Effectiveness of MI VarnishTM and PreviDentâ Varnish in Non-cavitated Interproximal Lesions: A Randomized Clinical Trial.

Background: the clinical investigations regarding the ecacy of different uoride products are limited. Aim: to compare the effectiveness of Colgateâ PreviDentâ and MI TM varnishes to the standard (1.23%) acidulated phosphate uoride gel in the remineralization of non-cavitated proximal incipient lesions. Design: this randomized clinical trial included 91 lesions which were assigned to three groups. We performed an initial examination, and 3- and 6-month follow-ups. Result: teeth treated with both MI TM and Colgateâ PreviDentâ varnishes showed statistically signicant improvements in caries progression. In the Colgateâ PreviDentâ group, nine surfaces with white spots with dryness remained unchanged, one surface changed to a white spot without dryness, and one surface improved to a sound surface. In the MI TM group, nine surfaces with white spots with dryness remained unchanged, and one surface changed to a white spot without dryness. Only teeth treated with MI varnish TM showed signicant improvements radiographically. Teeth affected by outer enamel caries remained the same or improved to sound surfaces. Additionally, teeth with inner enamel caries remained the same. Conclusion: both MI TM and Colgateâ PreviDentâ varnishes are effective therapies to remineralize non-cavitated incipient lesions. However, there were no signicant differences in the radiographic outcomes among the three types of varnish applications.


Background
Dental caries is the most prevalent chronic disease that affects all populations and has become a pandemic. 1 In Saudi Arabia, the prevalence of dental caries is approximately 80% in primary teeth and 70% in permanent teeth. 2 Tooth decay is a multifactorial disease; the primary etiological factors are the presence of fermentable carbohydrates, acid-producing bacteria, a susceptible tooth surface, and time. 3 The principal causative bacteria associated with dental caries are Streptococcus sp. (Streptococcus mutans and Streptococcus sobrinus). 3 Dental caries is the result of variations in pH caused by bacterial organisms in the bio lm, leading to demineralization of the dental hard tissues. 4,5 Caries can profoundly affect children's quality of life due to pain that could progress to acute and chronic infections, altered sleeping and eating habits, as well as high treatment cost and loss of school days. Caries management should focus on minimum intervention and maximum prevention. 6 Incipient carious lesions are areas showing early features of tooth decay. Indeed, they can reverse, be arrested, or progress to cavitation.
Such early lesions can remineralize if diagnosed early and treated at the appropriate time. 6,7 Therefore, an adequate home-care regimen and accurate clinical intervention are essential to initiate the remineralization process. Fluoride treatment has been the foundation of non-invasive dental treatment for incipient caries. 5 Low levels of uoride inhibit the demineralization of sound enamel and initiate the remineralization of demineralized enamel. Additionally, uoride affects bacterial metabolism and decreases the ability of cariogenic bacteria to produce acid. [8][9][10] When the oral pH drops, high levels of uoride result in the temporary formation of a calcium-uoride-like material on the tooth surface that enhances the uptake of calcium and uoride by the hydroxyapatite crystals, forming uorapatite. 10 Fluorapatite is more acid resistant than hydroxyapatite. 8,9 Different types of uoride treatments are used in various applications, concentrations, durations, and frequencies. Newer products are continuously evolving, but the evidence for their e cacy is insu cient. MI varnish™ with RECALDENT, which consists of casein phosphopeptide-amorphous calcium phosphate (CCP-ACP), is recommended for the treatment of white-spot lesions in orthodontic patients. The active ingredients are CCP-ACP and 5% NaF (22,600ppm uoride), which show a synergistic effect and a deeper remineralization capability. Colgate→ PreviDent varnish is another product with 5% NaF (22,600-ppm uoride) and xylitol as active ingredients. Most of these new products have not been su ciently studied to recommend their use to the general public. Our objective was to compare the effectiveness of Colgate PreviDent and MI varnish™ with the standard 1.23% acidulated phosphate uoride (APF) gel in the remineralization of non-cavitated proximal incipient lesions.

