SECTION 1: GENERAL CONSIDERATIONS

SECTION 2: CAUSES OF DEVELOPMENTAL DISORDERS

SECTION 3: SPECIFIC CASES

SECTION 4: MANAGEMENT OF CONDITIONS

A. Definition of Mental Retardation: Diminished intellectual function associated with impaired adaptive behavior, which is first manifested during the developmental period.

B. Causes of Mental Retardation:

1. "Neuro-developmental Disorder": A condition which disrupts development of normal brain function: associated with measurable chemical or structural abnormalities, and which leads to various clinical symptoms, including mental retardation, seizures, neuromuscular dysfunction , and abnormal behaviors. Prevalence: 3:1000.

2. Psychosocial-Environmental: Prevalence: 20:1000. Associated with mild mental retardation (> 55 I.Q.)

SECTION 2: Classification of "Neuro-developmental disorders"

1. Inherited Disorders

A. Metabolic: Missing enzyme leads to build-up of toxic metabolite (secondary brain-damage)and/ or deficiency of essential end product needed for neuronal function (primary brain-damage).

"Secondary" Brain Damage

1. Amino Acids: phenylalanine accumulation(Phenylketonuria)

2. Purines/pyrimidines: Uric acid accumulation(Lesch-Nyhan Syndrome)

"Primary" Brain Damage

1. Lipids: Sulfatide-lipidosis (Metachromatic Leukodystrophy)

2. Mucopolysaccharides: (Sanfilippo Syndrome)

B. Structural: Cellular derangement or malfunction of brain tissue, usually with structural abnormalities of other body parts. No known enzyme deficiency or chromosomal abnormality.

1. Tuberous Sclerosis

2. Neurofibromatosis

C. Chromosomal: Triplet Repeat Disorders, microdeletions, and translocations.

1. Fragile X Syndrome

2. Myotonic Dystrophy

3. Translocation Down's Syndrome

4. William's Syndrome

5. Prader-Willi Syndrome

6. Angelman's Syndrome

2. Acquired Disorders

A. Prenatal Disorder

1. Infections (TORCH, Rubella)

2. Toxins (alcohol, maternal PKC)

3. Traumatic (ischemia/hypoxia)

4. Chromosomal Aneuploidies (not genetically transmitted) a. T-21 Down's Syndrome (autosomal aneuploidy) b. XXX (Sex chromosome aneuploidy)

5. Structural (Cellular derangement or malfunction of brain tissue, usually associated with structural abnormalities of other body parts). a. Lissencephaly Syndromes and other neuro-migrational disorders, b. Cornelia de Lange Syndrome, c. Sturge-Weber Syndrome, d. Rett Syndrome e. Septo-Optic Dysplasia.

6. Prematurity/Low Birth Weight

7. Hypothyroidism

B. Perinatal Disorder

1. Traumatic(hypoxia/ischemia)

2. Toxins (Kernicterus)

C. Post-Natal Disorder

1. Infection (measles)

2. Traumatic

3. Metabolic (hypothyroid)

4. Neoplasm

5. Toxins(lead)

1. Metachromatic Leukodystrophy .

2. Phenylketonuria

3. Tuberous Sclerosis

4. Neurofibromatosis

5. Fragile X Syndrome

6. Myotonic Dystrophy

7. Prader Willi Syndrome

8. Angelman's Syndrome

9. William's Syndrome

10. Congential Rubella Syndrome

11. Down Syndrome

12. Prematurity

13. Measles Encephalitis

14. Rett Syndrome

15. Lead Poisoning

16. Septo-Optic Dysplasia

17. Cornelia de Lange Syndrome

Case 1: Metachromatic Leukodystrophy

1. Etiology: Autosomal recessive Aryl Sulfatase-A deficiency, with accumulation of Sulfatide (a sphingolipid).

2. Clinical Manifestations: Ataxia, spasticity, intellectual impairment Age of Onset: a. Late infantile(l-2 years) b. Juvenile(5-l0 years) c. Adult Onset.

3. Clinical Stages of Progression: a. Weakness of legs with hypotonia. Requires support for walking. In juvenile type earliest sign may be failure of school work, emotional lability, and slight visual problems. (Lasts 1 year.) b. Patient can no longer stand. Loss of speech, hypotonicity replaced by hypertonicity, but with diminished DTR's. Mental regression increases. (Lasts 6 months.) c. Child is bedridden, quadriplegia with dystonic postures. Loss of vision (optic atrophy), and bulbar palsies (lasts 2 years). d. Loss of all contact with environment, blind, no speech, NG feedings. (Lasts years.)

4. Lab Diagnosis: a. Aryl Sulfatase A activity in urine and blood. b. Sural nerve biopsy - see metachromatic granules in the nerve (Sulfatide)

Ref: Treatment of Late Infantile Metachromatic Leukodystrophy with Bone Marrow Transplantation NEJM 322: 28-32, Jan. 1990 (Editorial same issue (p. 54) good review of Leukodystrophies)

Case 2: Phenylketonuria

1. Etiology: Autosomal recessive deficiency of hepatic Phenylalanine hydroxylose, with increased excretion of phenylpyruvic acid (positive ferric chloride test).

2. Incidence: 1:15,000 births

3. Clinical Features: Normal at birth; detect increased phenylalanine 2-3 days after initiation of milk feeding. Eczema begins in early infancy Agitated behavior by third month. Neurologic Symptoms: Hypertonicity, hyperactive reflexes, tremor, seizures (25%) by third month. Blonde hair, blue eyes, pale skin Musty odor (from phenylacetic acid)

4. Treatment: Restrict phenylalanine in diet (250-500 mg./day). Don't over restrict. Keep blood level less than 4 mg.%. (Phenylalanine and/or metabolite interfere with myelin production and/or neurotransmitter amine metabolism)

5. Maternal PKU Syndrome

6. Variant PKU (Biopterin deficiency). Need L-dopa + 5-hydroxytryptophan supplements.

7. Clinical Types: a. Classical PKU> 20 mg.% b. Atypical PKU 10-20mg% c. Benign hyperphenylalaninemia 4-10 mg.%

Ref: Isoleucine Leucine A New Treatment for PKU, AJDC 144: 539-543, May 1990. Valine Isoleucine and Leucine

Case 3: Tuberous Sclerosis

1. Definition: Inherited systemic disorder characterized by focal cellular derangement of any organ (especially brain, skin, kidney). Focal block of cellular differentiation may lend to neoplastic changes in those locations.

