bluuebunny/crossref_metadata_2025_split huggingface dataset:
Quantitative immunohistochemical detection of gamma-H2AX in paraffin-embedded human tumor samples at National Clinical Target Validation Laboratory
Authors: Yang S. X. , Nguyen D. , Steinberg S. M. , Ji J. , Parchment R. , Kinders R. , Kummar S. , Gutierrez M. , Murgo A. , Tomaszewski J. E. , Doroshow J. H. 
Published: March 2020

10565 Background: DNA double-strand breaks (DSBs) caused by exposure to DNA damaging agents initiate phosphorylation of histone H2AX to form gamma-H2AX, which is considered a surrogate marker of DSBs. However, it is a challenge to quantitatively measure gamma-H2AX in clinical samples such as tumor biopsies. The aim of this study was to develop an immunohistochemical detection method for gamma-H2AX and to quantitatively evaluate the levels of gamma-H2AX in paraffin-embedded tumor samples. Methods: Human breast cancer MCF-7 cells were treated with the topoisomerase I inhibitor irinotecan at 1 μM or vehicle for 1 h, and fixed in 10% neutral buffered formalin and embedded in paraffin. Staining with gamma-H2AX antibody was performed on sections of treated MCF-7 cells, tumor specimens, and biopsies at baseline and after doxorubicin-containing chemotherapy from cancer patients. Numbers of foci and level of gamma-H2AX expression per tumor nucleus were determined by manual counting under a light microscope and an Automated Cellular Imaging System. Results: There was a rise in the mean numbers of nuclear foci and intensity of gamma-H2AX in MCF-7 cells treated with irinotecan versus vehicle (19.9 ± 2.7 vs. 9.95 ± 3.6; P < 0.0001 and 61.2 ± 8.5 vs. 16.2 ± 13.6; P < 0.0001 by Wilcoxon rank sum test). The level of gamma-H2AX foci in human tumor samples was 18.8 ± 13.1, 44.8 ± 14.5, 51.2 ± 20.8, or 69.7 ± 21.2 in carcinomas of the breast, colon, ovary, or prostate. In a patient with stable disease, levels of gamma-H2AX foci were 62.7 ± 26.9 at baseline and 67.2 ± 25.3 after doxorubicin-containing regimen chemotherapy. Conclusions: Our data suggest that the quantitative immunohistochemical detection of gamma-H2AX levels is facilitated by a digital imaging system, and is a reliable method to measure the effects of DNA damaging agents in cells and paraffin-embedded human tumor samples. Its application may help evaluate tumor response to various DNA damaging agents currently in the clinic and those presently undergoing clinical development. No significant financial relationships to disclose.
Elevated phosphorylated mammalian target of rapamycin (p-mtor) expression in human neoplasms
Authors: Ross J. S. , Qian J. , Sheehan C. , Jones D. , Jennings T. 
