Allan Balmain

TGF-|[beta]| signaling in tumor suppression and cancer progression

Epithelial and hematopoietic cells have a high turnover and their progenitor cells divide continuously, making them prime targets for genetic and epigenetic changes that lead to cell transformation and tumorigenesis. The consequent changes in cell behavior and responsiveness result not only from genetic alterations such as activation of oncogenes or inactivation of tumor suppressor genes, but also from altered production of, or responsiveness to, stimulatory or inhibitory growth and differentiation factors. Among these, transforming growth factor β (TGF-β) and its signaling effectors act as key determinants of carcinoma cell behavior. The autocrine and paracrine effects of TGF-β on tumor cells and the tumor micro-environment exert both positive and negative influences on cancer development. Accordingly, the TGF-β signaling pathway has been considered as both a tumor suppressor pathway and a promoter of tumor progression and invasion. Here we evaluate the role of TGF-β in tumor development and attempt to reconcile the positive and negative effects of TGF-β in carcinogenesis.

A Mutant-p53/Smad Complex Opposes p63 to Empower TGFβ-Induced Metastasis

TGFbeta ligands act as tumor suppressors in early stage tumors but are paradoxically diverted into potent prometastatic factors in advanced cancers. The molecular nature of this switch remains enigmatic. Here, we show that TGFbeta-dependent cell migration, invasion and metastasis are empowered by mutant-p53 and opposed by p63. Mechanistically, TGFbeta acts in concert with oncogenic Ras and mutant-p53 to induce the assembly of a mutant-p53/p63 protein complex in which Smads serve as essential platforms. Within this ternary complex, p63 functions are antagonized. Downstream of p63, we identified two candidate metastasis suppressor genes associated with metastasis risk in a large cohort of breast cancer patients. Thus, two common oncogenic lesions, mutant-p53 and Ras, selected in early neoplasms to promote growth and survival, also prefigure a cellular set-up with particular metastasis proclivity by TGFbeta-dependent inhibition of p63 function.

The genetics and genomics of cancer

The past decade has seen great strides in our understanding of the genetic basis of human disease. Arguably, the most profound impact has been in the area of cancer genetics, where the explosion of genomic sequence and molecular profiling data has illustrated the complexity of human malignancies. In a tumor cell, dozens of different genes may be aberrant in structure or copy number, and hundreds or thousands of genes may be differentially expressed. A number of familial cancer genes with high-penetrance mutations have been identified, but the contribution of low-penetrance genetic variants or polymorphisms to the risk of sporadic cancer development remains unclear. Studies of the complex somatic genetic events that take place in the emerging cancer cell may aid the search for the more elusive germline variants that confer increased susceptibility. Insights into the molecular pathogenesis of cancer have provided new strategies for treatment, but a deeper understanding of this disease will require new statistical and computational approaches for analysis of the genetic and signaling networks that orchestrate individual cancer susceptibility and tumor behavior.

Reduction of p53 gene dosage does not increase initiation or promotion but enhances malignant progression of chemically induced skin tumors

The availability of p53 knockout mice generated by gene targeting has enabled us to investigate the functional role of the p53 tumor suppressor gene in initiation, promotion, and progression of carcinogenesis in vivo, using mouse skin as a model system. The number, size, and growth rate of benign papillomas were not increased in the p53 heterozygous mice in comparison with wild type. The p53 null mice showed a reduced yield of papillomas, but these underwent much more rapid malignant progression, with some poorly differentiated carcinomas developing after only 10 weeks of promotion. Progression rate was also greater in heterozygous than in wild-type mice and was associated with loss of the remaining wild-type allele. Most tumors from all groups had activating mutations in the H-ras gene. Absence of p53, therefore, does not augment the frequency of initiation or the rate of promotion but greatly enhances malignant progression.

Oncogene Activation in Chemical Carcinogenesis

Publisher Summary This chapter discusses the advances made in understanding the mechanisms of action of chemical carcinogens in the light of the recent evidence that oncogenes are directly implicated in the process of tumor development. It discusses various model systems used to address the question of cell transformation in vivo. The process of carcinogenesis in cell culture is compared with the various stages of tumor development in vivo . It is noted that oncogenes provide potential targets for activation by carcinogens. Indeed, the full spectrum of changes that have been noted in the DNA of cells treated by chemical carcinogens— point mutations, translocations, and gene amplification—has been implicated in the activation of protooncogenes to an oncogenic state. It is, therefore, logical to expect that some oncogenes can be activated directly by the interaction between the target genes and chemical carcinogens. Others may be activated indirectly during the progression of tumorigenesis by nontargeted genetic events in somatic cells.

