Viktor Kožich
Institute of Inherited Metabolic Disease1.LF UK and VFN Praha
Introduction to biochemical genetics
OutlineAims of the courseBasic features of IEM, population frequencyHistory of IEMGenetic origin of IEMPathogenetic mechanismsSmall molecules and complex moleculesClinical features of IEM
Aims of the course on biochemical genetics
Biochemical genetics = IEMs
genetics
clinical medicine
biochemistry
Case 1-Isovaleric aciduria
failure to thrive, vomitingrepeatedly alteredconsciousnessketoacidosissweaty feet odour
Case 2-adult cystinosis
http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=gene&part=ctns
Case 3- MPS I
http://deti.msk.ru/plaxin_egor.jpg
http://myweb.lsbu.ac.uk/dirt/museum/margaret/438-1811-2640151.jpg
http://eyepathologist.com/images/KL1771.jpg
Case 4-cystinuria
http://graphics8.nytimes.com/images/2007/08/01/health/adam/17046.jpghttp://www.nature.com/ki/journal/v73/n8/images/5002790f1.jpg
Basic features of IEMs
Inborn errors of metabolism
product
substrateusually AR, GR
clinicallyvariable
usually enzyme
Clinical features of IEMs-age
http://markandrich.googlepages.com/Old-woman.jpg/Old-woman-full.jpg
http://www.hrr.co.uk/acatalog/crocodile_toddler.jpghttp://www.co.shasta.ca.us/html/DSS/images/FosterParentingAdopt/infant.jpg
Clinical features of IEMs-organs
http://universe-review.ca/I10-82-organs.jpg
Clinical features: (non)specific signs
http://img.medscape.com/pi/emed/ckb/pediatrics_genetics/941088-943343-507.jpg
http://gatsome.com/images/iq.gifhttp://www.saratogaschools.org/AcademicServices/MiddleSchool
specific non-specific
e.g. NH3, uric acid
Clinical features-multisystemicinvolvement
http://www.istockphoto.com/file_thumbview_approve/5982111/2/istockphoto_5982111-human-internal-organs.jpg
Diagnostic procedures in IEMs
produkt
substrát
DNA/RNA Enzymes Metabolites
|Clinicsspecific-e.g..
smellurine color
nonspecific-e.g.comaPMRdysmorfic featureshepato/myopathieother
Frequency of IEMs
newborn screening 1:1000-1:4000selective screening at least 1:500-1:1000frequency of heterozygotes at least 1:15population specific examples
higher incidence in imbred populations (PKU Turkey, organicacidurias Middle East)tyrosinemia type I- Quebecaspartylglykosaminuria- Finlandlysosomal strorage disorders- Izrael
IEMs in the Czech Republic
peroxisomální4%
sacharidy12%beta-oxidace a
OAU14%
AMK bez HPA18%
HPA a PKU13%
lysosomální16%
mitochondriální20%
puriny/pyrimidiny3%
incidence ~ 1:1000so far ~150 different diseases
ČR, 2005, n=127
Treatment of IEMs
produkt
substrát
History of biochemicalgenetics
http://www.historiadelamedicina.org/imagenes/garrod.jpg
1857-1936
http://www.thetoque.com/010605/pics/urine.jpg
Trophîme Bigot, 16.-17.století
http://images.google.com/imgres?imgurl=http://upload.wikimedia.org/wikipedia/commons/thumb/8/8c/Bigot,_Trophime_-_A_Doctor_Examining_Urine.jpg/
1857 L.Pasteur demonstrates microbial origin of fermentation, R.Wirchow describes mitosis1867 dynamit1874 jeans1886 automobil=car1890 underground1895 cinematograf1902 periskop, el.typewriter; Nobel prize fordiscovery of malaria transmission1908 Nobel prize for discovery of immunity1936 tape recored; Nobel prize forneurotransmitter research
Lancet, vol ii, 1902, 1616-1620
Expansion of the field
1908 • 5 diseases(alkaptonuria, cystinuria, porphyria, pentosuria, congenitalsteatorhea)•1909 first textbook•incidence unknown•dx: 4 metabolites in urine•Rx none
2009•at least 500 diseases•Medline 122.797 papers (28/9/2009)•textbook-web based•incidence at least1:1.000•Dx: metabolites, enzymes, DNA•screening MS-MS•Rx for ~ 1/3 diseases
cca 1960 • ca 40-50 diseases•analyticalmethods•incidence of PKU•PKU- 1st screening program (R.Guthrie)•PKU-1st treatableIEM (H.Bickel)
Biochemical genetics in the ČRPKU screening 1975-Doc.Blehová, Prof.Hyánek, Ing.MrskošDx of other IEMs-leader Prof.Hyáneklabs in Prague, H.Králové, Brno, Olomouc, Ostravain 2009:
