Point-of-Care Ammonia Monitoring from a Single Drop of Blood

​Thomas Veltman, Ph.D., Aza Technology, Inc.
tv@azatechnology.com

​In an earlier article in this newsletter, I described my efforts, along with a team of other researchers, towards producing a point-of-care ammonia meter with functionality similar to the well-known blood glucose meter.  In June, we published a cover article in ACS Sensors (doi:10.1021/acssensors.0c00480) showcasing this technology and the performance of a prototype meter and consumable cartridges.  The technology demonstrated excellent agreement with the existing plasma ammonia test performed on real patients at Lucille Packard Children’s Hospital and Stanford Hospital. The cartridges enable rapid (~1 minute) and accurate ammonia measurement from a single drop of blood with values between 10 and 1000 µM.
 
Spurred by the success of our research, we founded Aza Technology to commercialize the technology and hired CEO Peter Karkantis. Peter is an Abbott Laboratories veteran executive with extensive experience in point-of care glucose testing. Thanks to the support of the OAA, Aza Technology was able to begin work on developing a cartridge design for manufacturability.  We have recently constructed a system for producing cartridge components in the thousands of units. In parallel, we have designed and built a feature-complete meter and have begun providing units to potential users for evaluation and feedback. Our new instrument and cartridges are designed to be manufacturable on scale, allowing us to reach the patients that need it most. We are currently developing strategies for FDA evaluation, and seeking partners and funding for full commercialization.

From the Fall 2020 OAA Newsletter

Point-of-Care Ammonia Monitoring from a Single Drop of Blood

Thomas R. Veltman, Department of Chemistry, Stanford University, Natalia Gomez-Ospina, and Matthew W. Kanan, Department of Pediatrics – Medical Genetics, Stanford School of Medicine
 

Hyperammonemia (elevated blood ammonia) frequently occurs secondary to organic acidemia and is responsible for a large portion of brain damage and death in these patients.[1] The underlying cause is thought to be the accumulation of organic acyl-CoA derivatives,[2],[3] which inhibit the enzyme N-Acetylglutamate Synthetase and deplete acetyl CoA. Both of these effects cause a reduction of N-acetylglutamate levels, which results in dysregulation of the normal nitrogen metabolism that is responsible for converting ammonia to urea for subsequent excretion.  Timely intervention to reduce ammonia levels is essential to improve neurodevelopmental outcomes,[4],[5],[6] but diagnosing hyperammonemic infants before an acute crisis is extremely challenging with the standard clinical ammonia test. The standard assay requires intravenous access for at least 1 mL of blood and must be performed in specialized clinical labs. Obtaining samples requires an extremely skilled phlebotomist since accurate values can only be obtained from draws performed without a tourniquet, and patient distress or a slow-flowing venipuncture can contribute to inaccuracies.  Additionally, blood is unstable and releases ammonia over time, and therefore samples must be transported on ice and processed rapidly to avoid false elevations. 

Patients diagnosed with acidemia require lifelong management to minimize hyperammonemic crises. Current clinical guidelines indicate that suspected patients should be sent to a metabolic treatment center as soon as possible, and transport teams should be equipped with ammonia scavengers and Carbaglu to continue management.[7] Thus, any suspected hyperammonemic crisis may result in transport to a metabolic center.  Frequently, patient families may be hours away from such a center. Finally, uncertainty in a patient’s ammonia status is a source of significant stress for parents. The symptoms of hyperammonemia are non-specific, and caloric management can be difficult when children are physically active. Illness can also be a trigger for hyperammonemia, and parents frequently report concern over the uncertainty of the wellbeing of their children.

Our work has focused on addressing all of the above issues through the development of a point-of-care ammonia testing device.  The availability of a user-friendly and accurate ammonia monitor is of longstanding interest to the organic acidemia community, but technical challenges have heretofore kept other proposed devices from achieving success.  Our breakthrough came with the recognition that ammonia could be measured in small volumes of whole blood simply by alkalizing the blood sample and monitoring the concentration of ammonia released from the sample.  We have built a series of prototypes to validate this detection scheme, culminating in a handheld device that produces accurate ammonia readings in less than a minute from a single drop of blood.  The device is as easy to use as a blood glucose meter, and because we are detecting ammonia in the gas phase, typical interferents in the blood that can cause inaccurate readings with the conventional plasma ammonia test do not influence our results.  Furthermore, the immediate readout eliminates complications from sample handling and permits the immediate evaluation of the patient’s status with respect to their ammonia treatment regimen.  With the device in hand, we were further motivated to devise a measurement protocol that would provide the easiest access to patient samples with a minimum of effort and as little discomfort as possible, as routine venipuncture can be challenging and traumatic for acidemia patients.  Fortunately, we discovered that sampling with a skin prick from the underside of the patient’s earlobe is completely painless and produces ammonia values that match those measured from a concurrent venous draw.  For patients who have an indwelling line placed, access is not as significant of a barrier, however indwelling lines come with an increased risk of infection, and therefore their use is not appropriate in all settings.  In the future, we envision shifting the treatment paradigm to closer monitoring of ammonia levels in acidemia patients with the objective of reducing hospitalization frequency and parent stress while improving the quality of life for affected families.

[1] Filipowicz, Heather R.; Ernst, Sharon L.; Ashurst, Carrie L.; Pasquali, Marzia; Longo, Nicola. Metabolic changes associated with hyperammonemia in patients with propionic acidemia. Molecular Genetics and Metabolism. 88 (2006) 123-130.

[2] Coude, Francois X.; Sweetman, Lawrence; Nyhan, William L. Inhibition by propionyl-coenzyme A of N-acetylglutamate synthetase in rat liver mitochondria. J. Clin. Invest. 64 (1979) 1544-1551.

[3] Stewart, Peter M.; Walser, Mackenzie. Failure of the normal ureagenic response to amino acids in organic acid-loaded rats: proposed mechanism for the hyperammonemia of propionic and methylmalonic acidemia. J. Clin. Invest. 66 (1980) 484-492.

[4] Msall, Michael; Batshaw, Mark L.; Suss, Richard; Brushlow, Saul W.; Mellits, E. David. Neurologic outcome in children with inborn errors of urea synthesis. N. Engl. J. Med. 310 (1984) 1500-1505.

[5] Picca, Stefano; Dionisi-Vici, Carlo; Abeni, Damiano; Pastore, Anna; Rizzo, Cristiano; Orzalesi, Marcello; Sabetta, Gaetano; Rizzoni, Gianfranco; Bartuli, Andrea. Extracorporeal dialysis in neonatal hyperammonemia: modalities and prognostic indicators. Pediatr. Nephrol. 16 (2001) 862-867.

[6] Vergano, Samantha A.; Crossette, Jonathan M.; Cusick, Frederick C.; Desai, Bimal R.; Deardorff, Matthew A.; Sondheimer, Neal. Improving surveillance for hyperammonemia in the newborn. Mol. Genet. Metab. 110 (2013) 102-105.

[7] Chapman, Kimberly A.; Gropman, Andrea; MacLeod, Erin; Stagni, Kathy; Summar, Marshall L.; Ueda, Keiko; Mew, Nicholas Ah; Franks, Jill; Island, Eddie; Matern, Dietrich; Pena, Loren; Smith, Brittany; Sutton, V. Reid; Urv, Tiina; Venditti, Charles; Chakrapani, Anupam. Acute management of propionic acidemia. Mol. Genet. Metab. 105 (2012) 16-25.

​From the Spring, 2019 OAA Newsletter

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