bluuebunny/crossref_metadata_2025 huggingface dataset:
The inducible cyclic adenosine 3’,5’-monophosphate early repressor (ICER) enhances drug sensitivity in acute myeloid leukemia
Authors: Pigazzi M. , Manara E. , Baron E. , Beghin A. , Basso G.
Published: August 2020

e22045

Background: The Inducible Cyclic Adenosine 3’,5’-monophosphate early repressor (ICER) is a transcription factor that principally counteracts the cAMP response element binding protein (CREB) activity. CREB was previously demonstrated to be overexpressed in acute leukemia, whereas ICER was found rapidly degradated being unable to control gene transcription. ICER exogenous expression was demonstrated to repress CREB targets preventing leukemia progression. We hypothesized that ICER restoration deserves a special consideration for playing a role in CREB oncogenic feature and in modeling leukemic cell phenotype. Methods: We constructed an expression vector for ICER and induced its exogenous expression in HL60 cells (HL60+ICER). We monitored transcription and translation of a series of genes involved in different pathways by quantitative gene expression and western blot analysis. We investigate ICER's role in cell death after treatment with chemotherapic drugs. Results: We revealed that ICER was able to control gene expression in leukemia, principally of genes involved in cell death and survival. In particular, we focused on DUSP1 and DUSP4 phosphatases which were found significantly repressed. HL60+ICER after being treated with drugs increased apoptosis. Cell cycle analyses revealed a block in G2 phase, a lowered cell proliferation and clonogenic potential with respect to HL60 treated at the same conditions. DUSPs downstream targets (IL6, RB, RAS) were lowered expressed and p38 constitutively phosphorylated confirming their role in mediating cellular stress. ICER driven apoptosis was found to be caspases mediated. An enhanced activity of p38 was observed in treated HL60+ICER, and by the use of specific p38 inhibitors apoptosis decreased. Other possible apoptotic pathways (AKT, ERK, JNK) were excluded because found severely disrupted. Conclusions: Finally, we described an enhanced sensibility to drugs in HL60 induced by ICER exogenous expression and that p38 stress signaling pathway is the direct target of this phenomenon. The ability of ICER to modify leukemic cells phenotype respect to chemotherapeutic treatment gave an important information for targets and therapeutic possibilities.

No significant financial relationships to disclose.
Gene set enrichment analysis to evaluate expression of autoantigens in lung cancer
Authors: Johnson K. , Berger A. , Watkins T. , Cheadle C. , Casciola-Rosen L. , Levine S. M.
Published: August 2020

e22048

Background: There is a well-established association between certain autoimmune diseases and the development of specific malignancies. It has been demonstrated that myositis-specific autoantigens are expressed at higher levels in tumors associated with myositis compared to normal tissue, suggesting that immune responses to antigens expressed in nascent tumors may contribute to the autoimmune process. Whether this observation is a general feature of autoantigen expression in tumor tissue, and whether the relative expression of these antigens is enriched in relation to the rest of the tumor transcriptome is currently unknown. Methods: Tumor tissue from ten lung cancer biopsies (adenocarcinoma (4), carcinoid (4), and squamous cell carcinoma (2)) and normal lung from the same patients were obtained. Total RNA was extracted and hybridized to Illumina Sentrix BeadChips. Hiearchical clustering was used to visualize the expression levels of 146 known autoantigens. Gene Set Enrichment Analysis was performed using 20 disease-specific autoantigen gene sets and 1892 gene sets from the Molecular Signatures Database. Protein levels of selected autoantigens were assessed by immunoblotting detergent tissue lysates. Results: Single-linkage hierarchical clustering analysis reveals groups of autoantigens that are differentially expressed between normal lung tissue, carcinoid tumors, and adenocarcinomas. Adenocarcinoma tumor samples were significantly enriched for myositis (nominal p-value <0.001, false discovery rate (FDR) q-value 0.009) and SLE autoantigens (p-value 0.004, FDR 0.029). Scleroderma autoantigens were enriched in carcinoid tumors (p-value 0.003, FDR 0.053). Increased protein expression of the autoantigens Mi-2 and topoisomerase-1 in carcinoid tumors was confirmed by immunoblotting. Conclusions: This study reveals that autoantigens targeted in several autoimmune diseases are both transcribed and expressed at high levels in malignancies known to associate with those disorders. While autoantigens comprise only a small fraction of the total transcriptome, they are disproportionately expressed in tumors known to associate with autoimmunity, supporting the hyporthesis that autoimmunity to these proteins may arise via nascent anti-tumor responses.

No significant financial relationships to disclose.
A pilot study to compare 8 - FDG and F18 PET/CT study in delineating metastases in suspected skeletal disease
Authors: Shah S. A. , Rangarajan V. , Purandare N. C. , Sharma A. R. , Arora A. C. , Parasar D. S.
Published: August 2020

