Genetically modified mice and their importance in psychopharmacology

Genetically modified mice and their importance in psychopharmacology

Just as adding genes from a complex to a simpler organism (for example,
from man to a fruit fly) may be helpful in understanding the function of a
gene, so it may help to understand how a gene functions by eliminating it.
To date, most gene ‘‘knock-out’’ studies have been undertaken in mice
because of:
(a) the relative ease with which genes can be manipulated and eliminated;
(b) the relatively rapid rate at which mice breed;
(c) their well established and relatively complex behaviour.
The success, and also the limitations of the gene elimination strategy can be
illustrated by studies on the molecular basis of memory and learning. In the
early 1980s it had been shown that the glutamate NMDA receptor was an
essential component of memory formation, the term ‘‘long-term potentiation’’
(LTP) being applied to the molecular mechanism involved. The drugs
which were then available were limited in their specificity for the NMDA
receptor but by selectively deleting genes thought to be involved in memory
it was possible to identify the precise components of the NMDA-linked
messenger complex located in the hippocampus. Further studies enabled
genes ranging from those encoding neurotransmitter receptors, protein kinases and transcription factors to be identified. However, there are
limitations to these techniques which should be considered.
A major problem with ‘‘knock-out’’ technology relates to the need to
delete the gene at the very early stage of embryonic development. Often this
results in the death of the neonate. Even if the gene is not essential for
survival, it could have a key role to play in development that is unrelated to
neuronal plasticity. Thus the deficits in learning and memory seen in the
mature mouse could be the result of a developmental defect rather than a
specific abnormality in the NMDA receptor complex. Alternatively, the
deletion of a gene that from experimental studies might be expected to have
a major effect on learning and memory in practice may have no apparent
effect. This is due to the mechanism of compensation whereby other genes
take over the function of the deleted gene.
Thus developing ‘‘knock-out’’ mice to understand the function of a
particular gene gives little information on the timing when the gene
becomes active. Nor does it necessarily reflect the location of the gene in the
intact (wild-type) mouse or indeed, the long-term effect of the nervous
system on its function. Nevertheless, these are largely technical drawbacks
that will undoubtedly shortly be solved. In principle, studying the actions
of psychotropic drugs on genetically modified animals will allow the
detrimental effects of a deleted gene on the general health of the animal to
be avoided. Such an approach will also allow investigations of the
interactions between neuronal signalling pathways by assessing the
synergistic interactions between the behavioural and other biological effects
of the deleted gene and drugs.

