final report fragile x syndrome

All rights reserved 2010

DIGROUP 4

QUIWA, GELLI ANNERAMIREZ, BETTY JANERAQUIZA, KENNETHREBANAL, MARY REMILENNEREJANTE, TITO GUILLERMOREYES, NEIL JASON

Learning OBJECTIVES:

1.Describe the fragile x syndrome, its clinical features and manifestations.

2.Understand the cause and molecular perspective of the disease.

3.Explain the Sherman Paradox and its solution.

4.Discuss the symptomatology, treatment, inheritance and diagnosis of the disease.

CASE STUDY PRESENTATION

CASE REPORTCASE REPORT

• The patients were brothers, aged 2 yrs 9 mos and 1 yr 8 mos, and were referred to the hospital for evaluation for developmental delay.

ELDER BROTHER

• Born at term – uneventful pregnancy and delivery

• Birth weight – 3260 grams• Psychomotor

Development: Grossly retarded

• PE: Hyperactive boy with wt. of 13.2 kg, Ht. 91.5cm and head circumference of 47.9cm

YOUNGER BROTHER

• Born at 36 weeks of gestation

• Birth weight – 2760 grams

• Psychomotor Development: Moderately Retarded

• PE: Hyperactive boy with normal height and head circumference

ELDER BROTHER

Head Control: 5 monthsSit alone: 12 monthsWalk : 20 months

- Never spoke any meaningful words

YOUNGER BROTHER

Head Control : 4 monthsSit alone: 10 monthsStand up holding into

furniture: 20 months- Had not acquired any

meaningful words yet

ELDER BROTHER

Facial Abnormalities: • long and square-shaped

face with a high forehead

• ears were large and prominent

• absent left lower incisor

testes was not enlarged

YOUNGER BROTHER

Facial Abnormalities: • square-shaped face

with a prominent forehead

• everted ears • absent left lower incisor

ELDER BROTHER

Developmental Quotient (DQ): • 38% that of a normal child of

the same age

YOUNGER BROTHER

Developmental Quotient (DQ): • 47% of normal child of the

same age

ELDER BROTHER

• hyperactivity• short attention span• poor eye contact• excessive withdrawal

response to strange persons or environment

YOUNGER BROTHER

• short attention span but no social aversion or hand mannerisms

ELDER BROTHER

• Electroencephalography (EEG): normal

• Acoustic brainstem response(ABR): normal

YOUNGER BROTHER

• EEG and ABR: not remarkable (20 months)

• massive epileptic discharges during sleep had become evident on the bilateral parieto-occipital regions since one year and 6 months of age

General Information:• Born at term• Birth weight – 3080 grams• Pregnancy complicated by upper GI radiographic

exposure in 1st trimester

Delayed Developmental Milestones:Motor development: • almost normal but speech was grossly delayed

Pre pubertal

• Facial Abnormalities: long face with prognathism, ears were not malformed

• Speech disturbance characterized by lack of fluency, echolalia and inappropriate grammatical quotient

• Psychometric testing (I.Q): 45%• Menstruation started at 13 years of age and

remains regular

Post Pubertal• Physical examination: shy and mentally retarded

with normal height and head circumference (20 y/o)

Others…• Anticonvulsant drugs were administrated for EEG

abnormalities• Remaining maternal siblings appeared intellectually

normal although psychometric tests were not performed

No school problems except for one uncle who left high school to work as a truck driver

Mother: IQ of 92 by Wechsler Adult Intelligence Scale Test

Maternal grandmother: 6th of 7 sibship-denied the presence of Mental Retardation in her brothers and sisters.

General Information:• Born at term by caesarian section• Birth weight – 3462 grams

Motor development: • psychomotor development almost normal except

for speech

Pre Pubertal

• 18 mos –began to speak a few meaningful words• 2y/o & 10mos - PE: normal height and head

circumference• somewhat square face but appeared normal• I.Q: 85% that of a normal child

Family pedigree of the patients. Solid squares and circles indicate subjects with Mental Retardation, and half-circle shaded denotes a person with borderline intelligence

• Cytogenetic Study

The right-hand member in each of these two pairs of X chromosomes is a fragile X; the leader points to the fragile site at the tip of the long arm. Males hemizygous for this chromosome exhibit the fragile-X syndrome of mild to moderate mental retardation.

