Daniel Pipeleers

Four-year metabolic outcome of a randomised controlled CD3-antibody trial in recent-onset type 1 diabetic patients depends on their age and baseline residual beta cell mass

Aims/hypothesisThe aim of the study was to examine the 48xa0month outcome of treating recent-onset type 1 diabetic patients for 6xa0days with humanised CD3-antibody, ChAglyCD3.MethodsEighty patients, aged 12–39xa0years, were recruited for a phase 2 multicentre trial and randomised to placebo (nu2009=u200940) or ChAglyCD3 (nu2009=u200940) treatment by a third party member; participants and care-givers were blinded. The change in insulin dose (Uxa0kg−1u2009day−1) over 48xa0months was chosen as primary endpoint and compared in 31 placebo- and 33 ChAglyCD3-treated patients. Adverse events were followed in 35 and 38 patients, respectively.ResultsTreatment with ChAglyCD3 delayed the rise in insulin requirements of patients with recent-onset diabetes and reduced its amplitude over 48xa0months (+0.09 vs +0.32xa0Uu2009kg−1u2009day−1 in the placebo group). Using multivariate analysis this effect was correlated with higher baseline residual beta cell function and a younger age. It was associated with better outcome variables in subgroups selected according to these variables. In the ChAglyCD3 subgroup with higher initial beta cell function, 0/11 patients became C-peptide-negative over 48xa0months vs 4/9 in the corresponding placebo subgroup. In the subgroup aged <27xa0years old, antibody treatment preserved initial beta cell function for 36xa0months (vs >80% decline within 24xa0months in the placebo subgroup <27xa0years old), resulted in lower HbA1c concentrations and tended to reduce glycaemic variability (pu2009=u20090.08). No long-term adverse events were observed.Conclusions/interpretationA 6xa0day ChAglyCD3 treatment can suppress the rise in insulin requirements of recent-onset type 1 diabetic patients over 48xa0months, depending on their age and initial residual beta cell function. In younger patients this effect is associated with reduced deterioration of metabolic variables. These observations help to define inclusion criteria for prevention trials.Trial registration:ClinicalTrials.gov NCT00627146Funding:Center grants from the Juvenile Diabetes Research Foundation (4-2001-434, 4-2005-1327) and grants from the Belgian Fund for Scientific Research-Flanders and from Brussels Free University-VUB.

Sustained function of alginate-encapsulated human islet cell implants in the peritoneal cavity of mice leading to a pilot study in a type 1 diabetic patient

AbstractAims/hypothesisAlginate-encapsulated human islet cell grafts have not been able to correct diabetes in humans, whereas free grafts have. This study examined in immunodeficient mice whether alginate-encapsulated graft function was inferior to that of free grafts of the same size and composition.MethodsCultured human islet cells were equally distributed over free and alginate-encapsulated grafts before implantation in, respectively, the kidney capsule and the peritoneal cavity of non-obese diabetic mice with severe combined immunodeficiency and alloxan-induced diabetes. Implants were followed for in vivo function and retrieved for analysis of cellular composition (all) and insulin secretory responsiveness (capsules).ResultsFree implants with low beta cell purity (19u2009±u20091%) were non-functional and underwent 90% beta cell loss. At medium purity (50u2009±u20091%), they were functional at post-transplant week 1, evolving to normoglycaemia (4/8) or to C-peptide negativity (4/8) depending on the degree of beta cell-specific losses. Encapsulated implants immediately and sustainably corrected diabetes, irrespective of beta cell purity (16/16). Most capsules were retrievable as single units, enriched in endocrine cells that exhibited rapid secretory responses to glucose and glucagon. Single capsules with similar properties were also retrieved from a type 1 diabetic recipient at post-transplant month 3. However, the vast majority were clustered and contained debris, explaining the poor rise in plasma C-peptide.Conclusions/interpretationIn immunodeficient mice, i.p. implanted alginate-encapsulated human islet cells exhibited a better outcome than free implants under the kidney capsule. They did not show primary non-function at low beta cell purity and avoided beta cell-specific losses by rapidly establishing normoglycaemia. Retrieved capsules presented secretory responses to glucose, which was also observed in a type 1 diabetic recipient.n Trial registration: ClinicalTrials.gov NCT01379729n Funding: This study was supported by grants from the JDRF (centre grant 4-2005-1327), the Research Foundation Flanders (G.0801.10), the 6th and 7th Framework Program of the European Commission (numbers 512145 and 241883), and the Agency for Innovation by Science and Technology in Flanders (IWT-TBM7 090884).

