Administration of Extracellular Vesicles From Donor Human Milk in Preterm Infants

Official title

Administration of Extracellular Vesicles Isolated From Donor Human Milk as a Dietary Supplement for the Prevention of Necrotizing Enterocolitis in Preterm Infants

Overview

The AdVEMPrem study is exploring whether tiny particles called extracellular vesicles (EVs), which are naturally found in human milk, can help protect very premature babies from serious gut problems such as necrotizing enterocolitis (NEC). NEC is a dangerous condition that affects the intestines of preterm infants and can lead to long-term health issues. Human milk is the best nutrition for babies, but when a mother's own milk is not available, donor human milk (DHM) is used. EVs in milk carry proteins, fats, and genetic material that may support gut development, immunity, and brain growth. While laboratory studies suggest EVs are beneficial, their effects in premature babies have not yet been proven. In this study, 20 very preterm infants (\<32 weeks of gestation) will be enrolled during their stay in the Neonatal Intensive Care Unit (NICU). All babies in the study will receive oral supplementation with EVs isolated from donor human milk. Researchers will monitor feeding tolerance, growth, intestinal health, and early development. Blood and urine samples will also be collected to study how EVs affect metabolism and stress markers. The main goal is to see if EV supplementation is safe and well tolerated. Longer-term follow-up will explore whether EVs improve growth and neurodevelopment as the babies grow. This research could lead to new nutritional strategies to reduce NEC and improve outcomes for premature infants and their families.

Detailed description

Human milk is the optimal source of nutrition for infants, providing essential nutrients and bioactive components that promote growth and development. Very preterm infants (\<32 weeks gestation) are particularly vulnerable to feeding intolerance, impaired growth, and severe complications such as necrotizing enterocolitis (NEC). When a mother's own milk is insufficient or unavailable, pasteurized donor human milk (DHM) is the recommended alternative.

Extracellular vesicles (EVs) are nanosized particles naturally present in human milk that carry proteins, lipids, and nucleic acids involved in cell signaling, intestinal maturation, immune regulation, and neurodevelopment. Preclinical studies suggest that milk-derived EVs may reduce inflammation and support gut and brain development, but their role in clinical outcomes for very preterm infants has not yet been established.

The AdVEMPrem study (PI23/00202, ISCIII) is a prospective, single-arm pilot trial designed to evaluate the tolerance and safety of DHM-derived EV supplementation in very preterm infants. All enrolled infants will receive oral EV supplementation during hospitalization in the Neonatal Intensive Care Unit. Protocols for isolation and quality control of DHM-EVs will be established to ensure reproducible yields. The biochemical composition of milk and EVs will be characterized for product characterization and exploratory analyses, with emphasis on lipid profiles and functional properties. These analyses are not participant-level outcome measures. Clinical, nutritional, and developmental parameters will be monitored during the neonatal period, alongside biomarkers of redox balance and oxidative/nitrosative stress. Long-term follow-up will assess sustained effects on growth and neurodevelopmental trajectories.

Findings from this pilot study will provide foundational evidence for the potential of milk-derived EVs as a safe nutritional strategy to prevent NEC and improve outcomes in preterm infants. Results will inform the design of larger multicenter trials and may contribute to the development of standardized EV-based supplements or analogues from alternative sources, thereby addressing variability in donor milk composition. Ultimately, access to an efficient and safe nutritional supplement could reduce the incidence of NEC, improve infant and family outcomes, and deliver socio-economic and ecological benefits.

Interventions

• Dietary supplement Donor human milk extracellular vesicles (DHM-EVs)
  Infants born before 32 weeks of gestation will receive supplementation with extracellular vesicles isolated from donor human milk, in addition to standard nutritional care.

Primary outcome measures

• Number of participants with treatment-related serious adverse events [Time frame: From enrollment until term-equivalent age (i.e., up to 40 weeks postmenstrual age)]
• Tolerance of donor human milk EV supplementation [Time frame: From enrollment until term-equivalent age (i.e., up to 40 weeks postmenstrual age)]

Secondary outcome measures (12)

