Anette Duensing

Publications of Anette Duensing

• Daughter centriole elongation is controlled by proteolysis.

  Authors: Nina Korzeniewski, Rolando Cuevas, Anette Duensing, Stefan Duensing

  Molecular biology of the cell. 11/2010; 21(22):3942-51.

  The centrosome is the major microtubule-organizing center of most mammalian cells and consists of a pair of centrioles embedded in pericentriolar material. Before mitosis, the two centriolesThe centrosome is the major microtubule-organizing center of most mammalian cells and consists of a pair of centrioles embedded in pericentriolar material. Before mitosis, the two centrioles duplicate and two new daughter centrioles form adjacent to each preexisting maternal centriole. After initiation of daughter centriole synthesis, the procentrioles elongate in a process that is poorly understood. Here, we show that inhibition of cellular proteolysis by Z-L3VS or MG132 induces abnormal elongation of daughter centrioles to approximately 4 times their normal length. This activity of Z-L3VS or MG132 was found to correlate with inhibition of intracellular protease-mediated substrate cleavage. Using a small interfering RNA screen, we identified a total of nine gene products that either attenuated (seven) or promoted (two) abnormal Z-L3VS-induced daughter centriole elongation. Our hits included known regulators of centriole length, including CPAP and CP110, but, interestingly, several proteins involved in microtubule stability and anchoring as well as centrosome cohesion. This suggests that nonproteasomal functions, specifically inhibition of cellular proteases, may play an important and underappreciated role in the regulation of centriole elongation. They also highlight the complexity of daughter centriole length control and provide a framework for future studies to dissect the molecular details of this process.

• Genomic instability and cancer: lessons learned from human papillomaviruses.

  Authors: Nina Korzeniewski, Nicole Spardy, Anette Duensing, Stefan Duensing

  Cancer letters. 11/2010; 305(2):113-22.

  High-risk HPV E6 and E7 oncoproteins cooperate to subvert critical host cell cycle checkpoint control mechanisms in order to promote viral genome replication. This results not only in aberrantHigh-risk HPV E6 and E7 oncoproteins cooperate to subvert critical host cell cycle checkpoint control mechanisms in order to promote viral genome replication. This results not only in aberrant proliferation but also in host cellular changes that can promote genomic instability. The HPV-16 E7 oncoprotein was found to induce centrosome abnormalities thereby disrupting mitotic fidelity and increasing the risk for chromosome missegregation and aneuploidy. In addition, expression of the high-risk HPV E7 oncoprotein stimulates DNA replication stress as a potential source of DNA breakage and structural chromosomal instability. Proliferation of genomically unstable cells is sustained by several mechanisms including the accelerated degradation of claspin by HPV-16 E7 and the degradation of p53 by the high-risk HPV E6 oncoprotein. These results highlight the oncogenic potential of aberrant proliferation and opens new avenues for prevention of malignant progression, not only in HPV-associated cervical cancer but also in non-virally associated malignancies with disrupted cell cycle checkpoint control mechanisms.

• Tripeptidyl Peptidase II Is Required for c-MYC-Induced Centriole Overduplication and a Novel Therapeutic Target in c-MYC-Associated Neoplasms.

  Authors: Stefan Duensing, Sebastian Darr, Rolando Cuevas, Nadja Melquiot, Anthony G Brickner, Anette Duensing, Karl Münger

  Genes & cancer. 09/2010; 1(9):883-92.