e22052

Title: A pilot study to compare the 18 - FDG and F18 PET/CT study in delineating metastases in suspected skeletal disease. Background: Flourodeoxy glucose (FDG), positron emission tomography/computed tomography (PET/CT) scans have been used to identify metastatic disease including skeletal lesions. But the advent of Flourine -18 (F - 18), has necessitated a need to identify its accuracy over FDG scans. Aim: To evaluate and compare FDG PET/CT and F18 PET/CT studies in locating skeletal metastases in patients with suspected disease. Methods: A pilot study was carried out on 27 patients who were referred for a FDG PET/CT study for suspected skeletal disease. A whole body (skull to ankle) FDG PET/CT study followed by a F18 PET/CT bone scan within a period of 1 week was performed. A total of 150 lesions with increased tracer concentration on FDG and F18 scan were analyzed and the characteristics of the lesion on corresponding CT images were noted. Results: Of the 150 lesions noted, 49 were seen in both FDG and F18 scans. 11 were sclerotic,16 lytic, 17 mixed while CT was normal in 5 lesion. 95 of the 101 mismatched lesions were seen on F18 scan alone & were not appreciated on the FDG scan. 40% were sclerotic, 12% mixed and 11.5% were lytic. Degenerative changes comprised 12% lesions. Only 6 mismatched lesions were seen on FDG and not appreciated on F18 study.They showed no morphological abnormality on CT. 9 patients with a negative FDG scan showed lesions ranging from solitary to 16 on F18 scan, while 5 patients who had a single metastasis on FDG showed more than 6 lesions on a F18 scan. Conclusions: A F18 PET/CT study detects more skeletal lesions than FDG PET and can thus has a potential to impact patient management . Sclerotic lesions missed on FDG scans seem to be better picked on F18 scans. This pilot study provides the feasibility of a prospective study in a larger patient population to validate the impact of F18 scan in identifying skeletal metastases in various malignancies with a predisposition to bone spread.

No significant financial relationships to disclose.
Underweight and breast cancer recurrence and death
Authors: Moon H. , Han W. , Lee J. , Ko E. , Kim E. , Kim E. , Yu J. , Noh D.
Published: August 2020

e22054

Background: The association of body mass index or body weight and breast cancer outcome is controversial. Furthermore, the impact of underweight on breast cancer recurrence and death has not been adequately addressed. Methods: We investigated this issue in a large nation-wide database of 14,039 Korean breast cancer patients (Korean Breast Cancer Registry). Furthermore, to test whether the association was due to more frequent non-cancer death in underweight women, we used breast cancer recurrence data of 4,345 women from single institution (Seoul National University Hospital). Results: The results from Korean Breast Cancer Registry data showed significantly lower overall survival in underweight patients compared to normal weight after adjusting known prognostic factors [hazard ratio=1.36 (95% CI 1.08–1.72)], which was not observed in obese patients. The association between body mass index and breast cancer recurrence was further investigated in Seoul National University Hospital data. Underweight women had significantly higher risk of distant metastasis and local recurrence of breast cancer [hazard ratio=1.86 (95% CI 1.25–2.76) and 2.62 (95% CI 1.55–4.41), respectively]. Conclusions: These findings suggest that underweight could be considered as high risk factor of death and recurrence after breast cancer surgery.

No significant financial relationships to disclose.
Pattern of relapse and the risk of second malignancies in molecular subtypes of early breast cancer (BC)
Authors: Radecka B. , Hudala-Klecha J. , Badzio A. , Szwiec M. , Drosik K. , Jassem J.
Published: August 2020

e22057

Background: In a large, consecutive series of early BC patients (pts) we investigated clinical behavior of four major molecular subtypes, with particular focus on pattern of relapse and the risk of second malignancies. Methods: BC cases were divided into four phenotypes: triple negative BC (TNBC), characterized by negativity of hormone receptors (HR) and HER2 (0–2+ by immunohistochemistry), HR+/HER2-, HR+ /HER2+ and HR-/HER2+. All pts underwent curative surgery between January 2003 and December 2007, followed by radiation and/or systemic therapy according to current standards. Univariate risks of relapse and death were estimated using Kaplan-Meier survival curves and compared with log-rank test, and the multivariate analysis used stratified Cox model. Results: Of the 783 tumors, 520, 50, 48 and 165 were classified as HR+/HER2-, HR+/HER2+, HR-/HER2+ and TNBC, respectively. The TNBC and HR-/HER2+ subtypes were associated with higher tumor grade (p<0.001), higher T stage (p<0.001), and higher proportion of medullary type (p<0.001) compared to other two subtypes. After a median follow-up of 30 months, the risk of relapse in HR+/HER2-, HR+/HER2+, HR-/HER2+ and TNBC was 5.0%, 10.0%, 12.5% and 12.1%, respectively (p=0.007). Inferior prognosis of TNBC and HR-/HER2+ types remained significant after correction by T and N stage. Of the 57 relapses, 12 were locoregional, 37 distant and 8 concomitantly locoregional and distant. In TNBC isolated locoregional relapse constituted 45% of all relapses, compared to 8.1% in three other subtypes combined (p=0.001). TNBC was associated with almost doubled risk of second malignancy compared to all other types combined (12.7% vs. 6.5%, respectively; p=0.008). This risk was significantly increased for contralateral BC and endometrial cancer (p=0.002 and 0.024, respectively). Conclusions: Poor prognosis of TNBC is related to increased occurrence of both locoregional and distant relapses. Additionally, this subtype carries increased risk of second malignancies, most likely due to higher proportion of hereditary BC.

