GENETIC DISEASES

1
Classes of Biomolecules Affected in Disease

• M.Prasad Naidu
• MSc Medical Biochemistry,
• Ph.D.Research Scholar

2
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)

3
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)

4
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

5
Organelle Involvement

• Almost every cell organelle has been involved in
  the genesis of various diseases

6
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 (Wilsons disease), excess of iron
  (primary hemochromatosis), deficiency of
  a1-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

12
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

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Inborn Error of Metabolism
Increased phenylpyruvic acid
?
E
Increased phenylalanine ???? Decreased tyrosine
Block

• 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)

15
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

17
Methods to clone disease genes

• Functional approach
• gene identified on basis of biochemical defect
• e.g., found that phenotypic defect in HbS was
  Glu?Val, evident that mutation in gene encoding
  b-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

20
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., Hungtingtons 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

21
Molecular Medicine

• Knowledge of human genome will aid in the
  development of molecular diagnostics, gene
  therapy, and drug therapy

22
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-Hodgkins 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)

25
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 beactively 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