medical biochemistry biochemical and genetic basis of disease lecture 77 biochemical and genetic...
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Medical BiochemistryMedical Biochemistry
Biochemical and Genetic Basis of Disease
Lecture 77
Biochemical and Genetic Basis of Disease
Lecture 77
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Classes of Biomolecules Affected in Disease
• All classes of biomolecules found in cells are affected in structure, function, or amount in one or another disease– Can be affected in a primary manner (e.g., defect in
DNA) or secondary manner (e.g., structures, functions, or amounts of other biomolecules)
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Rate of Biochemical Alterations
• Biochemical alterations that cause disease may occur rapidly or slowly– Cyanide (inhibits cytochrome oxidase) kills within a
few minutes
– Massive loss of water and electrolytes (e.g., cholera) can threaten life within hours
– May take years for buildup of biomolecule to affect organ function (e.g., mild cases of Niemann-Pick disease may slowly accumulate sphingomyelin in liver and spleen)
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Deficiency or Excess of Biomolecules
• Diseases can be caused by deficiency or excess of certain biomolecules– deficiency of vitamin D results in rickets, excess
results in potentially serious hypercalcemia
– Nutritional deficiencies• primary cause - poor diet
• secondary causes - inadequate absorption, increased requirement, inadequate utilization, increased excretion
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Organelle Involvement
• Almost every cell organelle has been involved in the genesis of various diseases
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Different Mechanisms, Similar Effect
• Different biochemical mechanisms can produce similar pathologic, clinical, and laboratory findings– The major pathological processes can be produced by a
number of different stimuli– e.g., fibrosis of the liver (cirrhosis) can result from
chronic intake of EtOH, excess of copper (Wilson’s disease), excess of iron (primary hemochromatosis), deficiency of 1-antitrypsin, etc.
– different biochemical lesions producing similar end point when local concentration of a compound exceeds its solubility point (excessive formation or decreased removal) precipitation to form a calculus
• e.g., calcium oxalate, magnesium ammonium phosphate, uric acid, and cystine may all form renal stone, but accumulate for different biochemical reasons
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Genetic Diseases
• Many disease are determined genetically– Three major classes: (1) chromosomal disorders, (2)
monogenic disorders (classic Mendelian), and (3) multifactorial disorders (product of multiple genetic and environmental factors)
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Genetic Diseases
• Polygenic denotes disorder caused by multiple genetic factors independently of environmental influences
• Somatic disorders - mutations occur in somatic cells (as in many types of cancer)
• Mitochondrial disorders - due to mutations in mitochondrial genome
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Chromosomal Disorders
• Excess or loss of chromosomes, deletion of part of a chromosome, or translocation– e.g., Trisomy 21 (Down syndrome)
• Recognized by analysis of karyotype (chromosomal pattern) of individual (if alterations are large enough to be visualized)
• Translocations important in activating oncogenes– e.g., Philadelphia chromosome - bcr/abl)
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Monogenic Disorders
• Involve single mutant genes• Classification:
(1) autosomal dominant - clinically evident if one chromosome affected (heterozygote)
• e.g., Familial hypercholesterolemia
(2) autosomal recessive - both chromosomes must be affected (homozygous)
• e.g., Sickle cell anemia
(3) X-linked - mutation present on X chromosome• females may be either heterozygous or homozygous
for affected gene• males affected if they inherit mutant gene• e.g., Duchenne muscular dystrophy
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Multifactorial Disorders
• Interplay of number of genes and environmental factors– pattern of inheritance does not conform to classic
Mendelian genetic principles
– due to complex genetics, harder to identify affected genes; thus, less is known about this category of disease
– e.g., Essential hypertension
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Inborn Error of Metabolism
• A mutation in a structural gene may affect the structure of the encoded protein
• If an enzyme is affected, an inborn error of metabolism may result– A genetic disorder in which a specific enzyme is
affected, producing a metabolic block, that may have pathological consequences
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Inborn Error of Metabolism
• A block can have three results:(1) decreased formation of the product (P)
(2) accumulation of the substrate S behind the block
(3) increased formation of metabolites (X, Y) of the substrate S, resulting from its accumulation
• Any one of these three results may have pathological effects
S P Increased S Decreased PE
Normal Block
Increased X,Y
*E
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Inborn Error of Metabolism
• Phenylketonuria - mutant enzyme is usually phenylalanine hydroxylase– synthesize less tyrosine (often fair skinned), have
plasma levels of Phe, excrete phenylpyruvate and metabolites
• If structural gene for noncatalytic protein affected by mutation can have serious pathologic consequences (e.g., hemoglobin S)
Increased phenylalanine Decreased tyrosineBlock
Increased phenylpyruvic acid
*E
