Fourth Annual Symposium on Nephrogenic Systemic Fibrosis and Gadolinium-Based Contrast Agents

Web Sites

American College of Radiology
The latest American College of Radiology guidelines for the use of GBCAs can be found in the Updated ACR Screening Recommendations on Gadolinium-Based MR Contrast Agents, Renal Disease Patients, and Nephrogenic Systemic Fibrosis (NSF).

Array Biopharma
Biopharmaceutical company focused on the discovery, development, and commercialization of targeted small molecule drugs to treat patients with cancer and inflammatory diseases.

Bayer Schering Pharma Diagnostic Imaging
A leading company in the contrast media market.

GE Healthcare
Leading company in medical diagnostics instrumentation.

The Global Fibrosis Foundation
A not-for-profit organization whose mission is to help educate patients, families and the medical community about Nephrogenic Systemic Fibrosis and other organ specific fibrosing processes.

Guerbet
Maker of the MRI imaging agent gadoterate, not currently available in the U.S.

The International Center for Nephrogenic Systemic Fibrosis Research (ICNSFR)
The Yale registry collects data on NSF cases and provides information and support to patients and their families.

National Kidney Foundation
The National Kidney Foundation guide "Nephrogenic Systemic Fibrosis: Reducing Risk" for professionals on how to reduce NSF risk in their patients.

NSF Support Group at Yahoo.com
A point of communication about the disease and an electronic forum to share ideas and possible treatment measures.

Journal Articles

Ali Abu-Alfa

Abu-Alfa A. 2008. The impact of NSF on the care of patients with kidney disease. J. Am. Coll. Radiol. 5: 45-52.

Saab G, Abu-Alfa A. 2007. Are patients with moderate renal failure at risk for developing nephrogenic systemic fibrosis? Radiology 244: 930-931; author reply 931-932. Full Text

Saab G, Abu-Alfa A. 2007. Will dialysis prevent the development of nephrogenic systemic fibrosis after gadolinium-based contrast administration? AJR Am. J. Roentgenol. 189: W169.

Saab G, Abu-Alfa A. 2008. Nephrogenic systemic fibrosis—implications for nephrologists. Eur. J. Radiol. 66: 208-212.

Weinreb JC, Abu-Alfa AK. 2009. Gadolinium-based contrast agents and nephrogenic systemic fibrosis: why did it happen and what have we learned? J. Magn. Reson. Imaging 30: 1236-1239.

Richard Bucala

Bucala R. 2008. Circulating fibrocytes: cellular basis for NSF. J. Am. Coll. Radiol. 5: 36-39.

Quan TE, Bucala R. 2007. Culture and analysis of circulating fibrocytes. Methods Mol. Med. 135: 423-434.

Quan TE, Cowper S, Wu SP, et al. 2004. Circulating fibrocytes: collagen-secreting cells of the peripheral blood. Int. J. Biochem. Cell Biol. 36: 598-606.

Quan TE, Cowper SE, Bucala R. 2006. The role of circulating fibrocytes in fibrosis. Curr. Rheumatol. Rep. 8: 145-150.

Vakil V, Sung JJ, Piecychna M, et al. 2009. Gadolinium-containing magnetic resonance image contrast agent promotes fibrocyte differentiation. J. Magn. Reson. Imaging 30: 1284-1288.

Herzog EL, Bucala R. 2010. Fibrocytes in health and disease. Exp. Hematol. 38: 548-56.

Shawn Cowper

Cowper SE. 2003. Nephrogenic fibrosing dermopathy: the first 6 years. Curr. Opin. Rheumatol. 15: 785-90.

Cowper SE. 2008. Gadolinium—is it to blame? J. Cutan. Pathol. 35: 520-522.

Cowper SE. 2008. Nephrogenic systemic fibrosis: an overview. J. Am. Coll. Radiol. 5: 23-28.

Cowper SE, Rabach M, Girardi M. 2008. Clinical and histological findings in nephrogenic systemic fibrosis. Eur. J. Radiol. 66: 191-199.

DeHoratius DM, Cowper SE. 2006. Nephrogenic systemic fibrosis: an emerging threat among renal patients. Semin. Dial. 19: 191-194.

Deo A, Fogel M, Cowper SE. 2007. Nephrogenic systemic fibrosis: a population study examining the relationship of disease development to gadolinium exposure. Clin. J. Am. Soc. Nephrol. 2: 264-267. Full Text

Introcaso CE, Hivnor C, Cowper S, et al. 2007. Nephrogenic fibrosing dermopathy/nephrogenic systemic fibrosis: a case series of nine patients and review of the literature. Int. J. Dermatol. 46: 447-452.

Knopp EA, Cowper SE. 2008. Nephrogenic systemic fibrosis: early recognition and treatment. Semin. Dial. 21: 123-128.

Michael Edward

Abraham JL, Edward M. 2009. Free gadolinium is a likely trigger of nephrogenic systemic fibrosis. AJR Am. J. Roentgenol. 193: W354; author reply W355.

Edward M, Fitzgerald L, Thind C, et al. 2007. Cutaneous mucinosis associated with dermatomyositis and nephrogenic fibrosing dermopathy: fibroblast hyaluronan synthesis and the effect of patient serum. Br. J. Dermatol. 156: 473-479.

Edward M, Quinn JA, Mukherjee S, et al. 2008. Gadodiamide contrast agent 'activates' fibroblasts: a possible cause of nephrogenic systemic fibrosis. J. Pathol. 214: 584-593.

Jack Gauldie

Ask K, Bonniaud P, Maass K, et al. 2008. Progressive pulmonary fibrosis is mediated by TGF-β isoform 1 but not TGF-β3. Int. J. Biochem. Cell Biol. 40: 484-495. Full Text

Decologne N, Kolb M, Margetts PJ, et al. 2007. TGF-β1 induces progressive pleural scarring and subpleural fibrosis. J. Immunol. 179: 6043-6051. Full Text

Gauldie J, Kolb M. 2008. Animal models of pulmonary fibrosis: how far from effective reality? Am. J. Physiol. Lung Cell Mol. Physiol. 294: L151. Full Text

Moeller A, Gilpin SE, Ask K, et al. 2009. Circulating fibrocytes are an indicator of poor prognosis in idiopathic pulmonary fibrosis. Am. J. Respir. Crit. Care Med. 179: 588-594. Full Text

Tarantal AF, Chen H, Shi TT, et al. 2010. Overexpression of TGF-β1 in foetal monkey lung results in prenatal pulmonary fibrosis. Eur. Respir. J. 2010 Mar 29. [Epub ahead of print]

John Haylor

Haylor J, Vickers ME, Morcos SK. 2009. Interference of gadolinium-based contrast agents with the measurement of serum creatinine by the Jaffe reaction. Br. J. Radiol. 82: 438-439.

Whitney High

High WA, Ayers RA, Chandler J, et al. 2007. Gadolinium is detectable within the tissue of patients with nephrogenic systemic fibrosis. J. Am. Acad. Dermatol. 56: 21-26.

High WA, Ayers RA,Cowper SE. 2007. Gadolinium is quantifiable within the tissue of patients with nephrogenic systemic fibrosis. J. Am. Acad. Dermatol. 56: 710-712.

Thomas Hope

Hope TA, Herfkens RJ, Denianke KS, et al. 2009. Nephrogenic systemic fibrosis in patients with chronic kidney disease who received gadopentetate dimeglumine. Invest. Radiol. 44: 135-139.

Hope TA, High WA, Leboit PE, et al. 2009. Nephrogenic systemic fibrosis in rats treated with erythropoietin and intravenous iron. Radiology 253: 390-398.

Jean-Marc Idée

Idée JM, Port M, Dencausse A, et al. 2009. Involvement of gadolinium chelates in the mechanism of nephrogenic systemic fibrosis: an update. Radiol. Clin. North Am. 47: 855-869, vii.

