A phase 1b open-label dose-finding study of ustekinumab in young adults with type 1 diabetes

Aim We assessed the safety of ustekinumab (a monoclonal antibody used in psoriasis to target the IL-12 and IL-23 pathways) in a small cohort of recent-onset (<100 days of diagnosis) adults with type 1 diabetes (T1D) by conducting a pilot open-label dose-finding and mechanistic study (NCT02117765) at the University of British Columbia. Methods We sequentially enrolled 20 participants into four subcutaneous dosing cohorts: i) 45mg loading-weeks 0/4/16, ii) 45mg maintenance-weeks 0/4/16/28/40, iii) 90mg loading-weeks 0/4/16 and iv) 90mg maintenance-weeks 0/4/16/28/40. The primary endpoint was safety as assessed by an independent data and safety monitoring board (DSMB) but we also measured mixed meal tolerance test C-peptide, insulin use/kg, and HbA1c. Immunophenotyping was performed to assess immune cell subsets and islet antigen-specific T cell responses. Results Although several adverse events were reported, only two (bacterial vaginosis and hallucinations) were thought to be possibly related to drug administration by the study investigators. At 1 year, the 90mg maintenance dosing cohort had the smallest mean decline in C-peptide AUC (0.1pmol/mL). Immunophenotyping showed that ustekinumab reduced the percentage of circulating Th17, Th1 and Th17.1 cells and proinsulin-specific T cells that secreted IFN-γ and IL-17A. Conclusion Ustekinumab was deemed safe to progress to efficacy studies by the DSMB at doses used to treat psoriasis in adults with T1D. A 90mg maintenance dosing schedule reduced proinsulin-specific IFN-γ and IL-17A-producing T cells. Further studies are warranted to determine if ustekinumab can prevent C-peptide AUC decline and induce a clinical response

Treg gene signatures predict and measure type 1 diabetes trajectory

BACKGROUND: Multiple therapeutic strategies to restore immune regulation and slow type 1 diabetes (T1D) progression are in development and testing. A major challenge has been defining biomarkers to prospectively identify subjects likely to benefit from immunotherapy and/or measure intervention effects. We previously found that compared to healthy controls, Tregs from children with new-onset T1D have an altered Treg gene signature (TGS), suggesting this could be an immunoregulatory biomarker. METHODS: nanoString was used to assess the TGS in sorted Tregs (CD4+CD25hiCD127lo) or Peripheral Blood Mononuclear Cells (PBMC) from individuals with T1D or type 2 diabetes, healthy controls, or T1D recipients of immunotherapy. Biomarker discovery pipelines were developed and applied to various sample group comparisons. RESULTS: Compared to controls, the TGS in isolated Tregs or PBMCs is altered in adult new-onset and cross-sectional T1D cohorts, with sensitivity and specificity of biomarkers increased by including T1D-associated single nucleotide polymorphisms in algorithms. The TGS was distinct in T1D versus type 2 diabetes, indicating disease-specific alterations. TGS measurement at the time of T1D onset revealed an algorithm that accurately predicted future rapid versus slow C-peptide decline, as determined by longitudinal analysis of placebo arms of START and T1DAL trials. The same algorithm stratified participants in a phase I/II clinical trial of ustekinumab (αIL-12/23p40) for future rapid versus slow C-peptide decline. CONCLUSION: These data suggest that biomarkers based on measuring Treg gene signatures could be a new approach to stratify patients and monitor autoimmune activity in T1D

Phagocytosis of apoptotic cells by macrophages from NOD mice is reduced

Macrophages limit inflammatory responses by clearing apoptotic cells. Deficiencies in apoptotic cell phagocytosis have been linked to autoimmunity. In this study, we determined the efficiency with which macrophages from diabetes-prone NOD and diabetes-resistant NOR, Idd5, Balb/c, and C57BL/6 mice phagocytose apoptotic thymocytes and NIT-1 insulinoma cells. Peritoneal and bone marrow-derived macrophages from NOD mice engulfed fewer apoptotic thymocytes than macrophages from Balb/c mice (P < 0.05). Peritoneal macrophages from NOR and Idd5 NOD congenic mice were more proficient at engulfment than their NOD counterparts. Annexin V blockade diminished apoptotic thymocyte clearance and heat-labile serum factors augmented clearance. Binding of apoptotic thymocytes to NOD macrophages was also reduced, suggesting that the deficiency in phagocytosis may be partly attributable to a recognition defect. Peritoneal macrophages from female Balb/c and NOD mice were equally efficient in the engulfment of microspheres, suggesting that the phagocytic deficiency observed in NOD mice was specific for apoptotic cells. In summary, we have demonstrated a deficiency in phagocytic function of macrophages from NOD mice. Normal and diabetes-prone neonatal rodents have a wave of β-cell apoptosis coincident with the onset of target organ inflammation. A constitutive defect in the clearance of apoptotic β-cells may be contributory to the initiation of autoimmunity

In situ β cell death promotes priming of diabetogenic CD8 T lymphocytes

CTLs are important mediators of pancreatic β cell destruction in the nonobese diabetic mouse model of type 1 diabetes. Cross-presentation of Ag is one means of priming CTLs. The death of Ag-bearing cells has been implicated in facilitating this mode of priming. The role of β cell death in facilitating the onset of spontaneous autoimmune diabetes is unknown. Here, we used an adoptive transfer system to determine the time course of islet-derived Ag presentation to naive β cell-specific CD8 T cells in nonobese diabetic mice and to test the hypothesis that β cell death enhances the presentation of β cell autoantigen. We have determined that β cell death enhances autoantigen presentation. Priming of diabetogenic CD8 T cells in the pancreatic lymph nodes was negligible before 4 wk, progressively increased until 8 wk of age, and was not influenced by gender. Administration of multiple low doses of the β cell toxin streptozotocin augmented in situ β cell apoptosis and accelerated the onset and magnitude of autoantigen presentation to naive CD8 T cells. Increasing doses of streptozotocin resulted in both increased pancreatic β cell death and significantly enhanced T cell priming. These results indicate that in situ β cell death facilitates autoantigen-specific CD8 T cell priming and can contribute to both the initiation and the ongoing amplification of an autoimmune response

A deficiency in the in vivo clearance of apoptotic cells is a feature of the NOD mouse

Deficiencies in apoptotic cell clearance have been linked to autoimmunity. Here we examined the time-course of peritoneal macrophage phagocytosis of dying cells following the direct injection of apoptotic thymocytes into the peritoneum of NOD mice and BALB/c controls. Macrophages from NOD mice demonstrated a profound defect in the phagocytosis of apoptotic thymocytes as compared to control macrophages. Nonobese diabetic mice also demonstrated a decrease in the clearance of apoptotic cell loads following an apoptotic stimulus to thymocytes (dexamethasone) when compared to BALB/c or NOR controls. Further, NOD mice demonstrated an increase in apoptotic cell load following an apoptotic stimulus to keratinocytes (ultraviolet light, UVB) when compared to control strains. Animals deficient in macrophage phagocytosis of apoptotic debris often manifest an autoimmune phenotype characterized by the production of antinuclear autoantibodies (ANA). We determined whether increased apoptotic cell loads (through repeated exposure to UVB irradiation) could accelerate such autoimmune phenomena in young NOD mice. Following repeated UVB irradiation, NOD mice, but not BALB/c or NOR controls, developed ANA. We propose that abnormalities in apoptotic cell clearance by macrophages predispose NOD mice to autoimmunity. © 2005 Elsevier Ltd. All rights reserved