BIFI-OBESE: Clinical Trial in Paediatric Obesity
Condition: Obesity, Childhood
MeSH terms: Obesity; Pediatric Obesity
Study type: Interventional
Phase: Phase 4

Summary: 
      Obesity is a major, public health concern that affects at least 400 million individuals and
      is associated with severe disorders including diabetes and cancers. Worldwide, the prevalence
      of overweight and obesity combined in children, adolescents and youth, between 1980 and 2013,
      increased to 47.1%, with alarming data also in developing countries. Obesity is often caused
      by imbalance between excessive caloric intake and reduced physical activity.

      Recently, microbial changes in the human gut was proposed to be another possible cause of
      obesity and it was found that the gut microbes from fecal samples contained 3.3 million
      non-redundant microbial genes. However, it is still poorly understood how the dynamics and
      composition of the intestinal microbiota are affected by diet or other lifestyle factors.
      Moreover it has been difficult to characterize the composition of the human gut microbiota
      due to large variations between individuals.

      The role of the digestive microbiota in the human body is still largely unknown, but the
      bacteria of the gut flora do contribute enzymes that are absent in humans for food digestion.
      Moreover, the link between obesity and the microbiota is likely to be more sophisticated than
      the simple phylum-level Bacteroidetes: Firmicutes ratio that was initially identified, and it
      is likely to involve a microbiota-diet interaction.

      Obese and lean subjects presented increased levels of different bacterial populations. It is
      hypothesized that the obese microbiome is set up to extract more calories from the daily
      intake when compared to the microbiome of lean counterparts. In addition, a caloric diet
      restriction impacted the composition of the gut microbiota in obese/overweight individuals
      and weight loss.

      In lean subjects there are Coriobacteriaceae, Lactobacillus, Enterococcus, Faecalibacterium
      prausnitzii, Prevotella, Clostridium Eubacterium, E. coli and Staphilococcus. By contrast,
      Bifidobacterium, Methanobrevibacter, Xylanibacter, Bacteroides characterize the composition
      of lean gut microbiota.

      For this reason, in a cohort of obese paediatric subjects with visceral adiposity, the aim of
      the study is to assess the efficacy of a supplementation with probiotic bifidobacteria with
      respect to a conventional treatment on weight loss and improvement of cardio-metabolic risk
      factors.
    

Detailed description: 
      Study design: A single-center pilot open-label randomized control trial. Population: The
      study will comprise a total of 100 subjects of both sexes, between 6 and 18 years of age,
      obese, according to the IOTF criteria and with visceral adiposity, as waist circumference ≥
      90th percentile, pubertal stage ≥ 2 according to the Tanner stage, HOMA-IR > 2,5 or insulin >
      15 µU/ml, diet naïve or with failure of weight loss (defined as -1 kg/m2 BMI in 1 year).

      Inclusion/ Exclusion criteria (see Eligibility Criteria). Intervention: In the first part of
      the study (Study 1, V0-V1) patients will be randomized in a open-label, into two groups
      homogeneous for number and sex of the subjects. One group will receive a supplementation of
      probiotic containing Bifidobacterium breve B632 and Bifidobacterium breve BR03, 15 gtt/die
      (3x108 CFU/die) and one group will receive a placebo for a total of 2 months of treatment.
      Both group receives a Standard Diet according to routine care and practice. For patients who
      wants to continue the study there will be a cross-over study (study 2, V2-V3) after one month
      of wash-out.

      Dietary restriction: The standard diet will be distributed with 55-60% of carbohydrates
      (45-50% complex and no more than 10% refined and processed sugars), 25-30% lipids and 15%
      proteins, and will be performed in accordance with the calories of an isocaloric balanced
      diet calculated throughout the Italian LARN Guidelines for age and gender.

      Physical activity: all subjects will receive general recommendations about performing
      physical activity. Exercise will be conducted daily and will consist of 30 minutes of aerobic
      physical activity.

      Randomization: Participants will be randomly assigned in a 1:1 to probiotic intervention
      group or placebo group.

      Timing: Patients will be evaluated firstly at time of enrollment (V0) and, at the end of the
      first part of study (Study 1, V1), biochemical evaluations will be completed. Next there will
      be one month of wash-out when the patients don't take any probiotic or placebo. In the second
      part of the study 2, patients will be evaluated at V2 and, after 2 months of treatment (Study
      2, V3). The following anthropometric measures, biochemical and ultrasound evaluations and
      questionnaires will be obtained:

        1. Anthropometric measures:

             -  height (V0, V1, V2, V3);

             -  weight (V0, V1, V2, V3);

             -  body mass index (BMI; Kg/m2) (V0, V1, V2, V3);

             -  waist and hip circumferences (V0, V1, V2, V3) for the calculation of the following
                ratios: waist/hip, waist/height;

             -  Tanner stage (V0, V1, V2, V3);

