Telomeres, crisis and cancer

Eukaryotic chromosomes terminate in specialized nucleic acid-protein complexes known as telomeres. Disruption of telomere structure by erosion of telomeric DNA or loss of telomere binding protein function activates a signal transduction program that closely resembles the cellular responses generated upon DNA damage. Telomere dysfunction in turn induces a permanent proliferation arrest known as senescence. Senescence is postulated to perform a tumor suppressor function by limiting cellular proliferative capacity, thus imposing a barrier to cellular immortalization. Genetic or epigenetic silencing of components of the DNA damage pathway, allows cells to proliferate beyond senescence limits. However, these cells eventually reach a stage of extreme telomere dysfunction known as crisis that is characterized by cell death and the concomitant appearance of cytogenetic abnormalities. Telomeric crisis produces significant chromosomal instability, a hallmark of human cancer, and may thus be relevant to carcinogenesis by increasing the occurrence of genetic alterations that would favor neoplastic transformation. The following review examines the relationship of telomere function during crisis in accelerating chromosomal instability and cancer.     

Evolving views of telomerase and cancer

The maintenance of telomeres, nucleoprotein structures that constitute the ends of eukaryotic chromosomes, regulates many crucial cellular functions and might, in multicellular organisms, participate in the control of complex phenotypes such as aging and cancer. Stabilization of telomere length is strongly associated with cellular immortalization, and constitutive telomerase activation occurs in most human cancers. Such observations form the basis for the prevailing model that postulates that alterations in telomere biology both suppress and facilitate malignant transformation by regulating genomic stability and cell life span. However, recent findings suggest that telomere maintenance might not be an obligate requirement for initial tumor formation in some settings and that telomerase activation contributes to tumorigenesis independently of its role in maintaining telomere length. These recent developments indicate that our understanding of telomere biology remains incomplete and implicate additional complexity in the relationships among telomeres, telomerase and cancer.     

Short dysfunctional telomeres impair tumorigenesis in the INK4a(delta2/3) cancer-prone mouse.

Maintenance of telomere length is predicted to be essential for bypass of senescence and crisis checkpoints in cancer cells. The impact of telomere dysfunction on tumorigenesis was assessed in successive generations of mice doubly null for the telomerase RNA (mTR) and the INK4a tumor suppressor genes. Significant reductions in tumor formation in vivo and oncogenic potential in vitro were observed in late generations of telomerase deficiency, coincident with severe telomere shortening and associated dysfunction. Reintroduction of mTR into cells significantly restored the oncogenic potential, indicating telomerase activation is a cooperating event in the malignant transformation of cells containing critically short telomeres. The results described here demonstrate that loss of telomere function in a cancer-prone mouse model possessing intact DNA damage responses impairs, but does not prevent, tumor formation.     

Abolition of cyclin-dependent kinase inhibitor p16Ink4a and p21Cip1/Waf1 functions permits Ras-induced anchorage-independent growth in telomerase-immortalized human fibroblasts

Human cells are more resistant to both immortalization and malignant transformation than rodent cells. Recent studies have established the basic genetic requirements for the transformation of human cells, but much of this work relied on the expression of transforming proteins derived from DNA tumor viruses. We constructed an isogenic panel of human fibroblast cell lines using a combination of gene targeting and ectopic expression of dominantly acting mutants of cellular genes. Abolition of p21(Cip1/Waf1) and p16(Ink4a) functions prevented oncogenically activated Ras from inducing growth arrest and was sufficient for limited anchorage-independent growth but not tumorigenesis. Deletion of the tumor suppressor p53 combined with abolition of p16(Ink4a) function failed to mimic the introduction of simian virus 40 large T antigen, indicating that large T antigen may target additional cellular functions. Ha-Ras and Myc cooperated only to a limited extent, but in the absence of Ras, Myc cooperated strongly with the simian virus 40 small t antigen to elicit aggressive anchorage-independent growth. The experiments reported here further define specific components of human transformation pathways.     

Differentiation genes: are they primary targets for human carcinogenesis?