  Centrosome aberrations are frequently detected in c-MYC-associated human malignancies. Here, we show that c-MYC-induced centrosome and centriole overduplication critically depend on the proteaseCentrosome aberrations are frequently detected in c-MYC-associated human malignancies. Here, we show that c-MYC-induced centrosome and centriole overduplication critically depend on the protease tripeptidyl peptidase II (TPPII). We found that TPPII localizes to centrosomes and that overexpression of TPPII, similar to c-MYC, can disrupt centriole duplication control and cause centriole multiplication, a process during which maternal centrioles nucleate the formation of more than a single daughter centriole. We report that inactivation of TPPII using chemical inhibitors or siRNA-mediated protein knockdown effectively reduced c-MYC-induced centriole overduplication. Remarkably, the potent and selective TPPII inhibitor butabindide not only potently suppressed centriole aberrations but also caused significant cell death and growth suppression in aggressive human Burkitt lymphoma cells with c-MYC overexpression. Taken together, these results highlight the role of TPPII in c-MYC-induced centriole overduplication and encourage further studies to explore TPPII as a novel antineoplastic drug target.

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• Bortezomib: killing two birds with one stone in gastrointestinal stromal tumors.

  Authors: Stefan Duensing, Anette Duensing

  Oncotarget. 05/2010; 1(1):6-8.

• Targeted therapies of gastrointestinal stromal tumors (GIST)--the next frontiers.

  Authors: Stefan Duensing, Anette Duensing

  Biochemical pharmacology. 04/2010; 80(5):575-83.

  Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal (GI) tract and are caused by activating KIT or PDGFRA mutations. GISTs can be successfullyGastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal (GI) tract and are caused by activating KIT or PDGFRA mutations. GISTs can be successfully treated with the small molecule kinase inhibitor imatinib mesylate (Gleevec, Novartis) with response rates of up to 85%. However, complete responses are rare, and most patients will develop imatinib resistance over time. Recent results have shown that although imatinib effectively stimulates apoptotic cell death in sensitive GIST cells, a considerable proportion of cells does not undergo apoptosis, but instead enters a state of quiescence. Quiescence is characterized by a reversible withdrawal from the cell division cycle, during which the cells remain alive and metabolically active. It is conceivable that quiescence not only plays a pivotal role in the emergence of residual disease but also in creating a pool of tumor cells that survive continuous small molecule therapy and may hence represent the "seeds" for the outgrowth of resistant clones. This review will summarize the current knowledge about GIST biology and treatment response to imatinib including the induction of cellular quiescence in GIST. In addition, we will highlight future strategies to design more effective treatment options to overcome these problems with an aim towards cure of this hitherto untreatable tumor entity.

• Gamma-tubulin and 53BP1 as candidate biomarkers of human papillomavirus-associated anal dysplasia

  Authors: Ken Ho, Richard Day, Amy Perkins, Jia Xu, Shih-Fan Kuan, Anette Duensing, Ross Cranston, Stefan Duensing

  Infectious Agents and Cancer. 01/2010;

• Centrosomes, polyploidy and cancer.

  Authors: Anette Duensing, Stefan Duensing

  Advances in experimental medicine and biology. 01/2010; 676:93-103.

  Cancer cells are frequently characterized by ploidy changes including tetra-, poly- or aneuploidy. At the same time, malignant cells often contain supernumerary centrosomes. Aneuploidy and centrosomeCancer cells are frequently characterized by ploidy changes including tetra-, poly- or aneuploidy. At the same time, malignant cells often contain supernumerary centrosomes. Aneuploidy and centrosome alterations are both hallmarks of tumor aggressiveness and increase with malignant progression. It has been proposed that aneuploidy results from a sequence of events in which failed mitoses produce tetra-/polyploid cells that enter a subsequent cell division with an increased number of centrosomes and hence with an increased risk for multipolar spindle formation and chromosome missegregation. Although this model attempts to integrate several common findings in cancer cells, it has been difficult to prove. Findings that centrosome aberrations can arise in diploid cells and the uncertain proliferative potential of polyploid cells suggest that alternative routes to chromosomal instability may exist. We discuss here recent results on centrosome biogenesis and the possible link between ploidy changes, centrosome aberrations and cancer.

• A novel role of the aryl hydrocarbon receptor (AhR) in centrosome amplification - implications for chemoprevention.

