DOI: 10.1212/WNL.0b013e3181b8799a 2009;73;962-969 Neurology

Scala, M. Schaer, P. Gressens, B. Gerard and A. Verloes Hyon, B. Isidor, A. Mégarbané, U. Moog, S. Odent, K. Hernandez, N. Pouvreau, I.

C.Billette de Villemeur, O. Boespflug-Tanguy, L. Burglen, E. Del Giudice, F. Guimiot, S. Passemard, L. Titomanlio, M. Elmaleh, A. Afenjar, J. -L. Alessandri, G. Andria, T.

mutationsASPMdue to Expanding the clinical and neuroradiologic phenotype of primary microcephaly

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Expanding the clinical and neuroradiologicphenotype of primary microcephaly due toASPM mutations

S. Passemard, MD*L. Titomanlio, MD, PhD*M. Elmaleh, MDA. Afenjar, MDJ.-L. Alessandri, MDG. Andria, MD, PhDT. Billette de Villemeur,

MDO. Boespflug-Tanguy,

MD, PhDL. Burglen, MD, PhDE. Del Giudice, MDF. Guimiot, PhDC. Hyon, MDB. Isidor, MDA. Megarbane, MDU. Moog, MD, PhDS. Odent, MD, PhDK. HernandezN. PouvreauI. Scala, MD, PhDM. Schaer, MD, PhDP. Gressens, MD, PhDB. Gerard, PharmD, PhDA. Verloes, MD, PhD

ABSTRACT

Objective: To determine the spectrum of clinical, neuropsychological, and neuroradiologic features inpatients with autosomal recessive primary microcephaly (MCPH) due to ASPM gene mutations.

Methods: ASPM was sequenced in 52 unrelated MCPH probands. In patients with ASPM muta-tions, we evaluated the clinical phenotype, cognition, behavior, brain MRI, and family.

Results: We found homozygous or compound heterozygous ASPM loss-of-function mutations in11 (22%) probands and 5 siblings. The probands harbored 18 different mutations, of which 16were new. Microcephaly was severe after 1 year of age in all 16 patients, although in 4 patientsthe occipital-frontal circumference (OFC) at birth was decreased by only 2 SD. The OFC Z scoreconsistently decreased after birth. Late-onset seizures occurred in 3 patients and significantpyramidal tract involvement in 1 patient. Intellectual quotients ranged from borderline-normal tosevere mental retardation. Mild motor delay was noted in 7/16 patients. Language developmentwas delayed in all patients older than 3 years. Brain MRI (n � 12) showed a simplified gyral patternin 9 patients and several malformations including ventricle enlargement (n � 7), partial corpuscallosum agenesis (n � 3), mild cerebellar hypoplasia (n � 1), focal cortical dysplasia (n � 1), andunilateral polymicrogyria (n � 1). Non-neurologic abnormalities consisted of short stature (n � 1),idiopathic premature puberty (n � 1), and renal dysplasia (n � 1).

Conclusions: We provide a detailed description of features associated with ASPM mutations.Borderline microcephaly at birth, borderline-normal intellectual efficiency, and brain malforma-tions can occur in ASPM-related primary hereditary microcephaly. Neurology® 2009;73:962–969

GLOSSARYMCPH � primary hereditary microcephaly; MR � mental retardation; MSG � microcephaly with simplified gyral pattern; MV �microcephalia vera; OFC � occipital-frontal circumference.

Autosomal recessive primary microcephaly, or MCPH (for microcephaly primary heredi-tary, MIM 251200), formerly called microcephalia vera (MV), is clinically defined by 4criteria: congenital microcephaly ��3 SD; mild to moderate mental retardation (MR);no other neurologic anomalies, except mild epilepsy; and normal height, weight, andappearance.1-4 Five genes are known to be involved in autosomal recessive microcephaly:MCPH1 (8p23), ASPM (MCPH5; 1q31), CDK5RAP2 (MCPH3; 9q34), CENPJ(MCPH6; 13q12.2), and STIL (MCPH7; 1p32.3-p33). Two additional loci have beenmapped: MCPH2 (19q13.1–13.2) and MCPH4 (15q15-q21). About one-third of familiesare still unlinked. ASPM is considered responsible for about half the cases of MCPH.2,5

Fewer than 100 cases of MCPH due to ASPM mutations have been published. In a recent

*These authors contributed equally.

