Neonatal diagnosis and treatment of Menkes disease.

BACKGROUND: Menkes disease is a fatal neurodegenerative disorder of infancy caused by diverse mutations in a copper-transport gene, ATP7A. Early treatment with copper injections may prevent death and illness, but presymptomatic detection is hindered by the inadequate sensitivity and specificity of diagnostic tests. Exploiting the deficiency of a copper enzyme, dopamine-beta-hydroxylase, we prospectively evaluated the diagnostic usefulness of plasma neurochemical levels, assessed the clinical effect of early detection, and investigated the molecular bases for treatment outcomes. METHODS: Between May 1997 and July 2005, we measured plasma dopamine, norepinephrine, dihydroxyphenylacetic acid, and dihydroxyphenylglycol in 81 infants at risk. In 12 newborns who met the eligibility criteria and began copper-replacement therapy within 22 days after birth, we tracked survival and neurodevelopment longitudinally for 1.5 to 8 years. We characterized ATP7A mutations using yeast complementation, reverse-transcriptase-polymerase-chain-reaction analysis, and immunohistochemical analysis. RESULTS: Of 81 infants at risk, 46 had abnormal neurochemical findings indicating low dopamine-beta-hydroxylase activity. On the basis of longitudinal follow-up, patients were classified as affected or unaffected by Menkes disease, and the neurochemical profiles were shown to have high sensitivity and specificity for detecting disease. Among 12 newborns with positive screening tests who were treated early with copper, survival at a median follow-up of 4.6 years was 92%, as compared with 13% at a median follow-up of 1.8 years for a historical control group of 15 late-diagnosis and late-treatment patients. Two of the 12 patients had normal neurodevelopment and brain myelination; 1 of these patients had a mutation that complemented a Saccharomyces cerevisiae copper-transport mutation, indicating partial ATPase activity, and the other had a mutation that allowed some correct ATP7A splicing. CONCLUSIONS: Neonatal diagnosis of Menkes disease by plasma neurochemical measurements and early treatment with copper may improve clinical outcomes. Affected newborns who have mutations that do not completely abrogate ATP7A function may be especially responsive to early copper treatment. (ClinicalTrials.gov number, NCT00001262.)

Differences in ATP7A gene expression underlie intrafamilial variability in Menkes disease/occipital horn syndrome.

BACKGROUND: Pronounced intrafamilial variability is unusual in Menkes disease and its variants. We report two unrelated families featuring affected members with unusually disparate clinical and biochemical phenotypes and explore the underlying molecular mechanisms. METHODS: We measured biochemical markers of impaired copper transport in five patients from two unrelated families and used RNase protection, quantitative reverse transcription (RT)-PCR, Western blot analysis and yeast complementation studies to characterise two ATP7A missense mutations, A1362D and S637L. RESULTS: In two brothers (family A) with A1362D, RNase protection and Western blot analyses revealed higher amounts of ATP7A transcript and protein in the older, mildly affected patient, who also had a higher plasma copper level and lower cerebrospinal fluid dihydroxyphenylalanine : dihydroxyphenylglycol ratio. These findings indicate greater gastrointestinal absorption of copper and higher activity of dopamine-beta-hydroxylase, a copper-dependent enzyme, respectively. In family B, three males with a missense mutation (S637L) in an exon 8 splicing enhancer showed equally reduced amounts of ATP7A transcript and protein by quantitative RT-PCR and western blot analysis, respectively, despite a more severe phenotype in the youngest. This patient's medical history was notable for cardiac arrest as a neonate, to which we attribute his more severe neurodevelopmental outcome. CONCLUSIONS: These families illustrate that genetic and non-genetic mechanisms may underlie intrafamilial variability in Menkes disease and its variants.

Safety of intracerebroventricular copper histidine in adult rats.

Classical Menkes disease is an X-linked recessive neurodegenerative disorder caused by mutations in a P-type ATPase (ATP7A) that normally delivers copper to the developing central nervous system. Infants with large deletions, or other mutations in ATP7A that incapacitate copper transport to the brain, show poor clinical outcomes and subnormal brain copper despite early subcutaneous copper histidine (CuHis) injections. These findings suggest a need for direct central nervous system approaches in such patients. To begin to evaluate an aggressive but potentially useful new strategy for metabolic improvement of this disorder, we studied the acute and chronic effects of CuHis administered by intracerebroventricular (ICV) injection in healthy adult rats. Magnetic resonance imaging (MRI) after ICV CuHis showed diffuse T(1)-signal enhancement, indicating wide brain distribution of copper after ICV administration, and implying the utility of this paramagnetic metal as a MRI contrast agent. The maximum tolerated dose (MTD) of CuHis, defined as the highest dose that did not induce overt toxicity, growth retardation, or reduce lifespan, was 0.5mcg. Animals receiving multiple infusions of this MTD showed increased brain copper concentrations, but no significant differences in activity, behavior, and somatic growth, or brain histology compared to saline-injected controls. Based on estimates of the brain copper deficit in Menkes disease patients, CuHis doses 10-fold lower than the MTD found in this study may restore proper brain copper concentration. Our results suggest that ICV CuHis administration have potential as a novel treatment approach in Menkes disease infants with severe mutations. Future trials of direct CNS copper administration in mouse models of Menkes disease will be informative.

Brachial artery aneurysms in Menkes disease.

Generalized vascular tortuosity caused by deficiency of the copper enzyme lysyl oxidase is frequently noted in Menkes disease, but reported examples of peripheral aneurysms are rare. We describe bilateral brachial artery aneurysms in a 10-month-old infant with classical Menkes disease.

Functional copper transport explains neurologic sparing in occipital horn syndrome.

OBJECTIVE: A range of neurologic morbidity characterizes childhood-onset copper transport defects, including severe Menkes disease and milder occipital horn syndrome. Both phenotypes are caused by mutations in ATP7A, which encodes a copper-transporting adenosine triphosphatase, although defects causing occipital horn syndrome are rarely reported and nearly always involve exon-skipping (six of eight prior reports). Our objective was to characterize a novel occipital horn syndrome mutation (N1304S) not associated with aberrant splicing and to determine whether functional copper transport was associated with this allele. METHODS: We studied two brothers with typical occipital horn syndrome and used yeast complementation and timed growth assays, exploiting a Saccharomyces cerevisiae mutant strain, to assess in vitro N1304S copper transport. RESULTS: We documented that N1304S has approximately 33% residual copper transport, a result not inconsistent with a similar patient we reported with an exon-skipping mutation whose cells showed correctly spliced mRNA transcripts 36% of normal. CONCLUSION: These patients' mild neurologic phenotypes, together with our yeast complementation and growth experiments, imply that N1304S does not completely block copper transport to the developing brain early in life. The findings suggest that neurologic sparing in untreated occipital horn syndrome is associated with approximately 30% residual functional activity of ATP7A.