The impact of molecular neurobiology on psychopharmacology: from genes to drugs

The impact of molecular neurobiology on psychopharmacology: from genes to drugs

About 150 years ago, Charles Darwin observed that ‘‘those who make many
species are the ‘splitters’, and those who make few are the ‘lumpers’’’.
Today, the ‘‘splitters’’ dominate research in the life sciences. Such
researchers can generate massive quantities of data on genes and their
networks, proteins and their pathways and the numerous cascades of
messenger molecules that ultimately result in a physiological response.
Technological progress in recent years has enabled the genome of species as
diverse as the nematode worm Caenorhabditis elegans and the fruit fly
Drosophila melanogaster to the mouse and man to be unravelled, thereby
opening up the possibility not only of identifying genes that are responsible
for physiological processes but also those that are aberrant and cause
genetically based diseases.
Few would deny the importance of such research, but the very success
of the ‘‘splitters’’ has had a seriously detrimental effect on the equally
important role of the ‘‘lumpers’’, who attempt to integrate the
molecular/cellular approach with the behavioural/psychological consequences.
As a consequence, the ‘‘lumpers’’ are becoming a threatened
species of researchers. There are several reasons for this, not the least of
which is the widespread opposition to vivisection and the lack of
training in behavioural pharmacology in university courses. As a
consequence, research (and funding for behavioural research) has
declined in prestige. This has had an adverse impact not only in areas
of basic life science research but also in the pharmaceutical industry
where the ultimate validation of the therapeutic potential of a new
molecule depends on behavioural pharmacology. As a senior neuropharmacologist
has recently remarked ‘‘Many can genotype but few can
phenotype’’.
Despite this unfortunate disparity between molecular neurobiology and
behavioural pharmacology, it is essential that the neuropharmacologist and
biological psychiatrist are fully conversant with the basic concepts of the
subject in order to appreciate both its success and limitations.
To understand the basis of cloning, it isnecessary to consider how bacteria have evolved to resist infection by
external sources of genetic material. It has long been recognized that if a
virus could infect one strain of bacteria, it could then also infect other
bacteria of the same strain but not those of a different strain. Thus virus
infection was shown to be restricted to a particular strain, a restriction now
known to be due to two classes of enzyme, namely the methylases, which
modify bacterial DNA marking them as ‘‘self’’, and the destruction
enzymes, which act as molecular ‘‘scissors’’ and can destroy foreign DNA.
Restriction enzymes are sequence-specific in that they cut DNA at specific
locations along the nucleotide chain. While some of these enzymes yield
‘‘blunt’’ ends to the resulting DNA fragment, others make staggered cuts in
the DNA chain to produce ‘‘sticky’’ ends. Over 250 restriction enzymes are
now commercially available.
Cloning would not be possible without restriction enzymes. DNA chains
with a ‘‘sticky’’ end act like molecular ‘‘Velcro’’, thereby enabling two
pieces of DNA with complementary nucleotide sequences to be joined
together. The linking of the DNA strands is brought about by the enzyme
DNAligase which permanently joins the assembled DNA sequences with
covalent bonds, thereby producing a recombinant DNA molecule.
The next stage is to ensure that the recombinant DNA molecule is
copied by the enzymes which synthesize nucleic acids. These DNA and
RNA polymerases synthesize an exact copy of either DNA or RNA from
a pre-existing molecule. In this way the DNA polymerase duplicates the
chromosome before each cell division such that each daughter cell will
have a complete set of genetic instructions which are then passed to the
newly formed RNA by RNA polymerase. While both DNA and RNA
polymerase require a preformed DNA template, some viruses (such as
HIV) have an RNA genome. To duplicate that genome, and incorporate
it into a bacterial or mammalian cell, the viruses encode a reverse
transcriptase enzyme which produces a DNA copy from an RNA
template.
Thermostable DNA polymerases have now been produced for
polymerase chain reaction (PCR) studies in which specific segments of
the DNA molecule can be mass produced from minute quantities of
material. RNA polymerases are then used to create RNA transcripts from
cloned genes in vitro. Reverse transcriptases have their specific uses in
molecular biology. These enzymes are used to form ‘‘cDNA libraries’’
which are batteries of molecules each one representing a single gene
expression. Such DNA libraries can then be analysed to determine which
genes are active under different conditions and in different tissues. cDNA
libraries are now used experimentally in microarray assemblies to detect
gene changes following drug treatment.
In a typical experimental situation, the gene of interest is incorporated
into a plasmid, which is a natural vector used by either a bacterium or other
cell type. To transfer the DNA fragment of a gene, the plasmids are digested
with one or two restriction enzymes and the desired fragment joined into a
single DNA recombinant molecule using DNA ligase. To express the new
gene in vitro, the plasmid containing the recombinant DNA is then
incubated with an RNA polymerase to form new RNA which is then
used to programme an in vitro system which translates the information
necessary for the synthesis of a new protein.
The foregoing is only intended to give a brief overview of the
mechanisms behind cloning. So far, the impact on diseases in man has
been limited to experimental approaches to the treatment of cystic
fibrosis and rare conditions in which a recessive gene is responsible.
However, cloning techniques have provided important information in
producing animals, usually mice, which have been manipulated to
express or remove genes that are implicated in psychiatric disorders.
Such ‘‘knock-out’’ and ‘‘knock-in’’ mice now provide important
information in which specific genes can be studied for their effects
on behaviour, which may ultimately be an important contribution to
understanding the genetic basis of psychiatric and neurological
diseases.

Genetics of attention deficit/hyperactivity disorder (ADHD)

