bluuebunny/crossref_metadata_2025 huggingface dataset:
Blinded comparator study of immunohistochemistry (IHC) versus a 92-gene cancer classifier in the diagnosis of primary site in metastatic tumors.
Authors: Weiss Lawrence M. , Chu Peiguo , Schroeder Brock , Singh Veena , Zhang Yi , Erlander Mark G. , Schnabel Catherine A.
Published: March 2020

e21019

Background: Metastatic tumors of uncertain or unknown origin pose diagnostic and therapeutic challenges. Accurate tumor classification is fundamental to inform predictive biomarker testing and optimize therapy. Standard of care employs IHC analysis for tumor classification that is varied in approach and interpretation. More recently, gene expression-based tests have been proposed as diagnostic aids. This study directly compared the diagnostic accuracy of IHC vs molecular classification with a 92-gene RT-PCR assay (CancerTYPE ID) for determination of primary tumor site. Methods: In this prospectively-defined, blinded study, 132 high grade, metastatic cases were selected by City of Hope (COH). Cases were reviewed and reference diagnoses established by clinical correlation (eg, patient history, imaging). Blinded FFPE sections were evaluated by either IHC or the 92-gene RT-PCR assay. Results were scored within a clinically-relevant categorization system designated by COH for main tumor type and subtype. The primary endpoint was concordance with the reference diagnosis. Study unblinding and data analysis were conducted by an independent third party. Results: Final analysis included 123 cases (mean age±SD: 61.2±14.4y, 52% female; grades 3/4; 90% metastatic); 9 cases were excluded (8: RNA quality; 1: insufficient tumor tissue). The 92-gene assay demonstrated an overall accuracy of 78% for tumor classification vs 68% for IHC (P = 0.017). For tumor subclassification, the 92-gene assay correctly identified 72% of tumors compared to 61% with IHC (P=0.029). Sensitivities were similar for GI (77%) and kidney (77%). The 92-gene assay demonstrated higher sensitivities for lung (75% vs 67%), urinary bladder (75% vs 42%), and breast (73% vs 55%). Mean IHC use was 7.8 stains per case (median: 8, range: 2-15). Conclusions: This is the first study to directly compare the diagnostic accuracy of IHC vs molecular classification. Results from this blinded series of high grade metastatic cases demonstrate superior accuracy with the 92-gene assay vs standard of care IHC, and strongly support the diagnostic utility of molecular classification in difficult to diagnose metastatic cancer.
Antibody-independent ApoStream technology isolates folate receptor alpha (FRA)–positive circulating tumor cells from blood of non-small cell lung adenocarcinoma patients.
Authors: Davis Darren W. , Neal Christopher , Gupta Vishal , Melnikova Vladislava O. , Woo Jacky , Somers Elizabeth Brooke , O'Shannessy Daniel
Published: March 2020

e21028

Background: The ability to enrich and interrogate circulating tumor cells (CTCs) from blood may allow for the analysis of metastatic dissemination and potential use of CTCs as surrogates for monitoring drug efficacy. We developed ApoStream, a dieletrophoresis field-flow fractionation based platform, for antibody-independent CTC isolation. We demonstrated that ApoStream successfully isolates non-small cell lung adenocarcinoma (NSCLC) CTCs that express Folate Receptor alpha (FRA), a GPI-anchored receptor, which has emerged as a cancer biomarker and potential therapeutic target in multiple cancer types. Methods: ApoStream technology was used to enrich CTCs from NSCLC patients’ blood. CTC enrichment by ApoStream was compared to that of the FDA cleared CellSearch CTC kit. A multiplexed immunofluorescent assay was developed to enable CTC enumeration (Cytokeratin+/CD45-/DAPI+ cells) and analysis of FRA expression (detection by murine antibody clone 26B3) using single cell quantitative laser scanning cytometry (LSC). Results: In a side-by-side comparison with the CellSearch CTC kit, ApoStream isolated significantly higher numbers of CTCs in 9 metastatic adenocarcinoma NSCLC patients (Apostream: mean =139, range 3-487 per 7.5 mL of blood, versus CellSearch kit: mean =2, range 0-8 per 7.5 mL of blood, n= 9, p=0.041). All patients were found to be CTC-positive by ApoStream, while only 3 of 9 (33%) patients were CTC-positive based on the CellSearch kit. LSC analysis demonstrated that 8-33% of all CTCs isolated expressed FRA and that FRA expression was confined to CTCs only. No false positive CTCs and no FRA-expressing cells were isolated by ApoStream from normal donor blood (n=15). Conclusions: All NSCLC adenocarcinoma patients analyzed had FRA-positive CTCs, suggesting that FRA may play a key role in metastasis and that screening of patients with the ApoStream CTC isolation system may identify patients who could benefit from FRA-targeted therapy.
Comprehensive analysis of serum N-glycans as a biomarker for breast cancer.
Authors: Ueno Takayuki , Sato Fumiaki , Shinkura Nobuhiko , Aihara Taichi , Fukushima Masao , Sekijima Masaru , Toi Masakazu
Published: March 2020