No significant financial relationships to disclose.
The antitumor effect of orally active heat shock protein 90 Inhibitor, 17-DMAG, on the growth of gefitinib-resistant non-small cell lung cancer
Authors: Kubo T. , Kobayashi N. , Jida M. , Hayashi T. , Yamamoto H. , Otani H. , Kiura K. , Sano Y. , Toyooka S.
Published: August 2020

e22064

Background: It has been reported that NSCLC cell lines with activating mutations of EGFRgene were significantly sensitive to Heat Shock Protein 90 (Hsp 90 ) ihibitors regardless of the secondary T790M mutation. Methods: We examined the antitumor effect of 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG), the orally active Hsp90 inhibitor, on the growth of NSCLC cell lines in vitro and in vivo. Results: In MTS assay, the IC50 values for 3 cell lines with EGFR mutation (PC-9, RPC-9, and NCI-H1975) ranged from 0.15 to 0.28 μM. RPC-9 is a gefitinib-resistant lung cancer cell line with the secondary T790M mutation established from PC-9. By contrast, those of 7 EGFR-wild type cell lines ranged from 1.63 to 28.5 μM. Western blot analysis revealed that mutant EGFRs were more readily depleted than wild-type EGFRs after the treatment of 17-DMAG. Furthermore, expression of phospho-Akt, phospho-MAPK, cdk4, and cyclin D1 was depleted after exposure to low concentrations of 17-DMAG in EGFR-mutant cells. Celeaved PARP expression confirmed apoptosis in response to 17-DMAG in EGFR-mutant cells. In animal models, 17-DMAG significantly reduced the growth of EGFR-mutant cell lines PC-9 and RPC-9 compared with controls and induced degradation of mutant EGFR. In contrast, 17-DMAG did not inhibit the growth of EGFR wild type cell lines compared with controls. Conclusions: These results suggested that 17-DMAG could become a novel therapeutic agent for patients with lung tumors expressing mutant EGFR and having developed clinical resistance to gefitinib.

No significant financial relationships to disclose.
Immune self-tuning in renal cell carcinoma patients
Authors: Busse A. , Asemissen A. , Schmittel A. , Zimmermann K. , Miller K. , Rietz A. , Ochsenreither S. , Fusi A. , Thiel E. , Keilholz U.
Published: August 2020

e22069

Background: Renal cell carcinoma (RCC) cells can inhibit protective antitumor immunity by secretion of immunosuppressive factors leading to the induction of regulatory T cells. The objective of this study was to investigate the prognostic impact of mRNA expression levels of IL10, TGFβ and forkhead box transcription factor (FoxP3) mRNA in peripheral blood mononuclear cells (PBMCs) of metastatic RCC patients before receiving treatment with sorafenib. Methods: PBMCs of 46 patients were assessed for their expression levels of TGFβ, IL10 and FoxP3 by quantitative RT-PCR. Clinical features considered included ECOG performance status, hemoglobin, alkaline phosphatase, and calcium concentrations. Disease evaluation was performed every 8 weeks following RECIST criteria. Relationship between pre-treatment factors and survival were examined in univariate analyses and subsequently by multivariate analysis using a stepwise Cox regression model. Results: In contrast to FoxP3, mRNA expression levels of IL10 and TGFβ were significantly higher in the 46 RCC patients compared to healthy volunteers: Median expression levels [ratio marker /housekeeping gene PBGD] were 5.56E-05 vs 2.05E-04 (P=0.034) for IL10 and 7.38E-02 vs 3.04E- 01 (P=0.023) for TGFβ. Univariate analysis revealed a negative prognostic influence of IL10 on progression free survival (p=0.04) and on overall survival, although not significant (P= 0.063). Surprisingly, high TGFβ and FoxP3 expression levels had a positive influence on progression free (P<0.001 and P=0.047, respectively) and overall survival (P<0.001 and P= 0.031, respectively). In the multivariate analysis low ECOG performance status and high TGFβ mRNA expression levels were independently associated with worse progression free (P=0.001 and P=0.054,) and worse overall survival (P=0. 006 and P< 0.001, respectively). Conclusions: RCC caused an immune-suppressive phenotype in PBMC characterized by increased mRNA expression levels of IL10 and TGFβ. Surprisingly, in contrast to IL10, a high TGFβ mRNA expression level was an independent good prognostic factor. Whether this observation can be attributed to recently described immune promoting functions of TGFβ needs to be determined.

No significant financial relationships to disclose.
Investigation of infiltrating regulatory T lymphocytes in tumor areas in oral cancer lesions
Authors: Sukhotska I. , García-Alonso L. , Sirera R. , Sarrión G. , Jantus Lewintre E. , Gil M. , Cabrera A. , Berrocal A. , Bagán J. , Camps C.
Published: August 2020

e22070

Background: HLA-G is a human non-classical MHC molecule, mainly expressed in the trophoblast whose main function is to suppress immunologic activity that allows maternal tolerance to phoetus. On the other hand, infiltrating regulatory T lymphocytes (Treg) could promote peripheral inmunotolerance fo oncogenic transformation. Methods: We performed real-time PCR in frozen oral cancer specimens from untreated patients who had undergone surgical resection (n=22) and in normal oral mucosa from healthy subjects (n=10). Samples were processed for mRNA extraction and quantification of gene expression was expressed as relative concentration by an endogenous gene. To asses the presence of infiltrating Treg we analysed CTLA-4, Foxp-3, IL-10, TGF-beta, CD4, CD8, CXCR4, CD127 and CD25 and also we determined HLA-G levels. We correlate the expression of immunologic mediators with clinical variables. Results: Patients presented squamous carcinomas of the tongue (n=12) or gum (n=10) and stages ranged from I to IV (stage I=4; II=9; III=4; IV=5). 10 patients presented well-differentiated lesions and the other 12 moderately-differentiated cells. Eight patients received post- surgery chemo and radiotherapy. Our results show that tumor samples had significant higher expression of the CTLA-4, Foxp-3, TGF-beta and CD127 genes than normal tissue. However, those data showed no correlation between the levels of expression and clinico-pathologic variables. When patients were grouped according to tumor size, there was a trend in the way that bigger tumoral lesions expressed relative higher amounts of Treg. By contrast we could not observe an increase of the expression of HLA-G in patients. Conclusions: Our results reveal that there is an increase in the expression of Treg in oral cancer patients. These Treg in tumoral tissues might contribute to the impairment of immunological rejection of the neoplasic transformation. Conversely we have not been able to demonstrate tumoral expression of HLA-G as a strategy to escape from immunosurveyance. Further analysis of this cells and their function is important in order to develop new therapeutic strategies.

