WO2021146561A1

WO2021146561A1 - Feeding protocol for optimal growth of preterm infants

Info

Publication number: WO2021146561A1
Application number: PCT/US2021/013642
Authority: WO
Inventors: Melinda J. ELLIOTT; Andrea Marie MARKELL
Original assignee: Prolacta Bioscience, Inc.
Priority date: 2020-01-17
Filing date: 2021-01-15
Publication date: 2021-07-22

Abstract

Provided herein are methods and feeding protocols for the treatment of infants affected by premature birth or very low birth weight (VLBW) as well as for ameliorating the conditions and reducing the risks associated with premature birth or VLBW. The provided methods and protocols relate, at least in part, to an exclusive human milk diet, wherein the subject only receives enteral or oral nutrition containing human milk proteins, fats, and carbohydrates. In some aspects, the provided methods and protocols result in one or more of improved growth with respect to length, head circumference, and weight; improved health outcomes and a reduced risk of adverse events; and improved neurodevelopmental outcomes.

Description

FEEDING PROTOCOL FOR OPTIMAL GROWTH OF PRETERM INFANTS

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority from U.S. provisional application No. 62/962,785, filed January 17, 2020, U.S. provisional application No. 62/968,752, filed January 31, 2020, U.S. provisional application No. 62/994,016, filed March 24, 2020, U.S. provisional application No. 63/074,079, filed September 3, 2020, and U.S. provisional application No. 63/090,374 filed October 12, 2020, the contents of each of which are incorporated by reference in its entirety.

FIELD OF THE INVENTION

[0002] Provided herein are methods and feeding protocols for the treatment of infants affected by premature birth or very low birth weight (VLBW) as well as for ameliorating the conditions and reducing the risks associated with premature birth or VLBW. The provided methods and protocols relate, at least in part, to an exclusive human milk diet, wherein the subject only receives enteral or oral nutrition containing human milk proteins, fats, and carbohydrates. In some aspects, the provided methods and protocols result in one or more of improved growth with respect to length, head circumference, and weight; improved health outcomes and a reduced risk of adverse events; and improved neurodevelopmental outcomes.

BACKGROUND OF THE INVENTION

[0003] In 2017, approximately one out of ten infants bom in the United States were bom prematurely. Premature infants are at increased risk of early growth retardation, infectious disease, developmental delay, and death during infancy and childhood. Very-low- birth- weight (VLBW) infants, those infants bom weighing less than 1.5 kg, are particularly vulnerable to these adverse outcomes. Providing adequate nutrition early in life for this patient population is remains challenging. What is needed in the art are improved interventions and protocols to improve the feeding of preterm infants and thereby improve their immediate and longer-term health and well-being.

SUMMARY OF THE INVENTION

[0004] Provided herein are methods and protocols for providing nutrition to a subject that is a premature or very low birth weight infant. The provided methods and protocols relate, at least in part, to an exclusive human milk diet, where the subject only receives, enterally or orally, human milk proteins, fats, and carbohydrates for the duration of the method or protocol. In some aspects, the provided methods and protocols result in improved growth with respect to length, head circumference, and weight; improved health; and improved neurodevelopmental outcomes as compared to similar subjects provided nutrition by alternative methods and protocols.

[0005] Provided herein is a method for providing nutrition to a premature or very low birthweight (VLBW) infant, comprising: enterally administering a feeding volume of a first or a second human milk composition to a subject that is a premature or VLBW infant on each day of a feeding protocol; wherein the feeding protocol begins within the first three days of the subject’s life; wherein the feeding volume at day 1 of the protocol is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 10 mL/kg/day and 30 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the feeding protocol beginning at day 5 until a feeding volume of at least 160 mL/kg/day is achieved; wherein the first human milk composition is administered on day 1 of the feeding protocol and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the second human milk composition is administered daily during the feeding protocol beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first human milk composition comprises (i) mother’s own milk (MOM) or a mixture of MOM and pasteurized human donor milk, or (ii) a first standardized human milk formulation comprising at least 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL); and wherein the second human milk composition comprises (i) fortified MOM comprising (a) MOM or a mixture of MOM and pasteurized human donor milk and (b) a human milk-based human milk fortifier, wherein the human milk fortifier is added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 5 kcal/ounce (16.9 kcal/100 mL) or 6 kcal/ounce (20.2 kcal/100 mL); or (ii) a second standardized human milk formulation, wherein the second standardized human milk formulation comprises at least about 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL) or 26 kcal/ounce (87.9 kcal/100 mL).

[0006] Also provided are human milk compositions, e.g., a first and a second human milk composition, for use in a method of treating malnourishment in a premature or very low birth weight (VLBW) infant, wherein the method comprises any of the methods, treatment regimens, or feeding protocols described herein. In addition, provided herein are human milk compositions, e.g., a first and a second human milk composition, for use in a method of treating premature or very low birthweight infants, wherein the method comprises any of the wherein the method comprises any of the methods, treatment regimens, or feeding protocols described herein. Also provided herein are human milk compositions, e.g., a first and a second human milk composition, for use in a method for transitioning a premature or VLBW infant from parenteral nutrition (PN), comprising any of the methods, treatment regimens, or feeding protocols described herein.

[0007] In some embodiments, all of the proteins, fats, and carbohydrates that are enterally administered during the feeding protocol are derived from, originate from, and/or are native to human milk. In particular embodiments, the feeding volume of 60 mL/kg/day is achieved no later than day 5 of the feeding protocol and/or the subject’s life. In certain embodiments, the feeding volume of 60 mL/kg/day is achieved on day 2 of the feeding protocol and/or the subject’s life.

[0008] In particular embodiments, the methods comprise enterally administering a feeding volume of (i) mother’s own milk (MOM) or the mixture of MOM and pasteurized human donor milk or (ii) fortified MOM to the subject on each day of the feeding protocol; wherein the feeding volume at day 1 of the protocol is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 10 mL/kg/day and 30 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the feeding protocol beginning at day 5 until the feeding volume of at least 160 mL/kg/day is achieved; wherein the MOM or the mixture is administered on day 1 of the feeding protocol and is administered daily until the feeding volume of 60 mL/kg/day is achieved, and wherein the fortified MOM is administered daily during the feeding protocol beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the fortified MOM comprises (a) MOM or the mixture of MOM and pasteurized human donor milk and (b) the human milk fortifier, wherein the human milk fortifier has been added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 5 kcal/ounce (16.9 kcal/100 mL) or 6 kcal/ounce (20.2 kcal/100 mL).

[0009] In certain embodiments, the methods comprise: enterally administering a feeding volume of the first or the second standardized human milk formulation to the subject on each day of the feeding protocol; wherein the feeding volume at day 1 of the protocol is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 10 mL/kg/day and 30 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the feeding protocol beginning at day 5 until the feeding volume of at least 160 mL/kg/day is achieved; wherein the first standardized human milk formulation is administered on day 1 of the feeding protocol and is administered daily until the feeding volume of 60 mL/kg/day is achieved, and wherein the second standardized human milk formulation is administered daily during the feeding protocol beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first standardized human milk formulation comprises at least 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL); and wherein the second standardized human milk formulation comprises at least about 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL) or 26 kcal/ounce (87.9 kcal/100 mL).

[0010] In some embodiments, the feeding volume at day 1 of the protocol is about 10 mL/kg/day, wherein the feeding volume increases by about 10 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by about 20 mL/kg/day on each day of the feeding protocol beginning at day 5 until the feeding volume of at least 160 mL/kg/day is achieved. In particular embodiments, the first human milk composition is administered on day 1 until day 4 of the feeding protocol, and wherein the second human milk composition is administered daily during the feeding protocol beginning on day 5 of the feeding protocol. In certain embodiments, the subject is a high-risk premature or VLBW infant. In some embodiments, the subject has one or more of: an early gestational age, low birthweight, significant intrauterine growth restriction of less than or equal to the third percentile at birth, a low APGAR score, need for cardiopulmonary resuscitation (CPR) after delivery, maternal drug use, need for pressor support of blood pressure, of a presence of hemodynamically significant patent ductus arteriosus. In particular embodiments, the feeding volume at day 1 of the protocol is about 20 mL/kg/day, wherein the feeding volume increases by about 20 mL/kg/day on each day of the feeding protocol beginning from day 1 of the feeding protocol until the feeding volume of at least 160 mL/kg/day is achieved. In certain embodiments, the subject is a small premature infant, and/or wherein the subject has a birthweight below 1000 g or 75 Og.

[0011] In some embodiments, the feeding volume at day 1 of the protocol is about 30 mL/kg/day, wherein the feeding volume increases by about 30 mL/kg/day on each day of the feeding protocol beginning from day 1 of the feeding protocol until the feeding volume of at least 160 mL/kg/day is achieved. In particular embodiments, the subject is a large premature infant and/or wherein the subject has a birthweight greater than 750 g, 1000 g, or 1250 g.

[0012] In certain embodiments, the first human milk composition is administered on day 1 of the feeding protocol and wherein the second human milk composition is administered daily during the feeding protocol beginning on day 2 of the feeding protocol.

[0013] In some embodiments, the first human milk composition is MOM and the second composition is the fortified MOM. In particular embodiments, the pasteurized donor milk comprises about 20 kcal/ounce (67.6 kcal/100 mL). In certain embodiments, the fortified MOM comprises between 20% and 40% of the human milk fortifier. In some embodiments, the human milk-based fortifier comprises between 1.2 kcal/mL and 1.6 kcal/mL. In particular embodiments, the human milk-based fortifier comprises between 50 mg/mL and 70 mg/mL human milk protein In certain embodiments, the human milk fortifier comprises between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat. In some embodiments, the human milk fortifier comprises about 1.44 kcal/mL, about 59 mg/mL human milk protein, and about 93 mg/mL human milk fat.

[0014] In particular embodiments, the human milk-based fortifier has been added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 6 kcal/ounce (20.3 kcal/100 mL). In certain embodiments, the first human milk composition is the first standardized human milk formulation and wherein the second human milk composition is the second standardized human milk formulation. In some embodiments, the first standardized human milk formulation comprises between about 23 mg/mL and 26 mg/mL human milk protein. In particular embodiments, the first standardized human milk formulation comprises between about 22 kcal/ounce and 26 kcal/ounce (74.4 kcal/100 mL and 87.9 kcal/100 mL). In certain embodiments, the first standardized human milk formulation comprises between 22 kcal/ounce and 26 kcal/ounce (74.4 kcal/100 mL and 87.9 kcal/100 mL)., between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat. In some embodiments, the second standardized human milk formulation comprises between about 25 mg/mL and 28 mg/mL human milk protein. In particular embodiments, the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL). [0015] In certain embodiments, the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL), between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat. In some embodiments, the first standardized human milk formulation comprises between 24 kcal/ounce and 26 kcal/ounce (81.2 kcal/100 mL and 87.9 kcal/100 mL), between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat; and wherein the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL)„ between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat. In particular embodiments, the first standardized human milk formulation comprises about 24 kcal/ounce (81.2 kcal/100 mL), about 24 mg/mL human milk protein, and about 45 mg/mL human milk fat. In certain embodiments, the first standardized human milk formulation comprises about 26 kcal/ounce (87.9 kcal/100 mL), about 26 mg/mL human milk protein, and about 51 mg/mL fat. In some embodiments, the second standardized human milk formulation comprises about 26 kcal/ounce (87.9 kcal/100 mL), about 26 mg/mL human milk protein, and about 51 mg/mL fat. In particular embodiments, the feeding protocol continues at the feeding volume of at least 160 mL/kg/day after the feeding volume of 160 mL/kg/day is achieved, and wherein the feeding protocol continues until the subject reaches at least 34 weeks gestational age.

[0016] In certain embodiments, the method further comprises increasing the total calorie content of the second human milk composition by between about 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) by adding an amount of a human cream composition to the second human milk composition prior to enteral administration on each day of the protocol after a feeding volume of at least 100 mL/kg/day has been achieved, wherein the human cream composition comprises about 2.5-2.6 kcal/mL and about 25% fat, optionally wherein the total calorie content is increased by about 2 kcal/ounce (6.7 kcal/100 mL).

[0017] In some embodiments, the method further comprises increasing the total calorie content of the second human milk composition by between about 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) by adding an amount of a human cream composition to the second human milk composition prior to enteral administration on each day of the protocol if the subject’s growth rate is less than 15 g/kg/day for three consecutive days after the feeding volume of 160 mL/kg/day is achieved, wherein the human cream composition comprises about 2.5-2.6 kcal/mL and about 25% fat, optionally wherein the total calorie content is increased by about 2 kcal/ounce (6.7 kcal/100 mL).

[0018] In particular embodiments, the subject begins parenteral nutrition (PN) on the first day of life; wherein the subject receives PN until the enteral feeding volume is between 100 mL/kg/day and 120 mL/kg/day; and on days where the subject receives PN, the total volume intake from the PN and the enteral administration is between 150 mL/kg/day and 160 mL/kg/day. In certain embodiments, the PN is discontinued by day 10 of life.

[0019] Provided herein is a method for providing nutrition to a premature or very low birthweight (VLBW) infant, comprising: enterally or orally administering a feeding volume of a first or a second human milk composition to a subject that is a premature or VLBW infant on each day of a treatment regimen; wherein the treatment regimen begins on day 1 of the subject’s life and lasts for at least 10 days; wherein the feeding volume at day 1 of the subject’s life is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 0 mL/kg/day and 30 mL/kg/day from day 1 to day 2 of the treatment regimen, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the treatment regimen following day 2 until a feeding volume of 160 mL/kg/day is achieved; wherein the first human milk composition is administered on day 1 of the treatment and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the second human milk composition is administered daily during the treatment regimen beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first human milk composition comprises (i) mother’s own milk (MOM) or a mixture of MOM and pasteurized human donor milk, or (ii) a first standardized human milk formulation comprising at least 20 mg/mL human milk protein and greater than 20 kcal/ounce (67.6 kcal/100 mL); and wherein the second human milk composition comprises (i) fortified MOM comprising (a) MOM or a mixture of MOM and pasteurized human donor milk and (b) a human milk-based human milk fortifier (also referred to herein as a human milk-based fortifier), wherein the human milk based fortifier is added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 5 kcal/ounce (16.9 kcal/100 mL) or (ii) a second standardized human milk formulation, wherein the second standardized human milk formulation comprises at least about 20 mg/mL human milk protein and greater than 20 kcal/ounce (67.6 kcal/100 mL) and at least about 20 mg/mL human milk protein. [0020] In some embodiments, the feeding volume of 60 mL/kg/day is achieved no later than day 5 of the subject’s life. In particular embodiments, the feeding volume of 60 mL/kg/day is achieved on day 2 of the subject’s life. In certain embodiments, the first human milk composition and the second human milk composition are administered enterally. In some embodiments, the first human milk composition and the second human milk composition are different.

[0021] In particular embodiments, the first human milk composition is MOM and the second composition is the fortified MOM. In certain aspects, provided herein is a method for providing nutrition to a premature or very low birth weight (VLBW) infant when mother’s own milk (MOM) is available, comprising: enterally administering a feeding volume of a first or a second human milk composition to a subject that is a premature or VLBW infant on each day of a treatment regimen; wherein the treatment regimen begins on day 1 of the subject’s life and lasts for at least 10 days; wherein the feeding volume at day 1 of the subject’s life is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 0 mL/kg/day and 30 mL/kg/day from day 1 to day 2 of the treatment regimen, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the treatment regimen following day 2 until a feeding volume of 160 mL/kg/day is achieved; wherein the first human milk composition is administered on day 1 of the treatment regimen and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the second human milk composition is administered daily during the treatment regimen beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first human milk composition comprises mother’s own milk (MOM) or a mixture of MOM and pasteurized human donor milk comprising about 20 kcal/ounce (67.6 kcal/100 mL); and wherein the second human milk composition comprises a fortified MOM comprising (a) MOM or a mixture of MOM and pasteurized human donor milk and (b) a human milk-based fortifier, wherein the human milk based fortifier is added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 5 kcal/ounce (16.9 kcal/100 mL).

[0022] In some embodiments, the pasteurized donor milk comprises 20 kcal/ounce (67.6 kcal/100 mL). In particular embodiments, the pasteurized donor milk comprises is donated human milk pooled from at least 25, 50, or 100 different individual donors. In certain embodiments, the fortified MOM comprises at least 25 kcal/ounce (84.5 kcal/100 mL). and at least about 24 mg/ml human milk protein. In some embodiments, the fortified MOM comprises between 25 mg/mL and 33 mg/mL human milk protein. In particular embodiments, the fortified MOM comprises between 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL). In certain embodiments, the fortified MOM comprises between 20% and 40% human milk fortifier. In some embodiments, the human milk-based fortifier comprises between 1.2 kcal/mL and 1.6 kcal/mL. In particular embodiments, the human milk-based fortifier comprises between 50 mg/mL and 70 mg/mL human milk protein.

[0023] In certain embodiments, the human milk-based fortifier comprises between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat. In some embodiments, the fortified MOM comprises between about 60% and 80% of the MOM and between about 20% and 40% of the human milk-based fortifier. In particular embodiments, the fortified MOM comprises about 70% of the MOM and about 30% of the human milk-based fortifier.

[0024] Provided herein is a method for providing nutrition to a premature or very low birthweight (VLBW) infant, comprising: enterally administering to a subject that is a premature or VLBW infant mother’s own milk (MOM) and/or donor milk at a feeding volume of between 10 mL/kg/day and 30 mL/kg/day on day 1 of the feeding protocol and/or of the subject’s life, beginning at day 2 of the feeding protocol and/or of the subject’s life , enterally administering MOM or a fortified MOM, wherein the feeding volume is increased between 0 mL/kg/day and 30 mL/kg/day from day 1 to day 2 of the feeding protocol and/or of the subject’s life, wherein MOM is administered if the feeding volume is under 60 mL/kg/day and the fortified MOM is administered if the feeding volume is at least 60 mL/kg/day; and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day beginning on day 3 of the feeding protocol and/or the subject’s life each day of the treatment regimen following day 2 until a feeding volume of 160 mL/kg/day is achieved; wherein the fortified MOM comprises (i) about 70% of the MOM and/or donor milk and (ii) about 30% of a human milk-based fortifier comprising at between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat.

[0025] In certain embodiments, the human milk-based fortifier comprises about 1.44 kcal/mL, about 59 mg/mL human milk protein, and about 93 mg/mL human milk fat. In some embodiments, the human milk-based fortifier is formulated or manufactured from pooled human milk, wherein the pooled human milk is pooled from at least 25, 50, 100, or more individual donors. In particular embodiments, the human milk-based fortifier comprises at least 10, 20, 50, 100, 150, or more human milk oligosaccharides (HMOs) and wherein the human milk oligosaccharides comprise one or more sialylated HMOs.

[0026] In certain embodiments, the first human milk composition is a first standardized human milk formulation and wherein the second human milk composition is a second standardized human milk formulation.

[0027] Provided herein is a method for providing nutrition to a premature or very low birthweight (VLBW) infant when mother’s own milk (MOM) is not available, comprising: enterally administering a feeding volume of a first or a second standardized human milk formulation to a subject that is a premature or VLBW infant on each day of a treatment regimen; wherein the treatment regimen begins on day 1 or between days 1 and 3 of the subject’s life and lasts for at least 10 days; wherein the feeding volume at day 1 of treatment regimen and/or the subject’s life is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 0 mL/kg/day and 30 mL/kg/day from day 1 to day 2 of the treatment regimen, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the treatment regimen following day 2 until a feeding volume of 160 mL/kg/day is achieved; wherein the first standardized human milk formulation is administered on day 1 of the treatment regimen and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the second standardized human milk formulation is administered daily during the treatment regimen beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first standardized human milk formulation comprises at least 20 mg/mL human milk protein and greater than 20 kcal/ounce (67.6 kcal/100 mL); and wherein the second standardized human milk formulation comprises at least about 20 mg/mL human milk protein and greater than 20 kcal/ounce (67.6 kcal/100 mL) and at least about 20 mg/mL human milk protein.

[0028] In particular embodiments, the first standardized human milk formulation comprises between about 23 mg/mL and 26 mg/mL human milk protein. In particular embodiments, the first standardized human milk formulation comprises between about 22 kcal/ounce and 26 kcal/ounce (74.4 kcal/100 mL and 87.9 kcal/100 mL). In certain embodiments, the first standardized human milk formulation comprises between 22 kcal/ounce and 26 kcal/ounce (74.3 kcal/100 mL and 87.9 kcal/100 mL), between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat. In some embodiments, the second standardized human milk formulation comprises between about 25 mg/mL and 28 mg/mL human milk protein. In particular embodiments, the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL). In certain embodiments, the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL), between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat.

[0029] Provided herein is a method for providing nutrition to a premature or very low birthweight (VLBW) infant when mother’s own milk (MOM) not available, comprising: enterally administering a feeding volume of a first or a second standardized human milk formulation to a subject that is a premature or VLBW infant on each day of a treatment regimen; wherein the treatment regimen begins on day 1 or between days 1 and 3 of the subject’s life and lasts for at least 10 days; wherein the feeding volume at day 1 of the treatment regimen and/or the subject’s life is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 0 mL/kg/day and 30 mL/kg/day from day 1 to day 2 of the treatment regimen, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the treatment regimen following day 2 until a feeding volume of 160 mL/kg/day is achieved; wherein the first standardized human milk formulation is administered on day 1 of the treatment regimen and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the second standardized human milk formulation is administered daily during the treatment regimen beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first standardized human milk formulation comprises between 22 kcal/ounce and 26 kcal/ounce (74.4 kcal/100 mL and 87.9 kcal/100 mL)„ between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat; and wherein the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL), between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat.

[0030] In some embodiments, the first standardized human milk formulation comprises about 24 kcal/ounce (81.2 kcal/100 mL), about 24 mg/mL human milk protein, and about 45 mg/mL human milk fat.

[0031] In particular embodiments, the second standardized human milk formulation comprises about 26 kcal/ounce (87.9 kcal/100 mL), about 26 mg/mL human milk protein, and about 51 mg/mL fat. In particular embodiments, the method additionally comprises, prior to at least one administration of the first and/or second human milk compositions, increasing the total calorie content of the first and/or the second human milk composition by between about 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) by adding an amount of a human cream composition to the first and/or second human milk composition, wherein the human cream composition comprises about 2.5-2.6 kcal/mL and about 25% fat.

[0032] In certain embodiments, the subject’s gestational age at birth is less than 32 weeks. In some embodiments, the subject’s gestational age at birth is less than 28 weeks. In particular embodiments, the subject’s birthweight is less than 1500 g. In certain embodiments, the subject’s birthweight is less than 1250 g. In some embodiments, the subject’s birthweight is less than 1000 g.

[0033] Provided herein is a method for transitioning a subject from parenteral nutrition to oral or enteral nutrition, comprising providing nutrition to a subject according to any of the methods provided herein. In particular embodiments, the subject is removed from parenteral nutrition by day 10 of life.

[0034] Provided herein is a method for providing nutrition to a premature or very low birthweight infant, comprising a feeding protocol depicted in any of Tables 2-4 or E1-E6, or a feeding protocol depicted in any of Tables 5-8 or E7-E12.

[0035] Also provided is a method for providing nutrition to a premature or very low birthweight (VLBW) infant, comprising: (A) administering to a subject that is a premature or VLBW infant a first human milk composition, wherein the first human milk composition is administered orally or enterally at between about 10 mL/kg/day and 30 mL/kg/day on the first day of a first treatment phase, wherein the first human milk composition comprises (i) mother’s own milk (MOM) and/or donor milk, or (ii) a first standardized human milk formulation comprising at least 20 mg/mL human milk protein and at least 0.7 kcal/mL, wherein the first human milk composition is administered daily for the first treatment phase, and wherein the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day during the first treatment phase; and (B) administering to the subject a second human milk, wherein the second human milk composition is administered orally or enterally, wherein the second human milk composition is administered daily during a second treatment phase which begins immediately after the first treatment phase ends, wherein the second human milk composition is administered at between about 50 mL/kg/day and 120 mL/kg/day on the first day of the second treatment phase, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day during the second treatment phase until a feeding volume of about 160 mL/kg/day is achieved, and wherein the second human milk composition is (i) fortified MOM comprising (a) MOM and (b) a human milk-based fortifier, wherein the fortified MOM comprises at least about 0.84 kcal/mL and at least about 24 mg/ml human milk protein or (ii) a second standardized human milk formulation, wherein the second standardized human milk formulation comprises at least 0.7 kcal/mL and at least about 20 mg/mL human milk protein; and wherein the standardized human milk formulation is only administered to the subject if MOM is not available for administration. In some embodiments, the first treatment phase is from day 1 to day 3 of the treatment regimen and/or the subject’s life. In certain embodiments, the second treatment phase is from day 4 to at least day 10 of the treatment regimen and/or the subject’s life.

[0036] Also provided herein is a method for providing nutrition to a premature or very low birthweight infant, comprising: (a) administering to a subject that is a premature or very low birth weight infant a first human milk composition on days 1-3 of a treatment regimen and/or the subject’s life, wherein the first human milk composition is administered orally or enterally at between about 20 mL/kg/day and 30 mL/kg/day; and wherein the first human milk composition is mother’s own milk (MOM) or a first standardized human milk formulation comprising at least 20 mg/mL human milk protein and at least 0.7 kcal/mL; and (b) administering to the subject a second human milk composition on days 4-10 of the treatment regimen and/or the subject’s life, wherein the second human milk composition is administered orally or enterally, wherein the second human milk composition is administered on day 4 of the treatment regimen and/or the subject’s life at between about 40 mL/kg/day and 60 mL/kg/day, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 5 through 10 of the treatment regimen and/or the subject’s life or until a feeding volume of about 160 mL/kg/day is achieved, and wherein the second human milk composition is (i) fortified MOM comprising (a) MOM and (b) a human milk-based fortifier, wherein the fortified MOM comprises at least about 0.84 kcal/mL and at least about 24 mg/ml human milk protein or (ii) a second standardized human milk formulation, wherein the second standardized human milk formulation comprises at least 0.7 kcal/mL and at least about 20 mg/mL human milk protein; and wherein the standardized human milk formulation is only administered to the subject when MOM is not available for administration. [0037] In some embodiments, the first human milk composition and the second human milk composition is administered enterally. In particular embodiments, the first human milk composition is the MOM and wherein the second human milk composition is the fortified MOM.

