Megalencephaly and hemimegalencephaly: Breakthroughs in molecular etiology

American Journal of Medical Genetics Part C (Seminars in Medical Genetics) 166C:156–172 (2014)

A R T I C L E

Megalencephaly and Hemimegalencephaly:Breakthroughs in Molecular EtiologyGHAYDA M. MIRZAA* AND ANNAPURNA PODURI**

Grant sponsThe authorsDr. Ghayda

Research Institsyndromes.

Dr. AnnapuNeurology, whService. She isfamilial and de

*CorresponIntegrative Bra

**CorrespoBoston Childre

DOI 10.100Article first p

� 2014 Wil

Megalencephaly (MEG) is a developmental disorder characterized by brain overgrowth that occurs due to eitherincreased number or size of neurons and glial cells. The former may be due to either increased neuronalproliferation or decreased apoptosis. The degree of brain overgrowthmay be extensive, ranging from generalizedMEG affecting the entire cortex–as with mutations in PTEN (phosphatase and tensin homolog on chromosometen)–to unilateral hemispheric malformations–as in classic hemimegalencephaly (HME). On the other hand,some lesions are more focal or segmental. These developmental brain abnormalities may occur in isolation insome individuals, whereas others occur in the context of a syndrome involving dysmorphic features, skin findings,or other organ system involvement. Brain overgrowth disorders are often associated with malformations ofcortical development, resulting in increased risk of epilepsy, intellectual disability, and autistic features, and someare associated with hydrocephalus. The past few years have witnessed a dramatic leap in our understanding ofthe molecular basis of brain overgrowth, particularly the identification of mosaic (or post-zygotic) mutations incore components of key cellular pathways such as the phosphatidylinositol 3-kinase (PI3K)-vakt murine thymomaviral oncogene homolog (AKT)-mTOR pathway. These molecular insights have broadened our view of brainovergrowth disorders that now appear to span a wide spectrum of overlapping phenotypic, neuroimaging, andneuropathologic features andmolecular pathogenesis. Thesemolecular advances also bring to light the possibilityof pathway-based therapies for these often medically devastating developmental disorders.© 2014 Wiley Periodicals, Inc.

KEYWORDS:megalencephaly; hemimegalencephaly; polymicrogyria; somatic mosaicism; overgrowth; PI3K-AKT-mTOR pathway; Ras/MAPKpathway

How to cite this article: Mirzaa GM, Poduri A. 2014. Megalencephaly and hemimegalencephaly:Breakthroughs in molecular etiology. Am J Med Genet Part C 166C:156–172.

INTRODUCTION

The etiologies of focal malformations ofcortical development have long been apuzzle. Unlike most forms of lissence-phaly and microcephaly where brainmorphology appears affected globally inmost forms, suggesting a genetic disor-der affecting the entire cortex, focalmalformations such as focal corticaldysplasia (FCD), hemimegalencephaly

or: NINDS; Grant number: NS06978have no conflicts of interest to discM. Mirzaa is clinical and molecular gute. Her research interests focus on

rna Poduri is a clinician-scientist at Bere she is the director of the Hospita Co-Investigator of the NINDS-suppnovo causes of early onset epilepsydence to: Ghayda M. Mirzaa, M.Din Research, Seattle Children's Resendence to: Annapurna Poduri, M.Dn's Hospital, 300 Longwood Avenu2/ajmg.c.31401ublished online in Wiley Online Lib

ey Periodicals, Inc.

(HME), and focal megalencephaliessuggest another pattern in which onlypart of the developing brain appears tohave experienced a genetic aberration.Brain overgrowth phenotypes rangefrom very localized lesions to morediffuse multifocal forms. These areconditions were asymmetry is the rule,and where etiology had long beenelusive. Non-genetic etiologies werepreviously sought to explain the pres-

4.lose.eneticist in the Department of Human Genetics at Sethe clinical and molecular spectrum of developm

oston Children's Hospital in the Division of Epilepsyal's Epilepsy Genetics Program and a member of thorted Center without Walls Epi4K. Her research intand brain malformations.

., Department of Pediatrics, Division of Genetic Mearch Institute, 1900 9th Avenue, Seattle, WA 98101., M.P.H., Division of Epilepsy and Clinical Electrophe, Boston, MA 02115. E-mail: annapurna.poduri@c

rary (wileyonlinelibrary.com): 28 May 2014

ence of a severely overgrown anddysplastic portion of the brain with theneighboring cortex seemingly normal.However, one of the earliest and firstrecognized neurogenetic syndromes,tuberous sclerosis complex (TSC), fea-tured just this type of patchy malforma-tion, the cortical tuber. For over adecade, it has been known that germlineloss of function mutations in TSC1and TSC2, causing hyperactivation of

attle Children's Hospital and Seattle Children'sental brain disorders and overgrowth genetic

and Clinical Electrophysiology, Department ofe Translational Research Program Investigatorerests focus on the discovery and modeling of

dicine, University of Washington, Center for. E-mail: [email protected], Department of Neurology, Fegan 9,hildrens.harvard.edu

Megalencephaly (MEG) isclassically defined as an

oversized and overweight brainthat exceeds the age-relatedmean by 2 or more standarddeviations. Clinically, the

distinction betweenmegalencephaly (enlargedbrain) and macrocephaly

(enlarged head overall) relieson neuroimaging examinationof the brain and recognition ofenlarged cerebral structures.

ARTICLE AMERICAN JOURNAL OF MEDICAL GENETICS PART C (SEMINARS IN MEDICAL GENETICS) 157

the mammalian target of rapamycin(mTOR) signaling cascade, are associat-ed with cytomegaly and disorganizedlamination within the cerebral cortex,the hallmark features of TSC. Further-more, loss of function mutations ofPTEN, an upstream phosphatase thatinhibits the PI3K-AKT-mTOR path-way, are well known causes of general-ized megalencephaly (MEG) insyndromic (Cowden and Bannayan–Riley–Ruvalcaba syndromes) and non-syndromic phenotypes (MEG withautism). In this context, the identifica-tion of somatic mosaicism in HME andMEG phenotypes makes sense but hasonly recently come to attention. Mosai-cism, as exemplified by these disorders,provides a new set of mechanismspreviously under-appreciated as causesof neurological disease [Poduri et al.,2013]. Single cell sequencing of corticaltubers and the detection of somaticTSC1/2 mutations was a foreshadowingof our very rapidly improved under-standing of the genetics of these dis-orders [Crino et al., 2010].

In this review, we will highlightgenetic etiologies of brain overgrowthdisorders broadly and then review theclinical spectrum and molecular patho-genesis of a recently emerging class ofbrain overgrowth phenotypes associatedwith mutations in the PI3K-AKT-mTOR pathway. Identification of newgenes is ongoing. However, the biologiceffects of these mutations on brain andbody overgrowth and genotype–pheno-type correlations are still under investi-gation, and we anticipate that these willbe areas of continued discovery in thecoming years.