Published: March 2020

10571 Background: Mammalian target of rapamycin (mTOR) regulates several cell functions critical to tumorigenesis including proliferation, growth, mobility and survival. mTOR expression and dysregulation have been reported carcinomas of breast, head & neck, liver and kidney, but have not yet been studied in non small cell lung cancers (NSCLC), colorectal adenocarcinomas (CRC), nor brain tumors including gemistocytic astrocytomas (GA) and glioblastomas (GBM). Methods: Formalin-fixed paraffin-embedded tissue sections from 121 NSCLC [45 squamous cell carcinomas (SCC), 45 pure adenocarcinomas (AC), and 31 bronchioloalveolar carcinomas (BAC) including both pure BAC and adenocarcinomas with BAC features], 108 CRC, 8 GA and 24 GBM were immunostained by an automated method (Ventana Medical Systems, Tucson, AZ) using monoclonal rabbit anti-human p-mTOR antibody (Cell Signaling, Danvers, MA). Immunohistochemical assessment included intensity and percentage of positive cells in both benign and carcinomatous elements. Results were correlated with morphologic and prognostic variables in NSCLC and CRC and GBM. Results: Elevated p-mTOR expression was noted in 33% SCCs, 64% ACs, and 65% BAC, 78% CRC, 75% GA and 96% GBM. For NSCLC, increased expression of p-mTOR correlated with histologic subtype (p=0.004). Within SCC greater than 3 cm in size, only 20% had an increase in p-mTOR, while 53% of SCC ≤ 3 cm showed an increase in p-mTOR expression (p=0.03). 100% high grade vs. 43% low grade (p=0.05) and 100% advanced stage (p=0.037) vs. 54% low stage BAC showed an increase in p-mTOR. Consistently, 100% BACs with positive lymph node status vs. 54% node negative BACs showed an increased level of p-mTOR (p=0.037). For CRC, p-mTOR over-expression correlated with high tumor grade (80% Grade 3 vs 50% Grade 1 p=0.042). Majority of GA (6/8) and nearly all GBM (23/24) were positive for p-mTOR (p=0.135). Conclusions: Activated mTOR protein, defined by phosphorylation of mTOR at Serine-2448 (p-mTOR), is increased in NSCLC, CRC and both GA and GBM. These findings suggest a potential role for mTOR dysregulation in lung, colon and brain tumorigenesis and support the currently ongoing clinical trials investigating the therapeutic relevance of rapamycin analogs in the treatment of these tumors. No significant financial relationships to disclose.
Targeting the mitotic spindle in the treatment of adrenocortical cancer
Authors: Stephan E. A. , Gonzales P. , Han H. , Grant C. S. , Gately S. , Von Hoff D. D. , Trent J. M. , Demeure M. J. 
Published: March 2020

10572 Background: Adrenocortical carcinoma (ACC) is a rare and lethal cancer with an average incidence of 1 in 1.4 million. Response to current standard therapy including mitotane is approximately 22%. Herein we use gene expression profiling to identify novel targets for therapeutic intervention. Methods: The transcriptomes of 29 ACC tumors and 4 normal adrenal glands were profiled on Affymetrix Human Genome and Agilent 22K element expression arrays in order to identify genes that could represent potential therapeutic targets. Agents known to inhibit the selected targets were tested in comparison with mitotane in two ACC cell lines (H295R and SW-13) in vitro and are currently being tested in a Scid mouse xenograft system. Results: Expression of numerous kinesin family members is dysregulated within the ACC tumor population. Of interest in the ACC cells EG5 (KIF11, KSP), a kinesin important for mitotic spindle formation, is up-regulated 5.81+ 4.42 (μ+SD) and 6.48 + 0.99 fold on the Agilent and Affymetrix platforms respectively. In addition, SPARC, an albumin binding matrix associated protein is up-regulated in ACC tumors by 3.69 + 2.37 (μ+SD) and 1.56 + 0.44 on the two platforms. The anti-microtubular drugs paclitaxel and albumin bound paclitaxel (abraxane) exhibited in vitro inhibition of H295R and SW-13 cells at IC50 concentrations of 0.33 μM and 0.0078 μM for paclitaxel and 0.35 μM and 0.0087 μM for Abraxane compared to mitotane concentrations of 15.9 μM and 46.4 μM. Treatment effects of monastrol, paclitaxel, and Abraxane are currently being tested in xenograft murine models. Testing of the EG5 inhibitor S-trityl-L- cysteine is ongoing in the in vitro system. Conclusions: KIF11 inhibitors have been proposed in the treatment of cancer. These compounds may offer better anti-mitotic activity than other tubulin interacting agents with less neurotoxic side effects. Based on biological insight garnered through expression profiling of ACC tumors, the current ongoing research is investigating the use of abraxane and EG5 inhibitors in the in vitro and in vivo treatment of ACC. No significant financial relationships to disclose.
