1. Growth Factors, Receptors, and Signal Transduction I
Tumor Biology: Cellular and Molecular
Aspects of the Transformed Cell
Growth Factors, Receptors,
and Signal Transduction I
Rebecca Riggins

2. Overview Peptide growth factors Growth factor receptors
- structure and function
Receptor activation and its consequences
Growth factor and receptor defects in cancer
Targeted therapies to growth factors, receptors
Intracellular signaling “downstream” of receptors
- key pathways: Ras/MAPK, Src, PI3K
Intracellular signaling defects in cancer
Targeted therapies to intracellular signaling molecules
TNF and TRAIL

3. Polypeptide Growth Factors
small peptides, or proteins
interact specifically with a receptor on the cell surface
secreted by multiple cell types
most are secreted as inactive precursors that need
to be cleaved or fragmented before they can act
some are plasma membrane-bound or found in the
extracellular matrix
transported by blood and lymphatic systems by several
different routes…3 major ones we will discuss are
Endocrine
Paracrine
Autocrine

4. Endocrine: Synthesis by specific cells, target (receptor) is far away
Autocrine
Paracrine
Endocrine
Juxtacrine
BLOOD VESSEL
Intracrine
Different modes of action for growth factors (Bafico and Aaronson, Cancer Medicine, 2002)

5. Paracrine: Synthesis by many cell types, receptor is nearby
Autocrine
Paracrine
Endocrine
Juxtacrine
BLOOD VESSEL
Intracrine
Different modes of action for growth factors (Bafico and Aaronson, Cancer Medicine, 2002)

6. Autocrine: Synthesis by many cell types, receptor is on same cell
that synthesized
the growth factor
Autocrine
Paracrine
Endocrine
Juxtacrine
BLOOD VESSEL
Intracrine
CANCER!
Different modes of action for growth factors (Bafico and Aaronson, Cancer Medicine, 2002)

7. GROWTH FACTOR FUNCTIONS IN VIVO
1. Early development
2. Tissue differentiation
3. Wound healing and tissue repair
4. Immune responses
5. Stromal mediators of sex and other hormones
. Aaronson, Growth factor and receptor tyrosine kinases. Sci. STKE 2005, tr6 (2005).

8. GROWTH FACTOR FUNCTIONS IN VITRO
1. Proliferation
2. Differentiation
3. Chemo-attraction
4. Cell death
5. Cell migration
. Aaronson, Growth factor and receptor tyrosine kinases. Sci. STKE 2005, tr6 (2005).

9. REGULATION OF GROWTH FACTOR/RECEPTOR INTERACTIONS
1. Determined by growth factor availability and receptor expression levels
2. Different modes of growth factor action - autocrine, paracrine, other
3. Secretory properties - secretory signal; proteoglycan or serum protein binding
4. More than one member of same growth factor gene family may act on the same receptor
5. Same growth factor may cluster more than one receptor member of the same receptor family
6. Interactions regulated by alternative growth factor/receptor products
. Aaronson, Growth factor and receptor tyrosine kinases. Sci. STKE 2005, tr6 (2005).

10. ErbB/HER Family Epidermal Growth Factor Receptor (EGFR)
Seminars in Cancer Biology, Volume 14, Issue 4, August 2004, Pages

11. Other Growth Factors platelet-derived growth factor (PDGF)
nerve growth factor (NGF)
hepatocyte growth factor (HGF)
insulin-like growth factor (IGF)
fibroblast growth factor (FGF)
vascular endothelial growth factor (VEGF)
named according to what they do or where they act
most stimulate growth, or are mitogenic

12. Transforming Growth Factor (TGF)-b
subgroup of a large family…25 members
TGF-b1 is the most studied in cancer
in some cells, TGF-b1 is mitogenic, while in others it
actually inhibits growth
TGF-b can cooperate with PDGF…
Smooth muscle (mesenchymal)
TGF-b increases PDGF,
cells proliferate
Keratinocyte (epithelial)
TGF-b inhibits
cell proliferation
No PDGFR

13. Tumor Necrosis Factor (TNF)
again, in some cells, TNF is mitogenic, but in
most it promotes cell death, or apoptosis
this growth factor is not soluble, but is inserted into
the plasma membrane
Juxtacrine
Intracrine

