Association between treatment effect on C-peptide preservation and HbA1c in meta-analysis of glutamic acid decarboxylase (GAD)-alum immunotherapy in recent-onset type 1 diabetes.

BACKGROUND

Preservation of insulin secretion in type 1 diabetes (T1D) lowers the risk of complications such as retinopathy and severe hypoglycaemia. , , C‐peptide preservation—a biomarker for endogenous insulin secretion—is used as a surrogate endpoint in clinical trials assessing beta cell‐preserving therapies. Showing efficacy on clinical outcomes such as severe hypoglycaemia has proven challenging given their rarity shortly after diagnosis. Here, we show that treatment effect on C‐peptide correlates with treatment effect on HbA1c, a validated surrogate endpoint, in individuals treated with recombinant human glutamic acid decarboxylase 65 kDa in alum (GAD‐alum).

A retrospective post hoc meta‐analysis and prospective phase IIb trial, where the human leukocyte antigen (HLA)‐specific analyses of the topline results were specified before database lock, have shown the efficacy of GAD‐alum for stimulated C‐peptide preservation in the genetic subpopulation of T1D patients carrying HLA DR3‐DQ2. , Efficacy for reducing HbA1c and added benefit of more doses have also been observed in the HLA DR3‐DQ2–carrying probable responder population. A phase III trial in recent‐onset T1D patients carrying HLA DR3‐DQ2 (DIAGNODE‐3, NCT05018585) with the co‐primary endpoints of C‐peptide and HbA1c is ongoing.

Our research objective was to assess whether 15‐month treatment effects on preservation of endogenous insulin production were correlated with treatment effects on blood glucose measured by HbA1c.

METHODS

We carried out individual person meta‐analysis of four phase II‐III randomized controlled trials of subcutaneous or intralymphatic GAD‐alum versus placebo in recent‐onset T1D (n = 627).

All four studies were randomized placebo‐controlled trials in recent‐onset T1D patients receiving standard‐of‐care diabetes treatment including insulin replacement therapy. Study NCT00435981 was a multisite phase II clinical trial of subcutaneous GAD‐alum including 70 patients in Sweden. Study NCT00529399 was a phase II clinical trial of subcutaneous GAD‐alum conducted by Trialnet, which included 145 patients at 15 sites in the United States. Study NCT00723411 was a phase III clinical trial of subcutaneous GAD‐alum including 334 patients at sites in Finland, France, Germany, Italy, The Netherlands, Slovenia, Spain, Sweden and the UK. Study NCT03345004 (DIAGNODE‐2) was a phase IIb clinical trial of intralymphatic GAD‐alum that included 109 patients from multiple sites in the Czech Republic, The Netherlands, Spain and Sweden.

For analysis, patients were classified according to HLA DR3‐DQ2 presence, and as having received two injections or three to four injections of either GAD‐alum or placebo. A covariate term for study was included in the analysis to account for minor differences in design. Stimulated C‐peptide was assessed using the mixed meal tolerance test. Mean C‐peptide area under the curve was calculated by the trapezoid rule and natural log‐transformed. HbA1c was analysed at a central laboratory and not transformed. Mean change from baseline was analysed using a restricted maximum likelihood‐based mixed model for repeated measures (MMRM) adjusted for the fixed effects of baseline C‐peptide (or baseline HbA1c), study, treatment, HLA subgroup, visit, country, sex and age, as well as the interaction of baseline C‐peptide (or HbA1c) by visit and treatment by HLA subgroup by visit. Patient identification number and country were included as categorical random effects. The primary comparison was the contrast between treatments at month 15 for active treatment versus placebo. The change from baseline to month 15 was chosen as 15 months was the longest follow‐up available in all studies. Further details on the methods and the included studies are presented in the Appendix.

RESULTS

Figure 1 shows trial‐level scatterplots of the relationship between treatment effects of GAD‐alum versus placebo on C‐peptide (x‐axis) and HbA1c (y‐axis). For each trial, effects were estimated by MMRM analysis in probable responders (DR3‐DQ2–positive) and non‐responders (DR3‐DQ2–negative), depicted in opaque and transparent circles, respectively, with diameters proportional to sample size. An association between effects on C‐peptide and HbA1c is apparent for participants who had received three to four doses (B), while no association is apparent for two doses (A). The combined analysis of two to four doses (C) reflects the association driven by the three‐to‐four‐doses group. The treatment effect of GAD‐alum was foremost significant in DR3‐DQ2 patients receiving three to four rather than two doses. C‐peptide preservation of 40% from three to four doses of GAD‐alum corresponde to 3 mmol/mol lower HbA1c (regarding change from baseline compared with placebo) (Figure 1B). While the relationship between treatment benefit for C‐peptide and HbA1c is apparent across trials and HLA groups, Figure 1 reinforces the previous findings of treatment benefit in those participants with HLA DR3‐DQ2, while participants lacking DR3‐DQ2 do not appear to benefit from GAD‐alum. ,

