropyl, butyl, isobutyl, or t-butyl, C1-C4 alkyloxy (i.e., alkoxy), amino, C1-C4 alkylamino and C1-C4 dialkylamino (e.g., methylamino, dimethylamino), nitro, hydroxyl, and halo (i.e., fluoro, chloro, bromo, or iodine). Some specific examples of derivatives of naturally occurring amino acids that do not exist in nature include norvaline (Nva), norleucine (Nle), and hydroxyproline (Hyp). 【0017】 Another example of amino acid modification in peptides useful in the methods of the present invention is the derivatization of the carboxyl group of an aspartic acid or glutamic acid residue in the peptide. An example of derivatization is amidation using ammonia or a primary or secondary amine, such as methylamine, ethylamine, dimethylamine, or diethylamine. Another example of derivatization includes esterification using methyl or ethyl alcohol, for example. 【0018】 Other such modifications include the derivatization of the amino group of a lysine, arginine, or histidine residue. For example, such an amino group can be acylated. Some suitable acyl groups include, for example, benzoyl or alkanoyl groups containing any of the above C1-C4 alkyl groups, such as acetyl or propionyl groups. One exemplary embodiment described herein involves the amidation of the C-terminus of a peptide sequence, which can help make the peptide more resistant to hydrolysis by carboxypeptidases. 【0019】 The carboxyl group, particularly the terminal carboxyl group of the C-terminal amino acid, can be amidated with ammonia, for example, to form a C-terminal amide. Alternatively, the terminal carboxyl group of the C-terminal amino acid may be amidated with any primary or secondary amine. The primary or secondary amine may be, for example, alkyl, particularly branched or unbranched C1-C4 alkyl, or allylamine. Thus, the C-terminal amino acid of the peptide may be converted to an amide, N-methylamide, N-ethylamide, N,N-dimethylamide, N,N-diethylamide, N-methyl-N-ethylamide, N-phenylamide, or N-phenyl-N-ethylamide group. 【0020】 The free carboxylate groups of asparagine, glutamine, aspartic acid, and glutamic acid residues that are not present at the C-terminus of the aromatic-cationic peptide of the present invention may be amidated at any location within the peptide. These internal amidations may be carried out using ammonia or either of the primary or secondary amines mentioned above. 【0021】 Some exemplary embodiments described herein may include polypeptides composed of as few as four amino acids. 【0022】 Some exemplary embodiments described herein may include polypeptides comprising six or fewer amino acids. 【0023】 One exemplary embodiment described herein is directed to a water-soluble peptide sequence selected from polypeptides consisting of 4 to 6 amino acids (in D or L configuration) having alternating aromatic-cationic motifs. 【0024】 Some exemplary embodiments described herein are D-Arg-L-(2'6'-dimethylTyr)-L-Lys-L-Phe-NH2(SS-31) D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys-NH2(SPN02); D-Trp-D-Arg-D-Trp-D-Lys-OH (SPN07); L-Trp-L-Arg-L-Trp-L-Lys-NH2(SPN10); D-Trp-D-Arg-D-Trp-D-Lys-NH2(SPN13); and L-Trp-L-Arg-L-Trp-L-Lys-OH (SPN14) This is directed towards water-soluble peptide sequences, including but not limited to these. 【0025】 One exemplary embodiment described herein can provide a method for conjugating D-biotin to a short (i.e., 4-6 amino acid), water-soluble, cell-permeable, mitochondrial target peptide sequence for targeted delivery of biotin to mitochondria. 【0026】 Certain exemplary peptide sequences described herein are defined by their N-terminus and C-terminus, and may contain an α-amine at the N-terminus. A particular exemplary embodiment described herein can provide a method for conjugating D-biotin to the N-terminal α-amine of a mitochondrial target peptide sequence. 【0027】 Some exemplary peptide sequences described herein may include a lysine residue having an ε-amine at its C-terminus. Some exemplary embodiments described herein can provide a method for conjugating D-biotin to the ε-amine of a lysine residue at the C-terminus of a mitochondrial target peptide sequence (ε-N-[d-biotinyl]-L-lysine). 【0028】 One exemplary embodiment described herein can provide a method for conjugating D-biotin to both the N-terminal α-amine and the C-terminal ε-amine on a lysine residue of a mitochondrial target peptide sequence. 【0029】 Some exemplary embodiments described herein are directed to at least one therapeutically effective composition of a substance and / or a method for preparing and / or using one or more such compositions and / or components thereof, the composition is D-biotin-D-Arg-L-(2'6'-dimethylTyr)-L-Lys-L-Phe-NH2(SPN05); D-biotin-D-Trp-D-Arg-D-Trp-D-Lys-OH (SPN08); D-Trp-D-Arg-D-Trp-D-Lys(biothinyl)-OH (SPN09); L-Trp-L-Arg-L-Trp-L-Lys(biothinyl)-NH2(SPN11); D-biotin-L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH2(SPN12); D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biothinyl)-NH2(SPN15); and D-biotin-D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH2(SPN16) It comprises one or more compounds selected from the polypeptide group consisting of the following. 【0030】 One or more exemplary compositions described herein may contain one or more peptide-conjugated biotin molecules adapted to stimulate intracellular ATP production. The exemplary compositions may also contain vitamins and / or amino acids and / or metabolic supplements in concentrations effective for promoting ATP synthesis. 【0031】 One or more exemplary compositions described herein may include one or more peptide-conjugate biotin molecules adapted to maintain mitochondrial potential. 【0032】 One exemplary embodiment described herein relates to a novel composition and / or method adapted to promote cell viability when cells are cultured ex vivo in the absence of serum. One or more exemplary compositions described herein may contain one or more peptide-conjugated biotin molecules adapted to stimulate mitochondrial function, such as promoting cell proliferation. One or more exemplary compositions described herein may contain vitamins and / or amino acids and / or metabolic supplements in effective concentrations that exhibit additive or synergistic activity in maintaining cell growth. 【0033】 Due to the limited number of stem cells, extensive ex vivo proliferation is required to obtain a sufficient number of cells to treat clinical signs in order to carry out cell therapy. Culture media containing fetal bovine serum (FBS) provide a supportive environment for the isolation and proliferation of mesenchymal stem cells. FBS provides adhesion factors, growth factors, and numerous other nutrients. However, in addition to the inherent variability and limited comprehensive supply of FBS, serum products may also contain serum proteins that can be sources of pathogens and / or may trigger an immune response in human recipients. 【0034】 Serum depletion causes cell cycle arrest and apoptosis, severely limiting the yield of stem cells or primary cells for clinical use. To enable stem cell proliferation, various growth factors must be added to serum-free cultures, often involving the use of human-derived supplements that may be contaminated with human pathogens. Some exemplary media described herein contain chemically defined but serum-free and xeno-free components that support the growth and adhesion of human primary cells and subcultures, enabling large-scale production of primary mammalian cells for clinical use. 【0035】 Large-scale production of primary mammalian cells can be crucial for laboratory production of meat as a substitute for traditional livestock-derived meat. Culture of animal myoblasts using FBS is not sustainable. Some exemplary media described herein contain chemically defined, serum-free components to support the proliferation of animal myoblasts for large-scale production of laboratory-grown meat. 【0036】 Some exemplary embodiments described herein relate to novel compositions and / or methods adapted to promote cell or tissue survival for transplantation in humans. One or more exemplary compositions described herein may contain one or more peptide-conjugated biotin molecules adapted to stimulate mitochondrial intracellular ATP production, which promotes cell survival. One or more exemplary compositions described herein may contain vitamins and / or amino acids and / or metabolic supplements in effective concentrations that exhibit additive or synergistic activity in maintaining organ survival. 【0037】 Islet transplantation is an approach to β-cell replacement therapy for type 1 diabetes. Adding one or more peptide-conjugated biotin molecules that can improve cell viability throughout the harvesting and purification procedures can significantly improve islet yield and / or increase the success of blood glucose control in recipients. 【0038】 Ischemia and hypoxia are unavoidable events during organ preservation prior to transplantation. After organs are deprived of a normal blood supply, depletion of mitochondrial ATP synthesis leads to cell death. Storage duration varies from 4-6 hours for the heart and lungs to up to 36 hours for the kidneys. Adding one or more peptide-conjugated biotin molecules that improve cell viability in organ preservation solutions can extend organ survival time, allowing for wider distribution to suitable recipients. Improved transplant organ quality can also reduce post-transplant organ dysfunction and failure. 【0039】 Some exemplary embodiments described herein relate to novel compositions and / or methods adapted to promote tissue health and / or prevent tissue damage. Some exemplary methods described herein provide systemic administration of exemplary compositions to mammals. The compositions may contain one or more peptide-conjugated biotin molecules adapted to stimulate mitochondrial intracellular ATP production and promote cellular function. The compositions may contain vitamins and / or amino acids and / or metabolic supplements in effective concentrations that exhibit additive or synergistic activity in maintaining tissue health. 【0040】 Aging is associated with a decrease in the proliferative capacity of many cell types, including skin and epithelial membranes. Intestinal epithelium can completely regenerate within 5 days, while lung epithelium can take up to 6 months to regenerate. Aging reduces epithelial regenerative capacity, leading to progressive epithelial damage. 【0041】 Most tissues exhibit a progressive decline in regenerative capacity with age, leading to tissue degeneration, dysfunction, and pathological conditions. Certain methods described herein can enhance tissue health during aging and prevent and / or reduce many of these age-related functional impairments. 【0042】 The exemplary methods described herein relate to novel compositions and / or methods adapted for promoting tissue health that can promote the proliferation of resident stem / progenitor cells in vivo in aged and / or damaged tissues. 【0043】 One exemplary embodiment described herein relates to a novel composition and / or method adapted to promote tissue repair and regeneration. The exemplary method described herein provides systemic administration of the exemplary composition to a mammal after tissue injury. The composition may contain one or more peptide-conjugated biotin molecules adapted to stimulate mitochondrial intracellular ATP production to promote cell proliferation and / or tissue regeneration. The composition may contain vitamins and / or amino acids and / or metabolic supplements in effective concentrations that exhibit additive or synergistic activity in maintaining tissue health. 【0044】 Tissue damage may include damage to the skin and / or soft tissues (e.g., muscles, tendons, ligaments, nerves, blood vessels), and / or hard tissues (e.g., bones, teeth), and / or parenchymal organs (e.g., heart, kidneys, lungs, liver, spleen, intestines, etc.). Causes of tissue damage may include, for example, trauma, hypoxia (i.e., hypoxic supply), ischemia (i.e., low blood flow), infectious pathogens, drugs, chemicals, and / or toxins, etc. 【0045】 Some exemplary embodiments described herein relate to novel compositions and / or methods adapted for the treatment of wounds and / or tissue damage, and / or for promoting tissue regeneration and faster wound healing. Some exemplary methods described herein provide direct application of compositions to wounds, such as stimulating (i.e., promoting) the proliferation (i.e., increasing the number of cells) of adjacent cells around a wound, thereby promoting and / or achieving wound closure. The compositions may contain one or more peptide-conjugated biotin molecules adapted to stimulate mitochondrial intracellular ATP production to promote cell proliferation and / or migration (i.e., cell movement). The compositions may contain vitamins and / or supplements and / or metabolic supplements in effective concentrations that exhibit additive or synergistic activity in stimulating wound healing, such as for postoperative wounds and / or non-healing chronic wounds, e.g., pressure ulcers and / or diabetic ulcers and / or venous ulcers in patients. 【0046】 The exemplary methods described herein relate to novel compositions and / or methods adapted for treating tissue injury that can promote the proliferation of resident stem / progenitor cells in vivo in damaged tissue. 【0047】 One exemplary embodiment is a pharmaceutical composition that may be useful in promoting tissue repair and / or regeneration, (a) One or more compounds that promote ATP production and cell proliferation; (b) at least one vitamin; and / or (c) at least one amino acid; and / or (d) at least one metabolic supplement A composition containing the above can be provided. 【0048】 One exemplary embodiment described herein is a method for enhancing mitochondrial ATP production in mammals, wherein the formula is: D-biotin-D-Arg-L-(2'6'-dimethylTyr)-L-Lys-L-Phe-NH2(SPN05); D-biotin-D-Trp-D-Arg-D-Trp-D-Lys-OH (SPN08); D-Trp-D-Arg-D-Trp-D-Lys(biothinyl)-OH (SPN09); L-Trp-L-Arg-L-Trp-L-Lys(biothinyl)-NH2(SPN11); D-biotin-L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH2(SPN12); D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biothinyl)-NH2(SPN15); and / or D-biotin-D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH2(SPN16) This includes administering a therapeutically effective amount of a compound having the above properties to a mammal. The compound can be provided as a composition containing a pharmaceutically acceptable carrier, and a method of administration to mammals can be provided. 【0049】 One exemplary embodiment described herein provides a composition designed to increase cell proliferation for primary cells cultured ex vivo without serum, comprising an active ingredient comprising one or more peptide-conjugated biotin molecules that stimulate mitochondrial ATP production and cell proliferation, and a second ingredient comprising at least one vitamin and / or amino acid and / or metabolic supplement that enhances ATP production and cell proliferation. 【0050】 One exemplary embodiment described herein provides a composition designed to increase cell and organ survival in ex vivo, comprising an active ingredient comprising one or more peptide-conjugated biotin molecules that stimulate mitochondrial ATP production and cell proliferation, and a second ingredient comprising at least one vitamin and / or amino acid and / or metabolic supplement that enhances ATP production and cell proliferation. 【0051】 One exemplary embodiment described herein provides a composition designed to increase cell proliferation, such as in a wound area, comprising an active ingredient comprising one or more peptide-conjugate biotin molecules and / or a second ingredient comprising a mixture of at least one vitamin and / or one amino acid and / or one metabolic supplement that enhance cell proliferation and migration. 【0052】 Examples of amino acids that can be used with certain exemplary embodiments described herein include L-isomers of all natural amino acids, including essential and depletable amino acids (isoleucine, leucine, alanine, asparagine, lysine, aspartic acid, methionine, cysteine, phenylalanine, glutamic acid, threonine, glutamine, tryptophan, glycine, valine, proline, serine, tyrosine, arginine, histidine), as well as taurine, which is naturally derived from cysteine. 【0053】 Examples of metabolic supplements that can be used with certain exemplary embodiments described herein include pyruvate, carnitine, acetylcarnitine, creatine, alpha-ketoglutaric acid, alpha-lipoic acid, and coenzyme Q 10 It contains nicotinamide riboside and nicotinamide mononucleotide. 