Idée JM, Port M, Medina C, et al. 2008. Possible involvement of gadolinium chelates in the pathophysiology of nephrogenic systemic fibrosis: a critical review. Toxicology 248: 77-88.

Idée JM, Port M, Raynal I, et al. 2006. Clinical and biological consequences of transmetallation induced by contrast agents for magnetic resonance imaging: a review. Fundam. Clin. Pharmacol. 20: 563-576.

Idée JM, Port M, Robic C, et al. 2009. Role of thermodynamic and kinetic parameters in gadolinium chelate stability. J. Magn. Reson. Imaging 30: 1249-1258.

Port M, Idée JM, Medina C, et al. 2008. Stability of gadolinium chelates and their biological consequences: new data and some comments. Br. J. Radiol. 81: 258-259. Full Text

Port M, Idée JM, Medina C, et al. 2008. Efficiency, thermodynamic and kinetic stability of marketed gadolinium chelates and their possible clinical consequences: a critical review. Biometals 21: 469-490.

Emanuel Kanal

Kuo PH, Kanal E, Abu-Alfa AK, et al. 2007. Gadolinium-based MR contrast agents and nephrogenic systemic fibrosis. Radiology 242: 647-649. Full Text

Philip Leboit

Cowper SE, Bucala R, Leboit PE. 2006. Nephrogenic fibrosing dermopathy/nephrogenic systemic fibrosis—setting the record straight. Semin. Arthritis Rheum. 35: 208-210.

Cowper SE, Bucala R, LeBoit PE. 2005. Case 35-2004: nephrogenic fibrosing dermopathy. N. Engl. J. Med. 352: 1723-1724.

Cowper SE, Su LD, Bhawan J, et al. 2001. Nephrogenic fibrosing dermopathy. Am. J. Dermatopathol. 23: 383-393.

Cowper SE, Robin HS, Steinberg SM, et al. 2000. Scleromyxoedema-like cutaneous diseases in renal-dialysis patients. Lancet 356: 1000-1001.

LeBoit PE. 2003. What nephrogenic fibrosing dermopathy might be. Arch. Dermatol. 139: 928-930.

Benjamin Newton

Newton BB, Jimenez SA. 2009. Mechanism of NSF: New evidence challenging the prevailing theory. J. Magn. Reson. Imaging 30: 1277-1283.

Susanne Nicholas

Collins AR, Schnee J, Wang W, et al. 2004. Osteopontin modulates angiotensin II-induced fibrosis in the intact murine heart. J. Am. Coll. Cardiol. 43: 1698-1705.

Nicholas SB, Liu J, Kim J, et al. 2010. Critical role for osteopontin in diabetic nephropathy. Kidney Int. 77: 588-600.

Wolak T, Kim H, Ren Y, et al. 2009. Osteopontin modulates angiotensin II-induced inflammation, oxidative stress, and fibrosis of the kidney. Kidney Int. 76: 32-43.

Hubertus Sieber

Pietsch H, Jost G, Frenzel T, et al. Efficacy and safety of lanthanoids as X-ray contrast agents. Eur. J. Radiol. [Epub ahead of print]

Pietsch H, Lengsfeld P, Jost G, et al. 2009. Long-term retention of gadolinium in the skin of rodents following the administration of gadolinium-based contrast agents. Eur. Radiol. 19: 1417-1424.

Pietsch H, Lengsfeld P, Steger-Hartmann T, et al. 2009. Impact of renal impairment on long-term retention of gadolinium in the rodent skin following the administration of gadolinium-based contrast agents. Invest. Radiol. 44: 226-233.

Sieber MA, Lengsfeld P, Frenzel T, et al. 2008. Preclinical investigation to compare different gadolinium-based contrast agents regarding their propensity to release gadolinium in vivo and to trigger nephrogenic systemic fibrosis-like lesions. Eur. Radiol. 18: 2164-2173.

Sieber MA, Pietsch H, Walter J, et al. 2008. A preclinical study to investigate the development of nephrogenic systemic fibrosis: a possible role for gadolinium-based contrast media. Invest. Radiol. 43: 65-75.

Sieber MA, Steger-Hartmann T, Lengsfeld P, et al. 2009. Gadolinium-based contrast agents and NSF: evidence from animal experience. J. Magn. Reson. Imaging 30: 1268-1276.

Steger-Hartmann T, Raschke M, Riefke B, et al. 2009. The involvement of pro-inflammatory cytokines in nephrogenic systemic fibrosis—a mechanistic hypothesis based on preclinical results from a rat model treated with gadodiamide. Exp. Toxicol. Pathol. 61: 537-552.

Sponsors

Presented by:

Bronze

• AMAG Pharmaceuticals
• Bracco
• Covidien

Academy Friends

• This event was funded in part by the Life Technologies™ Foundation
• This symposium was endorsed by New York Society of Nephrology
• Amgen
• Bayer
• Department of Dermatology, Yale School of Medicine
• General Electric
• Guerbet
• Lantheus
• The Global Fibrosis Foundation

Grant Support
The project described is supported by Award Number R13DK088440 from the National Institute Of Diabetes And Digestive And Kidney Diseases. The content of this program is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute Of Diabetes And Digestive And Kidney Diseases or the National Institutes of Health.

Magnetic resonance imaging (MRI) has done much to revolutionize the practice of medicine, allowing physicians to see structures within the body that were previously invisible unless revealed by surgery. In order to get the best possible images, radiologists rely on compounds known as contrast agents, which enhance the MRI signal and provide clearer, more detailed pictures. Many of these contrast agents contain metals that have been chosen for their paramagnetic properties, including manganese, iron, or, most commonly, the rare earth metal gadolinium.

These compounds have been carefully engineered for safety and have exemplary safety records; however, they are not completely without risk. A recent symposium at the New York Academy of Sciences brought together researchers studying a very rare syndrome that appears to be caused by the use of gadolinium-based contrast agents (GBCAs) in individuals with severely impaired kidney function, a disorder known as nephrogenic systemic fibrosis (NSF).

NSF is an idiopathic fibrosing disorder in which the skin hardens, sometimes to a density resembling wood, due to the uncontrolled production of connective tissue. This extreme hardening leads to characteristic skin lesions, pain, and joint contractures that can significantly impair mobility. NSF also affects the underlying muscle, and can cause damage to organs such as the esophagus, lungs, and heart, eventually leading to death in some cases. The exact pathogenic mechanism of NSF is unknown, and effective treatments are lacking.

NSF was first identified as a rare syndrome in kidney patients in the late 1990s, but it was not until several years later that the association with GBCAs was recognized. In 2007, the Food and Drug Administration (FDA) issued a public health advisory on the use of GBCAs, and mandated label changes contraindicating their use in patients with acute or chronic severe renal insufficiency. Since that time, the incidence of new NSF cases has declined considerably, as patients got recommended prescreening for decreased kidney function when the use of these agents is under consideration.

While these actions appear to have contained the threat, there are a number of important questions that remain to be answered. Perhaps most critically, how can the thousand or more individuals who have already contracted NSF be treated for this debilitating and sometimes fatal condition? What is different about these individuals from the vast majority of kidney patients who received GBCAs without contracting NSF? NSF sometimes has a latency of several months or more: how can we be sure that no one else has been affected? And finally, what can this syndrome teach us about the many other conditions that involve fibrosis, including such common conditions as atherosclerosis?

