Prebiotic GOS and Lactoferrin With Iron Supplements

Study Overview
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Brief Summary
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The ultimate goal of this research is to develop a means to safely administer iron supplements to infants in settings with a high infection burden. The investigators will conduct a randomized clinical trial in 6 month-old Kenyan infants in conjunction with mechanistic microbiota studies using a novel long-term continuous polyfermenter platform inoculated with immobilized fecal microbiota from Kenyan infants. Oral iron supplements are associated with a significant 15% increase in the rate of diarrhea in children in malaria-endemic areas. The most recent studies have shown that prebiotic galacto-oligosaccharides (GOS) can provide partial amelioration of the adverse effects of iron supplementation by enhancing the growth of barrier populations of bifidobacteria and lactobacilli. The investigators hypothesize that the combination of GOS with bovine lactoferrin, adding iron sequestration as well as antimicrobial and immunomodulatory activities, will provide almost complete protection against the adverse effects of added iron on the intestinal microbiota.

Detailed Description
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Iron deficiency, the principal cause of anemia globally, affects more than two billion individuals, predominantly infants, children and women of childbearing age. Iron deficiency impairs cognitive and behavioral development in childhood, compromises immune responsiveness, decreases physical performance, and when severe, increases mortality among infants, children and pregnant women. Effective prevention and treatment of iron deficiency uses iron supplements or fortificants to increase oral iron intake. Generally, only a small fraction of the added iron is absorbed in the upper small intestine, with 80% or more passing into the colon. Because iron is an essential micronutrient for growth, proliferation, and persistence for most intestinal microbes, the increase in iron availability has profound effects on the composition and metabolism of intestinal microbiota. In particular, iron is a prime determinant of colonization and virulence for most enteric gram-negative bacteria, includingmSalmonella, Shigella and pathogenic Escherichia coli. Commensal intestinal microorganisms, principally of the genera Bifidobacterium and Lactobacillus, require little or no iron, provide a barrier effect and can inhibit pathogen growth by a variety of methods, including sequestration of iron, competition for nutrients and for intestinal epithelial sites stabilization of intestinal barrier function, and production of antibacterial peptides and organic acids that lower the pH. Increases in unabsorbed iron can promote the growth of virulent enteropathogens that overwhelm barrier strains and disrupt the gut microbiota. We hypothesize that the combination of prebiotic GOS with bovine lactoferrin (bLF), adding iron sequestration, antimicrobial and immunomodulatory activities, will provide virtually complete protection against the adverse effects of added iron on the intestinal microbiota. Our research has two specific aims: to conduct a randomized, controlled double-blind 9-month clinical trial in 6-month old Kenyan infants comparing the effects on gut microbiome composition among groups receiving in-home fortification for 6 months with micronutrient powders containing 5 mg iron (as sodium iron EDTA [2.5 mg] and ferrous fumarate [2.5 mg]) and (i) galacto-oligosaccharides (GOS; 7.5 g), (ii) bovine lactoferrin (bLF, 1.0 g), (iii) GOS (7.5 g) and bLF (1.0 g), and (iv) no GOS or bLF. Each infant will then be followed for an additional 3 months to determine the longer-term effects of the treatments. to examine mechanisms of iron, prebiotic GOS and iron-sequestering bLF on microbiota composition, enteropathogen development, microbiota functions and metabolic activity, and inflammatory potential in vitro with treatments paralleling those in Specific Aim 1, using immobilized fecal microbiota from Kenyan infants to inoculate our established long-term continuous polyfermenter intestinal model (PolyFermS) to mimic Kenyan infant colon conditions, together with cellular studies. Combining in vivo clinical and in vitro approaches will help guide formulation of safer iron supplements and fortificants and improve our understanding of the mechanisms whereby prebiotic GOS and iron-sequestering bLF support commensal microbiota to prevent iron-induced overgrowth by opportunistic enteropathogens.

Official Title
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Prebiotic GOS and Lactoferrin for Beneficial Gut Microbiota With Iron Supplements

Conditions
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Iron-deficiency

Intervention / Treatment
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* Dietary Supplement: Galacto-oligosaccharides
* Dietary Supplement: Bovine lactoferrin
* Dietary Supplement: Multiple micronutrient powders with 5 mg iron


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: vaginal or cesarean delivery an infant age of 6 months (±3 weeks) mother ≥15 years of age infant still breastfeeding anticipated residence in the area for the study duration. Exclusion Criteria: inability to provide informed consent hemoglobin < 70 g/L Z scores for weight-for-age (WAZ) or weight-for-height (WHZ) <3, any maternal or infant chronic illness administration of any infant vitamin or mineral supplements for the past 2 months history of infant antibiotic treatment within 7 days before study enrollment.