  Authors: Nina Korzeniewski, Sarah Wheeler, Payel Chatterjee, Anette Duensing, Stefan Duensing

  Molecular cancer. 01/2010; 9:153.

  Centrosome aberrations can cause genomic instability and correlate with malignant progression in common human malignancies such as breast and prostate cancer. Deregulation of cyclin/cyclin-dependentCentrosome aberrations can cause genomic instability and correlate with malignant progression in common human malignancies such as breast and prostate cancer. Deregulation of cyclin/cyclin-dependent kinase 2 (CDK2) activity has previously been shown to be critically involved in centrosome overduplication. We therefore test here whether small molecule CDK inhibitors derived from the bis-indole indirubin can be used to suppress centrosome aberrations as a novel approach to chemoprevention of malignant progression. As expected, we found that the CDK inhibitor indirubin-3'-oxime (IO) suppresses centrosome amplification in breast cancer cells. However, we made the unexpected discovery that indirubin-derived compounds that have been chemically modified to be inactive as kinase inhibitors such as 1-methyl-indirubin-3'-oxime (MeIO) still significantly reduced centrosome amplification. All indirubins used in the present study are potent agonists of the aryl hydrocarbon receptor (AhR), which is known for its important role in the cellular metabolism of xenobiotics. To corroborate our results, we first show that the coincidence of nuclear AhR overexpression, reflecting a constitutive activation, and numerical centrosome aberrations correlates significantly with malignancy in mammary tissue specimens. Remarkably, a considerable proportion (72.7%) of benign mammary tissue samples scored also positive for nuclear AhR overexpression. We furthermore provide evidence that continued expression of endogenous AhR is critical to promote centriole overduplication induced by cyclin E and that AhR and cyclin E may function in the same pathway. Overexpression of the AhR in the absence of exogenous ligands was found to rapidly disrupt centriole duplication control. Nonetheless, the AhR agonists IO and MeIO were still found to significantly reduce centriole overduplication stimulated by ectopic AhR expression. Our results indicate that continued expression of endogenous AhR promotes centrosome amplification in breast cancer cells in a pathway that involves cyclin E. AhR agonists such as indirubins inhibit centrosome amplification even when stimulated by ectopic expression of the AhR suggesting that these compounds are potentially useful for the chemoprevention of centrosome-mediated cell division errors and malignant progression in neoplasms in which the AhR is overexpressed. Future studies are warranted to determine whether individuals in which nuclear AhR overexpression is detected in benign mammary tissue are at a higher risk for developing pre-cancerous or cancerous breast lesions.

• Proapoptotic Activity of Bortezomib in Gastrointestinal Stromal Tumor Cells.

  Authors: Sebastian Bauer, Joshua A Parry, Thomas Mühlenberg, Matthew F Brown, Danushka Seneviratne, Payel Chatterjee, Anna Chin, Brian P Rubin, Shih-Fan Kuan, Jonathan A Fletcher, Stefan Duensing, Anette Duensing

  Cancer research. 12/2009;

  Gastrointestinal stromal tumors (GIST) are caused by activating mutations in the KIT or PDGFRA receptor tyrosine kinase genes. Although >85% of GIST patients treated with the small-molecule inhibitorGastrointestinal stromal tumors (GIST) are caused by activating mutations in the KIT or PDGFRA receptor tyrosine kinase genes. Although >85% of GIST patients treated with the small-molecule inhibitor imatinib mesylate (Gleevec) achieve disease stabilization, complete remissions are rare and a substantial proportion of patients develop resistance to imatinib over time. Upregulation of soluble, non-chromatin-bound histone H2AX has an important role in imatinib-induced apoptosis of GIST cells. Additionally, H2AX levels in untreated GIST are maintained at low levels by a pathway that involves KIT, phosphoinositide 3-kinase, and the ubiquitin-proteasome system. In this study, we asked whether bortezomib-mediated inhibition of the ubiquitin-proteasome machinery could lead to upregulation of histone H2AX and GIST cell death. We show that bortezomib rapidly triggers apoptosis in GIST cells through a combination of mechanisms involving H2AX upregulation and loss of KIT protein expression. Downregulation of KIT transcription was an underlying mechanism for bortezomib-mediated inhibition of KIT expression. In contrast, the nuclear factor-kappaB signaling pathway did not seem to play a major role in bortezomib-induced GIST cell death. Significantly, we found that bortezomib would induce apoptosis in two imatinib-resistant GIST cell lines as well as a short-term culture established from a primary imatinib-resistant GIST. Collectively, our results provide a rationale to test the efficacy of bortezomib in GIST patients with imatinib-sensitive or -resistant tumors. Cancer Res; 70(1); 150-9.