Cytogenetic Studies

• Useful to exclude sex chromosomes and subtle autosomal abnormalities frequently associated with nonspecific mental retardation

• The fragile site, fra(X)(q27.3), can be induced under specific culture conditions

Cytogenetic Studies

Cytogenetic Studies

As shown in table 3: All had fra(X)(q27.3)• Elder brother, younger brother,(III-1 and III-2)• Younger sister, (III-3)• Mother (II-3)• Maternal aunt (II-6)

• The remaining family members did not have the fragile site.

WWhat is fragile X hat is fragile X syndromesyndrome

aka MARTIN-BELL SYNDROME most common cause of inherited mental

impairment Second only to Down syndrome as an

etiology for MR X linked recessive Only the mother has to be a carrier for

the fetus to be at risk for fragile X syndrome

Incidence: 1:1250 (males)

1:2500 (females)

1:800 men are carriers 1:260 women are carriers

Appears in all socio-economic background

SYNDROMEGroup of unusual physical, behavioral and/or intellectual features, which occur in a pattern in an individual and together describe the characteristics of a particular condition

Mental RetardationLong face with large everted earsMacro-orchidism

WWhy fragile Xhy fragile X

•Particular feature at the end of some of their X chromosomes when examined under a high powered microscope in a cytogenetics laboratory using special techniques.

•This feature is called a 'fragile site' (fra X) because it appears as a narrowing at the end of the chromosome.

• Mutations in the FMR1 gene at fragile site Xq 27.3 (FRAXA)

WWhathat causes fragile X syndrome causes fragile X syndrome

Mutation in FMR1 geneMutation in FMR1 gene

•Each chromosome is made up of genes. •Genes contain the information used by other parts of the cell to make proteins, the body's building blocks. •Proteins make up the structure of your organs and tissues

• Each protein performs a specific job in different types of cells, and the information for making at least one protein is contained in a single gene.

• Genes are made up of various lengths of DNA, which contains four chemicals: adenine (A), guanine (G), cytosine (C), and thymine (T).

• Triplet: information in our gene is made up of 3 letters= trinucleotides

Additional Additional

• In some genes, the same triplet is repeated within the DNA sequence.

• This is a sequence of ‘triplet repeats’.

• The number of times that a triplet code is repeated in a gene can be critical

• The triplet that is repeated in the FMR-1 gene is made up of the letters ‘CGG’.

• When the number of ‘CGG’ repeats in the FMR-1 gene increases over a critical number, the gene becomes so long that it becomes faulty.

FMR1 geneFMR1 gene

CGG lies within the 5’ untranslated region of the first exon of FMR- 1

CPG island: DNA region of high deoxycytidine and deoxyguanosine content linked by a phosphodiester located near the protein coding region of a gene and related to its regulation of gene expression

Hypermethylation: a chemical alteration of DNA introduced by the attachment of a methyl group to deoxycytidine, thereby producing 5-methyl deoxycytidine in the CG sequence where cellular regulation of gene expression or X-inactivation is believed to occur.

Normally , thymine is the one methylated. In here, cytidine is the one being methylated.

Types of mutation for the Types of mutation for the FMR1 gene within FXSFMR1 gene within FXS

NormalNormal

The genetic code for the FMR-1 gene usually contains a limited repetition of CGG sequences.The normal range is 5-50 repeats.

Premutation

When the number of CGG repeats is between 50 and 200 the individual is a premutation carrier of fragile X syndrome. Carriers are not usually affected by fragile X syndrome, but they are at risk of having affected children.

Full mutation

If the number of repeats If the number of repeats exceeds 200, usually this exceeds 200, usually this disrupts the code and disrupts the code and prevents the production of prevents the production of the FMR protein. These the FMR protein. These individuals have the full individuals have the full mutation and usually are mutation and usually are affected by fragile X affected by fragile X syndrome.syndrome.

CCG repeats prone to CCG repeats prone to METHYLATIONMETHYLATION

TRANSCRIPT METHYLATION cause TRANSCRIPT METHYLATION cause ‘GENE SILENCING’‘GENE SILENCING’

Type No. Of CGG repeats

Level of methylation Male Female

Normal 6-50

Premutation 53-230 Unmethylated Men usually unaffected geneticcarriers

Women usually unaffectedgenetic carriers

Full mutation >230 Completely methylated

100% affected Women may be unaffectedor affected less severely thanaffected males

Premutation

Intellectual impairmentFragile X syndrome

Premutation carriersFMR-1 gene working>50yrs: Ataxia syndrome/tremorWomen: Premature ovarian failure before 40

Full Mutation( male)Intellectual disabilitySpeech delayDelay in physcial skills Coordination difficulties

Behavioral Attention problemsHand flapping and bittingGaze aversionRepetitive speechSensory problemsAnxiety and mood instability

FRAGILE X SYNDROMEFRAGILE X SYNDROME

In FXS, the expression of FMR1 gene is shut off, thus, In FXS, the expression of FMR1 gene is shut off, thus, there is less or no production of FMR1 protein. FMR1 there is less or no production of FMR1 protein. FMR1 protein is important for maturation of dendritic spines.protein is important for maturation of dendritic spines.

