Safety, Acceptability, and Feasibility of Enterade® in Children at Risk for Environmental Enteric Dysfunction in Kakamega County, Kenya
Phase: NA
Conditions: Stunting, Environmental Enteric Dysfunction
Sponsor: PATH

 
SAFETY, ACCEPTABILITY, AND FEASIBILITY OF ENTERADE® IN CHILDREN AT RISK FOR 
ENVIRONMENTAL ENTERIC DYSFUNCTION IN KAKAMEGA COUNTY, KENYA 
Short title: Safety, Acceptability, and Feasibility of enterade® (SAFE) 
Study Protocol 
Version 1.7 
28 August 2018 
 
Principal Investigators   
 
Co-Investigators 
 
 
 
Michael Arndt1  
 
 
Gwen Ambler1  
James Mukabi2  
 
 
Peninah Murunga2 
 
 
 
 
 
 
 
 
 
Jaclyn Delarosa1 
 
 
 
 
 
Nick Aduro Kidaha3 
 
 
 
 
 
 
 
Alfred Ochola2  
 
 
 
 
1. PATH 
Mailing address: PO Box 900922, Seattle, WA 98109, USA 
Street address: 2201 Westlake Avenue, Suite 200, Seattle, WA 98121 
Telephone: +1-206-285-3500 
2. PATH 
Mailing address: 76634-00508, Nairobi, Kenya 
Telephone: +254-20-387-7177 
3. Kakamega County General Hospital 
Mailing address: P.O. Box 15- G.P.O.- 50100 Kakamega, Kenya 
Telephone: +254-72-278-5487 
Study Sponsor: PATH 
Funding Source: Department for International Development (DFID) 
Location of Research: Kakamega County in Western Kenya 
 
 
Proposed Project Dates: October 2018–January 2019 (4 months) 
 
 
____________________________________        
 
 28 August 2018 
 
Co-Principal Investigator’s Signature 
 
 
 
Date 
 
____________________________________        
 
 28 August 2018 
 
Co-Principal Investigator’s Signature 
 
 
 
Date 
Confidentiality Statement 
This document is confidential and is to be distributed for review only to investigators, potential investigators, 
consultants, study staff, and applicable regulatory authorities, independent ethics committees or institutional review 
boards. The contents of this document shall not be disclosed to others without written authorization from PATH (or 
others, as applicable), unless it is necessary to obtain informed consent from potential study participants.

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Table of Contents 
Title page ............................................................................................................................................. 1 
Abbreviations and Acronyms .............................................................................................................. 3 
Study Overview ................................................................................................................................... 5 
a. Research Study Personnel ............................................................................................................... 7 
b. Abstract ........................................................................................................................................... 8 
c. Study Protocol ................................................................................................................................. 9 
i. Background and significance of the research study ..................................................................... 9 
ii. Aims of proposed research study .............................................................................................. 13 
iii. Study design, population, and study procedures ..................................................................... 14 
iv. Control or comparison group use ............................................................................................. 26 
v. Number and age range of research study participants/groups whose data shall be used ....... 26 
vi. Inclusion criteria for each group of research study participants .............................................. 26 
vii. Exclusion criteria for each group .............................................................................................. 26 
viii. Gender, race, or ethnicity use as variables in participant selection ....................................... 27 
ix. Time period in which these data shall be collected and/or stored and study time frame ....... 27 
x. Monitoring conduct of the study to ensure safety, confidentiality, and data integrity ............ 27 
xi. Data management and analysis plan ........................................................................................ 33 
xii. Study limitations and the expected results .............................................................................. 35 
xiii. Itemized budget ...................................................................................................................... 35 
xiv. Research study data collection forms and other references .................................................. 36 
d. Ethical Considerations ................................................................................................................... 37 
i. Risks to research study participants/groups ............................................................................... 37 
ii. Access to protected health information required by proposed research study ....................... 40 
iii. Sources of data, specimens, and records to be used in the proposed research study ............ 41 
iv. Maintaining confidentiality of research study data and protecting the participant privacy .... 41 
v. Storage of data and specimens being collected ........................................................................ 42 
vi. Duration of data storage, ensuring data security, and custodian and staff with data access .. 42 
vii. Plan for controlling access to stored research study data ....................................................... 42 
viii. Viewing of identifiable research study data by non-applicant or study team member ......... 42 
ix. Signing of agreement(s) by study team members with access to confidential data collected 42 
x. Potential benefits of research study to community and/or society .......................................... 42 
xi. Consent issues: how consent will be obtained ......................................................................... 43 
e. References ..................................................................................................................................... 44 
f. Appendix 8 – Toxicity Table............................................................................................................ 46 

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ABBREVIATIONS AND ACRONYMS  
AA-ORS: amino acid–based oral rehydration solution 
AE: adverse event 
CGH: Kakamega County General Hospital 
CRF: case report form 
CWC: child welfare clinic 
DFID: Department for International Development 
DO: direct observation 
DSMB: Data and Safety Monitoring Board 
EC: ethics committee 
EED: environmental enteric dysfunction 
ELISA: enzyme-linked immunosorbent assays 
EPI: Expanded Program on Immunization 
GCP: good clinical practices 
GRAS: generally regarded as safe 
HCP: health care provider 
ICH: International Council on Harmonisation 
IDI: in-depth interview 
IRB: institutional review board 
ISM: independent study monitor 
LAZ: Length-for-age z-score 
LRS: low-resource settings 
MAL-ED: Etiology, Risk Factors and Interactions of Enteric Infections and Malnutrition and the 
Consequences for Child Health and Development 
MOH: Ministry of Health 
MUERC: Maseno University Ethics Review Committee 
MUAC: mid-upper arm circumference 
ORS: oral rehydration solution 
PI: principal investigator 

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PID: Patient Identification Number  
PPB: Pharmacy and Poisons Board 
RA: research assistant 
REC: research ethics committee 
SAE: serious adverse event 
SOP: standard operating procedure 
SUSAR: suspected unexpected serious adverse reaction  
Vit. A: vitamin A 
WHO: World Health Organization 
 

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STUDY OVERVIEW 
Title: 
Safety, Acceptability, and Feasibility of enterade® in children at risk 
for environmental enteric dysfunction in Kakamega County, Kenya 
Sponsor: 
Drug Development, PATH 
Investigators: 
USA: Michael Arndt (co-PI), Gwen Ambler, Jaclyn Delarosa 
 
Kenya: Dr. James Mukabi (co-PI), Peninah Murunga, Alfred Ochola 
Ethics committees: 
Maseno University Ethics and Research Committee (MUERC) 
 
PATH Research and Ethics Committee (REC) 
Regulatory authority: 
Kenyan Ministry of Health (MOH) Pharmacy and Poisons Board (PPB) 
Phase: 
I 
Study goals: 
Determine the safety, acceptability, and pediatric dosing of 
enterade® solution, an amino acid–based oral rehydration solution 
(AA-ORS), for potential use in the management of environmental 
enteric dysfunction (EED) among children aged 12–24 months in 
Kenya.  
Study objectives: 
1. Primary: To determine the safety of a 2-week course of AA-ORS 
among children with length-for-age Z-scores (LAZ) between –1 
and –3. 
 
2.  Primary: To determine the feasibility and best tolerated dose of 
AA-ORS among children with LAZ between –3 and –1. 
 
3. Secondary: To determine the perceptions among caregivers on 
the acceptability of AA-ORS as a potential intervention for EED. 
 
4.  Exploratory: To determine the impact of AA-ORS on markers of 
metabolism, gut dysfunction, systemic inflammation, and 
micronutrient status among children with LAZ between –3 and 
– 1. 
Study endpoints: 
1. Primary: Adverse events and serious adverse events through 21 
days of follow-up, and changes to dietary and digestive habits 
through day 14 of follow-up. 
 
2.  Primary: Average daily product volume consumed through 14 
days of follow-up, time trends in average daily volume 
consumed, and factors that influence children’s willingness and 
ability to ingest the product as intended.  
 
3. Secondary: Key factors that influence caregivers’ willingness to 
have the product used by their children, and their perception of 
their children’s response to its consumption. 
 
4.  Exploratory: Average plasma/serum concentrations of metabolic, 
gut dysfunction, systemic inflammation, and micronutrient 
biomarkers at baseline and day 15 of follow-up. 
Study design: 
 
This is a mixed-methods pilot study to assess the safety, 
acceptability, and feasible dosage of AA-ORS among children with 
mild to moderate stunting (LAZ between –1 and –3) who are at risk 
for EED and to assess the acceptability of this fluid supplementation 
for their caregivers. During the main study, child participants will be 

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randomized to receive 14 days of AA-ORS or placebo solution daily, 
dosing of up to 237 ml over a 3-hour period. Study staff and 
participants will be blinded to the study product allocation. 
Caregivers will store and return unused study product. Data 
collection methods to include direct observation of product 
consumption, adverse event assessments, structured surveys with 
caregivers, average daily volume consumed, and measurement of 
plasma/serum markers of metabolism, gut dysfunction, systemic 
inflammation, and micronutrient status pre- and post-
administration of product.  
Study population: 
 
This study will recruit subjects from a health facility in Western 
Kenya. The study population is children 12–24 months of age with 
mild to moderate stunting (–3 < LAZ < –1), who are presenting to 
health facilities for routine immunizations, healthy child visits, or 
vitamin A supplementation. Caregivers of enrolled children will also 
be consented for interviews and direct observation. 
Sample size: 
N = 132 total individuals 
AA-ORS: 33 children; 1 dose of up to 237 ml × 14 days 
Placebo solution: 33 children; 1 dose of up to 237 ml × 14 days 
 
Study Arm 
 
AA-ORS  
Placebo solution 
Children 
33; receive 1 dose of up 
to 237 ml x 14 days 
33; receive 1 dose of 
up to 237 ml x 14 days 
Caregivers 
33; interviews and 
direct observation 
33; interviews and 
direct observation 
 
Study duration: 
4 months 

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A. RESEARCH STUDY PERSONNEL  
Statement of compliance: The trial will be conducted in accordance with the International Council on 
Harmonisation (ICH) Good Clinical Practices (GCP) E6(R2), the Declaration of Helsinki, and applicable 
local regulatory requirements. The principal investigator (PI) and site PI will assure that no deviation 
from or changes to the protocol will take place without prior agreement from the sponsor and 
documented approval from the relevant ethics committees (ECs) and regulatory body, except where 
necessary to eliminate an immediate hazard(s) to the trial participants. All personnel involved in the 
conduct of this study have completed Human Subjects Protection Training. 
Dr. Michael Arndt, PhD, MPH.  
Roles and Responsibilities: Lead protocol and questionnaire development, conduct site 
evaluation and study initiation visits, and direct quantitative analyses. Project oversight to 
complete deliverables in a thorough, thoughtful, and timely manner. Direct quantitative 
analyses. Responsible for notifying the PATH Research and Ethics Committee of serious adverse 
events (SAEs) per its reporting requirements. 
Dr. James Mukabi, MD.  
Roles and Responsibilities: Review protocol and questionnaires; provide scientific and strategic 
input for study design and implementation. Responsible for overseeing the study nurse and 
pediatrician, reviewing adverse events (AEs) and SAEs submitted by the pediatrician, and 
notifying the Maseno University Ethics and Research Committee and local regulatory authority 
of SAEs per their reporting requirements. 
Gwen Ambler, MPH.  
Roles and Responsibilities: Lead SAFE protocol and questionnaire development, and direct 
qualitative analyses. Ensure project is completed within scope, on time, and within budget. 
Jaclyn Delarosa, MPH.  
Roles and Responsibilities: Contribute to SAFE protocol and questionnaire development; ensure 
compliance with ethical, regulatory, and scientific review guidelines; and assist with qualitative 
analyses. 
Peninah Murunga, MPH.  
Roles and Responsibilities: Contribute to SAFE protocol, CRF, and questionnaire development; 
oversee qualitative data collection; obtain necessary approvals.  
Alfred Ochola, MPH.  
Roles and Responsibilities: Oversee day-to-day project activities in Western Kenya. 
Dorothy Brown, Kenya Registered Nurse.  
Roles and Responsibilities: Conduct weekly check-in phone calls with caregivers to answer any 
medical questions or provide advice. Responsible for monitoring and evaluating subject safety 
in consultation with study pediatrician. 
Dr. Nick Aduro Kidaha, MBChB, MMed (Paeds). 

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Roles and Responsibilities: Conduct medical evaluations at enrollment and post-dosing visits. 
Responsible for medical oversight: monitoring, evaluating, recording, and treating all adverse 
events and serious adverse events that occur during follow-up. 
Research assistant(s), to be determined.  
Roles and Responsibilities: Conduct direct observations, caregiver interviews, data entry, and 
data management. 
B. ABSTRACT 
Environmental enteric dysfunction (EED) is an intestinal disorder common among people living in low-
resource settings (LRS), which in children has been associated with increased risk of growth stunting, 
reduced cognitive development, and reduced oral vaccine responsiveness. An effective EED 
therapeutic would offer an opportunity to improve child growth and development in LRS. One 
promising intervention, enterade® (an amino acid–based oral rehydration solution [AA-ORS]), is a 
medical food product already sold in the United States. It consists of oral rehydration salts and a 
proprietary blend of amino acids designed to restore gut function, improve nutrient and electrolyte 
absorption, and improve barrier integrity. There is evidence that this AA-ORS reduces inflammation 
and promotes healing of damaged intestinal epithelium in murine models of intestinal damage 
(irradiated gut), and it may provide benefit to pediatric EED patients. Supplementation of amino acids 
may lessen or improve intestinal injury related to enteric illnesses commonly experienced in settings 
of poor hygiene and sanitation infrastructure. We will conduct a randomized, double-blinded, 
placebo-controlled pilot study in Kakamega County, Kenya, to assess the safety, feasible dosage, and 
acceptability/tolerability of the product in the target population (children aged 12–24 months) and 
their caregivers.

