Causes

1
Causes

• cancer cells arise due to progressive mutations
  of DNA
• causes
• inherited mutations
• viruses, chemicals, radiation
• mutations involve 2 types of genes
• proto-oncogenes dormant forms of cancer-causing
  genes
• tumor-suppressor genes required for
  differentiation into mature cell types

2
Normal vs. Cancer Cells

• Specialized with specific form and function
• Enter the cell cycle about 50x, then die
• Normal nuclei with 46 chromosomes
• Adhere to neighbor cells exhibit contact
  inhibition
• Require growth factors for survival

• Not specialized dont contribute to normal
  functioning of body part
• Enter cycle repeatedly are immortal
• Abnormal nuclei with deletions / amplif. of
  genes abnormal chrom.
• No restraint will pile on top of one another
• Reduced need for growth factors

3
Hormonal regulation of tumor cell proliferation

• Hormonal regulation of cancer cell proliferation
• Estrogen-dependent mammary tumors
• Melatonin
• GnRH

4
GnRH

• Gonadotropin-releasing hormone
• Role in hypothalamic-pituitary-gonadal axis
• Extrapituitary expression of GnRH receptors
• Presence of GnRH receptors in breast tumor cells
  (including MCF-7 cells)
• Antiproliferative effect of GnRH on mammary tumor
  cells
• Indirect via decreased estrogen production
• Direct induction of apoptosis in tumor cells

5
nutrition
stress
day length
Brain (hypothalamus)
GnRH

LH, FSH
Pituitary

Gonads
gametes, sex hormones
6
Types of cell death

• Apoptosis
• Programmed cell death
• Internal vs. external stimuli
• Necrosis
• Response to tissue damage
• Cells swell, burst, release intracellular contents

7
Characteristics of cells undergoing apoptosis

• Shrinkage in cell, nuclear volume
• Loss of adhesion
• Formation of blebs at surface
• Chromatin (DNA) fragmentation
• Figure 15.32

8
Role of caspaces in cell death

• Caspaces proteolytic enzymes activated during
  apoptosis
• Targets
• Protein kinases
• Lamins
• Cytoskeleton
• CAD (caspace-activated DNAse)
• DNA repair enzymes

9
Role of Bcl-2 family of proteins in apoptosis

• Made of 2 subgroups of proteins
• Those that promote apoptosis (Bad, Bax)
• Those that antagonize apoptosis (Bcl-x, Bcl-w,
  Bcl2)

10
Melatonin

• Hormone secreted by pineal gland
• Driven by circadian clock (in suprachiasmatic
  nucleus of hypothalamus)
• Circadian pattern of secretion
• Physiological roles
• Sleep, mood, oncostatic properties

11
Effects of melatonin on MCF-7 breast cancer cells

• Inhibits proliferation
• Prevention of replication or induction of
  apoptosis?
• Indirect (antiestrogen) or direct effect?
• P53 effect disruption of mitochondrial
  respiration
• Effective only in absence of estrogen
• Clinical studies
• Increased efficacy of chemotherapy in patients
  with poor clinical status
• 1 year survival rate 42/124 vs. 19/126

12
Our studies

• Exploration of GnRH / melatonins effects on
  apoptosis
• Expt 1 induction of caspase activity
• Expt 2 cell proliferation assay
• Expt 3 DNA fragmentation assay
• Exploration of cell signaling pathway used by
  GnRH / melatonin to induce cell death??
• PKC inhibitor

13
Role of PKC pathway in GnRH-induced apoptosis of
MCF-7 cells

• Role of PKC in mediating GnRH effects in
  pituitary
• Same GnRH receptor is expressed in mammary tissue
• Role of phorbol esters in inhibiting MCF-7 cell
  proliferation

14
Assignment for next week

• Write introduction for lab report on research
  project, using papers on reserve at Cofrin
  Library and lecture notes
• Assignments must be typewritten
• Clear statement of hypotheses and expected
  outcomes at the end of introduction