Exosc9 Initiates SUMO-Dependent lncRNA TERRA Degradation to Impact Telomeric Integrity in Endocrine Therapy Insensitive Hormone Receptor-Positive Breast Cancer.

Long, noncoding RNAs (lncRNAs) are indispensable for normal cell physiology and, consequently, are tightly regulated in human cells. Yet, unlike mRNA, substantially less is known about the mechanisms for lncRNA degradation. It is important to delineate the regulatory control of lncRNA degradation, particularly for lncRNA telomeric repeat-containing RNA (TERRA), as the TERRA-telomere R-loops dictate cell cycle progression and genomic stability. We now report that the exosome complex component Exosc9 degrades lncRNA TERRA in human mammary epithelial cells. Heterochromatin protein 1 alpha (HP1α) recruits Exosc9 to the telomeres; specifically, the SUMO-modified form of HP1α supports interaction with Exosc9 and, as previously reported, lncRNA TERRA. The telomeric enrichment of Exosc9 is cell cycle-dependent and consistent with the loss of telomeric TERRA in the S/G2 phase. Elevated Exosc9 is frequently observed and drives the growth of endocrine therapy-resistant (ET-R) HR+ breast cancer (BCa) cells. Specifically, the knockdown of Exosc9 inversely impacts telomeric R-loops and the integrity of the chromosome ends of ET-R cells. Consistently, Exosc9 levels dictate DNA damage and the sensitivity of ET-R BCa cells to PARP inhibitors. In this regard, Exosc9 may serve as a promising biomarker for predicting the response to PARP inhibitors as a targeted monotherapy for ET-R HR+ BCa.

Objective Analysis of Breast Symmetry in Female Patients Undergoing Breast Reconstruction After Total Mastectomy.

Background: Satisfaction with the breast aesthetic outcome is an expectation of breast reconstruction surgery, which is an integral part of cancer treatment for many patients. We evaluated postreconstruction breast symmetry in 82 female patients using distance and volume measurements. Objectives: Clinical factors, such as reconstruction type (implant-based and autologous reconstruction), laterality, timing of reconstruction (immediate, delayed, and sequential), radiation therapy (RT), and demographic factors (age, BMI, race, and ethnicity), were evaluated as predictors of postoperative symmetry. Matched preoperative and postoperative measurements for a subset of 46 patients were used to assess correlation between preoperative and postoperative symmetry. Methods: We used standardized differences between the left and right breasts for the sternal notch to lowest visible point distance and breast volume as metrics for breast, positional symmetry, and volume symmetry, respectively. We performed statistical tests to compare symmetry between subgroups of patients based on reconstruction type, laterality, timing, RT, and demographics. Results: Overall, reconstruction type, reconstruction timing, and RT were observed to be factors significantly associated with postoperative symmetry, with implant reconstructions and immediate reconstruction procedures, and no RT showing better postoperative breast volume symmetry. Subgroup analyses, for both reconstruction type and laterality, showed superior volume symmetry for the bilateral implant reconstructions. No correlation was observed between preoperative and postoperative breast symmetry. Demographic factors were not significant predictors of postreconstruction symmetry. Conclusions: This comprehensive analysis examines multiple clinical factors in a single study and will help both patients and surgeons make informed decisions about reconstruction options at their disposal.

Breast Decisions: Recommender System for Appearance Counseling about Breast Reconstruction.

Appearance counseling is an important component of the consent process for breast reconstruction. The purpose of appearance counseling is to help the patient form realistic expectations of what she might look like after breast reconstruction. In this article, we introduce a recommender system, "BreastDecisions," for appearance counseling that suggests photographs of previous patients that are tailored to a specific patient to help her form realistic expectations of her own reconstruction. Methods: We present user specifications and algorithm parameters needed to incorporate the recommender system into the appearance counseling workflow. We demonstrate the system for a common counseling scenario using a knowledgebase of previous breast reconstruction patients. The medical appropriateness of the recommended photographs for use in appearance counseling was evaluated by experts using a four-point rating system. Results: The recommender system presents photographs that are medically appropriate for counseling a specific patient, depicts typical outcomes, and adapts to a variety of clinical workflows. For each of 33 patients taken as examples of breast reconstruction patients, we used the system to identify photographs for appearance counseling. The baseline average medical appropriateness of the recommended photographs was between mostly appropriate (some explanation needed) and medically appropriate (minimal explanation needed). We demonstrate filtering and ranking steps to reduce the number of recommended photographs and increase the average medical appropriateness. Conclusions: Our recommender system automatically suggests photographs of previous breast reconstruction patients for use in counseling a patient about appearance outcomes. The system is patient-specific and customizable to a particular surgeon's practice.