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C. STUDY PROTOCOL  
i. Background and significance of the research study 
Environmental Enteric Dysfunction (EED) 
Environmental enteric dysfunction (EED) is an intestinal disorder common among people (especially 
young children) living in low-resource settings (LRS) with a high enteric pathogen burden and poor 
sanitation and hygiene (1–3). EED is characterized by mucosal inflammation, reduced barrier integrity, 
and nutrient malabsorption, and it is associated with increased risk of growth stunting, poor cognitive 
outcomes, and reduced oral vaccine responsiveness in children in low- and middle-income countries 
(4–8).  
Growth stunting (length-for-age z-score [LAZ] < –2) is associated with increased risk of chronic health 
issues including, but not limited to, cognitive deficits and increased susceptibility to infections (9–14). 
EED thus directly and indirectly contributes to child mortality and morbidity due to other common 
childhood diseases such as pneumonia, acute diarrhea, and malaria. Furthermore, deficits in cognitive 
function can diminish school performance and have lasting impacts on a child’s future financial success 
and economic productivity (12, 14). Finally, women that were stunted as children are more likely to 
suffer from malnutrition in adulthood and when pregnant, leading to low birthweights and increased 
likelihood of their children being stunted, thus completing a vicious cycle (12, 13, 15, 16). In fact, 
children who have a LAZ from less than −1 SD to −2 SD, while technically not considered stunted, have 
an increased risk of death, especially from diarrhea and pneumonia, compared to those with LAZ > −1 
SD (13).  
In sub-Saharan Africa, limited published data are available on the prevalence and incidence of EED. 
However, studies in Tanzania have observed that elevations in 1) antibodies related to microbial 
translocation and 2) markers related to immunotolerance of gut-based immunogens were associated 
with growth faltering in young children (17, 18). In the multi-site Etiology, Risk Factors and Interactions 
of Enteric Infections and Malnutrition and the Consequences for Child Health and Development (MAL-
ED) birth cohort, LAZ declined most precipitously among children enrolled at the Tanzania site; 
stunting prevalence rose from 16% at birth to 72% at 24 months of age. Fecal markers of intestinal 
inflammation and barrier disruption as well as urinary measures of intestinal permeability are 
markedly higher in children from Tanzania and South Africa in comparison with Brazilian children, who 
more closely resemble child populations in developed settings (19–21). Additional sub-Saharan 
African data highlighting negative associations between local and systemic inflammation and pediatric 
linear growth come from Malawi and Zimbabwe (22, 23). Among children under 5 years of age in 
Kenya, an estimated 26% have moderate or severe stunting, 11% are underweight, and 4% have 
wasting (24). Western Kenya has high stunting prevalence; stunting affects an estimated 31% of males 
and 35% of females under age 5 years (25).  
There remain major challenges in establishing a formal case definition for EED; however, a recently 
published editorial in the American Journal of Tropical Medicine and Hygiene noted the following 
considerations in the development of therapeutics to treat EED (26): 
To advance the EED field, we now propose a definition construct analogous to the Jones 
criteria for acute rheumatic fever, intended for therapeutic trials, and which can be modified 
as new data emerge. In building this proposal, we recognize that EED is, except for poor 
growth, rarely accompanied by clinical signs or symptoms, so our criteria highly depend on 
laboratory assessment. Accordingly, our definition is based on three domains, which, taken 

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together, indicate an acquired intestinal inflammatory disorder with substantial clinical 
impact. Domain 1 includes age, presence of linear growth failure, negative celiac disease 
testing; domain 2 includes gut histopathology consistent with EED or at least two intestinal 
deficits assessed by less invasive biomarkers; domain 3 consists of biomarkers nonspecific to 
enteric dysfunction, but representing consequences of EED. 
EED is a multifactorial disease with a complicated etiology. The main goal of an effective EED 
therapeutic is to lessen or improve intestinal injury related to enteric illnesses commonly experienced 
in settings of poor hygiene and sanitation infrastructure. Viable therapeutic candidates should have 
evidence from human and/or animal studies that the product(s) address at least one of the two issues 
that appear to play a big role in EED: repeated bouts of infection and impaired gut healing. Evidence 
from one clinical trial suggested that provision of zinc or albendazole (anthelminthic drug) may slow 
the progression of EED in children based on the L:M ratio (27). However, a subsequent clinical trial did 
not observe any effect of a combination intervention of albendazole, zinc, and multiple micronutrient 
supplements on EED progression (28). 
In contemplating development of an EED therapeutic with potential for widespread use, one 
important consideration is that, in contrast to diarrhea where symptoms immediately raise caregiver 
concern, EED manifests its symptoms slowly and chronically. Therefore, EED therapeutics may be even 
more difficult to promote than therapeutics for acute diarrhea. Furthermore, it is likely that EED 
therapeutics would need to be administered over long periods or even chronically to successfully treat 
the syndrome due to the complexity of reducing ongoing environmental exposures, thus complicating 
both patient compliance and clinical trial design. Formative, qualitative and quantitative research in 
the form of patient feasibility and acceptability studies could provide information critical to the 
development and implementation of effective EED interventions and generate data useful to inform 
the design of future clinical trials.  
Amino Acid–Based Oral Rehydration Solution 
As described in the Investigator’s Brochure (Appendix 4), enterade® (an amino acid–based oral 
rehydration solution [AA-ORS]) is a medical food product that consists of oral rehydration salts and a 
blend of amino acids that drive the uptake of water and electrolytes. The blend of amino acids appears 
to reduce intestinal damage and promote healing of the intestinal epithelium in irradiated mouse 
models commonly used to simulate gut damage encountered in inflammatory bowel diseases (29, 30). 
This AA-ORS contains water, a proprietary blend of amino acids (L-Valine, L-Aspartic Acid, L-Serine, L-
Threonine, and L-Tyrosine), sodium chloride, sodium citrate, potassium chloride, gum acacia, 
magnesium chloride, calcium chloride, and stevia leaf extract (sweetener). AA-ORS may be useful in 
treating children with EED, as low circulating levels of three of the five amino acids present in AA-ORS 
(valine, threonine, and serine) have been associated with stunting among children aged 6–59 months 
in rural Malawi (31). In an experimental irradiated mouse model, mice treated with a solution of amino 
acids similar to the AA-ORS for this trial (containing lysine, aspartic acid, glycine, isoleucine, threonine, 
tyrosine, valine, tryptophan, and serine) showed increased electrolyte absorption and decreased 
paracellular permeability, IL-1β levels, and plasma endotoxin levels in comparison to untreated 
controls. Treated mice also had increased weight gain and better survival following irradiation 
compared to untreated controls (29). In another study, a simpler amino acid– based oral rehydration 
solution (ORS) containing threonine, valine, serine, tyrosine, and tryptophan increased enterocyte 
proliferation, maturation, and differentiation and improved electrolyte and nutrient absorption in 
irradiated mice (30).  

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The safety and efficacy of AA-ORS have been tested in cohorts of adult cancer patients, including an 
ongoing study by the Dana-Farber Cancer Institute (NCT02919670). This AA-ORS was tested in more 
than 300 Indonesian adults with diarrhea, resulting in no observed serious adverse events (SAEs) or 
adverse events (AEs) (32). In a small hydration study among US adults comparing water, ORS, AA-ORS, 
and a commercial sports drink, AA-ORS and ORS both hydrated significantly better than water 
(NCT03262597). This AA-ORS has not yet been evaluated in children, save for a small ongoing study in 
five patients (2–14 years of age) with short bowel syndrome (NCT03105362). No AEs were observed 
in pediatric or adult studies to date (32). Pending receipt of study funding, this AA-ORS will be tested 
by icddr,b among children 6–36 months of age in Bangladesh for effectiveness in decreasing the 
duration and severity of diarrhea. AA-ORS is categorized by the US FDA as a medical food, like 
Pedialyte®, which is defined in section 5(b)(3) of the Orphan Drug Act (21 U.S.C. 360ee(b)(3)), as: 
‘a food which is formulated to be consumed or administered enterally under the supervision 
of a physician and which is intended for the specific dietary management of a disease or 
condition for which distinctive nutritional requirements, based on recognized scientific 
principles, are established by medical evaluation.’…The patient should generally see the 
physician on a recurring basis for, among other things, instructions on the use of the medical 
food as part of the dietary management of a given disease or condition. (33) 
This AA-ORS is not currently marketed or approved for use in Kenya. It would be classified as a “food 
supplement” according to local guidelines and would fall under the authority of Kenya’s Ministry of 
Health (MOH) regulatory body, the Pharmacy Poisons Board (PPB). The PPB is responsible for 
approving and overseeing the importation and use of unregistered products to be used in clinical trial 
settings in Kenya. A license for importation will be obtained from PPB prior to shipment of study 
product (i.e., AA-ORS and placebo) and any required regulatory approvals will be secured in 
accordance with PPB guidance before trial initiation.  
The active ingredients of the AA-ORS are described in Table 1, below. 
AA-ORS
Total mg
(in 237 ml) 
Aspartic Acid (g/L)
1.06
251.2
Threonine (g/L)
0.95
225.2
Tyrosine (g/L)
0.21
49.8
Valine (g/L)
1.17
277.3
Serine (g/L)
1.05
248.9
Sodium (mEq/L)
42.25
230.3
Potassium (mEq/L)
10
92.4
Citrate (mEq/L)
3.35
50.0
Magnesium (mEq/L)
1.2
3.4
Calcium Chloride (mEq/L)
1.2
15.6
 
The placebo solution for this study is the same as that being used in the ongoing Dana-Farber Cancer 
Institute (NCT02919670) study, and contains all the non-amino acid and non-electrolyte ingredients 
of the AA-ORS including purified water, trisodium citrate, citric acid, stevia, and natural flavors. All the 
non-amino-acid AA-ORS ingredients are generally regarded as safe (GRAS), a designation used by the 
US Food and Drug Administration (FDA) to classify components of food that are safe and suitable for 
extensive use in the United States population. Amino acids are not considered GRAS, but are governed 
by 21CFR172.320, a food additive regulation that sets out the conditions under which amino acids 

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used as food additives may safely be added to foods. Section F of this regulation refers to medical 
foods like this AA-ORS: 
(f) The food additive amino acids added as nutrients to special dietary foods that are intended 
for use solely under medical supervision to meet nutritional requirements in specific medical 
conditions and comply with the requirements of part 105 of this chapter are exempt from the 
limitations in paragraphs (c) and (d) of this section and may be used in such foods at levels not 
to exceed good manufacturing practices. 
For children at age 1-2 years, the World Health Organization (WHO) reports the average daily intake 
requirement of threonine and valine as 23 mg/kg and 36 mg/kg, respectively (34). At 12 months of 
age, healthy female children average 9 kg. Such a child would therefore require a minimum of 207 mg 
of threonine and 324 mg of valine. Estimated requirements for overall daily protein intake in children 
under 2 years of age range from 1 to 1.2 g/kg, and therefore a 9-kg child needs between 9 and 10.8 g 
of protein daily (35). 
Table 2. Amino acids in breastmilk and amino acid–based oral 
rehydration solution (AA-ORS) based on expected daily intake for 
a 10–12 month-old child (36). 
Amino Acid
Breastmilk 
(mg in 800 ml) 
AA-ORS
(mg in 237 ml)
Total daily mg 
Aspartic Acid
682
251.2
933.2
Threonine
364
225.2
589.2
Tyrosine
394
49.8
443.8
Valine
438.4
277.3
715.7
Serine
352.8
248.9
601.7
 
Most doctors recommend that women breastfeed their baby for at least 1 year (37). While breastmilk 
is the beverage recommended by the American Academy of Pediatrics guidelines for infants under 1 
year of age (38), children aged 12 months and older are consuming many other beverages in 
considerable volumes. A 2004 survey among several thousand parents in the United States reported 
that more than two-thirds of children between 12 and 14 months were consuming non-milk beverages 
in addition to breastmilk (39). The low total caloric content of AA-ORS makes this product unlikely to 
satiate children and reduce their consumption of breastmilk. After 6 months of age, an increasing 
proportion of calories in the young child diet come from solid foods. 
Oral rehydration solution (ORS) recommended for treatment of dehydration and acute diarrhea has 
similar osmolality to AA-ORS, and the two share some common ingredients, including potassium and 
sodium. However, a key difference (aside from its amino acid content) is that this AA-ORS lacks 
glucose. Glucose is widely held to be a key component in the rehydration mechanism of ORS, although 
according to some researchers, it has detrimental effects as well (40, 41). Table 3, below, from a poster 
created by researchers at the University of Florida compares the ingredients of ORS, a sports drink, 
and AA-ORS. The beverages (including water) and their corresponding osmolalities are as follows: 
distilled water (~0 mmol/kg), AA-ORS (195 mmol/kg), ORS (270 mmol/kg), and a sports drink (330 
mmol/kg). This poster can be found here. 

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Table 3. Comparison of the test beverage composition. 
 
