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| ID | Type | Description | Link |
|---|---|---|---|
| PI23/00202 | Other Identifier | Instituto de Salud Carlos III (ISCIII) |
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| Name | Class |
|---|---|
| Hospital Universitario La Fe | OTHER |
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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.
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.
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| Label | Type | Description | Intervention Names |
|---|---|---|---|
| EV supplementation in very preterm infants | Infants born before 32 weeks of gestation will receive own mother's milk supplemented with extracellular vesicles isolated from donor human milk. |
|
| Name | Type | Description | Arm Group Labels | Other Names |
|---|---|---|---|---|
| Donor human milk extracellular vesicles (DHM-EVs) | Dietary Supplement | Infants born before 32 weeks of gestation will receive supplementation with extracellular vesicles isolated from donor human milk, in addition to standard nutritional care. |
| Measure | Description | Time Frame |
|---|---|---|
| Number of participants with treatment-related serious adverse events | Safety will be evaluated by the number of participants experiencing one or more treatment-related serious adverse events, defined as any of the following: (i) necrotizing enterocolitis (Bell stage ≥ II); (ii) metabolic or renal complications requiring medical intervention, (iii) cholestasis, or (iv) culture-proven sepsis. Participants experiencing multiple events will be counted once. | From enrollment until term-equivalent age (i.e., up to 40 weeks postmenstrual age) |
| Tolerance of donor human milk EV supplementation | Feeding tolerance is evaluated by the number of participants without clinical signs of gastrointestinal symptoms and successful progression of enteral feeding. | From enrollment until term-equivalent age (i.e., up to 40 weeks postmenstrual age) |
| Measure | Description | Time Frame |
|---|---|---|
| Infant weight | Measurements of weight (grams) | From enrollment until term-equivalent age (weekly) and at 3, 6, 12, 18, and 24 months of corrected age |
| Infant length | Measurements of length (cm) |
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Inclusion Criteria:
Exclusion Criteria:
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Preterm infants at risk of developing necrotizing enterocolitis (NEC), recruited between 14 and 28 days of life during their stay in the Neonatal Intensive Care Unit (NICU). A total of 20 infants will be enrolled to receive oral administration of extracellular vesicles (EVs) isolated from donor human milk (DHM) as a dietary supplement.
| Name | Role | Phone | Extension | |
|---|---|---|---|---|
| Julia Kuligowski, PhD | Contact | +34961246661 | julia.kuligowski@uv.es | |
| María Gormaz, PhD, MD | Contact | +34/961245686 | gormaz_mar@gva.es |
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| Facility | Status | City | State | ZIP | Country | Contacts |
|---|---|---|---|---|---|---|
| Hospital Universitario y Politécnico La Fe | Recruiting | Valencia | Valencia | 46026 | Spain |
| PubMed Identifier | Type | Citation | Retractions |
|---|---|---|---|
| 39818239 | Background | 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.freeradbiomed.2025.01.020. Epub 2025 Jan 14. | |
| Background | 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. | ||
| 36836757 |
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| ID | Term |
|---|---|
| D047928 | Premature Birth |
| D020345 | Enterocolitis, Necrotizing |
| ID | Term |
|---|---|
| D007752 | Obstetric Labor, Premature |
| D007744 | Obstetric Labor Complications |
| D011248 | Pregnancy Complications |