From the Department of Genetics (S.P., L.T., C.H., K.H., N.P., B.G., A.V.), Service de Neuropediatrie (S.P., L.T., P.G.), Service de Radiologie (M.E.),Service de Foetopathologie (F.G.), and INSERM (S.P., P.G., A.V.), U676, Hopital Robert Debre, APHP, Paris, France; Department of Pediatrics (L.T.,G.A., E.D., I.S.), Federico II University, Naples, Italy; Service de Genetique Clinique (A.A., L.B.) and Service de Neuropediatrie (T.B.d.V.), HopitalTrousseau, APHP, Paris; Universites Paris VI/XII (T.B.d.V.); Service de Reanimation Neonatale et Pediatrique (J.-L.A.), CHD Felix Guyon, Saint Denis dela Reunion; Service de Genetique Clinique (O.B.-T.), Universite de Clermont Ferrand; Service de Genetique Clinique (B.I.), Universite de Nantes, France;Unite de Genetique Medicale (A.M.), Faculte de Medecine, Universite de Saint Joseph, Beirut, Lebanon; Institute of Human Genetics (U.M.), HeidelbergUniversity, Germany; Service de Genetique Clinique (S.O.), Universite de Rennes, France; and Department of Psychiatry (M.S.), University of GenevaSchool of Medicine, Switzerland.

Supported by the GIS-Maladies Rares, Fondation Lejeune, Fondation pour la Recherche Medicale, and Mariani Foundation (4 nonprofitorganizations).

Disclosure: Author disclosures are provided at the end of the article.

Address correspondence andreprint requests to Pr. AlainVerloes, Department of ClinicalGenetics, Robert DebreUniversity Hospital, 48 BdSerurier, 75019 Paris, Francealain.verloes@rdb.aphp.fr

962 Copyright © 2009 by AAN Enterprises, Inc. at Universitaet Heidelberg on October 1, 2009 www.neurology.orgDownloaded from

study,2 ASPM mutations were only foundin individuals meeting MCPH criteria (3/13). In those who did not meet the criteria(n � 32), no ASPM mutations were found.ASPM mutations have been associated withmicrocephaly with simplified gyral patternor without simplified gyration. A simplifiedgyration pattern was noted in 5 patients1,6,7

and frontal predominance of brain under-development was suggested.7 Although allprevious reports mention the presence ofMR, only 2 of them specify IQ values.7,8

To shed light on the phenotypic spectrumof ASPM mutations, we reviewed the clinicaland brain imaging data from 16 previouslyunreported patients with ASPM mutationsidentified among 52 pedigrees with MCPH.

METHODS All patients referred to one of the participatingcenters and diagnosed with MCPH between November 2004and December 2008 were considered eligible for ASPM se-quencing. Clinical and neuroradiologic data were collected byeach center. Genetic analysis was performed in France (Départe-ment de Genetique Clinique, APHP-Hopital Robert Debre, Paris).

Patient selection. Patients with MCPH were evaluated bygeneticists belonging to a network of French Reference Cen-ters for Developmental Anomalies, except those from Ger-many and Lebanon, who were evaluated by correspondinggeneticists.

Inclusion criteria were as follows: microcephaly ��2 SD(before 1 year) and ��3 SD (after 1 year of age), height ��3SD or at least 2 SD higher than the OFC, absence of gross CNSmalformations such as lissencephaly or generalized pachygyria,and absence of multiple congenital malformations. All patientshad normal karyotype, no subtelomeric rearrangements, normalmetabolic and TORCH screens, and a normal maternal phe-nylketonuria screen. No patient had an identifiable cause of mi-crocephaly. To avoid selection bias, the inclusion criteria werechecked by a pediatric neurologist (S.P. or L.T.) and a geneticist(A.V.) prior to DNA screening. Patients with mutations werereassessed to refine their phenotypic description.

The patients were divided into 3 subgroups. Group 1 metthe strict diagnostic criteria for MCPH, i.e., severe congenitalmicrocephaly (OFC ��3 SD since birth), subnormal motordevelopment, MR, otherwise normal neurologic examination,normal stature, MV or microcephaly with simplified gyral pat-tern (MSG) type 1, and absence of other neuroradiologic anom-alies. Group 2 was similar to group 1 but with an OFC at birthbetween �2 and �3 SD. Group 3 included all patients notmeeting criteria for MCPH, such as those with epilepsy, spastic-ity, brain malformations (e.g., partial corpus callosum agenesis,cerebellar hypoplasia, or focal cortical defect), or minor extracra-nial malformations.

