Ethical considerations of population screening for late-onset genetic disease.

Population-based genetic screening has been a mainstay of public health in the United States for many years. The goal of genetic screening is to identify individuals at increased risk for treatable diseases. The evolution of genetic testing to include multi-disease panels allows for new screening applications which challenge the traditional model of clinical genetics care by the identification of late-onset disorders in an asymptomatic fetus, child, or adult. We present two unique examples of individuals referred to a biochemical genetics clinic due to the detection of late-onset Pompe disease by population-based screening modalities. We review early experiences in counseling and management of pre-symptomatic individuals and highlight some of the primary ethical factors warranting consideration as we enter the era of genomic medicine.

Absence of pyruvate decarboxylase activity in man: a cause of congenital lactic acidosis.

A complete deficiency in the pyruvate dehydrogenase system activity contributed to the death of a 6-month-old infant with congenital lactic acidosis. The enzymatic block could be isolated to the first component, pyruvate decarboxylase (E1) of the pyruvate dehydrogenase complex. This enzymatic deficiency allowed a demonstration of an "intercomplex" exchange of the components of the mammalian pyruvate dehydrogenase system and indicated that the first component is normally present in an apparent excess.

Detection of the carrier state in combined immunodeficiency disease associated with adenosine deaminase deficiency.

A large pedigree containing a child with severe combined immunodeficiency disease (CID) associated with adenosine deaminase (ADA) deficiency was investigated to ascertain if heterozygotes could be detected by measuring red cell ADA activity. 9 of 17 individuals in three generations who were at risk for being heterozygous had decreased red cell ADA activity. This genetic information establishes one form of CID as an autosomal recessive disorder. The identified heterozygote population had a mean ADA value of 19.2 U/g hemoglobin (0.95 confidence interval; 14.0 to 24.4 U/g hemoglobin), which was approximately one-half the mean, 36.1 U/g hemoglobin, of a randomly selected control population (0.95 confidence interval; 22.5-58.1 U/g hemoglobin). Statistical comparisons of the heterozygotes to the normal population indicates that within a high-risk family heterozygotes may be identified with 90% confidence.

Three point mutations in the factor IX genes of five hemophilia B patients. Identification strategy using localization by altered epitopes in their hemophilic proteins.

In five patients with hemophilia B and detectable Factor IX antigen, altered reactivity to a specific polyclonal antibody fraction or monoclonal anti-Factor IX antibodies was noted. Amplification of selected portions of their Factor IX genes by polymerase chain reaction allowed rapid identification of a single base transition in each of the five families tested. In a patient with severe hemophilia and an altered calcium binding domain, a G to A transition in exon II changed the codon for Glu-27 to Lys (Factor IXSeattle 3). Patients from two families with mild hemophilia with decreased reactivity to a MAb that binds to a site within the sequence coded for by exon IV had a G to A transition changing the codon for Gly-60 to Ser (Factor IXDurham). Two unrelated patients with moderately severe hemophilia lacked reactivity to another murine monoclonal anti-Factor IX which binds to an epitope in the amino-terminal half of the heavy chain of Factor IXa. In these patients, exon VIII contained a G to A transition changing Arg-248 to Gln (Factor IXSeattle 4).

Homocystine-induced arteriosclerosis. The role of endothelial cell injury and platelet response in its genesis.

The atherogenic mechanism of homocystinemia has been defined by measuring endothelial cell loss and regeneration, platelet consumption, and intimal lesion formation in a primate model. Three groups of baboons were studied: (a) 8 control animals; (b) 15 animals after 3 mo of continuous homocystinemia; and (c) 11 animals after 3 mo of combined homocystinemia and oral treatment with dipyridamole. Experimental homocystinemia caused patchy endothelial desquamation comprising about 10% of the aortic surface despite a 25-fold increase in endothelial cell regeneration. Neither endothelial cell loss nor regeneration was changed significantly by dipyridamole. Homocystine-induced vascular deendothelialization produced a threefold increase in platelet consumption that was interrupted by dipyridamole inhibition of platelet function. All homocystinemic animals developed typical arteriosclerotic or preatherosclerotic intimal lesions composed of proliferating smooth muscle cells averaging 10-15 cell layers surrounded by large amounts of collagen, elastic fibers, glycosaminoglycans, and sometimes lipid. Intimal lesion formation was prevented by dipyridamole therapy. We conclude that homocystine-induced endothelial cell injury resulted in arteriosclerosis through platelet-mediated intimal proliferation of smooth muscle cells that can be prevented by drug-induced platelet dysfunction.