These questions and others were addressed from a variety of perspectives at the Fourth Annual Symposium on Nephrogenic Systemic Fibrosis and Gadolinium-Based Contrast Agents, held at the New York Academy of Sciences on May 14–15, 2010. Clinicians who played seminal roles in the identification and characterization of the disease, including Philip Leboit of the University of California, San Francisco, and Shawn Cowper of Yale School of Medicine, described the early history of the disease and its definitive diagnostic characteristics. The history of regulatory actions regarding GBCAs and NSF was given by Ira Krefting of the FDA. Biophysical experts, including Whitney High of the University of Colorado and Stephen Cramer of the University of California, Davis, described their work detecting and quantitating gadolinium in the tissues of NSF patients and patients with renal impairment. Jack Gauldie of McMaster University provided a perspective on NSF within the larger realm of fibrotic disorders, setting the stage for a number of speakers who described work on individual aspects of the mechanism of fibrosis: Michael Edward of the University of Glasgow, who studies the effects of GBCAs on fibroblast function, Richard Bucala of Yale School of Medicine, who studies the role of cellular precursors of fibroblasts known as fibrocytes, and Susanne Nicholas of the University of California, Los Angeles, who described a potential role for the ubiquitous messenger protein osteopontin.

A number of speakers from both academic and industrial settings described their experiences developing animal models and testing potential theories of NSF etiology, including John Haylor of the University of Sheffield, Sundararaman Swaminathan of the University of Arkansas, Thomas Hope of the University of California, San Francisco, Jean-Marc Idée of Guerbet (a French producer of GBCAs), Martin Sieber of Bayer-Schering Pharmaceuticals, and Benjamin Newton of GE Healthcare. They described animal modeling of the potential influences of renal insufficiency, erythropoietin and intravenous iron supplementation, and high serum phosphate levels on the development of NSF in patients with renal impairment. April Cox of Array Biopharma described her work testing imatinib, an FDA approved therapy for cancer and a number of other disorders involving abnormal cell proliferation, and a new compound, AR768, in an animal model of NSF. Jonathan Kay of the University of Massachusetts Medical School described his early success testing imatinib use in NSF patients.

Two speakers presented a primarily clinical point of view: Emanuel Kanal, of the University of Pittsburgh, who provided a radiologist's perspective on the importance of continuing, careful use of GBCAs, and Ali Abu-Alfa of Yale School of Medicine, who provided an epidemiologic update on NSF as well as a number of specific clinical precautions that can be used in radiology departments to help reduce the risk of new NSF cases. Ali Abu-Alfa, a co-chair of the meeting with Shawn Cowper, described NSF as "a unique disease that brings many disciplines together" in his introductory remarks. The meeting brought together 26 faculty from 7 countries and over 20 institutions and companies, attesting to the importance of gaining a better understanding of this rare but troubling complication of GBCA use.

Moderator:
Henrik Thomsen, University of Copenhagen

Speakers:
Shawn Cowper, Yale School of Medicine
Emanuel Kanal, University of Pittsburgh Medical Center
Jonathan Kay, University of Massachusetts Medical School
Whitney High, University of Colorado
Ira Krefting, U.S. Food and Drug Administration

Highlights

• NSF has specific clinical and histopathological characteristics that should be used to make a definitive diagnosis.
• GBCAs are valuable tools for radiologists with an exemplary safety record when they are used appropriately.
• Imatinib, an FDA-approved targeted cancer therapy, has shown efficacy against NSF in preliminary human studies.
• Gadolinium can be detected at high levels in many tissues from patients with renal insufficiency in general, and in skin lesions from NSF patients.
• Despite differences in stability, the FDA has thus far declined to assign levels of risk of NSF to different types of GBCAs, citing lack of sufficient evidence.
• Early warnings about NSF were hampered by difficulties getting the attention of public health officials.

NSF: Context, definition, direction

Speakers in this session provided an overview of the current status of NSF, including the discovery and history of NSF, clinical and histopathologic findings necessary for a definitive diagnosis, state-of-the-art research on the role of gadolinium, potential new therapies, and the perspectives of the FDA and a practicing radiologist who relies on GBCAs to make critical diagnoses.

The first case of NSF was identified in January 1997 at Sharp HealthCare, a managed health care organization in San Diego, CA. In 1999, Shawn Cowper of Yale School of Medicine established a registry to collect data on NSF cases, facilitating the study of this very rare syndrome. Cowper outlined the clinical characteristics and histopathologic findings that must be present for an individual to be diagnosed with NSF. These include certain distinctive clinical features, such as superficial pink or red plaques or papules on the skin, joint contractures, and abnormally hardened skin areas that show a cobblestone or peau d'orange (orange skin-like) appearance.

One characteristic of NSF is the presence of superficial pink plaques on the skin.

Cowper has combined these and other clinical findings with a group of distinct histopathologic characteristics to produce a diagnostic scoring grid that allows clinicians to differentiate NSF from a number of other fibrotic skin disorders, including scleroderma and scleromyxedema. In order to effectively research the causes and treatment of NSF, it is important that only accurately diagnosed cases are included in the case registry and in research studies. Cowper described several cases of misdiagnosis of NSF that have appeared in the literature.

Delineating the history of NSF, Cowper reviewed studies published from 2006 through the present that established the connection between NSF and the administration of GBCAs for MRI imaging. Analysis of the data collected by the registry shows that, thus far, verified cases of NSF have only been identified in patients with kidney disease. The vast majority (79%) are patients on dialysis, but another 17% are patients with acute kidney injury or stage 4 or 5 chronic kidney disease (CKD). Some of these patients had received kidney transplants.

Since GBCAs are cleared from the body largely by the kidneys, the current hypothesis is that individuals with reduced renal clearance are exposed to these agents longer, leading to NSF in individuals who are vulnerable to the disease. The exact mechanisms that lead from GBCA administration to the development of NSF are unknown, and the subject of considerable research as detailed by investigators at this symposium.

Radiology revelations

Radiologists rely on contrast agents to produce MRI images that are as informative as possible, and such enhanced images can play a key role in the diagnostic process. GBCAs are commonly used to image blood vessels for magnetic resonance angiography, and to enhance the imaging of brain tumors. Emanuel Kanal of the University of Pittsburgh Medical Center provided a working radiologist's perspective on the characteristics, uses, and risk/benefit ratios of these agents.

In its dissociated form as a free metal ion, gadolinium is highly toxic to animals and humans. The introduction of gadolinium as an agent for MRI contrast thus depended on the development of chelating agents that could tightly bind the metal ion and keep it in a biologically inert form. GBCAs are classified according to the chemical structure of the chelating agent, linear or macrocyclic, and ionic or non-ionic. The macrocyclic agents are more stable chemically, and thus are thought to be safer in general because they are less likely to release free gadolinium. The macrocyclic agents are cleared from the body almost exclusively by the kidney. The linear non-ionic agents, which are thought to be the least stable, are cleared by renal, and for some, by hepatobiliary mechanisms. The largely renal excretion pathways for these agents means that patients with reduced renal function will clear these agents more slowly, allowing longer exposure to the agents and more time for the chelated compound to potentially dissociate within the body.

GBCAs are nominally prescription drugs, but in practice they are often used interchangeably and patients' charts do not always indicate which specific agents were given. The lack of information on specific agents used in specific cases has made it more difficult to assess the risk of NSF associated with the different forms of GBCAs. Based on available evidence however, the European counterpart of the FDA, the European Agency for the Evaluation of Medicines (EMEA) has developed three risk categories for these agents with respect to NSF. Linear agents are in the high and medium risk categories while macrocyclic agents are in the low risk category. In the U.S., the FDA considers that there is not enough evidence to differentiate among these agents and has treated them as a single class of medications for regulatory purposes, mandating a black box warning for all GBCAs that are approved for use in the U.S.

Kanal provided numerous examples of the critical roles played by GBCAs in MRI-based diagnoses. He showed images of tumors and other serious abnormalities that were almost undetectable when a contrast agent was not used. For patients with suspected brain tumors or other serious conditions, he made it clear that the benefits of receiving the contrast agent far outweigh the potential risks.