2. Genetics: Autosomal dominant with variable expression.

3. Incidence: 1:30,000 births, 0.5-1.0 % of patients in institutions.

4. Clinical and Pathological Features: a. CNS: Cortical nodules (best detected with MRI) Periventricular nodules (usually around foramina of Munro, calcified, cause increased intracranial pressure, seizures, mental retardation). Gliomatous change occurs in some. CAT scan best test. b. Skin: Facial angiofibromata, hypopigmented macules (87%), shagreen patches, cafe-au lait spots, Subungual fibromata (17%) c. Renal: Hamartomas(80%). May undergo malignant change d. Cardiac: Rhabdomyomata (10-30%) e. Eye: Retinal hamartomata or angiomata f. Lung: Cystic fibromyomatosis (sensitive to estrogen)

5. Recognition of gene "carrier." a. Look for skin lesions with Wood's lamp (hypopigmented macules 87 %) b. CT Scan - Periventricular calcification (may be only finding) c. Renal Scan- Renal tumors (80%)

6. Example of Variable Expression in a single family. Various combinations of mental retardation, seizures, renal tumors, skin lesions, cardiac rhabdomyomata.

7. Causes of death - CNS and renal usually.

Refs:1. Renal Cell Carcinoma in Patients with Tuberous Sclerosis, Pediatrics 80: 898-903, Dec. 1987. 2. Causes of Death in Patients with Tuberous Sclerosis. Mayo Clinic Proc. 66: 792-796 Augsut 1991. (Editorial pp. 868-872)

Case 4: Neurofibromatosis

1. Etiology: Autosomal dominant disorder of nerve tissue growth .Chromosome 17 (50% inherited, 50% new mutations). Gene product: GAP protein - regulates nerve cell growth.

2. Incidence 1:3000 births

3. Clinical features of NF-1 (NF-2 - very rare. Bilateral aucoustic neuromas)

a) Intracranial neoplasms: Neuromas of 5th cranial nerve, optic nerve gliomas, astrocytomas, meningiomas Malignant change - 2-8% b) May see Moya-Moya like disorder associated with seizures c) Aqueductal sterosis secondary to gliosis may lead to macrocephaly d) Mental retardation rare (50% with some learning problems) e) Pheochromocytomas f) Short stature - ? etiology g) Scoliosis

4. Clinical Grades

I (48%) only cafe au lait spots and a few neurofibromas. II (31%) mild learning problems, macrocephaly, mild scoliosis, multiple subcutaneous neurofibromas. III (8%) mild mental retardation, scoliosis, short stature, precocious puberty. IV (13%) mental retardation, intracranial tumors, malignancy.

NIH Criteria (Need at least 2)

1. 6 Cafe au lait spots > 1.5 cm

2. 2 Neurofibromas

3. 2 Lisch Nodules

4. Axillary freckling

5. Bone lesions

6. Family History

Ref: Neurofibromatosis Type 1: Neurologic and Cognitive Assessment with Sibling Controls AJDC 143: 833-837, .July 1989

Case 5: Fragile X Syndrome

1. Etiology: X-linked pattern of inheritance (males more severely affected). Cytogenetic testing reveals "Fragile" site on X chromosome demonstrated by incubation in folate deficient medium.

2. Frequency: 1:1350 males (10 % of all MR) 3. Clinical Features: a) long narrow face b) macroorchidism c) large ears d) autistic behavior in some (e.g. poor eye contact, stereotypic behavior)

4. Diagnosis:

a) Cytogenetic techniques: Show "fragile" site on X Chromosome when incubated in folate deficient medium.

b) Molecular analysis: These studies have revealed that 20-50% of male carriers are asymptomatic. The mutation must be "processed" in the female offspring before the deletertious effects can be seen in the progeny. Blood is tested for the FMR-1 gene expansion characteristic of the fragile-X syndrome, by Southern Blot and Polymerase Chain Reaction(PCR). Southern Blot would reveal a large expansion of the FMR-1 region. It is generally accepted that a person with less than 40 CGG repeats is normal. An individual with 60-230 repeats is a carrier,persons with greater than 230 Cgg repeats will be clinically affected. A positive PCR assay would show an amplified band at the control locus and no band from the FMR-1(CGG)n region, which means there is an expanded FMR-1 allele which is refractory to PCR amplification.

Ref: 1. Institutional Screening for the Fragile X Syndrome AJDC 142: 1216-1221, Nov. 1988. 2. Unraveling the Genetics of Fragile X Syndrome Science 252: 1070, 1991

Clinical Features: Muscle weakness, atrophy and myotonia of face and jaw and distal muscles, multi-colored subcortical cataracts, Type-II Diabetes Mellitus, mental retardation, recurrent pneumonia from ineffective air-way clearance, malfunction of pharynx, esophagus, intestines, cardiac conduction defects.

The Role of Large Triplet Repeat Length in the Development of Mental Retardation

Objective: To describe mental retardation and microcephaly as initial clinical signs in myotonic dystrophy (MD) with high trinucleotide repeats on chromosome 19 (at 19q13.3).

Patients and Methods: Two patients with maternally inherited MD were examined. Southern blot analysis was performed and trinucleotide repeat expansions were related to the findings of clinical and magnetic resonance imaging investigations.

Results: Both patients had the large GCT trinucleotide repeat expansions often seen in congenital MD, but they lacked the typical clinical signs. Mental retardation and microcephaly were the leading features present in infancy. Muscular weakness, in contrast, developed after age 35 years. Although there was no evidence for perinatal asphyxia or sleep apnea, magnetic resonance imaging disclosed reduced brain volume and subcortical demyelination.

Conclusions: Mental retardation preceding the development of muscle weakness suggests that the cerebral involvement in MD is a direct consequence of the genetic disorder and not mediated by muscle disease. Careful clinical examination of the parents for signs of MD should be considered in patients with cognitive deficits even without apparent muscular involvement. Abstract from the Archives of Neurology. 1997;54:251-254

COMMENT: The clinical course of the disease can be variable from mild to severe. Mental retardation is typically a feature, but why? How is this related to the muscle weakness? The paper reported here suggests that mental retardation is a consequence of the disease itself and not secondary to the muscle disorder.The Brain MRI that I obtained on one patient with a milder clinical course (as well as the patient reported here) had "bright areas" of demyelination subcortically and this probably explains the cognitive difficulties, but what is the mechanism of the de-myelination?

