Paediatric formulations for phenylketonuria and tyrosinaemia and specific free amino acid mixture

Abstract

This invention relates to infant formulae or follow-on formulae suitable for children under the age of 2 years, and their use in the dietary management of phenylketonuria and tyrosinaemia. In particular, the invention relates infant formulae or follow-on formulae comprising casein glycomacropeptide and free amino acids as well as their use in the dietary management of phenylketonuria and tyrosinaemia, and the symptoms thereof. The invention relates to a mixture of free amino acids for use in manufacturing the infant formulae or follow-on formula.

Description

[0001] PAEDIATRIC FORMULATIONS FOR PHENYLKETONURIA AND TYROSINAEMIA AND SPECIFIC FREE AMINO ACID MIXTURE

[0002] FIELD OF THE INVENTION

[0003] This invention relates to infant formulae or follow-on formulae suitable for children under the age of 2 years, and their use in the dietary management of phenylketonuria and tyrosinaemia. In particular, the invention relates infant formulae or follow-on formulae comprising casein glycomacropeptide and free amino acids as well as their use in the dietary management of phenylketonuria and tyrosinaemia, and the symptoms thereof. The invention also relates to a mixture of free amino acids for use in manufacturing the infant formulae or follow-on formulae.

[0004] BACKGROUND OF THE INVENTION

[0005] Phenylketonuria and tyrosinaemia are diseases caused by inborn errors of metabolism of the aromatic amino acids phenylalanine and tyrosine, respectively.

[0006] Phenylketonuria (PKU) is an autosomal recessive disorder of phenylalanine metabolism, in which especially high phenylalanine concentrations can cause brain dysfunction. If untreated, this brain dysfunction results in severe intellectual disability, epilepsy and behavioural problems (van Spronsen et al. (2021) Nature Reviews Disease Primers, 7, 36). PKU is characterised by a deficiency of the liver enzyme phenylalanine hydroxylase (PAH), necessary for the conversion of the essential amino acid phenylalanine (Phe) to tyrosine (Tyr). A low protein diet is prescribed to reduce the intake of phenylalanine to very low levels and prevent irreversible intellectual disability, microcephaly, motor deficits, autism, seizures, developmental problems, aberrant behaviour and psychiatric symptoms.

[0007] Dietary intervention with a low-phenylalanine diet can prevent the major cognitive and neurological deficits, if started immediately after birth (Rocha and MacDonald (2018) Expert Opinion on Orphan Drugs, 6, 667-681). This generally prevents a gold-standard breast milk diet and requires at least some infant formula to be administered (van Rijn et al. (2003) European Journal of Pediatrics 162, 323-326).

[0008] Tyrosinaemia encompasses several diseases that manifest with elevated levels of the amino acid tyrosine in the blood (Chinsky et al (2017) Genetics in Medicine 19, 1380-1395; Beyzaei et al (2022) Orphanet Journal of Rare Diseases 17, 424). These conditions are rare and include hereditary tyrosinaemia, and tyrosinaemia that results from nitisinone treatment in alkaptonuria (AKU). There are three types of hereditary tyrosinaemia - Type I, Type II and Type III, which are characterised by their symptoms and genetic cause. To lower tyrosine levels, a low-phenylalanine, low-tyrosine diet is prescribed to reduce the intake of Tyr and Phe (i.e., as Phe is a precursor of Tyr) to a safe minimum level.

[0009] Dietary interventions for people aged 3 years and older with PKU or tyrosinaemia are known in the art, but research has focused on PKU due to its higher prevalence and relatively homogenous genetic cause (most cases are linked to mutations on the PAH gene (OMIM 612,349) with some linked to tetrahydrobiopterin-associated genes such as GCH1 ‘GTPC , PTS, PCBD1, QDPR, and SPR genes).

[0010] Casein glycomacropeptide (GMP) is often included in dietary interventions for people aged 3 years and older with PKU because it contains only -2 mg of Phe per gram of GMP protein (-0.2% by weight). GMP can be used alone (e.g., JPH04126051A) or in combination with essential amino acids (e.g., WO 1998 / 008402).

[0011] WO 2019 / 239439 describes a nutritional formulation suitable to manage the diet for phenylketonuria, which comprises GMP as the primary source of proteins and additional complementary essential amino acids, to complete the required protein profile, supplied in a mixture comprising arginine, cystine, histidine, tryptophan, tyrosine and leucine.

[0012] WO 2016 / 003263 and WO 2016 / 003273 describe amino acid-based nutritional compositions with improved taste, yet not compromising the (regulatory) dietary restrictions on essential amino acids and conditionally essential amino acids.

[0013] US 9,414,619 B describes a ready to drink liquid nutritional formulation to treat PKU, tyrosinaemia or other metabolism errors. The formulation comprises GMP, as a primary source of proteins, and complementary essential amino acids to complete the required protein profile. The beverage also includes vitamins and minerals to meet the daily needs of these elements and, when used as a complete diet, the formulation also includes a source of carbohydrates and a source of fats.

[0014] FR 3,002,831 Al describes a food product ready for consumption for patients with a metabolic pathology of one or several amino acids. It does not disclose that the food product is suitable for children under the age of 2 years.

[0015] WO 2010 / 144821 describes a medical food based on GMP for the nutritional management of PKU and other metabolic disorders, which, in addition to GMP, includes additional quantities of arginine, leucine and optionally other amino acids, such as tyrosine.

[0016] WO 2000 / 022937 describes a protein material with a low digestion rate, for the preparation of an enteral composition which makes it possible to modulate the levels of postprandial amino acids in the plasma. WO 1998 / 008402 describes a nutritional formula for patients suffering from PKU which has, as a source of protein, a mixture of GMP and complementary essential amino acids other than phenylalanine.

[0017] PKU sphere® has a GMP-dominant protein source supplemented with amino acids for the dietary management of PKU from 4 years of age.

[0018] Ahring et al. (J. Nutr. Metab. (2018) 6352919) compare absorption of CGMP-20 to free-synthetic amino acids with the aim of evaluating short-term effects on plasma AAs as well as biomarkers related to food intake. The patients are 15-48 years old and the formulation was not described as suitable for children under the age of 2 years.

[0019] US 2018 / 0064784 describes liquid metabolic formulas for dietary management of tyrosinemia, including nutritional formulas and hydration beverages (sport drinks). It highlights that not all compositions are suitable for children and does not disclose formulas suitable for children under the age of 2 years.

[0020] Most of the above-mentioned formulae are not tailored for children under the age of 2 years with PKU or tyrosinaemia. WO 1998 / 008402 highlights that amino acid Arg, Cys, Gin, His, He, Leu, Lys, Met, Thr, Trp, Tyr, Phe and Vai are considered essential or conditionally essential for children under the age of 2 years. It suggests the protein source should contain about 40% to about 60% of both GMP and complementary free amino acids in specific amounts, including excess Tyr compared to human milk.

[0021] In infancy, this low-phenylalanine diet is provided by a combination of low phenylalanine formula in conjunction with prescribed and controlled quantities of breast milk / standard infant formula.

[0022] For children under the age of 2 years, the low-phenylalanine diet is usually based on free amino acid formulae.

[0023] WO 2011 / 119023 discloses free amino acid infant formulae for treating PKU, maple syrup urine disease (MSUD) or tyrosinaemia. It is suggested to GMP is removed from the protein source to advantageously reduces the threonine content.

[0024] PKU Anamix Infant (Nutricia) is a free amino acid formula with prebiotic fibre and maintains phenylalanine control in line with an established infant protein substitute without prebiotics and maintains levels of bifidobacteria and lowers stool pH (MacDonald et al. (2011) Molecular Genetics and Metabolism 104, S55-S59).

[0025] PKU Start® (Vitaflo International Ltd) is a free amino acid formula that is well- tolerated, helps maintain good metabolic control, and normal growth in infants with PKU (Yilmaz et al. (2023) Orphanet Journal of Rare Diseases 18, 16). These formulae are recommended for use in infants with PKU, but difficulties with theses formulae can occur in terms of gastrointestinal side effects, such as constipation, and issues with adherence have been reported, perhaps due to the known bitter and / or acid tastes of free amino acid formulae.

[0026] WO 1998 / 008402 describes two infant formulae for patients suffering from PKU with -50% GMP and -50% free amino acids. However, the effect of these formulae on infants with PKU is not described.

[0027] Therefore, there are currently no infant formula recommended for infants with PKU or tyrosinaemia with proven palatability, tolerance and low gastrointestinal side effects that can maintain metabolic control of Tyr and Phe levels whilst supporting normal growth.

[0028] SUMMARY OF THE INVENTION

[0029] The present inventors have developed an infant formula (or follow-on formula) based on GMP and free amino acids suitable for dietary intervention and management in children under the age of 2 years, preferably an infant, with PKU or tyrosinaemia. The formula has excellent organoleptic properties and stability and is an ideal candidate for dietary intervention and management for children under the age of 2 years with an inborn error of Tyr or Phe metabolism.

[0030] Accordingly, in a first general embodiment, the invention provides a formula for children under the age of 2 years, preferably an infant formula, comprising intact casein glycomacropeptide (GMP) and free amino acids for use in the dietary management of phenylketonuria (PKU) or tyrosinaemia.

[0031] In a second general embodiment, the invention provides the use of intact GMP and free amino acids in the manufacture of a formula for children under the age of 2 years, preferably an infant formula, for the dietary management of PKU or tyrosinaemia in an individual in need thereof.

[0032] In a third general embodiment, the invention provides a method of providing dietary management of PKU or tyrosinaemia comprising administering a formula for children under the age of 2 years, preferably an infant formula, comprising intact GMP and free amino acids to an individual in need thereof.

[0033] In a fourth general embodiment, the invention provides a complete infant formula for the dietary management of PKU or tyrosinaemia comprising intact GMP and free amino acids, wherein the GMP to free amino acids by weight are present in a proportion of 1 :4 to In a fifth general embodiment, the invention provides a mixture of free amino acids comprising Arg, Asp+Asn, Cys, Gly, His, He, Leu, Lys, Met, Trp and Vai, and optionally Tyr, wherein the mixture of free amino acids contains less than 0.1 wt% of any of Ala, Gin, Glu, Pro, Phe, Ser or Thr.

[0034] DETAILED DESCRIPTION OF THE INVENTION

[0035] A. Definitions

[0036] The following definitions apply to the terms used throughout this specification, unless otherwise limited in specific instances.

[0037] The term “manage”, “managing” and “management”, particularly in the context of “dietarily manage”, “dietarily managing” and “dietary management”, refer to the prevention or reduction in severity or frequency of one or more symptom of a disease or condition, including ameliorating one or more existing symptoms of a disease or condition, preventing one or more existing symptoms of disease or condition, preventing one or more underlying causes of a disease or condition, ameliorating one or more underlying cause of a disease or condition, reducing the prevalence of one or more symptoms of a disease or condition, and / or reducing the occurrence of one or more symptoms of a disease or condition. It will be understood to include stabilising a disease or condition and preventing progression of a disease or condition. It will also be understood that management of a disease or condition, more specifically dietary management, includes meeting the nutritional requirements of a subject suffering from a disease or condition that prevents the meeting of nutritional requirements in the context of a normal or non-specialized diet. In the absence of management, more specifically dietary management, the subject may experience adverse effects such as malnutrition due to one or more of the following: limited, impaired or disturbed capacity to take, digest, absorb, metabolise or excrete ordinary foods or certain nutrients contained therein or metabolites; and / or symptoms of the disease or condition caused by the limited, impaired or disturbed capacity to take, digest, absorb, metabolise or excrete ordinary foodstuffs or certain nutrients contained therein or metabolites. Management, more specifically dietary management, may include medically-determine nutrient requirements and / or an alteration of a non-controlled and / or normal diet and may include supervision by a medical professional. The skilled person will be familiar with the diagnosis and management, more specifically dietary management, of such diseases or conditions (see, e.g. Commission Directive 1999 / 21ZEC including modifications thereto; Commission Notice on the classification of Food for Special Medical Purposes (2017 / C 401 / 01); Commission Delegated Regulation (EU) 2016 / 128 of 25 September 2015 supplementing Regulation (EU) No 609 / 2013 of the European Parliament and of the Council). In the context of a diseases or conditions that may be classified as an inborn error of metabolism, the subject may lack the ability to utilise specific nutrients and / or consume a normal diet. Management, more specifically dietary management, of the disease or condition may be the only effective therapy preventing further disability or death of a subject. (See, e.g. Berry et al., Peds 145(3): March 2020:e20192261). For example, for an infant starting dietary management at or close to birth, the dietary management is likely to prevent or stabilise the PKU or tyrosinaemia. For an infant starting dietary management when overt symptoms are present, dietary management is likely to reduce the severity and / or frequency of one or more symptoms of the PKU or tyrosinaemia. Symptoms of PKU or tyrosinaemia that may be dietarily managed are described below.