【0054】 One exemplary embodiment described herein provides a method for enhancing the viability and proliferation of primary mammalian cells in serum-free, chemically defined media by adding a formulation comprising an effective amount of one or more peptide-conjugate biotin molecules and a mixture of at least one vitamin and / or one amino acid and / or one metabolic supplement to a culture medium. 【0055】 In one or more exemplary embodiments of the methods described herein, primary cells may include bone marrow stem cells or mesenchymal stem cells for autologous, allogeneic, or xenogeneic regenerative medicine applications. 【0056】 In one or more exemplary embodiments of the methods described herein, the primary cells may be stem cells obtained from placenta or umbilical cord blood for allogeneic transplantation. 【0057】 In one or more exemplary embodiments of the methods described herein, the primary cells can be hematopoietic cells such as T cells for chimeric antigen receptor (CAR) T cell therapy. 【0058】 In one or more exemplary embodiments of the methods described herein, primary cells may be mammalian cells cultured for the production of therapeutic proteins such as monoclonal antibodies and / or biologics, and / or for the development and / or production of viral vaccines. 【0059】 In one or more exemplary embodiments of the methods described herein, the primary cells can be animal cells such as myoblasts cultured for laboratory growth and in vitro production of no-slaughter meat. 【0060】 In one or more exemplary embodiments of the methods described herein, primary cells can be islet cells harvested from a donor pancreas and purified in the laboratory before transplantation. 【0061】 One exemplary embodiment described herein can provide a method for optimizing and / or improving stem cell transplantation and / or mitochondrial migration for tissue regeneration by treating stem cells and / or mitochondria with a solution comprising a therapeutically effective amount of one or more peptide-conjugated biotin molecules and a mixture of at least one vitamin and / or one amino acid and / or one metabolic supplement prior to transplantation. 【0062】 In one or more exemplary embodiments of the methods described herein, the cells may be resident stem / progenitor cells in damaged tissue and can be systemically administered to a mixture of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement. 【0063】 In one or more exemplary embodiments of the methods described herein, mitochondrial survival and function can be optimized by systemically administering one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement to a subject after mitochondrial transplantation. 【0064】 In one or more exemplary embodiments of the methods described herein, one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement may be added to a serum-free medium for culturing laboratory-grown meat as a replacement for livestock-derived meat. 【0065】 One exemplary embodiment described herein can provide a method for enhancing an organ preservation solution by adding a preparation comprising an effective amount of one or more peptide-conjugate biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement to the preservation solution. 【0066】 In one or more exemplary embodiments of the methods described herein, the organs may include the kidneys, liver, heart, lungs, pancreas, skin, intestines, cornea, trachea, and / or blood vessels. 【0067】 In one or more exemplary embodiments of the methods described herein, systemic administration of an effective amount of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement to a transplant recipient can improve initial graft function and enhance graft survival. 【0068】 One exemplary embodiment described herein can provide a method for promoting tissue health in a subject by administering a micronutrient preparation comprising an effective amount of one or more peptide-conjugate biotin molecules and a mixture of at least one vitamin and / or one amino acid and / or one metabolic supplement. 【0069】 One exemplary embodiment described herein can provide a method for promoting tissue health in a subject by administering a micronutrient preparation comprising an effective amount of one or more peptide-conjugated biotin molecules and a mixture of acetylcarnitine and / or α-ketoglutaric acid. 【0070】 One exemplary embodiment described herein may provide a method for promoting tissue health in a subject by providing a micronutrient preparation comprising an effective amount of one or more peptide-conjugated biotin molecules and a mixture of acetylcarnitine, α-ketoglutaric acid, and / or taurine. 【0071】 In one or more exemplary embodiments of the methods described herein, tissue health may include keratinous tissue (e.g., skin, hair, and / or nails), and / or muscles, and / or joints, and / or bones, and / or heart, and / or lungs, and / or kidneys, and / or brain, and / or vision and / or hearing health. 【0072】 In one or more exemplary embodiments of the methods described herein, tissue damage may be caused by aging. All cells may experience aging changes. Many cells may lose their ability to function, waste products may accumulate within the cells, connective tissue may harden, and many tissues may lose mass. 【0073】 One exemplary embodiment described herein may provide a method for reducing, mitigating, and / or reversing age-related damage in a subject by administering a composition comprising an effective amount of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement, the administration of which occurs orally, sublingually, and / or subcutaneously, etc. Pharmaceuticals for oral administration may be in the form of solutions, suspensions, or solid forms such as tablets, capsules, and powders, etc. Pharmaceuticals for sublingual administration or subcutaneous injection may be prepared by mixing such compositions with non-toxic, therapeutically inactivated, and / or liquid carriers commonly used in sublingual and / or subcutaneous formulations. 【0074】 One exemplary embodiment described herein can provide a method for enhancing skin wound healing in a subject by directly administering a topical solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement to a wound area. 【0075】 One exemplary embodiment described herein can provide a method for enhancing skin wound healing in a subject by directly applying a topical solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and taurine to a wound area. 【0076】 In one or more exemplary embodiments of the methods described herein, the wound may be a cranial ulcer, surgical wound, burn, trauma, and / or a wound that has been exposed to one or more chemicals and / or therapeutic radiation, etc. 【0077】 For topical administration to the skin, some exemplary compositions described herein may be prepared as sprays, ointments, creams, and / or gels. Pharmaceuticals for topical administration to the skin may be prepared by mixing such compositions with non-toxic, therapeutically inactivated, solid, and / or liquid carriers commonly used with pharmaceuticals administered topically. 【0078】 One exemplary embodiment described herein can provide a method for enhancing gingival and / or periodontal healing in a subject by directly administering a topical solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement to a wound area. 【0079】 One exemplary embodiment described herein can provide a method for enhancing gingival and / or periodontal healing in a subject by administering a topical solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and taurine. 【0080】 For topical administration to the oral mucosa, some exemplary compositions described herein may be prepared as sprays, ointments, gels, mouthwashes, and / or toothpastes, etc. Pharmaceuticals for topical administration to the mucosa may be prepared by mixing such compositions with non-toxic, therapeutically inactivated, and / or liquid carriers commonly used with pharmaceuticals administered topically. 【0081】 One exemplary embodiment described herein can provide a method for enhancing the repair of eye damage in a subject by directly administering a topical solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement to the eye. 【0082】 One exemplary embodiment described herein can provide a method for enhancing the repair of eye damage in a subject by administering a topical solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and taurine. 【0083】 In some exemplary embodiments of one or more methods described herein, the injury to the eye may be one or more of, for example, acute corneal abrasion, subconjunctival hemorrhage, and / or retinal detachment, and / or chronic eye diseases, for example, age-related macular degeneration, diabetic retinopathy, glaucoma, and / or dry eye. 【0084】 One exemplary embodiment can provide a method for enhancing bone and soft tissue healing in a subject by directly administering a solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement to a wound site. 【0085】 In one or more exemplary embodiments of the methods described herein, the injury can be acute and / or chronic, such as trauma, arthritis, tendinitis, one or more ligament tears, and / or nerve compression. 【0086】 For direct application to soft tissue, some exemplary compositions described herein can be prepared as sterile solutions for injection into one or more joints, tendons, muscles, and / or nerves, etc. Pharmaceuticals for injection can be prepared by mixing such compositions with non-toxic, therapeutically inactivated, and / or liquid carriers commonly used in such formulations, such as polyethylene glycol and hyaluronic acid. 【0087】 One exemplary embodiment described herein can provide a method for enhancing organ repair in a subject by administering a solution containing a therapeutically effective amount of one or more peptide-conjugated biotin molecules and at least one vitamin and / or one amino acid and / or one metabolic supplement intravenously, intramuscularly, subcutaneously and / or orally, etc. Pharmaceuticals for injection can be prepared by mixing such compositions with non-toxic, therapeutically inactivated, and / or liquid carriers commonly used in injectable pharmaceuticals. Pharmaceuticals for oral administration can be administered, for example, in the form of solutions, suspensions, and / or solid forms such as one or more tablets, capsules, and / or powders, etc. 【0088】 In one or more exemplary embodiments of the methods described herein, tissue damage can be caused by acute diseases, such as trauma, reduced blood flow, reduced oxygen supply, infectious pathogens, drugs, and / or toxins, to one or more organs, structures, and / or systems, such as the heart, brain, kidneys, liver, intestines, and / or limbs. 【0089】 In one or more exemplary embodiments of the methods described herein, tissue damage may be caused by chronic diseases, such as heart failure, chronic kidney disease, inflammatory bowel disease, diabetic complications, seizures, macular degeneration, and / or neurodegenerative diseases including Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, chronic traumatic encephalopathy, and / or Alzheimer's disease. 【0090】 In one or more exemplary embodiments of the methods described herein, a mixture of one or more peptide-conjugate biotin molecules may be administered to subjects having progressive multiple sclerosis, frontotemporal dementia, Parkinson's disease, and / or Alzheimer's disease. 【0091】 In one or more exemplary embodiments of the methods described herein, a mixture of one or more peptide-conjugate biotin molecules and thiamine may be administered to subjects having biotin-thiamine-responsive basal ganglia disease. 【0092】 In one or more exemplary embodiments of the methods described herein, a mixture of one or more peptide-conjugated biotin molecules may be administered to a subject having inflammatory bowel disease. [Examples] 【0093】 Short aromatic-cationic peptide sequences increase intracellular biotin uptake. Certain peptide sequences described herein may be water-soluble polypeptides consisting of 4 to 6 amino acids that can have a common aromatic-cationic motif, meaning that the peptide sequence can be, for example, [aromatic-cationic-aromatic-cationic] or [cationic-aromatic-cationic-aromatic]. 【0094】 Certain "short" peptide sequences described herein can be polypeptides consisting of a minimum of 4 amino acids and a maximum of 6 amino acids. 【0095】 Amino acids can occur naturally. Naturally occurring amino acids include the 20 most common amino acids commonly found in proteins: alanine (Ala), arginine (Arg), asparagine (Asn), aspartic acid (Asp), cysteine ​​(Cys), glutamine (Gln), glutamic acid (Glu), glycine (Gly), histidine (His), isoleucine (Ile), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), and valine (Val). Aromatic amino acids can include Phe, Tyr, and Trp. Cationic amino acids can include Lys, Arg, and His. Amino acids can be naturally occurring in a D-configuration. In certain peptides, the carboxyl terminus can be amidated. 【0096】 Amino acids can not exist in nature. These are typically amino acids that are not synthesized through the normal metabolic processes of living organisms and are not naturally present in proteins. These naturally occurring amino acids may include derivatives of naturally occurring amino acids in either the L- or D-configuration. 【0097】 One exemplary embodiment may provide the administration of the composition to a subject for therapeutic purposes. In one exemplary therapeutic application, the composition and / or drug may be administered to a subject suspected of or already suffering from such a disease and / or condition in an amount sufficient to cure, or at least partially cessate, the symptoms of the disease and / or condition, including its complications and intermediate pathological phenotypes at the onset of the disease and / or condition. 【0098】 In one exemplary embodiment, the treatment method includes the administration of a composition in combination with one or more activators. In one exemplary embodiment, the peptide administration is over a long period of time. 【0099】 In some exemplary embodiments, the peptide may be administered in combination with one or more platelet-lytic agents. In some exemplary embodiments, one or more platelet-lytic agents may be selected from the group consisting of tissue plasminogen activator, urokinase, prourokinase, streptokinase, acylated plasminogen, acylated plasmin, and acylated streptokinase-plasminogen complexes. 【0100】 In some exemplary embodiments, the treatment method may include the administration of a composition in combination with one or more antihypertensive drugs. In some exemplary embodiments, one or more antihypertensive drugs may include diuretics, adrenergic receptor blockers, calcium channel blockers, renin inhibitors, angiotensin-converting enzyme (ACE) inhibitors, angiotensin II receptor antagonists, aldosterone antagonists, vasodilators, and / or alpha-2 agonists. 【0101】 In some exemplary embodiments, the diuretic may include loop diuretics, thiazide diuretics, thiazide-like diuretics, and / or potassium-sparing diuretics. In some exemplary embodiments, the diuretic may include bumetanide, ethacrine, furosemide, torsemide, epitizide, hydrochlorothiazide, chlorothiazide, bendorflumesiazide, indapamide, chlorthalidone, metrazone, amiloride, triamterene, and / or spironolactone. 【0102】 In some exemplary embodiments, the adrenergic receptor blocker may include a beta-blocker, an alpha-blocker, or a mixed alpha-beta-blocker. In some exemplary embodiments, the adrenergic receptor blocker may include atenolol, metoprolol, nadolol, oxprenolol, pindolol, propranolol, timolol, doxazosin, phentolamine, indramine, phenoxybenzamine, prazosin, terazosin, trazoline, bucindolol, carvedilol, and / or labetalol. 【0103】 In some exemplary embodiments, the calcium channel blocker may include dihydropyridine and / or non-dihydropyridine. In some exemplary embodiments, the calcium channel blocker may include amlodipine, felodipine, isradipine, relcanidipine, nicardipine, nifedipine, nimodipine, nitrendipine, diltiazem, and / or verapamil. 【0104】 In one exemplary embodiment, the renin inhibitor may include aliskiren®. 【0105】 In one exemplary embodiment, the angiotensin-converting enzyme (ACE) inhibitor may include captopril, enalapril, fosinopril, lisinopril, perindopril, quinapril, ramipril, trandolapril, and / or benazepril. 【0106】 In one exemplary embodiment, the angiotensin II receptor antagonist may include irbesartan®. 【0107】 In one exemplary embodiment, the aldosterone antagonist may include eplerenone and / or spironolactone. 【0108】 In one exemplary embodiment, the vasodilator antagonist may include sodium nitroprusside and / or hydralazine. 【0109】 In one exemplary embodiment, the alpha-2 agonist antagonist may include clonidine, guanabenz, methyldopa, moxonidine, guanethidine, and / or reserpine. 