Myotonic Dystrophy is one example of a growing number of "triplet repeat" diseases. Normal individuals have 5-30 GCT repeats at chromosome 19q13.3, while affected people have 50 to several thousand GCT repeats at the same site on chromosome 19. Other examples include Fragile X and Huntingdon's disease. In each case there is an increased number of DNA "triplets". A triplet (or Codon) is a linked series of 3 nucleotides. Nucleotides are the building blocks of DNA and RNA and chemically are phosphate esters of nucleosides(adenosine, guanosine, cytidine, thymidine, uridine). These are abbreviated By A, G, C, T, U, respectively. Each amino acid requires a specific triplet in order for it to be incorporated into protein. For example the triplet for phenylalanine is UUU. This UUU triplet "carries" or transfers phenylalanine to the ribosome where it is incorporated into protein.

For some reason, with the triplet repeat disorders, a specific DNA triplet proliferates. This may be a primary process that causes the clinical disease, or a secondary phenomenon, possibly related to a disorder in provision of specific amino acids to the ribosome.

In contrast, the Angelman syndrome (AS) is a non-progressive genetic disorder characterized by more severe developmental delay and mental retardation, absence of speech, ataxia,seizures, macrostomia, and brachycephaly. In addition patients with AS have ataxic limb movements and gait, as well as inappropriate bouts of laughter (Happy Puppet Syndrome).

Although these two syndromes have virtually no overlapping clinical features, they are both caused by abnormalities in the same region of chromosome 15 at q11.2-q12 . Approximately 70% of patients with PWS have a cytogenetic deletion of chromosome 15q 11.2-12. On molecular analysis this deletion is found on the paternally derived chromosome 15. About 20 to 25% of patients with PWS have two copies of the maternally and no copy of the paternally derived chromosome 15 . This condition is known as uniparental disomy (UPD).

Point mutations undetectable by cytogenetic or current DNA analyses are thought to account for the other 5% of patients with PWS. About 60% of patients with AS have a deletion of the maternal copy of chromosome 15 and 5% have UPD for the paternally derived chromosome. The other 35% have point mutations or very small deletions that are undetectable by current methods . Like PWS, AS seems to be associated with few phenotypic differences resulting from UPD in comparison with a chromosome 15 deletion.

METHODOLOGIES of DELETION DETECTION: From the Mayo Clinic.

1. High-Resolution Cytogenetics: Analysis of high-resolution banded chromosomes on a blood specimen has been the traditional method for identifying persons with PWS or AS. In a metaphase spread at the 550-band stage the del(15)(q11.2-12) is often evident. In addition, cases of familial PWS due to a chrornosome 15 translocation and de novo structural anomalies of chromosome15 have been reported that would be best detected by high-resolution cytogenetic analysis. In the absence of a 15q11.2-q12 deletion, high-resolution chromosome analysis can be used to detect other chromosome abnormalities that cause features that overlap with the PWS or AS phenotype.

2. Fluorescence in Situ Hybridization: Fluorescence in situ hybridization (FISH) analysis, a relatively new technology. is beginning to have a major role in cytogenetic analyses. In the case of PWS or AS, DNA probes specific for the PWS-AS region are hybridized to metaphase cells. These cells can be from the same preparation used for cytogenetic analysis. The DNA probes are labeled with digoxigenin; thus on hybridization to a complementary site, they can be detected with antidigoxigenin coupled to a fluorophore such as fluorascein (green) or rhodamine(red). Additionally, another probe for the distal region of 15q is included in the mixture to act at a hybridization control.

Metaphase cells from a normal person show a FISH signal at 15q11.2-q12 on both chromosomes 15, whereas a patient with PWS or AS caused by a deletion will lack a probe signal on one chromosome 15 . In up to 20% of patients with PWS or AS, the deletion cannot be detected by high-resolution chromosome analysis, but it is delectable by FISH. Patients with PWS or AS who have UPD, however, have two morphologically normal chromosomes, a situation that results in a normal signal pattern by FISH analysis.

3. Southern Blot testing: Another approach for the assessment of PWS or AS involves southern blot analysis . With this method, we utilize the finding that paternal and maternal DNA at the "PWS-AS locus" is methylated differently . With use of the probe PW71B and two restriction endonucleases (HindIII and the methylation-sensitive HpaII) a normal person has both a 6.0-kb and a 4.4-kb fragment. The 6.0-kb fragment is maternally derived and the 4.4-kb fragment is paternally derived.

When PWS is caused by either a deletion or maternal UPD, the paternal PWS region is absent. Therefore, Southern blot analysis of patients with PWS demonstrates only the maternal 6.0-kb fragment. For patients with AS the banding pattern is the opposite. In AS caused by a deletion or paternal UPD the maternal allele is absent; thus only the paternal 4.4-kb band is present. Consequently southern blot analysis is capable of identifying both (but cannot distinguish) deletions and UPD for both PWS and AS.

4. Polymerase Chain Reaction: Although Southern blot analysis cannot distinguish between a deletion and UPD, the polymerase chain reaction (PCR) can be used to distinguish between these two possibilities. Normally occurring polymorphisms on the chromosomes 15 allow one chromosome to be distinguished from the other. For the highest degree or accuracy, a blood sample is needed from both parents and the affected person. By analysis of the chromosome 15 markers, one can determine whether the affected person has both a maternal and a paternal chromosome 15, two maternal chromosomes 15, or two paternal chromosomes 15.

Because three of the markers used are within the "PWS-AS region", a deletion can be distinguished from UPD.

TESTING STRATEGIES: In order to keep the cost of genetic testing to a minimum, our goal is to produce the most informative results with the least amount of work. This testing strategy must focus on the incidence of the disease or syndrome, the prevalence of the various types of mutations, and the referral pattern. 44 cases were reviewed by the Mayo Clinic Cytogenetics and Molecular Genetics Laboratories for PWS or AS. Of these 44 cases, 37 patients (84%) had a normal karyotype by high-resolution cytogenetic analysis and results by Southern blot analysis were normal. Although some of these patients may have been part of the 5% of those with PWS or the 35% of these with AS who have a mutation undetectable by these methods, the most likely explanation is that they did not have PWS or AS. In five patients (12%), a del(15)(ql1.2-12) was evident by cytogenetic analysis, FISH testing, and Southern blot analysis. In one patient (2%), the karyotype was normal, but the results of Southern blot testing indicated the presence of either UPD or a del(15)(q11.2-q12); FISH analysis of metaphase cells indicated that one chromosome 15 was deleted for the PWS-AS region. Finally, one patient (2%) had a normal karyotype and normal results by FISH analysis, but results by Southern blot analysis were abnormal; PCR analysis indicated that this patient had maternal UPD. This series included no specimens from patients with a chromosome abnormality other than a del(15) that could have produced features overlapping with the PWS or AS phenotype. On the basis of a review of a larger series of 106 specimens analyzed by the Mayo Clinic Cytogenetics Laboratory, such an overlap would be expected to occur in about 4% of specimens referred for "rule out" PWS or AS (unpublished data).