Transient Epstein-Barr virus reactivation in CD3 monoclonal antibody-treated patients

Here we report a unique situation in which an early and synchronized Epstein-Barr virus (EBV) reactivation was induced by a 6-day course of treatment with a humanized CD3-specific monoclonal antibody in patients with recent onset of type 1 diabetes. The virologic and immunologic analysis demonstrated that this reactivation was transient, self-limited, and isolated, associated with the rapid advent of an EBV-specific T-cell response. The anti-CD3 antibody administration induced short-lasting immunosuppression and minor yet clear-cut signs of T-cell activation that preceded viral reactivation. Early posttransplant monitoring of renal and islet allograft recipients showed that no comparable phenomenon was observed after the administration of full-dose immunosuppressive therapy. This EBV reactivation remains of no apparent clinical concern over the long term and should not preclude further development of therapeutic anti-CD3 antibodies. This phenomenon may also direct new research avenues to understand the still ill-defined nature of stimuli triggering EBV reactivation in vivo.

Seasonality in clinical onset of Type 1 diabetes in Belgian patients above the age of 10 is restricted to HLA-DQ2/DQ8-negative males, which explains the male to female excess in incidence

Aims/hypothesisType 1 diabetes arises from an interplay between environmental and genetic factors. The reported seasonality at diagnosis supports the hypothesis that currently unknown external triggers play a role in the onset of the disease. We investigated whether a seasonal pattern is observed at diagnosis in Belgian Type 1 diabetic patients, and if so whether seasonality varies according to age, sex and genetic risk, all known to affect the incidence of Type 1 diabetes.MethodsThe seasonal pattern at clinical diagnosis was assessed in 2176 islet antibody-positive diabetic patients aged 0 to 39 years diagnosed between 1989 and 2000. Additional stratification was performed for age, sex and HLA-DQ genotype.ResultsOverall, a significant seasonal pattern at clinical diagnosis of diabetes was observed (p<0.001). More subjects were diagnosed in the period of November to February (n=829) than during the period of June to September (n=619) characterised by higher averages of maximal daily temperature and daily hours of sunshine. However, the seasonal pattern was restricted to patients diagnosed above the age of 10 (0–9 years: p=0.398; 10–19 years: p<0.001; 20–29 years: p=0.003; 30–39 years: p=0.015). Since older age at diagnosis is associated with a male to female excess and a lower prevalence of the genetic accelerator HLA-DQ2/DQ8, we further stratified the patients aged 10 to 39 years (n=1675) according to HLA-DQ genotype and sex, and we found that the seasonal pattern was largely restricted to male subjects lacking DQ2/DQ8 (n=748; p<0.001 vs all others: n=927; p=0.031).Conclusions/interpretationIn a subgroup of male patients diagnosed over the age of 10, the later stages of the subclinical disease process may be more driven by sex- and season-dependent external factors than in younger, female and genetically more susceptible subjects. These factors may explain the male to female excess in diabetes diagnosed in early adulthood.

Screening for insulinoma antigen 2 and zinc transporter 8 autoantibodies: a cost‐effective and age‐independent strategy to identify rapid progressors to clinical onset among relatives of type 1 diabetic patients

