bluuebunny/crossref_metadata_2025 huggingface dataset:
Practice patterns in the use of adjuvant therapy for post-menopausal early-stage breast cancer in the pre- and post-Oncotype DX era.
Authors: Latif Asma , Small Alexander C. , Moshier Erin L. , Adelson Kerin B. , Raptis George , Godbold James H. , Oh William K. , Galsky Matt D.
Published: March 2020

e21032

Background: Personalized oncology offers the promise of selectively applying therapeutics to patients most likely to benefit, while sparing those unlikely to benefit from potentially toxic therapies. Oncotype DX is a 21-gene assay utilized to identify hormone-receptor positive (HR+), node negative, breast cancer (Br CA) patients who may be successfully treated with adjuvant hormonal therapy alone. We hypothesized that practice patterns with adjuvant therapy have changed since the commercial availability of Oncotype DX in 2004. Methods: The Public National Cancer Database was queried to identify patients age ≥ 50 with stage I or II Br CA diagnosed from 2000 to 2008. Patients were classified by adjuvant therapy including hormone, chemotherapy, hormone and chemotherapy, and no hormone or chemotherapy. Log-binomial regression was used to estimate prevalence ratios for the proportion of patients receiving adjuvant therapies from 2000-2003 compared to 2004-2008. Results: 833,018 patients age ≥ 50 with stage I or II Br CA were identified. The application of adjuvant therapies for the periods pre- and post-availability of Oncotype DX are detailed in the Table. Conclusions: There has been significant increase (13%) in the use of hormonal therapy alone as adjuvant therapy for patients age ≥ 50 with HR+ stage I-II Br CA since commercial availability of Oncotype DX. While this has been slightly offset by a decrease in the use of chemotherapy plus hormonal therapy, there has been a larger decrease in the use of “no adjuvant therapy”. Data regarding predictive biomarkers should be captured by registries in an effort to determine the true impact of these tests on treatment utilization. [Table: see text]
HER3 expression in the primary tumor as marker of metastatic potential in breast cancer patients.
Authors: Berghoff Anna Sophie , Bago-Horvath Zsuzsanna , Preusser Matthias , Steger Guenther G. , Birner Peter , Zielinski Christoph , Bartsch Rupert
Published: March 2020

e21033

Background: Breast cancer (BC) is the most common cancer related cause of death in women. Therefore, identification of novel prognostic markers is an important field of research. Recently, HER3 expression was postulated as risk factor for metastatic spread to the brain. We aimed to analyze the predictive value of HER3 expression in patients with metastatic BC treated between 1993 and 2010 at the Medical University of Vienna. Methods: Patients of each subtype (luminal, HER2 positive, triple negative) with metastatic breast cancer were identified from a breast cancer data base. Tissue of the primary tumor was raised from a breast cancer bio-bank. Immunohistochemical staining of estrogen-receptor, progesterone-receptor, HER2 and HER3 was performed. Chi2 test was used to correlate two factors. Overall survival (OS) and time to progression (TTP) were estimated using the Kaplan-Meier method. Results: A total of125 patients (39 luminal, 42 HER2 positive, 44 triple negative) with metastatic BC were available for this analysis. 71 patients (56.6%) had visceral metastases and 67 patients (46.4%) had brain metastases. 7.4% of specimens showed a positive, uniform, intense membranous staining of > 30% of invasive tumor cells (Her3 3+). Less intense, incomplete, membranous staining was observed in a further 34.4%. HER3 expression in the primary tumor did not correlate with BC subtype (luminal 12.8%; HER2 positive 2.4%; triple negative 6.8%; n.s.; Chi2 test). Furthermore, no correlation with the likelihood of developing brain metastases (5% vs. 9.8%; n.s.; Chi2 test) or visceral metastases (7.3% vs. 7.7%; p:n.s.; Chi2 test) was found. Counter intuitively, patients with HER3 overexpression in the primary tumor had a significantly longer TTP (21 vs. 47 months; p=0.03; log rank test) and a trend towards longer OS was observed as well (53 vs. 112 months; p=0.073; log rank test). Conclusions: In contrast to previous reports, we could not validate that HER3 expression predicts for metastatic potential in general and also with regard to brain metastases of BC in a population that was equally distributed between BC subtypes. Further studies are therefore needed to evaluate the prognostic value HER3 expression in BC.
Retrospective study of systemic chemotherapy in combination with consolidative stereotactic body radiation therapy (SBRT) in patients with five or fewer oligometastases from breast, colorectal, and lung cancers.
Authors: Ulhoa-Cintra Alice , Monga Dulabh K. , Kirichenko Alexander V.
Published: March 2020

