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Background/Objective Breast biopsy and analysis of sentinel lymph nodes (SLNs) accurately predict tumor status in the affected basin and help in avoiding unnecessary axillary lymph node dissection, which is associated with remarkable morbidity risk. Blue dye and radioisotope are the most widely used mapping agents, but non-radioactive tracers of comparable accuracy warrant further investigation. This study aimed to investigate utilization of indocyanine green (ICG) fluorescence in sentinel node localization compared with blue dye and to assess the incremental value of ICG. Methods A total of 39 consecutive patients underwent sentinel lymph node biopsy (SLNB) (40 cases: 38 unilateral and 1 bilateral) with combined blue dye and ICG for localization. The obtained fluorescence images of the lymphatic system were investigated. Results All 84 lymph nodes removed in 40 procedures were identified by ICG, but only 37 were identified by blue dye. The ICG method identified an average of 2.1 SLNs in 39 of 40 cases with a detection rate of 97.5%, but only 0.93 SLN per case with blue dye. Subcutaneous lymphatic channel patterns were also detected by fluorescent imaging in 37 procedures, which all revealed lymphatic drainage toward the axilla except in one case with internal mammary pathway. Conclusion This study demonstrated the accuracy and safety of ICG for SLNB and its superiority to blue dye method in SLN localization. Therefore, ICG fluorescence method is safe and effective addition in breast clinical settings, wherein blue dye alone is used.

Elevation of naphthoquinones and estrogen quinones, which are reactive metabolites of naphthalene and estrogen, is thought to be an important indicator of naphthalene- and estrogen-induced carcinogenesis. We compared background levels of naphthalene and estrogen quinone-derived adducts in serum albumin (Alb) from 143 women with breast cancer and 119 healthy controls. Cysteinyl adducts of naphthoquinones, including 1,2-naphthoquinone (1,2-NPQ) and 1,4-naphthoquinone (1,4-NPQ), and estrogen quinones, including estrogen-2,3-quinones (E2-2,3-Q) and estrogen-3,4-quinones (E2-3,4-Q), were characterized after adduct cleavage. Levels of estrogen quinones and naphthoquinones were positively correlated in healthy controls, but not in breast cancer patients (p 2-3,4-Q were elevated by two- to ten-fold in cancer patients (p 2-3,4-Q-2-S-Alb to (E2-2,3-Q-4-S-Alb plus E2-3,4-Q-2-S-Alb) in patients and controls. These two groups were separable using albumin adducts of estrogen quinones and naphthoquinones, with 99.6% overall correct classification rate (overall accuracy). The findings of this study suggest that differences in the disposition of estrogen and naphthalene, and the subsequent elevation of cumulative E2-3,4-Q and 1,2-NPQ may serve as biomarkers of breast cancer risk.

Background Vitiligo is an acquired disease that involves low pigment variation in the skin. The use of cultured melanocytes for treatment of recalcitrant vitiligo has become a well-established treatment modality. In vitro cultured melanocytes present an effective autologous transplantation treatment modality for vitiligo. The present study investigated the in vitro culture of epidermal melanocytes sampled from multiple body parts, as well as the differences in total propagation time. Methods Skin specimens were collected from 30 participants (14 males and 16 females) who supplied normal colored skin sections from various regions of their body, including the face, chest, abdomen, buttock, and extremities. Subsequently, all of these specimens were treated with an identical melanocyte purification and culturing process. Results Melanocytes from the face demonstrated the most rapid growth and longest total propagation time. Melanocytes from the buttock, abdomen, and extremities displayed similar results to one another, and melanocytes from the chest and back had the slowest growth and shortest total propagation time. Conclusion Selecting the most favorable site to obtain epidermal melanocytes will reduce the required quantity of skin and culturing time, and maximize the growth and total propagation time of melanocytes. Therefore, care should be exercised when selecting the region of skin when culturing epidermal melanocytes.

Tamoxifen is one of the most common hormone therapy drug for estrogen receptor (ER)-positive breast cancer. Tumor cells with drug resistance often cause recurrence and metastasis in cancer patients. Luteolin is a natural compound found from various types of vegetables and exhibit anticancer activity in different cancers. This study demonstrated that luteolin inhibits the proliferation and induces apoptosis of tamoxifen-resistant ER-positive breast cancer cells. Luteolin also causes cell cycle arrest at the G2/M phase and decreases mitochondrial membrane potential. Besides, luteolin reduces the levels of activated PI3K/AKT/mTOR signaling pathway. The combination treatment of luteolin and PI3K, AKT, or mTOR inhibitors synergistically increases apoptosis in tamoxifen-resistant ER-positive breast cancer cells. Ras gene family (K-Ras, H-Ras, and N-Ras), an activator of PI3K, was transcriptionally repressed by luteolin via induction of tumor suppressor mixed-lineage leukemia 3 (MLL3) expression. MLL3 increases the level of monomethylation of Histone 3 Lysine 4 on the enhancer and promoter region of Ras genes, thus causes repression of Ras expressions. Our finding implies that luteolin was a promising natural agent against tamoxifen resistance of breast cancer.

Objectives Flow cytometry (FC) is critical for the diagnosis and monitoring of hematologic malignancies. Machine learning (ML) methods rapidly classify multidimensional data and should dramatically improve the efficiency of FC data analysis. We aimed to build a model to classify acute leukemias, including acute promyelocytic leukemia (APL), and distinguish them from nonneoplastic cytopenias. We also sought to illustrate a method to identify key FC parameters that contribute to the model’s performance. Methods Using data from 531 patients who underwent evaluation for cytopenias and/or acute leukemia, we developed an ML model to rapidly distinguish among APL, acute myeloid leukemia/not APL, acute lymphoblastic leukemia, and nonneoplastic cytopenias. Unsupervised learning using gaussian mixture model and Fisher kernel methods were applied to FC listmode data, followed by supervised support vector machine classification. Results High accuracy (ACC, 94.2%; area under the curve [AUC], 99.5%) was achieved based on the 37-parameter FC panel. Using only 3 parameters, however, yielded similar performance (ACC, 91.7%; AUC, 98.3%) and highlighted the significant contribution of light scatter properties. Conclusions Our findings underscore the potential for ML to automatically identify and prioritize FC specimens that have critical results, including APL and other acute leukemias.

Melanoma is the cause of most deaths from skin cancer. The extracellular signal-regulated kinase 1/2 (ERK1/2) pathway has been reported to participate in progression of melanoma in fair skinned populations. ERK1/2 is found in both the cytoplasm and nucleus of cells, and phosphorylated ERK1/2 has been implicated in tumor progression. We investigated the relation between melanoma progression and expression of cytoplasmic and nuclear phosphorylated ERK1/2. We examined 34 surgically resected melanomas and investigated their clinicopathologic characteristics. We found immunostaining of phosphorylated ERK1/2 in all melanomas and faint staining in benign nevi. We found expression of cytoplasmic phosphorylated ERK1/2 in most melanomas; however, nuclear phosphorylated ERK1/2 expression was found in only five melanomas. Expression of cytoplasmic phosphorylated ERK1/2 was related to the tumor stage in melanoma. Nine of 10 cases of distant metastasis were positive for cytoplasmic phosphorylated ERK1/2. Our findings suggest that phosphorylated ERK1/2 expression is relevant to clinical pathology and that in melanoma patients, phosphorylated ERK1/2 expression is found in both the cytoplasm and nucleus. Our findings suggest that cytoplasmic phosphorylated ERK1/2 participates in progression of melanoma and that it could be a useful target for clinical treatment of melanoma.