In first‐degree relatives of type 1 diabetic patients, we investigated whether diabetes risk assessment solely based on insulinoma antigen 2 (IA‐2) and zinc transporter 8 (ZnT8) antibody status (IA‐2A, respectively, ZnT8A) is as effective as screening for three or four autoantibodies [antibodies against insulin (IAA), glutamate decarboxylase 65u2009kDa (GAD) glutamate decarboxylase autoantibodies (GADA) and IA‐2A with or without ZnT8A] in identifying children, adolescents and adults who progress rapidly to diabetes (within 5 years). Antibodies were determined by radiobinding assays during follow‐up of 6444 siblings and offspring aged 0–39 years at inclusion and recruited consecutively by the Belgian Diabetes Registry. We identified 394 persistently IAA+, GADA+, IA‐2A+ and/or ZnT8A+ relatives (6·1%). After a median follow‐up time of 52 months, 132 relatives developed type 1 diabetes. In each age category tested (0–9, 10–19 and 20–39 years) progression to diabetes was significantly quicker in the presence of IA‐2A and/or ZnT8A than in their joint absence (Pu2009<u20090·001). Progression rate was age‐independent in IA‐2A+ and/or ZnT8A+ relatives but decreased with age if only GADA and/or IAA were present (Pu2009=u20090·008). In the age group mainly considered for immune interventions until now (10–39 years), screening for IA‐2A and ZnT8A alone identified 78% of the rapid progressors (versus 75% if positive for ≥u20092 antibodies among IAA, GADA, IA‐2A and ZnT8A or versus 62% without testing for ZnT8A). Screening for IA‐2A and ZnT8A alone allows identification of the majority of rapidly progressing prediabetic siblings and offspring regardless of age and is more cost‐effective to select participants for intervention trials than conventional screening.

Sex- and season-dependent differences in C-peptide levels at diagnosis of immune-mediated type 1 diabetes

Aims/hypothesisThe incidence of type 1 diabetes varies according to age, sex and season of diagnosis. We investigated whether these and other clinical, biological and anthropometric parameters were correlated with residual beta cell function in newly diagnosed patients, since it is possible that the nature of external and/or genetic disease accelerators may be (partly) reflected in the inaugural disease presentation.Materials and methodsThe correlates of random C-peptide levels sampled shortly after diagnosis (median [interquartile range]: 3 [0–14]xa0days) were studied by multivariate analysis in 1,883 islet-antibody-positive diabetic patients aged <40xa0years who were diagnosed between 1989 and 2000.ResultsHigher C-peptide levels (above percentile 50 of patients) were associated with older age at diagnosis, female sex, diagnosis in the high-incidence season (October to March), less-decreased BMI (expressed as a standard deviation score), lower insulin requirements after stabilisation, lower prevalence of ketonuria and a less-increased glycaemia at diagnosis (all p<0.001). C-peptide levels were not correlated with calendar year at diagnosis, duration of symptoms prior to diagnosis, HLA-DQ2/DQ8 genotype or islet antibody status.Conclusions/interpretationSex- and season-dependent differences in residual functional beta cell mass and/or insulin resistance have been identified at diagnosis of type 1 diabetes. They may reflect differences in disease-precipitating external or lifestyle factors and should be further investigated longitudinally in prediabetes to further identify putative aetiological factors, which may provide targets for prevention.

HLA-A*24 Is an Independent Predictor of 5-Year Progression to Diabetes in Autoantibody-Positive First-Degree Relatives of Type 1 Diabetic Patients

We investigated whether HLA-A*24 typing complements screening for HLA-DQ and for antibodies (Abs) against insulin, GAD, IA-2 (IA-2A), and zinc transporter-8 (ZnT8A) for prediction of rapid progression to type 1 diabetes (T1D). Persistently Ab+ siblings/offspring (n = 288; aged 0–39 years) of T1D patients were genotyped for HLA-DQA1-DQB1 and HLA-A*24 and monitored for development of diabetes within 5 years of first Ab+. HLA-A*24 (P = 0.009), HLA-DQ2/DQ8 (P = 0.001), and positivity for IA-2A ± ZnT8A (P < 0.001) were associated with development of T1D in multivariate analysis. The 5-year risk increased with the number of the above three markers present (n = 0: 6%; n = 1: 18%; n = 2: 46%; n = 3: 100%). Positivity for one or more markers identified a subgroup of 171 (59%) containing 88% of rapid progressors. The combined presence of HLA-A*24 and IA-2A+ ± ZnT8A+ defined a subgroup of 18 (6%) with an 82% diabetes risk. Among IA-2A+ ± ZnT8A+ relatives, identification of HLA-A*24 carriers in addition to HLA-DQ2/DQ8 carriers increased screening sensitivity for relatives at high Ab- and HLA-inferred risk (64% progression; P = 0.002). In conclusion, HLA-A*24 independently predicts rapid progression to T1D in Ab+ relatives and complements IA-2A, ZnT8A, and HLA-DQ2/DQ8 for identifying participants in immunointervention trials.