Potential of Intraoperative 3D Photography and 3D Visualization in Breast Reconstruction.

Although pre- and postoperative three-dimensional (3D) photography are well-established in breast reconstruction, intraoperative 3D photography is not. We demonstrate the process of intraoperative acquisition and visualization of 3D photographs for breast reconstruction and present clinicians' opinions about intraoperative visualization tools. METHODS: Mastectomy specimens were scanned with a handheld 3D scanner during breast surgery. The 3D photographs were processed to compute morphological measurements of the specimen. Three visualization modalities (screen-based viewing, augmented reality viewing, and 3D printed models) were created to show different representations of the 3D photographs to plastic surgeons. We interviewed seven surgeons about the usefulness of the visualization methods. RESULTS: The average time for intraoperative acquisition of 3D photographs of the mastectomy specimen was 4 minutes, 8 seconds ± 44 seconds. The average time for image processing to compute morphological measurements of the specimen was 54.26 ± 40.39 seconds. All of the interviewed surgeons would be more inclined to use intraoperative visualization if it displayed information that they are currently missing (eg, the target shape of the reconstructed breast mound). Additionally, the surgeons preferred high-fidelity visualization tools (such as 3D printing) that are easy-to-use and have minimal disruption to their current workflow. CONCLUSIONS: This study demonstrates that 3D photographs can be collected intraoperatively within acceptable time limits, and quantitative measurements can be computed timely to be utilized within the same procedure. We also report surgeons' comments on usability of visualization methods and of measurements of the mastectomy specimen, which can be used to guide future surgical practice.

Goldilocks Principle: Preference for Change in Breast Size in Breast Cancer Reconstruction Patients.

Patients' preferences regarding changing or maintaining their breast size after mastectomy and reconstruction are important but understudied determinants of post-surgical satisfaction and quality of life. The goal of this study was to identify factors associated with preferences for changing or maintaining breast size for women undergoing breast reconstruction at The University of Texas MD Anderson Cancer Center in the United States from 2011 to 2014. The average age of participants was 45.7 ± 9.1 years. At baseline, mean average breast volumes were 755.7 ± 328.4 mL for all women (n = 48), 492.3 mL ± 209.3 for 13 women who preferred to be "bigger than now," 799.2 mL ± 320.9 for 25 women who preferred to remain "about the same," and 989.3 mL ± 253.1 for 10 women who preferred "smaller than now." Among the 23 women who preferred to change their breast size, 19 desired to shift toward the mean. Women with the smallest and largest 20% of baseline breast size were more likely to desire a change toward the mean (p = 0.006). Multinomial logistic regression models found average breast volume and satisfaction with breast size to be the most important factors associated with preferences for changing or maintaining breast size for women undergoing breast reconstruction. This study provides preliminary evidence for a "Goldilocks principle" in women's preferences for breast size change in the context of breast reconstruction, and identifies hypotheses for future studies of the associations among preference for change in breast size, preference achievement, and post-reconstruction body image.

Review of Quantitative Imaging for Objective Assessment of Fat Grafting Outcomes in Breast Surgery.

This article presents a comprehensive review of the use of quantitative measurements based on breast images to assess fat retention, breast aesthetics, and fat necrosis after autologous fat grafting. Breast volume measured from MRI and 3-dimensional surface images is widely used as a measure of fat retention. Breast aesthetics are assessed from a combination of anthropometric (distance and contour) measurements on breast surface images. Examination of radiologic images (ultrasonography, mammography, and MRI) is utilized to assess fat necrosis. The article discusses implementation guidelines for objective outcome assessment to support robust quantification and enable investigations of fat grafting efficacy. Level of Evidence: 4.

Multifunctional Bio-Nanocomposite Coatings for Perishable Fruits.

Hunger and chronic undernourishment impact over 800 million people, which translates to ≈10.7% of the world's population. While countries are increasingly making efforts to reduce poverty and hunger by pursuing sustainable energy and agricultural practices, a third of the food produced around the globe still is wasted and never consumed. Reducing food shortages is vital in this effort and is often addressed by the development of genetically modified produce or chemical additives and inedible coatings, which create additional health and environmental concerns. Herein, a multifunctional bio-nanocomposite comprised largely of egg-derived polymers and cellulose nanomaterials as a conformal coating onto fresh produce that slows down food decay by retarding ripening, dehydration, and microbial invasion is reported. The coating is edible, washable, and made from readily available inexpensive or waste materials, which makes it a promising economic alternative to commercially available fruit coatings and a solution to combat food wastage that is rampant in the world.

Novel Silicon Titanium Diboride Micropatterned Substrates for Cellular Patterning.