e21036

Background: The oligometastatic state is one of limited metastases, amenable to local therapies of curative potential. Stereotactic body radiation therapy (SBRT) is a non-surgical treatment with a primary goal of rapid local tumor control and low toxicity allowing prompt return to systemic therapy. Breast (BRC), colorectal (CRC) and non-small cell lung cancers (NSCLC) have oligometastases as first presentation. In these patients, consolidative SBRT offers effective local control and low toxicity, with the potential for improved survival benefit without affecting quality of life. We report our experience on such patients. Methods: 31 patients with BRC (9), CRC (16) and NSCLC (6) with ≤ 5 unresectable metastases to one to three organ sites were identified. Data were obtained from chart review. Primary end point was infield local control. Lesions with at least 6 months of radiographic follow-up were assessable. Secondary end points were toxicity and survival. Results: All patients completed SBRT to 38 lesions (≤2/patient). 29 received prior chemotherapy for metastatic disease. 14 were treated with at least 1 prior regimen and 15 at least 2 (0-8). At a median follow-up of 16 months (4-32), 22 patients (8 BRC, 10 CRC and 4 NSCLC) were assessable. At 6 months, 20 patients had infield local control and 3 demonstrated distant metastases. At 1 year, this was seen in 19 and 7 patients respectively. The most common toxicity was grade ≤2 fatigue. All BRC had infield local control at 6 months and 1 year. Among CRC, 9 had infield local control at 6 months and 7 at 1 year. 3 NSCLC patients had infield local control at 6 months and 1 year. No BRC demonstrated distant progression at 6 months; at 1 year, it was seen in 2 patients. 3 CRC had evidence of distant progression at 6 months and 1 year. 1 NSCLC patient had distant progression at 6 months and 2 at 1 year. 6 patients died from their underlying malignancy; the median survival for 5 of these patients was 20 months (5-34). Conclusions: The combination of systemic chemotherapy and consolidative SBRT is safe and effective for patients with oligometastatic disease
Modulation of angiogenic biomarkers in patients receiving high-dose TRC105.
Authors: Liu Yingmiao , Starr Mark , Brady John C , Pang Herbert , Dellinger Andrew , Leigh Bryan R. , Theuer Charles P. , Hurwitz Herbert , Nixon Andrew B.
Published: March 2020

e21038

Background: TRC105, an anti-CD105 monoclonal antibody, has completed phase 1 testing and is being studied in multiple phase 2 trials. We previously reported that low dose TRC105 (0.01-1.0 mg/kg) modulated the expression of soluble angiogenic biomarkers in patients [J Clin Oncol 29: 2011 (suppl; abstr 10565)]. In this report, we evaluated angiogenic biomarkers in patients receiving higher doses of TRC105 including the recommended phase 2 dose. Methods: Patients with advanced refractory solid tumors were treated with escalating doses of TRC105 until disease progression. Serial plasma samples were analyzed via an optimized multiplex ELISA platform. 36 biomarkers related to tumor growth, angiogenesis, and inflammation were assayed at baseline (BL), after 1 month (C2D1), concurrent with radiological restaging near the end of cycle 2 (C2D22), and at end of study (EOS). Results: 32 patients treated with TRC105 at doses of 0.3 to 15 mg/kg were evaluated for biomarker expression. Wilcoxon signed rank tests indicated that the following analytes were significantly different at C2D1 when compared with baseline (p<0.05): Ang-2, IGFBP-3, total PAI-1, PDGF-AA, PDGF-BB, TSP-1, VEGF-D were all down-regulated; and E-Cadherin, soluble CD105, E-Selectin, IL-6, OPN, TSP-2, vWF were all up-regulated. At EOS, significant increases from C2D1 were observed for the following analytes: Ang-2, CRP, ICAM-1, IGFBP-1, IL-6, TSP-2, and VCAM-1 (p<0.05). Additionally, dose-dependent increases in soluble CD105 at C2D1 were observed (p<0.0001, r=0.9), suggesting direct target modulation by TRC105. Conclusions: TRC105 therapy is associated with early down-regulation of certain key angiogenic biomarkers and a dose-dependent increase in sCD105. Increases in many initially down-regulated angiogenic factors were observed at the time of progression, suggesting possible mechanisms for acquired resistance to TRC105. Further analysis of TRC105 in specified patient populations is underway and may provide more insight into the biological effects of this drug.
Prognostic impact of circulating endothelial cell counts in patients with advanced non-small cell lung cancer.
Authors: DeJoubner Nutan Jyothi , Kang Hyunseok , Li Qunna , Harris Wayne A C , Stewart Rena , Price Alicia , Quyyumi Arshed , Kono Scott A. , Ramalingam Suresh S. , Owonikoko Taofeek Kunle , Khuri Fadlo Raja , Waller Edmund K.
Published: March 2020

e21045

Background: Blood progenitor cells expressing the stem cell marker CD34 have been associated with prognosis in cancer. We hypothesized that non-small cell lung cancer (NSCLC) patients with higher numbers of circulating CD34+ endothelial progenitor cells (CD34+/CD146+/CD45-, CEC) would have worse post-treatment outcomes and patients with more hematopoietic progenitor cells (CD34+/CD45+/CD45dim/CD133+, HPC) would have better outcomes. Methods: Blood samples from patients with advanced NSCLC were analyzed at 3 time points: prior to chemotherapy, after cycle one, and at completion of treatment or progression of disease, in an IRB-approved protocol. CEC, HPC, and immune subsets were measured by flow cytometry optimized for rare event detection. Primary outcomes were progression and death from the time of study entry. Patients were categorized and compared for progression free survival (PFS) and overall survival (OS) based on median cell counts. Differences between groups were tested using log-rank test and Cox regression. Results: A total of 62 patients were enrolled with mean age of 64 (37-87 years). There were 29 deaths after a median follow-up of 8.1 months. 40 patients progressed at a median of 5.4 months. Median numbers of CEC, HPC, and CD4+ T-cells were (0.05/uL, 0.41/uL, and 637/uL, respectively). Lower CEC at baseline was associated with a favorable PFS compared to those with higher counts (median PFS of 7.3 months versus 4.3 months, p=0.049). In a multivariate analysis after adjusting for age, gender, smoking, race, TNM stage, pathology, performance status and CD4 counts at baseline, patients with high CEC had almost 3-fold risk of death (HR=2.70, p=0.04) and disease progression (HR=3.03, p < 0.01) compared with patients with low numbers of CEC. In the same model, patients with fewer CD4+ T cells had > 3 fold risk of death compared with patients with higher numbers (HR=3.79, p=0.01). HPC content was not associated with OS or PFS. Conclusions: In patients with advanced NSCLC, higher CEC and lower CD4 counts at diagnosis are associated with worse outcomes. Higher CEC may serve as a cellular biomarker for extent of disease or tumor biology. Patients with poor T-cell immunity prior to treatment have worse survival.
Variability of gene expression profile in breast cancer core biopsy: Impact on prognostic and predictive multigene signatures.
Authors: Jarzab Michal , Bal Wieslaw , Kowalska Malgorzata , Oczko-Wojciechowska Malgorzata , Kowal Monika , Rembak-Szynkiewicz Justyna , Dyla Alicja , Chmielik Ewa , Stobiecka Ewa , Bobek-Billewicz Barbara , Tarnawski Rafal
Published: March 2020

