Genetic Prion Diseases

[Transmissible Spongiform Encephalopathies (TSEs). Includes: Familial Creutzfeldt-Jakob Disease (fCJD), Fatal Familial Insomnia, Gerstmann-Sträussler-Scheinker Disease]

Clinical Diagnosis

Genetic prion diseases constitute a continuum of clinical manifestations, originally labeled as familial Creutzfeldt-Jakob disease (fCJD), Gerstmann-Sträussler-Scheinker (GSS) syndrome, and fatal familial insomnia (FFI). Note: A fourth clinical phenotype, known as Huntington disease like-1 (HDL-1) has been proposed, but this is based on a limited number of cases, and the underlying pathologic features would categorize it as a GSS [Moore et al 2001]. It is now known that these three phenotypes are not distinct entities but rather constitute a spectrum of clinical and pathologic manifestations of genetic prion disease; nonetheless, certain aspects of these phenotypes are useful in diagnosis and care. The diagnosis of genetic prion disease requires a combination of the following:

• Clinical features comprising varying combinations of adult-onset neurologic signs and symptoms, including: dementia, psychiatric symptoms, dyscoordination of movements (ataxia, dysarthria), myoclonus (muscle jerks), weakness and/or spasticity, chorea, stroke-like episodes, seizures, and autonomic disturbances

• Neuropathologic findings that include spongiform degeneration and astrogliosis diffusely distributed throughout the cortex and deep nuclei of the brain (fCJD); multiple amyloid plaques to which anti-prion protein (PrP) antibodies bind (GSS); a relative lack of spongiform degeneration and presence of neuronal dropout and gliosis primarily within the thalamus and inferior olivary nucleus of the brain stem (FFI) [DeArmond & Prusiner 1997]

• Family history consistent with autosomal dominant inheritance

• A PRNP disease-causing mutation (see Molecular Genetic Testing)

Other studies including electroencephalogram (EEG), brain imaging, or examination of cerebrospinal fluid (CSF) may be helpful in supporting the diagnosis, but none is diagnostic on its own. Often such studies are performed to evaluate for other potentially treatable diseases of the central nervous system (see Differential Diagnosis.) It should be emphasized that these tests have been best studied and are most helpful in the diagnosis of non-genetic prion disease (i.e., sporadic CJD). Reliance, therefore, on these studies for the diagnosis of genetic prion disease is cautioned.

• EEG. Characteristic EEG findings of periodic sharp wave complexes (PSWCs), consisting of triphasic or sharp wave bursts every 0.5 to 2.0 seconds, can suggest the diagnosis of prion disease. Although PSWCs are observed in a relatively small percentage of individuals with genetic prion disease, their presence appears to be highly dependent on the associated causal mutation and resultant clinical phenotype; those mutations that produce a CJD-like clinical phenotype and spongiform degeneration pathology appear more likely to have a positive EEG.
  
  Note: Initially, the PSWCs may be unilateral, but with disease progression, they typically spread to both brain hemispheres. In late stages of the disease, the periodic activity may disappear.

• Brain imaging

  • Magnetic resonance imaging (MRI) may show mild to moderate generalized atrophy at the time of presentation or within a short interval after presentation. T2-weighted images may demonstrate hyperintensity of the basal ganglia.

  • Diffusion-weighted MRI (DWI) appears to be more sensitive and specific in this regard. In some reports, signal hyperintensity appears within the cortical ribbon before it appears in the deeper structures in sCJD. A small number of reports suggest similar findings in genetic prion disease, especially those with CJD-like phenotypes.

  • PET or SPECT scanning, with the exception of FFI, appears to be of limited usefulness in diagnosing genetic prion disease, as the findings show nonspecific and diffuse cortical hypometabolic activity, sometimes with frontal predominance.

Note: (1) In some reports, a reduction in perfusion to specific brain regions correlates with the clinical symptoms observed in the individual (e.g., left frontal or occipital cortex affected in language or visual deficits, respectively). (2) In the FFI phenotype the PET scan demonstrates a significant and selective reduction in activity within the thalamus, often early in the disease.

• CSF (cerebrospinal fluid). An elevation of CSF protein concentration by approximately 10% is common, and may be attributed at least in part to release of the normal neuronal 14-3-3 protein into the CSF following neuronal death; however, this finding is not specific for prion disease.
  
