Telomeres, p21 and the cancer-aging hypothesis.
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Aging↗
Biomedical subjects
Norman E Sharpless
Publications and source records attributed to Norman E Sharpless.
The regulation of INK4/ARF in cancer and aging.
Loss of the INK4a/ARF/INK4b locus on chromosome 9p21 is among the most frequent cytogenetic events in human cancer. The products of the locus--p15(INK4b), p16(INK4a), and ARF--play widespread and independent roles in tumor suppression. Recent data also suggest that expression of p16(INK4a) induces an age-dependent decrease in the proliferative capacity of certain tissue-specific stem cells and unipotent progenitors. Here, we discuss the regulation and role of p16(INK4a), ARF, and p15(INK4b) in cancer and aging.
ADP-Ribosylation Factors↗
EGFR targeted therapy: view from biological standpoint.
Activating mutations in the kinase domain of the epidermal growth factor receptor (EGFR) in nonsmall cell lung cancers (NSCLCs) correlate with responsiveness to EGFR kinase inhibitors. In vitro cell culture studies have demonstrated that EGFR kinase domain mutants but not wild type (wt) EGFR are transforming and essential for cancer cell survival. We and others have recently demonstrated that the induction of EGFR kinase domain mutants specifically in murine lung epithelium in vivo led to development of adenocarcinoma with bronchioloalveolar carcinoma (BAC) features. These tumors depend completely on the sustained expression of EGFR kinase domain mutants for tumor maintenance. The murine tumors with EGFR kinase domain mutations are sensitive to EGFR targeted therapy similarly to NSCLC patients whose tumors harbor EGFR mutations. In contrast, initial results suggest that overexpression of wt EGFR in murine lungs does not seem to be transforming. We therefore divide EGFR targeted therapy in NSCLC patients into two parts: "EGFR mutant targeted therapy" and "wt EGFR targeted therapy". The "EGFR mutant targeted therapy" targets the oncogene essential for tumor initiation and maintenance and is frequently correlated with effective clinical outcome. In contrast, "wt EGFR targeted therapy" likely targets the proto-oncogene product wt EGFR, which is not directly involved in tumor initiation and maintenance, and in these cases, the response has been considerably less dramatic.
Adenocarcinoma↗
Increasing p16INK4a expression decreases forebrain progenitors and neurogenesis during ageing.
Mammalian ageing is associated with reduced regenerative capacity in tissues that contain stem cells. It has been proposed that this is at least partially caused by the senescence of progenitors with age; however, it has not yet been tested whether genes associated with senescence functionally contribute to physiological declines in progenitor activity. Here we show that progenitor proliferation in the subventricular zone and neurogenesis in the olfactory bulb, as well as multipotent progenitor frequency and self-renewal potential, all decline with age in the mouse forebrain. These declines in progenitor frequency and function correlate with increased expression of p16INK4a, which encodes a cyclin-dependent kinase inhibitor linked to senescence. Ageing p16INK4a-deficient mice showed a significantly smaller decline in subventricular zone proliferation, olfactory bulb neurogenesis, and the frequency and self-renewal potential of multipotent progenitors. p16INK4a deficiency did not detectably affect progenitor function in the dentate gyrus or enteric nervous system, indicating regional differences in the response of neural progenitors to increased p16INK4a expression during ageing. Declining subventricular zone progenitor function and olfactory bulb neurogenesis during ageing are thus caused partly by increasing p16INK4a expression.
Aging↗
p16INK4a induces an age-dependent decline in islet regenerative potential.
The p16INK4a tumour suppressor accumulates in many tissues as a function of advancing age. p16INK4a is an effector of senescence and a potent inhibitor of the proliferative kinase Cdk4 (ref. 6), which is essential for pancreatic beta-cell proliferation in adult mammals. Here we show that p16INK4a constrains islet proliferation and regeneration in an age-dependent manner. Expression of the p16INK4a transcript is enriched in purified islets compared with the exocrine pancreas, and islet-specific expression of p16INK4a, but not other cyclin-dependent kinase inhibitors, increases markedly with ageing. To determine the physiological significance of p16INK4a accumulation on islet function, we assessed the impact of p16INK4a deficiency and overexpression with increasing age and in the regenerative response after exposure to a specific beta-cell toxin. Transgenic mice that overexpress p16INK4a to a degree seen with ageing demonstrated decreased islet proliferation. Similarly, islet proliferation was unaffected by p16INK4a deficiency in young mice, but was relatively increased in p16(INK4a)-deficient old mice. Survival after toxin-mediated ablation of beta-cells, which requires islet proliferation, declined with advancing age; however, mice lacking p16INK4a demonstrated enhanced islet proliferation and survival after beta-cell ablation. These genetic data support the view that an age-induced increase of p16INK4a expression limits the regenerative capacity of beta-cells with ageing.
Aging↗
Stem-cell ageing modified by the cyclin-dependent kinase inhibitor p16INK4a.