Should AA-ORS prove to be effective for treating EED alone or in combination with other interventions, 
additional research will be necessary to optimize the dosing schedule for children of different ages, 
improve the formulation (e.g., powder sachets rather than bottles of solution), and target/prioritize 
subgroups most likely to benefit. The amino acid components of AA-ORS may be sufficient to elicit a 
therapeutic benefit, even when not formulated as an aqueous solution. Such questions would need 
to be answered in order to optimize the intervention for deployment in LRS. 
ii. Aims of proposed research study 
We propose to conduct a small pediatric pilot study to determine patient safety, feasible product 
volume (trends in dosage consumed over 14 days, etc.), and caregiver (i.e., parent or legally acceptable 
representative) acceptability of AA-ORS in the target population at risk for EED. To assess the product’s 
safety, we will randomize children to receive active product or placebo and compare the rate of 
adverse events (AEs) in the two groups. We will also assess the extent to which the product might 
meet the needs of the target population and setting. Results from this study will be used to inform:  
a) The design of a randomized controlled trial to test the efficacy of AA-ORS for treating pediatric 
EED. 
b) Future EED product development efforts. 
c) Program implementation strategies for EED interventions.  
Objectives  
PRIMARY OBJECTIVE: To determine the safety of AA-ORS among children with mild to moderate 
stunting, administered as a daily dose over 2 weeks, ad libitum up to 237 ml in 3 hours. 
Safety assessment in children will compare adverse event (AE) and serious adverse event (SAE) 
rates and changes to dietary and digestive habits between children receiving AA-ORS versus an 
inactive placebo solution containing no amino acids.  
PRIMARY OBJECTIVE: To determine the feasibility and well-tolerated dose of AA-ORS among children 
with mild to moderate growth stunting administered as a daily dose over 2 weeks, ad libitum up to 
237 ml in 3 hours. 
User preferences among children include average daily volume consumed over 14 days, trends in 
volume consumed, and factors that influence children’s willingness and ability to ingest the 
product as intended.  
SECONDARY OBJECTIVE: To determine the perceptions among caregivers on the use and acceptability of 
this product as a potential intervention for EED. 
Acceptability by caregivers includes factors that influence their willingness to have the product 
used by their children and their perception of their child’s response to its consumption.  
EXPLORATORY OBJECTIVE: To determine the impact of AA-ORS on gut dysfunction, metabolism, gut and 
systemic inflammation, and micronutrient status. 

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Impact of AA-ORS on gut dysfunction, systemic inflammation, and micronutrient status will be 
assessed by comparing non-invasive biomarker levels in blood samples collected following the 
intervention. These biomarkers will be assessed in both groups at baseline in order to have setting-
specific descriptive data with which to power a future EED efficacy trial. 
Endpoints 
Primary: Adverse events and serious adverse events through 21 days of follow-up, and changes to 
dietary and digestive habits through day 14 of follow-up.  
Primary: Average product volume consumed daily through 14 days of follow-up, time trends in 
average daily volume consumed, and factors that influence children’s willingness and ability to ingest 
the product as intended.  
Secondary: Key factors that influence caregiver willingness to have the product used by their children, 
and their perception of their child’s response to its consumption. 
Exploratory: Average plasma/serum concentrations of metabolic, gut dysfunction, systemic 
inflammation, and micronutrient biomarkers at baseline and day 15 of follow-up. 
iii. Study design, population, and study procedures 
Study Approach and Study Population 
To achieve the objectives listed above, we will closely monitor adverse events (AE) and serious adverse 
events (SAE) among study participants, conduct direct observations (DOs) and semi-structured in-
depth interviews (IDIs) with potential end users (caregivers) in Kenya. IDIs will be undertaken to obtain 
an in-depth understanding of acceptability among end users (caregivers of pediatric patients 12–24 
months of age) after AA-ORS use (post-deployment). The DOs will involve the study team observing 
initial administration of AA-ORS or placebo solution to a participating child. We will also gather and 
test blood samples in order to understand enteric dysfunction, systemic inflammation, and 
micronutrient deficiency in this child population. 
The study population is 66 child/caregiver pairs. Eligible children will be 12–24 months of age without 
diarrheal or febrile illness, within the prior 7 days, who are experiencing mild to moderate stunting 
(LAZ between –3 and –1). Children with mild to moderate stunting at this age do not fall under the 
WHO guidelines for exclusive breastfeeding (< 6 months of age), are at increased risk for morbidity 
and mortality compared to children without stunting (13, 42), and may still be able to regain linear 
growth. That is, when children are under the age of 24 months, their growth is more plastic. Thus, 
stunting is more easily reversed than in older children. On an individual level, growth stunting is not 
always caused by EED, and those suffering from EED may not always have stunting. However, we have 
taken a practical or proxy case definition for “at risk for EED” based on stunting. This will allow us to 
determine how AA-ORS is tolerated in a population that at least overlaps considerably with true EED 
cases. Caregiver/child pairs will be enrolled and randomized into one of two groups receiving a daily 
dose of either (1) AA-ORS or (2) placebo solution (no amino acids) for 2 weeks, ad libitum over 3 hours 
up to 237 ml each day.  
The study will be double-blinded as both study staff and participants will be blinded to study product 
allocation. Active and control product bottles will be labeled with identical product information. Each 
bottle will be also labeled with a randomization number assigned to a particular participant according 
to the appropriate allocation arm. The randomization list where randomization number and allocation 
code are provided will be maintained in a secure place by an independent statistician. The participants 

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will thus be blinded to whether they receive AA-ORS or the control solution. Study staff administering 
the study product and laboratory staff performing any laboratory testing will also be blinded to the 
product allocation. 
This study will provide critical information about the safety of AA-ORS in this at-risk pediatric 
population and qualitative and quantitative information on the user-preferred product volume for this 
age group, palatability, and ease of administration. The study will also describe whether the product 
can be feasibly consumed for a 2-week period and if there are differences in gut health between AA-
ORS and placebo groups based upon diarrhea frequency and plasma biomarkers of systemic 
inflammation and gut damage/repair at the end of follow-up. However, the study has not been 
statistically powered or designed to answer such gut health questions. Even though such exploratory 
comparisons will be underpowered, the baseline and follow-up biomarker data will be useful for 
informing study design and sample size of future EED clinical trials in this setting. 
The questionnaire guides are a mix of structured and semi-structured questions. Structured questions 
are necessary to standardize responses to questions such as the experience with diarrheal disease 
treatment. Semi-structured questions allow for flexibility and richness in responses, especially those 
on perceptions and experiences around AA-ORS use and acceptability. Semi-structured questions are 
open-ended and allow for probing a participant’s answers for in-depth responses. The DO checklist 
includes a list of questions to assess the administration of AA-ORS and explore ease of use, palatability, 
and acceptance of the product by the user(s). 
Study Site 
The study will be conducted in Kakamega County in Western Kenya. This county was selected as 
containing both rural and urban settings and is suitable for this research due to the high prevalence 
of malnutrition and PATH Kenya’s established relationships with health facilities in the area. In former 
Nyanza province, the stunting (LAZ < –2) prevalence in children under 5 years ranges from 2.5% in 
Migori County to 5.14% in Kisumu County. In neighboring Western province, where Kakamega is 
located, the prevalence is thought to be much higher.  
PATH has been involved in the establishment of the APHIAplus program in Western Kenya. APHIAplus 
promotes HIV testing and improved access to ORS for children experiencing diarrhea through ORS 
corners in several hundred health facilities in the region. Participants will be recruited from Kakamega 
County General Hospital (CGH). This facility draws from a peri-urban catchment area, has been 
participating in APHIAplus programming, and has set up an ORS corner for treatment of pediatric 
diarrhea. Therefore, health care providers in the facility are quite familiar with the use of ORS. 
Key Activities 
Following the training of research assistants (RAs), piloting study IDI guide(s) and questionnaires, and 
subsequent IDI guide and questionnaire revision, the key activities will include the following:  
1. Screen potential participants at routine immunization visit or vitamin A (Vit. A) supplementation 
visit after the child is 12 months of age and schedule enrollment visit at least 1 week later. OR: 
identify potentially eligible children from using the CGH child welfare clinic (CWC) register of 
patients who received immunizations in the first year of life. Call the caregivers of prospective 
participants using the telephone number provided, and schedule screening visit with interested 
caregivers (and their child). Schedule enrollment visit any time following the screening visit. 
2. Following informed consent, participants will be randomized to receive active product or placebo 
using sealed, unmarked envelopes with numbers generated previously by a biostatistician. These 

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envelopes will contain randomization numbers that are linked to numbers printed on a set of 
bottles containing either AA-ORS or placebo solution. Once the randomization number has been 
attached to a patient ID number, that participant will only receive bottles labeled with his/her 
randomization number.  
3. At enrollment, blood samples will be collected (between 1 and 1.5 ml) using venipuncture into 
heparinized collection tubes. Samples will be centrifuged into plasma or serum within 30 minutes 
of collection and stored in cryo tubes at –20°C until they can be analyzed.  
4. Study RAs will conduct a baseline questionnaire with caregivers before study product use and will 
perform IDIs with caregivers after using AA-ORS or placebo with their child at enrollment and then 
after daily study product use for 2 weeks. The RA will explain product directions to caregivers 
(recommend using a cup and spoon, provided) and give an instruction sheet to take home. RAs 
will conduct DOs with caregivers and their children, observing the administration of the solution 
for the first dose provided, for up to 3 hours. Any solution that has not been consumed after 3 
hours will be measured, recorded, and discarded. Patients will be monitored in the clinic for 30 
minutes following the end of product consumption (after 3 hours, or the 237 ml is consumed, 
whichever occurs first). A study nurse will be available on site in case the child needs medical 
attention.  
5. Interim visits: RAs will visit the home on days 2, 4, 7, and 14 to collect unconsumed solution, review 
instructions, provide additional bottles (on days 2 and 7 only), and weigh the children (on days 7 
and 14 only). 
6. Adverse event check-ins: Periodically during dosing (days 4, 7, 14, and 21), and one week after the 
end of dosing (day 21), a study nurse will contact the caregiver by phone and check in with the 
caregiver to assess any potential AE or SAE, complete the routine interval medical history (with 
feeding and breastfeeding assessment)/stool frequency questionnaire, answer any medical 
questions, and provide advice, as necessary. 
7. On the fifteenth day of follow-up (or within 2 days after), following the 2-week dosing period, 
patients and their caregivers will return to the clinic for medical evaluation, collection of blood 
samples, and caregiver IDIs. Samples will be processed and stored for laboratory analyses 
following the same procedures from the enrollment blood collection.  
Study Flow Diagram  
 
Enrollment visit: 
medical exam, 
randomization, DO
• Signing of informed 
consent
• Medical examination
• Collection of blood 
samples
• Pre-administration 
questionnaire with 
caregivers regarding 
stunting and EED 
intervention perceptions
• Observe administration 
of the first dose by 
caregivers
• Assess potential adverse 
events up to 30 minutes 
after first dose
• Post-administration IDI 
with caregivers
Interim visits: days 
2, 4, 7, 14
• Collect unconsumed 
solution
• Provide additional 
bottles (days 2, 7)
• Review instructions
• Weigh the children (days 
7 and 14)
Adverse event 
assesments: days 4, 
7, 14, 21
• Complete the routine AE 
and interval medical 
history questionnaires
• Answer any medical 
questions, and provide 
advice, as necessary
Study exit and 
medical exam visit: 
day 15 (+ 2 days)
• Medical examination
• Collection of blood 
samples
• Study exit IDI with 
caregivers
Two weeks of daily dosing 

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Study Products 
AA-ORS is an amino acid–based, glucose-free, medical food/beverage that is lightly sweetened with 
stevia leaf extract. AA-ORS provides select amino acids and electrolytes (sodium and potassium)—
nutrients intended to rebuild and protect the gastrointestinal tract and deliver hydration. 
The AA-ORS for use in this trial, enterade®, will be commercially produced by Entrinsic Health 
Solutions, Inc. in accordance with US FDA standards for medical food products and good 
manufacturing practices. The AA-ORS will be used as supplied, meaning that other than packaging and 
labeling, there is no further study product preparation required. The corresponding placebo solution 
will be produced by Entrinsic Health Solutions for clinical trial use only.  
In this study, AA-ORS and placebo solution will be formulated in bottles with 237 ml of clear liquid. All 
bottles will be clearly labeled as for research use only. Labels will meet all national and local 
requirements. The label must also include the product expiry date, batch number, and manufacture 
date. 
The AA-ORS is stable at room temperature and has a 2-year shelf life when stored unopened in 
ambient temperatures below 25oC. Long-term (6-month) stability of the AA-ORS has also been 
demonstrated at 40oC with 65% relative humidity. The AA-ORS is stable for the entire study period, 
even at higher environmental temperatures that may be encountered in participants’ homes during 
the study. The placebo solution is presumed stable. All study product not already dispensed to 
participants will be stored securely, only accessible to the study staff.  
The study investigators will be required to maintain complete records of all study products received 
from the sponsor and/or manufacturer and will be responsible for maintaining an accurate record of 
the randomization codes and inventory and an accountability record for this study. The study 
investigators will also be responsible for ensuring the security of these documents, maintaining them 
under lock and key and/or electronic encryption.  
At the completion of the study, the study investigators and site PI (or designee) will conduct and 
document a final reconciliation of all study product shipped, received, dispensed, consumed, and 
remaining. Any discrepancies identified will be investigated, resolved, and documented before any 
unused study product is destroyed. After all accounting and reconciliation procedures are complete 
and approved by the sponsor, all unused study product will be destroyed on site and documented in 
the master study files.  
Study Procedures 
All IDI guides and data collection instruments will be pretested prior to data collection for gauging 
time required and flow of questions. IDI guides will be revised and adapted accordingly prior to data 
collection. Therefore, research assistant (RA) training will be composed of 1) review of guides and 
instruction on carrying out recruitment and consenting procedures, enrollment and interim 
questionnaires, weighing of children, and IDIs; 2) pretesting of IDI guides; and 3) revision/adaptation 
of guides with investigators, as necessary. RAs will field/pilot test the IDI and other questionnaires 
with a small number of caregivers in Kakamega, Kenya, before finalizing the instrument. Results from 
initial pretesting will be used to refine the data collection instruments before starting the study. 
Specifically, testing will be used to confirm target respondents are able to understand the questions 
being asked, that questions are understood in the same way by all respondents, and that respondents 

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are willing and able to answer questions. Table 4 presents an overview of data collection procedures 
and timing.