A protocol for specimens referred to the laboratory for PWS or AS testing that required only a single blood specimen from the patient was developed at the Mayo Clinic. Blood is collected in an acid-citrate-dextrose Vacutainer. For the first round of testing, this strategy used a combination of high-resolution cytogenetic and Southern blot analyses. The cytogenetic analysis will detect most deletions at 15 ql1-q12 and other chromosome anomalies that may cause features that overlap with the PWS or AS phenotype. Because the cytogenetic analysis may overlook some deletions and all instances of UPD, Southern blot analysis is also part of the first round of testing. This test will detect both the chromosome 15 deletions and the UPD responsible for PWS or AS. With use of this strategy and the previously mentioned referral pattern, the testing will be complete in about 96% of cases. In a small percentage of cases with positive Southern blot results, a distinction cannot be made between UPD or a deletion. In such cases however, the diagnosis of PWS or AS will still be correct.

The second round of testing should cover the remaining 4% of cases and would be required only if the referring physician thought it was necessary to distinguish between a deletion and UPD as the cause of PWS or AS. These specimens would be tested by FISH (the same specimen used for the cytogenetic analysis) to distinguish between these two possibilities. If UPD testing by PCR were required , a blood specimen would be needed from both parents of the affected person for further testing.

Conclusion: We believe that a combination of Southern blot and cytogenetic analyses provides the best strategy for the diagnosis of PWS or AS.

A Brief Course in Angeman's Syndrome Molecular Genetics by Dr. Williams

Chromosome 15 - The chromosome that is abnormal in Angelman syndrome. Normal people have 23 pairs of chromosomes, one derived from each parent. There are 22 pairs that are numbered numerically from 1 to 22, the final pair is an X and Y. We receive one chromosome 15 from our mother and one chromosome 15 from our father. Chromosomes contain millions of molecules that are condensed together at the time of cell division and thus are able to be seen under the microscope.

15q11-13 Region - Chromosomes are divided into short arms and long arms and have a central segment called a centromere. The short arm is call "p" and the long arm is called "q". The "q" region is divided numerically into several segments and the q11 - 13 segment refers to an area that is toward the middle of the number 15 chromosome. It spans about 5-10 million nucleic acid molecules so this region includes many genes. It is the region that is crucial in Angelman syndrome but also contains other genes such as those causing the Prader Willi syndrome.

15q11-13 Deletion - This usually refers to a spontaneous chromosome defect whereby a large common region spanning between 5-10 million nucleic acid molecules is missing from chromosome 15. Deletions can also be microdeletions and involve even smaller segments, but this is unusual in Angelman syndrome. Very tiny deletions can affect small regions such as the imprinting center or the region where the UBE3A gene is located (see below).

Gene - A gene is a small piece of the genetic code that contains sufficient information to produce a protein. Genes are located within chromosomes and are composed of molecules of nucleic acid hooked together in a specific sequence. Thousands of these sequences form the DNA code and it is the main component of chromosomes. Four nucleic acids are used in the DNA code and are designated C, G, A and T. They can be changed into a second code termed RNA which uses the code sequences U, C, A and G. RNA can then be used by the cell to directly translate the code into a series of amino acids which ultimately form proteins.

Imprinting - A process (not completely understood) whereby a gene is inactivated or silenced. The result is that only one of the normal two genes is active. Imprinting occurs during development of the egg and sperm. The Angelman syndrome chromosome region is imprinted meaning that some genes are only active on the chromosome derived from the mother They are silenced on the chromosome derived (inherited) from the father.

Imprinting Center - This is a small area of DNA on chromosome 15 within the 15q11-13 region. It is believed to have control over large regions of 15q11-13. When the AS gene is inherited from your father it is believed to be in a turned off state but when inherited from your mother it is in the turned on state. This requires that the gene change it's active status as it passes through generations. In a way that is yet unknown the Imprinting Center is able to imprint as well as "unimprint" the 15q11-13 region, thereby allowing it to undergo it's normal turning on and turning off process.

Meiosis - This process occurs in germ cells (testes, ovaries) whereby a parent's normal dose of DNA is processed and reduced to a half dose. The half dose is needed during the time of conception when two parental half doses come together to make the usual normal dose of genetic material. Meiosis occurs during the time of sperm and egg development. This is also the time when some genes are evidently imprinted, thus establishing their active and inactive status.

Methylation - This term is often used to describe DNA which has methyl (CH3) groups attached to certain regions of it. The extent of methylation can be associated with gene inactivation, so it may play an important role in the imprinting process.

Mutation - Any condition which causes a detrimental change in DNA. This can involve a change in a single molecule or involve large deletions and other abnormalities. The net affect of a mutation is that it ultimately changes the way the protein is made, sometimes preventing its production or creating an abnormal protein which does not function properly.

UBE3A Gene - A gene that has been shown to be disrupted in some children with Angelman syndrome. The precise function of this gene is not yet known but it is presumed to affect the function of ubiquitin in the brain.

Ubiquitin - A small molecule that is present inside all cells. It can be attached to molecules that are old and ready to be degraded or that need remove for whatever reason. This removal system is called the ubiquitin degradation pathway. The Angelman gene, UBE3A, is a component of this ubiquitin pathway but it remains unclear exactly how or if this gene helps degrade proteins in the brain.

Uniparental Disomy(UPD) - This occurs when both of the chromosomes from a particular pair are inherited from the same parent. In Angelman syndrome, the presence of two of the paternally derived number 15 chromosomes results in Angelman syndrome (i.e., uniparental disomy of chromosome is UPD-15).

II. Genetic Testing Definitions: High Resolution Chromosome (Karyotype) - This was a term applied to a chromosome analysis which examined the number 15 chromosome under high resolution in order to detect large deletions of 15q11-13. This test is rarely used today because some large deletions were actually missed using this method.

FISH - This test uses the process of fluorescent in situ hybridization which means that chromosomes are directly visualized under the microscope while a molecular fluorescent probe is used. The probes are specific for areas within 15q11-13 and when a small area is missing the probe fails to light up. In the normal state each chromosome fluoresces with the probe, but in Angelman syndrome, due to a large common deletion, only the paternal chromosome shows fluorescence.

