Compositions and methods for diagnosing and treating chronic kidney disease

Abstract

The present invention relates to methods and compositions for enabling the diagnosis and treatment of chronic kidney disease in felines. In one embodiment, a method for diagnosing chronic kidney disease (CKD) in a feline may include measuring a normalized relative abundance or absolute quantification of a first serum biomarker and determining whether the feline has CKD, early CKD, or late CKD based on the normalized relative abundance or absolute quantification.

Description

Detailed Description of the Invention

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of and priority to U.S. Provisional Patent Application No. 63 / 293,855, filed December 27, 2021, the disclosure of which is incorporated by reference in its entirety into this specification. [Background technology]

[0002]

[0002] The kidneys have five main functions: they filter waste products (e.g., urea and creatinine) from the body, regulate electrolytes (e.g., potassium, calcium, phosphorus, and sodium), produce erythropoietin (which stimulates the bone marrow to produce red blood cells), produce renin (which controls blood pressure), and produce and concentrate urine.

[0003] Chronic kidney disease ("CKD") is a progressive kidney disease with four stages: loss of renal reserve, renal dysfunction, azotemia, and uremic. The kidneys have a large inherent reserve because normal renal function requires only about 30% of the renal capacity. Renal capacity decreases over time for various reasons, such as aging, kidney-damaging diseases, and medicines. Renal dysfunction is characterized by a decline in renal function and is generally observed when about 70% of renal function has been lost (i.e., when only about 30% of the renal capacity is available). Typically, the clinical signs between the stages of loss of renal reserve and renal dysfunction are not obvious, making CKD difficult to detect. Stage 1 CKD does not have azotemia and generally cannot be diagnosed because there are no obvious clinical symptoms. Stage 2 CKD is mild azotemia with no or mild clinical signs. Stage 3 is moderate azotemia with clinical signs. Stage 4 is severe azotemia with clinical signs.

[0004]

[0004] CKD is a terminal disease and one of the leading causes of death in felines. Thus, there is a need for compositions and methods for diagnosing and preventing CKD in felines, particularly for diagnosing early CKD. There is also a need for compositions and methods for treating CKD that result in partial or complete relief. Summary of the Invention

[0005]

[0005] The present disclosure generally relates to compositions and methods for diagnosing and treating chronic kidney disease in felines. In one embodiment, a method for diagnosing chronic kidney disease (CKD) in a feline may include measuring a normalized relative abundance or absolute quantification of a first serum biomarker selected from the serum biomarkers listed in Tables 1-3 as described in Example 1. The method may further include determining that the feline has CKD if the normalized relative abundance or absolute quantification of the first serum biomarker falls outside a range of values ​​in a non-CKD column, determining that the feline has early CKD if the normalized relative abundance or absolute quantification of the first serum biomarker has a value within a range of an early CKD column, or determining that the feline has late CKD if the normalized relative abundance or absolute quantification of the first serum biomarker has a value within a range of a late CKD column.

[0006]

[0006] In another embodiment, a method for enabling treatment of CKD or delaying the progression of CKD in a feline may include diagnosing CKD in a feline as discussed herein and recommending a composition for the feline, wherein the composition treats or delays the progression of CKD in the feline.

[0007] In yet another embodiment, feline serum biomarkers for diagnosing early stage CKD in felines are 4-ureidobutyrate, erythronate, 6-oxopiperidine-2-carboxylate, N-acetyl-isoptoreanine, retinol (vitamin A), pseudouridine, homocitrulline, ribonate, 1-methylhistidine, 3-hydroxy-2-ethylpropionate, 5,6-dihydrouridine, 3,4-dihydroxybutyrate, formimiminoglutamate, gamma-carboxyglutamate, (S)-3-hydroxybutyrylcarnitine, hydroxyasparagine, palmitoylcarnitine, aconitate [cis or trans]. , N1-methylinosine, orotidine, 7-methylguanine, N-acetylserine, succinylcarnitine, 2R,3R-dihydroxybutyrate, 2-O-methylascorbic acid, O-sulfo-L-tyrosine, N-acetyltaurine, 5-hydroxymethylcytidine, arabitol / xylitol, arachidoylcarnitine, lyxonate, N6,N6-dimethyllysine, 2,3-dihydroxy-5-methylthio-4-pentenoate (DMTPA), hexanoyltaurine, allantoin, 3-hydroxy-3-methylglutarate, allantoic acid, myo-inositol, urea, and dimethylglycine.

[0008]

[0008] Additional features and advantages are described herein, and will be apparent from the detailed description that follows. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0009] definition

[0009] As used in this disclosure and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context dictates otherwise. Thus, for example, reference to "a composition" or "the composition" includes two or more compositions. The term "and / or" used in the context of "X and / or Y" should be interpreted as "X" or "Y" or "X and Y." As used herein, the terms "example" and "for example," particularly when followed by a list of terms, are used merely exemplary and illustratively and are not exclusive or inclusive.

