bluuebunny/crossref_metadata_2025 huggingface dataset:
Gyrification abnormalities in presymptomatic c9orf72 expansion carriers
Authors: Caverzasi Eduardo , Battistella Giovanni , Chu Stephanie A , Rosen Howie , Zanto Theodore P , Karydas Anna , Shwe Wendy , Coppola Giovanni , Geschwind Daniel H , Rademakers Rosa , Miller Bruce L , Gorno-Tempini Maria Luisa , Lee Suzee E
Published: May 2019

Objective

To investigate in-vivo cortical gyrification patterns measured by the local gyrification index (lGI) in presymptomatic c9orf72 expansion carriers compared with healthy controls, and investigate relationships between lGI and cortical thickness, an established morphometric measure of neurodegeneration.

Methods

We assessed cortical gyrification and thickness patterns in a cohort of 15 presymptomatic c9orf72 expansion carriers (age 43.7 ± 10.2 years, 9 females) compared with 67 (age 42.4 ± 12.4 years, 36 females) age and sex matched healthy controls using the dedicated Freesurfer pipeline.

Results

Compared with controls, presymptomatic carriers showed significantly lower lGI in left frontal and right parieto-occipital regions. Interestingly, those areas with abnormal gyrification in presymptomatic carriers showed no concomitant cortical thickness abnormality. Overall, for both presymptomatic carriers and healthy controls, gyrification and cortical thickness measures were not correlated, suggesting that gyrification captures a feature distinct from cortical thickness.

Conclusions

Presymptomatic c9orf72 expansion carriers show regions of abnormally low gyrification as early as their 30s, decades before expected symptom onset. Cortical gyrification represents a novel grey matter metric distinctive from grey matter thickness or volume and detects differences in presymptomatic carriers at an early age.
Abstract DD01-04: NVP-BKM120: A selective inhibitor of type I PI3 kinases
Authors: Sellers William R.
Published: May 2019

AACR 2010 - poster I

Biological characterization of NVP-BKM120, a novel inhibitor of phosphoinosotide 3-kinase in Phase I/II clinical trials

The PI3K/Akt/mTor signaling pathway plays an important role in controlling cell growth, proliferation and survival. Through various mechanisms, the pathway is frequently dysregulated in human cancers, suggesting the use of pathway modulators as novel targeted anticancer therapeutic agents. To this end, substantial drug discovery efforts have been devoted both in pharmaceutical companies and in academia to identify and develop therapeutic agents able to specifically down regulate this pathway in tumors cells. Following the discovery of NVP-BEZ235, our first dual pan-PI3K/mTOR clinical compound, we sought to identify additional PI3K inhibitors from different chemical classes with more stringent selectivity profiles. The key to achieve these objectives was to pursue a structure-based design approach coupled with intensive pharmacological evaluation of selected compounds during the medicinal chemistry optimization process. Here we report on the biological characterization of the pan-PI3K pyrimidine-derived inhibitor NVP-BKM120. This compound inhibits all four Class I PI3K isoforms (IC50 values in the 35 to 248 nM range) with at least 50-fold selectivity (compared to p110[[Unsupported Character - Symbol Font ]]) towards protein kinases. The compound is also active against the most common somatic PI3K[[Unsupported Character - Symbol Font ]] mutations (H1047R, E542K and E545K). NVP-BKM120 does not significantly inhibit the related Class III (Vps34) and Class IV (mTOR, DNA-PK) PI3K kinases. Consistent with its mechanism of action, NVP-BKM120 decreases the cellular levels of p-Akt in mechanistic and relevant tumor cell lines (e.g., IC50 for S473P-Akt in Rat1-p110[[Unsupported Character - Symbol Font ]] cells of 93 nM). This biological activity correlates with inhibition of various Akt downstream signaling pathway components, and with its anti-proliferative activity. Thus, the compound demonstrates significant, concentration dependent cell growth inhibition and induction of apoptosis in a variety of tumor cancer cells, particularly for those harboring p110[[Unsupported Character - Symbol Font ]] mutants and/or over-expressing erbB2. In addition, NVP-BKM120 demonstrates significant, dose dependent in vivo pharmacodynamic activity as measured by inhibition of p-Akt in relevant xenograft models. The pharmacological, biological and preclinical

safety profile of NVP-BKM120 supports its clinical development and the compound is currently undergoing Phase 1/II clinical trials in cancer patients.

Citation Format: William R. Sellers. NVP-BKM120: A selective inhibitor of type I PI3 kinases [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr DD01-04
Abstract SS02-04: How much can we afford to pay for new drugs, and how do we make those decisions?
Authors: Smith Thomas J.
Published: May 2019

Cancer and overall medical care costs are rising at an unsustainable rate. At the same time there are a plethora of new cancer drugs with some activity, all coming to market with a cost of $3000-10,000/month, independent of their absolute benefit. And the cost of drugs does not fall when there are competitors. How should patients, payers, physicians and society choose, in the real world of limited resources? We will explore several alternatives including US and Canadian models and the UK NICE (National Institute for Health and Clinical Excellence) model.