In spite of extensive research in molecular carcinogenesis, genes that can be considered primary targets in human carcinogenesis remain to be identified. Mutated oncogenes or cellular growth regulatory genes, when incorporated into normal human epithelial cells, failed to immortalize or transform these cells. Therefore, they may be secondary events in human carcinogenesis. Based on some experimental studies we have proposed that downregulation of a differentiation gene may be the primary event in human carcinogenesis. Such a gene could be referred to as a tumor-initiating gene. Downregulation of a differentiation gene can be accomplished by a mutation in the differentiation gene, by activation of differentiation suppressor genes, and by inactivation of tumor suppressor genes. Downregulation of a differentiation gene can lead to immortalization of normal cells. Mutations in cellular proto-oncogenes, growth regulatory genes, and tumor suppressor genes in immortalized cells can lead to transformation. Such genes could be called tumor-promoting genes. This hypothesis can be documented by experiments published on differentiation of neuroblastoma (NB) cells in culture. The fact that terminal differentiation can be induced in NB cells by adenosine 3',5'-cyclic monophosphate (cAMP) suggests that the differentiation gene in these cells is not mutated, and thus can be activated by an appropriate agent. The fact that cAMP-resistant cells exist in NB cell populations suggests that a differentiation gene is mutated in these cancer cells, or that differentiation regulatory genes have become unresponsive to cAMP. In addition to cAMP, several other differentiating agents have been identified. Our proposed hypothesis of carcinogenesis can also be applied to other human tumors such as melanoma, pheochromocytoma, medulloblastoma, glioma, sarcoma, and colon cancer.     

Pregnancy-associated homeostasis and dysregulation: lessons from genetically modified animal models

Unlimitedly proliferating cells need to acquire the telomere DNA maintenance mechanism, to counteract possible shortening through multiple rounds of replication and segregation of linear chromosomes. Most human cancer cells express telomerase whereas the other cells utilize the alternative lengthening of telomeres (ALT) pathway to elongate telomere DNA. It is suggested that ALT depends on the recombination between telomere repetitive DNAs. However, the molecular details remain unknown. Recent studies have provided evidence of special structures of telomere DNA and genes essential for the phenotypes of ALT cells. The molecular models of the ALT pathway should be validated to elucidate recombination-mediated telomere maintenance and promote the applications to anti-cancer therapy.     

How do mutated oncogenes and tumor suppressor genes cause cancer?

In recent decades we have been given insight into the process that transforms a normal cell into a malignant cancer cell. It has been recognised that malignant transformation occurs through successive mutations in specific cellular genes, leading to the activation of oncogenes and inactivation of tumor suppressor genes. The further study of these genes has generated much of its excitement from the convergence of experiments addressing the genetic basis of cancer, together with cellular pathways that normally control important cellular regulatory programmes. In the present review the context in which oncogenes such as proliferation, cell death/apoptosis, differentiation and senescence will be described, as well as how these cellular programmes become deregulated in cancer due to mutations.     

Telomere and telomerase in oncology

Telomere and cell replicative senescenceTelomeres, which are located at the end of chro-mosome, are crucial to protect chromosome againstdegeneration, rearrangment and end to end fusion[1].Human telomeres are tandemly repeated units of thehexanucleotide TTAGGG. The estimated length oftelomeric DNA varies from 2 to 20 kilo base pairs,depending on factors such as tissue type and hu-man age. The buck of telomeric DNA is double-stranded, but the end of telomeric DNA consists of3' overhang of…     

The role of p53-mediated apoptosis as a crucial anti-tumor response to genomic instability: lessons from mouse models