[0010]

[0010] As used herein, "about" is understood to refer to a number within a numerical range, for example, within a range of -10% to +10%, within a range of -5% to +5%, or in one embodiment within a range of -1% to +1% of the referenced number, and in a specific embodiment within a range of -0.1% to +0.1% of the referenced number. Furthermore, all numerical ranges herein are to be understood to include all integers, whole or fractional, within the range. Furthermore, these numerical ranges are to be interpreted as supporting claims directed to any number or subset of numbers within the range. For example, a disclosure of 1 to 10 is to be interpreted as corresponding to ranges of 1 to 8, 3 to 7, 1 to 9, 3.6 to 4.6, 3.5 to 9.9, etc.

[0011]

[0011] All percentages expressed herein are by weight of the composition on a dry matter basis unless otherwise stated. Those skilled in the art will understand that the term "dry matter basis" means that the concentration or percentage of an ingredient in a composition is measured or specified after all free moisture in the composition has been removed. When reference is made to pH, the value corresponds to the pH measured at 25°C with standard equipment. "Amount" may be the total amount of the referenced ingredient per serving of the composition or the total amount per discrete unit of the composition, and / or may be the weight percent of the referenced ingredient by dry weight. Additionally, "amount" includes zero; for example, a recitation of an amount of a compound does not necessarily mean that the compound is present unless accompanied by a range excluding zero.

[0012]

[0012] The terms "food", "food product" and "food composition" refer to a product or composition intended for ingestion by an animal and providing the animal with at least one nutrient. Moreover, in this respect, these terms refer to the product or composition being in a form ready for ingestion and not merely an intermediate product from which an ingestible product or composition is made, although in some embodiments other food compositions can be added. The term "pet food" refers to any food composition intended for ingestion by a pet. The term "pet" refers to any animal that can benefit from or enjoy the compositions provided by the present disclosure. For example, a pet may be an animal such as a bird, a bovine, a canine, an equine, a feline, a caprine, a wolf, a murine, an ovine or a porcine, although a pet may be any suitable animal.

[0013]

[0013] The term "complete and balanced" when referring to a food composition means a food composition that contains all known necessary nutrients in appropriate amounts and proportions based on the recommendations of recognized authorities in the field of animal nutrition, and thus can serve as the sole source of dietary intake to sustain life or promote production without the need for additional nutritional sources. Nutritionally balanced pet food and animal food compositions, such as complete and balanced food compositions formulated according to standards established by the Association of American Feed Control Officials (AAFCO) as of January 1, 2021, are widely known and widely used in the art.

[0014]

[0014] The term "companion animal" means a dog or a cat. In one embodiment, the compositions and methods disclosed herein involve senior cats.

[0015] As used herein, "chronic kidney disease" or "CKD" is defined as the presence of structural or functional abnormalities in one or both kidneys for more than three months. In one embodiment, CKD can be diagnosed as one of four stages as defined by the International Renal Research Association (IRIS) guidelines (revised 2019) (http: / / www.iris-kidney.com / guidelines / staging.html).

[0016] As used herein, "early stage chronic kidney disease" or "early stage CKD" refers to stage 1 and / or stage 2 chronic kidney disease.

[0017] As used herein, "late stage chronic kidney disease" or "late stage CKD" refers to stage 3 and / or stage 4 chronic kidney disease.

[0018]

[0018] As used herein, "normalized relative abundance" refers to the area under the curve of the ion counts of each biomarker measured by liquid chromatography and mass spectrometry, which is further transformed using base 2 logarithm and autoscaled to achieve zero mean and unit variance. Such measurements are known in the art and described by Metabolon Inc. For example, details and further information can be found in Ford et al. Appl Lab Med 2020 and Li et al. J Am Heart Assoc 2021.

[0019]

[0019] The term "vitamin B" refers to any B vitamin, including derivatives, acid forms and salts thereof. Such B vitamins include, but are not limited to, vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin, nicotinic acid, nicotinamide), vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine, pyridoxal, pyridoxamine), vitamin B7 (biotin), vitamin B8 (myo-inositol), vitamin B9 (folic acid), vitamin B12 (cobalamin compounds, such as methylcobalamin, hydroxocobalamin, and cyanocobalamin).

[0020]

[0020] "Wet food" refers to pet food with a moisture content of about 50% to about 90%, and in one embodiment, about 70% to about 90%. "Dry food" refers to pet food with a moisture content of less than about 20%, and in one embodiment, less than about 15%, and in a specific embodiment, less than about 10%. "Semi-moist food" refers to pet food with a moisture content of about 20% to about 50%, and in one embodiment, about 25% to about 35%.

[0021]

[0021] "Kibble" is used synonymously with "chunk" herein, either term meaning a dry or semi-moist pet food piece that may have a pellet shape or any other shape, and can be made by slicing the food composition into individual pieces. Non-limiting examples of kibbles include granules; pellets; chunks of pet food, dehydrated meat, meat analogs, vegetables, and combinations thereof; and pet snacks such as meat or vegetable jerky, rawhide, and biscuits. "Meat analogs" are meat emulsion products that resemble natural meat pieces in appearance, texture, and physical structure.

[0022]

[0022] The term "effective amount" in "therapeutically effective amount" refers to an amount of a compound of the present invention that (i) treats or prevents a particular disease, condition, or disorder, (ii) relieves, ameliorates, or eliminates one or more symptoms of a particular disease, condition, or disorder, or (iii) prevents or delays the onset of one or more symptoms of a particular disease, condition, or disorder described herein. In some embodiments, the particular disease, condition, or disorder can be chronic kidney disease.