Hyperglycaemic clamp test for diabetes risk assessment in IA-2-antibody-positive relatives of type 1 diabetic patients

Aims/hypothesisThe aim of the study was to investigate the use of hyperglycaemic clamp tests to identify individuals who will develop diabetes among insulinoma-associated protein-2 antibody (IA-2A)-positive first-degree relatives (IA-2A+ FDRs) of type 1 diabetic patients.MethodsHyperglycaemic clamps were performed in 17 non-diabetic IA-2A+ FDRs aged 14 to 33xa0years and in 21 matched healthy volunteers (HVs). Insulin and C-peptide responses were measured during the first (5–10xa0min) and second (120–150xa0min) release phase, and after glucagon injection (150–160xa0min). Clamp-induced C-peptide release was compared with C-peptide release during OGTT.ResultsSeven (41%) FDRs developed diabetes 3–63xa0months after their initial clamp test. In all phases they had lower C-peptide responses than non-progressors (pu2009<u20090.05) and HVs (pu2009<u20090.002). All five FDRs with low first-phase release also had low second-phase release and developed diabetes 3–21xa0months later. Two of seven FDRs with normal first-phase but low second-phase release developed diabetes after 34 and 63xa0months, respectively. None of the five FDRs with normal C-peptide responses in all test phases has developed diabetes so far (follow-up 56 to 99xa0months). OGTT-induced C-peptide release also tended to be lower in progressors than in non-progressors or HVs, but there was less overlap in results between progressors and the other groups using the clamp.Conclusions/interpretationClamp-derived functional variables stratify risk of diabetes in IA-2A+ FDRs and may more consistently identify progressors than OGTT-derived variables. A low first-phase C-peptide response specifically predicts impending diabetes while a low second-phase response may reflect an earlier disease stage.Trial registration:ClinicalTrials.gov NCT00654121Funding:The insulin trial was financially supported by Novo Nordisk Pharma nv.

The age at diagnosis of type 1 diabetes continues to decrease in Belgian boys but not in girls: a 15-year survey

The age at clinical onset of type 1 diabetes is decreasing. Preliminary Belgian data suggested that this anticipation occurred preferentially in boys. We investigated whether this gender‐specific anticipation could be confirmed over a 15‐year observation period.

Twenty-Year Progression Rate to Clinical Onset According to Autoantibody Profile, Age, and HLA-DQ Genotype in a Registry-Based Group of Children and Adults With a First-Degree Relative With Type 1 Diabetes

OBJECTIVE We investigated whether islet autoantibody profile, HLA-DQ genotype, and age influenced a 20-year progression to diabetes from first autoantibody positivity (autoAb+) in first-degree relatives of patients with type 1 diabetes. RESEARCH DESIGN AND METHODS Persistently islet autoAb+ siblings and offspring (n = 462) under 40 years of age were followed by the Belgian Diabetes Registry. AutoAbs against insulin (IAA), GAD (GADA), IA-2 antigen (IA-2A), and zinc transporter 8 (ZnT8A) were determined by radiobinding assay. RESULTS The 20-year progression rate of multiple-autoAb+ relatives (n = 194) was higher than that for single-autoAb+ participants (n = 268) (88% vs. 54%; P < 0.001). Relatives positive for IAA and GADA (n = 54) progressed more slowly than double-autoAb+ individuals carrying IA-2A and/or ZnT8A (n = 38; P = 0.001). In multiple-autoAb+ relatives, Cox regression analysis identified the presence of IA-2A or ZnT8A as the only independent predictors of more rapid progression to diabetes (P < 0.001); in single-autoAb+ relatives, it identified younger age (P < 0.001), HLA-DQ2/DQ8 genotype (P < 0.001), and IAA (P = 0.028) as independent predictors of seroconversion to multiple positivity for autoAbs. In time-dependent Cox regression, younger age (P = 0.042), HLA-DQ2/DQ8 genotype (P = 0.009), and the development of additional autoAbs (P = 0.012) were associated with more rapid progression to diabetes. CONCLUSIONS In single-autoAb+ relatives, the time to multiple-autoAb positivity increases with age and the absence of IAA and HLA-DQ2/DQ8 genotype. The majority of multiple-autoAb+ individuals progress to diabetes within 20 years; this occurs more rapidly in the presence of IA-2A or ZnT8A, regardless of age, HLA-DQ genotype, and number of autoAbs. These data may help to refine the risk stratification of presymptomatic type 1 diabetes.