Signaling Pathways, Cell Fate Determination, and Cancer

Signaling Pathways, Cell Fate Determination, and Cancer

April 28 – May 4, 1996

by Spyros Artavanis-Tsakonas

 

Participants:

Monique Arpin (Institut national de la Santé et de la Recherche Médicale (INSERM), Institut Curie, Paris, France), Spyros Artavanis-Tsakonas (Yale University New Haven, USA), David Baltimore (Massachusetts Institute of Technology, Cambridge, USA), Anton Berns (The Netherlands Cancer Institute, Amsterdam, The Netherlands), Irwin Bernstein (Fred Hutchinson Cancer Center, Seattle, USA), Michael Bishop (University of California, Hooper Foundation, San Francisco, USA), Jacques Camonis (INSERM, Institut Curie, Paris, France), Kathleen (Cho Johns Hopkins University, Baltimore, USA), Jose Costa (Yale University, New Haven, USA), Catherine Dargemont (INSERM, Institut Curie, Paris, France), Eric Fearon (University of Michigan Medical Center, Ann Arbor, USA), Walter J. Gehring (Universität Basel, Biozentrum, Basel, Switzerland), Iswar Hariharan (Massachusetts General Hospital Cancer Center, Charlestown, USA), Ed Harlow( Massachusetts General Hospital Cancer Center, Charlestown, USA), Nick Hastie (Medical Research Center, Human Genetics Unit, Edinburgh, UK), Daniel Louvard (Institut Curie, Paris, France), Matthias Peter( Institut suisse de recherche expérimentale sur le cancer (ISREC), Epalinges, Switzerland), Bruce Ponder (University of Cambridge, GB), Gerald Rubin (University of California, Howard Hugues Medical Institute, USA), Hugues de Thé (Institut Pasteur, Paris, et Hôpital Saint-Louis, Paris, France), Xu Tian (Yale University School of Medicine, New Haven, USA)

Compte rendu:

A new era of therapeutic approaches

The past decade has witnessed a revolution in the molecular analysis of the processes governing the development of multicellular organisms.  The advent of recombinant DNA technology, along with unprecedented genetic analyses of model organisms, has uncovered fundamental principles and identified many of the molecular underpinnings of eukaryotic development.  Moreover, the astonishing conservation of basic biological mechanisms across species barriers is establishing flies, worms and even a unicellular organism, such as yeast, as useful models for human biology.  This gives us truly novel tools to approach and think about pathogenesis by allowing us to view medical problems in a mechanistic way.  There are many indications that a new era of therapeutics, based on the mechanistic understanding of disease, is coming closer. True to the spirit of the Treilles Biological Colloquia, which has been exploring novel biological avenues for many years now, a group of developmental geneticists, molecular biologists and physicians have explored and discussed issues related to how signaling mechanisms dictate cell fate choices, especially as it is related to the genesis of tumors.

Cancer and Developmental Genetics

If one considers cancer as a developmental biological problem, then some common properties of neoplastic cells seem to be emerging.  Notwithstanding the fact that each type of cancer may have a distinct molecular and consequently clinical profile, all cancerous cells belong to proliferative, non-terminally differentiated populations in which normal growth and differentiation programs are improperly implemented.  In addition, there is little doubt that neoplastic cells use normal cell signaling and proliferative mechanisms to maintain themselves.  It seems clear that the development of more rational therapies for cancer will require an understanding of the biochemical anomalies that arise from tumorigenic mutations.  It is just as important, however, to understand the cellular mechanisms responsible for maintaining neoplastic cells in a cancerous state. Therapeutically, it may not be possible to correct an oncogenic mutation, but we may still be able to interfere with the mechanisms which are responsible for maintenance of a neoplastic cell.  One can, for example, induce a cancerous cell to acquire a different differentiation state, thus eliminating their neoplastic qualities. This is the case in acute promyelocytic leukemia where the undifferentiated neoplastic cells are induced to differentiate with retinoic acid, resulting in a complete clinical remission of the disease (Hugues de The). Cancer research today has broadened to include themes that only a few years ago would not be considered relevant.  Using invertebrates to discover novel tumor suppressor genes, using yeast to study cell cycle and signaling pathways, or using Drosophila genetics to analyze pathways involved in human carcinogenesis are all novel approaches that offer a fresh look at fundamental biological and medical problems. The themes covered in the meeting included better molecular characterization of specific cancers, the use of biochemical approaches that afford us a more profound understanding of how certain oncogenes act, and the use of genetic model organisms to dissect and understand mechanisms relevant to carcinogenesis.  One of the most interesting, and potentially most important, links between developmental biology and cancer biochemistry is coming from the study of the mechanisms governing the transition of developmentally uncommitted cells to a more differentiated, mature state.  The combination of developmental genetics, molecular biology and biochemistry is uncovering the biochemical nature of the mechanisms dictating developmental cell fate choices, as well as links between various cell signaling mechanisms hitherto unsuspected.