SYNDROMES AND GENESASSOCIATED WITH BRAINOVERGROWTH: AGENERAL OVERVIEW

Megalencephaly (MEG) is classicallydefined as an oversized and overweightbrain that exceeds the age-related meanby 2 or more standard deviations.Clinically, the distinction between meg-alencephaly (enlarged brain) and macro-cephaly (enlarged head overall) relies onneuroimaging examination of the brain

and recognition of enlarged cerebralstructures. Whereas MEG is associatedwith specific syndromes, macrocephalycan be caused by a myriad of causes suchas hydrocephalus or ventriculomegaly,enlarged extra-axial spaces, and thick-ened skull bones. Therefore, the distinc-tion between MEG and macrocephalyis clinically helpful towards accuratediagnosis.

MEG has long been classified basedon pathogenesis into metabolic andnon-metabolic (or anatomic) subtypes[DeMyer, 1972, 1986]. Cellular hyper-trophy due to cellular edema or accu-mulation of metabolic substrates cancause MEG in a wide range of neuro-metabolic syndromes, such as Canavandisease, glutaric aciduria type I, lyso-somal storage disorders, among others(Table I). A growing number of devel-opmental (or non-metabolic) geneticsyndromes are known to be associatedwith generalized or focal MEG, includ-ing HME. Table II represents a broadoverview of genetic disorders whereMEG is a defining/diagnostic or com-mon feature. Brain overgrowth in thesesyndromes varies widely in severity,distribution and co-occurrence of othermalformations of cortical developmentfrom a mild (and often relatively)enlarged brain with a normal cortex to

bilateral MEG with diffuse corticaldysplasia, as discussed below. Finally,reciprocal copy number changes areknown to be associated with braingrowth dysregulation (i.e., MEG andmicrocephaly) (Table III).

Other important overgrowth con-ditions include Sotos syndrome, Weaversyndrome, Simpson Golabi Behmelsyndrome, and nevoid basal cell carcino-ma syndrome. While they are beyondthe scope of this review, many of thesame principles apply to these disordersas to the conditions that affect the brainpredominantly.

MOLECULAR PATHWAYSOF MEGALENCEPHALYANDHEMIMEGALENCEPHALY

Cellular growth of neuronal elements isan intricately orchestrated process, asdiscussed by Drs. Alcantara and O’Dris-coll in this series. Dysregulation of anumber of critical pathways is known tobe associated with human brain over-growth phenotypes, as highlighted inTable II. Dysregulation of two particularcritical cellular pathways, the Ras/mitogen-activated protein kinase(MAPK) pathway and the PI3K-AKT-mTOR pathway, appear to account forthe largest number of known MEG/HME syndromes. Both pathways areassociated with multiple diverse cellularfunctions including cellular prolifera-tion, differentiation, cell cycle regula-tion, survival, and metabolism. Notsurprisingly, these two pathways arefunctionally related. Given their criticaldevelopmental roles, germline muta-tions of genes in both pathways arebelieved to be embryonic lethal. Whendysregulated, regardless of the specificgene or protein alteration, the ensuingsyndromes exhibit numerous overlap-ping phenotypic features spanning manyorgan systems. Furthermore, both path-ways have been extensively studied inthe cancer field and constitute veryattractive targets for pathway (smallmolecule) inhibitor therapy to treatvarious malignancies.

Whereas most mutations in theRASopathies are germline, the emerging

TABLE I. Genes, Clinical Features, and Metabolic Abnormalities of Neurometabolic Syndromes Associated with MEG

Syndrome Gene Clinical features Neuroimaging findings Metabolic abnormalities

Cerebral organic acid disorders and disorders of lysine metabolism

N-Acetylaspartic

aciduria

(Canavan disease)a

ASPA Progressive severe ID, SZ,

OA, spasticity,

opisthotonus

Diffuse symmetric WM

abnormalities

#Aspartoacylase (ASPA),

"N-acetyl-aspartic acid

(NAA)

Glutaric aciduria (GA)

type IaGCDH Neonatal MAC, ID,

dyskinesia,

choreoathetosis,

dystonia

Frontotemporal atrophy

(95%), delayed

myelination, high signal

intensity in the dentate

nucleus, subdural

effusion/hemorrhage

#Glutaryl-CoA dehydrogenase,

"Glutaryl-CoA,"Acylcarnitines:freecarnitine, "Urinary

dicarboxylic acids

L-2-Hydroxyglutaric

aciduria

L2HGDH Progressive MAC (50%),

ID, SZ, extrapyramidal

signs

Swollen subcortical WM,

progressive loss of

arcuate fibers, severe

cerebellar atrophy, signal

intensities in the dentate

nuclei and globi pallidi,

low signal intensities in

the thalami

#L-2-hydroxyglutaratedehydrogenase, "L-2-

hydroxyglutaric acid (CSF>

plasma),

"hydroxydicarboxylic acids(CSF), "Lysine (CSF, blood)

D-2-Hydroxygylatric

aciduria

D2HGDH Neonatal epileptic

encephalopathy with

severe ID, hypotonia,

CM to mild DD/no

symptoms

Delayed and abnormal

gyration, myelination

and opercularization,

VMEG, cysts over head

of the caudate nucleus

#D-2-hydroxyglutaric acid

dehydrogenase, "D-2-

hydroxyglutaric acid

Lysosomal storage diseasesaDisorders of Sphingolipid Metabolism

Generalized

gangliosidosis

GM1 (early

infantile)a

GLB1 ID, HSM, SZ, tone

abnormalities, DYSM,

HSM, macular cherry

red spot

Diffuse hypomyelination,

mild T2 hyperintensities

of the caudate nucleus

and putamen

#b-galactosidase, "GM1

ganglioside, asialo-GA1

(neurons), "oligosaccharide,minor glycolipids,

glycopeptides (visceral

organs)

GM2 gangliosidosis

Tay-Sachs disease

(infantile)aHEXA Hypotonia, motor

weakness, SZ,

hyperacusis, macular

cherry red spot,

blindness, spasticity,

MAC by 18 months of

age

Similar to GM1 #Hexosaminidase A, "GM2-

ganglioside (neurons)

Sandhoff diseasea HEXB Organomegaly and bony

abnormalities less

common

Similar to GM1 #Hexosaminidase A and B,

"GM2-ganglioside, asialo-

GM2 (neurons),

"Globosides,oligosaccharides (viscera)

Krabbe disease (globoid

cell leukodystrophy)

(early infantile)a

GALC PN, opisthotonus, SZ,

hyperpyrexia, blindness,

loss of bulbar functions,

hypotonia

Diffuse WM

abnormalities, diffuse

cerebral atrophy,

calcifications (thalamus,

BG, periventricular

WM)

#Galactosylceramidase,

"Galactosylceramide

(globoid cells),

"Galactosylphingosine(oligodendrocytes, Schwann

cells)

158 AMERICAN JOURNAL OF MEDICAL GENETICS PART C (SEMINARS IN MEDICAL GENETICS) ARTICLE

TABLE (Continued )