No significant financial relationships to disclose.
Plasma thermogram profiling: A novel biomarker for lung cancer
Authors: Xiang D. , Garbett N. C. , Chaires J. , Laber D. L. , Kloecker G. H.
Published: August 2020

e22074

Background: Currently, there are no clinically useful biomarkers for lung cancer (LC). We developed a novel approach by using differential scanning calorimetry (DSC) to analyze the plasma proteome of LC patients. Our assay produces a signature snapshot of the plasma proteome based on the thermal properties of the constituent proteins within the mixture. Each individual protein has a unique and characteristic melting temperature and melting enthalpy. These biophysical properties are as intrinsic and unique for each protein as are its sequence, mass, and charge. The denaturation of plasma results in a composite signature thermogram that represents the sum of all of the individual proteins within the proteome, weighted according to their concentration within the plasma or serum. Our goal is to identify unique thermogram profiles as biomarkers for lung cancer evaluation. Methods: Plasma samples were collected from LC patients and healthy volunteers. Sample analysis was performed using an automated capillary DSC. Comparison of plasma thermograms from controls and diseased individuals was done using quantile-quantile plots and the Kolmogorov-Smirnov test. Results: One hundred samples were obtained from healthy volunteers. DSC thermograms from control plasma were highly reproducible and yielded a characteristic signature with a well-defined shape and temperature maxima. Twenty samples from LC patients were obtained at diagnosis. Plasma from LC individuals yielded a unique thermogram that differs from that of healthy controls. Preliminary results suggested that DSC was sensitive to properties of the plasma proteins other than their charge and mass. We hypothesize that changes in thermogram shapes and positions in LC can arise from changes in the concentrations of plasma proteins, from disease-induced protein-protein interactions, or from the secretion into plasma of disease-related peptides that can bind to the most abundant plasma proteins. Conclusions: Lung cancer may have a characteristic signature thermogram that may serve as a biomarker for LC; further studies are needed to elucidate the biophysical and molecular mechanisms of different thermograms between normal individuals versus those with disease or cancer.

No significant financial relationships to disclose.
Effect of Notch inhibition on radiation in an explant model of glioblastoma multiforme
Authors: Hovinga K. E. , Wang R. , Shimizu F. , Moayedparadazi H. , Menon J. , Correia A. , Major T. , Tabar V.
Published: August 2020

e22080

Background: Glioblastoma Multiforme (GBM) is highly radioresistant, possibly due to a subpopulation of Brain Tumor “Stem-like‘ Cells (BTSC) capable of repopulating the tumor. The Notch signaling pathway is essential for maintaining normal neural stem cells. We asked whether Notch inhibition can overcome radioresistance by allowing BTSCs to exit their stem cell state. Methods: To maintain the BTSCs in their original niche we used an organotypic culture method (explants). GBM explants are cultured on a semiporous membrane in an air-medium interface. They are treated with the Notch inhibitor DAPT, radiation or a combination thereof. Proliferation and neurosphere forming capacity are assessed. Results: The explant model faithfully maintained the cytoarchitecture of the tumor (preservation of blood vessels and pericytes) and its high proliferation rate (%ki67+ cells: 14.3% -17.1%). The effect of DAPT or radiation treatment alone differed among tumors (from no change to a two-fold and nine-fold decrease post DAPT or radiation respectively). However, in all tumors tested the combination of DAPT and radiation dramatically decreased the proliferation rate compared to either treatment alone (%ki67 + cells: 0.19%-2.24%). Interestingly the rate of neurosphere formation was highest following radiation alone (threefold increase). Neurosphere formation after DAPT treatment differed amongst the tumors from a decrease (fourfold) to an increase (twofold). The combination treatment resulted in a modest increase over control (twofold). Conclusions: Our data demonstrates that inhibition of proliferation does not correlate with the status of neurosphere forming ability, arguably a surrogate for cancer stem cell function. Combining radiation with notch inhibition has a profound effect on GBM proliferation, most likely due to selectively inhibiting the BTSC repopulating ability. Ongoing studies will determine the impact of this approach on tumor growth and progression in in vivo tumor models.

