Treatment Options For Type 2 Diabetes In Adolescents And Youth Research Articles

165-LB: Circulating Metabolomic Biomarkers of Glycemic Control in Youth with Type 2 Diabetes (T2D)

There are few biomarkers of glycemic response among youth with T2D, despite increasing disease prevalence and suboptimal response to approved therapies. We hypothesized that plasma metabolites may predict glycemic outcomes in youth-onset T2D. We measured 480 metabolites in fasting plasma samples in the Treatment options for T2D in Adolescents and Youth (TODAY) study (n=391), in which youth with T2D aged 10-17 years were randomized to metformin alone, metformin + rosiglitazone, or metformin + intensive lifestyle intervention. Metabolite associations with loss of glycemic control (defined as HbA1c ≥8% for 6 months or need for insulin therapy) were modeled using Cox proportional hazards regression adjusted for baseline age, sex, race/ethnicity, BMI, treatment group, and fasting glucose. Loss of glycemic control was observed in 150 of 391 youth (mean 2.6 years). Baseline levels of 11 metabolites were associated with loss of glycemic control (FDR<0.05, Fig. 1A). Treatment group modified the association of hexose and xanthurenic acid levels with glycemic control. For both compounds, youth with higher baseline levels had a lower risk of treatment failure when randomized to metformin therapy alone (Fig. 1B). Thus, metabolomics provides insight into circulating analytes associated with loss of glycemic control, and may highlight different effects of specific treatments in youth with T2D. Disclosure Z. Chen: None. C. Lu: None. X. Shi: None. S. Zheng: None. D. Wolfs: None. P. Bjornstad: Advisory Panel; AstraZeneca, Novo Nordisk, Lilly, Horizon Therapeutics plc, Boehringer Ingelheim (Canada) Ltd., LG Chem, Consultant; Bayer Inc., Bristol-Myers Squibb Company. R. E. Gerszten: None. E. M. Isganaitis: None. Funding National Institutes of Health (K23DK127073)

Maternal Diabetes in Youth-Onset Type 2 Diabetes Is Associated With Progressive Dysglycemia and Risk of Complications.

Prenatal exposures, including undernutrition, overnutrition, and parental diabetes, are recognized risk factors for future cardiometabolic disease. There are currently no data on effects of parental diabetes on disease progression or complications in youth-onset type 2 diabetes (T2D). We analyzed effects of parental diabetes history on glycemic outcomes, β-cell function, and complications in a US cohort of youth-onset T2D. Participants (N = 699) aged 10 to 17 years with T2D were enrolled at 15 US centers and followed for up to 12 years as part of the TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) and TODAY2 follow-up studies. Information about diabetes diagnosis in biological mothers was available for 621 participants (never = 301; before or during pregnancy = 218; after pregnancy = 102) and in biological fathers for 519 (no diabetes = 352; paternal diabetes = 167). Maternal, but not paternal, diabetes was associated with loss of glycemic control over time, defined as glycated hemoglobin A1c greater than or equal to 8% for more than 6 months (P = .001). Similarly, maternal, but not paternal, diabetes was associated with increased risk of glomerular hyperfiltration (P = .01) and low heart rate variability (P = .006) after 12 years of follow-up. Effects were largely independent of age, sex, race/ethnicity, and household income. Maternal diabetes during vs after pregnancy had similar effects on outcomes. Maternal diabetes, regardless of whether diagnosed during vs after pregnancy, is associated with worse glycemic control, glomerular hyperfiltration, and reduced heart rate variability in youth with T2D in TODAY. The strong associations of diabetes outcomes with maternal diabetes suggest a possible role for in utero programming.

