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JrMed Genet 1998;35:579-589 579 Syndrome of the month J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from

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Bert B A de Vries, Dicky J J Halley, Ben A Oostra, Martinus F Niermeijer

Abstract an X linked disorder. 30% of The Approximately fragile X syndrome is characterised the obligate carrier were mildly or by mental retardation, behavioural fea- moderately retarded and a large portion tures, and physical features, such as a long (±50%) had no cytogenetic fragile X expres- face with large protruding ears and sion. Also, mentally normal grandfathers link- macro-orchidism. In 1991, after identifi- ing two family branches with the fragile X syn- cation of the fragile X mental retardation drome were observed. These "normal (FMRI) , the cytogenetic marker (a transmitting males" without any clinical or fragile site at Xq27.3) became replaced by cytogenetic features could apparently transmit molecular diagnosis. The fragile X syn- the fragile X syndrome. drome was one of the first examples of a The gene involved in the fragile X syndrome, "novel" class of disorders caused by a tri- the Fragile X Mental Retardation (FMRl) nucleotide repeat expansion. In the nor- gene, was identified in 1991 .9-l' The gene mal population, the CGG repeat varies defect was the first expansion of a trinucleotide from six to 54 units. Affected subjects have repeat to be discovered and a whole class of expanded CGG repeats (>200) in the first disorders is now known to be associated with exon ofthe FMR1 gene (the full ). this type of mutation.'2 Phenotypically normal carriers of the fragile X syndrome have a repeat in the 43 to 200 range (the premutation). Clinical studies The cloning ofthe FAlR1 gene led to the The original "Martin-Bell family" was restud- ied in characterisation of its product 198113 and the typical cytogenetic and http://jmg.bmj.com/ FMRP, encouraged further clinical stud- clinical features were found. The "Martin-Bell ies, and opened up the possibility of more " was proposed as an eponym for the accurate family studies and fragile X phenotype of affected males. Additional clini- screening programmes. cal features include high arched palate (52%), (JMed Genet 1998;35:579-589) (36%), hyperextensible metacar- pophalangeal joints (67%), hand calluses Keywords: fragile X syndrome; FMR1 gene; mental (29%), double jointed thumbs (53%), retardation single on September 26, 2021 by guest. Protected copyright. palmar creases (25%), pes planus (71%), and heart murmur or click (18%) (figs 1 and 2, History table 1).8 '4 The facial features are often less In 1943, Martin and Bell' described sex linked noticeable, particularly in affected females and mental retardation without dysmorphic fea- children (fig 3). The macro-orchidism often tures in a family in which both affected males develops during or after puberty and is and females were observed. Thirty-six years frequently absent in young patients.7 8 later, Lubs2 a reported marker X are observed in approximately 20% of young (later to be known as the fragile X chromo- affected males, with a lower prevalence in some) as an inconsistent finding in cytogenetic adults.8 15 16 studies in leucocytes of some mentally retarded Fragile X infants often have relative macro- males. The folic acid and thymidine depleted cephaly persisting into adult life.'7 However, culture medium was identified as the the adult height of affected males is below the essential factor for the induction of this norm.'7 18 A few patients present with either Department of Clinical heritable fragile site at Xq27.3 During the sev- , University overweight or general persisting overgrowth enties, the combination of X linked mental which can be confused with either Prader-Willi Hospital Dijkzigt and retardation and Erasmus P macro-orchidism was syndrome or Sotos syndrome.'9-24 Associations University, recognised.4-6 Later, other clinical 0 Box 1738, 3000 DR characteris- with other syndromes are reported; examples Rotterdam, The were established, for example, a long face are the Robin sequence (micrognathia, glos- Netherlands with large, protruding ears, and behavioural soptosis, and cleft soft the FG B B A de Vries features, palate),25 including avoidance of eye contact, syndrome (congenital , macro- D J J Halley hyperactivity, hand flapping, and B A Oostra perseverative cephaly, distinctive face, and imperforate speech.7 8 M F Niermeijer anus),26 27 and the DiGeorge anomaly (defects The fragile X syndrome was diagnosed at of , and that time parathyroids, great vessels).28 Correspondence to: by cytogenetic detection of the fragile However, there is no definite evidence for a Dr de Vries. site at Xq27.3. It some had special features for causal relationship between the FMR1 gene 580 De Vries, Halley, Oostra, et al