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Table 4. Table of procedures (N=66 child/caregiver pairs). 
 
Screening 
Enrollment 
(Day 1) * 
Day 2 
Day 4 
Day 7 
 
Day 14 
Day 15 
Day 21 
Comments 
Review of medical records for child 
eligibility 
X 
 
 
 
 
 
 
 
 
 
Obtain consent 
 
X 
 
 
 
 
 
 
 
 
Physical exam 
X 
X 
 
 
 
 
 
X 
 
 
Blood draw (1–2 mL) 
 
X 
 
 
 
 
 
X 
 
 
Enrollment questionnaire (caregiver) 
 
X 
 
 
 
 
 
 
 
 
Teaching caregiver to administer 
study product 
 
X 
 
 
 
 
 
 
 
 
Direct observation of feeding 
 
X 
 
 
 
 
 
 
 
 
AE assessment 30 mins after dose 
 
X 
 
 
 
 
 
 
 
 
In-depth interview (caregiver) 
 
X 
 
 
 
 
X 
 
 
 
Supply product to home/pick up 
unused product 
 
 
X 
X# 
X 
 
X 
 
 
 
Review product administration with 
caregiver 
 
 
X 
 
X 
 
X 
 
 
 
Text message to remind caregiver to 
start and stop study product 
 
 
X 
X 
X 
 
X 
 
 
Twice daily during 2-week daily product 
administration. 
Weight assessment 
X 
 
 
 
X 
 
X 
 
 
Done at the household, but may be done at 
the clinic, if needed. 
Check in on adverse events; 
administer AE/interval medical 
history questionnaire 
 
X  
 
X 
X 
 
X 
 
 
Done by study nurse in person on day 1, and 
then by phone all others. 
Final recording of SAEs and close out 
of AEs 
 
 
 
 
 
 
 
 
X 
Done by study nurse, as needed, by phone 
or in person. 
Report SAEs/SUSARs to regulatory 
entities 
 
X 
X 
X 
X 
 
X 
X 
X 
 
*Enrollment will be at least 1 week post routine immunization, healthy child visit, or vitamin A supplementation visit at the study site, if recruited in-person rather than through 
CGH register review. 
#Day 4 is for pick-up of unconsumed product (bottles) only. 
Abbreviations: AE, adverse event; SAE, serious adverse event; suspected unexpected serious adverse reaction, SUSAR.  
 

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Screening visit: 
During the screening visit, potentially eligible participants will be identified by clinic staff through the 
clinic records and routine information gathered at Expanded Program on Immunization (EPI)/Vit. A 
supplementation visits. Caregivers and their children will be referred to a study RA by clinic staff so 
the study RA can inform the caregivers about the study activities. Alternatively, potentially eligible 
children and their caregivers will come to the clinic after being identified from the CWC register and 
contacted by study staff (as described in Recruitment and Consent Procedures, below). As part of 
routine care, the child will receive a physical exam by a clinician or nurse. The routine physical 
examination conducted at EPI and Vit. A supplementation visits includes assessment of weight, weight 
categories (underweight, severe underweight, overweight, obese), height/length, stunted (stunted, 
severe stunted), and mid-upper arm circumference (MUAC) (green, yellow, red). If appropriate, the 
children identified from the CWC register will be given Vit. A supplements and any missing 
immunizations by clinic staff. If the caregiver is interested in participating in the study, he/she will be 
given a copy of the consent form and patient information leaflet to take home and review before 
coming back to the clinic for enrollment. In the case of an illiterate caregiver, the RA will confirm that 
there is at least one literate adult living in the child’s household, and the RA will offer to read the 
consent form in full with the caregiver before sending the caregiver home. Any review of medical 
records prior to completion of informed consent will be done solely to identify potential participants 
for this study. Only data relevant to screening criteria will be recorded. If the caregiver declines 
enrollment in the study, any medical record data collected by study staff will be shredded and 
discarded.  
 
Enrollment visit: 
Upon return to the clinic (≥ 1 week post screening visit) the RA will review the consent form with the 
parent or legally acceptable representative and answer any questions. The caregiver will be asked if 
they are still interested in continuing with participation in the study; if they agree, then the RA will 
complete the informed consent process and obtain their written signature on the consent form. 
Once the consent form is signed and a copy has been given to the participant, the study nurse or 
clinician will conduct a medical examination, and a trained phlebotomist will draw blood from the 
child and collect a bloodspot on filter paper (see “Blood specimen collection and processing“). The RA 
will then proceed with the enrollment questionnaire and direct observation of the oral solution. For 
all children who join the study and are still breastfeeding, the enrollment questionnaire will include 
questions to establish a baseline breastfeeding rate (how many times a day). After direct observation 
of dosing with the solution, an RA will complete an IDI with the caregiver.  
 
Training of caregivers 
Caregivers will receive one 237-ml bottle of AA-ORS or placebo at enrollment along with a small cup 
and spoon to aid administration. The RA will train the caregiver to give the solution to their child and 
show them how to properly seal the bottle (containing unused solution) following solution provision 
(Appendix 6). The caregiver will administer the solution to their child in a private room in the clinic, 
under direct observation by an RA for 3 hours or until the solution has been finished, whichever comes 
first. Breastfeeding and solid feeding is permitted whenever desired during the 3-hour period.  
 
Direct observation: 
The goal of DOs is to directly witness the administration of AA-ORS or placebo solution and instruct 
caregivers to properly reseal bottles with unused solution. Target observations during the DOs include: 

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 
Perceived palatability of study product by children. 
 
Perceived acceptable volume of study product by children.  
 
Ease of administration of study product by caregivers to their child. 
 
Interim visits: 
The following day after enrollment, the RA will visit the caregiver in their home to review the 
instructions and give the caregiver six additional 237-ml bottles. The RA will instruct the caregiver to 
give doses using the same technique and time limit used in the clinic for the next 20 days, to reseal 
unconsumed solution in bottles, and store bottles in a safe location (Appendix 6). The RA will help the 
caregiver to select an appropriate storage location for new and used bottles of AA-ORS/placebo that 
is out of reach of children and protected from heat and sunlight. 
 
Each day during follow-up, an automated text message (SMS) will be sent to remind caregivers to 
administer the study product, and a second message (3 hours later) will remind them to mark and 
reseal bottles using tape and markers provided at enrollment. If automated SMS proves unfeasible, 
RAs will send SMS reminders to caregivers manually. The RA will visit the home on days 2, 4, 7, and 14 
to collect unconsumed solution, review instructions, provide additional bottles (on day 2 and 7), and 
on days 7 and 14 only weigh the children. All empty or partial bottles will be collected, and 
unconsumed solution will be measured as part of the longitudinal volume feasibility assessment.  
Interval medical history interviews: 
Caregivers will receive study nurse contact information and can contact the nurse by phone at any 
time during the course of the study. Periodically during dosing (days 4, 7, 14, and 21) and 1 week after 
the end of dosing (day 21), a study nurse will contact the caregiver by phone and check in with the 
caregiver to assess any potential adverse events or serious adverse events, complete the routine 
interval medical history/stool frequency questionnaire, ask if the child is still breastfeeding (for 
children who were still breastfeeding at enrollment), answer any medical questions, and provide 
advice, as necessary. If breastfeeding has stopped, the caregiver will be asked if the child instituted 
the stoppage or if the mother decided to wean. All AEs and SAEs reported by the nurse will be 
reviewed by the study clinician, who will provide medical advice and care as needed. 
 
Post-dosing visit: 
Upon return to the clinic (within 3 days of the final dose) the study nurse or clinician will conduct a 
medical examination, and a trained phlebotomist will draw blood from the child and collect a 
bloodspot on filter paper (see “Blood specimen collection and processing“). After the medical 
examination and blood draw, an RA will complete an IDI with the caregiver. For all children who were 
still breastfeeding at baseline, the IDI will include questions to determine the breastfeeding rate (how 
many times a day) at the end of the study. The IDI will also assess whether breastfeeding has stopped 
in the prior week, and if so, if the child instituted the stoppage or if the mother decided to wean.   
 
On day 21, a study nurse will contact the caregiver by phone for a final check-in with the caregiver to 
assess any potential adverse events 1 week after study product dosing ended. 
 
In-depth interviews 
 
Sixty-six IDIs with caregivers, after initial study product use and after prolonged use:  
The goals of the IDIs are to understand the experiences of the caregivers when administering study 
product to their child, how they perceive the value of the product, the potential advantages and 

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disadvantages of study product, and the factors relating to uptake. Topics explored in the IDIs will 
include:  
o Perceived palatability of solutions by children. 
o Ease of use, including how easy it was to understand use instructions and administer to the 
child. 
o Behavioral intention to use. 
o Perceived acceptability of study product by caregivers of children. 
Please see Appendix 2 for IDI guide and DO checklist. 
Blood Specimen Collection and Processing  
Between 1 and 1.5 ml venous blood will be collected from each child twice—at enrollment and at the 
post-dosing visit, 15 days after onset of the intervention. A trained phlebotomist will perform 
venipuncture using a technique appropriate for children who are younger than 2 years old, and a small 
drop of blood will be blotted from the insertion site and dried on filter paper. Blood samples will be 
centrifuged into plasma and/or serum within 30 minutes of collection and stored in cryo tubes at less 
than –20°C until they can be analyzed. Commercial enzyme-linked immunosorbent assays (ELISA) and 
a Quansys Biosciences Q-Plex™ panel will be used to quantify plasma/serum micronutrients, EED 
markers (intestinal fatty acid-binding protein [I-FABP], soluble CD14 [sCD14], glucagon-like peptide 2 
[GLP-2], insulin-like growth factor 1 [IGF-1], and fibroblast growth factor 21 [FGF21]), and systemic 
inflammation (alpha-1-acid glycoprotein [AGP] and C-reactive protein [CRP]). Mass-spectroscopy will 
be used to evaluate metabolic markers from dried bloodspot (e.g., acylcarnitine profile) to ascertain 
whether there is a tendency for child metabolism to switch to an oxidative state when supplementing 
a subset of essential amino acids. Detail provided in Table 5, below. 
Table 5. Plasma/serum biomarkers assessed in SAFE at enrollment and post-dosing visit.
Biomarker 
Assay name 
Company 
μl of plasma required 
(per well) 
I-FABP 
Human I-FABP ELISA 
Hycult 
25 
GLP-2 
GLP-2 ELISA Kit 
ALPCO 
25 
IGF-1 
Human IGF-I Quantikine ELISA Kit 
R&D Systems 
20 
sCD14 
Human CD14 Quantikine ELISA Kit 
R&D Systems 
10 
 
 
 
 
FGF21 
Human FGF-21 Quantikine ELISA Kit 
R&D Systems 
50 
AGP 
Q-Plex™ Human Micronutrient (7-Plex) 
Quansys 
10 
CRP 
Ferritin 
sTfR 
RBP4 
Thyroglobulin 
HRP2 
Acylcarnitine 
Acylcarnitine profile analysis 
Mass 
spectrometry 
40 (80 µl whole blood)
Total 
320 
 

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Discontinuation of Study Product Use and Withdrawal From the Study 
Participants have the right to decline study procedures for any reason and at any time during the study 
with no penalty. If a participant decides to discontinue study procedures, including AA-ORS or placebo 
administration, this will be recorded as a study deviation and the reason will be clearly documented.  
Participants have the right to withdraw from the study at any time and for any reason, without penalty. 
If a caregiver does not wish to remain in the study and/or decides to stop administering the solution 
altogether to his/her child, the participant can choose to withdraw consent and be withdrawn from 
the study. That decision will be documented, and study staff will attempt to schedule a final post-
dosing IDI and medical examination as soon as possible to confirm the welfare of the child. 
The principal investigator and site PI may, at their discretion, withdraw a subject from continuing in 
the study if it is considered to be in the participant’s best interest to do so, or if the participant is not 
willing or able to comply with the study requirements. The reason for withdrawal will be documented. 
Withdrawn subjects will not be replaced in the study. The data collected before withdrawal will be 
analyzed. 
Withdrawal from further study product administration may be at the discretion of the PI or site PI, in 
consultation with the study pediatrician if it is in the interest of the subject. In addition, participants 
will be withdrawn from further study product administration for the following reasons: 
 
Ineligibility (either arising during the study or retrospectively having been overlooked at 
enrollment). 
 
Significant protocol deviation. 
 
Significant noncompliance with the study product regimen or study requirements. 
 
Significant weight loss: if a child loses more than 5% of his or her body weight from that measured 
at enrollment, the child will be withdrawn from the study. The study team will refer the withdrawn 
child and caregiver to his or her primary health care provider or CGH, as appropriate.  
 
An AE (including intercurrent illness, diseases, or medical treatment) that requires discontinuation 
of study product use or results in inability to continue to comply with study procedures (Appendix 
2). The study team will refer anyone that develops an acute illness during the study to his or her 
primary health care provider or CGH, as appropriate. 
 