Rethinking the role of nodal status in TNM staging of node-positive breast cancer
Authors: Anderson S. J. , Bryant J. L. , Jeong J. , Tang G. , Costantino J. P. , Mamounas E. , Geyer C. E. , Wolmark N. 
Published: March 2020

10584 Background: Among TNM staging criteria, the number of positive lymph nodes [PNOD] is the strongest predictor of patient outcome. Based on SEER data, Vinh Hung (2004) asserted that the ratio of the number of positive nodes to the number of nodes removed [NODR] may better predict outcome. We evaluated this assertion in patients from 13 node-positive NSABP phase III trials. Methods: The impact of PNOD, its logarithm (LPNOD) and NODR on survival (S) and recurrence free survival (RFS) were assessed via Cox PH models in 19,768 patients with node-positive breast cancer in 13 NSABP clinical trials. The cohort was split into 2 groups: 9 older trials (Protocols B-05, B-07, B-08, B-09, B-10, B-11, B-12, B-15 and B-16) consisting of 9,915 patients and 4 newer trials (B-22, B-25, B-28 and B-31) consisting of 9,853 patients. Only randomized eligible patients with follow-up were considered. Models were fit to the outcomes for the older trials and validated in the newer trials. Results were adjusted for age, protocol and tumor size. We used Wald z-scores, Nagelkerke R2, and residual analyses to assess the strength of the relationship between nodal status and outcomes. Results: Median time on study was 19.8 years in the older trials and 10.6 years in the newer trials. Ten-year models across the protocols revealed that the relative hazard ratio for each of the outcomes studies was logarithmic according to the number of positive nodes for = 30 positive nodes. Using LPNOD and NODR compared to PNOD improved the strength of association but did not greatly improve goodness of fit ( Table 1 1). For all endpoints, the strength of association and goodness of fit associated with nodal status and other TNM factors attenuated in newer protocols. Conclusions: Nodal status and other TNM factors currently used for breast cancer staging continue to be strong indicators of patient prognosis. However, these factors should be augmented to account for new genetic and biological markers. [Table: see text] No significant financial relationships to disclose.
CDK2 and CDK1 specific activities: A novel prognostic indicator in early breast cancer
Authors: Kim S. , Masuda N. , Inaji H. , Yoshidome K. , Tsujimoto M. , Akiyama F. , Ishihara H. , Hortobagyi G. N. , Ueno N. T. , Noguchi S. 
Published: March 2020

10592 Background: Cyclin-dependent kinases (CDKs) are expressed almost constantly but their activities change according to cell cycle phase. We studied the specific activity (SA; activity/expression) of CDKs to accurately evaluate their role in cell proliferation. We focused on the ratio of CDK2 SA to CDK1 SA because this ratio has been associated with rapid tumor growth in human breast cancer xenografts. Our goal was to investigate the prognostic significance of CDK2/1 ratio in node-negative breast cancer. Methods: We used a novel assay to measure SAs of CDK2 and CDK1 in 365 primary breast tumors. All tumors were histologically confirmed invasive breast cancer without lymph node involvement. The primary endpoint was relapse-free survival (RFS). Cut-off values for CDK2/1 ratio were defined as those points that best discriminated groups according to RFS. Tumors were grouped as low (136pts), intermediate (84pts), and high (145pts) CDK2/1 ratio. The median follow-up was 59 months (range 4–102). Results: Pts characteristics are described as follows: menopausal status: pre- 42%, post- 58%; tumor size: =20mm 48%, >20mm 52%; histologic grade (HG): I 28%, II 47%, III 23%; ER: (+) 57%, (-) 42%; PR: (+) 51%, (-) 48%. Pts with HG III or high CDK2/1 ratio showed significantly lower 5y-RFS rates than those with HG I/II or low/intermediate CDK2/1 ratio, respectively (HG: I 97%, II 91%, III 85%, P=0.040; CDK2/1 ratio: low 97%, intermediate 92%, high 85%, P=0.017). In univariate analysis, PR (- vs +, P=0.088) had a tendency to associate with relapse, and HG (III vs I, P=0.024) and CDK2/1 ratio (high vs low, P=0.011) also had a significant correlation with relapse. However, only CDK2/1 ratio showed a significant independent prognostic indicator in multivariate analysis (hazard ratio 2.86, 95% CI 1.25–6.58, P=0.01). More important, among women (n=185) with hormone receptor (HR) positive disease given adjuvant hormone therapy alone, who have had no useful factors to predict their outcomes, high CDK2/1 ratio was also associated with worse prognosis than low CDK2/1 ratio (5y-RFS 84% vs 100%, P=0.007). Conclusions: For patients with node- negative disease, especially those with HR positive tumors given adjuvant hormone therapy alone, the CDK2/1 ratio might be useful as a routine laboratory test to predict outcome. No significant financial relationships to disclose.