Purine nucleoside phosphorylase deficiency. Evidence for molecular heterogeneity in two families with enzyme-deficient members.

Purine-nucleoside phosphorylase (NP) deficiency is associated with severely defective thymus-derived (T)-cell and normally functioning bone marrow-derived (B)-cell immunity. In this study, two unrelated families with a total of three NP deficient members were investigated. High pressure liquid chromatography of the plasma of the three patients showed inosine levels greater than 66 muM. This nucleoside was absent from the plasma of their parents and control samples.NP was purified from normal human erythrocytes by affinity chromatography and an antiserum prepared in rabbits was used to study the NP variants in the two families. In family M the patient had no detectable erythrocyte NP activity and no detectable immunological-reacting material (irm) to the NP antibody. The parents, who are second cousins, had less than one-half of normal enzyme activity and approximately 14% irm attributable to a variant protein. Their electrophoretic patterns revealed a series of isozymes with slower than normal migration. In family B the patients had 0.5% residual enzyme activity and about one-half normal irm. Their electrophoretic pattern showed faintly staining bands which migrated faster than normal NP. The mother of the patients had one-half normal enzyme activity, 11% irm attributable to her variant protein, and a normal electrophoretic pattern. The father had less than one-half normal enzyme activity, equal amounts of normal and variant irm, and an electrophoretic pattern that showed increased activity of the more rapidly migrating isozyme bands.The combined use of immunological and electrophoretic techniques has shown the presence of three separate mutations; one in family M and two in family B associated with severely defective T-cell function.

An intragenic deletion of the factor IX gene in a family with hemophilia B.

A family of seven patients severely afflicted with hemophilia B has been studied for their factor IX genes through the use of factor IX cDNA and genomic DNA probes. The patients had detectable (less than 10% of normal) factor IX antigen in urine and no detectable inhibitors in sera to factor IX protein. Based on the DNA hybridization analysis, these patients showed a partial intragenic deletion in their factor IX gene. The deletion included two exons (exons V and VI) coding for the amino acid sequence from number 85 to 195 of the factor IX protein. The deleted portion of the gene contained the entire factor IX activation peptide. The length of the deletion was estimated to be 10 +/- 0.3 kilobase pairs. This specific gene has been named FIXSeattle. In this family both the deletion and a Taq 1 restriction fragment length polymorphism can be used as a useful marker for accurate detection of female carriers of the deficient factor IX gene.

Adenosine deaminase deficiency: disappearance of adenine deoxynucleotides from a patient's erythrocytes after successful marrow transplantation.

Accumulation of adenine deoxynucleotides (dATP and dADP) in the erythrocytes of a patient with adenosine deaminase (ADA) deficiency was confirmed. The patient, now 18 mo old, was treated with a bone marrow transplantation from his HLA identical sister at 7 mo of age. Before and after the transplant, his erythrocyte and lymphocyte ADA activities, as well as his erythrocyte nucleotide profiles, were measured. 10 wk after the marrow transplant, no ADA activity could be detected in his erythrocytes, whereas there was a mixture of donor and patient lymphocytes as measured by ADA assays and karyotyping. At the same time, both dATP and dADP had disappeared from his erythrocytes, which were entirely of patient origin. These findings indicate that partial engraftment of donor lymphocytes into an ADA-deficient patient is capable of "correcting" alterations of deoxynucleotide concentrations in the patient's ADA-deficient erythrocytes.

Fabry disease: baseline medical characteristics of a cohort of 1765 males and females in the Fabry Registry.

