The Hardy-Weinberg Law states that the genotypes in the population from one generation to another generation will be in equilibriym.

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 (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

7. 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)

8. 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

9. 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)

10. 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

11. 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

13. 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 P
E
Normal Block
Increased X,Y
*E

14. 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 tyrosine
Block
Increased phenylpyruvic acid
*E

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)

16. 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
-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

18. 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

19. Identifying defect in disease
gene
 Once disease gene identified, still can be
arduous task identifying actual genetic defect
Mutations in CFTR gene
Structure of CFTR gene and
deduced protein

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., 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

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-
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

23. 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

24. 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 be
actively growing for
integration into genome
 usually performed ex vivo

26. 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

27. 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

28. 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