Kanal emphasized as well the exemplary safety record enjoyed by the GBCAs. These agents have a very low adverse events rate of around 3%. Before NSF was discovered and linked to them, they were the preferred choice for patients with kidney disease over iodinated compounds, which are used as contrast agents for X-ray and CT scan imaging. Iodinated compounds are associated with a risk of contrast-induced nephropathy, a form of acute kidney injury that occurs much more frequently in patients with pre-existing chronic kidney disease. Alternative contrast agents also exist for use in MRI, including iron-based agents, which are associated with a high rate of anaphylactic reactions; manganese-based agents; and specialized agents that provide contrast for images of the gut. Kanal said that new contrast agents are in development but it will be at least 10 years before they are fully studied and ready for use.

Gadolinium is a rare earth element, and radiologists are the only source by which humans are exposed to this metal. As such, Kanal emphasized that these clinicians have the ability and the responsibility to ensure that patients who are at high risk for NSF are not exposed to GBCAs unnecessarily, and that patients in need of imaging receive the optimal contrast agent.

Update on imatinib for NSF treatment

Current treatments for NSF are largely ineffective. They include potent topical steroids, immunosuppressive therapy, plasmapheresis, and a wide range of other therapies, most of which have significant side effects. A number of new treatments that are under development have shown some efficacy against NSF progression, including extracorporeal photophoresis using UVA light, pentoxifylline, sodium thiosulfate, and rapamycin. Jonathan Kay of the University of Massachusetts Medical School in Worcester, MA, and his colleagues are studying the use of imatinib mesylate, a targeted anti-cancer agent that was first developed for the treatment of chronic myelogenous leukemia, and is now in use for a number of conditions that involve abnormal cell proliferation.

Proposed pathogenesis of NSF and the mechanism by which imatinib could act as a therapeutic.

Imatinib belongs to a class of compounds known as tyrosine kinase inhibitors, which also includes the similar compounds dasatinib and nilotinib. All three are FDA-approved agents for the treatment of cancer. Imatinib inhibits a number of tyrosine kinases that play important roles in signaling pathways involved in cell proliferation and differentiation. Fibrotic conditions, including NSF, often involve abnormal increases in cell number and inappropriate differentiation, and evidence from previous preclinical and clinical studies suggests that imatinib can inhibit fibrotic processes.

Kay and his collaborators have tested imatinib in small numbers of NSF patients and seen significant improvement in some patients, including reduction of skin tethering and thickening, which is a first for any therapy tried for this disorder. Imatinib is associated with a number of side effects, including edema, weight gain, and gastrointestinal disturbances, which sometimes necessitates dose reductions or causes patients to drop out of treatment. But the results of these trials are encouraging and it is likely that they will continue. In the future, Kay said it is likely that dasatinib and nilotinib will be tried in NSF patients as well.

Gadolinium detection in tissues

Once the association with GBCAs was established, the well known toxicity of free gadolinium made it a likely suspect as the causative agent of NSF. Researchers began to look for the presence of gadolinium in the skin and other tissues affected by the disorder. However, gadolinium can be difficult to detect and requires very specialized methods to quantitate accurately. Whitney High of the University of Colorado has spent considerable effort developing highly sensitive and quantitative methods to detect gadolinium in tissues to facilitate investigation of its role in NSF etiology.

Several biophysical techniques, including energy dispersive spectroscopy, X-ray microscopy, and inductively coupled plasma mass spectrometry (ICP-MS), can be used to detect gadolinium in tissues, although these techniques vary in their sensitivity. All three have shown very high levels of gadolinium in skin and other tissues from NSF patients. Because the first two methods are only semi-quantitative, High used ICP-MS methods to quantitate gadolinium in multiple types of tissues. He has found that patients with NSF can accumulate quite high levels of gadolinium in specific tissues, including the skin. However, the significance of this accumulation is unclear since most of these patients do not go on to develop NSF.

High said that until recently it was thought to be highly unlikely that gadolinium would be able to dissociate from the chelating agents used in GBCAs, particularly since excess chelating agent is included in commercial preparations to limit this possibility. It appears, however, that conditions may exist in patients with reduced renal function that might allow the gadolinium to be released. High suggested that the process might involve transmetallation, in which another metal, such as iron or zinc, knocks gadolinium off the chelating compound and takes its place. Patients with kidney disease have multiple clinical characteristics that might promote this process, for example, high or low pH levels in the blood, high levels of other metals, endothelial injury or inflammation, and the use of aggressive iron supplementation. Phosphate is of particular interest since kidney patients often have high levels in their blood and gadolinium phosphate is a highly insoluble compound.

High described several new methods that he and his group are using, including synchrotronic X-ray fluoroscopy, that can map exactly where gadolinium is deposited in tissues. They have found that in NSF patients, most of the gadolinium is deposited in areas of fibrosis, and it is more deeply distributed in patients where the disease had reached deeper layers of the skin. They also detected gadolinium in the heart tissue of an NSF patient who had developed myocardial fibrosis. Using another technique, X-ray absorption near edge spectroscopy, or XANES, they have shown the presence of gadolinium in eye tissue, lymph nodes, and the liver and heart of NSF patients at autopsy. These findings suggest that gadolinium toxicity might be detected less often than it occurs and may be leading to previously unappreciated morbidity and mortality in patients with reduced kidney function.

An FDA perspective

Ira Krefting of the U.S. Food and Drug Administration (FDA) provided a regulatory perspective on GBCAs and their role in NSF, detailing the history of events from the agency's perspective. In 2007, when it became clear that NSF was strongly associated with GBCAs, the FDA issued public health advisories on these agents and mandated changes in their labeling. The changes included a "black box" safety warning for all approved GBCAs that warned against their use in patients with acute or chronic severe renal insufficiency unless GBCA-enhanced diagnostic imaging was essential to their care. The FDA chose not to distinguish among the different GBCAs with regard to NSF risk, although another section of the label refers to the fact that the scientific evidence on many of these agents is still incomplete.

At the same time, the FDA asked the drug sponsors (generally the companies involved in developing, producing, and/or marketing a specific drug) to make a voluntary commitment to do postmarketing studies of NSF risk in renal failure patients. The sponsors agreed to do prospective trials enrolling 1000 patients with moderate to severe renal failure who received GBCAs, and who would be followed for two years to look for signs of developing NSF. Because NSF is so rare, it would be necessary to follow large numbers of patients to detect a significant number of cases. The sponsors expressed concern that they would not be able to enroll this many patients, particularly given the drastically reduced use of GBCAs in renally impaired individuals since the warnings came out. In 2008, the Food and Drug Administrative Amendment Act (FDAAA), was implemented, allowing the FDA to mandate postmarketing studies when safety issues are raised about drugs that are already on the market. This new law allowed the FDA to change the voluntary post-marketing commitments to post-marketing requirements, which are legally binding, enforceable contracts. The future of these studies is still uncertain, however, as the sponsors have continued to inform the FDA of significant difficulties with patient enrollment.

As described above, in 2009, the EMEA instituted the three-tier risk system for different GBCAs and instituted patient testing requirements for the highest risk agents. The FDA, by contrast, continues to treat all GBCAs the same, requiring screening of all patients for renal impairment by history or by laboratory testing. In December 2009, an advisory committee on NSF convened to examine available data. The committee found that data were still too scarce, particularly on the newer agents, to distinguish among them in terms of NSF risk. They recommended that potentially vulnerable populations be screened by history and with laboratory testing. They also warned that risk may be especially high in patients with acute renal failure because the most commonly used measure of kidney function, estimated glomerular filtration rate (eGFR), is not reliable in such patients.

The FDA collects reports of adverse events associated with marketed drugs through its Adverse Events Reporting System (AERS). In addition to data available in the literature and the NSF registry, the AERS has collected data on NSF that will aid in making future determinations about the safety and appropriate uses of GBCAs. Because the system is voluntary and reported by many different types of entities, the data on NSF are somewhat inconsistent and likely to be incomplete. However, the data do show a marked increase in the reporting of NSF cases since 2006 but a leveling off of the total number of cases, suggesting that the medical community as a whole has become aware of the FDA warnings and of the potential for this rare adverse event.