FIGURE 1

Scatter plots illustrating the association between treatment effect (change from baseline to month 15 compared with placebo) on C‐peptide on the x‐axis and HbA1c on the y‐axis. The panels depict effect correlations for A, Two doses, B, Three to four doses, or C, Two to four doses. Values larger than 1 on the x‐axis, and lower than 0 on the y‐axis, indicate a beneficial treatment effect of GAD‐alum compared with placebo. Circles represent randomized controlled trials with diameters proportional to sample size. Each trial was split into the probable responder population (present HLA DR3‐DQ2, opaque circles) and non‐responder population (absent HLA DR3‐DQ2, transparent circles). The grey line represents the linear regression line and 95% confidence interval. Study labels: SWE Ph2—NCT00435981, Tn08—NCT00529399, EU Ph3—NCT00723411, DIAGNODE‐2—NCT03345004. GAD, glutamic acid decarboxylase; GMR, geometric mean ratio

Several sensitivity meta‐analyses were conducted and are reported in full in the Appendix. One analysis limited the age and country range of the four clinical trials in the meta‐analysis to correspond to the planned confirmatory phase III trial (Figure 1). A second analysis used an alternative C‐peptide treatment effect estimator (the quantitative response metric) (Figures A2 and A3). A third sensitivity analysis additionally adjusted for insulin dose (Figure A4). These sensitivity analyses confirmed the associations and the treatment benefits seen in HLA DR3‐DQ2–positive individuals who had received three or four injections.

FIGURE A2

Scatter plots illustrating the association between treatment effect (change from baseline to Month 15 compared to placebo) on C‐peptide on the x‐axis and HbA1c on the y‐axis. Panels (a) 2 injections, (b) 3‐4 injections, (c) 2‐4 injections. Values larger than 1 on the x‐axis, and lower than 0 on the y‐axis, indicate a beneficial treatment effect of GAD‐alum compared to placebo. Circles represent randomized controlled trials with diameters proportional to sample size. Each trial was split into the responder population (present HLA DR3‐DQ2, opaque circles) and non‐responder population (absent HLA DR3‐DQ2, transparent circles). The grey line represents the linear regression line and 95% confidence interval. Study labels: SWE Ph2 ‐ NCT00435981, Tn08 ‐ NCT00529399, EU Ph3 ‐ NCT00723411, DIAGNODE‐2 ‐ NCT03345004

FIGURE A3

Scatterplot illustrating good agreement between observed C‐peptide (y‐axis) and modified QR‐predicted C‐peptide at Month 15 in study participants with recent‐onset T1D assigned to placebo treatment in the DIAGNODE‐2 trial

FIGURE A4

Scatter plots illustrating the association between treatment effect (change from baseline to Month 15 compared to placebo) on C‐peptide on the x‐axis and HbA1c on the y‐axis. Panels (A) 2 injections, (B) 3–4 injections, (C) 2–4 injections. Similar to Figure 1 of the main text but C‐peptide effects on the x‐axis have been estimated using the modified QR metric instead of MMRM

We repeated the main analysis substituting HbA1c for the secondary endpoints of insulin dose‐adjusted HbA1c (IDAA1c) and insulin dose to assess whether a similar association between treatment effects on C‐peptide and these endpoints might exist. In both cases, there was a similar trend for an association between beneficial effects on C‐peptide being associated with beneficial effects on lower IDAA1c and lower insulin dose, respectively, in HLA DR3‐DQ2 individuals, and particularly in those who had received three or four doses (Figures A5 and A6).

FIGURE A5

Scatter plots illustrating the association between treatment effect (change from baseline to Month 15 compared to placebo) on C‐peptide on the x‐axis and HbA1c on the y‐axis; as in Figure 1 of the main text but with additional adjustment for insulin dose in the MMRM model. Upper panel: 2–4 injections. Lower panel: 3 or 4 injections

FIGURE A6

Scatter plots illustrating the association between treatment effect (change from baseline to Month 15 compared to placebo) on C‐peptide on the x‐axis and IDAA1c on the y‐axis. Upper panel: 2–4 injections. Lower panel: 3 or 4 injections

DISCUSSION

The correlation between 15‐month effects on C‐peptide and HbA1c suggests that therapeutically preserved C‐peptide in recent‐onset T1D might improve glycaemic control, probably at least for antigen‐specific immunotherapies, as immunomodulatory drugs with consistent C‐peptide effects have not shown convincing effects on HbA1c. , , Whether this discrepancy is a result of disease heterogeneity requiring subgroup‐targeted approaches, as for GAD‐alum, or because HbA1c is affected by complex factors, remains open.