The obesity paradox: Retinopathy, obesity, and circulating risk markers in youth with type 2 diabetes in the TODAY Study

AimTo understand the relationship of obesity and 27 circulating inflammatory biomarkers to the prevalence of non-proliferative diabetic retinopathy (NPDR) in youth with type 2 diabetes. MethodsYouth with type 2 diabetes who participated in the TODAY (Treatment Options for Type 2 Diabetes in Adolescents and Youth) study were followed for 2–6.5 years. Digital fundus photographs were obtained in the last year of the study. Blood samples during the study were processed for inflammatory biomarkers, and these were correlated with obesity tertiles and presence of retinopathy. ResultsHigher BMI was associated with an increase in circulating levels of metabolic biomarkers including high sensitivity C-reactive protein (hsCRP), plasminogen activator inhibitor 1 (PAI-1), fibrinogen, LDL-cholesterol (LDL-C) and Apolipoprotein B (ApoB), tumor necrosis factor receptors 1 and 2 (TNFR-1 and -2), interleukin 6 (IL-6), E-selectin, and homocysteine, as well as a decrease in the metabolic risk markers HDL-cholesterol (HDLC), and insulin-like growth factor binding protein 1 (IGFBP-1). Although NPDR risk decreased with increasing obesity, it was not associated with any of the measured biomarkers. ConclusionsCirculating levels of measured biomarkers did not elucidate the “obesity paradox” of decreased NPDR in the most obese participants in the TODAY study.Trial Registration: clinicaltrials.govNCT00081328.

Evaluation of the longitudinal change in health behavior profiles across treatment groups in the TODAY clinical trial.

Individual health behaviors (ie, eating habits and sedentary lifestyle) are associated with type 2 diabetes (T2D). Health behavior profiles specific to adolescents with T2D have not been described. To identify health behavior profiles in adolescents with T2D and examine how these profiles change over time. Diet (via food frequency questionnaire) and activity behaviors (via 3-day physical activity recall) examined at baseline, 6 months, and 24 months from participants in the the Treatment Options for T2D in Adolescents and Youth (TODAY) study were used for this analysis. Latent profile analysis identified profiles of health behaviors within three time points, and latent transition probabilities were estimated to examine the change from baseline to 6 months (n = 450) and baseline to 24 months (n = 415). Multinomial logistic regressions were used to examine if the assigned TODAY treatment group (Metformin [Met], Met + Rosiglitazone [Rosi], or Met + Lifestyle) predicted change in health behavior profiles. Three profiles emerged: "most sedentary," "healthy eaters," and "active and eat most." At 6 months, 50% of males and 29% of females in the Met + Lifestyle treatment group improved in their health behavior profile. Among males only, the Met + Lifestyle treatment group were more likely to improve their profiles from baseline to 6 months (P = .01). Three health behavior profiles emerged and shifted over time. A high quality, lifestyle intervention had little effect on improving health behavior profiles. Optimizing outcomes in youth with T2D might require more robust and multifaceted interventions beyond family-level lifestyle, including more extensive psychosocial intervention, novel medication regimen, or bariatric surgery.

Adherence to multiple medications in the TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) cohort: effect of additional medications on adherence to primary diabetes medication.

Background Non-adherence to diabetes medication leads to poor outcomes and increased healthcare costs. Multiple factors affecting adherence in adults with type 2 diabetes (T2D) have been identified, but pediatric data is sparse. We aimed to determine whether initiation of additional oral medications or insulin affects adherence to primary study medication (PSM) in the Treatment Options for type 2 Diabetes in Adolescents and Youth (TODAY) study. Methods Six hundred and ninety-nine youth (aged 10-17 years) with recent-onset T2D were randomized in the TODAY study. Participants were categorized as adherent (≥80% taken by pill count) or non-adherent (<80%), and adherence was compared between those on additional medications or not. Subgroup analyses to assess influence of race/ethnicity, gender, medication type, or depression were performed. Results At 36 months, 46.3% of participants were taking additional oral medications and 31.9% were on insulin. There was no difference in study medication adherence with additional oral medications (55.1%, 67.1%, and 56.7% at month 36 in those prescribed 0, 1, or 2+ additional medications; p = 0.16). Girls on oral contraceptives (OC) had higher adherence (65.2% vs. 55.8% at month 36; p = 0.0054). Participants on insulin had lower adherence (39.7% vs. 59.3% at 36 months; p < 0.0001). There was decreased adherence in participants with baseline depression (p = 0.008). Conclusions Additional oral medications did not influence adherence to diabetes medications in TODAY. Addition of insulin led to reduced adherence. In subgroup analyses, OC use was associated with higher adherence in girls, while baseline depression was associated with lower adherence overall. Further studies examining potentially modifiable risk factors of adherence in pediatric T2D are needed.