Written informed consent was obtained from all patients (orguardians of patients) participating in the study, and the studywas approved by the local ethics committee.

Microsatellite analysis. Primers for MCPH1– 6 loci weredesigned (available on request). PCR products were analyzed

on ABI 3100 using Genescan software. Microsatellites wereused in multiplex and/or consanguineous families to sort thecandidate loci.

Sequencing. The DNA was extracted using the Qiagen kit(Qiagen, Hilden, Germany). DNA preamplification was per-formed using the Genomiphi kit (GE Healthcare BiosciencesAB, Uppsala, Sweden). Direct sequencing of the coding se-quence and exon/intron junctions of ASPM (NM 018136)was achieved using 36 primer pairs (available on request). PCRproducts were sequenced using the BigDye kit (Applied, FosterCity, CA) and analyzed on an ABI 3130xl sequencer. Data wereanalyzed using Seqcape software (Applied). All mutations werecontrolled in a second run of sequence analysis on native DNA(C.H. and B.G.).

Quantitative PCR. Quantitative PCR was performed to con-firm deletions, using standard procedures on the Albi 7900 realtime PCR system (AME Bioscience, Toroed, Norway).

Clinical and developmental assessment of ASPM-mutated patients and affected siblings. Except for themembers of family 11, all patients with ASPM mutations wereclinically reassessed by a geneticist, a child neurologist, and aneuropsychologist. The cognitive evaluation included a generalevaluation with one of the Wechsler batteries depending on age(Wechsler Preschool and Primary Scale of Intelligence–III: 2years 9 months to 7 years; Wechsler Intelligence Scale for Chil-dren–IV: 6 to 16 years; or Wechsler Adult Intelligence Scale–III:�16 years). The Brunet-Lezine scale or Bayley Scales of InfantDevelopment were used for the youngest children and thePsycho-Educational Profile when the Wechsler batteries werenot valid (IQ �40); the results of these tests are reported asdevelopmental age.

Brain imaging. MRI scans were available for all index patientsand their living affected siblings except members of family 11(who underwent CT). All MRI scans were reviewed by a pediat-ric neuroradiologist (M.E.). Gyration was compared to age-matched controls. The gyral pattern was considered simplifiedwhen the number of gyri was reduced, the sulci were very shal-low (less than half the normal depth) on several MRI sections(frontal, parietal and occipital coronal sections, and axial sec-tions), and no tertiary gyri were visible.

RESULTS Mutation screening of the entire cohort.We enrolled 52 unrelated patients. All of them wereconfirmed eligible, included in the study, and ana-lyzed for ASPM sequencing. Among them, 9 (21%)had siblings with microcephaly. Parental consan-guinity was noted in 13 patients (9 sporadic and 4multiplex cases). In 5 nonconsanguineous families,more than one individual was affected. All patientshad had CNS imaging (usually MRI and occasion-ally CT). Four consanguineous and/or multiplexfamilies were linked to MCPH5. They were shown toharbor ASPM mutations. Four multiplex familieswere potentially linked to another MCPH locus butnot to ASPM. One family was not linked to anyknown MCPH locus.

We found ASPM mutations in 11 unrelated pa-tients, who harbored 2 known and 16 previously un-reported mutations (table 1). All mutations were

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predicted to result in absence or truncation of theASPM protein: 1 was a large gene deletion, 10were small out-frame deletions, and 7 were stopcodon mutations. The mutations were homozy-gous in 4 patients. Parental heterozygosity waschecked in these patients. Consanguinity was doc-umented in 2 of the 4 homozygous patients. Anal-ysis of microsatellites flanking the MCPH5 locusin the 2 other homozygous patients showed ho-mozygous haplotypes in the vicinity of theMCPH5 locus, suggesting remote consanguinity(data not shown).

Phenotypic spectrum of patients with ASPM muta-

tions. Pedigree data. We studied the 11 index patientsand 5 affected siblings. Two patients each had oneaffected sibling, who was evaluated. The other 3 af-fected siblings belonged to the same family (family11); the 4 patients in this family were unavailable forMRI or neuropsychological evaluation. In anotherfamily, MCPH recurrence was suspected during thethird trimester of pregnancy based on sonogramfindings and confirmed by fetal brain MRI at 32

weeks’ gestation; the pregnancy was terminated. Noinformation was available on the biologic family ofan adopted child.