The Fabry Registry is a global observational research platform established to define outcome data on the natural and treated course of this rare disorder. Participating physicians submit structured longitudinal data to a centralized, confidential database. This report describes the baseline demographic and clinical characteristics of the first 1765 patients (54% males (16% aged < 20 years) and 46% females (13% < 20 years)) enrolled in the Fabry Registry. The median ages at symptom onset and diagnosis were 9 and 23 years (males) and 13 and 32 years (females), respectively, indicating diagnostic delays in both sexes. Frequent presenting symptoms in males included neurological pain (62%), skin signs (31%), gastroenterological symptoms (19%), renal signs (unspecified) (17%), and ophthalmological signs (11%). First symptoms in females included neurological pain (41%), gastroenterological symptoms (13%), ophthalmological (12%), and skin signs (12%). For those patients reporting renal progression, the median age at occurrence was 38 years for both sexes, but onset of cerebrovascular and cardiovascular events was later in females (median 43 and 47 years, respectively) than in males (38 and 41 years, respectively). This paper demonstrates that in spite of the considerable burden of disease in both sexes that begins to manifest in childhood or adolescence, the recognition of the underlying diagnosis is delayed by 14 years in males and 19 years in females. The Fabry Registry provides data that can increase awareness of common symptoms in all age groups, as well as insight into treated and untreated disease course, leading to improved recognition and earlier treatment, and possibly to improved outcomes for affected individuals.

The Gaucher registry: demographics and disease characteristics of 1698 patients with Gaucher disease.

BACKGROUND: The Gaucher Registry, the largest database of patients with Gaucher disease (GD) worldwide, was initiated to better delineate the progressive nature of the disorder and determine optimal therapy. This report describes the demographic and clinical characteristics of 1698 patients with GD before they received enzyme replacement therapy. METHODS: Physicians worldwide who treat patients with GD were invited to submit prospective and retrospective data for an ongoing registry, using standardized data collection forms, for central processing and review. RESULTS: Most patients were from the United States (45%) and Israel (17%), but patients are from 38 countries. Most (94%) had type 1 GD, fewer than 1% had type 2, and 5% had type 3. Mutant allele frequency data, available for 45% of patients, showed the most common alleles to be N370S (53%), L444P (18%), 84GG (7%), and IVS2+1 (2%). Twenty-five percent of L444P homozygotes (13 of 52 patients) had type 1 GD phenotype. Mean age at diagnosis in patients with the N370S/N370S genotype was 27.2 years (SD, 19.7 years); in L444P/L444P patients, 2. 3 years (SD, 3.2 years). Histories of bone pain and radiological bone disease were reported by 63% and 94% of patients, respectively; both were more likely in asplenic patients than in patients with spleens. Mean spleen and liver volumes were 19.8 and 2.0 multiples of normal, respectively. Anemia and thrombocytopenia were present in 64% and 56%, respectively. Thrombocytopenia was present in 13% of asplenic patients. CONCLUSIONS: The Gaucher Registry permits a comprehensive understanding of the clinical spectrum of GD because of the uniquely large sample size. The Registry will be useful in evaluating the effects of specific therapies in GD and the possible influences of environment, ethnicity, and genotype on the natural history of the disorder.

Gaucher disease: recommendations on diagnosis, evaluation, and monitoring.

BACKGROUND: Timely diagnosis and continued monitoring of patients with type I Gaucher disease is critical because skeletal involvement can permanently disable patients and visceral organ involvement can lead to abdominal pain and secondary hematologic and biochemical complications. OBJECTIVE: To seek clinical consensus for minimum recommendations for effective diagnosis and monitoring of patients with type I Gaucher disease. PARTICIPANTS, EVIDENCE, AND CONSENSUS PROCESS: Contributing authors collaborated in quarterly meetings over a 2-year period to synthesize recommendations from peer-reviewed publications and their own medical experiences. These physicians care for most patients with Gaucher disease in the United States and serve as the US Regional Coordinators for the International Collaborative Gaucher Group Registry, the world's largest database for this disorder. CONCLUSIONS: The definitive method of diagnosis is enzyme assay of beta-glucocerebrosidase activity. Schedules differ for monitoring complications of type I Gaucher disease, depending on symptoms and whether enzyme replacement therapy is used. Hematologic and biochemical involvement should be assessed by complete blood cell count, including platelets, acid phosphatase, and liver enzymes, at baseline and every 12 months in untreated patients and every 3 months and at enzyme replacement therapy changes in treated patients. Visceral involvement should be assessed at diagnosis using magnetic resonance imaging or computed tomographic scans. Skeletal involvement should be assessed at diagnosis using T1- and T2-weighted magnetic resonance imaging of the entire femora and plain radiography of the femora, spine, and symptomatic sites. Follow-up skeletal and visceral assessments are recommended every 12 to 24 months in untreated patients, and every 12 months and at enzyme replacement therapy changes in treated patients.