e21049

Background: Gene expression profiling has been shown to be effective in the analysis of postoperative tumor samples in breast cancer. However, when small specimens like core biopsy samples are considered, the small size of the collected material makes it difficult to obtain high tumor percentage in the specimen. Thus, we tested the strategy of obtaining three independent cores for molecular analysis and assessed the influence of the stability of different prognostic and predictive signatures. Methods: We prospectively collected breast core biopsies from 82 consecutive patients treated by pre-operative chemotherapy. In each patient, three cores from different tumor areas were sampled for molecular testing, independent of the cores collected for histopathological analysis. RNA was isolated by QIAGEN RNeasy kits, gene expression profiling was carried out using Affymetrix HG-U133 Plus 2.0 microarrays. Results: In 27 patients, three independent specimens were analyzed by microarray gene expression profiling (in total 81 samples). When this training set was analyzed by unsupervised Principal Component Analysis, the overall concordance between the three tumor cores was observed, but no such concordance was seen for three tumors (for each with one significant outlier). When we assessed the set of chosen 20 prognostic and predictive signatures, we confirmed that the homogeneity of gene expression within the signatures is significantly better than for random sets of genes, but we observed the notable differences between the signatures. The heterogeneity decreased when genes with low between-patient variance were excluded; they also showed a higher intra-patient variability of gene expression. We also assessed the heterogeneity of single expression markers and furher validated our findings on an independent set of 55 patients. Conclusions: The use of three independent cores for gene expression testing in breast cancer may be a successful strategy to overcome tumor heterogeneity and sampling error and result in more stable results of prognostic and/or predictive signatures. Supported by Polish National Science Center grant N402 6861 40
Determining the metastatic potential of circulating tumor cells.
Authors: Menen Rhiana , Pinney Emmett , Kolostova Katarina , Bobek Vladimir , Suetsugu Atsushi , Zhang Nan , Hoffman Robert M.
Published: March 2020

e21051

Background: Knowledge of the metastatic potential of circulating tumor cells would be useful for cancer prognosis and also be a rationale for targeting metastatic CTCs for therapy. Methods: Using immunomagnetic beads, CTCs were rapidly isolated from the circulation of mice orthotopically implanted with human PC-3 prostate cancer cells stably expressing green fluorescent protein (GFP). The PC-3–GFP CTCs were then expanded in culture in parallel with the parental PC-3–GFP cell line. Both cell types were then inoculated onto the chorioallentoic membrane (CAM) of chick embryos. Eight days later, embryos were harvested and the brains were processed for frozen sections. The IV-100 intravital laser scanning microscope enabled rapid identification of fluorescent metastatic foci within the chick embryonic brain. Results: Inoculation of embryos with PC-3–GFP CTCs resulted in a 3 to 10-fold increase in brain metastasis when compared to those with the parental PC-3–GFP cells (p<0.05 in all animals). Thus, PC-3–GFP CTCs have increased metastatic potential compared to their parental counterparts. Conclusions: The chick embryo represents a rapid, sensitive, imageable assay of metastatic potential for CTCs. The chick embryo assay has future clinical application for individualizing patient therapy based on the metastatic profile of their CTCs.
Effect of orally administered S-1 on circulating endothelial cells in patients with metastatic breast cancer.
Authors: Tsuji Wakako , Ishiguro Hiroshi , Ueno Takayuki , Toi Masakazu
Published: March 2020

e21052

Background: S-1 is an oral cytotoxic preparation combining tegafur, 5-chloro-2, 4-dihydroxypyridine, and potassium oxonate at a molar ratio of 1:0.4:1, respectively. Tegafur is metabolized into 5-fluorouracil (5-FU), γ-butyrolactone (GBL), and γ-hydroxybutyric acid (GHB). GBL and GHB are known to suppress VEGF-mediated angiogenic activity. Circulating endothelial cells (CECs) have been suggested to be useful for predicting response to anti-angiogenic agents. We evaluated the effects of S-1 on CEC counts in patients with metastatic breast cancer. Methods: Patients with metastatic breast cancer received S-1 orally in 28-day cycles. The dose of S-1 was 80-120 mg/day depending on body surface area. Cycles were repeated every 42 days. CEC counts were measured with the CellSearch system (Veridex, LLC Co. Ltd., NJ) on days 1 (before the 1st cycle of S-1), 15, 29, 43 (before the 2nd cycle of S-1), and 85 (before the 3rd cycle of S-1). Plasma GBL and 5-FU levels were measured by a gas chromatographic-mass spectrometric method at one point each on days 1 (before the 1st cycle of S-1), 15 (1 hour after S-1 administration), 43 (before the 2nd cycle of S-1), and 57 (1 hour after S-1 administration). VEGF levels on days 1, 15, 43, and 57 were measured by an enzyme-linked immunosorbent assay. Results: A total of 18 patients were enrolled. Plasma GBL levels on days 15 and 57 were 42.0 ± 16.2 ng/mL and 43.2 ± 10.5 ng/mL, respectively. Both levels exceeded 25.8 ng/mL, the previously reported (Angiogenesis 2001). Both CEC and CD34+CEC levels significantly decreased on day 15, and low levels were maintained low until day 85. Plasma levels of VEGF also decreased similarly, but not significantly. Conclusions: The results of this exploratory study suggest that S-1 not only has cytotoxic effects on tumor tissue, but also anti-angiogenic activity in patients with metastatic breast cancer.
In vivo real-time imaging of tumor microcirculatory alterations in colorectal liver metastatic xenografts by chemotherapy using multiphoton microscopy.
Authors: Tanaka Koji , Okigami Masato , Toiyama Yuji , Morimoto Yuhki , Matsushita Kohei , Kawamura Mikio , Hashimoto Kiyoshi , Saigusa Susumu , Okugawa Yoshinaga , Hiro Junichiro , Inoue Yasuhiro , Uchida Keiichi , Araki Toshimitsu , Mohri Yasuhiko , Mizoguchi Akira , Kusunoki Masato
Published: March 2020