Syndrome Gene Clinical features Neuroimaging findings Metabolic abnormalities

Mucopolysaccharidoses (MPS)

Hurler syndrome

(type IH)

IDUA HSM, CNS, DM, DYS,

OPH, CAR

WM abnormalities,

cerebral atrophy,

cervical myelopathy

#Iduronidase, "Heparan sulfate,

"Dermatan sulfate

Hunter syndrome

(type II)

IDS HSM, CNS, DM, DYS,

OPH, CAR, SK

WM abnormalities,

cerebral atrophy,

cervical myelopathy

#Iduronate-2-sulfatase,"Heparin sulfate,

"Dermatan sulfate

Sanfilippo syndrome

(type III)

SGSH(IIIA), NAGLU

(IIIB),

HGSNAT(IIIC), GNS

(IIID)

CNS, DM (þ/�),

DYS (þ/�)

WM abnormalities,

cerebral atrophy,

cervical myelopathy

#Heparan N- sulfatase (IIIA),

#N-acetyl-glucosaminidase

(IIIB), #Acetyl CoAglucosamine N-acetyl

transferase (IIIC),

#N-acetyl-glucosamine-6-

sulfatase (IIID), "Heparan

sulfate

Morquio syndrome

(type IV]

GALNS(IVA), GLB1(IVB) DM, CAR, OPH (þ/�) WM abnormalities,

cerebral atrophy,

cervical myelopathy

#N-acetylgalactosamine-6-

sulfatase (IVA),

#b-galactosidase (IVB),

"Keratan sulfate

Maroteaux-Lamy

syndrome

(type VI)

ARSB HSM, DM, DYS, OPH,

CAR

WM abnormalities,

cerebral atrophy,

cervical myelopathy

#N-acetyl-galactosamine-4-

sulfatase, "Dermatan sulfate

Mucolipidosesb

Mucolipidosis type II

(I-cell disease)

GNPTAB HSM, CNS, DM, DYS,

OPH, CAR

Cerebral atrophy, WM

abnormalities

(occasionally)

#Transferasec

Mucolipidosis type III GNPTAB (a/b), GNPTG

(g)

HSM (þ/�), CNS (þ/�),

DM, DYS (þ/�), CAR

Cerebral atrophy, WM

abnormalities

(occasionally)

#Transferasec

Mannosidosis MAN2B1 (a), MANBA

(b)

HSM, DM, DYS, CAR,

CNS (þ/�)

Partially empty sella

turcica, cerebellar

atrophy, WM

abnormalities (a)

#a-mannosidase (a),

"a-mannosides (a),

#b-mannosidase (b),

"b-mannosides (b)

Leukoencephalopathiesa

aAlexander disease

(infantile and juvenile

forms)

GFAP ID, SZ, paraparesis, feeding

problems

WM abnormalities

(frontally-predominant),

calcification of the BG,

cerebellar changes,

HYD

—

Megalencephalic

leukoencephalopathy

with subcortical cysts

MLC1, HEPACAM Progressive spasticity, ataxia Extensive symmetric, WM

changes with subcortical

cyst

—

BG, basal ganglia; CAR, cardiovascular involvement; CC, corpus callosum; CM, cardiomyopathy; CNS, central nervous system regression;ID, developmental delay; DM, dysostosis multiplex; DYS, dysmorphic features; HL, hearing loss; HSM, hepatosplenomegaly; MAC,macrocephaly; OA, optic atrophy; OPH, ocular anomalies [corneal clouding, ophthalmoplegia]; PN, peripheral neuropathy; SK,dermatological findings; SZ, seizures; VMEG, ventriculomegaly; WM, white matter.Inheritance: Most of these disorders are AR in inheritance with the exception of Hunter syndrome (XL), Alexander disease (AD), andmegalencephalic leukoencephalopathy with subcortical cysts due to HEPACAM mutations.aOther leukoencephalopathies associated with megalencephaly (as indicated in the table): Canavan disease, glutaric aciduria type I, infantilegeneralized, or GM1, gangliosidosis, infantile GM2 gangliosidosis (Tay-Sachs; Sandhoff diseases), infantile Krabbe disease.bMay not be true MAC.cLysosomal UDP-N-acetylglucosamine-I-phosphotransferase.Reference: Mirzaa et al. [2012].

I.


TABLEII.Gen

es,Syn

dromes,an

dPathwaysAssociated

withMeg

alen

cephalyan

dHem

imeg

alen

cephaly

Gene

Proteinfunctio

nInheritance

Synd

rome

Clin

icalfeatures

Neurologicfin

dings

MRIfin

dings

PI3K

-AKT-MTOR

PTEN

Pho

sphatase,tumor

suppressor

Deno

vo/dom

inant

Megalenceph

aly-

autism

synd

rome

Mild

DYSM

(fron

talbo

ssing,

midface

hypo

plasia,biparietal

narrow

ing)

ASD

,ID

MEG

Cow

densynd

rome

Mucocutaneous

lesio

ns,malignancy

risk

(breast,thyroid,

endo

metrium

)

ID(10%

)Cerebellardysplastic

ganglio

cytoma

(Lherm

itte-Duclos

disease)

Bannayan–

Riley–

Ruvalcaba

synd

rome

Overgrowth,hamartomatou

sintestinalpo

lypo

sis,lipom

as,

penile

pigm

entedmacules,

malignancyrisk

similarto

CS

Autisticfeatures,ID

(70%

),SZ

(25%

),proxim

almyopathy

(60%

)

—

HME

Macroceph

aly(asymmetric;

maybe

seen

inHME)

ID(severe),SZ

(intractable),

hemiparesis

HME:VMEG,MCD,

WM

abno

rmalities

(ipsilateral)

PIK3C

AKinase

Postzygotic/m

osaic

(raregerm

line)

MCAPsynd

rome

MEG,capillarymalform

ations,d

igit

anom

alies(polydactyly,

synd

actyly),segm

entalsomatic

overgrow

th,conn

ectivetissue/

skin

laxity

ID,SZ

,hypo

tonia

(variable)

HYD,VMEG,CBTE,

PMG,thickCC

HME

Asabove

Asabove

Asabove

Somaticovergrow

th:

CLO

VES,

Fibroadipo

sehypo

plasia,Isolated

macrodactyly

Variable

segm

entalsomatic

overgrow

th,digitalanom

alies,

spinalanom

alies,cutaneou

svascular

malform

ations;also

isolatedmacrodactylyin

some

cases

Somehave

IDHMEandChiari

malform

ation

repo

rted

insome

individu

als

Klippel–Trenaun

aysynd

rome(KTS)

Cutaneous

VM

(capillary,veno

us,

lymph

atic),varicose

veins,

unilateralhypertrophyof

bones

andsofttissues

ID/SZ

(rare)

HYD,calcificatio

ns,

HMErepo

rted

insomeindividu

als

PIK3R

2Kinase

Deno

vo/dom

inant

MPP

Hsynd

rome

Postaxialpo

lydactyly,MEG

ID,epilepsy,tone

abno

rmalities

MEG,perisylvian

PMG,HYD,mega

CC


TABLE

(Continued)