Table 1 Clinical features and medical problems offragile Although several neuroimaging and a few X males * neuropathological studies have been done on the central nervous system in fragile X patients, knowledge on specific structural and cellular J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from Clinlicalfeaturest defects is still limited. Reduced size of the pos- Long face Prominent ears terior on MRI in male High arched palate patients and increased size of the caudate Hyperextensible finger joints nucleus, thalamus, and hippocampus have Double jointed thumbs Single palmar creases been reported.47 48 These findings have neither Hand calluses been confirmed nor refuted by others. studies of three fragile X Heart murmur or click Neuropathological Macro-orchidism males' brains showed immature, thin, and long Medical problemsf dendritic spines in different regions, with a Vomiting but with Failure to thrive in infancy reduction in mean synaptic contact Strabismus preservation of neuronal density in the Myopia or hyperopia neocortex." 49 5() Similar abnormal dendritic Hernia Joint dislocation spines were observed in fragile X knockout Orthopaedic problems mice.' Interestingly, in normal mouse brain, high FMRP expression is observed in the sy- Seizures naptoneurosomes which could be related to the Mitral valve prolapse high transcriptional activity necessary for func- Apnoea tion of neuronal spines.52 However, abnormally ADHD long, thin dendritic spines are also seen in cases Motor tics with chromosomal abnormalities53 or unspeci- fied mental retardation.'4 Apparently, these are *Majority of items scored in 100-280 patients. tAdapted from Merenstein et al.'-' non-specific neuropathological changes, as is tAdapted from Hagerman.' the neuronal heterotopia observed in two affected males and linked to their . In mutation and these . Several obser- the testes, macro-orchidism seems to be vations of the co-occurrence of the fragile X associated with increased tubular length and syndrome and sex chromosomal abnormalities interstitial oedema.5 4 5 like 47,XXY or 45,XO/46,XX25 34 suggest an increased rate of non-disjunction related to the The FMRl gene fragile . The identification of the FMR1 gene5' princi- Scales for facilitating diagnosis based on pally elucidated the special characteristics of physical and behavioural features have been the transmission of the defect in fragile X fami- developed and found to be effective.35 lies rather than explaining the mental retarda- Recognition of behavioural features also assists tion itself. The FMR1 gene has a size of 38 kb http://jmg.bmj.com/ in the diagnosis. In the younger child, hand with 17 exons57 and an untranslated polymor- flapping, hand biting, tactile defensiveness, phic CGG repeat in the first exon.58 In the nor- poor eye contact, hyperactivity, and persevera- mal population, this CGG repeat varies from tive speech are suggestive ofthe diagnosis.8 The six to 54 units58 (fig 4) with an average of 30 "autistic-like" behaviour of many fragile X units. The in the FMR1 gene can be males initiated the debate about whether divided into a major and a minor class (table

autism is associated with the fragile X 2). The major class of FMR1 gene mutations on September 26, 2021 by guest. Protected copyright. syndrome."4 42 affects the CGG repeat, whereas mutations in Although in premutation carriers an the coding region are infrequently observed. increased incidence of premature has been reported,43 45 a recent study indicated CGG REPEAT EXPANSIONS that the premutation does not seem to be a On the basis of the size of the CGG repeat and major risk for early menopause.4" its effect on protein and phenotypic expression,

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Figure 1 Three affected males with characteristic facialfeatures of the fragile X syndrome. Note the long, narrow face with large, everted ears and the poor eye contact. Thefragile X syndrome 581 J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from

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Figure 2 Some clinicalfeatures in fragile X subjects. (A) High arched palate, (B) dental crowding, (C) hyperextensible MPjoints, (D) hand calluses, and (E) pes planus.