14. Growth Factor Receptors
all are transmembrane proteins
3 major features:
extracellular domain (ectodomain)
transmembrane region
intracellular domain
where, when, and
how they are expressed
determines their biological
function

15. Figure 5.10 The Biology of Cancer (© Garland Science 2007)

16. Growth Factor Receptors
catalytic domain has kinase activity
kinases add phosphate groups to (phosphorylate)
specific amino acids
Receptor Tyrosine kinases phosphorylate Tyrosine
Receptor Serine/Threonine kinases phosphorylate
Serine and/or Threonine
ATP-binding
Phospho-transferase
ATP P
Amino Acid P
ADP
Autophosphorylation: receptor phosphorylates itself

17. Growth Factor Receptors
Transphosphorylation: receptor phosphorylates its
binding partner
Figure The Biology of Cancer (© Garland Science 2007)

18. “Different” Growth Factor Receptors
Some receptors have no catalytic domain…
TNF receptors rely instead on other intracellular
proteins to transmit their signal
Adhesion receptors (integrins) also do this

19. Integrins bind Extracellular Matrix (ECM)

20. G protein-coupled receptors
Another Example:
G protein-coupled receptors
Have 7 transmembrane domains
Ligand may bind to extracellular OR transmembrane region (depending on receptor)
Also do not have a catalytic domain

21. GPCR - Inactive State
Figure 5.25a The Biology of Cancer (© Garland Science 2007)

22. GPCR – Activation Process
Figure 5.25bc The Biology of Cancer (© Garland Science 2007)

23. However, “classical” growth factor receptors have catalytic domains
that must be activated

24. Receptor Tyrosine Kinase (RTK) Activation
Homodimer: 2 identical receptors cluster together
Heterodimer: 2 different receptors from the same
family cluster together
Nature Reviews Cancer, Vol. 5, May 2005, pg

25. RECEPTOR ACTIVATION BY GROWTH FACTORS
1. Growth factors induce receptor clustering a. High affinity binding b. Ligand mediated receptor cross-linking c. Ligand/receptor crystal structures
2. Receptor activation by homodimer or heterodimer formation a. Activation requires the ability to cross-link
. Aaronson, Growth factor and receptor tyrosine kinases. Sci. STKE 2005, tr6 (2005).

26. Tyrosine Kinase Receptor Activation by Dimerization
EGFR
Figure 1. Ligand Binding Stabilizes the Formation of Activated Dimers(A) Inactive receptor monomers (green) are in equilibrium with inactive (green) or active (blue) receptor dimers. The active receptor dimers exist in a conformation compatible with trans-autophosphorylation and stimulation of PTK activity (blue). Ligand binding stabilizes active dimer formation and hence PTK activation.(B) Inactive disulfide bridged insulin-receptor (IR) dimers (green) are in equilibrium with active dimers (blue). Insulin binding stabilizes the active dimeric state leading to PTK activation.
Insulin receptor
Schlessinger J., Cell Oct 13;103(2):

27. Consequences of RTK activation
GROWTH
FACTOR
PIP3
PIP3
RTK
SOS
Grb2
RAS
RAS P P P
PI3K
p85
p110
Akt
PDK1
Raf P
MEK P P
GSK3 P P P P
BAD
NF-ĸB
MDM2
p70S6K
FKHR P
ERK
PROLIFERATION
CELL SURVIVAL
PROTEIN SYNTHESIS
. Aaronson, Growth factor and receptor tyrosine kinases. Sci. STKE 2005, tr6 (2005).

28. RTK Signaling in Cancer
Autocrine Transforming Loops
Receptor Gene Amplification
Receptor Gene Mutation
Paracrine Acting Growth Factors in Tumor Progression
. Aaronson, Growth factor and receptor tyrosine kinases. Sci. STKE 2005, tr6 (2005).