• Human Papillomavirus 16 E7 Oncoprotein Attenuates DNA Damage Checkpoint Control by Increasing the Proteolytic Turnover of Claspin.

  Authors: Nicole Spardy, Kathryn Covella, Elliot Cha, Elizabeth E Hoskins, Susanne I Wells, Anette Duensing, Stefan Duensing

  Cancer research. 09/2009;

  The human papillomavirus (HPV) 16 E7 oncoprotein has been reported previously to stimulate DNA damage and to activate host cell DNA damage checkpoints. How HPV-16 E7 maintains proliferation despiteThe human papillomavirus (HPV) 16 E7 oncoprotein has been reported previously to stimulate DNA damage and to activate host cell DNA damage checkpoints. How HPV-16 E7 maintains proliferation despite activated DNA damage checkpoints is incompletely understood. Here, we provide evidence that cells expressing the HPV-16 E7 oncoprotein can enter mitosis in the presence of DNA damage. We show that this activity of HPV-16 E7 involves attenuation of DNA damage checkpoint control by accelerating the proteolytic turnover of claspin. Claspin mediates the activation of CHK1 by ATR in response to replication stress, and its degradation plays a critical role in DNA damage checkpoint recovery. Expression of a nondegradable mutant of claspin was shown to inhibit mitotic entry in HPV-16 E7-expressing cells. Multiple components of the SCF(beta-TrCP)-based claspin degradation machinery were found deregulated in the presence of HPV-16 E7, including cullin 1, beta-TrCP, Aurora A, and Polo-like kinase-1 (PLK1). In contrast, no difference in the expression level of the claspin deubiquitinating enzyme USP7 was detected. Levels of Aurora A and PLK1 as well as phosphorylated PLK1 at threonine 210, a prerequisite for DNA damage checkpoint recovery, remained detectable following replication stress in HPV-16 E7-expressing cells but not in control cells. In summary, our results suggest that the HPV-16 E7 oncoprotein alleviates DNA damage checkpoint responses and promotes mitotic entry by accelerating claspin degradation through a mechanism that involves deregulation of components of the SCF(beta-TrCP)-based claspin degradation machinery. [Cancer Res 2009;69(17):7022-9].

• Cullin 1 functions as a centrosomal suppressor of centriole multiplication by regulating polo-like kinase 4 protein levels.

  Authors: Nina Korzeniewski, Leon Zheng, Rolando Cuevas, Joshua Parry, Payel Chatterjee, Brittany Anderton, Anette Duensing, Karl Münger, Stefan Duensing

  Cancer research. 09/2009; 69(16):6668-75.

