VS002A in the Treatment of Acute Watery Diarrhea in Infants and Young Children
Condition: Acute Watery Diarrhoea
MeSH terms: Diarrhea
Study type: Interventional

Summary: 
      Background (brief):

        1. Burden: Diarrhea is the second deadliest disease for under-five children globally and
           the situation is more serious in developing countries. It was responsible for 688
           million illnesses and 499,000 deaths worldwide in children less than 5 years of age in
           2015 (Kotloff, 2017). Majority of the deaths from diarrhea occur before the second
           birthday of the children and it contributes to more than 20% of overall death in infants
           and young children.

        2. Knowledge gap: Oral rehydration solution (ORS) is being used as a standard treatment for
           acute watery diarrhea for long time and which is one of the best inventions in the
           history of medicine. The ORS currently recommended by the WHO/UNICEF contains glucose,
           sodium chloride, potassium chloride, and tri-sodium citrate dehydrate, which is optimal
           for rehydration of patients of all ages with dehydration from acute diarrhea of any
           aetiology. However, oral rehydration therapy (ORT) with the present ORS formulation has
           certain limitations - it does not reduce the volume, frequency or the duration of
           diarrhea. Additionally, the failure of present standard ORS to reduce dramatically stool
           output likely contributes to the relatively limited use of ORS by mothers as they do not
           feel that ORS is helping their child from the episode of diarrhea. Thus, it warrants the
           development of newer and improved formulation of ORS to become more effective against
           diarrhea.

        3. Relevance: It has been reported that the glucose contained in standard ORS may fail to
           absorb fluid and electrolytes adequately from gut and worsen diarrhea in different
           patho-physiological ways as the present WHO ORS is hypo-osmolar. Whereas, certain
           neutral amino acids (e.g. glycine, L-alanine, L-glutamine) are able to enhance the
           absorption of sodium ions and water from gut. By using this concept, the University of
           Florida (UF), developed a sugar-free, shelf-stable amino acid-based hydration medicinal
           food named 'VS002A' that effectively rehydrates, and improves barrier function of the
           bowel following infections targeting the gastrointestinal tract. So, the investigators
           ought to know whether VS002A will be superior or not to WHO-ORS in the treatment of
           acute non-cholera watery diarrhea in infants and young children.

      Hypothesis (if any): The amino acid-based ORS "VS002A" will reduce duration of non-cholera
      watery diarrhea in infants and young children when compared to treatment with standard
      WHO-ORS.

      Objectives: To compare the efficacy of amino acid-based ORS "VS002A" compared to standard
      Glucose-based WHO ORS in infants and young children suffering from acute non-cholera watery
      diarrhea.

      Methods:

        -  It will be a randomized, double-blind, two cell clinical trial at Dhaka Hospital of
           icddr,b. Total 312 (156 in each arm) male children aged 6-36 months old with acute
           (onset <48 hours) non-bloody watery diarrhea will be included in this study. However,
           patients with severe malnutrition, any systemic illness, cholera, unwilling to comply
           with study protocol, remain significantly dehydrated 4 hours after intravenous fluid
           infusion (if required at start), has documentation of taking antibiotics or
           antidiarrheal 48 hours before admission will be excluded.

        -  Intervention arm participants will get amino acid ORS (VS002A) and control arm will get
           standard glucose based WHO-ORS. Other aspects of clinical managements for diarrhea as
           per hospital guideline will be similar in both cases and controls
    

Detailed description: 
      Hypothesis to be tested:

      The amino acid-based ORS "VS002A" will reduce the duration of non-cholera, watery diarrhea in
      infants and young children when compared to treatment with standard WHO-ORS.

      Specific Objectives:

        1. To evaluate and compare clinical responses (duration of diarrhea, stool output, ORS
           intake, and clinical success) in infants and young children suffering from acute
           non-cholera watery diarrhea treated with standard Glucose-based WHO ORS and amino
           acid-based ORS "VS002A".

        2. To evaluate and compare electrolyte imbalance as possible complications in infants and
           young children suffering from acute non-cholera watery diarrhea treated with standard
           Glucose-based WHO ORS and amino acid-based ORS "VS002A".