e21053

Background: Tumor endothelium is a distinct target for anticancer treatments of metastatic colorectal cancer. Various chemotherapeutics themselves have anti-angiogenic effects on tumor microenvironment by targeting proliferative endothelial cells during tumor angiogenesis. In this study, we imaged the dynamics of tumor microcirculation of colorectal liver metastasis in living mice in vivo real-time using two-photon laser scanning microscopy (TPLSM), and evaluated the microcirculatory alterations in tumor microenvironment by chemotherapy. Methods: Red fluorescent protein expressing human colorectal cancer cell line (HT29) was inoculated to the spleen of green fluorescent protein expressing nude mice. 5-fluorouracil or irinotecan was administered three times a week for more than three weeks for metronomic scheduling. Intravital TPLSM was performed at multiple time points for time-series imaging of liver metastatic xenografts in the same mice. The alterations in tumor microcirculation during chemotherapy was evaluated by measuring blood flows at tumor vessels of colorectal liver metastasis and hepatic sinusoids of adjacent normal liver. Results: At the first TPLSM imaging, the blood flow, as determined from the movement of platelets, was heterogeneous and non-directional in the tumor vessels of liver metastatic xenografts. The blood flow was relatively slower in tumor vessels than normal hepatic sinusoids. At the second TPLSM imaging after chemotherapy, platelet aggregation was observed in tumor vessels of the same mice. Aggregated platelets were frequently adhered to the tumor endothelium, suggesting tumor vessel damage or intratumoral coagulation abnormality by chemotherapy. There was no difference in chemotherapeutics with regard to these findings. Conclusions: Intravital TPLSM imaging of tumor microcirculation at metastatic tumor xenografts is a useful tool to evaluate anti-angiogenic drugs in preclinical models.
Expressions of KLF2 and chemokine receptor CCR7 in pulmonary adenocarcinoma.
Authors: Itakura Meiji , Shingoji Masato , Iizasa Toshihiko
Published: March 2020

e21054

Background: Chemokines and chemokine receptors not only have the powerful ability in cancer metastasis and tumorigenesis, but also act as anti-tumorgenic ability. Lung Krueppel-like factor (LKLF, KLF2) is a member of the family of the Krueppel-like factors (KLFs). KLF2 was initially described as a lung-specific transcription factor. KLF2 is reported to regulate some malignant cells. We examined and evaluated the effect of KLF2 on pulmonary adenocarcinoma and the relationship of their mRNA expression with CCR7, EGFR and p53 genetical mutations in pulmonary adenocarcinoma. Methods: 120 patients of stage I to IV with pulmonary adenocarcinoma were included in this retrospective analysis. The expression of CCR7 and KLF2 mRNA expression in surgically resected pulmonary adenocarcinoma specimens were examined and evaluated the relation to prognosis, the effect of EGFR and p53 genetical mutations. Results: High mRNA expression of KLF2 in lung cancer patients indicated significantly good prognosis than the groups of low expressions (p= 0.0066, HR= 2.008, 95% CI of ratio 1.215 to 3.319). The expression of KLF2 mRNA had relationships with CCR7 mRNA expression in pulmonary adenocarcinoma. Moreover the mRNA expression of KLF2 in pulmonary adenocarcinoma specimens was influenced by the mutation of p53 mutation in lung cancer specimens. There was no significant difference within KLF2 mRNA expression quantities of EGFR mutation positive and negative group. Conclusions: We propose KLF2 as clinical good prognostic factors and that KLF2 has strong relation with CCR7 and p53 genetical mutation mRNA expression in pulmonary adenocarcinoma.
ApoStream, an antibody-independent platform, compared to CellSearch for enumeration of circulating tumor cells (CTCs) in patients with metastatic prostate cancer.
Authors: Poklepovic Andrew Stewart , Wan Wen , Wu Weiguo , Woo Jacky , Melnikova Vladislava , Jouravleva Elena , Anderes Kenna Lynn , Davis Darren
Published: March 2020