Moderator:
Michael Girardi, Yale School of Medicine

Speakers:
Philip Leboit, University of San Francisco
Christian Bull, Northwestern University
Stephen Cramer, University of California, Davis
Sundararaman Swaminathan, University of Arkansas

Highlights

• Most new diseases are man-made, often because of defective manufacturing processes.
• The true incidence of NSF is currently unknown.
• Gadolinium particles deposited in tissues from NSF patients consist largely of gadolinium phosphate.

Keynote presentation: the discovery of NSF

Sharp Health Care in San Diego, California, is one of the oldest managed care organizations in the U.S. In 1997, physicians in the renal transplant service at Sharp noticed a group of patients with progressively hardening skin. Many of these patients had received renal transplants that failed, or had had difficult postoperative courses, receiving multiple MRIs in the course of their treatment. These cases were felt to be sufficiently interesting to send to Philip Leboit, a noted dermatopathologist at the University of San Francisco, to investigate.

Leboit noted that, microscopically, these cases did not look exactly like other, better known fibrotic skin diseases such as scleromyxedema. Scleromyxedema is very rare and it would be quite unexpected to see ten cases in one city in one hospital. It also often involves the face, which the new syndrome did not, and there were other clinical and histopathological differences. Nor did they resemble another known fibrotic skin disease, scleroderma.

Leboit suspected that this new syndrome might be man-made, based on previous experiences with toxic sclerotic syndromes that were caused by defective manufacturing processes. These included the so-called Spanish cooking oil epidemic, which was really caused by defective canola oil, and eosinophilia-myalgia syndrome, which was caused by tryptophan supplements that were contaminated during the manufacturing process. These syndromes were fatal in many cases.

Leboit and his colleagues named the syndrome as a new disease, nephrogenic fibrosing dermopathy, in a publication in 2001. After some initial difficulty getting the attention of the Centers for Disease Control (CDC) to report this public health threat, they were assigned a case officer, who was then abruptly pulled from the case by the events of Sept. 11, 2001. Eventually, however, the CDC published a public health dispatch in Morbidity and Mortality Weekly Report in January 2002. In the meantime, the NSF registry was established at Yale in 1999, eventually leading to the association of the disorder with exposure to GBCAs.

Leboit described several lessons learned from the early years of investigating this disorder: first, most new diseases are man-made; second, getting the attention of public health officials may be difficult; and third, it is important to take a comprehensive approach to investigating new syndromes but this can be difficult if resources are lacking.

Platform presentations

The organizers of the meeting solicited poster abstracts from attendees. Most of these were selected as poster presentations while a select few were chosen for short platform presentations. Christian Bull of Northwestern University described a comprehensive study in which he analyzed available data on NSF from the Yale registry, the AERS, and from several institutions and GBCA manufacturers. He highlighted some of the difficulties with available data, particularly those from the AERS which is highly variable in aspects such as event date. Individuals reporting the data have variously taken "event date" to mean the date of GBCA exposure, the date of NSF diagnosis, or other interpretations. Some cases of NSF were also reported to the AERS more than once by different entities. Bull cautioned that, because of such inconsistencies, the true incidence of NSF is currently unknown. However, data culled from several institutions and GBCAs' manufacturers seem to confirm a significant drop in new cases since FDA warnings were issued.

Two speakers presented data that may shed light on contributing factors in patients with renal impairment. Stephen Cramer of the University of California, Davis, used X-rays from synchrotron radiation in several biophysical methods to investigate the chemical form and structures of the gadolinium particles deposited in tissues from NSF patients. He has concluded that they consist largely of gadolinium phosphate. Sundararaman Swaminathan of the University of Arkansas presented evidence that iron-metabolizing pathways may be involved in NSF pathogenesis, specifically the CD163 iron-scavenging pathway that is mobilized in human tissue culture cells on exposure to the GBCA gadodiamide.

Moderator:
Richard Bucala, Yale School of Medicine

Speakers:
Jack Gauldie, McMaster University
Susanne Nicholas, University of California at Los Angeles
Michael Edward, University of Glasgow
Richard Bucala, Yale School of Medicine

Highlights

• Fibrotic diseases are promoted by the actions of pro-fibrotic cytokines and growth factors, which may, in some cases, be limited to specific organs by their strong association with the extracellular matrix.
• The multifunctional protein osteopontin may be involved in fibrotic diseases, including NSF, perhaps representing a new therapeutic target or a potential biomarker.
• Cultured fibroblasts are stimulated to produce extracellular matrix components by serum from NSF patients and by GBCAs.
• Fibrocytes, circulating precursor cells, are stimulated by GBCAs to differentiate into fibroblasts.

The pathophysiology of fibrosis

This session focused on the status of basic scientific research that is intended to elucidate the cells, molecules, and pathways that are involved in NSF, and related this research to known properties of other fibrotic disorders. Increased knowledge of the underlying molecular mechanisms of this disorder will lead to the identification of new therapeutic targets and potentially to more effective therapies.

Jack Gauldie of McMaster University is an expert on molecular aspects of inflammation and immunity, and provided an overview of the overlapping mechanisms of fibrotic diseases as an introduction to the field. He said that 45% of U.S. deaths can be attributed to some kind of fibrotic outcome. There are a large number of major organ fibrotic syndromes, including interstitial lung disease, liver cirrhosis, kidney disease, heart disease, and fibrotic diseases of the eye. In addition, there are fibroproliferative disorders, which include local and systemic scleroderma, excess scarring associated with burns or trauma, and radiation- and chemotherapy-induced fibrosis. Thus while NSF is rare, understanding its etiology has the potential to promote our understanding of a large number of debilitating and/or deadly conditions.

Under normal circumstances, fibrosis is a part of the wound healing process. It is intended to restore connective tissue after it has been damaged by injury or disease. This well-studied process involves interactions between a large number of cell types, including epithelial and endothelial cells, multiple types of immune cells, and fibroblasts. Multiple pro-fibrotic messenger molecules have been identified as well, including the chemokines TGF-beta and interleukin-13. Studying the interactions between these cells, as mediated by these messenger molecules, is likely to shed light on the chronic, progressive production of connective tissue that is characteristic of fibrotic conditions. Some fibrotic conditions are limited to specific organs while others are systemic, and it is likely each type is regulated somewhat differently.

Gauldie and his coworkers are studying idiopathic pulmonary fibrosis (IPF), a highly lethal condition in which fibrosis is limited to the lungs. There are currently no treatments for IPF but a number of drug candidates are in clinical trials. Gauldie and his group have developed an animal model of IPF that allows them to study the role of cytokines and other peptides in the development of this condition. They are particularly interested in understanding how this disease progresses, and what limits it specifically to the lungs.

Using an adenovirus vector, they express TGF-beta in rat or mouse lungs, creating a fibrotic condition similar to IPF. They have used this system to identify many of the genes and proteins whose expression is turned on by the presence of TGF-beta. One key finding is that many of the messenger molecules whose expression is promoted by TGF-beta are strongly associated with the extracellular matrix, the sticky mixture of proteins and complex sugars that holds cells together in tissues. This strong association limits the messengers' mobility, restricting pro-fibrotic signaling to specific sites within the lungs, and perhaps explaining why the condition is limited only to this organ.

TGFβ1 induces progressive pleural fibrosis.

Gauldie also described considerable work from his lab and others that links the actions of cytokines, including IL-1, IL-13, and IL-17, to the Smad intracellular signaling pathway, which in turn is linked to the wound repair signaling process. Much work has also implicated circulating fibroblast precursor cells, known as fibrocytes, as well as macrophages in the fibrotic process. Such findings are likely to lead the way to new potential drug targets for fibrotic conditions. Many molecules, including immunomodulators such as interferon-gamma, anti-TGF-beta antibodies, and inhibitors of growth factors such as PDGFR and EGFR, are in clinical trials for these conditions, but most of the current agents are plagued by detrimental side effects. New ideas are needed, and it is hoped that basic scientific study of NSF will identify multiple new avenues for therapeutic research.