Genetics of attention deficit/hyperactivity disorder (ADHD) ADHD is the most common psychiatric disorder with onset in childhood. The condition is characterized by inattention and/or hyperactivity and impulsivity which is associated with cognitive, social and academic impairments. It has been estimated that in up to 60% of patients, these impairments persist into adulthood. Males are affected more than females (4:1). ADHD is a familial disorder with a population-based prevalence of about 10% and a prevalence rate in siblings of approximately 25%. Detailed studies of the disorder suggest that there is a five- to sixfold increase in firstdegree relatives of affected persons. However, as with other psychiatric disorders, the finding of familial aggregation alone does not necessarily lead to the conclusion that the disorder is of genetic origin as such studies do not separate genetic from environmental factors. Twin studies have been useful in discriminating the genetic from environmental factors. The concordance rate for monozygotic versus dizygotic twins has been estimated at 51% and 33% respectively, and it has been estimated that approximately half the variance in the trait factors of hyperactivity and inattentiveness are accounted for by the genetic basis of the disorder. There is evidence that, in ADHD, there is an incompletely penetrant autosomal-dominant gene; the penetrance of the gene being calculated as 46% in boys and 31% in girls. Molecular genetic studies have been particularly fruitful in evaluating the neurochemical basis of the disorder. Genes involved in the dopaminergic system were considered to be important as the most effective symptomatic treatment of the condition has been methylphenidate and dextroamphetamine, drugs which potentiate the release, and inhibit the reuptake, of dopamine.Of the genes for the dopamine receptors which have been studied, the candidate gene for the dopamine D4 receptor has been shown to be positively associated with ADHD. However, not all investigators have verified this finding. One of the major limitations in studies of the genetics of behavioural disorders in children arises from the overlap with other conditions. For example, nearly 50% of the patients with ADHD also have co-morbid conduct disorders. In addition, a subtype of the disorder may exist in those children in which the disorder persists into adulthood. An additional problem arises from the overlap between ADHD and bipolar disorder; this has been estimated to be as high as 16%. In CONCLUSION, although positive genetic studies have been reported and subsequently replicated, the results must be treated with caution as they are based on small sample sizes with restricted statistical power andcomplicated by co-morbid illnesses. Nevertheless, preliminary evidence suggests that ADHD, like many major psychiatric disorders, does have a genetic basis.

Genetics of panic disorder

Genetics of panic disorder Studies assessing the diagnosis of panic disorder by the direct interview of family members, a method commonly employed in the 1980s, showed that the risk of developing the disorder is higher in the first-degree probands. This has been estimated to be 17% for first-degree relatives compared to 2% for the control population. In addition, relatives of probands had a higher prevalence of generalized anxiety disorder and major depression. This was confirmed in another study of anxiety disorders in female relatives and showed a morbidity risk for all anxiety disorders to be 32% among firstdegree relatives of agoraphobic patients with panic attacks and 33% in those with panic disorder. The risk was found to be even greater in the offspring when both had anxiety disorders. Overall, family studies have shown that first-degree relatives of probands with panic disorder have a three- to 21- fold higher risk of developing panic disorder than first-degree relatives of healthy probands. The relationships between panic disorder and other types of anxiety disorder have also been the subject of study. Thus some investigators have reported that relatives of those with agoraphobia have a higher risk for the disorder than relatives of patients with panic disorder, leading to the suggestion that agoraphobia should be considered a more severe form of panic disorder which has an independent genetic transmission. Other studies, however, have found that the diagnosis of separation anxiety and agoraphobia in probands increased the risk of both panic disorder and agoraphobia in the relatives of the patients. Such observations support the hypothesis that panic disorder and agoraphobia are two phenotypic expressions of the same condition due to a different degree of genetic penetrance. The analysis of the relationship between panic disorder and major depression has produced conflicting results. The possible link between these disorders has been provided by the frequent occurrence of major depression in patients with panic disorder and agoraphobia, bothconditions responding to antidepressant treatments. Whereas most of the genetic analyses available suggest that there is an independent genetic basis for these conditions, it is noteworthy that relatives of patients with both panic disorder and agoraphobia are more likely to develop depression, phobias and alcoholism when compared with relatives of probands with panic disorder alone. There does not appear to be an association between generalized anxiety disorder and panic disorder, suggesting that they are separate entities. Generalized anxiety disorder appears to be more prevalent (approximately 20%) in relatives of probands with the disorder than in those of relatives with panic disorder (5%) or agoraphobia (4%). Twin studies have demonstrated only a moderate concordance in monozygotic versus dizygotic pairs. Using the hypersensitivity to carbon dioxide as a challenge test, it has been shown that there is a significantly higher concordance rate in monozygotic (56%) than in dizygotic (13%) twins. In recent years, emphasis has tended to switch from family and twin studies to molecular genetics. Linkage analysis in families with a psychiatric disorder has often proven to be a fruitful method to determine the degree of inheritance of a disorder. The method of parametric linkage estimates the likelihood of the distribution of genetic markers of an illness by a predetermined model and expressed as the logarithm of the odds score (the so-called lod score). For simple Mendelian inheritance of a disease, a lod score of 3.0 or greater, obtained by scanning the genome for markers of the disease, is considered to be a statistically significant linkage. However, most psychiatric disorders do not follow classical Mendelian genetics so such an approach is of limited value. For this reason, other linkage methods have been developed. For example, the allelic-sharing method is based on detecting the frequency of the inheritance of the same genetic marker from each parent. The presence of a gene that causes the disorder is revealed when the allele that is shared between the siblings is greater than 50%. To date, all the linkage studies in panic disorder have been inconclusive. Neither has the search for candidate genes for neurotransmitters (for example, GABA, tyrosine hydroxylase, serotonin receptors, dopamine receptors, adrenoceptors, opioid receptors) proven to be any more fruitful. However, there is some preliminary evidence for an association between the cholecytokinin (CCK) promoter gene and panic disorder which, if replicated, would support the hypothesis that the CCK-B receptor is hypersensitive in panic disorder.