No significant financial relationships to disclose.
False-positive diagnosis of breast cancer by diffused optical tomography with ultrasound
Authors: Wang X. , Li Y. , Cao X.
Published: August 2020

e22085

Background: Breast cancer is one of the most common cancer in women. Early detection, early diagnosis and early treatment play key role in fighting against breast cancer. OPTIMUS system is a system of diffused optical tomography with ultrasound. It provides dual modality images for early diagnosis of breast cancer. The aim of this study was to evaluate the OPTIMUS system on diagnosis of breast disease. Methods: OPTIMUS system was applied to 160 breast tumor patients. All patients had received surgical treatment and had definite pathological diagnosis. OPTIMUS system was evaluated as diagnostic tool of breast tumor in this study. Results: There were 42 cases diagnosed as benign breast disease and 118 cases diagnosed as breast cancer by OPTIMUS system. Pathology confirmed 60 cases of benign disease and 100 cases of breast cancer. False positive rate of breast cancer was 30% (18/60). False negative rate of breast cancer was 0% (0/100). The pathology of false positive cases was mild and severe papillomatosis (6/18), non-typical hyperplasia (4/18), chronic inflammation (3/18), fibroadenoma (3/18) and fat necrosis (2/18). Papillomatosis and non-typical hyperplasia are precancerous lesions and often difficult for clinical diagnosis. In this study the false positive diagnostic rate of mammography and ultrasonography is 13% and 11.1% respectively. Conclusions: OPTIMUS system is a non- invasive and highly effective diagnostic tool for breast disease. Its sensitivity is reached to 100% and specificity is about 70% on the diagnosis of breast cancer. OPTIMUS system could be used as assistant diagnostic tool for breast tumor.

No significant financial relationships to disclose.
Use of novel optical proteomics to profile breast cancer patients leading to individualised prognosis and tailored treatment
Authors: Kelleher M. T. , Festy F. , Barber P. R. , Gillett C. , Ofo E. , Coolen A. , Pinder S. , Patel G. , Vojnovic B. , Ng T. , Ellis P. A.
Published: August 2020

e22090

Background: Optical proteomics quantifies interactions between proteins and post-translational modifications by measuring Förster resonance energy transfer (FRET) quantified by fluorescence lifetime imaging microscopy (FLIM). This project aims to derive multiple high throughput optical proteomic markers, to predict metastatic risk at first diagnosis, and to perturb ‘high risk' protein-protein interactions using targeted therapeutics. This initial step develops robust FRET/FLIM assays, suitable for use in formalin fixed paraffin embedded (FFPE) tissue to be correlated with patient outcome. Methods: Fluorophore-conjugated antibodies to proteins involved in cell migration and survival, were applied to tissue microarrays (TMA), created from archived FFPE invasive ductal breast carcinoma samples. Where fluorophores are located within nanometer proximity, FRET occurs, thus allowing quantification of protein-protein interaction. Ezrin and PKCα phosphorylation, distribution, and interaction were imaged on four TMAs (patients diagnosed with early breast cancer 1984 -1987: 20 years follow-up data). Results: 71 patient samples were optically imaged. Patients were clustered based on the pairwise distances between 18 optical variables ‘input data'. Data are represented on self organising maps and dendrograms and correlated with clinical outcome ‘output data', displaying a heatmap distribution. Conclusions: Ezrin and PKCα phosphorylation, distribution, and interaction imaged optically within FFPE contain prognostic information regarding metastatic outcome in breast cancer, thus stepping ever closer to individualising prognosis. These advanced optics-based parameters informing on metastatic potential will be validated in prospective studies in conjunction with FRET/FLIM assays measuring HER2/HER3 dimerisation, and EGFR and HER2 ubiquitination in order to improve patient selection for targeted therapy.

No significant financial relationships to disclose.
Use of laser capture microdissection to derive poor prognosis gene expression signatures of epithelial abdominal malignant mesothelioma
Authors: Borczuk A. C. , Chen J. , Parikh F. , Powell C. A. , Taub R. N.
Published: August 2020

e22098

Background: Prior work in abdominal malignant mesothelioma (AMM) has established a poor prognosis profile of biphasic AMM when compared to more favorable prognosis epithelial AMM. It is evident, however, that epithelial AMM shows heterogeneity in biologic behavior and prognosis. Given improvements in therapy and survival in AMM, stratification of patients with epithelial AMM into favorable and poor prognosis subgroups is desirable and gene expression profiles may provide poor prognosis signatures to further guide therapy. Methods: Favorable and poor prognosis subgroups were defined as 1000 days of survival time (based on median survival of 90 patients with AMM at Columbia Presbyterian as of Sept 2007). Fourteen frozen epithelial AMM (7 favorable prognosis, 7 poor prognosis) with high quality RNA were laser capture microdissected using a PALM Zeiss microscope and analyzed using Affymetrix U133 Plus 2 microarrays. Unsupervised hierarchical clustering and supervised subgroup analysis were performed using BRB tools version 3.2. Results: Unsupervised hierarchical clustering revealed 2 main clades that contained 6 of 7 poor prognosis and 6 of 7 favorable prognosis tumors, respectively. Supervised class comparison revealed a 39-gene signature that distinguished the 2 classes (permutation values of p < .001). High expression of Nde1, UHRF1, and EZH2 by qPCR on a set of 32 AMM (using an upper quartile cutoff) was associated with poor survival (Log rank statistic p<.01, p<.003 and p<.03 respectively). Immunohistochemistry performed on tissue microarrays of 122 AMM for p16 loss and increased EZH2, Col3A1 and UHRF1 immunoreactivity was associated with poor survival (Log rank statistic p<.0001, p<.009, p<.0001 and p<.012, respectively). Conclusions: Using a training set of laser captured favorable and poor prognosis epithelial AMM, a 39-gene signature was obtained that is associated with prognosis. Validation of several of the candidate genes by qPCR expression and immunohistochemistry was performed, confirming favorable and poor prognosis strata among the epithelial AMM. Identification of poor prognosis epithelial AMM at the time of initial biopsy/tumor cytoreduction could lead to more aggressive primary therapeutic intervention.