A role for osteopontin?

Susanne Nicholas of the University of California, Los Angeles, presented evidence that osteopontin, a multifunctional protein that is expressed at high levels in a number of chronic inflammatory and autoimmune diseases, might be involved in the etiology of NSF. Osteopontin is upregulated in many chronic kidney diseases, including nephrolithiasis and glomerulonephritis.

Osteopontin is a small, 305 amino acid protein that is found in almost all body fluids and tissues. It was first identified in bone, where it is particularly abundant. Considerable study of osteopontin has revealed roles in a diverse range of physiological functions, including biomineralization, leukocyte function, macrophage recruitment, wound repair, and cancer biology. Osteopontin levels are regulated by a wide range of hormones and other substances, including parathyroid hormone, estrogen, vitamin D, phosphate, growth factors, cytokines, and certain drugs.

Osteopontin has also been implicated in a number of disease processes that involve fibrosis, including atherosclerosis, chronic kidney disease, and cancer. Studies of a mouse model of atherosclerosis show that mice lacking a functional copy of the osteopontin gene have fewer atherosclerotic lesions and less cardiac fibrosis. Mice lacking osteopontin also show less kidney damage in mouse models of chronic kidney disease and in models of type I and type II diabetes. These findings suggest that osteopontin plays a pro-fibrotic role, however, lack of osteopontin can also lead to excess fibrosis under some conditions, suggesting that its role in fibrosis is complex and requires further elucidation.

Specific evidence that osteopontin may be involved in NSF comes from a mouse model of the disease, which shows greatly increased serum osteopontin levels. Osteopontin deficiency has also been shown to accelerate wound healing. It remains to be determined whether osteopontin is an appropriate therapeutic target for NSF, as well as whether it might have value as a biomarker for the disease. In the meantime, osteopontin-targeted therapeutics are already being tested for other conditions. An anti-osteopontin monoclonal antibody is currently being tested to treat collagen-induced arthritis, and an osteopontin antagonist has been shown to block the growth and metastasis of breast cancer cells. These therapeutics could be put to use in NSF patients if an appropriate rationale for targeting osteopontin can be developed.

Impact of GBCAs on fibroblast proliferation, differentiation, and extracellular matrix production

One of the hallmarks of NSF is a marked increase in the number of fibroblasts in areas where fibrotic skin lesions have formed. Fibroblasts are normal components of the skin that are involved in the response to foreign material, and as such might be involved in a pathogenic response to free gadolinium, GBCAs, or other forms of the metal that are deposited there. Michael Edward of the University of Glasgow is studying cultured fibroblasts from NSF patients and normal individuals in order to shed some light on the potential mechanisms by which GBCAs might trigger or promote NSF.

In addition to increased fibroblasts, NSF lesions are characterized by an abnormal accumulation of components of the extracellular matrix, the protein and carbohydrate glue that holds cells together and provides structural support for tissues. Resident fibroblasts are largely responsible for secreting and maintaining the extracellular matrix. Edward and his group have found that fibroblasts derived from skin biopsies from NSF patients secrete unusually high levels of extracellular matrix compounds, including the complex polysaccharide hyaluronan and the fiber-forming protein collagen. Moreover, serum from NSF patients stimulates increased secretion of these compounds by normal fibroblasts, as does serum from dialysis patients, although to a lesser extent.

Since gadolinium is also present in NSF skin lesions, Edward asked whether gadolinium, in the form of GBCAs, affects the proliferation of fibroblasts or their synthesis of extracellular matrix components. He found that both linear and macrocyclic GBCAs stimulated fibroblast proliferation, and that linear GBCAs modestly stimulated hyaluronan synthesis, although they did not stimulate collagen production. Other compounds that contain gadolinium but that are not contrast agents—including a gadolinium-EDTA chelate and gadolinium chloride—also stimulate fibroblast proliferation. However their stimulatory effects were not as great as that of gadodiamide with excess chelate, which was also more active than gadodiamide on its own.

Furthermore, the linear GBCA gadodiamide caused normal fibroblasts to assume a myofibroblast-like phenotype, similar to the phenotype found in differentiated fibroblasts from NSF lesions. The free chelating agents, which are present in excess amounts in the GBCAs to minimize the possibility of exposure to free gadolinium, were unable to produce these effects. Thus though free gadolinium is the most obvious suspect in promoting NSF, Edward's results suggest that if free gadolinium were important, gadodiamide with excess chelate should not have been more active than gadodiamide on its own.

Impact of GBCAs on fibrocyte function

Another type of cell that accumulates in NSF skin lesions is the fibrocyte. Fibrocytes are fibroblast-like cells that normally circulate in the bloodstream and are recruited to sites of tissue injury to participate in the healing process. They are one of the many cell types that are found in the white blood cell, or leukocyte, component of blood. They can be identified by their expression of the membrane-bound marker proteins CD34 and CD45, and by the fact that they produce collagen types I and II, which is very unusual for a circulating cell type. These cells have been implicated in a number of fibrotic disorders, including interstitial pulmonary fibrosis, asthma, and renal and hepatic fibrosis. Understanding how they are recruited and how their differentiation is regulated could lead to insight into the disease process of NSF as well as the identification of new drug targets.

Richard Bucala of Yale School of Medicine is credited with the discovery of fibrocytes, and has worked out much of their biology, including details of how they are recruited to the site of a tissue injury and the molecular signals that prompt them to differentiate to fibroblasts and myofibroblasts once there. He and his group are studying the effects of GBCAs on circulating fibrocytes in NSF patients. NSF patients have higher levels of circulating fibrocytes than both normal controls and patients on hemodialysis. Bucala and his group have also found that the GBCA gadodiamide promotes fibrocyte differentiation, in both normal individuals and NSF patients. The fibrocytes from NSF patients are much more sensitive to this effect, even though they are less sensitive to other promoters of fibroblast differentiation.

To uncover the molecular basis of the increased sensitivity of NSF fibrocytes to gadolinium, Bucala and his group have compared gene expression in fibrocytes isolated from normal individuals that have been exposed to low, clinically relevant concentrations of gadolinium. Using whole genome microarray analysis, they found that exposure to gadolinium changed the expression of only 287 genes, which was unexpected since usually thousands of changes are detected by this method.

One particularly interesting class of genes whose expression was reduced in the presence of gadolinium was the group encoding the metallothioneins. Metallothioneins are a large family of cysteine-rich proteins responsible for binding metals that are needed for physiological functions, such as zinc, as well as for binding and sequestering potentially toxic xenobiotic metals. It is an unanswered question why these proteins would be downregulated rather than upregulated by the presence of the metal gadolinium. Further investigation will be needed to determine whether similar changes occur in patients who have received GBCAs, and whether these changes contribute to the pathogenic process in NSF.

Moderators:
Ali Abu-Alfa, Yale School of Medicine
Phillip Kuo, University of Arizona

Speakers:
John Haylor, University of Sheffield
Thomas Hope, University of California at San Francisco
Jean-Marc Idée, Guerbet
Martin Sieber, Bayer-Schering Pharmaceuticals
Benjamin Newton, GE Healthcare
April Cox, Array Biopharma

Highlights

• Rats with reduced kidney function accumulate gadolinium in their tissues in proportion to their degree of residual renal function.
• Multiple injections of high doses of GBCAs lead to NSF-like skin lesions in both normal and renally impaired rats.
• GBCAs have different physicochemical properties that can affect whether and how much gadolinium is deposited in the tissues of animal models of NSF and the extent of skin lesion development.
• The presence of erythropoietin and intravenous iron promotes the development of NSF-like skin lesions in rats with normal renal function who receive high doses of GBCAs.
• The chelating agent caldiamide, a component of the GBCA gadodiamide, does not by itself appear to promote the formation of NSF-like skin lesions.
• High serum phosphate levels may promote gadolinium deposition and the appearance of skin lesions.
• Intact gadolinium-chelate complexes stimulate activation of macrophages and fibroblasts in culture, suggesting a potential role for the GBCAs in NSF that does not involve the release of free gadolinium.
• The tyrosine kinase inhibitor imatinib and the PDGFR inhibitor AR768 can reverse NSF-like changes in a rat model.