Ethical considerations and registrations
This clinical trial was registered at ClinicalTrials.gov (no. NCT03925740), and and the rst registration date was on 24/04/2019. Ethical approval was obtained from the institutional ethical committee at King Abdulaziz University (REC-013-01019). The study started in January 2019 at King Abdulaziz University Dental Hospital (KAUDH) in Jeddah. Informed consents were signed by the parents or guardians of all patients. All methods were carried out in accordance to the ethical committee at King Abdulaziz University guidelines and regulations.

Study design
This double-blind, randomized-clinical trial was conducted in three arms. The patients were allocated to one of the three groups to compare the e cacy of two different uoride varnishes with a standard uoride gel. The examiners were blinded during the clinical and radiographic assessment, and the patients and their parents were not aware of the patient's allocation group. However, blinding of the clinicians during the application of the material was not possible due to the different packaging of the materials.

Sample size calculation
The minimum sample size was calculated at www.openepi.com. Ninety-one incipient carious lesionswere considered su cient for this study. Patients were recruited from the postgraduate and undergraduate clinics of XXX.

Random allocation and blinding
The three arms were de ned as the Colgateâ PreviDentâ, MI varnish TM , and control (APF gel) groups.
Patients were randomly allocated to the Colgateâ PreviDentâ and MI varnish TM groups by a coin toss.
After obtaining the consents and answers to the questionnaire, the examiners assessed the lesions clinically and radiographically using the International Caries Detection and Assessment System (ICDAS) criteria, and each lesion was scored before random allocation. Randomization was performed separately after the assessment of each lesion. 11 A two-digit code was allotted to each patient; the rst digit represented the patient's serial number, and the second digit represented the group. At the 6-month followup, radiographic re-assessment with bitewing and periapical radiographs was performed. We identi ed each radiograph with the patient's two-digit code along with the medical record number. After the radiographs of all patients were obtained, they were mixed together before interpretation to ensure blindness. Two trained and calibrated general dentists performed the randomization and assessment.
Inter-and intra-examiner reliability were assessed at two different time points with 90% agreement and a kappa score of 0.613 (substantial).

Patient selection:
A total of 18 children (6 to 15 years old) who sought dental treatment inKAUDH were evaluated for inclusion in the study according to the inclusion/exclusion criteria.
Inclusion criteria: The inclusion criteria were as follows: anterior or posterior, primary or permanent teeth with proximal incipient caries; clinical and radiographic ICDAS score of 1 or 2; and lesions detected by visual and tactile examination with the aid of ample light and mouth mirrors and probes, as well as by bitewing radiographs for posterior teeth and periapical for anterior teeth. 11 Exclusion criteria: The exclusion criteria were as follows: presence of active initial carious lesions (ICDAS scores ≥ 3) or deep caries crossing the dentinoenamel junction on bitewing radiographs 11 ; and chronic medical conditions and negative dental behavior. Children who presented with other needs for dental treatment were referred to complete their treatment. After excluding patients according to the inclusion and exclusion criteria, 18 patients (91 lesions) were included in the study (Fig. 1).

5% NaF MI varnish TM and 5% NaF Colgateâ PreviDentâ groups
Patients were randomly assigned to two groups (MI varnish TM or Colgateâ PreviDentâ) by a coin toss after we obtained the consent of their parents/guardians. We assigned three patients with 30 lesions to the MI varnish TM group and ve patients with 33 lesions to the Colgateâ PreviDentâ group. Oral hygiene assessment was performed using the Greene and Vermillion oral hygiene index. 12 The decay-missinglled-by surface (DMFS) index was recorded. Oral prophylaxis was performed with plaque removal using a polishing brush attached to a low-speed handpiece and dental oss. Orthodontic separators or wedges were placed between the teeth with incipient lesions on X-rays, and the patient was asked to return the next day. The teeth were dried, and the material (according to the group of the particular patient) was directly applied in the interproximal areas with the incipient lesions and then to the rest of the teeth. Patients were instructed to not rinse or drink water for 30 minutes and to avoid hard and sticky foods and eat only soft foods for the next 2 hours, according to the manufacturer's instructions. Detailed oral hygiene instructions, including brushing twice with a uoridated toothpaste (1100 ppm) and ossing the site of the lesion with waxed dental oss, and an oral hygiene checklist were provided. The next follow-up visit was scheduled after 3 months.