60% have mild intellectual disability

hyperactivity or a shy personality

selective lack of speech (mutism)

emotional and behavioural characteristics

Full mutation( female)

SSummaryummary

DEFECT IN FMR1 gene (Xq27.3)DEFECT IN FMR1 gene (Xq27.3)

PREMUTATION: CCG repeats >55 n <200 (FXTAS) PREMUTATION: CCG repeats >55 n <200 (FXTAS)

FULL MUTATION: CCG repeats 200 and aboveFULL MUTATION: CCG repeats 200 and above

HIGH NUMBER OF REPEATS causes transcript methylation HIGH NUMBER OF REPEATS causes transcript methylation

GENE SILENCING—NO PRODUCTION of FMR1GENE SILENCING—NO PRODUCTION of FMR1

FRAGILEFRAGILE XX SYNDROME!SYNDROME!

•Southern Blotting•Polymerase Chain Reaction (PCR)•Prenatal Diagnosis

REBANAL, Mary Remilenne E.

DNA blotDiagnostic method of choice

Determine the extent of CGG repeat expansion

Methylation status of the CPG islandif the gene has a full mutation and its approximate size.

Gene I

Genomic DNA

Restriction endonucle

ase

Gel Electrophore

sis

Long DNA fragments

Short DNA fragments

Agarose gel

DNA fragments

-

+

Eag1(methylation

sensitive) EcoR1, Eag1

DNA fragments

Agarose gel

-

+

1. Denature in alkali

2. Blot-transfer bake ssDNA

fragments

nitrocellulose

Radioactive RNA or denatured DNA containing sequences complementary to gene I (radioactive probe)

1. Hybridize nitrocellulose with radioactive probe

2. Wash

DNA digestion with EcoR1 + Eag1Hybridization with Pstl

pE5.1 Pfxa3 Pfxa3

Normal Male Female

2.4 + 2.82.4 + 2.8 + 5.2

2.82.8 + 5.2

1.01.0

Premutation Male Female

2.4 + (2.9 -3.3)2.4 + 2.8 + (2.9 -3.3) + 5.2

(2.9 -3.3)2.8 +(2.9 -3.3) +5.2

(1.1-1.6)1.0 + (1.1-1.6)

Full mutation Male Female

>5.72.4 + 2.8 + 5.2+ >5.7

>5.72.8 + 5.2 + >5.7

>1.61.0 + >1.6

•determine the actual number of repeats•If a premutation• not the test of choice to diagnose a full mutation•quite accurate.

•DNA template

•DNA polymerase

•Primers

•Nucleotides (dNTPs or deoxynucleotide

triphosphates

REQUIREMENTS:

PROCESS:

SOUTHERN BLOTTINGPOLYMERASE

CHAIN REACTION

Determine the actual number of repeatsIf a premutation.quite accurate.

Determine the extent of CGG repeat expansion.if the gene has a full mutation and its approximate size.Methylation status of the CPG island

CHORIONIC VILLUS SAMPLING (CVS)

AMNIOCENTESIS

@ 10 weeks of pregnanc

y

CHORIONIC VILLUS SAMPLING (CVS)

AMNIOCENTESIS

16-20 weeks of pregnancy

Inheritance


AN INFECTIOUS DISEASE

A PROBLEM WITH NUTRITION

FRAGILE X SYNDROME IS…

NOT

NOT

IT’S IN THE DNA!!!

POINTS TO REMEMBER IN FRAGILE X SYNDROME

INHERITANCE:•Fragile X in an "X-linked" condition, which means that the gene is on the X chromosome.

•If she has a premutation, and it is passed on (to either males or females), it can remain a premutation or it can expand to a full mutation.

•If she has a full mutation and it is passed on (to either males or females), it will remain a full mutation.

•Males with the premutation will pass it on to all of their daughters and none of their sons .

•In other X linked conditions all males who carry the gene are affected, in fragile X, unaffected males can carry the gene in the premutation form and have no symptoms of fragile X syndrome

•In many X linked conditions only males who inherit the abnormal gene are affected, in fragile X, females can also be affected.

What is SHERMAN PARADOX?