[0038] Also provided herein is a method for providing nutrition to a premature or very low birthweight infant, comprising: (a) enterally administering to a subject that is a premature or very low birth weight infant mother’s own milk (MOM) between about 20 mL/kg/day and 30 mL/kg/day on days 1-3 of a treatment regimen and/or the subject’s life; and (b) enterally administering to the subject a fortified MOM on days 4-10 of the treatment regimen and/or the subject’s life, wherein the fortified MOM is administered enterally on day 4 of life at between about 40 mL/kg/day and 60 mL/kg/day, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 5 through 10 of the treatment regimen and/or the subject’s life or until a feeding volume of about 160 mL/kg/day is achieved, wherein the fortified MOM comprises (i) MOM and (ii) a human milk-based fortifier comprising at least about 0.84 kcal/mL and at least about 24 mg/ml human milk protein.

[0039] In addition, provided herein is a method for providing nutrition to a premature or very low birthweight (VLBW) infant, comprising: (a) enterally administering to a subject that is a premature or VLBW infant mother’s own milk (MOM) and/or donor milk between 10 mL/kg/day and 30 mL/kg/day on day 1 of a treatment regimen and/or the subject’s life, wherein the MOM and/or donor milk is administered on days 1-3 of the treatment regimen and/or the subject’s life, and wherein the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day during days 1-3 of the treatment regimen and/or the subject’s life; and (b) enterally administering to the subject a fortified MOM on day 4 to at least day 10 of the treatment regimen and/or the subject’s life, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 4 through 10 of the treatment regimen and/or the subject’s life or until a feeding volume of about 160 mL/kg/day is achieved, wherein the fortified MOM comprises (i) MOM and (ii) a human milk-based fortifier comprising at least about 0.84 kcal/mL and at least about 24 mg/ml human milk protein.

[0040] In certain embodiments, the fortified MOM comprises between 25 mg/mL and 33 mg/mL human milk protein. In some embodiments, the fortified MOM comprises between 0.85 kcal/mL and 0.95 kcal/mL. In particular embodiments, the human milk-based fortifier comprises between 1.2 kcal/mL and 1.6 kcal/mL. In certain embodiments, the human milk-based fortifier comprises between 50 mg/mL and 70 mg/mL human milk protein. In some embodiments, the human milk-based fortifier comprises between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat. In particular embodiments, the human milk-based fortifier comprises about 1.44 kcal/mL, about 59 mg/mL human milk protein, and about 93 mg/mL human milk fat. In certain embodiments, the fortified MOM comprises between about 60% and 80% of the MOM and between about 20% and 40% of the human milk-based fortifier. In some embodiments, the fortified MOM comprises about 70% of the MOM and about 30% of the human milk-based fortifier.

[0041] Additionally, provided herein is method for providing nutrition to a premature or very low birthweight (VLBW) infant, comprising: (a) enterally administering to a subject that is a premature or VLBW infant mother’s own milk (MOM) and/or donor milk between 10 mL/kg/day and 30 mL/kg/day on day 1 of a treatment regimen and/or the subject’s life, (b) enterally administering on days 2 and 3 of the treatment regimen and/or the subject’s life: (i) MOM and or donor milk, or (ii) a fortified MOM and/or a fortified donor milk, wherein the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day during days 1-3 of the treatment regimen and/or the subject’s life; and (c) enterally administering to the subject the fortified MOM on day 4 to at least day 10 of the treatment regimen and/or the subject’s life, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 4 through 10 of the treatment regimen and/or the subject’s life or until a feeding volume of about 160 mL/kg/day is achieved, wherein the fortified MOM and/or the fortified donor milk comprises (i) about 70% of the MOM and/or donor milk and (ii) about 30% of a human milk-based fortifier comprising at between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat.

[0042] Also provided herein is a method for providing nutrition to a premature or very low birthweight infant, comprising: (a) enterally administering to a subject that is a premature or very low birth weight infant mother’s own milk (MOM) between about 20 mL/kg/day and 30 mL/kg/day on days 1-3 of a treatment regimen and/or the subject’s life; and (b) enterally administering to the subject a fortified MOM on days 4-10 of the treatment regimen and/or the subject’s life, wherein the fortified MOM is administered enterally on day 4 of the treatment regimen and/or the subject’s life at between about 40 mL/kg/day and 60 mL/kg/day, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 5 through 10 of the treatment regimen and/or the subject’s life or until a feeding volume of about 160 mL/kg/day is achieved, wherein the fortified MOM comprises (i) about 70% of the MOM and (ii) about 30% of a human milk-based fortifier comprising at between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat.

[0043] In particular embodiments, the human milk-based fortifier is formulated or manufactured from pooled human milk, wherein the pooled human milk is pooled from at least 10, 25, 50, 100, or more individual donors. In certain embodiments, the human milk- based fortifier comprises at least 10, 20, 50, 100, 150, or more human milk oligosaccharides (HMOs) and wherein the human milk oligosaccharides comprise one or more sialylated HMOs.

[0044] In some embodiments, the first human milk composition is the first standardized human milk formulation and wherein the second human milk composition is the second standardized human milk formulation.

[0045] Also provided herein is a method for providing nutrition to a premature or very low birthweight infant when mother’s own milk is not available, comprising: (a) enterally or orally administering to a subject that is a premature or very low birth weight infant a first standardized human milk formulation on days 1-3 of a treatment regimen and/or the subject’s life at between about 20 mL/kg/day and 30 mL/kg/day; and wherein the first standardized human milk formulation comprises at least 20 mg/mL human milk protein and at least 0.7 kcal/mL.

[0046] In particular embodiments, the method further comprises enterally or orally administering a second standardized human milk formulation on days 4 through 10 of the treatment regimen and/or the subject’s life wherein the second standardized human milk formulation comprises at least 0.7 kcal/mL and at least about 20 mg/mL human milk protein.

[0047] Also provided is method for providing nutrition to a premature or very low birthweight infant when mother’s own milk is not available, comprising: (a) enterally or orally administering to a subject that is a premature or very low birth weight infant a first standardized human milk formulation on day 1 of a treatment regimen and/or the subject’s life at between about 20 mL/kg/day and 30 mL/kg/day; and wherein the first standardized human milk formulation comprises at least 20 mg/mL human milk protein and at least 0.7 kcal/mL. In some embodiments, the method further comprises enterally or orally administering a second standardized human milk formulation on days 2 through at least day 10 of the treatment regimen and/or the subject’s life, wherein the second standardized human milk formulation comprises at least 0.7 kcal/mL and at least about 20 mg/mL human milk protein.

[0048] In particular embodiments, the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 5 through 10 of the treatment regimen and/or the subject’s life or until a feeding volume of about 160 mL/kg/day is achieved. In certain embodiments, the first standardized human milk formulation comprises between about 23 mg/mL and 26 mg/mL human milk protein. In some embodiments, the first standardized human milk formulation comprises between about 0.75 kcal/mL and 0.87 kcal/mL. In particular embodiments, the first standardized human milk formulation comprises between about 0.75 kcal/mL and 0.87 kcal/mL, between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat. In certain embodiments, the first standardized human milk formulation comprises about 0.82 kcal/mL, about 24 mg/mL human milk protein, and about 45 mg/mL human milk fat. In some embodiments, the second standardized human milk formulation comprises between about 25 mg/mL and 28 mg/mL human milk protein. In particular embodiments, the second standardized human milk formulation comprises between about 0.83 kcal/mL and 0.95 kcal/mL. In certain embodiments, the second standardized human milk formulation comprises between about 0.83 kcal/mL and 0.95 kcal/mL, between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat. In some embodiments, the second standardized human milk formulation comprises about 0.89 kcal/mL, about 26 mg/mL human milk protein, and about 51 mg/mL fat.

[0049] In addition, provided herein is a method for providing nutrition to a premature or very low birth weight infant when mother’s own milk (MOM) is not available, comprising: (a) enterally administering to a subject that is a premature or very low birth weight infant a first standardized human milk formulation on days 1 of a treatment regimen and/or the subject’s life at between about 10 mL/kg/day and about 30 mL/kg/day, wherein the first standardized human milk formulation comprises between about 0.75 kcal/mL and 0.87 kcal/mL, between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat; and (b) enterally administering the first standardized human milk formulation or a second standardized human milk formulation on days 2 to 3 of the treatment regimen and/or the subject’s life, wherein the feeding volume is advanced daily on days 1-3 of the treatment regimen and/or the subject’s life by between about 10 mL/kg/day and about 30 mL/kg/day, wherein the second standardized human milk formulation comprises between about 0.83 kcal/mL and 0.95 kcal/mL, between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat; and (c) enterally administering the second standardized human milk formulation on day 4 to at least day 10 of the treatment regimen and/or the subject’s life, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 4 through 10 of the treatment regimen and/or the subject’s life or until a target feeding volume of about 160 mL/kg/day is achieved.

[0050] Also provided herein is a method for providing nutrition to a premature or very low birthweight infant when mother’s own milk (MOM) is not available, comprising: (a) enterally administering to a subject that is a premature or very low birth weight infant a first standardized human milk formulation on days 1-3 of the treatment regimen and/or the subject’s life at between about 20 mL/kg/day and 30 mL/kg/day, wherein the first standardized human milk formulation comprises between about 0.75 kcal/mL and 0.87 kcal/mL, between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat; and (b) enterally administering a second standardized human milk formulation on days 4-10 of the treatment regimen and/or the subject’s life, wherein the second human milk composition is enterally administered on day 4 of the treatment regimen and/or the subject’s life at between about 40 mL/kg/day and 60 mL/kg/day, wherein the feeding volume is advanced daily by between about 20 mL/kg/day and 30 mL/kg/day on days 5 through 10 of the treatment regimen and/or the subject’s life or until a feeding volume of about 160 mL/kg/day is achieved, and wherein the second standardized human milk formulation comprises between about 0.83 kcal/mL and 0.95 kcal/mL, between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat.

[0051] In some embodiments, prior to at least one administration of the fortified MOM, the method includes increasing the total calorie content of the fortified MOM by between about 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) by adding an amount of a human cream composition to the fortified MOM, wherein the human cream composition comprises about 2.5-2.6 kcal/mL and about 25% fat. In certain embodiments, prior to at least one administration of the first and/or the second standardized human milk formulation, the method includes increasing the total calorie content of the first and/or the second standardized human milk formulation by between about 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) by adding an amount of a human cream composition to the fortified MOM, wherein the human cream composition comprises about 2.5-2.6 kcal/mL and about 25% fat.

[0052] In particular embodiments, the first and/or the second standardized human milk formulation is formulated or manufactured from pooled human milk, wherein the pooled human milk is pooled from at least 10, 25, 50, 100, or more individual donors. In certain embodiments, the first and/or the second standardized human milk formulation comprises at least 10, 20, 50, 100, 150 or more human milk oligosaccharides (HMOs), wherein the human milk oligosaccharides comprise one or more sialylated HMOs.

BRIEF DESCRIPTION OF THE DRAWINGS

[0053] FIG. 1 provides a flow diagram for an exemplary feeding protocol for enteral feedings.

DETAILED DESCRIPTION

[0054] Provided herein are methods and protocols for providing enteral and/or oral nutrition to a premature or very low birth weight infant. In some aspects, the provided feeding protocols are designed, at least in part, to provide enteral or oral nutrition in more complete amounts and at earlier stages in the subject’s, e.g., the premature infant’s, life, such as compared to alternative feeding protocols. In certain embodiments, the provided methods and protocols administer enteral nutrition as early as day 1 of life and advance the volume and nutritional content as early as day 3 or 4 of the subject’s life. In some aspects, the provided feeding protocols provide nutrition that is consistent with an exclusive human milk diet. In certain embodiments, the provided methods and protocols provide enteral or oral nutrition that is free or essentially free of non-human milk proteins, fats, and carbohydrates.

[0055] Nutritional support of the premature infant is of great importance since short term survival and long-term growth and development are at stake. While improvements in the field of nutrition have been made via the introduction of parenteral nutrition and milk fortifiers, malnourishment still continues to contribute to morbidity and mortality in this patient population. What is needed in the art are new feeding protocols to improve growth and health outcomes of premature and very low birthweight infants.

[0056] The provided feeding protocols address these needs. The provided feeding protocols safely begin to deliver enteral nutrition to a premature or very low birthweight infant as early as day 1 of life. In addition, the protocols contain steps for progressively and safely increasing feeding volume to provide an optimal caloric and protein intake over the first days of life, resulting in optimal growth and functional development not observed with alternative feeding protocols. In addition, the provided methods avoid the use of bovine- based or other non-human milk-based formulas and fortifiers, thus reducing the risk of adverse events that may slow growth or development or jeopardize the infant’s health.

[0057] Human milk is generally the food of choice for preterm and term infants because of its nutritional composition and immunologic benefits. The nutritional value of raw or conventionally-processed donor milk, however, varies and, in most instances, is not sufficient to meet the needs of preterm or very low birth weight infants. For these and other reasons, use of milk from the infant's own mother has become the preferred nutritional approach in many modem neonatal intensive care units (NICUs). Even the mother's own milk, however, is not nutritionally sufficient for the premature infant. Thus, in some embodiments, the provided feeding protocols administer human milk from the preterm infant’s mother that is fortified with human milk-based fortifiers, such as to provide additional human milk proteins, fats, and carbohydrates. In certain embodiments, the present inventors have not only recognized that higher levels of fortified human milk can be safely delivered at earlier timepoints than alternative existing protocols, but that this strategy yields surprisingly superior growth e.g., as reflected by head circumference, length, and weight gain.

[0058] Also provided herein are feeding protocols for preterm infants when mother’s own milk (MOM) is unavailable. Particular embodiments contemplate that MOM for the premature or very low birth weight infant has a higher protein content that milk from the mother of a term infant. When MOM is not available, the protein content of donor milk, e.g., pasteurized donor milk, might not provide adequate protein levels to meet the nutritional needs, e.g., protein, of the premature or very low birth weight infant. In some aspects, the provided feeding protocols utilizes standardized, high calorie, high protein human milk formulations instead of donor milk or term, and even instead of fortified donor milk. The standardized, high calorie, high protein human milk formulations consistently deliver nutrients across feedings.

[0059] In some aspects, the provided methods and protocols result in improved gains in growth metrics, e.g., improved increases in head circumference, length, and weight, as compared to the gains seen in alternative methods and protocols, e.g., protocols that utilize non-human milk based nutrition (e.g., bovine nutrition) or with slower volume advances and/or fewer calories or protein that are administered early on. In certain embodiments, the provided methods and protocols result in improved neurodevelopment as compared to alternative methods and protocols for providing enteral nutrition. In certain embodiments, the provided methods and protocols result in improved long term metabolic characteristics, such as reduced risked of metabolic diseases later in life (e.g., obesity or diabetes).

[0060] In some aspects, owing in part to transient gut immaturity, many preterm and very low birth weight infants receive parenteral nutrition (PN) in the first few weeks of life. Yet even with PN, providing enough protein and energy to sustain optimal growth in such infants has remained challenging. Furthermore, PN is associated with a risk for complications such as parenteral nutrition-associated liver disease (PNALD) and central line- associated bloodstream infections (CLASBIs), which carry a high morbidity and mortality.

In some embodiments, the provided feeding protocols allow for a faster transition away from PN than alternative feeding protocols, and thus reduces risks for developing these complications and improving clinical outcomes. In some aspects, the earlier transition from PN made possible by the provided feeding protocols is earlier than what would have been considered safe or possible based on current standard or existing feeding protocols. In some aspects, this early removal of PN is also associated with improved short and long-term neurodevelopment. In certain embodiments, premature and VLBW infants transition off of PN at an earlier age under the provided feeding protocol as compared to alternative feeding protocols.

[0061] The provided methods include oral or enteral administration of human milk compositions that may include human milk-based fortifier or standardized human milk formula that are produced or manufactured from human milk pooled from multiple donors. In certain aspects, at least in part because these human milk compositions are produced or manufactured from pooled human milk, the human milk compositions contain human milk oligosaccharides with a greater degree of diversity than what would be expected from any milk produced from a single donor. In some aspects, the diversity of HMOs present in the fortifier or standardized formula promote a healthy gut microbiome development, neurodevelopment, and reduce the risk of adverse events to a greater degree than human milk compositions originating from human milk compositions produced from human milk from pooled from fewer donors, e.g., ten or fewer donors, or a single donor. [0062] The provided protocols allow for feeding that begins on the first day of life if the premature infant is clinically stable. Particular embodiments contemplate that a clinician may select a specific feeding protocol among the provided protocols based on an assessment of risk factor, such as including but not limited to gestational age, birthweight, intrauterine growth restriction, APGAR score, need for cardiopulmonary resuscitation (CPR) after delivery, maternal substance abuse during pregnancy (e.g., cocaine or methamphetamine), need for pressor support of blood pressure, presence of hemodynamically significant patent ductus arteriosus (PDA). Such assessments may be performed by those of skill as a matter of routine.

[0063] All publications, including patent documents, scientific articles, and databases, referred to in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication were individually incorporated by reference. If a definition set forth herein is contrary to or otherwise inconsistent with a definition set forth in the patents, applications, published applications, and other publications that are herein incorporated by reference, the definition set forth herein prevails over the definition that is incorporated herein by reference.

[0064] The section heading used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

I. FEEDING PROTOCOLS

[0065] Provided herein are protocols (also referred to herein as treatment regimens) for orally or enterally delivering, providing, administering, nourishing, or feeding nutrition to a subject in need thereof, e.g., a premature or very low birth weight (VLBW) infant. In particular embodiments, the provided protocols allow for earlier, more complete oral or enteral nutrition than alternative existing protocols. In particular embodiments, the protocols result in one or more improvements on outcomes relating to short-term or long-term growth, development, or health.

[0066] In some embodiments, the protocols are or include oral or enteral administration of human milk based nutrition, e.g., one or more human milk compositions, during the first days of life, e.g., as early as day 1 of life. In certain embodiments, the protocols may be used to transition the subject, e.g., the premature or very low birth weight infant, off of parenteral nutrition (PN). [0067] The provided feeding protocols provide, deliver, or administer optimal oral or enteral nutrition to a subject in need thereof. In some embodiments, the provided protocols are useful for transitioning a subject in need thereof from parenteral nutrition (PN) to oral or enteral nutrition. In certain embodiments, the subject in need thereof is a preterm or premature infant. In particular embodiments, the subject is a late preterm infant, e.g., an infant bom between 34 and 36 weeks gestation; a moderately premature infant, e.g., an infant bom between 32 and 34 weeks gestation; a very preterm infant, e.g., an infant bom at or less than 32 weeks gestation; or an extremely preterm infant, e.g., an infant bom at or less than 25 weeks gestation.

[0068] In some embodiments, the subject in need thereof is one or more of a premature infant, a low birth weight (LBW) infant, a very low birth weight (VLBW), an extremely low birthweight (ELBW) infant, or a small for gestational age (SGA) infant, e.g., an SGA infant that has experienced intra-uterine growth retardation (IUGR). In particular embodiments, the subject is a VLBW infant, e.g., an infant having a birth weight less than

1.500 g. In certain embodiments, the subject is an ELBW infant, e.g., an infant with a birth weight of less than 1,000 g. In some embodiments, the subject has a birthweight of less than

2.500 g, less than 2,000 g, less than 1,800 g, less than 1,500 g, less than 1,250 g, less than 1,000 g, or less than 750 g. In particular embodiments, the subject has a birthweight of between 500 g and 1,250 g.

[0069] In particular embodiments, the provided feeding protocols begin or start on day 1 to 3 of the subject’s life. In certain embodiments, the feeding protocol begins, starts, or is initiated with PN. In particular embodiments, the feeding protocol begins, starts, or is initiated after PN has started, begun, or initiated. In particular embodiments, the feeding protocol begins on day 1 of life. In certain embodiments, the feeding protocol begins on day 2 of life. In certain embodiments, the feeding protocol begins on day 3 of life. In certain embodiments, the subject receives trophic feedings prior to the start, beginning, or initiation of the feeding protocol if the feeding protocol starts, begins, or initiates after day 1 of the subject’s life. In certain embodiments, the trophic feedings are enteral feedings. In some embodiments, the trophic feedings are enteral feedings with a feeding volume of less than 30 mL/kg/day, less than 20 mL/kg/day, or less than 10 mL/kg/day.

[0070] In some embodiments, the protocol is an exclusive human milk diet, thus, all macronutrients, e.g., proteins, fats, and carbohydrates, are from or originate or are enriched, isolated, or purified from human milk. In some embodiments, the protocol begins on day 1 of the feeding protocol and/or the subject’s life. In certain embodiments, protocol is initiated with feeds at an initial feeding volume of between 10 mL/kg/day and 30 mL/kg/day. In certain embodiments, the feeding volume is increased between 0 mL/kg/day and 30 mL/kg/day daily each day for the duration of the protocol until a target feeding volume, e.g., of at least 160 mL/kg/day, is achieved. In certain embodiments, the feeding volume is increased at a consistent rate, e.g., 10, 20, or 30 mL/kg/day, throughout the protocol. In some embodiments, the feeding volumes are increased at different rates during the protocol, e.g., an initial rate of at least 0, 10, 20 mL/kg/day and is then increased, such as once or twice or more than twice during the protocol, e.g., to a rate of at least 10, 20, or 30 mL/kg/day.

[0071] In certain embodiments, the protocols include one or more steps to orally or enterally deliver, provide, or administer nutrition, e.g., human milk-based nutrition, to the subject in need thereof, e.g., the premature or VLBW infant. In particular embodiments, the protocols are consistent with an exclusive human milk diet, in that all or essentially all of the proteins, fats, and carbohydrates that are enterally or orally administered, delivered, or provided to the subject are human milk proteins, fats, and carbohydrates. In certain embodiments, all oral and/or enteral nutrition provided, delivered, or administered to the subject in accordance with the provided protocol contains only proteins, fats, and carbohydrates that are human milk proteins, fats, and carbohydrates. In certain embodiments, all oral and/or enteral nutrition provided, delivered, or administered to the subject in accordance with the provided protocol is free or essentially free of proteins, fats, and carbohydrates that are non-human milk proteins, fats, and carbohydrates.

[0072] In some embodiments, minerals, vitamins, or both are added to the nutrition, e.g., the human milk-based nutrition. In certain embodiments, the minerals, vitamins, or both are from a non-human milk source. In particular embodiments, the minerals, vitamins, or both are from a non-human milk source but are identical to minerals or vitamins that are found in or that may naturally be present in human milk. Minerals may include, but are not limited to, calcium, phosphorus, magnesium, iron, zinc, copper, sodium, potassium, and chloride.

[0073] In particular embodiments, PN is administered to the subject at the beginning of the feeding protocol or treatment regimen. In certain embodiments, the PN and enteral feedings are administered to the subject during the treatment regimen or feeding protocol to maintain a total fluid intake of at least 100 mL/kg/day. In certain embodiments, the PN and enteral feedings are administered to the subject to maintain a total fluid intake of between about 120 mL/kg/day and 160 mL/kg/day. In some embodiments, the PN and enteral feedings are administered to the subject to maintain a total fluid intake of between about 150 mL/kg/day and 160 mL/kg/day. In certain embodiments, when the enteral feeding volumes are increased during the feeding protocol or treatment regimen, the PN is decreased by the same volume to maintain the total intake volume, e.g., of at least 100 mL/kg/day, of 120-160 ml/kg/day, or 150-160 mL/kg/day. In certain embodiments, when the target or desired fluid or nutritional intake is achieved from the enteral feeding, the PN is discontinued. In some embodiments, the PN is discontinued when the subject is enterally administered a feeding volume that is greater than 100 mL/kg/day. In particular embodiments, the PN is discontinued when the subject is enterally administered a feeding volume that is at least 120 mL/kg/day. In certain embodiments, the PN is discontinued when the subject is enterally administered a feeding volume that is between 100 mL/kg/day and 120 mL/kg/day.

[0074] In some embodiments, PN begins at the beginning of the feeding protocol. In certain embodiments, PN begins on day 1 of the subject’s life. In certain embodiments, the PN and enteral feedings are administered to the subject during the treatment regimen or feeding protocol to maintain a total protein or amino acid intake of at least 3 mg/kg/day. In certain embodiments, the PN and enteral feedings are administered to the subject during the treatment regimen or feeding protocol to maintain a total protein or amino acid intake of between about 3 mg/kg/day and about 4 mg/kg/day. In some embodiments, the PN and enteral feedings are administered to the subject during the treatment regimen or feeding protocol to maintain a total protein or amino acid intake of about 3 mg/kg/day. In particular embodiments, the PN and enteral feedings are administered to the subject to maintain a total protein or amino acid intake of about 4 mg/kg/day. In certain embodiments, when the subject’s protein or amino intake from enteral feeding is increased during the feeding protocol or treatment regimen, the PN is decreased to maintain the total protein or amino acid intake, e.g., between 3 mg/kg/day and 4 mg/kg/day. In particular embodiments, when the target or desired protein or amino acid intake is achieved from the enteral feeding, the PN is discontinued.

