Enzyme replacement therapy (ERT) reverses the resting hyperperfusion and the abnormal regional vascular reactivity response induced by acetazolamide. ERT also normalizes the blunted cerebral perfusion response to ascorbate and reduces 3-nitrotyrosine staining of dermal blood vessels. The function of the peripheral nervous system is also improved by ERT, with reduction in neuropathic pain and an improvement in the detection threshold for cold and warm sensation in the hands and feet. Improvement in sweating and heat tolerance has also been recorded following ERT. Despite these positive results, ERT does not completely normalize the function of the peripheral nervous system, and a reduction in the incidence of stroke remains to be demonstrated.

Introduction

Enzyme replacement therapy (ERT) is the first specific therapy for Fabry disease. It has been available only for the past 5–6 years, so it is a little early to reach any definitive conclusions about whether this therapy can reduce the morbidity associated with CNS disorders, such as stroke. In order to appreciate fully the difficulty in assessing the effect of ERT on the neurological aspects of Fabry disease, one needs to understand the nature of these abnormalities (see also Chapter 22). In general, neurological abnormalities (as well as non-neurological abnormalities) associated with Fabry disease can be divided into potentially reversible deficits/abnormalities, such as hypohidrosis, neuropathic pain or thermal sensation deficits, and non-reversible but preventable deficits, such as ischaemic stroke [1]. One might expect that it would be easier to assess the effect of ERT on potentially reversible aspects of Fabry disease than on, for example, reduction of stroke risk. The latter can be demonstrated prospectively only by observational studies on a relatively large cohort of patients over a prolonged period of time. The relatively low rate of stroke events makes the demonstration of the effect of a preventive therapy particularly difficult in a rare disease with few affected patients. Furthermore, general therapies, such as antiplatelet agents and statins, are highly effective in stroke prevention, making it very difficult to identify the specific effects of ERT [2, 3].

In this chapter, we concentrate on the effect of ERT on vascular dysfunction, small-fibre neuropathy (pain and thermal threshold), hypohidrosis and the incidence of stroke in patients with Fabry disease.

Effect of ERT on the vasculopathy of Fabry disease

Because of the expected difficulty in demonstrating an effect of ERT on stroke risk, we hypothesized that the vasculopathy of Fabry disease is associated with abnormalities in blood components, blood flow and vascular wall or endothelial function (Virchow's triad), resulting in vascular dysfunction and representing a vascular diathesis. If so, this vascular diathesis should improve with disease-specific therapy and potentially serve as a surrogate indicator of vascular risk reduction, including ischaemic stroke [4]. As described in Chapter 22, we found that a prothrombotic state exists in patients with Fabry disease, with cerebral hyperperfusion and an excessively prolonged vasodilatory response to acetazolamide (altered arterial wall reactivity) [5], increased nitrotyrosine staining in dermal blood vessels and a delayed decrease in cerebral perfusion in response to ascorbate [4].

The effect of ERT on cerebral vascular perfusion and function was examined as part of two 6-month randomized controlled trials of agalsidase alfa conducted at the National Institutes of Health (NIH; Bethesda, MD, USA). In our initial study, using H₂ ¹⁵O and positron emission tomography (PET), we found a significant decrease in resting cerebral blood flow in patients on ERT compared with the placebo group (Figure 1). This indicates at least a partial reversal of the cerebral hyper-perfusion seen in this disorder [4]. The decrease in resting cerebral blood flow was confirmed using two other methods, one for measuring blood perfusion and the other for measuring cerebral blood flow velocity (transcranial Doppler) [6, 7].

Figure 1

(Left panel) Regional cerebral blood flow (rCBF) standard parametric t-mapping (SPM{t}) showing significantly increased blood flow in patients with Fabry disease (n = 26) compared with controls (n = 10) after template co-registration of individual positron (more...)

In the initial study, we also examined the response of the cerebral vasculature to acetazolamide, a drug that maximally dilates the cerebral blood vessels by decreasing the pH of the extracellular space of the vessel wall [8]. Cerebral blood vessels of patients with Fabry disease remained maximally dilated significantly longer than controls [5]. We found that agalsidase alfa reverses the excessive reactivity to acetazolamide compared with placebo after 6 months of treatment (Figure 2). This finding suggested an altered cerebral vasomotor tone in the blood vessels of patients with α-galactosidase A deficiency, and may be related to the mechanism of the dilated cerebral vasculopathy or dolichoectasia noted in Fabry disease. As evidence of cellular access of intravenously infused α-galactosidase A beyond the vascular endothelial cells is lacking, the improved response to acetazolamide suggests a role for vascular endothelial cells in the CNS aspects of Fabry disease.