Citation Format: Thomas J. Smith. How much can we afford to pay for new drugs, and how do we make those decisions? [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SS02-04
Abstract SY02-03: Studying tumor evolution in mouse models of cancer
Authors: Jacks Tyler
Published: May 2019

Like other solid tumors, lung adenocarcinoma progresses through multiple stages including tumor initiation, expansion in the lung, and metastasis of the primary tumor to distant organs. Dissection of this process requires quantitative and tractable models that recapitulate the human disease at the genetic and histological level. We have employed autochthonous mouse models of lung adenocarcinoma that recapitulate tumor initiation, progression and metastasis in the human disease both genetically, and pathologically. We have established oncogenic Kras as the initiating event in lung adenoma and amplification of the oncogenic signaling is associated with progression to adenocarcinoma. The p53 tumor suppressor limits tumor progression in this model. Restoration of the Trp53 locus has pinpointed the adenoma-adenocarcinoma transition as the stage in which p53 function is critical for tumor suppression. Expression of oncogenic Kras in the absence of p53 leads to the development of distant metastases after long latency. Identification of the lentiviral integration site in each tumor allows the clonal relationship between metastases and the primary tumors to be unequivocally established. Gene expression and DNA copy number analyses of metastasis-derived cell lines and primary lung tumors have uncovered a metastasis associated gene expression signature and recurrent genomic alterations. Integration of these datasets, other genome-wide analyses and human tumor data are providing insights into the molecular mechanism and biology of lung cancer progression and metastasis.

Citation Format: Tyler Jacks. Studying tumor evolution in mouse models of cancer [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY02-03
Abstract SY09-02: Can sitruin activation treat cancer
Authors: Guarente Leonard P.
Published: May 2019

SIR2 and related genes are NAD-dependent deacetylases that slow aging in yeast, C. elegans, and Drosophila. In yeast and flies, SIR2 genes are also involved in the longevity conferred by dietary or calorie restriction (CR). The mammalian SIR2 homologs termed SIRT genes, or sirtuins, are involved in changes in stress resistance and metabolism and are known to be associated with CR. Critically, the CR diet in rodents not only extends life span, but also protects against many diseases of aging. In this talk, I will describe recent findings in the field regarding SIRT1 function in specific mammalian tissues in relation to murine disease models, with an emphasis on cancer. Our findings indicate that SIRT1 can influence many of the major diseases of aging, including metabolic diseases like diabetes, neurodegenerative diseases (Alzheimer's and Huntington's), cancer and osteoporosis. Therefore small molecules that alter the activity of SIRT1 (i.e. CR mimetic drugs) offer a new approach to prevent and possibly treat the major diseases of aging.

Supported by NIA, The Glenn Foundation, and The AAF. Guarente consults for Sirtris/GSK

Guarente, L. (2007). Sirtuins in aging and disease. CSH Symp. Quant. Biol. 72, 483-488.

Guarente, L. (2008). Mitochondria - a nexus for aging, calorie restriction and sirtuins?. Cell 132, 171-175.

Citation Format: Leonard P. Guarente. Can sitruin activation treat cancer [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY09-02
Abstract SY11-03: Turning enzymes on with small molecules
Authors: Wells James A. , Wolan Dennis , Zorn Julie , Datta Debajyoti , Gray Daniel , Sadowsky Jack
Published: May 2019

Most drug discovery efforts focus on generating inhibitors leading to loss-of-function phenotypes. My lab has been focusing on discovering ways to activate enzymes leading to gain-of-function phenotypes. Many enzymes such as proteases and kinases are stored in dormant “off” forms that are subsequently activated through post-translational modification. Work will be presented for discovery of small molecule activators for the apoptotic proteases known as caspases, as well a Ser-Thr kinase PDK-1. Our group is also developing split-proteins that can be activated by orthogonal small molecule dimerizers to phenocopy the effects seen from the small molecule activators. Such protein activators allow one to ignite signaling from specific nodes and thus probe the sufficiency of enzyme activity to drive a cellular response. Protein activation, as opposed to inhibition, expands the landscape for drug discovery.

Citation Format: James A. Wells, Dennis Wolan, Julie Zorn, Debajyoti Datta, Daniel Gray, Jack Sadowsky. Turning enzymes on with small molecules [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY11-03
Abstract SY13-02: Transcriptional control of tumor-induced myeloid-derived suppressor cells
Authors: Bronte Vincenzo
Published: May 2019

We have generated myeloid-derived suppressor cells (MDSCs) from bone marrow precursors (BM-MDSCs) as a tool to investigate the molecular pathways involved in their immunosuppressive program. BM-MDSCs suppress in vitro both antigen-specific and unspecific CD8+ T cells. The tolerogenic properties of BM-MDSC were further confirmed in vivo following their adoptive transfer in a setting of allogeneic pancreatic insulae transplantation in pharmacologically-induced diabetic mice. The cytokines leading to generation of MDSCs are normally produced during an inflammatory response and act on a common transcription factor which is considered a master regulator of the emergency granulopoyesis. We investigated the effect of genetic knock out of this transcription factor in vivo by creating mice lacking it in myelomonocytic cells. In these mice tumor growth did not cause dysfunction in CD8+ T cells, suggesting that this transcription factor is crucial for their tolerogenic activity and a potential target for new approaches aimed at controlling tumor-induced tolerance.