Metastasis is driven by sequential elevation of H-ras and Smad2 levels.

Metastasis is a multistep process that involves local tumour invasion followed by dissemination to, and re-establishment at, distant sites. Here we show that during multistage tumorigenesis, discrete expression thresholds of activated Smad2 and H-ras are sequentially surpassed, driving tumour progression through distinct phases from a differentiated squamous carcinoma to a motile invasive stage, followed by an overt change from epithelial to mesenchymal cell type, finally culminating in metastatic tumour spread. Smad2 activation alone induces migration of tumour cells. Elevated H-ras levels, however, are required for nuclear accumulation of Smad2, both of which are essential for the epithelial–mesenchymal transition (EMT). Having undergone EMT, fibroblastoid carcinoma cells with elevated levels of activated Smad2, gain the capability to spread to a wide variety of tissues by a further increase in Smad2 expression. These findings have far-reaching implications for the prevention of tumour growth, invasion and metastasis.

Fbxw7 / Cdc4 is a p53-dependent, haploinsufficient tumour suppressor gene

The FBXW7/hCDC4 gene encodes a ubiquitin ligase implicated in the control of chromosome stability. Here we identify the mouse Fbxw7 gene as a p53-dependent tumour suppressor gene by using a mammalian genetic screen for p53-dependent genes involved in tumorigenesis. Radiation-induced lymphomas from p53+/- mice, but not those from p53-/- mice, show frequent loss of heterozygosity and a 10% mutation rate of the Fbxw7 gene. Fbxw7+/- mice have greater susceptibility to radiation-induced tumorigenesis, but most tumours retain and express the wild-type allele, indicating that Fbxw7 is a haploinsufficient tumour suppressor gene. Loss of Fbxw7 alters the spectrum of tumours that develop in p53 deficient mice to include a range of tumours in epithelial tissues such as the lung, liver and ovary. Mouse embryo fibroblasts from Fbxw7-deficient mice, or wild-type mouse cells expressing Fbxw7 small interfering RNA, have higher levels of Aurora-A kinase, c-Jun and Notch4, but not of cyclin E. We propose that p53-dependent loss of Fbxw7 leads to genetic instability by mechanisms that might involve the activation of Aurora-A, providing a rationale for the early occurrence of these mutations in human cancers.

FBXW7 Targets mTOR for Degradation and Cooperates with PTEN in Tumor Suppression

The enzyme mTOR (mammalian target of rapamycin) is a major target for therapeutic intervention to treat many human diseases, including cancer, but very little is known about the processes that control levels of mTOR protein. Here, we show that mTOR is targeted for ubiquitination and consequent degradation by binding to the tumor suppressor protein FBXW7. Human breast cancer cell lines and primary tumors showed a reciprocal relation between loss of FBXW7 and deletion or mutation of PTEN (phosphatase and tensin homolog), which also activates mTOR. Tumor cell lines harboring deletions or mutations in FBXW7 are particularly sensitive to rapamycin treatment, which suggests that loss of FBXW7 may be a biomarker for human cancers susceptible to treatment with inhibitors of the mTOR pathway.

Stem-cell hierarchy in skin cancer.

Tumour architecture mimics many of the features of normal tissues, with a cellular hierarchy that regulates the balance between cell renewal and cell death. Although many tumours contain cells with the characteristics of stem cells, the identity of the normal cells that acquire the first genetic hits leading to initiation of carcinogenesis has remained elusive. Identification of the primary cell of origin of cancers and the mechanisms that influence cell-fate decisions will be crucial for the development of novel non-toxic therapies that influence tumour-cell behaviour.

v-ras genes from harvey and BALB murine sarcoma viruses can act as initiators of two-stage mouse skin carcinogenesis

Activated Harvey murine sarcoma virus ras genes were introduced into epidermal cells in vivo by direct application of retroviruses to mouse skin. Subsequent treatment with the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate (TPA) induced benign papillomas, some of which progressed to invasive carcinomas. Initiation with virus was irreversible for at least 4 months, since TPA treatment after this latency period produced papillomas within 4 weeks. Analysis of viral integration sites showed that carcinomas are clonal in origin. Both papillomas and carcinomas express virus-specific ras mRNA and the viral form of ras P21 protein. The results show that activated ras genes can replace chemical carcinogens in initiation of mouse skin carcinogenesis. This system presents a novel approach to in vivo analysis of the biological role of oncogenes in epithelial tumorigenesis.