[0023]

[0023] The term "dietary supplement" refers to a product that is intended to be consumed in addition to the normal animal diet. Dietary supplements may be in any form, such as solids, liquids, gels, tablets, capsules, and powders. In one aspect, dietary supplements may be provided in convenient dosage forms. In some embodiments, dietary supplements may be provided in bulk home packages, such as bulk powders, liquids, gels, or oils. In other embodiments, supplements may be provided in bulk amounts for inclusion in other food items, such as snacks, treats, supplement bars, and beverages.

[0024]

[0024] The term "long-term administration" refers to a period of repeated administration or intake that exceeds one month. For certain embodiments, periods of more than two, three, or four months may be employed. Longer periods may also be employed, including more than five, six, seven, eight, nine, or ten months. Periods of more than eleven months or one year may also be employed. Longer term use extending beyond one, two, three, or more years is included in the present invention. For certain elderly animals, the animal will continue regular intake for the remainder of its life. This may also be referred to as intake over "extended" periods.

[0025] The term "regularly" refers to administration of the composition or ingestion of the composition at least monthly, and in one aspect, at least weekly. In certain embodiments, more frequent administration or ingestion can occur, such as two or three times per week. Additionally, in other embodiments, a dosing regimen including ingestion at least once per day can be used.

[0026]

[0026] The compositions disclosed herein may be free of any element not specifically disclosed herein. Thus, the disclosure of an embodiment using the term "comprising" includes the disclosure of an embodiment "consisting essentially of" the specified components, and an embodiment "consisting of" the specified components. Similarly, the methods disclosed herein may be free of any step not specifically disclosed herein. Thus, the disclosure of an embodiment using the term "comprising" includes the disclosure of an embodiment "consisting essentially of" the specified steps, and an embodiment "consisting of" the specified steps. Unless otherwise indicated and directly stated, any embodiment disclosed herein may be combined with any other embodiment disclosed herein.

[0027]

[0027] Unless otherwise specified, all technical and scientific terms and any abbreviations used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any compositions, methods, products, or other techniques or materials similar or equivalent to those described herein can be used in the practice of the present invention, the preferred compositions, methods, products, or other techniques or materials are described herein.

[0028]

[0028] All patents, patent applications, publications, and other references cited or referred to in this specification are hereby incorporated by reference to the extent permitted by law. The discussion of these references is intended only to summarize the assertions made in the references. No admission is made that such patents, patent applications, publications, or references, or any portion thereof, are relevant prior art to the present invention, and the right to challenge the accuracy and pertinence of such patents, patent applications, publications, and other references is expressly reserved.

[0029] Embodiment

[0029] In one embodiment, a method for diagnosing chronic kidney disease (CKD) in a feline may include measuring a normalized relative abundance or absolute quantification of a first serum biomarker selected from the serum biomarkers listed in Tables 1-3 as described in Example 1. The method may further include determining that the feline has CKD if the normalized relative abundance or absolute quantification of the first serum biomarker falls outside (greater or smaller than) a value within a range in the non-CKD column, determining that the feline has early stage chronic kidney disease if the normalized relative abundance or absolute quantification of the first serum biomarker has a value within a range in the early CKD column, or determining that the feline has late stage CKD if the normalized relative abundance or absolute quantification of the first serum biomarker has a value within (less than or greater than) a value within a range in the late CKD column.

[0030]

[0030] Furthermore, a method for enabling treatment or slowing the progression of chronic kidney disease in a feline can include the steps of diagnosing CKD in a feline as discussed herein and recommending a composition for the feline, wherein the composition treats or slows the progression of CKD in the feline.

[0031] Further feline serum biomarkers for diagnosing early stage CKD in felines include 4-ureidobutyrate, erythronate, 6-oxopiperidine-2-carboxylate, N-acetyl-isoptoreanine, retinol (vitamin A), pseudouridine, homocitrulline, ribonate, 1-methylhistidine, 3-hydroxy-2-ethylpropionate, 5,6-dihydrouridine, 3,4-dihydroxybutyrate, formimiminoglutamate, gamma-carboxyglutamate, (S)-3-hydroxybutyrylcarnitine, hydroxyasparagine, palmitoylcarnitine, aconitate [cis or trans]. ], N1-methylinosine, orotidine, 7-methylguanine, N-acetylserine, succinylcarnitine, 2R,3R-dihydroxybutyrate, 2-O-methylascorbic acid, O-sulfo-L-tyrosine, N-acetyltaurine, 5-hydroxymethylcytidine, arabitol / xylitol, arachidoylcarnitine, lyxonate, N6,N6-dimethyllysine, 2,3-dihydroxy-5-methylthio-4-pentenoate (DMTPA), hexanoyltaurine, allantoin, 3-hydroxy-3-methylglutarate, allantoic acid, myo-inositol, urea, and dimethylglycine.