two types of mutations within the CGG repeat The CGG repeat is generally interspersed http://jmg.bmj.com/ can be distinguished: the premutation (size 43 with two AGG triplets6'-66 and their position at to 200 units) apparently without clinical effect the 3' end influences repeat instability, with an and the full mutation (size >200 units) which, instability threshold of 34-38 uninterrupted if fully methylated, is associated with mental CGG repeats.6' 65 The potential predictive retardation in all male and in 50-70% offemale value of the size of the uninterrupted CGG patients. At the protein level, FMR1 is absent repeat for risk prediction in the offspring of from cells in cases with a fully methylated

female premutation carriers has not been on September 26, 2021 by guest. Protected copyright. mutation only. adequately evaluated until now. The mutations in the CGG repeat are In the 43 to 55 CGG repeat range, there is an dynamic. They may change from generation to overlap between normal and premutated alle- generation as well as within a single person les, often referred to as the "grey zone". Only during early embryogenesis. Stable with the observation of intrafamilial instability iden- a CGG repeat <55 units are defined as normal, tifies these "intermediate" alleles as premuta- and unstable alleles ranging from 43 to 200 tion alleles.6' 62 64 However, such premutation repeat units are regarded as premutation alleles can also be stably inherited through alleles. many generations.67 68Alteration in one genera- The premutation and its intergenerational tion from alleles of <43 to >43 repeats is instability unknown, suggesting that such changes are The premutation with repeats in the range of very gradual. However, regressions from pre- 43 to 200 units causes no phenotypic abnor- mutation to normal size (<43 repeats) have malities in (male and female) carriers58 (fig 4). been observed within one generation.6-7' The intergenerational (in)stability of the pre- The timing of any transition from pre- to full mutation is dependent on the sex of the trans- mutation is not fully understood. The presence mitting parent and on the size of the repeat. of full mutations in the ovary of female , Only women with a premutation have a risk of without evidence of a premutation, and simula- having affected offspring with a full mutation. tion studies support the hypothesis of precon- This risk of expansion into a full mutation ceptional enlargement.72 73 This seems to con- depends on the size of their premutation; it is flict with the finding of only premutations in less than 20% for smaller premutation alleles the of four males with a full mutation in (<70 CGG repeats), but more than 80% for somatic cells.74 The latter observation might be larger premutation alleles (>80 CGG explained by regression of a full mutation to a repeats).58 60 premutation in a limited number of cell 582 De Vries, Halley, Oostra, et al J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from

Figure 3 Two affected males and one female with less obvious facialfeatures.

lineages. In the germline, we have then to (>60%) of leucocytes with an unmethylated assume a selection for those cells with a full mutation have been reported.92 premutation. OTHER MUTATIONS The full mutation Mutations other than CGG repeat expansions Subjects with the fragile X syndrome have are most often large deletions (partial or com- CGG repeats above 200 units, the "full muta- plete) of the FMR1 gene, with or without sur- tion" (fig 4). The expansion into full mutation rounding regions. Several unrelated cases have is usually accompanied by hypermethylation of been reported.'°" 112 the repeat and its flanking regions, resulting in The first intragenic mutation in the FMR1 a shut down of transcription and the absence of gene, a de novo (an Ile304Asn the FMR1 protein.75 7' The latter causes the substitution) in exon 1 1, was reported in a pro- mental retardation. All males and a majority of foundly retarded male with fragile X features. "3 the females with a hypermethylated full muta- The resided in one of the tion are mentally retarded.79 8 important domains of the FMR1 protein."' In somatic cells of fragile X patients, the Intragenic mutations may result in an absence mitotic instability of the repeat of the full of FMRP and thereby cause the fragile X syn- mutation causes longer and shorter expan- drome. One single de novo in sions, so all fragile X patients are somatic exon 5 and an inherited two base pair substitu- http://jmg.bmj.com/ mosaics. The instability occurs during early tion in exon 2 resulting in a loss of protein pro- embryogenesis and results in this somatic duction have been described."' A C to T point heterogeneity."4 85 mutation in the 14th nucleotide of intron 10 of In fragile X patients, two special subclasses the FMR1 gene causing a deletion of exon 10 of mosaicism can be distinguished on the basis was reported in three unrelated fragile X of size and pattern. patients.'' All five male patients showed the (1) "Size mosaics" are those patients with mental retardation and physical features char- both a full and a premutation, which is acteristic of the fragile X syndrome. on September 26, 2021 by guest. Protected copyright. observed in 20-40% of the male patients.'"' Also, males with somatic mosaicism for a full mutation and a (partial) deletion of the FMR1 Molecular diagnosis gene have been reported.87 9' Before the cloning of the FMR1 gene, cytoge- (2) "Methylation mosaics" are subjects with netic detection of the fragile site at Xq27.3 variations between cells in the methylation sta- (FRAXA) was the only method for (prenatal) tus of a full mutation. The proportion of leuco- diagnosis and carrier detection. However, this cytes with an unmethylated full mutation may method was inadequate for carrier detection vary from low (10%) to 100%. A few intellec- and other fragile sites localised near FRAXA tually normal males with a high proportion caused diagnostic confusion, like FRAXE Table 2 Mutations in the FMR1 gene and their effect