[0038] The term “dietary intervention” is defined as switching from a diet with a standard amino acid profile (i.e., breast milk or standard infant formula) to a diet including a formula described herein in order manage (specifically dietarily manage), prevent and / or treat one or more symptoms of PKU or tyrosinaemia. Where possible, infants should maintain the maximal possible amount of breast milk. However, a dietary intervention based entirely on an infant formula described herein may be required to stabilise or reduce the levels of Phe or Tyr in the blood and ensure the PKU or tyrosinaemia is ameliorated. Follow-on formula may be given alongside a carefully selected diet low in Phe and / or Tyr.

[0039] The terms “treat”, “treating” or “treatment” relates to the full or partial treatment of one or more symptom of PKU and / or tyrosinaemia, described below. Equally, “treat”, “treating” or “treatment” may relate to the prevention of a known symptom that the patient does not yet suffer from, but is expected to suffer from, based on their age or disease severity. Prevention and treatment may be achieved through dietary intervention and / or dietary management.

[0040] The term “infant” is generally applied to very young children under the age of 1 year, preferably under 6 months of age.

[0041] The term "infant formula" as used herein refers to a nutritional composition intended for infants and as defined in Codex Alimentarius, (Codex STAN 72-1981) and / or Infant Specialities (incl. Food for Special Medical Purpose) as defined in Codex Alimentarius, (Codex STAN 72-1981). It may also refer to a foodstuff intended for particular nutritional use by infants during the first months of life and satisfying by itself the nutritional requirements of this category of person (Article 2(c) of the European Commission Directive 91 / 321 / EEC 2006 / 141 / EC of 22 December 2006 on infant formulae and follow-on formulae).

[0042] The term “starter formula” is a formula for infants from birth as breast-milk substitute.

[0043] The term “follow-on formula” is a formula given from the 6th month onwards when an infant starts weaning. It constitutes the principal liquid element in the progressively diversified diet of this category of person.

[0044] The term “liquid” is defined a liquid that is a nearly incompressible fluid that conforms to the shape of its container but retains a nearly constant volume independent of pressure. It is one of the four fundamental states of matter, and is the only state with a definite volume but no fixed shape. Generally, the liquids described herein are a solution or emulsion with a viscosity of below 200 mPa s, measured at 20° C at a shear rate of 100 s ' . The viscosity may be determined by methods known to a skilled person, e.g., by using a rheometer (e.g., a Netzsch Kinexus Pro+) equipped with a plate / plate geometry (60 mm diameter) and 1 mm gap.

[0045] The term “reconstitutable powder” is defined as a powder that can be dissolved into a liquid formula with vigorous shaking by an adult in less than one minute. The reconstituted liquid formula should pass through a standard infant formula bottle teat, so must be free of particles larger than the teat hole, e.g., 1 mm in diameter or greater.

[0046] The term “spoonable food” is defined as liquid, gel or semi-solid with a viscosity of below between 400 and 4000 mPa s, measured at 20° C at a shear rate of 100 s '. The viscosity may be determined by methods known to a skilled person, e.g., by using a rheometer (e.g., a Netzsch Kinexus Pro+) equipped with a plate / plate geometry (60 mm diameter) and 1 mm gap.

[0047] The term “shelf-stable” means that the formula, is chemically stable and maintains the same (low) level of bacterial contamination when stored at 20° C in a sealed, sterile container for a period of at least 3 months, preferably at least 6 months, more preferably at least 9 months and most preferably at least 1 year. The state of the formula should not change during this time such that it cannot be converted to a homogenous form (e.g., a liquid, gel or powder) with minimal shaking or mixing.

[0048] The term “shelf-stable liquid” means that the formula, is chemically stable and maintains the same (low) level of bacterial contamination when stored at 20° C in a sealed, sterile container for a period of at least 3 months, preferably at least 6 months, more preferably at least 9 months and most preferably at least 1 year. The state of the formula should not change during this time such that it cannot be converted to a homogenous form (e.g., a liquid solution or emulsion) with minimal shaking or mixing.

[0049] The term “phenylketonuria” is defined as abnormally high Phe levels in the blood of >360 pmol / L prior to starting dietary intervention. Traditionally, the Guthrie test was employed (Guthrie and Susi (1963) Pediatrics 32; 338-43), but more modern systems based on tandem mass spectrometry are often employed in developed countries. PKU may be confirmed by testing for pathogenic mutations in the PAH or DNAJC12 genes.

[0050] The term “tyrosinaemia” is defined as abnormally high Tyr levels in the blood of >200 pmol / L prior to starting dietary intervention. However, clinical manifestations typically do not become apparent until plasma levels >500 pmol / L. Tyrosinaemia may be confirmed by testing for pathogenic mutations in the FAH gene (expressing fumarylacetoacetase; Type I tyrosinaemia), TAT gene (expressing tyrosine aminotransferase; Type II tyrosinaemia) or HPD gene (expressing 4-hydroxyphenylpyruvate dioxygenase; Type III tyrosinaemia).

[0051] The term “hyperphenylalaninaemia” is defined as abnormally high Phe levels in the blood of 120-360 pmol / L prior to starting dietary intervention and is suggestive of possible PKU. In a patient with hyperphenylalaninaemia, PKU may be confirmed by continued monitoring for Phe levels in the blood and / or by testing for pathogenic mutations in the PAH or DNAJC12 genes. Dietary intervention may be appropriate for children under the age of 2 years with hyperphenylalaninaemia.

[0052] The term “good metabolic control” is defined as partial or total control of amino acid concentrations in the blood. For PKU, Phe levels in the blood of 120-360 pmol / L are considered good metabolic control. Normal Phe levels in the blood are 35-120 pmol / L and are preferable. For tyrosinaemia, Tyr levels in the blood of 200-400 pmol / L are considered good metabolic control. Moderately elevated Tyr levels in the blood are 120-200 pmol / L and are preferable. Normal Tyr levels in the blood are 20-120 pmol / L and are more preferable.

[0053] The term “improved gastrointestinal tolerance” is defined by a caregiver questionnaire recording occurrence of vomiting, regurgitation, abdominal discomfort or colic symptoms, flatulence, number of daily stools, and a description of their characteristics [loose, soft, hard], any feeding difficulties (Yilmaz et al. (2023) Orphanet Journal of Rare Diseases 18, 16). Generally, the improvement is an improvement of overt symptoms described above. However, as gastrointestinal damage may be insidious, the improvement in tolerance encompasses reducing the prevalence of one or more symptoms of gastrointestinal intolerance and / or reducing the occurrence of one or more symptoms of gastrointestinal intolerance that would have occurred without dietary intervention and management. The term “casein glycomacropeptide”, “glycomacropeptide” or “GMP” is a casein-derived peptide formed during milk renneting which is retained in the soluble fraction known as “sweet whey”. In particular, GMP corresponds to the 64 carboxy-terminal amino acids of K-casein (methionine- 106 to valine-169). Within the meaning of this application, GMP preferably also includes the non-glycosylated “caseinomacropeptide” or “CMP”, which is found in approximately equal proportions in whey (Neelima et al. (2013) Diary Science and Technology 93, 21-43). Commercially available sources of GMP may contain a ratio of glycosylatedmon-glycosylated GMP of about 1 : 1.

[0054] The term “intact GMP” refers to GMP that has not been further hydrolysed, especially not fully hydrolysed into free amino acids. As discussed above, it includes both glycosylated and non-glycosylated GMP.

[0055] The term “free amino acids” refers to the 20 canonical amino acids found in human proteins when not in the form of peptides or proteins.

[0056] The terms “Asp+Asn” and “Glu+Gln” related to the total amounts of these amino acids within a sample. Extraction procedures frequently employ dilute acid which partially converts Gin and Asn to Glu and Asp. Therefore, it is common practice to quantify these amino acids in pairs. The standard three-letter code for amino acids are known to the skilled person.

[0057] Other variations to the disclosed embodiments can be understood and effected by those skilled in the art in practicing the claimed invention, from a study of the disclosure, and the appended claims. In the claims, the word "comprising" does not exclude other elements or steps, and the indefinite article "a" or "an" does not exclude a plurality. The mere fact that certain measures are recited in mutually different dependent claims does not indicate that a combination of these measures cannot be used to advantage.

[0058] B. Formula comprising casein glycomacropeptide (GMP) and free amino acids for the dietary management of phenylketonuria (PKU) or tyrosinaemia

[0059] In a first general embodiment, the invention provides a formula for children under the age of 2 years, preferably an infant formula, comprising intact casein glycomacropeptide (GMP) and free amino acids for use in the dietary management of Phenylketonuria (PKU) or tyrosinaemia.

[0060] In a second general embodiment, the invention provides the use of intact GMP and free amino acids in the manufacture of a formula for children under the age of 2 years, preferably an infant formula, for the dietary management of PKU or tyrosinaemia in an individual in need thereof. The infant formula may be classed as a medicament that may be used for treatment of a patient.

[0061] In a third general embodiment, the invention provides a method of providing dietary management of PKU or tyrosinaemia comprising administering a formula for children under the age of 2 years, preferably an infant formula, comprising intact GMP and free amino acids to an individual in need thereof.

[0062] These general embodiments can be readily combined with the specific compositions, patient populations and symptoms described below. These general embodiments for dietary management can be used prevent or treat PKU and tyrosinaemia in patient populations and symptoms described below.

[0063] Dietary Intervention / Dietary Management

[0064] The dietary intervention and management ensures good metabolic control of PKU and tyrosinaemia compared to before dietary intervention - that is target blood phenylalanine concentrations (120-360 pmol / L) are achieved for children under the age of 2 years with PKU and target blood tyrosine concentrations (200-400 pmol / L) are achieved for children under the age of 2 years with tyrosinaemia.

[0065] The dietary management ensures normal growth and development in children under the age of 2 years with PKU and tyrosinaemia compared to dietary management with an amino acid based formula without GMP - that is maintenance or increase in the percentiles for length for age, weight for age and / or body mass index (BMI) for age (compared to the WHO 50thpercentile for that age: WHO. Global Database on Child Growth and Malnutrition,' WHO: Geneva, Switzerland, 1997).

[0066] The dietary management ensures improved gastrointestinal tolerance compared to dietary management with an amino acid based formula without GMP - that is improved gastrointestinal tolerance as qualified by a caregiver questionnaire recording occurrence of vomiting, regurgitation, abdominal discomfort or colic symptoms, flatulence, number of daily stools, and a description of their characteristics [loose, soft, hard], any feeding difficulties (Yilmaz et al. (2023) Orphanet Journal of Rare Diseases 18, 16).

[0067] Therefore, in one embodiment the dietary management results in good metabolic control as compared to before dietary intervention, and / or normal growth and / or improved gastrointestinal tolerance as compared to dietary management with an amino acid based formula without GMP. In a specific embodiment, the dietary management results in good metabolic control. Preferably, the good metabolic control achieves target blood phenylalanine concentrations (120-360 pmol / L). This is particularly preferred for a child under the age of 2 years with PKU. Preferably, the good metabolic control achieves target blood tyrosine concentrations (200-400 pmol / L) for a child under the age of 2 years with tyrosinaemia. This is particularly preferred for a child under the age of 2 years with tyrosinaemia.

