Combined vitamin D, ibuprofen and glutamic acid decarboxylase-alum treatment in recent onset Type I diabetes: lessons from the DIABGAD randomized pilot trial

Johnny Ludvigsson, Indusmita Routray, Sriramulu Elluru, Per Leanderson, Helena E Larsson, Björn Rathsman, Ragnar Hanås, Annelie Carlsson, Torben Ek, Ulf Samuelsson, Torun Torbjörnsdotter, Jan Åman, Eva Örtqvist, Karun Badwal, Craig Beam, Rosaura Casas, Johnny Ludvigsson, Indusmita Routray, Sriramulu Elluru, Per Leanderson, Helena E Larsson, Björn Rathsman, Ragnar Hanås, Annelie Carlsson, Torben Ek, Ulf Samuelsson, Torun Torbjörnsdotter, Jan Åman, Eva Örtqvist, Karun Badwal, Craig Beam, Rosaura Casas

Abstract

Aim: Double-blind placebo-controlled intervention using glutamic acid decarboxylase (GAD)-alum, vitamin D and Ibuprofen in recent onset Type I diabetes (T1D).

Methods: 64 patients (T1D since <4 months, age 10-17.99, fasting sC-peptide ≥0.12 nmol/l, GADA-positive) were randomized into Day(D) 1-90 400 mg/day Ibuprofen, D1-450 vitamin D 2000 IU/day, D15, 45 sc. 20 μg GAD-alum; as A but placebo instead of Ibuprofen; as B but 40 μg GAD-alum D15, 45; placebo.

Results: Treatment was safe and tolerable. No C-peptide preservation was observed. We observed a linear correlation of baseline C-peptide, HbA1c and insulin/per kilogram/24 h with change in C-peptide AUC at 15 months (r = -0.776, p < 0.0001).

Conclusion: Ibuprofen, vitamin D + GAD-alum did not preserve C-peptide. Treatment efficacy was influenced by baseline clinical and immunological factors and vitamin D concentration. Clinical Trial Registration: NCT01785108 (ClinicalTrials.gov).

Keywords: C-peptide; GAD-alum; Type I diabetes; ibuprofen; immune response; vitamin D.

Conflict of interest statement

Financial & competing interests disclosure This trial was generously funded by Barndiabetesfonden (Swedish Child Diabetes Foundation), FORSS (the Research Council of Southeast Sweden), ALF (Region Östergötland) and unrestricted grants from Diamyd Medical. The funders had no role in designing or performing the study, nor in interpreting or presenting the results. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. No writing assistance was utilized in the production of this manuscript.

© 2020 Johnny Ludvigsson.

Figures

Figure 1.. Flow chart showing the recruitment…
Figure 1.. Flow chart showing the recruitment and distribution of patients into the different groups.
Figure 2.. C-peptide response to treatment and…
Figure 2.. C-peptide response to treatment and C-peptide change predicted by baseline clinical end points.
(A) Mean (95% CI) arithmetic change in C-peptide AUC/120 from 6 to 30 months-PP population. (B) Score values generated by CCA are plotted against arithmetic change in C-peptide AUC/120 from baseline to visit eight (30 months). The correlation between the score and change is estimated to be -0.7764 and statistically significant. (C) Weights assigned to each end point by canonical correlation analysis (CCA). Bar chart showing the numeric weights assigned by CCA to each variable and predicting change in C-peptide AUC/120. GAD, baseline AUC/120 and maximum baseline C-peptide are most prominent in determining the clinical score. The score increases with increases in GAD and AUC while decreasing with the other variables.
Figure 3.. The effect of vitamin D…
Figure 3.. The effect of vitamin D treatment on vitamin D concentrations and association between vitamin D increase and C-peptide.
(A) Increase of vitamin D concentrations in the three arms treated with 2000 U/day. (B) Association between increasing vitamin D from baseline and reduced loss of C-peptide. Arithmetic change in C-peptide AUC/120 min for each subject and from baseline to visit six and visit seven. Treatment group assignments indicated by different colors and alphabetic letter. Trend lines fit within each treatment group are superimposed. Treatment group key: A=Ibuprofen Diamyd® 20 mg×2 and vitamin D, B=Diamyd® 20 mg×2 and vitamin D, C=Diamyd® 40 mg×2 and vitamin D, D = placebo. Significant differences are indicated by p-values.
Figure 4.. Baseline cytokines levels in serum.
Figure 4.. Baseline cytokines levels in serum.
(A) Median levels (horizontal line) of IL-1 (pg/ml) at baseline and 180 days for A (Ibuprofen + Diamyd® 20 mg×2 + vitamin D; black circles); B (Diamyd® 20 mg×2 + vitamin D, black squares); C (Diamyd® 40 mg×2 + Vitamin D, black triangles); D (Placebo, open circles) were detected by Luminex. (B) Relative contribution (%) of the cytokines at baseline. Median values are indicated by horizontal lines. Significant differences are indicated by p-values.
Figure 5.. Effect of the treatment and…
Figure 5.. Effect of the treatment and the immune response.
Proliferative response to GAD65 at (A) 3 and (B) 6 months. Proliferative response to PMA/Ionomycin at (C) 3 and (D) 6 months. Proliferation is expressed as stimulation index (SI), calculated from the mean of triplicates divided by the mean of triplicates with medium alone. (E–H) GAD65-induced cytokine secretion at 6 months upon in vitro PMBC stimulation. Cytokines were detected by Luminex in supernatants collected after 7 days culture in presence of medium or GAD65 (5 μg/ml). GAD65-induced cytokine secretion is given after subtraction of spontaneous secretion from each individual. Median levels (horizontal line) of cytokine (pg/ml) at baseline and 180 days for A (Ibuprofen + Diamyd® 20 mg×2 + Vitamin D; black circles); B (Diamyd® 20 mg×2 + vitamin D, black squares); C (Diamyd® 40 mg×2 + vitamin D, black triangles); D (placebo, open circles). Median values are indicated by horizontal lines. Significant differences are indicated by p-values.

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