Signaling and Cell Fates

A fundamental cell interaction mechanism controlling the fate of precursor cells is defined by the Notch signaling pathway, which controls the ability of non-terminally differentiated cells to respond to differentiation and possibly to proliferation signals (Spyros Artavanis-Tsakonas, Michael Bishop, Irwin Bernstein). The molecular dissection of this pathway has been accomplished to a large extent in fruit flies but has been shown to be fully conserved in humans, where malfunction can lead to abnormalities in cell differentiation, including neoplastic conditions.  The manipulation of Notch activity may offer a novel tool for differentiation therapy approaches since it appears to provide an opportunity to effectively manipulate undifferentiated cells.  By the same token, Notch activity may also prove to be useful for inducing the renewal of stem cells. For example, the ex vivo expansion of hematopoietic stem cells would have substantial practical value for generating increased numbers of stem cells in order to overcome myeloablative therapies such as radiation, or, upon genetic modification, for treating genetic diseases.  We heard that this pathway offers a good paradigm of how lower eukaryotic model systems can be used in human biology. The most important experimental tool offered by such lower eukaryotes is the ability to carry out genetic analysis. The studies described by Iswar Hariharan and Matthias Peter who use, respectively, Drosophila and yeast to identify and study genes involved in the arrest of cell cycle, illustrated the power of this approach.  The Drosophila analysis led to the identification of novel genes involved in the control of cell proliferation, while the yeast analysis showed how a gene involved in cell cycle arrest is also capable of controlling the orientation of cell polarity within a morphogenetic gradient, providing a link between signal transduction and cell cycle.  The interplay between distinct signaling pathways, the synergistic interaction between gene activities in the cell, and the pleiotropic developmental action of single genes were recurring themes throughout the colloquium. The current models of carcinogenesis emphasize the successive accumulation of genetic alterations and their role in gene­ rating clonal dominance.  However, little is known about the genetic variation that can be found in tumors during the earliest stages of carcinogenesis and about the selective forces that determine which combination of genes will eventually act in tumors (Jose Costa).  Examination of the synergy between the various genetic factors necessary to attain an oncogenic state is the subject of intensive study. Anton Berns described how insertional mutagenesis, using a particular leukemia virus, is used to accelerate tumorigenesis in mice by inducing successive mutations. Molecular cloning of the corresponding genes allows the identification of genes whose mutant synergistic action can lead to cancer.  The accumulation of multiple genetic alterations required for the induction of solid tumors was also supported by the study of cervical cancers associated with human papillomavirus infections presented by Kathleen Cho.

From Flies to Humans

A gene which has been associated with solid tumors is ras, which encodes a protein essential for the transmission of developmental signals in a variety of cells.  In more than 30% of solid tumors, ras is mutated.  Hence the search for genes integrated in ras-mediated cell signaling has been an area of exceptionally intense study. Most of the ras signaling cascade has been elucidated in flies, worms and yeast, and subsequently shown to be fully conserved in humans. The search for ras pathway genes is important because they can potentially be targets for drugs that interfere with ras signaling in tumors.  Gerald Rubin presented a genetic screen in flies that led to the identification of a novel ras pathway member, showing again the unique power of the genetic approach.

Fruit flies are not only used for dissecting pathways. As we heard from Tian Xu, the ability to genetically manipulate this organism can also serve as an ingenious gene discovery tool. The ability to examine the activity of mutant genes in small tissue patches overcomes the difficulty of analyzing the developmental effects of lethal mutations, which per definition kill the animal and limit our ability to examine mutant phenotypes. The efficient generation of such mosaic animals, where the overproliferation phenotype can be readily identified in patches of mutant cells, is used to identify novel tumor suppressor genes or negative regulators of cell proliferation. Several novel tumor suppressor genes have thus been identified and their human homologues have been isolated (Tian Xu).

Connections between cancer and development continued to be obvious in the discussion of specific genes that have been directly or indirectly associated with tumors. Wilms tumor is a developmental abnormality in kidneys that arises through loss of function of a tumor suppressor gene encoding for proteins involved in transcription as well as in splicing (Nick Hastie). Eric Fearon explained that the involvement of the gene called DCC (Deleted in Colon Cancer) in tumorigenesis is unclear. Moreover, its biochemical role is unclear.  He did demonstrate however that when the human gene is expressed in the developing fly eye, it can disrupt normal development. Such ectopic expressions of heterologous genes can, at least theoretically, provide a unique tool to explore genes of unknown function and help identify and dissect their genetic circuitry.