[0068] In a specific embodiment, the dietary management results normal growth for the child with PKU or tyrosinaemia compared to dietary management with an amino acid based formula without GMP. Preferably, the normal growth results in maintenance of length for age, weight for age and / or BMI for age compared to dietary management with an amino acid based formula without GMP. More preferably, the normal growth results in an increase of length for age, weight for age and / or BMI for age compared to dietary management with an amino acid based formula without GMP.

[0069] In a specific embodiment, the dietary management results in improved gastrointestinal tolerance compared to dietary management with an amino acid based formula without GMP, wherein the improved gastrointestinal tolerance is selected from the group consisting of reduced vomiting, regurgitation, abdominal discomfort or colic symptoms, flatulence, proportion or loose or hard stools, and / or difficulty feeding.

[0070] In the most preferred embodiment, the dietary management results in good metabolic control compared to the child before dietary intervention, as well as normal growth and improved gastrointestinal tolerance as compared to dietary management with an amino acid based formula without GMP.

[0071] The dietary management is generally continued until the infant or child reaches the age of 2 years. Preferably, the dietary management lasts at least 3 months, more preferably at least 6 months, even more preferably at least 1 year.

[0072] Given the presence of intact GMP and improved amino acid profile, it is predicted that the dietary intervention and management of the invention will result in improved metabolic control, normal growth and / or improved gastrointestinal tolerance as compared to standard free amino acid infant and follow-on formulae currently used for dietary intervention and management PKU and tyrosinaemia.

[0073] Diseases and their symptoms

[0074] Phenylketonuria Phenylketonuria (PKU; also known as phenylalanine hydroxylase (PAH, EC 1.14.16.1) deficiency (OMIM # 261600) and Folling disease) is primarily caused by pathogenetic variants in the PAH gene that result in an inborn error of Phe metabolism. PAH is responsible for the conversion of Phe to Tyr, in a reaction that requires the co-substrate tetrahydrobiopterin (BH4).

[0075] PKU (or the related but less severe condition hyperphenylalaninaemia) may also be caused by defects in tetrahydrobiopterin (BH4) metabolism or pathogenetic variants in DNAJC12.

[0076] Mutation that can cause PKU are known to the skilled person and can be confirmed by genetic testing.

[0077] In infants, PKU is diagnosed by detecting high Phe levels in the blood (i.e., >360 pmol / L), normally detected as part of a “heel prick test” in the first days or weeks after birth. Hyperphenylalaninaemia is an intermediate state between normal Phe levels in the blood (35-120 pmol / L) and PKU, where Phe levels in the blood of 120-360 pmol / L.

[0078] Therefore, in one embodiment the formula for a child aged under 2 years, preferably an infant formula, is for use in the dietary management of PKU.

[0079] Symptoms of phenylketonuria

[0080] If untreated, PKU leads to global intellectual disability, significant delays in developmental milestones, hyperactive behaviour with autistic features, seizures, eczema, musty body odour, and light pigmentation (eyes, hair, and skin), causing substantial lifelong disability (Camp et al. (2014) Molecular Genetics and Metabolism, 112, 87-122; Rocha and MacDonald (2018) Expert Opinion on Orphan Drugs, 6, 667-681).

[0081] Therefore, in one embodiment the formula is for use in the dietary management of a symptom selected from the group intellectual disability, delays in developmental milestones, hyperactive behaviour, seizures, eczema, musty body odour, and light pigmentation (e.g., of the eyes, hair, and / or skin) in a child with PKU.

[0082] If the infant is diagnosed with PKU via newborn screening (NBS) and starts the dietary management within 28 days from birth (preferably 21 days, more preferably 14 days, even more preferably 7 days), the symptoms may not be overt and may be prevented. If the infant or child is diagnosed with PKU at a later stage or the dietary management is not started within 28 days from birth (preferably 21 days, more preferably 14 days, even more preferably 7 days), the symptoms may be overt and may be treated or ameliorated. In a specific embodiment, the symptom is intellectual disability. For example, the Bayley Scale of Infant Development (3rded.) may be employed from birth to 42 months.

[0083] In a specific embodiment, the symptom is a delay in one or more developmental milestone. For example, anthropometry milestones such as length for age, weight for age and / or BMI may be measured. For example, developmental milestones known to the skilled paediatrician may be assessed individually or globally at a particular age. For example, development milestones at 2 months may be selected from the group consisting of turning head towards sounds; paying attention to faces; beginning to act bored (e.g., cries, fussy) if activity doesn’t change; and / or able to raise head. For example, development milestones at 4 months may be selected from the group consisting of beginning to babble; reaching for toy with one hand; holding head steady unsupported; and / or able to roll over from tummy to back. For example, development milestones at 6 months may be selected from the group consisting of responding to sounds by making sounds; beginning to sit without support; and / or looking around at nearby objects. For example, development milestones at 9 months may be selected from the group consisting of understanding the word “no”; playing games such as “peek-a-boo”; pointing at objects; and / or crawling. For example, development milestones at 12 months may be selected from the group consisting of saying simple words; starting to use objects correctly; pulling up to stand; and / or following simple directions.

[0084] In a specific embodiment, the symptom is hyperactive behaviour. For example, Rutter assessment may be used to predict hyperactivity in infants (Ostergaard et al. (2016 PLoS ONE, 11, eO 157352).

[0085] In a specific embodiment, the symptom is seizures.

[0086] In a specific embodiment, the symptom is eczema.

[0087] In a specific embodiment, the symptom is musty body odour (e.g., in the breath, skin and / or urine). This symptom is linked to high levels of Phe in saliva, sweat and / or urine, and may be detected by the parent, medical professional or caregiver. This symptom can be unpleasant and distressing for parents; it represents an overt sign of disease.

[0088] In a specific embodiment, the symptom is light pigmentation (e.g., of the eyes, hair, and / or skin). This symptom may be detected by the parent, medical professional or caregiver (with reference to the skin tone of parents and / or siblings) and can result in sensitivity to the sun.

[0089] It is apparent that more overt symptoms will become recognised the later the dietary management is initiated. Tyrosinaemia

[0090] Tyrosinaemia is an error of metabolism, usually inborn, in which the body cannot effectively break down the amino acid tyrosine. Due to its low prevalence, tyrosinaemia is often not included as part of a “heel prick test” NBS after birth. Therefore, the disease is often diagnosed when overt clinical symptoms occur. There are three recognised types linked to different genetic causes and with different symptoms.

[0091] Mutation that can confirm tyrosinaemia are known to the skilled person and are described further below.

[0092] In children, tyrosinaemia is diagnosed by detecting high Tyr levels in the blood (i.e., >200 pmol / L), generally after symptoms start. Symptoms normally become apparent at plasma levels >500 pmol / L. Target blood tyrosine concentrations are 200-400 pmol / L in patients. Moderately elevated Tyr levels in the blood are 120-200 pmol / L and normal Tyr levels in the blood are 20-120 pmol / L.

[0093] Therefore, in one embodiment, the formula is for use in the dietary management of tyrosinaemia.

[0094] It is apparent that more overt symptoms will become recognised if the dietary management is delayed.

[0095] Type I Tyrosinaemia

[0096] Type I tyrosinaemia is linked to pathogenic mutations in the FAH gene (expressing fumarylacetoacetase). It is associated with the substrate fumarylacetoacetate and is the most prevalent and severe form of tyrosinaemia. Build up of fumarylacetoacetate can result in toxicity to the kidney and liver as well as brain, eyes, heart and endocrine system. Lack of treatment of Type I tyrosinaemia can result in liver failure (Chinsky et al. (2017) Genetics in Medicine, 19, 1380). Whilst Type I tyrosinaemia may be treated with Nitisinone (NTBC), this in turn can increase tyrosine levels meaning that dietary management is still required and the patient may display symptoms of Type II tyrosinaemia, described below.

[0097] Therefore, in one embodiment, the formula is for use in the dietary management of Type I tyrosinaemia.

[0098] In one embodiment, the formula is for use in the dietary management of a symptom selected from liver toxicity, kidney toxicity, nervous system toxicity, and endocrine dysfunction in a child with Type I tyrosinaemia.

[0099] In a specific embodiment, the symptom is liver toxicity. For example, the symptom may be a haemophiliac-like symptom such as impaired clotting, increased bruising, bleeding inside the joins or bleeding inside the brain. For example, the symptom may be acute liver failure. For example, the symptom may be hypoalbuminemia. For example, the symptom may be hepatocarcinoma.

[0100] In a specific embodiment, the symptom is kidney toxicity. For example, the symptom may be hypophosphatemic rickets. For example, the symptom may be impaired kidney function. For example, the symptom may be renal tubular damage. For example, the symptom may be kidney failure.

[0101] In a specific embodiment, the symptom is neurotoxicity. For example, the symptom may be porphyria. For example, the symptom may be peripheral neuropathy.

[0102] In a specific embodiment, the symptom is endocrine dysfunction. For example, the symptom may be dopamine, norepinephrine and / or thyroxine dysfunction.

[0103] Symptoms associated with Type I tyrosinaemia may be treated with Nitisinone (NTBC). Therefore, the formula may be for use in the dietary management of Type I tyrosinaemia in a patient also treated with Nitisinone (NTBC).

[0104] Where the Nitisinone (NTBC) treatment is successful, the symptoms associated with Type I tyrosinaemia may be consistent with symptoms of Type II tyrosinaemia, described below.

[0105] Type II tyrosinaemia is linked to pathogenic mutations in the TAT gene (expressing tyrosine aminotransferase). Symptoms include eye toxicity, skin toxicity and mentally disabled.

[0106] Therefore, in one embodiment, the formula is for use in the dietary management of Type II tyrosinaemia.

[0107] In one embodiment, the formula is for use in the dietary management of a symptom selected from eye toxicity, skin toxicity and / or mentally disabled in a child with Type II tyrosinaemia (and / or a child with Type I tyrosinaemia treated with Nitisinone (NTBC)).

[0108] In a specific embodiment, the symptom is eye toxicity. For example, the symptom may be epiphora (excessive tearing). For example, the symptom may be photophobia (abnormal sensitivity to light). For example, the symptom may be eye pain. For example, the symptom may be hyperaemia (red eye).

[0109] In a specific embodiment, the symptom is skin toxicity. For example, the symptom may be palmoplantar hyperkeratosis (painful skin lesions).

[0110] In a specific embodiment, the symptom is mentally disabled. Type III Tyrosinaemia

[0111] Type III tyrosinaemia is linked to pathogenic mutations in the HPD gene (expressing 4-hydroxyphenylpyruvate dioxygenase). Symptoms include mental retardation, seizures, and intermittent ataxia (periodic loss of balance and coordination).

[0112] Therefore, in one embodiment, the formula is for use in the dietary management of Type III tyrosinaemia.

[0113] In one embodiment, the formula is for use in the dietary management of a symptom selected from mental retardation, seizures, and intermittent ataxia in a child with Type III tyrosinaemia.

[0114] In a specific embodiment, the symptom is mental retardation.

[0115] In a specific embodiment, the symptom is seizures.

[0116] In a specific embodiment, the symptom is intermittent ataxia.

[0117] Infant and child under the age of 2 years

[0118] Of the inborn errors of metabolism described herein, PKU is most prevalent, followed by Type I tyrosinaemia, then Type II tyrosinaemia and finally Type III tyrosinaemia.

[0119] The child is a child with an inborn error in Phe and / or Tyr metabolism, preferably PKU, Type I tyrosinaemia, Type II tyrosinaemia or Type III tyrosinaemia. Preferably, the child is a child with PKU, Type I tyrosinaemia or Type II tyrosinaemia. More preferably, the child is a child with PKU or Type I tyrosinaemia. Most preferably, the child is a child with PKU.

