This open pilot RCT will enroll healthy children aged 12 to 42 months coming to the Odonto-stomatology Unit at Bambino Gesù Children's Hospital and Research Institute (OBG) with their parents, who seek help in stopping their children continue the NNS habit. This RCT will last 3 years. In the first year, patient enrollment will be completed, and the analyses needed to measure the primary and secondary outcomes conducted. The RCT will test the efficacy of NNS in improving balance and gait after a two-year follow-up. Patient registries will follow internal ad hoc OBG procedures for good-clinical practice (GCP) in trial conduction (Prot. 01.01.99 published in the OBG intranet site on 5th April 2017) including patients' data registered in case report forms (CRF) in electronic and paper records. After children's parents give their consent signing the information sheet approved by the OBG Institutional Review Board (OBG-IRB), namely Comitato Etico, every eligible child will be coded with a computerized randomization code and identified in the CRF with this code, their name, surname and date of birth (RCT protocol approved on 10th September 2018, OBG-IRB Prot. N. 960). The RCT includes two parallel arms, the intervention arm and the control arm. Children in the intervention arm will suck a pacifier of identical shape and type produced by Artsana S.P.A., and will be allowed other NNS habits (finger-thumb). Children in the control arm will be induced by the OBG odonto-stomatologists (DDSs) to stop sucking and their parents will receive from the DDSs an ad hoc information sheet advising them how to stop their children's NNS habit. Children in both arms will undergo a computerized photometric postural balancing measure and gait control assessment, done by a physiotherapist (PT) delegated by the PI, every 6 months over 24 months at the Neuro-rehabilitation Unit in the OBG site at Palidoro (Rome). After enrollment, children in both arms will also undergo an electroencephalogram (EEG) while sucking, done by an EEG technician (EEG tech) delegated by the PI. Two years later, children will undergo a second EEG. Parents will report in a diary their children's time spent on pacifier or finger-thumb sucking, crying time, afternoon- and night-time sleep induction, awakenings, and daily sleeping time duration. Every 6 months, parents will bring their children and diaries to the DDSs. DDSs will visit the child and report data from oral examination findings and from parents' diaries in hospital electronic medical records and paper CRF. In accordance with the European Medicine Agency (EMA), Committee for Medicinal Products for Human Use (CHMP), International Conference on Harmonization (ICH) guidelines for GCP(EMA/CHMP/ICH/135/1995), Italian Ministry of Health law (15th July 1997) and the OBG-IRB approved RCT protocol, the OBG (acting as the Promoter for this clinical trial) and the PI are responsible for the quality assurance plan including all the enrolled children's data validation and patient registry. The PT, DDSs and the EEG tech will record also the results in an ad hoc sheet. All the variables included in the PT evaluation, orthognathic findings, parents' diary and EEGs will be recorded in an Excel file by the PT, DDSs and the EEG tech delegated by the PI. The PI is responsible for comparing and validating the data recorded and the data registered in an Excel file for the statistical analysis.
The postural balance and gait assessment, oral diagnostic examinations and EEG results detailed in every electronic and paper CRF for the children enrolled will be reviewed without undue delay by the research team including the PI, DDS members, a PT and an EEG tech delegated by the PI. These personnel will check concordance between the results and source data (electronic medical records). The PI will check, sign and date the signature on the paper CRF. If the results concern the patient's safety, the research team will inform the PI. In accordance with the OBG hospital's internal GCP protocol (Prot. OBG 01.01.11), the PI will notify adverse events to the Health Management Board and OBG-IRB. The research team will be responsible for creating a data dictionary that contains detailed descriptions of each variable used in the RCT study and CRF, including the source of the variable, coding information used and relevant ranges for outcome measures. Following the standard operating GCP procedures approved by the OBG promoter (Prot. OBG 01.01.11 and 01.01.99), the PI will archive all the documents including authorizations, contracts and all the essential documents in a trial master file in electronic and paper versions. The PI and delegate collaborators will be responsible for maintaining the CRF trace in electronic and paper forms as a locked archive in the Epidemiology Unit where the PI works. According to the internal OBG GCP procedures and always in the presence of the PI, the OBG-IRB will monitor the CRF and all documentation archived with independent monitoring activities on-site, so as to survey and assess periodically this RCT protocol and study conduction. These on-site audits will ensure that all activities related to the RCT protocol and all other data will be registered, analyzed and accurately maintained in accordance with the OBG GCP standard operating procedures specified in the foregoing. The PI will also record in the CRF every change in trial management and update the protocol in the clinicaltrial.gov website. As requested by the Italian Drug Agency, Agenzia Italiana Farmaco (AIFA), the European Medical Agency (EMA) and Food and Drug Administration (FDA), all procedures and their documentation referred to in this protocol will be made available to inspection. As specified in the aforementioned OBG GCP protocols, the electronic and paper investigator file including all the trial documentations will be kept for at least 15 years.