The Shape of the Glucose Response Curve During an Oral Glucose Tolerance Test: Forerunner of Heightened Glycemic Failure Rates and Accelerated Decline in β-Cell Function in TODAY.

Obese youth without diabetes with monophasic oral glucose tolerance test (OGTT) glucose response curves have lower insulin sensitivity and impaired β-cell function compared with those with biphasic curves. The OGTT glucose response curve has not been studied in youth-onset type 2 diabetes. Here we test the hypothesis that the OGTT glucose response curve at randomization in youth in the TODAY (Treatment Options for Type 2 Diabetes in Adolescents and Youth) study forecasts heightened glycemic failure rates and accelerated decline in β-cell function. OGTTs (n = 662) performed at randomization were categorized as monophasic, biphasic, or incessant increase. Demographics, insulin sensitivity (1/fasting insulin), C-peptide index (△C30/△G30), and β-cell function relative to insulin sensitivity (oral disposition index [oDI]) were compared among the three groups. At randomization, 21.7% had incessant increase, 68.6% monophasic, and 9.7% biphasic glucose response curves. The incessant increase group had similar insulin sensitivity but significantly lower C-peptide index and lower oDI, despite similar diabetes duration, compared with the other two groups. Glycemic failure rates were higher in the incessant increase group (58.3%) versus the monophasic group (42.3%) versus the biphasic group (39.1%) (P < 0.0001). The 6-month decline in C-peptide index (32.8% vs. 18.1% vs. 13.2%) and oDI (32.2% vs. 11.6% vs. 9.1%) was greatest in incessant increase versus monophasic and biphasic with no difference in insulin sensitivity. In the TODAY study cohort, an incessant increase in the OGTT glucose response curve at randomization reflects reduced β-cell function and foretells increased glycemic failure rates with accelerated deterioration in β-cell function independent of diabetes duration and treatment assignment compared with monophasic and biphasic curves. The shape of the OGTT glucose response curve could be a metabolic biomarker prognosticating the response to therapy in youth with type 2 diabetes.

Cardiac Biomarkers in Youth with Type 2 Diabetes Mellitus: Results from the TODAY Study

To examine cardiac biomarkers over time in youth-onset type 2 diabetes, and relate serum concentrations to cardiovascular disease risk factors, and left ventricular structure and function. TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) was a multicenter randomized trial of 3 treatments including 521 participants with type 2 diabetes, aged 10-17 years, and with 2-6 years of follow-up. Participants were 36% male, obese, and ethnically diverse. Annual serum concentrations of brain natriuretic peptide, troponin, tumor necrosis factor (TNF)-α, receptors 1 and 2 were related to blood pressure, body mass index, hemoglobin A1c, and left ventricular ejection fraction, diastolic function, relative wall thickness, and mass. Elevated concentrations of brain natriuretic peptide (≥100 pg/mL), TNF-α (≥5.6 pg/mL) and troponin (≥0.01 ng/mL), were present in 17.8%, 18.3%, and 34.2% of the cohort, respectively, at baseline, and in 15.4%, 17.1%, and 31.1% at the end of the study, with wide variability over time, without persistence in individuals or clear relationship to glycemia or cardiovascular structure/function. TNF receptors concentrations were increased at baseline and not significantly different from end-of-study concentrations. Adverse echocardiographic measures were more likely in the highest TNF receptor tertile (all P < .05): higher left ventricular mass (39.3 ± 9.0 g/m2.7), left atrial internal dimension (3.7 ± 0.4 cm) and E/Em ratio, a measure of diastolic dysfunction (6.2 ± 1.9). After adjustment for body mass index, these relationships were no longer significant. Elevated serum concentrations of cardiac biomarkers were common in youth with type 2 diabetes, but their clinical significance is unclear and will require further long-term study. ClinicalTrials.govNCT00081328.

