Highlights

Focus on Boston Children's Proteomics Center

Biomarkers: The Clue to Individualized Patient Care

Hanno Steen

Dr. Hanno Steen at Children’s Proteomics Center in collaboration with clinicians is unlocking the secrets behind disease states.

With healthcare costs on the rise, the government, consumers and payors are pushing to find ways to reduce costs. Pharmaceutical drugs in development currently have a very low success rate and response rates to existing medications in large patient populations is on average low. Improving the ability to pinpoint correct treatments for a specific patient through a personalized medicine approach could help decrease high costs and improve treatment effectiveness. “Theranostics” — the combination of a targeted therapeutic and a companion diagnostic takes into account an individual’s genetic makeup or levels of specific proteins expressed in tissue or fluids that coincide with a disease. The ability to direct drugs to the appropriate patients can increase drug R&D productivity by improving clinical trial design and its probability of success, decreasing trial size, reducing attrition rates, and increasing the speed to market. Genentech’s Herceptin drug and companion HER2 genetic test were the first approved theranostics combination approved in 1998. After Genentech’s success, many more pharmaceutical and biotech companies have started developing similar theranostics.

Proteomics represents a new frontier theranostic approach. Sensitive measurement of protein levels using proteomics, the science of analyzing the protein complement of a cell, tissue or body fluid, can often be a very effective way to identify protein biomarkers that correlate with different disease states or disease susceptibility. Understanding the proteome with a level of sophistication will ultimately translate to effective real time treatment decisions that could make a difference in a patient’s life. However, understanding the proteome is technically challenging based on the selectivity and sensitivity of the measurements that are needed. At Children’s Hospital Boston’s unique Proteomics Center, the only center at a New England pediatric research enterprise, researchers are accelerating the pace of understanding and discovery of proteomic biomarkers.

The Proteomics Center at Children’s

Directed by Hanno Steen, PhD, a leading researcher in the field of proteomics, Children’s Proteomics Center has developed an advanced multidisciplinary effort that leverages the hospital’s resources, including the Center's own computational team, its unique patient population, and its leading clinicians and scientists with an emphasis on translating these findings into clinical use. Children’s state-of-the-art proteomics facility uses customized and cutting-edge proteomics screening and analysis such as the latest equipment for protein separation and several mass spectrometers, which detect, quantify and analyze proteins to help determine their structure and characteristics. Researchers at the center analyze the massive data output with powerful bioinformatics tools developed by Dr. Steen's team. The algorithms help process what would otherwise be overwhelming sets of data so that clinical meaning can be abstracted. The Center typically collaborates with Children’s own investigators, where a clinician or researcher will have a disease he or she wishes to study, and Dr. Steen will recommend a technology or method to achieve the desired results. The output from these collaborations is usually a short list of protein biomarkers that are clinically relevant and could yield big dividends in both pediatric and adult medicine. “The mix of research and clinical expertise helps drive the projects, as they learn to understand each other better and broaden each other’s knowledge,” said Dr. Steen. Some recent findings from collaborations with the Center are highlighted below.

Studies at Children’s



Appendicitis Diagnostic

Appendicitis is the most common childhood surgical emergency, but the diagnosis can be challenging, often leading to either unnecessary surgery when appendicitis isn’t present, or a ruptured appendix and serious complications when the condition is missed. The lifetime risk is 5-10 percent for the disease and even at top tier hospitals, there is still a false discovery rate of 5-10 percent. The consequences of misdiagnosis are severe, and increasingly expensive diagnostic tests have become the standard of care.

In a collaborative project with Emergency Medicine, Alex Kentsis, MD, PhD, fellow in Hematology/Oncology and once a clinician-in-residence at the Proteomics Center, Richard Bachur, MD, chief of Emergency Medicine, and Dr. Steen, the center has identified protein biomarkers in patient urine that distinguish acute appendicitis from other conditions. The marker discovery was based on deep proteomic analysis of patient urine, assessing over 2,000 proteins by mass spectroscopy. The strongest single biomarker, leucine-rich alpha-2-glycoprotein (LRG), showed a tremendous increase in abundance during the progression of appendix inflammation.

With an award from Children’s Technology Development Fund, the team will translate the LRG test to an antibody based platform, and will further validate this biomarker for appendicitis. The clinical laboratory and point-of-care immunoassay tests would be valuable in reducing expensive diagnostic scans, unnecessary surgeries and the number of cases that progress to rupture before surgery, all leading to improved patient outcome. The goal is to develop a dip stick test that will be able to diagnose appendicitis in minutes. “Many hospitals around the U.S. do not have CT or ultrasound, and in those areas, a dipstick test would be of great value,” said Dr. Steen.