【0110】 In some exemplary embodiments, any method known to those skilled in the art for contacting cells, organs, and / or tissues with the peptide may be used. Suitable methods may include in vitro, ex vivo, and / or in vivo methods. When used in vivo for therapeutic purposes, the composition may be administered to the subject in an effective amount (i.e., an amount that produces the desired therapeutic effect). The dose and administration regimen may depend on the extent of the subject's injury, the characteristics of the particular composition used, e.g., its therapeutic index, the subject, and / or the subject's medical history. 【0111】 The effective dose can be determined during preclinical and / or clinical trials by methods well known to physicians and / or clinicians. The effective dose of the peptide useful in the method can be administered to the mammal in need by any of several well-known methods for administering the drug. The peptide can be administered systemically and / or topically. 【0112】 Peptides may be formulated as pharmaceutically acceptable salts. Pharmaceutically acceptable salts can be derived from pharmaceutically acceptable inorganic or organic bases and from pharmaceutically acceptable inorganic or organic acids. If a peptide contains both a base portion, such as an amine, pyridine, or imidazole, and an acidic portion, such as a carboxylic acid or tetrazole, it can form a zwitterion, which is included in the term “salt” as used herein. Salts derived from pharmaceutically acceptable inorganic bases include salts of ammonia, calcium, copper, ferric iron, ferrous iron, lithium, magnesium, manganese digane, manganese monogyne, potassium, sodium, and zinc, as well as similar salts. Salts derived from pharmaceutically acceptable organic bases include salts of primary, secondary, and tertiary amines, including substituted amines, cyclic amines, naturally occurring amines and similar substances, such as arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydravamin, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purine, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, and similar substances. Salts derived from pharmaceutically acceptable inorganic acids include salts of boric acid, carbonic acid, hydrohalic acids (hydrobromic acid, hydrochloric acid, hydrofluoric acid, or hydroiodic acid), nitric acid, phosphoric acid, sulfamic acid, and sulfuric acid.Salts derived from pharmaceutically acceptable organic acids include aliphatic hydroxy acids (e.g., citric acid, gluconic acid, glycolic acid, lactic acid, lactobionic acid, malic acid, and tartaric acid), aliphatic monocarboxylic acids (e.g., acetic acid, butyric acid, formic acid, propionic acid, and trifluoroacetic acid), amino acids (e.g., aspartic acid and glutamic acid), aromatic carboxylic acids (e.g., benzoic acid, p-chlorobenzoic acid, diphenylacetic acid, centisic acid, hippuric acid, and triphenylacetic acid), and aromatic hydroxylic acids (e.g., o-hydroxybenzoic acid, p-hydroxybenzoic acid, 1-hydroxynaphthalene). This includes salts of xynafoleci (2-carboxylic acids and 3-hydroxynaphthalene-2-carboxylic acids), ascorbic acid, dicarboxylic acids (e.g., fumaric acid, maleic acid, oxalic acid, and succinic acid), glucuronic acid, mandelic acid, mucoic acid, nicotinic acid, orotic acid, pamoic acid, pantothenic acid, sulfonic acids (e.g., benzenesulfonic acid, camphorsulfonic acid, edisylic acid, ethanesulfonic acid, isethionic acid, methanesulfonic acid, naphthalenesulfonic acid, naphthalene-1,5-disulfonic acid, naphthalene-2,6-disulfonic acid, and p-toluenesulfonic acid), xynafoleci (hydroxynaphthalene-2-carboxylic acid and 3-hydroxynaphthalene-2-carboxylic acid), ascorbic acid, dicarboxylic acids (e.g., fumaric acid, maleic acid, oxalic acid, and succinic acid), xynafoleci (hydroxynaphthalene-2-carboxylic acid and 3-hydroxynaphthalene-2-carboxylic acid), and similar substances. In some embodiments, the salt is an acetate or trifluoroacetate. 【0113】 The compositions described herein, or pharmaceutically acceptable salts thereof such as acetates or trifluoroes, may be incorporated into pharmaceutical compositions for single or combined administration to subjects for the treatment and / or prevention of the disorders described herein. Such compositions may contain activators and pharmaceutically acceptable carriers, which may include one or more of physiological salines, solvents, dispersions, coatings, antimicrobial and antifungal agents, isotonic and absorption retarders, and the like, that are compatible with the administration of the pharmaceutical. Co-active compounds may also be incorporated into some exemplary compositions. 【0114】 Pharmaceutical compositions can be formulated to suit their intended route of administration. Exemplary routes of administration include parenteral (e.g., intravenous, intradermal, intraperitoneal, or subcutaneous), oral, sublingual, nasal, inhalation, transdermal (topical), intraocular, iontophoresis, and transmucosal administration. Solutions or suspensions used for parenteral, intradermal, or subcutaneous application may contain one or more of the following components: sterile diluents such as water for injection, physiological saline, fixative oil, polyethylene glycol, glycerin, propylene glycol, or other synthetic solvents; antimicrobial agents such as benzyl alcohol or methylparaben; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetic acid, citric acid, or phosphoric acid; and osmotic regulators such as sodium chloride or glucose. In some exemplary embodiments, the pH can be adjusted with an acid or base such as hydrochloric acid or sodium hydroxide. Parenteral formulations can be sealed in glass and / or plastic ampoules, disposable syringes, and / or multi-dose vials. For the convenience of the patient and / or the treating physician, the administered formulation may be provided in a kit containing any or all of the necessary supplies (e.g., vials of the drug, vials of diluents, syringes, and / or needles, etc.) for the course of treatment (e.g., a 7-day treatment). 【0115】 Pharmaceutical compositions suitable for injection may include sterile aqueous solutions (if water-soluble) and / or dispersions and / or sterile powders for the immediate preparation of sterile injections and / or dispersions. For intravenous administration, suitable carriers may include physiological saline, bacteriostatic water, Cremofor EL® (BASF, Parsippany, NJ), and / or phosphate-buffered saline (PBS). Compositions for parenteral administration may be sterile and / or liquid to facilitate the use of syringes. Certain exemplary compositions may be stable under manufacturing and / or storage conditions and / or resistant to contamination by microorganisms such as bacteria and / or fungi. 【0116】 The composition may contain a carrier, which may be a solvent or dispersion medium containing, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, and / or liquid polyethylene glycol, and the like), and suitable mixtures thereof. Appropriate fluidity can be maintained, for example, by the use of coatings such as lecithin, by maintaining the required particle size in the case of dispersion, and / or by the use of surfactants. Prevention of microbial action can be achieved by various antimicrobial and / or antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, and the like. Glutathione and / or other antioxidants may be included to prevent oxidation. Some exemplary compositions may contain isotonic agents, such as sugars and / or polyalcohols such as mannitol and sorbitol and / or sodium chloride. Long-term absorption of the injectable composition can be achieved by including absorption-delaying agents in the composition, such as aluminum monostearate and / or gelatin. 【0117】 Sterile injectable solutions can be prepared by incorporating the active compound in the required amount, along with one or a combination of the components listed above, in a suitable solvent, followed by filtration sterilization. Dispersions can be prepared by incorporating the active compound in a sterile medium containing a basic dispersion medium and other desired components, such as one or more of the components listed above. In the case of sterile powders for the preparation of sterile injectable solutions, exemplary preparation methods include vacuum drying and / or freeze-drying, which can yield a powder of the active component plus any additional desired components from a solution that has been previously sterile filtered. 【0118】 Oral compositions may include inactivating diluents and / or edible carriers. For oral therapeutic administration, the active compound may be incorporated with excipients and / or used in the form of tablets, lozenges, or capsules, such as gelatin capsules and / or powders soluble in diluents such as water. Oral compositions may be formulated using liquid carriers for use as mouthwashes. Pharmaceutically compatible binders and / or adjuvant materials may be included as part of the composition. Exemplary tablets, pills, capsules, lozenges, and similar products may contain any of the following ingredients or compounds of similar properties: binders such as microcrystalline cellulose, tragacanth gum, and / or gelatin; excipients such as starch and / or lactose; disintegrants such as alginic acid, Primogel, and / or corn starch; lubricants such as magnesium stearate and / or sterol; lubricants such as colloidal silicon dioxide; sweeteners such as sacrose and / or saccharin; or flavoring agents such as peppermint, methyl salicylate, and / or orange flavor; etc. 【0119】 For administration by inhalation, the compound can be delivered in the form of an aerosol spray from a pressurized container and / or dispenser containing a suitable pressurized gas, such as carbon dioxide, and / or from a nebulizer. 【0120】 Systemic administration of the therapeutic compounds described herein may be by transmucosal and / or transdermal means. For transmucosal or transdermal administration, a penetrating agent suitable for the permeable barrier may be used in the formulation. Such penetrating agents may include, for example, in transmucosal administration, cleansing agents, bile salts, and / or fusidic acid derivatives. Transmucosal administration can be achieved through the use of an intranasal spray. For transdermal administration, the active compound may be formulated in ointments, plasters, gels, and / or creams. In some exemplary embodiments, transdermal administration may be carried out by ion implantation, microneedling, and / or electroporation. 【0121】 Therapeutic proteins and / or peptides can be formulated into a carrier system. The carrier can be a colloidal system. The colloidal system can be a liposome and / or a phospholipid bilayer medium. In one exemplary embodiment, the therapeutic peptide can be encapsulated in a liposome while maintaining peptide integrity. The activator can be loaded into particles prepared from pharmaceutically acceptable components including, but not limited to, soluble, insoluble, permeable, impermeable, biodegradable, and / or gastric retention polymers and / or liposomes. Such particles can include nanoparticles, biodegradable nanoparticles, microparticles, biodegradable microparticles, nanospheres, biodegradable nanospheres, microspheres, biodegradable microspheres, capsules, emulsifiers, liposomes, micelles, and / or viral vector systems. 【0122】 The carrier can be a polymer, for example, a biodegradable and / or biocompatible polymer matrix. In one exemplary embodiment, the therapeutic peptide can be embedded in the polymer matrix while maintaining protein integrity. The polymer can be natural, such as polypeptides, proteins, or polysaccharides, or synthesized, such as poly-α-hydroxy acids. Examples include carriers made from collagen, fibronectin, elastin, cellulose acetate, nitrocellulose, polysaccharides, fibrin, gelatin, and combinations thereof. In one exemplary embodiment, the polymer can be polylactic acid (PLA) and / or copolymer lactic acid / glycolic acid (PGLA). The polymer matrix can be prepared and / or isolated in a wide variety of forms and / or sizes, including microspheres and nanospheres. Polymer formulations can provide long-lasting therapeutic effects. 【0123】 In one exemplary embodiment, the therapeutic compound may be formulated with a carrier that protects the therapeutic compound from rapid elimination from the body, such as a sustained-release formulation including an implantable tablet and a microcapsule-encapsulated delivery system. Biodegradable and / or biocompatible polymers such as ethylene vinyl acetate, polyanhydride, polyglycolic acid, collagen, polyorthoester, and / or polylactic acid can be used. Such formulations can be prepared using known techniques. Materials are also commercially available, for example, from Alza Corporation and Nova Pharmaceuticals. Liposome suspensions (containing liposomes directed to specific cells with monoclonal antibodies against cell-specific antigens) can be used as pharmaceutically acceptable carriers. 【0124】 Therapeutic compounds can be formulated to enhance intracellular delivery. For example, ribosome delivery systems and / or membrane-fused liposomes can be used to deliver proteins to cells in vivo and / or in vitro. 【0125】 The dosage, toxicity, and therapeutic efficacy of a therapeutic agent can be determined, for example, by standard pharmaceutical procedures in cell cultures and / or experimental animals to determine the LD50 (lethal dose for 50% of the population) and / or ED50 (therapeutably effective dose for 50% of the population). The dose ratio between the toxic effect and the therapeutic effect is called the "therapeutic index" and can be expressed as the ratio LD50 / ED50. Compounds exhibiting a high therapeutic index are sometimes preferred. While compounds exhibiting toxic side effects can be used, delivery systems can be carefully designed to direct such compounds to the affected tissue site to minimize potential damage to uninfected cells, thereby reducing side effects. 【0126】 Data obtained from cell culture assays and / or animal studies can be used to formulate dosage ranges for human use. Doses of such compounds can be within the range of circulating concentrations, including ED50, with little to no toxicity. Doses can vary within this range depending on the dosage form used and / or the route of administration utilized. For some exemplary compounds, the therapeutically effective dose can first be assessed from a cell culture assay. Once the dose is determined in cell culture, it can be formulated in animal models to achieve the circulating plasma concentration range, including IC50 (i.e., the concentration of the test compound that achieves the median inhibitory concentration of the symptoms). Using such information, useful doses in humans can be determined more accurately. Plasma levels can be measured, for example, by high-performance liquid chromatography. 【0127】 The effective amount of the composition sufficient to achieve a therapeutic or preventive effect can range from about 0.000001 mg per kg of body weight per day to about 10,000 mg per kg of body weight per day. The dosage range can be from about 0.0001 mg per kg of body weight per day to about 100 mg per kg of body weight per day. For example, the dosage can be about 1 mg / kg of body weight or about 10 mg / kg of body weight daily, every two days, or every three days, or in the range of about 1 to about 10 mg / kg of body weight per week, every two weeks, or every three weeks. In one exemplary embodiment, the single dose of the peptide can range from about 0.1 to about 10,000 micrograms per kg of body weight. In one exemplary embodiment, the concentration of the composition in the carrier can range from about 0.2 to about 2000 micrograms per milliliter delivered. The exemplary treatment regimen may require administration once daily or once weekly. In some therapeutic applications, relatively high doses at relatively short intervals may be required until disease progression is reduced and / or terminated, and / or until the subject shows partial or complete remission of the disease symptoms. A prophylactic regimen may then be administered to the patient. 【0128】 In one exemplary embodiment, the therapeutically effective amount of the composition is approximately 10% in the target tissue. -12 ~about 10 -6 Moles, for example, approximately 10 -7 This can be defined as the peptide concentration in moles. This concentration can be delivered by a systemic dose of approximately 0.01 to approximately 100 mg / kg or an equivalent dose based on body surface area. Therapeutic concentrations in target tissues can be maintained by optimizing the administration schedule, including single daily or weekly doses, but also including continuous administration (e.g., parenteral infusion and / or transdermal application). 