Clinical data. Clinical data are detailed in table 2.OFC. OFC ranged from �3.5 SD to �9 SD at

study inclusion (mean, �6.7 SD). OFC at birthranged from �2 SD to �6 SD (mean, �3.6 SD)and was between �2 and �3 SD in 4 probands.OFC decreased with age in all patients; the meandecline for 12 patients was �0.36 SD/y (SEM,0.25).

Height. Four probands had short stature at birth(between �2 SD and �3 SD). Among them, 3achieved normal height (��2 SD) within a fewmonths after birth and 1 was still short (�3 SD) at 3years of age; his parents were of normal height(mother �1.5 SD and father �2 SD).

Extraneurologic abnormalities. One patient had unilat-eral multicystic kidney. One girl had precocious pu-berty onset at 8 years of age.

Dysmorphic features. The typical dysmorphism (nar-row bitemporal distance, sloping forehead, oxyceph-aly) was noted in the patients with the lowest OFC

Table 1 ASPM gene mutations

Family

Geographicorigin ofparents Consanguinity

Patientno. (sex) Exon ASPM mutation (nt)

Predicted proteinfull-length mRNA

LargestpredictedASPMprotein size

Patientgroup

1 Algeria Yes 1-1 (M) 18 �c.7782_7783delGA� homozygous p.Lys2595fsX6 2,595 2

1-2 (M) 1

2 France No 2-1 (M) 1 �c.77delG� � p.Gly26AlafsX42* 2,077 2

18 �c. 6232 T�C� p.Arg2078Stop*

3 France Likely* 3-1 (M) 18 �c.6651_6654delAACA� homozygous p.thr2218TyrfsX8* 2,217 2

4 France No 4-1 (M) 18 �c. 8190_8191delAG� � p.Glu2731LysfsX18* 2,757 3

18 �c.8273 T�A� p. Leu2758Stop*

5 France No 5-1 (F) 3 �c.1630_1634delATCTT� � �deletionof ASPM gene�

p.Tyr544SerfsX9* 543 3

6 Germany No 6-1 (M) 17 �c.3945_3946delAG� � p.Arg1315SerfsX2* 2,731 3

18 �c.8191_8194delGAAA� p.Arg2732LysfsX4*

7 France No 7-1 (M) 22 �c.9319 C�T� � p.Arg3107Stop* 3,169 3

7-2 (M) 23 �c.9507delG� p.Ile3170LeufsX9* 3

8 Reunion Island Likely* 8-1 (F) 16 �c.3811 C�T� homozygous p.Arg1271Stop 1,270 3

9 ? (Morocco, adopted) No 9-1 (F) 6 �c.2389 C�T� � p.Arg797Stop* 1,357 3

17 �c. 4074 G�A� p.Trp1358Stop*

10 Lebanon/Algeria No 10-1 6 �c.2389 C�T� � p.Arg797Stop* 2,228 3

18 �c.6686_6689delGAAA� p.Arg2229ThrfsX10*

11 Lebanon Yes 11-1 (M) 24 �c.9686_9690delTTAAA�homozygous

p.Ile3229SerfsX10* 3,228 3

11-3 (M) 3

11-2 (F) 3

11-4 (M) 3

*Remote consanguinity inferred from common haplotype shared for both mutations.

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but not in patients with less severe decreases in brainsize.

Neurologic and neurodevelopmental data. Neurologicand neurodevelopmental data are detailed in table 3.

All patients but one had normal neurologic findings.One patient (10-1) had pyramidal syndrome in thelower limbs with spasticity and tiptoe walking. Epilepsywas noted in 3 patients. One patient (11-3) experiencedthe onset of poorly controlled, generalized, tonic-clonicepilepsy at 8 years of age. Her brother (patient 11-2)had onset of generalized tonic-clonic epilepsy at 18months of age and responded to topiramate, with onlyvery few residual seizures. The third patient (7-2) hadone generalized tonic-clonic seizure at 14 years of ageand a second one 3 years later, with no further recur-rences under valproate therapy.