[0120] The child is preferably a human child, more preferably a child under the age of 2 years, even more preferably the child is an infant is up to the age of 1 year, most preferably an infant up to the age of 6 months.

[0121] Formula

[0122] The formula is suitable for consumption by a child under the age of 2 years.

[0123] The formula described herein may be in the form of a liquid, a reconstitutable powder or a spoonable food. Preferably, the formula is in the form of a liquid or a reconstitutable powder that can be administered to the child through a teated bottle in a liquid form. More preferably, the formula is in the form of a liquid. Even more preferably, the formula is a shelf-stable liquid. Most preferably, the formula is a packaged, shelf-stable liquid.

[0124] The formula described is preferably shelf stable when sealed in a container (i.e., packaged). In this context, stability means that, once sterilised, the formula is in a stable physical state (a liquid, a reconstitutable powder or a spoonable food) for an extended period of time. Moderate shaking or mixing may be required to ensure homogeneity or uniformity. The formula is preferably shelf stable for at least 3 months, preferably at least 6 months, more preferably at least 9 months and most preferably at least 1 year.

[0125] To ensure an optimal milk or single cream consistency, the liquid or reconstituted liquid formula may have a viscosity of below 200 mPa s, preferably of from 5 to 180 mPa s, preferably 20 to 150 mPa s, measured at 20° C at a shear rate of 100 s ' . The viscosity may be determined by methods known to a skilled person, e.g., by using a rheometer (Netzsch Kinexus Pro+) equipped with a plate / plate geometry (60 mm diameter) and 1 mm gap.

[0126] The density of the liquid or reconstituted liquid formula may be in the range of 1.02 to 1.2 g / L, preferably 1.03 to 1.25 g / L, more preferably 1.05 to 1.25 g / L.

[0127] The pH of the liquid or reconstituted liquid formula may be in the range of 7 to 8, preferably 7 to 7.5.

[0128] The formula preferably has a neutral flavour, optionally flavoured with a flavouring, such as vanilla. Flavourings are generally only used in compositions for a child older than 6 months. Therefore, when the formula is for an infant aged less than 6 months, the formula is preferably unflavoured.

[0129] The formula may have a caloric density of 0.6 to 3.0 kcal / mL when in a liquid form. Preferably, the formula may have a caloric density of 0.6 to 2.0 kcal / mL, more preferably 0.6 to 1.0 kcal / mL, when in a liquid form.

[0130] The formula contains a protein source, described in detail below. The protein may be present in an amount of 1 to 15 g / 100 mL, preferably 1 to 10 g / 100 mL.

[0131] The formula preferably has an amino acid balance that meets the minimum requirements in Table 36 of the “Protein and Amino Acid Requirements in Human Nutrition WHO / FAO Consultation 2007”. For an infant under the age of 1 year this includes at least: 20 mg / g His, 32 mg / g He, 66 mg / g Leu, 57 mg / g Lys, 28 mg / g sulphur amino acids (Cys + Met), 52 mg / g aromatic amino acids (Phe + Trp + Tyr), 31 mg / g Thr, 8.5 mg / g Trp and 43 mg / g Vai. For a child under 2 years, this includes at least: 18 mg / g His, 31 mg / g He, 63 mg / g Leu, 52 mg / g Lys, 26 mg / g sulphur amino acids (Cys + Met), 46 mg / g aromatic amino acids (Phe + Trp + Tyr), 27 mg / g Thr, 7.4 mg / g Trp and 42 mg / g Vai.

[0132] The formula may contain a lipid source. The lipid may be from animal, vegetable and / or microbial sources. Preferably, the lipid comprises a fat selected from the group consisting of milk fat, sunflower oil, rapeseed oil, low erucic acid rapeseed oil, safflower oil, canola oil, olive oil, coconut oil, palm kernel oil, soybean oil, fish oil, palm oleic, high oleic sunflower oil and high oleic safflower oil, and microbial fermentation oil containing long chain, polyunsaturated fatty acids. Preferably, the fat comprises docosahexaenoic acid and / or arachidonic acid.

[0133] The formula may contain a digestible carbohydrate source. Preferred digestible carbohydrate are selected from the group consisting of lactose, glucose, sucrose, fructose, galactose, maltose, starch and maltodextrin.

[0134] The formula may contain a vitamin source. Vitamins may comprise Vitamin A, Vitamin D3, Vitamin E, Vitamin C, Vitamin K, Thiamin (Bl), Riboflavin (B2), Niacin (B3), Vitamin B6, folic acid, Vitamin B12, Biotin and / or Pantothenic acid. Preferably, the vitamin source comprises Vitamin A, Vitamin D3, Vitamin E, Vitamin C, Vitamin K, Thiamin (Bl), Riboflavin (B2), Niacin (B3), Vitamin B6, folic acid, Vitamin B12, Biotin and Pantothenic acid.

[0135] The formula may contain a mineral source. Minerals may comprise sodium, potassium, chloride, fluoride, calcium, phosphorus, magnesium, iron, copper, zinc manganese, iodine, molybdenum, selenium, and / or chromium. Preferably, the mineral source comprises sodium, potassium, chloride, fluoride, calcium, phosphorus, magnesium, iron, copper, zinc manganese, iodine, molybdenum, selenium, and chromium.

[0136] The formula preferably comprises protein, fat, carbohydrate, vitamins and minerals. More preferably, the formula is a complete formula.

[0137] The formula may contain other nutrients selected from the group consisting of choline, taurine, L-carnitine and inositol.

[0138] The formula may contain other substances with beneficial effects selected from the group consisting of prebiotics, probiotics, fibres and / or nucleotides.

[0139] The formula may contain thickeners. Preferably, the thickener is selected from the group consisting of guar gum, locust bean gum, xanthan gum and pectins. More preferably, the thickener is xanthan gum.

[0140] The formula may contain additives. Preferably, the additive is selected from the group consisting of starches and carrageenan.

[0141] The formula may contain emulsifiers. Preferably, the emulsifier is selected from the group consisting of lecithins, mono- and di -glycerides, and citric and fatty acid esters of glycerol. More preferably, the emulsifier is mono- and di-glycerides.

[0142] The formula may contain acidity regulators. Preferably, the acidity regulator is selected from the group consisting of sodium hydrogen carbonate, sodium carbonate, potassium hydroxide, potassium hydrogen carbonate, potassium carbonate, calcium hydroxide, L(+) lactic acid, citric acid, sodium dihydrogen citrate, trisodium citrate, potassium citrate, sodium dihydrogen phosphate, disodium hydrogen phosphate, trisodium phosphate, potassium dihydrogen phosphate, dipotassium hydrogen phosphate and tripotassium phosphate.

[0143] The formula may contain antioxidants. Preferably, the antioxidant is selected from the group consisting of tocopherols and ascorbyl palmitate.

[0144] The formula may be a follow-on or an infant formula, preferably an infant formula.

[0145] Infant Formula

[0146] The infant formula may be administered to weaned infants who do not use the formula for complete nutrition, the infant formula may preferably provide 1.0 to 3.5 g / 100 kcal of protein, 3.0 to 8.0 g / 100 kcal of fat and 6.0 to 20.0 g / 100 kcal of digestible carbohydrate. As the infant formula may be intended as a Food for Special Medical Purpose, it more preferably provides 1.8 to 3.0 g / 100 kcal of protein, 4.4 to 6.0 g / 100 kcal of fat and 9.0 to 14.0 g / 100 kcal of digestible carbohydrate.

[0147] The infant may be in the form of a liquid, a reconstitutable powder or a spoonable food. Preferably, the infant formula is in the form of a liquid or a reconstitutable powder (e.g., that can be administered to the infant through a teated bottle in a liquid form). More preferably, the infant formula is a liquid. Even more preferably, the infant formula is a shelfstable liquid. The shelf-stable liquid infant formula is preferably shelf stable for at least 3 months, preferably at least 6 months, more preferably at least 9 months and most preferably at least 1 year. Shelf-stable liquid are generally stored in sealed containers.

[0148] The liquid infant formula may have an energy density of 60 to 75 kcal / 100 mL.

[0149] The protein source typically provides 7 to 15% of the total calories of the infant formula. Preferably, the liquid infant formula comprises a protein source that provides 9 to 13% of the total calories. More preferably, the liquid infant formula comprises a protein source that provides 10 to 12% of the total calories.

[0150] The protein concentration in the infant formula is determined by the sum of protein, peptides and free amino acids. Based on dry weight, the infant formula preferably comprises from 10 to 20 wt% protein, more preferably from 12 to 19 wt%, even more preferably 14 to 18 wt%. Based on a liquid or reconstituted infant formula, the infant formula preferably comprises from 1 to 2.5 g protein / 100 mL, more preferably of 1.2 to 2.3 g / 100 mL, even more preferably of 1.5 to 2.1 g / 100 mL. Protein source

[0151] The formula comprises a mixture of GMP and free amino acids in certain proportions as a source of protein. The protein fraction includes all proteins, peptides and amino acids within the formula.

[0152] The formula may comprise GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1. Preferably, the formula comprises GMP to free amino acids by weight in a proportion of 1 :3 to 3 : 1. More preferably, the formula comprises GMP to free amino acids by weight in a proportion of 1 :2 to 2: 1. In one embodiment, formula comprises GMP to free amino acids by weight in a proportion of about 1 : 1. In a preferred embodiment, the protein fraction consists of a mixture of GMP and free amino acids.

[0153] For the infant formula, smaller proportions of GMP to free amino acids are favoured, so GMP to free amino acids by weight in a proportion of 1 :4 to 1 :2 is especially preferred. In spite of the high proportion of free amino acids, these infant formulae had a neutral taste when combined with the free amino acid mixture described below.

[0154] Therefore, in a fourth general embodiment, the invention provides a complete infant formula for the dietary management of PKU or tyrosinaemia comprising intact GMP and free amino acids, wherein the GMP to free amino acids by weight are present in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2: 1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2.

[0155] The complete infant formula for the dietary management of PKU or tyrosinaemia is particularly effective when the free amino acids are based on the free amino acid mixture described below.

[0156] Free amino acid mixture

[0157] It is well known that free amino acids can impart bitter and / or acidic flavours on formulae and food. The specific amino acid mixtures for use in the infant formula described above and below displayed a surprisingly neutral flavour.

[0158] Therefore, in a fifth general embodiment, the invention provides a mixture of free amino acids comprising Arg, Asp+Asn, Cys, Gly, His, He, Leu, Lys, Met, Trp and Vai, and optionally Tyr, wherein the mixture of free amino acids contains less than 0.1 wt% of any of Ala, Gin, Glu, Pro, Phe, Ser or Thr.

[0159] The mixture of free amino acids may be used in the formula described above and below. In a preferred embodiment, the mixture of free amino acids comprises as a total percentage by weight of free amino acids in the mixture: Arg (9 to 13 wt%), Asp+Asn (8 to 12 wt% total), Cys (2 to 6 wt%), Gly (12 to 16 wt%), His (4 to 8 wt%), He (2 to 6 wt%), Leu (13 to 17 wt%), Lys (5 to 9 wt%), Met (0.5 to 4 wt%), Trp (1 to 5 wt%), Vai (5 to 9%), and Tyr (13 to 17 wt%). This mixture of free amino acids is particular suitable for combining with GMP (preferably with GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2: 1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2.) to produce an infant formula, preferably for use in the dietary management of PKU.

[0160] In a more preferred embodiment, the mixture of free amino acids comprises as a total percentage by weight of free amino acids in the mixture: Arg (10 to 12 wt%), Asp+Asn (9 to 11 wt% total), Cys (3 to 5 wt%), Gly (13 to 15 wt%), His (5 to 7 wt%), He (3 to 5 wt%), Leu (14 to 16 wt%), Lys (6 to 8 wt%), Met (1 to 3 wt%), Trp (2 to 4 wt%), Vai (6 to 8%), and Tyr (14 to 16 wt%). This mixture of free amino acids is particular suitable for combining with GMP (preferably with GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2: 1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2.) to produce an infant formula, preferably for use in the dietary management of PKU.