Predictive value for survival of a risk model of two serological markers, beta-2-microglobulin (B2M) and lactic dehydrogenase (LDH), in diffuse large cell lymphoma (DLCL)
Authors: Rodriguez M. A. , Temple S. , Fayad L. , Hagemeister F. , McLaughlin P. , Romaguera J. , Cabanillas F. 
Published: March 2020

10599 Background: The International Prognostic Index (IPI), the standard tumor risk model in patients (pts) with diffuse large cell lymphoma (DLCL), has 5 factors (age, LDH, performance status, extra nodal sites, and stage). We previously proposed a simpler two-factor model, based on LDH and B2M (JCO10;1989). B2M is a component of HLA class I antigens, expressed in lymphocytes, and a known prognostic indicator in some lymphoid malignancies. The benefits of this model are: objective measures; lab method widely available; simplicity; biologic marker. Methods: We applied the model to a large cohort of DLCL patients with prospective baseline B2M, and treated with doxorubicin-based (chemo) regimens, with and without rituximab. 718 pts with DLCL were sequentially treated at MDACC by IRB approved chemo protocols from 1988–2000. In 2001, rituximab plus chemo (RCHOP) became standard. 311 DLCL pts were sequentially treated with RCHOP from 2001–2005. Cox regression analyses for univariate and multivariate models of IPI factors and B2M were done. Kaplan-Meier survival projections were in three risk categories: low (normal [nl] LDH and B2M); intermediate (either LDH or B2M > nl); or high (LDH and B2M > nl). Results: In both treatment groups, IPI factors and B2M were significant as univariate factors. In the RCHOP group, however, the IPI multivariate model showed age, stage, and extra-nodal sites were not significant risk factors, while B2M and LDH remained highly significant (p<0.01). The 5 year survival projections by risk category were: (*) combines intermediate low and intermediate-high categories Conclusions: This simple two-factor model predicts risk for patients with DLCL, treated with or without rituximab, comparably to the IPI. B2M should be considered an important prognostic indicator in DLCL, particularly in rituximab treated patients. Exploratory analyses to revise the IPI model are indicated. [Table: see text] No significant financial relationships to disclose.
Clinical significance of Cdh1-Skp2 cascade in breast cancer
Authors: Fujita T. , Wu G. , Wood R. D. , Wan Y. 