  Note: (1) Because a significant number of neurons die in individuals with prion disease, the concentration of the 14-3-3 protein in CSF may increase substantially in some but not all cases. (2) Because of the generally slower rate of progression of genetic prion disease, the detection of the 14-3-3 protein in the CSF of these individuals appears less consistent than with sporadic CJD, suggesting that the increased rate of neuronal death leads to the measurable increase in 14-3-3 protein. (3) The 14-3-3 protein is released into the CSF in herpes encephalitis and hypoxic brain damage resulting from stroke, Hashimoto's encephalopathy, Alzheimer disease, and, on occasion, multiple sclerosis [Zerr et al 1998, Satoh et al 1999, Huang et al 2003]. (4) Van Everbroeck et al [2005] reported a more specific antibody to the gamma isoform of 14-3-3 that is more reliable against false positive tests. (5) Recent studies comparing the level of 14-3-3 protein with the levels of neuron specific enolase (NSE) and total tau suggest that each are comparable in their elevations in non-genetic prion disease; however, data are not sufficient to draw conclusions about genetic prion disease [Sanchez-Juan et al 2006].

Molecular Genetic Testing

GeneReviews designates a molecular genetic test as clinically available only if the test is listed in the GeneTests Laboratory Directory by either a US CLIA-licensed laboratory or a non-US clinical laboratory. GeneTests does not verify laboratory-submitted information or warrant any aspect of a laboratory's licensure or performance. Clinicians must communicate directly with the laboratories to verify information.—ED.

Gene. PRNP is the only gene known to be associated with genetically transmissible human prion disease.

Clinical testing

• Sequence analysis. Sequence analysis of the coding and flanking exon/intron boundaries of the PRNP gene is available on a clinical basis.

Table 1 summarizes molecular genetic testing for this disorder.

Table 1. Molecular Genetic Testing Used in the Diagnosis of Genetic Prion Diseases

Gene Symbol	Test Method	Mutations Detected	Mutation Detection Frequency by Test Method	Test Availability
PRNP	Sequence analysis	Sequence variants	Unknown 1	Clinical
	Targeted mutation analysis	Duplication of one to nine additional octapeptide repeats (Pro-His-Gly-Gly-Gly-Trp-Gly-Gln) 2	Unknown

1. By definition, "genetically transmissible" prion disease requires the presence of a disease-causing PRNP mutation; however, it is possible that sequence analysis does not detect all disease-causing mutations. Thus, the absence of a disease-causing mutation does not rule out the diagnosis.

2. Normal alleles have five such octapeptide repeat units (see Molecular Genetics).

Interpretation of test results

• Because identification of a PRNP mutation is required to make the diagnosis of a genetically transmissible human prion disease, it is important to consider possible outcomes of sequence analysis.

• For issues to consider in interpretation of sequence analysis results, click here.

Testing Strategy

To establish the diagnosis of genetically transmissible prion disease in a proband, a PRNP mutation must be identified.

Predictive testing for at-risk asymptomatic adult family members requires prior identification of the disease-causing mutation in the family.

Genetically Related (Allelic) Disorders

No phenotypes other than the genetic prion diseases are known to be associated with mutations in PRNP.

A single case of a diffuse Lewy body disease phenotype associated with the PRNP mutation p.Met232Arg has been reported. Whether the mutation and the phenotype are causally related remains uncertain [Koide et al 2002], but the lack of follow up suggests that it is unlikely.

Natural History

Genetic prion diseases generally manifest with cognitive difficulties, ataxia, and myoclonus (abrupt jerking movements of muscle groups and/or entire limbs); however, the order and/or predominance of these features and associated neurologic and psychiatric findings vary with genetic prion disease subtype and/or PRNP mutation. The age at onset ranges from the third to ninth decade of life. Of note, a 13-year-old with a p.Pro105Thr mutation developed symptoms suggestive of prion disease; however, pathology has not been reported [Rogaeva et al 2006].

The course ranges from a few months to several years (typically five to seven years, but in rare cases more than ten years). Death often results from infection, either by pneumonia (typically from aspiration) or urosepsis.

The classic phenotypes associated with genetic prion disease (fCJD, GSS, and FFI), were defined by clinical and neuropathologic findings long before the molecular basis of this group of disorders was discovered. The recognition that some families with clinical features of Huntington disease had a PRNP gene mutation has expanded the clinical spectrum of prion disease. While it is recognized that these phenotypes are part of a continuum and have overlapping features, it can be helpful to think of genetic human prion disease at least in part in terms of these phenotypes when providing individuals and families with information about the expected clinical course.