[0032]

[0032] Generally, the method of the present invention for diagnosing a feline with CKD includes measuring the relative abundance or absolute quantification of serum biomarkers and determining that the feline has CKD, and in some embodiments, determining whether the feline has early CKD or late CKD based on the specific ranges disclosed herein. Such diagnosis is possible based on the novel biomarkers and ranges discovered by the inventors. Generally, the values ​​of the biomarkers range from lower values ​​(non-CKD) to higher values ​​(CKD) for the various ranges, although the reverse may also be true. All ranges are provided in Tables 1-3, including smaller (<) and larger (>) ranges.

[0033]

[0033] The markers have distinct ranges that allow diagnosis between early CKD and late CKD, but some markers have undetermined ranges that are not determined to be at a particular stage. Thus, the method of the present invention includes the use of multiple markers to allow diagnosis between early CKD and late CKD. Thus, in one embodiment, the method may further include the steps of measuring the normalized relative abundance or absolute quantification of a second serum biomarker when the normalized relative abundance of a first serum biomarker has a value within the range of the undetermined stage column, and determining that the feline has early CKD if the normalized relative abundance or absolute quantification of the second serum biomarker has a value within the range of the early CKD column, or determining that the feline has late CKD if the normalized relative abundance or absolute quantification of the second serum biomarker has a value within the range of the late CKD column (less than or greater than the value).

[0034]

[0034] Additionally, the method may further include the steps of measuring a normalized relative abundance or absolute quantification of a third serum biomarker when the normalized relative abundance or absolute quantification of the first serum biomarker and the second serum biomarker have values ​​within the range of the undetermined stage column, and determining that the feline has early stage chronic kidney disease if the normalized relative abundance or absolute quantification of the third serum biomarker has a value within the range of the early CKD column, or determining that the feline has late stage CKD if the normalized relative abundance or absolute quantification of the third serum biomarker has a value within the range of the late CKD column (smaller or greater than the value).

[0035]

[0035] The method may further include measuring a normalized relative abundance or absolute quantification of a fourth serum biomarker when the normalized relative abundance or absolute quantification of the first, second, and third serum biomarkers have values ​​within the ranges of the undetermined stage column, and determining that the feline has early stage chronic kidney disease if the normalized relative abundance or absolute quantification of the fourth serum biomarker has a value within the ranges of the early CKD column, or determining that the feline has late stage CKD if the normalized relative abundance or absolute quantification of the fourth serum biomarker has a value within the ranges of the late CKD column (less than or greater than). Other markers, such as a fifth marker, sixth marker, seventh marker, etc., may also be measured in a similar manner.

[0036]

[0036] Therefore, the determining step of the present invention can be based on one marker, two markers, three markers, four markers, five markers, six markers, seven markers, eight markers, nine markers, ten markers, etc. When multiple markers are used, the diagnosis between early and late can be based on a simple majority vote if the markers indicate more than one stage, for example, if seven markers are used and five of the seven markers indicate early and two of the seven markers indicate late, a diagnosis of early can be made.

[0037]

[0037] Generally, a method for enabling, treating or slowing the progression of chronic kidney disease in a feline can include diagnosing chronic kidney disease (CKD) in a feline as discussed herein and recommending or providing a composition for the feline, wherein the composition treats or slows the progression of CKD in the feline.

[0038]

[0038] Such a composition can be any composition useful for providing nutrition to a feline with CKD. In one embodiment, the composition can include a combination of glycine, methionine, cysteine, and glutamine. In one aspect, an amino acid blend of glycine, methionine, cysteine, and glutamine can have a synergistic effect for treating a renal cat. In another aspect, the amino acid blend can be free amino acids. In yet another aspect, the amino acid blend can further include at least one of protein, carbohydrate, fat, and fiber. In one embodiment, the composition can include medium chain triglycerides. In another embodiment, the composition can include less than 1% phosphorus and / or phosphate compounds. In yet another embodiment, the composition can include less than 1% potassium. In yet another embodiment, the composition can have a protein to phosphorus ratio of 5:1 to 15:1. In one embodiment, the composition can include arginine, eicosapentaenoic acid, docosahexaenoic acid, vitamin E, and B vitamins. Such B vitamins include any combination of vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin, nicotinic acid, nicotinamide), vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine, pyridoxal, pyridoxamine), vitamin B7 (biotin), vitamin B8 (myo-inositol), vitamin B9 (folic acid), and vitamin B12 (cobalamin compounds, such as methylcobalamin, hydroxocobalamin, and cyanocobalamin).

[0039]

[0039] Generally, the composition may be administered sufficiently to be effective for treatment. In one aspect, administration may be periodic. In another aspect, administration may be chronic. Administration of the composition may include any delivery method. In one embodiment, the composition may be administered in conjunction with a pet food composition. In another embodiment, the composition is a pet food. In yet another embodiment, the composition may be a sachet or supplement administered in conjunction with a pet food. In yet another embodiment, the composition may be a sachet or supplement administered separately from other food compositions.

[0040]

[0040] Generally, the protein may be any crude protein material and may include vegetable proteins such as soybean meal, soybean protein concentrate, corn gluten meal, wheat gluten, cottonseed, pea protein, canola meal, and peanut meal, or animal proteins such as casein, albumin, and meat proteins. Examples of meat proteins useful herein include beef, pork, lamb, horse, poultry, fish, and mixtures thereof. The composition may also optionally include other materials such as whey and other dairy by-products. In one aspect, the protein includes collagen, whey, or mixtures thereof. In one embodiment, the food composition may comprise protein in an amount of about 10%, 20%, 30%, 35%, 40%, 45%, 50%, or even 55% by weight to about 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, or even 70% by weight, including various subranges within these ranges. In one aspect, the protein may be about 20% to about 60% by weight of the food composition. In another aspect, the protein may be about 25% to about 50% by weight of the food composition.