29. Figure 5.12b The Biology of Cancer (© Garland Science 2007)

30. Figure 5.12a The Biology of Cancer (© Garland Science 2007)

31. Historical Perspective

32. In the beginning… Oncogenic retroviruses caused tumors in animal
models (monkeys, chickens, etc.)
These viruses contained transforming genes that
had nothing to do with the viral life cycle…

33. Growth Factor Oncogenes
eg. PDGF
Cancer cells, and cells in culture that were transformed
by simian sarcoma virus (SSV), were less dependent
on PDGF for growth
PDGF-B was cloned…its sequence was very close to
that of the v-sis oncogene from SSV
Thus, cellular PDGF-b (or c-sis-B) is a protooncogene
Other oncogenic growth factors include the FGFs:
FGF3 – activated by mouse mammary tumor virus insertion
FGF4 – cloned from Kaposi’s sarcoma virus (skin tumors assoc.
with HIV/AIDS)
FGF5 – originally isolated from DNA of bladder tumors

34. Receptor Oncogenes Mechanisms
v-erbB = mutated EGFR (avian erythroblastosis virus)
v-fms = viral form of CSF-1 receptor
v-kit = viral form of c-kit receptor
Mechanisms
deletions, and/or insertions (v-erbB, v-kit)
chromosomal translocation, fusions with other proteins
“Neu” mutation of erbB2 = change of Val → Glu in
the transmembrane domain…constitutive dimerization,
activation

35. Table 5.1 The Biology of Cancer (© Garland Science 2007)

36. Table 5.2 The Biology of Cancer (© Garland Science 2007)

37. Table 5.3 The Biology of Cancer (© Garland Science 2007)

38. Receptors can be good targets:
Easy Access!
Seminars in Cancer Biology, Volume 14, Issue 4, August 2004, Pages

39. Targeted Therapy in Clinical Use
Nature Reviews Cancer, Vol. 5, May 2005, pg

40. Understanding Targeted Therapy:
ErbB receptor family
Herceptin: anti-ErbB2
Erbitux: anti-EGFR
Iressa, Tarceva :
EGFR kinase inhibitor

41. Consequences of Inhibitor Action
in vitro, these drugs can individually inhibit cell growth
and induce tumor shrinkage in mouse models
However, cooperation between ErbB family members
in vivo means one inhibitor is usually not enough
metastatic breast cancer, positive for ErbB2
expression…Herceptin used alone = 34% response rate
Herceptin + cytotoxic chemotherapy drugs = better than
either drug alone…BUT there are many side effects
in cell culture studies, Herceptin + Tarceva yield much
better response rates

42. UCLA Clinical Trial open to women with metastatic breast cancer
must demonstrate amplified ErbB2 (excess gene copies)
will be treated with Herceptin and Tarceva
Phase I study: 14 patients, all received H+T…T at
various doses to determine the best
3 patients of 14 showed partial response or
stable disease…21%
Phase II study: standard Herceptin + 150mg/day Tarceva

43. Eastern Cooperative Oncology Group Trial
open to women with metastatic breast cancer
must demonstrate amplified ErbB2 (excess gene copies)
will be treated with Herceptin and Iressa
Phase I study: standard Herceptin + 250mg/day Iressa is
both active and well-tolerated by patients
Phase II study: still ongoing

44. New Developments 2nd-generation tyrosine kinase inhibitors that target
BOTH EGFR and ErbB2…pan-ErbB inhibitors
ATP-binding
Phospho-transferase
Highly conserved between EGFR and ErbB2
Canertinib, Lapatinib, others…target ATP binding site
Lapatinib is active in breast cancer cells that have
acquired resistance to Herceptin
In phase I/II trials, Lapatinib has also shown some effect
in patients resistant to Herceptin and Iressa

45. More New Developments
blocking the interaction of receptor family members
Pertuzumab = antibody against ErbB2, prevents
dimerization with other receptors, ie. EGFR
This may be active in tumors without over-expressed or
amplified ErB2…currently used in trials for ovarian,
prostate, islet cell cancer

46. A few words about VEGF…
VEGF interactions with its receptor are critical for
angiogenesis (formation of new blood vessels by tumors)
anti-VEGF antibodies block the growth factor, not the
receptor
Could be effective in treating both the primary tumor
and the metastases that develop elsewhere in the body
Currently, Bevacizumab is approved for treatment of
metastatic colon cancer, and is undergoing phase I/II
clinical trials in breast and other cancers

47. On Wednesday Intracellular signaling “downstream” of receptors
- key pathways: Ras/MAPK, Src, PI3K
Intracellular signaling defects in cancer
Targeted therapies to intracellular signaling molecules
TNF