Familial Creutzfeldt-Jakob disease (fCJD). Progressive confusion and memory impairment occur first, followed by ataxia and myoclonus. The disease typically manifests between ages 30 and 50 years, although a few individuals present before age 30 years or as late as the upper 80s. The course from onset to death ranges from a few months to five years. At the end stage of disease, the individual is generally bed-bound, mute, and immobile (akinetic), except for myoclonic jerks.

The cognitive impairment observed may initially be mild confusion or it may be specific for a particular cortical function, such as language or constructional abilities; however, the resultant picture is one of global dementia. As the disease progresses, neurobehavioral symptoms may vary considerably. Psychiatric features, including delusions and hallucinations, may also occur.

Ataxia may be either truncal or appendicular, manifesting either as an unsteady gait, clumsiness while carrying out commonly performed tasks (e.g., picking up the salt shaker while dining), or progressive dysarthria (slurred speech). As the ataxia progresses, the individual may fall repeatedly, necessitating the use of a wheelchair to prevent injury.

Myoclonus generally, but not always, occurs after cognitive impairment is evident. Myoclonus may begin focally in a single limb but eventually becomes generalized. "Startle myoclonus" may be elicited by simple acts such as clapping the hands or turning on the room lights. Even if warned of an impending noise, the individual cannot suppress the startle response.

Other neurologic signs and symptoms such as focal or generalized weakness, rigidity, bradykinesia, tremor, chorea, alien hand syndrome, stroke-like symptoms, visual disturbances, and seizures have been reported.

Gerstmann-Sträussler-Scheinker syndrome (GSS) typically begins in the fourth to sixth decade with the insidious onset of cerebellar dysfunction, manifest as unsteady gait and mild dysarthria. Cognitive dysfunction is generally not apparent early on; however, with progression, bradyphrenia (slowness of thought processing) may become evident. Pyramidal involvement with spasticity and/or extrapyramidal involvement with bradykinesia, increased muscle tone with or without cogwheeling, and masked facies are also common. Psychiatric or behavioral symptoms are atypical. The disease progresses at a relatively slow but relentless pace over the course of a few to seven or more years. Cerebellar dysfunction results in severe dysarthria, gait and appendicular ataxia, ocular dysmetria, and lack of coordination in swallowing. A decline in cognitive abilities, particularly of concentration and focus, becomes apparent with progression into the late stage of disease. In the terminal stage, the individual is bedridden from the disabling ataxia, unable to eat because of severe lack of coordination in swallowing, and unable to communicate because of the profound dysarthria; yet insight into his/her condition may remain. This pattern of progression relates to the cerebellar nature of this disease, with progression into the brain stem and eventually the cerebrum.

Fatal familial insomnia (FFI) typically presents in midlife (40s to 50s) with the insidious or subacute onset of insomnia, initially manifest as a mild, then more severe, reduction in overall sleep time. When sleep is achieved, vivid dreams are common. A disturbance in autonomic function then emerges; it may manifest as elevated blood pressure, episodic hyperventilation, excessive lacrimation, sexual and urinary tract dysfunction, and/or a change in basal body temperature. Signs of brain stem involvement including decreased ability to gaze upward, double vision, jerky eye pursuit movements, or dysarthric speech may also appear in some individuals. With continued progression over the next few months, individuals develop truncal and/or appendicular ataxia.

The speed of thought processing may be reduced, as is common in subcortical dementing states, and memory impairment may be variable; however, compared with other more prominent features of disease, cognitive capacity is relatively spared until late in the course. Advancing disease results in progressively greater loss of total sleep time, worsening ataxia, and more profound confusion, leading ultimately to an awake but stuporous state as death approaches. As with other forms of prion disease, debilitation leading to feeding difficulties and loss of airway protection is the most common immediate cause of death. The typical duration of disease is 12 to 16 months, with a range of a few months to five years.

Neuropathology. The various genetic prion disease syndromes have relatively characteristic neuropathologic changes including abundant deposition of amyloid plaques that are stained by PrP antibodies in GSS, focal thalamic neuronal loss and gliosis in FFI, and diffusely distributed spongiform change with neuronal destruction in fCJD. Note that HDL-1 associated pathology suggests a variable picture, with prion protein containing plaque deposits present in some and spongiform degeneration in others. This phenotype is not well characterized at this time.