e21058

Background: Development of CTC detection platforms is a rapidly advancing field. The CellSearch technique relies on cell surface expression of EpCAM to select for rare tumor cells in whole blood specimens. Consequently, the use of EpCAM-based enrichment platforms to recover CTCs is limited to EpCAM expressing cells and is poorly suited for recovery of EpCAM negative cells like those having undergone epithelial mesenchymal transition (EMT). ApoStream is an antibody independent enrichment platform which utilizes the principle of continuous flow dielectrophoresis field-flow fractionation (DEP-FFF) to isolate and enrich for CTCs. A head to head comparison of CellSearch to ApoStream for recovery of CTCs in patients with metastatic prostate cancer was performed. Methods: Two 7.5 mL bloodsamples were collected at a single time point for each patient with stage IV prostate cancer. One sample was analyzed by CellSearch CTC enumeration kit, and one sample was analyzed by ApoStream. CTCs recovered by both devices were immunophenotyped using antibodies against cytokeratin (CK), CD45 and DAPI. CTCs were defined as CK+/CD45-/DAPI+ intact cells. CTCs recovered by ApoStream were further analyzed by quantitative laser scanning cytometry (LSC). A paired t-test was used to compare the cell counts in the two devices. Results: The ApoStream CTC enrichment platform isolated a greater number of CTCs from blood of eight patients compared to CellSearch (p= 0.0027). All cell counts obtained by the ApoStream technique were higher than CellSearch, and all patients had detectable CTCs by ApoStream, while only 75% of patients had detectable CTCs with CellSearch. Conclusions: The ApoStream platform is differentiated from EpCAM dependent platforms and is well suited for detection and recovery of CTCs in advanced stage disease where tumor cell heterogeneity is common and expression of EpCAM may be low or lost. [Table: see text]
The novel receptor targeted (CD206) 99mTc-labeled tilmanocept versus the currently employed Tc99m-sulfur colloid in intraoperative lymphatic mapping (ILM) on key performance metrics breast cancer.
Authors: Cope Frederick Oliver , Metz Wendy , Potter Beth , Abbruzzese Bonnie , Blue Michael , Shuping Jo , Christman Lori , King Dennis
Published: March 2020

e21066

Background: ILM is a key diagnostic component for solid tumor excision. ILM utilizes the injection of a color agent and/or a radiopharmaceutical agent (RPA) which may be tracked intraoperatively w/ a γ-probe. ILM is designed to aid the surgeon in tracing lymph nodes that may have a connection to the tumor bed. ILM in the U.S. employs a non-standard RPA, S-colloid (TcSC). This agent was recently approved by the FDA for breast cancer ILM using a literature review. In contrast, CD206-targeted Tc99m-tilmanocept (TCP) was studied in 2 prospective Ph-3 trials. These trials compared TCP to blue dye, the same dye utilized in the FDA review of TcSC. Methods: For a statistical review and contrast of TCP vs TcSC, we used a meta-analysis and pooled analysis based on the FDA-review of TcSC. We focused on the functional endpoints: 1] Localization rate (% of pts w/ a found hot node) and 2] degree of localization (# of found hot nodes/pt). Both metrics define the potential for agent performance for ILM and nodal metastasis. Results: The contrast of the localization rate/pt for TcSC at 94% was based on TcSC’s reviewed value. The results for the proportion of TCP patients were localization rates of 99.91% by meta-analysis and 98.65% by pooled analysis; p<0.0001 and p<0.008, repsectively (1-sided test, H0 p=/< 0.94). The contrast of the # hot nodes/pt. This value was benchmarked at 1.6 nodes/pt based on TcSC’s review value. The results for the nodes/pt of TCP patients were 2.08 by meta-analysis and 2.16 by pooled analysis; p<0.0001 and p<0.0001, repsectively (1-sided test, H0 p=/< 1.6). Conclusions: TCP provided significantly greater performance in key metrics, node localization/pt, and degree of localization (nodes found/pt) in breast cancer. All values are highly statistically significant for the key clinical performance parameters. These data suggest that receptor targeted TCP may provide improved diagnostic ILM over the currently employed nonspecific RPA lymphatic mapping agent TcSC.
Effect of matrix metalloproteinase-2 on CD8+ T cell and NK cell responses.
Authors: Gallois Anne , Godefroy Emmanuelle , Manches Olivier , Bhardwaj Nina
Published: March 2020

e21081

Background: Matrix metalloproteinase 2 (MMP-2) cleaves many components of the extracellular matrix and is over-expressed in several cancers including melanoma. High levels of MMP-2 expressed by tumors are associated with later tumor stages, increased dissemination and poorer survival/prognosis. We recently identified a pathway whereby MMP-2 functions as a human endogenous “conditioner” that skews CD4+ T cells towards a detrimental/inefficient TH2 phenotype through OX40L expression and inhibition of IL-12p70 production. We unraveled the underlying mechanism: MMP-2 degrades the type I IFN receptor (IFNAR1), thereby preventing STAT1 phosphorylation necessary for IL-12 production. Here, we described IFNAR1 as a novel MMP-2 substrate. This finding opens the way to characterize a possible new immunosuppressive effect of MMP-2 on other IFNAR1+ immune cells such as NK cells and CD8+ T cells. Methods: CD8+ T cells and NK cells were purified from PBMCs and incubated with MMP-2 overnight. IFNAR1 expression was assessed by FACS and Western Blot. Granzyme B, TNF alpha and IFN gamma expression was determined by intracellular staining of memory T cells. NK cells cytotoxicity was measured by Lamp-1 expression after 4h of coculture with K562 cells. Results: We first found that IFNAR1 is expressed by NK and CD8+ T cells and confirmed that MMP-2 degraded it. Memory CD8+ T cells significantly down-regulate granzyme B expression in the presence of MMP-2. Furthermore, memory CD8+ T cells pre-incubated with MMP-2 were significantly impaired with respect to IFN-gamma and TNF-alpha production. Finally, our data suggest that NK cells’ cytotoxicity might also be affected by MMP-2 through modulation of Granzyme B production. Conclusions: Our data strongly suggest that MMP-2 regulates memory CD8+ T cells not only by down regulating cytolytic activity through inhibition of granzyme B, but also by inhibiting cytokine production such as IFN-gamma and TNF-alpha. Elucidating the underlying mechanisms of MMP-2-dependent immune regulation should lead to the development of innovative immune therapies bypassing tumor escape to treat cancer patients.
Enhancement of human cancer cell migration and invasion by anaphylatoxin C5a viaaberrantly expressed C5a-receptor.
Authors: Nitta Hidetoshi , Takamori Hiroshi , Imamura Takahisa , Baba Hideo
Published: March 2020