Published: March 2020

10609 Background: Skp2 (S-phase kinase-associated protein 2) is substrate-recognizing subunit of an ubiquitin E3 ligase SCF (Skp1, Cullin and F-box) complex and has a major role in p27 regulation. Overexpression of Skp2 is frequently seen in human tumors, often correlating with poor prognosis. Anaphase-promoting complex/cyclosome (APC/C) is another most prominent ubiquitin E3 ligase in cell cycle control, and its function is tightly controlled by activating subunit Cdh1 (Fizzy-related). APC-Cdh1 complex is supposed to be involved in Skp2 proteolysis and have essential role for S phase entry, however clinical significance of Cdh1 have not evaluated yet. Herein we examined the significance of Cdh1 in breast cancer. Methods: Firstly, MCF7 human breast cancer cells and MCF10A normal breast epithelial cells were analyzed in vitro and in vivo. Thereafter, using tissue microarray, we evaluate the expression profile of Cdh1, Skp2 and p27 in both breast cancer and normal breast epithelial tissue. Moreover, clinicopathological significance (age, tumor size, lymph node metastasis, distant metastasis, histological grade, and stage) of Cdh1 was analyzed from 126 breast cancer patients. Chi square-test, Fisher's exact test were used for statistical analysis of immunostaining results and clinicopathological data. p<0.05 was considered statistically significant. Results: Overexpression of Cdh1 induced attenuation of Skp2 and increased p27 protein expression, resulted in growth suppression. Moreover, knockdown of Cdh1 promoted higher Skp2 expression and S-phase population, reduced p27 and consequently induced cell transformation and proliferation in vitro and in vivo. Tissue microarray results appeared that positive Cdh1 and p27 was more frequently seen in normal breast tissue and statistically significant. On the other hand, Skp2 was less in normal tissue. Furthermore, Cdh1 positive breast cancer was more frequently seen in low histological grade tumors and statistically significant (p=0.04). Conclusions: Skp2 protein expression is regulated by Cdh1 in breast cancer. Cdh1 expression could be possible novel biomarker in patient with breast cancer. No significant financial relationships to disclose.
Combined MEK and mTOR suppression is synergistic in human NSCLC and is mediated via inhibition of protein translation
Authors: McDaid H. M. , Legrier M. , Yang C. H. , Yan H. G. , Lopez-Barcons L. , Keller S. M. , Horwitz S. B. 
Published: March 2020

10615 Background: Lung cancer is a heterogeneous disease characterized by the acquisition of somatic mutations in numerous protein kinases, including components of RAS and AKT. These intersect at various points rendering the network highly redundant. Hence, dual suppression of both, using MEK and mTOR inhibitors may be a promising rational drug combination that can overcome the intrinsic plasticity of this signaling network. Methods: The concurrent combination of the MEK inhibitor CI-1040 and the mTOR inhibitor AP23573 was evaluated in non-small cell lung cancer (NSCLC) cell lines, using the combination index method of Chou and Talalay. The concurrent combination of Rapamycin and the MEK inhibitor PD0325901 was also evaluated in animal models of human lung cancer. Results: The MEK inhibitors, CI-1040 or PD0325901, in combination with the mTOR inhibitor rapamycin, or its analog AP23573, were synergistic in human lung cancer cell lines. Statistically significant enhancement of cell death by the combination of MEK and mTOR inhibitors was not observed. Rather, synergism was associated with suppression of proliferation and inhibition of protein translation. Concurrent suppression of both MEK and mTOR inhibited ribosomal biogenesis by forty percent after twenty-four hours and was associated with a block in the initiation phase of translation. Furthermore, the combination of PD0325901 and rapamycin was significantly superior to either drug alone, or PD0325901 at the maximum tolerated dose, in xenograft and human heterotransplanted lung tumor models grown in nude mice. Conclusions: These data support therapeutic strategies in lung cancer that simultaneously suppress RAS and mTOR signaling networks, and provide evidence that both pathways converge on factors that regulate protein translation initiation. No significant financial relationships to disclose.
Phase II study of docetaxel, carboplatin and trastuzumab (THC) in patients with locally advanced breast cancer
Authors: Dirix L. Y. , Prové A. , Sweldens C. , Van Dam P. , Vermeulen P. , Benoy I. 