【0129】 In some exemplary embodiments, the dosage of the composition can be provided at “low,” “intermediate,” or “high” dose levels. In some exemplary embodiments, the low dose can be provided in the range of about 0.01 to about 0.5 mg / kg / h, for example, about 0.0001 to about 0.1 mg / kg / h. In some exemplary embodiments, the intermediate dose can be provided in the range of about 0.001 to about 1.0 mg / kg / h, for example, about 0.01 to about 0.5 mg / kg / h. In some exemplary embodiments, the high dose can be provided in the range of about 0.005 to about 10 mg / kg / h, for example, about 0.01 to about 2 mg / kg / h. 【0130】 In some exemplary embodiments, certain factors, including but not limited to the severity of the disease or disorder, previous treatments, the subject's overall health and / or age, and / or other pre-existing conditions, may influence the dosage and / or timing required to effectively treat the subject. Furthermore, treatment of a subject with a therapeutically effective amount of the therapeutic composition described herein may include a single treatment or a series of treatments. 【0131】 Mammals treated according to a certain exemplary method may include, for example, livestock such as sheep, pigs, cattle, and horses; pet animals such as dogs and cats; and / or any mammals including laboratory animals such as rats, mice, and rabbits. In a certain exemplary embodiment, the mammal may be a human. 【0132】 One exemplary embodiment is a biologically active composition of a substance comprising a first D-biotin that is water-soluble, cell-permeable, and conjugated to lysine located at the C-terminus of a mitochondrial target peptide sequence, Mitochondrial target peptide sequences contain a minimum of 4 amino acids and a maximum of 6 amino acids; Mitochondrial target peptide sequences typically possess alternating aromatic-cationic motifs; The mitochondrial target peptide sequence is, D-Arg-L-(2'6'-dimethylTyr)-L-Lys-L-Phe-NH2; D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys-NH2; D-Trp-D-Arg-D-Trp-D-Lys-OH; L-Trp-L-Arg-L-Trp-L-Lys-NH2; D-Trp-D-Arg-D-Trp-D-Lys-NH2; and / or L-Trp-L-Arg-L-Trp-L-Lys-OH; Selected from; The second D-biotin is conjugated to the N-terminal α-amine of the mitochondrial target peptide sequence; composition, D-biotin-D-Arg-L-(2'6'-dimethylTyr)-L-Lys-L-Phe-NH2; and D-biotin-D-Trp-D-Arg-D-Trp-D-Lys-OH; Includes one or more of the following; The first D-biotin is conjugated to the ε-amine of lysine at the C-terminus of the mitochondrial target peptide sequence; composition, D-Trp-D-Arg-D-Trp-D-Lys(biothinyl)-OH; L-Trp-L-Arg-L-Trp-L-Lys(biothinyl)-NH2; and D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biothinyl)-NH2; Includes one or more of the following; The first D-biotin is conjugated to the ε-amine of lysine at the C-terminus of the mitochondrial target peptide sequence, and the second D-biotin is conjugated to the α-amine at the N-terminus; and / or composition, D-biotin-L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH2; and D-biotin-D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH 2; Includes one or more of the following; A composition is provided. 【0133】 One exemplary embodiment is a biologically active composition of a substance comprising a first D-biotin that is water-soluble, cell-permeable, and conjugated to lysine located at the N-terminus of a mitochondrial target peptide sequence, wherein the mitochondrial target peptide sequence is It contains a minimum of 4 amino acids and a maximum of 6 amino acids; It has a common alternating aromatic-cationic motif; A composition is provided. 【0134】 One exemplary embodiment is: One or more biologically active, water-soluble, cell-permeable, mitochondrial-targeting compounds selected from the biotinylated polypeptide group; One or more biologically active, water-soluble, cell-permeable, and pharmaceutically acceptable carriers for each of the mitochondrial target compounds; Vitamin B1 (thiamine); Vitamin B2 (riboflavin); Vitamin B3 (nicotinic acid, niacinamide); Vitamin B5 (pantothenic acid); Vitamin B6 (pyridoxine); Vitamin B7 (biotin); Vitamin B9 (folic acid); Vitamin B12 (cyanocobalamin); and Vitamin C (ascorbic acid) One or more vitamins selected from, Pyruvate; Carnitine; Acetylcarnitine; Creatine; α-Ketoglutaric acid; Alpha-lipoic acid; Coenzyme Q Nicotinamide riboside; and Nicotinamide mononucleotide; One or more metabolic supplements selected from; and / or Leucine; Isoleucine; Valin; glutamine; Serine; Arginine; Methionine; Tryptophan; glycine; Trimethylglycine Methyl β-hydroxy-β-butyrate; and Taurine One or more amino acids selected from A therapeutically effective formulation containing, Each of the biotinylated polypeptide groups exhibits biological activity, water solubility, cell permeability, and mitochondrial targeting properties. Multiple amino acids arranged in a common alternating aromatic-cationic motif; A minimum of 4 amino acids and a maximum of 6 amino acids; The first D-biotin is conjugated to the lysine located at the C-terminus or N-terminus of the biotinylated polypeptide. Defined by, Biotinylated polypeptides D-biotin-D-Arg-L-(2'6'-dimethylTyr)-L-Lys-L-Phe-NH2; D-biotin-D-Trp-D-Arg-D-Trp-D-Lys-OH; D-Trp-D-Arg-D-Trp-D-Lys(biothinyl)-OH; L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH2; D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biothinyl)-NH2; D-biotin-L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH2; and D-biotin-D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH2 Consists of, The present invention provides a composition of substances containing therapeutically effective formulations. 【0135】 Table 1 identifies an exemplary short, water-soluble, alternating aromatic-cationic peptide sequence. 【0136】 [Table 1] 【0137】 An alternating aromatic-cationic peptide sequence useful in an exemplary method described herein can be chemically synthesized by any of the methods described in the following U.S. patent documents, each of which, by reference, is incorporated herein by reference in whole and / or any part relating to the synthesis of the peptide and / or the use of the synthesized peptide: U.S. Patent No. 4749742; U.S. Patent No. 5026773; U.S. Patent No. 7576061; U.S. Patent No. 9388212; U.S. Patent No. 9695214; U.S. Patent No. 10125163; U.S. Patent Application Publication No. 2019 / 0202861; U.S. Patent Application Publication No. 2019 / 0015521; and U.S. Patent Application Publication No. 2012 / 0149868. 【0138】 A representative example of an alternating aromatic-cationic peptide sequence, listed in Table 1, is water-soluble. 【0139】 A representative alternating aromatic-cationic peptide sequence, shown in Table 1, is water-soluble but can permeate the cell membrane. 【0140】 A certain exemplary alternating aromatic-cationic peptide sequence, listed in Table 1, is water-soluble but can permeate the mitochondrial outer membrane. 【0141】 Biotin (Figure 1A) can be conjugated at its N-terminal α-amine to an alternating aromatic-cationic peptide sequence (Figure 1B) or at its C-terminus to the ε-amino group of lysine (Figure 1C). 【0142】 In one exemplary embodiment, biotin can be conjugated to a polypeptide via chemical synthesis using one or more methods described in any of the following U.S. patent applications, each of which is incorporated herein by reference in whole and for the incitement of such methods: U.S. Patent No. 5,391,711; U.S. Patent No. 5,416,016; and U.S. Patent Application Publication No. 2006 / 0149035. 【0143】 In one exemplary embodiment, the α-amino N-terminus of a peptide can be preferentially labeled with biotin. In one exemplary embodiment, biotination can be easily achieved by activating the carboxyl group of biotin to react with the free amino group of the peptide. In one exemplary embodiment, biotinylating reagents such as D-biotin-N-hydroxy-succinimide ester or biotinyl-p-nitrophenyl ester can be used. The activated ester can incorporate the biotin residue into the desired molecule if it reacts with the amino group under mild conditions. In one exemplary embodiment, a polymer can be biotinylated using D-biotin-N-hydroxy-succinimide ester and / or an exogenous molecule can be biotinylated using biotinyl-ε-nitrophenyl ester as the biotinylated agent. Other reagents such as D-biotinyl-ε-aminocaproic acid N-hydroxy-succinimide ester, in which ε-aminocaproic acid acts as a spacer link to reduce steric hindrance, can also be used in one exemplary embodiment. 【0144】 In one exemplary embodiment, biocitin can be formed by biotinylation with ε-aminolysine. In one exemplary embodiment, a biocitin-containing peptide can be achieved via solid-phase synthesis using Fmoc-Lys(ε-biotinyl) at the C-terminal position, which yields Lys(biotinyl) or biocitin after acid cleavage of the resin. For example, to produce biocitin, one exemplary embodiment may utilize one or more methods described in U.S. Patent No. 2,720,527, which is incorporated herein by reference in whole and for the instructive use of such methods. 【0145】 An example of a peptide-conjugated biotin molecule and its corresponding mitochondrial target peptide sequence are listed in Table 2. 【0146】 [Table 2] 【0147】 Intracellular uptake and localization of peptide-conjugated biotin molecules were assessed in HK-2 human renal epithelial cells and ARPE-19 human retinal pigment epithelial cells (ATCC, Manassas, VA). HK-2 cells were cultured in Dulbecco's modified Eagle medium (DMEM) containing 1 g / L glucose, 10% fetal bovine serum (FBS), 100 units / ml penicillin, and 100 μg / ml streptomycin. ARPE-19 cells were cultured in DMEM / F12 medium containing 1 g / L glucose, 10% fetal bovine serum (FBS), 100 units / ml penicillin, and 100 μg / ml streptomycin. Cells were incubated in a humidified incubator at 37°C with 5% CO2. HK-2 and ARPE-19 cells were initially densified at 5 × 10⁶ cells in a 35 mm dish. 4 Cells were seeded. FBS was removed from the culture medium for 3 days to deplete endogenous biotin in the cells. The cells were then incubated in serum-free medium containing biotin or peptide-conjugated biotin molecules for 12 hours (HK-2 cells) or 1 hour (ARPE-19 cells). All compounds were used at 1 μM. 【0148】 Biotin uptake was determined using streptavidin binding. Streptavidin has high affinity for biotin. By using Alexa Fluor 594 conjugated streptavidin, biotin uptake can be visualized using a fluorescence microscope. Cells were fixed with 4% PFA for 10 minutes at RT, permeable in 0.1% triton X-100 / PBS for 10 minutes at RT, and incubated with 3.2 μg / ml streptavidin-Alexa Fluor 594 (Jackson ImmunoResearch, West Grove, PA) and 5 μg / ml Hoechst 33342 (Novus Biologicals, Centennial, CO) for 30 minutes at RT. Hoechst 33342 is a fluorescent stain for labeling DNA and is used for nuclear staining. After adding a live cell image buffer, Alexa-594 fluorescence (Ex / Em = 591 / 614 nm) and Hoechst fluorescence (Ex / Em = 490 / 461 nm) were observed using a Nikon Eclipse Ti2 fluorescence microscope (100× oil objective lens). Ten random fields were obtained from each sample, and streptavidin fluorescence was quantified using NIS-Elements imaging software (Nikon). This was then normalized to Hoechst fluorescence to explain the number of cells per field. 【0149】 Figure 2 shows representative microscopic images of HK-2 human renal epithelial cells after 12-hour incubation with 1 μM free biotin or with peptide-conjugated biotin molecules selected from Table 2. Intracellular biotin is visualized with streptavidin-AlexaFluor594 (red fluorescence). The nucleus is visualized with Hoechst 33342 dye (blue fluorescence) (1000× magnification). Minimal intracellular streptavidin fluorescence was observed when HK-2 cells were incubated with 1 μM biotin alone. In contrast, all peptide-conjugated biotin compounds (SPN05, SPN08, SPN09, SPN11, and SPN12) showed strong red fluorescence, indicating significant intracellular biotin uptake. 【0150】 Figure 3 is a graph quantifying biotin uptake in HK-2 human renal epithelial cells after 12-hour incubation with 1 μM biotin or peptide-conjugated biotin molecules selected from Table 2. Peptide-conjugated biotin molecules delivered significantly higher intracellular biotin content compared to free biotin in HK-2 cells. Total streptavidin fluorescence, normalized to cell number (Hoechst fluorescence), was calculated from a 10-point random field and averaged for each treatment. All data were then normalized to treatments using free biotin, with free biotin arbitrarily set to 1.0. All peptide-conjugated biotin molecules resulted in significantly higher intracellular biotin compared to free biotin (*P<0.05, ***P<0.001). Uptake of peptide-conjugated biotin molecules was 2–2.5 times that of free biotin. 【0151】 Figure 4 shows representative microscopic images of ARPE-19 human retinal pigment epithelial cells after 1 hour incubation with 1 μM biotin or peptide-conjugate biotin molecules selected from Table 2. Intracellular biotin is visualized with streptavidin-AlexaFluor594 (red fluorescence). The nucleus is visualized with Hoechst 33342 dye (blue fluorescence) (1000× magnification). All peptide-conjugate biotin compounds (SPN08, SPN09, SPN11, SPN12, SPN15, and SPN16) showed strong red staining with a perinuclear distribution. 【0152】 Figure 5 is a graph showing biotin uptake in ARPE-19 human retinal pigment epithelial cells after 1 hour incubation with 1 μM biotin or peptide-conjugated biotin molecules selected from Table 2. Peptide-conjugated biotin molecules delivered significantly higher intracellular biotin content compared to free biotin in ARPE-19 cells. The ratio of streptavidin fluorescence to Hoechst fluorescence was calculated from a random field of 10 and averaged for each treatment. All data were then normalized to treatments using free biotin, with the ratio arbitrarily set to 1.0. All peptide-conjugated biotin molecules resulted in significantly higher intracellular biotin staining compared to free biotin (***P<0.001). Uptake of peptide-conjugated biotin molecules was 1.5–2 times that of free biotin. 【0153】 These peptides enter cells by simple diffusion and, once inside the cell, are found only in mitochondria. This is confirmed in Figures 2 and 4 for the identified SPN peptides. The distribution pattern shown for these peptides (a fibrous network starting from the perinucleus) is very different in mitochondria. Note that the nucleus is not stained, and staining is not present everywhere within the cell. This significantly reduces the chance of side effects from compounds acting on other organelles. 【0154】 These results demonstrate that the intracellular uptake of biotin can be significantly increased when biotin is conjugated to exemplary short, water-soluble, aromatic-cationic peptide sequences. Microscopic images in Figures 2 and 4 show that the conjugation of biotin with SS-31, SPN02, SPN07, and SPN10 significantly enhances biotin uptake into two different cell lines (renal cells and retinal epithelial cells). All four of these short peptides follow an alternating aromatic-cationic motif, with SS-31 and SPN02 having a "cationic-aromatic-cationic-aromatic" sequence order, and SPN07 and SPN10 having an "aromatic-cationic-aromatic-cationic" sequence order. All exemplary peptides are water-soluble. SPN02 demonstrates that a pentapeptide can function as well as a tetrapeptide and is water-soluble. SS-31 shows that a naturally occurring amino acid (2'6'-dimethylTyr) can be used as a substitute for a naturally occurring amino acid. These peptides also support the use of Tyr, Phe, or Trp as aromatic amino acids, and Arg or Lys as cationic amino acids. Exemplary examples demonstrate that amino acids can be in D- or L- configurations, and that C-terminal amidation does not affect mitochondrial delivery at all, but can improve peptide stability against carboxypeptidase degradation in vivo. 【0155】 Common peptides consisting of 4-6 amino acids, having alternating aromatic-cationic motifs made from naturally occurring or non-naturally occurring amino acids, in a D- or L- configuration, and with or without C-terminal amidation, can act as water-soluble delivery vectors to enhance intracellular biotin uptake in mammalian cells. [Examples] 【0156】 Short aromatic-cationic peptide sequences selectively deliver biotin to the inner mitochondrial membrane. A certain exemplary alternating aromatic-cationic peptide sequence, listed in Table 1, can selectively target and localize to the inner mitochondrial membrane. 【0157】 The perinuclear distribution patterns for all peptide-conjugate biotin molecules (Figures 2 and 4) suggest that this molecule is localized in the mitochondrial filamentous network. 