Both hard material photolithography and soft lithography are widely used for patterned cell culture. Soft lithography techniques enable bioactive molecule incorporation, however complex surface modifications are required to introduce specific ligands or proteins in conventional photolithography. In this study, we demonstrate human umbilical vein cell (HUVEC) and adult bone marrow derived mesenchymal stem cell (MSC) patterning on titanium diboride (TiB2) layers deposited on silicon (Si) substrates by electron-beam evaporation and micropatterned using photolithography. Micropatterned cell growth specificity on geometric shapes of circle and/or lines is achieved via differential growth factors adsorption in the presence of heparin. Specifically, the deposited films of TiB2 showed increased stiffness, hardness, hydrophilicity and surface charge when compared to background Si. These substrates were found to be compatible with HUVEC and MSC viability, based on biomarker expression and RNA-sequence transcriptome analysis. Cell-type dependent, micropattern selective cell growth, such as contact guidance, alignment, and durotaxis, were observed. In addition, MSC clustering was achieved, enabling a three-dimensional (3D) aggregate based microenvironment during culture. This study clearly demonstrates the potential of microfabricated Si and TiB2 biomaterials for patterned cell culture in vitro, independent of any additional surface modification.

Natural Breast Symmetry in Preoperative Breast Cancer Patients.

Plastic surgeons aim to achieve breast symmetry during cosmetic and reconstructive breast surgery. They rely on measures of breast size, position, and projection to determine and achieve breast symmetry, but normative data on symmetry in preoperative breast reconstruction patients are scarce. METHODS: A statistical evaluation was performed to examine the relationship of breast symmetry to demographic and clinical factors such as age, body mass index (BMI), race, and cancer status in a sample population of 87 patients who were scheduled to undergo mastectomy and breast reconstruction. The sternal notch to nipple (SN-N) distance and breast volume were measured on three-dimensional images, and distance and volume ratios across the left and right breasts were compared to determine symmetry. Ptosis grades were recorded and grade agreement (match) across the left and right breasts was assessed to determine shape symmetry. RESULTS: A substantial portion of women (41.4%) showed SN-N distance differences >5 mm and 50.6% exhibited a volume difference >50 mL between their right and left breasts. Multiple linear regression modeling did not show any association between age, BMI, and cancer status and the SN-N and volume ratios. Race showed an association with volume symmetry but not with SN-N symmetry. A higher BMI increased the likelihood of ptosis disagreement. Additionally, tumor size did not impact overall breast symmetry. CONCLUSION: This study provides normative data on the extent of breast asymmetry in preoperative patients that can guide physicians in setting realistic goals for reconstruction procedures and manage patients' expectations related to outcomes.

Transcriptional regulation of core autophagy and lysosomal genes by the androgen receptor promotes prostate cancer progression.

AR (androgen receptor) signaling is crucial for the development and maintenance of the prostate as well as the initiation and progression of prostate cancer. Despite the AR's central role in prostate cancer progression, it is still unclear which AR-mediated processes drive the disease. Here, we identified 4 core autophagy genes: ATG4B, ATG4D, ULK1, and ULK2, in addition to the transcription factor TFEB, a master regulator of lysosomal biogenesis and function, as transcriptional targets of AR in prostate cancer. These findings were significant in light of our recent observation that androgens promoted prostate cancer cell growth in part through the induction of autophagy. Expression of these 5 genes was essential for maximal androgen-mediated autophagy and cell proliferation. In addition, expression of each of these 5 genes alone or in combination was sufficient to increase prostate cancer cell growth independent of AR activity. Further, bioinformatic analysis demonstrated that the expression of these genes correlated with disease progression in 3 separate clinical cohorts. Collectively, these findings demonstrate a functional role for increased autophagy in prostate cancer progression, provide a mechanism for how autophagy is augmented, and highlight the potential of targeting this process for the treatment of advanced prostate cancer.

Identification of vascular disruptor compounds by analysis in zebrafish embryos and mouse embryonic endothelial cells.

To identify vascular disruptor compounds (VDCs), this study utilized an in vivo zebrafish embryo vascular model in conjunction with a mouse endothelial cell model to screen a subset of the U.S. Environmental Protection Agency (EPA) ToxCast Phase I chemical inventory. In zebrafish, 161 compounds were screened and 34 were identified by visual inspection as VDCs, of which 28 were confirmed as VDCs by quantitative image analysis. Testing of the zebrafish VDCs for their capacity to inhibit endothelial tube formation in the murine yolk-sac-derived endothelial cell line C166 identified 22 compounds that both disrupted zebrafish vascular development and murine endothelial in vitro tubulogenesis. Putative molecular targets for the VDCs were predicted using EPA's Toxicological Prioritization Index tool and a VDC signature based on a proposed adverse outcome pathway for developmental vascular toxicity. In conclusion, our screening approach identified 22 novel VDCs, some of which were active at nanomolar concentrations.