selective screening ~ 100 pacientů/rok (incidence 1:1000)newborn screening- 10 IEMs since October 1 2009, expected incidence 1:4000
First IEM:
Alkaptonuria
http://www.mja.com.au/public/issues/184_12_190606/sha20286_fm-1b.jpghttp://www.scielo.br/img/revistas/rbr/v46n5/a14fig02.jpg
</< td>
http://bjo.bmj.com/content/vol83/issue6/images/large/98532.f1.jpeg
1859 Boedeker- alkapton in urine1891 alkapton=homogentisic acid
http://www.bioinformatics.nl/webportal/background/images/mendelexperiment.gif
Garrodův revolutionary concept
Chemical individuality is determined by genes
Inborn errors of metabolism= disturbanceof chemical individuality
Alkaptonuria in 2009retrospective reasearch of mummy HARWA 1500 př.n.l.pateints known before Garrod1902 Garrod1909 tyrosine degradationpathway described1958 enzyme defect in alkaptonuria established1996 HGO gene description2003 treatment with nitisone
http://www.mja.com.au/public/issues/184_12_190606/sha20286_fm-1b.jpghttp://www.scielo.br/img/revistas/rbr/v46n5/a14fig02.jpg
</< td>
http://bjo.bmj.com/content/vol83/issue6/images/large/98532.f1.jpeg
CoxarthrosisValvularinvolvementUrolithiasis
http://www.natuurlijkerwijs.com/english/b4f4ca00.gif
http://ourworld.compuserve.com/homepages/sbrillanti/liver2.jpg
Liver HGO deficiencyIncidence 1:250 000 až 1:1 000 000High incidence SlovakiaLate complicationsfrequentRx inefficient
http://en.wikipedia.org/wiki/
http://www.natuurlijkerwijs.com/english/b4f4ca00.gif
nitison (NTBC)
Novel treatment in alkaptonuria
Genetic origin of IEMs
Genetic origin of IEMMonogenic Mendelian inheritance
AR inheritanceGR/GD inheritancerarely AD inheritance
X
Mitochondrial inheritanceEpigenetic changesX-inactivationEpsistasisGene x environment interactionother
Common types of mutations
Molecular consequences of mutations
Protein amountsdecreasedsameincreased
Protein propertieschanges of isolated functionsglobal changesmisfolding
Nonprotein gene productssiRNA
Role of mutations in evolutionSource of variability(less important thanmeiotic recombinationand combination of gametes)
Favourable mutationsare rare and alreadycontained in genomes
Genetic diseases= tip of iceberg (of geneticvariability)
Mutations affect different domains
e.g. PAHcytosolic enzymehomotetramerBH4 as cofaktorcomplicatedmechanism of catalysis
Erlandsen H and Stevens RC, The Structural Basis of Phenylketonuria. Mol Genet Metab. 1999 Oct; 68(2):103-125.
Cellular consequences of mutations (e.g.CFTR)
http://student.biology.arizona.edu/honors97/group7/Hallick1.gif
Genotype-phenotype (M.Gaucher)
http://www.nature.com/embor/journal/v4/n7/thumbs/embor873-f4.gif
severesplenomegalyanemiabone disorderCNS
mildsplenomegalyanemia
unclear
Phenotype
Health-disease threshold
environmentgenes
clinical manifestation
genetic test
preclinical test
IEM treatment- modifying environment
environmentgenes
clinical manifestation
genetic test
preclinical test
Gene x environment interaction (PKU)
http://www.pkunews.org/adults/image005.gif
Disease modification by environment (alpha1-antitrypsin)
Current genetics is not Mendelian
classical genetics1 gene=1 disease
1 gene = more diseases1 gene = diseases withdifferent types of inheritance several genes = same diseasedigenic inheritancetransgenerationaltransmission of epigeneticmarksother ……
Genetics has been and will be full of suprises
Patophysiologicalmechanisms in IEMs
Patophysiology
product
byproduct
enzyme transporter
substrate
precursor
Patophysiology
byproductsubstrate
precursor
Examples:Phe a Phe-derivativesammoniacystine in cystinosiscystine in cystinuriamucopolysaccharides
Patophysiology
product
Examples:glucose in GSDketone bodies in beta-oxidation defectsplasmalogens in peroxisom.diseasescysteine in CBS deficiencyAdoMet in RMATP in mitochondrialdiseases
Local vs. systemic consequences
Local consequence, e.g.:MPS-bones, tendons, spleen, liver, CNScystinuria-urinary tract
Local vs. systemic consequences
produkt
substrát