【0158】 To demonstrate the mitochondrial localization of peptide-conjugated biotin molecules, ARPE-19 human retinal pigment epithelial cells were incubated in serum-free DMEM / F12 medium with 1 μM SPN05, SPN12, or SPN15 for 2 hours. The cells were fixed with 4% PFA for 10 minutes at RT, and then permeable with 0.1% Triton X-100 for 10 minutes at RT. The cells were blocked with 2% BSA (bovine serum albumin) for 30 minutes at RT, and then incubated with a primary antibody against cytochrome c oxidase subunit 4 (COX4), a major protein complex expressed on the inner mitochondrial membrane. Rabbit polyclonal COX4 antibody (Invitrogen PA5-29992, Waltham, MA) was used at a 1:500 dilution in 2% BSA overnight at 4°C. Next, the cells were incubated for 30 minutes at RT with a secondary antibody (goat anti-rabbit Alexa Fluor 488, 1:500 dilution, Invitrogen A-11008), streptavidin-Alexa Fluor 594 (Jackson ImmunoResearch, West Grove, PA), and 5 μg / ml Hoechst 33342 (Novus Biologicals, Centennial, CO). Live cell imaging buffer was added, and the fluorescence of Alexa Fluor-594, Alexa Fluor-488, and Hoechst was observed using a Nikon Eclipse Ti2 fluorescence microscope (100× oil objective lens). 【0159】 Figure 6 shows the co-localization of peptide conjugate biotin molecules and cytochrome c oxidase. The upper panel shows representative microscopic images of SPN15 (stained red), cytochrome c oxidase (COX) (stained green), and combined images (yellow / orange). The lower panel shows images of SPN05 and SPN12 combined with COX staining (yellow / orange). All images are shown at 600× magnification. 【0160】 These results demonstrate that short aromatic-cationic peptide sequences can act as delivery vectors to direct biotin to mitochondria. COX4 is a subunit of cytochrome c oxidase, the terminal complex (complex IV) of the electron transport chain. The COX complex is a major regulatory site for oxidative phosphorylation on the inner mitochondrial membrane. Colocalization of biotin staining and COX staining from SPN05, SPN12, and SPN15 demonstrates that these short aromatic-cationic peptide sequences can act as mitochondrial target sequences to deliver biotin to the inner mitochondrial membrane. [Examples] 【0161】 Peptide-conjugated biotin molecules can promote cell proliferation under prolonged serum starvation. Serum removal can cause a decrease in cellular ATP, cell cycle arrest, and / or apoptosis. Serum deficiency can inhibit the ability of cultured cells to proliferate. Incubation with certain exemplary peptide conjugate biotin molecules can promote cell viability under serum-free conditions. 【0162】 In the first model, HK-2 cells were cultured in serum-free DMEM alone or in serum-free DMEM containing 10 nM SPN11 or SPN12 in 96-well plates for 11 days. The culture medium was changed every 3 days. 【0163】 Cell viability was measured using the Alamar Blue cell proliferation assay (Bio-Rad) according to the manufacturer's protocol. Alamar Blue measures the reducing power of viable cells using resazurin. Weakly fluorescent resazurin is reduced to highly fluorescent resorphine when reduced by cellular metabolism. Briefly, Alamar Blue reagent (10 μl) was added to the culture medium and incubated at 37°C for 1 hour. Fluorescence from resorphine (Ex / Em 530 / 590 nm) was detected using a microplate reader (SpectraMax iD3, Molecular Devices). 【0164】 Figure 7 is a graph showing the cell viability of HK-2 cells after 11 days of serum starvation. Treatment with as little as 10 nM of SPN11 or SPN12 significantly improved cell viability by 35% in the absence of serum (**P<0.005). 【0165】 In the second model, ARPE-19 cells were grown for 30 days in serum-free DMEM, either in the absence or in the presence of peptide-conjugated biotin molecules. Serum-free medium supplemented with 10 nM SPN12, SPN15, or SPN16 compounds (all at 10 nM) was replaced every 3 days. The culture medium was replaced every 3 days. Figure 8 is a representative microscopic image showing that all three SPN compounds increased cell number after prolonged serum deprivation. 【0166】 Figure 9 shows that treatment with SPN12, SPN15, and SPN16 significantly doubled cell viability after 30 days of serum deprivation compared to the control. Data represent the mean ± SEM from six samples for each treatment (**P<0.005; ***P<0.001, compared to the control). 【0167】 These results demonstrate that the addition of as little as 10 nM of peptide-conjugated biotin molecules can significantly double cell viability under serum-free conditions. These results also demonstrate that exemplary mitochondrial-targeting biotin molecules are biologically active (i.e., these biotin molecules alter cell biology) and protect cell viability in at least two different mammalian cell lines. [Examples] 【0168】 Peptide-conjugated biotin molecules increased cellular ATP content in serum-free and nutrient-deficient cultures. Serum depletion can cause a decrease in cellular ATP. Certain exemplary peptide-conjugated biotin molecules can increase cellular ATP levels under serum-deficient and / or nutrient-deficient conditions. 【0169】 Human renal epithelial cells (HK-2) were cultured in DMEM containing 1 g / L glucose, 10% fetal bovine serum (FBS), 100 units / ml penicillin, and 100 μg / ml streptomycin. The cells were incubated in a humidified incubator with 5% CO2 at 37°C. HK-2 cells were initially cultured in 96-well plates at a density of 5 × 10⁴. 3 Cells were seeded. FBS was removed from the culture medium, and the cells were incubated for 7 days in serum-free DMEM alone (control group) or serum-free DMEM containing 10 nM biotin or peptide-conjugated biotin molecules. All procedures were performed in N=6 for each experiment. The culture medium was changed every 3 days. 【0170】 Human retinal pigment epithelial cells (ARPE-19) were cultured in DMEM / F12 medium containing 1 g / L glucose, 10% fetal bovine serum (FBS), 100 units / ml penicillin, and 100 μg / ml streptomycin. The cells were incubated in a humidified incubator with 5% CO2 at 37°C. ARPE-19 cells were initially cultured in 96-well culture plates at a density of 5 × 10⁴. 3Cells were seeded. FBS was removed from the culture medium, and the cells were incubated for 7 days in serum-free DMEM / F12 alone (control group) or in serum-free DMEM / F12 containing 10 nM biotin or different peptide conjugate biotin molecules. All procedures were performed with N=5-6 in each experiment. The culture medium was changed every 3 days. 【0171】 ATP levels were measured using the ApoSENSOR ATP Bioluminescence Assay Kit (BioVision) according to the manufacturer's protocol. This kit utilizes luciferase, which catalyzes the formation of light from ATP and luciferin. Briefly, cells were treated with 100 μl of nuclear release buffer with gentle shaking for 5 minutes at RT. 10 μl of ATP monitoring enzyme was added to the cell lysate, and luminescence was measured using a microplate reader (SpectraMax iD3, Molecular Devices). ATP levels were normalized to a serum-free DMEM control group (arbitrarily set to 1.0). 【0172】 After 7 days under serum-free conditions, all peptide-conjugated biotin molecules produced higher ATP concentrations compared to free biotin in HK-2 cells (Figure 10). The data shown are mean ± SEM from 6 samples per treatment. SPN08, SPN09, SPN11, SPN12, SPN15, and SPN16 increased ATP levels by 35%–100% under serum-free conditions compared to free biotin (**P<0.005; ***P<0.001). 【0173】 Peptide-conjugated biotin molecules significantly increased ATP concentrations in ARPE-19 cells after 7 days of serum starvation. The data shown are mean ± SEM from 6 samples per treatment. Peptide-conjugated biotin molecules increased ATP levels by 60–75% compared to free biotin (***P<0.001) (Figure 11). 【0174】 The second model uses an extreme starvation model in which HK-2 cells are cultured in serum-free 5% DMEM in PBS (phosphate-buffered saline) in a 96-well plate for 3 days. The cells were treated with 10 nM free biotin or peptide-conjugated biotin molecules. All treatments were performed with N=6 in each experiment. 【0175】 All peptide-conjugated biotin molecules resulted in significantly higher ATP concentrations compared to free biotin under extreme starvation. The data shown are mean ± SEM from 6 samples per treatment. (*P<0.05; **P<0.01; ***P<0.001), SPN11 and SPN12 doubled ATP content (Figure 12). 【0176】 These results demonstrate that exemplary mitochondrial target biotin molecules are biologically active and promote mitochondrial ATP synthesis under serum- and nutrient-deficient conditions in at least two different mammalian cell lines. [Examples] 【0177】 Peptide-conjugated biotin molecules can restore mitochondrial potential and prevent mitochondrial fragmentation caused by serum starvation. Serum deficiency can induce mitochondrial depolarization, preceded by a decrease in ATP synthesis. A certain exemplary peptide-conjugated biotin molecule can restore mitochondrial potential in cells cultured in serum-free medium. 【0178】 Electron transfer along the electron transport chain on the inner mitochondrial membrane causes proton pumping from the mitochondrial matrix to the intermembrane space. This creates an electrical potential across the inner mitochondrial membrane, and the proton gradient drives ATP synthase (complex V) to produce ATP from ADP. When serum or nutrients are drawn up, the mitochondrial potential decreases, and ATP production decreases. 【0179】 Human retinal pigment epithelial cells (ARPE-19) were cultured in DMEM / F12 medium containing 1 g / L glucose, 10% FBS, 100 units / ml penicillin, and 100 μg / ml streptomycin. The cells were incubated in a humidified incubator with 5% CO2 at 37 °C. ARPE-19 cells were seeded at an initial density of 5×10 4 cells on a 35 mm glass plate. FBS was removed from the culture medium for 3 days to deplete endogenous biotin. Next, the cells were incubated for 2 hours in serum-free DMEM / F12 alone (control group) or DMEM / F12 containing 1 μM SPN12 or SPN15. 【0180】 To determine the mitochondrial potential, live cells were incubated in DMEM without phenol red together with 5 nM tetramethylrhodamine methyl ester (TMRM, #70017, Biotium, Fremont, CA). TMRM is a potential-dependent fluorescent dye (Ex / Em = 548 / 573). TMRM accumulates in negatively charged polarized mitochondria and can be detected as red fluorescence. Total mitochondria were imaged with a mitochondria potential-independent fluorescent dye (100 nM MitoView Green; Ex / Em = 490 / 523; #70054, Biotium, Fremont CA). Hoechst 33342 (10 μg / ml, Novus Biologicals, Centennial, CO) was added to stain the nuclei. Live cells were covered with phosphate buffer and imaged using a Nikon Eclipse Ti fluorescence microscope using a 60× water objective lens. 【0181】 Figure 13 (upper panel) shows representative fluorescence microscopy images of ARPE-19 cells cultured for 3 days in DMEM / F12 with 10% FBS. Figure 13 (lower panel) shows representative images of ARPE-19 cells cultured for 3 days in serum-free DMEM / F12. TMRM (red) indicates the dramatic disappearance of mitochondrial potential under serum-free conditions compared to serum controls. Mitogreen (green) staining also shows that serum-free conditions fragment mitochondria, causing them to aggregate into perinuclear patterns without a distinct filamentous network. When the images are combined, mitochondrial depolarization can be seen in any serum-free cell. 【0182】 Figure 14 shows representative fluorescence microscopy images of ARPE-19 cells cultured for 3 days after 2-hour incubation in 10% FBS (serum control) or in serum-free medium alone or with 1 μM SPN12 or SPN15. The images shown are combined images of TMRM (red) and MitoGreen (green). Two-hour incubation with SPN compounds restored mitochondrial potential in all cells and was sufficient to restore mitochondrial filamentous network in ARPE-19 cells after 3 days of serum starvation. 【0183】 These results indicate that mitochondria are completely depolarized after 3 days of serum starvation, and the mitochondrial network in cells is fragmented. Peptide-conjugated biotin molecules (SPN12 and SPN15) can rescue mitochondrial potentials within 2 hours of serum starvation. These results confirm that biotin conjugated to aromatic-cationic peptide sequences is directed towards the inner mitochondrial membrane and is biologically active in protecting potentials across the inner mitochondrial membrane. [Examples] 【0184】 Peptide-conjugated biotin molecules are more effective than free biotin in wound healing. Certain exemplary peptide-conjugated biotin molecules can accelerate wound healing in cell cultures. The in vitro scratch assay is a simple and well-developed method for measuring cell proliferation and migration on a "wound area" in vitro. This assay involves creating a "scratch" in a cell monolayer membrane with a pipette tip, and then examining the rate at which cell proliferation and migration close the scratch. 【0185】 HK-2 cells were cultured in DMEM (1 g / L glucose) containing 10% FBS, 100 units / ml penicillin, and 100 μg / ml streptomycin. One day prior to the experiment, 3 × 10⁶ cells per well were cultured in a 6-well plate in DMEM at 37°C in a humidified incubator with 5% CO₂. 5 Cells were seeded. To mimic the wound microenvironment in vitro, serum-free medium was used in a scratch assay. On the day of the scratch assay, the medium was replaced with serum-free DMEM, and a linear scratch was made across the cell monolayer using a p1000 pipette tip. Cells were washed to remove cell debris and replaced with DMEM alone (control) or DMEM containing 10 nM biotin or peptide-conjugated biotin molecules. 【0186】 Scratch areas were examined immediately (day 0) using a Nikon Eclipse Ti2 fluorescence microscope. Six different fields were captured for each sample using a 4× symmetric lens. Scratch area (cell-free zone) was calculated using ImageJ software, an open-source Java image processing program suggested by NIH imaging (National Institutes of Health). Scratch areas were re-examined 24 hours later (day 1), and the areas were normalized to the areas determined on day 0 for the same sample. All results were calculated as change from untreated control and averaged across the six fields. 【0187】 Figure 15 is a representative microscopic image showing that treatment with 10 nM SPN11 or SPN12 significantly accelerated scratch closure in HK-2 cells 24 hours after mechanical scratching. The scratch areas determined at 24 hours are normalized to the areas determined immediately after scratching (day 0) for each treatment group. Biotin had only a slight effect on reducing the scratch area, and the scratch area was almost completely closed within 24 hours with SPN11 and SPN12. 【0188】 Figure 16 summarizes the effects of biotin, SPN11, and SPN12 on reducing scratch areas 24 hours after mechanical scratching in HK-2 cells. All treatments significantly accelerated the closure of scratch areas (***P<0.001, compared to control). In reducing scratch areas, biotin conjugate peptides (SPN11 and SPN12) were twice as effective as biotin (P<0.001). 【0189】 Figure 17 shows that other peptide conjugate biotin molecules, SPN15 and SPN16, also significantly accelerated scratch region closure after 24 hours in another experiment using HK-2 cells (***P<0.001), and their effect was three times greater than that of free biotin (P<0.001). 【0190】 Peptide-conjugated biotin molecules (SPN12, SPN15, and SPN16) accelerated wound healing in ARPE-19 cells (Figure 18). Figure 19 is a graph summarizing the effect of peptide-conjugated biotin molecules on scratch regions (**P<0.01; ***P<0.001). Peptide-conjugated biotin molecules were significantly more effective than free biotin (1.5 to 2 times) (P<0.001). 【0191】 These results demonstrate that peptide-conjugated biotin molecules are not only biologically active but also superior to free biotin in promoting wound healing in vitro. [Examples] 【0192】 Peptide-conjugated biotin molecules can promote cell proliferation in wound areas. Epithelial regeneration in a wound site requires cell proliferation at the wound edge, which is often compromised in most wounds due to a lack of blood flow that can deliver nutrients and / or growth factors necessary for cell proliferation. Cell proliferation at the wound edge can be monitored by proliferating cell nuclear antigen (PCNA) staining. To mimic the wound microenvironment in vitro, serum-free medium was used for scratch assays in cell cultures. 