DNA Methylation (Southern Blot Testing)- This test detects changes in the methylation status of nucleic acids in the 15q11-13 Angelman region. Within this area the paternal region contains a different number of methyl (CH3) molecules than the maternal region. When this region is then chopped up using enzymes that cut DNA, it creates different size fragments. Normal individuals show two distinct fragments, one from each of their parent's 15q11-13 region. If something disturbs the methylation status or the methylated region is completely missing, then the methylation pattern is abnormal. In about 70% of Angelman syndrome children, only the methylation pattern from the father's chromosome is seen indicating that the 15q11-13 region is either missing due to a large maternal deletion, or has two paternal 15 chromosomes or has a defect in the imprinting center.

UBE3A Direct Mutation Analysis - This involves examining DNA sequences inside the UBE3A gene. This test is not available at present and is currently in the research phase.Preliminary studies indicate that a small percentage of Angelman children who are normal by the above three tests may have mutations within this gene.

Normal Genetic Tests - This is not a test but represents a situation where a child is believed to have the clinical manifestations of Angelman syndrome but typically has normal high resolution chromosome, FISH and methylation studies. Most children with normal genetic studies have not had UBE3A mutation testing but it appears that most would still be normal even with this testing today. Accordingly, there remains a group of "genetically negative" children who show all the classical features of Angelman syndrome.

III. Inheritance of Angelman Syndrome: (Note: estimating recurrence risk In AS can be very complicated and professional genetic counseling is advised)

Large Common Deletion q11-13 - This is almost uniformly a sporadic occurrence without apparent increased risk in future offspring. Large common deletions occur in about 70% of cases of AS and are usually diagnosed by FISH tests using probes that map to region q11-13.

Microdeletion in 15q11-13 (molecular microdeletion) - This can occur in several molecular regions of 15q11-13 including the imprinting center and the area where the UBE3A gene resides. These microdeletions can either be sporadic (non-inherited) or they can be inherited from an apparently normal mother. If these microdeletions are also present in the mother then there is a 50% theoretical recurrence risk in future offspring.

UBE3A Mutations - This is a newly described group that has mutations within the putative AS gene. It appears that most of these mutations are sporadic and non-inherited although there is a risk, not yet well defined, that UBE3A mutations can be inherited from an apparently normal mother. If so, the theoretical risk would be 50%.

Uniparental Disomy for 15 - These appear to be sporadic. Recurrence risk for this disorder appears very low at less than 1%.

Normal Genetic Studies - This is a group that has completely normal genetic studies induding negative studies for the UBE3A gene. Sometimes affected siblings are included in this group and when this occurs, the theoretical recurrence risk is 50%. Accurate recurrence risks are difficult to establish for the overall group because of the varied nature of patients in this category.

Other Categories - There are a few AS individuals with unusual chromosome 15 rearrangements. In these cases, recurrence risk is variable depending on the chromosomal findings in the parents.Charles A. Williams, M.D. 7/4/97

RENAL ABNORMALITIES: Biesecker et al. (1987) described a 19-year-old patient with Williams syndrome who had renal cystic dysplasia and gradual deterioration of renal function, with recurrent episodes of dehydration secondary to a concentrating defect. They suggested that this is a more frequent complication than previously realized. In studies of 40 persons with Williams syndrome who were assessed at an average age of about 7 years, Pober et al. (1993) found renal abnormalities in 7: nephrocalcinosis in 2, marked asymmetry in kidney size in 2, small kidneys in 1, solitary kidney in 1, and pelvic kidney in 1. Renal artery stenosis was sought in 9 persons who underwent abdominal angiography during cardiac catheterization. Unilateral or bilateral mild renal artery narrowing was found in 4 persons and normal renal arteries in the remaining 5. Persistent hypertension was found in only 2 individuals and did not correlate with renal artery status.

CEREBROVASCULAR ABNORMALITIES: Kaplan et al. (1995) pointed out that stenoses in the cerebral arteries can cause strokes with brain damage and chronic hemipareses in children with Williams syndrome. Increased irritability, loss of consciousness, and seizures were initial signs in 2 patients. One patient, aged 22 years, had episodes of cerebral vascular insufficiency beginning at the age of 3 years at which time moyamoya was diagnosed.

EYE ABNORMALITIES: A stellate pattern was noted in the irides of 51% of the Williams syndrome patients and in 12% of the control subjects. The pattern was more difficult to detect or was absent in heavily pigmented irides. Hotta et al. (1990) reported on the iris pattern in 3 cases. Winter et al. (1996) assessed the frequency and severity of ophthalmologic features in 152 patients with Williams-Beuren syndrome. Eighty-two (54%) had strabismus, while 149 had esotropia. Blue irides were present in 117 (77%), green irides in 10 (7%), and brown irides in 25 (16%). A typical stellate iris pattern of the anterior stroma was found in 112 (74%).

CALCIUM METABOLISM: Taylor et al. (1982) investigated the effects of pharmacologic doses of vitamin D2 given for 4 days to normal children and to children with Williams disease and their sibs. The results indicated an exaggerated increase in serum 25-OH-D in response to challenge with vitamin D in patients with the Williams syndrome and in some of their sibs with no clinical features of the syndrome. Despite the increases in serum 25-OH-D, none of the patients became hypercalcemic. Garabedian et al. (1985) found high plasma concentrations of 1,25-(OH)2D in 4 children with hypercalcemia and elfin facies. The levels were higher than in 3 children with elfin facies but without hypercalcemia or dysmorphia. In Williams syndrome, a low calcium diet controlled the hypercalcemia. They suggested that an abnormal synthesis or degradation of 1,25-(OH)2D is present in this syndrome.

BIOCHEMICAL GENETIC FEATURES: Dutly and Schinzel (1996) carried out molecular genetic studies in 15 families with Williams Syndrome. They demonstrated a microdeletion of the elastin gene in all of the probands. The 15 families consisting of patients, parents, and paternal or maternal grandparents were genotyped using microsatellites adjacent to the centromeric or telomeric end of the elastin gene (ELN). They demonstrated that in 10 out of 15 William's Syndrome families with a de novo deletion within the elastin gene region of chromosome 7 (7q11.23), the segment flanking the deleted region contained recombined haplotypes. These recombination events indicated that deletion was the result of an unequal crossing over event between the chromosome 7 homologs during gametogenesis.