Acute leukemia often comes with life-threatening prognosis outcome and remains a critical clinical issue today. The implementation of measurable residual disease (MRD) using flow cytometry (FC) is highly effective but the interpretation is time-consuming and suffers from physician idiosyncrasy. Recent machine learning algorithms have been proposed to automatically classify acute leukemia samples with and without MRD to address this clinical need. However, most prior works either validate only on a small data cohort or focus on one specific type of leukemia which lacks generalization. In this work, we propose a transfer learning approach in performing automatic MRD classification that takes advantage of a large scale acute myeloid leukemia (AML) database to facilitate better learning on a small cohort of acute lymphoblastic leukemia (ALL). Specifically, we develop a knowledge-reserved distilled AML pre-trained network with ALL complementary learning to enhance the ALL MRD classification. Our framework achieves 84.5% averaged AUC which shows its transferability across acute leukemia, and our further analysis reveals that younger and elder ALL patient samples benefit more from using the pre-trained AML model.

Background The aim of the current study was to report a single-institution experience using breast-conserving surgery after neoadjuvant chemotherapy (NACT), focusing on the association between microscopic resection margin status and locoregional recurrence (LRR). Methods Our institutional prospectively maintained database was reviewed to identify patients who were treated with NACT between January 2008 and April 2018. Results Among the main partial mastectomy specimens available for analysis (n = 161), 28 had margins 2 mm. LRR occurred in 16 patients (9.9%) and distant metastases were detected in 27 (16.8%) patients. There was no significant difference in the LRR between the > 2 mm margin group with a 60-month cumulative survival of 85.2% compared with 76.2% for the ≤2 mm group (P = 0.335) in the Kaplan-Meier analysis. When we stratified patients by margin widths of ≥1 mm or P = 0.161) with a 60-month cumulative LRR-free survival of 84.9% compared with 69.5% for the P = 0.150). Conclusions In the absence of multiple scattered microscopic tumour foci, a negative margin of no ink on tumour maybe sufficient for stage I–III invasive breast cancer treated with NACT and breast-conserving surgery.

Identification of minimal residual disease (MRD) is important in assessing the prognosis of acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). The current best clinical practice relies heavily on Flow Cytometry (FC) examination. However, the current FC diagnostic examination requires trained physicians to perform lengthy manual interpretation on high-dimensional FC data measurements of each specimen. The difficulty in handling idiosyncrasy between interpreters along with the time-consuming diagnostic process has become one of the major bottlenecks in advancing the treatment of hematological diseases. In this work, we develop an automatic MRD classifications (AML, MDS, normal) algorithm based on learning a deep phenotype representation from a large cohort of retrospective clinical data with over 2000 real patients’ FC samples. We propose to learn a cytometric deep embedding through cell-level autoencoder combined with specimen-level latent Fisher-scoring vectorization. Our method achieves an average AUC of 0.943 across four different hematological malignancies classification tasks, and our analysis further reveals that with only half of the FC markers would be sufficient in obtaining these high recognition accuracies.

The prognosis management is crucial for highrisk disease like Acute Myeloid Leukemia (AML) in order to support decisions of clinical treatment. However, the challenges of accurate and consistent forecasting lie in the high variability of the disease outcomes and the complexity of the multiple clinical measurements available over the course of the treatment. In order to capture the multi-dimensional and longitudinal aspect of these comprehensive clinical parameters, we utilize an attention-based bi-directional long shortterm memory (Att-BLSTM) network to predict AML patient's survival and relapse. Specifically, we gather a 10-year worth of real patient's clinical data including blood test, medication, HSCT status, and gene mutation information. Our proposed Att-BLSTM framework achieves 77.1% and 67.3% AUC in tasks of predicting the next 2-year mortality and disease relapse with these comprehensive clinical parameters, and our further analysis demonstrates that a next 0 to 3 months prediction performs equally well, i.e., 74.8% and 67% AUC for mortality and relapse respectively.

BACKGROUND Hypoxic-ischemic brain injury in the perinatal period is a main cause of perinatal mortality and neurologic complications in neonates and children. Recent studies have focused on the neuroprotective effect of anesthetic drugs. The volatile anesthetic isoflurane has been shown to exert neuroprotective effects in cerebral ischemia. Mangiferin is a natural polyphenol with various pharmacological properties, including antioxidant and ant-tumor effects. This study aimed to determine whether mangiferin potentiates the neuroprotective effects of isoflurane and also if mangiferin when administered alone exerts neuroprotective effects following hypoxic-ischemic brain injury. MATERIAL AND METHODS Sprague-Dawley rats were subjected to cerebral hypoxic ischemia on postnatal day 10 (P10). Mangiferin (50, 100, or 200 mg/kg b.w.) was intragastrically administered from P3 to P12 and 1 h prior to insult on the day of ischemic induction. At 3 h after hypoxia-ischemia (HI) insult, separate groups of rat pups were exposed to isoflurane (1.5%) for 6 h. Following 48 h of HI, the rats were sacrificed and brain tissues were used for analysis. RESULTS Mangiferin treatment attenuated neuronal apoptosis and reduced cerebral infarct volume. The expression of cleaved caspase-3 and apoptotic cascade proteins were regulated. The levels of reactive oxygen species (ROS) and malondialdehyde were reduced by mangiferin and/or isoflurane exposure. The levels of antioxidant glutathione were considerably raised under HI injury, which was modulated by mangiferin and isoflurane exposure. The PI3K/Akt signaling pathway, which was downregulated following HI insult, was activated by mangiferin and/or isoflurane. CONCLUSIONS This study reveals the potent neuroprotective efficacy of mangiferin against HI-induced brain injury via effectively modulating apoptotic pathways, ROS levels, and PI3K/Akt cascades while potentiating protective effects of isoflurane.

Background and purposeEngorged medullary vein (EMV) in patients with intracranial dural arteriovenous fistula (DAVF) suggests venous congestion. The aim of this study is to investigate its prevalence, pattern, and correlation with clinical findings.Materials and methodsCT angiography (CTA) raw data of DAVF were used for multiplanar reconstruction and then analyzed for the presence and pattern of EMV, which is defined as a dilated vein in the cerebral white matter. Patients with EMV were divided into two groups: regional and extensive. Regional type is defined as EMV limited to one cerebral hemisphere or cerebellum without evidence of subcortical calcification. Extensive type is defined as EMV involvement of more than one cerebral hemisphere or both the cerebrum and cerebellum. Descriptive analysis of clinical information, DAVF characteristics, and other imaging findings was conducted. Clinical information, including demographic data, clinical presentation, and hemorrhage, were correlated with both types of EMV.ResultsAmong 192 eligible patients with DAVF, 71 (37%) had EMV. Patients with EMV were older (63 years vs 56 years, P=0.02), with DAVF more often at the transverse and sigmoid sinus (PConclusionsEMV in patients with DAVF is associated with an aggressive manifestation. Regional type EMV is associated with a higher risk of a hemorrhagic presentation.