Gene

Proteinfunctio

nInheritance

Synd

rome

Clin

icalfeatures

Neurologicfin

dings

MRIfin

dings

AKT1

Kinase

Post-zygotic/

mosaic

Proteussynd

rome

(associatedwith

HME)

Asymmetricanddispropo

rtionate

hamartomatou

sovergrow

thof

multip

letissues,conn

ectivetissue

andepidermalnevi,dysregulated

adiposetissue,

VM,hyperostosis

ID(20%

),SZ

(13%

)Calcifications,

abno

rmalities

ofthe

CC,HYD,HME

repo

rted

insome

individu

als

AKT3

Kinase

Deno

vo/dom

inant

MPP

HAsabove

Asabove

Asabove

HME

Asabove

Asabove

Asabove

STRADA/

LYK5

STE20-related

kinase

adaptor

(“pseudo

kinase”)

Recessive

Polyhydram

nios,

MEG,symptom

atic

epilepsy(PMSE

)synd

rome

DYSM

,strabism

us,skeletalmuscle

hypo

plasia,neph

rocalcinosis

ID,hypo

tonia,SZ

,ASD

VEMG

(mild),

subepend

ymal

dysplasia,WM

abno

rmalities

TSC

1,TSC

2Tu

mor

suppressor

Deno

vo/dom

inant

(som

aticmosaicism

described)

Tuberous

sclerosis

complex

(TSC

)(associatedwith

HME,FC

D)

Skin

(hypom

elanoticmacules,facial

angiofibromas,shagreen

patches,

fibrous

facialplaques,un

gal

fibromas),angiom

yolipom

as,

rhabdo

myomas

SZ(80%

),ID

(50%

),ASD

/PDD

(40–

50%),ADHD

SEN,corticaltubers,

SEGAs,WM

abno

rmalities,

HME/FCD

TBC1D

7GTPase-R

HEB

Recessive

ID,macroceph

aly,

patellardislo

catio

n,celiacdisease

Osteoarticular

prob

lems,celiac

disease,

myopia,astig

matism

ID(m

ild),behavioral

abno

rmalities,LD

Cerebralcalcificatio

ns

MTOR

Kinase

Post-zygotic/

mosaic

HME

Asabove

Asabove

Asabove

CCND2

Cyclin

;cellcycle

control

Deno

vo/dom

inant

MPP

HAsabove

Asabove

Asabove

Ras/m

itogen-activated

proteinkinase

(MAPK

)pathway

“theRASo

pathies”

(associatedwith

absoluteor

relativemacrocephaly)

NF1

RasGAP

Deno

vo/dom

inant

Neurofib

romatosis1

CALs,axillaryfreckling,

cutaneou

sneurofibromas,shortstature

LD(50-75%),(severe

ID3%

),ADHD,

headaches(20%

),SZ

(10%

)

Optic

glioma(15%

),UBOs,CC

abno

rmalities,HYD

SPRED1

Sprouty-related

Deno

vo/dom

inant

Legius

synd

rome

CALs,freckling,

lipom

as,

macroceph

aly,no

tumor

manifestations

ID/LD,ADHD,

headaches,SZ

—

HRAS

GTPase

Deno

vo/dom

inant

Costello

synd

rome

FTT,

shortstature,

coarse

facial

features,fine,curly

orsparse

hair,

papillomata,HCM,CHD,

malignancyrisk

(15%

)

ID(�

100%

),hypo

tonia(m

ost),

SZ(20-50%)

CBTE,VMEG/H

YD

II.


TABLE

(Continued)

Gene

Proteinfunctio

nInheritance

Synd

rome

Clin

icalfeatures

Neurologicfin

dings

MRIfin

dings

BRAF

Kinase

Deno

vo/dom

inant

Cardiofaciocutaneou

s(C

FC)synd

rome

Cardiac

abno

rmalities

(VHD,

HCM,dysrhythmias),DYSM

,multip

lecutaneou

sabno

rmalities

ID(80%

),SZ

(50%

),hypo

tonia

HYD/V

MEG,cortical

atrophy,ACC,N

MD

Noo

nansynd

rome

(NS)

Shortstature,

CHD

(PVS,

HCM),

characteristicfacies,webbed

neck,coagulationdefects,

lymph

atic

dysplasias

ID(variable),

language

delay

VMEG,CBTE

MAP2

K1

Kinase

Deno

vo/dom

inant

CFC

synd

rome

Asabove

Asabove

Asabove

Noo

nansynd

rome

Asabove

Asin

theabove

Asin

theabove

MAP2K

2Kinase

Deno

vo/dom

inant

CFC

synd

rome

Asabove

Asabove

Asabove

KRAS

GTPase

Deno

vo/dom

inant

CFC

synd

rome

Asabove

Asabove

Asabove

Noo

nansynd

rome

Asabove

Asabove

Asabove

PTPN11

Phosph

atase

Deno

vo/dom

inant

Noo

nansynd

rome

Asabove

Asabove

Asabove

NRAS

GTPase

Deno

vo/dom

inant

Noo

nansynd

rome

Asabove

Asabove

Asabove

RAF1

Kinase

Deno

vo/dom

inant

Noo

nansynd

rome

Asabove

Asabove

Asabove

SOS1

RasGEF

Deno

vo/dom

inant

Noo

nansynd

rome

Asabove

Asabove

Asabove

RIT1

GTPase

Deno

vo/dom

inant

Noo

nansynd

rome

Asabove

Asabove

Asabove

SHOC2

Scaffolding

Deno

vo/dom

inant

Noo

nansynd

rome

with

looseanagen

hair

Skin

andhairabno

rmalities

(sparse,

thin,slo

wgrow

inghairwith

pigm

entabno

rmalities),features

ofNS

Asabove

Asabove

Transcriptio

nalregulatio

nNSD

1Histon

emethyltransferase

Deno

vo/dom

inant

Sotossynd

rome

Prenatalandpo

stnatalovergrow

th,

characteristicfacialgestalt,

advanced

bone

age

Hypoton

ia,ID

/behavioral

prob

lems(very

common

),SZ

(25%

)

Prom

inenttrigon

e,VMEG/H

YD,X

AX,

CC

abno

rmalities,

CSP

EZH2

Histon

emethyltransferase

Deno

vo/dom

inant

Weaversynd

rome

Characteristic

facies

(prominent

hypertelorism

,micrognathia,

deep

horizontalskin

crease),

camptod

actyly

ID(81%

)Pachygyria,VMEG,

cystsof

theseptum

pellucidu

m(rare)

MED12

Mediatorcomplex

X-linked

Opitz–Kaveggia(FG)

synd

rome

Imperforateanus,characteristic

facialfeatures,broadthum

bsID

(97%

),hypo

tonia

(90%

),SZ

(70%

)Abn

ormalities

ofCC,

VMEG,HET

Lujan(Lujan–Fryns)

synd

rome

Marfano

idhabitus,maxillary

hypo

plasia,palate

anddental

prob

lems,long

hand

s,

ID(m

ild-m

od),

behavioral

abno

rmalities

Dysgenesis

oftheCC

II.


