Urine studies yield valuable diagnostic insights

June 22, 2016

As Leslie Williams’ article acknowledges, urinalysis is something of an old standby of the clinical laboratory. But interesting recent studies suggest that urine testing may have unanticipated diagnostic value, with significant future applications. Here’s a review of three examples of recent research.

Urine and preeclampsia

A urine sample could be used to diagnose a complex and serious pregnancy disorder: a team of Russian scientists has developed an approach for a method of non-invasive testing for pregnant women with preeclampsia. They have found potential biomarkers in the urine of pregnant women. In the future, this urine-based diagnostic method may enable specialists to detect the disease in its early stages. A paper detailing the results was recently published in the Journal of Proteomics.

Preeclampsia is a multisystem disorder that occurs in the second half of pregnancy and is characterized by high blood pressure and proteinuria (a significant increase in the normal level of protein in the urine). Women with the condition suffer headaches, swelling, fainting, and other unpleasant and often dangerous symptoms. Ultimately, preeclampsia puts the health and life of an expectant mother and her baby at serious risk. The symptoms of the condition are ambiguous, so doctors rarely diagnose “pure” preeclampsia. However, even if this diagnosis is made, a new problem arises: It is still not known what causes the disorder, and therefore it’s difficult to control it effectively.

That means that doctors are only able to prescribe supportive care and treat the symptoms, while trying to delay the birth of the child. A balance has to be struck between allowing enough time for full fetal growth (although the baby is also affected by improper development of the placenta as a result of the condition) and preserving the mother’s health.

If something is wrong with all the systems in the body, it is logical to assume that the “fault” is at the molecular level. Protein is found even in the urine of a healthy person, although only in very small concentrations. From a doctor’s point of view, one of the main symptoms of preeclampsia is a marked increase in this concentration (from the normal 0.03 g/l to the critical 0.3 g/l and above). The Russian researchers wondered whether peptides (pieces of proteins) that are produced by women with preeclampsia could carry information about the disease. This approach is very much in line with the current focus on non-invasive testing, i.e., analyzing available biomaterials such as urine, saliva, or exhaled air.

To identify potential peptide biomarkers, the researchers compared samples of three groups of ten women: women with a normal pregnancy, and with mild and severe preeclampsia. The researchers were not only interested in comparing healthy women and patients with preeclampsia; they also wanted to examine how certain biomarkers are associated with the severity of the condition. Following the experiments, 35 potential peptide biomarkers of preeclampsia were identified. They included fragments of alpha-1-antitrypsin (14 peptides), collagen alpha-1(I) and alpha-1(III) chains (6 peptides), and uromodulin (7 peptides).

“We were able to confirm a number of markers previously proposed by our colleagues abroad, and also identify some new ones,” says Evgeniy Nikolaev, a study co-author. “This non-invasive method has proven effective; it can be used as a basis to develop a clinical method.”

Urine and high-grade prostate cancer

An experimental urine test that detects genetic changes associated with prostate cancer identified 92 percent of men with elevated PSA (prostate-specific antigen) levels who had high-grade cancers, according to a study published in JAMA Oncology online.

“The test has the potential to be a significant improvement over PSA alone in distinguishing between low- and high-grade prostate cancer, especially in the PSA gray zone patient,” says first author James McKiernan, MD. In addition, the test is the only urine-based assay that does not require a digital rectal exam prior to collection and is easily integrated in the clinic environment.

Although the PSA blood test is commonly used to screen for prostate cancer, its value has come under question. An elevated PSA level—above 4 ng/mL—only indicates that a patient may have cancer and does not reliably distinguish between low-grade cancer, which can be monitored without active treatment, and high-grade disease, which requires aggressive treatment with surgery or radiation therapy. And, because PSA tests yield a high number of false positive results—only 25 percent of men with an elevated PSA level have prostate cancer—the U.S. Preventive Services Task Force recommends against PSA-based screening.

The only way to definitively diagnose prostate cancer is with a tissue biopsy, a painful procedure that carries a substantial risk of bleeding and infection and a very small chance of death.

In the study, prostate biopsy results from 774 men with PSA levels between 2 and 10 were compared with a composite score based on results of the urine test plus other risk factors, including PSA level, age, race, and family history of the disease.

Using data from 255 of the men, the researchers first set a cut-off score of 15.6 that identified over 90 percent of men found to have high-grade cancer. The researchers then assessed the test’s performance among the remaining 519 men using the designated cut-off score. A score above 15.6 was used to predict the presence of high-grade cancer (Gleason Score greater or equal to 7); a score equal to or below 15.6 was used to predict the presence of low-grade cancer or no disease.

