Effect of Fibre Supplements on Gestational Diabetes

Aim: To evaluate the effect of a soluble fibre supplement in glucose metabolism in pregnant women at high risk of developing gestational diabetes.

Study methodology: 12 week randomised, placebo-controlled, single-blinded study.

Intervention: Women will first be categorised according to their glycaemic status into two groups: GDM and glucose tolerant women. Afterwards, each group will be randomised into two groups and given either

1. Fibre supplement (Guar gum)

2. Placebo (Cellulose) The participants will take the dietary supplement three times a day for 12 weeks. They will receive regular antenatal control according to NICE guidelines recommendations. Compliance will be addressed by weekly phone calls.

Number of volunteers: The number of participants will be arbitrarily define as no data of this type of intervention exists in this cohort. 40 participants will be recruited. Previous studies, outside pregnancy, have observed metabolic effects in samples of 15-40 subjects.

Recruitment: Women will be recruited in the antenatal clinic at Queen Charlotte and Chelsea's Hospital. An information sheet will be provided to eligible women in the first antenatal visit to explain the purposes of the study. Women who decide to take part will be asked to sign a consent form.

Randomisation: Initially women will be categorised according to glycaemic status in two cohorts: Glucose tolerant and Early gestational diabetes (GDM) cohort. Each of the two cohorts will then be independently randomised to either intervention or standard care using the sealed envelope website. Guar gum (intervention) and cellulose (placebo) will be provided in identical packed sachets.

PRIMARY OUTCOME MEASURES

- Glucose values at 0 and 120 minutes during an OGTT performed at 28 weeks in the glucose tolerant group.

- Glycaemic control assessed by need of insulin treatment, days to insulin treatment and total insulin dose.

SECONDARY OUTCOME MEASURES

Assessing insulin sensitivity and insulin secretion at 28 weeks of pregnancy in the glucose tolerant cohort of women through validated indices using glucose and insulin (or C-peptide) values derived from the OGTT.

• Insulin sensitivity will be assessed using the Matsuda formula

• Insulin secretion will be assessed by second-void fasting UCPCR. In the gestational diabetes cohort, where a second OGTT will not be performed, parameters of insulin resistance (HOMA_IR index) and secretion could be assessed using urinary C-peptide obtained from a fasting second-void urine sample at 28 weeks gestation, although these measurements have not been validated in pregnancy.

6. ASSESSMENT AND FOLLOW UP Women will be assess by the researchers at four different visits. At the first antenatal visit, the general care team will mentioned the study, if the patient consents, she will then be approached by the researchers to explain the study protocol. At the second visit, when the OGTT is performed, women who decided to take part in the study will sign the consent from. At the next antenatal visit where results from the OGTT are explained, women included will be randomised. The last research visit will take place at 28 weeks gestation. Additionally, they will be contacted weekly, by telephone to assess compliance and confirm willingness to participate in the study. All participating women will continue to attend routine follow up by the Obstetrics team at the antenatal clinic. Women diagnosed with GDM in early pregnancy will continue their routine care under the Obstetric team at Queen Charlotte's Hospital. We will be collecting information regarding treatment from the notes taken at this follow-up visits.

1. st visit (≤16 weeks gestation) Women with a previous history of GDM will be considered at high risk of recurrence and will be referred to the antenatal clinic before 16 weeks of pregnancy. At this first visit, the direct care team will inform them about the study and ask them if they are willing to speak with a member of the research team. If they agree, a research team member will explain the protocol and invite them to participate in the study. An information sheet will be provided and women will have time to consider if they are willing to participate and inform the research team at their next antenatal visit.

2. nd visit Women who decide to participate will inform the research team when they attend their second antenatal visit, when an OGTT is to be performed. In this visit, women willing to participate will be asked to sign a consent form.

As part of the standard pregnancy care, these women will be booked for GDM early screening. The following investigations will be performed as part of their usual NHS care:

- Blood tests: full blood count, renal function. GAD antibodies can be requested in women in whom type 1 diabetes is suspected.

- Urine sample to test for proteinuria, glycosuria, leucocytes and nitrates.

- Women willing to participate in the study will be additionally requested to provide a second urine sample after the 120 min during the OGTT. When bloods are taken will also request HbA1c as part of the study's investigations.