Mutation No of cases FMRP MR Physical features Referenices * CGG repeat expanision Premutation (43-200 CGGs) Innumerable Normal - - 58, 79, 198 Full mutation (>200 CGGs) >60% methylated Innumerable Reduced/absent + + 75-77, 79 + premutation Innumerable Reduced + + 80, 86, 87, 93, 157, 158 + deletion 8 Absent + + 87-91 <40% methylated 16 >60% - Absent or minor 92-99 Other nmutationls 2 base pair change (exon 2) 1 Absent + + 115 1 base pair deletion (exon 5) 1 Absent + + 115 1 base pair change (exon 11) 1 Present + + (less typical) 113 1 base pair change (intron 10) 3 Truncated + + 116 Partial or complete deletion of FMR1 gene 14 Absent + + (some atypical) 100-112 *For additional references see text. The fragile X syndrome 583

(=0.6 Mb distal to FRAXA) and FRAXF (1-2 sue. Nevertheless, testing of methylation status Mb distal to FRAXA)."7 19 may be useful in amniotic cells.

The methods described above can identify J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from DNA ANALYSIS repeat amplifications in the CGG repeat and The actual diagnostic standard is the size ofthe large deletions in the FMR1 gene. Subtle CGG repeat amplification in patients and car- mutations outside the CGG repeat will not be riers. Two major methods (Southern blots and detected. At present, only a limited number of PCR) are used. For Southern blotting different mutations in the coding region of the FMR1 restriction enzymes may be used, such as Hin- gene have been found using sequencing dIII, BglII, and PstI, each with their specific techniques."3 115 116 It is conceivable that other fragment sizes.'20 A double digest, including a rare mutations in the FMR1 gene will be methylation sensitive restriction enzyme, ena- detected in the future. bles detection of the methylation of the CpG island in the CGG region. This is also valuable FMRP TEST for identification of males with incompletely Recently, an FMR1 protein antibody test was methylated full mutations ("methylation mosa- developed for detecting the presence or ab- ics"). The double digest may also be used in sence of FMRP in lymphocytes.'26 This rapid distinguishing between an unmethylated large detection method allows the diagnosis of frag- premutation and a small methylated full muta- ile X syndrome from a blood smear; cells from tion. fragile X males with a methylated full mutation An alternative method for determining CGG produce no FMRP.'26 The proportion of cells repeat size is the polymerase chain reaction or with FMRP production allows a distinction PCR analysis.58 However, CGG repeats of between males with the fragile X syndrome more than =150 units are generally difficult to (<30% staining cells) and normal subjects amplify and will often not give a detectable (>40% staining cells).'27 The test does not product after the PCR. detect dysfunctional , neither does it Most laboratories use both methods (restric- differentiate between normal and premutation tion enzyme and PCR analysis) for postnatal alleles. and prenatal diagnosis.'21-'25 When a premuta- As a result of lyonisation, females with a full tion is found in a chorionic villus sample, con- mutation may have staining ofup to 80% ofthe firmation in amniotic fluid or a cord blood lymphocytes, overlapping with 40% of the nor- sample might be considered to ascertain the mal females who have staining in less than 80% stability of the premutation during fetal life.'25 of their lymphocytes.'27 The test is therefore In the (near) future, when sufficient infor- not suitable for detecting females with a full mation about stability becomes available, this mutation or a premutation. precaution may become unnecessary. Absence The main application for this rapid and of methylation of the fully expanded mutation cheap test is screening for the fragile X http://jmg.bmj.com/ in chorionic villi at 11 weeks of gestational age syndrome among mentally retarded males or has been observed in a with hypermeth- male neonates. DNA studies are needed to ylation of the full mutation in fetal tissue.76 123 confirm the diagnosis and to ascertain carrier Methylation status is therefore unreliable in an status in their relatives. early (< 12 weeks ofpregnancy) chorionic villus Recently, the antibody test was successfully sample because it may differ from the fetal tis- used in the prenatal diagnosis of at risk male on September 26, 2021 by guest. Protected copyright.

