Using Statistical Modeling To Establish Reference Intervals for Pediatric Patients

Establishing accurate reference intervals for pediatric patients is notoriously difficult. Traditional direct studies require drawing blood samples from healthy children who don't have a medical need, which is extremely challenging. However, age-based reference intervals are essential to accurately diagnose and treat pediatric patients.

To solve this challenge, Kelly Doyle, PhD, DABCC, FADLM, and fellow colleagues at ARUP Laboratories have turned instead to statistical modeling.

"At ARUP, we've tested tens of thousands of pediatric patients. We can use novel, state-of-the-art statistical modeling techniques to look at the distribution of results in each of these age brackets," Doyle said.

Using these techniques, ARUP Laboratories has validated age-specific reference intervals for pediatric patients who have glucose-6-phosphate dehydrogenase (G6PD) deficiency. G6PD is an enzyme that protects red blood cells from oxidative stress, and G6PD deficiency is one of the most common genetic deficiencies, affecting around 400 million people worldwide, said Doyle. Many of those affected don't experience symptoms unless they are exposed to a trigger, such as certain foods or medications.

"Individuals who have G6PD deficiency cannot generate sufficient amounts of antioxidant in their red blood cells, which are then susceptible to damage and rupture, leading to hemolytic anemia," Doyle said.

Identifying G6PD deficiency early on can help these individuals avoid triggers that would cause symptoms.

In newborns, G6PD deficiency can have more severe consequences, such as bilirubin neurotoxicity, which occurs when bilirubin-a byproduct of damaged red blood cells-builds up in the brain.

"These cases of hyperbilirubinemia in pediatric patients can cascade or precipitate into much more severe forms of harm," Doyle said. One example is a condition called kernicterus, in which "the large amount of bilirubin in the brain can severely affect neurological function-not just in the short term; it can lead to irreversible damage."

For these newborns, it's critical to identify G6PD deficiency early and initiate therapy to prevent lasting damage.

Newborns have higher amounts of G6PD activity than adults, which presents a problem when using adult ranges to measure potentially abnormal values in a newborn.

"If the reference interval is shifted lower, but pediatric patients typically show higher levels of activity, then a partially affected individual will appear normal by an adult range," Doyle said.

As an X-linked genetic disorder, G6PD deficiency presents with varying levels of severity and affects male patients more than female patients. Without age-specific reference intervals, it may be easier to miss a diagnosis for a patient who is only moderately affected.

"These new ranges allow us to detect abnormally low individuals accurately per each of the age brackets," Doyle said. The age brackets include four divisions for infants under one year old, as well as a bracket for children who are 1-17 years old and one for adults who are 18 years or older.

As part of the study, Doyle and his colleagues were able to overlay phenotype data with genotype findings for over 100 pediatric patients, several of whom had known deficiencies, to verify the accuracy of the proposed ranges.

"By overlaying these findings on top of the established reference intervals, we can show that we're calling more individuals appropriately with these new ranges," Doyle said.

ARUP is an ISO 15189-accredited laboratory, and regular review of reference intervals is one of the criteria to achieve and maintain accreditation.

"Many think it's an impossibility to do routine reference interval studies because it's so expensive and so challenging," Doyle said. "This novel method provides a key opportunity to improve reference intervals for pediatric patients."

Doyle has already collaborated on several studies to establish reference intervals using indirect methods, including fibrinogen ratio cutoff limits and fractionated metanephrines.

On December 17, Doyle will present a webinar, "Development of Pediatric G6PD Reference Intervals Through Integration of Indirect Methods and Molecular Data," on the development of pediatric G6PD reference intervals using indirect statistical methods. Registration details are available here.

Kellie Carrigan, [email protected]