This was a multicentre prospective controlled observational cohort study adapted from the Dutch protocol used from the study of safety and immunogenicity of the bivalent HPV vaccine in children with Juvenile idiopathic arthritis (68 patients), childhood systemic lupus erythematous (6 patients) and JDM (6 patients) with a real-world approach (11, 12).
In the present study a 3-dose schedule (0, 1 or 2, and 6 months) of the qHPV vaccine (against HPV6, HPV11, HPV16, HPV18) was used in patients who met the Bohan and Peter’s criteria for JDM (13), from 9 to 20 years old, and age-matched healthy controls (HC). The doses of the qHPV vaccine used in the study were received by donation from the local Special Immunobiological Reference Centers of NIP.
Participants who were eligible and willing to receive the qHPV vaccine were enrolled in the study from March 2014 until March 2016. Moreover, JDM patients who had already received one or two doses of the qHPV vaccine before inclusion were also allowed to participate, as a care standard to reach three doses, which is indicated for immunosuppressed patients (14), since this is a real-life study.
Patients were recruited in 10 pediatric rheumatology units from tertiary centres of different Brazilian regions. Patients were selected regardless of medication used to constitute a real-life setting. HCs were recruited from patient peer groups in two study sites. The protocol under the code U1111-1211-2150, was approved by all the local ethics committees and informed consent was obtained from each participant and their guardians. Study visits were planned before the first dose, one month after the second and third doses, and one year after the starting dose.
Main Outcome Measures
For safety evaluation of the qHPV vaccine, participants were asked to complete a diary for 14 days after each dose, about the occurrence of possible local and/or systemic adverse events following vaccination (AEFV). The local AEFV addressed included redness, bruising, edema, induration, and pain. Systemic AEFV included fever, skin abnormalities, itchiness, headache, nausea, vomiting, fatigue, fainting, and muscular and articular pain.
Another outcome considered for safety evaluation in JDM patients included the assessment of disease activity at each study visit, using muscular and cutaneous parameters. For muscular evaluation the Childhood Myositis Activity Score (CMAS) and Manual Muscle Testing (MMT) were used. The CMAS ranges from 0 (high disease activity) to 52 (no disease activity) (15), and the MMT ranges from 0 (high disease activity) to 80 (no disease activity) (16). To verify whether disease activity had changed post vaccination, the CMAS and MMT values were compared between visits, considering as stable disease when the scores changed less than 20%; worsening if the scores decreased at least 20%; and improvement when both scores increased at least 20%.
The most usual cutaneous manifestations of JDM, cutaneous rash, heliotrope of the upper eyelids and Gottron papules, were evaluated in each visit according to their intensity by the same Pediatric rheumatologist and compared as follow: improvement, if the manifestation has subsided; stable, if it remained unchanged; worsening, if it had aggravated.
Comparison of the medications in use at each visit, which can indirectly quantity disease activity intensity, was used as an additional parameter, as follows: stable disease, if the medication remained the same between the visits; improvement if it had been withdrawn and worsening if a new treatment had been added or if previous treatment doses were increased.
In order to compare the changes in the measured values, considering CMAS and MMT scores, cutaneous manifestations, and use of medications, according to the established criteria already specified, three comparisons were made: Comparison 1: after two doses versus baseline; Comparison 2: after three doses versus baseline; and Comparison 3: after three doses versus after two doses.
As good practice in clinical trials, all participants received the investigators’ contact details and were guided to contact the hospital if any symptom occurred during the study period, or in case of any doubt regarding the study protocol. Each participant centre had the autonomy to decide whether their patients would continue to receive the qHPV vaccination in case of disease worsening.
For immunogenicity evaluation, blood drawing was performed at each study visit. Serum was collected and frozen under -70ºC in Brazil and subsequently shipped to the Netherlands for serologic antibody concentration testing using a virus-like particle based multiplex Luminex assay (12). Seropositivity for HPV16 and HPV18 was defined as an antibody concentration higher than 9 Luminex Units/ml and 13 Luminex Units/ml, respectively.
Data were organized in the Microsoft Excel program v16.23 according to participant group (JDM or HC), and whether the participant completed the protocol or not. Descriptive analyses were performed for the qualitative variables. Data were presented in numbers (percentages) for categorical variables and in median (range) for continuous variables.
Graphs and tables were prepared in Affinity Designer 1.6.1 and GraphPad Prism v7. The statistical significance of the categorical data was tested with the Fisher’s exact test, using IBM SPSS statistics v21. P values less than 0.05 were considered as significant.
In addition, the McNemar test (17) and the Kappa concordance coefficient (18, 19) were applied to compare the disease activity parameters scores used for JDM patients between pos the qHPV doses and before the vaccination. The hypothesis tested was the vaccination no induce flare or worsening of disease activity. The software used for the analyses was SAS 9.4 (20).
The null hypothesis was considered for these tests when the frequency of variables considered to evaluated disease activity were the same in the period pre and post vaccination.