Virchows Arch (2006) 448: 337–343DOI 10.1007/s00428-005-0089-x

ORIGINAL ARTICLE

Raphael Schiffmann . Amy Rapkiewicz .Mones Abu-Asab . Markus Ries . Hasan Askari .Maria Tsokos . Martha Quezado

Pathological findings in a patient with Fabry disease who diedafter 2.5 years of enzyme replacement

Received: 22 June 2005 / Accepted: 31 August 2005 / Published online: 29 November 2005# Springer-Verlag 2005

Abstract We describe the postmortem findings of a 47-year-old man with Fabry disease, an X-linked glycolipidstorage disorder, who was on enzyme replacement therapywith recombinant α-galactosidase A for more than 2 years.The patient had widespread atherosclerotic coronary arterydisease that culminated in a massive acute myocardial in-farction. Atherosclerotic lesions were seen in the right andleft coronary systems, aorta, and the basilar artery. TypicalFabry cardiomyopathy and glomerular nephropathy werefound. With the exception of vascular endothelial cells,extensive glycolipid storage deposits were seen in allvascular and nonvascular cells and organ systems. Weconclude that, at least in this patient, repeated infusionswith α-galactosidase A over a prolonged period did notappreciably clear storage material in cells other than vas-cular endothelial cells. These findings also illustrate ac-celerated atherosclerosis in susceptible patients with Fabrydisease.

Keywords Fabry disease . Lysosomes .Atherosclerosis . Myocardial infarction . Enzymereplacement therapy . Glycolipids

Introduction

Fabry disease, an X-linked systemic disorder caused by adeficiency of the lysosomal enzyme α-galactosidase A,results in a failure to metabolize α-D-galactosyl moieties[4]. This abnormality leads to the systemic deposition ofglycosphingolipids, particularly globotriaosylceramide (Gb3)[5]. In adulthood, progressive renal failure, strokes, andheart disease develop, leading to death at a median age of50–55 years. In the past 5 years, enzyme replacement ther-apy (ERT) with recombinant α-galactosidase A has beenadded to the standard therapeutic regimen [8, 17, 18, 22].Targeted primarily to reverse vasculopathy of Fabrydisease, ERT has been shown to markedly reduce thecellular storage deposition in vascular endothelial cells[22]. Its long-term clinical effects, however, remain to befully assessed. No extensive pathological analysis inpatients on long-term ERT has been reported thus far. Wepresent in this report the autopsy results of a 47-year-oldman who was on ERT for more than 2 years and diedsuddenly.

Case report

The patient was a 47-year-old man who was diagnosedwith Fabry disease at age 19 with α-galactosidase Aactivity of 1.3% of normal controls. Since 5 years of age, hesuffered from episodes of fever and acroparesthesia,hypohidrosis, poor heat and exercise tolerance, diarrheawith abdominal cramps, and bouts of depression. At 35, hedeveloped an acute onset of right-sided weakness anddifficulty with word finding. A CT scan revealed a largeanterior parietal infarct, and a left carotid angiogramshowed marked narrowing of mid-sylvian branches of theleft middle cerebral artery (MCA) with an area of avas-cularity in the left anterior parietal lobe. He was put on oralwarfarin therapy with no apparent stroke recurrence. Hismost recent head MRI scan showed old left encephalo-malacia in the left frontal lobe along with white matter

R. Schiffmann . M. Ries . H. AskariDevelopmental and Metabolic Neurology Branch,National Institute of Neurological Disorders and Stroke,National Institutes of Health,Bethesda, MD, USA

R. Schiffmann (*)National Institutes of Health,Room 3D03, Building 10, 9000 Rockville Pike,Bethesda, MD 20892-1260, USAe-mail: RS4e@nih.govTel.: +1-301-4961465Fax: +1-301-4808354

A. Rapkiewicz . M. Abu-Asab . M. Tsokos . M. QuezadoLaboratory of Pathology, National Cancer Institute,National Institutes of Health,Bethesda, MD, USA

hyperintense lesions on T2 and fluid-attenuated inverserecovery (FLAIR) images on the left.

He had no hypertension. His renal function was normaluntil age 37 when his serum creatinine was found to be1.4 mg/dL (normal 0.9–1.4) with a creatinine clearance of78 mL/min. He had mild-to-moderate proteinuria withvariable 24-h protein excretion of up to 1.2 g/24 h. At age43, his serum creatinine level was 1.5 mg%. His total bloodcholesterol at age 20 was 189 mg/dL. At age 36, it was201 mg/dL, and his HDL and triglyceride levels were87 mg/dL and 92 mg/dL, respectively. At age 43, totalcholesterol was 241 mg/dL, LDL level was 161 mg/dL, andHDL level was 24 mg/dL.

Other abnormalities noted were bilateral pedal edema,mild-to-moderate bilateral high-frequency hearing loss,sinus bradycardia and left ventricular hypertrophy in theelectrocardiogram, and mild mitral and tricuspid valveregurgitation.