[0041]

[0041] Additionally, the compositions of the present invention may include isoflavones. In various embodiments, the isoflavones include at least one of daidzein, 6-O-malonyldaidzein, 6-O-acetyldaidzein, genistein, 6-O-malonylgenistein, 6-O-acetylgenistein, glycitein, 6-O-malonylglycitein, 6-O-acetylglycitein, biochanin A, or formononetin. In certain embodiments, the isoflavones or metabolites thereof may be derived from soybean (Glycine max). When present, the one or more metabolites preferably include equol. In one embodiment, the food composition may include isoflavones in an amount of about 300 mg, 400 mg, 500 mg, 600 mg, 700 mg, 800 mg, 900 mg, or even 1,000 mg per kg of food composition to about 500 mg, 600 mg, 700 mg, 800 mg, 900 mg, 1,000 mg, 1,100 mg, 1,200 mg, 1,300 mg, 1,400 mg, or even 1,500 mg per kg of food composition, including various subranges within these amounts. In one aspect, the isoflavones may be present in an amount of about 300 mg to 1,500 mg per kilogram of the pet food composition. In another aspect, the isoflavones may be present in an amount of about 700 mg to 1,200 mg per kilogram of the pet food composition.

[0042] Generally, any type of carbohydrate can be used in the food composition. Examples of suitable carbohydrates include grains or cereals such as rice, corn, millet, sorghum, alfalfa, barley, soybean, canola, oats, wheat, rye, triticale, and mixtures thereof. In one embodiment, the carbohydrate comprises about 10% to about 70% by weight of the food composition. In another embodiment, the carbohydrate comprises about 20% to about 60% by weight of the food composition. In other aspects, the carbohydrate can be present in an amount from about 10%, 20%, 30%, 40%, or even 50% by weight to about 20%, 30%, 40%, 50%, 60%, or even 70% by weight.

[0043]

[0043] Generally, the food composition includes fat. Examples of suitable fats include animal fats and vegetable fats. In one embodiment, the fat source can be an animal fat source, such as tallow, lard or poultry fat. Vegetable oils, such as corn oil, sunflower oil, safflower oil, grape seed oil, soybean oil, olive oil, fish oil, and other oils rich in monounsaturated fatty acids, as well as n-6 polyunsaturated fatty acids and n-3 polyunsaturated fatty acids, can also be used. In one embodiment, the food composition may include fat in an amount of about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or even 50% by weight to about 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, or even 60% by weight, including various subranges within these amounts. In one aspect, the fat comprises about 10% to about 40% by weight of the food composition. In another aspect, the fat comprises about 20% to about 35% by weight of the food composition.

[0044]

[0044] In addition, the composition may include omega-3 fatty acids. Non-limiting examples of suitable omega-3 fatty acids include eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), alpha linolenic acid (ALA), and mixtures thereof. In one embodiment, the omega-3 fatty acids may range from about 0.2%, 0.5%, 1%, 2%, or even 3% by weight of the composition to about 1%, 2%, 3%, 4%, or even 5% by weight. In some embodiments, the omega-3 fatty acids are present in the food composition in an amount of about 1% to about 5% by weight. In some embodiments, the omega-3 fatty acids are present in the food composition in an amount of about 1% to about 2% by weight.

[0045]

[0045] The compositions of the present invention may include omega-6 fatty acids in addition to the fats and fatty acids discussed herein. Non-limiting examples of suitable omega-6 fatty acids include linoleic acid (LA), gamma-linolenic acid (GLA), arachidonic acid (AA, ARA), eicosadienoic acid, docosadienoic acid, and mixtures thereof. In one embodiment, the omega-6 fatty acids may range from about 0.2%, 0.5%, 1%, 2%, or even 3% to about 1%, 2%, 3%, 4%, or even 5% by weight of the composition. In some embodiments, the omega-6 fatty acids are present in the food composition in an amount of about 1% to about 5% by weight. In some embodiments, the omega-6 fatty acids are present in the food composition in an amount of about 1% to about 2% by weight.

[0046] Administration of the pet food composition can be done periodically or intermittently, as needed and desired. In one aspect, the food composition can be administered to the animal periodically. In one aspect, administration can be at least once a week. In certain embodiments, more frequent administration or consumption can be performed, such as two or three times a week. In one aspect, the administration regimen can include at least one daily consumption.

[0047]

[0047] According to the disclosed methods, administration can span the period from birth through adulthood of the animal, including administration as part of a dietary regimen. In certain embodiments, the animal can be a juvenile or growing animal. In other embodiments, administration can be on a regular or extended regular basis, beginning, for example, when the animal has reached more than about 10%, 20%, 30%, 40%, or 50% of its estimated or expected lifespan. In some embodiments, the animal may have reached 40%, 45%, or 50% of its expected lifespan. In still other embodiments, the animal may be older, having reached 60%, 66%, 70%, 75%, or 80% of its expected lifespan. Calculations of lifespan can be based on actuarial tables, calculations, estimates, or the like, and can take into account past, present, and future influences or factors known to affect lifespan positively or negatively. Considerations such as species, sex, body size, genetic factors, environmental factors and stressors, current and past health status, past and current nutritional status, and stressors may also affect and may be taken into account when calculating lifespan.