Figure 2

Regional cerebral blood flow (rCBF) standard parametric t-mapping (SPM{t}) after an acetazolamide challenge, showing significantly increased blood flow in patients with Fabry disease (n = 26) compared with normal controls (n = 10). (Left panel) The rCBF (more...)

In another randomized placebo-controlled trial, we tested the hypothesis that reactive oxygen species (ROS) contribute to the cerebral hyperperfusion in Fabry disease by assessing the response of cerebral blood flow to the intravenous infusion of 1 g ascorbate over 4 minutes, a known scavenger of ROS [9]. This study, using quantitative arterial spin tagging and magnetic resonance imaging, confirmed the cerebral hyperperfusion seen with PET, and also demonstrated that healthy controls and patients on ERT responded similarly to ascorbate infusion by a decrease in cerebral blood flow [6]. The patients on placebo had a significant delay in the reduction of cerebral blood flow, again suggesting a defect in vessel reactivity, possibly due to an excess production of ROS in Fabry disease (Figure 3) [6]. This finding was supported by the observation of significantly increased staining for 3-nitrotyrosine in dermal and cerebral blood vessels of patients with Fabry disease [4] and in the increased 3-nitrotyrosine and myeloperoxidase in the blood of patients with the disease compared with controls [6]. In the initial NIH randomized controlled trial, we found a significant reduction in dermal 3-nitrotyrosine staining in patients receiving ERT, suggesting reduction in the potentially toxic effect of ROS secondary to peroxynitrite formation (Figure 4) [4].

Figure 3

Comparison between the change in regional cerebral blood flow (rCBF) after infusion of ascorbate between (a) patients with Fabry disease given placebo and normal control individuals, and (b) patients with Fabry disease given enzyme replacement therapy (more...)

Figure 4

Dermal nitrotyrosine (a, c, e) and laminin (b, d, f) immunohistochemical localization in control skin and in patients with Fabry disease before and after enzyme replacement therapy (ERT). (a) Control skin showing nitrotyrosine staining (rhodamine filter, (more...)

Despite the significant improvement in the function of the cerebral vasculature, four of 25 patients in our original study followed for 4.5 years on ERT developed non-debilitating strokes and one patient had a transient ischaemic attack. One stroke involved a large vessel (vertebral artery occlusion) and the others were small-vessel strokes. Based on our experience and that of others, strokes also continue to occur in patients on agalsidase beta [10], and there is no indication of a marked reduction in stroke risk in the first few years following initiation of ERT in adulthood. To our knowledge, however, no formal study has addressed this question. The lack of an observed clinical effect can be explained in a number of ways. First, there may be a small therapeutic effect that cannot be detected in the relatively small number of patients studied. It is also possible that the infused α-galactosidase A does not have access to the entire thickness of cerebral vessels and therefore leaves untreated a significant component of the vessel wall (Figure 5). In addition, the intermittent nature of ERT and the possibility of pre-existing irreversible structural changes in the vasculature may limit the effect of ERT when initiated in adulthood. It remains to be demonstrated whether better stroke prevention can be obtained when ERT is begun in childhood.

Figure 5

Effect of 28 months of enzyme replacement therapy (ERT) using agalsidase beta on cerebral vasculature. Sections from the brain of a patient who died of an acute myocardial infarction after long-term ERT are stained with Luxol Fast Blue. (a) Blood vessel (more...)

Effect of ERT on the peripheral nervous system

Our studies, and those of others, focused on neuropathic pain scores, sensory detection threshold for cold, warmth and vibration, sweat function and epidermal innervation density. There was also an attempt to study the effect of ERT on hearing in Fabry disease, which we shall briefly describe.