TABLE

(Continued)

Gene

Protein

functio

nInheritance

Synd

rome

Clin

icalfeatures

Neurologicfin

dings

MRIfin

dings

hyperextensib

ledigits

Glypicans

GPC

3Glypican;

cell

surfaceheparan

sulfate

proteoglycans

X-linked

Simpson

-Golabi-

Behmel

synd

rome

Prenatalovergrow

th,characteristic

facies

(macroglossia,

macrostom

ia,centralgroo

veof

lower

lip,ocular

hypertelorism

),supernum

erarynipples

Hypoton

ia,ID

(variable),SZ

HYD,CBTE,ACC

(all

rare)

Mito

ticregulatio

n,centrosomeandmicrotubu

leassembly

KIF7

Kinesin

Deno

vo/dom

inant

Acrocallosalsynd

rome

Polysynd

actyly,hypertelorism

SZ,ID

(very

common

)ACC

OFD

1Centriole-

associated

X-linked

Simpson

-Golabi-

Behmel

synd

rome

(type2)

Ciliarydyskinesia,respiratory

prob

lems,DYSM

,shortfin

gers

Severe

ID,hypo

tonia

VMEG

�

DIS3L

2Exo

ribo

nuclease

Recessive

Perlm

ansynd

rome

Fetalgigantism

,renalhamartomas,

neph

roblastomatosis,

risk

for

Wilm

stumor

ID(m

ost),(poo

rsurvival)

Abn

ormalities

ofthe

CC,HET,

WM

abno

rmalities,

cerebralatrophy

SonicHedgeho

g(Shh

)sig

nalin

gPT

CH1

Patched;

receptor

forsonic

hedgehog

Deno

vo/dom

inant

Nevoidbasalcell

carcinom

a(G

orlin

)synd

rome

Jaw

keratocysts,basalcell

carcinom

as(BCCs),coarse

facial

features,facialmilia,skeletal

anom

alies(bifidribs,wedge-

shaped

vertebrae)

—Ectop

iccalcificatio

ns(in

falx

>90%),

medulloblastoma

(PNET)(5%)

GLI3

Zincfin

ger

Recessive

Acrocallosalsynd

rome

Asabove

Asabove

Asabove

Deno

vo/dom

inant

Greig ceph

alosyndactyly

Polydactyly(pre-,po

st-axial,

mixed),ocular

hypertelorism

,craniosyno

stosis

ID,SZ

(<10%)

HYD

(uncom

mon

)

Other

lesscommon

genesanddisorders

RIN

2Raseffector

protein

Recessive

MACSsynd

rome

(macroceph

aly,

alop

ecia,cutis

laxa,

andscoliosis)

Coarsefacialfeatures,ging

ival

hyperplasia,severe

joint

hyperm

obility,softredu

ndant

skin,sparse

hair,

shortstature,

severe

scoliosis

——

II.


TABLE

(Continued)

Gene

Proteinfunctio

nInheritance

Synd

rome

Clin

icalfeatures

Neurologicfin

dings

MRIfin

dings

RAB39

Rab

GTPase;

cellular

endo

cytosis

X-linked

XLID,autism,

epilepsyand

macroceph

aly

Macroceph

aly

ID/M

R,ASD

,SZ

—

Notes1.

Thistableinclud

eson

lygene-kno

wnMEG/H

MEdisordersanddo

esno

tinclude

otherswhere

underly

inggenetic

etiology

isun

know

nasof

writin

gthismanuscript(such

asMacroceph

aly,megalocornea,motor

andmentalretardatio

n(M

MMM)syndrom

e,Macrosomia,obesity,macroceph

aly,andocularabno

rmalities(M

OMO),forexample).T

hislist

alsodo

esno

tinclud

eskeletaldysplasiaskn

ownto

beassociated

with

MEG

(suchasacho

ndroplasiaandthanatop

horicdysplasia).

2.HMEhasalso

been

repo

rted

with

othersomaticmanifestations

such

ashypo

melanosisof

Itoandlin

earnevussebaceou

ssynd

rome.

Reference:M

irzaaet

al.[2012].

ACC,agenesis

ofthecorpus

callosum;A

DHD,attentio

n-deficit-hyperactivity

disorder;A

SD,autism

spectrum

disorder;C

AL,

caféau

laitmacules;C

BTE,cerebellartonsillarectopia;CC,

corpus

callosum;CHD,congenitalheartdisease;

CLO

VES,

congenitallipom

atou

sasym

metricovergrow

thof

thetrun

k,lymph

atic,capillary,veno

us,andcombined-type

vascular

malform

ations,epiderm

alnevi,skeletaland

spinalanom

alies;CSP,cavum

septum

pellucidu

m;D

YSM

,dysmorph

icfeatures;F

CD,focalcorticaldysplasia;F

TT,

failu

reto

thrive;H

CM,

hypertroph

iccardiomyopathy;HET,

heterotopias;H

ME,h

emim

egalenceph

aly;HYD,h

ydroceph

alus;ID,intellectuald

isability;LD

,learningdisability;MEG,m

egalenceph

aly;MPP

H,

megalenceph

aly-po

lymicrogyria-po

lydactyly-hydrocephalussynd

rome;

NMD,neuron

almigratio

ndisorder;PD

D,pervasivedevelopm

entaldisorder;PM

G,po

lymicrogyria;

PNET,

prim

itive

neuroectod

ermaltumor;PV

S,pu

lmon

aryvalvestenosis;

SZ,seizures;UBO,un

identifiedbright

object;VHD,valvular

heartdisease;

VM,vascular

malform

ation;

VMEG,

ventriculomegaly;

WM,w

hite

matter;XAX,enlargedextra-axialspace;X

LID,X

-linkedintellectuald

isability.

II.


spectrum of PI3K-AKT-mTOR path-way phenotypes (particularly in upstreamcomponents such as PIK3CA) includespredominantly post-zygotic mutationspresent in a mosaic pattern. The relatedMEG syndromes involve almost everyorgan system in the body including thebrain (intellectual disability, autism,epilepsy, hydrocephalus, Chiari malfor-mation), heart and vascular system(conduction defects, heart-great vesselanomalies), skin (capillary malforma-tions, epidermal nevi), connective tissue(skin laxity, joint hypermobility), skele-ton (polydactyly, syndactyly), and others.Mutations within the PI3K-AKT-mTOR pathway are associated withthe most severe brain overgrowth phe-notypes including marked brain over-growth (occipito-frontal circumference,OFC, more than 4 standard deviationsabove the mean) and HME, a seriousmedical condition typically associatedwith severe early onset intractable epi-lepsy and poor developmental outcome.While some syndromes in both pathwaysare considered cancer syndromes, asidentified mutations are “activating”causing enhanced pathway activation,some of the novel germline and mosaicmutations are not as robustly activating asthose associated with oncogenesis andrequire further study. We will specificallyfocus on an area of rapidly increasingknowledge related to PI3K-AKT-mTOR-related brain overgrowth phe-notypes, their clinical and neuroimagingfeatures, and molecular pathogenesis.