EDITOR: Williams syndrome is another example of a "micro-deletion disease". Prader-Willi syndrome is caused by a micro-deletion of chromosome 15. In the case of Williams syndrome, the micro-deletion is found in the "elastin" gene region of chromosome 7. To diagnose micro-deletions one might order a F.I.S.H. test (fluorescent in-situ hybridization). This utilizes a fluorescent DNA "probe" which normally would bind to the specific gene in question. If no fluorescence is seen, this means the gene is missing, and the F.I.S.H test would be positive for the disease.

At one Developmental Center there is a 35 year old man who turned out to have Williams syndrome, having been diagnosed by the geneticist after referral to rule out a genetic syndrome (positive family history of mental retardation). This individual has mild renal failure of undetermined etiology. Dynamic renal scans were normal. Perhaps he has subtle nephrocalcinosis.

Case 10: Congenital Rubella Syndrome

1. Etiology: Susceptible mother infected with Rubella virus during first trimester of pregnancy. Rubella virus inhibits mitosis, causes chromosomal breaks and neuronal damage, independent of inflammation.

2. Clinical Features: a) Congenital heart defects (septal defect, PDA) b) Peripheral pulmonic stenosis c) Hearing loss d) Visual loss (microphthalmia, "salt and pepper" retinopathy, cataract, glaucoma) e) Diabetes mellitus - 20% by 3rd decade f) Thyroid disease - 5% with Hashimoto's thyroiditis g) Growth retardation - ? etiology h) Mental retardation

Ref: A Profile of Mothers Giving Birth To Infants With Congenital Rubella Syndrome AJDC 1·14: 118-123, Jan. 1990

Case 11: Down Syndrome

1. Etiology: T-21 or Translocation

2. Frequency 1:1000 births

3. Clinical Features: a) Heart disease - VSD, ASD, mitral valve prolapse (40%). b) Prone to Hepatitis B Carrier State (25%) - monitor for cirrhosis and hepatoma c) Thyroid disease (40% with some abnormality by adulthood) d) Atlanto-axial instability (100%)? significance ? management. e) Alzheimer changes f) Leukemia (in infancy only) g) Optic- cataracts, keratoconus h) Hearing loss i) Amenorrhea (1/3 of women)

Ref: Correction of Atrioventricular Septal Defect: Results Influenced by Down Syndrome? AJDC 143: 1361-1365, Nov. 1989.

Case 12: Prematurity/low birth weight.

1. Etiology: Increased risk of intra or periventricular hemorrhage in infant of less that 1500 g. Intraventricular hemorrhage may damage brain cells directly or indirectly by causing hydrocephalus. In addition, premature infants (less than 1500 g) are more prone to the damaging effect of hypoxia-ischemia, hypoglycemia and hyperbilirubinemia.

2. The dilemma of the neonatal ICU.

Ref: Neurodevelopmental Performance of Very-Low-Birth-Weight Infants with Mild Periventricular, Intra-Ventricular Hemorrhage: Outcome at 5 to 6 years of age. AJDC : 1242-1245, Nov. 1990

Case 13: Measles Encephalitis

1. Etiology: Post natal (less than 6 months) infection with measles virus. Measles virus is neurotrophic. Does not affect children less than 6 months because of maternal antibody protection.

2. Incidence: 0.1 - 0.2 % of children who get measles.

3. Clinical Features: a) Usually affects children 6 mos. - 2 years. b) 50% have full clinical recovery, although EEG changes may persist for months. It is known that patients with even uncomplicated measles have EEG changes. c) Symptoms include lethargy and irritability which progress to coma or seizures, associated with fever, headache, vomiting.

1 - 3 days of symptoms, but may last a month. Approximately 1/3 are left with neurologic sequelae (mental retardation, paralysis, seizures).

4. Mechanism of Disease - 3 theories: a) Direct viral invasion b) Autoimmune or allergic demyelination c) Slow viral infection (SSPE)

Ref: Measles Outbreak Among Unvaccinated Preschool-Aged Children. Pediatrics 83: 369-373, March 1989.

Case 14: Rett Syndrome

1. Normal development until 6-18 months.

2. Followed by slowing of rate of head growth, loss of skills and cognitive function.

3. Gait and truncal apraxia/ataxia.

4. Repetitive hand movements.

5. May see: Breathing abnormalities, seizures, abnormal sleep, poor circulation of lower extremities, teeth grinding.

6. May progress over decades to quadriplegia and vegetative state.

Ref: Rett Syndrome: Natural History and Management Pediatrics 82: 1-10, July 1988

Case 15: Lead Encephalopathy

1. In children, lead causes necrosis of endothelial cells in developing blood vessels; this leads to transudation of fluid, hemorrhage and edema of the brain. Permanent neurological sequelae result (mental retardation, paresis, seizures). This may happen acutely or with chronic low level exposure.

2. Adults get peripheal neuropathy.

3. Organic lead is toxic at lower levels than inorganic lead.

Ref: Six Children with Lead Poisoning AJDC 144: 1039-1044, Sept. 1990.

16. Septo-optic dysplasia : The association of optic-nerve hypoplasia with absence of the corpus callosum and septum pellucidum combined with pituitary hormone deficiency is known as Septo-Optic Dysplasia (SOD), or the Syndrome of de Morsier .

Septo-Optic Dysplasia is an infrequently diagnosed disorder that typically presents in early infancy with blindness and growth retardation. SOD appears to be a disorder of embryological development of brain midline structures . There is evidence for both genetic and environmental etiologic factors, but the majority of evidence favors an environmental etiology .

While various pituitary hormone deficiencies have been reported, growth hormone deficiency is the the most frequent pituitary hormone deficiency associated with SOD. Growth hormone deficiency appears to be secondary to a lack of Growth Hormone Releasing Hormone which is normally produced by the hypothalamus . Therefore, a defective hypothalamus encountered in cases of SOD is not able to adequately stimulate production and release of Growth Hormone from an otherwise normal pituitary gland. Since Growth Hormone is important in maintaining glucose homeostasis, it has been previously suggested that the Growth Hormone deficiency associated with SOD is responsible for severe recurrent bouts of hypoglycemia which are known to occur in newborns with the disorder . It has also been suggested that young children with SOD should be given Growth Hormone injections to prevent growth retardation .

In conclusion, while SOD may be infrequently diagnosed, individuals with mental retardation who are visually impaired and are of short stature are commonly seen. When encountered by primary care physicians who serve these patients, neuroimaging studies should be performed . If midline developmental abnormalities are discovered , such as absence of corpus callosum , septum pellucidum and hypoplasia of the optic system, a complete evaluation to determine adequacy of pituitary function should be determined.