| D005261 | Female Urogenital Diseases and Pregnancy Complications |
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The study will retain biological samples collected from preterm infants, own mother's milk (OMM), and donor human milk (DHM) to support biochemical and functional analyses. Specifically:
| From enrollment until term-equivalent age (weekly) and at 3, 6, 12, 18, and 24 months of corrected age |
| Infant head circumference | Measurement of head circumference (cm) | From enrollment until term-equivalent age (weekly) and at 3, 6, 12, 18, and 24 months of corrected age |
| Analysis of redox status biomarkers | Evaluation of the ratio reduced/oxidized glutathione | 21 days of life |
| Concentration of TFN alpha (inflammatory biomarker) | Evaluation of TFN alpha in plasma employing an Enzyme-Linked Immunosorbent Assay (ELISA) kit (nM) | 21 days of life |
| Concentration of IL-6 (inflammatory biomarker) | Evaluation of IL-6 in plasma employing an ELISA kit (nM) | 21 days of life |
| Concentration of calprotectin (inflammation biomarker) | Evaluation of calprotectin in plasma employing an ELISA kit (nM) | 21 days of life |
| Ratio of meta-tyrosine/phenylalanine | Evaluation of the ratio of meta-tyrosine/phenylalanine in urine samples by Liquid Chromatography coupled to tandem Mass Spectrometry (LC-MS/MS) as an indicator of oxidative damage to proteins | 14, 21, and 28 days of life |
| Ratio of ortho-tyrosine/phenylalanine | Evaluation of the ratio of ortho-tyrosine/phenylalanine in urine samples by LC-MS/MS as an indicator of oxidative damage to proteins | 14, 21, and 28 days of life |
| Ratio of 8-hydroxy-2'-deoxyguanosine/2'-deoxyguanosine | Evaluation of the ratio of 8-hydroxy-2'-deoxyguanosine/2'-deoxyguanosine in urine samples by LC-MS/MS as an indicator of oxidative damage to DNA | 14, 21, and 28 days of life |
| Concentrations of 2,3-dinor-iPF2α-III | Evaluation of 2,3-dinor-iPF2α-III in urine samples by LC-MS/MS as an indicator of oxidative damage to lipids (n mol/g creatinine) | 14, 21, and 28 days of life |
| Concentrations of 5-iPF2α-VI | Evaluation of 5-iPF2α-VI in urine samples by LC-MS/MS as an indicator of oxidative damage to lipids (n mol/g creatinine) | 14, 21, and 28 days of life |
| Bayley Scales of Infant and Toddler Development, Fourth Edition (Bayley-IV) - Motor Composite Score | Motor development will be assessed using the Motor Composite Score of the Bayley-IV. Higher scores indicate better motor development. | 6 and 24 months corrected age |
| Bayley Scales of Infant and Toddler Development, Fourth Edition (Bayley-IV) Language Composite Score | Language development will be assessed using the Language Composite Score of the Bayley-IV. Higher scores indicate better language development. | 6 and 24 months corrected age |
| Bayley Scales of Infant and Toddler Development, Fourth Edition (Bayley-IV) - Cognitive Composite Score | Neurodevelopment will be assessed using the Cognitive Composite Score of the Bayley Scales of Infant and Toddler Development, Fourth Edition (Bayley-IV). Higher scores indicate better cognitive development. | 6 and 24 months of corrected age |
| Ages and Stages Questionnaire, Third Edition (ASQ-3) Total Score | Developmental screening will be assessed using the Ages and Stages Questionnaire, Third Edition (ASQ-3). The ASQ-3 is a parent-completed developmental screening tool consisting of five domains (communication, gross motor, fine motor, problem solving, and personal-social skills). Higher scores indicating better overall development. | 6 and 24 months of corrected age |
| Background |
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| 35954189 | Background | 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. |