【0193】 Proliferating cells were identified by immunostaining with antibodies against proliferating cell nuclear antigen (PCNA) according to the standard procedure provided by IHC World. PCNA staining was performed using cells from the same plate after determining the scratch area. HK-2 cells were fixed in ethanol:methanol (1:1) at -20°C for 30 minutes, permeabilized with 0.1% Triton X-100 / PBS for 10 minutes at RT, blocked with 2% goat serum for 30 minutes at RT, and incubated with primary mouse anti-PCNA antibody (Dako Agilent, Santa Clara, CA) and secondary goat anti-mouse IgG-BI for 30 minutes at RT. The cells were then incubated with streptavidin-Alexa Fluor 594 (Jackson ImmunoResearch, West Grove, PA) and Hoechst (Novus Biologicals, Centennial, CO) and imaged with a Nikon Eclipse Ti2 fluorescence microscope (20× objective lens). All cell nuclei stain blue with Hoechst, but the nuclei of proliferating cells stain red. The number of proliferating cells per field was quantified by the intensity of red nucleus staining and normalized to the intensity of blue nucleus staining (determined using Nikon NIS-Element Imaging Software). 【0194】 Figure 20 shows representative microscopic images (40x magnification) of a monolayer of HK-2 human renal epithelial cells 24 hours after mechanical scratching in serum-free medium alone (control) or in serum-free medium containing an exemplary SPA compound (10 nM). Cells positive for the proliferating cell nuclear antigen are shown in red. All other nuclei are shown in blue. Compared to free biotin, SPN11 and SPN12 increased the number of proliferating cells at the edge of the scratch. 【0195】 Figure 21 is a graph summarizing the number of proliferating cells in a monolayer of HK-2 human renal epithelial cells 24 hours after mechanical scratching, in serum-free medium alone (control) or in serum-free medium supplemented with 10 nM biotin or peptide-conjugated biotin molecules. The number of proliferating cells was normalized to all nuclei. The effect of biotin was small and did not reach statistical significance. Incubation with SPN11 and SPN12 doubled the number of proliferating cells at the scratch edge compared to control or biotin (***P<0.001). 【0196】 Figure 22 shows representative microscopic images (40x magnification) of a monolayer of ARPE-19 human retinal pigment epithelial cells 24 hours after mechanical scratching in serum-free medium alone (control) or in serum-free medium supplemented with 10 nM free biotin or peptide-conjugated biotin molecules. Cells positive for the proliferating cell nuclear antigen are shown in red. All other nuclei are shown in blue. Incubation with SPN08, SPN09, SPN11, or SPN12 increased the number of proliferating cells at the edge of the scratch, and 10 nM biotin had no effect on cell proliferation. 【0197】 Figure 23 is a graph summarizing the number of proliferating cells in a monolayer of ARPE-19 human retinal pigment epithelial cells 24 hours after mechanical scratching in serum-free medium alone (control) or in serum-free medium containing 10 nM biotin or peptide-conjugated biotin molecules. The number of proliferating cells was normalized to all nuclei. Incubation with SPN09, SPN11, SPN12, SPN15, and SPN16 each resulted in a significant increase of 35–100% in the percentage of proliferating cells at the edge of the scratch, but biotin itself had no effect (*P<0.05; **P<0.01; ***P<0.001, compared to control). 【0198】 These results indicate that peptide-conjugated biotin molecules are more effective than biotin alone in promoting cell proliferation in wound conditions, likely due to increased biotin delivery to mitochondria. [Examples] 【0199】 Peptide-conjugated biotin molecules protect mitochondrial potentials in cells at wound sites. Rapidly dividing cells may require ATP to support the synthesis of essential building blocks. Tissue damage can cause rapid mitochondrial depolarization, impairing ATP production and / or leading to cell death. The loss of ATP can further impair cell proliferation and / or tissue repair. Mitochondrial potential can be monitored in cells at the edge of a scratch region using the cell-permeable voltage-gated dye TMRM (tetramethylrhodamine methyl ester). TMRM can accumulate in negatively polarized mitochondria and can be detected as red fluorescence. 【0200】 To mimic the wound microenvironment in vitro, serum-free medium was used in the scratch assay. On the day of the scratch assay, the medium was replaced with serum-free DMEM, and a linear scratch was made across the HK-2 cell monolayer using a p1000 pipette tip. The cells were washed to remove cell debris and replaced with DMEM alone (control) or DMEM supplemented with 10 nM free biotin or peptide-conjugated biotin molecules for 24 hours. Next, the HK-2 cells were incubated with 5 nM TMRM in DMEM without phenol red for 30 minutes. The cells were then covered with live cell imaging buffer and imaged using a Nikon Eclipse Ti2 fluorescence microscope (20× target lens). 【0201】 Figure 24 shows representative microscopic images (200× magnification) of mitochondrial potentials in HK-2 cells 24 hours after mechanical scratching, in serum-free medium alone (control) or in serum-free medium containing 10 nM biotin, SPN11, or SPN12. Mitochondrial potentials were detected using TMRM, which stains red, and nuclear staining blue, which is detected using Hoechst. Under control conditions, there were almost no cells showing TMRM staining at the edge of the scratch, indicating that many cells had undergone mitochondrial depolarization. The addition of 10 nM SPN11 or SPN12 increased the number of cells with good mitochondrial potentials, while biotin had no effect. 【0202】 Figure 25 is a graph summarizing the ratio of normalized mitochondrial potential (red fluorescence) to nucleus number (blue fluorescence) in HK-2 cells 24 hours after mechanical scratching, in serum-free medium alone (control) or in serum-free medium containing 10 nM biotin or peptide-conjugated biotin molecules. Incubation with SPN11 or SPN12 significantly increased mitochondrial potential in cells at the edge of the scratch (***P<0.001, compared to control). The effect of free biotin was not statistically significant. 【0203】 These results demonstrate that biotin conjugate peptides (SPN11 and SPN12) are superior to free biotin in preserving mitochondrial potential in wound conditions, thanks to their targeted delivery to the inner mitochondrial membrane. This can be rephrased as a better increase in ATP production and cell proliferation. [Examples] 【0204】 Uptake and distribution of SPN15 into the mouse retina after intraperitoneal administration. To demonstrate that peptide conjugate biotin molecules can be administered in vivo, we investigated the uptake and distribution of SPN15 in mice following intraperitoneal (ip) administration. 【0205】 SPN15 (30 mg / kg) was administered intraperitoneally to 13-month-old male mice (C57BL / 6J strain). Two hours after administration, the mice were anesthetized by an overdose of ketamine / xylazine, and their eyeballs were removed. After removing the cornea and lens, the optic cups were fixed in 4% paraformaldehyde in 0.1 M Tris buffer for 1.25 hours. The optic cups were then rinsed three times in Tris buffer for 10 minutes each, and then treated with 10%, 20%, and 30% sacrose to prevent frost damage to the tissue before preparing 30 μm thin sections in a Leica 3050 cryostat. The sections were stored at -20°C until use. 【0206】 To label SPN15 with biotin, thin sections were incubated with streptavidin conjugate AlexaFluor 594 (1:500) (Jackson ImmunoResearch, West Grove, PA). Cell nuclei were stained using ToPro 3 (nuclear staining, 1:1000, Molecular Probes). Mitochondrial locations were determined using antibody against Cox IV (cytochrome c oxidase subunit 4; PA529992, 1:300, Invitrogen) and Mitotracker Red (1:1500 in Tris, Invitrogen). All incubations were performed in microwave (150w) except for Cox IV and Mitotracker labeling, which were performed overnight at 4°C with primary antibody followed by secondary antibody (AF488 conjugate donkey anti-rabbit, Jackson ImmunoResearch). Thin sections were imaged using an Olympus Fluoview 300 with a 40× oil immersion lens at a resolution of 1024×1024 and helium, argon, and neon laser light. 【0207】 Figure 26 (left panel) shows cryostat thin sections from control mice without peptide administration, revealing TOPRO (blue) labeled nuclei in different layers of the retina, where OS = outer segment; IS = inner segment; ONL = outer granular layer; OPL = outer plexiform layer; INL = inner granular layer; IPL = inner plexiform layer; RGC = retinal ganglion cells. 【0208】 Figure 26 (right panel) shows cryostat thin sections from control mice stained with COX4 (green), a protein expressed on the inner mitochondrial membrane; Mitotracker Red (red), a fluorescence that labels mitochondria; and TOPRO (blue), a nucleus-labeling fluorescence. COX4 staining of mitochondria is clearly visible in the photoreceptor IS, as well as in the INL, IPL, and RGC, along with abundant mitochondria. The staining with Mitotracker Red in the photoreceptor OS is surprising, as mitochondria are not present in the OS. 【0209】 Figure 26 (central panel) is a cryostat section from a mouse 2 hours after SPN15 administration. When visualized by streptavidin Alexa Fluor 594 (red) staining, SPN15 is clearly incorporated into different layers of the retina. SPN15 staining is concentrated in the inner and outer plexiform layers (IPL and OPL), where there are synapses between ganglion, bipolar, and amacrine cells and photoreceptors in the inner nuclear layer (INL). Streptavidin staining co-localizes with COX4 staining in RGC, IPL, INL, and photoreceptor IS. The exception is strong streptavidin staining in photoreceptor OS where there is no COX4 staining. The reason why SPN15 is abundantly distributed in OS containing high-density packed discs responsible for light transmission is not clear. 【0210】 Figure 27 is an extended photograph of photoreceptor IS and OS. The retinal pigment epithelium (RPE) is a single layer of cells above the OS. SPN15 staining is clearly visible in RPE cells. 【0211】 These results indicate that SPN15 is rapidly absorbed after ip administration in live mice and widely distributed throughout the retina. There are two blood supply sources to the mammalian retina, the central retinal artery and the choroidal vessels. The central retinal artery starts at the optic nerve and supplies the inner retina. Choroidal blood flow is extremely important for the maintenance of the outer retina (photoreceptors and retinal pigment epithelium (RPE)). Streptavidin staining indicates that SPN15 is distributed through both vasculatures. 【0212】 SPN15 staining is concentrated in the inner and outer plexiform layers (IPL and OPL). The outer nuclear layer (ONL) contains the cell bodies of rods and cones, and the inner nuclear layer (INL) contains the cell bodies of ganglion cells, horizontal cells, and amacrine cells. The OPL is where synapses between rods and cones, as well as vertically running bipolar cells and horizontally oriented cells, are present. The IPL functions as a relay station for bipolar cells to ganglion cells, where visual messages are transmitted along the optic nerve to the brain. 【0213】 SPN15 is also concentrated in the inner segment (IS), which contains very long, thin mitochondrial aggregates that support the metabolism of both rod and cone photoreceptors. The high concentration of SPN15 in the IS is expected due to the mitochondrial density. These mitochondria play a role in the biosynthesis of numerous lipid disks in the outer segment (OS), which contains the visual pigment molecule (rhodopsin) for visual signaling. The very high concentration of SPN15 in the OS is unexpected, as mitochondria are completely absent. This is confirmed by the lack of staining for COX4 in the OS. Staining of the OS with Mitotracker has been reported, but the reason is unclear and further research is needed. The concentration of SPN15 in the OS may be important for maintaining disk stability. 【0214】 The oscillous retinal disk undergoes photo-induced oxidative damage, is typically phagocytosed by the retinal pigment epithelium (RPE), and is degraded by lysosomal degradation, allowing for the reuse of rhodopsin and the production of ketone bodies that can be used as metabolic fuel by photoreceptors. The RPE is a monolayer of cells between the oscillous retinal disk and the choroid, which also forms the blood-retinal barrier. The distribution of SPN15 in the RPE layer is highlighted in Figure 27. 【0215】 These findings confirm that SPN15 can be distributed to high-mitochondrial-density areas of the retina after systemic administration. These findings suggest that SPN15 and other peptide-conjugated biotin molecules may be beneficial for a number of ophthalmic diseases. Targeting of RPE suggests that SPN15 can improve mitochondrial bioenergy therapy in aging and combat age-related macular degeneration and diabetic retinopathy. This is supported by results showing that SPN compounds conserve mitochondrial potential, increase ATP synthesis, improve cell viability, and increase cell proliferation in ARPE-19 cells, a human retinal pigment epithelial cell lineage. SPN15 may also increase the viability of retinal ganglion cells under increased pressure from glaucoma, which can also lead to optic nerve damage. 【0216】 definition When the following terms are substantially used herein, the accompanying definitions apply. These terms and definitions are provided unbiased and consistent with this application, and the right to redefine them through amendments made in the course of this application or any application claiming priority therein is reserved. For the purpose of describing the claims of any patent claiming priority therein, each definition in that patent serves as an explicit and unambiguous disavow of the subject matter outside of that definition. 【0217】 1 (a) - at least one. 【0218】 Activity - action, act, stage, and / or process or part thereof. 【0219】 To adapt - to design, make, set up, arrange, shape, form, and / or make suitable and / or conform to a particular purpose, function, use, and / or situation. 【0220】 To add - to combine and / or combine (something) with something else in order to increase its size, quantity, effect, and / or range. 【0221】 To administer - to give and / or apply. 【0222】 Alcohol is one of the classes of chemical compounds having the general formula ROH, where R represents an alkyl group and -OH represents a hydroxyl group, such as in methyl alcohol CH3OH or ethyl alcohol C2H5OH. 【0223】 Alternating - indicates or relates to every other element in a sequence. 【0224】 Amines – any of the group of nitrogenous organic compounds, such as ethylamines and C2H5NH2, which can be considered ammonia derivatives in which one or more hydrogen atoms are replaced by a hydrocarbon group. 【0225】 Amino acids are compounds in which at least one amino group and at least one carboxyl group are bonded to the same carbon skeleton, and the nitrogen atom of the amino group may form part of a ring. Such compounds include L- and D-isomers of natural amino acids. 【0226】 And together with -. 【0227】 And / or - used in conjunction with or instead of. 【0228】 Antibiotics are substances produced by and / or derived from certain microorganisms, including fungi and bacteria, that can destroy or inhibit the growth of other microorganisms, particularly bacteria, such as penicillin or erythromycin. Antibiotics are widely used for the prevention and treatment of infectious diseases. 【0229】 Anticonvulsants - Any type of drug used to prevent or eliminate seizures. 【0230】 Antioxidants—substances that reduce the generation of reactive oxygen species, inhibit the oxidation of other substances, delay the oxidation of other substances, and / or scavengers of free radical species, reactive oxygen species, hydroxyl radical species, oxidized lipids, and / or lipid peroxidation products. 【0231】 Any - without specifying, one, several, all, and / or all. 【0232】 Instrument - a device or apparatus for a specific purpose. 【0233】 Approximately and / or substantially the same, including for each value in a series of two or more values; within an acceptable error for a particular value as determined by one of ordinary skill in the art, the error depending in part on how the value is measured or determined; within one standard deviation; a range that is considered to include the recited value and, taking significant digits into account, is also considered to include the recited value rounded up or down, if no specific error (e.g., the standard deviation of the average value given in a data chart or table) is recited; and / or within ±20%, 15%, 10%, or 5% of the specified value. 【0234】 An organic compound having an unsaturated ring containing alternating double and single bonds, including compounds having an aromatic - benzene ring. 【0235】 Arrange - To place in a particular order. 【0236】 Associate - To join, connect to each other, and / or relate. 【0237】 At - At, on, and / or near. [[ID=十七]] 【0238】 At least - Above, and perhaps more, which applies to each value in any continuation of the value that the phrase "at least" precedes. 