Sampling Strategy 
Estimated sample size: We expect to enroll 132 participants, which includes 66 caregivers, and 66 
children (aged 12–24 months); half of these child-caregiver pairs will be randomized to receive AA-
ORS (n= 33) and half to receive placebo solution (n= 33). We anticipate that there will be a dropout 
rate not to exceed 10%. We have incorporated this expected withdrawal rate into the final sample 
size numbers.  
For the primary endpoint of safety in this study, 30 subjects in each arm would result in more than 
95% probability of observing at least one AE if the rate of that type of event in the subject population 
after being exposed to AA-ORS is 10%. To have a 95% probability of being able to detect at least one 
AE if the rate of that event is truly 5%, the sample size would have to be at least 60 per arm. For more 

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rare events, the sample size would have to be much larger. Table 6 below shows the probabilities of 
observing an event with various sample sizes and varying true rates of that event in the population.  
Table 6. Probability (P) of observing none, one, or more adverse events (AEs) at different true rates of 
AEs in the population.  
N (per arm) 
Rate of AE 
P of 0 AEs 
P of 1+ AEs 
P of 2+ AEs 
P of 3+ AEs 
20 
0.15 
3.9% 
96.1% 
82.4% 
59.5% 
0.2 
1.2% 
98.8% 
93.1% 
79.4% 
30 
0.1 
4.2% 
95.8% 
81.6% 
58.9% 
0.15 
0.8% 
99.2% 
95.2% 
84.9% 
40 
0.1 
1.5% 
98.5% 
92.0% 
77.7% 
0.15 
0.2% 
99.9% 
98.8% 
95.1% 
50 
0.1 
0.5% 
99.5% 
96.6% 
88.8% 
0.15 
0.03% 
99.97% 
99.7% 
98.6% 
60 
0.05 
4.6% 
95.4% 
80.8% 
58.3% 
0.1 
0.2% 
99.8% 
98.6% 
94.7% 
Gray highlighted cells are those with a probability of 95% or higher. Blue highlighted cells are those 
representing a true AE rate of 0.1 or lower. 
Increasing the per-arm sample size above 30 (but < 60) does not add much in terms of increasing the 
probability of detecting an AE with a true rate in the population of 10%. However, 30 subjects per arm 
has relatively low power for an efficacy endpoint using biomarkers. For example, a sample size of 30 
per arm provides approximately 80% power for a t test to detect a 376 ng/ml (21.9% reduction) 
difference in soluble CD14 (marker of bacterial translocation) between the arms assuming the mean 
sCD14 is 1,715 ng/ml in the placebo group. These calculations used data from an infant cohort in 
Zimbabwe (23) (SD = 520 ng/ml).  
Recruitment and Consent Procedures 
Recruitment of child and caregiver participant pairs:  
Children and their caregivers potentially eligible for the study will be identified by clinic staff from the 
child welfare clinic (CWC), which is the outpatient department where they are administered routine 
child immunizations (measles and yellow fever at 9 months of age, but sometimes postponed until the 
second year), healthy child visits, or receive routine Vit. A supplementation (at 12 and 18 months of 
age). Potential participants will be screened using medical records that include child anthropometry 
and medical history. Height and weight will be converted into z-scores using an algorithm.   
a. Recruitment at clinic visit: During the potential participant’s CWC visit, a study RA will 
introduce the study to the caregiver and ask if they are interested in learning more. Following 
a short recruitment script (Appendix 5), study staff will inform the caregivers that a research 
study on a product that may improve pediatric gut health will be tested. If the caregiver is 
interested in learning more about the study, the caregiver will be provided with a product 
information form and study consent form to take home and review before returning to the 
clinic for an enrollment visit. If the caregiver declines enrollment in the study, any medical 
record data collected will be discarded and shredded.  
b. Recruitment from CWC register: Study RAs will identify potentially eligible children from CGH 
CWC register that lists childrens’ names, age, caregiver contact information, exclusive 
breastfeeding status, and anthropometric measurements from patients receiving 
immunizations in the first year of life. The hospital has approved using retrospective data from 
the CWC register for this pre-screening activity. The CWC register is an alternative recruitment 

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mechanism whereby the caregivers of children who have been seen previously for routine EPI 
immunizations (such as measles and yellow fever vaccine at age 9 months) will be contacted 
for potential interest in study participation. The RA will record information into a pre-
screening log from the register for children who would currently meet age eligibility 
requirements, limited to CWC visit date, child age, name, and contact number. RAs will call 
the caregivers of prospective participants using the telephone number provided and introduce 
the study to the caregiver and ask if they are interested in learning more. Using a short 
telephone-specific recruitment script (Appendix 5), study staff will inform the caregivers that 
a research study on a product that may improve pediatric gut health will be tested. If the 
caregiver is interested in learning more about the study, they will be invited to set up a 
screening visit (to be conducted at the CWC) where they will be provided with a product 
information form and study consent form to take home and review before returning to the 
clinic for an enrollment visit. If the caregiver is not interested in the study during the phone 
call and declines to schedule a screening visit, their information in the pre-screening log will 
be blotted out using dark ink. If the caregiver declines enrollment in the study at the screening 
visit, any medical record or contact data previously collected will be shredded and discarded 
or blotted out, as applicable. 
 
Following the visit, those who are still interested and eligible will be scheduled to undergo the 
informed consent process and enrollment visit at CGH at a time convenient to them, 1–3 weeks 
after the immunization or Vit. A supplementation visit, or whenever is convenient for those who 
had been identified from the CWC register (but did not receive immunizations or Vit. A 
supplementation). The delayed enrollment visit is to avoid conflating potential immunization-
related AEs with those related to the study product. Caregivers will receive a reminder call from 
study staff the day before their scheduled enrollment visit. During the appointment reminder call, 
caregivers will be advised of changes that would make them ineligible (to avoid an extra trip to 
the hospital). These changes include diarrhea in household in past week, child acute illness, and 
child antibiotic treatment within the past 2 weeks. If the child is no longer eligible based on one 
of these changes, the enrollment visit will be postponed and rescheduled or cancelled, depending 
on the caregiver’s preference. The caregiver will be told that they have the option to decline to 
participate at the time they are contacted by the study team or any time thereafter.  
Children and their caregivers will be recruited as a dual enrollment participant pair. Upon their return 
to the facility, the RA will escort the caregiver and child to a private exam room where they will review 
the consent form and answer questions they may have. The RA will confirm that both the child and 
caregiver still meet both sets of eligibility criteria. The RA will then obtain written informed consent 
from willing and eligible caregivers prior to the child medical examination, blood draw, IDI, and 
administration of AA-ORS or placebo solution.  
All participants will be informed that the study does not offer any direct benefit to them and the use 
of this product is currently for exploratory research purposes only. The study does not aim to assess 
the reliability or compare the outcomes of AA-ORS to the standard of care. The information 
participants will provide throughout this study is not considered sensitive and will not pose any 
significant risk to them personally. Participants have the option to decline being interviewed for the 
study without any negative consequences.  

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Once informed consent is obtained, participants will be assigned an identification number that will be 
used to identify the paper forms, solution consumption data, and blood specimens collected from that 
patient during the study. No personal identifying information will be recorded on any of the data 
collection tools.  
iv. Control or comparison group use  
Controls will be children from the same patient population as those receiving AA-ORS. These children 
will receive a placebo solution containing all the non-amino acid and non-electrolyte components of 
the AA-ORS and therefore will be able to contribute useful information regarding the feasible dose of 
the solution. This placebo solution is produced by the manufacturer of the AA-ORS for the purpose of 
clinical studies. Because these children will not be receiving the amino acids, they will provide the 
background rate of AEs (and morbidity experience) of children in this population, allowing for the 
detection of an unusually high event rate in the AA-ORS group, should it arise. 
v. Number and age range of research study participants/groups whose data shall be used 
Sixty-six children between the ages of 12 and 24 months and their caregivers will be enrolled into the 
study.  
vi. Inclusion criteria for each group of research study participants 
Inclusion Criteria  
Pediatric and caregiver pairs (must meet inclusion criteria for both categories): 
Child: 
1. Is between 12 and 24 months of age. 
2. LAZ between –1 and –3 SD. 
3. At least one week post routine immunization or vitamin A supplementation visit at the study site. 
4. Has a parent or legally acceptable representative willing and able to provide informed consent. 
5. No plans for travel outside of the community for the duration of the study. 
Caregiver of child: 
1. Is a parent or legally accepted representative of a child eligible for this study. 
2. Is 18 years of age or older. 
3. Has a working mobile phone. 
4. Is willing and able to provide informed consent. 
5. If illiterate—there is at least one literate adult living in the child’s household. 
vii. Exclusion criteria for each group 
Exclusion Criteria  
Pediatric and caregiver pairs (must meet none of the exclusion criteria for either category): 
Child: 
1. Has any sign of acute illness, including but not limited to fever, cough, and diarrhea. 
2. Is wasted (weight for length z-score < –2 or mid-upper arm circumference [MUAC] < 12.4 cm) or 
has pitting edema. 
3. Is exclusively breastfed. 

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4. Is seeking medical attention at the health facility other than for routine, preventative care (e.g., 
immunization visit, vitamin supplementation). 
5. Has suffered within the prior week from illnesses that might impact nutritional status (e.g., severe 
diarrhea or pneumonia; vomiting; persistent diarrhea; cleft lip or palate; blindness; tuberculosis; 
jaundice; renal or cardiac disease; cerebral palsy; known metabolic disorders; and chromosomal 
disorders, including trisomy 21). 
6. Medical history of chronic health condition (i.e., HIV, hepatitis B or C, end stage renal disease, 
severe liver disease—absence of a diagnosis is sufficient). 
7. Participating in any other clinical trials.  
8. Recent (prior 2 weeks) use of antibiotics or any other medical treatments (including ORT, but not 
including vaccines or vitamin/mineral supplementation). 
9. Cannot give the necessary biological (blood) sample. 
Caregiver: 
Reports diarrhea in the household in the prior 7 days. 
viii. Gender, race, or ethnicity use as variables in participant selection  
Not applicable. 
ix. Time period in which these data shall be collected and/or stored and time frame for entire 
study 
Data will be collected for this study over a 4-month period, and a permanent master set of data will 
be kept by PATH for at least 5 years post-completion of the funded project, up to indefinitely. All 
plasma/serum specimens collected at enrollment and post-dosing visits will be stored in Kenya, 
analyzed, and destroyed within 5 years. We have included language in the consent form to allow for 
potential future research with the samples and data related to nutrition or infectious diseases. 
x. Monitoring conduct of the study to ensure participants’ safety, confidentiality, and data 
integrity 
Study Monitoring  
PATH investigators will monitor study activities at the health care facilities where DO and IDIs are 
conducted. An adverse event case report form (CRF) will be used to record any unanticipated adverse 
events (Appendix 2). On on-site independent study monitor (ISM) will be contracted to oversee 
compliance with safety monitoring and stopping rules. The ISM will conduct safety monitoring visits 
every two weeks during the first month of the study, and at least once per month for each subsequent 
month of the study. During monitoring visits, the ISM will review safety data for participants, including 
all AEs and SAEs. The ISM will confirm the severity and relatedness assessment initially determined 
for all AEs, assess weight loss and breastfeeding practices in accordance with the study stopping rules, 
and confirm that all appropriate AE follow-up and SAE reporting procedures have been followed. 
PATH will also regularly review study records and logs for quality assurance purposes. 
Study participant confidentiality will be respected. Site monitoring visits may be conducted to: 
 
Verify compliance with human subjects and other research regulations and guidelines, including 
confidentiality procedures, informed consent process, and regulatory documentation.  
 
Assess adherence to the study protocol and study-specific procedures manual. 

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 
Confirm the quality and accuracy of information collected at the study site and entered into the 
study database, including the validation of data reported on data collection forms.  
 
Inspect the study facility and documentation (e.g., informed consent forms, data collection forms) 
as well as observe the performance of study procedures. If a monitor is present during consent, 
participants will be asked their permission to accept or decline the monitor observing.  
To minimize protocol deviations, study staff will be thoroughly trained on study conduct, safety, and 
careful data management. In the event of a protocol deviation, the event will be documented in the 
corresponding protocol deviation CRF and the study team will notify the co-PIs within 24 hours. 
Protocol deviations will be reported to the appropriate ethics committees in accordance with their 
reporting requirements.  
Study procedures will begin only after the study has received necessary ethical and regulatory 
approval(s) from relevant parties and written informed consent from the participants.  
The Pharmacy and Poisons Board (PPB) may also inspect the clinical trial site and trial sponsor to 
ensure that the principles of GCP are being met. In order to demonstrate compliance with the protocol 
and applicable regulations, the PI and site PI will maintain a Clinical Trial Master File containing 
relevant documentation to allow effective supervision. At the request of a properly authorized officer 
of PPB, the site PI shall permit such officer to have access to, copy, and verify any records or reports 
made by the investigator. The site PI will notify the sponsor within 24 hours following contact by a 
regulatory authority. The sponsor will provide any needed assistance in responding to regulatory 
audits or correspondence. 
Safety Monitoring 
The following procedures/evaluations will be conducted as part of the study safety assessment: 
 
Medical examination at enrollment and day 15 of follow-up: assessment of height, weight, MUAC, 
vital signs (i.e., temperature, pulse, respiration rate), organ systems, breathing, hydration status, 
fontanelles, and skin irritation/rashes. 
 
Counseling procedures that need to be adhered to during study participation. 
 
Assessment of study intervention adherence via measurement of residual solution will be 
performed on each bottle (numbered 1–21).  
 
Assessment of adverse events including solicited AEs (i.e., vomiting, diarrhea, fever, constipation, 
irritability, allergic reactions, etc.) and unsolicited AEs by study nurse initially in person, and 
subsequently by phone (days 4, 7, 14, 21), and review of any AEs/SAEs by study clinician.  
 
Assessment of child weight by the RA, who will weigh the child at interim visits on day 7 and 14. 
To screen children for recent illness or chronic illness (e.g. HIV, hepatitis, etc.), medical charts created 
as part of regular medical care will be reviewed, as available. Confidentiality of medical records will be 
protected. 
Adverse Events and Serious Adverse Events 
All participants will be monitored closely for adverse events during the study at five time points by the 
study nurse. The nurse will do these assessments 60 minutes after the first administration (in person) 
and via phone questionnaire on days 4, 7, 14, and 21. All clinical personnel involved in the study will 
be trained in identifying adverse events and all participants will be educated regarding symptoms that 
require urgent medical attention. Any adverse events will be managed by the appropriate clinic site, 
and if necessary, hospitalization. The costs of care directly attributable to participation in the study 

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will be borne by the study sponsor. The clinical site involved in this study is a Provincial Hospital, which 
can provide emergency care if necessary. 
 