Citation Format: Vincenzo Bronte. Transcriptional control of tumor-induced myeloid-derived suppressor cells [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY13-02
Abstract SY14-03: Anaplastic lymphoma kinase (ALK): Normal biology and role in hematopoietic malignancies
Authors: Morris Stephan W.
Published: May 2019

Citation Format: Stephan W. Morris. Anaplastic lymphoma kinase (ALK): Normal biology and role in hematopoietic malignancies [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY14-03
Abstract SY15-04: Systematic reconstruction of regulatory networks in mammalian cells: Lessons from the pathogen response
Authors: Regev Aviv
Published: May 2019

Deciphering the regulatory networks that control dynamic and specific gene expression responses in mammalian cells remains a major challenge. While models inferred from genomic data have identified candidate regulatory mechanisms, such models remain largely unvalidated. Here, we present an unbiased strategy based on systematic gene perturbation and innovative multiplex detection to derive regulatory networks in mammalian cells. We apply this approach to decipher the network that controls the transcriptional response to pathogens in primary dendritic cells (DCs), testing the regulatory function of 125 transcription factors, chromatin modifiers, and RNA binding proteins. Our approach accurately assigned 32 known regulators (e.g. NFκB, IRFs, and STATs) to their target genes and discovered 68 additional functional regulators that were not previously implicated in this response. We quantify the contribution of each regulator to two major transcriptional programs (inflammatory and antiviral), identifying a core network of two dozen key regulators and 76 fine-tuners, that uses a combination of coherent feed-forward circuits, dominant activation, and cross-inhibition to control response specificity. Among these we discover a tier of chromatin modifiers that specifically repress interferon beta 1 (IFNβ1) expression upon bacterial but not viral stimulation, and a large circuit of cell cycle regulators that was co-opted to regulate the viral response. Our work establishes a broadly-applicable, comprehensive and unbiased approach to identifying the wiring and function of a regulatory network controlling a major transcriptional response in primary mammalian cells.

Citation Format: Aviv Regev. Systematic reconstruction of regulatory networks in mammalian cells: Lessons from the pathogen response [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY15-04
Abstract SY19-03: Single-cell signaling and pathology in primary cell cancers for mechanism and diagnostics
Authors: Nolan Garry P.
Published: May 2019

Intracellular assays of signaling systems has been limited by an inability to correlate functional subsets of cells in complex populations based on active kinase states or other nodal signaling junctions. Such correlations could be important to distinguish changes in signaling status that arise in rare cell subsets during functional activation or in disease manifestation. We have demonstrated the ability to simultaneously detect activated kinases and phosphoproteins in simultaneous pathways in subpopulations of complex cell populations by multi-parameter flow cytometric analysis, and now via high throughput mass spectrometry at the single cell level.

The focus of the presentation will be our mechanistic studies of human Follicular Lymphoma, as well as signaling of “stem cell” like cell subsets in Acute Myelogenous Leukemia and other myeloid dysplasias. The detailed, correlated datasets generated via single cell phospho-flow allows for ready representation via automated signaling network determination using Bayesian analysis. Our pursuit of deep analysis of these datasets, and the limitations of cloud computing or standard multi-CPU systems, has stimulated our development of unique computational approach using field programmable gate arrays and GPU multiprocessor architectures in the development of a ‘bioinformatics supercomputer’ and associated algorithms.

By analyzing the immune system, or cancer, as individual cells we now observe structured network interactions within these tissues at a new level of clarity. I will present our initial generations of comprehensive network topology maps of signaling within, and between, primary immune subsets in normal and pathologic disease tissues in cancer. My emphasis will be on the application of these approaches directly to human samples in near-patient settings for the development of point-of-care mechanistic referencing of disease and drug action.

Citation Format: Garry P. Nolan. Single-cell signaling and pathology in primary cell cancers for mechanism and diagnostics [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY19-03
Abstract SY19-04: A systems biology approach reveals master regulators of the mesenchymal phenotype of glioblastoma
Authors: Califano Andrea
Published: May 2019

A key challenge in cancer research is the identification of master regulators associated with oncogenesis and tumor progression, i.e., gene products that are necessary and/or sufficient to induce these events. We show that this problem can be successfully tackled by reverse-engineering and unbiased interrogation of context-specific maps of molecular interactions, commonly known as Interactomes.

Specifically, it has been shown that a mesenchymal phenotype is the hallmark of tumor aggressiveness in human malignant glioma. Yet, the regulatory programs determining the associated signature are unknown. We used the ARACNe reverse-engineering algorithm to infer a genome-wide transcriptional interactome, from a large collection of high-grade glioma samples. Interactome-based analysis of the mesenchymal signature of high-grade gliomas revealed a regulatory module causally related to its transcriptional activation.

