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Significant strides have been made in the development of precision therapeutics for cancer. Aberrantly expressed glycoproteins represent a potential avenue for therapeutic development. The MUC16/CA125 glycoprotein serves as a biomarker of disease and a driver of malignant transformation in epithelial ovarian cancer. Previously, we demonstrated a proof-of-principle approach to selectively targeting MUC16+ cells. In this report, we performed a synthetic lethal kinase screen using a human kinome RNAi library and identified key pathways preferentially targetable in MUC16+ cells using isogenic dual-fluorescence ovarian cancer cell lines. Using a separate approach, we performed high-content small-molecule screening of six different libraries of 356,982 compounds for MUC16/CA125-selective agents and identified lead candidates that showed preferential cytotoxicity in MUC16+ cells. Compounds with differential activity were selected and tested in various other ovarian cell lines or isogenic pairs to identify lead compounds for structure–activity relationship (SAR) selection. Lead siRNA and small-molecule inhibitor candidates preferentially inhibited invasion of MUC16+ cells in vitro and in vivo, and we show that this is due to decreased activation of MAPK, and non–receptor tyrosine kinases. Taken together, we present a comprehensive screening approach to the development of a novel class of MUC16-selective targeted therapeutics and identify candidates suitable for further clinical development.

Supplementary Figure from Dual Fluorescence Isogenic Synthetic Lethal Kinase Screen and High-Content Secondary Screening for MUC16/CA125-Selective Agents

Significant strides have been made in the development of precision therapeutics for cancer. Aberrantly expressed glycoproteins represent a potential avenue for therapeutic development. The MUC16/CA125 glycoprotein serves as a biomarker of disease and a driver of malignant transformation in epithelial ovarian cancer. Previously, we demonstrated a proof-of-principle approach to selectively targeting MUC16+ cells. In this report, we performed a synthetic lethal kinase screen using a human kinome RNAi library and identified key pathways preferentially targetable in MUC16+ cells using isogenic dual-fluorescence ovarian cancer cell lines. Using a separate approach, we performed high-content small-molecule screening of six different libraries of 356,982 compounds for MUC16/CA125-selective agents and identified lead candidates that showed preferential cytotoxicity in MUC16+ cells. Compounds with differential activity were selected and tested in various other ovarian cell lines or isogenic pairs to identify lead compounds for structure–activity relationship (SAR) selection. Lead siRNA and small-molecule inhibitor candidates preferentially inhibited invasion of MUC16+ cells in vitro and in vivo, and we show that this is due to decreased activation of MAPK, and non–receptor tyrosine kinases. Taken together, we present a comprehensive screening approach to the development of a novel class of MUC16-selective targeted therapeutics and identify candidates suitable for further clinical development.

In this paper, we present a compliant six-axis sensor for total ground reaction measurement, the calibration methodology, and preliminary data. The sensor is intended for gait analysis and is designed to have minimal effect on natural gait. A triaxial optical force sensor is combined with an array of pressure sensing films to form a wearable compliant six-axis force/moment sensor. Two sensor units were developed for the toe and heel and used in two types of experiments: stepping on the sensors that are mounted on the ground and attaching the sensors under the shoe while walking. The data from the sensors are compared with measurements obtained from a standard force plate. The deflections induced by the sensor compliance exhibit a slight nonlinear force–deflection relation. Regardless of the nonlinear effects, the sensor is accurately calibrated with a linear least squares method. To see how well these nonlinearities could be calibrated for, a nonlinear calibration with a neural network was used. For the sensors attached to the floor, a linear calibration achieved an RMSE of 4.49% while the neural network achieves 2.68%. For the wearable sensor, the linear calibration RMSE was 9.39% and the neural network RMSE was 5.21%. When the orientation data of the sensors (measured by a motion capture system) were added as an input to the neural network calibration, the RMSE was reduced to 3.25%.

