Analytical and clinical validation of biomarkers of beta cell damage in type 1 diabetes (IWT621)

Background and overall aim: Type 1 diabetes (T1DM) can be treated by transplantation of insulin-producing beta cells procured from donor pancreases. A major problem today is that such primary islet grafts are scarce. Moreover, transplantation techniques are suboptimal since up to 50% of grafted cells die shortly after implantation by hypoxia and inflammation. Final outcome of transplant therapy can be measured clinically by classical metabolic indices (HbA1c, glycemic variability) but there is a clear need for intermediate end-point markers that provide a more direct and real-time measurement of the on-going graft destruction. Use of such early intermediate end-point biomarker in clinical trials of cell therapy is strongly advocated by both EMEA and FDA, since it allows a more objective evaluation of transplant regimens, and more timely adaptation of immunosuppressive or anti-inflammatory therapy. Therefore our overall aim is to develop a simple, highly-sensitive clinical test for real-time quantification of beta cell destruction in vivo. This test will be routinely applied in islet transplantation programs. Key words describing its anticipated performance are: femtomolar sensitivity, robustness through biomarker multiplexing, miniaturization, ease of blood sampling and high-throughput. Prior work and embedment in the research context: The research will be conducted in the Diabetes Research Center (DRC) at VUB-University Medical Center (UMC). The DRC implements novel types of diabetes therapy through immune modulation (anti-CD3) and beta cell transplantation, and has taken a lead in the exploration of beta cell regenerative medicine. More specifically, our tests are developed in the Clinical Biology of Diabetes Unit, which serves as core laboratory for the DRC, in close collaboration with both clinicians and animal experimentalists, which ensures good access to both patient and animal samples. Current IWT project builds further on prior data collected with support by FWO, and that are now ready to translate into a clinical and economic finality. We have previously obtained proof-of-concept for our approach using a first biomarker, glutamic acid decarboxylase 65Kd (GAD65): we found that GAD65 was specifically discharged by damaged beta cells after transplantation. Its discharged level in plasma predicted long-term graft outcome, and thus correlated with the extent of graft destruction. For several reasons, GAD65 has limitations, which served as impetus to identify novel biomarkers using "OMICS"-technology. This resulted in two novel biomarkers for beta cell destruction (PPP1R1A and UCHL1) for which we also obtained proof-of-principle in vivo. With support by IWT we now want to translate these findings into a robust, sensitive high-throughput assay. Its intended first clinical use is the monitoring of islet transplantation. In addition its format and sensitivity will allow its evaluation as tool to detect the silent beta cell destruction in the preclinical phase of T1DM, which would further expand the test's clinical and commercial potential.
Technology and specific aims: As first line of research we will introduce novel technology to enhance the analytical sensitivity of our reference biomarker assay (GAD65). This will be done by developing bead-based immunoassays, and by introducing PCR amplification technology in the detection phase of GAD65. As second line of research, we will build sandwich immunoassays for the 2 novel biomarkers (UCHL1 and PPP1R1A) and check if their diagnostic sensitivity is superior to that of GAD65. Finally, we will merge both lines of research and propose a single assay that is robust through multiplexing of several biomarkers and shows femtomolar sensitivity. Combining both features will allow miniaturization on small sample volumes such as dried blood spots, which in turn will increase throughput and widely expand the application field through ease of sampling.

Keywords:Cell Therapy, Prevention, Transplantation, Diagnostic Tests, Immunology, Cell Death and Survival, Islet Cell Pathology, Islet Cell Biology, Beta Cell Transplantation, Diabetes

Disciplines:Cellular therapy, Immunology, Biochemistry and metabolism, Endocrinology and metabolic diseases, Laboratory medicine