All patients had normal motor development until1 year of age. Walking was mildly delayed: 3 patientswalked between 18 and 24 months of age and 4 be-tween 2 and 4 years of age. The first words wererecorded after 3 years of age in all but 2 patients.Sleep disturbances were noted for 2 young patients.Before 6 years of age, most of the patients were hy-perkinetic and many exhibited aggressive behaviortoward themselves and others. From 8 years of age toadulthood, attention deficit was predominant,whereas the hyperactivity abated. Cognitive develop-ment ranged from borderline-normal intelligence

(IQ 70–80) to severe mental retardation. Six pa-tients (1-1, 1-2, 3-1, 4-1, 7-1, and 7-2) were able toread, write, and count. Patient 2-2 had a job in anon-sheltered environment and patients 7-1 and 7-2in a sheltered environment.

CNS imaging. Brain MRI scans were available for12 patients (10 probands and 2 siblings) (table 4 andfigure). The remaining 4 patients, all from family 11,were investigated only by CT scan. Among the 12patients who underwent MRI, 3 (8-1, 9-1, and 10-1)had partial agenesis of the corpus callosum. The lat-eral ventricles were enlarged (colpocephaly) and dys-morphic in 7 patients including a pair of siblings(3-1, 5-1, 6-1, 7-1, 7-2, 9-1, and 10-1). The 2 sib-lings in family 7 had infratentorial abnormalities:mild, asymmetric cerebellar hypoplasia in patient 7-1and asymmetric pons hypoplasia in patient 7-2. Pa-tient 7-2 had extensive unilateral perisylvian polymi-crogyria and patient 10-1 had focal parietal corticaldysplasia. A simplified gyration pattern was noted in9 of the 12 patients and was concordant amongsiblings.

DISCUSSION We describe 16 patients from 11 fami-lies with MCPH and ASPM gene mutations. MCPHwas formerly called MV (true microcephaly). The termMV is now used in a more restrictive sense for nonsyn-dromal congenital microcephaly with decreased brain

Table 2 Clinical data of patients with ASPM gene mutations

Patientno. (sex)

OFC atbirth(SD)

Weight atbirth (SD)

Lengthat birth(SD)

Clinicalfindings

Age atlastfollow-up, y

LastOFC (SD)

Lastweight(SD)

Lastheight(SD)

Neurologicexamination Epilepsy

Patientgroup

1-1 (M) �2 M M 14 �4 �1 M N 2

1-2 (M) �5 2.95 49 24 �7 �1.5 �2 N 1

2-1 (M) �2 �2 �2.5 6 �5 �2 �1 N 2

3-1 (M) �2.5 �1.5 �2 13 �3.5 M �1 N 2

4-1 (M) �2 M �1.5 Unilateralmulticystickidney

9 �4.5 M �1 N 3

5-1 (F) �3.5 �1.5 NA Earlypuberty

13 �7 �1 �1.5 N 3

6-1 (M) �4 �1 �2 3.5 �7 �2 �3 N 3

7-1 (M) �4 �1 M 20 �6 �1.5 M N 3

7-2 (M) �4 �0.5 M 18 �5 �1.5 �0.5 N � 3

8-1 (F) �5 �1 �2 3 �8 �2.5 �2 N 3

9-1 (F) �5 M �1 7 �9 �1 M N 3

10-1 (M) �6 �1 �2 3 �7 M M Pyramidalsyndrome

3

11-1 (M) NA NA NA 24 �7 �2 N 3

11-3 (M) NA NA NA 20 �8 �0.5 N � 3

11-2 (F) NA NA NA 25 �7 �1 N � 3

11-4 (M) NA NA NA �40 NA NA N 3

OFC � occipital-frontal circumference; M � median; NA � not available.

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size but a roughly normal gyral pattern by MRI.9 MSGhas usually been considered different from MV and hasa different OMIM number (MIM 603802). In MSG,the brain has a reduced number of gyri with shallowsulci (usually less then half the normal depth), the ter-tiary sulci are absent, and the cortex is normal or thin.10

MSG has been tentatively subdivided into several sub-groups based on radiologic and clinical criteria atbirth.11 MSG is distinct from microlissencephaly, inwhich the simplified gyral pattern is associated with athick (lissencephalic) cortex.12

The phenotype of our patients appeared to bemore heterogeneous than previously described: somepatients had low-normal OFC at birth followed by a

decrease with age, and some had IQ values in the70–80 range. Furthermore, we confirmed MSG tobelong to the ASPM spectrum, and showed for thefirst time that loss of ASPM function can be associ-ated with cortical defects.