Aloe-emodin (AE) is derived from Aloe vera and rhubarb (Rheum palmatum) and exhibits anticancer activities via multiple regulatory mechanisms in various cancers. AE can also enhance the anticancer efficacy of cisplatin, doxorubicin, docetaxel, and 5-fluorouracil; however, its effects remain poorly characterized. MCF-7, MDA-MB-231, MDA-MB-468, BT-474, and HCC-1954 breast cancer cell lines were treated with the indicated conditions of AE, and cell viability assays were performed. The expression levels of signaling proteins were determined by western blot analysis, intracellular reactive oxygen species (ROS), cell cycle distributions, and rates of apoptosis as estimated by flow cytometry. In comparison with other cells, MCF-7 cells were more sensitive to AE treatment; AE enhanced the cytotoxicity of 9[Formula: see text][Formula: see text]g/ml tamoxifen by reducing EGFR, ER[Formula: see text], Ras, ERK, c-Myc, and mTOR protein expression and blocking PI3K and mTOR activation. Finally, although co-treatment of AE with tamoxifen increased intracellular ROS, there were no effects on cell cycle progression. Besides facilitating tamoxifen-induced cell death, AE also enhanced the antiproliferative activity of tamoxifen by blocking Ras/ERK and PI3K/mTOR pathways in breast cancer cells, thus demonstrating the chemosensitizing potential of AE.

Introduction Identification of gene mutation status prior treatment has improved our capability in risk stratifying acute myeloid leukemia (AML) patients greatly, but it is really a labor-exhausting work to identify these mutations. In this this study, we hypothesize immunophenotyping could predict gene mutation and we aim to develop machine learning algorithms that could predict the AML gene mutation status with the immunophenotype by clinical flow cytometry. Method Retrospective clinical data of patients with AML, including demographic (age & gender), molecular genetics, cytogenetics as well as flow cytometry (FC) data at National Taiwan University Hospital were collected. A total of 529 newly diagnosed de novo AML from 2009 to 2019 enrolled this study. The median age at diagnosis was 58 years (Table 1). In total, 428 NPM1, 415 FLT3, 331 CEBPA and 338 RUNX1 gene testing results and a total of 529 initial diagnostic FC data from these patients were used in developing the gene mutation prediction models. Each FC data sample contained 100,000 cells acquired on FACSCantoII machine 6 fluorescent channels with multiple fluorescent markers. The markers measured are listed in Table 2. There were 19 combinations of markers and fluorescent channels which were served as feature inputs of the machine learning framework. Our proposed machine learning framework can be divided as a phenotype representation learning paradigm and a classification model. To derive the phenotype representation, we trained a multivariate Gaussian Mixture Model (GMM) on the 19-dimension FC data to capture the training data distribution and characteristics in a probabilistic unsupervised manner. Then, a Fisher-scoring method was used to vectorize each sample as a high dimensional representation via differential computation in terms of the learned GMM parameters. This Fisher vectorization method transformed samples to a high dimensional feature space as phenotype vectors. We performed analysis of variance (ANOVA)-based feature selection on these representations which were finally fed into the support vector machine (SVM) classifier. To alleviate the negative effects of imbalance classes in gene mutation identification tasks, we applied synthetic minority oversampling technique (SMOTE) algorithm which augmented the minority class by interpolating samples near support vectors. We train independent SVM models to detect the occurrences of the four gene mutation. The algorithm is evaluated by randomly divided 5-fold cross validation which separates 80% data for training and 20% for testing. Results This gene mutation rate of this cohort for NPM1, FLT3, CEBPA and RUNX1 were 22.2% (95/428), 25.1% (104/415), 20.2% (67/331), and 13.6% (46/338), respectively. The average accuracies (ACC) of the prediction model performance for NPM1, FLT3, CEBPA and RUNX1 were 82.6%, 76.3%, 84.2% and 84.1%, respectively, whereas the area under the ROC curve (AUC) were 77.9%, 63.4%, 80.7% and 67.7%, respectively (Table. 3). Conclusions We demonstrated the potential of the correlation of recurrent AML gene mutation status with immunophenotype of AML through our preliminary gene mutation prediction model. Further study with larger cohorts followed by external validation are needed to further evaluate the feasibility of using machine learning based algorithm as one of triage tools to support physicians in aggressive AML clinical decision before receiving molecular genetic reports. Disclosures Ko: Roche: Honoraria.

Background Proper patient selection criteria for treatment of carotid chronic total occlusion (CTO) are unclear. This study was designed to predict procedure successful rate and 1-year patency after carotid artery stenting (CAS) for carotid CTO using pre-procedural CTA. Methods Patients with CTO detected on CTA who underwent recanalization within 3months were divided into those with occlusions at (or distal to) the clinoid segment of the internal carotid artery (group A) and those with occlusions proximal to the clinoid segment (group B) and outcomes were compared between groups. Results Technical success rates, major complications, and re-occlusions within 1-year were 52%, 22%, 91% in group A (N=23), and 89%, 0%, 0% in group B (N=19), respectively. Diabetes was more frequent in group A (43%) compared with group B (11%). Conclusion CTA may play a role in predicting successful rate and 1-year patency for endovascular recanalization in carotid CTO.

Synthetic quartz glass is an excellent transmittance material in ultraviolet (UV), visible and near-infrared bands. Homogenization process to synthetic quartz glass optical elements is done in order to make the composition and structure more uniform. The homogenization temperature is above 1800°C. After high-temperature homogenization, the UV transmittance of quartz glass appears a certain extent of decline in the edge partition., After high temperature homogenization process, it is found that the closer the layer to the graphite vessel, the stronger influence on UV transmittance will be. This conclusion is made by comparing the UV transmission of the quartz glass in different layers. Furthermore, the factors affecting the UV transmittance is also analyzed in this article.

Introduction Current risk stratification of Acute Lymphoblastic Leukemia (ALL) approach has significantly improve the outcome for pediatric and adolescent ALL. However, the outcome for adult ALL remains poor. With the emerging technologies that could drastically increase the parameters and scales of clinical data for treatment strategy formulation and more new therapeutic agents approved for ALL treatment, a better integrated and more systematic risk stratification method is needed for ALL. In our study, we propose to use artificial intelligence to develop an integral outcome prediction models that include all clinical exam and treatment history, aiming to further improve current risk stratification strategies. Method Retrospective clinical data of patients with ALL, including demographic (age & gender), laboratory results including complete blood count (CBC), white blood count (WBC), molecular genetics, cytogenetics, flow cytometry (FC) and treatment history at National Taiwan University Hospital were collected. A total of 513 newly diagnosed ALL from 2005 to 2017 were enrolled this study. The median age at diagnosis was 20 years and the median follow up duration was 49.1 months, 78.9% of them achieved complete remission (CR), 27.5% had relapsed, 31.6% had received HSCT (Table 1). In total, 243,464 CBC & WBC records, 32 types of anti-neoplastic medications in L01, L03 and L04 ATC code category with a total of 20,043 medication records from these 513 patients were utilized in developing the ALL outcome prediction model. A vectorized representation that captures the static-dynamic clinical aspects of an ALL patient can be learned directly from the collected data. The representation includes both static personal attributes (demographic, and genetic) and time-varying progression of patient's clinical assessment across time (laboratory results, HSCT, and medication). The time-dependent representation was derived from training a deep network architecture of bi-directional long short-term memory network (BLSTM). By taking 7 days as a time step, the BLSTM took the input of Fisher-vector encoded time series of a patient's CBC and WBC, medication, and HSCT records separately. The last output layer, which summarized the relapse/mortality risk exhibited in the measurements of patient's clinical conditions over time, of each separately-learned BLSTM was used as the encoded dynamic clinical representation. The concatenation of the ALL patient's static features and the time-dependent representations were fed into a deep neural network followed by a support vector machine to carry out the final prediction. The prediction models were conducted in 5-fold cross-validation experiments and further evaluated using metrics of accuracy (ACC). Results The median leukemia free survival and median overall survival of these 513 ALL patients are 29.3 months and 61.9 months respectively. The outcome prediction models developed achieved accuracy of next 3-month relapse, relapse or mortality, and mortality prediction reached 86.2%, 74.9% and 64.3% respectively (Table 2). Conclusions Our team has demonstrated the BLSTM-DNN model's potential that is capable in taking multiple and longitudinal clinical measurements into integrated relapse or mortality prediction in AML in 2018 (ASH 2018 #2811) and we have observed the same potential in ALL relapse and mortality prediction. More external and multi-national center studies are needed to further improve the performance of such integrated outcome prediction models. Disclosures Ko: Roche: Honoraria.