No significant financial relationships to disclose.
Apoptotic and immunomodulatory effects of green tea extracts (GTE) on chemoresistant human tumor cells
Authors: Braun D. P. , Johnson D. M. , Katsantonis N. G. , Bhesaniya K. , Staren E. D.
Published: August 2020

e22101

Background: GTE and Epigallocatechin 3-gallate (EGCG), the most abundant polyphenol in GTE, have been shown to exert inhibitory effects on carcinogenesis; modulatory effects on tumor proliferation and differentiation; and immunomodulatory effects on tumor immunity in different pre-clinical models. These pluripotent effects suggest that GTE may have clinical activity that could be exploited for treatment of chemo-insensitive but immunologically responsive tumors. Thus, the present study investigated the effects of EGCG on the proliferation and immunologic sensitivity of human renal cell cancer (RCC) and malignant melanoma (MM) cell lines. Methods: Human RCC (769-P) and MM (A375) cell lines were tested following incubation in media ± 21.8μM EGCG, a pharmacologically-achievable concentration produced by 8, 200mg capsules GTE daily. Tumor proliferation was assessed by MTS assay; lytic sensitivity to IL2-activated human peripheral blood lymphocytes (IL2PBL) by 51Chromium release assay; and gene expression by quantitative RT-PCR assay. Results: EGCG produced significant inhibition of proliferation of both RCC and MM cells (61.5% and 67.3% of media control values respectively; p<0.01 by 2-tailed, paired t test). EGCG also caused significantly increased expression (≥ 2 times (×) media control values) of pro-apoptotic genes in both RCC: BCL2L1 (5.4×); BCL2L10 (7.9×); BIK (2.9×); Caspase 5 (6.3×); and Caspase 14 (6.3×) and MM cells: Caspase8 (2X) and Card6 (2.5X). Unfortunately, EGCG pre-treatment also decreased the sensitivity of RCC (43% of control) and MM (53% of control) to IL2PBL-mediated lysis (p<0.01 respectively). This occured in association with significant upregulation of Fas ligand mRNA in RCC (6.3×) and TRAF2 mRNA in MM (33 ×). Conclusions: The results of this pre-clinical study demonstrate that EGCG exerts significant antiproliferative effects against human RCC and MM cells in vitro in association with increased expression of different pro-apoptotic genes. Unfortunately, EGCG-treated cells also demonstrated reduced sensitivity to lysis by IL2-activated human lymphocytes. Taken together, the results suggest that GTE may have activity in vivo in patients against chemoresistant RCC or MM but should not be used in conjunction with IL2 therapy.

No significant financial relationships to disclose.
Relationship between lymphovascular invasion (LVI) and prognostic markers in different subtypes of breast cancer
Authors: Panopoulos C. G. , Tzavara C. , Papadimitriou K. , Pistalmatzian N. , Lypas G. , Barbounis V. , Demiri S. , Koumakis G. , Apostolikas N. , Efremidis A.
Published: August 2020

e22109

Background: Prognostic and predictive factors for invasive breast cancer (BC) are well established. LVI was added to the list of prognostic factors for node negative patients (St.Gallen 2006). The aim of this study was to investigate the relationship of LVI with known prognostic markers and its role on different subtypes of breast cancer. Methods: 367 patients with primary BC were evaluated in this study. Tumoral and peritumoral breast tissue was examined for LVI. The patient's age, menopausal status, tumor histology, tumor size, histologic grade, lymph node (LN) involvement, hormone receptor (HR) status and HER-2 status were determined as markers of prognosis. The association of LVI with the aforementioned markers was evaluated using multiple logistic regression analysis. Results: The distribution of the patients was as follows: 40.9% were HR + and HER-2 - (luminal type), (32.4%) were triple negative, 26.7% were HER-2 Positive (9.5% of them were HER-2 + and HR -, while 17.2% were HER-2 + and HR+). Almost half of the patients (48.8%) had one or more positive nodes. Multiple analysis revealed that histologic grade, LN involvement and HER-2 positivity were independently associated with LVI. Patients with histologic grade II and III had 5.31 (95% CI: 1.14–24.75) and 7.03 (95% CI: 1.47- 33.53) times greater odds for LVI respectively compared to those with grade I. Patients with one or more than three nodes involved had 3.95 (95% CI: 2.13–7.33) and 6.29 (95% CI: 3.14–12.63) times greater odds for LVI respectively compared with node negative tumors. Furthermore, HER-2 positive patients had almost twofold likelihood for LVI (OR=1.80, 95% CI: 1.02–3.16). The effect of HER-2 positivity on LVI was independent of HR status. Age, menopausal status, tumor histology, tumor size and HR status were not significantly associated with LVI. Conclusions: LVI shows a close relationship with known markers of poor prognosis as histologic grade, and LN invasion. More aggressive tumors as HER-2 + tumors have a significantly higher probability of LVI, which may be used as an indicator of more aggressive behavior of the primary tumor.