Distant consequences-e.g.urea cycle disorders- comaorganic acidurias-encephalopathyCBS deficiency- thrombosis, connectivetissue disturbances
Disorders of small andcomplex molecules
Small molecules in biochemicalgenetics
definition: < 1500 Dagases, inorganic ionsamino acidsorganic acidssaccharidespolyolssimple lipidspurines, pyrimidinesvitaminsoligomers: peptides up to 5-10 AA, oligosaccharides
cytosol, mitochondrial stromablood, urine
Diseases of small molecules
usually dependent on exogenous supplymanifestation: (repeated) acute toxicity, usuallywith encephalopathy/comahepatopathy commoncommon disturbances in routine labs-ammonia, Astrup, ketone bodies, glycemia, uric acid...symptoms develop due to specific type of food, fasting, catabolismchronic course possible (if toxicity low)usually good therapeutic response to diet and/orvitamins
Isovaleric aciduria
FTT, vomiting, Kussmaulbreathingconsciousness: comawithin 24-48 h after onsetof symptomsmetabolic acidosis, ketonuriasweaty feet syndrome
http://images.google.com/imgres?imgurl=http://upload.wikimedia.org/wikipedia/commons/8/8b/Isovaleric_acid_structure.png
Cystinuria
http://graphics8.nytimes.com/images/2007/08/01/health/adam/17046.jpghttp://www.nature.com/ki/journal/v73/n8/images/5002790f1.jpg
Complex molecules in biochemicalgenetics
definition: > 1500 Daglykolipidssphingolipidsplasmalogensneutral polysaccharides (glycogen)mucopolysaccharides(other polymers: proteins, nucleic acids...)
usually associated with membranesconcentrations in blood/urine rather low, exceptions exist ( x MS/MS technologie)
Diseases of complex moleculesdisease progresses usually regardless of anyexogenous sources from foodtypical course is progressive (± symptom-freeperiod)dysmorphy at birth possiblefrequent involvement of nervous system andmusculatureorganomegaly due to storage in lysosomalstorage disordersusually untreatable by diet or vitamins
Mukopolysaccharidosis type I
http://deti.msk.ru/plaxin_egor.jpg
http://myweb.lsbu.ac.uk/dirt/museum/margaret/438-1811-2640151.jpg
http://eyepathologist.com/images/KL1771.jpg
endogenousexogenousOrigin
N YAcute toxicity
inefficiousefficiousRx-diet, vitamins
tissues (U)blood,urineDx
YNImpact on structure
membranescytosol, ECTLocalization
Y±Chronic progression
> 1500 Da< 1500 Da
Patophysiology IEM
product
vedl.produkt
substrate<1500 Da
>1500 Da
1 32
Clinical manifestation of IEMs
Clinical picture-age
http://markandrich.googlepages.com/Old-woman.jpg/Old-woman-full.jpg
http://www.hrr.co.uk/acatalog/crocodile_toddler.jpghttp://www.co.shasta.ca.us/html/DSS/images/FosterParentingAdopt/infant.jpg
Clinical features-organs
http://universe-review.ca/I10-82-organs.jpg
Selected common situations withhigh risk of IEM
Small moleculesacutelly ill newborn(repeated) atack of long-term uncosciousnessfailure to thrive
Complex moleculesprogressive CNS and musculature involvementfacial dysmorphyorganomegaly (liver, spleen, heart)
Food and IEMs (small molecules)
(sub)acute toxicitymilk (lactose)-hepatopathysaccharose/fructose/sorbitol- hepatopathyand hypoglycemiaexcess protein- vomiting, lethargy, coma(urea cycle disorders, organic acidurias)
Fasting and IEMs
hypoglycemia in GSDhypoglycemia with decreased production of ketone bodies (beta-oxidation defects)acidosis, ketonuria and metabolicencephalopathy in prolonged fasting (organicacidurias)respiratory alkalosis and encephalopathy (urea cycle disorders)
Abnormal urinary smell and colorsmell (small volatile molecules):
sweaty feet-isovaleratemaple syrup-branched ketoacidsboiled cabbage-methionine oxidfish-trimethylamineblackcurrant- organic acidsmouse-phenylacetate
colororange-urateblack upon oxidation-homogentisateblue-indoxyl derivatiesgreen-4-OH-butyrate
Common labs in IEMs
BloodglycemiacholesterolTGuric acidMAchyperammonemia, RAlkALT,ASTCKanemia/pancytopenia
Urineketone bodiesuric acidcrystaluriamyoglobinuria
http://www.historiadelamedicina.org/imagenes/garrod.jpg
1857-1936
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