The molecular foundations of Hürthle cell carcinoma (HCC) are poorly understood. Here, we describe a comprehensive genomic characterization of 56 primary HCC tumors that span the spectrum of tumor behavior. We elucidate the mutational profile and driver mutations and show that they exhibit a wide range of recurrent mutations. Notably, we report an extremely high number of disruptive mutations to both protein-coding and tRNA-encoding regions of the mitochondrial genome. We reveal unique chromosomal landscapes that involve whole-chromosomal duplications of chromosomes 5 and 7 and widespread loss of heterozygosity arising from haploidization and copy number–neutral uniparental disomy. We also identify fusion genes and disrupted signaling pathways that may drive disease pathogenesis.

Context:Hurthle cell cancer (HCC) is an understudied cancer with poor prognosis.Objective:Our objective was to elucidate the genomic foundations of HCC.Design and Setting:We conducted a large-scale integrated analysis of mutations, gene expression profiles, and copy number alterations in HCC at a single tertiary-care cancer institution.Methods:Mass spectrometry-based genotyping was used to interrogate hot spot point mutations in the most common thyroid oncogenes: BRAF, RET, NRAS, HRAS, KRAS, PIK3CA, MAP2K1, and AKT1. In addition, common oncogenic fusions of RET and NTRK1 as well as PAX8/PPARγ and AKAP9-BRAF were also assessed by RT-PCR. Global copy number changes and gene expression profiles were determined in the same tumor set as the mutational analyses.Results:We report that the mutational, transcriptional, and copy number profiles of HCC were distinct from those of papillary thyroid cancer and follicular thyroid cancer, indicating HCC to be a unique type of thyroid malignancy. Unsupervised hierarchical clustering of gene expression showed the 3 groups of Hurthle tumors (Hurthle cell adenoma [HA], minimally invasive Hurthle cell carcinoma [HMIN], and widely invasive Hurthle cell carcinoma [HWIDE] clustered separately with a marked difference between HWIDE and HA. Global copy number analysis also indicated distinct subgroups of tumors that may arise as HWIDE and HMIN. Molecular pathways that differentiate HA from HWIDE included the PIK3CA-Akt-mTOR and Wnt/β-catenin pathways, potentially providing a rationale for new targets for this type of malignancy.Conclusions:Our data provide evidence that HCC may be a unique thyroid cancer distinct from papillary and follicular thyroid cancer.

Aberrant Wnt signaling can drive cancer development. In many cancer types, the genetic basis of Wnt pathway activation remains incompletely understood. Here, we report recurrent somatic mutations of the Drosophila melanogaster tumor suppressor-related gene FAT1 in glioblastoma (20.5%), colorectal cancer (7.7%), and head and neck cancer (6.7%). FAT1 encodes a cadherin-like protein, which we found is able to potently suppress cancer cell growth in vitro and in vivo by binding β-catenin and antagonizing its nuclear localization. Inactivation of FAT1 via mutation therefore promotes Wnt signaling and tumorigenesis and affects patient survival. Taken together, these data strongly point to FAT1 as a tumor suppressor gene driving loss of chromosome 4q35, a prevalent region of deletion in cancer. Loss of FAT1 function is a frequent event during oncogenesis. These findings address two outstanding issues in cancer biology: the basis of Wnt activation in non-colorectal tumors and the identity of a 4q35 tumor suppressor.

Adenoid cystic carcinomas (ACCs) are among the most enigmatic of human malignancies. These aggressive salivary gland cancers frequently recur and metastasize despite definitive treatment, with no known effective chemotherapy regimen. Here we determined the ACC mutational landscape and report the exome or whole-genome sequences of 60 ACC tumor-normal pairs. These analyses identified a low exonic somatic mutation rate (0.31 non-silent events per megabase) and wide mutational diversity. Notably, we found mutations in genes encoding chromatin-state regulators, such as SMARCA2, CREBBP and KDM6A, suggesting that there is aberrant epigenetic regulation in ACC oncogenesis. Mutations in genes central to the DNA damage response and protein kinase A signaling also implicate these processes. We observed MYB-NFIB translocations and somatic mutations in MYB-associated genes, solidifying the role of these aberrations as critical events in ACC. Lastly, we identified recurrent mutations in the FGF-IGF-PI3K pathway (30% of tumors) that might represent new avenues for therapy. Collectively, our observations establish a molecular foundation for understanding and exploring new treatments for ACC.