[0161] In another preferred embodiment, the mixture of free amino acids comprises as a total percentage by weight of free amino acids in the mixture: Arg (11 to 15 wt%), Asp+Asn (10 to 14 wt% total), Cys (2 to 6 wt%), Gly (15 to 19 wt%), His (5 to 9 wt%), He (3 to 7 wt%), Leu (16 to 20 wt%), Lys (6 to 10 wt%), Met (0.5 to 4 wt%), Trp (2 to 6 wt%) and Vai (7 to 11%). This mixture of free amino acids is particular suitable for combining with GMP (preferably with GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2: 1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2.) to produce an infant formula, preferably for use in the dietary management of tyrosinaemia.

[0162] In another more preferred embodiment, the mixture of free amino acids comprises as a total percentage by weight of free amino acids in the mixture: Arg (12 to 14 wt%), Asp+Asn (11 to 13 wt% total), Cys (3 to 5 wt%), Gly (16 to 18 wt%), His (6 to 8 wt%), He (4 to 6 wt%), Leu (17 to 19 wt%), Lys (7 to 9 wt%), Met (1 to 3 wt%), Trp (3 to 5 wt%) and Vai (8 to 10%). This mixture of free amino acids is particular suitable for combining with GMP (preferably with GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2: 1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2.) to produce an infant formula, preferably for use in the dietary management of tyrosinaemia.

[0163] At a limiting extreme, the mixture of free amino acids may consist of the amino acids described above, such that less than 0.2 wt% of any other individual amino acid may be present.

[0164] The mixture of free amino acids may be used in the formula and dietary intervention / management described above and below.

[0165] C. Formulations

[0166] Shelf-stable liquid and solid infant formulae suitable for children under 1 year of age

[0167] Table 1: Batch scale production of dry ingredients for a liquid infant formula (or a powdered infant formula produced therefrom) according to the invention is shown below:

[0168] A powdered infant formula can be produced by mixing the ingredients listed above using methods know to in the art. If wet mixed, it can be dried for use. To produce a liquid infant formula, the ingredients or powdered infant formula (from 10 and 20 g, preferably 13 to 15 g) is reconstituted in water (100 mL) with shaking to form a homogeneous liquid emulsion. Optionally, the emulsion can be pasteurised (e.g., 86 °C for 30 seconds). The emulsion can be sterilised (e.g., UHT sterilised at 142.5 °C for 5 seconds) to form a shelfstable liquid emulsion, which, once packaged, is stable for at least 9 months, has a neural flavour with the consistency of milk to single cream and a density of approximately 1.05 g / L. 13.4 g of the mixed ingredients listed in Table 1 is reconstituted (13.4% solids; 86.6% water) to produce the liquid infant formula of Table 2. Table 2: An example liquid infant formula according to the invention is shown below:

[0169] The final sterilised product had a pH of 7.1 to 7.2, a density of 1.03, and a viscosity of 15 to 35 mPa.s (10 s’1@20°C).

[0170] The ingredients or powdered infant formula are diluted in water in amounts from 10 to 20 g / 100 mL, preferably 13 to 15 g / 100 mL, homogenised and sterilised (e.g., by direct steam injection). Alternatively, the powdered formula is reconstituted for direct use. The liquid infant formula may be packaged in sterile containers (stable for up to 1 year) or dried (e.g., spray dried) and packaged in sterile containers (stable for up to 2 years) to be reconstituted on demand. The liquid infant formula has a neural flavour with the consistency of milk to single cream and a density of approximately 1.05 g / L.

[0171] The infant formula contains 38% protein by weight of GMP and 62% protein by weight of free amino acids. The final amino acid profile (for the combined GMP and free amino acids) is: Ala (2.3%), Arg (7.2%), Asp+Asn (9.3%), Cys (2.4%), Glu+Gln (7.1%), Gly (9.4%), His (3.8%), He (6.7%), Leu (10.4%), Lys (6.5%), Met (1.8%), Pro (4.4%) Ser (2.8%), Thr (6.3%), Trp (2.1%), Tyr (9.8%) Vai (7.7%) and Phe (0.1%).

[0172] Modification of the amino acid by minimising tyrosine results in an equivalent formula suitable for an individual or a patient with PKU and / or tyrosinaemia.

[0173] D. Clinical Trial

[0174] Study design

[0175] A 2-part study in full-term infants diagnosed with PKU and identified by newborn screening. Infants are recruited from specialist Phenylketonuria and Tyrosinaemia Centres. Inclusion criteria include infants diagnosed with PKU by newborn screening and continuously treated with a low-protein diet supplemented with phenylalanine-free and / or tyrosine-free infant amino acid formula. Infants < 4 weeks or > 1 year of age, diagnosed with hyperphenylalaninaemia (blood phenylalanine concentrations < 360 pmol / L when untreated) or mild tyrosinaemia (blood phenylalanine concentrations < 600 pmol / L when untreated), or with a co-morbidity such as diabetes are excluded. Infants already established on solids are included in the study.

[0176] Part I: a 28-day short-term acceptability and tolerance study, infants replace their usual daily doses of infant amino acid formula (e.g., PKU Anamix Infant: Nutricia, or PKU Start: Vitaflo) with the same amount of study formula (based on shelf-stable liquid infant formula for children under 1 year of age in Table 2). Subject demographics are recorded at baseline, including PKU classification according to pre-treatment blood phenylalanine levels (classical PKU [> 1200 pmol / L]), moderate PKU [600-1200 pmol / L], or mild PKU [360— 600 pmol / L]), tyrosinaemia classification according to pre-treatment blood tyrosine levels (classical tyrosinaemia [> 1200 pmol / L]), moderate tyrosinaemia [600-1200 pmol / L], or mild tyrosinaemia [360-600 pmol / L]), infant’s gestational age, sex, ethnicity, relevant medical history, and any concurrent medications. Information on the usual infant amino acid formula (dose, type, daily volume, and the number of feeds each day), source and amount of natural protein (breast milk or standard infant formula), and time of commencement of solid food intake is recorded from pre-baseline. Prior to baseline and during part I (28-day study), parents / caregivers complete daily questionnaires to record gastrointestinal tolerance / intolerance: occurrence of vomiting, regurgitation, abdominal discomfort or colic symptoms, flatulence, number of daily stools, and a description of their characteristics [loose, soft, hard], any feeding difficulties, and concomitant medications, as well as disease symptoms. The last three retrospective blood phenylalanine and / or tyrosine concentrations, weight, length, dietary energy, and total protein intake are collected at baseline and week 4. At the end of 4 weeks, parents describe the ease of preparation and dissolving properties of the study formula.

[0177] Part II: an extended follow-up study is conducted with infants who continue to use the study formula for 12 months or until the study formula is discontinued. Dietary prescriptions (energy, total protein and natural protein intake, protein equivalent from protein substitute intake) and anthropometric measurements (weight and length) are documented by the dietitians at each clinic visit. Routine blood phenylalanine and / or tyrosine concentrations (twice weekly / weekly) are also recorded. Any relevant medications and gastrointestinal problems are reported by parents / caregivers.

[0178] Prevalence of symptoms described above are also monitored throughout the trial.

[0179] Study formula

[0180] The study formula (based on shelf-stable liquid infant formula for children under 1 year of age in Table 2) is an GMP and amino acid-based powdered phenylalanine-free infant formula containing essential and non-essential amino acids (but low levels of phenylalanine), carbohydrates, fat, vitamins, minerals, trace elements, and other nutrients described on Table 1, diluted at 13 to 15 g / 100 mL in water, homogenised, pasteurised and sterilised. It is provided as a shelf-stable ready -to-drink formula with 2 g / 100 ml of protein.

[0181] Stool characteristics, gastrointestinal symptoms and medications

[0182] Data on gastrointestinal symptoms (constipation, abdominal discomfort, colic, vomiting, regurgitation, flatulence), any feeding difficulties, and concomitant medications are collected for three days pre-baseline, 28-day in the short-term evaluation period, and at each clinic visit during the 12-month follow-up. Parents / caregivers record stool characteristics (three categories: soft, loose, and hard) pre-baseline and 28-day in the short-term evaluation period.

[0183] Anthropometry Length is measured using a Holtain Harpenden infantometer (Holtain Ltd., Crymych, UK), head circumference is measured by tape measure and weight on calibrated digital baby scales (Seca, Medical Measuring Systems and Scales, Model 875, UK). Weight is measured to the nearest 0.1 g and length and circumference to the nearest 0.1 cm. Data are converted to age-based z-scores for weight, length, head circumference, and body mass index (BMI) according to WHO / UK growth definitions (WMGRS Group (2006) Acta Paediatrica 95,76- 85).

[0184] Blood phenylalanine levels

[0185] Weekly / twice weekly morning fasting heel prick blood spots for phenylalanine and / or tyrosine are collected on filter cards, Perkin Elmer 226 (UK Standard NBS) by caregivers at home. The number of fasting hours depends on the number of infant night feeds and the interval between feeds. All caregivers receive blood spot training from a specialist nurse. All blood spot samples are sent by first class post to the hospital laboratories for phenylalanine and / or tyrosine analysis. The cards have a standard thickness, and the blood phenylalanine and / or tyrosine concentrations are calculated on a 3.2 mm punch by MS / MS tandem mass spectrometry.

[0186] Dietary assessment

[0187] Dietary intake is determined by a 24-h recall recorded at baseline, week 4, and at each clinic visit during a 12-month long-term follow-up. Prescribed energy, total protein, natural protein, and protein equivalent from protein substitutes are assessed using the computer software, e.g., nutritional analysis program Nutritics (Nutritics, R. Edition (v5. 09) [Computer Software]).

[0188] Symptomatic effects

[0189] Prevalence of symptoms described above are also monitored throughout the trial.

[0190] Statistical analysis

[0191] Sample size calculations are not performed due to the rarity of the conditions and the exploratory nature of the study. Only descriptive statistics are used to present the results of this study.

[0192] Ethical approval The study will be approved by a recognised Research Ethics Committee. Written informed consent will be obtained for all subjects from at least one caregiver with parental responsibility. Results

[0193] It is predicted that the formula will be well-tolerated, will help maintain good metabolic control, and normal growth in infants with PKU and tyrosinaemia. Good metabolic control is predicted to result in good dietary management of the diseases and their symptoms.

[0194] EMBODIMENTS OF THE INVENTION:

[0195] Embodiment 1. A formula for children under the age of 2 years comprising intact casein glycomacropeptide and free amino acids for use in the dietary management of phenylketonuria or tyrosinaemia in an individual in need thereof.

[0196] Embodiment 2. The formula for use according to embodiment 1, wherein the dietary management results in good metabolic control, normal growth and / or improved gastrointestinal tolerance.

[0197] Embodiment 3. The formula for use according to embodiment 1, wherein the dietary management results in good metabolic control, normal growth and improved gastrointestinal tolerance.

[0198] Embodiment 4. The formula for use according to embodiment 2 or embodiment 3, wherein the good metabolic control achieves blood phenylalanine concentrations of 120-360 pmol / L in an individual with phenylketonuria or blood tyrosine concentrations of 200- 400 pmol / L in an individual with tyrosinaemia.

[0199] Embodiment 5. The formula for use according to embodiment 2 or embodiment 3, wherein the normal growth results in maintenance, preferably an increase, of length for age, weight for age and / or BMI for age.

[0200] Embodiment 6. The formula for use according to embodiment 2 or embodiment 3, wherein the improved gastrointestinal tolerance is selected from the group consisting of reduced vomiting, regurgitation, abdominal discomfort or colic symptoms, flatulence, proportion or loose or hard stools, and / or difficulty feeding.

[0201] Embodiment 7. The formula for use according to any one of the previous embodiments, wherein the use is for the dietary management of phenylketonuria. Embodiment 8. The formula for use according to embodiment 7, wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of intellectual disability, delays in developmental milestones, hyperactive behaviour, seizures, eczema, musty body odour, and light pigmentation (e.g., of the eyes, hair, and / or skin).