- An OGTT to test for early GDM will be performed as part of routine care. During the OGTT blood samples are taken at 0 and 120 minutes. Additionally to glucose, which is normally measure during the OGTT, for study purposes we will request serum C-peptide at these time points.

Baseline insulin sensitivity and insulin secretion will be estimated through validated indices using serum glucose, insulin (or C-peptide) and urinary C-peptide.

After the participants have agreed to take part in the study and consent has been signed, data regarding anthropometrics, demographics and medical history will be collected. This information will be obtained by reviewing the participant's medical notes.

Anthropometric measurements will be recorded at the first antenatal routine visit, including: pre-pregnancy weight, weight at first visit and height. BMI will be calculated with this data.

Demographics, personal and medical history will be extracted from medical records, including: maternal age, ethnicity, education, smoking during pregnancy, medical history (hypertension, PCOS, other relevant condition), obstetric history (parity, stillbirth, macrosomia, pre-eclampsia).

3. rd visit (Randomisation)

After obtaining the results from the OGTT, women will be categorised by their glycaemic status and divided into two independent cohorts:

- Women diagnosed with gestational diabetes (according to NICE guidelines 2015 diagnostic criteria): early GDM cohort

- Women with normal glucose tolerance or impaired glucose tolerance but not fulfilling GDM criteria: Glucose tolerant cohort.

Women from each cohort will be randomised into two groups after the results of the OGTT are obtained:

- Intervention group: Guar gum fibre supplement

- Placebo group: Cellulose supplement Women will the fibre supplement 3 times a day (with each main meal) until the 28th week of pregnancy when they will be re-evaluated.

4. rd visit (28th weeks gestation)

Glycaemic status will be assessed at 28 weeks gestation in the two cohorts of women participating in the study:

- Women from the Glucose tolerant cohort will undergo a second OGTT. Blood and urine samples will be collected at 0 and 120 minutes. Glycaemic status will then be assessed with the endpoint being the diagnosis of GDM (yes/no). Insulin sensitivity and insulin secretion will be estimated using the same parameters as in the first visit. As part of the study, women will be weight to calculate BMI and gestational weight gain at 28 weeks gestation. The research team will have their last face to face visit with the participant to discontinue the intervention and answer any potential questions about the study.

- In women in the Early GDM cohort, glycaemic control will be assessed by the need of insulin treatment (yes/no) and in those requiring insulin the days-to-treatment (insulin) and total insulin dose will be recorded. Data will be obtained by reviewing medical records. As part of the study, women will be weight to calculate BMI and gestational weight gain at 28 weeks gestation. The research team will have their last face to face visit with the participant to discontinue the intervention and answer any potential questions about the study.

PROCEDURES Women will be advised to fast for 10 hours the night before the oral glucose tolerance test. They will also be advised no to restrict carbohydrate ingestion the three days prior to the OGTT. At the antenatal clinic, blood samples will be collected at baseline (0 min) and at 120 min after the administration of an oral load of 75 g of glucose.

In the baseline sample other parameters will also be determined. Creatinine and HbA1c will measure in all participants and GAD antibodies will be requested in women in whom type 1 diabetes is suspected.

Serum insulin and C-peptide will be collected at 0 and 120 min during the OGTT. Women will be advised to void their first urine before attending the antenatal clinic. Second-void fasting urine sample will be collected at 0 min during the OGTT. A second urine sample will be collected at 120 min. Urinary C-peptide will measure in these samples will be transfer to boric acid containers, the alliquoted into 1 ml cryotubes ans stored at -80 ºC until analysis. Serum creatinine will be collected at baseline (0 min) and measure in the central lab. The urinary C-peptide creatinine ratio (UCPCR) will be calculated using these data.

ASSESSMENTS

(i) Gestational diabetes Gestational diabetes will be diagnosed if fasting plasma glucose >5.6 mmol/L and/or 120 min plasma glucose >7.8 mmol/L after the glucose load administered in the OGTT. Women with fasting plasma glucose <5.6 mmol/L and 120 min plasma glucose <7.8 mmol/L will be classified as "Glucose tolerant".

(ii) Insulin sensitivity Insulin sensitivity will be assessed using the Matsuda index as originally described (18) (Equation 1) and using two modified Matsuda equations in which insulin is substitute by either serum or urinary C-peptide (21,22) (Equations 2 and 3).