Two transcription factors (C/EBP and Stat3) emerged as synergistic initiators and master regulators of mesenchymal transformation. Indeed, ectopic co-expression of C/EBPβ and Stat3 was sufficient to reprograms neural stem cells along an aberrant mesenchymal lineage. Conversely, co-silencing of these transcription factors in high-grade glioma cells led to collapse of the mesenchymal signature and systematic reduction in tumor aggressiveness. In human glioma, expression of C/EBPβ and Stat3 correlates with mesenchymal differentiation and predicts poor clinical outcome.

Citation Format: Andrea Califano. A systems biology approach reveals master regulators of the mesenchymal phenotype of glioblastoma [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY19-04
Abstract SY24-03: The signaling landscape of oncogenic ras
Authors: Bar-Sagi Dafna
Published: May 2019

Ras proteins are highly conserved membrane-bound nucleotide binding proteins. They are essential for the transduction of diverse extracellular signals that control cell growth, and abnormal activation of Ras proteins is implicated in the development of several types of human cancers. It is now well established that Ras proteins control cell growth through the activation of multiple effector pathways. These pathways form a signaling network that enables the cell to interpret biological inputs in a context-dependent manner. I will describe our efforts to define a molecular framework for the regulation of Ras-dependent signal output and to understand how this output is modified in cancer cells.

Citation Format: Dafna Bar-Sagi. The signaling landscape of oncogenic ras [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY24-03
Abstract SY28-02: Connections in the BRCA1-BRCA2 pathway of homologous recombination: Implications for breast cancer development and treatment
Authors: Powell Simon N.
Published: May 2019

The breast cancer susceptibility gene, BRCA2, mediates homologous recombination (HR) via its direct interaction with the Rad51 recombinase. There is ample genetic and biological evidence that BRCA1 and BRCA2 are functioning in the same pathway of DNA repair, but the functional connection between the two proteins is poorly understood. Interestingly, a mutation of BRCA1 (S988A) localized normally to ionizing radiation induced nuclear foci (IRIF), but the BRCA2 protein could not be recruited to BRCA1 at the damage site. Phosphorylation of BRCA1 at the S988 site is carried out by Chk2, which was also required for BRCA2 to localize to sites of DNA damage and support homologous recombination. These observations suggest that CHK2-BRCA1-BRCA2-Rad51 function in a single pathway of homologous recombinational repair to prevent the development of breast cancer. In familial breast cancer, one allele of BRCA1 or BRCA2 is germline mutant and the second allele is inactivated to allow tumor development. Thus, loss of function of BRCA1/BRCA2 is a tumor-specific finding, making the role of alternative repair pathways in these cells critical for their survival. We show that loss of Rad52 function is synthetic lethal with BRCA2 deficiency, whereas there is no impact on cell growth in BRCA2-complemented cells. The frequency of both spontaneous and double-strand break-induced homologous recombination, and ionizing radiation induced Rad51 foci, decreased by greater than 50%, when Rad52 was depleted in BRCA2-deficient cells, with no effect in BRCA2-complemented cells. Ionizing radiation-induced and S-phase associated Rad52-Rad51 foci form equally well in the presence or absence of BRCA2, indicating that Rad52 can respond to DNA double strand breaks and replication stalling independently of BRCA2. The observation that Rad52-inactivation is synthetic lethal with BRCA2 and that Rad52 acts independently of BRCA2 makes Rad52 a target for therapy in these tumors. Finally, we have evidence that the BRCA1-BRCA2 pathway can be functionally inactivated in sporadic breast cancer, suggesting that the pool of patients with defects in homologous recombinational repair is larger than initially expected.

Citation Format: Simon N. Powell. Connections in the BRCA1-BRCA2 pathway of homologous recombination: Implications for breast cancer development and treatment [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY28-02
Abstract SY29-02: The Mdm2/Mdm4 complex: A rheostat for p53 activity
Authors: Lozano Guillermina
Published: May 2019

The activity of the p53 tumor suppressor is regulated by multiple factors. Two of these, Mdm2 and Mdm4, function as p53 inhibitors in vivo in multiple cell types. Loss of Mdm2 or Mdm4 in mice leads to embryo and cell lethal phenotypes that are rescued by deletion of p53. Additionally, haplo insufficiency at these loci results in elevated p53 activity, delayed lymphomagenesis, and developmental defects in some cases. On the other hand, over expression of Mdm2 and Mdm4 occurs in many tumors most of which retain wild type p53. Mdm2 and Mdm4 transgenic mice are tumor prone. Additionally, a single nucleotide polymorphism (SNP) in the human MDM2 promoter increases the levels of MDM2 expression is associated with increased cancer risk. A mouse model with this SNP shows increased Mdm2 levels, decreased p53 activity, and an enhanced tumor phenotype. These data emphasize the exquisite sensitivity of the p53 pathway to changes in Mdm2 and Mdm4 gene dosage, and the consequences of those effects on development and tumorigenesis.