[0048]

[0048] Such administration can be carried out for a period of time necessary to achieve one or more of the objectives described herein, e.g., treatment of renal disease. Other dosage amounts may be appropriate and can be determined based on the animal's initial weight, as well as other variables such as species, sex, breed, age, the desired health effect, etc.

[0049] The moisture content of such pet food compositions varies depending on the nature of the food composition. The food composition may be a dry composition (e.g., kibble), a semi-moist composition, a moist composition, or any mixture thereof. In one embodiment, the composition may be a complete and nutritionally balanced pet food. In this embodiment, the pet food may be a "wet food", a "dry food", or a "medium moisture content" food. "Wet food" refers to pet food that is typically sold in cans or foil bags and has a moisture content that typically ranges from about 70% to about 90%. "Dry food" refers to pet food (typically in the form of small biscuit-like kibbles) that is similar in composition to wet food, but has a limited moisture content, typically ranging from about 5% to about 15% or 20%. In one embodiment, the composition may have a moisture content of about 5% to about 20%. Dry food products include a variety of foods with various moisture contents that are relatively shelf stable and resistant to microbial or fungal deterioration or contamination. Also, in one aspect, the dry food composition may be an extruded food product for companion animals.

[0050]

[0050] The food composition may also include one or more fiber sources. Such fiber sources include soluble, insoluble, fermentable, and non-fermentable fibers. Such fibers may be derived from plant sources such as marine plants, but microbial fiber sources may also be used. A variety of soluble or insoluble fibers may be used, as known to those skilled in the art. The fiber sources may be beet pulp (from sugar beet), gum arabic, gum talha, psyllium, rice bran, corn bran, wheat bran, oat bran, locust bean gum, citrus pulp, pectin, fructooligosaccharides, short chain oligofructose, mannan oligofructose, soybean fiber, arabinogalactan, galactooligosaccharides, arabinoxylan, cellulose, chicory, or mixtures thereof.

[0051] Alternatively, the fiber source may be a fermentable fiber. Fermentable fiber has previously been reported to be beneficial to the immune system of companion animals. Fermentable fiber or other compositions known to those skilled in the art that provide prebiotics and promote the growth of probiotics in the intestine may also be incorporated into the composition to help enhance the effects described herein or to help enhance the effect on the animal's immune system.

[0052]

[0052] In one embodiment, the food composition may comprise about 1% to about 15% total dietary fiber by weight. In some aspects, the total dietary fiber may be present in an amount of about 5% to about 15% by weight, or even about 8% to about 13% by weight. In another embodiment, the food composition may comprise about 1% to about 10% crude fiber by weight. In some aspects, the crude fiber may be present in an amount of about 3% to about 10% by weight, or even about 3% to about 7% by weight.

[0053] In some embodiments, the ash content of the food composition ranges from less than 1% to about 15%. In one aspect, the ash content may be from about 5% to about 10%.

[0054]

[0054] Generally, the food composition may be a meal, a meal component, a snack, a dietary supplement, or a treat. Such compositions may include complete foods intended to supply the necessary dietary requirements of an animal.

[0055] The pet food composition may further comprise one or more substances such as vitamins, minerals, antioxidants, probiotics, prebiotics, salts, and functional additives such as palatants, colorants, emulsifiers, and antimicrobials or other preservatives. Non-limiting examples of suitable preservatives include potassium sorbate, sorbic acid, sodium methyl para-hydroxybenzoate, calcium propionate, propionic acid, and combinations thereof. Minerals that may be useful in such compositions include, for example, calcium, phosphorus, potassium, sodium, iron, chloride, boron, copper, zinc, magnesium, manganese, iodine, and selenium. Examples of additional vitamins useful herein include fat-soluble vitamins such as vitamin A, vitamin D, vitamin E, and vitamin K, and water-soluble vitamins including vitamin B and vitamin C. In various embodiments, it may be useful to include inulin, amino acids, enzymes, coenzymes, and the like.

[0056]

[0056] The specific amount of each additional ingredient in the pet food compositions disclosed herein will depend on a variety of factors, such as the ingredients contained in the first food ingredient and any second food ingredient, the species of the animal, the age, weight, health, sex, and diet of the animal, the rate at which the animal eats and drinks, and the purpose for which the food product is administered to the animal. Thus, the ingredients and their amounts can vary widely.

[0057] For example, the amount of any of the above ingredients may be decreased or increased based on their predicted effect on feline CKD. EXAMPLES

[0058]

[0058] The following non-limiting examples are illustrative of embodiments of the present disclosure.

[0059] Example 1 Feline CKD Study Serum samples from 25 healthy cats and 31 cats with different stages of CKD (4 cats with stage 1, 20 cats with stage 2, 6 cats with stage 3, and 1 cat with stage 4) were collected for untargeted metabolomics analysis. Stages were diagnosed according to the International Renal Research Association (IRIS) guidelines (revised 2019) (http: / / www.iris-kidney.com / guidelines / staging.html).