As with other syndromes, individuals with CdLS strongly resemble one another. Common characteristics include: low birthweight (usually, but not always, under five pounds), delayed growth and small stature, and small head size (microcephaly). Typical facial features include thin eyebrows which frequently meet at midline (synophrys), long eyelashes, short upturned nose and thin, downturned lips. Other frequent findings include excessive body hair (hirsutism), small hands and feet, partial joining of the second and third toes, incurved fifth fingers, gastroesophageal reflux, seizures, heart defects, cleft palate, bowel abnormalities, feeding difficulties, and developmental delay. Limb differences, including missing limbs or portions of limbs, usually fingers, hands or forearms, are also found in some individuals.

CdLS is a congenital syndrome, meaning it is present from birth. Most of the signs and symptoms may be recognized at birth or shortly thereafter. A child need not demonstrate each and every sign or symptom for the diagnosis to be made.

The exact incidence is unclear, but it is thought to be between 1:10,000 and 1:30,000 live births.

Earlier, many children died of serious medical problems in infancy because their needs were not anticipated. This is no longer the case, and it is expected that most will live into adulthood.

Mental retardation is usally associated, ranging from mild to profound. The majority fall in the moderate to severe range.

The cause is not clearly known, although it is suspected that a gene may be responsible. At present there are several research programs underway which are attempting to find answers to the cause of CdLS. It is likely that if a gene is involved, it is simply a rare and random mutation. This mutant gene is almost never passed on to the next generation because affected individuals seldom have children of their own. There have been rare instances in which mildly affected individuals have had children with the syndrome.

There are tests which may help resolve some of the uncertainty felt by CdLS families in future pregnancies. High resolution ultrasound may be useful to monitor for unusually poor fetal growth or detectable limb abnormalities. Genetic counseling centers are able to provide current information on the development of other prenatal tests.

A thorough medical evaluation including a history and physical examination, family history, laboratory tests, X-rays and chromosome analysis is usually conducted before a diagnosis is made. Since there is no specific test for CdLS, this is best accomplished through a referral to a genetics specialist or clinic. If you suspect that your child has CdLS, you should arrange for an evaluation by a genetics specialist. Arrangements can usually be made through your local physician, hospital, or university medical center.

Each child will progress at his/her own rate, but you can generally expect a slower than average rate of development. The area of speech and communication is often delayed, even in the more mildly affected. Infant stimulation programs and other developmental and therapeutic interventions are strongly recommended. Growth and development charts are available through the Foundation.

Further information is available from:

The Cornelia de Lange Syndrome Foundation, Inc.

302 West Main Street , #100 Avon, CT 06001 - USA (860) 676-8166 (860) 676-8337 (Fax) 1-800-753-CdLS (U.S. and Canada) 1-800-223-8355 (U.S. and Canada)

1. Etiologic Diagnosis :"Developmental onset" chronic brain dysfunction is a devastating condition from personal, familial, societal, and economic perspectives, yet the precise etiology is often unknown. This is unfortunate because knowledge of the exact diagnosis can at times lead to appropriate therapy which can markedly improve quality of life. In addition, diagnostic evaluation can provide important genetics counseling information which can reassure anxious family members.

The primary-care Developmental Medicine Physician should attempt to establish a precise etiology by first obtaining careful medical history. This may require chart review as well as interviews with family members. If prenatal or perinatal pathology is clearly documented (e.g. rubella infection, very low birth weight, severe problems during delivery), the diagnosis can be established by history alone. However, further evaluation would be indicated if there were no evidence for early abnormal developmental problems, but subsequent failure to attain normal developmental milestones. If deterioration occurred after a period of normal development, then further evaluation would also be indicated.

Physical examination should attempt to reveal "syndromic" features (e.g. tuberous sclerosis, neurofibromatosis, William's Syndrome, Prader-Willi Syndrome). If the history and physical examination do not suggest a diagnosis, then certain laboratory tests should be ordered.

Laboratory testing: Brain MRI can reveal a demylinating process, mass lesions , obstruction to CSF flow, and migrational disorders.Chromosome studies can reveal aneuploidies, deletions, and translocations.DNA analysis can reveal "triplet-repeat" disorders, and biochemical tests can reveal metabolic disorders. Biochemical screening tests for metabolic disorders include: 24 hour urine for amino acids, serum quantitative amino acids, spot urine for organic acids, urinary excretion of mucopolysaccharides, plasma amnonia level, plasma asylsulfatase A and hexosaminadase A levels. If these screening tests are all normal, referral to a medical neurogeneticist should be made. A medical report detailing the history, physical examination and laboratory results should accompany the referral to the medical neurogeneticist.

2. Seizures: Many patients with developmental onset chronic brain disorders have seizures. These seizures are often incompletely or inaccurately described, and may have multiple clinical presentations. At times true seizures may be confused with abnormal involuntary movements (see below). The primary-care physician should provide a careful written description of the clinical presentation of the seizure (or seizures). Special attention should be made to the initial manifestation (i.e. how it starts). A video recording of the seizure(s) should be obtained to assist in the neurological description. A 24-hour ambulatory EEG should be obtained whenever possible. Brain MRI should be obtained to rule out mass lesions, mesial temporal sclerosis, or other focal abnormalities. Drug therapy should be monitored by "longitudinal graphic analysis" (*). Epileptology consultation should be obtained in cases of "refractory" seizures (as defined by greater than 25 seizures/year in spite of trials of Tegretol, Depakote, Dilantin, Phenobarbital, singly or in combination).