[0075] In some embodiments, the PN and enteral feedings are administered to the subject during the treatment regimen or feeding protocol to maintain a total fat or lipid intake, e.g., of at least 6 mg/kg/day. In certain embodiments, when the subject’s fat or lipid intake from enteral feeding is increased during the feeding protocol or treatment regimen, the PN is decreased to maintain the total fat or lipid intake, e.g., about 6 mg/kg/day. In some embodiments, when the target or desired fat or lipid intake is achieved from the enteral feeding, the PN is discontinued.

[0076] In some embodiments, orally or enterally providing, delivering, or administering human milk-based nutrition to a subject, e.g., a premature or VLBW infant, is or includes administering one or more human milk compositions. In certain embodiments, the human milk composition is or includes mother’s own milk (MOM), i.e., milk from the subject’s mother; fortified MOM such as MOM fortified with a human milk-based fortifier, and/or a standardized human milk formulation. Human milk-based fortifiers are known in the art and include any of the human milk-based fortifiers described herein, e.g., PROLACT +6™ human milk fortifier. In certain embodiments, the protocol includes steps for orally or enterally providing, delivering, or administering more than one human milk composition, e.g., a first human milk composition and a second human milk composition.

[0077] In certain embodiments, MOM is used if MOM is available. In particular embodiments, if MOM is available but in an insufficient amount to meet a target feeding volume, donor milk is used for feedings where MOM is unavailable. In certain embodiments, when MOM or donor milk is administered in a feeding volume of 60 mL/kg/day (e.g., when the feeding volume has advanced to at least 60 mL/kg/day) the MOM or donor milk is fortified with a human milk based fortifier, e.g., a fortifier containing 144 kcal/100 mL, 5.9 g/100 mL human milk protein, 9.3 g/100 mL human milk fat, and 8.9 g/100 mL of human milk carbohydrate such as PROLACT+6™, such as to about 26 kcal/ounce (87.9 kcal/100 mL) and/or to add 6 kcal/ounce (20.3 kcal/100 mL) to the MOM or donor milk. If MOM is not or will not be available, a standardized human milk formulation is used, for example a standardized human milk formulation containing 24 kcal/ounce, such as a standardized human milk formulation that contains 82.1 kcal/100 mL, 2.4 g/100 mL human milk protein,

8.1 g/100 mL human milk carbohydrate, and 4.5 g/100 mL human milk fat, e.g., PROLACT RTF24™, for feedings at under 60 mL/kg/day and a standardized human milk formulation containing 26 kcal/ounce such as a standardized human milk formulation that contains 88 kcal/100 mL, 2.6 g/100 mL human milk protein, 8.1 g/100 mL human milk carbohydrate, and

5.1 g/100 mL human milk fat, e.g., PROLACT RTF26™ for feedings over 60 mL/kg/day.

[0078] Particular embodiments contemplate that a premature or VLBW infant may tolerate feedings with a high calorie content, provided the source of the calories are human milk-based, e.g., from one or more of human milk protein, human milk carbohydrates, or human milk fats. In some embodiments, a human milk composition may contain a calorie content between about 18 kcal/ounce and 36 kcal/ounce (60.9 kcal/100 mL and 121.7 kcal/100 mL). In certain embodiments, the human milk composition has a calorie content of between about 18 kcal/ounce and 30 kcal/ounce (60.9 kcal/100 mL and 101.4 kcal/100 mL). In certain embodiments, the human milk compositions may be further supplemented, e.g., with a human cream composition, to further increase the calorie content, e.g., up to 34 kcal/ounce (115 kcal/100 mL) or higher.

[0079] In certain embodiments oral or enteral delivery or administration of the human milk-based nutrition is initiated within the first few days of the subject’s life, e.g., within the first five days (within the first 120 hours), within the first four days (within the first 96 hours), within the first three days (within the first 72 hours), within the first two days (within the first 48 hours), or on the first day (within the first 24 hours) of life.

[0080] In some embodiments, the human-milk based nutrition is administered or delivered in a feeding volume that is increased daily over a treatment regimen or treatment phase. In some embodiments, the protocol includes one or more steps for orally or enterally providing nutrition to the subject, and the feeding volume is increased over a period of two, three, four, five, six, or greater than six days.

[0081] In particular embodiments, the delivery or administration of human milk- based nutrition is or includes delivery or administration of one or more human milk compositions, e.g., one or more human milk compositions described herein, such as in Section-II. In some embodiments, a first human milk composition is enterally or orally administered, delivered, or fed to the subject for a treatment phase, e.g., for a first phase of the protocol, and then one or more subsequent human milk compositions are administered to the infant, such as in a second phase of the protocol. In certain embodiments, the feeding volume (e.g., mL/kg/day of enterally administered nutrition) remains consistent during the first phase of the feeding protocol. In some embodiments, the feeding volume increases during the second phase of the protocol. In certain embodiments, the protocol has a third phase where full enteral or oral feedings for the first or second human milk composition are administered to the subject.

[0082] In some embodiments, the protocol includes one or more steps for enterally or orally delivering, providing, or administering human milk-based nutrition, e.g., one or more human milk compositions, to a subject in need thereof, e.g., a premature of VLBW infant. In certain embodiments, the human milk-based nutrition, e.g., the one or more human milk compositions, are administered to the subject at the beginning (day 1) of the protocol. In some embodiments, the human milk-based nutrition, e.g., the one or more human milk compositions, are administered to the subject beginning for at least 5, 6, 7, 8, 9, or 10 consecutive days, e.g., of the protocol. In some embodiments, the protocol begins on day 1 to day 3 of the subject’s life. In certain embodiments, once the protocol begins, the enteral feeding volume is advanced, e.g., by 10-30 mL/kg/day, for at least 4, 5, 6 or more than 6 consecutive days.

[0083] In certain embodiments, the human milk-based nutrition, e.g., the one or more human milk compositions, are administered to the subject beginning at the first day (day 1) of the subject’s life. In some embodiments, the human milk-based nutrition, e.g., the one or more human milk compositions, are administered to the subject beginning for at least the first five days, the first seven days, or the first ten days the subject’s life. In certain embodiments, the one or more human milk compositions are enterally or orally delivered, provided, or administered for at least the subject’s first ten days of life.

[0084] In some embodiments, the feeding volume is advanced or increased until a target feeding volume is achieved. In some embodiments, the target feeding volume is between 110 and 200 mL/kg/day, 140 and 180 mL/kg/day, 160 and 180 mL/kg/day, or about 160 mL/kg/day. In certain embodiments, the target feeding volume is at least or about 160 mL/kg/day.

[0085] Feeding progresses so long as feeds are tolerated. If any signs of intolerance, fortification or feeding volume may be reduced. In some aspects, feeding intolerance may include, but is not limited to, any one or more of the following: gastric residuals, abdominal distension, onset of crises of apnea/bradycardia, inability to achieve a full enteral feeding volume of 140 ml/kg/d by 10 days after initiation of enteral feedings, emesis, visible bowel loops, or increased abdominal girth. Particular embodiments contemplate that feeding tolerance and intolerance may be determined or identified as a matter of routine by those of skill in the art.

[0086] In certain embodiments, the feeding volume is increased on one or more days. In certain embodiments, the feeding volume is increased on two or more consecutive days, e.g., as tolerated by the subject. In some embodiments, the feeding volume is gradually increased over a set treatment phase or until a target feeding volume is reached. In some embodiments, the feeding volume is increased daily for a treatment regimen or feeding protocol starting at day 1, day 2, day 3, day 4, day 5, day 7, or after day 7 until a target feeding volume is achieved. In certain embodiments, the feeding volume is increased daily beginning at day 1, day 2, day 3, or day 4 of life. In particular embodiments, the feeding volume is increased daily beginning at day 1 or day 2 of the feeding protocol. In some embodiments, the feeding volume is gradually increased over a set treatment phase that between 1 and 10, 3 and 10, or 3 and 7 days in length. In certain embodiments, the feeding volume is gradually increased until a target feeding volume is achieved, e.g., at least or about 160 mL/kg/day. In some embodiments the feeding volume is increased at a rate of no more than 30 mL/kg/day. In certain embodiments, the feeding volume is increased by less than 30 mL/kg/day. In particular embodiments, the feeding volume is increased at a rate of at least 10 mL/kg/day or at least 20 mL/kg/day. In certain embodiments, the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day or 20 mL/kg/day and 30 mL/kg/day.

[0087] In some embodiments, the feeding volume is increased at a rate of between 20 ml/kg/day and 30 mL/kg/day beginning on or after day 4 of the subject’s life. In particular embodiments, the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day beginning on day 1 of the feeding protocol. In some embodiments, the feeding volume is increased at a rate of between 20 mL/kg/day and 30 mL/kg/day beginning on day 1 of the feeding protocol.

[0088] In particular embodiments, the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day during a first phase of the feeding protocol, and then increased at a rate of between 20 mL/kg/day and 30 mL/kg/day at a second or subsequent phase of the feeding protocol. In certain embodiments, the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day during days 1-3 of the feeding protocol and/or during days 1-3 of life, and then is increased at a rate of between 20 mL/kg/day and 30 mL/kg/day, such as until day 10 of the feeding protocol and/or day 10 of life, or until a target feeding volume is achieved, e.g., at least or about 160 mL/kg/day.

[0089] In some embodiments, a human cream composition, e.g., a composition that includes cream from a pooled human milk source such as, for example, PROLACT CR, may be added to one or more enteral or oral feedings associated with any of the feeding protocols described herein. Unless otherwise stated, the human cream composition is included with the feeding in addition to any other fortifiers and does not replace any of the fortifiers used in the feeding. In certain embodiments, the human cream composition is added to a feeding as needed, e.g., based on the determination of a clinician. In particular embodiments, the human cream composition may be added to one or more feedings associated with a feeding protocol described herein if the subject displays slow or poor growth, if the subject has a high energy expenditure, if the subject has a high-volume milk producing mother, if MOM is low calorie, e.g., below 20 kcal/ounce (67.6 kcal/100 mL), if feeds are pumped longer than 30 minutes, or if the one or more feedings are or include continuous feeds. In certain embodiments, the human cream composition is administered if growth is slow, e.g., below a rate of 15 g/kg/day.

[0090] In particular embodiments, the human cream composition includes or contains between 2 kcal/mL and 3 kcal/mL. In certain embodiments, the human cream composition includes or contains between 20% and 30% human milk fat. In some embodiments, the human cream composition is a low protein composition, e.g., contains less than 2%, 1.5%, or 1% protein. In certain embodiments, the human cream composition is or contains about 2.5- 2.6 kcal/mL and about 25% human milk fat. In certain embodiments, the human cream composition has been treated to reduce bioburden, e.g., pasteurized.

[0091] In certain embodiments, the human cream composition may be added to, mixed with, or used to supplement an enteral or oral feeding of human milk, fortified human milk, or standardized human milk formulation. In particular embodiments, the human cream composition is added to one or more enteral or oral feedings in an amount sufficient to add between 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) or between 3 kcal/100 mL and 34 kcal/ounce (10.1 kcal/100 mL and 115 kcal/100 mL) In some embodiments, the human cream composition is added to one or more enteral or oral feedings in an amount sufficient to add about or at least 1 kcal/ounce (3.4 kcal/100 mL), 2 kcal/ounce (6.7 kcal/100 mL), 3 kcal/ounce (10.1 kcal/100 mL), 4 kcal/ounce (13.5 kcal/100 mL), 5 kcal/ounce (16.9 kcal/100 mL), 6 kcal/ounce (20.3 kcal/100 mL), 7 kcal/ounce (23.7 kcal/100 mL), 8 kcal/ounce (27.1 kcal/100 mL), 9 kcal/ounce (30.4 kcal/100 mL), 10 kcal/ounce (33.4 kcal/100 mL), 11 kcal/ounce (37.2 kcal/100 mL), 12 kcal/ounce (40.6 kcal/100 mL), or more.

[0092] Particular embodiments contemplate that lipids are typically removed from PN as a matter of standard practice when the enteral feeding volume reaches about 100 mL/kg/day. In some embodiments, the human cream composition is added to enteral feedings when lipids are removed from PN, e.g., when the enteral feeding volume reaches about 100 mL/kg/day. In some embodiments, the human cream composition is added to enteral feedings when an enteral feeding volume of at least 100 mL/kg/day is achieved.

[0093] Particular embodiments contemplate that the appropriate amounts of human cream composition to achieve a desired calorie content may be determined as a matter of routine by those of skill in the art. For example, in certain embodiments, about 2 mL of the human cream composition is added to 100 mL of the human milk composition (e.g., human milk, fortified human milk, or a standardized human milk formulation) for each kcal/ounce to be added to the total calorie content. In some embodiments, about 4 mL of the human cream composition is added to 100 mL of the human milk composition to increase the total calorie content by about 2 kcal/ounce (about 6.7 kcal/100 mL). In certain embodiments, about 8 mL of the human cream composition is added to 100 mL of the human milk composition to increase the total calorie content by about 4 kcal/ounce (about 13.5 kcal/100 mL). In particular embodiments, about 12 mL of the human cream composition is added to 100 mL of the human milk composition to increase the total calorie content by about 6 kcal/ounce (about 20.3 kcal/100 mL). In certain embodiments, about 16 mL of the human cream composition is added to 100 mL of the human milk composition to increase the total calorie content by about 8 kcal/ounce (about 27.1 kcal/100 mL). In particular embodiments, about 22 mL of the human cream composition is added to 100 mL of the human milk composition to increase the total calorie content by about 10 kcal/ounce (about 33.8 kcal/100 mL).

[0094] In some embodiments, a bolus of the human milk composition may be provided prior to the one or more oral or enteral feedings. In some embodiments, appropriate amounts of human cream composition to achieve a desired calorie content of the bolus may be determined as a matter of routine by those of skill in the art. For example, in some embodiments, the bolus may be calculated as summarized in Table 1 :

Table 1: Exemplary Calculations for Human Cream Bolus

_

[0095] In certain embodiments, human milk, e.g., mother’s own milk or donor milk, is analyzed or measured for nutritional content, e.g., with a human milk analyzer. In certain embodiments, the content or density of one or more of calories, proteins, lipids, carbohydrates, sugars, lactose, glucose, vitamins, or minerals of the human milk is analyzed or measured. In some embodiments, the nutritional content of the human milk is analyzed or measured prior to fortification or enteral feeding. In particular embodiments, the fortification of the human may be adjusted based on the measurements or analysis, such as to achieve a target range or value of the infant’s daily nutrient intake, e.g., from enteral feeding and parenteral nutrition. In certain embodiments, the parenteral nutrition may be adjusted (e.g., reduced or increased) based on the nutritional measurements or analysis of the human milk. Such adjustments may include fortification according to any of the methods described herein, such as increased fortification with a human milk based fortifier (e.g., to increase the protein or calorie content of enteral feedings) or with additional fortification with a human cream composition (e.g., to increase calorie, lipid, or fat intake of enteral feedings).

[0096] In some embodiments, the nutritional content of human milk is analyzed or measured with a human milk analyzer. Particular embodiments contemplate that human milk analyzers are known and include those that are commercially available, including but not limited to MIRIS HMA™ human milk analyzer (Miris AB, Uppsala, Sweden); CREAMATOCRIT PLUS ™breast milk analyzer (Stanbio, Boeme TX, USA); SPECTRASTAR analyzers (Unity Scientific, Milford MA, USA), and MAYA (Mao Foodtech, Tel-Aviv Israel).

[0097] In some embodiments the analysis or measurements are uploaded, sent, or inputted into an interface, e.g., a computer, personal digital assistant, tablet, or smartphone, optionally along with information about the subject, e.g., weight, length, head circumference, gestational age, growth velocity, health, and/or organ function. In some embodiments, information relating to the measurements or analysis is communicated to the interface over a network, e.g., a Wi-Fi or Bluetooth network. In some embodiments, the interface processes the measurements or analysis and generates or provides recommended adjustments for fortification and/or PN to achieve a target range or value of the infant’s daily nutrient and/or caloric intake, e.g., from enteral feeding and parenteral nutrition, such as based on the infant’s health or growth status.

[0098] In particular embodiments, fortification of the human milk for enteral feeding and/or the parenteral nutrition is adjusted based on the measurements or analysis of the human milk to achieve a total daily intake of proteins or amino acids of about 3 to 4 grams of proteins or amino acids per kg of the infant’s weight per day (g/kg/day). In certain embodiments, fortification of the human milk for enteral feeding and/or the parenteral nutrition is adjusted based on the measurements or analysis of the human milk to achieve a total daily intake of proteins or amino acids of about 3 g/kg/day, e.g., during the first 10 days of life. In particular embodiments, fortification of the human milk for enteral feeding and/or the parenteral nutrition is adjusted based on the measurements or analysis of the human milk to achieve a total daily intake of proteins or amino acids of about 4 g/kg/day, e.g., until PN is discontinued such as when an enteral feeding volume of between 100 and 120 mL/kg/day is achieved. In certain embodiments, fortification of the human milk for enteral feeding and/or the parenteral nutrition is adjusted based on the measurements or analysis of the human milk to achieve a total daily intake of lipids of about 6 grams of lipids per kg of the infant’s weight per day (g/kg/day). In some embodiments, fortification of the human milk for enteral feeding and/or the parenteral nutrition is adjusted based on the measurements or analysis of the human milk to achieve a total daily calorie intake of about 120 kcal/kg/day to 160 kcal/kg/day.

A. Feeding protocols when mother’s own milk is available

[0099] In some embodiments, the protocol includes one or more steps for administering, providing, or delivering nutrition, e.g., human milk-based nutrition, to a subject in need thereof, e.g., a preterm or VLBW infant, when MOM is available. In certain embodiments, the MOM is delivered, administered, or fed orally or enterally to the infant within the first few days of life, e.g., within the first three, first two, or on the first day of life. In particular embodiments, the MOM is orally or enterally delivered or administered to the infant throughout at least the first five, seven, or ten days of life. In particular embodiments, if initial feeds are tolerated, e.g., over the first three days of life and/or the protocol, the MOM is fortified with a human milk-based human milk fortifier, e.g., a human milk-based fortifier described herein such as in Section-II-B. In certain embodiments, if initial feeds are tolerated, e.g., over the first three days of life and/or the treatment regimen, the feeding volume is increased gradually over a treatment regimen or over a phase of the protocol or treatment regimen. In particular embodiments, the feeding volume is gradually increased until a target feeding volume is achieved, e.g., at least or about 160 mL/kg/day (mL MOM or fortified MOM per kg of the subject’s body weight per day).

[0100] Particular embodiments contemplate that MOM may be available, but in some cases may not be available in sufficient quantities to achieve daily feeding goals, particularly in the first few days of life, e.g., days 1-3 of life. In some embodiments, the feeding protocol may include feedings of donated breast milk (donor milk) to achieve the target daily feeding volume when sufficient amounts of MOM are not available. In particular embodiments, the donor milk is pasteurized. In some instances, the donor milk may be fortified with a human milk-based fortifier, such as to between 24 and 28 or 24 and 30 kcal/ounce (81.2 and 94.7 or 81.2 and 101.4 kcal/100 mL). In some embodiments, the donor milk is fortified to about 26 kcal/Oz with a human milk-based fortifier. In some embodiments, the donor milk is fortified with a human milk-based fortifier to increase the calorie content by about 4 to 8 kcal/ounce or 4 to 10 kcal/ounce (13.5 to 27.1 kcal/100 mL or 13.5 to 33.4 kcal/100 mL).

[0101] In certain embodiments, the feeding protocol is initiated on day 1 to day 3 of the subject’s life, e.g., the preterm or VLBW infant’s life. In certain embodiments, MOM is enterally administered to the subject beginning on day 1 of the protocol. In some embodiments, MOM is administered at an initial feeding volume of at least 10 mL/kg/day. In particular embodiments, MOM is administered at an initial feeding volume of at least 20 mL/kg/day. In some embodiments, MOM is administered at an initial feeding volume of more than 10 or 20 mL/kg/day. In certain embodiments, MOM is administered at an initial feeding volume of between 10 mL/kg/day and 30 mL/kg/day or between 20 mL/kg/day and 30 mL/kg/day. In particular embodiments, the initial feeding volume is maintained for the first three days of the subject’s life. In some embodiments, the feeding volume is increased after the first day of the feeding protocol and/or the first day of life.

[0102] In particular embodiments, the MOM is fortified with a human-milk based fortifier prior to enteral delivery or administration on one or more days. In certain embodiments, the MOM is fortified with a human milk-based fortifier prior to enteral delivery or administration on two or more consecutive days. In certain embodiments, MOM is fortified prior to all or substantially all enteral administrations or feedings after a time period in which unfortified MOM feedings are tolerated. In some embodiments, the time period is at least one, two, or three days. In certain embodiments, the MOM is fortified after one day of feeding. In some embodiments, the MOM is fortified after three days of feedings or administrations of unfortified MOM are tolerated. In some embodiments, unfortified MOM is enterally administered, delivered, or fed to the subject, e.g., the preterm or VLBW infant, for the first three days of the subject’s life (days 1-3 of life) and fortified MOM is enterally administered, delivered, or fed to the subject after the first three days of life. In particular embodiments, unfortified MOM is enterally administered, delivered, or fed to the subject, e.g., the preterm or VLBW infant, for the first day of the subject’s life (day 1 of life) and fortified MOM is enterally administered, delivered, or fed to the subject after the first day of life. In some embodiments, enteral feedings or administrations of the fortified MOM is initiated on day 4 of life. In particular embodiments, enteral feedings or administrations of the fortified MOM is initiated on day 2 of life. In certain embodiments, enteral feeding or administrations of the fortified MOM are initiated on day 4 of life and continue for the duration of the feeding protocol, e.g., at least to seventh, tenth, or fourteenth day of the subject’s life and/or the protocol.

[0103] In particular embodiments, the MOM is fortified beginning at day 2 of the feeding protocol. In some embodiments, the MOM is fortified beginning on a day when a minimum daily feeding volume has been achieved, e.g., a volume of at least 60 mL/kg/d (60 mL feeding volume per kg body weight per day). In certain embodiments, the MOM is fortified on a day when a feeding volume of 60 mL/kg/day is achieved.

[0104] In certain embodiments, the MOM is fortified beginning at day 4 of the feeding protocol and/or at day 4 of life. In some embodiments, the MOM is optionally fortified at days 2 and 3 of the feeding protocol and/or at days 2 and 3 of life. In particular embodiments, the MOM is fortified beginning on days 2, 3, or 4 if a minimum daily feeding volume has been achieved, e.g., a volume of at least 60 mL/kg/d (60 mL feeding volume per kg body weight per day).

[0105] In some embodiments, the fortified MOM is composed of MOM and human- milk based fortifier. In certain embodiments, the human milk-based fortifier is mixed with MOM prior to the enteral or oral delivery, administration, or feeding. In certain embodiments, the human milk-based fortifier is mixed with MOM, and the resulting fortified MOM is orally or enterally delivered, administered, or fed to the subject.

[0106] In certain embodiments, the fortified MOM has at least 20 mg/mL protein, e.g., human milk protein. In certain embodiments, the fortified MOM has at least 21 mg/mL, 22 mg/mL, 23 mg/mL, 24 mg/mL, 25 mg/mL, 26 mg/mL, 27 mg/mL, 28 mg/mL, 29 mg/mL, 30 mg/mL, 32 mg/mL, 34 mg/mL, or 36 mg/mL protein, e.g., human milk protein. In some embodiments, the fortified MOM has between or between about 20 mg/mL and 40 mg/mL protein, e.g., human milk protein. In certain embodiments, the fortified MOM has between or between about 25 mg/mL and 35 mg/mL protein, e.g., human milk protein. In some embodiments, the fortified MOM has between or between about 27 mg/mL and 32 mg/mL protein, e.g., human milk protein.

[0107] In particular embodiments, the fortified MOM has at least 0.8 kcal/mL. In particular embodiments, the fortified MOM has at least 0.82 kcal/mL, 0.84 kcal/mL, 0.86 kcal/mL, 0.88 kcal/mL, 0.90 kcal/mL, 0.92 kcal/mL, 0.94 kcal/mL, or 0.96 kcal/mL. In certain embodiments, the fortified MOM has between 0.80 kcal/mL and 1.1 kcal/mL, 0.85 kcal/mL and 0.95 kcal/mL, 0.90 kcal/mL and 1.00 kcal/mL, 0.95 kcal/mL and 1.05 kcal/mL, or 1.00 kcal/mL and 1.10 kcal/mL. In some embodiments, the fortified MOM has at least 0.85 kcal/mL. In certain embodiments, the fortified MOM has between 0.85 kcal/mL and 0.95 kcal/mL.

[0108] In certain embodiments, the fortified MOM has at least 23 kcal/ounce In particular embodiments, the fortified human milk composition has at least 24 kcal/ounce, 25 kcal/ounce, 26 kcal/ounce, 27 kcal/ounce, or 28 kcal/ounce In certain embodiments, the fortified MOM has between 24 kcal/ounce and 30 kcal/ounce, 24 kcal/ounce and 28 kcal/ounce, 25 kcal/ounce and 27 kcal/ounce, 27 kcal/ounce and 29 kcal/ounce, or 29 kcal/ounce and 31 kcal/ounce In some embodiments, the fortified MOM has at least 25 kcal/ounce In certain embodiments, the fortified MOM has between 25 and 28 kcal/ounce In particular embodiments, the fortified MOM has about 20 kcal/ounce.