Kawasaki Disease Diagnostic

In collaboration with Susan Kim, MD, MMSc, of Boston Children's Department of Rheumatology, the center began the hunt for biomarkers that act as a red flag for Kawasaki Disease by analyzing urine samples. Kawasaki disease is the most common cause of acquired pediatric heart disease in the industrialized world, including the United States. Without timely treatment, this condition can lead to inflammation of blood vessels, including the heart's arteries, creating the risk for aneurysms and possible death.

Unfortunately, there is no rapid test that can diagnose the condition. Because the disease's common clinical features, such as rash, lymph node enlargement, conjunctivitis and persistent fevers that resemble other infections, doctors often cannot distinguish Kawasaki disease from many other possible conditions. In addition, there may often be a delay in diagnosis, since not all children will show the classical clinical manifestations of Kawasaki disease. This is precious time needed for treatment and protecting the heart. "Without treatment, about 25 percent of children can develop coronary artery aneurysms," says Dr. Kim. A tool capable of diagnosing Kawasaki disease by day five of a fever or before all the clinical features are evident could improve a patient's outcome.

Drs. Steen and Kim will continue to build the molecular profile for Kawasaki disease by studying the urine of children with the condition, before and after treatment. They will compare these samples to urine samples from children with symptoms similar to Kawasaki disease, but stemming from a known, separate condition.

Urinary Markers of Urinary Tract Obstruction and Vesicoureteral Reflux

Vesicoureteral Reflux (VUR) is diagnosed in 17–37 percent of all prenatal ultrasounds. The condition is more prevalent in females and in children who have red hair. One-third of Urinary Tract Obstructions (UTI) in children are caused by vesicoureteral reflux. Unfortunately, there are no appropriate guidelines or indicators to determine which children are at risk for renal damage or who should be tested, and postnatal tests are invasive and involve radiation.

Richard Lee, MD, assistant in Urology and a former clinician-in-resident at the Proteomics Center, is focused on identifying clinically significant urinary markers of urinary tract obstruction (UTO) and vesicoureteral reflux (VUR) in urine. The biomarkers he discovered through Proteomics program will be helpful in determining which children with these conditions require either surgical or medical intervention or observation. The findings were originally studied in rats and now being studied in human samples. Through development, the group hopes to develop a high throughput immunoassay test. Through his research, he has also developed a translational platform for discovery based quantitative urinary proteomics that is faster and has improved results than existing platforms. Additionally, through his clinical interactions, he has started the PUPPI project, which is an assembled unique urinary specimen repository from healthy children and children with urological conditions.

Distinguishing Benign from Cancerous Prostate Disease

Marsha Moses, PhD, the newly appointed Director of the Folkman Research Institute, is aggressively researching cancer’s most dangerous stages – the earliest, when tumors first develop the ability to grow and spread, and the last, metastasis, when cancer spreads throughout the body. Dr. Moses aims to develop a simple, noninvasive urine test to distinguish benign from cancerous prostate disease. Her goal is to launch a large-scale, multi-institutional clinical trial. This singular achievement could become the first noninvasive test ever to distinguish cancer from benign disease.

Working with partners outside of the Center, Dr. Moses’ team has identified novel urinary proteins present at different levels in the urine of men with BPH and those with prostate cancer. All were startled that a remarkably small number of proteins – just 26 – significantly differentiated benign BPH and prostate cancer. More typically, such studies result in hundreds of differentiators, most of which turn out to be nonspecific to the condition under investigation. These exciting findings provide the first evidence that distinguishing between these two diseases may be possible, ultimately leading to more precise and specific treatment of each.

Dr. Moses’ next step is to validate these initial findings in a larger number of samples. To perform the validation studies, she and her team will systematically analyze each of the urine samples for the presence of the novel panel of potential markers. They will use state-of-the-art methodologies with which they have significant experience, including ELISA and immunoblot analysis, among others.

Moving the technology to the clinic

While these discoveries have the potential to improve diagnostic and therapeutic effectiveness, any findings that come out of the Center will need large scale validation studies to move the technology into the clinic. This represents a great opportunity for a partnership with industry partners. Children’s multidepartment knowledge sharing mixed with industry resources and development will streamline these exciting findings into validated products ready for clinical use. Both Children’s and a potential partner would each benefit from such collaborations. “We have access to the samples, are great at doing the discovery work, and have a regulatory platform (IRB), but after the discovery, a corporate partner is needed to provide the large high throughput screening validation platform for larger validation studies,” said Dr. Steen. Children’s is actively seeking such partnerships through its Technology & Innovation Development Office to facilitate validated biomarker development that could contribute to personalized medicine. Through such partnerships, Children’s Proteomics Center’s is in position to contribute heavily to the field of personalized medicine and to excellence in patient care.

Related Links

  1. Boston Children's Proteomics Center Website
  2. Introduction to Proteomics
  3. Virtual Proteomics Lab Tour
  4. A Urine Test for Appendicitis
  5. A Dipstick Test for Breast Cancer?
  6. Search for Boston Children's Proteomic Technologies
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