No significant financial relationships to disclose.
Correlation of education, age, and health insurance status with attitudes about and preferences for direct-to-consumer BRCA testing
Authors: Gray S. W. , Smith D. , Karp L. , O'Grady C. , Hornik R. , Armstrong K.
Published: August 2020

e22116

Background: Direct-to-consumer (DTC) cancer genetic testing is increasingly common. However, little is know about peoples' attitudes about or preferences for DTC testing as compared to clinic-based genetic testing. Methods: We surveyed a convenience sample of 257 women without prior BRCA testing/counseling experience about their attitudes about and intentions to get online BRCA testing (90% participation rate). Subjects completed baseline interviews, viewed a modified commercial DTC website and completed an online survey. Results: Sample characteristics: mean age 37 (range 19–71), 80% white, mean education 3 years college, 45% with a household income <$50,000, 3% with a history of breast/ovarian cancer and mean Frank risk 4.7%. 64% of women expressed interest in BRCA testing at baseline and 24% intended to get tested in the next year. After viewing the website, 22% of women said that they would prefer DTC testing, 60% said that they would prefer clinic-based testing and 18% were indifferent. Overall, women believed that DTC testing would be more convenient (p <0.001), offer greater privacy (p <0.01), and be less likely to lead to discrimination (p <0.001) while clinic testing would provide more information (p <0.001) and counseling (p <0.001). After adjusting for confounders, older and less highly educated women had more positive attitudes about DTC BRCA testing (adjusted odds ratio (OR) 4.20, p= 0.02 and OR 10.3, p=0.03 high school (HS) vs. college and OR 16.1, p=0.02 HS vs. graduate school). Women with no health insurance and more positive attitudes about DTC genetic testing were more likely to prefer DTC genetic testing over clinic testing (adjusted OR 3.1, p=0.03 and OR 3.7, p<0.01). There were no associations between race and outcomes. Conclusions: A substantial minority of women expressed a preference for DTC BRCA testing after viewing a modified commercial website. Preferential interest in DTC testing by less educated and uninsured populations makes it especially critical that these sites provide high quality services. Without high quality service, the availability of DTC testing may only exacerbate disparities in genetic services and prevention.

No significant financial relationships to disclose.
High expression levels of putative hepatic stem/progenitor cells biomarkers related to tumor angiogenesis and poor prognosis of hepatocellular carcinoma
Authors: Xu Y. , Fan J. , Yang X. , Zhou J. , Qiu S.
Published: August 2020

e22121

Background: To investigate the prognostic values of putative hepatic stem/progenitor cells (HSCs/HPCs) biomarkers in hepatocellular carcinoma (HCC) patients. Methods: Fourteen biomarkers related with HSCs/HPCs or tumor angiogenesis were assessed by qRT-PCR and then validated by tissue microarrays (TMAs) in three independent cohorts of HCC patients underwent curative resection (n=67, 314 and 73). Results: Most of the biomarkers were found over-expressed in recurrent HCC patients by qRT-PCR. HSCs/HPCs biomarkers cytokeratin 19, ABCG2, CD133, Nestin, CD44 and angiogenesis agents CD34, VEGF and PD-ECGF, were confirmed as significant predictors for overall survival (OS) and/or relapse-free survival (RFS) in TMAs analysis. Compared with the low HSCs/HPCs profile group, patients with high HSCs/HPCs profile had significantly lower OS and RFS (p<0.0001), expressed higher VEGF levels (p = 0.012) and microvessel density (MVD, determined by CD34 immunostaining, p = 0.030). Based on Cox regression, a simplified model including CD133, CD44, Nestin, and MVD was constructed and confirmed as an independent predictor for OS (p<0.0001) and RFS (p<0.0001), regardless of alpha-fetoprotein level, tumor stage and recurrence time (p<0.0001 for all). Conclusions: High expression levels of HSCs/HPCs biomarkers are related to tumor angiogenesis and poor prognosis of HCC. The simplified model based on HSCs/HPCs and tumor angiogenesis profile can be used to classify HCC patients with high risk of tumor recurrence after operation.

No significant financial relationships to disclose.
Prevalence of early-stage prostate and estrogen receptor positive (ER+) breast cancer patients receiving primary androgen deprivation therapy (ADT) and aromatase inhibitors (AIs) in the United States (U.S.)
Authors: Barlev A. , Yong M. , Cherkowski G. , Cetin K. , Fryzek J.
Published: August 2020

e22126

Background: AIs and ADT are used to prevent recurrence of breast and prostate cancers but have been shown to accelerate bone loss. We estimated the prevalence of early-stage ER+ breast and prostate cancer patients on hormone therapy in the U.S., as this is not well-described in the literature. Methods: Data from the Surveillance, Epidemiology, and End Results (SEER) Program, published literature, clinical practice, and a large claims database were used. We began with the American Cancer Society's estimated number of new breast and prostate cancer cases for the year 2008. We then assessed the number of patients with localized/regional disease and ER+ tumors and those receiving primary ADT (both chemical and surgical) or AI therapy by applying proportions from SEER, published literature, clinical practice, and the claims database. Using these incident case counts, we calculated the 5-year prevalence using appropriate cohort-specific survival rates to sum the number of new and surviving cases over a 5-year period. Results: The estimated 5-year prevalence of early-stage ER+ breast cancer for women aged ≥50 years in the U.S. was 607,411, of which 293,904 (48.4%) were on AI therapy based on the claims database. However, because this data source was limited to women aged <65 years, we also used estimates from clinical practice to capture AI use for women of all ages. Based on clinical practice, 402,637 (66.3%) to 460,156 (75.8%) of early-stage ER+ breast cancer patients were on AI therapy. For early-stage prostate cancer, the estimated 5-year prevalence for all ages was 1,024,238, of which 141,451 (13.8%) were on primary ADT. However, these figures may underestimate current usage of hormone therapies, as our data and the literature show increasing trends in ADT and AI use for early-stage disease. Conclusions: Based on a combination of population-based data and the published literature, approximately half of all early-stage ER+ breast cancer patients and a modest proportion of early-stage prostate cancer patients are on hormone therapy in the U.S.

