|
 |
|
Martin McMahon, PhD
Oncogenes, signal transduction and cancer
|
|
|
Work in my laboratory focuses on the role of oncogenes
and tumor suppressors in the aberrant proliferation of cancer cells.
The past 25 years have seen enormous progress in the elucidation of
the fundamental mechanisms by which normal cells are converted to
a tumorigenic phenotype. The general consensus is that activation
of oncogenes such as RAS, accompanied by loss of function of tumor
suppressor genes such as TP53 promotes the conversion of normal cells
to a neoplastic phenotype. Alterations in oncogenes and tumor suppressor
genes leads to subversion of the machinery that controls the cell
division cycle, cell senescence and the process of programmed cell
death (apoptosis).
The Ras-family of membrane associated GTPases transmit signals into
the interior of the cell by the activation of a number of cytosolic
signal transduction pathways. Prominent among these is the RafÆMEKÆERK
MAP kinase signaling pathway. Binding of Raf to activated Ras leads
to activation of Raf protein kinase activity. Activated Raf phosphorylates
to activate a second protein kinase MEK, which in turn phosphorylates
to activate the MAP kinases ERK1 and 2. Activated ERKs are pleiotropic
modulators of cell physiology that elicit their effects by phosphorylating
numerous proteins including several transcription factors. Using conditionally
active forms of Raf (DRaf:ER) that permit selective activation of
the ERK MAP kinase pathway in cells we have explored the regulation
of gene expression by this pathway. It is clear that the RafÆMEKÆERK
pathway can contribute to many of the phenotypes of the cancer cell
by regulating genes involved in the cell division cycle (cyclin D1,
p21Cip1), apoptosis (Mdm2, HB-EGF), cell invasion (avb3-integrin),
epithelial cell multilayering and angiogenesis (VEGF).
In 1997 an intriguing connection between oncogene activation and tumor
suppressor gene expression was uncovered. Although oncogenes such
as Ras and Raf came to view as agents of neoplastic transformation,
these genes can also have effects that run counter to oncogenic transformation,
such as the arrest of the cell division cycle and the induction of
cell senescence. Ras and Raf-induced senescence is mediated by proteins
such as p53 and p16INK4A, tumor suppressors that are frequently mutated
in human cancer cells expressing activated Ras or Raf proteins. It
is possible the effects of Ras and Raf on cell cycle arrest/senescence
may be a defense mechanism against neoplastic transformation of normal
cells when the RasÆRafÆMEKÆERK signaling pathway
is inappropriately active. Hence, in order for cancer cells that express
an activated form of Ras to progress, they must silence the expression
of tumor suppressors such as p53 and p16INK4A. The best examples of
this are in human pancreatic cancer and melanoma where the extremely
high frequency of Ras (~95%) or B-Raf mutation (~70%) respectively
is accompanied by an equally high frequency of mutation or gene silencing
of p16INK4A expression (~99%).
Although the molecular genetics of pancreatic cancer and melanoma
have been explored in some detail, there is a large gulf in our understanding
of how mutations in oncogenes and tumor suppressors influence the
aberrant behavior of these tumors. Furthermore, both pancreatic cancer
and melanoma are diseases for which there is an urgent need for new
diagnostic and therapeutic tools. Consequently we have initiated a
series of new projects to understand the role of oncogenes and tumor
suppressors in the initiation and progression of both pancreatic cancer
and melanoma in more detail. We are taking three main approaches to
explore fundamental aspects of the cell and molecular biology of these
diseases:
1. Understanding the cell of origin.
To understand the aberrant properties of the cancer cell, we need
to know something about the properties of the normal cells from which
the cancer cells are derived. To do this we have established conditions
that allow the in vitro culture of human pancreatic ductal epithelial
cells (PDEC) and melanocytes, the cells from which pancreatic cancer
and melanoma arise respectively. By expression of telomerase or other “immortalizing” genes, we are attempting to isolate long-term
cultures of these cells 8. Such cell lines will then be subjected
to an in-depth analysis of the regulation of gene expression with
particular emphasis on the control of the cell division cycle, apoptosis
and senescence. In addition, we will use these cell lines as recipients
in gene transfer experiments to explore the effects of activated Ras
and Raf on human cells.
2. Understand the altered properties of the cancer cell.
Using human pancreatic cancer and melanoma cell lines, we are using
high throughput profiling techniques to explore the genetic alterations
and gene expression changes that occur in the initiation and progression
of pancreatic cancer and melanoma. We are also assessing the effects
of inhibitors of various cell signaling pathways on cancer cell physiology.
Moreover, as new techniques become available to scan the proteome
of human cancer cells, we will apply the full spectrum of high throughput
profiling techniques to understand how alterations in the patterns
of mRNA and protein expression contribute to the aberrant properties
of these cells. Although the goal of this research is to understand
the biology of the cancer cell, we anticipate that this analysis may
ultimately lead to the identification of candidate diagnostic and
therapeutic targets to aid in the management of these diseases
3. Mouse models of pancreatic cancer and melanoma
To explore the initiation and progression of pancreatic cancer and
melanoma in a true in vivo setting, we are deriving transgenic mice
with tissue specific expression of oncogenes that will confer an inherited
pre-disposition either to pancreatic cancer or melanoma. These mice
will then be bred to other transgenic/knock-out mice to explore the
role of specific oncogenes and tumor suppressors in the genesis and
progression of pancreatic cancer and melanoma. These studies will
focus in particular on genes that regulate the cell division cycle,
apoptosis and senescence. Clearly, transgenic mouse models that accurately
recapitulate the features of human cancer will be very useful tools
in understanding the earliest stages of the initiation and progression
of these diseases. Such mice may also be useful in testing new therapies
to target these diseases.
|
|
McMahon, M. Steroid receptor fusion proteins
for conditional activation of Raf-MEK- ERK signaling pathway. Methods
Enzymol 332, 401-17 (2001).
Woods, D. et al. Raf-induced proliferation or cell
cycle arrest is determined by the level of Raf activity with arrest
mediated by p21Cip1. Mol Cell Biol 17, 5598-611 (1997).
Schulze, A., Lehmann, K., Jefferies, H. B., McMahon,
M. & Downward, J. Analysis of the transcriptional program induced
by Raf in epithelial cells. Genes Dev 15, 981-94. (2001).
Hansen, S. H. et al. Induced expression of Rnd3 is associated with
transformation of polarized epithelial cells by the RafÆMEKÆERK
pathway. Mol Cell Biol 20, 9364-75. (2000).
Ries, S. et al. Opposing effects of Ras on p53: transcriptional
activation of mdm2 and induction of p19ARF. Cell 103, 321-30. (2000).
Woods, D. et al. Induction of beta3-integrin gene expression by
sustained activation of the Ras-regulated Raf-MEK-extracellular
signal-regulated kinase signaling pathway. Mol Cell Biol 21, 3192-205.
(2001).
Zhu, J., Woods, D., McMahon, M. & Bishop, J. M. Senescence of
human fibroblasts induced by oncogenic Raf. Genes Dev 12, 2997-3007
(1998).
Venetsanakos, E. et al. Induction of tubulogenesis in telomerase-immortalized
human microvascular endothelial cells by glioblastoma cells. Exp
Cell Res 273, 21-33. (2002)
McMahon, M and Woods, D. Regulation of the p53 pathway by Ras, the
plot thickens. BBA Reviews on Cancer Online, 1461: M63-M71 (2001)
information last updated February 2003 |
|
|
|
|
|
|