Cumulative estrogen concentration is an important determinant of the risk of developing breast cancer. Estrogen carcinogenesis is attributed to the combination of receptor-driven mitogenesis and DNA damage induced by quinonoid metabolites of estrogen. The present study was focused on developing an improved breast cancer prediction model using estrogen quinone-protein adduct concentrations. Blood samples from 152 breast cancer patients and 71 healthy women were collected, and albumin (Alb) and hemoglobin (Hb) adducts of estrogen-3,4-quinone and estrogen-2,3-quinone were extracted and evaluated as potential biomarkers of breast cancer. A multilayer perceptron (MLP) was used as the predictor model and the resultant prediction of breast cancer was more accurate than other existing detection methods. A MLP using the logarithm of the concentrations of the estrogen quinone-derived adducts (four input nodes, 10 hidden nodes, and one output node) was used to predict breast cancer risk with accuracy close to 100% and area under curve (AUC) close to one. The AUC value of one showed that both data sets were separable. We conclude that Alb and Hb adducts of estrogen quinones are promising biomarkers for the early detection of breast cancer.

Breast cancer development and progression are influenced by insulin-like growth factor receptor 1 (IGF1R) and insulin receptor (InsR) signaling, which drive cancer phenotypes such as cell growth, proliferation, and migration. IGF1R and InsR form IGF1R/InsR hybrid receptors (HybRs) consisting of one molecule of IGF1R and one molecule of InsR. The specific signaling and functions of HybR are largely unknown, as HybR is activated by both IGF1 and insulin, and no cellular system expresses HybR in the absence of holo-IGF1R or holo-InsR. Here we studied the role of HybR by constructing inducible chimeric receptors and compared HybR signaling with that of holo-IGF1R and holo-InsR. We cloned chemically inducible chimeric IGF1R and InsR constructs consisting of the extracellular domains of the p75 nerve growth factor receptor fused to the intracellular β subunit of IGF1R or InsR and a dimerization domain. Dimerization with the drugs AP20187 or AP21967 allowed specific and independent activation of holo-IGF1R, holo-InsR, or HybR, resulting in activation of the PI3K pathway. Holo-IGF1R and HybR both promoted cell proliferation and glucose uptake, whereas holo-InsR only promoted glucose uptake, and only holo-IGF1R showed anti-apoptotic effects. We also found that the three receptors differentially regulated gene expression: holo-IGF1R and HybR up-regulated EGR3; holo-InsR specifically down-regulated JUN and BCL2L1; holo-InsR down-regulated but HybR up-regulated HK2; and HybR specifically up-regulated FHL2, ITGA6, and PCK2. Our findings suggest that, when expressed and activated in mammary epithelial cells, HybR acts in a manner similar to IGF1R and support further investigation of the role of HybR in breast cancer.

Background Magnetic resonance (MR) enterography has emerged as a new imaging modality in evaluating patients with Crohn's disease. However, whether interpretations of MR enterography findings are consistent among radiologists has not yet been fully investigated. The purpose of this study is to evaluate the intraobserver and interobserver agreement for identifying extraluminal manifestations of Crohn's disease with MR enterography. Methods MR enterography was performed with a 3-tesla system. We retrospectively enrolled patients with Crohn's disease that had MR enterography between November 2011 and March 2013. Three radiologists reviewed the images independently. Extraluminal findings of these patients were recorded and described. Intraobserver and interobserver agreement were calculated using the kappa statistic. Results A total of 22 sessions of MR enterography were performed on 15 patients (12 men and 3 women) with Crohn's disease. The mean age was 38.7 ± 11.4 years. We identified extraluminal manifestations such as engorged vasa recta, enlarged mesenteric lymph nodes, fibrofatty proliferation, fistulas, and abscesses. In addition, avascular necrosis of the femoral head as a complication of long-term steroid treatment was also depicted. The intraobserver agreement was almost perfect (κ = 0.83; p

Introduction The development of complex renal cysts after crizotinib treatment for non–small-cell lung cancer (NSCLC) is a reported side effect. However, its occurrence and characteristics have not been reported. Methods Medical records and computed tomography images of crizotinib-treated patients in three prospective clinical trials were reviewed. The size and Bosniak category of the renal cysts before and after crizotinib treatment were determined. Patients' clinical characteristics, tumor stage, treatment response, renal function, and outcomes were analyzed. Results During December 2010 to March 2013, we enrolled 32 patients who received crizotinib. There were 23 patients who had renal cysts before crizotinib. The median follow-up time was 493 days. Seven patients (22%, six with baseline renal cyst and one without baseline renal cyst) had significant renal cyst change. Four (13% of all) had new complex renal cysts. The median time from crizotinib treatment to first recognization of significant renal cyst change was 77 days. After stopping crizotinib, complex renal cysts regressed significantly. Patients with significant renal cyst change received more previous anticancer therapy (median, 5 lines versus 3 lines, p = 0.04) and received crizotinib for longer duration (median, 956 days versus 248 days, p = 0.007) compared with those without significant renal cyst change. Conclusions Change of renal cysts after crizotinib treatment is not uncommon. Development of complex renal cysts reverses after stopping crizotinib.

Background Tumor characteristics was sought to be related to axillary lymph node metastasis (ALNM), the paramount prognostic factor in patients with invasive breast cancer. This study was aimed to identify the ALNM-associated tumor characteristics and to determine the predictive clinical pathway. Material/Methods Data from 1325 patients diagnosed with invasive breast cancer between January 2004 and January 2010 were retrospectively reviewed. The structure equation model (SEM) was used to build the predictive clinical pathway. Results Among the factors found in the final model, the status of human epidermal growth factor receptor 2 is the primary influence on ALNM through histology grade (β=0.18), followed by tumor size (β=0.16). Tumor size was highly relevant to lymphovascular invasion (LVI) and influenced ALNM through LVI (β=0.26), the strongest predictor of ALNM in the final model (β=0.46) and the highest risk of ALNM (odds ratio=9.282; 95% confidence interval: 7.218–11.936). Conclusions The structure equation model presented the relation of these important predictors, and might help physicians to assess axillary nodal condition and appropriate surgical procedures.

Objective To observe the clinical efficacy and safety of Shuanghu Qinggan Granule (双虎清肝颗 粒, SQG) plus Yigan Yiqi Jieyu Granule (乙肝益气解郁颗粒, YYJG) combined with lamivudine (LAM) on chronic hepatitis B (CHB) patients.