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Regulatory role of p53 in cancer metabolism through SCO2 and TIGAR in human breast cancer
Authors: Choi S. , Song J. , Lim S. , Lee J.
Published: August 2020

e22128

Background: Cancer cells showed higher rate of anaerobic respiration than normal cells. The exact mechanisms for this higher rate of glycolysis in cancer cells remain to be elucidated. Recent researches suggest that p53, the most commonly mutated tumor suppressor gene, might play an important roles in the regulation of energy generating metabolic pathways that switch from oxidative phosphorylation to glycolysis through synthesis of cytochrome C oxidase 2 (SCO2) and TP53-induced glycolysis and apoptotic regulator (TIGAR). Methods: We investigated the expression of p53, SCO2, TIGAR and cytochrome C oxidase (COX) in 95 cases of invasive ductal carcinoma using immunohistochemistry. Results: Overexpression of p53, SCO2, TIGAR and COX was observed in 27.4% (26 cases), 82.1% (78 cases), 76.8% (73 cases), and 77.9% (74 cases) respectively. Overexpression of p53 was significantly associated with decreased expression of SCO2 (p=0.009), COX (p=0.001) and TIGAR (p=0.03). Conclusions: These results suggest that p53 can modulate the metabolic pathways via SCO2 and TIGAR in human breast cancer.

No significant financial relationships to disclose.
Quantitative estimates of the effects of concomitant chemotherapy on acute dysphagia in patients receiving radical treatment for head and neck cancer
Authors: Bhide S. , Gulliford S. , A'Hern R. , Hall E. , Newbold K. , Harrington K. , Nutting C.
Published: August 2020

e22134

Purpose: To generate quantitative parameters describing the effect of concomitant chemotherapy on incidence of grade 3 dysphagia (CTCAE v3.0, assisted feeding) using dose response curves in patients receiving radical treatment for head and neck cancer. Methods: Patients treated at a single centre in prospective phase I and II trials of concomitant chemo-IMRT (CRT) (n=85) and the phase III trial of IMRT vs. conventional radiotherapy (PARSPORT) (n=82) formed the basis of this non-randomized comparison. Patients in the PARSPORT trial received radiation alone (RT). Radiation dose for all patients was radiobiologically equivalent to at least 70Gy in 35 fractions. Concomitant chemotherapy was cisplatin (100 mg/m2) on days 1 and 29. G3 dysphagia was recorded prospectively. Dose volume histograms (DVH) were generated for the pharyngeal mucosa. The mean dose (converted to equivalent dose in 2Gy/fraction, MD2) was used as a univariate descriptor of the DVH, for the generation of the dose response curves. A logistic function of the form p=1/[1+(MD50/D)k] was fitted where, p is the probability of the incidence of toxicity, D is the mean dose, MD50 is the mean dose at which 50% of patients experience toxicity and k describes the increase in incidence with increasing dose. The dose response curves were fitted using non-linear logistic regression. Results: The mean MD2 to the pharyngeal mucosa were 56Gy and 55.8Gy respectively, in the CRT and RT groups. There was a statistically significant difference of 25% (95% CI: 10–38, p=0.002) in the incidence of G3 dysphagia between the CRT (68%) and RT (43%) groups. Fitting dose response curves to the clinical data yielded parameter values (95% CIs) of MD50=46 Gy (42–49), k=4.8 (2.3–7.2) for the CRT group and MD50= 58 Gy (55–61), k=3 (1.6-.45) for RT group. Dose response gradients for CRT and RT showed approximately 1.95% and 1.3% increase (respectively) in probability of G3 dysphagia resulting from an increase in mean dose of 1Gy between doses of 30Gy to 70Gy. Conclusions: Addition of concomitant chemotherapy increases the incidence of G3 dysphagia by 0.65% for every 1 Gy increase in radiation dose. The observed MD50 for G3 dysphagia is lower for RT alone (46 Gy vs. 58 Gy).

No significant financial relationships to disclose.
Novel genes mediating cisplatin resistance in head and neck cancer
Authors: Gasco M. , Syed N. , Coley H. , Colantonio I. , Merlano M. , Crook T. , Crook T.
Published: August 2020

e22135

Background: Chemo-radiotherapy with cisplatin-based regimens offers the possibility of cure to a subset of patients with surgically non-resectable squamous carcinomas of the head and neck (HNSCC), but outcome is frequently limited by acquired drug resistance. We have sought novel genes mediating cisplatin resistance in HNSCC. Methods: We derived in vitro cisplatin resistant variants of the HN5 HNSCC cell line and performed micro-array anaylsis to identify differentially expressed genes. Differences in gene expression were confirmed by qPCR and /or western blotting. Methylation-dependent transcriptional silencing of down-regulated genes was studied by bisulphate sequencing and methylation specific PCR. Selected genes were further analysed in a cohort of stage III and IV HNSCC patients treated with cisplatin-based chemo-radiotherapy. Results: We have identified a panel of genes in which changes in expression occur with acquisition of cisplatin resistance both in vitro and, some cases, in vivo. Up-regulated genes include TAOK1, BZW1 and RECQL, whereas down-regulated genes include FAM83D, PAFAH1B2, DLL1, ABPA1 and FH. Conclusions: We report the identification of a novel panel of genes which function as determinants of cisplatin sensitivity. Analysis of expression and/or epigenetic regulation of these genes may have clinical utility in prediction of patients likely to respond to highly toxic combined modality chemo-radiotherapy.

No significant financial relationships to disclose.