This analysis showed that the test correctly identified 92 percent of men with high-grade cancer. However, the test also predicted high-grade cancer in 66 percent of men whose biopsies revealed low-grade or no cancer. In clinical practice, use of the test would have spared 27 percent of men from having an unnecessary prostate biopsy.

Among the 138 men who received a low test score, predicting low-grade or no cancer, 91 percent had no cancer or low-grade cancer that didn’t require immediate treatment. “By adding this test to our evaluation, men who receive a low score could potentially choose to forgo a biopsy,” McKiernan says.

Of the 12 men who had high-grade cancer but received a low test score, nine had moderately aggressive cancer that would likely have been detected with follow-up monitoring. The remaining three had a higher risk of cancer.

The test, called ExoDxTM Prostate (IntelliScore), detects RNA from three genes (ERG, PCA3, and SPDEF) that have been linked to the development and progression of prostate cancer. The RNA is encapsulated in lipid membrane-coated structures called exosomes that are excreted by cancer cells into urine. The test does not require a digital rectal exam or prostate massage before sample collection. Additional clinical trials to obtain FDA approval are being planned.

Urine and tuberculosis

A low cost, easy-to-use urine test to diagnose tuberculosis (TB) among patients with HIV could help reduce the TB death rate of HIV-positive patients in the hospital, according to a new study published in The Lancet.

In Africa, nearly 40 percent of all adult deaths related to HIV or AIDS are due to TB, but almost half of the TB cases remain undiagnosed and untreated before death. The authors of the study, which was conducted in 10 hospitals in sub-Saharan Africa, say that if implemented more widely, this low-cost intervention could save thousands of lives per year.

“This is the first trial of any diagnostic test for tuberculosis to show a reduction in the number of deaths. The reduction in mortality is likely to be because urine-testing, in conjunction with routine testing, resulted in a greater proportion of patients starting tuberculosis treatment early,” explains senior author and project supervisor Professor Keertan Dheda from the University of Cape Town in South Africa.

He adds: “When used in conjunction with routine testing, urine-testing for tuberculosis reduced the TB death rate of HIV patients in the hospital. Importantly, we found that the test was particularly effective in identifying tuberculosis among patients with advanced HIV infection who are most vulnerable to advanced TB disease. The absolute reduction in mortality was small at four percent, but with 300,000 patients with HIV dying from tuberculosis in Africa every year, implementing this low-cost, rapid, bedside test could potentially save thousands of lives annually.”

Tuberculosis is the leading cause of death in people with HIV in low- and middle-income countries, and hospitals are often overwhelmed with HIV-positive individuals who have suspected TB. Diagnosis of TB usually includes taking a chest X-ray and microbiological examination of sputum. However, for severely ill patients with TB and HIV, it can be hard to produce sputum, and alternative approaches (sputum induction or alternative invasive sampling) are often unavailable. The test detects a glycolipid molecule (lipoarabinomannan, or LAM), which is linked to TB. It provides a result in 25 minutes and each test costs about $2.66.

Dheda says: “Since November 2015, a test measuring urine LAM has been recommended by WHO. However, the recommendation was conditional, meaning it is up to the doctor or health system to decide whether or not to use the test. These data make use of the test more compelling and suggest that scale up and roll out of the test is now required.”

In this study, the research team randomly allocated 2,528 patients with HIV from ten hospitals in four countries in sub-Saharan Africa (four in South Africa, two in Tanzania, two in Zambia, and two in Zimbabwe) to receive either routine testing (smear, Xpert MTP/RIF and culture) as well as the LAM urine-test (LAM group, 1257 patients) or routine testing alone (no-LAM group, 1271 patients).

Eight weeks after being discharged from the hospital, 21 percent (261) of patients in the LAM group had died compared to 25 percent (317) of patients in the non-LAM group—an absolute reduction of four percent. A greater proportion of patients in the LAM group (648; 52 percent) were treated for TB than in the no-LAM group (598; 47 percent). And, of those who started TB treatment, a higher proportion of patients were treated in the first three days in the LAM group (513/648; 79 percent) compared to the non-LAM group (413/598; 69 percent).

The effect of LAM testing on mortality varied by country, but the highest sensitivity was found in people with the lowest CD4 cell count (a measure of how badly the immune system is damaged in patients with HIV), who are the hardest to diagnose with TB using routine testing. The authors found that the LAM urine-test had a sensitivity of 46 percent (the proportion of people with TB correctly diagnosed) and specificity of 90 percent (the proportion of healthy people who are correctly identified as healthy).