Tumors located in the upper/upper inner quadrant of the breast warrant more attention. A small lesion relative to the size of breast in this location may be resolved by performing a level I oncoplastic technique. However, a wide excision may significantly reduce the overall quality of the breast shape by distorting the visible breast line. From June 2012 to April 2015, 36 patients with breast cancer located in the upper/upper inner quadrant underwent breast-conservation surgery with matrix rotation mammoplasty. According to the size and location of the tumor relative to the nipple-areola complex, 11 patients underwent matrix rotation with periareolar de-epithelialization (donut group) and the other 25 underwent matrix rotation only (non-donut group). The cosmetic results were self-assessed by questionnaires. The average weights of the excised breast lumps in the donut and non-donut groups were 104.1 and 84.5 g, respectively. During the 3-year follow-up period, local recurrence was observed in one case and was managed with nipple-sparing mastectomy followed by breast reconstruction with prosthetic implants. In total, 31 patients (88.6%) ranked their postoperative result as either acceptable or satisfactory. The treated breasts were also self-evaluated by 27 patients (77.1%) to be nearly identical to or just slightly different from the untreated side. Matrix rotation is an easy breast-preserving technique for treating breast cancer located in the upper/upper inner quadrant of the breast that requires a relatively wide excision. With this technique, a larger breast tumor could be removed without compromising the breast appearance.

Insulin and insulin-like growth factor I (IGF1) influence cancer risk and progression through poorly understood mechanisms. To better understand the roles of insulin and IGF1 signaling in breast cancer, we combined proteomic screening with computational network inference to uncover differences in IGF1 and insulin induced signaling. Using reverse phase protein array, we measured the levels of 134 proteins in 21 breast cancer cell lines stimulated with IGF1 or insulin for up to 48 h. We then constructed directed protein expression networks using three separate methods: (i) lasso regression, (ii) conventional matrix inversion, and (iii) entropy maximization. These networks, named here as the time translation models, were analyzed and the inferred interactions were ranked by differential magnitude to identify pathway differences. The two top candidates, chosen for experimental validation, were shown to regulate IGF1/insulin induced phosphorylation events. First, acetyl-CoA carboxylase (ACC) knock-down was shown to increase the level of mitogen-activated protein kinase (MAPK) phosphorylation. Second, stable knock-down of E-Cadherin increased the phospho-Akt protein levels. Both of the knock-down perturbations incurred phosphorylation responses stronger in IGF1 stimulated cells compared with insulin. Overall, the time-translation modeling coupled to wet-lab experiments has proven to be powerful in inferring differential interactions downstream of IGF1 and insulin signaling, in vitro.

Introduction While continuous advancement has been made in the treatment of AML, the overall 5-year survival rate achieved in current days is still less than 40%. The ability to accurately assess AML patient's risk of mortality and relapse is crucial to act on the most appropriate comprehensive treatment strategy. However, the surge of clinical parameters and the scale of data collection are becoming too complex for us to stratify risk and deriving predictive model systematically with conventional statistical methods. In our study, we propose to utilize artificial intelligence techniques in creating risk stratification and outcome prediction model through retrospective data analysis. Method Retrospective clinical data of patients with AML, including basic demographics (age & gender), laboratory results of complete blood count (CBC), white blood composition (WBC) and cytogenetics, and treatment history such as Hematopoietic Stem Cell Transplant (HSCT) and other medication history, was collected at the National Taiwan University Hospital. A total of 479 patients with newly diagnosed de novo AML were enrolled in this study. The median age at diagnosis was 50 years, and the median follow-up duration was 26.6 months, 77.7% of them achieved CR and, 53.4% had relapsed, and 47.2% had received HSCT. In total, 479 cytogenetics tests at diagnosis, 43,518 CBC & WBC records, 28 types of anti-neoplastic medications in L01 & L03 ATC code category with a total of 69,546 medication records from these 479 AML patients were used in deriving the outcome prediction model. A vectorized representation that captures the static-dynamic clinical aspects, i.e., demographics, laboratory results, and treatments, of an AML patient can be learned directly from the collected data. The representation includes both static personal attributes (demographic, and cytogenetic) and time-varying progression of patient's clinical assessment across time (laboratory results, HSCT, and medication). The time-dependent representation was derived from training a deep network architecture of bi-directional long short-term memory network (BLSTM). By taking 10 days as a time step, the BLSTM took the input of Fisher-vector encoded time series of a patient's CBC and WBC, medication, and HSCT records separately. The last output layer, which summarized the relapse/mortality risk exhibited in the measurements of patient's clinical conditions over time, of each separately-learned BLSTM was used as the encoded dynamic clinical representation. The concatenation of the AML patient's static features and the time-dependent representations were fed into a deep neural network followed by a support vector machine to carry out the final prediction. The prediction models were conducted in 5-fold cross-validation experiments and further evaluated using metrics of accuracy (ACC). Results The median leukemia-free survival and median overall survival of these 479 patients are 7.2 months and 49.0 months respectively. By using the CBC & WBC data, the accuracy of next 3-months relapse and mortality prediction at any time point reached 77.9 and 77.0% respectively, while models built with CBC, WBC combined with medication records reached accuracies of 78.4% and 84.6% respectively. Furthermore, incorporating static features, i.e., demographics and cytogenetics, together with time series representation of CBC & WBC, and medication records in our prediction model, we achieved further improved performances of 82.2% and 85.0% accuracies in next 3-month relapse and mortality prediction respectively (Figure 2). Conclusions The BLSTM-DNN model is a novel approach that is capable of jointly taking into account multiple heterogeneous clinical measurements throughout the clinical courses of AML patients in order to derive relapse and mortality prediction. It could be applied at any given time point using past 3-month data to predict the next 3-month relapse or mortality. We also observed the model accuracy increased as we increased the number of clinical exams included in our model. Our study results demonstrated a potential model in facilitating precise and personalized risk assessment that could support physicians for better risk-averse intervention in the future. We plan to integrate other exams including genomic and pathology reports with a larger patient's cohort into our model in the next phase of model development. Disclosures Ko: GNT Biotech & Medicals Crop.: Research Funding; Roche: Research Funding; Abbevie: Research Funding; Mumdipharma Taiwan: Consultancy.

Purpose: This study aimed to identify molecular determinants of sensitivity of non–small cell lung cancer (NSCLC) to anti–insulin-like growth factor receptor (IGF-IR) therapy. Experimental Design: A total of 216 tumor samples were investigated, of which 165 consisted of retrospective analyses of banked tissue and an additional 51 were from patients enrolled in a phase II study of figitumumab, a monoclonal antibody against IGF-IR, in stage IIIb/IV NSCLC. Biomarkers assessed included IGF-IR, epidermal growth factor receptor, IGF-II, IGF-IIR, insulin receptor substrate 1 (IRS-1), IRS-2, vimentin, and E-cadherin. Subcellular localization of IRS-1 and phosphorylation levels of mitogen-activated protein kinase and Akt1 were also analyzed. Results: IGF-IR was differentially expressed across histologic subtypes (P = 0.04), with highest levels observed in squamous cell tumors. Elevated IGF-IR expression was also observed in a small number of squamous cell tumors responding to chemotherapy combined with figitumumab (P = 0.008). Because no other biomarker/response interaction was observed using classical histologic subtyping, a molecular approach was undertaken to segment NSCLC into mechanism-based subpopulations. Principal component analysis and unsupervised Bayesian clustering identified three NSCLC subsets that resembled the steps of the epithelial to mesenchymal transition: E-cadherin high/IRS-1 low (epithelial-like), E-cadherin intermediate/IRS-1 high (transitional), and E-cadherin low/IRS-1 low (mesenchymal-like). Several markers of the IGF-IR pathway were overexpressed in the transitional subset. Furthermore, a higher response rate to the combination of chemotherapy and figitumumab was observed in transitional tumors (71%) compared with those in the mesenchymal-like subset (32%; P = 0.03). Only one epithelial-like tumor was identified in the phase II study, suggesting that advanced NSCLC has undergone significant dedifferentiation at diagnosis. Conclusion: NSCLC comprises molecular subsets with differential sensitivity to IGF-IR inhibition. Clin Cancer Res; 16(18); 4654–65. ©2010 AACR.