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Figure 4 Part of the FMRl gene and its CGG related mutations in thefirst exon of the FMRl gene that contains the CGG repeat (grey) (see textforfurther details). 584 De Vries, Halley, Oostra, et al

Table 3 Overview ofDNA screening programmes among institutes and schools for the mentally retarded*

Persons with grey zone Persons with

Persons studied premutation Persons with full mutation J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from M+F (CGG M F M+F M F range) M+F M F M+F Characteristics ofpopulation studied

England 219 - 219 - - - 6 6 (2.7%) Mentally retarded children in Webb et al (1986), Turner et al (1996) schools/institutions Australia 472 - 472 - - - - 10 - 10 (2.1%) Mentally retarded children in special Turner et al (1986), Turner et al (1996) schools England 180 74 254 - - 11 0 4 0 4 (1.6%) Children with special educational needs Jacobs et al (1992) (41-49) United States 299 140 439 - - - 1 (M) 1 3 4 (0.9%) Retarded/learning disabled children Hagerman et al (1994) England 103 51 154 - - - 0 4 0 4 (2.5%) School children with learning difficulties Slaney et al (1995) England 1013 - 1013 35 - 35 1 (M) 5 - 5 (0.5%) Boys with learning difficulties Murray et al (1996) (41-60) United States 888 391 1279 3 7 10 0 1 1 2 (0.2%) Children in education Meadows et al (1996) (50-60) programmes The Netherlands 870 661 1531 10 9 19 0 9 2 11 (0.7%) Mentally retarded children/adults in De Vries et al (1997) (43-60) institutions, special schools Subtotalt 3353 1317 4670 75 (1.8%) 2 (0.04%) 24 (0.7%) 6 (0.5%) 30 (0.6%) *For references see text. tWithout (reassessed) studies of Webb et al (1986) and Turner et al (1986). pregnancies using chorionic villi at 12.5 weeks identified (±1% of the tested patients have the of gestation'28 and uncultured amniotic fluid fragile X syndrome) may be significantly cells. 129 Further tests are needed to establish the increased by preselection based on physical reliability of this technique for prenatal diagno- features.35 36 38 39 155 However, the yield of iden- sis because of methylation differences between tified and informed (pre)mutation carriers sur- villi and fetal tissue at week 1. passes the number of identified affected males. Precise identification of the various allele classes may have major implications for screen- Prevalence and screening ing programmes in the general population. For Several genetic epidemiological studies of the accurate counselling of females identified with fragile X syndrome have been done using vari- a premutation sized allele, more knowledge of ous techniques (, DNA) and clini- the allele's stability is desirable. cal selection criteria, for example, level of men- tal retardation, macro-orchidism, or