Insulin Sensitivity and Diabetic Kidney Disease in Children and Adolescents With Type 2 Diabetes: An Observational Analysis of Data From the TODAY Clinical Trial

Diabetic kidney disease is a major cause of premature mortality in type 2 diabetes mellitus (T2DM). Worsening insulin sensitivity independent of glycemic control may contribute to the development of diabetic kidney disease. We investigated the longitudinal association of insulin sensitivity with hyperfiltration and increased albumin excretion in adolescents with T2DM. Observational prospective cohort study. 532 TODAY (Treatment Options for Type 2 Diabetes in Adolescents and Youth) participants aged 12 to 17 years with T2DM duration less than 2 years at baseline. The TODAY Study was a multicenter randomized clinical trial that examined the efficacy of 3 treatment regimens (metformin monotherapy, metformin plus rosiglitazone, or metformin plus an intensive lifestyle intervention program) to achieve durable glycemic control. Natural log-transformed estimated insulin sensitivity (reciprocal of fasting insulin), hemoglobin A1c concentration, age, race-ethnicity, treatment group, body mass index, loss of glycemic control, and hypertension. Hyperfiltration was defined as 99th percentile or higher of estimated glomerular filtration rate (≥140mL/min/1.73m2) when referenced to healthy adolescents (NHANES 1999-2002) and albumin-creatinine ratio≥ 30μg/mg at 3 consecutive annual visits. Hyperfiltration was observed in 7.0% of participants at baseline and in 13.3% by 5 years, with a cumulative incidence of 5.0% over 5 years. The prevalence of increased albumin excretion was 6% at baseline and 18% by 5 years, with a cumulative incidence of 13.4%. There was an 8% increase in risk for hyperfiltration per 10% lower estimated insulin sensitivity in unadjusted and adjusted models (P=0.01). Increased albumin excretion was associated with hemoglobin A1c concentration, but not estimated insulin sensitivity. Longer follow-up is needed to capture the transition from hyperfiltration to rapid glomerular filtration rate decline in youth-onset T2DM. Lower estimated insulin sensitivity was associated with risk for hyperfiltration over time, whereas increased albumin excretion was associated with hyperglycemia in youth-onset T2DM.

Structure to utilize interventionists' implementation experiences of a family-based behavioral weight management program to enhance the dissemination of the standardized intervention: The TODAY study.

Background For a 2- to 6-year period, interventionists for the TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) randomized clinical trial delivered a family-based, behavioral weight-loss program (the TODAY Lifestyle Program) to 234 youth with type 2 diabetes. Interventionists held at least a bachelor's degree in psychology, social work, education, or health-related field and had experience working with children and families, especially from diverse ethnic and socioeconomic backgrounds. This article describes the administrative and organizational structure of the lifestyle program and how the structure facilitated collaboration among study leadership and lifestyle interventionists on the tailoring of the program to best suit the needs of the trial's diverse patient population. Methods During the pilot phase and throughout the duration of the trial, the interventionists' experiences in delivering the intervention were collected in a variety of ways including membership on study committees, survey responses, session audio recordings, and feedback during in-person trainings. Results The experiences of interventionists conveyed to study leadership through these channels resulted in decisions to tailor the lifestyle intervention's delivery location and ways to supplement the standardized educational materials to better address the needs of a diverse patient population. Conclusion The methods used within the TODAY study to encourage and utilize interventionists' experiences while implementing the lifestyle program may be useful to the design of future multi-site, clinical trials seeking to tailor behavioral interventions in a standardized, and culturally and developmentally sensitive manner.

Correlates of Medication Adherence in the TODAY Cohort of Youth With Type 2 Diabetes.

To identify factors that predict medication adherence and to examine relationships among adherence, glycemic control, and indices of insulin action in TODAY (Treatment Options for Type 2 Diabetes in Adolescents and Youth). A total of 699 youth 10-17 years old with recent-onset type 2 diabetes and ≥80% adherence to metformin therapy for ≥8 weeks during a run-in period were randomized to receive one of three treatments. Participants took two study pills twice daily. Adherence was calculated by pill count from blister packs returned at visits. High adherence was defined as taking ≥80% of medication; low adherence was defined as taking <80% of medication. Depressive symptoms, insulin sensitivity (1/fasting insulin), insulinogenic index, and oral disposition index (oDI) were measured. Survival analysis examined the relationship between medication adherence and loss of glycemic control. Generalized linear mixed models analyzed trends in adherence over time. In this low socioeconomic cohort, high and low adherence did not differ by sex, age, family income, parental education, or treatment group. Adherence declined over time (72% high adherence at 2 months, 56% adherence at 48 months, P < 0.0001). A greater percentage of participants with low adherence had clinically significant depressive symptoms at baseline (18% vs. 12%, P = 0.0415). No adherence threshold predicted the loss of glycemic control. Longitudinally, participants with high adherence had significantly greater insulin sensitivity and oDI than those with low adherence. In the cohort, the presence of baseline clinically significant depressive symptoms was associated with subsequent lower adherence. Medication adherence was positively associated with insulin sensitivity and oDI, but, because of disease progression, adherence did not predict long-term treatment success.