Citation Format: Guillermina Lozano. The Mdm2/Mdm4 complex: A rheostat for p53 activity [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY29-02
Abstract SY31-01: The role of autophagy in cancer progression and therapy
Authors: White Eileen P.
Published: May 2019

Autophagy is induced by metabolic stress and can promote cell survival by maintaining energy homeostasis and by damage mitigation. Autophagy is negatively regulated by activation of mammalian target of rapamycin (mTOR) that links nutrient and growth factor availability to cell growth and catabolism. mTOR inhibitors have shown efficacy in renal cell carcinoma (RCC) but the role of autophagy stimulation and potentially counterproductive promotion of tumor cell stress tolerance was not known. We have found that the mTOR inhibitor CCI-779 (temsirolimus) promoted autophagy-mediated stress tolerance and that inhibition of autophagic degradation with chloroquine enhanced kidney tumor cell death in vitro and in vivo. Thus, autophagy inhibition may be a useful strategy to enhance the anti-tumor activity of mTOR inhibitors in RCC. Although therapeutic inhibitors of the mTOR pathway have been demonstrated to slow progression of advanced RCC in patients, and were approved by the FDA as a standard therapy for RCC, this beneficial effect is only temporary. One possible mechanism of resistance may be due to autophagy as a survival response. Additionally, metabolic stress caused by insufficient angiogenesis and the resulting mTOR inhibition activate tumor cell survival by autophagy, and tumor cells in these hypoxic regions respond most poorly to treatment. We reveal a new therapeutic paradigm of coordinate mTOR and autophagy inhibition to block the survival advantage and enhance tumor cell death to maximize therapeutic benefit.

Citation Format: Eileen P. White. The role of autophagy in cancer progression and therapy [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY31-01
Abstract SY31-02: Autophagy, cancer cell survival, and mitochondrial homeostasis
Authors: Debnath Jayanta
Published: May 2019

Autophagy is a tightly regulated lysosomal self-digestion process that serves as an important adaptive mechanism during stress or starvation. My laboratory recently discovered that autophagy is strongly induced in epithelial cells deprived of extracellular (ECM) contact, which protects them from apoptosis. In further work, we have uncovered that autophagy is robustly induced in epithelial cells transformed by mutationally active oncogenes, raising the hypothesis that autophagy functions as a survival mechanism in cancerous cells deprived of proper ECM contact. We are currently dissecting whether detachment-induced autophagy contributes to oncogenic cell survival and transformation in vitro, and promotes the dissemination and metastasis of tumor cells in vivo.

In addition, we are uncovering novel cellular functions for ATGs, the proteins that regulate autophagy. ATG12 is an ubiquitin-like modifier required for autophagy, which to date, has been proposed to possess a single conjugation target, another autophagy regulator called ATG5. Now, we identify ATG3 as a novel substrate for ATG12 conjugation. ATG3 is the E2-like enzyme necessary for ATG8/LC3 lipidation during autophagy. ATG12-3 complex formation requires ATG7 as the E1 enzyme and ATG3 autocatalytic activity as the E2, resulting in the covalent linkage of ATG12 onto a single lysine on ATG3. Mutation of this single lysine abolishes ATG12-ATG3 complex formation.

To uncover the biological functions of ATG12-ATG3, we reconstituted ATG3-deficient cells with wild type ATG3 (ATG3WT) or a mutant version unable to conjugate to ATG12 (ATG3KR). Remarkably, disrupting ATG12-3 complex formation has no observable effect on autophagy. In fact, during nutrient starvation and rapamycin- mediated mTORC1 inhibition, ATG3WT and ATG3KR-expressing cells exhibit equal rates of both LC3 lipidation (LC3-II) and turnover in the lysosome, as well as the robust proteolytic degradation of autophagy substrates like p62/SQSTM.

Instead, the most obvious consequence of disrupting ATG12-3 is an expansion in mitochondrial mass due to autophagy- and mitophagy-independent mechanisms. Lack of ATG12 conjugation to ATG3 also produces an increase in mitochondrial fragmentation, arising from a defect in mitochondrial fusion. Furthermore, cells lacking ATG12-3 exhibit resistance to the intrinsic apoptotic pathway. Most importantly, these unique functions of the ATG12-3 complex can be completely separated from the established roles of either individual ATG on autophagosome formation.

Overall, these results reveal unexpected novel roles for ATG12-3 in mitochondrial homeostasis and cell death, and for the first time, implicate the ATG12 conjugation system in new cellular functions distinct from the early steps of autophagosome formation.