autism.'30 131 Studies using DNA analysis indi- http://jmg.bmj.com/ cate a prevalence for males of 1/4425 to FMRP 1/6045.38 132 133 The combined results of studies The translation of the FMR1 gene starts distal from England, the United States, Australia, to the CGG repeat and results in a 70-80 kDa and The Netherlands (table 3), irrespective of protein.77 156 In lymphoblastoid cells the normal methodological differences, showed, in a total and premutated FMR1 alleles are transcribed of 4670 mentally retarded subjects, 30 (0.6%) equally, resulting in FMRI transcripts with a newly diagnosed fragile X patients.38 133 138 half life of 12 hours, and identical levels of In the general population, a premutation FMRI protein expression.'57 158 Alternative on September 26, 2021 by guest. Protected copyright. carrier frequency among females of ±1/250 has splicing in the FMR1 gene occurs in various been observed.'38 14 This reflects the propor- human tissues, leading to 24 distinct tional excess of relatively stable premutation transcripts. 156 151 168 WVhether those transcripts sized alleles over full mutation alleles. This high lead to functional proteins is unclear. The frequency of premutations might partially be localisation of FMRP is predominantly explained by a founder effect in the population cytoplasmic.77 studied (French-Canadian).'4' In normal human brain, FMRP is found in The identification of the gene paved the way nearly all neurones studied, but is absent or for screening programmes; however, it is still an reduced in non-neuronal cells.158 'I' In normal open question if large scale testing, for human testes, both the primordial germ cells example, among the mentally retarded or (prenatally) and spermatogonia (postnatally) young adult females, is desirable.'42 143 Testing show high FMRP expression, with low expres- mentally retarded subjects leads to diagnosis sion in the Sertoli and Leydig cells. Remark- and improved support for newly diagnosed ably, in fragile X patients a proportion of the fragile X patients. Such programmes have been primordial germ cells and all early spermatogo- in place since cytogenetic testing was nia show FMRP expression.1"1 This is concord- introduced.'I'"5' DNA testing is much more ant with the presence of a premutation allele in precise, especially for subsequent testing for the sperm of patients.74 (pre)mutation carriers in the family.38 132 136 138 High FMRP expression was also found in These programmes are welcomed by parents/ other tissues not obviously involved in the frag- guardians and well accepted by the staff of ile X phenotype,77 158 162 163 for example, divid- schools and institutes.38 15' An underdiagnosis ing mesodermal cells during wound healing of the fragile X syndrome of more than 50% and in cardiac hyperplasia, whereas in other even in well developed countries is found.38 circumstances dermis and skeletal and cardiac The relatively low "yield" of new cases muscle were negative for FMRP.'58 The fragile X syndronie 585

The FMRP has RNA binding capacity active X chromosome (which are unable to owing to characteristic sequence motifs in the produce FMRP) has an influence on cognitive protein: an RGG box and ribonucleoprotein development.63 163 185 Apparently, there is no J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from particle (RNP) K domains (desig- large scale transmission/passage of corrective nated KH domains).'' 1"7 A patient with a molecules from FMRP producing cells to non- severe phenotype and a missense mutation in FMRP producing neurones. one of these KH domains had quantitatively normal FMRP production in lymphoblastoid Management cells.''3 114 Mental retardation and behavioural problems FMRP binds its own FMR1 transcript and, dominate the clinical presentation. The mental with unknown selectivity, 4% of fetal brain retardation is not amenable to intervention. transcripts.'65 Recently, a more specific binding However, careful medical follow up and some- of FMRP was found via RNA with the 60S times intervention are required as the physical ribosomal subunit, suggesting a function of and behavioural problems of fragile X patients FMRP in ribosomal function and in translation are related to their stage of development (table of certain proteins.'68 172 A nuclear localisation 1)."'6 Seizures observed in approximately 20% signal encoded by the 3' part of the gene, as of males and 5% of females necessitate timely well as a nuclear export signal encoded by exon diagnosis and treatment. During infancy, con- 14, were seen. Eberhart et all'3 postulated an nective tissue abnormalities, such as congenital interaction of FMRP with mRNA(s) in the hip dislocations and inguinal hernia, may be nucleus, followed by export to the cytoplasm as present. In later life the connective tissue part of a mRNP particle where it associates dysplasia may lead to , flat feet, and with . A role of the FMRP in the mitral valve prolapse.'6 The mitral valve translation machinery could explain the high prolapse requires evaluation by a cardiologist expression in (some) actively dividing pre- and and a recommendation for prophy- postnatal tissues. laxis before surgical or dental procedures. Effects of the absence ofFMRP were studied Some children fail to thrive because of in knockout mice. These mice showed relevant gastro-oesophageal reflux, tactile defensive- learning deficits, hyperactivity, and macro- ness, or difficulties in sucking."'6 The latter of orchidism as observed in fragile X these problems requires attention from a patients.'4 ' The knockout mice brain and specialised speech therapist or physiotherapist, testes lacked pathological abnormalities, ex- while the gastro-oesophageal reflux can be cept for enlargement of the testes.'74 treated by dietary advice or medication or both. Surgery is rarely needed. The frequent Mental retardation otitis media and sinusitis in approximately 50% Fragile X males show some variability in intel- of affected children require adequate interven-