Adherence to Universal (barrier) Precautions during interventions on critically ill and injured emergency department patients.

In a study undertaken to determine compliance with Universal Precautions, we observed 129 personnel performing 1,274 interventions on 151 consecutive critically ill and injured patients in an emergency department setting in July 1988. Barrier precautions were fully adhered to 44.0% of the time. During interventions in patients with profuse bleeding, adherence was only 19.5% in contrast to 44.7% for those who were not bleeding. Adherence was 56.4% during minor interventions but only 16.7% during major procedures. Adherence rates varied among health care providers: residents, 58%; emergency staff physicians, 38%; consultant physicians, 43%; emergency nursing staff, 44%; paramedics, 8%; radiology technicians, 14%; and housekeeping, 91%. In a follow up questionnaire that ascertained reasons for lack of compliance, 47% of providers indicated that there was not always sufficient time to put on protective material, 33% felt that precautions interfered with skillful performance of procedures, and 23% stated that materials were uncomfortable. Only 2.7% felt that Universal Precautions did not work. Since there is no proven postexposure prophylaxis for human immunodeficiency virus, Universal Precautions must be rigorously followed until such time as they are shown not to be effective or an alternate approach is developed. Strategies to improve compliance and improvements in barrier technology need to be developed.

The neuropathology of the nonketotic and ketotic hyperglycinemias: three cases.

Abnormal dysmyelination constitutes a pathoanatomic basis for the mental retardation in two different aminoacidopathies, nonketotic hyperglycinemia and ketotic hyperglycinemia. In both conditions myelin is decreased in amount and vacuolated. Similar patterns of dysmyelination in different aminoacidopathies suggest that abnormal myelination results from inadequate synthesis of myelin proteins.

Severe type II Gaucher disease with ichthyosis, arthrogryposis and neuronal apoptosis: molecular and pathological analyses.

Severe infantile Gaucher disease associated with ichthyosis and neonatal death is a rare subgroup of Type II Gaucher disease. This group of infants has little, if any, detectable beta-glucocerebrosidase activity, and prior genetic analyses have been limited in detecting the mutations responsible for this phenotype. We document an Hispanic infant succumbing with arthrogryposis and collodion membrane covering the skin who had no detectable beta-glucocerebrosidase activity in tissue samples and who was homozygous for a rare recombinant allele, RecNciI. Microscopic evaluation demonstrated accumulation of Gaucher cells in visceral organs and extensive loss of neurons in the anterior horns, brainstem, and cortex of the nervous system. The apoptosis of neuronal cells from the anterior horns and brainstem are a reasonable explanation for the arthrogryposis and neonatal death, respectively.

Hawkinsinuria in two families.

Hawkinsinuria, a disorder of tyrosine metabolism has been documented in two families in the United States, in one of which there was clear evidence of autosomal dominant inheritance. Metabolic acidosis and failure to thrive appear to be confined to infancy. Tyrosyl metabolites and 5-oxoproline are also found only in infancy, while 4-hydroxycyclohexylacetic acid was present only with time. The disease may be detected by organic acid analysis or by staining an electropherogram for sulfur containing compounds.

Gaucher's disease in the presence of normal glucocerebrosidase activity.