The Sheffield experience: 5/6 subtotal nephrectomy as an animal model of NSF

John Haylor and his colleagues at the University of Sheffield are using an animal model with reduced kidney function as the starting point for a model of NSF. Reduced kidney function can be created in animals such as rats and mice by the simple expedient of surgically removing a large portion of the kidneys, here using a procedure known as 5/6 subtotal nephrectomy. This procedure results in animals with varying levels of kidney function, as detected by a measure known as the glomerular filtration rate.

Haylor and his group injected nephrectomized rats with the linear GBCA gadodiamide and found that the amount of gadolinium deposited in the skin depended on the remaining level of kidney function in each rat, as measured by the glomerular filtration rate. Multiple injections of gadodiamide also led to changes in the skin that were similar to NSF. Injection of the macrocyclic GBCA gadoterate, which is considered the lowest risk agent for NSF because of its high stability, resulted in lower levels of gadolinium deposition in multiple tissues including kidney, liver, bone, and skin. In addition, gadoterate did not cause many of the histological changes, resembling NSF, that were caused by gadodiamide. By studying the differences among these agents, the researchers hope to gain insight into the mechanism by which GBCAs promote the development of NSF.

The UCSF experience: effects of erythropoietin and intravenous iron

There is an increase in gadolinium deposition in rats treated with Epo, IV iron, and gadodiamide in comparison to those receiving gadodiamide alone.

Thomas Hope and his colleagues at the University of California, San Francisco, have taken a different tack in studying NSF in animal models. Because patients with kidney disease are often anemic, many receive high doses of erythropoietin and intravenous iron to promote the production of red blood cells. Hope is investigating the role of these agents in NSF by dosing rats with normal renal function with combinations of these agents and the GBCA gadodiamide. Groups of rats received gadodiamide alone, gadodiamide plus erythropoietin, gadodiamide plus intravenous iron, or all three agents. Of the six rats who received all three agents, three developed numerous skin lesions. All of the rats showed increased cell numbers in the superficial layers of the skin, but levels were highest in those who received all three agents and in the rats where skin gadolinium deposition was highest.

This model does not represent an exact match for NSF because Hope did not see the deep changes in cell numbers or fibrosis that are more characteristic of the disease, although it is possible that these changes would have developed if the rats were studied longer. Hope and his group are also testing the effects of these combinations of agents in a commercially available strain of rats with reduced kidney function. Their preliminary results suggest that erythropoietin and intravenous iron may play a role in the development of NSF, as has frequently been speculated due to the frequent exposure of kidney patients to these agents.

The Guerbet experience: role of GBCA structures and physicochemical properties

Jean-Marc Idée and his colleagues at Guerbet are investigating the physicochemical differences among the GBCAs in an effort to better understand whether the causative agent of NSF is free gadolinium, the gadolinium-chelate complex as a whole, or possibly even the excess chelating agent found in commercial GBCA formulations to improve their safety. They are also investigating the effects of physiological conditions found in dialysis patients, including excess phosphate in the blood, a condition known as hyperphosphatemia. As mentioned above, phosphate is of particular interest because gadolinium phosphate is an insoluble compound that would tend to draw gadolinium away from its chelating agent, possibly leading to increased tissue deposition. The currently prevailing theory is that gadolinium deposits provoke a response from fibroblasts, whose job it is to react to the presence of foreign material in the skin, setting off the chain of events that leads to NSF in some individuals.

The various GBCAs have different thermodynamic and kinetic stabilities that are likely to influence how much free gadolinium might be released into animal or human tissues after they enter the body. Using nephrectomized rats, Idée and his group measured the amount of free gadolinium deposited in tissues after multiple injections of the macrocyclic GBCA gadoterate, the linear GBCA gadodiamide as formulated for use in humans, and unformulated gadodiamide that does not contain the protective high levels of excess chelate found in the commercial preparations.

Idée's group saw a gradual in vivo dissociation of the linear GBCA gadodiamide after intravenous administration to renally-impaired rats while a macrocyclic compound remained stable. This effect was found in plasma, skin and bone. Interestingly, the "free" gadolinium released was still soluble and probably bound to extracellular proteins.

In addition, the group found that unformulated gadodiamide, lacking the protective effect of excess chelating agent, produced the most NSF-like skin changes in rats. Non-formulated gadodiamide also killed 8 of the 10 rats to which it was administered, while all of the other rats survived, demonstrating the well known high toxicity of free gadolinium. Gadodiamide, both formulated and unformulated, was also associated with higher serum, skin, and bone levels of gadolinium. Taken together, these findings suggest that repeated injections of gadodiamide lead to the gradual release of free gadolinium that is deposited in tissues, leading to NSF in susceptible individuals, as posited by the leading theory of NSF etiology.

Idée and his coworkers also tested the potential influence of hyperphosphatemia on this process by injecting formulated gadodiamide into rats that were given a normal or a high phosphate diet. Although levels of free gadolinium were similar between the two groups, the rats on the high phosphate diet developed more skin lesions with NSF-like characteristics.

One of the mysteries surrounding NSF is why only a few of the many patients with reduced kidney function who received multiple doses of GBCAs developed the condition. Idée's work provides further evidence that there are many contributing factors that may come into play before the condition develops.

The Bayer-Schering experience: implicating gadolinium complex stability

Martin Sieber of Bayer-Schering Pharmaceuticals summarized a great deal of animal work on GBCAs that has been done at his company over the past few years. Sieber and his colleagues have used repeated high dose injections of GBCAs in healthy rats to simulate the increased exposure levels of individuals with impaired renal function. Multiple injections of gadodiamide were found to produce skin lesions similar to lesions found in humans with NSF. Multiple injections of caldiamide, which is the excess chelating agent found in commercially formulated preparations of gadodiamide, did not cause such lesions. Moreover, the appearance of lesions correlated with high levels of gadolinium in the skin of some of the rats, and preparations containing higher levels of excess chelating agent led to less gadolinium deposition and fewer skin lesions.

Their results support the idea that the stability of the complex is important, implicating free gadolinium as the culprit and potentially absolving the chelating agent alone. One potential hypothesis has also been that the excess chelating agent found in the GBCAs causes a depletion of physiologically important trace elements, including zinc, leading to the development of NSF. The lack of deleterious effects from the chelating agent alone or excess chelating agent in these studies suggests this may no longer be a viable hypothesis.

Sieber and his colleagues also studied the differences among GBCAs when they were administered to rats. They found that the non-ionic linear GBCA gadodiamide led to the highest amount of gadolinium deposition in the skin, and the macrocyclic agents gadoterate, gadoteridol, and gadobutrol led to the lowest. The non-ionic linear GBCAs led to much higher gadolinium concentrations in the serum compared to the ionic linear, and levels of free gadolinium in the serum were too low to measure after administration of macrocyclic GBCAs.

They tested the effects of these agents in renally impaired rats as well. As also shown by Idée and others, the increased retention of the GBCAs in these animals led to significantly increased concentrations of gadolinium in the skin. This increase was most pronounced for the non-ionic linear GBCAs, and was further promoted by the presence of high levels of phosphate.

Sieber's results provide further evidence that the stability of the complex is important, and that the presence of gadolinium is associated with the development of NSF-like skin lesions. They have undertaken further studies of complex stability using chelates of other metals in the lanthanide series where gadolinium resides on the periodic table. In addition, they are studying the roles of different cell types, such as macrophages, and messenger molecules, such as the inflammatory cytokines, in the process of skin lesion formation in these animal models.

