May 12 , 2005

In This Issue . . .

Before the Nick of Time: Evaluating NGAL as a Biomarker for Acute Renal Failure
By Richard A. Pizzi

Acute renal failure (ARF) wreaks havoc on a wide variety of hospital populations—from 5% of all admitted patients, to up to 40% of post-cardiac surgery patients, and 30-50% of those in intensive care units—many of whom never recover. ARF’s high mortality rates are largely blamed on clinicians’ inability to diagnose the disorder in its earliest stages. Like other diagnostic quagmires, many clinicians believe that if a biomarker existed, physicians might be able to initiate preventive and therapeutic treatment within a few hours after ARF onset. In pursuit of such a biomarker, one candidate has emerged—neutrophil gelatinase-associated lipocalin (NGAL). This month, Strategies looks at some recent NGAL research and what laboratory directors need to know about its potential impact on the development of diagnostic tests for ARF.

Current clinical diagnosis of ARF involves measuring serum creatinine, even though this measurement can be unreliable because it will not accurately reflect kidney function until a steady state is reached, which usually takes several days. By that time, over 50% of kidney function may be lost.

The unreliability of serum creatinine prompted researchers at Cincinnati Children’s Hospital Medical Center (CCHMC) to evaluate NGAL as a potential biomarker, as reported in a recent issue of The Lancet. The CCHMC team was interested in NGAL because it is a protein product of kidney genes that had already been detected in urine after ischemic renal injury in rodents and humans.

“We identified NGAL as one of the most dramatically up-regulated genes and proteins in the kidney after ischemia,” noted Prasad Devarajan, MD, Director of Nephrology, CCHMC, and senior author of the study. The researchers had previously worked with animal models and utilized a genome-wide interrogation strategy to identify kidney genes that are induced very early after ischemia, he explained. They then decided to test their hypothesis that NGAL was an early biomarker in a human population.

ARF Following Cardiac Surgery

The CCHMC study involved 71 children undergoing cardiopulmonary bypass surgery who had normal renal function prior to surgery. The researchers thought that these children were an ideal test case because ARF complicates up to 10% of cardiac surgical procedures in children with congenital heart disease. Because the children had no additional comorbid conditions such as atherosclerotic disease or diabetes, there were no other major variables that might influence the results.

Following cardiac surgery, ARF occurred in 28%—or 20—of the children within a three-day period. A rise in serum creatinine levels was detected within one to three days after surgery, which is typical for the traditional diagnostic method. But by using Western blot analysis and an ELISA procedure to measure NGAL in urine and serum, the CCHMC researchers discovered substantial increases in NGAL levels only two hours after surgery. Those children who developed ARF had, according to the results, a striking rise in urinary NGAL at all timepoints at which measurement was taken. In tests performed prior to surgery, these same children had normal kidney function and showed almost undetectable levels of urinary NGAL, indicating that the post-surgery NGAL assays were able to detect ARF one to three days before an increase in serum creatinine was detected. The most powerful independent predictor of ARF in this group of children was urinary NGAL two hours after surgery, the researchers concluded.

NGAL Questions

The CCHMC study clearly shows that the concentration of NGAL in urine and serum was raised significantly in children suffering ARF after cardiopulmonary bypass. But how broadly can the results be generalized? In a Lancet editorial on the results of the study, Stefan Herget-Rosenthal, MD, of the Division of Nephrology of the University Duisburg-Essen, Germany, advised against generalizing these results. Herget-Rosenthal wrote that ARF is “mainly multifactorial in origin and occurs more commonly in adults with pre-existing chronic renal disease,” adding that little is known about the stability of NGAL in serum and urine; and about accuracy, precision, and interferences in NGAL measurement.

While Devarajan and his colleagues acknowledge that ARF is multifactorial and that their results will need to be confirmed in larger population studies, they insist the results of the study are widely applicable. “First, we now have evidence [from other studies] that adults with acute kidney injury also display a tremendous increase in urine NGAL,” Devarajan said. “Second, patients with chronic renal injuries also have increased urine NGAL, but to a less degree than those with acute injuries.”

Devarajan also emphasizes that NGAL may have therapeutic as well as diagnostic importance. “We have shown in animal models of ischemic and nephrotoxic acute kidney injuries that NGAL administered before, during, or even after the acute injury can protect the kidney from functional damage,” he explained. “We propose that NGAL will emerge not only as an early biomarker of acute kidney injury, but also as a novel therapeutic intervention.”

Devarajan and his colleagues are also confident that concerns about the stability of NGAL in serum and urine and questions about the accuracy and precision of NGAL measurement can be addressed. “NGAL is protease resistant and is stable in the urine and serum for several days when the samples are refrigerated and for several months when the samples are frozen,” he said. “In the ELISA method we use, the inter- and intra-assay coefficient variations are less than 5%, indicative of excellent reliability.”

But what about applying NGAL in clinical scenarios, such as a commercially available NGAL assay in a diagnostic kit? It might not be far off, according to Devarajan. His research group is working with biopharmaceutical companies and hopes to have a commercially available NGAL kit in two to three years. It is likely that NGAL will be one of a “collection of strategically selected proteins” that may provide the “elusive ARF panel for the early and rapid diagnosis of acute kidney injury,” he added.

Richard A. Pizzi is a freelance writer based in Portland, Maine.

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