Neuropathic pain

In the initial NIH 6-month randomized controlled study, we found a significant reduction in pain scores, using the Brief Pain Inventory, in patients on ERT compared with those on placebo [11]. When the patients on placebo crossed over to receive agalsidase alfa, they exhibited a similar benefit (Figure 6) [12]. When followed over a longer period of time, however, there was no further reduction in pain scores (Figure 6). For these studies, patients were selected for severe neuropathic pain, and medication for neuropathic pain was stopped 1 week prior to pain scoring [12]. Our clinical impression since then supports these initial findings. Neuropathic pain is reduced in patients, including children (authors' unpublished data); however, it is not usually completely eliminated and patients often need to continue with their pain medication, albeit at a lower dose.

Figure 6

Pain as assessed using the Brief Pain Inventory (BPI) in patients given enzyme replacement therapy (ERT) with agalsidase alfa for 24 months and those given placebo for the first 6 months followed by ERT for the next 18 months. Values are means ± (more...)

Peripheral nerve sensory function

Using a well-established biophysical method based on computerized automated sensory testing equipment (CASE IV, WR Medical, Rochester, MN, USA) to measure detection thresholds for warmth and cold in the foot, thigh and hand, patients with Fabry disease were found to have significantly elevated detection thresholds for warm and cold stimuli in the foot, and for cold stimuli in the hand, compared with controls. Warm sensation was found to be normal in the hand [12]. ERT with agalsidase alfa had no significant effect on these sensory parameters over the 6-month period of the randomized controlled trial. Over the 3 years of open-label treatment, however, there was a significant but modest reduction in the cold (Figure 7) and warm detection thresholds in the foot in patients receiving ERT and for warm perception in the thigh. There was also a trend for reduction of cold detection thresholds in the hand [12]. This effect took about 18 months to occur and sensory function seemed to stabilize thereafter. Similar results, looking particularly at heat pain thresholds, were obtained by a group treating patients with agalsidase beta [13]. These authors also described an improvement in vibration detection thresholds. In two separate studies, we found that patients with Fabry disease had a normal vibration detection threshold. However, over time there was no change in vibration threshold in the hand, but a significant increase in the foot threshold, possibly reflecting uraemic neuropathy in some patients. The vibration detection function, however, remained within the normal range for the patient group as a whole [12]. The functional improvement in cold perception of about 10% was not associated with an increase in epidermal innervation density [14].

Figure 7

Thermal sensation in patients with Fabry disease on enzyme replacement therapy (ERT). Kaplan–Meier curves show the proportion of patients not responding to cold stimulus in the foot as a function of the stimulus threshold. The solid line (a) represents (more...)

Overall, although these findings are encouraging, they do not suggest complete normalization of peripheral nerve function. It may be that early treatment before irreversible axonal damage, or higher and more frequent dosing, may be more effective. Alternatively, as mentioned above for the Fabry vascular diathesis, perhaps the infused enzyme has insufficient access to affected sensory nerves and ganglia.

Sweat gland function

Sweat gland function in Fabry disease is of particular interest, as it is possible to measure sweat gland function directly. Moreover, as the capillaries around the sweat glands are fenestrated, it might be expected that ERT would improve sweat gland function relatively early in the course of therapy [15]. We studied sweat gland function using the quantitative sudomotor sweat test (QSART) [16]. As we did not have this technique at our disposal at the start of our initial randomized controlled study, the study of sweat gland function was started at the 3-year time point for this patient cohort. Sweat gland function was found to improve 24–48 hours after enzyme infusion compared with pre-infusion values (Figure 8), while the QSART response normalized in four anhidrotic patients [12]. We also observed a significant improvement in sweat response over time in another study (Figure 9). To date, however, some patients have remained anhidrotic despite years of ERT.

Figure 8

Quantitative sudomotor axon reflex test (QSART) in patients with Fabry disease and controls. The QSART response is shown in controls (n = 38) and in patients 3 years after the start of enzyme replacement therapy both immediately before an infusion and (more...)

Figure 9

Quantitative sudomotor axon reflex test in a patient (a) prior to initiation of enzyme replacement therapy (ERT) with agalsidase alfa, (b) after 6 months of ERT and (c) after 3 years of ERT. This patient became extremely sensitive to acetylcholine stimulation (more...)

Conclusions

ERT has shown promise in ameliorating some of the neurological manifestations of Fabry disease. However, this form of therapy needs to be optimized in the future (e.g. by testing more frequent administration of the enzyme). Structural modification of the infused protein to allow better delivery of enzyme within organs and tissues should be attempted. Controlled trials should be performed to assess whether initiating ERT in early childhood will further improve the neurological outcome of patients with Fabry disease.

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