Continuous glucose measurement‐derived variables, such as time in the glycaemic target range, are not accepted as primary endpoints by regulatory authorities, and serious clinical outcomes such as severe hypoglycaemia are (fortunately) rare in individuals with recently diagnosed T1D receiving standard‐of‐care treatment. This poses challenges for clinical trials with regard to the required sample size and follow‐up to show efficacy. Therefore, many T1D trials assess a surrogate primary endpoint such as C‐peptide, which is assumed to predict effects on clinical outcomes. Currently, neither the US Food and Drug Administration nor the European Medicines Agency accept C‐peptide preservation as a single primary endpoint. Our findings provide important new evidence supporting the use of C‐peptide as a surrogate endpoint in clinical trials in T1D. This is particularly poignant given current efforts by international consortia (such as the Critical Path Institute′s Trial Outcome Markers Initiative, TOMI‐T1D; https://c-path.org/programs/tomi-t1d/overview/tomi-t1d-team/) and key opinion leaders to convince the regulatory agencies to accept C‐peptide as a surrogate primary endpoint. Both agencies have, amongst others, mentioned two aspects that have yet to be satisfactorily addressed: to show a quantitative relationship between the amount of preserved C‐peptide and a clinical outcome or validated surrogate; and to show that therapeutic C‐peptide preservation achieves clinical benefits (even although there is evidence based on naturally preserved C‐peptide). , , We believe that our novel findings address both aspects in that we show correlated benefits of therapeutically preserved C‐peptide on the validated surrogate endpoint HbA1c. While it is not possible to say if there is a direct causal relationship between preserved C‐peptide and lowered HbA1c (compared with placebo‐treated patients) among individuals who received three to four injections of GAD‐alum, our findings certainly suggest benefits of therapeutically preserved C‐peptide on blood glucose control within about 2 years of diagnosis.

The current analysis has certain limitations. It is a post hoc exploratory analysis whose results are considered hypothesis‐generating, not confirmative. No formal statistical significance testing was carried out and the observed associations should be considered preliminary until confirmation in a well‐powered prospective trial. The association between the treatment effect has not been reported for other investigational treatments, such as for other antigen‐specific therapies or immunosuppressive treatments. The findings may be specific to the antigen‐specific treatment used in the trials, and while all available randomized controlled trials of GAD‐alum with sufficient data have been included in the analysis, the findings still need to be independently validated in a separate dataset. The current meta‐analysis of previously published clinical trials of rhGAD65 also provides a concise summary of efficacy results of this treatment modality.

Overall, we show that preservation of C‐peptide using GAD‐alum correlates with effects on HbA1c in individuals with recent‐onset T1D carrying HLA DR3‐DQ2. These findings will be evaluated in a confirmatory phase III trial (DIAGNODE‐3) and support using C‐peptide as a clinically relevant surrogate endpoint for endogenous insulin production‐preserving therapies.

AUTHOR CONTRIBUTIONS

CN initiated, and CN, UH and JL planned and developed the work. CN carried out the analyses and wrote the first draft, and CN, UH and JL critically revised the manuscript. CN, UH and JL have verified the underlying data and all authors have had full access to the data.

CONFLICT OF INTEREST

UH and CN are employed by Diamyd Medical and own stock in the company. JL has received unrestricted grants from Diamyd Medical, was earlier a member of Provention Bio, Inc.′s Advisory Council and is a present member of Dompé Farmaceutici S.p.A.'s International Advisory Board.

PEER REVIEW

The peer review history for this article is available at https://publons.com/publon/10.1111/dom.14720.

HLA subgroup	NCT00435981	NCT00529399	NCT00723411	NCT03345004	Total
DR3‐DQ2 N (% study, % total)	34 (49%, 11%)	71 (51%, 23%)	161 (51%, 51%)	47 (44%, 15%)	313
Not DR3‐DQ2 N (% study, % total)	35 (51%, 11%)	68 (49%, 22%)	152 (49%, 48%)	59 (53%, 9.4%)	314