PI3K-AKT RELEATED MEGAND HME: THE CLINICALAND NEUROIMAGINGSPECTRUM

In addition to PTEN-related disorders,upstream mutations in core componentsof the PI3K-AKT-mTORpathway havenow been identified in classic HME anda variety of MEG syndromes includingthe MEG-capillary malformation syn-drome (MCAP) and the MEG-poly-microgyria-polydactyly-hydrocephalussyndrome (MPPH). Collectively, thesedisorders not only overlap molecularlybut also share clinical, neuroimaging,and neuropathologic features.

TABLE III. Reciprocal Copy Number Changes Associated with Brain Growth Dysregulation(Megalencephaly and Microcephaly)

Locus CNV Gene Size Syndrome/pathway Refs.

1q21.1 Duplication — MAC — Brunetti-Pierri et al. [2008]Deletion — MIC — Brunetti-Pierri et al. [2008]

1q43.44 Duplication AKT3 MEG PI3K-AKT; MEG and HME Poduri et al. [2012], Wang et al. [2013],Chung et al. [2014]

Deletion — MIC PI3K-AKT; postnatal MIC Ballif et al. [2012]2q24.3a Duplication MYCN MEG Regulates growth an apoptosis Malan et al. [2010]

Deletion — MIC Feingold syndrome Van Bokhoven et al. [2005]5q35.5 Duplication — MIC — Rosenfeld et al. [2013]

Deletion NSD1 MEG Sotos syndrome, transcriptional regulator Tatton-Brown et al. [2005]10q22–23 Duplication — MIC — Aalfs et al. [1995]

Deletion — MAC — Van Bon et al. [2011]16p11.2 Duplication — MIC — Shinawi et al. [2009]

Deletion — MAC — Shinawi et al. [2009]

HME, hemimegalencephaly; MAC, macrocephaly; MEG, megalencephaly; MIC, microcephaly.aThis disorder is associated with triphalangeal thumb, and dysmorphic facial features similar to Weaver syndrome.


PTEN-Related Disorders

Loss of function mutations of PTENhave been identified in a wide rangeof MEG phenotypes including twowell-known cancer predisposition syn-dromes: Cowden and Bannayan–Riley–Ruvalcaba syndrome (BRRS) syn-dromes [Liaw et al., 1997; Marshet al., 1997]. These disorders constitutea clinical spectrum associated withprenatal onset MEG, hamartomas, lipo-mas, intestinal polyps, and various typesof cutaneous vascular malformations[Gorlin et al., 1992; Tan et al., 2011].Neurologically, affected individuals havehypotonia, delayed gross motor skillsand, in some, proximal myopathy[Marsh et al., 1999]. Prenatal onset

Pten loss of function mutantmice develop macrocephalyand behavioral abnormalitiessuch as reduced social activity,increased anxiety and sporadicseizures, closely resemblingthe human phenotype ofPTEN-related disorders.

progressive MEG is typical and OFCsare usually 4–5 (and up to 8) standarddeviations above the mean, whereasbody overgrowth is typically mild(þ1–3 SD). PTEN mutation carriersare at increased risk for various tumors,most notably of the breast, thyroid, andendometrium. Finally, PTENmutationsconstitute the largest single gene defectsin autistic children with MEG, withestimates of mutations between 1% and17% in this cohort [Butler et al., 2005;Buxbaum et al., 2007]. While OFCs inthis cohort vary, one of the earliestreports showedOFCs ofþ7–8 SD abovethe mean [Butler et al., 2005]. Anaverage OFC of þ4.35 SD was recentlyreported in 6 PTEN mutation-positiveindividuals with autism [Hobert et al.,2014]. Pten loss of function mutant micedevelop macrocephaly and behavioralabnormalities such as reduced socialactivity, increased anxiety and sporadicseizures, closely resembling the humanphenotype of PTEN-related disorders[Kwon et al., 2001; Ogawa et al.,2007].

While most children with PTENmutations have uniform bilateral MEGwith grossly normal cortical cytoarchi-tecture (Fig. 1A–C), there are severalpublished cases of asymmetric or focal

brain phenotypes in individuals withgermline PTEN mutations. These in-clude HME and linear epidermal nevi ina child whose family has features ofBRRS, and FCD with focal intractableepilepsy in a child with features ofCowden syndrome [Merks et al., 2003;Elia et al., 2012] (Fig. 1D–F).

PIK3CA-Related Disorders

Post-zygotic gain-of-functionmutationsin PI3KCA have been recently identi-fied in a growing number of segmentalbrain and body overgrowth disorders(Table IV). These phenotypes includesomatic overgrowth disorders such asCLOVES syndrome (congenital lipoma-tous asymmetric overgrowth of thetrunk, lymphatic, capillary, venous, andcombined-type vascular malformations,epidermal nevi, skeletal and spinalanomalies), fibroadipose hyperplasia,isolated macrodactyly and Klippel–Tre-naunay syndrome, and predominantlybrain overgrowth phenotypes such asHME and MCAP syndrome [Kureket al., 2012; Lindhurst et al., 2012;Poduri et al., 2012; Rivière et al., 2012;Lee et al., 2012; Rios et al., 2013]. Theseverity and extent of brain and bodyovergrowth in these phenotypes is


variable. This is particularly true withbrain involvement that ranges frombilateral generalized MEG with PMG(as with classic MCAP), to HME todysplastic MEG (Fig. 1G–L). The tissuedistribution and types of post-zygoticmutations are expected to be among theprimary determinants of the develop-mental phenotypes that are often severe,as we discuss below.

The classic neuroimaging featuresof MCAP are marked diffuse MEG with

The classic neuroimagingfeatures of MCAP are

marked diffuse MEG withbilateral perisylvian

polymicrogyria (PMG),although this latter featuremay not be present in a largenumber of children. Some

individuals also have markedcerebellar enlargement, with alarge and crowded posteriorfossa and ensuing cerebellartonsillar ectopia that may

necessitate surgicaldecompression.

bilateral perisylvian polymicrogyria(PMG), although this latter featuremay not be present in a large numberof children. Some individuals also havemarked cerebellar enlargement, with alarge and crowded posterior fossa andensuing cerebellar tonsillar ectopia thatmay necessitate surgical decompression[Conway et al., 2007a,b; Mirzaa et al.,2012a]. Somatic manifestations ofMCAPinclude cutaneous vascular malforma-tions (typically capillary malformations),polydactyly, syndactyly, and focal somat-ic overgrowth—overlapping with buttypically milder than other somaticPIK3CA-related disorders [Clayton-Smith et al., 1997; Moore et al., 1997;Conway et al., 2007b; Mirzaa et al.,2012a].