      Background of the Project including Preliminary Observations:

      Diarrheal disorders constitute a leading cause of morbidity and mortality globally, and
      continue to be a major concern particularly for developing countries (Platts-Mills, 2015;
      WHO/UNICEF, 2013;Bern, 1992). Estimates show that in the year 2010, globally there were 1.7
      billion episodes of diarrhea and 7.6 million deaths in children less than 5 years of age,
      mostly in the developing world; 72% of the deaths occurred in the first 2 years of life (Liu,
      2012, 2015; Walker, 2013). In the Southeast Asian region, diarrhea is responsible for more
      than 20% of the total deaths among infants and young children (Liu, 2012, 2015).

      Acute infectious diarrheal diseases are caused by a number of microbial pathogens. The
      bacterial pathogens include Vibrio cholerae, enterotoxigenic Escherichia coli, Shigella,
      Salmonella, and Campylobacter. Rotavirus is the most prominent viral diarrheagenic pathogen
      and also the leading cause of infantile diarrhea, whereas Entamoeba histolytica, Giardia
      lamblia and Cryptosporidia are the important parasites causing diarrhea. Based upon the
      pathogenetic mechanisms, these organisms may be broadly divided into two groups - secretory
      and invasive. V. cholerae is the prototype pathogen causing secretory diarrhea through
      liberation of an enterotoxin which stimulates intestinal secretion via intracellular
      accumulation of cyclic AMP (Fasano, 2002). There is insignificant inflammation or structural
      change in the intestinal mucosa and the diarrhea is watery in nature. Sufficient loss of
      fluids and electrolytes leads to dehydration, and thus the mainstay of treatment of patients
      with acute infectious secretory diarrhea is fluid replacement. On the other hand, Shigella
      (the prototype invasive pathogen) invades the gut mucosa and induces an inflammatory
      reaction, producing a dysenteric illness (Levine, 1982). The amount of fluid loss is minimal,
      and antibiotics are generally required in their management. In the developing countries,
      recent studies show that the commonest attributable microbial causes of moderate-to-severe
      diarrheal illnesses in infants and young children are: rotavirus, Cryptosporidium,
      enterotoxigenic E. coli producing heat-stable toxin (ST-ETEC, with or without co-expression
      of heat-labile enetrotoxin), and Shigella (Kotloff, 2013; Liu, 2016).

      The standard clinical intervention in acute watery diarrhea is the use of oral rehydration
      solution (ORS), continued feeding, and oral zinc supplementation (WHO/UNICEF, 2013; WHO,
      1995). The development of ORS for the treatment of dehydration due to diarrhea is one of the
      most significant therapeutic advances in the history of medicine. It is based upon the
      observations that even in a secreting small intestine it is possible to achieve a positive
      gut balance of fluid and electrolytes by adding glucose to the salt solution (Pierce, 1968;
      Fordtran, 1975). ORT is now recognized as a major scientific advance of utmost practical
      importance. The ORT solution currently recommended by the WHO/UNICEF contains glucose, sodium
      chloride, potassium chloride, and trisodium citrate dehydrate, which is optimal for
      rehydration of patients of all ages with dehydration from acute diarrhea of any etiology. It
      is estimated that ORS alone can successfully rehydrate 90% of patients with dehydration from
      acute diarrhea who previously would have required intravenous therapy therapy (Hirschorn,
      1991).

      However, ORT with the present ORS formulation has certain limitations - it does not reduce
      the volume, frequency or the duration of diarrhea (Mahalanabis, 1996). This is a constraint
      on the acceptance of ORT as a treatment of diarrhea, since the mothers and the health-care
      providers expect earlier recovery and a reduced severity of the disease, which is partly
      responsible for the widespread use of ineffective (and often harmful) antibiotics and other
      anti-diarrheal agents. These limitations prompted the development of the concept of improved
      ORS (initially termed 'super ORS') (Mahalanabis, 1986). Conceptually, an improved ORS should
      be able to reduce stool volume (by enhancing better reabsorption of fluid secreted into the
      small intestine), shorten duration of diarrhea (by reducing ileal effluent flow and
      stimulating colonic salvage), and reduce failure rate of ORT in patients with high purging
      rate. Several strategies were used to develop and test improved ORS - including the use of
      amino acids in addition to glucose, replacing glucose with glucose-polymers, using cereals
      instead of glucose, adding non-absorbable fermentable products to the ORS, and making the ORS
      hypo-osmolar by reducing both glucose and sodium concentrations. The recently recommended
      hypo-osmolar ORS is the result of intense search for such an improved ORS. Though the new
      hypo-osmolar ORS is more efficacious than the previously standard ORS, the duration of
      diarrhea is still not shortened, and there are failures with ORT (Alam, 1999). Thus, there is
      scope for further improvements of ORT.