Sample size - Given the lack of studies investigating NNS efficacy for the primary outcome (evaluating the proprioceptive and exteroceptive benefits of NNS), to define the sample size, we analyzed the data for the incidence of instinctive NNS from all the children brought by their parents to the Odonto-stomatology Unit at Bambino Gesù Children's Hospital to investigate prolonged NNS during the year 2015. The power analysis, based on these data, estimated that 4000 children in the 12 to 42 months age group will yield an incidence rate of 2% of children (n = 80) who come for their first dental visit because of prolonged NNS. Estimating that gait (width, frequency and speed of steps, cadence, step/stride length, step/stance time, and number of falls during walking along a walkway that records spatio-temporal features) will improve in 30% of the children in the interventional arm and no children in the control arm, a study sample of 50 children will be needed to obtain a significant X2 test with α 0.05 and β of about 0.20. Assuming that 15% of the children enrolled will be lost to follow-up, sample size calculations estimate that the study needs to enroll 64 children (32 in each arm) to demonstrate a significant effect. Every 6 months after enrollment, the 64 children will attend for their dental visits. At one year after enrollment, it is estimated that reduced overbite and increased overjet, with or without stopping NNS habits, will be detected in 60 to 70% of children. It is also estimated that overbite, if present, will diminish every year by 3 mm in 40-56% of these children aged from 42 to 66 months, yielding an α of about 0.05 and β of about 0.20.
Data on outcome measures will be inserted in the RCT database progressively. Retrospective data for variables reported in the parents' diaries, or electronic and paper CRF will also be inserted in the RCT database. Missing data (unavailable, un-reported, uninterpretable or considered missing owing to data inconsistency or out-of-range results) in the RCT database will be retrieved whenever possible by the PI by interviewing parents or by re-evaluating outcome measures with PTs, DDSs or the EEG tech, and will be registered in electronic and paper CRF. Data remaining unavailable will be considered as missing.
Intellectual property and publications - Artsana S.P.A. and OBG (Promoter) signed a funding agreement (7 August 2018). They agreed that OBG is the sole owner of data and of direct and indirect RCT results. The PI is the only person responsible for the data entry and documentation archive and will follow the study as specified in the OBG-IRB protocol and its possible amendments, without disclosing any data analyses or results to Artsana S.P.A. before their publication. In future published papers, Artsana S.P.A. and OBG have agreed to state that "Artsana S.P.A. has helped to fund the RCT activities".
Practical implications and future developments - The findings obtained in this open pilot RCT should provide information on the proprioceptive, exteroceptive and muscle-relaxation benefits of NNS in children enrolled at 12 to 42 months of age and followed for 3 years (maximum age when the 3-year follow-up ends, 66 months = 5 years and 6 months). In all children follow-up will end before mixed dentition erupts.The information obtained from the EEG results should help in designing the following new research projects. For example, it will help in studies designed to identify and assess in children who prolong NNS, the pediatric population older than 36 months to enroll in a later study on diagnostic accuracy using functional magnetic resonance imaging (fMRI) without sedation (during NNS lasting at least 10 minutes) to define which neuroreceptor transmitters stimulate NNS. The fMRI findings should also help in developing a differentiated genomic analysis for girls and boys.
Statistical analysis plan - The statistical analysis will start by analyzing descriptive data for all the quantitative variables (means, medians and standard deviations) and for categorical data obtained from the electronic medical records and parents' diaries. Data frequencies and percentages will be calculated with STATA® 13 software. Various multivariate regression analyses will calculate the risk of malocclusion (Cox regression) considering as independent variables age at enrollment, sex, age at standing without support, postural balance, gait assessment and stratifying the RCT population according to possible malocclusion. The statistical significance of possible NNS-related changes in the initial malocclusion (reduced overbite and increased overjet) will be evaluated in children in whom NNS correlates with dental problems at 12, 24 and 36 months after enrollment. The statistical analysis will also assess the various types of malocclusions and correlation coefficients (r) for the psycho-social variables, including times for inducing and maintaining sleep and other variables collected in the database during a 48- to 66-month follow-up for the primary and secondary outcomes.