We obtained reliable birth OFC data from 10families. Although all patients had microcephaly atbirth, some of them (patients 1-1, 2-1, 3-1, and 4-1)had OFC values between �2 and �3 SD. In all pa-tients, postnatal head growth was slow, with a declinein the OFC Z score to less than �3 SD during thefirst year of life. Therefore, the definition ofMCPH should include patients with mild micro-cephaly at birth, and OFC values should be moni-

Table 3 Clinical and developmental data of patients with ASPM gene mutations

Patientno. (sex)

Age atwalking

Age atfirstwords, y Behavior

Sleepdisturbance

Attentiondeficit

Age atclinicalevaluation

Total IQ/DQ(test) Subtests*

Patientgroup

1-1 (M) 10 mo �2 Normal �� 13 y 8 Heterogeneousscale(WISC-IV)

76/61/62/78 2

1-2 (M) 12 mo �3 Normal �� 24 y 53 (WAIS-III) 55/55/53/50 1

2-1 (M) 20 mo 4 Hyperactivitydisorder (3–6 y),aggressiveness,self-mutilation

6 y 70 (BL-R) 4;1/5/5/4;2 2

3-1 (M) 13 mo 3 Hyperactivitydisorder,withdrawn oropposing (3–6 y)

�� 13 y 9 43 (WISC-IV) 47/47/56/69 2

4-1 (M) 13 mo 3 Hyperactivitydisorder (4–6 y),quiet �6 y

� NA NA NA 3

5-1 (F) 20 mo 4 Hyperactivitydisorder(4–12 y)

12 y 30 (PEP-R) 5/5;4/4;2/3;10 3

6-1 (M) 15 mo 2.5 Normal � 32 mo 104 (BSID-II) 34m/32m 3

7-1 (M) 13 mo 2.5 Hyperactivitydisorder (�6 y),apathetic (�8 y)

�� 20 y 55 (WAIS-III) 52/63/50/50 3

7-2 (M) 13 mo 3 �� 17 y 11 46 (WAIS-III) 49/47/50/50 3

8-1 (F) 17 mo Notacquired

Hyperactivitydisorder

��� Notassessable

Not assessableat age 3 y

3

9-1 (F) 25 mo 4 Hyperactivitydisorder (4–6 y)

6 y 36 (PEP-R) 3/2;4/2;2/2;3 3

10-1 (M) 14 mo Notacquired

Hyperactivitydisorder

��� 3 y 11 42 (BL-R) 2;3/1;9/1;7/1;6 3

11-1 (M) 4 y 3.5 NA NA Notassessable

Notassessable

3

11-3 (M) 4 y 4 NA NA Notassessable

Notassessable

3

11-2 (F) 3 y 3.5 NA NA Notassessable

Notassessable

3

11-4 (M) 3.5 y 3.5 NA NA Notassessable

Notassessable

3

Brunet-Lezine revised scale (BL-R): global motor development/hand-eye coordination/language/social behavior (years; months of development). Psycho-Educational Profile (PEP-R): global motor development/hand-eye coordination/language/cognitive performance (years; months of development).*Subtests of Wechsler batteries: Wechsler Intelligence Scale for Children (WISC)–IV and/or Wechsler Adult Intelligence Scale (WAIS)–III � verbal compre-hension/perceptual reasoning/working memory/processing speed index.BSID-II � Bayley Scales of Infant Development; NA � not available.

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tored closely over time. The gradual OFC declinenoted in our study has not been highlighted previ-ously. Future studies should include serial OFCmeasurements.

Of our 16 patients, 3 had nonspecific seizure dis-orders that started after infancy, in keeping with ear-lier data. Epilepsy is a feature in up to 10%–20% ofpatients with MCPH.2,6 One of our patients hasmarked pyramidal tract involvement of the lowerlimbs, which would have excluded a diagnosis ofMCPH according to the classic definition.

Although MR is a constant feature of MCPH, IQvalues have been published only for 2 cases.7,8 Neuro-psychological testing showed that IQ values variedwidely, from 30 to 70–80. Subscores were consistentwithin patients. Thus, MCPH is compatible withborderline-normal development and IQ values be-tween 70 and 80, in keeping with recent findingsregarding PERICENTRIN13,14 and MCPH1 muta-tions.15 Hyperactivity and attention deficit were ma-jor childhood problems in all patients and possiblycaused impairments in performance. Intrafamilialdiscordance was noted in family 1, with one siblinghaving an IQ of 53 and the other of 70. In this fam-ily, the younger patient, who had the higher IQ, re-ceived early educational interventions that had notbeen available for the older brother in his country of

origin. These early interventions may have contrib-uted to the higher IQ, although the OFC Z score wasalso better in the younger brother.