      The ORS takes advantage of the glucose-coupled intestinal sodium transport mechanism across
      the luminal membrane by the protein sodium glucose co-transporter 1 (SGLT1). Although
      nutrient independent sodium absorption across the intestinal epithelial brush border membrane
      is impaired in diarrhea, co-transport of sodium and glucose is preserved. This allows
      absorption of sodium and thus water as provided by oral rehydration solutions (Figure 1). Key
      in the success of use of ORS for rehydration is the fact that these physiological processes
      stay intact, even in the setting of severe diarrhea.

      However, although ORS substantially enhances glucose-stimulated sodium absorption and
      correction of dehydration, it does not significantly decrease stool output, which is one of
      the main reasons for underutilization of ORS (Greenough, 1993; Wagner, 2015). Although an
      adequate explanation for this has not been established, it could be explained if glucose also
      stimulated anion secretion. In fact, based upon in vitro studies on the effect of glucose on
      active chloride and fluid secretion in mouse small intestinal cells and human colorectal
      adenocarcinoma (Caco-2) cells, glucose not only enhanced sodium absorption via SGLT1, a
      well-established phenomenon, but also stimulated active electrogenic chloride secretion via
      an increase in intracellular Ca2+ (Yin, 2014, AJP Cell). In clinical or experimental
      conditions, glucose-stimulated chloride secretion is most likely to occur in the villous cell
      region. These observations raise the possibility that in those clinical conditions in which
      there is villous damage and impaired glucose-stimulated sodium absorption, glucose may have
      deleterious effects on overall fluid and electrolyte movement by virtue of its stimulation of
      active chloride secretion.

      In young children with acute non-cholera watery diarrhea, the rate of treatment failure due
      to clinically evident glucose intolerance is generally expected to be around 2% (Santosham M,
      1982). However, 30% of young Peruvian children with acute diarrhea had detectable glucose
      malabsorption leading to treatment failure in 7% of the patients (Salazar-Lindo E, 1986).
      Thus, it may be hypothesized that simple dietary modifications and/or changes in the ORS
      formulation, such that glucose is replaced with compounds that do not stimulate chloride
      secretion, could significantly alter the outcome of the disease process.

      Since certain neutral amino acids (e.g. glycine, L-alanine, L-glutamine) are also able to
      enhance the absorption of sodium ions and water, one approach to developing an improved ORS
      formulation was based on adding neutral amino acids or their dipeptides to WHO-ORS
      (Mahalanabis, 1983). The first studies of glycine-containing ORS were inconclusive; a
      meta-analysis of seven clinical trials conducted later showed that the ORS formulations
      containing glucose (or maltodextrin) and glycine (or glycyl-glycine) were not clinically
      superior to the standard WHO-ORS (International Study Group on Improved ORS, 1991). A summary
      of the eight clinical trials comparing L-alanine containing ORS with the WHO-ORS also showed
      variable results; in general, the differences in children with non-cholera diarrhea were not
      statistically significant (Bhan, 1994). Three clinical trials comparing glutamine-containing
      ORS with the WHO-ORS showed that it does not have any clinical advantage in children with
      non-cholera diarrhea (Bhan, 1994). One common thing among all the trials is that all the
      experimental amino acid-containing ORSs also contained glucose in variable amounts (9 g/L to
      20 g/L); also most of these solutions had relatively high osmolarity (320 - 400 mosmol/L).
      One study which compared glucose-free glutamine-containing ORS with the WHO-ORS showed
      similar efficacy in children with acute non-cholera diarrhea (Gutierrez, 2007); it is to be
      noted that glutamine behaves like glucose in stimulating anion secretion (Yin, 2014). In a
      study comparing L-isoleucine supplemented glucose-containing ORS with WHO-ORS in young
      children with acute diarrhea, the differences were not statistically significant (Alam,
      2011). However, in adult patients suffering from severe cholera, L-histidine-supplemented ORS
      was found superior to a histidine-free ORS; both the experimental and control solutions were
      rice-based and glucose-free (Rabbani GH, 2005). In another study, a glucose-free
      peptilose-based ORS was found more advantageous and acceptable than the WHO-ORS for treatment
      of mostly acute watery diarrhea among children (Simakachorn, 1993).