The genetic basis of obsessive–compulsive disorder (OCD)

The genetic basis of obsessive–compulsive disorder (OCD) There have been over 18 family studies of OCD during the past 70 years but, due to methodological differences, the familial aspects of the disorder remain controversial. Family studies look for the prevalence of the disorder among the biological relatives of the probands (i.e. individuals who are affected by OCD) and the prevalence is then compared with that seen in the general population or in a control group. The latter are usually unaffected subjects or relatives of those with OCD. Despite the limitations of the studies, most have found a significant increase in the rates of OCD among the first-degree relatives of the probands when compared with the general population. Similarly, there is evidence from family studies of patients with chronic motor tics and Tourette syndrome that the rates of these conditions, and OCD, are higher among the relatives of patients with Tourette syndrome. More recent studies have suggested that some of these cases are familial but unrelated to tics while in other cases there appears to be no family history of either OCD or tics. Twin studies comparing mono- and dizygotic twins have shown concordance rates of between 25 and 87%, depending on the study. Since none of the studies show a concordance rate of 100%, it is clear that nongenetic factors must influence the expression of OCD. Linkage analysis studies, in which the OCD is linked to a known polymorphic marker, are used to determine if there is a single gene locus causing susceptibility for the disorder. Using such an approach for a battery of candidate genes coding for receptors and enzymes involved in dopaminergic and serotonergic transmission (i.e. for the two neurotransmitters most likely to be involved in OCD), no evidence was found for a link. Association studies have proven to be more fruitful. In such studies, the allelic frequencies for specific marker genes are compared with a control population. When OCD patients were investigated for the association of tics with the dopamine receptor 2 marker it was shown that there was an increased frequency of homozygosity for the allele A2 of TaqIA at the D2 receptor locus. Not all investigators could replicate this finding however. No positive associations have been found between OCD and the D3 gene while there was some association found between OCD with tics and the D4

gene. Two other catecholaminergic markers have been investigated in
patients with OCD. Thus a positive association has been reported between
the low activity allele of catechol-O-methyl transferase in male patients and
a higher frequency of the low activity allele of monoamine oxidase type A
gene in female patients. There is also evidence that the serotonin transporter
gene is abnormal. In conclusion, evidence from genetic studies lends
support to the view that a single gene defect occurs in patients with OCD
while association studies of candidate genes, in spite of methodological
difficulties, have highlighted the loci for D2 and D4 receptor genes together
with those for catechol-O-methyl-transferase and monoamine oxidase A.

Summary of methods used to detect possible genetic defects in psychiatric disorders