e21084

Background: Accumulated evidence has revealed implications of chemokines and their receptors in the pathophysiology of cancer. Anaphylatoxin C5a, a fragment of the complement 5th component (C5), is a potent chemokine for leukocytes that express the C5a receptor (C5aR). Recent reports indicate production of C5a by proteases of various origins, besides a by-product of activation of the complement system, and expression of C5aR in non-myeloid cells, suggesting an involvement of C5a in these cell biology. However, little is known about roles of C5aR in human cancer. Aim of our study was to investigate the expression and the role of C5aR on cancer cells in proliferation and invasion. Methods: Surgically resected cancer tissues and cancer cell lines were investigated C5aR expression. The effect of C5a on cancer cell proliferation, cell shape and invasion activity were examined in vitro and in vivo. Results: By immunohistochemical analysis, C5aR was strongly expressed in human cholangiocarcinoma samples. Using RT-PCR, immunoblotting and flow cytometric analysis human cholangiocarcinoma cell line MEC but not HuCCT1 were found to express C5aR. Incubation of both cancer cells with C5a did not increase these cells, indicating no mitogenic effect of C5a on these cells. Interestingly, C5a stimulated invasion of MEC and C5aR-transgenic HuCCT1 when examined by Matrigel invasion assay, which was suppressed by anti C5aR antibody. F-actin staining and human MMPs array revealed that C5a induced actin polymerization and MMPs production, respectively, in MEC and C5aR-transgenic HuCCT1. In contrast, these C5a effects were not seen in HuCCT1. Conclusions: These results illustrate a novel activity of C5a-C5aR axis that promotes cancer cell invasion through cytoskeletal rearrangement and MMP release. Thus, targeting this signaling pathway may provide a useful therapeutic option for cancer treatment.
Identification of new liver tumor biomarkers using proteomics.
Authors: Maimonis Peter J. , Zen Yoh , Britton David J. , Brand Andrew , Ward Malcolm , Pike Ian , Quaglia Alberto
Published: March 2020

e21091

Background: It is crucial to identify new biomarkers that help diagnose cancers at an early stage and contribute to the development of new anti-cancer drugs. Here, we identify new liver tumor biomarkers using proteomics. Methods: We evaluated 9 types of liver tissue from 55 patients: normal liver parenchyma (n=7), hepatocellular carcinoma (HCC) (n=7), normal bile duct (n=6), peripheral cholangiocarcinoma (CC) (n=7), hilar CC (n=7), CC associated with primary sclerosing cholangitis (PSC) (n=7), metastatic colorectal cancer (n=7), and mixed HCC/CC after trans-arterial chemoembolization (n=7; areas of HCC and areas of CC separately examined). Protein extracted from microdissected, formalin-fixed paraffin-embedded tissue (0.15 mm3 in total) was reduced, alkylated and digested with trypsin in a stacking gel. Peptides were analyzed using nano-liquid chromatography-Mass spectrometry. The raw data were searched using Mascot. Normalized spectral counts for each protein among each tissue type were compared. For each comparison, an unrelated t-test was computed to obtain the p value. q values (adjusted p values) were computed using a direct False Discovery Rate approach (q < 0.05 considered statistically significant). Results: The mean number of proteins identified per sample was 762 +/- 119 S.D, resulting in a total of 2643 proteins identified. Numbers of proteins with significantly different expression levels among different tissue types are shown below. Some of these proteins are being reported here for the first time in the context of liver carcinogenesis. Conclusions: This study revealed proteins that are significantly over/underexpressed in particular types of liver tumors. Clinically useful new biomarkers may be present among these proteins and are now undergoing validation using immunohistochemisty. [Table: see text]
18F-fluorothymidine (FLT) PET-CT for early response assessment in advanced pancreatic cancer.
Authors: Challapalli Amarnath , Wasan Harpreet S , Al-Nahhas Adil , Aboagye Eric , Coombes Charles , Sharma Rohini
Published: March 2020

e21093

Background: Advanced pancreatic cancer has a poor prognosis with a median survival of 6-10 months. There is a need for early non-invasive assessment of treatment response. We evaluated FLT PET-CT combined with a kinetic spatial filtering method (FLT-PETKSF) for detecting response to gemcitabine-based chemotherapy in advanced pancreatic cancer. Methods: Dynamic FLT PET-CT data were collected from patients with confirmed locally advanced or metastatic pancreatic cancer before and 2 weeks after the first cycle of chemotherapy. Changes in tumor FLT-PET variables with treatment were determined. Standardized uptake value (SUV) reduction of 18% was taken as cut-off for response. Voxel quantification of each tumor volume was performed on the filtered data. Each voxel-intensity was normalised by injected dose, body weight and decay corrected to obtain the SUV for the voxel. Changes in high intensity voxels (HiVox: SUV ≥ 2) - were computed. Results: Results of the first 5 patients are discussed. There were 4 primary and 9 metastatic tumors. FLT-PETKSF improved tumor-to-background ratio and enabled visualisation of all the primary and metastatic tumors. The mean (± SD) average and maximum SUV at 60 min (SUV60, av & SUV60, max) of the primary lesions was 2.10 (±0.38) & 4.85 (±1.55) and that of the metastatic lesions was 3.74 (±1.49) & 6.90 (±2.05) respectively. The mean (± SD) percentage reduction in the SUV60, av & SUV60, max, HIVox was 26 (±44), 18 (±38) and 23 (±50) respectively. The changes in the voxel occurrences correlated strongly with the changes in both SUV60, av & SUV60, max (Pearson r-0.9, p=0.001) .Overall, there were 2 partial responders and 3 with stable disease. These responses concurred with response evaluation on mid-treatment CT scan. Conclusions: FLT-PET and FLT-PETKSF enables visualisation of the pancreatic tumors and the liver metastases, and could be used to monitor response to therapy.
Clinical significances of ErbB family receptors, Myc, and mitogen-activated protein kinase (MAPK) in patients with early-stage non-small cell lung cancer.
Authors: Ludovini Vienna , Bellezza Guido , Bianconi Fortunato , Ferri Ivana , Pistola Lorenza , Flacco Antonella , Baldelli Elisa , Chiari Rita , Minotti Vincenzo , Tofanetti Francesca Romana , Siggillino Annamaria , Ragusa Mark , Vannucci Jacopo , Puma Francesco , Sidoni Angelo , Crinò Lucio
Published: March 2020