  Abnormal centrosome and centriole numbers are frequently detected in tumor cells where they can contribute to mitotic aberrations that cause chromosome missegregation and aneuploidy. The molecularAbnormal centrosome and centriole numbers are frequently detected in tumor cells where they can contribute to mitotic aberrations that cause chromosome missegregation and aneuploidy. The molecular mechanisms of centriole overduplication in malignant cells, however, are poorly characterized. Here, we show that the core SKP1-cullin-F-box component cullin 1 (CUL1) localizes to maternal centrioles and that CUL1 is critical for suppressing centriole overduplication through multiplication, a recently discovered mechanism whereby multiple daughter centrioles form concurrently at single maternal centrioles. We found that this activity of CUL1 involves the degradation of Polo-like kinase 4 (PLK4) at maternal centrioles. PLK4 is required for centriole duplication and strongly stimulates centriole multiplication when aberrantly expressed. We found that CUL1 is critical for the degradation of active PLK4 following deregulation of cyclin E/cyclin-dependent kinase 2 activity, as is frequently observed in human cancer cells, as well as for baseline PLK4 protein stability. Collectively, our results suggest that CUL1 may function as a tumor suppressor by regulating PLK4 protein levels and thereby restraining excessive daughter centriole formation at maternal centrioles.

• Centrosome overduplication, chromosomal instability, and human papillomavirus oncoproteins.

  Authors: Anette Duensing, Nicole Spardy, Payel Chatterjee, Leon Zheng, Joshua Parry, Rolando Cuevas, Nina Korzeniewski, Stefan Duensing

  Environmental and molecular mutagenesis. 04/2009;

  Centrosome aberrations are a frequent finding in human tumors. However, very little is known about the molecular mechanisms leading to disruption of centrosome duplication control and the functionalCentrosome aberrations are a frequent finding in human tumors. However, very little is known about the molecular mechanisms leading to disruption of centrosome duplication control and the functional consequences of aberrant centrosome numbers. The high-risk human papillomavirus Type 16 (HPV-16) E6 and E7 oncoproteins are overexpressed in HPV-associated malignancies of the anogenital tract and have been instrumental in delineating different pathways of centrosome amplification. Whereas the E6 oncoprotein was found to provoke centrosome accumulation, the HPV-16 E7 oncoprotein triggers a genuine disruption of the centrosome duplication cycle. Importantly, the E7 oncoprotein can rapidly cause centrosome overduplication through a pathway that involves the concurrent formation of multiple daughters at single maternal centrioles (centriole flowers). Several lines of evidence suggest that cyclin E/CDK2 complexes and Polo-like kinase 4 (PLK4) are crucial players in this process. These findings underscore that the HPV-16 E7 oncoprotein is a unique tool to dissect normal and abnormal centriole biogenesis and the underlying molecular circuitry. Environ. Mol. Mutagen. 2009. (c) 2009 Wiley-Liss, Inc.

• HPV-16 E7 Reveals a Link between DNA Replication Stress, Fanconi Anemia D2 Protein, and Alternative Lengthening of Telomere-Associated Promyelocytic Leukemia Bodies.

  Authors: Nicole Spardy, Anette Duensing, Elizabeth E Hoskins, Susanne I Wells, Stefan Duensing

  Cancer research. 01/2009; 68(23):9954-63.

  Expression of the high-risk human papillomavirus (HPV-16) E7 oncoprotein extends the life span of primary human keratinocytes and partially restores telomere length in the absence of telomerase. TheExpression of the high-risk human papillomavirus (HPV-16) E7 oncoprotein extends the life span of primary human keratinocytes and partially restores telomere length in the absence of telomerase. The molecular basis of this activity is incompletely understood. Here, we show that HPV-16 E7 induces an increased formation of alternative lengthening of telomeres (ALT)-associated promyelocytic leukemia bodies (APBs) in early passage primary human keratinocytes as well as HPV-negative tumor cells. This activity was found to require sequences of HPV-16 E7 involved in degradation of the retinoblastoma tumor suppressor protein as well as regions in the COOH terminus. HPV-16 E7-induced APBs contained ssDNA and several proteins that are involved in the response to DNA replication stress, most notably the Fanconi anemia D2 protein (FANCD2) as well as BRCA2 and MUS81. In line with these results, we found that FANCD2-containing APBs form in an ATR-dependent manner in HPV-16 E7-expressing cells. To directly show a role of FANCD2 in ALT, we provide evidence that knockdown of FANCD2 rapidly causes telomere dysfunction in cells that rely on ALT to maintain telomeres. Taken together, our results suggest a novel link between replication stress and recombination-based telomere maintenance that may play a role in HPV-16 E7-mediated extension of host cell life span and immortalization. [Cancer Res 2008;68(23):9954-63].