Summary of methods used to detect possible genetic defects in psychiatric disorders The genetic basis of late onset Alzheimer’s disease conforms to the common disease–common variant hypothesis which states that genetic susceptibility is attributable to common variants present in the population at a high frequency. Significant associations have been demonstrated between several common polymorphisms such as APO E4 and Alzheimer’s disease, already referred to. Individuals differ from each other at many positions across the genome and a variation at a particular nucleotide is called a polymorphism. It has been calculated that approximately one nucleotide base in every 1000–2000 nucleotides differs between chromosomes. If, as seems likely, the common disease–common variant hypothesis applies to schizophrenia and bipolar disorder then it may be essential to undertake association studies which follow the inheritance of a gene within astudies are an attractive means to understanding complex psychiatric diseases because they have the power to identify causal associations between a particular allele and a heterogeneous disease and do not require the collection of large pedigrees containing multiple affected individuals. However, for such a method to be applied, the polymorphisms used in the assessments must have been identified. This necessitates the collection of a large group of polymorphisms to form a library of potential disease-causing alleles. Traditionally techniques for the detection of polymorphisms have involved gel-based screening methods or direct DNA sequencing of numerous individuals. Such methods are laborious and time-consuming. Microarray technology has revolutionized the approach by developing DNA microchips that contain a high density of oligonucleotides which are capable of rapidly detecting variations in nucleotide sequences. Microarrays consist of microchips of DNA attached to the surface of a solid support that may vary according to the density and the form, or size, of the DNA on the surface. There are different commercially available forms of microarray chips and the presence of many independent DNA molecules on the chip surface allows hybridization of many different species simultaneously and for the results to be similarly detected on a microscope slide. By labelling the final products formed with a fluorescent dye, the resulting hybridization pattern can be detected by a confocal microscope. The major question which arises now that the technology for rapid screening has become available is which genes should be chosen? If the possible cause of the disease is known then the answer is clear. However, for most of the major psychiatric disorders the causes are unknown although neurochemical, neurodevelopmental, autoimmune and environmental factors are probably involved. For this reason, association studies in schizophrenia and bipolar disorder have focused on genes related to the dopaminergic and serotonergic systems. Unfortunately, polymorphisms related to these genes have not convincingly been found to be associated with schizophrenia or bipolar disorder. However, several other lines of evidence suggest that schizophrenia may be a neurodevelopmental disorder in which the frontal association cortex, an area responsible for several important hig-level functions, has been shown to be malfunctional. Thus schizophrenia may result from a disorder of neuronal migration. If so, the discovery of polymorphisms in genes that regulate cortical development could prove to be a useful area of investigation. This approach has already identified the gene dsh (dishevelled, named after a gene identified in the fruit fly, Drosophila) in the mouse which, when absent, leads to a reduction in the startle response. Prepulse inhibition, the reduction in the startle that the animal shows to a previous stimulus, has been shown to be diminished in patients with schizophrenia. While it is premature to propose that dsh deficient mice are a model for schizophrenia, such findings do suggest thatsuch genes may play an important role in the early differentiation of the brain. During the normal development of the brain, a large number of genes are activated and become involved at different times and have different functions. In the early pattern formation of the brain, during the development of the neural tube, the homeotic genes (the so-called hox family genes) encode for transcriptional factors that bind to DNA and thereby regulate the expression of other genes. This process is believed to convey information on dorsoventral positioning to the cells of the developing neural tube. However, there is evidence that a mutation in a homeobox gene is associated with gross brain defects which makes it unlikely that a mutation in the early genes could be responsible for the subtle changes found in schizophrenia. There is evidence that the cortex of the brain of schizophrenic patients contains neurons that are in abnormal positions when compared to nonschizophrenic individuals. This suggests that the migration of neurons may be abnormal during the developmental period. The neuronal cell adhesion molecule (NCAM) is an immunoglobulin that mediates adhesion between neurons, thereby exerting a key role in morphogenesis, differentiation and the migration of neurons. NCAM is encoded by a single gene that undergoes alternative splicing to generate several alleles which exhibit different spatial and temporal patterns of expression in vertebrates. The NCAM gene is located on chromosome 11 and two polymorphisms are known to occur in the NCAM gene region. However linkage analysis of 71 families has failed to confirm that an abnormality in the NCAM gene occurs in schizophrenia. During brain development, neurons send out numerous dendrites in which growth factors play a prominent part. The neurotrophins comprise a family of structurally and functionally related growth factors that include nerve growth factor (NGF), brain derived neurotrophic factor (BDNF) and the neurotrophins 3 and 4/5. These peptides cause neuronal growth, increase the size of the body of the neuron and maintain the survival of the neurons of the dopaminergic, serotonergic and glutaminergic systems. As a decrease of 20–30% in BDNF mRNA has been reported to occur in the hippocampi of schizophrenic patients, it seems possible that a defect in this growth factor could be responsible for the occurrence of smaller neurons in these patients. The BDNF gene is located on chromosome 11 and preliminary findings from 72 nuclear families suggest that the frequency of the A2 allele is significantly higher, and that of the A1 allele significantly lower, in schizophrenic patients. The NT-3 gene, located on chromosome 12, and the promoter region of this gene contain a highly polymorphic marker yielding at least 11 dinucleotide repeat alleles. Of the casecontrolled association studies undertaken so far, abnormal alleles have been reported in two of the five studies.Lastly, studies on the different polymorphic forms of the synapsins, that organize the mobilization of neurotransmitter vesicles thereby regulating neurotransmitter release, could account for some of the subtle changes in neurotransmission that occur in schizophrenia. However, to date linkage analysis studies have failed to reveal any positive associations between the various polymorphisms of the synapsin gene and schizophrenia. population rather than within families. Association studies test if a polymorphism is more frequently found in those with the disease (called ‘‘cases’’) than in normal individuals (controls). In such tests, the transmission of a polymorphism from a heterozygous parent to an affected offspring is followed. If the polymorphisms are not associated with the disease, then the rate of transmission from parent to affected offspring in a population will be 50%. Significant deviations from this predicted transmission rate indicate a possible association with the disease.