e21094

Background: EGFR deregulation has been extensively studied in non small-cell lung cancer (NSCLC), but less is known about the expression and role of other ErbB receptors and their downstream signal transductions. Myc and MAPK are key downstream components of the EGFR pathway and have significant roles in cell survival, proliferation, and growth. The purpose of this study is to determine the prognostic value of EGFR, ErbB2, ErbB3, ErbB4, Myc and MAPK by immunohistochemistry (IHC) in early stage NSCLC. Methods: 109 NSCLC were evaluated: median age was 67 years (range 40–84); Male/Female:93/16; squamous (SCC)/adenocarcinoma (ADC)/BAC/other: 52/36/3/18; smoker/never smoker:100/9, and stage I/II/III:67/17/25. The tumors with ≥10% positive cells were classified positive, further confirmed by Receiver Operating Characteristic (ROC) analysis. Kaplan-Meier estimates of survival and time to recurrence were calculated for clinical and biologic variables using Cox model for multivariate analysis Results: EGFR and EbrB3 were associated with SCC (p<0.0001 and p=0.004, respectively) whereas ErbB2 and Myc with ADC (p=0.004 and p<0.0001, respectively) . EGFR and ErbB3 were significantly associated (p=0.003), as well as MAPK and ErbB4 (p=0.02). At a median follow-up of 75 months the contemporary overexpression of EGFR, ErbB2 and MAPK was associated with shorter disease free survival (DFS) (HR=5.4, p=0.002) and overall survival (OS) (HR=8.9, p<0.0001). At multivariate analysis adjusting for stage, the co-expression of EGFR, ErbB2 and MAPK was an independent predictor for worse DFS and OS (HR=5.7, p=0.004; HR=8.67, p<0.001, respectively). Conclusions: Our results suggest that in early stage NSCLC, the co-expression of EGFR, ErbB2 and MAPK predicted a worse prognosis. ErbB3, ErbB4 and Myc were not prognostic factors. Such features may have important implications for future targeted therapies. We thank Italian Association for Cancer Research (AIRC) for supporting the study.
The value of RCAS1 as a potential biomarker in non-small cell lung cancers.
Authors: Tsoukalas Nikolaos , Kostakis Ioannis , Siakavellas Spiros , Tolia Maria , Papakostidi Aristoula , Karameris Andreas , Tzovaras Alexandros , Sfiniadakis Ioannis , Tsiambas Evangelos , Ardavanis Alexandros , Manolis Evangelos , Kittas Christos , Theocharis Stamatios
Published: March 2020

e21098

Background: RCAS1 (Receptor-binding Cancer Antigen expressed on SiSo cells) is a membrane protein that is expressed in different types of cancer. It halts the cell cycle and/or induces the apoptosis of the immune system cells within the tumour microenvironment. Hence, it is possible that this molecule is involved in the mechanism of the tumour cells’ escape from the immune system surveillance (immunoescape). Methods: Patients with primary non small cell lung cancer, initially eligible for surgical treatment, were included. The tissue samples (paraffin cubes) were processed using the Tissue Micro-arrays Method. Immunohistochemical study was done with specific monoclonal antibodies for RCAS1 and Ki-67. The image analysis was feasible due to a special program (image analysis software). In addition, a database was created that included the clinical and pathological characteristics of the patients. After the surgery patients received chemotherapy and/or radiotherapy according to the guidelines. Associations between variables were analyzed with non-parametric tests (Mann-Whitney and Kruskal-Wallis test) and the survival analysis with the Cox proportional hazards model. Two tailed p values ≤ 0.05 were considered to be statistically significant. Results: In total, 112 patients were examined (93 men and 19 women), mean age 63,6 years old. Almost 53% of the cases were adenocarcinoma, 33% squamous cell, 9% large cell, and 5% other types. Statistical significance was identified correlating RCAS1 over-expression to lower overall survival (p=0.032), to grade III (p=0.006) and in a positive correlation between RCAS1 and Ki-67 (p<0.001). Moreover, there is a trend of RCAS1 over-expression in advanced or metastatic stages and in tumours with lymphovascular invasion. In contrast, protein expression was not strongly associated to tumour size, to histological type, to patient age or to gender. Conclusions: The most important conclusions of this study are the negative correlation between RCAS1 and overall survival, the over-expression of RCAS1 in grade III tumors and the positive correlation between RCAS1 and Ki-67 which means that when Ki-67 increases RCAS1 is higher. Consequently, this molecule RCAS1 probably could be considered as a marker of the tumor’s aggressiveness.
Novel tumor markers in non-small cell lung cancer detected both in serology and tissue.
Authors: Radhi Saba , Alalawi Raed , Mirandola Leonardo , Yu Yuefei , Pandey Apurva , D'Cunha Ruth , Saadeh Charles , Nguyen Diane Duyen Thuy , Hardwicke Fred L. , D'Cunha Nicholas C. , Tijani Lukman Adeoju , Jenkins Marjorie , Jumper Cynthia A. , Chiriva-Internati Maurizio , Cobos Everardo
Published: March 2020

