Lower blood 25 (OH) D concentration is associated with increased risk of islet autoimmunity (IA) in some cohorts; it is not known if 25 (OH) D metabolites are important after the onset of IA. We investigated novel vitamin D metabolites in development of IA and progression from IA to T1D in the Diabetes Autoimmunity Study in the Young (DAISY) . IA was defined as two or more consecutive visits positive for ≥ 1 autoantibody (IAA, IA-2A, GAD, ZnT8) . In a nested case-control study, we examined plasma vitamin D metabolites at a pre-seroconversion visit in 114 IA cases and 116 controls. We then followed 144 IA-positive children, of whom 46 developed T1D during a mean follow-up of 8.4 years. Plasma was quantified for 25 (OH) D3, 3-epi-25 (OH) D3, 25 (OH) D2, 24,25 (OH) 2D3, and 1,25 (OH) 2D3, using targeted LC-MS/MS. Logistic regression was used to examine pre-seroconversion metabolites and IA, and Cox regression was used to examine metabolites at seroconversion and progression to T1D. Both models adjusted for age, ethnicity, T1D family history and HLA-DR3/4. Higher pre-seroconversion level of 24,25 (OH) 2D3, an inactive catabolite of 25 (OH) D3, was associated with increased odds of IA (OR: 1.35; 95%CI: 1.03,1.78 per SD of 24,25 (OH) 2D3; p=0.031) . The association between 3-epi-25 (OH) D3 at seroconversion and progression from IA to T1D was modified by age at seroconversion. Higher 3-epi-25 (OH) D3 level was more protective against progression to T1D among children who seroconverted at an older age (HR: 0.47; 95%CI: 0.26, 0.85 for children at the 75th percentile of age at seroconversion (ie years)) compared to children who seroconverted at a younger age (HR: 0.88; 95%CI: 0.63, 1.24 for children at the 25th percentile (ie 3 years)) . Neither IA nor progression to T1D was associated with 25 (OH) D3, 25 (OH) D2 or 1,25 (OH) 2D3.Increased 24,25 (OH) 2D3 may play a role in IA development. The age-related heterogeneity of the relationship between 3-epi-25 (OH) D3 and T1D progression requires further investigation. Disclosure S.Kim: None. K.Kechris: None. O.Fiehn: None. M.Rewers: Consultant; Janssen Research & Development, LLC, Medscape, Provention Bio, Inc., Research Support; Dexcom, Inc., JDRF, Roche Diagnostics USA. J.M.Norris: None. N.L.Zwick: None. P.M.Carry: None. B.C.Defelice: None. R.K.Johnson: None. F.Dong: None. T.Buckner: None. L.A.Vanderlinden: None. B.I.Frohnert: Advisory Panel; Provention Bio, Inc. Funding National Institutes of Health (R01-DK104351, R01-DK32493, RI142483)