Definition of Adverse Event (AE) 
An adverse event (AE) is any untoward medical occurrence in a participant after administration of the 
study product and that does not necessarily have a causal relationship with the study product. An AE 
can therefore be any unfavorable and unintended signs (including abnormal laboratory findings), 
symptoms, physical examinations, or disease temporally associated with the use of the study product, 
whether related to the study product or not. This definition includes exacerbations of pre-existing 
conditions. Stable pre-existing conditions that do not change in nature or severity during the study are 
not considered AEs; however, these should be reported as part of the medical history. 
 
Solicited AEs are pre-specified adverse events that are actively monitored by study staff during the 
full 21 days of follow-up. Personal communication with the study product manufacturer indicated that 
no adverse events have been found to be associated with the AA-ORS in adult studies or in the small 
study (n=5) featuring children with short bowel syndrome. 
 
For this study, solicited AEs will be assessed by study nurse 30 minutes after the first administration 
(in person), and then by phone on days 4, 7, 14, and 21.  
 
The following specific solicited adverse events will be monitored for this trial: 
 
 
 
Vomiting. 
 
Diarrhea. 
 
Fever. 
 
Constipation. 
 
Lethargy. 
 
Irritability/restlessness. 
 
Allergic reactions: sudden swelling of the face or throat, itching, and rash. 
 
Other reactions: fits (convulsions/seizures); gas, abdominal cramps, or stomach pain; weight loss. 
Changes in the severity of an AE (e.g., diarrhea) will be documented to allow an assessment of the 
duration of the event at each level of severity to be performed. AEs characterized as intermittent 
require documentation of onset and duration of each episode.  
 
Stopping rule: RAs will weigh children in the household on a weekly basis (end of weeks 1 and 2). If a 
child loses more than 5% of his or her body weight from that measured at enrollment, he/she will be 
withdrawn from the study. The study team will refer the withdrawn child and caregiver to his or her 
primary health care provider or CGH, as appropriate. In addition, there will be a review of the first 10 
enrolled breastfeeding participants by the investigators and ISM. If two of the first 10 participants who 
are breastfeeding at enrollment stop breastfeeding during the 2-week course of the study, the study 
will halt the enrollment of children who are currently breastfeeding. If there is evidence of a negative 
impact on participants’ breastfeeding, PATH REC will be notified. 
 
Unsolicited AEs are any significant events not specified as a solicited AE by study personnel and can 
be reported spontaneously by the participant, observed by the study personnel during study visits, or 
identified during review of medical records or source documents. 

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Any medical condition that is present at the time that the participant is screened will be considered 
as baseline and not reported as an AE. However, if the study participant’s condition deteriorates at 
any time during the study, it will be recorded as an AE.  
 
Changes in the severity of an AE will be documented to allow an assessment of the duration of the 
event at each level of severity to be performed. AEs characterized as intermittent require 
documentation of onset and duration of each episode. 
 
Serious Adverse Event (SAE)  
A serious adverse event (SAE) is any adverse event that results in any of the following outcomes: 
1. Death. 
2. Is life-threatening (life-threatening means that the study participant was, in the opinion of the site 
PI or sponsor, at risk of death at the time of the event; it does not refer to an event that 
hypothetically might have caused death if it were more severe).  
3. Requires inpatient hospitalization or prolongation of existing hospitalization.  
4. Results in persistent or significant incapacity or substantial disruption of the ability to conduct 
normal life functions.  
5. Is an important medical event that may not result in one of the above outcomes but may 
jeopardize the health of the study participant or require medical or surgical intervention to 
prevent one of the outcomes listed in the above definition of serious adverse event.  
Suspected Unexpected Serious Adverse Reactions (SUSARs) 
A suspected unexpected serious adverse reaction (SUSAR) is a serious adverse reaction that is not 
identified in practice, severity, or frequency by the reference safety information. A SUSAR must meet 
the criteria for an SAE and also be determined to be related to the study product.  
Reporting Period and Parameter 
Safety events are reported from the time of the first study product administration through completion 
of the study 21 days later. Specifically, solicited AEs will be collected up to 30 minutes after the first 
administration and on days 4, 7, 14, and 21. Both solicited and unsolicited AEs and SAEs will be 
assessed through 7 days after the end of the study product dosing. The final AE and SAE assessment 
will be done via a phone call on day 21 (or within 3 days thereafter): 
 
1. The caregiver will be contacted via telephone and identity confirmed.  
2. Study nurse will ask about any medical event that would constitute an AE or SAE since the last 
visit.  
3. If an AE or SAE is reported, the nurse should record it on the appropriate CRF and consult the study 
clinician, who will notify the entities who require notification. The nurse will refer the participant 
for treatment, if warranted.  
After recording the information, the participant will be discharged from the study.  
 
Severity of Adverse Events 
The severity of all AEs will be assessed by the study clinician based on the following guidelines.  
• 
Mild—Events require minimal or no treatment and do not interfere with the participant’s daily 
activities.  

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• 
Moderate—Events result in a low level of inconvenience or concern. Moderate events may cause 
some interference with functioning and may require therapeutic measures. 
• 
Severe—Events that prevent a participant’s usual daily activity and require systemic drug therapy 
or other treatment. Of note, the term “severe” does not necessarily equate to “serious.” 
• 
Potentially Life-Threatening—Events that require emergency room visit or hospitalization. 
Potentially life-threatening symptoms causing incapacitation with intervention needed to prevent 
disability or death. 
 
A toxicity table for solicited AEs with guides for assessing severity of the specific event is found in 
Appendix 8. 
 
Causality of Adverse Event  
The study investigator/s will determine the causal relationship between the study product and the AE. 
The causality assessment is made on the basis of the available information at the time of reporting 
and can be subsequently changed according to follow-up information. Determining of causality is 
based on clinical judgment and should take into considerations the following factors: 
 
Is there a temporal (time-based) relationship between the event and administration of the study 
product? 
 
Is there a plausible biological mechanism for the study product to cause the AE? 
 
Is there a possible alternative etiology for the AE such as concurrent illness or concomitant 
medications? 
 
Are there previous reports of similar AEs associated with the study product? 
For this study, the investigator/s must classify the causality of the AE according to the categories 
defined below:  
Related: There is a reasonable possibility that the product caused the event. “Reasonable possibility” 
means that there is evidence to suggest a causal relationship between the study product and the AE. 
Not Related: There is not a reasonable possibility that the administration of the study product caused 
the event. 
Follow-up of Adverse Event 
All reported AEs should be followed until resolution or stabilization, or until the participant’s 
participation in the study ends. Participants who have an ongoing study product–related SAEs at study 
completion or at discontinuation from the study will be followed by the PI or his designee until the 
event is resolved or determined to be irreversible, chronic, or stable by the PI. 
The outcome of an adverse event will be assessed as at the time of last observation as per the 
following categories:  
 
Recovered/resolved without sequelae.  
 
Recovered/resolved with sequelae. 
 
Ongoing at the end of the study. 
 
Death. 
 
Unknown. The outcome of the AE is not known. 
Reporting of SAEs and SUSARs 
 

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STUDY PEDIATRICIAN REPORTING TO INVESTIGATORS 
Within 24 hours of the study pediatrician’s awareness of an SAE as defined in the protocol, an AE/SAE 
form must be completed and submitted to the study’s PIs by email. The ISM will also be notified. 
The initial AE/SAE form should be completed with all information known at the time and should 
include as much as possible the following: 
 
Name and contact of the investigator submitting the AE/SAE report. 
 
Participant ID number.  
 
Participant age. 
 
Date participant received study drug, including cohort group.  
 
Description of the serious adverse event.  
 
Date of event onset and date of event resolution. 
• 
Investigator’s assessment of severity, causality, and expectedness.  
• 
Action taken and current status. 
• 
If available, any diagnostic test reports or hospital records that may help the sponsor to evaluate 
the SAE.  
The site PI will be responsible for notifying the Maseno University Ethics and Research Committee and 
the PPB, and the PI will be responsible for notifying the PATH research ethics committee (REC) as per 
the respective ethical or regulatory authority’s reporting requirements.  
 
Notification and Review of SAEs 
The site PI is responsible for evaluating SAEs submitted by the study pediatrician within 24 hours to 
convene a safety review if the investigator reported the SAE as fatal or life-threatening and suspected 
to be related to study drug. Medical officer(s) from PATH serving as technical consultants will provide 
technical guidance regarding SAE management including classification and reporting.  
Sponsor Reporting to Regulatory Agency 
The sponsor has authorized the site PI to execute its responsibility for safety reporting to the 
appropriate regulatory authorities within specified time period of notification.  
The site PI will be responsible for reporting to the Kenya MOH PPB, in accordance with applicable 
regulatory requirements.  
• 
All fatal and life-threatening, unexpected adverse study product reactions (i.e., SUSARs) should be 
reported within 7 calendar days after first knowledge by the applicant. The initial notification must 
be followed by as complete a report as possible, within an additional 8 calendar days.  
• 
SUSARs that are not fatal or life-threatening must be reported as soon as possible and not later 
than 15 calendar days after first knowledge by the applicant. 
• 
PPB must be notified, within 15 calendar days after first knowledge by the applicant, when there 
is a suggestion of a change in the nature, severity, or frequency of expected adverse drug reactions 
or when new risk factors are identified. The basis on which these assessments are made should 
be included.  
• 
Any information that may in any way influence the benefit-risk assessment of a medicine or that 
would be sufficient to consider changes in the administration of the medicine or in the overall 
conduct of a clinical trial must be reported to PPB.  

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• 
A safety report will be submitted to PPB once a year throughout the clinical trial, or on request by 
PPB, that takes into account all new available safety information received during the reporting 
period. The aim of the annual safety report is to describe concisely all new safety information 
relevant for the clinical trial and to assess the safety conditions of subjects enrolled. The safety 
report shall include a summary of all SAEs and SUSARs. 
Safety Oversight 
The site PI and designated study staff will be responsible for safety monitoring of all study participants 
and for alerting the PI if unexpected concerns arise. This study will also have a Data and Safety 
Monitoring Board (DSMB), composed of two pediatric gastroenterologists (one as chair) and a 
biostatistician. The DSMB is responsible for safeguarding the interests of study participants, assessing 
the safety of study procedures, and for monitoring the overall conduct of the study. The DSMB is an 
independent group advisory to the study investigators. The responsibilities are outlined in detail in the 
DSMB Charter, which will be approved by the DSMB prior to the study’s start. 
DSMB meetings will be held remotely. The purpose of the first meeting held before the study begins 
recruitment is to review and discuss the Charter, to review the protocol and informed consent form, 
and to confirm the report contents and format for data reports during interim and final DSMB 
meetings. Enrollment in the study will not begin until the DSMB’s Charter has been accepted by the 
DSMB and IRB/regulatory approval has been obtained. The DSMB is expected to meet twice more 
over the course of the study. One meeting will be held to review interim data when approximately 
half of the participants have been enrolled or two months have elapsed after the beginning of 
recruitment, whichever is earlier. The final meeting will be held at the end of data collection, once 
topline data is available. Additional meetings or conference calls will be scheduled as needed. At the 
interim meeting, the DSMB will also review formal interim analyses of the primary (safety) endpoint.  
There are no a priori reasons for a particular safety concern or the possibility of serious toxicity with 
the study product. As a small, single-site study for a short duration, the trial pediatrician will be able 
to have close supervision and monitoring of all AEs. Additionally, an ISM will be contracted to oversee 
compliance with safety monitoring and stopping rules (e.g., weight loss ≥ 5%, cessation of 
breastfeeding by 2 of the first 10 breastfeeding participants). 
xi. Data management and analysis plan 
Data Management Procedures 
IDI data will be analyzed qualitatively, using transcripts and notes to identify key themes and topics.  
DOs will be manually coded and analyzed to identify the time taken to drink up to 237 ml of AA-ORS, 
ease of administration, and visible signs of palatability.  
Quantitative clinical data focused on demographics, anthropometrics, AEs, product volume consumed 
daily, plasma/serum micronutrients, metabolic markers (acylcarnitine profile), EED markers (gut 
dysfunction and growth hormone resistance), and systemic inflammation will be collected on 
standardized forms or exported to computerized databases by RAs and laboratory workers. Form data 
will be double entered into a password-protected Microsoft Access database by RAs blinded to the 
supplementation assignment (AA-ORS vs. placebo). After entering all data and resolving 
inconsistencies, the data set will be locked.  
All data will be stored on password-protected computers in PATH offices. Only members of the study 
team will have access to the raw transcripts, CRFs, and consent forms in a locked filing cabinet.  