A powerful reminder that genetic analysis without a thorough biochemical backing has limited value came from Ed Harlow and David Baltimore.  The analysis of the retinoblastoma protein, one of the best understood oncogenes, almost entirely relied on biochemical analysis, which is only now starting to use genetic techniques to further dissect its genetic circuitry (Ed Harlow).  Similarly, the NFkB and CD40 signaling cascades have been largely analyzed biochemically (David Baltimore). In fact, Catherine Dargemont used the biochemical paradigms developed during the study of NFkB signaling to study nucleocytoplasmic shuttling, a phenomenon of central importance in eukaryotic biology.  Finally, Monique Arpin and Daniel Louvard discussed the involvement of the 4.1 superfa­ mily of proteins in tumors. These proteins were shown to be involved in Hepatocyte growth factor (HGF) induced epithelial mesenchymal transition.  The 4.1 family seems to provide a link between surface receptors and the cytoskeleton, influencing cell adhesion and migration, phenomena intimately related to metastasis. In summary, the colloquium provided a platform for fertile discussion.  True to what by now is tradition, the Fondation des Treilles encouraged interdisciplinary dialogue and helped define scientific trends and avenues which are not immediately obvious.

 

Contributions presented

Monique Arpin – The morphogenetic signals transduced by HGF/SF in epithelial cells are mediated by ezrin, a membrane-cytoskeleton linker

Spyros Artavanis-Tsakonas – Notch signaling

David Baltimore – Signaling pathways: CD40 and 5H3

Anton Berns – Identification of cooperating oncogenes

Irwin Bernstein – Regulation of early hematopoietic development

Michael Bishop – Proto-oncogenes and cell signaling

Jacques Camonis – A two hybrid approach as an Ariadne’s thread in the transduction labyrinth: joy and beguilement

Kathleen Cho – The role of human papillomaviruses in cell fate determination in cervical cancer

José Costa – Microevolution and tumor formation

Catherine Dargemont – Nucleocytoplasmic transport: mechanisms and role in the regulation of transcription

Eric Fearon – Studies of the DDC gene in cancer and development

Walter Gehring – Induction on the eye morphogenetic pathway

Iswar Hariharan – Regulation of cell proliferation and differentiation in Drosophila

Ed Harlow – The retinoblastoma protein

Nick Hastie – Wilms tumor: a case of disrupted development

Daniel Louvard – Role of membrane-microfilament interactions in cell polarity and signaling

Matthias Peter – Signal transduction and cell cycle arrest in response to mating pheromones in yeast

Bruce Ponder – Tumor and developmental syndroms associated with mutation of the RET receptor tyrosine kinase

Gerald Rubin – Ras-mediated signaling in Drosophila

Hugues de The – Physiopathological models of acute promyelocytic leukemia

Tian Xu – Tumor suppressors and Drosophila genetics

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Articles disponibles dans les archives de la Fondation des Treilles / Articles available in the archives of the Fondation des Treilles 

Mulligan, Lois M. et al.
Germ-line mutations of the RET proto-oncogene in multiple endocrine neoplasia type 2A
Nature, Vol. 363, pp. 458-160, June 3, 1993

Mulligan, Lois M. et al.
Specific mutations of the RET proto-oncogene are related to disease phenotype in MEN 2A and FMTC
Nature Genetics, Vol. 6, pp. 70-74, January 1994

Mulligan, Lois M. et al.
Diverse phenotypes associated with exon 10 mutations of the RET proto-oncogene
Human Molecular Genetics, Vol. 3, N° 12, pp. 2163-2167, 1994

Eng, Charis et al.
Mutation of the RET Proto-oncogene in Sporadic Medullary Thyroid Carcinoma
Genes, chromosomes & cancer, Vol. 12, pp. 209-212, 1995

Eng, Charis et al.
A novel point mutation in the tyrosine kinse domain of the RET proto-oncogene in sporadic medullary thyroid carcinoma and in a family with FMTC
Oncogene, Vol. 10, pp. 509-513, 1995

Lorenzo, Maria Jesus et al.
Multiple mRNA isoforms of the human RET proto-oncogene generated by alternate splicing
Oncogene, Vol. 10, pp. 1377-1383, 1995

Eng, Charis et al.
Low frequency of germine mutations in the RET proto-oncogene in patients with apparently sporadic medullary thyroid carcinoma
Clinical Endocrinology, Vol. 43, pp. 123-127, 1995

Mak, Yin F. – Ponder, Bruce A. J.
RET oncogene
Current opinion in Genetics & Development, Vol. 6, pp. 82-86, 1996

Bishop, J. Michael
Cancer: the rise of the genetic paradigm
Genes and Development, Vol. 9, pp. 1309-1315, 1995

Doyle, Helen J. – Bishop, J. Michael
Torso, a receptor tyrosine kinase required for embryonic pattern formation, shares substrates with the Sevenless and EGF-R pathways in Drosophila
Genes and Development, Vol. 7, pp. 633-646, 1993

Bishop, J. Michael
Molecular Themes in Oncogenesis
Cell, Vol. 64, pp. 235-248, January 25, 1991 

Vetter, Monica L. – Bishop, J. Michael
β PDGF receptor mutants defective for mitogenesis promote neurite outgrowth in PC12 cells
Current Biology, Vol. 5, No 2, pp. 168-178, 1995

Reynolds, Lucinda F. – Eng, Charis
RET mutations in multiple endocrine neoplasia type 2 and Hirschsprung disease
Current opinion in Pediatrics, Vol. 7, pp. 702-709, 1995

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