1.23% APF gel (control) group
Ten patients (28 lesions) were recruited by scanning dental records, including radiographs of patients who were treated in the undergraduate clinics. APF application was performed using the tray technique as follows: The patient was seated in an upright position. After plaque removal with a polishing brush attached to a low-speed handpiece, an APF gel was dispensed on a disposable foam tray no more than 1/3 full, according to the manufacturer's instructions. The patient was instructed to not swallow the gel and exert slight pressure using the checks and tongue, as well as light biting force to allow the gel to ow interproximally for 4 minutes. A saliva ejector was used for salivary isolation and removal of excess gel.
The patient was instructed to not eat, drink, or rinse for at least 30 minutes.

Follow-up visits for the Colgateâ PreviDentâ and MI varnish TM groups
The rst follow-up visit was scheduled after 3 months (T1) and the second follow-up visit after 6 months (T2), according to the Caries Management by Risk Assessment criteria (CAMBRA). 13 At T1, scores for the oral hygiene index were determined, oral prophylaxis, including plaque removal, was performed, and orthodontic wedges or separators were placed interproximally. The patients were recalled the next day.
Thereafter, orthodontic wedges or separators were removed, teeth were dried, and the material was applied interproximally to the selected lesion and then to the rest of the teeth. The same instructions as T0 were provided to the patients and their parents. Oral hygiene instructions were reinforced, and another follow-up visit was scheduled after 3 months. At T1, no loss to follow-up was recorded for either group. We had two discontinued interventions in the Colgateâ PreviDentâ group because of a misdiagnosis by the physician (they had been restored by composite restoration) (Fig. 1).
At the 6-month visit, the same procedure was performed. Additionally, bitewing and periapical radiographs were obtained to assess the lesions. At T2, no loss to follow-up or discontinued intervention was recorded for either group (Fig. 1).

Follow-up visit for the APF gel (control) group
At the 6-month visit, the same procedure was performed as for the other two groups. No T1 visit was scheduled for patients in this group, since this was the protocol followed at the undergraduate dental clinics. At T2, no loss to follow-up or discontinued intervention was recorded for this group (Fig. 1).
Outcome assessment criteria: The primary outcome was to evaluate the caries lesion progression clinically and radiographically using direct visual examination and ICDAS scores at baseline and comparing them after 6 months. Clinical and radiographic success was considered as maintenance of the caries score after 6 months or decreased to sound surfaces free of caries.
Statistical analysis: Statistical analysis was performed using the SPSS version 20 software (Armonk, NY: IBM Crop). Descriptive statistics were displayed as frequencies and percentages for categorical variables and mean and standard deviation (±) for continuous variables. The chi-square test was performed, with statistical signi cance at p<0.05, and the post-hoc Bonferroni correction was applied. The Fischer's exact test was used when cells consisted of fewer than ve cells.
After 6 months of follow-up, both MI varnish™ and Colgate→ PreviDent groups showed statistically signi cant improvements in ICDAS scores for incipient caries. In the PreviDent group, ve surfaces of teeth changed to from carious to sound. One of them was a white spot with dryness con ned to the outer enamel radiographically, while the rest were white spots without dryness con ned to the inner enamel radiographically. After we performed the Bonferroni correction to adjust the p-value for results of the chisquare test, nine surfaces (81.8%) with white spots accompanied by dryness remained unchanged, one surface changed to a white spot without dryness, and one surface changed to sound (Table 3). In the MI varnish™ group, four surfaces changed to sound. One of them was a white spot without dryness and con ned in the inner enamel radiographically, while the rest were white spots with dryness in the outer enamel radiographically.  (Table 4). Table 5 shows statistically signi cant differences in the clinical outcomes among the three groups of varnishes (p = 0.004). After the Bonferroni correction, teeth in the control group showed statistically signi cantly increased frequency of failures compared to teeth in the experimental groups (p < 0.05). However, there were no statistically signi cant differences in the radiographic outcomes among the three groups of varnishes.