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Genetic Linkage Studies• The more distant two genes are from each other on the
same chromosome, the greater the chance of recombination occurring between them
• To identify disease-causing genes, perform linkage analysis using RFLP or other marker to study inheritance of the disease (marker)
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Genetic Linkage Studies
• Simple sequence repeats (SSRs), or microsatellites, small tandem repeat units of 2-6 bp are more informative polymorphisms than RFLPs; thus currently used more
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Methods to clone disease genes
• Functional approach– gene identified on basis of biochemical defect
– e.g., found that phenotypic defect in HbS was GluVal, evident that mutation in gene encoding -globin
• Candidate gene approach– genes whose function, if lost by mutation, could
explain the nature of the disease
– e.g., mutations in rhodopsin considered one of the causes of blindness due to retinitis pigmentosa
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Methods to clone disease genes
• Positional cloning– no functional information about gene product, isolated
solely by it chromosomal position (information from linkage analysis
– e.g., cloning CF gene based on two markers that segregated with affected individuals
• Positional candidate approach– chromosomal subregion identified by linkage studies,
subregion surveyed to see what candidate genes reside there
– with human genome sequenced, becoming method of choice
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Identifying defect in disease gene
• Once disease gene identified, still can be arduous task identifying actual genetic defect
Mutations in CFTR geneStructure of CFTR gene and
deduced protein
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Ethical Issues• Once genetic defect identified, no treatment options may
be available– Will patients want to know?– Is prenatal screening appropriate?– Will identification of disease gene
affect insurability?
• e.g., Hungtington’s disease - mutation due to trinucleotide (CAG) repeat expansion (microsatellite instability)– normal individual (10 to 30 repeats)– affected individual (38 to 120) - increasing length of
polyglutamine extension appears to correlate with toxicity
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Molecular Medicine
• Knowledge of human genome will aid in the development of molecular diagnostics, gene therapy, and drug therapy
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Gene expression in diagnosis• Diffuse large B-cell lymphoma
(DLBCL), a disease that includes a clinically and morphologically varied group of tumors that affect the lymph system and blood. Most common subtype of non-Hodgkin’s lymphoma.
• Performed gene-expression profiling with microarray containing 18,000 cDNA clones to monitor genes involved in normal and abnormal lymphocyte development
• Able to separate DLBCL into two categories with marked differences in overall patient survival.
• May provide differential therapeutic approaches to patients
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Treatment for Genetic Diseases• Treatment strategies
(1) correct metabolic consequences of disease by administration of missing product or limiting availability of substrate
• e.g., dietary treatment of PKU
(2) replace absent enzyme or protein or to increase its activity
• e.g., replacement therapy for hemophilia
(3) remove excess of stored compound
• e.g., removal of iron by periodic bleeding in hemochromatosis
(4) correct basic genetic abnormality• e.g., gene therapy
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Gene Therapy
• Only somatic gene therapy is permissible in humans at present
• Three theoretical types of gene therapy– replacement - mutant gene removed and replace
with a normal gene– correction - mutated area of affected gene would be
corrected and remainder left unchanged– augmentation - introduction of foreign genetic
material into cell to compensate for defective product of mutant gene (only gene therapy currently available)
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Gene Therapy
• Three major routes of delivery of genes into humans(1) retroviruses
• foreign gene integrates at random sites on chromosomes, may interrupt (insertional mutagenesis) the expression of host cell genes
• replication-deficient• recipient cells must be
actively growing forintegration into genome
• usually performed ex vivo
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Gene Therapy
(2) adenoviruses• replication-deficient• does not integrate into host cell genome
– disadvantage: expression of transgene gradually declines requiring additional treatments (may develop immune response to vector)
• treatment in vivo, vector can be introduced into upper respiratory tract in aerosolized form
(3) plasmid-liposome complexes
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Gene Therapy
• Conclusions based on recent gene therapy trials– gene therapy is feasible (i.e., evidence for expression of
transgene, and transient improvements in clinical condition in some cases
– so far it has proved safe (only inflammatory or immune reactions directed toward vector or some aspect of administration method rather than toward transgene
– no genetic disease cured by this method– major problem is efficacy, levels of transgene product
expression often low or transient
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Genetic Medicines
• Antisense oligonucleotides– complementary to specific mRNA
sequence
– block translation or promote nuclease degradation of mRNA, thereby inhibit synthesis of protein products of specific genes
– e.g., block HIV-1 replication by targeting gag gene
• Double-stranded DNA to form triplex molecule