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Data Analysis Procedures 
DEFINITIONS OF POPULATIONS TO BE ANALYZED 
The intention-to-treat population will include all participants who are assigned AA-ORS or placebo 
regimen and receive at least one dose, regardless of whether he/she completes the full 14-day 
regimen (or misses dose[s] of study product for a medical reason such as vomiting). The first primary 
(safety), secondary (caregiver acceptability), and exploratory (biomarker) analyses will be performed 
using this population.  
The per-protocol population will include all participants who are assigned a regimen with no major 
protocol violations that are determined to potentially interfere with the ad-libitum dosing of the study 
solution(s) (i.e., early withdrawal or product discontinuation, etc.). This population will serve as the 
primary analysis population for the volume consumption and consumption factor endpoint. As the 
primary objective is to accurately estimate the average daily volume consumed in the full sample (AA-
ORS + placebo), it is critical that all subjects in the population have completed a full dosing regimen.  
STUDY ENDPOINTS TO BE ANALYZED AND ANALYTICAL METHODOLOGY 
Primary: AEs and SAEs through 21 days of follow-up, and changes to dietary and digestive habits 
through day 14 of follow-up. 
For AEs and SAEs that occur within 21 days of enrollment, statistical comparisons will be made between 
the AA-ORS and the placebo solution groups using Fisher’s exact test. Changes to feeding habits 
(normal, less frequent/less volume, more frequent/more volume), diarrhea (1–2, 3–4, 5–7, or 7+ events 
in 24 hours), and vomiting (none, 1, 2–5, > 6 events in 24 hours, or hospitalization) will be made using 
chi-square tests. 
Primary: Average daily product volume consumed through 14 days of follow-up, time trends in daily 
volume consumed, and factors that influence children’s willingness and ability to ingest the product 
as intended.  
The average daily volume (and statistical distribution) of solution consumed by children over the 2-
week period will be calculated. These descriptive data will be pooled from AA-ORS and placebo arms 
as the solutions are designed to taste the same. Lowess curves will be used to describe trends in 
volumes consumed, and linear mixed effects models will be used to test for an association between 
day of follow-up and product volume consumed. We will use linear regression to identify factors (e.g., 
child weight, length, and age) associated with a child’s average daily intake. 
Secondary: Key factors that influence caregivers’ willingness to have the product used by their 
children, and their perception of their children’s response to its consumption. 
These qualitative analyses will use ATLAS.ti software to identify and group key themes and topics from 
IDI notes. 
Exploratory: Average plasma/serum concentrations of metabolic, gut dysfunction, systemic 
inflammation, and micronutrient biomarkers at baseline and day 15 of follow-up. 
Metabolic, EED, systemic inflammation, and micronutrient marker data collected at baseline will be 
pooled across intervention groups and used to generate descriptive statistics for the study population. 
The same descriptive statistics will be generated for day 15 of follow-up. Paired student’s t test will be 
used to compare these continuous parameters between baseline and day 15 of follow-up.  

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Unpaired student’s t test will be used to compare these continuous parameters between the two study 
groups at day 22. A difference with P < 0.05 will be considered statistically significant. However, there 
is limited statistical power to make such comparisons between study groups for biomarker outcomes.  
ISSUES WITH MISSING DATA AND MULTIPLICITY 
We will analyze whether there is differential missingness or loss to follow-up by supplementation arm 
for the primary endpoints. No imputation of missing outcome data is planned. We will adjust the 
significance level for multiple comparisons by using the Benjamini-Hochberg method to control for 
the false discovery rate for related hypotheses. 
Quality Assurance  
The study will be conducted in full compliance with the protocol and ICH GCP to provide public 
assurance that the rights, safety, and well-being of trial participants are protected and that the clinical 
trial data are credible. To ensure quality and standardization, the site will develop standard operating 
procedures (SOPs) for key protocol procedures and conduct the study guided by the study Manual of 
Procedures or other written guidelines. The site will also develop routine operational checks to verify 
that critical protocol requirements and procedures are executed correctly and completely at the time 
the work is being performed. Prior to the initiation of the study, the sponsor will conduct training on 
the protocol, including applicable SOPs, for study staff. 
Clinical and laboratory data for each participant will be abstracted from study CRFs into standardized 
project files. These data will be entered directly into computers at a centralized site. Information will 
be cross-checked for accuracy on a biweekly basis. All data, both hard- and soft-copy formats, will be 
stored in locked cabinets initially at CGH and will be transported to the PATH office in Kisumu after 
data entry with limited access by project staff only.  
Participants will be identified using a unique Patient Identification Number (PID). The code linking the 
PID to individual identifying information will be kept in a separate secure location by the project 
coordinator. Clinical staff at CGH will not be able to access identifying information linked to the PID, 
except to match laboratory results reported by the lab to individual patients. 
xii. Study limitations and the expected results 
Study Limitations 
This is an exploratory study with a small sample size that is not adequately powered for statistical 
comparisons of the EED outcomes (non-invasive biomarkers). As a result, the findings of this study 
may not be widely generalizable. This limitation will be mitigated during presentation of the study 
findings, which will avoid drawing conclusions beyond the scope of the study methods. The expected 
results from the study are intended to be useful for establishing a safety profile and reasonable 
pediatric dose and providing study design considerations (e.g., child EED biomarker data) for future 
research on the efficacy of AA-ORS and the deployment of similar EED interventions in LRS. 
xiii. Itemized budget and budget justification 
Budget 
Category
Amount (USD)
PERSONNEL
[redacted]

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TRAVEL
[redacted]
EQUIPMENT & SUPPLIES
[redacted]
WORKSHOPS & TRAINING 
[redacted]
PARTICIPANT SUPPORT COSTS
[redacted]
OTHER PROJECT COSTS
[redacted]
Overhead
[redacted]
Total
[redacted]
 
Budget justification  
The largest budget item for this study is personnel costs, totaling USD [redacted], which includes 
salaries and benefits for the study PIs and co-investigators as well as the research assistants, clinical 
staff, and lab personnel required to implement a quality study. Travel costs of USD [redacted] include 
international travel for non-Kenyan investigators to attend study trainings and site visits as well as 
domestic transportation for study staff to travel to the study site. Equipment and supply expenses 
include paper and printing costs for study materials, biomarker assays, and acylcarnitine mass-
spectrometry, totaling USD [redacted]. Workshops and trainings are budgeted to cost USD [redacted] 
and include the initial training of all study staff on the protocol and consenting process to be followed 
for this study. A total of USD [redacted] is budgeted for participant support costs to pay individuals 
according to the compensation scheme outlined in the informed consent document. Other project 
costs of USD [redacted] include fees for ethical and regulatory submissions, shipment and 
procurement of study product and placebo, legal and contracting support, among other miscellaneous 
expenses. 
xiv. Research study data collection forms and other references 
Appendix 1 – Informed Consent Form  
Appendix 2 – Case Report Forms (CRF) 
Appendix 3 – Patient Information Leaflet  
Appendix 4 – Investigator’s Brochure 
Appendix 5 – Recruitment Scripts for Caregivers 
Appendix 6 – Patient Instruction Sheet 
Appendix 7 – Letter of Support From Robert Bandsma 
Appendix 8 – Toxicity Table 
 
 

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D. ETHICAL CONSIDERATIONS  
This study will be conducted under the auspices of the following ethics committees (ECs):  
 
PATH Research and Ethics Committee (REC) 
2201 Westlake Avenue, Suite 200 
Seattle, WA 98121 
USA 
rec@path.org 
+1 206 285 3500 
 
Maseno University Ethics Review Committee (MUERC) 
Directorate of Research, Publications and Innovations 
Maseno University Main Campus 
Along Kisumu-Busia Road 
P.O. Box, Private Bag 
Maseno, Kenya 
muerc-secretariate@maseno.ac.ke 
+254 57 351 622 Ext. 3050 
 
The PI and site PI will be responsible for obtaining approval from the ECs listed above. These 
committees will review and approve the protocol, informed consent form, and any recruitment 
materials (advertising or informational material), including any modifications to these documents 
prior to or during the study. All changes to the protocol or consent form must be reviewed and 
approved prior to implementation, except where necessary to eliminate apparent immediate hazard 
to study participants. The PI and site PI also will be responsible for obtaining continuing review 
throughout the duration of the study in accordance with existing regulations. 
i. Risks to research study participants/groups 
Risk Considerations 
The proposed study is expected to be of minimal risk as the study AA-ORS is a medical food product 
regulated by the US FDA. As a result, we do not anticipate any adverse events for this study related to 
the administration of AA-ORS, but we will include a control arm in order to better assess the safety of 
this product in children with mild-moderate stunting. Children with mild to moderate stunting are at 
increased risk for morbidity and mortality, so we anticipate that participants may report diarrhea or 
respiratory infections at high rates, regardless of receipt of active AA-ORS or placebo. While there is 
mounting evidence that microbiome immaturity contributes to growth deficits in LRS, any changes to 
the microbiome composition and diversity from provision of AA-ORS are likely to be transient, based 
upon previous observations among infants and children provided probiotic-supplemented formula or 
ready-to-use supplementary food (43, 44).  
This study involves blood specimen collection. The collection of these samples involves venipuncture, 
which may cause discomfort, pain, introduction of infection, bleeding, fainting, or bruising. 
Precautions will be taken to avoid introduction of infection by disinfecting the site of venipuncture 
and using sterile equipment. The risk of bleeding and bruising will be minimized by immediate 
application of pressure after venipuncture. The child will be restrained appropriately by the caregiver 
to prevent kicking of the phlebotomist. A thorough description of potential risk considerations and 
mitigation strategies is outlined in Table 7. 

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Table 7. Known risks and mitigation strategies for SAFE study participants. 
Potential risk to 
participant 
Concern 
Study design feature 
intended to minimize 
risk 
Rationale for the 
necessity of exposing 
participants to risk 
Reduction in breastmilk 
consumption among 
those breastfeeding at 
enrollment.  
Consumption of solution 
(AA-ORS or placebo) 
could reduce amount of 
breastmilk consumed by 
child. 
Exclusion of exclusively 
breastfed children. 
Caregivers instructed to 
provide breastmilk as 
needed during the 3-
hour dosing period. 
Monitoring for 
decreased frequency of 
breastfeeding and for 
unplanned (by caregiver) 
weaning. Will stop the 
study if it is clear there is 
a major reduction in 
breastfeeding from the 
study drink. 
Ingestion required for 
product delivery. 
Overall caloric content 
of AA-ORS unlikely to 
satiate children given 
daily intake 
requirements. 
Use in pediatric 
populations who are 
acutely ill. 
Children experiencing 
mild-moderate stunting 
are at increased risk for 
morbidity and mortality 
and may develop 
intercurrent illnesses 
during the course of the 
study. 
Will recruit children with 
mild to moderate 
stunting (length-for-age 
z-score between –1 and 
–3), but not acute illness 
at enrollment. 
Safety monitoring 
procedures in place to 
identify and treat 
adverse events. 
Children with stunting, 
with or without acute 
illness, overlap 
considerably with the 
target population for an 
EED intervention. 
No safety data in 
children. 
Prior safety data on the 
study AA-ORS is from 
adult populations and 
may not apply to 
pediatric subjects. 
Will directly observe the 
first dose of the product 
in the clinic to identify 
any acute reaction. 
Will treat all participants 
who experience adverse 
events during follow-up. 
Study is needed to 
establish the safety 
profile of the product.  
May also assess whether 
children’s metabolism 
switches to oxidation. 
Spoilage/contamination. 
Open AA-ORS bottles 
could pose a health risk 
if consumed at a later 
time, due to 
contamination. 
Mark and reseal bottles 
using tape and markers 
provided at enrollment. 
Research assistant (RA) 
to visit home on second 
and fourth day of follow-
up to instruct and advise 
caregiver on safe 
storage and proper 
resealing of bottles after 
maximum of 3 hours. 
Need to measure and 
calculate the average 
daily volume consumed 
during follow-up in 
order to guide dosing 
recommendations in 
future clinical trial.  

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Potential risk to 
participant 
Concern 
Study design feature 
intended to minimize 
risk 
Rationale for the 
necessity of exposing 
participants to risk 
RAs to pick up partial 
and empty bottles 1–2 
times per week. 
Volume of solution too 
great. 
237 ml may be too much 
volume for children. 
Children to drink as 
much or as little of the 
solution (up to 237 ml) 
as desired over 3 hours. 
Gavage will not be done. 
Breastmilk will be 
permitted throughout 
this period. 
Need to measure and 
calculate the average 
daily volume consumed 
during follow-up in 
order to guide dosing 
recommendations in 
future clinical trial. 
Blood draw in children. 
Potential discomfort, 
bleeding, bruising, and 
minimal risk of infection. 
Precautions will be taken 
to avoid introduction of 
infection by disinfecting 
the site of venipuncture 
and using sterile 
equipment. The risk of 
bleeding and bruising 
will be minimized by 
immediate application of 
pressure after 
venipuncture. 
Only trained, 
experienced 
phlebotomist will 
perform venipunctures.  
Blood samples are 
needed in order to 
measure metabolic 
profile and biomarkers 
of gut dysfunction, 
systemic inflammation, 
and micronutrient 
status.  
Metabolic profile 
assessment will be an 
important part of 
exploratory analysis; it is 
helpful to determine if 
the absence of certain 
essential amino acids 
leads to increase in 
oxidative pathways. 
These biomarker 
measurements are 
important in order to 
have setting-specific 
descriptive data with 
which to power a future 
EED efficacy trial.  
A comparison of these 
non-invasive biomarkers 
following the 
intervention will also be 
done. 
Loss of confidentiality. 
Medical information 
from a participant’s 
medical record or other 
Access to study records 
will be limited to study 
staff with electronic 
passwords (data files) or 
Review of medical 
records will help ensure 
that only eligible 

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Potential risk to 
participant 
Concern 
Study design feature 
intended to minimize 
risk 
Rationale for the 
necessity of exposing 
participants to risk 
study documents are 
shared inappropriately. 
lock and key (physical 
records). 
Study staff will only 
review medical records 
for eligibility assessment 
and will not make copies 
or remove such records 
from the hospital 
premises. 
If the caregiver declines 
enrollment in the study, 
any medical record data 
collected will be 
discarded. 
participants are enrolled 
in the study. 
Detailed study records 
are critical for data 
analysis and reporting 
results accurately. 
 