Activation of the PI3K and epidermal growth factor receptor (EGFR) pathway is able to drive oncogenesis in multiple human cancers, including head and neck squamous cell carcinoma. Targeted agents such as cetuximab and erlotinib are currently used in patients with head and neck squamous cell carcinoma, but, in this disease, the genomic alterations that cause pathway activation and determine response to pharmacologic inhibition remain ill-defined. Here, we present a detailed dissection of the EGFR/PI3K pathway, composed of sequencing of the core pathway components, and high-resolution genomic copy number assessment. Mutations were found in PIK3CA (6%), but no point mutations were observed in other pathway genes such as PTEN and EGFR . In contrast, we observed frequent copy number alterations of genes in the pathway, including PIK3CA , EGFR , protein tyrosine phosphatase receptor S ( PTPRS ), and RICTOR . In total, activating genetic pathway alterations were identified in 74% of head and neck tumors. Importantly, intragenic microdeletions of the EGFR phosphatase PTPRS were frequent (26%), identifying this gene as a target of 19p13 loss. PTPRS loss promoted EGFR/PI3K pathway activation, modulated resistance to EGFR inhibition, and strongly determined survival in lung cancer patients with activating EGFR mutations. These findings have important implications for our understanding of head and neck cancer tumorigenesis and for the use of targeted agents for this malignancy.

Background: MUC16 is a tethered, cell surface mucin which is over expressed on ovarian cancer cells and encodes the CA125 antigen. The CA125 antigen is present in the serum of women with ovarian cancer and appears to be an independent adverse marker of prognosis. Elements of the MUC16 molecule appear to promote invasion, alter gene expression and will induce malignant transformation in NIH 3T3 cells. We have previously demonstrated that MUC16 expression alters the expression of invasion/metastasis genes and genes linked to drug resistance. We hypothesize that the forced expression of MUC16 induces changes that might be relected in altered kinase activities and provides an opportunity to specifically target the MUC16+ ovarian cancer cell. Methods: A human kinome siRNA library (2 siRNA's/well) was purchased from Qiagen. A2780 cells with and without MUC16 expression were labeled with GFP or mCherry Fluorescence proteins and plated in 96 well plates at 25000 cells/well. Each siRNA was plated into six replicate wells and analyzed for inhibition compared to negative controls and a proprietary “All Star Death” siRNA mix. If the candidate siRNA inhibited the proliferation of MUC16+ cells statistically (p Results: To date, we have screened siRNA's for a total of 160 of the planned 714 kinases. From the initial screen, we identified 28 kinase candidates for additional evaluation. Two additional siRNA's per kinase were obtained for confirmatory screening. From this initial group of kinases, inhibition of 3 kinases (CHEK1, ABL1 and CDK4) were significantly more inhibitory for MUC16+ cells than the syngeneic MUC16- comparator. Eight additional kinases (including CSK, CHUK, ILK and LTK) were more inhibited in the MUC16+ cells for 3 of 4 tested siRNA's for the target kinase. Although CDK4 protein levels were not altered in MUC16 transformed A2780 cells, CDK4 inhibitors were differentially active between the MUC16 +/- syngeneic pair and additional testing with other small molecule inhibitors is ongoing. Conclusions: 1)MUC16 expression alters the biologic behavior of ovarian cancer cells and 2)MUC16 targeted, syngeneic screening of kinase siRNA libraries can identify potential clinical target candidates for ovarian cancer selected for MUC16. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B222.