[0109] In certain embodiments, the fortified MOM has at least 77.8 kcal/100 mL. In particular embodiments, the fortified human milk composition has at least 81.2 kcal/100 mL, 84.5 kcal/100 mL, 87.9 kcal/100 mL, 91.3 kcal/100 mL, 94.6 kcal/100 mL, 98.1 kcal/100 mL, or 101.4 kcal/100 mL. In certain embodiments, the fortified MOM has between 81.2 kcal/100 mL and 101.4 kcal/100 mL, 81.2 kcal/100 mL and 94.7 kcal/100 mL, 84.5 kcal/100 mL and 91.3 kcal/100 mL, 91.2 kcal/100 mL and 98.1 kcal/100 ML, or 98.1 kcal/100 mL and 104.8 kcal/100 mL. In some embodiments, the fortified MOM has at least 84.5 kcal/100 mL. In certain embodiments, the fortified MOM has between 84.5 and 94.7 kcal/100 mL.

[0110] In particular embodiments, human milk based fortifier has been added to MOM or donor milk to increase the calorie content of the MOM or donor milk by at least 4 kcal/ounce (13.5 kcal/100 mL), 5 kcal/ounce (16.9 kcal/100 mL), 6 kcal/ounce (20.2 kcal/100 mL), 7 kcal/ounce (23.7 kcal/100 mL), 8 kcal/ounce (27.1 kcal/100 mL), 9 kcal/ounce (30.4 kcal/100 mL), or 10 kcal/ounce (33.8 kcal/100 mL). In some embodiments, the human milk- based fortifier has been added to MOM or donor milk to increase the calorie content of the MOM or donor milk by at least 4 kcal/ounce (13.5 kcal/100 mL). In certain embodiments, the human milk-based fortifier has been added to MOM or donor milk to increase the calorie content of the MOM or donor milk by at least 6 kcal/ounce (20.2 kcal/100 mL. In certain embodiments, the human milk-based fortifier has been added to MOM or donor milk to increase the calorie content of the MOM or donor milk by at least 8 kcal/ounce (27.1 kcal/100 mL). [0111] In certain embodiments, the human milk-based fortifier has been added to MOM or donor milk in an amount sufficient for the fortified MOM to have a calorie content that is at least 4 kcal/ounce (13.5 kcal/100 mL), 5 kcal/ounce (16.9 kcal/100 mL), 6 kcal/ounce (20.2 kcal/100 mL), 7 kcal/ounce (23.7 kcal/100 mL), 8 kcal/ounce (27.1 kcal/100 mL), 9 kcal/ounce (30.4 kcal/100 mL), or 10 kcal/ounce (33.8 kcal/100 mL) greater than the MOM or donor milk prior to fortification. In particular embodiments, the human milk-based fortifier has been added to MOM or donor milk in an amount sufficient for the fortified MOM to have a calorie content that is at least 4 kcal/ounce (13.5 kcal/100 mL) greater than the MOM or donor milk prior to fortification. In some embodiments, the human milk-based fortifier has been added to MOM or donor milk in an amount sufficient for the fortified MOM to have a calorie content that is at least 6 kcal/ounce (20.2 kcal/100 mL greater than the MOM or donor milk prior to fortification. In particular embodiments, the human milk- based fortifier has been added to MOM or donor milk in an amount sufficient for the fortified MOM to have a calorie content that is at least 8 kcal/ounce (27.1 kcal/100 mL) greater than the MOM or donor milk prior to fortification.

[0112] In some embodiments, the fortified MOM has at least 0.85 kcal/mL and at least 25 mg/mL protein, e.g., human milk protein. In certain embodiments, the fortified MOM has at least 0.85 kcal/mL and at least 27 mg/mL protein, e.g., human milk protein. In some embodiments, the fortified MOM has between 0.85 kcal/mL and 0.95 kcal/mL and between 27 and 32 mg/mL protein, e.g., human milk protein.

[0113] In particular embodiments, the fortified MOM has at least 24 kcal/ounce (81.2 kcal/100 mL) and at least 25 mg/mL protein, e.g., human milk protein. In certain embodiments, the fortified MOM has at least 25 kcal/ounce (84.5 kcal/100 mL) and at least 27 mg/mL protein, e.g., human milk protein. In some embodiments, the fortified MOM has between 25 kcal/ounce and 28 kcal/ounce (84.5 and 94.7 kcal/100 mL) and between 27 and 32 mg/mL protein, e.g., human milk protein. In particular embodiments, the fortified MOM has about or at least 26 kcal/ounce (87.9 kcal/100 mL) and about or at least 29 mg/ml human milk protein.

[0114] In particular embodiments, the protocol includes one or more steps of gradually increasing the feeding volume of a human milk composition, e.g., MOM or fortified MOM, over a treatment phase. In certain embodiments, the initial feeding volume is increased to a target feeding volume over a period time, e.g., a period of 4 days, 5 days, 6 days, 7 days, or more than 7 days. In some embodiments, an initial feeding volume of between 20 mL/kg/day and 30 mL/kg/day is increased to a target feeding volume of about 160 mL/kg/day over a period of 4 days, 5 days, 6 days, or 7 days, or more than 7 days. In some embodiments, the feeding volume is increased at a rate of between 20 mL/kg/day and 30 mL/kg/day over a period of between 4 days and 7 days.

[0115] In certain embodiments, exemplary feeding protocols for when MOM is available are summarized in Tables El, E2, E3, E7, E9, and Ell.

[0116] In some embodiments the protocol includes one or more steps for enterally feeding, delivering, or administering to a subject in need thereof, e.g., a premature or very low birthweight infant, between 20 mL/kg/day and 30 mL/kg/day MOM on the first three days (days 1-3) of the feeding protocol and/or the subject’s life. In certain embodiments, MOM is replaced with fortified MOM on day 4 of the feeding protocol and/or the subject’s life. In particular embodiments, fortified MOM is enterally fed, delivered, or administered to the subject beginning at day 4 of the feeding protocol and/or the subject’s life and is used until at least day 10 of the feeding protocol and/or the infant’s life. In some embodiments, the subject is enterally fed, delivered, or administered between 40 mL/kg/day and 60 mL/kg/day fortified MOM. In certain embodiments, the enteral delivery, administration, or feeding of the fortified MOM is increased from an initial feeding volume of between 40 mL/kg/day and 60 mL/kg/day to a target volume of about 160 mL/kg/day at a rate of between 20 mL/kg/day and 30 mL/kg/day. In particular embodiments, the fortified MOM has about or at least 25 kcal/ounce (84.5 kcal/100 mL) and about or at least 25 mg/ml human milk protein. In particular embodiments, the fortified MOM has about or at least 26 kcal/ounce (87.9 kcal/100 mL) and about or at least 29 mg/ml human milk protein.

[0117] In certain embodiments, the protocol includes one or more steps for enterally feeding, delivering, or administering to a subject in need thereof, e.g., a premature or very low birthweight infant, between 10 mL/kg/day and 30 mL/kg/day MOM on the first day of the feeding protocol and/or the subject’s life. In some embodiments, the feedings are advanced or increased by between 10 mL/kg/day and 30 mL/kg/day on days 1-3 of the feeding protocol and/or the subject’s life. In some embodiments, the MOM is optionally fortified MOM on days 2 and/or 3 of the feeding protocol and/or the subject’s life. In particular embodiments, fortified MOM is enterally fed, delivered, or administered to the subject beginning at day 4 of the feeding protocol and/or the subject’s life. In particular embodiments, the feedings are advanced or increased by between 20 mL/kg/day and 30 mL/kg/day starting at day 4 of the feeding protocol and/or the subject’s life until a target feeding volume, e.g., about or at least 160 mL/kg/day. In particular embodiments, the fortified MOM has about or at least 25 kcal/ounce (84.5 kcal/100 mL) and about or at least 25 mg/ml human milk protein. In particular embodiments, the fortified MOM has about or at least 26 kcal/ounce (87.9 kcal/100 mL) and about or at least 29 mg/ml human milk protein.

[0118] In particular embodiments, the protocol includes one or more steps for (i) enterally administering, delivering, or feeding between 20 mL/kg/day and 30 mL/kg/day MOM to a subject that is a premature or VLBW infant on days 1-3 on the feeding protocol and/or the subject’s life, (ii) enterally administering, delivering, or feeding between 40 mL/kg/day and 60 mL/kg/day fortified MOM having about or at least 25 kcal/ounce (84.5 kcal/100 mL) and about or at least 25 mg/ml human milk protein and continuing to administer, deliver, or feed the fortified MOM to the subject at least until day 10 of the feeding protocol and/or the subject’s life, and (iii) increasing the feeding volume of the fortified MOM beginning on day 4 of the feeding protocol and/or the subject’s life at a rate of between 20 mL/kg/day and 30 ml/kg/day until day 10 or to a target feeding volume of about 160 mL/kg/day.

[0119] In certain embodiments, the protocol includes one or more steps for (i) enterally administering, delivering, or feeding between 10 mL/kg/day and 30 mL/kg/day MOM to a subject that is a premature or VLBW infant on day 1 of the feeding protocol and/or the subject’s life; (ii) increasing or advancing the feedings at a rate of between 10 mL/kg/day and 30 mL/kg/day over days 1-3 of the feeding protocol and/or the subject’s life or until a feeding volume of 60 mL/kg/day is achieved; (iii) optionally fortifying MOM with a human milk-based fortifier to about or at least 25 kcal/ounce (84.5 kcal/100 mL) and about or at least 25 mg/ml human milk protein on days 2 and 3 of the feeding protocol and/or the subject’s life; (iv) administering, delivering, or feeding the fortified MOM of step (iii) to the subject from day 4 onward, and (iv) increasing the feeding volume of the fortified MOM beginning on day 4 of the feeding protocol and/or the subject’s life at a rate of between 20 mL/kg/day and 30 ml/kg/day until day 10 or until a target feeding volume is achieved, e.g., about or at least 160 mL/kg/day.

B. Feeding protocols when mother’s own milk is not available [0120] In certain embodiments, the protocol includes one or more steps for administering, providing, or delivering nutrition, e.g., human milk-based nutrition, to a subject in need thereof, e.g., a preterm or VLBW infant, when mother’s own milk (MOM) is not available. Particular embodiments contemplate that MOM may be unavailable for a variety of reasons, including but not limited to an inability to produce milk by the subject’s mother, e.g., such as due to an illness, condition, or a physical inability to produce milk, incarceration or illness or other circumstances that prevent the mother from being in proximity to the infant; a risk of contamination from the mother’s own milk, such as a risk of exposure to infectious disease e.g., HIV, through the milk, or chemical contamination such as a risk of exposure to prescription or illegal drugs through the milk.

[0121] In certain embodiments, if MOM is not available, a standardized human milk formulation is orally or enterally fed, delivered, or administered to the subject, e.g., the preterm or VLBW infant. In particular embodiments, the feeding protocol is or includes orally or enterally administering the standardized human milk formulation to the subject. In certain embodiments, the standardized human milk formulation is a ready to feed formulation, e.g., contains sufficient caloric and protein content and sufficient osmolarity to be directly fed to a premature or very low birth weight infant such as without first mixing or diluting with milk or any other substance. In particular embodiments, the standardized human milk formulation is delivered, administered, or fed orally or enterally to the infant within the first few days of life, e.g., within the first three, first two, or on the first day of life. In some embodiments, the feeding protocol that involves or includes administering the standardized human milk formulation begins within the first few days of life, e.g., within the first three, first two, or on the first day of life. In particular embodiments, the standardized human milk formulation is orally or enterally delivered or administered to the infant throughout at least the first five, seven, or ten days of the feeding protocol and/or the subject’s life. In particular embodiments, the standardized human milk formulation is any suitable standardized human milk formulation described herein, e.g., in Section II-C. In certain embodiments, if initial feeds are tolerated, e.g., over the first three days of life and/or the protocol, the feeding volume is increased gradually over a treatment phase. In particular embodiments, the feeding volume is gradually increased until a target feeding volume is achieved, e.g., at least or about 160 mL/kg/day (mL of the standardized human milk formulation per kg of the subject’s body weight per day).

[0122] In particular embodiments, if MOM is not available, a first standardized human milk formulation is orally or enterally fed, delivered, or administered to the subject, e.g., the preterm or VLBW infant. In particular embodiments, the first standardized human milk formulation is delivered, administered, or fed orally or enterally to the infant within the first few days of life, e.g., within the first three, first two, or on the first day of life. In particular embodiments, the feeding protocol begins within the first three days of life. Prior to the start of the protocol, the first standardized human milk formulation may be used for trophic feedings, e.g., enteral feedings under 30 mL/mg/day. In particular embodiments, the first standardized human milk formulation is any suitable standardized human milk formulation described herein, e.g., in Section II-C. In particular embodiments, the first standardized human milk formulation is orally or enterally delivered or administered to the infant for a treatment phase until a second standardized human milk formulation is enterally or orally administered, fed, or delivered. In some embodiments, after at least one, two, or three days of the feeding protocol and/or days of the subject’s life, the first standardized human milk formulation is replaced with a second standardized human milk formulation. In certain embodiments, if initial feeds are tolerated, e.g., over the first three days of life and/or the feeding protocol, then the first standardized human milk formulation is replaced with a second standardized human milk formulation. In particular embodiments, the second standardized human milk formulation is different from the first standardized human milk formulation. In some embodiments, the first and second standardized human milk formulations are the same. In some embodiments, the second standardized human milk formulation has a greater concentration of one or more of energy (e.g., greater amount of kcal/mL), human milk protein, and/or human milk fat than the first standardized human milk formulation. In some embodiments, the second standardized human milk formulation is any suitable standardized human milk formulation described herein, e.g., in Section II-C. In some embodiments, the feeding volume of the second standardized human milk formulation is increased gradually over a treatment phase. In particular embodiments, the feeding volume is gradually increased until a target feeding volume is achieved, e.g., 160 mL/kg/day (mL of the standardized human milk formulation per kg of the subject’s body weight per day).

[0123] In particular embodiments, the feeding protocol is initiated on day 1 of the subject’s life, e.g., the preterm or VLBW infant’s life. In certain embodiments, the standardized human milk formulation is enterally administered to the subject beginning on day 1 to 3 of the subject’s life. In some embodiments, the standardized human milk formulation (e.g., a first standardized human milk formulation) is enterally administered to the subject beginning at day 1 of the feeding protocol. In certain embodiments, the standardized human milk formulation is administered at an initial feeding volume of at least 10 mL/kg/day. In some embodiments, the standardized human milk formulation is administered at an initial feeding volume of at least 20 mL/kg/day. In particular embodiments, the standardized human milk formulation is administered at an initial feeding volume of more than 10 mL/kg/day or 20 mL/kg/day. In certain embodiments, the standardized human milk formulation is administered at an initial feeding volume of between 10 mL/kg/day and 30 mL/kg/day; 10 mL/kg/day and 30 mL/kg/day; or 20 mL/kg/day and 30 mL/kg/day. In particular embodiments, the initial feeding volume is maintained for the first three days of the feeding protocol and/or the subject’s life. In certain embodiments, the feeding volume is increased beginning at days 1 to 3 of the subject’s life and/or beginning on day 1 of the feeding protocol.

[0124] In certain embodiments, a standardized human milk formulation has at least 0.85 kcal/mL and at least 25 mg/mL protein, e.g., human milk protein. In some embodiments, a standardized human milk formulation has between 0.85 kcal/mL and 0.95 kcal/mL and between 27 and 32 mg/mL protein, e.g., human milk protein. In particular embodiments, a standardized human milk formulation has or has about 26 kcal/ounce or 0.88 kcal/mL and 26 mg/mL human milk protein.

[0125] In particular embodiments, the feeding volume is increased on one or more days. In some embodiments, the feeding volume is increased on two or more consecutive days, e.g., as tolerated by the subject. In certain embodiments, the feeding volume is gradually increased over a set treatment phase or until a target feeding volume is reached. In particular embodiments, the feeding volume is increased daily for a treatment phase starting at day 1, day 2, day 3, day 4, day 5, day 7, or after day 7 until a target feeding volume is achieved. In certain embodiments, the feeding volume is increased daily beginning at from day 1 to day 4 of life or from day 1 to day 3 of life and/or the feeding protocol. In some embodiments, the feeding volume is increased daily beginning at day 1 of life and/or the feeding protocol In some embodiments, the feeding volume is gradually increased over a set treatment phase that between 1 and 10, 3 and 10, or 3 and 7 days in length. In certain embodiments, the feeding volume is gradually increased until a target feeding volume is achieved, e.g., 160 mL/kg/day. In some embodiments the feeding volume is increased at a rate of no more than 30 mL/kg/day or less than 30 mL/kg/day. In particular embodiments, the feeding volume is increased at a rate of at least 20 mL/kg/day or more than 20 mL/kg/day. In certain embodiments, the feeding volume is increased at a rate of between 10 mL/kg/day and 20 mL/kg/day; 10 mL/kg/day and 30 mL/kg/day; or 20 mL/kg/day and 30 mL/kg/day. In some embodiments, the feeding volume is increased at a rate of between 20 ml/kg/day and 30 mL/kg/day beginning on or after day 4 of the subject’s life and/or the feeding protocol. In certain embodiments, the feeding volume is increased at a rate of between 20 ml/kg/day and 30 mL/kg/day beginning on or after day 1 of the feeding protocol and/or the subject’s life.

[0126] In particular embodiments, the protocol includes one or more steps of gradually increasing the feeding volume of the standardized human milk formulation over a treatment phase. In certain embodiments, the initial feeding volume is increased to a target feeding volume over a period time, e.g., a period of 4 days, 5 days, 6 days, 7 days, or more than 7 days. In some embodiments, an initial feeding volume of between 10 mL/kg/day and 30 mL/kg/day, between 10 mL/kg/day and 30 mL/kg/day; or between 20 mL/kg/day and 30 mL/kg/day is increased to a target feeding volume of about 160 mL/kg/day over a period of 4 days, 5 days, 6 days, or 7 days, or more than 7 days. In some embodiments, the feeding volume is increased at a rate of between 10 mL/kg/day and 30 mL/kg/day, between 10 mL/kg/day and 30 mL/kg/day; or between 20 mL/kg/day and 30 mL/kg/day over a period of between 1 day and 7 days, 4 days and 7 days, or until a target feeding volume, e.g., at least or about 160 mL/kg/day is achieved.

[0127] In some embodiments, more than one standardized human milk formulations are enterally or orally administered, delivered, or fed to the subject. In particular embodiments, a first standardized human milk formulation is orally or enterally fed, delivered, or administered to the subject for a treatment phase or during a first phase of the protocol, and then a second standardized human milk formulation is administered to the subject for a second treatment phase or during a second phase of the protocol. In certain embodiments, the second formulation may have greater amounts or concentrations of one or more of energy (e.g., kcal/mL), human milk protein, human milk fat, and HMO.

[0128] In certain embodiments, exemplary feeding protocols for when MOM is not available are summarized in Tables E4-E6, E8, E10, and E12.

[0129] In particular embodiments the protocol includes one or more steps for enterally feeding, delivering, or administering to a subject in need thereof, e.g., a premature or very low birth weight infant, between 20 mL/kg/day and 30 mL/kg/day of a first standardized human milk formulation, e.g., a standardized human milk formulation described herein such as in Section II-C, beginning within the first three days of the subject’s life, e.g., at day 1 of the feeding protocol. In particular embodiments, between 20 mL/kg/day and 30 mL/kg/day of a standardized human milk formulation is enterally delivered, administered, or fed on the first day of the feeding protocol and/or on the subject’s first day of life. In particular embodiments, the standardized human milk formulation is enterally fed, delivered, or administered to the subject beginning at day 1 of the feeding protocol and/or the subject’s life and is used until at least day 3 of the feeding protocol and/or the infant’s life. In some embodiments, the subject is enterally fed, delivered, or administered a second standardized human milk formulation at between 40 mL/kg/day and 60 mL/kg/day.

[0130] In particular embodiments, the subject is enterally fed a first standardized human milk formulation at the beginning of the protocol and until a daily feeding volume of 60 mL/kg/day is achieved. In certain embodiments, a second standardized human milk formulation is then administered for the remainder of the feeding protocol. In some embodiments, the second standardized human milk formulation is a standardized human milk formulation described herein, e.g., in Section II-C. In particular embodiments, the second standardized human milk formulation has a greater or larger calorie content and/or protein content than the first standardized human milk formulation. In certain embodiments, the enteral delivery, administration, or feeding of the second standardized human milk formulation is increased from an initial feeding volume of between 40 mL/kg/day and 60 mL/kg/day to a target volume of about 160 mL/kg/day at a rate of between 20 mL/kg/day and 30 mL/kg/day. In some embodiments, the first standardized human milk formulation has between about 0.75 kcal/mL and 0.87 kcal/mL, between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat. In certain embodiments, the second standardized human milk formulation has between about 0.83 kcal/mL and 0.95 kcal/mL, between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat.

[0131] In some embodiments, provided herein is a feeding protocol including one or more steps for providing nutrition to a premature or VLBW infant when MOM is not available. In some embodiments, the protocol includes a step of enterally administering to the infant a first standardized human milk formulation comprising between about 0.75 kcal/mL and 0.87 kcal/mL, between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat at day 1 of the feeding protocol and/or the subject’s life. In certain embodiments, the first standardized human milk formulation is initially (e.g., at day 1 of the protocol and/or the subject’s life) enterally administered in an amount between about 10 mL/kg/day and about 30 mL/kg/day. In certain embodiments, on the second and third day of the protocol, e.g., at days 2 and 3 of life, either the first standardized human milk formulation or a second standardized human milk formulation comprising between about 0.83 kcal/mL and 0.95 kcal/mL, between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat are enterally administered to the infant. In certain embodiments, the feeding volume is advanced between days 1 to 3 of the feeding protocol and/or the subject’s life by between about 10 mL/kg/day and about 30 mL/kg/day. In particular embodiments, the second standardized human milk formulation is enterally administered on day 4 to at least day 10 of the feeding protocol and/or the subject’s life. In certain embodiments, the feeding volume, e.g., of the second standardized human milk formulation, is advanced by between about 20 mL/kg/day and about 30 mL/kg/day from day 4 of the feeding protocol and/or the subject’s life until a target feeding volume of about 160 mL/kg/day is achieved.

C. Exemplary Feeding Protocols

[0132] An exemplary feeding protocol is summarized in Table 2.

Table 2: Exemplary Feeding Protocol

[0133] In the exemplary protocol depicted in Table 2, the standardized human milk formulations are only administered if MOM is unavailable.

[0134] An additional exemplary feeding protocol is shown in Table 3. In some embodiments, the protocol begins on day 1 to day 3 of life. In particular embodiments, the protocol begins on day 3 of life. In some embodiments, the protocol begins on day 2 of life. In certain embodiments, to protocol begins on day 1 of life. In some embodiments, the subject receives trophic feedings prior to the start of the protocol. In some embodiments, the trophic feedings are no more than 30 mL/kg/day. In certain embodiments, the trophic feedings are no more than 20 mL/kg/day. In particular embodiments, the trophic feedings are no more than 10 mL/kg/day.

Table 3: Exemplary Feeding Protocol

[0135] In the exemplary protocol depicted in Table 3, the standardized human milk formulations are only administered if MOM is unavailable.

[0136] Another exemplary feeding protocol is depicted in Table 4. In some embodiments, the protocol begins on day 1 to day 3 of life. In particular embodiments, the protocol begins on day 3 of life. In some embodiments, the protocol begins on day 2 of life. In certain embodiments, to protocol begins on day 1 of life. In some embodiments, the subject receives trophic feedings prior to the start of the protocol. In some embodiments, the trophic feedings are no more than 30 mL/kg/day. In certain embodiments, the trophic feedings are no more than 20 mL/kg/day. In particular embodiments, the trophic feedings are no more than 10 mL/kg/day.

Table 4: Exemplary Feeding Protocol

[0137] An additional exemplary feeding protocol is depicted in Table 5. In some embodiments, the protocol begins on day 1 to day 3 of life. In particular embodiments, the protocol begins on day 3 of life. In some embodiments, the protocol begins on day 2 of life. In certain embodiments, to protocol begins on day 1 of life. In some embodiments, the subject receives trophic feedings prior to the start of the protocol. In some embodiments, the trophic feedings are no more than 30 mL/kg/day. In certain embodiments, the trophic feedings are no more than 20 mL/kg/day. In particular embodiments, the trophic feedings are no more than 10 mL/kg/day. Table 5: Exemplary Feeding Protocol

[0138] In particular embodiments, mother’s own milk (MOM) is available but not in enough quantities to supply all feedings. In some such embodiments, the amounts of MOM may be combined with donor milk, e.g., pasteurized donor milk, to provide for a sufficient feeding, and optionally the combined MOM and DM is supplemented with a human milk based fortifier, e.g., to a final calorie content of approximately 26 kcal/ounce (87.9 kcal/100 mL). In certain such embodiments, mother’s milk is collected until there is a sufficient amount for a feeding, so that feedings with MOM or DM are interspersed throughout the treatment regimen. In particular embodiments, MOM may only be available in very small amounts for all or for a duration of the treatment regimen. In certain such embodiments, small amounts of MOM may be administered to the subject, while ready to feed standardized human milk formulations, e.g., containing about 26 kcal/ounce (87.9 kcal/100 mL), are used for all other feedings.

[0139] In some embodiments, the mother’s own milk fortified with about an additional 6 kcal/ounce (20 kcal/100 mL) with a human milk-based fortifier (MOM + Fort+6) is composed of about 70% MOM and about 30% of a human milk-based fortifier that contains between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat.

[0140] In certain embodiments, the mother’s own milk fortified with about an additional 8 kcal/ounce (27 kcal/100 mL) with a human milk-based fortifier (MOM + Fort+8) is composed of about 60% MOM and about 40% of a human milk-based fortifier that contains between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat.

