Efficacy and safety of intranasal application of humen milk

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1. Synopsis Presentation Thesis Topic: Efficacy and Safety of Intranasal Application of Human milk for Neuroprotection in Preterm Infants: A Randomized Controlled Trial Dr. Aditi Agarwal

2. Name and Designation of Guide/Co-Guides Co-Guides Dr. Ishan Kumar Associate Professor Department of Radiodiagnosis & Imaging Institute of Medical Sciences Dr. Kumari Divya Singh Assistant Professor (Child Psychology) Department of Pediatrics Institute of Medical Sciences Guide: Dr. Ashok kumar Professor Department of Pediatrics Institute of Medical Sciences Banaras Hindu University

3. Introduction Prematurity represents the largest single cause of neonatal morbidity and mortality worldwide (AJNR 2020) Preterm babies, especially those below 34 weeks of gestation are at high risk of neurodevelopmental impairments (Arch Neurol 2008) Nearly 75% of preterm infants show brain MRI changes at term equivalent age (AJNR 2020) No specific intervention is available which can help protect the brain of preterm infants after birth Intranasal application of human milk is a novel therapeutic strategy to ameliorate brain damage in preterm infants

4. Direct transport theory: From nose to brain Intranasal route involving several pathways (viz., olfactory, trigeminal, and rostral migratory stream-based pathway) has emerged as potential route to bypass the BBB and transport drug directly to the brain.(1) Intranasal route involves two distinct mechanistic transportation pathway: Extracellular and Intracellular pathway of nasal drug transmission

5. Why Breast Milk? Human breast milk contains numerous neurotrophic factors such as epidermal growth factor(EGF), brain-derived neurotrophic factor, glial-derived neurotrophic factor, nerve growth factor, insulin-like growth factor-1, and hepatic growth factor. Neurotrophins are considered to potently support the development, growth and survival of neurons. Breast milk contains lactoferrin in high concentrations which has neuroprotective effects on the immature brain. Stem cells with multipotent and even pluripotent properties have been detected in human breast milk ranging between < 1 and 30% of total breast milk cells. (3,4,5)

6. Research Question: PICO Format In preterm neonates with gestational age of less than 34 weeks (P), what is the efficacy of intranasal application of human milk (I), in comparison to the usual care (C), on brain MRI at term equivalent age (O)?

7. Materials and Methodology Study design : Open label randomized controlled trial. Ethical Approval : Ethical approval will be taken from Institute Ethics Committee, Institute of Medical Sciences, Banaras Hindu University Case enrolment will start after trial registration with CTRI Informed consent from parents before inclusion in study ​Period of study : October 2022 to July 2024

8. Inclusion and Exclusion criteria: Inclusion Criteria : • Preterm neonates of less than 34 weeks of gestation Exclusion Criteria: Major congenital malformation ● Microcephaly ● Dysmorphic features/ Chromosomal anomalies ● Critically sick newborns unlikely to survive beyond first week Suspected intrauterine infection ● Failure to obtain consent

9. Sample Size Calculation : Considering 75% incidence of brain MRI abnormalities at term equivalent age in control group and 50% incidence in experimental group, and accepting a two-sided type I error rate of <0.05 and a power of 80% %, we need to enrol 57 preterm infants in each group. Assuming an attrition rate of approximately 20% the final sample size will be 70 preterm infants in each group.

10. Outcome Measures: Primary Outcome Measure: • Brain MRI findings at term equivalent age of 40 ±2 weeks. Secondary Outcome Measures: • Neurodevelopmental assessment at corrected age of 6 ±1 months and 24 ±2 months. • Hospital mortality • Late onset sepsis after randomization • Length of hospital stay

11. Secondary Outcome Measures contd … • Safety of intranasal breast milk application, as indicated by fall in oxygen saturation of > 10% from baseline increase in oxygen requirement > 10 % from baseline bradycardia (heart rate <100 per minute) apnea or choking episode change in color within 2 minutes of milk application restlessness within 2 minutes of milk application escalation of respiratory support • Mother’s response and acceptance to intranasal human milk application. • Post intervention survey of physicians and nurses. Qualitative component of study

12. Eligible newborns will be randomized within 5 days of age Allocation of newborns to different study groups will be done using serially numbered, opaque and sealed envelopes. Small syringes (1 ml) will be used to administer 0.2 ml of breast milk per nostril 4 times a day. The breast milk will be freshly expressed and used within 2 hr of expression. Intranasal application of milk will be continued till discharge from hospital or 4 weeks whichever is earlier. Control group will receive usual care. No placebo will be used Administration of Intervention

13. Measurement of Outcome Variables Primary outcome variable : Brain MRI at term equivalent age as a surrogate measure of neuroprotection. Radiologist will be masked to group allocation. Secondary outcome variables : Development assessment using DASII at corrected age of 6±1 month and 24±2 months of age. Child Psychologist will be masked to group allocation. Other secondary outcome variables including safety issues will be measured using standard definitions. A qualitative study will be done to find out the attitudes and feelings of mothers, staff nurses and physicians regarding intranasal application of human milk.

14. References Kobata R, Tsukahara H, Ohshima Y, Ohta N, Tokuriki S, Tamura S,Mayumi M (2008) High levels of growth factors in human breast milk. Early Hum Dev 84:67–69 Dvorak B, Fituch CC, Williams CS, Hurst NM, Schanler RJ (2003) increased epidermal growth factor levels in human milk of mothers with extremely premature infants. Pediatr Res 54:15–19 Read LC, Upton FM, Francis GL, Wallace JC, Dahlenberg GW, Ballard FJ (1984) Changes in the growth-promoting activity of Human milk during lactation. Pediatr Res 18:133–139 Ginet V, van de Looij Y, Petrenko V, Toulotte A, Kiss J, Hüppi PS et al (2016) Lactoferrin during lactation reduces lipopolysaccharide-induced brain injury. BioFactors Oxf Engl 42:323–336 De Looij Y, Ginet V, Chatagner A, Toulotte A, Somm E, Hüppi PS, Sizonenko SV (2014) Lactoferrin during lactation protects the immature hypoxic-ischemic rat brain. Ann Clin Transl Neurol 1:955–967 Ballard O, Morrow AL (2013) Human milk composition: nutrients And bioactive factors. Pediatr Clin N Am 60:49–74 Hassiotou F, Beltran A, Chetwynd E, Stuebe AM, Twigger A-J, Metzger P, Trengove N, Lai CT, Filgueira L, Blancafort P, Hartmann PE (2012) Breastmilk is a novel source of stem cells with Multilineage differentiation potential. Stem Cells Dayt Ohio 30:2164–2174

15. Thank you