Computation of breast ptosis from 3D surface scans of the female torso.

Stereophotography is now finding a niche in clinical breast surgery, and several methods for quantitatively measuring breast morphology from 3D surface images have been developed. Breast ptosis (sagging of the breast), which refers to the extent by which the nipple is lower than the inframammary fold (the contour along which the inferior part of the breast attaches to the chest wall), is an important morphological parameter that is frequently used for assessing the outcome of breast surgery. This study presents a novel algorithm that utilizes three-dimensional (3D) features such as surface curvature and orientation for the assessment of breast ptosis from 3D scans of the female torso. The performance of the computational approach proposed was compared against the consensus of manual ptosis ratings by nine plastic surgeons, and that of current 2D photogrammetric methods. Compared to the 2D methods, the average accuracy for 3D features was ~13% higher, with an increase in precision, recall, and F-score of 37%, 29%, and 33%, respectively. The computational approach proposed provides an improved and unbiased objective method for rating ptosis when compared to qualitative visualization by observers, and distance based 2D photogrammetry approaches.

Inferior Breast-Chest Contour Detection in 3-D Images of the Female Torso.

Stereophotogrammetry is finding increased use in clinical breast surgery, both for breast reconstruction after oncological procedures and cosmetic augmentation and reduction. The ability to visualize and quantify morphological features of the breast facilitates pre-operative planning and post-operative outcome assessment. The contour outlining the lower half of the breast is important for the quantitative assessment of breast aesthetics. Based on this inferior breast contour, relevant morphological measures, such as breast symmetry, volume, and ptosis, can be determined. In this paper, we present an approach for automatically detecting the inferior contour of the breast in 3D images. Our approach employs surface curvature analysis and is able to detect the breast contour with high accuracy, achieving an average error of 1.64 mm and a dice coefficient in the range of 0.72-0.87 when compared with the manually annotated contour (ground truth). In addition, the detected contour is used to facilitate the detection of the lowest visible point on the breast, which is an important landmark for breast morphometric analysis.

Embryonic exposure to sodium arsenite perturbs vascular development in zebrafish.

Exposure to arsenic in its inorganic form, arsenite, causes adverse effects to many different organs and tissues. Here, we have investigated arsenite-induced adverse effects on vascular tissues in the model organism zebrafish, Danio rerio. Zebrafish embryos were exposed to arsenite at different exposure windows and the susceptibility to vascular tissue damage was recorded at 72hours post fertilization (hpf). Intersegmental vessel sprouting and growth was most perturbed by exposure to arsenite during the 24-48hpf window, while disruption in the condensation of the caudal vein plexus was more often observed at the 48-72hpf exposure window, reflecting when these structures develop during normal embryogenesis. The vascular growth rate was decreased by arsenite exposure, and deviated from that of control embryos at around 24-26.5hpf. We further mapped changes in expression of key regulators of angiogenesis and vasculogenesis. Downregulation of vascular endothelial growth factor receptor 1/fms-related tyrosine kinase 1 (vegfr1/flt1) expression was evident already at 24hpf, coinciding with the decreased vascular growth rate. At later time points, matrix metalloproteinase 9 (mmp9) expression was upregulated, suggesting that arsenite affects the composition of the extracellular matrix. In total, the expression of eight key factors involved in different aspects of vascularization was significantly altered by arsenic exposure. In conclusion, our results show that arsenite is a potent vascular disruptor in the developing zebrafish embryo, a finding that calls for an evaluation of arsenite as a developmental vascular toxicant in mammalian model systems.

Automated analysis of zebrafish images for screening toxicants.

An important factor facilitating the application of zebrafish in biomedical research is high throughput screening of vertebrate animal models. For example, being able to model the growth of blood vessel in the vasculature system is interesting for understanding both the circulatory system in humans, and for facilitating large scale screening of the influence of various chemicals on vascular development. Compared to other models, the zebrafish embryo is an attractive alternative for environmental risk assessment of chemicals since it offers the possibility to perform high-throughput analyses in vivo. However the lack of an automated image analysis framework restricts high throughput screening. In this paper, we provide a method for quantitative measurements of zebrafish blood vessel morphology since it is difficult to assess changes in vessel structure by visual inspection. The method presented is generalized, i.e. it is not restricted to any specific chemically treated zebrafish, and can be used with wide variety of chemicals.