【0239】 Be - To actually exist. 【0240】 Biologically active - Configured to alter cell biology. 【0241】 Biotin - A crystalline, water - soluble vitamin of the vitamin B complex, C 10 H 16 O3N2S, also sometimes called vitamin B7 and / or vitamin H. 【0242】 Biotinidase is an enzyme encoded in the BTD gene in humans that readily cleaves and / or degrades biotinamides to release free biotin and amines; its main substrate is biocitin, or biotin linked to lysine; biotinidase can also break down biotin esters. 【0243】 A biotinylated polypeptide has one or more biotin molecules located at its terminal ends. 【0244】 The C-terminus (also known as the carboxyl terminus, carboxyl terminus, C-terminal tail, C-terminus, or COOH terminus) is the end of an amino acid chain (protein or polypeptide) terminated by a free carboxyl group (-COOH). 【0245】 Can - Possible in at least some embodiments. 【0246】 A carrier-active component and / or activator is a substance added as a method for applying and / or transferring the active component and / or activator. 【0247】 Cationic ions—either ions or groups of ions—have a positive charge and are characterized by their movement towards the negative electrode during electrolysis. 【0248】 Cause - to bring about, induce, induce, generate, or elicit; to be the reason for, result in, and / or bring about. 【0249】 Caused by - resulting from. 【0250】 Cell - The smallest structural unit of living organisms, capable of functioning independently, consisting of cytoplasm, usually a single nucleus, and various other organelles, all surrounded by a semipermeable cell membrane. 【0251】 Cell permeable - the ability to move freely and passively into and out of cells. 【0252】 Cellular - related to or consisting of cells. 【0253】 Chemically specified culture media - Culture media in which all components are specified at precise concentrations, and the media does not contain added animal or human serum, growth factors, hormones, etc. 【0254】 Composition of substances - combinations, reaction products, compounds, mixtures, formulations, materials, and / or synthetic products formed by human and / or automation from two or more substances and / or elements. 【0255】 A compound is a pure, macroscopically homogeneous substance consisting of atoms or ions of two or more different elements in a definite proportion that cannot be separated by physical means. Compounds typically possess properties distinct from those of their constituent elements. 【0256】 Including - meaning that what is included is not limited. 【0257】 To conceive - to imagine, conceptualize, form, and / or develop in the mind. 【0258】 To configure - to design, arrange, set up, shape, and / or make suitable and / or conform to a particular purpose, function, use, and / or situation. 【0259】 Conjugate - the process of linking two compounds together through a covalent bond. 【0260】 Conservative substitutions – substitutions of amino acids in a polypeptide with functionally, structurally, and / or chemically similar natural or non-natural amino acids, such as those in the following groups. Each group contains natural amino acids in which each is a conservative substitution: 1) Glycine (Gly / G), Alanine (Ala / A); 2) Isoleucine (Ile / I), leucine (Leu / L), methionine (Met / M), valine (Val / V); 3) Phenylalanine (Phe / F), tyrosine (Tyr / Y), tryptophan (Trp / W); 4) Serine (Ser / S), threonine (Thr / T), cysteine ​​(Cys / C); 5) Asparagine (Asn / N), glutamine (Gln / Q); 6) Aspartic acid (Asp / D), glutamic acid (Glu / E); and / or 7) Arginine (Arg / R), Lysine (Lys / K), Histidine (His / H) And / or the following groups, each containing a natural amino acid that is a conserved substitution of the other: 1) Nonpolar: Ala, Val, Leu, Ile, Met, Pro (proline / P), Phe, Trp; 2) Hydrophobicity: Val, Leu, Ile, Phe, Trp; 3) Aliphatic: Ala, Val, Leu, Ile; 4) Aromatic: Phe, Tyr, Trp, His; 5) Non-charged polar or hydrophilic: Gly, Ala, Ser, Thr, Cys, Asn, Gln, Tyr; 6) Aliphatic hydroxyl- or sulfhydryl-containing: Ser, Thr, Cys; 7) Amides - containing: Asn, Gln; 8) Acidic: Asp, Glu; 9) Basicity: Lys, Arg, His; and / or 10) Small: Gly, Ala, Ser, Cys And / or the following groups: 1)Hydrophobicity: Val, Leu, Ile, Met, Phe, Trp; 2) Aromatic: Phe, Tyr, Trp, His; 3) Neutral hydrophilic: Gly, Ala, Ser, Thr, Cys, Asn, Gln; 4) Acidic: Asp, Glu; 5) Basicity: Lys, Arg, His; and / or 6) Residues that affect skeletal orientation: Pro, Gly 【0261】 To be related to a closed group (and its legal equivalent) that excludes anything that is not included. 【0262】 To consume - to eat and / or drink; to take into the body by mouth for digestion and / or absorption. 【0263】 Contains - Not limited to containing. 【0264】 To transform, adapt, and / or change. 【0265】 Corresponding - related, associated, accompanying, similar in purpose and / or location, matching in all respects and / or equivalent, and / or matching in quantity, content, size, quality and / or degree. 【0266】 To create - the act of bringing something into existence. 【0267】 Culture medium - A nutrient-rich substance, such as an agar gel or liquid medium, on which cultures of bacteria, fungi, animal cells, or plant cells are grown and / or cultured for scientific purposes. 【0268】 Missing - A state and / or condition that deviates from the desired state and / or condition. 【0269】 To define - to establish the meaning, relationship, outline, form, and / or structure; and / or to describe and / or specify accurately and / or clearly. 【0270】 Service - the act of carrying and / or transferring. 【0271】 To derive - to receive, obtain, and / or produce from a source and / or origin. 【0272】 To decide - typically by investigation, reasoning, and / or calculation, to find, obtain, calculate, decide, estimate, verify, and / or make a decision. 【0273】 Apparatus - machinery, products, and / or their collections. 【0274】 Each and every one of the groups that are considered individually. 【0275】 Effective - sufficient to bring about, cause, induce, and / or be the cause. 【0276】 Embodiments - performance, expression, and / or specific representation. 【0277】 Estimate, estimate - (noun) a calculated value that is close to the actual value; (verb) to calculate and / or determine roughly and / or tentatively. 【0278】 Excessive - exceeding normal, typical, reasonable, and / or appropriate limits. 【0279】 Exemplary—serving as an example, illustration, and / or demonstrative, but not necessarily preferable or advantageous to other embodiments or features. 【0280】 The first element listed in a set. 【0281】 Pharmaceutical preparations - Pharmaceutical preparations administered in specific forms such as tablets, syrups, ointments, or injections. 【0282】 From - Used to indicate the source, origin, and / or location of a supply. 【0283】 A specific sequence, such as a peptide sequence having alternating -ACACAC or CACACA residues (where A represents aromaticity and C represents cationicity), or the reverse thereafter, for example, having aromatic-cationic or cationic-aromatic properties. 【0284】 To create, generate, come into being, and / or be present. 【0285】 Larger than - at least. 【0286】 To gather, to assemble - (noun) several individuals and / or things that are considered together because of one or more similarities; (verb) to associate several individuals or things so that they are considered together and / or have similar characteristics. 【0287】 Having - including, but not limited to. 【0288】 Health – the overall state of a living being at any given time; in particular, physical and / or mental soundness; the absence of disease, injury, disability, and / or abnormality. 【0289】 To improve - to change to a better state and / or condition. 【0290】 Insufficient - lacking. 【0291】 Including - Having what follows, but not being limited to it. 【0292】 To initialize - to prepare something for use and / or for some future event. 【0293】 Damage - Damage and / or harm inflicted on and / or suffered by a person and / or thing. 【0294】 To install - to connect or place in a designated location in preparation for use. 【0295】 Ingestion - the act and / or stage of taking in, orally ingesting, and / or receiving. 【0296】 Intestinal malabsorption - incomplete and / or insufficient absorption of nutrients from the intestines. 【0297】 Inside the muscle - within the muscle. 【0298】 Inside the nasal cavity - inside the nose. 【0299】 Inside a vein - within a vein. 【0300】 "To exist" - the state of actually existing. 【0301】 Lysine is a crystalline, basic, essential amino acid with the structure H2N(CH2)4CH(NH2)COOH, and is mainly produced from many proteins by hydrolysis. 【0302】 Maintenance – Activities related to restoring and / or preserving the performance of goods and / or systems. 【0303】 Mammals—any of the various warm-blooded vertebrates within the class Mammalia, including humans, characterized by skin covered with hair, and in females, mammary glands that produce milk to nourish their offspring. 【0304】 Mammalian - and / or related to mammals. 【0305】 Maximum - Having the largest value. 【0306】 May - permitted and / or accepted in at least some embodiments. 【0307】 Medical condition – disease, injury, disability, and / or physical and / or mental disorder. 【0308】 A drug—a substance intended to relieve at least one symptom of a medical condition and / or to cure a medical condition. 【0309】 Metabolism - related to metabolism and / or metabolism. 【0310】 Metabolic supplements - naturally occurring compounds that enhance energy production. 【0311】 Metabolism is a chemical process that occurs within living cells or organisms that is necessary for maintaining life. In metabolism, some substances are broken down to produce energy for life-sustaining processes, and other substances necessary for life are synthesized. 【0312】 Method - One or more actions performed on an object that is transformed into a different state or object, and / or associated with a particular device, said one or more actions not being the main subject, nor preceding all use of the main subject. 【0313】 Having the minimum - the lowest value. 【0314】 To alleviate - to make something less severe, less serious, or less painful. 【0315】 Mitochondria are spherical or elongated organelles in the cytoplasm of almost all eukaryotic cells that contain genetic material and many enzymes important for cellular metabolism, such as those responsible for converting food into usable energy. 【0316】 This indicates that, once a mitochondrial-targeted substance (e.g., biotin) enters the cell, it is preferentially localized to mitochondria and transported to them so that it is not substantially distributed to other organelles or membranes. 【0317】 Furthermore, a quantifier meaning a larger size, quantity, degree, and / or extent. 【0318】 A motif is a repeating pattern of molecular structures, usually consisting of nucleotides or amino acids in a protein, or typically corresponding to a specific biological activity. 【0319】 The N-terminus (also known as the amino-terminus, NH2-terminus, N-terminal terminus, or amine-terminus) refers to the free amine group (-NH2) located at the end of a polypeptide, which is the starting point of a protein or polypeptide. Within a peptide, the amine group binds to another carboxyl group in the protein to form a peptide chain, but the terminal amino acid of the protein is only linked to the carboxyl terminus, so the remaining free amine group is called the N-terminus. 【0320】 No - the absence of and / or lack of both. 【0321】 One - a single unit, individual, and / or whole thing, article, and / or object, and / or reaching. 【0322】 Operable - capable of performing and / or suitable, ready, and / or adapted to perform its intended use and / or service. 【0323】 Or is a conjunction used to indicate an alternative, and typically appears only before the last item in a group of alternatives. 【0324】 Oral - of the mouth and / or related thereto. 【0325】 Organ - A differentiated part of an organism that performs a specific function, such as an eye, wing, or leaf. 【0326】 Patients, humans, and other mammalian subjects. 【0327】 Peptides - any of the various natural or synthetic compounds (including amino acid salts such as pharmaceutically acceptable salts) that contain at least one peptide linking the carboxyl group of one amino acid to the amino group of another amino acid, and / or two or more amino acids covalently linked by an amide bond. 【0328】 per - each and / or by. 【0329】 Pharmacopoeia – Substances suitable for use in contact with the target tissues and organs without excessive irritation, allergic response, immunogenicity, and / or toxicity, commensurate with a reasonable benefit / risk ratio, effective for its intended use, and / or compatible with any composition containing the substance and other components (e.g., active ingredient or excipient); Substances useful in preparing pharmaceutical compositions, generally safe, non-toxic, not biologically or otherwise harmful, and acceptable for veterinary and human pharmaceutical use; and / or substances approved and / or worthy of approval by federal and / or state regulatory agencies and / or corresponding agencies in countries other than the United States, and / or substances listed in the United States Pharmacopoeia and / or other generally approved pharmacopoeias for use in animals, more specifically in humans. 【0330】 Multiple - A state of having multiple and / or more than one. 【0331】 A chain of amino acids linked together by polypeptide-peptide bonds, with a molecular weight of up to approximately 10,000. 【0332】 Polypeptides—generally peptides and proteins—are referred to herein as the left-hand end of a polypeptide sequence, which is called the "amino (N) terminus," and the right-hand end of the sequence, which is called the "carboxyl I terminus." 【0333】 Part - a smaller portion, component, piece, percentage, ratio, and / or quantity than a larger whole. 【0334】 Pre - A prefix placed before an activity that has occurred in advance or / or beforehand. 【0335】 To decide in advance - to decide, determine, and / or establish beforehand. 【0336】 To store - to keep safely for later use. 【0337】 To prevent - to obstruct, prevent, resist, interfere, stop, and / or prevent something from happening. 【0338】 Probability - A quantitative representation of the likelihood of an event occurring. 【0339】 Products - things made by human and / or mechanical efforts. 【0340】 To estimate - to calculate, estimate, or predict. 【0341】 To promote - to contribute to the progress and / or growth of; to advance, encourage, and / or bring to market. 【0342】 Protein - A sequence of linked amino acid residues containing 50 or more amino acid residues. 【0343】 To provide - to provide, to furnish, to supply, to give, and / or to make available. 【0344】 Range - A standard for the degree and / or variation of a set of values. 【0345】 Ratio - The relationship between two quantities expressed as a quotient obtained by dividing one quantity by the other. 【0346】 To accept - to receive, take, acquire, and / or obtain as a signal. 【0347】 To recommend - to endorse, praise, commend, and / or approve of. 【0348】 To reduce - to make less and / or smaller and / or become. 【0349】 Regeneration - the regrowth of lost and / or destroyed parts and / or organs. 【0350】 To remove, to move - to take away, remove, and / or cause to disappear, and / or move from the place or location where it was located. 【0351】 To repair - to restore to a desired state. 【0352】 To repeat - to do something again and / or to perform it again. 【0353】 Repeatedly; over and over again. 【0354】 To request - to express a desire for and / or to ask for. 【0355】 As a result, occurring - (noun) the outcome and / or consequence of a particular action, operation, and / or process; (verb) the act of causing the outcome and / or consequence of a particular action, operation, and / or process. 【0356】 The aforementioned article, when used in a system or device claim, indicates a subsequent claim term that has been previously introduced. 【0357】 The second set of elements that follows the first element. 【0358】 To select - to choose or select from a set of options. 【0359】 Selected - chosen from multiple options. 【0360】 Array - Ordered set. 【0361】 Serum-free - Lacking the clear, yellowish liquid obtained when whole blood is separated into its solid and liquid components after coagulation. 【0362】 Group - related (multiple). 【0363】 A solution is a homogeneous mixture of two or more substances, which may be solids, liquids, gases, or combinations thereof. 【0364】 Species - A group of solids and / or objects that are grouped together due to their common attributes and assigned a general name; a division which is a subdivision of a genus. 【0365】 Storage - The act of storing or the state of being stored. 