[0202] Embodiment 9. The formula for use according to embodiment 8, wherein the symptom is intellectual disability.

[0203] Embodiment 10. The formula for use according to embodiment 8, wherein the symptom is a delay in one or more developmental milestone, preferably anthropometry milestones including length for age, weight for age and / or BMI; and / or development milestones at 2 months, 4 months, 6 months, 9 months and / or 12 months.

[0204] Embodiment 11. The formula for use according to embodiment 8, wherein the symptom is hyperactive behaviour.

[0205] Embodiment 12. The formula for use according to embodiment 8, wherein the symptom is seizures.

[0206] Embodiment 13. The formula for use according to embodiment 8, wherein the symptom is eczema.

[0207] Embodiment 14. The formula for use according to embodiment 8, wherein the symptom is musty body odour.

[0208] Embodiment 15. The formula for use according to embodiment 8, wherein the symptom is light pigmentation.

[0209] Embodiment 16. The formula for use according to any one of embodiment 1 to 6, wherein the use is for the dietary management of tyrosinaemia.

[0210] Embodiment 17. The formula for use according to embodiment 16, wherein the use is for the dietary management of Type I tyrosinaemia.

[0211] Embodiment 18. The formula for use according to embodiment 17, wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of liver toxicity, kidney toxicity, nervous system toxicity, and endocrine dysfunction.

[0212] Embodiment 19. The formula for use according to embodiment 18, wherein the symptom is liver toxicity; preferably selected from the group consisting of a haemophiliaclike symptom, such as impaired clotting, increased bruising, bleeding inside the joins or bleeding inside the brain; acute liver failure; hypoalbuminemia and hepatocarcinoma.

[0213] Embodiment 20. The formula for use according to embodiment 18, wherein the symptom is kidney toxicity; preferably selected from the group consisting of hypophosphatemic rickets; impaired kidney function; renal tubular damage; and kidney failure.

[0214] Embodiment 21. The formula for use according to embodiment 18, wherein the symptom is neurotoxicity; preferably selected from the group consisting of porphyria; and peripheral neuropathy.

[0215] Embodiment 22. The formula for use according to embodiment 18, wherein the symptom is endocrine dysfunction; preferably selected from the group consisting of dopamine dysfunction; norepinephrine dysfunction; and thyroxine dysfunction. Embodiment 23. The formula for use according to embodiment 16, wherein the use is for the dietary management of Type II tyrosinaemia, and / or Type I tyrosinaemia treated with Nitisinone (NTBC).

[0216] Embodiment 24. The formula for use according to embodiment 23, wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of eye toxicity, skin toxicity and mentally disabled.

[0217] Embodiment 25. The formula for use according to embodiment 24, wherein the symptom is eye toxicity; preferably selected from the group consisting of epiphora; photophobia; eye pain; and hyperaemia.

[0218] Embodiment 26. The formula for use according to embodiment 24, wherein the symptom is skin toxicity; preferably palmoplantar hyperkeratosis.

[0219] Embodiment 27. The formula for use according to embodiment 24, wherein the symptom is mentally disabled.

[0220] Embodiment 28. The formula for use according to embodiment 16, wherein the use is for the dietary management of Type III tyrosinaemia.

[0221] Embodiment 29. The formula for use according to embodiment 28, wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of mental retardation, seizures, and intermittent ataxia.

[0222] Embodiment 30. The formula for use according to embodiment 29, wherein the symptom is mental retardation.

[0223] Embodiment 31. The formula for use according to embodiment 29, wherein the symptom is seizures.

[0224] Embodiment 32. The formula for use according to embodiment 29, wherein the symptom is intermittent ataxia.

[0225] Embodiment 33. The formula for use according to any one of the previous embodiments, wherein the individual is a child under the age of 2 years, preferably the child is an infant, more preferably an infant up to the age of 6 months.

[0226] Embodiment 34. The formula for use according to any one of the previous embodiments, wherein the dietary management lasts at least 3 months, preferably at least 6 months, more preferably at least 1 year. Embodiment 35. The formula for use according to any one of the previous embodiment, wherein the formula is in the form of a liquid, a reconstitutable powder or a spoonable food; preferably in the form of a liquid or a reconstitutable powder.

[0227] Embodiment 36. The formula for use according to embodiment 35, wherein the formula is in the form of a liquid; more preferably in the form of a shelf-stable liquid; even more preferably a packaged, shelf-stable liquid.

[0228] Embodiment 37. The formula for use according to embodiment 36, wherein the formula has a viscosity of below 200 mPa s, preferably of from 5 to 180 mPa s, preferably 20 to 150 mPa s, measured at 20° C at a shear rate of 100 s '.

[0229] Embodiment 38. The formula for use according to any one of embodiments 35 to 37, wherein the formula is in the form of a liquid or a spoonable food, and has a density in the range of 1.02 to 1.2 g / L, preferably 1.05 to 1.25 g / L.

[0230] Embodiment 39. The formula for use according to any one of embodiments 35 to 38, wherein the formula is in the form of a liquid or a spoonable food, and has a caloric density of 0.6 to 3.0 kcal / mL, preferably 0.6 to 2.0 kcal / mL, more preferably 0.6 to 1.0 kcal / mL. Embodiment 40. The formula for use according to any one of embodiments 35 to 39, wherein the formula is in the form of a liquid or a spoonable food, and protein is present in an amount of 1 to 15 g / 100 mL, preferably 1 to 10 g / 100 mL.

[0231] Embodiment 41. The formula for use according to any one of the previous embodiments, wherein the formula comprises GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1, preferably in a proportion of 1 :3 to 3 : 1, more preferably in a proportion of 1 :2 to 2: 1.

[0232] Embodiment 42. The formula for use according to any one of the preceding embodiments, wherein the formula contains a lipid source, preferably the lipid source comprises a fat selected from the group consisting of milk fat, sunflower oil, rapeseed oil, low erucic acid rapeseed oil, safflower oil, canola oil, olive oil, coconut oil, palm kernel oil, soybean oil, fish oil, palm oleic, high oleic sunflower oil and high oleic safflower oil, and microbial fermentation oil containing long chain, polyunsaturated fatty acids, more preferably, the lipid source comprises docosahexaenoic acid and / or arachidonic acid. Embodiment 43. The formula for use according to any one of the preceding embodiments, wherein the formula contains a digestible carbohydrate source, preferably selected from the group consisting of lactose, glucose, sucrose, fructose, galactose, maltose, starch and maltodextrin. Embodiment 44. The formula for use according to any one of the preceding embodiments, wherein the formula contains a vitamin source, preferably comprising Vitamin A, Vitamin D3, Vitamin E, Vitamin C, Vitamin K, Thiamin (Bl), Riboflavin (B2), Niacin (B3), Vitamin B6, folic acid, Vitamin B 12, Biotin and / or Pantothenic acid.

[0233] Embodiment 45. The formula for use according to any one of the preceding embodiments, wherein the formula contains a mineral source, preferably comprising sodium, potassium, chloride, calcium, phosphorus, magnesium, iron, copper, zinc manganese, iodine, molybdenum, selenium, and / or chromium.

[0234] Embodiment 46. The formula for use according to any one of the preceding embodiments, wherein the formula comprises protein, fat, carbohydrate, vitamins and minerals.

[0235] Embodiment 47. The formula for use according to any one of the preceding embodiments, wherein the formula comprises prebiotics, probiotics, fibres and / or nucleotides; and / or choline, taurine, L-carnitine and inositol.

[0236] Embodiment 48. The formula for use according to any one of the preceding embodiments, wherein the formula comprises one or more thickener, additive, emulsifier, acidity regulators and / or antioxidants.

[0237] Embodiment 49. The formula for use according to any one of the preceding embodiments, wherein the formula is an infant formula, preferably wherein the infant formula provides 1.8 to 3.0 g / 100 kcal of protein, 4.4 to 6.0 g / 100 kcal of fat and 9.0 to 14.0 g / 100 kcal of digestible carbohydrate.

[0238] Embodiment 50. The infant formula for use according to embodiment 49, wherein the infant formula is a liquid and has an energy density of 60 to 75 kcal / 100 mL.

[0239] Embodiment 51. Use of intact GMP and free amino acids in the manufacture of a formula for children under the age of 2 years, preferably an infant formula, for the dietary management of PKU or tyrosinaemia in an individual in need thereof.

[0240] Embodiment 52. A method of providing dietary management of PKU or tyrosinaemia comprising administering a formula for children under the age of 2 years, preferably an infant formula, comprising intact GMP and free amino acids to an individual in need thereof.

[0241] Embodiment 53. A complete infant formula for the dietary management of PKU or tyrosinaemia comprising intact GMP and free amino acids, wherein the GMP to free amino acids by weight are present in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2: 1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2. Embodiment 54. A mixture of free amino acids comprising Arg, Asp+Asn, Cys, Gly, His, He, Leu, Lys, Met, Trp and Vai, and optionally Tyr, wherein the mixture of free amino acids contains less than 0.1 wt% of any of Ala, Gin, Glu, Pro, Phe, Ser or Thr.

[0242] Embodiment 55. A mixture of free amino acids according to embodiment 54, comprising as a total percentage by weight of free amino acids in the mixture: Arg (9 to 13 wt%), Asp+Asn (8 to 12 wt% total), Cys (2 to 6 wt%), Gly (12 to 16 wt%), His (4 to 8 wt%), He (2 to 6 wt%), Leu (13 to 17 wt%), Lys (5 to 9 wt%), Met (0.5 to 4 wt%), Trp (1 to 5 wt%), Vai (5 to 9%), and Tyr (13 to 17 wt%); preferably: Arg (10 to 12 wt%), Asp+Asn (9 to 11 wt% total), Cys (3 to 5 wt%), Gly (13 to 15 wt%), His (5 to 7 wt%), He (3 to 5 wt%), Leu (14 to 16 wt%), Lys (6 to 8 wt%), Met (1 to 3 wt%), Trp (2 to 4 wt%), Tyr (14 to 16 wt%) and Vai (6 to 8%).

[0243] Embodiment 56. A mixture of free amino acids according to embodiment 54, comprising as a total percentage by weight of free amino acids in the mixture: Arg (12 to 16 wt%), Asp+Asn (10 to 14 wt% total), Cys (2 to 6 wt%), Gly (15 to 19 wt%), His (5 to 9 wt%), He (3 to 7 wt%), Leu (16 to 20 wt%), Lys (6 to 10 wt%), Met (0.5 to 4 wt%), Trp (2 to 6 wt%) and Vai (7 to 11%); preferably: Arg (12 to 14 wt%), Asp+Asn (11 to 13 wt% total), Cys (3 to 5 wt%), Gly (16 to 18 wt%), His (6 to 8 wt%), He (4 to 6 wt%), Leu (17 to 19 wt%), Lys (7 to 9 wt%), Met (1 to 3 wt%), Trp (3 to 5 wt%) and Vai (8 to 10%).

[0244] Embodiment 57. A complete infant formula for use in the dietary management of PKU or tyrosinaemia comprising intact GMP and free amino acids according to embodiment 53, wherein the free amino acids consist of the mixture of free amino acids of any one of embodiments 54 to 56.

[0245] Embodiment 58. A complete infant formula comprising intact GMP and free amino acids according to embodiment 53, wherein the free amino acids consist of the mixture of free amino acids of embodiment 55 for use in the dietary management of phenylketonuria in an individual in need thereof.

[0246] Embodiment 59. A complete infant formula comprising intact GMP and free amino acids according to embodiment 53, wherein the free amino acids consist of the mixture of free amino acids of embodiment 56 for use in the dietary management of tyrosinaemia in an individual in need thereof.

[0247] Embodiment 60. Use of intact GMP and free amino acids in the manufacture of a formula for children under the age of 2 years, preferably an infant formula, for the dietary management of PKU or tyrosinaemia in an individual in need thereof, wherein the formula is according to any one of embodiments 35 to 50, 53, 58 or 59.