The Megalencephaly-Polymicrogyria- Polydactyly-Hydrocephalus (MPPH) Syndrome

This rare developmental syndrome ischaracterized predominantly by prenatalonset MEG and bilateral perisylvianPMG. Hydrocephalus and postaxialpolydactyly are more variable featuresseen in nearly half of reported individu-als [Kariminejad et al., 2012; Mirzaaet al., 2013]. A subset of children also hasa distinctly thick corpus callosum (mega-corpus callosum). De novo germlinemutations in three core PI3K-AKT-mTOR pathway genes are nowknown to be associated with MPPHincluding, in order of frequency,PIK3R2, CCND2, and AKT3 [Rivièreet al., 2012; Nakamura et al., 2014;Mirzaa et al., 2014] (Fig. 2A–C, G–L).These mutations are mostly germlinemutations with a narrow mutationalspectrum. For example, a single recur-rent PIK3R2 (p.Gly373Arg) mutationhas been reported in most MPPHchildren. Postaxial polydactyly is amore frequent feature in PIK3R2 versusCCND2-positive children.

Hemimegalencephaly

HME is a severe brain malformationcharacterized by overgrowth of all orpart of a cerebral hemisphere, often withipsilateral severe cortical dysplasia ordysgenesis, white matter hypertrophyand a dilated and dysmorphic lateralventricle. It is often an isolated congenitalabnormality, but there are sporadicassociations with neurocutaneous andovergrowth syndromes in the literatureincluding with Proteus syndrome, Klip-pel–Trenaunay syndrome, linear nevussebaceous (LNS) syndrome, TSC,neurofibromatosis type 1, and hypome-lanosis of Ito [Cristaldi et al., 1995;Sharma et al., 2009; Pavlidis et al., 2012].HME constitutes the most severe brainovergrowth phenotype not only mor-phologically but also because mostchildren with HME experience earlyonset intractable epilepsy, typically with-in the first few months of life. Childrenwith HME can present with focalseizures or epilepsy syndromes such as

infantile spasms. Developmental delayis often early and severe. Within theaffected hemisphere, neuroimaging re-veals regions of apparent PMG, pachy-gyria, subcortical, and periventriculargray matter heterotopia. However, vari-ous morphological abnormalities outsidethe involved cerebral hemisphere havebeen reported such as ipsilateral cerebralvascular dilatation, ipsilateral and bilateralcerebellar enlargement with dysplasticfolia, and ipsilateral olfactory nerveenlargement [Sato et al., 2007]. More-over, contralateral volume loss (orhemimicrencephaly) with white matterabnormalities have been reported [Shir-oishi et al., 2010]. Although it is notdetermined whether these abnormalitiesare developmental or acquired, these andother more widespread or asymmetricmalformations are believed to partiallyaccount for poor seizure control andpoor post-hemispherectomy outcome insome individuals.

Activating mosaic mutations inthree PI3K-AKT-mTOR pathwaygenes have now been reported inisolated HME including PIK3CA (fourpatients), AKT3 (two patients), andMTOR (one patient) [Lee et al., 2012;Poduri et al., 2012]. Further, duplica-tions of 1q encompassing AKT3 havebeen identified in two HME patientswith presumed activation of the gene[Poduri et al., 2012]. Duplications ofAKT3 have also been reported inchildren with macrocephaly, focalPMG, and intellectual disability [Wanget al., 2013; Chung et al., 2014].

Interestingly, other less commonpatterns of focal MEG with corticaldysplasia have been described in theliterature such as total or diffuse HME,localized MEG (hemi-hemimegalence-phaly), and multilobar cortical dysplasiathat share similar neuropathologicalfindings to HME including large neu-rons, cortical dyslamination, with orwithout dysmorphic and ectopic neu-rons, heterotopia, balloon cells, andabnormal white matter [Barkovich andChuang, 1991; Nakahashi et al., 2009;Blümcke and Mühlebner, 2011]. It istherefore expected that these more focalmanifestations may share the samemolecular pathogenesis.

Figure 1. The PI3K-AKT-mTOR associated neuroimaging spectrum (part I). A–C: T1-weighted mid-sagittal, T2-weighted axial,and T1-weighted coronal images of a child with PTEN-related hamartoma tumor syndrome due to a germline PTEN mutation (p.Gln17X). Note megalencephaly, large cerebellum, crowded posterior fossa and mild cerebellar tonsillar ectopia. This patient had a choroidplexus carcinoma and underwent placement of a left frontal approach ventricular catheter.D–F: Autopsy (D) and CT scan images (E,F) of achild with a maternally inherited PTENmutation (IVS5þ 1delG) showing segmental dysplastic megalencephaly with a markedly enlargedand cerebral hemisphere, periventricular cysts, and focal cortical dysplasia (adapted with permission fromMerks et al., J Med Genet, 2003).This child also had facial linear epidermal naevi ipsilateral to the severely affected cerebral hemisphere.G–I: T1-weighted mid-sagittal, andT2-weighted axial and coronal MRI images of a child with the megalencephaly-capillary malformation syndrome (MCAP) due to a mosaicPIK3CA mutation (p.Glu726Lys). Note generalized megalencephaly, enlarged cerebellum, crowded posterior fossa, moderate cerebellartonsillar ectopia, bilateral perisylvian polymicrogyria (arrowheads, H,I), and moderate to severe ventriculomegaly with stretching of thecorpus callosum. J–L: T1-weighted, and T2-weighted axial and coronal images of a child with a mosaic PIK3CAmutation (p.Glu545Lys)with bilateral asymmetric megalencephaly, severe bilateral cortical dysplasia, dysplastic, and enlarged ventricles. This child also had severesegmental somatic overgrowth (also published in Riviere et al., Nature Genetics, 2013).

TABLE IV. Summary of the PI3K-AKT Associated Developmental Brain Phenotypes

Gene Type of mutation Inheritance CNS phenotype Non-CNS phenotype

PTEN Loss of function De novo/dominant MEG-autism, HME, FCD Cowden, BRRSAKT3 Gain of function Post-zygotic/mosaic,

De novo/dominantHME, MPPH —

PIK3CA Gain of function Post-zygotic/mosaic Megalencephaly (MCAP),HME

Somatic overgrowth (CLOVES/FH, macrodactyly, MCAP)

PIK3R2 Gain of function De novo/dominant MPPH PolydactylyCCND2 Gain of function De novo/dominant MPPH PolydactylyAKT1 Gain of function Post-zygotic/mosaic HMEa Proteus syndrome

BRRS, Bannayan–Riley–Ruvalcaba syndrome; FH, fibro-adipose hyperplasia; HME, hemimegalencephaly; MCAP, megalencephaly-capillary malformation syndrome.aHME reported in Proteus syndrome, but no AKT1 mutations have been identified in affected brain tissues to our knowledge.