[0141] In some embodiments, the mother’s own milk fortified with about an additional 10 kcal/ounce (33.8 kcal/100 mL) with a human milk-based fortifier (MOM + Fort+8) is composed of about 50% MOM and about 50% of a human milk-based fortifier that contains between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat.

[0142] In certain embodiments, the standardized human milk formulation that has about 24 kcal/ounce (SHMF 24) has between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat.

[0143] In particular embodiments, the standardized human milk formulation that has about 26 kcal/ounce (SHMF 26) has between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat.

[0144] In some embodiments, the standardized human milk formulation that has about 28 kcal/ounce (SHMF 28) has between about 27 mg/mL and 30 mg/mL human milk protein, and between about 55 mg/mL and 60 mg/mL human milk fat.

[0145] In particular embodiments, a human cream composition is added to enteral feedings to add about 7 kcal/100 mL (about 2 kcal/ounce) when the enteral feeding volume reaches 100 mL/kg/day and/or when intravenous lipids are discontinued. In some embodiments, the human cream composition is or includes 20% to 30% human milk fat or lipids. In some embodiments, the human cream composition is or includes about 2.0 to 3.0 kcal/mL. In particular embodiments, the human cream composition is or includes about 25% human milk fat or lipids and 2.5 to 2.6 kcal/mL.

[0146] An exemplary feeding protocol for high risk premature infants is depicted in Table 6. In some embodiments, the feeding protocol begins at day 1 to day 3 of life. In some embodiments, the high risk infant has one or more risk factors, such as but not limited to an early gestational age, low birthweight, significant intrauterine growth restriction, e.g., less than or equal to the third percentile at birth, a low APGAR score, need for cardiopulmonary resuscitation (CPR) after delivery, maternal drug use, e.g., cocaine or methamphetamine, need for pressor support of blood pressure, presence of hemodynamically significant patent ductus arteriosus. Particular embodiments contemplate that a skilled clinician may determine if a premature infant is at high risk and/or should be administered a feeding protocol designated for high risk infants. Table 6: Exemplary Feeding Protocol for High Risk Premature Infants

[0147] An exemplary feeding protocol for small premature infants with standard risk factors for gestational age is depicted in Table 7. In some embodiments, the small premature infant has a birth weight below 750 g. In certain embodiments, the small premature infant has a birth weight below 1000 g. Table 7: Exemplary Feeding Protocol for Small Premature Infants

[0148] An exemplary feeding protocol for larger premature infants with standard risk factors for gestational age is depicted in Table 8. In certain embodiments, the larger premature infant has a birthweight of over 750 g. In some embodiments, the larger premature infant has a birthweight of over 1,000 g. In some embodiments, the larger premature infant has a birthweight of over 1,250 g. Table 8: Exemplary Feeding Protocol for Larger Premature Infants

D. Exemplary Outcomes

[0149] In some embodiments, the feeding protocols provided herein result in short term and long-term benefits as compared to existing, alternative protocols. In some embodiments, alternative protocols may include protocols including one or more of, but not limited to: (i) steps for administering non-human milk based, e.g., bovine-based formula, such as during the first 10 days, 2 weeks, or 3 weeks of the subject’s life; (ii) relying on total parenteral nutrition (TPN) for one or more days during the first 10 days, 2 weeks, or 3 weeks of the subject’s life; (iii) beginning enteral or oral feedings at a later time point (e.g., with respect to the protocols provided herein), such as later than day 1, 2, or 3 of life; (iv) beginning fortified feeds, e.g. fortified MOM, at a later time point (e.g., with respect to the protocols provided herein), such as later than day 4, 5, 6, or 7 or later of the subject’s life); (v) increase volume of feeds at a slower rate (e.g., with respect to the protocols provided herein), such as below 20 mL/kg/day; (vi) achieve full feedings at a later time point (e.g., with respect to the protocols provided herein), such as after 11, 12, 13, or 14 days of the subject’s life, or after 2 weeks or 3 weeks of the subject’s life; smaller initial enteral or oral feeding volumes (e.g., with respect to the protocols provided herein), such as below 20 mg/kg/day.

[0150] In particular embodiments, a subject, e.g., a preterm or VLBW infant, undergoing the provided feeding protocol transitions off or away from parenteral nutrition (PN) by or by about day 10 of life, day 9 of life, day 8 of life, day 7 of life, day 6 of life, day 5 of life, day 4 of life, or day 3 of life. In certain embodiments, the subject transitions off or away from PN by between day 6 and day 8 of life. In some embodiments, a subject, e.g., preterm or VLBW infant, undergoing the provided feeding protocol transition off of PN sooner than with an alternative feeding protocol. In certain embodiments, a subject, e.g., preterm or VLBW infant, undergoing the provided feeding protocol transition off of PN at least 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 1 week, 2 weeks, or 3 weeks younger or sooner than a subject undergoing an alternative protocol. Particular embodiments contemplate that earlier removal of PN improves short and long-term neurodevelopment and improves clinical outcomes as compared to existing, alternative protocols, e.g., where PN is removed later.

[0151] In some embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have a reduced probability or risk of mortality, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, 75%, 90%, 95%, 99%, or 99.9% less risk or probability for mortality, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In certain embodiments, there is a less than 5%, less than 1%, less than 0.5%, less than 0.1%, less than 0.05%, or less than 0.001% incidence or rate of mortality among the subjects that undergo the provided feeding protocol.

[0152] In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have improved or increased growth, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have an improved or increased rate of growth for head circumference (such as from birth to 34 weeks gestational age), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, subjects undergoing the protocol have at least a 10%, 20%, 30%, 40%, or 50% increase in the rate of growth for head circumference, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, the head circumference increases at a rate of about or at least 0.5 cm/week, 0.6 cm/week, 0.7 cm/week, 0.8 cm/week, 0.9 cm/week, or 1.0 cm/week when undergoing a feeding protocol provided herein.

[0153] In some embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have an improved or increased rate of gain in length (such as from birth to 34 weeks gestational age), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, subjects undergoing the protocol have at least a 10%, 20%, 30%, 40%, or 50% increase in the rate of gain in length, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, the gain in length increases at a rate of about or at least 0.7 cm/week, 0.8cm/week, 0.9 cm/week, 1.0 cm/week, 1.1 cm/week, or 1.2 cm/week when undergoing afeeding protocol provided herein.

[0154] In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have an improved or increased weight gain (such as during the period from birth to 34 weeks gestational age), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, subjects undergoing the protocol have at least a 10%, 20%, 30%, 40%, or 50% increase in the weight gain, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, weight gain is about or at least 15 g/kg/day, 20 g/kg/day, 25 g/kg/day, 30 g/kg/day, 35 g/kg/day, or 40 g/kg/day when undergoing a feeding protocol provided herein.

[0155] In some embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol reach full feeds sooner, e.g., sooner than subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol reach full feeds about or at least 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 1 week, or 2 weeks sooner, e.g., sooner than subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, the subjects reach full feeds within 14 days, within 10 days, within 9 days, within 8 days, or within 7 days of life when undergoing a feeding protocol provided herein.

[0156] In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have a reduced probability or risk for necrotizing enterocolitis (NEC), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, 75%, 90%, 95%, 99%, or 99.9% less risk or probability for NEC, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, there is a less than 1%, less than 0.5%, less than 0.1%, less than 0.05%, or less than 0.001% incidence of NEC among the subjects that undergo the provided feeding protocol.

[0157] In particular embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have a reduced probability or risk for sepsis such as late-onset sepsis (LOS), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, 75%, 90%, 95%, 99%, or 99.9% less risk or probability for sepsis, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, there is less than 0.5%, less than 0.1%, less than 0.05%, or less than 0.001% incidence of sepsis among the subjects that undergo the provided feeding protocol.

[0158] In some embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have a reduced probability or risk for Bronchopulmonary Dysplasia (BPD), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, or 50% less risk or probability for BPD, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, there is less than 50%, less than 40%, less than 25%, or less than 10%, or less than 1% incidence of BPD among the subjects that undergo the provided feeding protocol.

[0159] In some embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol that contract BPD that less severe, e.g., less severe than BPD experienced by subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In certain embodiments, the BPD is less severe with respect to one or more symptoms associated with BPD.

[0160] In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have a reduced probability or risk for Retinopathy of prematurity (ROP), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, or 90% less risk or probability for ROP, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, there is less than 10%, less than 5%, less than 1%, or less than 0.1%, incidence of ROP among the subjects that undergo the provided feeding protocol.

[0161] In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have a reduced probability or risk for metabolic disease (e.g., childhood or adult obesity), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, or 90% less risk or probability for childhood obesity, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, or 90% less risk or probability for adult obesity, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In certain embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, or 90% less risk or probability for type II diabetes (such as during childhood or adulthood), e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol.

[0162] In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have a reduced probability or risk for cerebral palsy, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, subjects who undergo the provided feeding protocol have at least a 10%, 20%, 25%, 30%, 40%, 50%, or 90% less risk or probability for cerebral palsy, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In particular embodiments, there is less than 10%, less than 5%, less than 1%, or less than 0.1%, incidence of cerebral among the subjects that undergo the provided feeding protocol.

[0163] In certain embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have improved scores on neurodevelopmental assessments, such as at or old than 1 year, 2 years, or 5 years of age, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol. In some embodiments, subjects, e.g., premature or VLBW infants, that undergo the provided feeding protocol have fewer neurodevelopmental delays, e.g., as opposed to subjects that undergo an alternative protocol and/or do not undergo the provided feeding protocol.

II. HUMAN MILK COMPOSITIONS

[0164] In certain embodiments, one or more human milk compositions are administered to the subject in need thereof, e.g., the premature or VLBW infant. In particular embodiments, the one or more human milk composition is or includes human milk that is mother’s own milk (MOM). In particular embodiments, the one or more human milk composition is or includes MOM that is fortified with a human milk-based human milk fortifier. In certain embodiments, the one or more human milk composition is or includes a human milk-based standardized human milk formulation.

A Mother’s own milk

[0165] In some aspects, human milk from women who deliver prematurely differs from that of women who deliver at term. Preterm milk is initially higher in calories, protein, fat, free amino acids, and sodium, than term milk, but over the first few weeks following delivery these levels decrease. In certain embodiments, the one or more human milk compositions administered to the subject in need thereof, e.g., the preterm or VLBW infant, is mothers’ own milk (MOM). In particular embodiments, the one or more compositions is preterm human milk, e.g., is produced by the mother of a preterm infant, e.g., the subject’s mother. Particular embodiments contemplate that the milk produced by the mother of a preterm or VLBW infant is more nutrient dense than milk produced by the woman following a term pregnancy. Thus, in some such embodiments, use of MOM or preterm human milk will insure that the subject receives more complete nutrition, e.g., higher levels of calories or human milk protein, from the provided feeding protocol as opposed to if donor milk (which is almost always term milk) is used.

[0166] In certain embodiments, the one or more human milk composition is not or does not include donor human milk. In particular embodiments, the one or more human milk composition is not or does not include donated human milk that is term human milk or milk donated from a mother who experienced a full-term pregnancy. In some embodiments, the one or more human milk composition is not or does not include donor human milk that is pasteurized. In some embodiments, the one or more human milk composition is not or does not include donor human milk, such as pasteurized donor human milk and/or term donor human milk, that is mixed with a human milk-based fortifier prior to administration or delivery to the subject, e.g., the preterm or VLBW infant.

B Human milk-based fortifier

[0167] In some aspects, human milk provides the optimal nutrition for term infants. In certain aspects, human milk is also recommended for preterm infants, but does not alone provide optimal nutrition. In some aspects, the growth and neurodevelopmental needs of premature infants are best met by appropriate fortification of human milk. In some embodiments, the one or more human milk composition administered to the subject, e.g., the preterm or VLBW infant, is or includes human milk, e.g., MOM, that is fortified with a human milk-based fortifier. In particular embodiments, the one or more human milk composition is or includes MOM that is fortified with a human milk-based human milk fortifier (also referred to herein as a human milk-based fortifier).

[0168] In some embodiments, the human milk composition is fortified MOM that is composed of MOM mixed with a human milk-based fortifier. In particular embodiments, the MOM is mixed with the human milk-based fortifier prior to enterally or orally administering, delivering, or feeding the subject, e.g., the preterm or VLBW infant.

[0169] In particular embodiments, fortified MOM and the human milk-based fortifier is consistent with an exclusive human milk diet, in that all or essentially all of the proteins, fats, and carbohydrates are human milk proteins, fats, and carbohydrates. In certain embodiments, the human milk-based fortifier contains only proteins, fats, and carbohydrates that are human milk proteins, fats, and carbohydrates. In certain embodiments, the human milk-based fortifier is free or essentially free of non-human milk proteins, fats, and carbohydrates. [0170] In some embodiments, the human milk-based fortifier is produced or manufactured from a source of human milk that was pooled from the milk of multiple donors, e.g., multiple qualified donors. Particular aspects contemplate that fortifier or standardized formulations produced or manufactured from a source of human milk from multiple qualified donors will provide a much more complete complement of biological factors including bioactive proteins, fatty acids, lipids, and HMOs than what would be achieved by a milk sample from a single donor (See, e.g., Meredith-Dennis et al, Journal of Human Lactation, 34(1): 120-129 (2018)). In certain aspects, no mother produces milk having a full complement of all possible fatty acids, immune proteins, and HMOs. By pooling milk from multiple qualified donors, a broader spectrum of all of these components is achieved. In certain embodiments, the human milk-based fortifier is manufactured or produced from a source of human milk that is pooled from the milk of at least ten donors, at least twenty donors, at least twenty-five donors, at least fifty donors, at least one hundred donors, at least one hundred and twenty-five donors, or at least one hundred and fifty or more donors. In certain embodiments, the donors are qualified, e.g., by any one or more of the qualification methods described herein.

[0171] In certain embodiments, the human milk-based fortifier contains about or at least 1.0 kcal/mL, 1.2 kcal/mL, 1.4 kcal/mL, 1.6 kcal/mL, or 1.8 kcal/mL. In some embodiments, the human milk-based fortifier contains between 1.0 kcal/mL and 2.0 kcal/mL, 1.2 kcal/mL and 1.6 kcal/mL, 1.3 kcal/mL and 1.5 kcal/mL, 1.4 kcal/mL and 1.5 kcal/mL, or 1.35 kcal/mL and 1.45 kcal/mL. In certain embodiments, the human milk based-fortifier contains about 1.40 kcal/mL and 1.50 kcal/mL. In particular embodiments, the human milk- based fortifier contains about 1.44 kcal/mL.

[0172] In particular embodiments, the human milk-based fortifier contains or contains about or at least 30 mg/mL, 40 mg/mL, 45 mg/mL, 50 mg/mL, 55 mg/mL, 60 mg/mL, 70 mg/mL, or 80 mg/mL human milk protein. In certain embodiments, the human milk-based fortifier contains between 40 mg/mL and 80 mg/mL, 50 mg/mL and 70 mg/mL, or 55 mg/mL and 65 mg/mL human milk protein. In some embodiments, the human milk-based fortifier contains between 55 mg/mL and 65 mg/mL human milk protein. In certain embodiments, the human milk-based fortifier contains about 59 mg/mL protein.

[0173] In particular embodiments, the human milk-based fortifier contains or contains about or at least 50 mg/mL human milk fat. In certain embodiments, the human milk-based fortifier contains between 60 mg/mL and 120 mg/mL, 75 mg/mL and 110 mg/mL, 90 mg/mL and 100 mg/mL, or 92 mg/mL and 97 mg/mL human milk fat. In some embodiments, the human milk-based fortifier contains between 90 mg/mL and 100 mg/mL human milk fat. In certain embodiments, the human milk-based fortifier contains about 93, 94, 95, 96, or 97 mg/mL human milk fat.

[0174] In some embodiments, the human milk-based fortifier contains about or at least 5 mg/mL, 10 mg/mL, 20 mg/mL, 25 mg/mL, 30 mg/mL, 40 mg/mL, or 50 mg/mL human milk oligosaccharides. In certain embodiments, the human milk-based fortifier contains between 5 mg/mL and 50 mg/mL, 10 mg/mL and 30 mg/mL, 20 mg/mL and 40 mg/mL, 25 mg/mL and 35 mg/mL, 25 mg/mL and 30 mg/mL, 27 mg/mL and 29 mg/mL, or 27 mg/mL and 32 mg/mL human milk oligosaccharides. In certain embodiments, the human milk-based fortifier contains between 25 mg/mL and 30 mg/mL human milk oligosaccharides. In some embodiments, the human milk-based fortifier contains between 25 mg/mL and 30 mg/mL human milk oligosaccharides. In particular embodiments, the human milk-based fortifier contains between 27 mg/mL and 29 mg/mL human milk oligosaccharides.

[0175] In some embodiments, the human milk-based fortifier contains a plurality of HMOs. In some embodiments, the human milk-based fortifier contains a plurality of, of about, or of at least 10, 25, 50, 75, 100, 125, 150 different individual HMO species, e.g., HMOs with different individual chemical formulas or chemical structures. In certain embodiments, the human milk-based fortifier contains at least 50 individual species of HMO. In certain embodiments, the human milk-based fortifier contains at least 100 individual HMO species. In certain embodiments, the human milk-based fortifier contains at least 150 individual HMO species. In some embodiments, the human milk-based fortifier includes some or all of 2'-fucosyl-lactose, 3 ’-fucosy 1-lactose, 3 ’-sialyl-lactose, 6'-sialyl-lactose, Lacto- N-tetraose, Lacto-N-neo-tetraose, Lacto-N-fucopentaose I, Lacto-N-fucopentaose II, Lacto- N-fucopentaose III, Sialyl-lacto-N-tetraose b, Sialyl-lacto-N-tetraose c, Lacto-N-difuco- hexaose I, Lacto-N-difuco-hexaose II, Lacto-N-hexaose, para-Lacto-N-hexaose, Disialyllacto-N-tetraose, Fucosyl-Lacto-N-hexaose, Difucosyl-Lacto-N-hexaose a, and Difucosyl-Lacto-N-hexaose b. In certain embodiments, the human milk-based fortifier contains at least 100, 125, or 150 individual HMO species which includes all of 2'-fucosyl- lactose, 3 ’-fucosy 1-lactose, 3 ’-sialyl-lactose, 6'-sialyl-lactose, Lacto-N-tetraose, Lacto-N-neo- tetraose, Lacto-N-fucopentaose I, Lacto-N-fucopentaose II, Lacto-N-fucopentaose III, Sialyl- lacto-N-tetraose b, Sialyl-lacto-N-tetraose c, Lacto-N-difuco-hexaose I, Lacto-N-difuco- hexaose II, Lacto-N-hexaose, para-Lacto-N-hexaose, Disialyllacto-N-tetraose, Fucosyl- Lacto-N-hexaose, Difucosyl-Lacto-N-hexaose a, and Difucosyl-Lacto-N-hexaose b.

[0176] In some aspects, one of the biggest variables in HMO diversity derives from the mother’s Lewis blood group and specifically whether or not she has an active fucosyltrasferase 2 (FUT2) and/or fucosyltrasferase 3 (FUT3) gene. When there is an active FUT2 gene, an al-2 linked fucose is produced, whereas fucose residues are al-4 linked when the FUT3 gene is active. The result of this “secretor status” is, generally, that “secretors” (i.e. those with an active FUT2 gene) produce a much more diverse profile of HMOs dominated by al-2 linked oligosaccharides, whereas “non-secretors” (i.e. those without an active FUT2 gene) may comprise a more varied array of, for example al,-4 linked oligosaccharides (as compared to secretors), but comprise an overall decrease in diversity since they are unable to synthesize a major component of the secretor’ s HMO repertoire. In particular embodiments, the human milk-based fortifier contains al-2 linked human milk oligosaccharides and al-4 linked human milk oligosaccharides.

[0177] In some embodiments, the human milk based-fortifier may be supplemented with minerals. In certain embodiments, the human milk based-fortifier may include one or more of: calcium concentration of about 4.0-5.5 mg/mL; chloride concentration of about 0.35-0.95 mg/mL; copper concentration of about 0.0005-0.0021 mg/mL; iron concentration of about 0.001-0.700 mg/mL; magnesium concentration of about 0.180-0.292 mg/mL; manganese concentration of about 0.010-0.092 micrograms/mL; phosphorus concentration of about 2.00-3.05 mg/mL; potassium concentration of about 1.90-2.18 mg/mL; sodium concentration of about 0.75-0.96 mg/mL; and zinc concentration of about 0.0200-0.0396 mg/mL. In certain embodiments, the minerals can include calcium concentration of about 2.00-2.9 mg/mL; chloride concentration of about 0.175-0.475 mg/mL; copper concentration of about 0.00025-0.001 mg/mL; iron concentration of about 0.0005-0.0025 mg/mL; magnesium concentration of about 0.090-0.170 mg/mL; manganese concentration of about 0.005-0.046 micrograms/mL; phosphorus concentration of about 1.00-1.50 mg/mL; potassium concentration of about 0.95-1.41 mg/mL; sodium concentration of about 0.375- 0.608 mg/mL; and zinc concentration of about 0.010-0.0198 mg/mL.

[0178] Human milk-based fortifiers are commercially available and include PROL ACT +4™, PROLACT+6™, PROLACT+8™, and PROLACT+10™, which are described in U.S. Patent Nos.: 8,545,920 and 9,149,052, and U.S. Patent App. Nos.: US20190075808, US0190090501, US20160050949, US20190053508, US20190075809, and US 20190327994, hereby incorporated by reference in their entirety.

[0179] In certain embodiments, the human milk-based fortifier is mixed with or added to human milk, e.g., MOM, prior to enteral or oral feeding or administration, such as to increase one or more of the nutrient, caloric, or protein content. In some embodiments, the human milk-based fortifier is mixed with human milk, e.g., MOM, in a ratio or percentage, e.g., by volume, of between 1% and 50% human milk-based fortifier and between 50% and 99% human milk, e.g., MOM. In certain embodiments, the human milk-based fortifier is mixed with MOM in a ratio of between 1% and 50% human milk-based fortifier and between 50% and 99% MOM. In some embodiments, the human milk-based fortifier is mixed with MOM in a ratio of between 30% and 50% human milk-based fortifier and between 50% and 70% MOM.

[0180] In some embodiments, the human milk-based fortifier has at least 1.0 kcal/mL and at least 30 mg/mL human milk protein. In certain embodiments, the human milk-based fortifier has between 1.0 kcal/mL and 2.0 kcal/mL and between 40 mg/mL and 80 mg/mL human milk protein. In certain embodiments, the human milk-based fortifier has between 1.0 kcal/mL and 2.0 kcal/mL, between 40 mg/mL and 80 mg/mL human milk protein, and at least 10 mg/mL HMO. In particular embodiments, the human milk-based fortifier has between 1.2 kcal/mL and 1.6 kcal/mL and between 50 mg/mL and 70 mg/mL human milk protein, and at least 10 mg/mL HMO.

[0181] In certain embodiments, the human milk-based fortifier has between 1.4 kcal/mL and 1.5 kcal/mL and between 55 mg/mL and 65 mg/mL human milk protein, between 90 mg/mL and 100 mg/mL human milk fat, and at least 20 mg/mL HMO. In some embodiments, the human milk-based fortifier has about 1.44 kcal/mL, about 59 mg/mL human milk protein, about 93 mg/mL human milk fat, and at least 20 mg/mL HMO. In certain embodiments, the human milk-based fortifier has between about 1.4 kcal/mL and 1.5 kcal/mL, between about 55 mg/mL and 65 mg/mL human milk protein, between about 90 mg/mL and 100 mg/mL human milk fat; and between about 55 mg/mL 65 mg/mL human milk sugars.

[0182] In some embodiments, the human milk composition is or includes a human cream composition. In particular embodiments, the human milk composition is or includes human milk fortified with a human cream composition. In certain embodiments, the human cream composition is or includes between 20% and 30% human milk fat. In particular embodiments, the human cream composition is or includes between 2.0 kcal/mL and 3.0 kcal/mL. In certain embodiments, the human cream composition is or includes 25% human milk fat. In some embodiments, the human cream composition is or includes 2.5 kcal/mL to 2.6 kcal/mL.

C. Standardized human milk formulations

[0183] In some embodiments, the standardized human milk formulations featured herein can be used in lieu of mother's own milk to feed the infants, e.g., premature infants. In some aspects, the standardized human milk formulations include various nutritional components for infant growth and development.

[0184] In some embodiments, the one or more human milk composition is or includes one or more human milk-based standardized human milk formulations (also referred to herein as standardized human milk formulation). In certain embodiments, the standardized human milk formulation is a ready -to-feed formulation. In particular embodiments, the standardized human milk formulation does not require addition or mixture of human milk or other human milk composition prior to feeding, delivering, or administering to the subject, e.g., the premature or VLBW infant. In certain embodiments, more than one standardized human milk formulation is administered, fed, or delivered to the subject, e.g., the premature or VLBW infant.

[0185] In particular embodiments, the one or more standardized human milk formulations are consistent with an exclusive human milk diet, in that all or essentially all of the proteins, fats, and carbohydrates are human milk proteins, fats, and carbohydrates. In some embodiments, the one or more standardized human milk formulations contain only proteins, fats, and carbohydrates that are human milk proteins, fats, and carbohydrates. In certain embodiments, the one or more standardized human milk formulations are free or essentially free of non-human milk proteins, fats, and carbohydrates.