• Imatinib mesylate induces quiescence in gastrointestinal stromal tumor cells through the CDH1-SKP2-p27Kip1 signaling axis.

  Authors: Ying Liu, Sophie A Perdreau, Payel Chatterjee, Linan Wang, Shih-Fan Kuan, Anette Duensing

  Cancer research. 12/2008; 68(21):9015-23.

  Gastrointestinal stromal tumors (GIST) are caused by activating mutations in the KIT or platelet-derived growth factor receptor alpha receptor tyrosine kinase genes. Approximately 85% of GISTGastrointestinal stromal tumors (GIST) are caused by activating mutations in the KIT or platelet-derived growth factor receptor alpha receptor tyrosine kinase genes. Approximately 85% of GIST patients treated with imatinib mesylate achieve disease stabilization, however, often in the presence of residual tumor masses. Complete remissions are rare and a substantial proportion of patients develop resistance to imatinib. Our study was designed to determine whether imatinib-associated responses may account for these clinical findings. We report here that imatinib stimulates cellular quiescence in a proportion of GIST cells as evidenced by up-regulation of the CDK inhibitor p27(Kip1), loss of cyclin A, and reduced BrdUrd incorporation. Mechanistically, these events are associated with an imatinib-induced modulation of the APC/CDH1 signaling axis. Specifically, we provide evidence that imatinib down-regulates SKP2 and that this event is associated with increased nuclear CDH1, an activator of the APC that has been shown to regulate SKP2 stability. We also show that those GIST cells that do not undergo apoptosis in response to imatinib overexpress nuclear p27(Kip1), indicating that they have withdrawn from the cell cycle and are quiescent. Lastly, we provide evidence that a fraction of primary GISTs with high SKP2 expression levels may have an increased risk of disease progression. Taken together, our results support a model in which GIST cells that do not respond to imatinib by apoptosis are removed from the proliferative pool by entering quiescence through modulation of the APC/CDH1-SKP2-p27(Kip1) signaling axis. These results encourage further studies to explore compounds that modulate this pathway as antitumor agents in GISTs.

• Soluble histone H2AX is induced by DNA replication stress and sensitizes cells to undergo apoptosis.

  Authors: Ying Liu, Joshua Parry, Anna Chin, Stefan Duensing, Anette Duensing

  Molecular cancer. 08/2008; 7(1):61.

  ABSTRACT: BACKGROUND: Chromatin-associated histone H2AX is a key regulator of the cellular responses to DNA damage. However, non-nucleosomal functions of histone H2AX are poorly characterized. WeABSTRACT: BACKGROUND: Chromatin-associated histone H2AX is a key regulator of the cellular responses to DNA damage. However, non-nucleosomal functions of histone H2AX are poorly characterized. We have recently shown that soluble H2AX can trigger apoptosis but the mechanisms leading to non-chromatin-associated H2AX are unclear. Here, we tested whether stalling of DNA replication, a common event in cancer cells and the underlying mechanism of various chemotherapeutic agents, can trigger increased soluble H2AX. RESULTS: Transient overexpression of H2AX was found to lead to a detectable fraction of soluble H2AX and was associated with increased apoptosis. This effect was enhanced by the induction of DNA replication stress using the DNA polymerase alpha inhibitor aphidicolin. Cells manipulated to stably express H2AX did not contain soluble H2AX, however, short-term treatment with aphidicolin (1 h) resulted in increased H2AX in the soluble nuclear fraction and enhanced apoptosis. We found that excessive soluble H2AX causes chromatin aggregation and inhibition of ongoing gene transcription as evidenced by the redistribution and/or loss of active RNA polymerase II as well as the transcriptional co-activators CBP and p300. CONCLUSIONS: Taken together, these results show that DNA replication stress rapidly leads to increased soluble H2AX and that non-chromatin-associated H2AX can sensitize cells to undergo apoptosis. Our findings encourage further studies to explore H2AX and the cellular pathways that control its expression as anti-cancer drug targets.