PI3K-AKT-MTORRELEATED MEG AND HME:MOLECULAR SPECTRUMAND INSIGHTS INTOMOLECULARPATHOGENESIS

Mutations of the above mentionedPI3K-AKT-mTOR pathway genes,whether loss of function mutations of

Mutations of the abovementioned PI3K-AKT-mTOR pathway genes,whether loss of function

mutations of PTEN or gainof function mutations ofPIK3CA, AKT3, and

PIK3R2, all share a commonfunctional endpoint, namelyactivation of the pathway.

PTEN or gain of function mutations ofPIK3CA, AKT3, and PIK3R2, all sharea common functional endpoint, namelyactivation of the pathway (Table IV).Mutations of PTEN, PIK3R2, andAKT3 have been predominantly germ-line.Whereas mutations of PIK3CA andthe Proteus syndrome gene,AKT1, havebeen mosaic, providing a molecularexplanation for the wide phenotypic

variability in their attendant overgrowthphenotypes [Lindhurst et al., 2011]. Thephenotypic spectrum of PIK3CA-relat-ed disorders is particularly wide and,while the exact mechanisms by whichmutations result in these manifestationsare currently under study, some prelimi-nary genotype–phenotype correlationscan be suggested. For example, the samePIK3CA mutation (p.Glu545Lys) hasbeen identified in the four childrenwith HME so far [Lee et al., 2012].The mutational spectrum of MCAPsyndrome, on the other hand, iswide and has not included any ofthe so-called mutation “hotspots” seenin cancer (p.Glu542Lys, p.Glu545Lys,p.His1047Arg, p.His1047Leu) [Samuelsand Ericson, 2006; Samuels andWaldman, 2010; Rivière et al., 2012;Mirzaa et al., 2013]. While thesemutations are all activating, they maynot as robustly activating as those seen incancer, given that the cancer risk in thesephenotypes does not appear to be muchincreased; although natural history, andparticularly cancer risk, data are lacking.

For more than a decade now,hyperactivation of the mTOR signalingdownstream of PI3K-AKT (due to lossof function mutations of TSC1 andTSC2, for example) have been con-sidered to provide a pathological linkbetween TSC, HME, and FCD byextensive studies [Crino, 2007; Limand Crino, 2013]. Further, mTORinhibition reversed neuronal hypertro-phy in Pten-deficient mice and amelio-

rated a subset of Pten-associatedabnormal behaviors, thereby substanti-ating evidence that the mTOR pathwaydownstream of PTEN is critical for itscomplex phenotype [Kwon et al., 2003;Zhou et al., 2009]. However, the recentidentification of mutations in CCND2in MPPH syndrome sheds a novelinsight into the molecular pathology ofthese phenotypes [Mirzaa et al., 2014].CCND2 is a member of the D-type

Recent data demonstrateaccumulation of degradation

resistant CCND2 inindividuals with MPPH and

also, interestingly, inlymphoblastoid cell lines ofindividuals with upstreammutations in PIK3CA,PIK3R2, and AKT3.

cyclin family critically required for G1/Stransition during the cell cycle [Mat-sushime et al., 1991; Inaba et al., 1992;Ross et al., 1996; Glickstein et al., 2006,2009]. Identified mutations withinCCND2 affect highly conserved termi-nal residues that include targets forglycogen synthase kinase 3b (GSK-3b)-phosphorylation and, ultimately,its’ ubuiquitin mediated degradation[Kida et al., 2007]. Recent data

Figure 2. The PI3K-AKT-mTOR associated neuroimaging spectrum (part II). A–C: T1-weighted mid-sagittal, and T2-weightedaxial and coronal images of a child withMPPH syndrome due to a de novo germline PIK3R2mutation (p.Gly373Arg) showing generalizedmegalencephaly and bilateral perisylvian polymicrogyria. D–F: T1-weighted and T2-weighted axial and coronal images of a childwith MPPH syndrome due to a germline AKT3 mutation (p.Arg465Trp) showing bilateral but asymmetric megalencephaly, bilateralperisylvian PMG, and mild enlargement of the lateral ventricles. G–I, T1-weighted mid-sagittal and T2-weighted axial and coronalimages of a child with isolated hemimegalencephaly due to a mosaic AKT3 (p.Glu17Lys) mutation. J–L: T2-weighted mid-sagittaland axial and T1-weighted coronal images of a child with MPPH syndrome due to a de novo germline mutation in CCND2(p.Thr289Ala) showing generalized megalencephaly, bilateral extensive perisylvian polymicrogyria, marked ventriculomegaly status postshunt placement.


Figure 3. Schematic of PI3K-AKT-mTOR associated Meg Phenotypes. Asimplified schematic diagram showing key PI3K-AKT-mTOR genes (PIK3CA,PIK3R2, PTEN, AKT3, and CCND2) with their associated phenotypes.


demonstrate accumulation of degradationresistant CCND2 in individuals withMPPH and also, interestingly, inlymphoblastoid cell lines of individualswith upstream mutations in PIK3CA,PIK3R2, and AKT3 [Mirzaa et al., 2014].These data implicate for the first time theinvolvement of another critical effectorpathway downstream of PI3K-AKT: thecell cycle pathway downstream of GSK3b(Fig. 3). Clearly, further investigationis necessary to determine the detailedmolecular mechanisms of brain over-growth in this pathway and how thesevarious critical downstream pathwaysinteract in these phenotypes.

THERAPIES AND FUTUREDIRECTIONS

The past few years have witnessedexciting advances in our understandingof the molecular pathogenesis of brainovergrowth. It follows that individuals

Some of the key functionaldeficits of these disorders, such

as intellectual disability,

autism, epilepsy,hydrocephalus, are naturallyattractive targets for treatmentthat today are still treated

empirically. For example, theepilepsy associated with someMEG syndromes is treatedwith anti-epileptic drugs thatdo not specifically address themolecular defects that we

now know to be the basis oftheir disease.

with disorders of the PI3K-AKT-mTOR and the interacting Ras/MAPK pathways may have the optionin the future of pathway-based rationaltreatment. Some of the key functionaldeficits of these disorders, such asintellectual disability, autism, epilepsy,hydrocephalus, are naturally attractivetargets for treatment that today are stilltreated empirically. For example, theepilepsy associated with some MEG

syndromes is treated with anti-epilepticdrugs that do not specifically address themolecular defects that we now know tobe the basis of their disease. The use ofnumerous small molecule PI3K-AKT-mTOR pathway inhibitors to alleviatesome of these developmental defectsis currently under study. While specifictherapy is already available for thedownstream TSC-mTOR pathway byeverolimus [Kingwell, 2013; Kruegeret al., 2013], it is clear that the brain andsomatic overgrowth phenotypes associ-ated with upstream PI3K-AKT-mTORpathway mutations result from increasedactivation of multiple pathways down-stream of AKT, leading us to predict thatsuccessful treatment strategies will needto downregulate more than one of thesepathways.

ACKNOWLEDGMENTS

The authors thank our patients andtheir families for their valuable andongoing contributions and support ofour research. A.P. was supported by theNINDS (NS069784).

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Megalencephaly and hemimegalencephaly: Breakthroughs in molecular etiology