Recent studies have indicated the existence of tumorigenesis barriers that slow or inhibit the progression of preneoplastic lesions to neoplasia. One such barrier involves DNA replication stress, which leads to activation of the DNA damage checkpoint and thereby to apoptosis or cell cycle arrest, whereas a second barrier is mediated by oncogene-induced senescence. The relationship between these two barriers, if any, has not been elucidated. Here we show that oncogene-induced senescence is associated with signs of DNA replication stress, including prematurely terminated DNA replication forks and DNA double-strand breaks. Inhibiting the DNA double-strand break response kinase ataxia telangiectasia mutated (ATM) suppressed the induction of senescence and in a mouse model led to increased tumour size and invasiveness. Analysis of human precancerous lesions further indicated that DNA damage and senescence markers cosegregate closely. Thus, senescence in human preneoplastic lesions is a manifestation of oncogene-induced DNA replication stress and, together with apoptosis, provides a barrier to malignant progression.

Two major mechanisms have been causally implicated in the establishment of cellular senescence: the activation of the DNA damage response (DDR) pathway and the formation of senescence-associated heterochromatic foci (SAHF). Here we show that in human fibroblasts resistant to premature p16INK4a induction, SAHF are preferentially formed following oncogene activation but are not detected during replicative cellular senescence or on exposure to a variety of senescence-inducing stimuli. Oncogene-induced SAHF formation depends on DNA replication and ATR (ataxia telangiectasia and Rad3-related). Inactivation of ATM (ataxia telangiectasia mutated) or p53 allows the proliferation of oncogene-expressing cells that retain increased heterochromatin induction. In human cancers, levels of heterochromatin markers are higher than in normal tissues, and are independent of the proliferative index or stage of the tumours. Pharmacological and genetic perturbation of heterochromatin in oncogene-expressing cells increase DDR signalling and lead to apoptosis. In vivo, a histone deacetylase inhibitor (HDACi) causes heterochromatin relaxation, increased DDR, apoptosis and tumour regression. These results indicate that heterochromatin induced by oncogenic stress restrains DDR and suggest that the use of chromatin-modifying drugs in cancer therapies may benefit from the study of chromatin and DDR status of tumours.

The accurate execution of DNA replication requires a strict control of the replication licensing factors hCdt1 and hCdc6. The role of these key replication molecules in carcinogenesis has not been clarified. To examine how early during cancer development deregulation of these factors occurs, we investigated their status in epithelial lesions covering progressive stages of hyperplasia, dysplasia, and full malignancy, mostly from the same patients.

Oncogene-induced reactive oxygen species (ROS) have been proposed to be signaling molecules that mediate proliferative cues. However, ROS may also cause DNA damage and proliferative arrest. How these apparently opposite roles can be reconciled, especially in the context of oncogene-induced cellular senescence, which is associated both with aberrant mitogenic signaling and DNA damage response (DDR)-mediated arrest, is unclear. Here, we show that ROS are indeed mitogenic signaling molecules that fuel oncogene-driven aberrant cell proliferation. However, by their very same ability to mediate cell hyperproliferation, ROS eventually cause DDR activation. We also show that oncogenic Ras-induced ROS are produced in a Rac1 and NADPH oxidase (Nox4)-dependent manner. In addition, we show that Ras-induced ROS can be detected and modulated in a living transparent animal: the zebrafish. Finally, in cancer we show that Nox4 is increased in both human tumors and a mouse model of pancreatic cancer and specific Nox4 small-molecule inhibitors act synergistically with existing chemotherapic agents.

Aims: To identify factors contributing to laboratory overutilisation in an academic medical department, and to assess the effect of an educational feedback strategy on inappropriate test-ordering behaviour. Methods: The records of 426 patients admitted during a 6-month period were reviewed. The usefulness of 25 investigations (haematology, basic biochemistry and arterial blood gases) was assessed according to implicit criteria. Trainees' acquaintance with investigation costs was assessed via a multiple-choice questionnaire. The medical staff was informed about their test-ordering behaviour, cost awareness and the factors associated with overuse of diagnostic tests. The test-ordering behaviour of the same doctors was reassessed on 214 patients managed during 6 months after the intervention. Results: Overall, 24 482 laboratory tests were ordered before the intervention (mean 2.96 tests/patient/ day). Among those, 67.9% were not considered to have contributed towards management of patients (mean avoidable 2.01 tests/patient/day). Patient age >65 years, hospitalisation beyond 7 days and increased case difficulty (death or inability to establish a diagnosis) were factors independently associated with overuse of laboratory tests. Senior trainees ordered more laboratory examinations, but the percentage of avoidable tests requested by junior trainees was higher. A moderate and disparate level of trainees' awareness about the cost of common laboratory examinations was disclosed. The avoidable tests/patient/day were significantly decreased after the intervention (mean 1.58, p = 0.002), but containment of unnecessary ordering of tests gradually waned during the semester after the intervention. Conclusion: Repeated audit, continuous education and alertness of doctors, on the basis of assessment of factors contributing to laboratory overutilisation, result in restraining the redundant ordering of tests in the hospital setting.T he overuse of laboratory investigations is widely prevalent in hospital practice, including academic departments.1-3 Reasons for excessive ordering of tests by doctors include defensive behaviour and fear or uncertainty, lack of experience, the use of protocols and guidelines, ''routine'' clinical practice, inadequate educational feedback and clinician's unawareness about the cost of examinations. [4][5][6] Inappropriate testing causes unnecessary patient discomfort, entails the risk of generating false-positive results, leads to overloading of the diagnostic services, wastes valuable healthcare resources and is associated with other inefficiencies in healthcare delivery, undermining the quality of health services.1 5 Interventions on inappropriate testing aim to reduce costs, along with improving the quality of care provided. Results, however, are not always consistent, usually owing to inherent limitations of the strategies proposed. In this study, we assessed the appropriateness of routine ordering of laboratory tests of the trainees in an academic internal medicine department, as well as their awa...