Although characteristic of each syndrome, the neuropathology is not 100% specific and variations from case to case are apparent [Gambetti et al 1999, Kovacs et al 2002, Liberski et al 2005].

Genotype-Phenotype Correlations

Detailed phenotype-genotype correlations of the various genetic prion syndromes can be found in Mastrianni [1998], Gambetti et al [1999], and Kovacs et al [2002].

Familial CJD phenotype. Several PRNP point mutations cause the fCJD phenotype (p. Asp178Asn with normal variant p.Val129, and mutations p.Val180Ile, p.Thr183Ala, p.Glu200Lys, p.Arg208His, p.Val210Ile, p.Met232Arg). See Table 2.

GSS phenotype. Causal mutations may include p.Pro102Leu, p.Pro105Leu, p.Ala117Val, p.Tyr145X, p.Gln160X, p.Phe198Ser, and p.Gln217Arg. See Table 2.

FFI phenotype. Only one haplotype causes FFI (p.Asp178Asn + normal variant p.M129).

p.His187Arg. Early onset and long duration have been associated with the p.His187Arg mutation [Hall et al 2005].

Insertional mutations are associated with the fCJD phenotype and the GSS phenotype. These insertions all lie within an unstable region of PRNP that is rich in proline, glycine, and glutamine (see Molecular Genetics).Normal PRNP alleles have one nonapeptide followed by four octapeptide repeat sequences each of which comprises the following amino acids: Pro-(His/Gln)-Gly-Gly-Gly-(-/Trp)-Gly-Gln. Because the nucleotide sequence encoding the octapeptide may vary, the repeat is described typically as an octapeptide rather than as a 24-nucleotide repeat.

From one to nine additional octapeptide repeat segments have been detected in familial forms of genetic prion disease. A correlation between the number of extra octapeptide repeats and phenotype seems to exist:

• Two to seven additional repeats are typically associated with the fCJD pathologic phenotype, but clinical presentation varies remarkably.

• Eight or nine extra repeats are associated with the GSS pathologic phenotype.

Of note, some families with octapeptide repeat insertions show significant phenotypic variability among affected individuals [Mead et al 2006]. (See Genomic Databases).

Normal / disease-modifying variant at codon 129. The common normal variant at codon 129 of PRNP (c.385A>G) codes for either the amino acid methionine (Met; p.M129) or valine (Val; p.V129). Approximately 50% of the Caucasian population is homozygous at this variation for either Met or Val, although 80%-90% of individuals with sporadic CJD are homozygotes.

• In general, the onset of genetic prion disease is earlier and its course shorter in individuals homozygous for p.Met129 compared to either heterozygotes for p.Met129Val or homozygotes for p.Val129.

• For alleles with the p.Asp178Asn mutation, the presence of p.Met129 vs. p.Val129 modifies the phenotype of disease.

  • If Val is encoded, the phenotype is almost always typical fCJD.

  • If Met is encoded, the phenotype is almost always FFI.

  • Even in nonfamilial forms of CJD, the p.Met129Val variant appears to have an effect on disease phenotype such that individuals with Met homozygosity present most often with dementia and a more rapid disease course, whereas those with a Val on one or both alleles display ataxia at onset and a slower disease course [Parchi et al 1999b].

Penetrance

The p.Glu200Lys and p.Val210Ile mutations of PRNP are commonly associated with a variable, but generally age-dependent penetrance, such that the older the individual, the greater likelihood of his/her manifesting the disease. Thus, it is not uncommon to encounter a situation in which the parents and other relatives of an affected individual may be unaffected but have a PRNP mutation [Kovacs et al 2005].

Most other PRNP mutations demonstrate complete penetrance.

Anticipation

Genetic anticipation has not been documented.

Nomenclature

Spastic pseudosclerosis is an older term used to describe a more fulminant course of CJD, as it appeared to have features suggestive of multiple sclerosis, with spasticity of gait and limbs.

Heidenhain’s variant is a term used to describe a specific presentation of CJD that occurs in about 10% of non-genetic forms. It begins with visual symptoms that may manifest as visual loss, blurring, and visual distortions (e.g., bending in the walls), related to the early involvement of the occipital lobes of the brain.

