Assessment of a Novel Dielectric Microsensor for Monitoring Coagulation Factor Therapy in Children with Hemophilia with and without Inhibitors

Introduction: Monitoring coagulation factor or bypassing agent therapy has important clinical implications for the management of children with hemophilia, especially those with inhibitors. For patients with hemophilia, conventional laboratory-based coagulation tests are time-consuming, labor-intensive, and costly. Moreover, no monitoring assay is currently available for use with bypassing agents in patients with hemophilia and inhibitors, except for thromboelastography or rotational thromboelastometry (ROTEM), both of which are not widely available clinically and are labor-intensive. There is a need to develop an easy-to-use, low-cost, portable global assay of coagulation for monitoring therapies in hemophilia. The aim of this study was to assess the clinical utility of monitoring replacement therapy using a new technique based on a novel dielectric microsensor (ClotChip) that can potentially enable global blood coagulation assessment in a miniaturized, portable measurement platform.

Methods: Patients accrued for the study were children with severe hemophilia A & B (n = 6), children with hemophilia and inhibitors (n = 4), and normal, healthy children without coagulopathy (n = 22). Blood samples were obtained by venipuncture into collection tubes containing 3.2% sodium citrate anticoagulant. Samples from patients with hemophilia were collected both prior to the initiation of replacement therapy and 15 minutes after administration of an appropriate coagulation factor VIII or IX for severe hemophilia A or B, respectively, or a bypassing agent (FEIBA or recombinant factor VIIa) for hemophilia with inhibitors. Whole blood samples were then tested with the ClotChip within 2 hours of collection, and coagulation was induced with CaCl₂ prior to the start of ClotChip measurements. The ClotChip is based on the fully electrical technique of dielectric spectroscopy (DS), and features a three-dimensional, parallel-plate, capacitive sensing structure integrated into a low-cost (material cost < $1), small- sized (26mm × 9mm × 3mm), and disposable microfluidic biochip with miniscule sample volume (< 10µL). The ClotChip curves were calculated as the real part of blood permittivity at 1MHz, and the time to reach a peak in permittivity (T_peak, Fig 1A) was taken as an indication of coagulation time. Samples from 8 of the 10 hemophilia patients were also tested with ROTEM using the non-activated TEM (NATEM) test according to the manufacturer's direction. The ROTEM clotting time (CT) parameter (Fig 1B) was used in this study as a clinically important parameter to monitor patients with hemophilia.

Results: We observed a significant decrease in the ClotChip T_peak parameter for post-therapy samples compared to baseline (pre-therapy) for children with hemophilia without inhibitors (Fig 1C, n = 6, p = 0.0001) and with inhibitors (Fig 1D, n = 4, p = 0.0024). Interestingly, baseline ClotChip measurements for 2 tolerized patients with inhibitors exhibited T_peak of 970s and 1,010s, which are within the range of T_peak for patients without inhibitors (750s - 1,320s), while the baseline ClotChip measurements of 2 non-tolerized patients with inhibitors exhibited higher T_peak of 1,810s and 2,100s (Fig 1E). Moreover, ClotChip measurements for all samples taken post-therapy exhibited T_peak that fell within the reference range of T_peak for normal, healthy children (180s - 720s, Fig 1E). Taken together, these data suggest that the ClotChip T_peak parameter is sensitive to detection and correction of coagulopathy in children with hemophilia with and without inhibitors, as indicated by T_peak reaching reference range levels after coagulation factor therapy. Finally, the ClotChip T_peak exhibited a very strong correlation (Fig 1F, Pearson's r = 0.94) to the clinically important CT parameter of ROTEM.

Conclusions: Our studies demonstrate the feasibility of monitoring coagulation factor therapy in children with hemophilia with and without inhibitors, using a novel, microfluidic, dielectric sensor (ClotChip), allowing whole blood assessment of hemostasis in a single disposable sensor. ClotChip has potential to fulfill an important unmet clinical need for monitoring therapy, especially for hemophilia with inhibitors. Further studies are underway to assess the ability of ClotChip to differentiate bleeding phenotypes and monitor outcomes for treatment of bleeds in children with hemophilia.

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