      Previous studies on VS002:

      Entrinsic Bioscience, a small business formed from intellectual property from the University
      of Florida (UF), developed a sugar-free, shelf-stable amino acid-based hydration medicinal
      food that effectively rehydrates, and improves barrier function of the bowel following
      infections targeting the gastrointestinal tract (GI). VS002's proprietary formulation of
      select amino acids contains no glucose. Studies in humans and animals demonstrate that
      sugar-free VS002 is an effective rehydration solution in a variety of circumstances.

      Irradiation increases paracellular permeability in the gut with associated nutrient
      malabsorption, fluid imbalance, and bacterial translocation. In addition, irradiation induces
      active chloride ion secretion and fluid loss, and reduces transport of many carbohydrates,
      proteins and electrolytes. In a series of studies with radiation-induced enteritis with
      associated villus atrophy, an optimal composition of a novel ORS was defined based upon the
      observations that the inclusion of specific amino acids caused tightening of the mucosal
      barrier and reversal of symptoms of radiation-induced enteric dysfunction (Yin, 2014). VS002
      thus consists of only the amino acids that improve electrolyte and water absorption and
      correct the increased gut permeability noted following irradiation and infectious disease.

      It has been observed that certain amino acid-based pathways for sodium transport become the
      dominant pathway to compensate during times of stress. These pathways can be utilized to
      rehydrate the intestinal epithelial cells and restore normal bowel function. Eight specific
      amino acids retained their absorptive capacity from the gut lumen following radiation and at
      the same time decreased paracellular permeability: Lysine, Aspartic acid, Glycine,
      Isoleucine, Threonine, Tyrosine, Valine, and Serine.

      Studies with VS002 have shown that VS002 administration i) rapidly rehydrates to increase
      fluid and electrolyte absorption, ii) tightens the mucosal barrier, thus decreasing local and
      systemic inflammation, and iii) increases crypt count and villus height, leading to an
      increase in the surface area of absorption. There is no toxicity associated with VS002
      administration and the results of studies in animals and humans suggest that VS002 can serve
      as a highly safe and effective supportive care in place of standard ORS (Yin, 2014;
      Cheuvront, 2018; Clarke, 2018; Sollanek, 2018).

      Results of In Vivo Studies Irradiated mice: Irradiated mice were treated either with saline
      or with VS002 for 14 days. Treatment with VS002 reduced mortality, preserved body weight,
      decreased pica (indication of nausea), and improved electrolyte and nutrient absorption (Yin,
      2014; 2017) Mice with Rotavirus enterotoxin: Mice were exposed to rotavirus enterotoxin
      (NSP4) and sections of gut were tested for transport in vitro, and glucose-coupled sodium
      transport was found significantly impaired along with increased chloride secretion in the
      presence of NSP4. Glucose in the presence of NSP4 further increased the chloride secretion
      (Yin, 2017) Healthy Humans: Trials in healthy humans demonstrated i) VS002 safety; and ii)
      rehydration efficacy compared to a variety of commercial, glucose-containing beverages
      (sports drink, ORS). These studies included adults with diarrhea, healthy soldiers, young
      college students, and older adults (> 60 years old). Thus, administration of VS002 using
      eight amino acids to increase intestinal water and electrolyte absorption was associated with
      safe rehydration. See below for further study details.

      VS002 was provided to > 250 adults treated for diarrhea in Indonesia and the Philippines. The
      consumption of VS002 was well tolerated by all subjects, with no adverse events (Olveda,
      2016; Abdulah, 2018).

      A study in healthy U.S. Soldiers and civilians (n = 26, males and females) examined the
      effect of rehydration via a common sports drink or VS002 after a bout of hypertonic or
      isotonic dehydration (3-4% body mass). Rehydration via VS002 resulted in improved rehydration
      of the extracellular space compared to the common sports drink, as well as greater fluid
      retention in the VS002 group (p < 0.05). The consumption of VS002 was well tolerated by all
      subjects, with no adverse events (Cheuvront, 2018).

      Another study evaluated the ability of VS002 to optimize hydration compared to water in
      healthy adults (n = 40, males and females). Consumption of VS002 resulted in improved
      retention compared to water (p < 0.05). The consumption of VS002 was well tolerated by all
      subjects, with no adverse events (Sollanek, 2018).

      The ability of VS002 to optimize hydration was also compared in younger and older adults (n =
      24, males and females). Consumption of VS002 resulted in improved retention compared to water
      (p < 0.05). The consumption of VS002 was well tolerated by all subjects, with no adverse
      events (Clarke, 2019).