• ERCC1-XPF endonuclease facilitates DNA double-strand break repair.

  Authors: Anwaar Ahmad, Andria Rasile Robinson, Anette Duensing, Ellen van Drunen, H Berna Beverloo, David B Weisberg, Paul Hasty, Jan H J Hoeijmakers, Laura J Niedernhofer

  Molecular and cellular biology. 07/2008;

  ERCC1-XPF endonuclease is required for nucleotide excision repair (NER) of helix-distorting DNA lesions. However, mutations in ERCC1 or XPF in humans or mice cause a more severe phenotype thanERCC1-XPF endonuclease is required for nucleotide excision repair (NER) of helix-distorting DNA lesions. However, mutations in ERCC1 or XPF in humans or mice cause a more severe phenotype than absence of NER, prompting a search for novel repair activities of the nuclease. In Saccharomyces cerevisiae, orthologs of ERCC1-XPF (Rad10-Rad1) participate in the repair of double-strand breaks (DSB). Rad10-Rad1 contributes to two error-prone DSB repair pathways: microhomology-mediated end-joining (a Ku86-independent mechanism) and single-strand annealing. To determine if ERCC1-XPF participates in DSB repair in mammals, mutant cells and mice were screened for sensitivity to gamma-irradiation. ERCC1-XPF-deficient fibroblasts were hypersensitive to gamma-irradiation and gammaH2AX foci, a marker of DSBs, persisted in irradiated mutant cells, consistent with a defect in DSB repair. Mutant mice were also hypersensitive to irradiation, establishing an essential role for ERCC1-XPF in protecting against DSBs in vivo. Mice defective in both ERCC1-XPF and Ku86 were not viable. However, Ercc1(-/-)Ku86(-/-) fibroblasts were hypersensitive to gamma-irradiation compared to single mutants and accumulated significantly greater chromosomal aberrations. Finally, in vitro repair of DSBs with 3'-overhangs led to large deletions in the absence of ERCC1-XPF. These data support the conclusion that like in yeast ERCC1-XPF facilitates DSB repair via an end-joining mechanism that is Ku86-independent.

• Analysis of centrosome overduplication in correlation to cell division errors in high-risk human papillomavirus (HPV)-associated anal neoplasms.

  Authors: Anette Duensing, Anna Chin, Linan Wang, Shih-Fan Kuan, Stefan Duensing

  Virology. 04/2008; 372(1):157-64.

  High-risk HPV-associated anal neoplasms are difficult to treat and biomarkers of malignant progression are needed. A hallmark of carcinogenic progression is genomic instability, which is frequentlyHigh-risk HPV-associated anal neoplasms are difficult to treat and biomarkers of malignant progression are needed. A hallmark of carcinogenic progression is genomic instability, which is frequently associated with cell division errors and aneuploidy. The HPV-16 E7 oncoprotein has been previously shown to rapidly induce centriole and centrosome overduplication and to cooperate with HPV-16 E6 in the induction of abnormal multipolar mitoses. Based on this function, it has been suggested that HPV-16 E7 may act as a driving force for chromosomal instability. However, a detailed analysis of centrosome overduplication in primary HPV-associated neoplasms has not been performed so far. Here, we determined the frequency of centrosome overduplication in HPV-associated anal lesions using a recently identified marker for mature maternal centrioles, Cep170. We detected centrosome overduplication in a small but significant fraction of cells. Remarkably, centrosome overduplication, but not aberrant centrosome numbers per se or centrosome accumulation, correlated significantly with the presence of cell division errors. In addition, our experiments revealed that in particular pseudo-bipolar mitoses may play a role in the propagation of chromosomal instability in high-risk HPV-associated tumors. These results provide new insights into the role of centrosome aberrations in cell division errors and encourage further studies on centrosome overduplication as a predictive biomarker of malignant progression in HPV-associated anal lesions.