Pharmacogenetics and psychopharmacology

Pharmacogenetics and psychopharmacology For more than 40 years, epidemiological studies have clearly demonstrated a tendency for diseases such as schizophrenia, bipolar disorder and autismto run in families. Thus it has been shown that such disorders are much more frequent in close relatives of patients than in the general population. For example, estimates of the increased risk of suffering from the disorder if the patient has a sibling with the disorder range from nine- to eleven-fold for schizophrenia and about sevenfold for those with bipolar disorder. These major psychiatric disorders show a significantly greater concordance rate in genetically identical twins. Thus the concordance rate for monozygotic twins in schizophrenia is approximately three times that observed in dizygotic twins. In bipolar disorder, the corresponding concordance rate is approximately eight times greater in monozygotic than in dizygotic twins. From such studies it has been calculated that between 60 and 80% of the liability of these two disorders is genetic in origin. However, it must be emphasized that these calculations do not identify specific genetic causes for the conditions but they do demonstrate that the genetic, as well as environmental components, play a significant role. The question arises regarding how the genes that contribute to major psychiatric disorders can be identified. Many of the medical conditions for which a genetic component has been identified follow a genetic pattern that clearly follows classical Mendelian inheritance. For example, in cases of Huntington’s disease and cystic fibrosis, the two parents who carry the recessive gene give rise to offspring in the ratio of 1:2:1, one expressing the disease, two not showing the symptoms but carrying the gene and one not carrying the gene for the disorder. Such conditions follow the Mendelian pattern of inheritance because the condition is caused by a mutation of a single gene. While the locating of the underlying gene in such situations is often difficult and time-consuming, the techniques of classical molecular genetic analysis, and linkage studies followed by positional cloning, are now well established. For monogenic diseases such as Huntington’s, a common pattern emerges in which a single gene, or small number of genes, that may harbour a number of rare mutations can be identified. Each mutation alone is then sufficient to produce a phenotype of the disease. Regarding bipolar disorder and schizophrenia, while there is some evidence that some families transmit the risk of the disease in a Mendelian fashion, the overall pattern of disease transmission is complex and it is unlikely that these conditions are due to a single gene. This suggests that there may be multiple genes involved, either many genes with strong alleles or common variants in many genes, each of which increases the risk of the disease in a modest way. An example of this would be Alzheimer’s disease, in which significant associations have been demonstrated between apolipoprotein (APO) E4 and the occurrence of the disease. An account of the genetic basis of Alzheimer’s disease can provide a useful example of the relationship between the genetic basis and the expression of the disease.Alzheimer’s disease exists in two major forms, the so-called early and late onset types. The former follows typical Mendelian inheritance while the latter shows a more complex, non-Mendelian, pattern of inheritance. The early onset form of the disease has permitted the identification of several genes which are causally related to the condition. In the elderly, in which Alzheimer’s disease has been estimated to occur in up to 20% of those aged 80 years, it has been shown that one allelic form of APO E is associated with an increased risk for developing the disease. Of the three APO E allelic forms in man, APO E4 is associated with the late onset form of the disease; this may account for up to 50% of the genetic risk for the late onset form whereas those carrying the less frequent E2 allele appear to be protected from the disease. In the early onset familial form of the disease, affecting approximately 5% of cases, there is a clear autosomal dominant pattern of inheritance. Mutations in three genes have been identified involving the beta amyloid precursor protein, presenilin-1 and presenilin-2. The function of these proteins is described in more detail in the chapter on the dementias . It has been estimated that mutations in these genes account for approximately 50% of the cases of the early onset disease.