      In a randomized, double-blind study to evaluate the efficacy of VS002, 587 adult patients
      received either WHO-ORS or VS002 to treat diarrhea. Safety results revealed that no patient
      consuming VS002 experienced any serious adverse events, whereas a small number of patients
      taking WHO-ORS did. Efficacy data for clinical cure rates were not clinically or
      statistically significant between treatment groups.

      Therefore, it is hypothesized that VS002 will also be superior to WHO-ORS in the treatment of
      acute watery non-cholera diarrhea in infants and young children.

      Advanced Amino acid-based ORS: VS002A Traditional oral rehydration solutions (ORS) contain
      sugars which stimulate intestinal sodium and water absorption through a variety of
      mechanisms. However, it has been under-appreciated that traditional ORS possess no
      anti-diarrheal functions and may exacerbate infectious diarrheal secretions (Yin et al.,
      2017).

      Safety data of Advanced amino acid based ORS: VS002A

      The food additive amino acids may be safely used as nutrients added to foods as individual
      amino acids in the free, hydrated, or anhydrous form, or as the hydrochloride, sodium, or
      potassium salts:

      (1) L-Alanine; (2) L-Arginine; (3) L-Asparagine; (4) L-Aspartic acid; (5) L-Cysteine; (6)
      L-Cystine; (7) L-Glutamic acid; (8) L-Glutamine; (9) Aminoacetic acid (glycine); (10)
      L-Histidine; (11) LIsoleucine; (12) L-Leucine; (13) L-Lysine; (14) DL-Methionine (not for
      infant foods); (15) LMethionine; (16) L-Phenylalanine; (17) L-Proline; (18) L-Serine; (19)
      L-Threonine; (20) LTryptophan; (21) L-Tyrosine; (22) L-Valine.

      EBS amino acid formulas are fully compliant with FDA 21 CFR requirements.

      Gut health biomarkers Although oral rehydration therapy has greatly reduced
      diarrhoea-associated mortality, burden of diarrheal disease still persists in children below
      5 years who are living under unsanitary condition and limited public health resources. This
      contributes to 43% of stunted growth and impaired cognitive development, affecting one-fifth
      of children worldwide and one-third of children in low- and middle-income countries (LMICs)
      (Guerrant, 2013; Kosek, 2013). Vicious cycle of enteric infection and malnutrition often
      leads to enteropathy for extended periods of time in young children when adequate water and
      sanitation are lacking (Guerrant, 2013; Owino, 2016). This type of enteropathy is known as
      Environmental Enteric Dysfunction (EED).

      EED, previously known as tropical enteropathy or environmental enteropathy, is a sub-acute
      inflammatory condition of the small intestinal mucosa of unknown etiology (Ramakrishna,
      2006). It is characterized by structural changes in the small intestine including villous
      atrophy and crypt hyperplasia compromising nutrient absorption and pathogenic barrier
      (increased permeability and inflammatory cell), impaired gut immune function, malabsorption,
      growth faltering, and generally asymptomatic, as distinct from diarrheal disease (Owino,
      2016; Sullivan, 1991).

      Data are limited from LMICs including Bangladesh on inflammatory and pathological changes in
      the gut wall like EED among children with diarrhea. In this study we will study plasma
      Citrulline (CIT) concentration as potential marker of functional enterocyte mass and
      absorptive functions of intestine, and Tryptophan-Kyanurenine (KT) ratio as a marker of gut
      inflammation.

      CIT is an amino acid produced by intestinal epithelial cells and found lower in patients with
      enteropathy. One animal study reported lower plasma CIT level in the neonatal calves with
      diarrhea compared to healthy control (Gultekin, 2019). Tryptophan (TRP), a plant-derived
      essential amino acid (EAA) is needed to support growth and health in humans. In response to
      infection, TRP is mostly catabolized by an enzyme indoleamine 2,3-dioxygenase (IDO) to a
      toxic metabolite kynurenine (KYN). Low plasma TRP, high KYN and elevated KT ratio are found
      to be associated with infections (e.g. patients with inflammatory bowel disease suffer from
      diarrhoea) and chronic immune activation (Kosek, 2016).
    

Intervention name(s): VS002A
Intervention type(s): Dietary Supplement
Intervention description(s): Amino acid based, glucose free ORS
Allocation: Randomized
Intervention model: Parallel Assignment
Intervention model description: Double blind, randomized, parallel group, clinical trial
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Treatment