His therapy included Coumadin and carbamazepine. Inaddition, he received intravenous α-galactosidase A infu-sions (agalsidase beta; Genzyme Corporation, Cambridge,

MA) at a dosage of 1 mg/kg every 2 weeks for the last 2.5years of his life.

He first complained of sharp chest pain at age 40, lastingabout 30 s (not precipitated by exertion). He complainedmostly of burning chest pain on and off in the years prior tohis death. One week prior to his death, he again reported aburning discomfort on his chest, which increased withphysical activity. He refused to seek medical attention andshortly thereafter was found dead.

Regarding his family history, his mother, two sisters, anda brother were affected by Fabry disease. Two of hismaternal uncles died of the same disease. One died of acardiac cause at age 39 and another maternal uncle died ofstroke at age 47. Another cousin with Fabry disease died ofmyocardial infarction at age 53.

Pathological findings

The postmortem interval was approximately 32 h before anunrestricted autopsy was performed. Only the pertinentfindings will be described.

The heart weighed 520 g with approximately 30–50 mLof pericardial fluid. The right dominant coronary arterialsystems showed marked atheromas with calcifications. Theleft ventricular wall thickness was 1.7 cm (normal 1.0–1.5 cm). Sections of the left coronary arterial systemshowed significant intimal atheromas with intimal prolif-eration and calcifications with reduction of the luminaldiameter of up to 95% in some regions (Fig. 1a). The rightcoronary artery also showed significant intimal atheromasand calcification with reduction of the interior diameter ofup to 50%. Microscopically, portions of the anterior freewall of the left ventricle showed an inflammatory infiltrateconsisting predominately of neutrophils with necrosis ofthe myocardium (Fig. 1b). Additional sections of theanterior wall distant from the inflammatory infiltrateshowed a significant patchy, fibrous scarring of themyocardium. Ultrastructural analysis showed the heartmuscle separated by thin strands of fibrosis (Fig. 2). Themyocytes showed focal loss of filaments and increasednumber of mitochondria containing tubulovesicular cristaeand dense aggregates. Numerous lamellar inclusions werefound in myocytes and fibroblasts. Inclusions were onlyrarely seen in endothelial cells (Fig. 2d).

The intimal surface of the aorta showed marked ath-erosclerotic ulcerative plaques. The orifices of major ar-teries were widely patent.

The lungs were heavy and consolidated with blood-tinged pulmonary edema. Pulmonary hemorrhage wasidentified in the peripheral hilar region of the right and leftlung. Microscopic examination showed significant con-gestion, with hemorrhage into the bronchioles and spillinginto the alveolar spaces with distal necrosis/infarction ofthe parenchyma. Other sections from the consolidated lungshowed areas of frothy, eosinophilic cell-free fluid fillingthe alveolar spaces and scattered, admixed hemosiderin-laden macrophages.

Fig. 1 Cardiovascular findings. a Section of the left coronary arteryshowing a marked intimal proliferation and stenosis of the lumen.Hematoxylin and eosin (H & E), ×20. b Representative section ofthe anterior portion of the left ventricle showing coagulativenecrosis of the muscle with wavy fibers and a dense polymorpho-nuclear leukocytic infiltrate. H & E, ×20

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The right kidney weighed 140 g and the left one weighed184 g. The pelvis and calices of both kidneys weremarkedly dilated with thinning of the renal cortical pa-renchyma. Microscopically, glomeruli were either normalwith occasional mesangial widening or sclerosis (Fig. 3).At the ultrastructural level, numerous lamellar inclusionswere present in the glomerular epithelium and tubularepithelial cells. Only an occasional inclusion was seen inendothelial cells of glomerular capillaries (Fig. 4). Glyco-

lipid deposits were not seen in peritubular capillaryendothelial cells and only rarely seen in mesangial cells(not shown).

The fixed brain weighed 1,520 g. The left cerebralhemisphere showed a cystic cavity area in the frontopari-etal lobes corresponding to an old infarct in the left MCAterritory. Microscopically, this area was characterized byabnormal brain parenchyma with abundant reactive astro-cytes, few foamy macrophages containing hemosiderin,

Fig. 2 Ultrastructure of the heart. a Large aggregates of concentriclamellated cytoplasmic inclusions displace cardiac myofibrils to theperiphery of the cell. Interstitial fibrosis and increased numbers ofmitochondria are secondary features of the cardiomyopathy. b High-er magnification of the lamellar inclusions and the accumulated

mitochondria. The latter exhibit tubulovesicular cristae and cell sizevariation. c The myelinoid inclusions sometimes consisted of parallel-stacked lamellae with a zebra-like pattern. d Inclusions were only rarelypresent in endothelial cells of adjacent interstitial capillaries (arrows)

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and calcification. The basilar artery was atherosclerotic.Luxol fast blue (LFB)-stained deposits were found in bloodvessels, mostly in the smooth muscle cells of the media andadventitia and little or none in endothelial cells (Fig. 5).Similar results were obtained by electron microscopy(Fig. 6). Neuronal storage was also observed in variousareas including the hypothalamus (Fig. 7a), brainstem, andspinal cord. Sections of the paraspinal ganglia showedprominent neuronal distention with accumulating storagedeposits (Fig. 7b).