• Infant weight [Time frame: From enrollment until term-equivalent age (weekly) and at 3, 6, 12, 18, and 24 months of corrected age]
• Infant length [Time frame: From enrollment until term-equivalent age (weekly) and at 3, 6, 12, 18, and 24 months of corrected age]
• Infant head circumference [Time frame: From enrollment until term-equivalent age (weekly) and at 3, 6, 12, 18, and 24 months of corrected age]
• Analysis of redox status biomarkers [Time frame: 21 days of life]
• Concentration of TFN alpha (inflammatory biomarker) [Time frame: 21 days of life]
• Concentration of IL-6 (inflammatory biomarker) [Time frame: 21 days of life]
• Concentration of calprotectin (inflammation biomarker) [Time frame: 21 days of life]
• Ratio of meta-tyrosine/phenylalanine [Time frame: 14, 21, and 28 days of life]
• Ratio of ortho-tyrosine/phenylalanine [Time frame: 14, 21, and 28 days of life]
• Ratio of 8-hydroxy-2'-deoxyguanosine/2'-deoxyguanosine [Time frame: 14, 21, and 28 days of life]
• Concentrations of 2,3-dinor-iPF2α-III [Time frame: 14, 21, and 28 days of life]
• Concentrations of 5-iPF2α-VI [Time frame: 14, 21, and 28 days of life]

Eligibility criteria

Inclusion criteria

• Preterm infants born at <32 weeks gestational age
• Age between 0 and 14 days of life at enrollment
• At risk of developing necrotizing enterocolitis (NEC)
• Written informed consent obtained from parent(s) or legal guardian(s)

Exclusion criteria

• Major congenital anomalies or chromosomal abnormalities
• Severe gastrointestinal malformations (e.g., gastroschisis, intestinal atresia)
• Conditions incompatible with enteral feeding or EV supplementation
• Participation in another interventional clinical trial

Criteria are shown verbatim from the registry (in English). Final eligibility is always assessed by the study center.

Healthy volunteers: No

Study design

Observational model

Other

Study locations

Spain · 1 center

• Hospital Universitario y Politécnico La Fe — Valencia

Publications

• Albiach-Delgado A, Pinilla-Gonzalez A, Cascant-Vilaplana MM, Solaz-Garcia A, Torrejon-Rodriguez L, Lara-Canton I, Parra-Llorca A, Cernada M, Gormaz M, Pertierra A, Tapia C, Iriondo M, Aguar M, Kuligowski J, Vento M. The effect of inhaled nitric oxide treatment on biomarkers of oxidative/nitrosative damage to proteins and DNA/RNA. Free Radic Biol Med. 2025 Feb 16;228:350-359. doi: 10.1016/j.freerad PMID 39818239
• Ramos-Garcia V, Ten-Doménech I, Moreno-Giménez A, Gormaz M, Parra-Llorca A, Shephard AP, et al. ATR-FTIR spectroscopy for the routine quality control of exosome isolations. Chemometrics and Intelligent Laboratory Systems 2021:104401. https://doi.org/10.1016/j.chemolab.2021.104401.
• Buratta S, Urbanelli L, Tognoloni A, Latella R, Cerrotti G, Emiliani C, Chiaradia E. Protein and Lipid Content of Milk Extracellular Vesicles: A Comparative Overview. Life (Basel). 2023 Feb 1;13(2):401. doi: 10.3390/life13020401. PMID 36836757
• Pisano C, Galley J, Elbahrawy M, Wang Y, Farrell A, Brigstock D, Besner GE. Human Breast Milk-Derived Extracellular Vesicles in the Protection Against Experimental Necrotizing Enterocolitis. J Pediatr Surg. 2020 Jan;55(1):54-58. doi: 10.1016/j.jpedsurg.2019.09.052. Epub 2019 Oct 25. PMID 31685268
• Miyake H, Lee C, Chusilp S, Bhalla M, Li B, Pitino M, Seo S, O'Connor DL, Pierro A. Human breast milk exosomes attenuate intestinal damage. Pediatr Surg Int. 2020 Feb;36(2):155-163. doi: 10.1007/s00383-019-04599-7. Epub 2019 Nov 12. PMID 31713717
• Liao Y, Du X, Li J, Lonnerdal B. Human milk exosomes and their microRNAs survive digestion in vitro and are taken up by human intestinal cells. Mol Nutr Food Res. 2017 Nov;61(11). doi: 10.1002/mnfr.201700082. Epub 2017 Aug 15. PMID 28688106
• Galley JD, Besner GE. The Therapeutic Potential of Breast Milk-Derived Extracellular Vesicles. Nutrients. 2020 Mar 11;12(3):745. doi: 10.3390/nu12030745. PMID 32168961
• Chutipongtanate S, Morrow AL, Newburg DS. Human Milk Extracellular Vesicles: A Biological System with Clinical Implications. Cells. 2022 Jul 30;11(15):2345. doi: 10.3390/cells11152345. PMID 35954189

Identifiers

NCT: NCT07402083 · PI23/00202 · PI23/00202

Primary sources (government registries)

• ClinicalTrials.gov (US NIH)

View this study on ClinicalTrials.gov ↗