[0186] In some aspects, naturally occurring term human milk varies widely in caloric content, nutrient content (particularly fat content), bioactive proteins, fatty acids, and human milk oligosaccharides (HMOs). Thus, in some aspects, the one or more standardized human milk formulations, owing at least in part to the standardization, deliver a more consistent amount of proteins, calories, fats, and/or HMOs across different feedings than unprocessed or non-standardized human milk. [0187] In certain embodiments, one or more standardized human milk formulation is produced or manufactured from a source of human milk that was pooled from the milk of multiple donors, e.g., multiple qualified donors. In certain embodiments, the one or more standardized human milk formulation is manufactured or produced from a source of human milk that is pooled from the milk of at least ten donors, at least twenty donors, at least twenty-five donors, at least fifty donors, at least one hundred donors, at least one hundred and twenty-five donors, or at least one hundred and fifty or more donors. In certain embodiments, the donors are qualified, e.g., by any one or more of the qualification methods described herein.

[0188] In certain embodiments, the one or more standardized human milk formulation has about or at least 20 mg/mL protein, e.g., human milk protein. In some embodiments, the one or more standardized human milk formulation has at least 24 mg/mL protein, e.g., human milk protein. In some embodiments, the standardized human milk formulation has about or at least 24 mg/mL, 25 mg/mL, 26 mg/mL, 27 mg/mL, 28 mg/mL, or 29 mg/mL protein, e.g., human milk protein. In particular embodiments, the standardized human milk formulation has between or between about 20 mg/mL and 40 mg/mL protein, e.g., human milk protein.

In certain embodiments, the standardized human milk formulation has between or between about 20 mg/mL and 30 mg/mL protein, e.g., human milk protein. In some embodiments, the standardized human milk formulation has between or between about 25 mg/mL and 38 mg/mL protein, e.g., human milk protein. In certain embodiments, the one or more standardized human milk formulation has about 26 mg/mL.

[0189] In particular embodiments, the standardized human milk formulation has at least 0.8 kcal/mL. In particular embodiments, the standardized human milk formulation has at least 0.82 kcal/mL, 0.84 kcal/mL, 0.86 kcal/mL, 0.88 kcal/mL, 0.90 kcal/mL, 0.92 kcal/mL, 0.94 kcal/mL, or 0.96 kcal/mL. In some embodiments, the standardized human milk formulation has between 0.80 kcal/mL and 1.0 kcal/mL, 0.85 kcal/mL and 0.95 kcal/mL, 0.90 kcal/mL and 0.95 kcal/mL, or 0.95 kcal/mL and 1.00 kcal/mL. In some embodiments, the standardized human milk formulation has at least 0.85 kcal/mL. In certain embodiments, the standardized human milk formulation has between 0.85 kcal/mL and 0.95 kcal/mL. In some embodiments, the standardized human milk formulation has between 0.85 kcal/mL and 0.90 kcal/mL.

[0190] In some embodiments, the one or more standardized human milk formulation has at least 24 kcal/ounce (81.2 kcal/100 mL). In particular embodiments, the fortified human milk composition has about or at least 25 kcal/ounce (84.5 kcal/100 mL, 26 kcal/ounce (87.9 kcal/100 mL), 27kcal/ounce (91.3 kcal/100 mL), or 28 kcal/ounce (94.7 kcal/100 mL). In certain embodiments, the one or more standardized human milk formulation has between 24 kcal/ounce and 28 kcal/ounce (81.2 kcal/100 mL and 94.7 kcal/100 mL, 25 kcal/ounce and 27 kcal/ounce (84.5 kcal/100 mL and 91.3 kcal/100 mL), or 27 kcal/ounce and 29 kcal/ounce (91.3 kcal/100 mL and 98.0 kcal/100 mL). In some embodiments, the one or more standardized human milk formulation has at least 25 kcal/ounce (84.5 kcal/100 mL). In certain embodiments, the one or more standardized human milk formulation has between 25 kcal/ounce and 28 kcal/ounce (84.5 kcal/100 mL and 94.7 kcal/100 mL). In particular embodiments, the one or more standardized human milk formulation has about 26 kcal/ounce (87.9 kcal/100 mL).

[0191] In certain embodiments, the one or more standardized human milk formulation has at least 0.85 kcal/mL and at least 25 mg/mL protein, e.g., human milk protein. In some embodiments, the one or more standardized human milk formulation has between 0.85 kcal/mL and 0.90 kcal/mL, and between 25 and 30 mg/mL protein, e.g., human milk protein. In particular embodiments, the one or more standardized human milk formulation has or has about 26 kcal/ounce or 0.88 kcal/mL and 26 mg/mL human milk protein.

[0192] In certain embodiments, the one or more standardized human milk formulations contain a plurality of HMOs. In some embodiments, the one or more standardized human milk formulations contains a plurality of, of about, or of at least 10, 25, 50, 75, 100, 125, 150 different individual HMO species. In certain embodiments the one or more standardized human milk formulations contains at least 50 individual species of HMO. In certain embodiments, the one or more standardized human milk formulations contain at least 100 individual HMO species. In certain embodiments, the one or more standardized human milk formulations contain at least 150 individual HMO species. In some embodiments, the one or more standardized human milk formulations include some or all of 2'-fucosyl-lactose, 3’-fucosyl-lactose, 3 ’-sialyl-lactose, 6'-sialyl-lactose, Lacto-N-tetraose, Lacto-N-neo-tetraose, Lacto-N-fucopentaose I, Lacto-N-fucopentaose II, Lacto-N- fucopentaose III, Sialyl-lacto-N-tetraose b, Sialyl-lacto-N-tetraose c, Lacto-N-difuco-hexaose I, Lacto-N-difuco-hexaose II, Lacto-N-hexaose, para-Lacto-N-hexaose, Disialyllacto-N- tetraose, Fucosyl-Lacto-N-hexaose, Difucosyl-Lacto-N-hexaose a, and Difucosyl-Lacto-N- hexaose b. In certain embodiments, the one or more standardized human milk formulations contain at least 100, 125, or 150 individual HMO species which includes all of 2'-fucosyl- lactose, 3 ’-fucosy 1-lactose, 3 ’-sialyl-lactose, 6'-sialyl-lactose, Lacto-N-tetraose, Lacto-N-neo- tetraose, Lacto-N-fucopentaose I, Lacto-N-fucopentaose II, Lacto-N-fucopentaose III, Sialyl- lacto-N-tetraose b, Sialyl-lacto-N-tetraose c, Lacto-N-difuco-hexaose I, Lacto-N-difuco- hexaose II, Lacto-N-hexaose, para-Lacto-N-hexaose, Disialyllacto-N-tetraose, Fucosyl- Lacto-N-hexaose, Difucosyl-Lacto-N-hexaose a, and Difucosyl-Lacto-N-hexaose b.

[0193] In some embodiments, the one or more standardized human milk formulations are supplemented with vitamins and/or minerals. In certain embodiments, the one or more formulations may include: calcium concentration of about 0.40-1.50 mg/mL; chloride concentration of about 0.30-0.80 mg/mL; copper concentration of about 0.0005-0.0021 mg/mL; iron concentration of about 0.001-0.005 mg/mL; magnesium concentration of about 0.03-0.13 mg/mL; manganese concentration of about 0.01-0.092 mg/mL; phosphorus concentration of about 0.15-0.631 mg/mL (e.g., about 0.15-0.60 mg/mL); potassium concentration of about 0.60-1.20 mg/mL; sodium concentration of about 0.20-0.60 mg/mL; and zinc concentration of about 0.0025-0.0120 mg/mL.

[0194] Standardized human milk formulations are commercially available and include PROLACT RTF24™, PROLACT RTF26™, and PROLACT RTF28™, which are described in U.S. Patent Nos.: 8,545,920 and 9,149,052, and U.S. Patent App. Nos.: US20190075808, US20190090501, US20160050949, US20190053508, US20190075809, and US 20190327994, hereby incorporated by reference in their entirety.

[0195] In particular embodiments, at least one of the one or more standardized human milk formulation has between about 23 mg/mL and 26 mg/mL human milk protein. In certain embodiments, the standardized human milk formulation has between about 0.75 kcal/mL and 0.87 kcal/mL. In particular embodiments, the standardized human milk formulation has between about 0.75 kcal/mL and 0.87 kcal/mL, between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat. In some embodiments, the standardized human milk formulation has about 0.82 kcal/mL, about 24 mg/mL human milk protein, and about 45 mg/mL human milk fat. In some embodiments, the standardized human milk formulation has about 24 kcal/ounce (81.2 kcal/100 mL)

[0196] In some embodiments, at least one of the one or more standardized human milk formulations has between about 25 mg/mL and 28 mg/mL human milk protein. In certain embodiments, the standardized human milk formulation has between about 0.83 kcal/mL and 0.95 kcal/mL. In some embodiments, the standardized human milk formulation has between about 0.83 kcal/mL and 0.95 kcal/mL, between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat. In certain embodiments, the standardized human milk formulation has about 0.89 kcal/mL, about 26 mg/mL human milk protein, and about 51 mg/mL fat. In some embodiments, the standardized human milk formulation has about 26 kcal/ounce (87.9 kcal/100 mL).

[0197]

III. METHODS OF MAKING HUMAN MILK COMPOSITIONS

[0198] In some embodiments, the human milk compositions described herein are produced from whole human milk. In certain embodiments, donated human milk is pooled to provide a pool of human milk. In some embodiments, a pool of human milk comprises milk from two or more (e.g., ten or more) donors. In some embodiments, the pooled human milk contains milk from at least ten, at least twenty-five, at least fifty, at least seventy -five, at least one hundred, or at least one hundred fifty individual human milk donors.

Obtaining human milk donors

[0199] In some embodiments, the human milk is provided by donors, and the donors are pre-screened and approved before any milk is processed. In some aspects, various techniques are used to identify and qualify suitable donors. In some embodiments, a potential donor must obtain a release from her physician and her child’s pediatrician as part of the approval process. This helps to insure, inter alia, that the donor is not chronically ill and that her child will not suffer as a result of the donation(s). Methods and systems for qualifying and monitoring milk collection and distribution are described, e.g., in U.S. Patents 8,545,920; 7,943,315; 9,149,052; 7,914,822 and 8,278,046, which are incorporated herein by reference in its entirety. Donors may or may not be compensated for their donation.

[0200] In certain embodiments, the donor screening includes a comprehensive lifestyle and medical history questionnaire that includes an evaluation of prescription and non-prescription medications, testing for drugs of abuse, and testing for certain pathogens. In some embodiments, a biological sample, e.g., a blood sample and/or a milk sample, may be screened for the presence of an infectious agent such as a bacteria or virus by any suitable routine technique, e.g., qPCR or ELISA. Such infections agents may include, but are not limited to, human immunodeficiency virus Type 1 (HIV-1), HIV-2, human T-lymphotropic virus Type 1 (HTLV- 1), HTLV-II, hepatitis B virus (HBV), hepatitis C virus (HCV), and syphilis. [0201] In some embodiments, donors may continuously provide samples over a treatment phase, e.g., about or at least one month, three months, six months, a year, or over a year. In some embodiments, during the treatment phase, the donor may be requalified. In some aspects, a donor who does not requalify or fails qualification is deferred until such time as they do, or permanently deferred if warranted by the results of requalification screening.

In the event of the latter situation, all remaining milk provided by that donor is removed from inventory and destroyed or used for research purposes only.

[0202] In some embodiments, a donor may donate at a designated facility (e.g., a milk bank office) or, in a preferred embodiment, express milk at home. If the donor will be expressing milk at home, she will measure the temperature in her freezer with, e.g., a supplied thermometer to confirm that it is cold enough to store human milk in order to be approved.

Testing Donor Identity

[0203] In some embodiments, once the donor has been approved, donor identity matching may be performed on donated human milk such as to ensure that the donated milk was expressed from the qualified donor and not another, e.g., when the milk was expressed by a donor away from the milk banking facility. In particular embodiments, the donor’s milk may be sampled for genetic markers, e.g., DNA markers, to guarantee that the milk is truly from the approved donor. Such subject identification techniques are known in the art (see, e.g., US Patent No.: U.S. 7,943,315, which is incorporated herein by reference in its entirety). In some embodiments, milk may be stored (e.g., at -20°C or colder) and quarantined until the test results are received.

[0204] In certain embodiments, a biological reference sample may be obtained from a potential human breast milk donor. Such sample may be obtained by methods known in the art such as, but not limited to, a cheek swab sample of cells, or a drawn blood sample, milk, saliva, hair roots, or other convenient tissue. Samples of reference donor nucleic acids (e.g., genomic DNA) may be isolated from any convenient biological sample including, but not limited to, milk, saliva, buccal cells, hair roots, blood, and any other suitable cell or tissue sample with intact interphase nuclei or metaphase cells. In certain embodiments, the sample is labeled with a unique reference number. In certain aspects, the sample can be analyzed for one or more markers that can identify the potential donor. In some embodiments, the analysis is performed at or around the time the sample is obtained from the subject, and the results may be stored, e.g., on a computer-readable medium. In some embodiments, the sample can be stored and analyzed for identifying markers at a later time.

[0205] In some aspects, it is contemplated that the biological reference sample may be DNA typed by methods known in the art such as STR analysis of STR loci, HLA analysis of HLA loci or multiple gene analysis of individual genes/alleles. The DNA-type profile of the reference sample is recorded and stored, e.g., on a computer-readable medium.

[0206] It is further contemplated that the biological reference sample may be tested for self-antigens using antibodies known in the art or other methods to determine a self antigen profile. The antigen (or another peptide) profile can be recorded and stored, e.g., on a computer-readable medium.

[0207] In particular embodiments, a test sample of human milk is taken for identification of one or more identity markers. The sample of the donated human milk is analyzed for the same marker or markers as the donor’s reference sample. The marker profiles of the reference biological sample and of the donated milk are compared. The match between the markers (and lack of any additional unmatched markers) would indicate that the donated milk comes from the same individual as the one who donated the reference sample. Lack of a match (or presence of additional unmatched markers) would indicate that the donated milk either comes from a non-tested donor or has been contaminated with fluid from a non-tested donor.

[0208] The donated human milk sample and the donated reference biological sample can be tested for more than one marker. For example, each sample can be tested for multiple DNA markers and/or peptide markers. Both samples, however, need to be tested for at least some of the same markers in order to compare the markers from each sample.

[0209] Thus, the reference sample and the donated human milk sample may be tested for the presence of differing identity marker profiles. If there are no identity marker profiles other than the identity marker profile from the expected subject, it generally indicates that there was no fluid (e.g., milk) from other humans or animals contaminating the donated human milk. If there are signals other than the expected signal for that subject, the results are indicative of contamination. Such contamination will result in the milk failing the testing.

[0210] The testing of the reference sample and of the donated human milk can be carried out at the donation facility and/or milk processing facility. The results of the reference sample tests can be stored and compared against any future donations by the same donor. Screening for Contaminants

[0211] The milk is also tested for pathogens. In some embodiments, the milk is genetically screened, e.g., by polymerase chain reaction (PCR), to identify, e.g., viruses, such as HIV-1, HBV and HCV. In some embodiments, a microorganism panel that screens for various bacterial species, fungus and mold via culture may also be used to detect contaminants. In some embodiments, a microorganism panel may test for aerobic count, Bacillus cereus, Escherichia coli, Salmonella, Pseudomonas, coliforms, Staphylococcus aureus, yeast, and mold. In some embodiments, pathogen screening may be performed both before and after pasteurization.

[0212] In addition to screening for pathogens, the donor milk may also be tested for drugs of abuse (e.g., including but not limited to cocaine, opiates, synthetic opioids (e.g. oxycodone/oxymorphone) methamphetamines, benzodiazepine, amphetamines, and THC) and/or adulterants such as non-human proteins. In certain embodiments, an ELISA may be used to test the milk for a non-human protein, such as bovine proteins, to ensure, e.g., that cow milk or cow milk infant formula has not been added to the human milk, for example to increase donation volume when donors are compensated for donations.

[0213] In certain embodiments, adulterants may include any non-human milk fluid or filler that is added to a human milk donation, thereby causing the donation to no longer be unadulterated, pure human milk. Particular adulterants to be screened for include non-human milk and infant formula. In particular embodiments, the adulterants that are screened for include cow milk, cow milk formula, goat milk, soy milk, and soy formula.

[0214] In some embodiments, methods that are known and routine by those of skill in the art may be adapted to detect non-human milk proteins, e.g., cow milk and soy proteins, in a human milk sample. In particular, immunoassays that utilize antibodies specific for a protein found in an adulterant that is not found in human milk can be used to detect the presence of the protein in a human milk sample. In some embodiments, an enzyme-linked immunosorbent assay (ELISA), such as a sandwich ELISA, may be used to detect the presence of an adulterant in a human milk sample. An ELISA may be performed manually or be automated. In some embodiments, the protein detection assay is a western blot or immunoblot. In some embodiments, flow cytometry is used to detect an adulterant in a human milk sample. ELISA, western blot, and flow cytometry protocols are well known in the art and related kits are commercially available. In some embodiments, infrared spectroscopy and in particular mid-range Fourier transform infrared spectrometry (FTIR) is used to detect adulterants in human milk.

[0215] In some embodiments, the human milk may be pooled prior to screening. In some embodiments, the human milk is pooled from more than one donation from the same individual. In certain embodiments, the human milk is pooled from two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more individual donors. In particular embodiments, the human milk is pooled from ten or more individuals. The human milk may be pooled prior to obtaining a sample by mixing human milk from two or more individuals. Alternatively, human milk samples may be pooled after they have been obtained, thereby keeping the remainder of each donation separate.

[0216] In some embodiments, the screening step will yield a positive result if the adulterant is present in the human milk sample at about 20% or more, about 15% or more, about 10% or more, about 5% or more, about 4% or more, about 3% or more, about 2% or more, about 1% or more, or about 0.5% or more of the total volume of the milk donation.

[0217] In some embodiments, the screening of the donated human milk for one or more adulterants can be carried out at the donation facility and/or milk processing facility.

[0218] In certain embodiments, human milk that has been determined to be free or negative of an adulterant, is selected and may be stored and/or further processed. Human milk that contains an adulterant will be discarded and the donor may be disqualified. In some embodiments, if an adulterant is found in two or more human milk samples from the same donor, the donor is disqualified. In certain embodiments, if an adulterant is found in one or more human milk samples from the same donor, the donor is disqualified.

Processing Human Milk

[0219] In certain embodiments, the human milk compositions described herein, e.g., human milk based-fortifiers and standardized human milk formulations, may be made according to any suitable method or technique known in the art. In some aspects, suitable methods and techniques include those described in U.S. Patent Nos.: 8,545,920 and 9,149,052, and U.S. Patent App. Nos.: US20190075808, US20190090501, US20160050949, US20190053508, US20190075809, and US 20190327994, hereby incorporated by reference in their entirety. Exemplary methods and techniques for producing the human milk compositions are briefly summarized herein.

[0220] In some embodiments, once the human milk has been screened, it is processed to produce a human milk-based fortifier or a standardized human milk formulation. In some embodiments, the donor milk is received frozen, and when desired, is thawed and pooled. In some embodiments, donor milk is then screened, e.g., to identify contaminants, by one or more of the methods discussed herein.

[0221] In some embodiments, the pooled milk is filtered, e.g., through about a 200- micron filter. In some embodiments, the pooled milk is heated, e.g., at about 63°C or greater for about 30 minutes or more. In some embodiments, the milk is transferred to a separator, e.g., a centrifuge, to separate the cream from the skim. In some embodiments, the cream may go separation once again to yield more skim. In some embodiments, a desired amount of cream is added to the skim prior to ultra-filtration. In certain embodiments, material that that did not pass through the filter is collected as the retentate fraction, and material that passes through the filter is collected as the permeate fraction. In some embodiments, post wash of the filters using permeate may be carried out to recover any nutrients, e.g., protein, remaining on the filters. The post wash may be added to the retentate to increase protein content.

[0222] In some embodiments, the caloric, protein, fat, and/or carbohydrate content of the milk fractions, e.g., cream, skim, permeate, and/or retentate, may be analyzed or verified, such as by routine methods. In certain embodiments, two or more of cream, skim, permeate, and retentate may be combined, mixed, or formulated together to achieve a human milk composition, e.g., a fortifier or standardized formulation, with target calorie, protein, fat, and/or carbohydrate contents. In some embodiments, cream, permeate, and retentate is combined, mixed, or formulated together to achieve a human milk composition, e.g., a fortifier or standardized formulation, with target calorie, protein, fat, and/or carbohydrate contents. In certain embodiments, cream, permeate, and retentate is combined, mixed, or formulated together to achieve a human milk-based fortifier, such as one described herein, e.g., in Section II-B. In some embodiments, cream, permeate, and retentate is combined, mixed, or formulated together to achieve a standardized human milk formulation, such as one described herein, e.g., in Section II-C.

[0223] In some embodiments, a sample of the human milk formulation is analyzed for mineral content. In certain embodiments, the human milk composition may be supplemented or fortified with additional minerals, such as including but not limited to one or more of calcium, chloride, copper, iron, magnesium, manganese, phosphorus, potassium, sodium, and zinc.

[0224] In certain embodiments, the human milk composition is treated to reduce bioburden. In certain embodiments, methods of reducing bioburden are known, and include but are not limited to techniques such as pasteurization, e.g., high-temperature, short-time (HTST).

[0225] In some embodiments, e.g., after treatment to reduce bioburden, the human milk composition generally processed and packaged aseptically. In certain embodiments, various samples of the human milk composition are taken for nutritional and bioburden analysis, e.g., to ensure safety and nutritional content of the human milk composition.

V. DEFINITIONS

[0226] Unless defined otherwise, all terms of art, notations and other technical and scientific terms or terminology used herein are intended to have the same meaning as is commonly understood by one of ordinary skill in the art to which the claimed subject matter pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and/or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art.

[0227] As used herein, the term “exclusive human milk diet” refers to a diet when all of the protein, fat and carbohydrates in the diet are naturally found in or derived from human milk. If the infant is receiving nutrition by PN in addition to enteral or oral administration, the “exclusive human milk diet” applies to the nutrition administered enterally or orally and not the nutrition delivered by PN. Thus, an infant’s diet may be considered to fall under an exclusive human milk diet, even if the infant if receiving one or more non-human milk proteins, fats, or carbohydrates via PN.

[0228] As used herein, the terms “subject” and “subject in need thereof’ are used interchangeably unless otherwise noted.

[0229] As used herein, the terms “preterm” and “premature” are used interchangeably and refer to infants that are bom less than 37 weeks gestational age and/or with birth weights less than 2500 gm.

[0230] As used herein, the term “very low birth weight” or “VLBW” infant refers to infants bom with a birth weight of under 1,500 g. Unless otherwise noted, as used herein “very low birth weight” or “VLBW” infants are meant to include extremely low birth weight or ELBW infants.

[0231] As used herein, the term “extremely low birth weight” or “ELBW” infants refer to infants bom with a birth weight of under 1,000 g. [0232] As used herein, the term “small for gestational age” infants refer to infants that may be full-term but are underweight, also known as small-for-date infants. Such infants may have low birth weight due to an event or condition that had slowed or halted their growth in the uterus (e.g., intra-uterine growth retardation, IUGR). Some babies are both premature and have suffered IUGR, and these babies are particularly at high risk for health problems.

[0233] As used herein, the term “human milk based human milk fortifier” (also referred to herein as “human milk-based fortifier”) refers to a human milk fortifier that is completely originated, derived, or made from human milk ingredients. Human milk-based human milk fortifiers contain proteins, fats, and carbohydrates are human milk proteins, human milk fats, and human milk carbohydrates. Human milk based human milk fortifiers are free or essentially free of proteins, fats, and carbohydrates that are not derived from or naturally occurring in human milk. Unless otherwise stated, the human milk fortifiers used within the methods, feeding protocols, and treatment regimens described herein are human milk-based human milk fortifiers. Human milk based human milk fortifiers may contain added minerals and/or vitamins to support growth and development of a premature or very low birthweight infant.

[0234] Unless otherwise indicated, a standardized human milk formulation refers to a ready to feed formula that is completely originated, derived, or made from human milk ingredients. Standardized human milk formulations contain proteins, fats, and carbohydrates are human milk proteins, human milk fats, and human milk carbohydrates and are free or essentially free of proteins, fats, and carbohydrates that are not derived from or naturally occurring in human milk. Standardized human milk formulations may contain added minerals and/or vitamins to support growth and development of a premature or very low birthweight infant.

[0235] As used herein, the term “standardized” in reference to a formulation, indicates that the formulation has very little variance from product to product or lot to lot with respect to nutritional content. In some aspects, the variation is less than 20%, 10%, 5%, or 1% from the state value of a given nutrient content.

[0236] As used herein, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. For example, "a" or "an" means "at least one" or "one or more." It is understood that aspects and variations described herein include "consisting" and/or "consisting essentially of aspects and variations. [0237] Unless otherwise indicated, the terms “protocol”, “feeding protocol”, and “treatment regimen” may be used interchangeably.

[0238] Unless otherwise indicated, the term “subject” may be used interchangeably with “subject in need thereof’ (e.g., referring to need to be administered, delivered, fed, or provided with a nutrition).

[0239] Throughout this disclosure, various aspects of the claimed subject matter are presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the claimed subject matter. Accordingly, the description of a range should be considered to have specifically disclosed all the possible sub-ranges as well as individual numerical values within that range. For example, where a range of values is provided, it is understood that each intervening value, between the upper and lower limit of that range and any other stated or intervening value in that stated range is encompassed within the claimed subject matter. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the claimed subject matter, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the described or claimed subject matter. This applies regardless of the breadth of the range.

[0240] Throughout this disclosure, ranges that are presented or expressed as “between” two endpoints, e.g., “between A and B” are understood to include the endpoints, e.g. “A” and “B”, unless otherwise indicated.

[0241] Unless otherwise noted, the terms “days of life” and “days of the subject’s life” are used interchangeably herein. For example, unless otherwise noted the term “day 1 of life” refers to the first day of a subject’s life unless otherwise noted.