A simplified gyration pattern has been noted in 5patients from the literature, including 2 siblings,1,6,7

and frontal predominance of brain underdevelop-ment was suggested in one family.7 Our study pointsup that MV and MSG type 1 constitute a radiologiccontinuum in patients with ASPM mutations. Al-though marked gyral simplification is readily de-tected, quantification of gyration on plain MRI scansremains subjective, as no simple gyration index isavailable. We used a previously published definitionof MSG9,10,16 to assess the gyral pattern in our pa-tients. As recently noted,7 most patients showedmore severe underdevelopment of the frontal lobes,but patient 2-1 had a reverse pattern, with more se-vere underdevelopment of the occipital lobes. Wefound gyral simplification in 9 of the 12 patients whounderwent MRI. This high prevalence of gyral sim-plification in MCPH was unexpected. Siblings hadsimilar degrees of gyration, even when their OFC Zscores differed. MSG did not seem associated withlower OFC values at birth or later on, or with worseMR. Enlarged and dysmorphic ventricles were com-mon, as was partial agenesis of the corpus callosum.Two siblings had mild asymmetric abnormalities of

Table 4 MRI findings in patients with ASPM gene mutations

Patient no.Corpuscallosum Ventricles

Cerebellumand brainstem Gyration pattern

Patientgroup

1-1 (M) Normal Normal Normal Simplification of the frontaland occipital gyral pattern

2

1-2 (M) Thick CC Normal Normal Simplification of the frontal andoccipital gyral pattern

1

2-1 (M) Normal Normal Normal Normal gyral pattern 2

3-1 (M) Normal Occipital horns of theLV enlarged

Normal Simplification of the frontal andoccipital gyral pattern

2

4-1 (M) Normal Normal Normal Normal gyral pattern 3

5-1 (F) Normal Occipital horns ofthe LV enlarged

Normal Simplified frontal gyral pattern 3

6-1 (M) Normal Occipital horns of the LVenlarged; dysmorphic frontalventricles

Normal Simplified frontal gyral pattern 3

7-1 (M) Normal Occipital horns of the LVenlarged; dysmorphic frontalventricles.

Mild asymmetriccerebellar hypoplasia

Simplified frontal and occipitalgyral pattern

3

7-2 (M) Normal LV enlarged; dysmorphicfrontal ventricles.

Ipsilateral ponshypoplasia

Extensive unilateral perisylvianpolymicrogyria from the frontalpole to the occipital pole;contralateral simplified frontal andoccipital gyral pattern

3

8-1 (F) Agenesis ofCC splenium

Normal Normal Normal gyral pattern (at 1 month) 3

9-1 (F) Agenesis of CCrostrum; largepituitary

LV enlarged Normal Coarse gyri, simplified frontaland occipital gyral pattern

3

10-1 (M) Agenesis ofCC rostrum

Occipital horns ofthe LV enlarged

Normal Simplified gyral pattern posteriorfocal parietal cortical dysplasia

3

CC � corpus callosum.

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brainstem and cerebellum size without neurologicimpairment. More surprisingly, 2 patients had corti-cal dysgenesis: patient 7-2 exhibited a large area ofpolymicrogyria extending from the frontal pole tothe occipital pole, and patient 10-1 had focal corticaldysplasia. Cortical dysgenesis has not been reportedpreviously in MCPH (possibly because this findingwould have led to patient exclusion).

The ASPM protein is nuclear or centrosomal dur-ing the interphase in Hela and HT1080 cells.17 It isredistributed to the poles of the mitotic spindle in adynein-dynactin-dependent manner early in mitosis,suggesting that microcephaly in patients with ASPMmutations may be chiefly due to impaired mitoticspindle regulation in cortical progenitors.17-21 TheCNS anomalies found in our patients suggest thatASPM may be involved not only in mitosis control,but also as an accessory factor in other developmentalprocesses such as migration and cortical layering.