Bevacizumab, an antiangiogenic monoclonal anti- body against vascular endothelial growth factor, was designed to normalize tumor vasculature and reduce intratumoral pres- sure. It can create a 'normalization window' during which the cancer can be attacked the most effectively, and the effects of chemotherapeutic drugs are enhanced. Representative trials (E2100, AVADO, RIBBON-1, RIBBON-2 and TURANDOT) have shown that the addition of bevacizumab to chemotherapy has significant benefits on progression‑free survival for metastatic breast cancer, but not on overall survival. The present study describes two patients with meta- static triple-negative breast cancer who received 6 courses of bevacizumab-containing chemotherapy. Each course comprised 5-7.5 mg /kg bevacizumab administered on days 1 and 15, and 20-24 h after bevacizumab delivery, 80 mg/m 2 paclitaxel was administered for 3 weeks on days 2, 9 and 16, followed by 1 week of rest. Following sequential treatment with bevacizumab and paclitaxel, the results of computed tomog- raphy showed that the tumors were rapidly reduced in size. Based on the imaging findings from three‑dimension power Doppler ultrasonography in one of the breast cancer patients who received neoadjuvant chemotherapy with bevacizumab, the possible timing of the normalization window was 20-24 h after the administration of bevacizumab. The normalization window may provide an opportunity to enhance the effect of chemotherapy with the aid of bevacizumab.

HER-2 belongs to the ErbB family of receptor tyrosine kinases, which has been implicated in a variety of cancers. Overexpression of HER-2 is seen in 25–30% of breast cancer patients and predicts a poor outcome in patients with primary disease. Trastuzumab (Herceptin), a monoclonal antibody to HER-2, is specifically approved for HER-2-positive breast cancer but is active only in a subset of these tumors. Blocking HER-2 function by a small molecule kinase inhibitor, therefore, represents an attractive alternate strategy to inhibit the growth of HER-2-positive tumors. HKI-272 is a potent inhibitor of HER-2 and is highly active against HER-2-overexpressing human breast cancer cell lines in vitro. It also inhibits the epidermal growth factor receptor (EGFR) kinase and the proliferation of EGFR-dependent cells. HKI-272 reduces HER-2 receptor autophosphorylation in cells at doses consistent with inhibition of cell proliferation and functions as an irreversible binding inhibitor, most likely by targeting a cysteine residue in the ATP-binding pocket of the receptor. In agreement with the predicted effects of HER-2 inactivation, HKI-272 treatment of cells results in inhibition of downstream signal transduction events and cell cycle regulatory pathways. This leads to arrest at the G1-S (Gap 1/DNA synthesis)-phase transition of the cell division cycle, ultimately resulting in decreased cell proliferation. In vivo, HKI-272 is active in HER-2- and EGFR-dependent tumor xenograft models when dosed orally on a once daily schedule. On the basis of its favorable preclinical pharmacological profile, HKI-272 has been selected as a candidate for additional development as an antitumor agent in breast and other HER-2-dependent cancers.

The swelling behaviour and elastic properties of gelatin gels are reported. Ampholytic swelling properties have been investigated. Antipolyelectrolyte performance is observed when gels are swollen in different salt solutions. Swelling behaviour in high salt solutions is different from that in low salt solutions; this may be attributed to the formation of ion pairs. The elastic properties of the gel and their relationship with swelling character are also discussed. ©1997 SCI

The Nb2 PRL receptor (PRL-R) is known to mediate PRL signaling to the interferon (IFN) regulatory factor-1 (IRF-1) gene via the family of signal transducers and activators of transcription or Stats. To analyze the components of the PRL-R/Stat/IRF-1 signaling pathway, various PRL-R, Stat, and IRF-1-CAT reporter constructs were transiently cotransfected into COS cells. First, mutations in the IFNgamma-activated sequence (GAS), either multimerized or in the context of the 1.7-kb IRF-1 promoter, failed to mediate a PRL response, showing that the IRF-1 GAS is a target of PRL signaling. Next, pairwise alanine substitutions into conserved residues in the proline-rich motif or Box 1 region and two tyrosine mutations, Y308F and Y382F, in the PRL-R intracellular domain all impaired PRL signaling to multimerized GAS or to the 1.7-kb IRF-1 promoter. Furthermore, these PRL-R mutants mediated reduced Stat1 binding to the IRF-1 GAS. Transfection of Stat1 further enhanced PRL signaling to the IRF-1 promoter, suggesting that Stat1 is a positive mediator of PRL action. These studies show that both membrane proximal and distal residues of the PRL-R are involved in signaling to the IRF-1 gene. Further, Stat1 and the GAS element are important for PRL activation of the IRF-1 gene.

Pylephlebitis (septic thrombophlebitis of the portal venous system) is a rare complication of intra-abdominal infection. We aimed to investigate the recent trend of its etiology, clinical manifestation, and prognosis.We retrospectively studied the etiology, clinical manifestation, and outcome by reviewing the medical records of all imaging-confirmed pylephlebitis cases diagnosed during the period 2002-2011 in a university hospital in Taiwan. To identify the risk factors for pylephlebitis, we randomly selected 160 patients with intra-abdominal infections but without pylephlebitis as the comparison group.We identified 35 cases of pylephlebitis. Most patients were men [29/35 (83 %)]. The median age of the patients was 57 years (range 35-90 years). Unspecified abdominal pain (18/35) and fever (10/35) were the most common clinical manifestations. Klebsiella pneumoniae liver abscess (7/35) and cholangitis (7/35) were the most common etiologies. Liver abscess was a risk factor for pylephlebitis (13/35 vs. 27/160, P = 0.01). With antibiotic therapy, there was no in-hospital mortality, but pylephlebitis was still associated with an excess hospital stay (22.2 ± 17.6 vs. 9.8 ± 7.1 days, P 0.001).Our study results suggested a different pattern of pylephlebitis from previous Western literature. K. pneumoniae liver abscess (7/35) is an emerging etiology of pylephlebitis in Taiwan.

Interferon regulatory factor-1 (IRF-1) is a major immediate early gene induced by prolactin (PRL) in a biphasic, cell cycle-dependent manner in Nb2 T cells. This biphasic expression (30 min and 10 h) is mediated in part by an interferon-gamma activation sequence (GAS) in the IRF-1 promoter which binds factors belonging to the Signal Transducers and Activators of Transcription (Stat) family. By electrophoretic mobility shift assays (EMSA), Stat1 alpha was found to be the major and Stat5a a minor component of the 30 min complex. At 10 h, Stat-like factors were again found at the IRF-1 GAS. Western blot analyses show that Stat5a was rapidly induced by PRL to enter the nucleus, but unexpectedly, Stat1 alpha and the alternatively-spliced Stat1 beta were already present in the uninduced nucleus. Further, Stat1 alpha but not Stat1 beta is preferentially tyrosine phosphorylated in response to PRL stimulation. Our studies suggest that multiple Stat complexes may contribute to the biphasic transcription of the IRF-1 gene in PRL-stimulated T cells.