[0248] Embodiment 61. A method of providing dietary management of PKU or tyrosinaemia comprising administering a formula for children under the age of 2 years, preferably an infant formula, comprising intact GMP and free amino acids to an individual in need thereof, wherein the formula is according to any one of embodiments 35 to 50, 53, 58 or 59.

[0249] Embodiment 62. A formula for children under the age of 2 years comprising intact casein glycomacropeptide and free amino acids for use in the treatment or prevention of phenylketonuria or tyrosinaemia in a patient in need thereof.

[0250] Embodiment 63. The formula for use according to embodiment 62, wherein the treatment or prevention results in good metabolic control, normal growth and / or improved gastrointestinal tolerance.

[0251] Embodiment 64. The formula for use according to embodiment 62, wherein the treatment or prevention results in good metabolic control, normal growth and improved gastrointestinal tolerance.

[0252] Embodiment 65. The formula for use according to embodiment 63 or embodiment 64, wherein the good metabolic control achieves blood phenylalanine concentrations of 120-360 pmol / L in a patient with phenylketonuria or blood tyrosine concentrations of 200- 400 pmol / L in a patient with tyrosinaemia.

[0253] Embodiment 66. The formula for use according to embodiment 63 or embodiment 64, wherein the normal growth results in maintenance, preferably an increase, of length for age, weight for age and / or BMI for age.

[0254] Embodiment 67. The formula for use according to embodiment 63 or embodiment 64, wherein the improved gastrointestinal tolerance is selected from the group consisting of reduced vomiting, regurgitation, abdominal discomfort or colic symptoms, flatulence, proportion or loose or hard stools, and / or difficulty feeding.

[0255] Embodiment 68. The formula for use according to any one of the previous embodiments 62 to 67, wherein the use is for the treatment or prevention of phenylketonuria.

[0256] Embodiment 69. The formula for use according to embodiment 68, wherein the treatment or prevention results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of intellectual disability, delays in developmental milestones, hyperactive behaviour, seizures, eczema, musty body odour, and light pigmentation (e.g., of the eyes, hair, and / or skin). Embodiment 70. The formula for use according to embodiment 69, wherein the symptom is intellectual disability.

[0257] Embodiment 71. The formula for use according to embodiment 69, wherein the symptom is a delay in one or more developmental milestone, preferably anthropometry milestones including length for age, weight for age and / or BMI; and / or development milestones at 2 months, 4 months, 6 months, 9 months and / or 12 months.

[0258] Embodiment 72. The formula for use according to embodiment 69, wherein the symptom is hyperactive behaviour.

[0259] Embodiment 73. The formula for use according to embodiment 69, wherein the symptom is seizures.

[0260] Embodiment 74. The formula for use according to embodiment 69, wherein the symptom is eczema.

[0261] Embodiment 75. The formula for use according to cl embodiment aim 69, wherein the symptom is musty body odour.

[0262] Embodiment 76. The formula for use according to embodiment 69, wherein the symptom is light pigmentation.

[0263] Embodiment 77. The formula for use according to any one of embodiments 62 to 67, wherein the use is for the treatment or prevention of tyrosinaemia.

[0264] Embodiment 78. The formula for use according to embodiment 77, wherein the use is for the treatment or prevention of Type I tyrosinaemia.

[0265] Embodiment 79. The formula for use according to embodiment 78, wherein the treatment or prevention results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of liver toxicity, kidney toxicity, nervous system toxicity, and endocrine dysfunction.

[0266] Embodiment 80. The formula for use according to embodiment 79, wherein the symptom is liver toxicity; preferably selected from the group consisting of a haemophiliaclike symptom, such as impaired clotting, increased bruising, bleeding inside the joins or bleeding inside the brain; acute liver failure; hypoalbuminemia and hepatocarcinoma.

[0267] Embodiment 81. The formula for use according to embodiment 79, wherein the symptom is kidney toxicity; preferably selected from the group consisting of hypophosphatemic rickets; impaired kidney function; renal tubular damage; and kidney failure. Embodiment 82. The formula for use according to embodiment 79, wherein the symptom is neurotoxicity; preferably selected from the group consisting of porphyria; and peripheral neuropathy.

[0268] Embodiment 83. The formula for use according to embodiment 79, wherein the symptom is endocrine dysfunction; preferably selected from the group consisting of dopamine dysfunction; norepinephrine dysfunction; and thyroxine dysfunction.

[0269] Embodiment 84. The formula for use according to embodiment 77, wherein the use is for the treatment or prevention of Type II tyrosinaemia, and / or Type I tyrosinaemia treated with Nitisinone (NTBC).

[0270] Embodiment 85. The formula for use according to embodiment 84, wherein the treatment or prevention results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of eye toxicity, skin toxicity and mentally disabled.

[0271] Embodiment 86. The formula for use according to embodiment 85, wherein the symptom is eye toxicity; preferably selected from the group consisting of epiphora; photophobia; eye pain; and hyperaemia.

[0272] Embodiment 87. The formula for use according to embodiment 85, wherein the symptom is skin toxicity; preferably palmoplantar hyperkeratosis.

[0273] Embodiment 88. The formula for use according to embodiment 85, wherein the symptom is mentally disabled.

[0274] Embodiment 89. The formula for use according to embodiment 77, wherein the use is for the treatment or prevention of Type III tyrosinaemia.

[0275] Embodiment 90. The formula for use according to embodiment 89, wherein the treatment or prevention results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of mental retardation, seizures, and intermittent ataxia.

[0276] Embodiment 91. The formula for use according to embodiment 90, wherein the symptom is mental retardation.

[0277] Embodiment 92. The formula for use according to embodiment 90, wherein the symptom is seizures.

[0278] Embodiment 93. The formula for use according to embodiment 90, wherein the symptom is intermittent ataxia. Embodiment 94. The formula for use according to any one of the previous embodiments, wherein the patient is a child under the age of 2 years, preferably the child is an infant, more preferably an infant up to the age of 6 months.

[0279] Embodiment 95. The formula for use according to any one of the previous embodiments, wherein the treatment or prevention lasts at least 3 months, preferably at least 6 months, more preferably at least 1 year.

[0280] Embodiment 96. The formula for use according to any one of the previous embodiments, wherein the formula is in the form of a liquid, a reconstitutable powder or a spoonable food; preferably in the form of a liquid or a reconstitutable powder.

[0281] Embodiment 97. The formula for use according to embodiment 96, wherein the formula is in the form of a liquid; more preferably in the form of a shelf-stable liquid; even more preferably a packaged, shelf-stable liquid.

[0282] Embodiment 98. The formula for use according to embodiment 97, wherein the formula has a viscosity of below 200 mPa s, preferably of from 5 to 180 mPa s, preferably 20 to 150 mPa s, measured at 20° C at a shear rate of 100 s '.

[0283] Embodiment 99. The formula for use according to any one of embodiments 96 to 98, wherein the formula is in the form of a liquid or a spoonable food, and has a density in the range of 1.02 to 1.2 g / L, preferably 1.05 to 1.25 g / L.

[0284] Embodiment 100. The formula for use according to any one of embodiments 96 to 99, wherein the formula is in the form of a liquid or a spoonable food, with a caloric density of 0.6 to 3.0 kcal / mL, preferably 0.6 to 2.0 kcal / mL, more preferably 0.6 to 1.0 kcal / mL.

[0285] Embodiment 101. The formula for use according to any one of embodiments 96 to 100, wherein the formula is in the form of a liquid or a spoonable food, and wherein protein is present in an amount of 1 to 15 g / 100 mL, preferably 1 to 10 g / 100 mL.

[0286] Embodiment 102. The formula for use according to any one of the previous embodiments, wherein the formula comprises GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1, preferably in a proportion of 1 :3 to 3 : 1, more preferably in a proportion of 1 :2 to 2: 1.

[0287] Embodiment 103. The formula for use according to any one of the preceding embodiments, wherein the formula contains a lipid source, preferably the lipid source comprises a fat selected from the group consisting of milk fat, sunflower oil, rapeseed oil, low erucic acid rapeseed oil, safflower oil, canola oil, olive oil, coconut oil, palm kernel oil, soybean oil, fish oil, palm oleic, high oleic sunflower oil and high oleic safflower oil, and microbial fermentation oil containing long chain, polyunsaturated fatty acids, more preferably, the lipid source comprises docosahexaenoic acid and / or arachidonic acid. Embodiment 104. The formula for use according to any one of the preceding embodiments, wherein the formula contains a digestible carbohydrate source, preferably selected from the group consisting of lactose, glucose, sucrose, fructose, galactose, maltose, starch and maltodextrin.

[0288] Embodiment 105. The formula for use according to any one of the preceding embodiments, wherein the formula contains a vitamin source, preferably comprising Vitamin A, Vitamin D3, Vitamin E, Vitamin C, Vitamin K, Thiamin (Bl), Riboflavin (B2), Niacin (B3), Vitamin B6, folic acid, Vitamin B 12, Biotin and / or Pantothenic acid.

[0289] Embodiment 106. The formula for use according to any one of the preceding embodiments, wherein the formula contains a mineral source, preferably comprising sodium, potassium, chloride, calcium, phosphorus, magnesium, iron, copper, zinc manganese, iodine, molybdenum, selenium, and / or chromium.

[0290] Embodiment 107. The formula for use according to any one of the preceding embodiments, wherein the formula comprises protein, fat, carbohydrate, vitamins and minerals.

[0291] Embodiment 108. The formula for use according to any one of the preceding embodiments, wherein the formula comprises prebiotics, probiotics, fibres and / or nucleotides; and / or choline, taurine, L-carnitine and inositol.

[0292] Embodiment 109. The formula for use according to any one of the preceding embodiments, wherein the formula comprises one or more thickener, additive, emulsifier, acidity regulators and / or antioxidants.

[0293] Embodiment 110. The formula for use according to any one of the preceding embodiments, wherein the formula is an infant formula, preferably wherein the infant formula provides 1.8 to 3.0 g / 100 kcal of protein, 4.4 to 6.0 g / 100 kcal of fat and 9.0 to 14.0 g / 100 kcal of digestible carbohydrate.

[0294] Embodiment 111. The infant formula for use according to embodiment 110, wherein the infant formula is a liquid and has an energy density of 60 to 75 kcal / 100 mL.

[0295] Embodiment 112. Use of intact GMP and free amino acids in the manufacture of a medicament for children under the age of 2 years, preferably an infant formula, for the treatment or prevention of PKU or tyrosinaemia in a patient in need thereof. Embodiment 113. A method of treatment or prevention of PKU or tyrosinaemia comprising administering a formula for children under the age of 2 years, preferably an infant formula, comprising intact GMP and free amino acids to a patient in need thereof.

[0296] Embodiment 114. A complete infant formula for the treatment or prevention of PKU or tyrosinaemia comprising intact GMP and free amino acids, wherein the GMP to free amino acids by weight are present in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2: 1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2.

[0297] Embodiment 115. A mixture of free amino acids comprising Arg, Asp+Asn, Cys, Gly, His, He, Leu, Lys, Met, Trp and Vai, and optionally Tyr, wherein the mixture of free amino acids contains less than 0.1 wt% of any of Ala, Gin, Glu, Pro, Phe, Ser or Thr.

[0298] Embodiment 116. A mixture of free amino acids according to embodiment 115, comprising as a total percentage by weight of free amino acids in the mixture: Arg (9 to 13 wt%), Asp+Asn (8 to 12 wt% total), Cys (2 to 6 wt%), Gly (12 to 16 wt%), His (4 to 8 wt%), He (2 to 6 wt%), Leu (13 to 17 wt%), Lys (5 to 9 wt%), Met (0.5 to 4 wt%), Trp (1 to 5 wt%), Vai (5 to 9%), and Tyr (13 to 17 wt%); preferably: Arg (10 to 12 wt%), Asp+Asn (9 to 11 wt% total), Cys (3 to 5 wt%), Gly (13 to 15 wt%), His (5 to 7 wt%), He (3 to 5 wt%), Leu (14 to 16 wt%), Lys (6 to 8 wt%), Met (1 to 3 wt%), Trp (2 to 4 wt%), Tyr (14 to 16 wt%) and Vai (6 to 8%).