Parenting style, parent−youth conflict, and medication adherence in youth with type 2 diabetes participating in an intensive lifestyle change intervention.

Parenting behaviors and family conflict relate to type 1 diabetes outcomes in youth. Our purpose was to understand these relationships in parents and youth with type 2 diabetes (T2DM). The TODAY (Treatment Options for Type 2 Diabetes in Adolescents and Youth) trial enrolled youth (10-17 years) with T2DM and parent/guardian. For this ancillary study, we enrolled a sample of youth-parent pairs (N = 137) in 1 study arm (metformin plus lifestyle intervention). They completed questionnaires measuring parenting style related to normative (e.g., completing homework) and diabetes self-care (e.g., testing blood glucose) tasks, and parent-youth verbal conflict (baseline, 6, and 12 months). Parenting style was consistent across normative and diabetes tasks, with gradual increases in autonomy perceived by youth. Conversations were generally calm, with greater conflict regarding normative than diabetes tasks at baseline (youth: p < .001, parent: p = .01), 6 months (youth: p = .02, parent: p > .05), and 12 months (youth: p > .05., parent: p = .05). A permissive parenting style toward normative tasks and a less authoritarian style toward diabetes tasks, at baseline, predicted better medication adherence (8-12 months) (normative: adjusted R2 = 0.48, p < .001; diabetes: adjusted R2 = 0.47, p < .001). Parent-youth conflict did not predict medication adherence. Youth with T2DM who perceive more autonomy (less parental control) in day-to-day and diabetes tasks are more likely to adhere to medication regimens. It may be valuable to assess youth perceptions of parenting style and help parents understand youths' needs for autonomy.

“TODAY” Reflects on the Changing “Faces” of Type 2 Diabetes

The diabetes landscape has seen unprecedented changes over the recent past. On the one hand, there have been consistent and impressive advancements in clinical care, which include new therapeutic agents, novel dietary approaches, technical advances in glycemic monitoring and closed-loop technology, and a heightened awareness of individuals at risk. The tools required to treat and halt the progression of diabetes complications and the clinical evidence in support of effective management strategies have been established. On the other hand, the need for these developments has never been greater given the global burden of the diabetes epidemic. A particularly disturbing observation is that the “faces” of those who develop type 2 diabetes are becoming younger by the year as evidenced by the reports demonstrating the increased frequency of both type 1 and type 2 diabetes in youth (1–3). Until recent data were made available, it was really not known how adolescents would respond to therapies normally reserved for the adult population. We also had no evidence regarding the rate and severity of the complications in this age-group or the prevalence and progression of other risk factors. In this regard, the TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) study has succeeded in filling a major gap in knowledge (4–10). Thus, given the importance of the problem of type 2 diabetes in youth and the need to effectively disseminate the information, our editorial team has elected to feature the TODAY study in this issue of Diabetes Care . Specifically, articles from the TODAY study researchers featured in this issue provide new data on the efficacy and safety of clinical treatment and longitudinal observations of specific risk factors and complications for the new “faces” of the type 2 diabetes epidemic—namely, adolescents! As I reflect on the problem …

Crisis in Care: Limited Treatment Options for Type 2 Diabetes in Adolescents and Youth