The rat homolog of sheep mammary gland factor (MGF)/Stat5 has been isolated and used to study the regulation of Stat5 during mammary gland development and PRL regulation in COS cells transfected with Stat5a and the PRL receptor. Two alternatively spliced isoforms, designated Stat5a1 and Stat5a2, were identified, the latter encoding a carboxy-terminal truncated protein. A polyclonal antibody to a carboxy-terminal peptide of Stat5a1 was generated and used to measure the level of this isoform during mammary gland development and after PRL induction in COS cells transiently transfected with Stat5a and the long form of the PRL receptor. Surprisingly, Stat5a mRNA and protein were readily detected both in virgin rats and after mammary gland involution. The levels of Stat5a increased during pregnancy, were highest in late pregnancy, and then, unexpectedly, decreased during lactation, the time at which the highest levels of milk protein gene expression are observed. Electrophoretic mobility shift assays using the specific anti-Stat5a1 antisera demonstrated that Stat5a1 comprises part of the heterogeneous, PRL-inducible, protein-DNA complex associated with the beta-casein GAS site. Immunocytochemical analysis detected considerable cytoplasmic and some nuclear staining for Stat5a1 during late pregnancy and predominantly nuclear staining during early lactation. The lack of correspondence of Stat5a gene expression and beta-casein gene expression suggests that Stat5 activation may facilitate the interaction of other factors binding within composite response elements identified recently in the milk protein gene promoters that are then responsible for the stable expression of milk protein genes in terminally differentiated mammary epithelial cells.

Insulin-like growth factor I (IGF1) plays an important role in breast cancer initiation and progression due to its regulation of cell proliferation, migration, and invasion. These characteristics make IGF1R an attractive therapeutic target. While numerous clinical trials have sought to inhibit IGF1R action, these were largely unsuccessful due to a lack of biomarkers for positive therapeutic response, poor patient selection, and potential compensatory signaling by the highly similar insulin receptor (InsR). To identify biomarkers of response, our laboratory identified a set of genes regulated by IGF1 (IGF-sig) that revealed a correlation between activation of the IGF-sig and poor prognosis in estrogen receptor (ER)-negative breast cancer. Further, we showed that ER-negative breast cancer cells are sensitive to IGF1R inhibition both in vitro and in vivo. In an effort to better understand the IGF1 and insulin signaling networks in breast cancer, we performed a reverse phase protein array (RPPA) using 134 antibodies on lysates from twenty-one breast cancer cell lines stimulated with a six point time-course of IGF1 or insulin. We developed a time-dependent model to predict differential mediators of IGF1 and insulin signaling using perturbation analysis. The model predicted that alterations in levels of E-cadherin (CDH1), a major component of the adherens junction, affect IGF1 induced Akt activation. In breast cancer, E-cadherin is genetically lost in invasive lobular cancer (ILC), a subtype that accounts for ∼10-15% of breast cancers. Alternatively, invasive ductal cancers may lose E-cadherin via EMT. I confirmed this in silico prediction, showing that shRNA reduction of E-cadherin enhances the ability of IGF1 to induce Akt signaling, and additionally IGF1R, ERK and S6 ribosomal protein activation. Supporting the clinical relevance of our observations, we found a correlation between loss of E-cadherin (CDH1) mRNA expression and the activation of the IGF-sig in ER-negative tumors within The Cancer Genome Atlas (TCGA). Therefore, we hypothesize that loss of E-cadherin potentiates IGF1 signaling to enhance breast cancer progression, and that loss of E-cadherin expression in ILC and ER-negative tumors may highlight those susceptible to IGF1R inhibition. These studies will investigate how E-cadherin modulates IGF1 signaling and the interaction with EMT with the goal of better defining breast cancers that may respond to IGF1R inhibitors. Citation Format: Alison M. Nagle, Cemal Erdem, YuFen Wang, Kevin Levine, Tiffany Katz, D. Lansing Taylor, Adrian V. Lee, Timothy R. Lezon. Determining the role of E-cadherin in regulating IGF1 signaling in breast cancer: An interaction predicted by large-scale modeling. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1880.

We have previously demonstrated that PRL acts as an autocrine growth factor in the pituitary tumor GH3 cell line. In this study we present evidence that the 235-1 cell line also uses PRL as an autocrine growth factor, and that in this case interaction between PRL and its receptor occurs intracellularly. First, the PRL produced by 235-1 cells was shown to be capable of initiating a proliferative response by placement in the Nb2 bioassay. Second, PRL receptors were demonstrated to be present, but, unusually, primarily in the Golgi complex by light and electron microscope immunocytochemistry. Incubation of the cells in anti-PRL had no effect on 235-1 cell proliferation until interleukin-1 treatment caused the PRL receptors to move to the cell surface, whereupon cell proliferation was inhibited. Specific interference with PRL biosynthesis in noninterleukin-1-treated cells also inhibited cell proliferation. Thus, the essential elements of an autocrine loop are present and can be demonstrated to be functional by antibody inhibition of growth after movement of the receptors to the cell surface and by antisense inhibition of growth with the receptors in their normal intracellular location.

Background Epidemiological evidence suggests that moderately elevated levels of circulating insulin-like growth factor-I (IGF-I) are associated with increased risk of breast cancer in women. How circulating IGF-I may promote breast cancer incidence is unknown, however, increased IGF-I signaling is linked to trastuzumab resistance in ErbB2 positive breast cancer. Few models have directly examined the effect of moderately high levels of circulating IGF-I on breast cancer initiation and progression. The purpose of this study was to assess the ability of circulating IGF-I to independently initiate mammary tumorigenesis and/or accelerate the progression of ErbB2 mediated mammary tumor growth. Methods We crossed heterozygous TTR-IGF-I mice with heterozygous MMTV-ErbB2 mice to generate 4 different genotypes: TTR-IGF-I/MMTV-ErbB2 (bigenic), TTR-IGF-I only, MMTV-ErbB2 only, and wild type (wt). Virgin females were palpated twice a week and harvested when tumors reached 1000 mm3. For study of normal development, blood and tissue were harvested at 4, 6 and 9 weeks of age in TTR-IGF-I and wt mice. Results TTR-IGF-I and TTR-IGF-I/ErbB2 bigenic mice showed a moderate 35% increase in circulating total IGF-I compared to ErbB2 and wt control mice. Elevation of circulating IGF-I had no effect upon pubertal mammary gland development. The transgenic increase in IGF-I alone wasn't sufficient to initiate mammary tumorigenesis. Elevated circulating IGF-I had no effect upon ErbB2-induced mammary tumorigenesis or metastasis, with median time to tumor formation being 30 wks and 33 wks in TTR-IGF-I/ErbB2 bigenic and ErbB2 mice respectively (p = 0.65). Levels of IGF-I in lysates from ErbB2/TTR-IGF-I tumors compared to ErbB2 was elevated in a similar manner to the circulating IGF-I, however, there was no effect on the rate of tumor growth (p = 0.23). There were no morphological differences in tumor type (solid adenocarcinomas) between bigenic and ErbB2 mammary glands. Conclusion Using the first transgenic animal model to elevate circulating levels of IGF-I to those comparable to women at increased risk of breast cancer, we showed that moderately high levels of systemic IGF-I have no effect on pubertal mammary gland development, initiating mammary tumorigenesis or promoting ErbB2 driven mammary carcinogenesis. Our work suggests that ErbB2-induced mammary tumorigenesis is independent of the normal variation in circulating levels of IGF-I.