[0060]

[0060] Metabolites that differ in concentration between healthy cats and cats with CKD were identified, adjusting for the effects of sex and age. A signature of metabolite biomarkers was selected based on a combination of the following criteria:

[0061] 1. A Student's t-test with pooled variance was performed to compare healthy cats and cats with early CKD. p-values ​​were corrected for multiple testing errors. Metabolites with adjusted p-values ​​less than 0.05 and fold changes greater than 1.5 between cats with early CKD and healthy cats were selected.

[0062] 2. For each metabolite, the area under the curve (AUC) value of the receiver operating characteristic (ROC) curve, which indicates the accuracy of classification, was calculated between healthy cats and cats with CKD. Metabolites with an AUC value of 0.8 or more were selected.

[0063] 3. Random forest machine learning was used to identify metabolites that distinguish between healthy and CKD cats. Metabolites with a mean reduction in accuracy score greater than 0.001 were selected.

[0064]

[0064] 4. The intersection of the three metabolite groups from steps 1 to 3 above.

[0065]

[0065] As shown in Table 1, normalized relative abundances were calculated for serum metabolites distinguishing between CKD cats and healthy cats. [Table 1]

[0066] In one embodiment, the serum biomarkers can be selected from the group consisting of 4-ureidobutyrate, erythronate, 6-oxopiperidine-2-carboxylate, N-acetyl-isoptoreanine, retinol (vitamin A), and pseudouridine. In another embodiment, the serum biomarkers can be selected from the group consisting of 4-ureidobutyrate, erythronate, 6-oxopiperidine-2-carboxylate, N-acetyl-isoptoreanine, retinol (vitamin A), pseudouridine, homocitrulline, ribonate, 1-methylhistidine, 3-hydroxy-2-ethylpropionate, 5,6-dihydrouridine, 3,4-dihydroxybutyrate, formimiminoglutamate, gamma-carboxyglutamate, (S)-3-hydroxybutyrylcarnitine, and hydroxyasparagine. In one aspect, the method can include using allantoin as a biomarker.

[0067] Other markers that can be used to diagnose CKD include N,N-dimethyl-pro-pro, 5-methylthioribose, N1-methyl-2-pyridone-5-carboxamide, glutaryl carnitine, histidine, arabonate / xylonate, thioproline, sulfate, 3-hydroxyisobutyrate, suberoyl carnitine, 3-methylglutaconate, adipoyl carnitine, N-acetylalanine, stearoyl carnitine, N-acetyl-β-alanine, equol sulfate, hexanoyl carnitine, N,N,N-trimethyl-alanyl proline betaine (TMAP), methyl succinate, butyryl glycine, 4-acetamidobutanoate, α-ketoglutaramate, These include N6,N6-dimethyllysine, arachidonoylcarnitine, N6,N6,N6-trimethyllysine, creatinine, laurylcarnitine, glucuronate, tryptophan, succinoyltaurine, oleoylcarnitine, eicosenoylcarnitine, 3-hydroxybutyrate (BHBA), gamma-glutamylleucine, dihomo-linoleoylcarnitine, 4-ethylcatechol sulfate, N1-methyladenosine, isovalenylcarnitine, dimethylarginine (SDMA+ADMA), proline, arachidate, hexanoylglycine, linoleoylcarnitine, N-acetylmethionine, acetylcarnitine, butyrylcarnitine, N-acetylthreonine, and gamma-glutamylvaline.

[0068] Based on the biomarkers in Table 1, the biomarkers of the present invention can be used to diagnose CKD in cats, as well as for the specific diagnosis of early CKD and late CKD. The method may include measuring the normalized relative abundance of the biomarkers, and determining that the feline has CKD if the normalized relative abundance of the markers is greater than the range in the non-CKD column according to Table 1. Furthermore, if the markers fall within the range of early CKD or late CKD, further diagnosis can be made. The method may also include measuring additional markers and determining whether the feline has early CKD or late CKD according to the ranges listed in Table 1 when the first marker(s) falls within the range listed in the indeterminate stage.

[0069]

[0069] Eleven markers were quantified by LC-MS / MS. Feline serum samples were spiked with internal standards and subjected to protein precipitation. After centrifugation, an aliquot of the supernatant was injected into an Agilent 1290 / AB SCIEX QTRAP 5500 LC-MS / MS system equipped with an amide UPLC column. The mass spectrometer was operated in positive or negative HILIC mode using electrospray ionization (ESI). The peak areas of the product ions are measured relative to the peak areas of the respective internal standard product ions. Quantification is performed using a weighted linear least-squares regression analysis generated from calibration standards prepared immediately before each measurement. The quantitative results of the eleven metabolites are summarized in Table 2. [Table 2]

[0070] Additional serum samples were collected from 30 privately owned cats in London, UK: healthy cats (n=10), cats with stage 2 CKD (n=10), and cats with stage 3 CKD (n=10). Samples were frozen and stored in a -80°C freezer until use. Additional markers identified according to the protocol described herein are summarized in Table 3. [Table 3]

[0071]

[0071] It should be understood that various changes and modifications to the presently preferred embodiments described herein will be apparent to those skilled in the art. Such changes and modifications can be made without departing from the spirit and scope of the present subject matter and without diminishing its intended advantages. Accordingly, such changes and modifications are intended to be covered by the appended claims.