Ages Eligible for Study
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Minimum Age: 21 Weeks
Maximum Age: 27 Weeks

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Treatment
Allocation: Randomized
Intervention Model: Factorial Assignment
Interventional Model Description: The study is a single-center, double-blinded, randomized, 9-month clinical trial with a 2X2 factorial design to determine the efficacy of galacto-oligosaccharides and bovine lactoferrin in preserving a beneficial gut microbiota during iron supplementation in Kenyan infants.
Masking: Quadruple


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Active Comparator: Study group A: GOS<br>This study group will receive daily in-home fortification for 6 months with multiple micronutrient powders with 5 mg iron (as sodium iron EDTA [2.5 mg] and ferrous fumarate [2.5 mg]) and galacto-oligosaccharides (GOS), 7.5 mg. | Dietary Supplement: Galacto-oligosaccharides<br>* Galacto-oligosaccharides are classified as Generally Recognized As Safe (GRAS) by the U.S. Food and Drug Administration, are components of cow's milk and have been used repeatedly in clinical trials without adverse effects.<br>Dietary Supplement: Multiple micronutrient powders with 5 mg iron<br>* The multiple micronutrient powders are composed of Vitamin A, 400 μg; Vitamin D, 5 μg; Tocopherol Equivalents, 5 mg; Thiamine, 0.5 mg; Riboflavin, 0.5 mg; Vitamin B6, 0.5 mg; Folic Acid, 90 μg; Niacin, 6 mg; Vitamin B12, 0.9 μg; Vitamin C, 30 mg; Copper, 0.56 mg; Iodine, 90 μg; Selenium, 17 μg; Zinc, 4.1 mg; Phytase, 190 FTU; Iron, 5 mg [(as Ferrous fumarate, 2.5 mg and sodium iron ethylenediaminetetraacetate (NaFeEDTA), 2.5 mg].<br>|
| Active Comparator: Study group B: bLF<br>This study group will receive daily in-home fortification for 6 months with multiple micronutrient powders with 5 mg iron (as sodium iron EDTA [2.5 mg] and ferrous fumarate [2.5 mg]), bovine lactoferrin (bLF), 1.0 g. | Dietary Supplement: Bovine lactoferrin<br>* Bovine lactoferrin is classified as Generally Recognized As Safe (GRAS) by the U.S. Food and Drug Administration, is a component of cow's milk and has been used repeatedly in clinical trials without adverse effects.<br>Dietary Supplement: Multiple micronutrient powders with 5 mg iron<br>* The multiple micronutrient powders are composed of Vitamin A, 400 μg; Vitamin D, 5 μg; Tocopherol Equivalents, 5 mg; Thiamine, 0.5 mg; Riboflavin, 0.5 mg; Vitamin B6, 0.5 mg; Folic Acid, 90 μg; Niacin, 6 mg; Vitamin B12, 0.9 μg; Vitamin C, 30 mg; Copper, 0.56 mg; Iodine, 90 μg; Selenium, 17 μg; Zinc, 4.1 mg; Phytase, 190 FTU; Iron, 5 mg [(as Ferrous fumarate, 2.5 mg and sodium iron ethylenediaminetetraacetate (NaFeEDTA), 2.5 mg].<br>|
| Active Comparator: Study group C: GOS + bLF<br>This study group will receive daily in-home fortification for 6 months with multiple micronutrient powders with 5 mg iron (as sodium iron EDTA [2.5 mg] and ferrous fumarate [2.5 mg]), galacto-oligosaccharides (GOS), 7.5 mg, and bovine lactoferrin (bLF), 1.0 g. | Dietary Supplement: Galacto-oligosaccharides<br>* Galacto-oligosaccharides are classified as Generally Recognized As Safe (GRAS) by the U.S. Food and Drug Administration, are components of cow's milk and have been used repeatedly in clinical trials without adverse effects.<br>Dietary Supplement: Bovine lactoferrin<br>* Bovine lactoferrin is classified as Generally Recognized As Safe (GRAS) by the U.S. Food and Drug Administration, is a component of cow's milk and has been used repeatedly in clinical trials without adverse effects.<br>Dietary Supplement: Multiple micronutrient powders with 5 mg iron<br>* The multiple micronutrient powders are composed of Vitamin A, 400 μg; Vitamin D, 5 μg; Tocopherol Equivalents, 5 mg; Thiamine, 0.5 mg; Riboflavin, 0.5 mg; Vitamin B6, 0.5 mg; Folic Acid, 90 μg; Niacin, 6 mg; Vitamin B12, 0.9 μg; Vitamin C, 30 mg; Copper, 0.56 mg; Iodine, 90 μg; Selenium, 17 μg; Zinc, 4.1 mg; Phytase, 190 FTU; Iron, 5 mg [(as Ferrous fumarate, 2.5 mg and sodium iron ethylenediaminetetraacetate (NaFeEDTA), 2.5 mg].<br>|