【0366】 To store - to keep, preserve, deposit, keep securely, and / or set aside for future use. 【0367】 Subcutaneous tissue - just below the skin. 【0368】 Target animals include, but are not limited to, mammals such as primates (e.g., humans, chimpanzees, or monkeys), rodents (e.g., rats, mice, guinea pigs, gerbils, or hamsters), rabbits (e.g., rabbits), cattle (e.g., cows), suids (e.g., pigs), goats (e.g., sheep), horses (e.g., horses), canids (e.g., dogs), and / or felines (e.g., cats). 【0369】 Sublingual - Located directly beneath and / or on the underside of the tongue. 【0370】 In effect - to a considerable extent and / or degree. 【0371】 To suffer—to feel pain or distress; to endure injury and / or injury; to be tolerant of, to be afflicted with, and / or to be ill with; to be accurately diagnosed with. 【0372】 Supplement - (noun) A product taken to supplement a diet, such as by containing one or more vitamins, herbs, enzymes, amino acids, and / or other ingredients, and by providing missing nutrients. 【0373】 A system is a collection of mechanisms, devices, machines, products, processes, compositions, data, and / or instructions, where the collection is designed to perform one or more specific functions. 【0374】 To target - to interact with. 【0375】 Therapeutic - the medical treatment of, or related to, a disease, injury, disability, and / or abnormality. 【0376】 Therapeutic effective dose – an amount of the substance sufficient, when administered to a subject, to prevent, reduce the risk of developing, delay the onset, slow the rate of progression, and / or reverse the medical condition being treated, and / or to alleviate, to some extent, the symptoms and / or complications of the medical condition and / or one or more of the symptoms and / or complications of the medical condition in at least some of the subjects taking the substance, and / or to induce a biological and / or medical response in cells, tissues, organs, systems, animals and / or humans that is being sought by researchers, veterinarians, physicians and / or clinicians. 【0377】 Tissue - A collection of morphologically similar cells and related intracellular substances that work together to perform one or more specific functions in the body. 【0378】 Tissue regeneration - the regrowth of tissue through cell proliferation, completely restoring a portion of damaged tissue to its normal state. 【0379】 Tissue repair—restoration of tissue structure and function after damage; said restoration includes tissue regeneration and tissue replacement. 【0380】 Tissue replacement - a type of healing in which damaged tissue is repaired by implanting connective tissue or scar tissue. 【0381】 Localized - a specific part of the body, and typically the skin. 【0382】 Transcutaneous - through or via the skin. 【0383】 To deform - to change form, appearance, properties, and / or characteristics to a measurable degree. 【0384】 To transmit - to send as a signal, to provide, to supply, and / or to give. 【0385】 To transport - to move and / or transport something from one place to another. 【0386】 To treat - to alleviate, reduce, reduce in remission, inhibit, reverse, prevent, and / or suppress the progression of a medical condition and / or one or more causes, symptoms, and / or complications associated with that condition; to handle and / or deal with a person and / or an object. 【0387】 Intake - to ingest, consume, and / or use. 【0388】 To use - to operate. 【0389】 To be used - to be used to accomplish something. 【0390】 via and / or by means of. 【0391】 Vitamins are a variety of fat-soluble or water-soluble organic substances that are essential in trace amounts for the normal growth and activity of living organisms, and such substances are synthesized by bacteria and / or plants and / or obtained by animals mainly in their diet; and / or are essential components of the food of animals to which vitamins are administered, other than proteins, carbohydrates, or lipids, including B group vitamins such as B1 (thiamine), B2 (riboflavin), B3 (niacin, niacinamide), B5 (pantothenic acid), B6 ​​(pyridoxine), B7 (biotin), B9 (folic acid), and B12 (cyanocobalamin), vitamin C (ascorbic acid), vitamin D, and vitamin K. 【0392】 Water-soluble - Can dissolve in water. 【0393】 Gravity / weight - the force that attracts an object to Earth or another celestial body, equal to the product of the object's mass and its gravitational acceleration; and / or a factor and / or value assigned to a number in a calculation, such as when determining the mean, in order to reflect its importance, significance, priority, influence, etc., to the effect of the number on the calculation. 【0394】 Among them (wherein) - concerning; and; and / or, in addition. 【0395】 "With" - accompanied by. 【0396】 Zone - an area and / or quantity having at least one predetermined boundary. Note 【0397】 Various substantially and particularly practical and useful exemplary embodiments of the claimed subject matter, including, if any, the best form known to the inventor, are described herein in text and / or graph. References herein of “in one embodiment,” “in an embodiment,” or similar are not necessarily of the same embodiment. 【0398】 Any of the numerous possible variations (e.g., modifications, enhancements, embellishments, refinements, and / or strengthenings), details (e.g., types, aspects, nuances, and / or refinements), and / or equivalents (e.g., substitutions, replacements, combinations, and / or alternatives), of one or more embodiments described herein, may become apparent to a person skilled in the art, relying on their expertise and / or knowledge of the art as a whole, by reading this document without any unnecessary experimentation. The inventors anticipate that anyone skilled in the art, after obtaining the inventors' consent, will implement such variations, details, and / or equivalents as necessary, and therefore the inventors intend that the claimed subject matter will be implemented in ways other than those specifically described herein. Accordingly, as permitted by law, the claimed subject matter includes and encompasses all variations, details, and equivalents of the claimed subject matter. Furthermore, as permitted by law, every combination of the features, functions, activities, substances, and / or structural elements described herein, as well as all conceivable variations, details, and equivalents thereof, are included in the claimed subject matter unless otherwise expressly indicated herein, unless the claim is clearly and specifically waived, or otherwise clearly inappropriate, impractical, or inconsistent with the context. 【0399】 Any examples or illustrative language provided herein (e.g., "etc.") are intended solely to illustrate one or more embodiments and, unless otherwise stated, do not limit the scope of any claimed subject matter. The language herein should not be construed as indicating subject matter not claimed as essential to the performance of the claimed subject matter. 【0400】 Therefore, regardless of the content of any part of this document (e.g., title, field, background, summary, description, abstract, diagrams, etc.), any claim in this document and / or any document claiming this priority, whether first presented or otherwise, is not explicitly stated to be contrary by clear definition, assertion, or reasoning, or is not clearly inconsistent with the context: Without requiring the inclusion of any specific described feature, function, activity, substance, or structural element, any specific sequence of activities, any specific combination of substances, or any specific interrelationship of elements; The features, functions, activities, materials, or structural elements described are not "essential." Within any of the described embodiments, between and between: Any two or more of the listed substances may be mixed, combined, reacted, separated, and / or isolated; Any of the features, functions, activities, substances, components, and / or structural elements described herein, or any combination thereof, may, in particular, include, replicate, exclude, combine, rearrange, reconfigure, integrate, and / or isolate; Any described interaction, arrangement, and / or dependence between any of the described features, functions, activities, substances, components, and / or structural elements may be removed, altered, changed, and / or rearranged; Any of the activities described may be performed manually, semi-automatically, and / or automatically; Any of the activities described may be repeated, carried out by multiple entities, and / or carried out by multiple jurisdictions. 【0401】 In the context of describing various embodiments (particularly in the context of the following claims), the use of the terms “a,” “an,” “the,” “the,” and / or similar referents should be interpreted as encompassing both singular and plural forms unless otherwise indicated herein or clearly contradicts the context. 【0402】 The terms “comprising,” “having,” “including,” and “containing” should be interpreted as unrestricted terms (i.e., “including, but not limited to”) unless otherwise specified. 【0403】 Where any number or range is described herein, unless otherwise explicitly stated, that number or range is an approximation. Unless otherwise indicated herein, any enumeration of ranges of values ​​is intended solely as a shorthand notation for referring individually to each separate value that falls within the range, and each separate value and each separate subrange limited by such separate value are incorporated into the specification as if they were individually listed herein. For example, where the range 1 to 10 is described, even if the specific value or specific subrange is not explicitly stated, the range includes all intermediate values ​​such as 1.1, 2.5, 3.335, 5, 6.179, 8.9999, etc., and all intermediate subranges such as 1 to 3.65, 2.8 to 8.14, 1.93 to 9, etc. 【0404】 If any phrase (i.e., one or more words) appearing in a claim is followed by a figure element number, that figure element number is illustrative and not limiting to the scope of the claim. 【0405】 The claims or claim elements in this document are not intended to evoke 35 USC 112(f) unless the exact phrase “means” is followed by a gerund. 【0406】 Any information in any material incorporated herein by reference (e.g., U.S. patents, U.S. patent applications, books, articles, web pages, etc.) is incorporated herein by reference to the maximum extent permitted by law, but only to the extent that there is no conflict between such information and any other definitions, statements, and / or figures expressed herein. In the event of a conflict that is considered to invalidate any claim herein or to seek priority therein, any such conflicting information in such material is not specifically incorporated herein by reference. No specific information in any part of any material incorporated herein by reference that identifies, criticizes, or compares any prior art is incorporated herein by reference. 【0407】 The applicant intends that each claim presented herein and at any point during the execution of this application and in any application claiming priority hereto define a different patentable invention, and that the scope of the invention must change if the scope of the claims changes during the execution of the application and in proportion thereto. Therefore, any reference to any claimed subject matter within this document and during the execution of any patent application relating thereto is intended to refer only to the then undetermined exact language of the claimed subject matter at that particular point in time. 【0408】 Therefore, all parts of this document (e.g., title, field, background, summary, description, abstract, illustrations, etc.), other than the definitions provided for the claims themselves and any terms used therein, should be considered factually descriptive and not restrictive. As would be reasonably understood by the context of this document, the scope of subject matter protected by any claim of any patent arising therefrom is defined and limited only by the exact language of that claim (and all its legal equivalents) and any definitions provided for any terms used therein.

Claims

[Claim 1] A water-soluble, cell-permeable, first D-biotin is conjugated directly and exclusively via an amide bond to the ε-amine of the C-terminal lysine residue of a mitochondrial target peptide sequence to form a biotinylated polypeptide sequence, wherein the mitochondrial target peptide sequence is: It contains multiple amino acids arranged in a common alternating aromatic-cationic motif, and A biologically active composition of a substance containing a minimum of 4 amino acids and a maximum of 6 amino acids. [Claim 2] The aforementioned mitochondrial target peptide sequence is D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys-NH 2 ; D-Trp-D-Arg-D-Trp-D-Lys-OH; L-Trp-L-Arg-L-Trp-L-Lys-NH 2 (Sequence ID 1); D-Trp-D-Arg-D-Trp-D-Lys-NH 2 and L-Trp-L-Arg-L-Trp-L-Lys-OH(Sequence ID 2); The composition according to claim 1, selected from the polypeptide group comprising: [Claim 3] The composition according to claim 1, wherein a second D-biotin is conjugated to the N-terminal α-amine of the mitochondrial target peptide sequence. [Claim 4] The aforementioned mitochondrial target peptide sequence is D-biotin-D-Trp-D-Arg-D-Trp-D-Lys-OH; The composition according to claim 1. [Claim 5] The biotinylated polypeptide D-Trp-D-Arg-D-Trp-D-Lys(biothinyl)-OH; L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH 2 and D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH 2 ; A composition according to claim 1, selected from the group consisting of the following. [Claim 6] The composition according to claim 1, wherein a second D-biotin is conjugated to an N-terminal α-amine. [Claim 7] The biotinylated polypeptide D-biotin-L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH 2 and D-biotin-D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH 2 . A composition according to claim 1, selected from the group consisting of the following. [Claim 8] It comprises one or more biologically active, water-soluble, cell-permeable, mitochondrial-targeting compounds selected from the biotinylated polypeptide group, Each of the biotinylated polypeptides in the group is a biologically active, water-soluble, cell-permeable, and mitochondrial targeting compound. Multiple amino acids arranged in a common alternating aromatic-cationic motif; A minimum of 4 amino acids and a maximum of 6 amino acids; and D-biotin is conjugated directly to the ε-amine of the C-terminal lysine residue via an amide bond. A pharmaceutically acceptable and therapeutically effective composition as defined by [the relevant definition]. [Claim 9] The biotinylated polypeptide group is D-Trp-D-Arg-D-Trp-D-Lys(biothinyl)-OH; L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH 2 ; D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH 2 ; D-biotin-L-Trp-L-Arg-L-Trp-L-Lys(biotinyl)-NH 2 and D-biotin-D-Arg-L-Tyr-D-Arg-L-Phe-L-Lys(biotinyl)-NH 2 ; A pharmaceutically acceptable and therapeutically effective composition according to claim 8, comprising the above. [Claim 10] Vitamin B1 (thiamine); Vitamin B2 (riboflavin); Vitamin B3 (nicotinic acid, niacinamide); Vitamin B5 (pantothenic acid); Vitamin B6 (pyridoxine); Vitamin B7 (biotin); Vitamin B9 (folic acid); Vitamin B12 (cyanocobalamin); and Vitamin C (ascorbic acid); One or more vitamins selected from the vitamin group consisting of the following: A pharmaceutically acceptable and therapeutically effective composition according to claim 8, further comprising: [Claim 11] Pyruvate; Carnitine; Acetylcarnitine; Creatine; α-Ketoglutaric acid; Alpha-lipoic acid; Coenzyme Q Nicotinamide riboside; and Nicotinamide mononucleotide; Select one or more metabolic supplements from the group of metabolic supplements consisting of the following: A pharmaceutically acceptable and therapeutically effective composition according to claim 8, further comprising: [Claim 12] Leucine; Isoleucine; Valin; glutamine; Serine; Arginine; Methionine; Tryptophan; glycine; Trimethylglycine; and Taurine; One or more amino acids selected from the group of amino acids consisting of the following: A pharmaceutically acceptable and therapeutically effective composition according to claim 8, further comprising: [Claim 13] The pharmaceutically acceptable and therapeutically effective composition according to claim 8, wherein an additional D-biotin is conjugated to the N-terminal α-amine of a biologically active, water-soluble, cell-permeable, mitochondrial-targeting compound. [Claim 14] The pharmaceutically acceptable and therapeutically effective composition according to claim 8, comprising a pharmaceutically acceptable carrier.

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