Prevalence

Genetic prion diseases are rare. The general worldwide yearly incidence of genetic and non-genetic prion disease is approximately one case per million. Thus, in the US, approximately 300 new cases of prion disease are observed per year. The genetic forms of prion disease represent approximately 10% of the total number of cases of prion disease.

The most common disease-associated mutations of PRNP are p.Glu200Lys, the largest focus being present in the Middle East (Libyan Jews) and Eastern Europe (Slovakia), and p.Asp178Asn, which is found worldwide. Other mutations are relatively rare [Kovacs et al 2005].

In Italy nearly 18% of all cases of prion disease had a PRNP mutation, with p.Val210Ile and p.Glu200Lys being the most common mutations observed [Ladogana et al 2005].

Evaluations Following Initial Diagnosis

To establish the extent of disease in an individual diagnosed with genetic prion disease, physical examination, with focus on the neurologic features of cognition, motor function, and coordination is recommended.

Treatment of Manifestations

Therapy is aimed at controlling symptoms that may cause discomfort.

• If present, seizures may be treated with general antiepileptic drugs (AEDs) including diphenylhydantoin or carbamazepine.

• Myoclonus can sometimes be mitigated by clonazepam.

• Issues related to dysphagia are often difficult to resolve. Since the disease is terminal, families are often faced with the difficult decision of whether or not to place a permanent feeding tube. The timing of this decision differs depending on the type of prion disease.

Evaluation by a social worker is mandatory to assist the family in management planning, as many decisions are required during the course of disease and at the end of the disease process.

Surveillance

Affected individuals are examined at regular intervals for complications related to swallowing difficulties, infections, and other disease manifestations.

Testing of Relatives at Risk

See Genetic Counseling for issues related to testing of at-risk relatives for genetic counseling purposes.

Therapies Under Investigation

Government-sponsored clinical trials are currently underway in the US and the UK to test quinacrine, an antimalarial agent that showed promise in tissue culture, although results in rodents were less convincing. Trials are nearly at the mid-point of study but to date no progress reports are available.

Search ClinicalTrials.gov for access to information on clinical studies for a wide range of diseases and conditions.

Other

Individuals with genetic prion disease do NOT need to be quarantined. While all prion diseases are potentially transmissible through ingestion or injection of infectious tissue (neural), they are not contagious by typical means of close contact with affected individuals. It is advisable, however, that body fluids of symptomatic individuals be handled as biohazard waste.

Antiviral therapies have been tested and anecdotal reports do not support them to be efficacious.

In rodent studies, amphotericin B, pentosan polysulfate, and various other agents displayed promise when administered prior to infecting animals with prions; however, no successful clinical results have been reported.

Genetics clinics are a source of information for individuals and families regarding the natural history, treatment, mode of inheritance, and genetic risks to other family members as well as information about available consumer-oriented resources. See the GeneTests Clinic Directory.

Support groups have been established for individuals and families to provide information, support, and contact with other affected individuals. The Resources section may include disease-specific and/or umbrella support organizations.

Mode of Inheritance

Genetic human prion disease is inherited in an autosomal dominant manner.

Risk to Family Members

Parents of a proband

• Most individuals diagnosed with genetic human prion disease have an affected parent. However, a proband with a genetic human prion disease may have the disorder as the result of a de novo gene mutation.

• The proportion of cases caused by de novo gene mutations is unknown.

• In addition, families in which penetrance appears to be reduced have been observed; thus the parent with a disease-causing mutation is unaffected while the child is affected.

Sibs of a proband

• The risk to the sibs of the proband depends on the genetic status of the proband's parents.

• If a parent of the proband is affected or has a disease-causing PRNP mutation, the risk to each sib of inheriting the allele is 50%.

• If the parents are clinically unaffected and do not have a PRNP disease-causing mutation, the risk to the sibs of a proband appears to be low.

• If a PRNP mutation cannot be detected in the DNA of either parent, it is presumed that the proband has a de novo gene mutation and the risk to the sibs of the proband depends on the spontaneous mutation rate of PRNP and the probability of germline mosaicism. Although no instances of germline mosaicism have been reported, it remains a possibility.

Offspring of a proband. Each child of an individual with a disease-causing PRNP mutation has a 50% chance of inheriting the mutation.

Other family members. The risk to other family members depends on the status of the proband's parents. If a parent is affected or has a disease-causing PRNP mutation, his or her family members are at risk.