e21105

Background: Cancer testis antigens (CTA) are a class of tumor associated antigens, showing a restricted expression in cancer, a strong immunogenicity and weak expression or absence in normal tissues. CTA are a growing family of proteins involved in signal transduction control. Sp17/AKAP4/PTTG1 have been previously investigated, showing promising results as a target antigens. Our aim was to investigate the expression of Sp17/AKAP4/PTTG1 in lung cancer patients. Methods: We analyzed 2 lung cancer cell lines, one normal bronchus cell line, a panel of normal tissues and patient’s cells by RT-PCR, flow-cytometry, immunocytochemistry (ICC), and immunofluorescence (IF). CTA immunogenicity was investigated by measuring circulating specific antibodies in the sera of lung cancer patients. PCR was performed by 35 amplification cycles. RNA integrity in each sample was checked by amplification of ß-actin. We studied the pattern of expression in patients with NSCLC according to histology, gender, and age. Results: Specimens from 11 patients were examined.CTA, Sp17, AKAP-4 and PTTG-1 are aberrantly expressed in lung cancer cell lines and primary cells from patients by RT-PCR, immunocytochemistry, and flow cytometry. ELISA analyses show the presence of circulating CTA-specific antibodies in the sera of lung cancer patients, indicating the immunogenicity of Sp17, AKAP-4 and PTTG-1. The mean age of patients was 60 years(range 57-76). Fifty eight percent were males(N=7). Seventy two percent (n=8) had adenocarcinoma. Six patients were stage IV at diagnosis. There was no difference in the pattern of expression between gender, stage or histology. Conclusions: We showed that CTA, Sp17, AKAP-4 and PTTG-1 can be detected in both sera and tissue of patients with NSCLC. They are potential therapeutic targets for immunotherapies. These may me useful in determining response to treatment as well as asses possible recurrence.
Evaluation of the pattern of SOX2 expression in non-small cell lung cancer (NSCLC) and correlation with clinicopathologic (CP) features and prognosis.
Authors: Velcheti Vamsidhar , Cheng Huan , Yao Xiaopan , Deng Yanhong , Gettinger Scott N. , Rimm David
Published: March 2020

e21106

Background: SOX2 is an embryonic stem cell transcription factor that plays a crucial role in differentiation and maintaining pluripotency. We examine the pattern of SOX2 expression and assess correlation with CP features in NSCLC. Methods: Using the AQUA method of automated quantitative immunofluorescence we measured SOX2 expression in two cohorts (in a tissue microarray format), from Yale New Haven Hospital (YTMA 79) and an independent cohort from Patras University Hospital in Greece (YTMA 140) (196 and 342 cases, respectively). YTMA 140 had high percentage of Squamous cell carcinoma (SCC) (48.8%) as opposed to 17.9% in YTMA 79. There were more stage III & IV pts in YTMA 140 (41.3%) compared to YTMA79 (30.6%). 88% of pts in the YTMA 140 were male compared to 52.5% in YTMA79. Survival analysis was done using Kaplan-Meier analysis with log-rank test. The associations between SOX2 expression level and CP features were evaluated by using non-parametric Kruskal-Wallis test. Results: In both cohorts, we found significantly higher SOX2 expression in SCC compared to adenocarcinoma. There was no correlation with gender or stage. Using the threshold of specific signal detection as the cut point, we divided the cohorts into expressers and non-expressers. YTMA 140 cohort which is enriched in SCC showed that 28/161(17.3%) pts with SCC were SOX2 non-expressers. Pts who were SOX2 non-expressers had a worse median overall survival (15 months) compared to pts who were SOX2 expressers (23.5 months) (p=0.015). Conclusions: Our data reveals that pts who have SCC have significantly higher expression of SOX2 and higher SOX2 expression correlates with better outcome. Further studies to define the molecular characteristics of SCC may elucidate more effective markers and effective classification for NSCLC.
Primary breast cancer tryptase expression as novel molecular drug target.
Authors: Ranieri Girolamo , Ammendola Michele , Brandi Mario , Misino Andrea , Goffredo Veronica , Patruno Rosa , Di Lecce Valentina , Gadaleta Caldarola Gennaro , De Sarro Giovambattista , Sacco Rosario , Gadaleta Cosmo Damiano
Published: March 2020

e21113

Background: Tryptase, a serine protease stored and released from mast cells granules has been identified as a new non-classical angiogenetic factor. Mast cells can release tryptase following c-Kit receptor activation. We have evaluated the correlations among the number of MCs positive to tryptase (MCDPT), the number of c-Kit receptor expressing cells (C-KREC) and microvascular density (MVD) in a series of 88 primary T1-3, N0-2 M0 female breast cancer by means of immunohistochemistry and image analysis methods. Methods: Six-micrometers thick serial sections of formalin-fixed and paraffin-embedded bioptic tumor samples were microwaved at 500 W for 10 min. and treated with a 3% hydrogen peroxide solution. Sections were incubated with primary antibodies: anti-tryptase (AA1; Dako, Glostrup, Denmark), anti-c-Kit receptor (A4502;Dako, Glostrup, Denmark) and anti-CD34 (QB-END 10; Bio-Optica Milan, Italy). In serial sections MVD, MCDPT and C-KREC were counted by means of image analysis at x400. Results: Data demonstrated a significantly (r= ranging from 0.71 to 0.91; p: ranging from 0.001 to 0.003 by Pearson’s analysis respectively) correlation between MVD, MCDPT and C-KREC to each other. Conclusions: Published in vitro data suggest that tryptase induce angiogenesis in vascular endothelial cells and breast cancer cells lines. According to these data we shown that MVD, MCDPT and C-KREC paralleled to each other suggesting a role in in vivo breast cancer angiogenesis. In this context several tryptase inhibitors such as gabexate mesilate or nafamostat mesilate might be evaluated in clinical trials as a new antiangiogenetic approach.