In all, 84 ASPM mutations have been identifiedto date.2,15,22-24 Most of them result in a prematurestop codon, although a few are large deletions. ASPMmRNA is not subject to nonsense-mediated RNAdecay. Kouprina et al17 showed that truncated ASPMprotein was expressed in a lymphoblast cell line de-rived from a patient with homozygous frameshiftmutation in ASPM exon 24. To date, subcellular lo-calization of ASPM truncated protein is not known.Reported mutations are spread throughout theASPM gene and are predicted to result in truncatedproducts ranging in size from 116 (R116X) to 3,357amino acids (K3328fsX29). Two major translatedisoforms of ASPM are known: a full-length mRNAof 10,434 nt encoding a 3,477-amino acid proteinand an alternatively spliced mRNA lacking exon 18and encoding a 1,892-amino acid protein. Twoother weakly expressed isoforms have been de-scribed.17 Patients with at least 1 mutation in exon18 are still able to synthesize 1 of the 2 wild typeisoforms, i.e., the exon-18 spliced mRNA. If this iso-form exerts physiologic effects, then patients still ca-pable of producing it may have a milder phenotype.In our series, the mean IQ value was 64 for patientswho had at least 1 mutation in exon 18 vs 42 forpatients whose mutations were both located outsideexon 18 (p � 0.13, Mann-Whitney U test). In anearlier study,25 severe MR was reported only in pa-tients with mutations outside exon 18. In contradic-tion with this hypothesis, moderate MR (IQ � 50)was reported in a girl with 2 exon 18 mutations andmild MR in children with homozygous mutationssparing exon 18.5,8 In these last families, IQ valueswere not available (the reported “social quotient” wasobtained using the Vineland scale, which does notstrictly evaluate cognitive abilities8). The trend ob-

Figure MRI findings in patients with ASPM gene mutations compared toage-matched control

(A) Sagittal T1-weighted image showing the typical MRI features of ASPM-related primary he-reditary microcephaly (MCPH), with poorly developed frontal lobes in patients 7-1, 7-2, 1-1,and 1-2 compared to age-matched controls (a). Note cerebellar hypoplasia in patient 7-1. (B)Axial T1-weighted images showing the typical appearance of ASPM-related MCPH. The gyriare scarce and the sulci very shallow compared to age-matched controls (a). No tertiary gyri arevisible. Colpocephaly is common (patients 7-1 and 7-2). Note unilateral perisylvian polymicro-gyria extending to the occipital pole (short white arrows) in patient 7-2. (C) Coronal T1- andT2-weighted images showing marked simplification of the gyration pattern (patients 7-1, 7-2,1-1, and 1-2), a common finding in ASPM-related MCPH. The number of gyri is reduced and thesulci are shallow compared to the age-matched controls (a).

968 Neurology 73 September 22, 2009 at Universitaet Heidelberg on October 1, 2009 www.neurology.orgDownloaded from

served in our small sample may nevertheless indicatethat exon 18 mutations convey a lower risk of severeMR. To confirm or reject this hypothesis would re-quire a larger number of patients. Confirmation ofthe lower risk would argue in favor of a physiologicrole for the shorter ASPM isoform.

ACKNOWLEDGMENTThe authors thank the families for their patience and contribution to this

study and Christelle Desire for technical skills in DNA sequencing.

DISCLOSUREDr. Passemard receives research support from Fondation pour la Recher-

che, Medicale and Fondation Lejeune. Dr. Titomanlio receives research

support from Fondazione Mariani. Drs. Elmaleh, Afenjar, Alessandri, An-

dria, Billette de Villemeur, Boespflug-Tanguy, Burglen, Del Giudice,

Guimiot, Hyon, Isidor, Megarbane, Moog, Odent, Hernandez, Pouv-

reau, Scala, and Schaer report no disclosures. Dr. Gressens serves as editor

of Pediatric Research; receives research support from Servier (France), In-

serm, Paris 7 University, the ELA Foundation (France), Grace de Monaco

Foundation, and Fondation Motrice (France). Drs. Gerard and Verloes

report no disclosures.

Received March 11, 2009. Accepted in final form June 25, 2009.

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DOI: 10.1212/WNL.0b013e3181b8799a 2009;73;962-969 Neurology

Scala, M. Schaer, P. Gressens, B. Gerard and A. Verloes Hyon, B. Isidor, A. Mégarbané, U. Moog, S. Odent, K. Hernandez, N. Pouvreau, I.

C.Billette de Villemeur, O. Boespflug-Tanguy, L. Burglen, E. Del Giudice, F. Guimiot, S. Passemard, L. Titomanlio, M. Elmaleh, A. Afenjar, J. -L. Alessandri, G. Andria, T.

mutationsASPMdue to Expanding the clinical and neuroradiologic phenotype of primary microcephaly

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