Combinational targeting offsets antigen escape and enhances effector functions of adoptively transferred T cells in glioblastoma.

Preclinical and early clinical studies have demonstrated that chimeric antigen receptor (CAR)-redirected T cells are highly promising in cancer therapy. We observed that targeting HER2 in a glioblastoma (GBM) cell line results in the emergence of HER2-null tumor cells that maintain the expression of nontargeted tumor-associated antigens. Combinational targeting of these tumor-associated antigens could therefore offset this escape mechanism. We studied the single-cell coexpression patterns of HER2, IL-13Rα2, and EphA2 in primary GBM samples using multicolor flow cytometry and immunofluorescence, and applied a binomial routine to the permutations of antigen expression and the related odds of complete tumor elimination. This mathematical model demonstrated that cotargeting HER2 and IL-13Rα2 could maximally expand the therapeutic reach of the T cell product in all primary tumors studied. Targeting a third antigen did not predict an added advantage in the tumor cohort studied. We therefore generated bispecific T cell products from healthy donors and from GBM patients by pooling T cells individually expressing HER2 and IL-13Rα2-specific CARs and by making individual T cells to coexpress both molecules. Both HER2/IL-13Rα2-bispecific T cell products offset antigen escape, producing enhanced effector activity in vitro immunoassays (against autologous glioma cells in the case of GBM patient products) and in an orthotopic xenogeneic murine model. Further, T cells coexpressing HER2 and IL-13Rα2-CARs exhibited accentuated yet antigen-dependent downstream signaling and a particularly enhanced antitumor activity.

Androgens promote prostate cancer cell growth through induction of autophagy.

Androgens regulate both the physiological development of the prostate and the pathology of prostatic diseases. However, the mechanisms by which androgens exert their regulatory activities on these processes are poorly understood. In this study, we have determined that androgens regulate overall cell metabolism and cell growth, in part, by increasing autophagy in prostate cancer cells. Importantly, inhibition of autophagy using either pharmacological or molecular inhibitors significantly abrogated androgen-induced prostate cancer cell growth. Mechanistically, androgen-mediated autophagy appears to promote cell growth by augmenting intracellular lipid accumulation, an effect previously demonstrated to be necessary for prostate cancer cell growth. Further, autophagy and subsequent cell growth is potentiated, in part, by androgen-mediated increases in reactive oxygen species. These findings demonstrate a role for increased fat metabolism and autophagy in prostatic neoplasias and highlight the potential of targeting underexplored metabolic pathways for the development of novel therapeutics.

Automated Identification of Fiducial Points on 3D Torso Images.

Breast reconstruction is an important part of the breast cancer treatment process for many women. Recently, 2D and 3D images have been used by plastic surgeons for evaluating surgical outcomes. Distances between different fiducial points are frequently used as quantitative measures for characterizing breast morphology. Fiducial points can be directly marked on subjects for direct anthropometry, or can be manually marked on images. This paper introduces novel algorithms to automate the identification of fiducial points in 3D images. Automating the process will make measurements of breast morphology more reliable, reducing the inter- and intra-observer bias. Algorithms to identify three fiducial points, the nipples, sternal notch, and umbilicus, are described. The algorithms used for localization of these fiducial points are formulated using a combination of surface curvature and 2D color information. Comparison of the 3D co-ordinates of automatically detected fiducial points and those identified manually, and geodesic distances between the fiducial points are used to validate algorithm performance. The algorithms reliably identified the location of all three of the fiducial points. We dedicate this article to our late colleague and friend, Dr. Elisabeth K. Beahm. Elisabeth was both a talented plastic surgeon and physician-scientist; we deeply miss her insight and her fellowship.

3D Symmetry Measure Invariant to Subject Pose During Image Acquisition.

In this study we evaluate the influence of subject pose during image acquisition on quantitative analysis of breast morphology. Three (3D) and two-dimensional (2D) images of the torso of 12 female subjects in two different poses; (1) hands-on-hip (HH) and (2) hands-down (HD) were obtained. In order to quantify the effect of pose, we introduce a new measure; the 3D pBRA (Percentage Breast Retraction Assessment) index, and validate its use against the 2D pBRA index. Our data suggests that the 3D pBRA index is linearly correlated with the 2D counterpart for both of the poses, and is independent of the localization of fiducial points within a tolerance limit of 7 mm. The quantitative assessment of 3D asymmetry was found to be invariant of subject pose. This study further corroborates the advantages of 3D stereophotogrammetry over 2D photography. Problems with pose that are inherent in 2D photographs are avoided and fiducial point identification is made easier by being able to panoramically rotate the 3D surface enabling views from any desired angle.