[0299] Embodiment 117. A mixture of free amino acids according to embodiment 115, comprising as a total percentage by weight of free amino acids in the mixture: Arg (12 to 16 wt%), Asp+Asn (10 to 14 wt% total), Cys (2 to 6 wt%), Gly (15 to 19 wt%), His (5 to 9 wt%), He (3 to 7 wt%), Leu (16 to 20 wt%), Lys (6 to 10 wt%), Met (0.5 to 4 wt%), Trp (2 to 6 wt%) and Vai (7 to 11%); preferably: Arg (12 to 14 wt%), Asp+Asn (11 to 13 wt% total), Cys (3 to 5 wt%), Gly (16 to 18 wt%), His (6 to 8 wt%), He (4 to 6 wt%), Leu (17 to 19 wt%), Lys (7 to 9 wt%), Met (1 to 3 wt%), Trp (3 to 5 wt%) and Vai (8 to 10%).

[0300] Embodiment 118. A complete infant formula for use in the treatment or prevention of PKU or tyrosinaemia comprising intact GMP and free amino acids according to embodiment 114, wherein the free amino acids consist of the mixture of free amino acids of any one of embodiments 115 to 117.

[0301] Embodiment 119. A complete infant formula comprising intact GMP and free amino acids according to embodiment 114, wherein the free amino acids consist of the mixture of free amino acids of embodiment 116 for use in the treatment or prevention of phenylketonuria in a patient in need thereof. Embodiment 120. A complete infant formula comprising intact GMP and free amino acids according to embodiment 114, wherein the free amino acids consist of the mixture of free amino acids of embodiment 117 for use in the treatment or prevention of tyrosinaemia in a patient in need thereof. Embodiment 121. Use of intact GMP and free amino acids in the manufacture of a medicament for children under the age of 2 years, preferably an infant formula, for the treatment or prevention of PKU or tyrosinaemia in a patient in need thereof, wherein the formula is according to any one of embodiments 96 to 111, 114, 119 or 120.

[0302] Embodiment 122. A method of treating or preventing PKU or tyrosinaemia comprising administering a formula for children under the age of 2 years, preferably an infant formula, comprising intact GMP and free amino acids to a patient in need thereof, wherein the formula is according to any one of embodiments 96 to 111, 114, 119 or 120.

Claims

CLAIMS:

1. A formula for children under the age of 2 years comprising intact casein glycomacropeptide and free amino acids for use in the dietary management of phenylketonuria or tyrosinaemia in an individual in need thereof.

2. The formula for use according to claim 1, wherein the dietary management results in(a) good metabolic control, normal growth and / or improved gastrointestinal tolerance; or(b) good metabolic control, normal growth and improved gastrointestinal tolerance.

3. The formula for use according to claim 2, wherein(a) the good metabolic control achieves blood phenylalanine concentrations of 120- 360 pmol / L in an individual with phenylketonuria or blood tyrosine concentrations of 200-400 pmol / L in an individual with tyrosinaemia;(b) the normal growth results in maintenance, preferably an increase, of length for age, weight for age and / or BMI for age; or(c) the improved gastrointestinal tolerance is selected from the group consisting of reduced vomiting, regurgitation, abdominal discomfort or colic symptoms, flatulence, proportion or loose or hard stools, and / or difficulty feeding.

4. The formula for use according to any one of the previous claims, wherein(a) the use is for the dietary management of phenylketonuria; or(b) the use is for the dietary management of phenylketonuria and wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of intellectual disability, delays in developmental milestones, hyperactive behaviour, seizures, eczema, musty body odour, and light pigmentation (e.g., of the eyes, hair, and / or skin).

5. The formula for use according to any one of claims 1 to 3, wherein the use is for the dietary management of(a) tyrosinaemia;(b) tyrosinaemia, wherein the tyrosinaemia is Type I tyrosinaemia.

6. The formula for use according to claim 5, wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of liver toxicity, kidney toxicity, nervous system toxicity, and endocrine dysfunction.

7. The formula for use according to claim 6, wherein the symptom is(a) liver toxicity; preferably selected from the group consisting of a haemophiliaclike symptom, such as impaired clotting, increased bruising, bleeding inside the joins or bleeding inside the brain; acute liver failure; hypoalbuminemia and hepatocarcinoma;(b) kidney toxicity; preferably selected from the group consisting of hypophosphatemic rickets; impaired kidney function; renal tubular damage; and kidney failure;(c) neurotoxicity; preferably selected from the group consisting of porphyria; and peripheral neuropathy; or(d) endocrine dysfunction; preferably selected from the group consisting of dopamine dysfunction; norepinephrine dysfunction; and thyroxine dysfunction.

8. The formula for use according to claim 5(a), wherein the use is for the dietary management of Type II tyrosinaemia, and / or Type I tyrosinaemia treated with Nitisinone (NTBC).

9. The formula for use according to claim 8, wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of eye toxicity, skin toxicity and mentally disabled.

10. The formula for use according to claim 9, wherein the symptom is(a) eye toxicity; preferably selected from the group consisting of epiphora; photophobia; eye pain; and hyperaemia;(b) skin toxicity; preferably palmoplantar hyperkeratosis; or(c) mentally disabled.

11. The formula for use according to claim 5(a), wherein the use is for the dietary management of Type III tyrosinaemia.

12. The formula for use according to claim 11, wherein the dietary management results in prevention or reduction in severity or frequency of one or more symptom selected from the group consisting of mental retardation, seizures, and intermittent ataxia.

13. The formula for use according to any one of the previous claims, wherein the dietary management lasts at least 3 months, preferably at least 6 months, more preferably at least 1 year.

14. The formula for use according to any previous claim, wherein one or more of the following applies:(a) the formula is in the form of a liquid, a reconstitutable powder or a spoonable food; preferably in the form of a liquid or a reconstitutable powder;(b) the formula is in the form of a liquid; more preferably in the form of a shelf-stable liquid; even more preferably a packaged, shelf-stable liquid(c) the formula has a viscosity of below 200 mPa s, preferably of from 5 to 180 mPa s, preferably 20 to 150 mPa s, measured at 20° C at a shear rate of 100 s ';(d) wherein the formula is in the form of a liquid or a spoonable food, and has a density in the range of 1.02 to 1.2 g / L, preferably 1.05 to 1.25 g / L;(e) the formula is in the form of a liquid or a spoonable food, and has a caloric density of 0.6 to 3.0 kcal / mL, preferably 0.6 to 2.0 kcal / mL, more preferably 0.6 to 1.0 kcal / mL; and(f) wherein the formula is in the form of a liquid or a spoonable food, and protein is present in an amount of 1 to 15 g / 100 mL, preferably 1 to 10 g / 100 mL.

15. The formula for use according to any one of the previous claims, wherein the formula comprises GMP to free amino acids by weight in a proportion of 1 :4 to 4: 1, preferably in a proportion of 1 :3 to 3 : 1, more preferably in a proportion of 1 :2 to 2: 1.

16. The formula for use according to any preceding claim, wherein one or more of the following applies:(a) the formula contains a lipid source, preferably the lipid source comprises a fat selected from the group consisting of milk fat, sunflower oil, rapeseed oil, low erucic acid rapeseed oil, safflower oil, canola oil, olive oil, coconut oil, palm kernel oil, soybean oil, fish oil, palm oleic, high oleic sunflower oil and high oleic safflower oil, and microbial fermentation oil containing long chain, polyunsaturated fatty acids, more preferably, the lipid source comprises docosahexaenoic acid and / or arachidonic acid;(b) the formula contains a digestible carbohydrate source, preferably selected from the group consisting of lactose, glucose, sucrose, fructose, galactose, maltose, starch and maltodextrin;(c) the formula contains a vitamin source, preferably comprising Vitamin A, Vitamin D3, Vitamin E, Vitamin C, Vitamin K, Thiamin (Bl), Riboflavin (B2), Niacin (B3), Vitamin B6, folic acid, Vitamin B 12, Biotin and / or Pantothenic acid;(d) the formula contains a mineral source, preferably comprising sodium, potassium, chloride, calcium, phosphorus, magnesium, iron, copper, zinc manganese, iodine, molybdenum, selenium, and / or chromium;(e) the formula comprises protein, fat, carbohydrate, vitamins and minerals;(f) the formula comprises prebiotics, probiotics, fibres and / or nucleotides; and / or choline, taurine, L-carnitine and inositol; and(g) the formula comprises one or more thickener, additive, emulsifier, acidity regulators and / or antioxidants.

17. The formula for use according to any one of the preceding claims, wherein(a) the formula is an infant formula, preferably wherein the infant formula provides1.8 to 3.0 g / 100 kcal of protein, 4.4 to 6.0 g / 100 kcal of fat and 9.0 to 14.0 g / 100 kcal of digestible carbohydrate; or(b) the formula is an infant formula, preferably wherein the infant formula provides 1.8 to 3.0 g / 100 kcal of protein, 4.4 to 6.0 g / 100 kcal of fat and 9.0 to 14.0 g / 100 kcal of digestible carbohydrate, and wherein the infant formula is a liquid and has an energy density of 60 to 75 kcal / 100 mL.

18. A complete infant formula for use in the dietary management of PKU or tyrosinaemia comprising intact GMP and free amino acids, wherein the GMP to free amino acids by weight are present in a proportion of 1 :4 to 4: 1, preferably 1 :4 to 3 : 1, more preferably 1 :4 to 2:1, more preferably 1 :4 to 1 :2 and most preferably 1 :3 to 1 :2.

19. A mixture of free amino acids comprising Arg, Asp+Asn, Cys, Gly, His, He, Leu, Lys, Met, Trp and Vai, and optionally Tyr, wherein the mixture of free amino acids contains less than 0.1 wt% of any of Ala, Gin, Glu, Pro, Phe, Ser or Thr.

20. A mixture of free amino acids according to claim 19, comprising as a total percentage by weight of free amino acids in the mixture:(a) Arg (9 to 13 wt%), Asp+Asn (8 to 12 wt% total), Cys (2 to 6 wt%), Gly (12 to 16 wt%), His (4 to 8 wt%), He (2 to 6 wt%), Leu (13 to 17 wt%), Lys (5 to 9 wt%), Met (0.5 to 4 wt%), Trp (1 to 5 wt%), Vai (5 to 9%), and Tyr (13 to 17 wt%) preferably: Arg (10 to 12 wt%), Asp+Asn (9 to 11 wt% total), Cys (3 to 5 wt%), Gly (13 to 15 wt%), His (5 to 7 wt%), He (3 to 5 wt%), Leu (14 to 16 wt%), Lys (6 to 8 wt%), Met (1 to 3 wt%), Trp (2 to 4 wt%), Tyr (14 to 16 wt%) and Vai (6 to 8%); or(b) Arg (12 to 16 wt%), Asp+Asn (10 to 14 wt% total), Cys (2 to 6 wt%), Gly (15 to 19 wt%), His (5 to 9 wt%), He (3 to 7 wt%), Leu (16 to 20 wt%), Lys (6 to 10 wt%), Met (0.5 to 4 wt%), Trp (2 to 6 wt%) and Vai (7 to 11%); preferably: Arg (12 to 14 wt%), Asp+Asn (11 to 13 wt% total), Cys (3 to 5 wt%), Gly (16 to 18 wt%), His (6 to 8 wt%), He (4 to 6 wt%), Leu (17 to 19 wt%), Lys (7 to 9 wt%), Met (1 to 3 wt%), Trp (3 to 5 wt%) and Vai (8 to 10%).

21. A complete infant formula for use in the dietary management of PKU or tyrosinaemia comprising intact GMP and free amino acids according to claim 18, wherein the free amino acids consist of the mixture of free amino acids of claim 19 or claim 20.

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