Related Genetic Counseling Issues

Molecular genetic testing of symptomatic individuals from families with no history of neurologic disease. Goldman et al [2004] stress the importance of pretest counseling for families of symptomatic individuals with no family history of neurologic disease to better prepare families and to allow them to make informed decisions about receiving genetic test results.

Testing of at-risk asymptomatic adults. Testing of at-risk asymptomatic adults for genetic human prion diseases is available using the same techniques described in Molecular Genetic Testing. Such testing is not useful in predicting age of onset, severity, type of symptoms, or rate of progression in asymptomatic individuals. When testing at-risk individuals for genetic prion diseases, an affected family member must be tested first to confirm the molecular diagnosis in the family.

Testing for the disease-causing mutation in the absence of definite symptoms of the disease is predictive testing. At-risk asymptomatic adult family members may seek testing in order to make personal decisions regarding reproduction, financial matters, and career planning. Others may have different motivations including simply the "need to know." Testing of asymptomatic at-risk adult family members usually involves pre-test interviews in which the motives for requesting the test, the individual's knowledge of genetic prion diseases, the possible impact of positive and negative test results, and neurologic status are assessed. Those seeking testing should be counseled about possible problems that they may encounter with regard to health, life, and disability insurance coverage, employment and educational discrimination, and changes in social and family interaction. Other issues to consider are implications for the at-risk status of other family members. Informed consent should be procured and records kept confidential. Individuals with a positive test result need arrangements for long-term follow-up and evaluations.

Testing of at-risk individuals during childhood. Consensus holds that individuals younger than age 18 years who are at risk for adult-onset disorders should not have testing in the absence of symptoms. The principal arguments against testing asymptomatic individuals during childhood are that it removes their choice to know or not know this information, it raises the possibility of stigmatization within the family and in other social settings, and it could have serious educational and career implications. In addition, no preventive treatment for prion diseases is available.

See also the National Society of Genetic Counselors resolution on genetic testing of children and the American Society of Human Genetics and American College of Medical Genetics points to consider: ethical, legal, and psychosocial implications of genetic testing in children and adolescents (Genetic Testing; pdf)

Considerations in families with an apparent de novo mutation. When neither parent of a proband with an autosomal dominant condition has the disease-causing mutation or clinical evidence of the disorder, it is likely that the proband has a de novo mutation. However, possible non-medical explanations including alternate paternity or undisclosed adoption could also be explored.

Family planning

• The optimal time for determination of genetic risk is before pregnancy. Similarly, decisions about testing to determine the genetic status of at-risk asymptomatic family members are best made before pregnancy.

• It is appropriate to offer genetic counseling (including discussion of potential risks to offspring and reproductive options) to young adults who are at risk.

DNA banking. DNA banking is the storage of DNA (typically extracted from white blood cells) for possible future use. Because it is likely that testing methodology and our understanding of genes, mutations, and diseases will improve in the future, consideration should be given to banking DNA, particularly when the sensitivity of currently available testing is less than 100%. See for a list of laboratories offering DNA banking.

Prenatal Testing

No laboratories offering molecular genetic testing for prenatal diagnosis of genetic prion diseases are listed in the GeneTests Laboratory Directory. However, prenatal testing may be available for families in which the disease-causing mutation has been identified. For laboratories offering custom prenatal testing, see .

Requests for prenatal testing for late-onset conditions such as genetic prion diseases are not common. Differences in perspective may exist among medical professionals and within families regarding the use of prenatal testing, particularly if the testing is being considered for the purpose of pregnancy termination. Although most centers would consider decisions about prenatal testing to be the choice of the parents, discussion of these issues is appropriate.

Preimplantation genetic diagnosis (PGD) may be available for families in which the disease-causing mutation has been identified [Meiner et al 2006]. For laboratories offering PGD, see .

Literature Cited

Published Statements and Policies Regarding Genetic Testing

Suggested Readings

Author Notes

University of Chicago Memory Center Web site: memorycenter.bsd.uchicago.edu

Revision History

• 18 December 2008 (me) Comprehensive update posted live

• 7 October 2005 (cd) Revision: targeted mutation analysis for common mutations no longer clinically available

• 16 May 2005 (me) Comprehensive update posted to live Web site

• 4 March 2004 (cd) Revisions: Testing

• 27 March 2003 (me) Review posted to live Web site

• 12 April 2002 (jm) Original submission