Until two decades ago, children and adolescents were automatically assumed to have insulin-dependent type 1 diabetes. However, type 2 diabetes emerged as a “new type” of childhood diabetes in the 1990s in association with the epidemic of childhood obesity. It quickly became apparent that this new pediatric disease disproportionally affected disadvantaged minority children and was associated with comorbidities that increased the risk of future cardiovascular disease. After more than 20 years, the optimal approach to the treatment of childhood type 2 diabetes remains largely unknown. Besides insulin, metformin remains the only other antidiabetic medication that is approved by the U.S. Food and Drug Administration (FDA) and the European Medicines Agency (EMA) for use in youth with type 2 diabetes. Glimepiride and rosiglitazone failed noninferiority tests versus metformin as initial monotherapy in company-sponsored clinical trials. While the primary study results of the randomized phase of the TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) study showed that combination therapy with metformin plus rosiglitazone was more effective than metformin plus intensive lifestyle intervention and metformin alone (1), rosiglitazone will not be used with any frequency in young patients with type 2 diabetes because of concerns about the cardiovascular and other adverse effects of this class of medications. Once again, pediatric diabetes practitioners are left with just metformin and insulin for adolescents with type 2 diabetes. Why haven’t glucagon-like peptide 1 (GLP-1) agonists and dipeptidyl peptidase 4 (DPP-4) inhibitors that have been approved for use in adults with type 2 diabetes been approved for the treatment of adolescents …

Learning Needs of Youth With Type 2 Diabetes

The purpose of this study was to examine reports of diabetes learning needs by youth with type 2 diabetes and their family support person (FSP) and to examine correlations of these perceptions with demographic variables. Data were analyzed from the Participant Needs Assessment (PNA), a routinely self-administered questionnaire designed to assess learning needs, in 191 youth (mean age = 14.0 ± 1.9 years, duration 8.78 ± 6.30 months; 36% black, non-Hispanic; 40% Hispanic; 20% white) enrolled in the TODAY (Treatment Options for Type 2 Diabetes in Adolescents and Youth) study. Youth and their FSPs completed the PNA at scheduled study visits, rating their learning needs on a 5-point scale, with higher scores indicating higher needs. Paired t tests and ANOVA were used. There were no differences in reported learning needs by gender, although scores on the Teen Issues scale approached significance, with girls reporting higher needs than boys. Youth and FSPs were similar on their reports of needs on 3 scales (Ordinary Day, Nutrition, and Living With Diabetes), but youth reported higher needs than FSPs on Focus and Teen Issues. Controlling for duration of diabetes, these differences between youth and FSPs persisted. In general, learning needs for both youth and FSPs decreased over time. Youth with type 2 diabetes and their FSPs have high levels of ongoing learning needs despite completion of a standard diabetes education program. Continued assessment of learning needs, follow-up education, and behavioral approaches are warranted for these high-risk youth.

Metformin monotherapy in youth with recent onset type 2 diabetes: experience from the prerandomization run-in phase of the TODAY study.

TODAY (Treatment Options for type 2 Diabetes in Adolescents and Youth) is a federally funded multicenter randomized clinical trial comparing three treatments of youth onset type 2 diabetes. To describe the experience of youth participating in a 2-6 month run-in period in preparation for randomization into TODAY. An ethnically diverse sample of 927 youth, 65.4% females, aged 13.7 ± 2.0 yr old, with type 2 diabetes for a median of 2 months (0.7-7.8 months, 25th-75th percentiles). A run-in period was conducted to achieve HbA1c <8% with metformin monotherapy and diabetes education, and to evaluate adherence to pill taking, visit attendance, and other procedures. At entry, mean body mass index (BMI) and z-BMI were 35.6 ± 7.7 and 2.3 ± 0.4, respectively, mean HbA1c was 7.7 ± 2.2%, only 42.5% were on a hypoglycemic treatment, and 35.6% had HbA1c ≥8%. Comorbid conditions were common; 18.8% had hypertension, 24.2% had elevated cholesterol, and 6.5% had abnormal liver enzymes. After a median 71 d of run-in, 90.9% had HbA1c <8%, 77.9% had HbA1c <7%, and 46.4% had HbA1c <6%. Of the 772 youth achieving the target HbA1c <8%, 704 (91.2%) were randomized; non-adherence to metformin treatment was the main cause for non-randomization. Youth proceeding to randomization decreased weight by 0.68 kg and HbA1c by 1.45% compared to a weight gain of 0.71 kg and HbA1c decrease of 0.74% in the non-randomized youth (p = 0.01 in both cases). However, change in z-BMI was not significantly different between the two groups. Most youth with recent onset type 2 diabetes can achieve target HbA1c <8.0% with short-term metformin monotherapy and standard diabetes education (ClinicalTrials.gov identifier: NCT00081328).