| Placebo Comparator: Study group D<br>This study group will receive daily in-home fortification for 6 months with multiple micronutrient powders with 5 mg iron (as sodium iron EDTA [2.5 mg] and ferrous fumarate [2.5 mg]) alone, with no galacto-oligosaccharides (GOS), and no bovine lactoferrin (bLF). | Dietary Supplement: Multiple micronutrient powders with 5 mg iron<br>* The multiple micronutrient powders are composed of Vitamin A, 400 μg; Vitamin D, 5 μg; Tocopherol Equivalents, 5 mg; Thiamine, 0.5 mg; Riboflavin, 0.5 mg; Vitamin B6, 0.5 mg; Folic Acid, 90 μg; Niacin, 6 mg; Vitamin B12, 0.9 μg; Vitamin C, 30 mg; Copper, 0.56 mg; Iodine, 90 μg; Selenium, 17 μg; Zinc, 4.1 mg; Phytase, 190 FTU; Iron, 5 mg [(as Ferrous fumarate, 2.5 mg and sodium iron ethylenediaminetetraacetate (NaFeEDTA), 2.5 mg].<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Ratio of harmful to beneficial bacterial genera in fecal microbiota as determined by quantitative polymerase chain reaction (qPCR) at 1 month | The primary outcome measure will be the ratio of the abundances of potentially harmful (enteropathogenic and/or enterotoxigenic E. coli, C. difficile, members of the C. perfringens group, B. cereus, S. aureus, sum of Shigella spp., and Salmonella) to beneficial (bifidobacteria and the group of Lactobacillus/Leuconostoc/Pediococcus spp.) bacterial genera in fecal microbiota as determined by quantitative polymerase chain reaction (qPCR) at 1 month. | 1 month | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Ratio of harmful to beneficial bacterial genera in fecal microbiota as determined by quantitative polymerase chain reaction (qPCR) at 6 months | A key secondary outcome measure will be the ratio of the abundances of potentially harmful (enteropathogenic and/or enterotoxigenic E. coli, C. difficile, members of the C. perfringens group, B. cereus, S. aureus, sum of Shigella spp., and Salmonella) to beneficial (bifidobacteria and the group of Lactobacillus/Leuconostoc/Pediococcus spp.) bacterial genera in fecal microbiota as determined by quantitative polymerase chain reaction (qPCR) at 6 months. | 6 months | 
| Ratio of harmful to beneficial bacterial genera in fecal microbiota as determined by quantitative polymerase chain reaction (qPCR) at 9 months | A key secondary outcome measure will be the ratio of the abundances of potentially harmful (enteropathogenic and/or enterotoxigenic E. coli, C. difficile, members of the C. perfringens group, B. cereus, S. aureus, sum of Shigella spp., and Salmonella) to beneficial (bifidobacteria and the group of Lactobacillus/Leuconostoc/Pediococcus spp.) bacterial genera in fecal microbiota as determined by quantitative polymerase chain reaction (qPCR) at 9 months. | 9 months | 
| Microbiota composition as determined by quantitative polymerase chain reaction (qPCR). | A secondary outcome measure will be the microbiota composition among study groups as determined by quantitative polymerase chain reaction (qPCR) measures of the abundances of potentially harmful (enteropathogenic and/or enterotoxigenic E. coli, C. difficile, members of the C. perfringens group, B. cereus, S. aureus, sum of Shigella spp., and Salmonella) and of beneficial (bifidobacteria and the group of Lactobacillus/Leuconostoc/Pediococcus spp.) bacterial genera at 1, 6, and 9 months. | 1, 6 and 9 months | 
| Diarrhea | A secondary outcome measure will be the prevalence of diarrhea among study groups | 1, 6 and 9 months | 
| Malaria | A secondary outcome measure will be the prevalence of malaria among study groups | 1, 6 and 9 months | 
| Anemia | A secondary outcome measure will be the prevalence of anemia among study groups | 1, 6 and 9 months | 
| Iron deficiency | A secondary outcome measure will be the prevalence of iron deficiency among study groups | 1, 6 and 9 months | 
| Iron deficiency anemia | A secondary outcome measure will be the prevalence of iron deficiency anemia among study groups | 1, 6 and 9 months | 
| Inflammation | A secondary outcome measure will be the prevalence of inflammation among study groups | 1, 6 and 9 months | 
| Respiratory tract infections | A secondary outcome measure will be the prevalence of inflammation among study groups | 1, 6 and 9 months | 
| Other illnesses | A secondary outcome measure will be the prevalence of other illnesses among study groups | 1, 6 and 9 months | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Iron-deficiency, Iron supplementation, Galacto-oligosaccharides, Lactoferrin, Ferrous fumarate, Clinical trial, Microbiome, PolyFermS, Polyfermenter intestinal model