Discussion

We describe herein the pathological abnormalities of a 47-year-old man with Fabry disease who died of acutemyocardial infarction associated with nonhypertensive

atherosclerotic coronary artery disease. Other significantfindings at autopsy were the characteristic Fabry cardio-myopathy with increased cardiac mass by both weight andarterial wall thickness, as well as microscopic fibrosis andmarked cellular storage. Typical primary and secondaryfindings of Fabry disease were also found in the kidney, thebrain, and the paraspinal sensory ganglia.

A number of autopsy and pathological reports of patientswith Fabry disease have been published [6, 7, 10, 20, 21].The pathological findings in this patient showed an almostcomplete absence of glycolipid deposits in vascularendothelial cells attributable to the agalsidase beta infu-sions. Similar findings of predominant clearing of capillaryendothelial cells were described in kidney biopsy materialin the pivotal trial of agalsidase beta [21]. The presence ofmarked storage in other cells, including smooth musclecells and pericytes of the vascular system, as well as in

Fig. 3 Representative section of the renal cortex showing aglomerulus with focal mesangial sclerosis. H & E, ×40

Fig. 5 Intracerebral blood vessels with Luxol fast blue (LFB)-stained deposits within vacuolated smooth muscle/adventitial cells.LFB–period acid Schiff (PAS), ×100 oil

Fig. 4 Ultrastructure of the kidney. a Abundant lamellar inclusions in the glomerular epithelial cells (podocytes) and tubular epithelial cells(not shown). b On the other hand, only an occasional inclusion was identified in the glomerular capillary endothelial cells (arrow)

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nonvascular cells, after more than 2 years of enzyme in-fusions suggests that the infused enzyme has limited accessto nonvascular endothelial cells. If so, this may constitute asignificant limitation of this therapeutic approach. We can-not however exclude the fact that greater storage clearanceis achieved in other patients. Efforts to modify recombinantα-galactosidase A or the development of other therapeuticstrategies may achieve more complete clearing of storagematerial. The partial clearance of glycolipid deposits fromthe vascular tree may explain in part the clinical impressionthat the incidence of strokes is not reduced in the first fewyears after α-galactosidase A infusions.

The involvement of the coronary arteries in Fabry diseasehas been previously recognized [1, 7, 9, 12], althoughminimized by others [19]. In addition, the possibility ofaccelerated atherosclerosis in Fabry disease has beenpreviously mentioned [11, 13, 15]. Reported autopsieshave shown that these coronary lesions are similar to typical

atherosclerosis except that they are concentric with a whitediscoloration rather than the eccentric yellowish character-istics of atherosclerotic plaques [1].

The severe atherosclerotic disease seen in this patientthat culminated in a fatal coronary event could be co-incidental to the Fabry disease process. Nevertheless, weincreasingly recognize fixed coronary artery disease amongour Fabry disease patient population. For example, fivemen with Fabry disease from a group of 38 patients whoparticipated in enzyme replacement trials (age range 19–49, not including this patient) were found to havesymptomatic fixed coronary artery disease requiring cor-onary bypass or angioplasty (three patients). In addition,we found that random (not during acute events) bloodlevels of myeloperoxidase (MPO) in patients with Fabrydisease are significantly higher than controls [15]. MPOhas been associated with the atherosclerosis process, en-dothelial dysfunction, and acute coronary events, and it

Fig. 6 Ultrastructure of the brain. The smooth muscle cells in thewall of a cerebral vessel shown in a semithin section in a (toluidineblue/fuchsin stain) are loaded with lamellar inclusions exhibitingconcentric and zebra-like patterns at the ultrastructural level (b).

Another semithin section (c) shows a capillary identified within thecerebral tissue, which exhibits many Fabry-type inclusions in per-icytes (arrows) but lacks endothelial cell inclusions (d)

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was recently shown to modify apo A-1 HDL cholesterol tomake it more likely to accumulate in macrophages [16].Evidence for excess production of reactive oxygen species,such as superoxide in Fabry disease, has also accumulated[14, 15]. Recently, another group has shown that aorticatherosclerosis in mice deficient in both apo E and α-galactosidase A is markedly accelerated compared witheither of those mouse models alone [3]. Together, thisclinical and experimental evidence suggests that theprocess of atherosclerosis may be accelerated in Fabrydisease. It is likely therefore that other factors in com-bination with α-galactosidase A deficiency render certainpatients with Fabry disease susceptible to atherosclerosis.Indeed, we have recently shown that polymorphisms ofgenes known to be risk factors for stroke modify thelikelihood of developing cerebral ischemic lesions in Fabrydisease [2]. Therefore, a number of genetic factors maymodulate the interaction between the α-D-galactosyl gly-colipid accumulation and vascular plaque formation.Additional investigation is necessary to confirm and furthercharacterize this process.

Acknowledgements We thank Dr. Kristen Carr for her participa-tion in the autopsy of this patient. This work was supported by theIntramural program of National Institute of Neurological Disordersand Stroke, NIH.

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