Genomic instability is a major force driving human cancer development. A cellular safeguard against such genetic destabilization, which can ensue from defects in telomere maintenance, DNA repair, and checkpoint function, is activation of the p53 tumor suppressor protein, which commonly responds to these DNA damage signals by inducing apoptosis. If, however, p53 becomes inactivated, as is typical of many tumors and pre-cancerous lesions, then cells with compromised genome integrity pathways survive inappropriately, and the accrual of oncogenic lesions can fuel the carcinogenic process. Studies of mouse models have been instrumental in providing support for this idea. Mouse knockouts in genes important for telomere function, DNA damage checkpoint activation and DNA repair - both non-homologous end joining and homologous recombination - are prone to the development of genomic instability. As a consequence of these DNA damage signals, p53 becomes activated in cells of these mutant mice, leading to the induction of apoptosis, sometimes at the expense of organismal viability. This apoptotic response can be rescued through crosses to p53-deficient mice, but has dire consequences: mice predisposed to genomic instability and lacking p53 are susceptible to tumorigenesis. Thus p53-mediated apoptosis provides a crucial tumor suppressive mechanism to eliminate cells succumbing to genomic instability.     

靶向端粒酶的肿瘤治疗方案相关进展

近年来,端粒作为真核细胞染色体末端的保护染色体免受核酸酶降解的特殊的DNA重复结构,成为现代生物学的研究热点。端粒与基因表达调控、细胞生长、肿瘤发生、衰老有着密切的关系。端粒维持过程中有2类重要的蛋白,即端粒相关蛋白和端粒酶。端粒酶,特别是其催化亚基hTERT,在端粒延长过程中起着不可替代的作用,与细胞永生化和癌变密切相关。近年来,靶向端粒酶的肿瘤治疗在逆转录酶抑制剂尤其是反义核酸和免疫治疗方面都取得了突破性的进展。肿瘤干细胞在肿瘤的发生发展过程中起重要作用,将很有希望成为未来靶向端粒酶的肿瘤治疗的一个重要靶标。     

Oncogenic transformation of human cells: shortcomings of rodent model systems

Long-standing difficulties in the in vitro transformation of human cells have been overcome. Using telomerase, several successful oncogene-mediated transformations of human cells have been reported and the following cellular requirements for human cell transformation have been proposed: the maintenance of telomere sequences, the inactivation of Rb and p53 pathways, the perturbation of protein phosphatase 2A (PP2A) and the expression of activated Ras. Even when all of these requirements are fulfilled, however, the transformed phenotypes of human cells seem to be much less malignant than those of rodent cells meeting the same requirements. This suggests the existence of undefined cell-autonomous mechanisms that render human cells resistant to malignant transformation.     

Different telomere damage signaling pathways in human and mouse cells

Programmed telomere shortening in human somatic cells is thought to act as a tumor suppressor pathway, limiting the replicative potential of developing tumor cells. Critically short human telomeres induce senescence either by activating p53 or by inducing the p16/RB pathway, and suppression of both pathways is required to suppress senescence of aged human cells. Here we report that removal of TRF2 from human telomeres and the ensuing de-protection of chromosome ends induced immediate premature senescence. Although the telomeric tracts remained intact, the TRF2(DeltaBDeltaM)-induced premature senescence was indistinguishable from replicative senescence and could be mediated by either the p53 or the p16/RB pathway. Telomere de-protection also induced a growth arrest and senescent morphology in mouse cells. However, in this setting the loss of p53 function was sufficient to completely abrogate the arrest, indicating that the p16/RB response to telomere dysfunction is not active in mouse cells. These findings reveal a fundamental difference in telomere damage signaling in human and mouse cells that bears on the use of mouse models for the telomere tumor suppressor pathway.     

Degradation of p53, not telomerase activation, by E6 is required for bypass of crisis and immortalization by human papillomavirus type 16 E6/E7