• Centrosome-mediated chromosomal instability and steroid hormones as co factors in human papillomavirus-associated cervical carcinogenesis: small viruses help to answer big questions.

  Authors: Anette Duensing, Stefan Duensing

  Advances in experimental medicine and biology. 02/2008; 617:109-17.

• The human papillomavirus type 16 E7 oncoprotein activates the Fanconi anemia (FA) pathway and causes accelerated chromosomal instability in FA cells.

  Authors: Nicole Spardy, Anette Duensing, Domonique Charles, Nathan Haines, Tomomi Nakahara, Paul F Lambert, Stefan Duensing

  Journal of virology. 01/2008; 81(23):13265-70.

  Fanconi anemia (FA) patients have an increased risk for squamous cell carcinomas (SCCs) at sites of predilection for infection with high-risk human papillomavirus (HPV) types, including the oralFanconi anemia (FA) patients have an increased risk for squamous cell carcinomas (SCCs) at sites of predilection for infection with high-risk human papillomavirus (HPV) types, including the oral cavity and the anogenital tract. We show here that activation of the FA pathway is a frequent event in cervical SCCs. We found that FA pathway activation is triggered mainly by the HPV type 16 (HPV-16) E7 oncoprotein and is associated with an enhanced formation of large FANCD2 foci and recruitment of FANCD2 as well as FANCD1/BRCA2 to chromatin. Episomal expression of HPV-16 oncoproteins was sufficient to activate the FA pathway. Importantly, the expression of HPV-16 E7 in FA-deficient cells led to accelerated chromosomal instability. Taken together, our findings establish the FA pathway as an early host cell response to high-risk HPV infection and may help to explain the greatly enhanced susceptibility of FA patients to squamous cell carcinogenesis at anatomic sites that are frequently infected by high-risk HPVs.

• Histone H2AX is a mediator of gastrointestinal stromal tumor cell apoptosis following treatment with imatinib mesylate.

  Authors: Ying Liu, Michelle Tseng, Sophie A Perdreau, Ferdinand Rossi, Cristina Antonescu, Peter Besmer, Jonathan A Fletcher, Stefan Duensing, Anette Duensing

  Cancer research. 04/2007; 67(6):2685-92.

  Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract and are caused by activating mutations of the KIT or platelet-derived growth factorGastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract and are caused by activating mutations of the KIT or platelet-derived growth factor receptor alpha (PDGFRA) tyrosine kinases. GISTs can be successfully treated with imatinib mesylate, a selective small-molecule protein kinase inhibitor that was first clinically approved to target the oncogenic BCR-ABL fusion protein kinase in chronic myelogenous leukemia, but which also potently inhibits KIT and PDGFR family members. The mechanistic events by which KIT/PDGFRA kinase inhibition leads to clinical responses in GIST patients are not known in detail. We report here that imatinib triggers GIST cell apoptosis in part through the up-regulation of soluble histone H2AX, a core histone H2A variant. We found that untreated GIST cells down-regulate H2AX in a pathway that involves KIT, phosphoinositide-3-kinase, and the ubiquitin/proteasome machinery, and that the imatinib-mediated H2AX up-regulation correlates with imatinib sensitivity. Depletion of H2AX attenuated the apoptotic response of GIST cells to imatinib. Soluble H2AX was found to sensitize GIST cells to apoptosis by aberrant chromatin aggregation and a transcriptional block. Our results underscore the importance of H2AX as a human tumor suppressor protein, provide mechanistic insights into imatinib-induced tumor cell apoptosis and establish H2AX as a novel target in cancer therapy.