lectual : in most prepubertal boys it is tion ( or polyethylene tubes or http://jmg.bmj.com/ moderate (IQ=35-55), and in adults moderate both)."' Approximately 30-50% of cases need to severe (IQ=20-40).6 176 This apparent de- ophthalmological help for strabismus, myopia, cline in IQ probably reflects a slowing of cogni- or hyperopia.' ' tive development, stabilising after puberty.' hl1 Behavioural problems include attention defi- In affected males, a methylated, full muta- cit and hyperactivity at a young age. Although tion will lead to mental impairment whatever fragile X patients are generally friendly, some the repeat size of the mutation. I7 It has been may show aggressive behaviour in adulthood. suggested that males with a "size " DNA Influencing these behavioural problems is diffi- on September 26, 2021 by guest. Protected copyright. pattern (premutation together with the full cult, although behavioural therapy and avoid- mutation with an associated residual FMR1 ance of overwhelming stimuli may alleviate protein production) might have better mental some of the symptoms. In some countries, functioning than fragile X males with a full pharmacological intervention for the behav- mutation only. 14 311 16I2 However, others found ioural problems is common.'66 However, ad- no mean IQ differences."' 67 Apparently, the equately controlled studies on their effective- proportion of brain neurones producing ness in the fragile X syndrome are scarce. FMRP in "size mosaic" males is insufficient for The need for and training, normal cognitive functioning. This is also especially in the younger child, is of primary shown by the presence of mental retardation in importance. Speech therapists and physio- a male mosaic for a partial deletion of the CGG therapists can help with language and motor repeat and a full mutation, who had FMRP development. 162192 production in 28% of his lymphocytes.9" An indication of the proportion of cells Conclusions expressing FMRP needed for normal mental Since the original observation of the fragile X functioning is shown in mentally normal males syndrome as a familial sex linked mental retar- with incompletely methylated full mutations in dation syndrome in 1943, its clinical and more than 60% of their leucocytes. 6 6397 66 molecular diagnosis has greatly improved. The An insufficient proportion of FMRP produc- biological basis for the unique phenomena of ing cells also explains cognitive impairment normal transmitting males and the "Sherman (IQ<85) in females who have a full mutation; paradox"'63 1(4 became evident after the cloning 52-82% of the women with a full mutation of the FMR1 gene in 1991, as well as the nature were shown to have mental impairment of the CGG repeat at the 5' end of the FMR1 (IQ<85).7 61 X31S3 In these heterozygotes, the gene and its intergenerational instability. The proportion of abnormal FMR1 alleles on the precise mechanism of the repeat expansion is 586 De Vries, Halley, Oostra, et al