We encountered an infant with clinical and histopathologic features of Gaucher's disease (infantile, type 2) with normal glucocerebrosidase (D-glucosyl-N-acylsphingosine glucohydrolase, E.C.3.2.1.45) activity. Biochemical analysis was performed on leukocytes, cultured skin fibroblasts, and liver. Normal activity of glucocerebrosidase previously has been reported in an older child with juvenile onset (type 3) Gaucher's disease and attributed to a deficiency of a sphingolipid activator protein. These rare cases illustrate and expand our concept of Gaucher's disease and may have both diagnostic and therapeutic implications.

Variable severity of pulmonary disease in adults with identical cystic fibrosis mutations.

Adults with CF followed in a university center were assessed for the presence of the most common CF gene mutation, delta-F508. Excluding one member of a sibling pair, 29 of 55 subjects had two copies of delta-F508 (homozygotes), 23 had one copy of delta-F508 with the other CF mutation not identified (complex heterozygotes) and three were lacking delta-F508. A wide range of clinical severity was seen among individuals carrying two copies of the delta-F508 gene, who are genetically identical at the CF gene locus. The number of individuals diagnosed with CF as adults was significantly lower in the homozygote group (1 of 29) as compared with the heterozygote group (7 of 24). No differences were detected between groups in pulmonary function, non-pulmonary complications or overall clinical severity. These results suggest that environmental or background genetic factors contribute significantly to the variability in pulmonary and other complications seen among individuals with CF.

Use of activated coagulation time to monitor heparin during cardiac surgery.

Activated coagulation time (ACT) for protamine reversal was monitored in 28 consecutive patients (Group 1) and a standard heparin-protamine protocol was used for an earlier series of 28 patients (Group 2). Although Group 1 received a significantly higher total heparin dose than Group 2 (p less than 0.01), the protamine dose for reversal was significantly less for the ACT group than for the controls (p less than 0.0005). The mean ratio of protamine to total heparin was 1 : 1 (range, 0.33 to 1.44) for the ACT group and 2 : 1 (range, 1.42 to 2.59) for the controls. There were no significant differences between the two groups in operative and postoperative blood loss, transfusion requirements, hematocrit, and partial thromboplastin time. This study shows that the ACT test did not reduce postoperative bleeding significantly when compared with our standard protocol. It also indicates that there is wide individual sensitivity to heparin and that significantly less protamine is required for reversal.

Characterization of phosphoglycerate kinase from human spermatozoa.

We have confirmed the presence of a unique electrophoretic isoenzyme of phosphoglycerate kinase (PGK) in homogenate extracts of human sperm. No genetic variation was found in 30 individual specimens tested. However, immunologic neutralization studies indicate that the sperm PGK is different from red cell PGK.

Lactose metabolism and time to pregnancy.

OBJECTIVE: To investigate whether, in the absence of galactosemia, relatively high intestinal lactase activity or low activity of an enzyme involved in galactose catabolism reduces fertility, as it does in the presence of galactosemia. DESIGN: Retrospective cohort study. SETTING: Healthy women selected from the community. PATIENTS: Fifty-three married women. INTERVENTION: Urinary galactose after an oral lactose challenge (a measure of intestinal lactase activity), erythrocyte galactose-1-phosphate uridyltransferase (transferase) activity, and transferase polymorphisms by isoelectric focusing. MAIN OUTCOME MEASURE: Pregnancy rate (number of pregnancies divided by number of months at risk) in the 12 months after stopping use of birth control to become pregnant. RESULTS: Relatively high urinary galactose was not related to a decreased rate of pregnancy during the first 12 months (> or = 24.6 compared with < or = 14.3 mg: relative risk [RR] = 1.9; 95% confidence interval [CI] = 0.86 to 4.0). Relatively high transferase activity was not related to an increased rate of pregnancy (> or = 19.5 compared with < or = 17.2 mumol/h per g hemoglobin: RR = 1.1; 95% CI = 0.56 to 2.4). Low-activity transferase polymorphisms were not related to a decreased rate (RR = 1.2; 95% CI = 0.58 to 2.5). CONCLUSION: Our study does not support the hypothesis that the biologic variation in galactose metabolism that exists in the general population influences infertility.