Citation Format: Jayanta Debnath. Autophagy, cancer cell survival, and mitochondrial homeostasis [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY31-02
Abstract SY33-04: Role of shelterin in cancer and aging: Generation of conditional knockout mice for TRF1, TPP1, and RAP1
Authors: Blasco Maria
Published: May 2019

Shelterin is a highly conserved chromosome end-capping complex, which encompasses TRF1, TRF2, TPP1, POT1, TIN2 and RAP1. Although extensive cell culture studies suggest that shelterin is essential for telomere regulation, its role in telomere biology and disease in the context of the organism remained elusive due to lack of viable loss-of-function mouse models. I will report on mice and cells conditionally deleted for TRF1, TPP1 and RAP1. While deletion of TRF1 and TPP1 in mouse stratified epithelia results in perinatal mortality, severe skin atrophy and hyperpigmentation, and widespread epithelial dysplasia and tumors in the absence of p53, RAP1 deletion does not affect mouse viability and only leads to mild skin hyperpigmentation at aduldhood. This differential impact of TPP1 and TRF1 deletion compared to RAP1 deletion is in agreement with severe telomere uncapping produced by either TPP1 or TRF1 abrogation, which is not observed upon RAP1 deletion. These results argue that RAP1 is not an essential telomere capping protein in mammals. Instead, we show here that both TPP1 and RAP1 are important to recruit telomerase to telomeres and to maintain telomere length. In addition, we demonstrate that RAP1 is important for silencing of genes located near the telomeres, demonstrating a conserved role of RAP1 in subtelomeric gene silencing from yeast to mammals.

Citation Format: Maria Blasco. Role of shelterin in cancer and aging: Generation of conditional knockout mice for TRF1, TPP1, and RAP1 [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY33-04
Abstract SY34-04: Do we need to know what pO2 hypoxia is
Authors: Koch Cameron J.
Published: May 2019

Despite its widespread use in the literature, the word ‘hypoxia’ is often missing from dictionaries. It means “a deficiency of oxygen reaching the tissues of the body” (Encyclopaedia Britannica). Since many molecular, biological and chemical phenomena are related to the actual oxygen concentration (most commonly estimated in units of partial pressure for tissue) it seems reasonable to suggest that one must carefully define the precise nature of ‘hypoxia’ in tissue. In addition to many other possible applications, this first step is critical before attempting to overcome hypoxia's many deleterious effects by therapeutic intervention. Among its multiple complexities, and consistent with its vague definition, hypoxia in tumors can be caused by several distinct mechanisms. Although most of these mechanisms have been predicted to cause oxygen-deficits in isolated small volumes or thin inter-connected shells of tissue, we have recently described a new mechanism that leads to what we have termed ‘extended regional hypoxia’, whereby entire zones of hypoxic tissue, having multi-millimeter dimensions, may form.

In order to address the biological and/or therapeutic importance of the complexity described above, we clearly need to be able to distinguish between the causes of hypoxia, and then determine the relevance of this to either the biology or therapy of tumors and normal tissue. Unfortunately, the development of validated methods to determine the type and degree of hypoxia is far from complete. This talk will discuss various approaches to understanding tumor ‘hypoxia’ using the hypoxia marker EF5 (a 2-nitroimidazole), both from the point of view of invasive and non-invasive imaging. A novel modification of the EF5 binding technique has allowed an initial description of the mRNA transcriptional changes that accompany ‘hypoxia’ in a syngeneic rat tumor model.

Citation Format: Cameron J. Koch. Do we need to know what pO2 hypoxia is [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY34-04
Abstract 1210: Mifepristone abrogates repopulation of ovarian cancer cells in between courses of cisplatin treatment
Authors: Freeburg Elizabeth M , Goyeneche Alicia A , Telleria Carlos M.
Published: May 2019

Repopulation of cancer cells that escape cytotoxic chemotherapy is a critical factor that negatively impacts treatment success. Thus, any strategy that prevents this phenomenon from occurring would be beneficial for the outcome of the treatment. One such strategy to inhibit tumor cell repopulation is the use of cytostatic compounds among courses of cytotoxic therapy. In this study, we tested the hypothesis that mifepristone, a steroid compound displaying a potent cytostatic effect in ovarian cancer as recently demonstrated in our laboratory (Clin Cancer Res 2007, 13, 3370-3379), should be efficacious to induce cytostasis and to prevent ovarian cancer repopulation if given in between rounds of cisplatin therapy. We established an in vitro approach in which OV2008 cisplatin-sensitive ovarian cancer cells were exposed to three rounds of a cytotoxic dose (20 μM) of cisplatin for 1 h, twelve days apart. Every four days the number of cells, their viability, and their distribution within the cell cycle were assessed by micro-cytometry using exclusion fluorochromes and propidium iodide, respectively. Further, the colony-forming capacity of the viable cells was evaluated along the experiment. Although cisplatin killed the vast majority of OV2008 cells, there were cells that escaped cisplatin toxicity and repopulated the culture. This phenomenon occurred each one of the three times the cells were challenged by cisplatin during the 36 days the study lasted. Post-cisplatin repopulation was evidenced by the increase in number and clonogenic capacity of the viable cells, the normalization in the distribution of cells within the different phases of the cell cycle (S, G1 and G2/M), and the concomitant reduction in sub-diploid DNA content (as marker of cytotoxicity). Instead, when the cells were exposed to cisplatin for 1 h, but 5, 10 or 20 μM mifepristone was present in the culture medium after removal of the platinum agent, both number of cells and their clonogenic capacity were reduced in a dose-related manner. Remarkably, the cultures exposed to cisplatin for 1 h followed by chronic exposure to 20 μM mifepristone did not have remaining viable cells to allow performing the clonogenic survival assay by day 12 of the study, suggesting an added lethality to the cytostatic effect of mifepristone. In conclusion, this study demonstrates that presence of cytostatic concentrations of mifepristone prevents the repopulation of ovarian cancer cells surviving the lethality of cisplatin, and further suggests mifepristone may potentiate cisplatin toxicity. The scheduling of mifepristone between courses of cisplatin-based therapy for human ovarian cancer has potential for improving treatment success. Supported by NIH grant CA121991.
Abstract 1582: Identification of BCAR genes relevant for breast cancer progression and endocrine therapy resistance
Authors: Van Agthoven Ton , Veldscholte Jos , Sieuwerts Anieta , Smid Marcel , Meijer-Van Gelder Marion E , Look Maxime P , Foekens John A , Dorssers Lambert C. J.
Published: May 2019