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| 33715285 | Background | Chen W, Chen X, Qian Y, Wang X, Zhou Y, Yan X, Yu B, Yao S, Yu Z, Zhu J, Han S. Lipidomic Profiling of Human Milk Derived Exosomes and Their Emerging Roles in the Prevention of Necrotizing Enterocolitis. Mol Nutr Food Res. 2021 May;65(10):e2000845. doi: 10.1002/mnfr.202000845. Epub 2021 Apr 22. |
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| 17641064 | Background | Admyre C, Johansson SM, Qazi KR, Filen JJ, Lahesmaa R, Norman M, Neve EP, Scheynius A, Gabrielsson S. Exosomes with immune modulatory features are present in human breast milk. J Immunol. 2007 Aug 1;179(3):1969-78. doi: 10.4049/jimmunol.179.3.1969. |
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| 32340495 | Background | Poulimeneas D, Bathrellou E, Antonogeorgos G, Mamalaki E, Kouvari M, Kuligowski J, Gormaz M, Panagiotakos DB, Yannakoulia M; NUTRISHIELD Consortium. Feeding the preterm infant: an overview of the evidence. Int J Food Sci Nutr. 2021 Feb;72(1):4-13. doi: 10.1080/09637486.2020.1754352. Epub 2020 Apr 27. |
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| 40861903 | Result | Hondl N, Neubauer L, Ramos-Garcia V, Kuligowski J, Bishara M, Sevcsik E, Lendl B, Ramer G. Method for Mid-IR Spectroscopy of Extracellular Vesicles at the Subvesicle Level. ACS Meas Sci Au. 2025 Apr 3;5(4):469-476. doi: 10.1021/acsmeasuresciau.5c00001. eCollection 2025 Aug 20. |
| Result | Simultaneous Screening and Quantitation of Human Milk Oligosaccharides by Liquid Chromatography - Mass Spectrometry, Víctor Navarro-Esteve, Anna Zöchner, Marta Roca, Anna Parra-Llorca, Alba Moreno-Giménez, Laura Campos-Berga, María Jesús Vaya, Máximo Vento, Pilar Sáenz-González, María Gormaz, Isabel Ten-Domenech, Julia Kuligowski, Guillermo Quintás, Carbohydrate Polymer Technologies and Applications 9 (2024) 100644. doi: 10.1016/j.carpta.2024.100644 |
| Result | Normalization approaches for extracellular vesicle-derived lipidomic fingerprints - A human milk case study, Isabel Ten-Doménech, Victoria Ramos-Garcia, Abel Albiach- Delgado, Jose Luis Moreno-Casillas, Alba Moreno-Giménez, María Gormaz, Marta Gómez-Ferrer, Pilar Sepúlveda, Máximo Vento, Guillermo Quintás, Julia Kuligowski, Chemometrics and Intelligent Laboratory Systems 246 (2024) 105070. doi: https://doi.org/10.1016/j.chemolab.2024.105070 |
| 39490927 | Result | Ten-Domenech I, Moreno-Gimenez A, Campos-Berga L, Zapata de Miguel C, Lopez-Nogueroles M, Parra-Llorca A, Quintas G, Garcia-Blanco A, Gormaz M, Kuligowski J. Impact of maternal health and stress on steroid hormone profiles in human milk: Implications for infant development. J Lipid Res. 2024 Dec;65(12):100688. doi: 10.1016/j.jlr.2024.100688. Epub 2024 Oct 26. |
| 39182979 | Result | Kuligowski J, Moreno-Torres M, Quintas G. Improving insights from metabolomic functional analysis combining multivariate tools. Anal Chim Acta. 2024 Sep 22;1323:343062. doi: 10.1016/j.aca.2024.343062. Epub 2024 Aug 5. |
| 38862207 | Result | Albiach-Delgado A, Moreno-Casillas JL, Ten-Domenech I, Cascant-Vilaplana MM, Moreno-Gimenez A, Gomez-Ferrer M, Sepulveda P, Kuligowski J, Quintas G. Oxylipin profile of human milk and human milk-derived extracellular vesicles. Anal Chim Acta. 2024 Jul 18;1313:342759. doi: 10.1016/j.aca.2024.342759. Epub 2024 May 21. |
| 42368742 | Derived | Brattini M, D'Urso M, Mariotto S, Butturini E. Bovine Milk-Derived Extracellular Vesicles as Emerging Drug Delivery Platforms: Current Advances, Applications and Challenges. J Extracell Biol. 2026 Jun 27;5(7):e70160. doi: 10.1002/jex2.70160. eCollection 2026 Jul. |
| D000091642 | Urogenital Diseases |
| D004760 | Enterocolitis |
| D005759 | Gastroenteritis |
| D005767 | Gastrointestinal Diseases |
| D004066 | Digestive System Diseases |
| D007410 | Intestinal Diseases |