Equation 1. ISOGTT= 10,000 / √ {[FPG x fasting insulin] x [mean glucose x mean insulin during OGTT]} Equation 2. ISOGTTC-pep=500,000/√{[FPGxFsC-pep] x [mean glucose x mean sC-pep during the OGTT]} Equation 3. ISOGTT-UCPCR=500,000/√{[FPG×FUCPCR pmol/mmol] x [mean glucose x mean UCPCR during the OGTT]}

(iii) Insulin secretion Insulin secretion will be estimated by the UCPCR obtained using the fasting second-void urine sample, as it is strongly correlated with serum insulin, serum C-peptide (19) and 24-h urinary C-peptide (20).

Insulin response will be estimated using the UCPCR calculated with 120 min post-OGTT urine sample, as it is strongly correlated with the C-peptide and insulin area under the curve (19).

(iv) Need of insulin treatment, Days to insulin treatment and Total insulin dose Medical records of women in the Early GDM cohort will be review in order to determine the need of insulin treatment (yes/no) at 28 weeks gestation. In women requiring insulin treatment at any time during follow up, days-to-insulin treatment, calculated from randomisation day to the day insulin treatment is prescribed, and total insulin dose, calculated as the total dose of insulin international units (IU) divided by weight in Kg (insulin UI/Kg), will be recorded

PRE-RANDOMISATION EVALUATIONS Women will be evaluated by the mid-wife at the first visit in the antenatal clinic. An OGTT will be book to test for early GDM in women with previous GDM considered at high risk of recurrence. Personal and medical history will be recorded. When women attend for the OGTT, blood and urine samples will be collected at 0 and 120 minutes.

WITHDRAWAL CRITERIA Participants will be free to withdraw at any time and are not required to give a reason.

ADVERSE EVENTS Adverse Event (AE): Any untoward medical occurrence in a patient or clinical study subject.

Serious Adverse Event (SAE): Any untoward and unexpected medical occurrence that: results in death, is life- threatening, requires hospitalisation, results in persistent or significant disability or incapacity or a congenital abnormality or birth defect

Medical judgement should be exercised in deciding whether an AE is serious in other situations. Important AEs that are not immediately life threatening or do not result in death or hospitalisation but may jeopardise the subject or may require intervention to prevent one of the other outcomes listed in the definition above, should also be considered serious.

REPORTING PROCEDURES All adverse events should be reported. Depending on the nature of the event the reporting procedures below should be followed. Any questions concerning adverse event reporting should be directed to the Chief Investigator in the first instance.

Non serious AEs All such events, whether expected or not, should be recorded.

Serious AEs An SAE form should be completed and faxed to the Chief Investigator within 24 hours. However, relapse, death and hospitalisations for elective treatment of a pre-existing condition do not need reporting as SAEs. All SAEs should be reported to the sponsor.

All SAEs should be reported to the Bromley Research Ethics Committee where in the opinion of the Chief Investigator the event was:

'Related', i.e. resulted from the administration of any of the research procedures; and 'Unexpected', i.e. an event that is not listed in the protocol as an expected occurrence.

Reports of related and unexpected SAEs should be submitted within 15 days of the Chief Investigator becoming aware of the event, using the NRES SAE form for non-IMP studies.

Local investigators should report any SAEs as required by their Local Research Ethics Committee and/ or Research and Development Office.

STATISTICS AND DATA ANALYSIS Data handling and analysis will be carried out by SPSS version 20.0 version. No power calculation to decide sample size will be used. The number of participants has been arbitrarily defined, as it is a pilot study. We aim to recruit 40 women.

Before randomisation, women will be categorised according to their glycaemic status in two cohorts: Glucose tolerant and GDM. Each of these two cohorts will be randomly allocated into the intervention (soluble fibre supplement) and control (placebo) group. Randomisation will be by sealed envelopes. Participants but not researchers will be blinded as to the type of intervention prescribed.

Differences between group en each cohort will be assessed using Student t test or Mann-Whitney U test for numeric variables depending on distribution; X2 will be used for categorical variables.

In the Glucose tolerant women cohort:

For the primary outcome, GDM (yes/no), logistic regression analysis will be performed.

For the secondary outcomes, insulin sensitivity and insulin secretion, linear regression analysis will be performed.

In the early GDM cohort:

For the primary outcome, need for insulin treatment (yes/no), logistic regression analysis will be performed. Additionally days-to-treatment analysis will be performed for those participants starting insulin therapy during follow up.