SHERMAN PARADOX

•It indicates the variation in the propensity of premutation to become full mutations maybe related to:

1.Size of the premutation

2. Gender of the carrier

SHERMAN PARADOX

Analyze FMR-1 gene in fragile X families with transmitting maleTransmitting males = premutation

Daughters = same number of CGG repeats from fatherUnstable during oogenesis= full mutation on sons

Brothers= full mutationsPremutations >80 repeats of CGG= full mutations( mothers)

Resolving

GENETIC DISEASES

ASSOCIATED WITH

DYNAMIC MUTATIONS


Disease Chromosome Location

Repeated Sequence

Sex Bias of Parent

Contributing Severe Form

Normal number of

copies

No. of Copies assoc with the disease

Change in Gene function

Fragile X syndrome

Xq27.3 CGG Maternal 6-50 Premutation: 50-230Full mutation 230-2000

Loss

Spinobulbar muscular atrophy

(Kennedy disease)

Xq11-12 CAG ? 11-31 40-62 Gain

Myotonic Dystrophy

19q13.3 CTG Maternal 5-35 Premutation: 50-80Full mutation 80-2000

mRNA disability

Huntington disease

4p16.3 CAG Paternal 9-37 Premutation: 30-38Full mutation 37-121

Gain

Disease Chromosome

Location

Repeated Sequence

Sex Bias of Parent

Contributing Severe

Form

Normal number of copies

No. of Copies assoc with the disease

Change in Gene

function

Spinocerebellar Ataxia Type I

6p22-23 CAG Paternal(Possibly)

25-36 43-81 Gain

Fragile XE mental

retardation

Xq28 CGG ? 6-25 Premutation: 24-2000Full mutation >200

Loss

Denatorubral

pallidoluysian

atrophy

12p12.13 CAG Paternal (mainly)

7-23 49-75 Gain

Machado-Joseph Disease

14q 32.1 CAG ? 13-36 68-79 Gain

Therapeutic Options


•Carbamazepine (Tegretol)•Valproic acid or divalproex (Depakote)•Lithium carbonate•Gabapentin (Neurontin)•Lamotrigine (Lamictal)•Topiramate (Topomax), tiagabine(Gabitril), and vigabatrin (Sabril)•Phenobarbital and primidone (Mysoline)

GENETIC MEDICATIONS(BRAND NAMES IN PARENTHESES)

SEIZURES and MOOD INSTABILITY

Therapeutic Options

•Methylphenidate (Ritalin, Concerta) and dexamethamphetamine (Adderall, Dexedrine)•L-acetylcarnitine•Venlafaxine (Effexor) and nefazodone (Serzone)•Amantadine (Symmetrel)•Folic acid

ATTENTION DEFICIT(WITH OR W/out HYPERACTIVITY)

Therapeutic Options

•Clonidine (Catapres TTS patches)•Guanfacine (Tenex)

HYPERAROUSALSENSORY OVER-STIMULATION

(OFTEN OCCURS WITH ADD/ADHD)

Therapeutic Options

AGGRESSIONINTERMITTENT EXPLOSIVE DISORDEROBSESSIVE-COMPULSIVE DISORDER

Therapeutic Options

•Fluoxetine (Prozac)•Sertraline (Zoloft) and citalopram (Celexa)•Paroxetine (Paxil)•Fluvoxamine (Luvox)•Risperidone (Risperdal)•Quetiapine (Seroquel)•Olanzepine (Zyprexa)

SLEEP DISTURBANCES

Therapeutic Options

•Trazadone•Melatonin

MANAGEMENT

•Assistive Technology

•Speech and language therapy

•Therapy for behavioral disorders

•Medical treatment of aggression

•Behavioral treatment of aggression

•Cognitive development intervention

•Intervention for sensory integration disorders

•Gross motor skill development

•Daily living skills

•Medical Follow-up and Pharmacotherapy

•Toilet Training

COMPREHENSIVE APPROACH

EDUCATION

Birth-Three: Early Intervention for Infants and Toddlers with

Fragile X

Early Childhood/Preschool Services for Children with Fragile X.

Elementary School Aged Children with Fragile X

High School and the Transition to Adulthood for Adolescents

with Fragile X

SPECIAL EDUCATION

In summary..

FRAGIILEXSYNDROME..•Is a X linked disorder.•Can be a premutation and full mutation.•Caused by mutation in the trinucleotide sequence causing methylation.•Loss of FMRP causing mental retardation etc.•Can be diagnosed molecularly.•Worsen with each passing generations.•Has no cure, but can be manage.

… the end …

final report fragile x syndrome

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