The GE Healthcare experience: direct effects of GBCAs

Much of the recent work in animal models has been taken to show that the deposition of free gadolinium in tissues is an important proximal cause of NSF, although the chelated complexes have also been implicated in some studies. Benjamin Newton of GE Healthcare Medical Diagnostics described work that provides further evidence for a direct role of the gadolinium-chelate complex as a whole, as shown by its effects on cell types involved in the fibrotic process.

Newton maintains that animal models that have linked levels of gadolinium in tissues to NSF produce lesions that are not really the same as NSF lesions. Moreover, many studies in both animals and humans did not determine whether the gadolinium deposited in tissues was free or chelated, since making this determination is technically difficult and requires instrumentation that not all investigators can access.

Newton and his group are testing the effects of GBCAs on normal human macrophages and fibroblasts in culture. Their results suggest that chelated gadolinium is not the biologically inert compound it has been thought to be. They have found that the GBCA gadodiamide can activate pro-inflammatory signaling pathways, inducing the release of cytokines, in macrophages, under conditions in which free gadolinium is unlikely to be present. Newton also reviewed evidence from other groups showing that a number of GBCAs stimulate monocytes to produce growth factors and cytokines as well. Newton has found that fibroblasts can also be directly activated by gadodiamide.

These results suggest that the GBCAs themselves could directly influence the initiation of fibrosis, without the need to postulate the deposition of free gadolinium in tissues as a triggering mechanism. According to this alternate hypothesis, the gadolinium-chelate complex as a whole is internalized and processed by macrophages and fibroblasts, leading to the release of pro-inflammatory and pro-fibrotic cytokines, which in turn stimulate fibroblasts to initiate overproduction of extracellular matrix components, leading to fibrosis. Further research will be needed to confirm whether intact GBCAs do in fact play a role in the NSF disease process.

The Array experience: testing potential therapies in an animal model

April Cox of Array Biopharma rounded out the session on animal models by discussing her company's work testing potential therapies for NSF. Like other models, hers seeks to replicate gadolinium exposure in renally impaired individuals by administering repeated high doses of gadodiamide. Therapies are then administered prophylactically, overlapping in time with gadodiamide administration, or much later, after gadodiamide-induced changes have had time to fully develop, in a therapeutic mode.

This animal model has been used to test the compounds AR768, a small molecule inhibitor of the platelet derived growth factor receptor (PDGFR), and the tyrosine kinase inhibitor imatinib. Cox presented results showing that the gadodiamide-treated rats developed skin lesions with many similarities to NSF in humans, including alterations in skin texture and pigmentation, increased numbers of macrophages, fibroblasts, and other cells, and systemic involvement in the form of lung fibrosis. Both therapies improved a number of these measures of NSF-like pathology, in both the prophylactic and therapeutic settings. The observed efficacy of AR768 suggests that PDGFR might play an important role in the establishment and/or progression of NSF, representing a potential new therapeutic target for this and other fibrotic conditions.

Moderator:
Jeffrey Weinreb, Yale School of Medicine

Speaker:
Ali Abu-Alfa, Yale School of Medicine

Highlights

• Patients with chronic kidney disease stages 4 and 5 are at highest risk for NSF.
• Patients with acute kidney injury are also at high risk.
• Physicians can mitigate these risks with careful screening for kidney disease, appropriate selection of GBCA type and dose, and consideration of the patient's overall history of GBCA exposure.
• NSF risk may also be mitigated by post-imaging dialysis to remove the GBCA from the circulation as soon as possible.

Development of clinical recommendations

While researchers continue to work to unravel the causes of NSF and identify potential treatments, it is vitally important that practicing clinicians are kept informed on how to ensure that their patients are protected from the risk of NSF, while at the same time making use of necessary and important diagnostic imaging procedures. Ali Abu-Alfa of Yale School of Medicine closed out the conference with a presentation on current clinical recommendations for the uses of GBCAs in patients with renal insufficiency, particularly highlighting some of the areas of where clinical decision making might be more difficult.

Abu-Alfa began by reviewing the epidemiology of kidney disease, since this is the population at risk for NSF. There are about 26 million people in the U.S. who have chronic kidney disease (CKD), most of whom have less severe stages of the disease known as CKD stages 1 through 3. To date, no cases of NSF have been definitively identified in individuals with stages 1–3 or in patients with normal renal function. About 1 million of these 26 million people have CKD stages 4 and 5, and these individuals are among those at the highest risk for developing NSF.

Another high risk population is represented by individuals with acute kidney injury, such as those that occur with trauma or in cases of severe liver disease. Abu-Alfa noted that trauma patients are at particularly high risk because they are likely to have unrecognized acute kidney injury while at the same time may have a need for imaging procedures.

Considerations for risk reduction in patients with chronic kidney disease.

As mentioned above, about 90% of NSF patients are patients who are on dialysis. The prevalence of NSF among exposed dialysis patients varies in different reports but is generally around 5%. The overall prevalence is around 0.5% to 1% for all dialysis patients. Prior to the FDA warnings, exposure to GBCAs was quite common in dialysis patients, with a study done approximately every 3 years. Patients on dialysis often suffer from co-morbidities, such as anemia, hypertension, and diabetes. These patients often have physiologic abnormalities as well, including high or low pH of the blood, high calcium, and high phosphate levels. However, careful study of available data on NSF cases has yet to identify specific characteristics that represent additional risk factors for NSF.

Abu-Alfa described a number of strategies that can be used to reduce the risk of NSF in advanced stage CKD patients who must undergo imaging procedures. Patients should be screened by history and laboratory tests for CKD, using estimated values for glomerular filtration rate.  Although there is currently no consensus on how low this value must be to make GBCA use questionable, it is generally accepted to be less than 30 ml/min/m² at present. Patient self-reporting alone is likely to be unreliable as many individuals with CKD do not know they have it.

In patients with identified CKD or acute kidney injury, clinicians should consider using alternative contrast agents or non-enhanced MR imaging. In cases where GBCA administration is deemed necessary, it is important to use the lowest dose possible of a GBCA and to choose the GBCA that is associated with a reduced risk for NSF, such as a macrocyclic agent, although data are still emerging in this area. In making the decision to use a GBCA, the clinician should also consider the patient's past history of exposure to GBCAs. Intervals between receiving GBCAs should be extended if possible, and total lifetime dose of GBCAs may also be an important consideration.

If GBCA use is deemed necessary in a high risk patient, Abu-Alfa presented data showing that GBCA are removed by hemodialysis, though evidence that post-exposure risk is reduced by hemodialysis soon after administration of a GBCA is still lacking. Nonetheless, immediate hemodialysis should be considered within 2 hours in patients already on hemodialysis, patients maintained on peritoneal dialysis, patients with acute kidney injury, and potentially in CKD stage 5 patients. Initiation of hemodialysis should be individualized given its own associated risks.

Abu-Alfa emphasized that recommendations in the area of NSF risk reduction are still emerging as research continues, and advised radiologists and other clinicians to stay tuned as experts strive to reach a consensus on the best way to prevent this debilitating condition.

Why have only a small subset of the patients with severe kidney disease who received GBCAs developed NSF?

What precipitating factors act together with the GBCAs and reduced kidney function to promote NSF in susceptible individuals?

What is the most clinically relevant, efficient, and cost effective way to screen patients for reduced kidney function before administering a GBCA?

Are some GBCAs safer than others with regard to the risk of NSF?

What is the mechanism by which GBCAs promote fibrosis?

Is there a role for the entire gadolinium-chelate complex in NSF or is the disease caused only by the presence of free gadolinium?

What studies need to be done to fully implicate osteopontin in NSF, and is there a role for osteopontin-targeted therapeutics or osteopontin as a biomarker?

Will it be possible to develop therapeutics that effectively and safely stop the progression or perhaps even reverse the clinical course of NSF?