Claims

1. 1. A method for diagnosing chronic kidney disease (CKD) in a feline, comprising: measuring a first serum biomarker selected from the serum biomarkers listed in Table 1 or Table 3 by normalized relative abundance, or measuring a first serum biomarker selected from the serum biomarkers listed in Table 2 by absolute quantification; 【Table 1】 【Table 2】 【Table 3】 determining that the feline has CKD if the normalized relative abundance or absolute quantification has a value outside the range of the non-CKD column, determining that the feline has early stage chronic kidney disease if the normalized relative abundance or absolute quantification of the first serum biomarker has a value within the range of the early CKD column, or determining that the feline has late stage CKD if the normalized relative abundance or absolute quantification of the first serum biomarker has a value within the range of the late CKD column; A method comprising:

2. 2. The method of claim 1, further comprising the steps of: measuring a normalized relative abundance or absolute quantification of a second serum biomarker when the normalized relative abundance or absolute quantification of the first serum biomarker has a value within a range of the undetermined stage column; and determining that the feline has early stage chronic kidney disease if the normalized relative abundance or absolute quantification of the second serum biomarker has a value within a range of the early CKD column, or determining that the feline has late stage CKD if the normalized relative abundance or absolute quantification of the second serum biomarker has a value within a range of the late CKD column.

3. 3. The method of claim 2, further comprising the steps of: measuring a normalized relative abundance or absolute quantification of a third serum biomarker when the normalized relative abundance or absolute quantification of the first and second serum biomarkers have values ​​within the ranges of the undetermined stage column; and determining that the feline has early stage chronic kidney disease if the normalized relative abundance or absolute quantification of the third serum biomarker has a value within the ranges of the early CKD column, or determining that the feline has late stage CKD if the normalized relative abundance or absolute quantification of the third serum biomarker has a value within the ranges of the late CKD column.

4. 4. The method of claim 3, further comprising the steps of: measuring a normalized relative abundance or absolute quantification of a fourth serum biomarker when the normalized relative abundance or absolute quantification of the first, second, and third serum biomarkers have values ​​within a range of the undetermined stage column; and determining that the feline has early stage chronic kidney disease if the normalized relative abundance or absolute quantification of the fourth serum biomarker has a value within a range of the early CKD column, or determining that the feline has late stage CKD if the normalized relative abundance or absolute quantification of the fourth serum biomarker has a value within a range of the late CKD column.

5. The method of claim 1 , wherein the determining step is based on five biomarkers.

6. The method of claim 1 , wherein the determining step is based on six biomarkers.

7. The method of claim 1 , wherein the determining step is based on seven biomarkers.

8. 11. A method for enabling the treatment or slowing the progression of chronic kidney disease in a feline, comprising the steps of diagnosing chronic kidney disease (CKD) in the feline according to claim 1 and recommending a composition for the feline, wherein the composition treats or slows the progression of CKD in the feline.

9. 9. The method of claim 8, wherein the composition comprises a combination of glycine, methionine, cysteine, and glutamine.

10. The method of claim 8 , wherein the composition comprises medium chain triglycerides.

11. 9. The method of claim 8, wherein the composition comprises arginine, eicosapentaenoic acid, docosahexaenoic acid, vitamin E, and B vitamins.

12. 9. The method of claim 8, wherein the composition contains less than 1% potassium and less than 1% phosphorus and phosphate compounds.

13. 9. The method of claim 8, wherein the composition has a protein to phosphorus ratio of 5:1 to 15:

1.

14. 9. The method of claim 8, wherein the composition is a pet food composition or is administered in conjunction with a pet food composition.

15. 4-Ureidobutyrate, Erythronate, 6-Oxopiperidine-2-carboxylate, N-Acetyl-isopthreanine, Retinol (Vitamin A), Pseudouridine, Homocitrulline, Ribonate, 1-Methylhistidine, 3-Hydroxy-2-ethylpropionate, 5,6-Dihydrouridine, 3,4-Dihydroxybutyrate, Formiminoglutamate, γ-Carboxyglutamate, (S)-3-Hydroxybutyrylcarnitine, Hydroxyasparagine, Palmitoylcarnitine, Aconitate [cis or trans], N1-Methylinosine, Orotidine, 7-Methylguanine, N-Acetylserine, Succinimide 1. A feline serum biomarker for diagnosing early stage chronic kidney disease (CKD) in a feline selected from the group consisting of: arachidyl carnitine, 2R,3R-dihydroxybutyrate, 2-O-methylascorbic acid, O-sulfo-L-tyrosine, N-acetyltaurine, 5-hydroxymethylcytidine, arabitol / xylitol, arachidoylcarnitine, lyxonate, N6,N6-dimethyllysine, 2,3-dihydroxy-5-methylthio-4-pentenoate (DMTPA), hexanoyltaurine, allantoin, 3-hydroxy-3-methylglutarate, allantoic acid, myo-inositol, urea, and dimethylglycine.

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