3. Neuromotor Disorders:

a) "muscle rigidity": Muscle rigidity in the developmentally disabled might be caused by damage to either: (1) pyramidal (spastic), or (2) extra-pyramidal (cogwheel) systems. Pyramidal (spastic) type rigidity is by far the most common type encountered in patients with developmental onset chronic brain disorders. If untreated, pyramidal type rigidity leads to impaired mobility by causing ataxic movement and contractures. Impaired mobility is highly correlated with increased morbidity and mortality. Pressure sores, gastroesophageal reflux, constipation, and aspiration pleumonia are often associated with impaired mobility secondary to muscle rigidity. Before treatment is implemented, baseline video-analysis and goniometric (joint angle) measurements should be obtained. Treatment may include medical (e.g. Baclofen, Valium, Botox) or surgical (e.g. selective dorsal rhizotomy or percutaneous tendon release) modalities. Recent evidence suggests intra-thecal Baclofen may be highly efficacous treatment of spastic rigidity for some individuals. Regardless of the modalities chosen, efficacy should be documented by careful follow-up periodic video-analysis and goniometric measurements.

b) "dyskinesia "(abnormal involuntary movements): Abnormal involuntary movements are frequently seen in patients with developmental onset chronic brain disorders. They may appear as dystonia, choreoathetosis, myoclonus, tremor, or combinations of these. Stereotypies and tics may be closely related to involuntary movements (some call these "unvoluntary"). Treatment depends on an accurate diagnosis. MRI of the brain should be considered in cases of involuntary movement to rule out disease of the basal ganglia or cerebellum . Video-analysis of the dyskinesia should precede treatment. In addition, treatment efficacy should be documented by follow-up video analysis.

4. Severe Abnormal Behavior: Severe abnormal behavior may be symptomatic of: (1) A medical problem, (eg. drug toxicity and epilepsy), (2) a psychiatric disorder( eg. schizophrenia or OCD), (3) environmental toxicity (e.g. too much "stimulation"), or (4) a "personality" disorder (e.g. conduct disorder, manipulation). Accurate diagnosis requires coordinated interdisciplinary video and graphic analysis. Treatment efficacy should be documented by periodic follow-up video-graphic studies (*). The "Physician" (Primary Care Physician or Psychiatrist), along with the psychologist, should be responsible for the formulation of a "neurobehavioral" analysis (working hypothesis) before "treatment" is started. In addition, the physician and psychologist should be responsible for treatment efficacy documentation. 'Ihere is evidence that ventricular enlargement of the frontal lobes may be an indication for neuroleptic medication; therefore, a brain MRI scan might be considered in cases of severe refractory abnormal behaviors.

5. Recurrent Pneumonia (three times in a five year period): Recurrent pneumonia in developmentally disabled individuals can be caused by: (1) aspiration, (2) ineffective airway clearance secondary to accessory muscle malfunction, or (3) immune deficiency secondary to drug toxicity. Aspiration as cause of pneumonia, while perhaps the most common etiology, may be over-diagnosed. If aspiration from oropharyngeal dysfunction is suspected, a swallowing study (preferably videofluoroscopic) should be performed. Feeding gastrostomy would be indicated if feeding becomes extremely difficult for the patient, if there is significant weight loss, or if clinical aspiration occurs. If recurrent pneumonia is caused by gastroesophageal reflux, as documented by esophagogastroduodenoscopy (EGD), then medical (prokinetic agents, acid blockers) and/or surgical (fundoplication) treatments would be indicated.

6.Gastointestinal Disorders:

a.Recurrent gastrointestinal bleeding: the most common causes of gastrointestinal bleeding in the developmentally disabled population are: (1) esophagitis and (2) gastritis. Esophagitis is secondary to gastroesophageal reflux which in turn is secondary to either gastroesophageal dyskinesia (dyskinetic GERD), or rumination (self-induced GERD). Barrett's esophagus (a premalignant lesion) can complicate long standing esophagitis. If Barrett's esophagus is demonstrated, then careful surveillance is indicated. "Dyskinetic" GERD can be treated with prokinetic agents, acid blockers, or fundoplication. "Self-induced" GERD (rumination) can be treated by behavior modification and medically as well (especially acid blockers). Efficacy of treatment and surveillance of Barrett's esophagus should be documented by yearly endoscopy. Gastritis can be caused by "duodeno-gastric reflux" (DGR), i.e. reflux of bile salts and pancreatic enzymes into the stomach, or by Helicobacter pylori infection. DGR can be diagnosed by isotopic gastric emptying studies, while H. pylori is diagnosed by endoscopic biopsy with CLO test and histopathological examination. DGR can be treated with prokinetic agents (and possibly cholestyramine) while H. pylori is treated with antibiotics. Efficacy of treatment should be documented by repeat endoscopy, since H. pylori infection may recur.

b. GI Motility Disorders: Chronic brain injury is often associated with abnormal motility of oropharynx, esophagus, stomach, and intestines. Oropharyngeal dysfunction and esophageal spasm may lead to chronic pulmonary aspiration as previously discussed . Gastroesophageal reflux and duodeno-gastric reflux (DGR) are associated with G.I. bleeding (as discussed above) and also with pulmonary aspiration. Chronic constipation can be caused by: (1) Anticholinergic medication (e.g. neuroleptics), (2) immobility (e.g. from contractures), and (3) the underlying brain disorder. Constipation can lead to fecal impaction and volvulous and may also indirectly contribute to an increased risk of pulmonary aspiration. The first steps in treatment of constipation are directed towards improving "mobility" by treating somatomotor disorders and promoting exercise, avoidance of anticholinergic drugs if possible, and addition of fiber and fluid to the diet. If these measures are tried and are ineffective, the addition of oral lactulose (or Sorbitol), followed by laxatives, and/or enemas would be indicated. The prokinetic agent Cisapride (Propulsid) may also be effective in refractory cases.

7. Osteoporosis: Impaired mobility, phenytoin, amenorrhea in women, and hypogonadism in men are risk factors for osteoporosis. Individuals who possess these risk factors should have baseline bone density studies and/or urine pyridinoline cross-links determination. Regardless of treatment chosen (eg. Didronel, Fosamax, Prempro, Calcimar) efficacy of treatment should be documented by yearly urine cross-links assessment and bone densitometric studies every three years. Calcium and vitamin D supplements alone are probably inadequate treatment for the osteoporosis encountered in patients with developmental disabilities.

8. Hepatitis B Carrier State: The Hepatitis B carrier state is common in developmentally disabled individuals, especially if there has been a history of institutionalization or if the individual has Down Syndrome. Risk of infectivity and presence of hepatitis should be ascertained by determination of Hepatitis B viral DNA, e antigen, and hepatic enzymes in all cases of Hepatitis B surface antigen carrier state. Treatment (eg. with Intron A and/or Epivir) should be considered in those patients who demonstrate active viral proliferation and/or hepatitis. Those patients who continue to be "active" (positive viral DNA, e antigen, elevated SGPT) should undergo yearly abdominal sonograms and/or alpha-fetoprotein to rule out hepatic carcinoma.

(*) "Longitudinal Graphic/video analysis" refers to an objective and systematic graphic and/or video correlation of a targeted behavior or symptom with a "treatment" intervention, which is designed to determine "efficacy" of the intervention .