(100)
Results are based on two-sided tests with a signi cance level of 0.05, using the Bonferroni correction. For each signi cant pair, the key of the category (A, B, or C).
**The frequency is signi cantly higher than the number under the category that appears next to it.
*p-value is signi cant at 0.05 ICDAS, International Caries Detection and Assessment System; APF, acidulated phosphate uoride

Discussion
The aim of this clinical trial was to validate a potential cost-effective treatment modality for incipient proximal carious lesions in children. Using dental oss to remove the bio lm from proximal areas is the simplest and most effective way to control caries progression. 14 Indeed, the method and frequency of ossing by the patients affects the outcome of caries management. 15 Furthermore, children's poor compliance to ossing renders the proximal areas di cult to clean. 16 Thus, early interventions for incipient carious lesions are essential to prevent cavitations. Fluoride treatments can remineralize incipient carious lesions, and uoride products containing CCP-ACP and xylitol may have an impact on the clinical and radiographic outcomes of incipient caries. In recent studies, CCP-ACP has shown promising outcomes in the treatment of incipient caries. [17][18][19][20][21][22] The addition of CCP-ACP to uoride has a synergistic effect. 22 However, some studies have found that the addition of CCP-ACP to uoride has no clinical bene t over uoride alone. [23][24][25][26][27] This notable discrepancy in the studies regarding the clinical relevance of CCP-ACP can be attributed to the study design, duration of use, differences in the severity and activity of the lesions, and the possible variations between orthodontic and non-orthodontic incipient carious lesions.
A previous systematic review showed that uorides, combined with CCP-ACP, have a clinical advantage over uoride monotherapy on the occlusal surface.
In contrast, uoride monotherapy may have the same effect on smooth surface lesions. 28 Our study showed that both MI varnish™ and Colgate→ PreviDent signi cantly improved the clinical ICDAS scores of incipient carious lesions. However, radiographically, only lesions treated with MI varnish™ showed signi cant improvements. Nonetheless, there were no signi cant differences in the radiographic outcomes among the three types of varnishes used. The results of this study support the previous systematic review, 28 since we also analyzed smooth surface lesions (proximal). Although there were no signi cant differences in the radiographic outcomes among the three types of varnishes, further longterm studies may be required to validate the results of this study. A study that evaluated the e cacy of a 3-month CCP-ACP application regime with a follow-up duration of 12 months showed that CCP-ACP is effective. 29 There are several limitations to this study. This randomized clinical trial included patients with incipient carious lesions and compared three uoride products. Hence, this evidence is not relevant to the population free of carious lesions. A major limitation was the control group, which we recruited from undergraduate clinics. Therefore, the initial clinical caries assessment was not performed by the same research examiners. Another limitation was the use of visual-tactile rather than instrumental diagnostic methods such as laser uorescence and quantitative light-induced uorescence. However, the use of these technologies in our multisite trial was not possible due to economic constraints.
Nonetheless, this randomized clinical trial has several strengths. The outcome is a parameter that is crucial to dentists and patients. Moreover, all groups were comparable at recruitment by randomization, and the results were consistent when adjusted for sex, age, and initial severity. Regarding assessment, a 6-month follow-up duration was chosen to include the time required to visualize signi cant radiographic improvements while minimizing loss of follow-up. A period of at least 6 months is required to show evident changes in caries regression. 30 All examiners and patients were blinded during the assessment.
Additionally, the patient compliance was high. No patient dropped out of the trial due to adverse effects such as allergy, in ammation of the gingiva, or plaque accumulation. We recommend a longer follow-up period and the use of quantitative light-induced uorescence in future studies to substantiate the results of this study.

Conclusions
Prevention of dental caries is vital to preserve natural teeth in dental practice. Incipient carious lesions treated with both MI varnish TM and Colgateâ PreviDentâ showed statistically signi cant improvements in clinical ICDAS scores. However, radiographically, only teeth treated with MI varnish TM showed signi cant improvements. Nonetheless, there were no signi cant differences in the radiographic outcomes among the three types of varnishes used. Previous studies have reported that CPP-ACP has a synergistic effect when added to uoride, and this randomized controlled trial validated this nding. This ongoing registered trial has many future prospects, including longer follow-up periods and larger sample sizes. This study provides preliminary evidence to support the paradigm shift in pediatric dentistry from standard restorative treatment to disease prevention and conservation of tooth structure.