[0242] The term "about" as used herein refers to the usual error range for the respective value readily known to the skilled person in this technical field. Reference to "about" a value or parameter herein includes (and describes) embodiments that are directed to that value or parameter per se. For example, description referring to "about X" includes description of "X". In some aspects, “about” a value means within a range of ±25%, ±10%, ±5%, ±1%, ±0.1%, or ±0.01% of the stated value.

[0243] Unless otherwise specified, as used herein the term “malnutrition” refers to nutrition that is not adequate to provide for optimal growth. Human milk is generally the food of choice for preterm and term infants because of its nutritional composition and immunologic benefits. However, the nutritional value of mother’s own milk or conventionally -processed donor milk is not sufficient to meet the nutritional needs of preterm or very low birth weight infants. Thus, unless otherwise specified, “malnutrition” as used herein with respect to VLBW or premature infants may refer to nutrition resulting from the intake of unfortified MOM or donor milk alone, or feedings with similar nutritional content, since the growth and development of VLBW infants fed a diet without fortification is slower and/or less optimal than what would be expected to be observed in infants fed fortified human milk and/or fed according to any of the protocols or treatment regimens provided herein.

[0244] By “human milk oligosaccharide(s)” (also referred to herein as “HMO(s)”) is meant a family of structurally diverse unconjugated glycans that are found in human breast milk. Human milk oligosaccharides are carbohydrates that contain lactose at the reducing end and, typically, fucose, sialic acid or N-acetylglucosamine at the non-reducing end (Morrow et ak, J. Nutri. 2005 135:1304-1307). These terminal sugars are the residues that most strongly influence the selective growth of bacteria and the interaction of oligosaccharides with other molecules or cells, including bacterial pathogens in the gut lumen. Furthermore, sialic acids are structural and functional components of brain gangliosides and have been implicated in neurological development of infants.

[0245] Glycans in milk are found as oligosaccharides or conjugated to milk proteins as glycoproteins, or lipid as glycolipids etc. HMO are free glycans that constitute the third most abundant component of human milk, after lactose and lipid (Morrow, 2005). The majority of HMO, however, are not metabolized by the infant and can be found in infant feces largely intact.

[0246] By “milk” is meant the fluid that is produced by the mammary gland of a mammal and expressed by the breast. Milk includes all lactation products including, but not limited to colostrum, whole milk and skim milk taken at any point post parturition or during the cycle of lactation, in the production of hind milk and foremilk. Unless otherwise specified, as used herein “milk” refers typically to whole human milk.

[0247] By “substantially” as in “substantially reduced content” is meant that the reduction in the level of minerals and/or lactose represents a statistical difference when compared to concentrated permeate that has not been subject to the current methods. By way of example, in some embodiments, the HMO compositions with substantially reduced lactose comprise lactose levels of < 5%.

[0248] By “consisting essentially” of, as used herein refers to compositions containing particular recited components while excluding other major bioactive factors.

EXAMPLES

[0250] The following examples are included for illustrative purposes only and are not intended to limit the scope of the invention.

EXAMPLE 1 : EXEMPLARY FEEDING PROTOCOL FOR PREMATURE OR VERY LOW BIRTH WEIGHT INFANTS WHEN MOTHER’S OWN MILK IS AVAILABLE

[0251] The feeding protocol begins on day 1 of the subject’s life. The subject is a premature infant or a very low birth weight infant. On day 1, the subject receives parenteral nutrition (PN) in addition to enteral feedings. In general, the PN is administered at between 2 g/kg/day and 3 g/kg/day of amino acids or protein on day 1 and then between 3 g/kg/day and 3.5 g/kg/day on day - 3.5 starting from day 2. The PN decreases as enteral feeds increase to keep the subject’s total protein or amino acid intake at 4 g/kg/day.

[0252] On days 1, 2, and 3 of life, between 20 and 30 mL/kg/day of mother’s own milk (MOM) is enterally administered to the subject. If MOM is available but not in a sufficient amount to supply the full volume for feeding, then the MOM is supplemented with donated breast milk (donor milk).

[0253] Optionally on day 4, and then on days 5-10 of life, the MOM is fortified with a human milk-based fortifier prior to administration and the feeding is advanced between 20 and 30 mL/kg/day. The human milk-based fortifier, (PROLACT +6™ human milk fortifier) contains about 1.4-1.5 kcal/mL and 58-60 mg/mL human milk protein, 90-95 mg/mL human milk fat, and 85-95 mg/mL human milk carbohydrates (60-65 mg/mL human milk sugars (e.g., lactose). An exemplary human milk-based fortifier contains 1.44 kcal/mL, 59 mg/mL human milk protein, 93 mg/mL human milk fat, and 61 mg/mL human milk carbohydrates. The human milk-based fortifier is mixed with MOM at a ratio of 3:7 (fortifier to MOM) to produce fortified human milk. An exemplary fortified human milk composition may contain approximately 0.9 kcal/ml (26 kcal/ounce) and about 29 mg/mL human milk protein, 78 mg/mL human milk carbohydrate, and 53 mg/mL human milk fat.

[0254] In general, the PN and enteral feedings are administered to maintain a total fluid intake of between about 150 mL/kg/day and 160 mL/kg/day (unless the subject’s clinician identifies a medical reason to lower amount). When the enteral feeding volumes are increased, the PN decreases by the same volume feeds increase by to maintain the goal total volume of 150-160 ml/kg/day and total protein or amino acid intake of 4 g/kg/day protein intake. [0255] For the duration of the feeding protocol, the subject receives an exclusive human milk diet in that the subject only receives human milk based enteral nutrition and all enterally administered proteins, carbohydrates, and fats are human milk proteins, carbohydrates, and fats and the diet is free or essentially free of non-human milk proteins, carbohydrates, and fats. None or essentially none of the enteral or oral nutrition provided to the subject originates from a non-human milk source or is not naturally found in human milk.

[0256] The feeding protocol is summarized in Table EL

Table El: Summary of Exemplary Feeding Protocol

[0257] PN is discontinued when the feeding volume of the enteral feeds reach between 100 mL/kg/day and 120 mL/kg/day.

[0258] The final fortification provides 4 g/kg/day protein. If growth is appropriate, the subject remains on the exclusive human milk diet (e.g., MOM+ PROLACT+6) until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier. In some cases, further increase in concentration, volume, or caloric supplement may be needed to meet individual growth goals

[0259] When full feeds, e.g., 160 mL/kg/day, are tolerated, 0.5 ml infant vitamin drops may be administered b.i.d. (1 ml/day). In addition, supplemental iron (2-4 mg Fe/kg/day) may be administered at full feeds after two weeks of age.

[0260] A human milk cream composition, e.g., PROLACT CR™ containing 25% fat and 2.5-2.6 kcal/mL, may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce) if growth is not appropriate for three days after the full feeds are reached, a human cream composition. PROLACT CR™ may also be added to fortified MOM if a loss of fat is observed, e.g., trapped in the feeding tube in addition to increasing the duration of feeds to at least 30 minutes.

[0261] If there is concern that the MOM has a low-calorie content, e.g., below 20 kcal/ounce (67.6 kcal/100 mL), and/or if poor growth is observed, e.g., weight gain of less than or slower than 15 g/kg/day, or if the infant has a high energy expenditure, e.g., if the infant has infant respiratory distress syndrome (RDS), bronchopulmonary dysplasia (BPD), or patent ductus arteriosus (PDA), then the concentration, volume, or caloric supplement of the fortified MOM may be increased according to the recommendations of the subject’s clinician or nutritionist to meet individual growth goals. Exemplary adjustments may first include the addition of the human cream composition, e.g., PROLACT CR™, to the fortified MOM and then replacing 160 ml/kg/day feedings of MOM fortified with PROLACT+6™, with 140 ml/kg/day feedings of MOM fortified with PROLACT+8™, or with 115 ml/kg/day feedings of MOM fortified with PROLACT+10™. Similar adjustments may be made for subjects that are fluid restricted.

EXAMPLE 2: EXEMPLARY FEEDING PROTOCOL FOR PREMATURE OR VERY LOW BIRTH WEIGHT INFANTS WHEN MOTHER’S OWN MILK IS AVAILABLE

[0262] The feeding protocol begins on day 1 to day 3 of the subject’s life. The subject is a premature infant or a very low birth weight infant. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. PN is administered beginning at day 1 of life as described in Example 1.

[0263] On day 1 of the protocol, between 10 and 30 mL/kg/day of mother’s own milk (MOM) is enterally administered to the subject. If MOM is available but not in a sufficient amount to supply the full volume for feeding, then donor milk is also administered to achieve to total feeding volume.

[0264] On days 2 and 3 of the protocol, the feeding volume is advanced between 10 and 30 mL/kg/day. Feedings include MOM, or donor milk if MOM is available but not in sufficient amounts. When feeding volume reaches at least 60 mL/kg/day (e.g. at days 2 or 3), MOM and donor milk may be fortified with the human milk-based fortifier PROLACT +6™ (described in Example 1) to add approximately 6 kcal/ounce (20 kcal/100 mL).

[0265] Optionally on day 4, and then on days 5-10 of the protocol, the MOM is fortified with the PROLACT +6™ human milk-based fortifier to add approximately 6 kcal/ounce (20.3 kcal/100 mL) prior to administration. Starting on day 4, feedings are advanced between 20 and 30 mL/kg/day until day 10 or until a target feeding volume, e.g., at least or about 160 mL/kg/day, is achieved.

[0266] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 1.

[0267] The feeding protocol is summarized in Table E2.

Table E2: Summary of Exemplary Feeding Protocol

[0268] PN is discontinued when the feeding volume of the enteral feeds reach between 100 mL/kg/day and 120 mL/kg/day.

[0269] In some cases, further increase in concentration, volume, or caloric supplement may be needed to meet individual growth goals

[0270] The final fortification provides about 4 g/kg/day protein. If growth is appropriate, the subject remains on the exclusive human milk diet (e.g., MOM+ PROLACT+6) until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier.

[0271] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 1

[0272] A human milk cream composition, e.g., PROLACT CR™ containing 25% fat and 2.5-2.6 kcal/mL may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce) similar to as described in Example 1. EXAMPLE 3: EXEMPLARY FEEDING PROTOCOL FOR PREMATURE OR VERY LOW BIRTH WEIGHT INFANT WHEN MOTHER’S OWN MILK IS AVAILABLE

[0273] The feeding protocol begins on day 1 to 3 of the subject’s life. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. The subject is a premature infant or a very low birth weight infant. PN is administered as described in Example 1.

[0274] On day 1 of the protocol, between 10 and 30 mL/kg/day of mother’s own milk (MOM) is enterally administered to the subject. If MOM is available but not in a sufficient amount to supply the full volume for feeding, then donor milk is also administered to achieve to total feeding volume.

[0275] On day 2 of the protocol, the clinician may decide to keep the feeding volume the same as day 1 or to advance the feeding volume by up to 30 mL/kg/day. Feedings include MOM, or pasteurized donor milk (e.g., standardized pasteurized donor milk containing 20 kcal/ounce (67.6 kcal/100 mL)) if MOM is available but not in sufficient amounts. When feeding volume reaches at least 60 mL/kg/day (e.g. at days 2, 3, or 4), MOM and donor milk may be fortified with the human milk-based fortifier PROLACT +6™ (described in Example 1) to approximately 26 kcal/Oz (87.9 kcal/100 mL).

[0276] Once the feedings reach 60 mL/kg/day, MOM is fortified with the PROLACT +6™ human milk-based fortifier to add approximately 6 kcal/ounce (20.3 kcal/100 mL) prior to administration. Starting on day 4, feedings are advanced between 20 and 30 mL/kg/day until day 10 or until a target feeding volume, e.g., at least or about 160 mL/kg/day, is achieved.

[0277] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 1.

[0278] The feeding protocol is summarized in Table E3.

Table E3: Summary of Exemplary Feeding Protocol

[0279] PN is discontinued when the feeding volume of the enteral feeds reach 100 mL/kg/day.

[0280] The final fortification provides about 4 g/kg/day protein. If growth is appropriate, the subject remains on the exclusive human milk diet (e.g., MOM+ PROLACT+6) until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier. In some cases, further increase in concentration, volume, or caloric supplement may be needed to meet individual growth goals

[0281] When full feeds, e.g., 160 mL/kg/day, are tolerated, 0.5 ml infant vitamin drops may be administered b.i.d. (1 ml/day). In addition, supplemental iron (2-3 mg Fe/kg/day) may be administered at full feeds after two weeks of age.

[0282] A human milk cream composition, e.g., PROLACT CR™ containing 25% fat and 2.5-2.6 kcal/mL, may be added feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce) similar to as described in Example 1.

EXAMPLE 4: EXEMPLARY FEEDING PROTOCOL FOR PREMATURE OR VERY LOW BIRTH WEIGHT INFANT WHEN MOTHER’S OWN MILK IS UNAVAILABLE

[0283] In some circumstances, mother’s own milk (MOM) from the infant’s mother might not be available. Reasons may include, but are not limited to, inability of the infant’s mother to produce milk, absence of the mother (e.g., such as due to incarceration), or risk of exposure by MOM to the infant to infectious diseases (e.g., viral infection such as with HIV) or chemical contamination (e.g., illegal or prescription drugs).

[0284] The feeding protocol begins on day 1 of the subject’s life. The subject is a premature infant or a very low birth weight infant. On day 1, the subject receives parenteral nutrition (PN) in addition to enteral feedings. In general, the PN is administered at between 2 g/kg/day and 3 g/kg/day or amino acids or protein on day 1 and then between 3 g/kg/day and 3.5 g/kg/day on day - 3.5 starting from day 2. The PN decreases as enteral feeds increase to keep the subject’s total protein or amino acid intake at 4 g/kg/day. [0285] On days 1, 2, and 3 of life, between 20 and 30 mL/kg/day of a first standardized, ready to feed human milk based formula containing approximately 24 kcal/ounce (81.2 kcal/100 mL) is enterally administered to the infant (PROLACT RTF24™). The first human milk-based standardized formula, PROLACT RTF24™, contains about 0.81- 0.83 kcal/mL and 23-25 mg/mL human milk protein, 44-49 mg/mL human milk fat, and 80- 85 mg/mL human milk carbohydrates (65-70 mg/mL human milk sugars e.g., lactose). An exemplary human milk based standardized formula contains 0.821 kcal/mL, 24 mg/mL human milk protein, 45 mg/mL human milk fat, and 81 mg/mL human milk carbohydrates.

[0286] On day 4 of life, between 40 and 60 mL/kg/day of a second standardized, ready to feed human milk based formula containing approximately 26 kcal/ounce (87.9 kcal/100 mL) is enterally administered to the infant (PROLACT RTF26™). On days 4-10 of life, the feeding is advanced between 20 and 30 mL/kg/day. The second human milk based standardized formula, PROLACT RTF26™, contains about 0.88-0.9 kcal/mL and 25-28 mg/mL human milk protein, 50-55 mg/mL human milk fat, and 80-85 mg/mL human milk carbohydrates (65-70 mg/mL human milk sugars e.g., lactose). An exemplary human milk based standardized formula contains 0.887 kcal/mL, 26 mg/mL human milk protein, 51 mg/mL human milk fat, and 81 mg/mL human milk carbohydrates.

[0287] In general, the PN and enteral feedings are administered to maintain a total fluid intake of between about 150 mL/kg/day and 160 mL/kg/day (unless the subject’s clinician identifies a medical reason to lower amount). When the enteral feeding volumes are increased, the PN decreases by the same volume feeds increase by to maintain the goal total volume of 150-160 ml/kg/day and total protein or amino acid intake of about 4 g/kg/day protein intake.

[0288] For the duration of the feeding protocol, the subject receives an exclusive human milk diet in that the subject only receives human milk based enteral nutrition and all enterally administered proteins, carbohydrates, and fats are human milk proteins, carbohydrates, and fats and the diet is free or essentially free of non-human milk proteins, carbohydrates, and fats. None or essentially none of the enteral or oral nutrition provided to the subject originates from a non-human milk source or is not naturally found in human milk.

[0289] The feeding protocol is summarized in Table E4.

Table E4: Summary of Exemplary Feeding Protocol

[0290] PN is discontinued when the feeding volume of the enteral feeds reach between 100 mL/kg/day and 120 mL/kg/day.

[0291] The final feeding volume provides about 4 g/kg/day protein. The subject remains on the exclusive human milk diet (e.g., PROLACT RTF26™) until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier. If growth is not appropriate for three days after the full feeds are reached, a human cream composition, e.g., PROLACT CR™, may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0292] When full feeds are tolerated, infant vitamin drops and supplemental may be administered similar to as described in Example 1.

[0293] A human milk cream composition, PROLACT CR™, may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce) if growth is not appropriate for three days after the full feeds are reached, a human cream composition. PROLACT CR™ may also be added to feedings if a loss of fat is observed, e.g., trapped in the feeding tube in addition to increasing the duration of feeds to at least 30 minutes.

[0294] If poor growth is observed, e.g., weight gain of less than or slower than 15 g/kg/day, or if the infant has a high energy expenditure, e.g., if the infant has infant respiratory distress syndrome (RDS), then the concentration, volume, or caloric supplement of the fortified MOM may be increased according to the recommendations of the subject’s clinician or nutritionist to meet individual growth goals. Exemplary adjustments may first include the addition of PROLACT CR™ to provide an additional 2 kcal/ounce to feedings, and then replacing 160 ml/kg/day PROLACT RTF26™ with 140 ml/kg/day PROLACT RTF28™. Similar adjustments may be made for subjects that are fluid restricted. EXAMPLE 5: EXEMPLARY FEEDING PROTOCOL FOR PREMATURE OR VERY LOW BIRTH WEIGHT INFANT WHEN MOTHER’S OWN MILK IS UNAVAILABLE

[0295] The feeding protocol for when MOM is not available begins on day 1 to 3 of the subject’s life. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. The subject is a premature infant or a very low birth weight infant. The subject receives parenteral nutrition (PN) beginning at day 1 of life in addition to enteral feedings similar to as described in Example 1.

[0296] On day 1 of the protocol, between 10 and 30 mL/kg/day of the first standardized, ready to feed human milk based formula described in Example 4 (PROLACT RTF24™) is enterally administered to the infant.

[0297] On days 2 and 3 of the protocol, the feeding volume is advanced between 10 and 30 mL/kg/day. When the feeding volume is at least 60 mL/kg/day, e.g., at day 2 or day 3, a second human milk-based standardized formula containing approximately 26 kcal/ounce (87.9 kcal/100 mL), PROLACT RTF26™ (described in Example 4), replaces the first standardized ready to feed human milk based formula. Starting from day 4 of the protocol, PROLACT RTF26™ is administered, and feedings are advanced between 20 and 30 mL/kg/day until day 10 of the protocol or until a target feeding volume, e.g., at least or about 160 mL/kg/day, is achieved.

[0298] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 4.

[0299] The feeding protocol is summarized in Table E5.

Table E5: Summary of Exemplary Feeding Protocol

[0300] PN is discontinued when the feeding volume of the enteral feeds reach between 100 mL/kg/day and 120 mL/kg/day.

[0301] The final feeding volume provides about 4 g/kg/day protein. The subject remains on the exclusive human milk diet (e.g., PROLACT RTF26™) until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier.

[0302] If growth is not appropriate for three days after the full feeds are reached, a human cream composition, e.g., PROLACT CR™, may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0303] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

[0304] A human milk cream composition, PROLACT CR™, may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce) similar to as described in Example 4.

EXAMPLE 6: EXEMPLARY FEEDING PROTOCOL FOR PREMATURE OR VERY LOW BIRTH WEIGHT INFANTS WHEN MOTHER’S OWN MILK IS UNAVAILABLE

[0305] The feeding protocol for when MOM is not available begins on day 1 to 3 of the subject’s life. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. The subject is a premature infant or a very low birth weight infant. The subject receives parenteral nutrition (PN) beginning at day 1 of life in addition to enteral feedings similar to as described in Example 1.

[0306] On day 1 of the protocol, between 10 and 30 mL/kg/day of the first standardized, ready to feed human milk based formula described in Example 3 (PROLACT RTF24™) is enterally administered to the infant.

[0307] On day 2 of the protocol, the clinician may decide to keep the feeding volume the same as day 1 or to advance the feeding volume by up to 30 mL/kg/day. When the feeding volume is at least 60 mL/kg/day, e.g., at days 2, 3, or 4, a second human milk-based standardized formula containing approximately 26 kcal/ounce (87.9 kcal/100 mL),

PROLACT RTF26™ (described in Example 4), replaces the first standardized ready to feed human milk based formula. Beginning from day 2 of the protocol, feedings are advanced between 20 and 30 mL/kg/day until a target feeding volume of about 160 mL/kg/day, is achieved. In some cases, the clinician may adjust the target feeding volume to an amount higher than 160 mL/kg/day.

[0308] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 4.

[0309] The feeding protocol is summarized in Table E6.

Table E6: Summary of Exemplary Feeding Protocol

*RTF26 is fed on days where the feeding volume is at least 60 mL/kg/day

[0310] PN is discontinued when the feeding volume of the enteral feeds reach 100 mL/kg/day.

[0311] The final feeding volume provides about 4 g/kg/day protein. If growth is appropriate, subject remains on the exclusive human milk diet (e.g., PROLACT RTF26™) until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier. In some cases, further increase in concentration, volume, or caloric supplement may be needed to meet individual growth goals

[0312] If growth is not appropriate for three days after the full feeds are reached, a human cream composition, e.g., PROLACT CR™, may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0313] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

[0314] A human milk cream composition, PROLACT CR™, may be added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce) similar to as described in Example 4. EXAMPLE 7 : EXEMPLARY FEEDING PROTOCOL FOR HIGH RISK PREMATURE INFANTS

[0315] The feeding protocol for critically ill, premature infants begins on day 1 to 3 of the subject’s life. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. The subject is a premature infant or a very low birth weight infant with one or more of the following: early gestational age, extremely low birthweight, significant intrauterine growth restriction of less than or equal to the third percentile at birth, a low APGAR score e.g., 5 or lower, need for cardiopulmonary resuscitation (CPR) after delivery, maternal drug use, e.g., cocaine or methamphetamine, need for pressor support of blood pressure, or presence of hemodynamically significant patent ductus arteriosus.

[0316] PN is administered beginning at day 1 of life as described in Example 1.

[0317] The subject receives parenteral nutrition (PN) in addition to enteral feedings similar to as described in Example 1.

[0318] On day 1 of the protocol, 10 mL/kg/day of MOM is enterally administered to the infant. MOM is administered on days 1-4 of the protocol, and feeding volume is increased at a rate of 10 mL/kg/day.

[0319] Beginning on day 5 of the protocol, MOM is fortified with the human milk- based fortifier PROLACT +6™ (described in Example 1) to add approximately 6 kcal/ounce (20.3 kcal/100 mL). The feeding volume is increased at a rate of 20 mL/kg/day starting at day 5 until a target feeding volume of 160 mL/kg/day is achieved at day 10 of the protocol. In some cases, the clinician may adjust the target feeding volume to an amount higher than 160 mL/kg/day. Beginning at day 7 of the protocol, a human cream composition, e.g.,

PROLACT CR™, is added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0320] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 4.

[0321] The feeding protocol is summarized in Table E7.

Table E7: Summary of Exemplary Feeding Protocol

[0322] PN is discontinued when the feeding volume of the enteral feeds reach 100 mL/kg/day.

[0323] The final feeding volume provides about 4 g/kg/day protein. If growth is appropriate, subject remains on the exclusive human milk diet until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier.

In some cases, further increase in concentration, volume, or caloric supplement may be needed to meet individual growth goals

[0324] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

EXAMPLE 8 : EXEMPLARY FEEDING PROTOCOL FOR CRITICALLY III INFANTS WHEN MOTHER’S OWN MILK IS UNAVAILABLE

[0325] The feeding protocol for critically ill, premature infants when MOM is not available begins on day 1 to 3 of the subject’s life. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. The subject is a premature infant or a very low birth weight infant with one or more of the following: early gestational age, extremely low birthweight, significant intrauterine growth restriction of less than or equal to the third percentile at birth, a low APGAR score e.g., 5 or lower, need for cardiopulmonary resuscitation (CPR) after delivery, maternal drug use, e.g., cocaine or methamphetamine, need for pressor support of blood pressure, or presence of hemodynamically significant patent ductus arteriosus The subject receives parenteral nutrition (PN) beginning at day 1 of life in addition to enteral feedings similar to as described in Example 1.

[0326] On day 1 of the protocol, 10 mL/kg/day of a standardized, ready to feed human milk based formula containing 24 kcal/ounce (81.2 kcal/100 mL) or 26 kcal/ounce (87.9 kcal/100 mL) described in Example 4 (PROLACT RTF24 or RTF26) is enterally administered to the infant. On days 1-4 of the protocol, either PROLACT RTF24 or RTF26 is enterally administered, and feeding volume is increased at a rate of 10 mL/kg/day. [0327] A feeding volume of 60 mL/kg/day is achieved on day 5 of the protocol. Beginning at day 5, PROLACT RTF26 is administered and the feeding volume is increased at a rate of 20 mL/kg/day. Feeding volumes increase until a target feeding volume of 160 mL/kg/day is achieved at day 10 of the protocol. In some cases, the clinician may adjust the target feeding volume to an amount higher than 160 mL/kg/day. Beginning at day 7 of the protocol, a human cream composition, e.g., PROLACT CR™, is added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0328] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 4.

[0329] The feeding protocol is summarized in Table E8.

Table E8: Summary of Exemplary Feeding Protocol

[0330] PN is discontinued when the feeding volume of the enteral feeds reach 100 mL/kg/day.