still unknown.15 185 The FMR1 protein shows 8 Hagerman RJ. Physical and behavioural phenotype. In: Hagerman RJ, Cronister A, eds. Fragile-X syndromtie: widespread expression in nearly all tissues diagnosis, treatmentand research. Baltimore: The Johns Hop- studied, with both nuclear and cytoplasmic kins University Press, 1996. 9 Verkerk AJ, Pieretti M, Sutcliffe JS, etal. Identification of a J Med Genet: first published as 10.1136/jmg.35.7.579 on 1 July 1998. Downloaded from localisation. The protein seems to shuttle gene (FMR-1) containing a CGG repeat coincident with a between nucleus and cytoplasm and it can bind breakpoint cluster region exhibiting length variation in fragile X syndrome. Cell 1991;65:905-14. to ribosomes via mRNA. Why the major clini- 10 Yu S, Pritchard M, Kremer E, et al. Fragile X cal are restricted to the brain and characterized by an unstable region of DNA. Scienzce 1991; symptoms 252:1179-81. testis remains unresolved. 11 Oberle I, Rousseau F, Heitz D, etal. Instability of a 550-base The is the most con- pair DNA segment and abnormal methylation in fragile X syndrome. Science 1991;252:1097-102. sistent clinical symptom in males with the 12 Wells RD, Warren ST. Genetic instabilities and hereditary, methylated full mutation. FMR1 protein ex- neurological . New York: Academic Press, 1998. 13 Richards BW, Sylvester PE, Brooker C. Fragile X-linked pression in less than 30% of lymphocytes is mental retardation: the Martin-Bell syndrome. YMentDefic associated with mental retardation and other Res 1981;25:253-6. 14 Merenstein SA, Sobesky WE, Taylor AK, Riddle JE, Tran clinical symptoms.87 9" Size mosaicism (com- HX, Hagerman RJ. Molecular-clinical correlations in bined presence of and full muta- males with an expanded FMR1 mutation. Amn YMed Geniet premutation 1996;64:388-94. tion) does not seem to mitigate mental retarda- 15 Partington MW The fragile X syndrome II: preliminary tion while it is likely that at least 50% of (brain) data on growth and development in males. AmnY Med Genet 1984;17: 175-94. cells expressing FMRP will be required to 16 Wisniewski KE, Segan SM, Miezejeski CM, Sersen EA, function at a normal IQ level. Rudelli RD. The fra(X) syndrome: neurological, electro- physiological, and neuropathological abnormalities. Am]Y The identification of the gene defect enabled Med Genet 1991;38:476-80. screening programmes to be implemented, but 17 Butler MG, Brunschwig A, Miller LK, Hagerman RJ. Standards for selected anthropometric measurements in the practicability and desirability of screening males with the fragile X syndrome. 1992;89: the mentally retarded or young adult females is 1059-62. 18 Loesch DZ, Lafranchi M, Scott D. Anthropometry in still under debate.'42 149 187 Testing mentally Martin-Bell syndrome. Ami YMed Genet 1988;30: 149-64. retarded subjects leads to diagnosis and 19 Fryns JP, Haspeslagh M, Dereymaeker AM, Volcke P, Van den Berghe H. A peculiar subphenotype in the fra(X) improved support for newly diagnosed fragile syndrome: extreme obesity--stubby hands and X patients and allows identification of all feet-diffuse hyperpigmentation. Further evidence of dis- turbed hypothalamic function in the fra(X) syndrome? Clin premutation carriers in the proband's Genet 1987;32:388-92. family.'32 136 138 Subsequently, a fragile X diag- 20 De Vries BB, Fryns JP, Butler MG, et al. Clinical and molecular studies in fragile X patients with a Prader-Willi- nosis assists the parents in coping with like phenotype. YMed Geniet 1993;30:761-6. problems and eventually accepting their men- 21 Schrander-Stumpel C, Gerver WJ, Meyer H, Engelen J, Mulder H, Fryns JP. 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YMed Getnet 1995;32:764-9. strategies, one should consider that protein 25 Lachiewicz AM, Hoegerman SF, Holmgren G, Holmberg production probably needs to be restored in at E, Arinbjarnarson K. Association of the Robin sequence with the fragile X syndrome. Am3Med Genet 1991 ;41:275- least ±50% of the cells (based on studies in 8. blood) to achieve normal cognitive function. 26 Loesch DZ, Hay DA, Sheffield LJ. Fragile X family with unusual digital and facial abnormalities, cleft lip and Before this long term aim, efforts should be and Ami ] Med 1992;44:543-50. palate, epilepsy. Genet on September 26, 2021 by guest. Protected copyright. concentrated on improving the diagnostic rate 27 Piussan C, Mathieu M, Berquin P, Fryns JP. Fragile X for mutation and FG syndrome-like phenotype. Anm Y Med and subsequently the conventional support Genet 1996;64:395-8. fragile X syndrome patients. In the meantime, 28 Macpherson JN, Curtis G, Crolla JA, et al. Unusual concentrate on (CGG)n expansion and recombination in a family with (basic) research can increasing fragile X and DiGeorge syndrome. ] Med Genet 1995;32: our understanding of the actual mechanism(s) 236-9. of the neuronal the 29 Kupke KG, Soreng AL, Muller U. Origin of the supernu- dysfunction underlying merary X chromosome in a patient with fragile X and mental retardation. . Anm Y Med Genet 1991,38:440-4. 30 Voelckel MA, Pellissier MC, Piquet C, et al. Fragile X syn- drome in an extended family with special reference to an We are grateful to Professor H Galjaard and Professor P J Wil- affected male with Klinefelter syndrome. Amii Y Med Geniet lems for their comments on the manuscript, and to the Rotter- 1991 ;38:374-7. dam Foundation of Clinical Genetics for its financial support. 31 Tejada MI, Mornet E, Tizzano E, Molina M, Baiget M, We would like to thank Dr A M W van den Ouweland, Dr R Boue A. 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