Stem-cell ageing is thought to contribute to altered tissue maintenance and repair. Older humans experience increased bone marrow failure and poorer haematologic tolerance of cytotoxic injury. Haematopoietic stem cells (HSCs) in older mice have decreased per-cell repopulating activity, self-renewal and homing abilities, myeloid skewing of differentiation, and increased apoptosis with stress. Here we report that the cyclin-dependent kinase inhibitor p16INK4a, the level of which was previously noted to increase in other cell types with age, accumulates and modulates specific age-associated HSC functions. Notably, in the absence of p16INK4a, HSC repopulating defects and apoptosis were mitigated, improving the stress tolerance of cells and the survival of animals in successive transplants, a stem-cell-autonomous tissue regeneration model. Inhibition of p16INK4a may ameliorate the physiological impact of ageing on stem cells and thereby improve injury repair in aged tissue.
Aging↗
The mighty mouse: genetically engineered mouse models in cancer drug development.
Deficiencies in the standard preclinical methods for evaluating potential anticancer drugs,such as xenograft mouse models, have been highlighted as a key obstacle in the translation of the major advances in basic cancer research into meaningful clinical benefits. In this article, we discuss the established uses and limitations of xenograft mouse models for cancer drug development, and then describe the opportunities and challenges in the application of novel genetically engineered mouse models that more faithfully mimic the genetic and biological evolution of human cancers. Greater use of such models in target validation, assessment of tumour response, investigation of pharmacodynamic markers of drug action, modelling resistance and understanding toxicity has the potential to markedly improve the success of cancer drug development.
Animals↗
The impact of human EGFR kinase domain mutations on lung tumorigenesis and in vivo sensitivity to EGFR-targeted therapies.
To understand the role of human epidermal growth factor receptor (hEGFR) kinase domain mutations in lung tumorigenesis and response to EGFR-targeted therapies, we generated bitransgenic mice with inducible expression in type II pneumocytes of two common hEGFR mutants seen in human lung cancer. Both bitransgenic lines developed lung adenocarcinoma after sustained hEGFR mutant expression, confirming their oncogenic potential. Maintenance of these lung tumors was dependent on continued expression of the EGFR mutants. Treatment with small molecule inhibitors (erlotinib or HKI-272) as well as prolonged treatment with a humanized anti-hEGFR antibody (cetuximab) led to dramatic tumor regression. These data suggest that persistent EGFR signaling is required for tumor maintenance in human lung adenocarcinomas expressing EGFR mutants.
Adenocarcinoma↗
RAS unplugged: negative feedback and oncogene-induced senescence.
Many normal cells respond to certain stresses, such as oncogene activation, by undergoing a permanent form of growth arrest known as senescence, an intrinsic tumor suppressor program. The predominant view has been that senescence is caused in some settings through a mutant oncogene's ability to induce activation of high levels of sustained MAP kinase and PI3 kinase signaling. A new study in this issue of Cancer Cell has challenged this model with the surprising finding that aberrant activation of the RAS/RAF pathway can induce a negative feedback loop that globally attenuates MAPK and PI3K signaling and that the reduction of signaling in these pathways is required for senescence.
Animals↗
ROS as a tumour suppressor?
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Cell Division↗
Targeting of C-terminal binding protein (CtBP) by ARF results in p53-independent apoptosis.
ARF encodes a potent tumor suppressor that antagonizes MDM2, a negative regulator of p53. ARF also suppresses the proliferation of cells lacking p53, and loss of ARF in p53-null mice, compared with ARF or p53 singly null mice, results in a broadened tumor spectrum and decreased tumor latency. To investigate the mechanism of p53-independent tumor suppression by ARF, potential interacting proteins were identified by yeast two-hybrid screen. The antiapoptotic transcriptional corepressor C-terminal binding protein 2 (CtBP2) was identified, and ARF interactions with both CtBP1 and CtBP2 were confirmed in vitro and in vivo. Interaction with ARF resulted in proteasome-dependent CtBP degradation. Both ARF-induced CtBP degradation and CtBP small interfering RNA led to p53-independent apoptosis in colon cancer cells. ARF induction of apoptosis was dependent on its ability to interact with CtBP, and reversal of ARF-induced CtBP depletion by CtBP overexpression abrogated ARF-induced apoptosis. CtBP proteins represent putative targets for p53-independent tumor suppression by ARF.
ADP-Ribosylation Factors↗
p16(Ink4a) inhibits histologic progression and angiogenic signaling in min colon tumors.
The Ink4a/Arf tumor suppressor locus is widely inactivated in cancer but little is known about the tumor biology of its two products, p16(Ink4a) (p16) and Arf. Both the p16 and Arf promoters are methylated in a significant fraction of human colon carcinomas, implying a functional role. We have demonstrated previously that Ink4a/Arf-null colon tumors display increased growth and vascularity in C57Bl6 mice carrying the Min (multiple intestinal neoplasia) mutation. We present here an analysis in a mixed genetic background of Min colon tumors (N=215) in mice with or without selective deficiencies in p16 or Arf, respectively. Absence of Arf did not significantly alter tumor formation. In contrast, tumors in mice lacking p16 were moderately larger and redder. Histological analysis demonstrated that these tumors contained significantly more pockets of necrosis (p=0.02), a marker of carcinoma in situ; less apoptosis (p=0.02); and higher red blood cell density (p=0.02, 0.006 within vessels). Biochemical analyses demonstrated increased levels of vascular endothelial cell growth factor (VEGF, p<0.01). Exogenous p16 expression in human colon tumor cells in vitro inhibited VEGF production. These results suggest that p16 constrains colon tumor progression, in part through inhibiting angiogenic signaling.
