Normal haemopoiesis
ABNORMALITIES IN THE HEMOPOIETIC SYSTEM
• CAN LEAD TO
• HEMOGLOBINOPATHIES
• HEMOPHILIA
• DEFECTS IN HEMOSTASIS/THROMBOSIS
• HEMATOLOGICAL MALIGNANCY
MUTATIONS AND DNA
• VARIOUS TYPES OF MUTATIONS CAN OCCUR LEADING TO DISEASE PHENOTYPE
• POINT MUTATIONS• INSERTIONS OR DELETIONS• TRANSLOCATIONS• COMPLEX CHROMOSOMAL
REARRANGEMENTS
EXAMPLE OF COMMON MUTATIONS IN HUMAN DISEASE
Sickle cell disease
Sickle cell disease, morphology and molecular
VARIABILITY IN GENETIC DISEASES
•ONE DISEASE, ONE GENE, ONE MUTATION
•ONE DISEASE, ONE GENE, MANY MUTATIONS
•ONE DISEASE, MORE THAN ONE GENE, MANY •MUTATIONS
HAEMOPHILIA
X LINKED RECESSIVE DISORDER
HAEMOPHILIA A – MUTATIONS IN FACTOR VIII GENE
HAEMOPHILIA B – MUTATIONS IN FACTOR IX GENE
SIMPLE AND COMPLICATED MUTATIONS
THE FLIP TIP MUTATION
F8A
E1 E22 E23 E26
F8A
E1 E22 E23 E26
TEL CEN
CEN
CENTEL
TEL
INVERSION 22
E1E22 E23 E26
A
B
C
FIGURE 4 THE IVS 22 MUTATION IN HAEMOPHILIA A.
F8B
Genetic factors and deep vein thrombosis
• FACTOR V LEIDEN MUTATION• PROTHROMBIN MUTATION• ? OTHER FACTORS IN THE PROTEIN C
PATHWAY• FVL LEADS TO SIGNIFICANT INCREASE IN
RISK OF DVT, PARTICULARLY IN ASSOCIATION WITH OTHER ENVIRONMENTAL FACTORS EG OCP
CANCER DEVELOPMENT: ITS IN THE GENES
HOW DOES A CELL BECOME TUMORIGENIC?
• THREE PROCESSES ARE INVOLVED
• IMMORTALISATION
• TRANSFORMATION
• METASTASIS
IMMORTALISATION
• PROCESS BY WHICH THE CELLS ARE INDUCED TO GROW INDEFINITELY
TRANSFORMATION
• CELLS ARE NOT CONSTRAINED IN TERMS OF GROWTH CHARACTERISTICS AND TEND TO BECOME FACTOR INDEPENDENT
METASTASIS
• CANCER CELLS GAIN THE ABILITY TO INVADE NORMAL TISSUE AND ESTABLISH OTHER FOCI OF MALIGNANCY
WHAT CAUSES CELL TRANSFORMATION?
• ENVIRONMENTAL• CARCINOGENS(INITIATORS AND
PROMOTERS)
• GENETIC• SOMATIC MUTATIONS• MENDELIAN INHERITANCE
ONCOGENES• NORMAL CELLULAR
COUNTERPARTS(PROTO-ONCOGENES)
• MUTATION/ACTIVATION LEADS TO TUMOR FORMATION
• HUNDREDS OF ONCOGENES IDENTIFIED
• GAIN OF FUNCTION
Tumour suppressor genes
• Originally known as recessive oncogenes
• Need to have both copies of the gene affected to promote a malignant phenotype
• Knudsons 2 hit hypothesis
• First mutation makes cells susceptiple to development of cancer
• 2nd hit leads to a malignant phenotype
TRANSLOCATIONS AND CANCER
• SEEMS PARTICULARLY RELEVANT IN HEMATOLOGICAL MALIGNANCIES
• CHRONIC MYELOID LEUKEMIA
• ACUTE PROMYELOCYTIC LEUKEMIA
• BURKITTS LYMPHOMA
• NON HODGKINS LYMPHOMA
Leukaemia, the current hypothesis
• Defect in maturation of white blood cells• May involve a block in differentiation and/or a
block in apoptosis• Acquired genetic defect• Initiating events unclear• Transformation events involve acquired genetic
changes• Chromosomal translocation implicated in many
forms of leukaemia
Chronic Myeloid Leukaemia• Malignancy of the haemopoietic system• Transformation of the pluripotent stem cell• 9;22 translocation giving rise to the Philadelphia
(Ph’) chromosome• Creation of a leukaemia specific mRNA (BCR-
ABL)• Resistance to apoptosis, abnormal signalling and
adhesion• Molecular diagnostics• Molecular and cellular therapeutics
Cytogenetic Abnormality of CML:The Ph Chromosome
1 2 3 4 5
6 7 8 10 119 12
13 14 15 16 17 18
19 20 21 22 x Y
The Ph Chromosome: t(9;22) Translocation
22
bcr
abl
Ph ( or 22q-)
bcr-abl
FUSION PROTEINWITH TYROSINEKINASE ACTIVITY
9 9 q+
Prevalence of the Ph Chromosome in Haematological Malignancies
Leukaemia % of Ph+ Patients
CML 95
ALL (Adult) 15–30
ALL (Paediatric) 5
AML 2
Faderl S et al. Oncology (Huntingt). 1999;13:169-184.
bcr-abl Gene and Fusion Protein Tyrosine Kinases
Adapted from Melo JV. Blood. 1996;88:2375-2384.
p210Bcr-Abl
p185Bcr-Abl2-11
2-11
Chromosome 9
c-bcr
Chromosome 22
c-abl
Exons
Introns
CML Breakpoints
ALL Breakpoints
1
2-11
NON HODGKINS LYMPHOMA
• B CELL FOLLICULAR LYMPHOMA
• t(14;18)(q21;q14)
• BCL 2 AND IMMUNOGLOBULIN GENES INVOLVED
• DYSREGULATION OF BCL 2
• FAILURE OF APOPTOSIS
Detecting Cancer – where to begin?
Detecting cancer, the need for a marker of disease
Detecting Cancer – different markers for different diseases?
Cancer Molecular Diagnostics – discriminating cancers at the gene level
How Cancer Molecular Diagnostics?
• Chromosome analysis
• Gene analysis
• Gene expression analysis
• Protein analysis
• Gene chip analysis
Leukaemia diagnostics
• Morphology
• Cytogenetics
• Fluorescent In Situ Hybridisation (FISH)
• Immunophenotyping
• PCR of chromosomal translocations
New developments in Cancer Molecular DiagnosticsThe Gene Chip
The Gene Chip, a Molecular snap shot of the cell
MOLECULAR MEDICINE
• A new approach to medicine
• New Diagnostics
• New Therapeutics
• A number of agents now in clinical trials
• Molecular medicine will help identify new targets and permit rational drug development
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