             -  blood pressure and heart rate (V0, V1, V2, V3). Biochemical evaluations (after a
                12-h overnight fast): CBC with formula, serum insulin-like growth factor 1 (IGF1,
                ng/mL), 25-hydroxy (OH) vitamin D (ng/mL), uric acid (mg/dL), alkaline phosphatase
                (U/L), ACTH (pg/mL), cortisol (microg/dL), TSH (uuI/mL), fT4 (ng/dL) (V0, V1, V2,
                V3); aspartate aminotransferase (AST, IU/L), alanine aminotransferase (ALT, IU/L);
                AST-to-ALT ratio will be calculated as the ratio of AST (IU/L) and ALT(IU/L) (V0,
                V1, V2, V3); serum creatinine concentration (mg/dL) will be measured with the
                enzymatic method; according to the NKF-K/DOQI Guidelines for CKD in children and
                adolescents, the eGFR will be calculated using updated Schwartz's formula: eGFR
                (mL/min/1.73 m2) = [0.413 x patient's height (cm)] / serum creatinine (mg/dL) (V0,
                V1, V2, V3); glucose (mg/dL), insulin (μUI/mL); insulin-resistance (IR) will be
                calculated using the formula of Homeostasis Model Assessment (HOMA)-IR: (insulin
                [mU/L] x glucose [mmol/lL) / 22.5) (V0, V1, V2, V3); lipid profile: total
                cholesterol (mg/dL), High-Density Lipoprotein (HDL)-cholesterol (mg/dL),
                triglycerides (mg/dL); Low-Density Lipoprotein (LDL)-cholesterol will be calculated
                by the Friedwald formula and non-HDL (nHDL)-cholesterol will be also calculated(V0,
                V1, V2, V3); oral glucose tolerance test (OGTT: 1.75 g of glucose solution per kg,
                maximum 75 g) and samples willbe collected for the determination of glucose and
                insulin every 30 min. The area under the curve (AUC) for parameters after OGTT will
                be calculated according to the trapezoidal rule. Insulin sensitivity at fasting and
                during OGTT will be calculated as the formula of the Quantitative
                Insulin-Sensitivity Check Index (QUICKI) and Matsuda index (ISI). The stimulus for
                insulin secretion in the increment in plasma glucose as insulinogenic index will be
                calculated as the ratio of the changes in insulin and glucose concentration from 0
                to 30 min (InsI). Βeta-cell compensatory capacity will be evaluated by the
                disposition index defined as the product of the ISI and InsI (DI) (V0, V1, V2, V3);
                a collection at rest of first-morning urine sample. Physical and chemical
                urinalysis; urine albumin (mg/L) will be determined by an advanced
                immunoturbidimetric assay and urine creatinine (mg/dL) will be measured using the
                enzymatic method. Urine albumin to creatinine ratio (u-ACR - mg/g), will be
                calculated using the following formula: [urine albumin (mg/dL) / urine creatinine
                (mg/dL)] x 1000. For these calculations both albumin and creatinine will be in the
                same unit. The subjects whose urine will be found positive, they will undergo a
                collection of two more samples and will be considered the u-ACR mean value of these
                (V0, V1, V2, V3). A sample of feces will be taken for microbic count (V0, V1, V2,
                V3). LPS (V0, V1, V2, V3). LPS will be measured with commercial kits (Limulus
                amoebocyte lysate assay) with standard procedures. Citokines IL1, IL1β, IL6, IL10,
                TNFα will be evaluated (V0, V1, V2, V3) (ELISA kit).

        2. A health diary will be taken during the 2 months of treatment: each patient will
           complete the diary with collateral effects or antibiotic treatment ecc.

        3. NGS (Next Generation Sequencing) will be analized for fecal analysis (V0, V1, V2, V3)

        4. Metabolomic analysis will be taken with mass spectrometry on fecal samples (V0, V1, V2,
           V3)

        5. SCFA analysis on fecal samples (V0, V1, V2, V3).

      Outcomes (see Outcome Measures). Information retrieval: A case report form (CRF) will be
      completed for each subject included in the study. The source documents will be the hospital's
      or the physician's chart.

      Statistical e sample size: A sample of 16 individuals has been estimated to be sufficient to
      demonstrate a difference of 10 mg/dl in the basal glucose concentration with 90% power and a
      significance level of 95% and a drop-out rate of 10% at the 8th weeks of treatment. A sample
      of 34 individuals in each group has been estimated to be sufficient to demonstrate a
      difference of 1,4 point in the HOMA-IR index with 90% power and a significance level of 95%
      and a drop-out rate of 10% at the 8th weeks of treatment. Statistical significance will be
      assumed at P< 0.05. The statistical analysis will be performed with SPSS for Windows version
      17.0 (SPSS Inc., Chicago, IL, USA).

      Organization characteristics: The study will be conducted at the Pediatric Endocrine Service
      of Division of Pediatrics.

      All blood samples will be measured evaluated using standardized methods in the Hospital's
      Chemistry Laboratory, in Maggiore della Carità hospital, in Novara, previously described.
      Fecal analysis will be measured in the Department of Sciences and Technologies, University of
      Bologna, in Bologna.

      Good Clinical Practice: The protocol will be conducted in accordance with the declaration of
      Helsinki. Informed consent will be obtained from all parents prior to the evaluations after
      careful explanations to each patient.
    

Intervention name(s): Bifidobacterium breve BR03 and Bifidobacterium breve B632; Placebos
Intervention type(s): Drug; Drug
Other intervention name(s): probiotic; Placebo
Allocation: Randomized
Intervention model: Crossover Assignment
Intervention model description: In the first part of the study (Study 1, V0-V1) patients will be randomized in a open-label, into two groups homogeneous for number and sex of the subjects. One group will receive a supplementation of probiotic containing Bifidobacterium breve B632 and Bifidobacterium breve BR03, 15 gtt/die (3x108 CFU/die) and one group will receive a placebo for a total of 2 months of treatment. For patients who wants to continue the study there will be a cross-over study (study 2, V2-V3) after one month of wash-out.
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Treatment
Keywords: obesity; paediatric; probiotic