Endocrine therapy of breast cancer is effective and widely applied. In advanced disease, about half of the patients will experience a clinical benefit from tamoxifen treatment, but ultimately the disease will progress into a resistant phenotype. The molecular basis of this general failure of endocrine therapy is not understood yet. Although gene expression profiling experiments provide gene lists to identify patients who may benefit from specific treatments, these genes mostly do not define the underlying molecular mechanisms. We have executed a functional screen for the identification of BCAR genes causing anti-estrogen resistance in a human breast cancer model and evaluated their role in a large breast cancer cohort.

Estrogen-dependent ZR-75-1 human breast cancer cells were subjected to insertion mutagenesis with replication-defective retroviruses and selected for proliferation in the presence of 4-hydroxy tamoxifen. In our panel of 79 tamoxifen-resistant cell lines, the target genes of the retroviruses have been identified and were functionally tested following transfection into ZR-75-1 cells. Seven BCAR genes (AKT1, AKT2, BCAR1, BCAR3, EGFR, GRB7 and TRERF1) causing the tamoxifen-resistant phenotype were identified. Furthermore, additional putative BCAR genes and loci have been pinpointed.

To determine the relevance of the identified genes in clinical breast cancer, we have studied 561 estrogen receptor-positive primary breast cancers for tumor aggressiveness and tamoxifen therapy resistance. Quantitative RT-PCR of ten genes was performed to measure the tumor mRNA levels. The endpoint for tumor aggressiveness was metastasis-free survival (MFS) of a cohort of lymph node-negative patients. Therapy resistance was evaluated in a group of patients treated with tamoxifen as first-line therapy for recurrent disease and the endpoint of this analysis was progression-free survival (PFS). Statistically significant associations with MFS were found for AKT2, EGFR, TLE3 and TRERF1 mRNA levels. In multivariate analyses including the traditional prognostic factors, AKT2, EGFR and TRERF1 were independent prognostic factors for MFS and thus are associated with tumor aggressiveness. In the analyses of PFS, mRNA levels of BCAR3, ERBB2, GRB7, TLE3 and TRERF1 were associated with the duration of clinical benefit of tamoxifen therapy, independently of the classical predictive factors.

Our functional screen identified a set of clinically relevant BCAR genes offering insight in the molecular and cellular pathways of breast tumor aggressiveness and tamoxifen resistance, and thereby provides novel targets for personalized treatment and prevention strategies.

Supported by the Dutch Cancer Society, the Susan G Komen Breast Cancer Foundation and the Association for International Cancer Research.
Abstract 1604: Dietary energy balance modulates multistage epithelial carcinogenesis in mouse skin
Authors: Moore Tricia , Carbajal Steve , Jiang Anna , Beltran Linda , Hursting Stephen , DiGiovanni John
Published: May 2019

Dietary energy balance refers to the balance between caloric intake and energy expenditure. Findings from both epidemiological and experimental studies suggest chronic positive energy balance, which can lead to obesity, heightens the risk of developing multiple cancers, while a negative energy balance state, as induced by calorie restriction (CR), decreases these risks. In the present study, we used the two-stage skin carcinogenesis model to examine the effect of dietary energy balance manipulation on both the promotion and progression stages of skin tumorigenesis. Groups of female ICR mice (n=30) were initiated with 25 nmol DMBA and then placed on one of four dietary treatment regimens: 30% CR, 15% CR, 10Kcal% fat (control), or 60Kcal% fat [diet-induced obesity (DIO) diet]. Following four weeks on diet, mice were then promoted with either acetone or 6.8 nmol TPA twice-weekly for the duration of the study. Chronic CR (30% CR) led to a statistically significant inhibition of papilloma formation in the TPA treated group compared to the other TPA treated diet groups. In contrast, while there was a slightly higher number of papillomas per mouse in the DIO diet group treated with TPA, this was not statistically different from the control and 15% CR groups treated with TPA. This experiment is currently being carried to carcinoma development to determine the impact of dietary energy balance manipulation on the conversion of papillomas to carcinomas in this model of epithelial multistage carcinogenesis. Mechanistic studies have been performed to examine the impact of dietary energy balance manipulation on epidermal cell signaling pathways related to TPA-mediated skin tumor promotion. In this regard, CR reduced whereas DIO increased activation of both epidermal Akt and mTOR (assessed by Western blot analysis of phosphorylated proteins) both in the absence and presence of TPA. Furthermore, activation of both the EGFr and IGF-1R was modulated in a similar manner by the different diets. These results suggest that dietary energy balance may modulate tumor promotion in part through modulation of critical signaling pathways downstream of cell surface receptors such as the EGFr and IGF-1R.
Abstract 1795: Noninvasive genetic detection of head and neck squamous cell carcinoma
Authors: Sethi Seema , Benninger Michael S , Lu Mei , Havard Shaleta , Worsham Maria J
Published: May 2019

Purpose of the study: To develop and evaluate a noninvasive approach for head and neck squamous cell carcinoma (HNSCC) based on genomic alterations of loss and gain of gene copy number.