Animals↗
INK4a/ARF: a multifunctional tumor suppressor locus.
The INK4a/ARF locus encodes two physically linked tumor suppressor proteins, p16(INK4a) and ARF, which regulate the RB and p53 pathways, respectively. The unusual genomic relationship of the open reading frames of these proteins initially fueled speculation that only one of the two was the true tumor suppressor, and loss of the other merely coincidental in cancer. Recent human and mouse genetic data, however, have firmly established that both proteins possess significant in vivo tumor suppressor activity, although there appear to be species- and cell-type specific differences between the two. For example, ARF plays a clear role in preventing Myc-induced lymphomagenesis in mice, whereas the role for p16(INK4a) is human carcinomas is more firmly established. In this review, I discuss the evolutionary history of the locus, the relative importance of these tumor suppressor genes in human cancer, and recent information suggesting novel biochemical and physiologic functions of these proteins in vivo.
Amino Acid Sequence↗
Cancer: crime and punishment.
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ADP-Ribosylation Factors↗
How disruption of cell cycle regulating genes might predispose to sun-induced skin cancer.
The Ink4a/Arf (CDKN2a) locus encodes two proteins that regulate two of the most important tumor suppressor pathways represented by p53 and Rb.(1) Loss of either p16(INK4a) or p19(ARF) was recently reported to reduce the ability of mouse cells to repair UV-induced DNA damage and to induce a UV-mutator phenotype. This observation was independent of cell cycle effects incurred by either p16(INK4a) and/or p19(ARF) loss, as it was demonstrable in unirradiated cells using UV-treated DNA. We suggest that this might explain why germ line mutations of INK4a/ARF predispose mainly to malignant melanoma, a UV-induced skin cancer, and provides a molecular explanation for the link between melanomagenesis and impaired DNA repair. It also further demonstrates that regulation of cell cycle check points and DNA repair in response to genomic insults, such as ultraviolet irradiation are intricately interwoven processes. Differences in the apoptotic response to ultraviolet light between melanocytes and keratinocytes might explain why INK4a/ARF mutations predispose to malignant melanoma, but not to keratinocyte-derived skin cancers.
Animals↗
Impaired processing of DNA photoproducts and ultraviolet hypermutability with loss of p16INK4a or p19ARF.
Reduced DNA repair has been linked to an increased risk of cutaneous malignant melanoma, but insights into the molecular mechanisms of that link are scarce. The INK4a/ARF (CDKN2a) locus, which codes for the p16(INK4a) and p19ARF proteins, is often mutated in sporadic and familial malignant melanoma, but it has not been directly associated with reduced DNA repair. We transfected unirradiated mouse fibroblast cells with UV-treated DNA to measure DNA repair in normal, p16INK4a mutant, p19ARF mutant, or double mutant mouse host cells. Loss of either p16(INK4a) or p19ARF reduced the ability of the cells to process UV-induced DNA damage, independent of cell cycle effects incurred by the loss. These results may further explain why INK4a/ARF mutations predispose to malignant melanoma, a UV-induced tumor.
Animals↗
Medical problems in patients with malignancy: case 2. Wernicke's encephalopathy: an unusual acute neurologic complication of lymphoma and its therapy.
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Adult↗
The differential impact of p16(INK4a) or p19(ARF) deficiency on cell growth and tumorigenesis.
Mounting genetic evidence suggests that each product of the Ink4a/Arf locus, p16(INK4a) and p19(ARF), possesses tumor-suppressor activity (Kamijo et al., 1997; Krimpenfort et al., 2001; Sharpless et al., 2001a). We report the generation and characterization of a p19(ARF)-specific knockout allele (p19(ARF)-/-) and direct comparison with mice and derivative cells deficient for p16(INK4a), both p16(INK4a) and p19(ARF), and p53. Like Ink4a/Arf-/- murine embryo fibroblasts (MEFs), p19(ARF)-/- MEFs were highly susceptible to oncogenic transformation, exhibited enhanced subcloning efficiency at low density, and resisted both RAS- and culture-induced growth arrest. In contrast, the biological profile of p16(INK4a)-/- MEFs in these assays more closely resembled that of wild-type cells. In vivo, however, both p19(ARF)-/- and p16(INK4a)-/- animals were significantly more tumor prone than wild-type animals, but each less so than p53-/- or Ink4a/Arf-/- animals, and with differing tumor spectra. These data confirm the predominant role of p19(ARF) over p16(INK4a) in cell culture-based assays of MEFs, yet also underscore the importance of the analysis of tumor suppressors across many cell types within the organism. The cancer-prone conditions of mice singly deficient for either p16(INK4a) or p19(ARF) agree with data derived from human cancer genetics, and reinforce the view that both gene products play significant and nonredundant roles in suppressing malignant transformation in vivo.
Alleles↗