The information participants will provide in the context of this study is not considered sensitive and 
will not pose any significant risk to them personally. There is a minimal risk of loss of confidentiality. 
All study staff will be trained in the protection of human subjects, including maintaining participant 
confidentiality. Study procedures will begin only after the appropriate ethics committees grant 
approval. 
As the sponsor of this study, PATH will secure clinical trials insurance, which will cover the costs 
associated with any study-related injuries. 
Benefits 
No direct benefit is anticipated to participants for their inclusion and participation in this study. 
Information collected during this study may benefit the participants’ community in the future for 
potential EED interventions administered in an aqueous solution, including but not limited to the study 
AA-ORS. 
Rights of the Study Participants 
Provisions will be made to protect the participants in this study. Confidentiality, anonymity, and 
storage of data and specimens will be explained to all participants or their caregivers (i.e., parent or 
legally acceptable representative). The participants will be informed that they may withdraw from this 
study at any time with no negative repercussions for themselves or the child participant. All data 
collection tools and biological samples will be identified only with the individual study participant 
number and study title. It will be made clear that only the study team will have access to the data that 
emerges from this study and the data will be only used for research purposes.  
ii. Access to protected health information required by proposed research study 
The study will have access to protected health information. The use of patient medical records will be 
necessary to help ensure that only eligible participants are enrolled in the study (e.g., confirm the child 
does not have chronic health conditions such as HIV, hepatitis B, or hepatitis C). 

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iii. Sources of data, specimens, and records to be used in the proposed research study  
Data Collection Procedures 
Data collection tools (e.g., IDI guides and questionnaires) will be administered by study staff in the 
language of the caregiver’s preference. Participants will be allowed to ask questions in order to obtain 
clarification on study procedures.  
Clinical information  
Adverse events will be assessed via clinician-directed physical examination (screening, enrollment 
visit, and day 15 visit) and signs and symptoms reported by participants (days 1, 4, 7, and 14). Data 
collected at interim visits will use a structured questionnaire to record interval medical history. All 
clinical information collected as part of this study will be stored on password-protected computers in 
PATH offices. 
IDIs: 
Interviews will be conducted by an RA trained in qualitative methods. During the IDIs, the interviewer 
will guide the flow of the discussion. Any other member of the IDI team present will take note of topics 
that generate more animated discussions, participants’ attitude, body language, and interactions 
among the interviewer and interviewee. Notes will be documented in an unstructured format and 
saved as part of the participant study record. Discussions will be guided by a semi-structured data 
collection guide (Appendix 2), with responses hand-written contemporaneously by the interviewer.  
The interview guides will be a mix of structured and semi-structured questions. Structured questions 
will be necessary for standardizing responses to particular questions. Semi-structured questions will 
allow for flexibility and richness in responses, especially those on perceptions of facilitators and 
barriers for AA-ORS uptake. Semi-structured questions are open-ended and will allow for probing a 
participant’s answers for in-depth responses. 
Post-dosing IDIs with caregivers will be conducted after use of study product with their child in a 
private setting (Appendix 2). Caregiver interviews will last between 45 to 60 minutes at enrollment 
and post-dosing visits.  
DO of product administration: 
DO will be conducted with caregivers and their child during the very first study product administration 
at the enrollment visit. AA-ORS or placebo solution will be provided to participants by study staff. The 
DO will last up to three hours, depending on how quickly the child drinks the product. The researcher 
will take notes contemporaneously during the DO.  
Blood samples: 
All samples collected as part of this study will be stored in freezers at Kakamega CGH. Samples may be 
shipped to Kilifi and/or Nairobi for analysis at laboratories operated by Kenya Medical Research 
Institute (KEMRI). 
iv. Maintaining confidentiality of research study data and protecting the participant privacy 
All participants will be assigned unique identification numbers in place of their names to maintain 
confidentiality. All data will be stored on password-protected computers in PATH offices. Only 
members of the study team will have access to the raw transcripts and consent forms in a locked filing 
cabinet.  

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v. Storage of data and specimens being collected 
All data will be stored on password-protected computers in PATH offices in Kenya and the United 
States. All plasma/serum specimens will be labeled with alphanumeric patient and sample 
identification numbers and stored in a freezer at Kakamega CGH until shipment. Specimens will be 
prepared for shipment on dry ice in compliance with recognized standards for air transport and will 
be transported to KEMRI labs in Nairobi and/or Kilifi, Kenya by a third-party shipper with experience 
with biological specimens. Once the samples have arrived at the KEMRI laboratory, they will be 
unloaded into a freezer (temperature ≤20° C), before being assessed using the immunoassays 
described in Table 5.  Dried bloodspots will be shipped to KEMRI or another third-party laboratory in 
Kenya with the necessary mass spectrometry equipment to run metabolomic analyses. All 
plasma/serum specimens collected will analyzed for the study, and remaining specimens will be 
destroyed within 5 years. We have included language in the consent form to allow for potential future 
research with the samples and data related to nutrition or infectious diseases. 
vi. Duration of data storage, ensuring data security, and custodian and staff with data access 
A permanent master set of data will be kept by PATH at least 5 years post-completion of the funded 
project, but may be stored indefinitely. All data will be stored on password-protected computers in 
PATH offices. Only members of the study team will have access to the raw transcripts and consent 
forms in a locked filing cabinet. De-identified patient data will be shared with the manufacturer of the 
product.  
vii. Plan for controlling access to stored research study data 
All data will be stored on password-protected computers in PATH offices. De-identified patient data 
will be shared in an electronic format with the manufacturers of the AA-ORS. Explicit permission from 
PATH is required before the manufacturers can license the de-identified study data to a third party. 
viii. Viewing of identifiable research study data by persons unlisted as applicant or study team 
member 
Research assistants, study nurse, and study pediatrician will be contracted to conduct the IDIs and 
other study procedures as outlined above. These individuals shall collect and or view research study 
data with identifiers during enrollment and data collection activities in order to carry out visits and 
project activities. The RAs will not have access to the identifiable data once it has been entered into 
an electronic format and data cleaning and analysis is complete. 
ix. Signing of agreement(s) by research study team members with access to confidential data 
collected 
All study staff will have signed agreements protecting confidentiality of research data prior to 
conducting any study activities. 
x. Potential benefits of research study to community and/or society 
Benefits 
No direct benefit is anticipated to participants for their inclusion and participation in this study. 
Information collected during this study may benefit the participants’ community in the future for 
potential EED interventions. 

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xi. Consent issues: how consent will be obtained 
Consent Process 
Informed consent is the process of ensuring that potential participants fully understand what will and 
may happen to them while participating in a research study. An information sheet describing the study 
product (Appendix 3) will be provided to participants before the consent process begins. All consent 
materials, including any non-English translations, will be approved by the PATH REC and MUERC prior 
to use.  
 
In the region where this study is conducted, Luhya is the predominant ethnic group. However, there 
is no single Luhya language and there are over 10 different Luhya dialects spoken and mutually 
understood. Kiswahili is commonly spoken and understood as well, so documents translated into the 
local language will be translated into Kiswahili. Study staff will be conversant in local dialects in 
addition to Kiswahili to help ensure adequate comprehension of the consent materials. 
 
All study staff will be trained and certified in the protection of human subjects. In obtaining and 
documenting informed consent, the study staff will comply with applicable local and domestic 
regulatory requirements and will adhere to GCP and to the ethical principles that have their origin in 
the Declaration of Helsinki.  
 
Before an individual is enrolled in this study, it is the site investigators’ responsibility to ensure that 
informed consent is obtained from the participant after adequate explanation of the aims, methods, 
and potential risks and benefits of the study. The consent form will be reviewed and discussed with 
potential participants in a private space where others cannot easily overhear the conversation. The 
informed consent process will give individuals all the relevant information they need to decide 
whether to participate in this study. Potential research participants will be permitted to ask questions 
and to exchange information freely with the study team prior to providing written consent. We will 
not restrict enrollment to literate individuals, as we intend to recruit patients with caregivers from a 
variety of education levels. However, the requirement for caregivers to own a cellphone is closely 
associated with basic literacy. In the case of an illiterate potential participant, an independent witness 
must be present for the entire consenting process and will have to sign and date the informed consent 
form. In addition, the illiterate participant will also put a thumbprint on the consent form. Consent 
will be documented by having the participant sign and date the appropriate consent form. The study 
staff obtaining consent will also sign and date the consent form. A signed and dated copy of the 
consent form will be given to the participant.  
Cost/Volunteer Benefits 
There is no financial cost to the participants in this study. Caregivers in this study will receive a nominal 
compensation for their time and involvement. Compensation values and distribution mechanism will 
be documented in the informed consent form, and the value for the enrollment/consent visit will be 
greater than that of the day 15 visit based upon the longer duration of the initial visit. PATH will 
provide appropriate treatment or reimbursement for the treatment of injuries directly attributable to 
participation in this study. 
 
 

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F. APPENDIX 8 – TOXICITY TABLE 
Solicited symptoms grading scale1 
Parameter 
Grade 1 
Mild 
Grade 2 
Moderate 
Grade 3 
Severe 
Grade 4 
Potentially Life-
Threatening 
CLINICAL 
Clinical adverse 
event NOT 
identified 
elsewhere in the 
grading table 
Mild symptoms 
causing no or 
minimal 
interference with 
usual social & 
functional 
activities with 
intervention not 
indicated 
Moderate 
symptoms causing 
greater than 
minimal 
interference with 
usual social & 
functional activities 
with intervention 
indicated 
Severe symptoms 
causing inability to 
perform usual 
social & functional 
activities with 
intervention or 
hospitalization 
indicated 
Potentially life-
threatening 
symptoms causing 
inability to perform 
basic self-care 
functions with 
intervention 
indicated to 
prevent permanent 
impairment, 
persistent 
disability, or death 
Dermatologic 
Pruritus [Itching] 
(without skin 
lesions) 
Itching causing no 
or minimal 
interference with 
usual social & 
functional 
activities 
Itching causing 
greater than 
minimal 
interference with 
usual social & 
functional activities 
Itching causing 
inability to perform 
usual social & 
functional activities 
NA 
Rash 
 
Localized rash 
Diffuse rash OR 
Target lesions 
Diffuse rash AND 
Vesicles or limited 
number of bullae 
or superficial 
ulcerations of 
mucous membrane 
limited to one site 
Extensive or 
generalized bullous 
lesions OR 
Ulceration of 
mucous membrane 
involving two or 
more distinct 
mucosal sites OR 
Stevens-Johnson 
syndrome OR Toxic 
epidermal 
necrolysis 
 
 
1 Adapted from the Division of AIDS Table for Grading the Severity of Adult and Pediatric Adverse Events, version 
2.1, March 2017, of the United States National Institutes of Health. This table is available at https://rsc.tech-
res.com/clinical-research-sites/safety-reporting/daids-grading-tables.  

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Parameter 
Grade 1 
Mild 
Grade 2 
Moderate 
Grade 3 
Severe 
Grade 4 
Potentially Life-
Threatening 
Gastrointestinal 
Bloating or 
Distension  
Symptoms causing 
no or minimal 
interference with 
usual social & 
functional 
activities 
Symptoms causing 
greater than 
minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
inability to perform 
usual social & 
functional activities 
NA 
Constipation 
NA 
Persistent 
constipation 
requiring regular 
use of dietary 
modifications, 
laxatives, or 
enemas 
Obstipation with 
manual evacuation 
indicated 
Life-threatening 
consequences 
(e.g., obstruction) 
Diarrhea 
< 1 year of age 
Liquid stools (more 
unformed than 
usual) but usual 
number of stools 
Liquid stools with 
increased number 
of stools OR Mild 
dehydration 
Liquid stools with 
moderate 
dehydration 
Life-threatening 
consequences 
(e.g., liquid stools 
resulting in severe 
dehydration, 
hypotensive shock) 
Vomiting 
Transient or 
intermittent AND 
No or minimal 
interference with 
oral intake 
Frequent episodes 
with no or mild 
dehydration 
Persistent vomiting 
resulting in 
orthostatic 
hypotension OR 
Aggressive 
rehydration 
indicated (e.g., IV 
fluids) 
Life-threatening 
consequences 
(e.g., hypotensive 
shock) 
Neurologic 
Seizures 
< 18 years of 
age (includes 
new or pre-
existing febrile 
seizures) 
 
Seizure lasting < 5 
minutes with < 24 
hours postictal 
state 
 
Seizure lasting 5 to 
< 20 minutes with 
< 24 hours 
postictal state 
 
Seizure lasting ≥ 20 
minutes OR > 24 
hours postictal 
state 
 
Prolonged and 
repetitive seizures 
(e.g., status 
epilepticus) OR 
Difficult to control 
(e.g., refractory 
epilepsy) 
Systemic 

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Parameter 
Grade 1 
Mild 
Grade 2 
Moderate 
Grade 3 
Severe 
Grade 4 
Potentially Life-
Threatening 
Acute Allergic 
Reaction 
Localized urticarial 
(wheals) with no 
medical 
intervention 
indicated 
Localized urticarial 
with intervention 
indicated OR Mild 
angioedema with 
no intervention 
indicated 
Generalized 
urticarial OR 
Angioedema with 
intervention 
indicated OR 
Symptoms of mild 
bronchospasm 
Acute anaphylaxis 
OR Life-threatening 
bronchospasm OR 
Laryngeal edema 
Lethargy 
Symptoms causing 
no or minimal 
interference with 
usual social & 
functional 
activities 
Symptoms causing 
greater than 
minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
inability to perform 
usual social & 
functional activities 
Incapacitating 
symptom of 
lethargy causing 
inability to perform 
basic self-care 
functions 
Fever  
38.0°C to < 38.6 °C 
≥ 38.6 °C to < 
39.3°C 
≥ 39.3 °C to < 
40.0°C 
≥ 40.0 °C 
Weight Loss  
NA 
5% to < 9% loss in 
body weight from 
baseline 
≥ 9% to < 20% loss 
in body weight 
from baseline 
≥ 20% loss in body 
weight from 
baseline OR 
Aggressive 
intervention 
indicated (e.g., 
tube feeding, total 
parenteral 
nutrition)