Experimental Procedures: The study cohort comprised 37 subjects, 27 HNSCC patients and 10 healthy controls. DNA extracted from saliva of the study subjects was interrogated by multiplex ligation-dependent probe amplification (MLPA) assay for gene copy number alterations (losses and gains) using two panels of 82 gene probes with known associations to HNSCC. MLPA permits evaluation of 41 different genes in one PCR reaction using scant amounts of DNA. The data were analyzed using the Classification and Regression Tree (CART) to determine whether gene alterations detected in saliva DNA discriminated HNSCC patients from normal controls. Results were validated using the leave-one-out validation approach.

Results: Of the 82 genes analyzed, 11 genes, ERBB2, ABCC4, UTY, DNMT1, PMAIP1, CDKN2B, CDKN2D, NFKB1, TP53, PTPN1, DCC, had the highest individual predictive ability for HNSCC. CART analysis revealed gain of PMAIP1 (18q21.31) solely or in conjunction with gain of PTPN1 (20q13.13) genes classified all 27 HNSCC patients with 100% sensitivity and 100% specificity. Validation results showed a sensitivity and specificity of 96% and 90% respectively.

Conclusions: Gain of PMAIP1 (18q21.31) and PTPN1 (20q13.13) genes definitively separated HNSCC patients from normal controls, providing proof of concept that saliva based gene algorithms may have clinical utility for noninvasive HNSCC detection.

Keywords: Genetic alterations; Head and neck squamous cell carcinoma (HNSCC); Multiplex Ligation-dependent Probe Amplification Assay (MLPA); Classification and Regression Tree (CART).

Support:

HFHS Dev Grant 06 (SS), R01 NIH DE 15990 (MJW).
Abstract 2029: Identification and characterization of cancer stem cells in ovarian cancer
Authors: Mor Gil , Chen Rui , Fu Han Hsuan , Montagna Michele , Visintin Irene , Silasi Dan , Schwartz Peter , Azodi Masoud , Rutherford Thomas , Alvero Ayesha
Published: May 2019

Introduction: One of the major burdens in the treatment of epithelial ovarian cancer (EOC) is the high percentage of recurrence characterized by chemoresistance. The biology underlying the tumor’s high capacity for recurrence has not been elucidated. Tumors are made of a heterogeneous cell population consisting of both cancer and non-cancer cells. New data suggest that even the cancer cell population is heterogeneous and contains a small subset of cells, the cancer stem cells (CSC), which constitute a reservoir that can self-renew and therefore maintain the tumor. CSC can divide and expand their pool and also differentiate into non-CSC, which constitute the bulk of the tumor. Contrary to CSC, the non-CSC are rapidly dividing and therefore sensitive to therapies, which target highly proliferative cells. In the present study we identified and characterized the CSC of EOC.

Methods: EOC cells were isolated from malignant ovarian cancer ascities and solid tumors (n=80). Marker expression was determined using Flow Cytometry, Western Blots and Immunocytochemistry. Xenograft nude mice model was used for tumor growth by injecting cancer cells either s.c. or i.p. Isolation of CD44+ population was done using FACS. Cancer cells were maintained in culture as previously described {Kamsteeg, 2003 #358}

Results: CSC were identified in EOC cells isolated form ascites and solid tumors with the following characteristics: 1) cellular markers: CD44+, MyD88+, constitutive NFκB activity and cytokine and chemokine production, high capacity for repair, chemoresistance to conventional chemotherapies, resistance to TNFα-mediated apoptosis, capacity to form spheroids in suspension, and unique microRNA phenotype; 2) tumor formation in animals: 100% CD44+ cells formed tumors which contained 10% CD44+ and 90% CD44-negative cells. Re-injection of isolated CD44+ cells from previous engraftment was able to again recapitulate the original tumor phenotype. Isolation and in vitro treatment of CD44+ cells from fresh samples showed resistance to carboplatin and paclitaxel. In contrast, the sorted CD44-negative cell population from the same sample/patient was chemosensitive.

Conclusion: Present chemotherapy modalities eliminate the bulk of the tumor but it leaves a core of cancer cells with high capacity for repair and renewal. CSC corresponds to the core of malignant cells that promotes recurrence and chemoresistance. Identification of these cells represents the first step in the development of therapeutic modalities that can eliminate not only the bulk of the tumor but its source. Prevention of recurrence will be achieved only with therapies targeting this cell population.