Published: March 2020

1062 Background: Preclinical and clinical data suggest that docetaxel, trastuzumab and platinum compounds act synergistically. Studies in the metastatic disease are conflicting. In this phase II study the activity of this regimen was assessed and correlated with changes in circulating biomarkers. Methods: Patients with locally advanced, FISH+ HER2 amplified, breast cancer were treated in this protocol with TCH as initial neo-adjuvant treatment. Patients received Docetaxel 75 mg/sqm, carboplatinum AUC 6mg/mL.min and trastuzumab 6 mg/kg (with a loading dose in cycle 1). Six cycles were intended provided no progression occured. No prophylactic G-CSF or antibiotics were administered. Measurement of circulating ECD HER2, CTC (RT-PCR mammaglobin and CK-19), and total plasma DNA was performed prior to the start of therapy, after cycle 1 and after 3 and 6 cycles. A cardiac MUGA exam was performed every three cycles. All patients were intented to undego mastectomy and axillary clearance after the end of chemotherapy. Results: 40 patients have been treated with a median age of 56 year. No patient suffered from progression of disease while on THC. All patients received 6 cycles, 4 patients received respectively 7, 7, 8 and 9 cycles. In total 247 cycles were administered. No cases of CHF were observed. 5 patients suffered from a asymptomatic decline > 10% in LVEF. A clinical CR and PR was obtained in 32 (80%). A pathological CR both in breast axilla was obtained in 16/39 (41%). Tumor response was predicted by changes in both CTC and ECD HER-2. Conclusions: THC was confirmed to be a very active and safe regimen in patients with HER-2 amplified breast cancer. Changes in biomarkers rapidly predicted response. No significant financial relationships to disclose.
Chemoprotection by VEGF: Rationale for combination nab-paclitaxel and bevacizumab
Authors: Trieu V. , Ran S. , Bivens C. , Desai N. 
Published: March 2020

1064 Background: Nanoparticle albumin-bound (nab-) paclitaxel (ABX) has shown greater efficacy and less toxicity than solvent-based paclitaxel (TAX) in xenograft models and clinical trials. The goal of this study was to determine the effects of VEGF modulation in human MDA-MB-231 breast tumor cell line and the effects of ABX and VEGF-neutralizing antibody bevacizumab (AVA) combination on the growth and metastasis of orthotopically implanted MDA-MB-231 tumors. Methods: VEGF expression was evaluated by ELISA in MDA-MB-231 tumor extract one week after treatment (qdx5) with saline, doxorubicin (10 mg/kg), TAX (10 mg/kg), or ABX (15 mg/kg). VEGF-receptor expression in MDA-MB-231 was quantitated by RT-PCR. MDA-MB-231 cytotoxicity with ABX, VEGF, AVA alone or in combination was measured by cytotoxicity and clonogenic assays. Implanted MDA-MB-231 tumors expressing luciferase were treated with saline, 2 cycles of ABX (10 mg/kg, two qdx5 cycles separated by 1 week, N=5) alone or in combination with AVA (2, 4 and 8 mg/kg, 2/wkx6). Tumor lymph node and pulmonary metastasis was determined by measuring luciferase activity. Results: Compared with saline, MDA-MB-231 tumors following chemotherapies exhibited significant tumor shrinkage (p≤0.006, t-test) and VEGF induction (p<0.0001, t-test). MDA-MB-231 was shown to express VEGFR2. Exogenous VEGF had a protective effect on MDA-MB-231 tumor cells by reducing cytotoxicity of ABX in both cytotoxicity and clonogenic assays. Sequestration of VEGF with AVA increased cytotoxicity of ABX in vitro. Treatment of MDA-MB-231 breast tumors with ABX and AVA combination resulted in greater than additive antitumor response and significantly reduced metastasis to the lungs (p=0.025 vs control) and LN (p=0.022) at the highest AVA dose. Conclusions: Chemotherapies induced VEGF expression in MDA-MD-231 breast tumor in vivo. In vitro, VEGF exerted a protective effect against ABX chemotherapy in VEGFR2-expressing MDA-MD-231 cells, which was abrogated by addition of AVA. In vivo, ABX and AVA combination significantly inhibited the metastatic potential of MDA-MB-231 tumor cells. These data provide a rational basis for the combination of nab- paclitaxel and bevacizumab in VEGF-receptor expressing tumors. [Table: see text]