[0331] The final feeding volume provides about 4 g/kg/day protein. If growth is appropriate, subject remains on the exclusive human milk diet until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier.

In some cases, further increase in concentration, volume, or caloric supplement may be needed to meet individual growth goals.

[0332] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

EXAMPLE 9 : EXEMPLARY FEEDING PROTOCOL FOR SMALL PREMATURE INFANTS [0333] The feeding protocol for small premature infants begins on day 1 to 3 of the subject’s life. The subject is a premature infant with a birthweight of less than or equal to 750 g and has standard risk factors for the subject’s gestational age. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. PN is administered beginning at day 1 of life as described in Example 1.

[0334] On day 1 of the protocol, 20 mL/kg/day of MOM is enterally administered to the infant. MOM is administered on days 1 and 2 of the protocol. Throughout the feeding protocol, the feeding volume is increased at a rate of 20 mL/kg/day until a target feeding volume of 160 mL/kg/day is achieved at day 8 of the protocol. In some cases, the clinician may adjust the target feeding volume to an amount higher than 160 mL/kg/day.

[0335] Beginning on day 3 of the protocol, MOM is fortified with the human milk- based fortifier PROLACT +6™ (described in Example 1) to add approximately 6 kcal/ounce. Beginning at day 5 of the protocol, a human cream composition, e.g., PROLACT CR™, is added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0336] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 1.

[0337] The feeding protocol is summarized in Table E9.

Table E9: Summary of Exemplary Feeding Protocol

[0338] PN is discontinued when the feeding volume of the enteral feeds reach 100 mL/kg/day.

[0339] The final feeding volume provides about 4 g/kg/day protein. If growth is appropriate, subject remains on the exclusive human milk diet until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier. In some cases, further increase in concentration, volume, or caloric supplement may be needed to meet individual growth goals

[0340] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

EXAMPLE 10: EXEMPLARY FEEDING PROTOCOL FOR SMALL PREMATURE INFANTS WHEN MOTHER’S OWN MILK IS UNAVAILABLE

[0341] The feeding protocol for small premature infants when MOM is not available begins on day 1 to 3 of the subject’s life. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. The subject is a premature infant with a birthweight of less than or equal to 750 g and has standard risk factors for the subject’s gestational age. PN is administered beginning at day 1 of life as described in Example 1.

[0342] On day 1 of the protocol, 20 mL/kg/day of a standardized, ready to feed human milk based formula containing 24 kcal/ounce or 26 kcal/ounce described in Example 4 (PROLACT RTF24 or RTF26) is enterally administered to the infant. PROLACT RTF24 or RTF26 is also enterally administered on day 2. Throughout the feeding protocol, the feeding volume is increased at a rate of 20 mL/kg/day until a target feeding volume of 160 mL/kg/day is achieved at day 8 of the protocol. In some cases, the clinician may adjust the target feeding volume to an amount higher than 160 mL/kg/day.

[0343] Beginning on day 3 of the protocol, PROLACT RTF26 is administered, and beginning at day 5 of the protocol, a human cream composition, e.g., PROLACT CR™, is added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0344] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 4.

[0345] The feeding protocol is summarized in Table E10.

Table E10: Summary of Exemplary Feeding Protocol

[0346] PN is discontinued when the feeding volume of the enteral feeds reach 100 mL/kg/day.

[0347] The final feeding volume provides about 4 g/kg/day protein. If growth is appropriate, subject remains on the exclusive human milk diet until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier.

[0348] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

EXAMPLE 11 : EXEMPLARY FEEDING PROTOCOL FOR LARGER PREMATURE INFANTS

[0349] The feeding protocol for larger premature infants begins on day 1 to 3 of the subject’s life. The subject is a premature infant with a birthweight of at least 750 g, or fromlOOO g to 1500 g or greater and has standard risk factors for the subject’s gestational age. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. PN is administered beginning at day 1 of life as described in Example 1.

[0350] On day 1 of the protocol, 30 mL/kg/day of MOM is enterally administered to the infant. Throughout the feeding protocol, the feeding volume is increased at a rate of 30 mL/kg/day until a target feeding volume of 160 mL/kg/day is achieved at day 6 of the protocol. In some cases, the clinician may adjust the target feeding volume to an amount higher than 160 mL/kg/day.

[0351] Beginning on day 2 of the protocol, MOM is fortified with the human milk- based fortifier PROLACT +6™ (described in Example 1) to add approximately 6 kcal/ounce (20.3 kcal/100 mL). Beginning at day 4 of the protocol, a human cream composition, e.g., PROLACT CR™, is added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0352] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 1.

[0353] The feeding protocol is summarized in Table Ell. Table Ell: Summary of Exemplary Feeding Protocol

[0354] PN is discontinued when the feeding volume of the enteral feeds reach over 100 mL/kg/day or between 100 and 120 mL/kg/day.

[0355] The final feeding volume provides about 4 g/kg/day protein. If growth is appropriate, subject remains on the exclusive human milk diet until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier.

[0356] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

EXAMPLE 12 : EXEMPLARY FEEDING PROTOCOL FOR LARGER INFANTS WHEN MOTHER’ S OWN MILK IS UNAVAILABLE

[0357] The feeding protocol for larger premature infants when MOM is not available begins on day 1 to 3 of the subject’s life. The subject is a premature infant with a birthweight of more than or equal to 1000 g to 1500 g or greater and has standard risk factors for the subject’s gestational age. The subject may receive trophic feedings before the start of the protocol if the protocol does not begin at day 1. PN is administered beginning at day 1 of life as described in Example 1.

[0358] On day 1 of the protocol, 30 mL/kg/day of a standardized, ready to feed human milk based formula containing 24 kcal/ounce or 26 kcal/ounce described in Example 4 (PROLACT RTF24 or RTF26) is enterally administered to the infant. Throughout the feeding protocol, the feeding volume is increased at a rate of 30 mL/kg/day until a target feeding volume of 160 mL/kg/day is achieved at day 6 of the protocol. In some cases, the clinician may adjust the target feeding volume to an amount higher than 160 mL/kg/day. [0359] Beginning on day 2 of the protocol, PROLACT RTF26 is administered, and beginning at day 4 of the protocol, a human cream composition, e.g., PROLACT CR™, is added to feeds to provide an additional 7 kcal/100 mL (2 kcal/ounce).

[0360] For the duration of the feeding protocol, the subject receives an exclusive human milk diet as defined in Example 4.

[0361] The feeding protocol is summarized in Table E10.

Table E10: Summary of Exemplary Feeding Protocol

[0362] PN is discontinued when the feeding volume of the enteral feeds reach 100 mL/kg/day or between 100 and 120 mL/kg/day.

[0363] The final feeding volume provides about 4 g/kg/day protein. If growth is appropriate, subject remains on the exclusive human milk diet until 34 weeks post menstrual age (PMA). At 34 weeks PMA, the subject may be transitioned off the exclusive human milk diet over a 4-day period to promote tolerance to non-human milk based formula or fortifier.

[0364] Supplemental infant vitamin drops and supplemental iron may be administered similar to as described in Example 4.

EXAMPLE 13: PROVIDING ENTERAL NUTRITION TO VERY LOW BIRTH WEIGHT INFANTS

[0365] VLBW infants are fed an exclusive human milk diet (EHMD) according to a feeding protocol illustrated in FIG. 1 and are compared to VLBW infants that underwent current nutritional practices (as shown by, e.g., national or institutional statistics, such as 2015-2017 Japanese Neonatal Research Network (NRN) data). VLBW infants (< 1500 g) are enrolled in a single cohort study and receive an EHMD until 34 weeks gestational age. The EHMD includes mom’s own milk (MOM), donor human milk (DM), human milk (HM)- derived HM fortifiers, and HM-derived caloric fortifiers.

[0366] MOM will be utilized and supplemented with DM as needed. One HM- derived HM fortifier that will be mixed with the MOM, DM, and/or MOM supplemented with DM provides an additional 6 kcal/ounce of HM (PROLACT +6™). PROLACT +6™ contains 144 kcal/100 mL, 5.9 g/100 mL human milk protein, 9.3 g/100 mL human milk fat, and 8.9 g/100 mL of human milk carbohydrate and is added to human milk at a ratio of 7:3 human milk to PROLACT +6™.

[0367] A second HM-derived HM fortifier that will be mixed with the MOM, DM, and/or MOM supplemented with DM provides an additional 8 kcal/ounce of HM (PROLACT +8™). PROLACT +8™ contains 144 kcal/100 mL, 5.9 g/100 mL human milk protein, 9.3 g/100 mL human milk fat, and 8.9 g/100 mL of human milk carbohydrate and is added to human milk at a ratio of 3:2 human milk to PROLACT +8™

[0368] A HM caloric fortifier (PROLACT CR™) will also be used in this study. PROLACT CR™ is a HM cream composition that contains 2.5 to 2.6 kcal/mL, 25% human milk fat (weight/volume), 7% human milk carbohydrate (weight/volume), and 0.8% human milk fat (weight/volume).

[0369] Growth and safety of the VLBW infants receiving the EHMD feeding protocol are assessed. The primary outcome of the study may include weight velocity, measured in g/kg/day as determined by the exponential method of Patel et al. (Pediatrics 2005; 116(6): 1466-1473) and secondarily change in weight z-score (from standard World Health Organization growth charts) from birth to discharge. Superiority with respect to a minimum standard weight velocity of 10 g/kg/day from birth to discharge will be evaluated.

[0370] Other growth outcomes such as length and head circumference velocity and the various co-morbidities of prematurity may be compared with national or institutional statistics, e.g., the 2015-2017 Japanese Neonatal Research Network (NRN) databases, of infants that were fed according to standard feeding protocols. For example, the standard of care in Japan is to feed VLBW infants with MOM fortified with cow milk-derived fortifier or cow milk-derived formula if no MOM or DM is available to achieve full feeds.

[0371] Study infants will also be monitored for adverse events. The adverse events may include medical necrotizing enterocolitis (NEC); late-onset sepsis as identified by positive blood cultures; and feeding intolerance resulting in nil per os (NPO) status at least 24 hours (not due to a clinical procedure), change to infant formula, bloody stools (not due to anal fissures), bilious emesis, and severe abdominal distension that results in a change with clinical nutritional management of the patient. Serious adverse events may include death and surgical NEC.

[0372] The study may also assess other benefits such as better growth, demonstrated safety and tolerability, as well as improved feeding tolerance, and improved body composition. Secondary objectives to be examined may include the relationship of an EHMD to a morbidity and mortality index consisting of the following components: death, late-onset sepsis, NEC (Bell Stage II or greater), NEC requiring surgery or peritoneal drainage, bloody stools, bronchopulmonary dysplasia (BPD, defined as oxygen support at 36 weeks gestational age (GA)), severe retinopathy of prematurity (ROP, defined as requiring intervention, Stage 3, with plus disease, or in zone 1). Additional secondary endpoints may include the following: days from birth to full feeds; time from birth to regain birthweight; full feeds achieved by two weeks of age (yes/no); days on parenteral nutrition (PN, both initially and total days); number of days on antibiotics after 72 hours of life; incidence of feeding intolerance (feeding interruption for 24 hours or more not due to a clinical procedure); number of ventilator days; focal intestinal perforation (FIP, including age in days of occurrence); length gain (cm/week) from birth to discharge and change in z-score; patent ductus arteriosus (PDA); body composition (fat mass, fat-free mass, lean mass) by PEA POD at 37-40 weeks GA; weekly BUN and albumin levels; CBC, Na, K, Cl, Ca, P, alkaline phosphatase, AST, and ALT levels evaluated every two weeks; and prealbumin and IF-1 levels evaluated every four weeks.

[0373] An exploratory objective may examine effects of the EHMD on quality of life (QOL) and other aspects of the care of VLBW infants including length of stay, cost of care (COC), and neurodevelopmental outcomes at 18-22 months of age.

[0374] All study infants will be fed according to the EHMD feeding protocol shown in 1. The start of enteral feeding will occur within 24 hours of birth with MOM or DM at a volume of 10-20 mL/kg/day, divided Q 2-3 hours. Feeds will be given via gravity bolus or maximum 30-minute infusion time, unless medically contraindicated. Feeds will increase by 10-30 mL/kg/day as tolerated. When the enteral feed reaches 60 mL/kg/day, MOM or DM will be fortified with PROLACT +6™ (30 mL PROLACT +6™ mixed with 70 mL MOM or DM). Feeding volume will continue to be gradually advanced to 150 or 160 mL/kg/day (defined as full feeds) or more total feed volume as ordered by the treating team. As an example, 160 mL/kg/day PROLACT +6™ with termHM or DM provides about 4 g/kg/day protein.

[0375] Infants who require greater than 30 minutes or continuous feed infusion times will have PROLACT CR™ (4 mL PROLACT CR™ added to 100 mL PROLACT +6™ fortified milk) automatically added to feeds, once feeding volume of 100 mL/kg/day is reached.

[0376] The use of PROLACT CR™ caloric fortifier will be added to feeds if a subject is predominantly on DM once the infant reaches 100 mL/kg/day. However, if a subject is on MOM alone, the infant will receive PROLACT CR™ if weight gain is less than 15 g/kg/day for 3 days after reaching full feeding volume.

[0377] If weight gain is inadequate on PROLACT +6™ and PROLACT CR™ fortification, increase calories to +8 (40 mL PROLACT +8™ mixed with 60 mL MOM or DM) and continue PROLACT CR™. PROLACT CR™ may be increased to a maximum of 8 mL per 100 mL fortified milk if needed.

[0378] PN will be started within a few hours of birth and stopped no later than when the enteral feed reaches 120 mL/kg/day and the infant’s glucose is stable.

[0379] Infants may be transitioned off the study feeding protocol to a standard diet as they near discharge or at 34 weeks gestation, whichever comes first. The transition period will be over minimum period of 3 days.

[0380] Results of the study may indicate improved growth as measured by one or more metrics by VLBW infants that receive the EHMD feeding protocol as compared to VLBW infants that were treated with standard of care feeding protocols, e.g., protocols including human milk fortified with cow’s milk based fortifier or feedings with formula derived from cow’s milk. VLBW infants receiving the EMHD feeding protocol may experience better survival and/or fewer or less severe adverse events, e.g., sepsis, NEC, bloody stools, BPD, or ROP, than VLBW infants that were treated with standard of care feeding protocols. VLBW infants that received the EMHD feeding protocol may transition away from PN at an earlier age than VLBW infants on standard of care feeding protocols.

Claims

1. A method for providing nutrition to a premature or very low birthweight (VLBW) infant, comprising: enterally administering a feeding volume of a first or a second human milk composition to a subject that is a premature or VLBW infant on each day of a feeding protocol; wherein the feeding protocol begins within the first three days of the subject’s life; wherein the feeding volume at day 1 of the protocol is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 10 mL/kg/day and 30 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the feeding protocol beginning at day 5 until a feeding volume of at least 160 mL/kg/day is achieved; wherein the first human milk composition is administered on day 1 of the feeding protocol and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the second human milk composition is administered daily during the feeding protocol beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first human milk composition comprises (i) mother’s own milk (MOM) or a mixture of MOM and pasteurized human donor milk, or (ii) a first standardized human milk formulation comprising at least 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL); and wherein the second human milk composition comprises (i) fortified MOM comprising (a) MOM or a mixture of MOM and pasteurized human donor milk and (b) a human milk- based human milk fortifier, wherein the human milk fortifier is added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 5 kcal/ounce (16.9 kcal/100 mL) or 6 kcal/ounce (20.2 kcal/100 mL); or (ii) a second standardized human milk formulation, wherein the second standardized human milk formulation comprises at least about 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL) or 26 kcal/ounce (87.9 kcal/100 mL).

2. The method of claim 1, wherein all of the proteins, fats, and carbohydrates that are enterally administered during the feeding protocol are derived from, originate from, and/or are native to human milk.

3. The method of claim 1 or 2, wherein the feeding volume of 60 mL/kg/day is achieved no later than day 5 of the feeding protocol and/or the subject’s life.

4. The method of any of claims 1-3, wherein the feeding volume of 60 mL/kg/day is achieved on day 2 of the feeding protocol and/or the subject’s life.

5. The method of any of claims 1-4, comprising: enterally administering a feeding volume of (i) mother’s own milk (MOM) or a mixture of MOM and pasteurized human donor milk or (ii) fortified MOM to the subject on each day of the feeding protocol; wherein the feeding volume at day 1 of the protocol is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 10 mL/kg/day and 30 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the feeding protocol beginning at day 5 until a feeding volume of at least 160 mL/kg/day is achieved; wherein the MOM or the mixture is administered on day 1 of the feeding protocol and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the fortified MOM is administered daily during the feeding protocol beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the fortified MOM comprises (a) MOM or the mixture of MOM and pasteurized human donor milk and (b) the human milk fortifier, wherein the human milk fortifier has been added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 5 kcal/ounce (16.9 kcal/100 mL) or 6 kcal/ounce (20.2 kcal/100 mL).

6. The method of any of claims 1-4, comprising: enterally administering a feeding volume of the first or the second standardized human milk formulation to the subject on each day of the feeding protocol; wherein the feeding volume at day 1 of the protocol is between 10 mL/kg/day and 30 mL/kg/day, wherein the feeding volume increases by between 10 mL/kg/day and 30 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by between 20 mL/kg/day and 30 mL/kg/day on each day of the feeding protocol beginning at day 5 until the feeding volume of at least 160 mL/kg/day is achieved; wherein the first standardized human milk formulation is administered on day 1 of the feeding protocol and is administered daily until a feeding volume of 60 mL/kg/day is achieved, and wherein the second standardized human milk formulation is administered daily during the feeding protocol beginning on the day the feeding volume of 60 mL/kg/day is achieved; wherein the first standardized human milk formulation comprises at least 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL); and wherein the second standardized human milk formulation comprises at least about 20 mg/mL human milk protein and at least 24 kcal/ounce (81.2 kcal/100 mL) or 26 kcal/ounce (87.9 kcal/100 mL).

7. The method of any of claims 1-6, wherein the feeding volume at day 1 of the protocol is about 10 mL/kg/day, wherein the feeding volume increases by about 10 mL/kg/day from day 1 to day 4 of the feeding protocol, and wherein the feeding volume is increased by about 20 mL/kg/day on each day of the feeding protocol beginning at day 5 until the feeding volume of at least 160 mL/kg/day is achieved.

8. The method of claim 7, wherein the first human milk composition is administered on day 1 until day 4 of the feeding protocol, and wherein the second human milk composition is administered daily during the feeding protocol beginning on day 5 of the feeding protocol.

9. The method of claim 7 or 8, wherein the subject is a high-risk premature or VLBW infant.

10. The method of claim 9, wherein the subject has one or more of: an early gestational age, low birthweight, significant intrauterine growth restriction of less than or equal to the third percentile at birth, a low APGAR score, need for cardiopulmonary resuscitation (CPR) after delivery, maternal drug use, need for pressor support of blood pressure, of a presence of hemodynamically significant patent ductus arteriosus.

11. The method of any of claims 1-6, wherein the feeding volume at day 1 of the protocol is about 20 mL/kg/day, wherein the feeding volume increases by about 20 mL/kg/day on each day of the feeding protocol beginning from day 1 of the feeding protocol until the feeding volume of at least 160 mL/kg/day is achieved.

12. The method of claim 11, wherein the subject is a small premature infant, and/or wherein the subject has a birthweight below 1000 g or 750g.

13. The method of any of claims 1-6, wherein the feeding volume at day 1 of the protocol is about 30 mL/kg/day, wherein the feeding volume increases by about 30 mL/kg/day on each day of the feeding protocol beginning from day 1 of the feeding protocol until the feeding volume of at least 160 mL/kg/day is achieved.

14. The method of claim 13, wherein the subject is a large premature infant and/or wherein the subject has a birthweight greater than 750 g, 1000 g, or 1250 g.

15. The method of any of claims 13-14, wherein the first human milk composition is administered on day 1 of the feeding protocol and wherein the second human milk composition is administered daily during the feeding protocol beginning on day 2 of the feeding protocol.

16. The method of any of claims 1-4 or 7-15, wherein the first human milk composition is MOM and the second composition is the fortified MOM.

17. The method of any of claims 1-5 or 7-16, wherein the pasteurized donor milk comprises about 20 kcal/ounce (67.6 kcal/100 mL).

18. The method of any of claims 1-5 or 7-17, wherein the fortified MOM comprises between 20% and 40% of the human milk fortifier.

19. The method of any of claims 1-5 or 7-18, wherein the human milk fortifier comprises between 1.2 kcal/mL and 1.6 kcal/mL.

20. The method of any of claims 1-5 or 7-19, wherein the human milk fortifier comprises between 50 mg/mL and 70 mg/mL human milk protein.

21. The method of any of claims 1-5 or 7-20, wherein the human milk fortifier comprises between 1.4 and 1.5 kcal/mL, between 55 and 65 mg/mL human milk protein, and between about 90 mg/mL and 100 mg/mL human milk fat.

22. The method of any of claims 1-5 or 7-21, wherein the human milk fortifier comprises about 1.44 kcal/mL, about 59 mg/mL human milk protein, and about 93 mg/mL human milk fat.

23. The method of any of claims 1-5 or 7-22, wherein the human milk-based fortifier has been added to the MOM or the mixture in an amount sufficient to increase the calorie content of the MOM or the mixture by at least 6 kcal/ounce (20.3 kcal/100 mL).

24. The method of any of claims 1-4 or 6-15, wherein the first human milk composition is the first standardized human milk formulation and wherein the second human milk composition is the second standardized human milk formulation.

25. The method of any of claims 1-4, 6-15, or 24, wherein the first standardized human milk formulation comprises between about 23 mg/mL and 26 mg/mL human milk protein.

26. The method of any of claims 1-4, 6-15, 24, or 25, wherein the first standardized human milk formulation comprises between about 22 kcal/ounce and 26 kcal/ounce (74.4 kcal/100 mL and 87.9 kcal/100 mL).

27. The method of any of claims 1-4, 6-15, or 24-26, wherein the first standardized human milk formulation comprises between 22 kcal/ounce and 26 kcal/ounce (74.4 kcal/100 mL and 87.9 kcal/100 mL)., between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat.

28. The method of any of claims 1-4, 6-15, or 24-27, wherein the second standardized human milk formulation comprises between about 25 mg/mL and 28 mg/mL human milk protein.

29. The method of any of claims 1-4, 6-15, or 24-28, wherein the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL).

30. The method of any of claims 1-4, 6-15, or 24-29, wherein the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL), between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat.

31. The method of any of claims 1-4, 6-15, or 24-30, wherein the first standardized human milk formulation comprises between 24 kcal/ounce and 26 kcal/ounce (81.2 kcal/100 mL and 87.9 kcal/100 mL), between about 23 mg/mL and 26 mg/mL human milk protein, and between about 40 mg/mL and 50 mg/mL human milk fat; and wherein the second standardized human milk formulation comprises between about 24 kcal/ounce and 30 kcal/ounce (81.2 kcal/100 mL and 101.4 kcal/100 mL), between about 25 mg/mL and 28 mg/mL human milk protein, and between about 47 mg/mL and 56 mg/mL human milk fat.

32. The method of any of claims 1-4, 6-15, or 24-31, wherein the first standardized human milk formulation comprises about 24 kcal/ounce (81.2 kcal/100 mL), about 24 mg/mL human milk protein, and about 45 mg/mL human milk fat.

33. The method of any of claims 1-4, 6-15, or 24-31, wherein the first standardized human milk formulation comprises about 26 kcal/ounce (87.9 kcal/100 mL), about 26 mg/mL human milk protein, and about 51 mg/mL fat.

34. The method of any of claims 1-4 or 23-33, wherein the second standardized human milk formulation comprises about 26 kcal/ounce (87.9 kcal/100 mL), about 26 mg/mL human milk protein, and about 51 mg/mL fat.

35. The method of any of claims 1-34, wherein the feeding protocol continues at the feeding volume of at least 160 mL/kg/day after the feeding volume of 160 mL/kg/day is acheived, and wherein the feeding protocol continues until the subject reaches at least 34 weeks gestational age.

36. The method of any of claims 1-35, wherein the method further comprises increasing the total calorie content of the second human milk composition by between about 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) by adding an amount of a human cream composition to the second human milk composition prior to enteral administration on each day of the protocol after a feeding volume of at least 100 mL/kg/day has been achieved, wherein the human cream composition comprises about 2.5-2.6 kcal/mL and about 25% fat, optionally wherein the total calorie content is increased by about 2 kcal/ounce (6.7 kcal/100 mL).

37. The method of any of claims 1-36, wherein the method further comprises increasing the total calorie content of the second human milk composition by between about 1 kcal/ounce and 10 kcal/ounce (3.4 kcal/100 mL and 33.8 kcal/100 mL) by adding an amount of a human cream composition to the second human milk composition prior to enteral administration on each day of the protocol if the subject’s growth rate is less than 15 g/kg/day for three consecutive days after the feeding volume of 160 mL/kg/day is achieved, wherein the human cream composition comprises about 2.5 -2.6 kcal/mL and about 25% fat, optionally wherein the total calorie content is increased by about 2 kcal/ounce (6.7 kcal/100 mL).

38. The method of any of claims 1-37, wherein the subject begins parenteral nutrition (PN) on the first day of life; wherein the subject receives PN until the enteral feeding volume is between 100 mL/kg/day and 120 mL/kg/day; and on days where the subject receives PN, the total volume intake from the PN and the enteral administration is between 150 mL/kg/day and 160 mL/kg/day.

39. The method of claim 38, wherein the PN is discontinued by day 10 of life.

40. A method for providing nutrition to a premature or very low birthweight infant, comprising a feeding protocol depicted in any of Tables 2-8 or E1-E12.