Charge isomers of monomer PRL have been described for all species thus far examined. For the rat, cow, and chicken, at least a major proportion of the more acidic isomers have been shown to be phosphorylated. The physiological relevance of this posttranslational phosphorylation, however, remains unclear. In this study we have compared the growth-promoting activities of dephosphorylated and standard rat PRL (partially phosphorylated) in the widely used Nb2 bioassay. Dephosphorylated PRL was produced by treatment of standard rat PRL with acid phosphatase, resulting in the conversion of a 91.9% nonphosphorylated/8.1% phosphorylated preparation to a 98.6% nonphosphorylated preparation. These two preparations were added separately or in combination to stationary Nb2 cells. Cell number was assessed 3 days later using a colorimetric assay. Dephosphorylated PRL showed significantly higher growth-promoting activity than standard PRL at concentrations up to 10 ng/ml. In the 1-5 ng/ml concentration range, dephosphorylated PRL was twice as active. Given that the conversion of 6.7% phosphorylated PRL to nonphosphorylated PRL resulted in a doubling of activity, one can deduce not only that phospho-PRL acted as an antagonist to nonphosphorylated PRL in this assay, but also that it did not do so on an equimolar basis. Titration of the two PRL preparations produced data confirming this latter deduction. These data are of importance in our understanding of PRL-promoted cell proliferation.

Insulin receptor (IR) and insulin-like growth factor 1 receptor (IGF1R) have been shown to have a role in breast cancer, although the specific importance of hybrid IR/IGF1R (Hybrid-R)is unknown. I proposed to use an inducible dimerization system to study Hybrid-R, and then optimize an assay to detect Hybrid-R in breast tumor samples. I constructed chimeric IR and IGF1R plasmids with different dimerization domains and developed stable MCF10A clones expressing only chimeric IR or chimeric IGF1R allowing homodimerization. Homodimerizer AP20187 treatment induced activation of chimeric IGF1R and IR in a dose dependent manner and activated AKT and ERK. However, I encountered technical difficulties in establishing cells expressing both chimeric IR and chimeric IGF1R and was unable to induce specific Hybrid-R. To measure the IR/IGF1R hybrid receptor on paraffin embedded specimens, a proximity ligation assay (PKLA) was optimized using both anti-IR and anti- IGF1R antibodies. The presence of PLA signals was increased in cells had IR or IGF1R overexpression and decreased in cells with IGF1R knock down. The variability of signals was also observed in a tissue microarray with normal breast tissue and breast tumor. This study shows the presence of Hybrid-R in breast cancer cell lines and human tumors and suggests that more studies are needed to determine its function and importance in breast cancer.

The effects of different concentrations of various K-current blockers on prolactin-induced proliferation and membrane K-currents in malignant lymphocytes (Nb2 cells) were investigated. Membrane currents were measured with the whole cell patch-clamp technique, and lymphocyte density was quantified by both spectrophotometric and conventional methods. K-current blockers tested (quinidine, 4-aminopyridine, barium, and tetraethylammonium) exhibited similar rank order potency for K-current block and inhibition of prolactin-induced proliferation of malignant lymphocytes. Because Nb2 cells proliferate independently of a transmembrane Ca-influx, these results suggest that K-currents per se rather than K-current modulation of Ca-influx is an essential event for lymphocyte proliferation.

Capsaicin (trans-8-methyl-N-vanillyl-6-nonenamide), a significant pungent ingredient in a variety of red peppers of the genus Capsicum, is a type of vanilloid. It has been shown to exert biological activities (anticarcinogenic, antimutagenic and chemopreventive) in many cancer cell lines. It was found that capsaicin induces dose-dependent growth inhibition of MCF-7 cells, which does not express caspase-3. In this study, we investigated the molecular mechanism of capsaicin-induced apoptosis in MCF-7 cells. Treatment with capsaicin for 24 h resulted in dose-dependent apoptosis in these cells. After the addition of capsaicin, the levels of reactive oxygen species were reduced slightly in the earlier stage of treatment. Interestingly, an elevation of intracellular calcium ion concentration was detected in the MCF-7 cells. In time course and dosage studies, the mitochondrial membrane potential of MCF-7 cells decreased. However, the change was not significant. It is worth noting that the apoptosis-inducing factor translocated into the cytosol and nucleus from the mitochondria. Our results suggest that capsaicin induces cellular apoptosis through a caspase-independent pathway in MCF-7 cells, and that reactive oxygen species and intracellular calcium ion fluctuation has a minimal role in the process.

BackgroundThe roles of mitochondria in energy metabolism, the generation of ROS, aging, and the initiation of apoptosis have implicated their importance in tumorigenesis. In this study we aim to establish the mutation spectrum and to understand the role of somatic mtDNA mutations in esophageal cancer.MethodsThe entire mitochondrial genome was screened for somatic mutations in 20 pairs (18 esophageal squamous cell carcinomas, one adenosquamous carcinoma and one adenocarcinoma) of tumor/surrounding normal tissue of esophageal cancers, using temporal temperature gradient gel electrophoresis (TTGE), followed by direct DNA sequencing to identify the mutations.ResultsFourteen somatic mtDNA mutations were identified in 55% (11/20) of tumors analyzed, including 2 novel missense mutations and a frameshift mutation in ND4L, ATP6 subunit, and ND4 genes respectively. Nine mutations (64%) were in the D-loop region. Numerous germline variations were found, at least 10 of them were novel and five were missense mutations, some of them occurred in evolutionarily conserved domains. Using real-time quantitative PCR analysis, the mtDNA content was found to increase in some tumors and decrease in others. Analysis of molecular and other clinicopathological findings does not reveal significant correlation between somatic mtDNA mutations and mtDNA content, or between mtDNA content and metastatic status.ConclusionOur results demonstrate that somatic mtDNA mutations in esophageal cancers are frequent. Some missense and frameshift mutations may play an important role in the tumorigenesis of esophageal carcinoma. More extensive biochemical and molecular studies will be necessary to determine the pathological significance of these somatic mutations.

The components of the Wnt-signaling pathway are reported to be mutated in human cancer cells, but the relationship between the components and oral squamous carcinoma (SCC) is still unknown. In this study, we analyzed the epigenetic changes and expression patterns of four member proteins of the Wnt-signaling pathway and analyzed the mutations of beta-catenin and AXIN 1 genes, in order to explore the roles of the pathway in the development of oral cancer. The results showed that there are no beta-catenin and AXIN 1 gene mutations and no methylation of the CpG island of beta-catenin, AXIN I and GSK3beta genes in oral cancer cells; methylation of the CpG island of APC occurs in the precancerous stage and it is a dynamic change; the aberrant expressions or abnormal localization of the Wnt-signaling pathway proteins have no relationship with methylation status or mutation. From our results, we suggest that the Wnt pathway related genes play a very limited role in the development of oral SCC.