Background Despite over 2 decades of research, the ability to prevent work-related low back pain (LBP) and disability remains elusive. Recent research suggests that interventions that are focused at the workplace and incorporate the principals of participatory ergonomics and return-to-work (RTW) coordination can improve RTW and reduce disability following a work-related back injury. Workplace interventions or programs to improve RTW are difficult to design and implement given the various individuals and environments involved, each with their own unique circumstances. Intervention mapping provides a framework for designing and implementing complex interventions or programs. The objective of this study is to design a best evidence RTW program for occupational LBP tailored to the Ontario setting using an intervention mapping approach. Methods We used a qualitative synthesis based on the intervention mapping methodology. Best evidence from systematic reviews, practice guidelines and key articles on the prognosis and management of LBP and improving RTW was combined with theoretical models for managing LBP and changing behaviour. This was then systematically operationalized into a RTW program using consensus among experts and stakeholders. The RTW Program was further refined following feedback from nine focus groups with various stakeholders. Results A detailed five step RTW program was developed. The key features of the program include; having trained personnel coordinate the RTW process, identifying and ranking barriers and solutions to RTW from the perspective of all important stakeholders, mediating practical solutions at the workplace and, empowering the injured worker in RTW decision-making. Conclusion Intervention mapping provided a useful framework to develop a comprehensive RTW program tailored to the Ontario setting.

Background Some studies in adults indicate a positive correlation between eating later in the day and overall energy intake as well as body weight status. Thus, the time of food intake may be a risk factor in childhood obesity. This study was designed to describe the proportion of energy consumed in the time from 4 pm to midnight measured in two-hour increments and to determine a potential association between the time of proportion of energy consumed and body weight status. Methods Dietary, anthropometric, and socio-demographic data of 2–18 year olds (N = 11,072) of the National Health and Nutrition Examination Survey (NHANES) 1999–2004 was examined to describe the proportion of total energy consumed within two-hour time periods between 4 pm and midnight. To examine the potential association between eating later in the day and body weight status, generalized estimating equations (GEE) models were used to quantify the effect of time trends (proportion of total energy consumed in each 2-hour time period from 4 pm to 11.59 pm) on body weight status. Analysis was conducted in the total sample and in subgroups stratified by sex, ethnic group (Non-Hispanic white, Non-Hispanic black, Mexican American, Other Hispanic, and Other Race including multi-racial) and age group (2–5, 6–11, and 12–18 year olds). Complex sample survey analysis were used to assess differences at a significance level of p-value < 0.05. Results Proportion of energy consumed varied by sex, ethnic group, and age groups between 4 pm and 11.59 pm. Compared to healthy weight children, overweight school-age children consumed significantly higher while overweight adolescents consumed significantly lower proportions of total daily energy with each advancing two-hour time increment. Conclusion The association between the circadian rhythm of eating and body weight status needs to be investigated further to examine the effect of time of consumption on the risk of childhood obesity. Especially longitudinal studies in diverse child populations would help elucidate the importance of time of eating on obesity.

Summary Papillary thyroid carcinoma (PTC) is the most common type of thyroid cancer. Here, we describe the genomic landscape of 496 PTCs. We observed a low frequency of somatic alterations (relative to other carcinomas) and extended the set of known PTC driver alterations to include EIF1AX , PPM1D , and CHEK2 and diverse gene fusions. These discoveries reduced the fraction of PTC cases with unknown oncogenic driver from 25% to 3.5%. Combined analyses of genomic variants, gene expression, and methylation demonstrated that different driver groups lead to different pathologies with distinct signaling and differentiation characteristics. Similarly, we identified distinct molecular subgroups of BRAF -mutant tumors, and multidimensional analyses highlighted a potential involvement of oncomiRs in less-differentiated subgroups. Our results propose a reclassification of thyroid cancers into molecular subtypes that better reflect their underlying signaling and differentiation properties, which has the potential to improve their pathological classification and better inform the management of the disease.