Bypass of two arrest points is essential in the process of cellular immortalization, one of the components of the transformation process. Expression of human papillomavirus type 16 E6 and E7 together can escape both senescence and crisis, processes which normally limit the proliferative capacity of primary human keratinocytes. Crisis is thought to be mediated by telomere shortening. Because E6 stimulates telomerase activity and exogenous expression of the TERT gene with E7 can immortalize keratinocytes, this function is thought to be important for E6 to cooperate with E7 to bypass crisis. However, it has also been reported that E6 dissociates increased telomerase activity from maintenance of telomere length and that a dominant-negative p53 molecule can substitute for E6 in cooperative immortalization of keratinocytes with E7. Thus, to determine which functions of E6 are required to allow bypass of crisis and immortalization of keratinocytes with E7, immortalization assays were performed using specific mutants of E6, in tandem with E7. In these experiments, every clone expressing an E6 mutant capable of degrading p53 was able to bypass crisis and immortalize, regardless of telomerase induction. All clones containing E6 mutants incapable of degrading p53 died at crisis. These results suggest that the ability of E6 to induce degradation of p53 compensates for continued telomere shortening in E6/E7 cells and demonstrate that degradation of p53 is required for immortalization by E6/E7, while increased telomerase activity is dispensable.     

Murine tumor suppressor models

Tumor suppressor genes have been shown to be necessary for proper maintenance of cell growth control. Inactivation of these genes in the germline of humans is linked to inherited cancer predisposition. Moreover, sporadically arising human tumors often have somatic mutations in tumor suppressor genes. During the past few years, advances in molecular and cellular biology have led to the creation of animal models that have germline mutations of various tumor suppressor genes. Such mice potentially represent important animal models for familial cancer predisposition syndromes, and the study of the tumorigenesis process has been greatly assisted by their development. Such models have also demonstrated the importance of tumor suppressor function in embryonic development. In this review, we describe mice with inactivated germline tumor suppressor genes that are genetically analogous to 10 different inherited cancer syndromes in humans. We describe the variable usefulness of the mutant mice as models for human disease.     

Functional integration of microRNAs into oncogenic and tumor suppressor pathways

A large body of evidence has documented abnormal microRNA (miRNA) expression patterns in diverse human malignancies. Given that miRNA expression is tightly regulated during development and cellular differentiation, aberrant miRNA expression in cancer cells is likely to be in part a consequence of the loss of normal cellular identity that accompanies malignant transformation. Nevertheless, it is now clear that miRNAs function as critical effectors of several canonical oncogenic and tumor suppressor pathways, including those controlled by Myc and p53. Gain- and loss-of-function of these factors in cancer cells contributes to miRNA dysregulation, directly influencing neoplastic phenotypes including cellular proliferation and apoptosis.     

Senescence: a telomeric limit to immortality or a cellular response to physiologic stresses?

Cells entering a state of senescence undergo a irreversible cell cycle arrest, associated by a set of functional and morphological changes. Senescence occurs following telomeres shortening (replicative senescence) or exposure to other acute or chronic physiologic stress signals (a phenomenon termed stasis: stress or aberrant signaling-induced senescence). In this review, I discuss the pathways of cellular senescence, the mechanisms involved and the role that these pathways have in regulating the initiation and progression of cancer. Telomere-initiated senescence or loss of telomere function trigger focal recruitement of protein sensors of the DNA double-strand breaks leading to the activation of the DNA damage checkpoint responses and the tumour suppressor gene product, p53, which in turn induces the cell-cycle inhibitor, p21(WAF1). Loss of p53 and pRb function allows continued cell division despite increasing telomere dysfunction and eventually entry into telomere crisis. Immortalisation is an essential prerequisite for the formation of a tumour cell. Therefore, a developing tumour cell must circumvent at least two proliferative barriers--cellular senescence and crisis--to achieve neoplastic transformation. These barriers are regulated by telomere shortening and by the p16(INK4a)/Rb and p53 tumour suppressor pathways. Elucidation of the genes and emerging knowledge about the regulatory mechanisms that lead to senescence and determine the pattern of gene expression in senescent cells may lead to more effective treatments for cancer.     

鉴定肿瘤驱动基因的实验模型

人体肿瘤的形成是一个复杂的过程.在这个过程中会发生许多基因突变,这些突变中只有较少一部分具有驱动肿瘤发生的作用,大部分突变作为伴随性变化对肿瘤的发生并无明确的贡献.要确定哪些变异具有驱动肿瘤发生的作用及其作用机制,需要通过实验验证.伴随着新的研究技术的出现,鉴定肿瘤驱动基因的手段也不断演变.从早期主要是从动物诱癌实验、...