e21031

Background: Early detection of breast cancer is an important aspect to improve therapeutic outcome. Serum biomarkers have a number of advantages including less invasiveness and possible application for monitoring. Glycans are involved in many biological processes including cancer progression and promising candidates for cancer biomarker. Methods: Sera were collected from 54 patients with operable breast cancer and 118 healthy volunteers. N-glycans were isolated by BlotGlyco (Sumitomo Bakelite Co., LTD, Japan) and N-glycan profile was obtained by MALDI-TOF MS. The principal component analysis (PCA) was performed by SIMCA-P+ ver.12 (Umetrics). N-glycan compositions were estimated from mass spectra using 'GlycoMod' database. Results: Fifty-four pts (age: 31-78) and 118 healthy females (age: 29-73) were enrolled. The tumor size was between 9 and 107mm (median 29mm). Twenty six pts (48%) had lymph node involvement. Fifty three peaks of N-glycans were observed in more than 95% of the samples and utilized for the analyses. By PCA, no distinction was observed in sera form breast cancer pts in terms of the tumor size, hormone receptor status and HER2 status. The N-glycan signature to identify cancer pts was created based on the PCA. The signature categorized samples into three groups: cancer, healthy and border groups. Half of the samples (even registration number) were used for training and the rest for validation. In the validation cohort, 18 (66.7%) of 27 pts were categorized into the cancer group and 9 were into the border group. Forty-seven (79.7%) of 59 volunteers were categorized into the healthy group and 11 were into the border group. When the border group was included in the cancer group, the sensitivity was 100% (27 of 27 pts in the cancer group) and the specificity was 79.7% (47 of 59 volunteers in the healthy group). Conclusions: The comprehensive analysis of N-glycans in sera enabled the creation of the N-glycan signature to distinguish between breast cancer pts and healthy females with high sensitivity and high specificity. Confirmative studies are warranted.
PTHrP(12-48) for the diagnosis of breast cancer bone metastasis.
Authors: Washam Charity L. , Byrum Stephanie D. , Leitzel Kim , Suhail Ali M. , Lipton Allan , Suva Larry J.
Published: March 2020

e21039

Background: Bone metastasis of breast cancer significantly compromises patient morbidity and mortality. Currently, no reliable methods detect or predict patients at increased risk for developing bone metastasis. We utilized 3 independent cohorts of breast cancer patients to validate a highly discriminatory plasma-based proteomic profile that identifies breast cancer bone metastasis. The identity of the most discriminatory protein component identified was a parathyroid hormone-related protein fragment, PTHrP(12-48). Methods: Plasma samples collected from 21 breast cancer patients with clinical evidence of a bone metastasis and 21 patients with no evidence of bone metastasis from time of diagnosis to clinical outcome were evaluated. A novel mass spectrometry-based assay using human serum spiked with synthetic PTHrP(12-48) was used to measure PTHrP(12-48) concentrations (pg/μl). Statistical significance was assessed by one-way ANOVA. ROC curves evaluated the diagnostic potential of PTHrP(12-48) and a simple logistic regression derived from the combined measurement of PTHrP(12-48) and NTx. Results: PTHrP(12-48) concentrations ranged between 11.6 and 92.1 pg/μl in bone metastasis patients and between 4.5 and 34.2 pg/μl in patients without bone metastases. PTHrP(12-48) was significantly increased in bone metastasis plasma (p < 0.05). No significant correlation was identified between PTHrP(12-48) and NTx. ROC analysis of PTHrP(12-48), threshold 18 pg/μl, classified the two groups with high accuracy. Class prediction by the PTHrP(12-48)/NTx logistic regression model increased diagnostic specificity. Conclusions: The measurement of PTHrP(12-48) in patient plasma has potential as a viable clinical measure of bone metastasis. In combination with serum NTx, PTHrP(12-48) may assist in identifying bone metastases in patients presenting with low to normal bone turnover markers.
Investigation of telomerase activity in inflammatory and noninflammatory breast cancer.
Authors: Salem Ayman Maher , El-Shinawi Mohamed , Sabet Salwa Farouk
Published: March 2020

e21055

Background: Inflammatory breast cancer (IBC) is an aggressive type of breast cancer disease that has a high incidence in Egypt than western countries. It is characterized by rapid progression, involvement of dermal lymphatic emboli and extensive lymph node involvement. Basic and translational studies needed to define IBC disease biology and identify specific biomarkers have been limited by the paucity of patient samples. Hence, the current study aimed to introduce the telomerase activity level as a novel diagnostic marker for breast cancer and specifically for IBC to be differentiated from non-IBC. Methods: Breast cancer patients were enrolled from Ain Shams University hospitals in Cairo, divided into two groups: IBC (n=26) and non-IBC (n=27). Tissue samples were collected during modified radical mastectomy. TRAP (Telomerase repeat amplification protocol) assay was used to assess the telomerase activity in inflammatory and non inflammatory breast cancer tissue samples and their representative cell lines, SUM149 and MCF-7 respectively. Immunohistochemistry was used to investigate the expression of hTERT subunit of telomerase in paraffin embedded tissue samples of both types of patients. MTT assay was used to compare the effect of AZT (3’-azido-3’-deoxythymidine) on the growth of cell lines SUM149 and MCF-7. Results: : IBC showed Telomerase activity ranged from 12 to 376 units with a mean value of 75 and a median value of 43, while telomerase activity in non-IBC ranged from 6 to 109 units with a mean value of 35 and a median value of 24. On the other hand, normal tissues showed telomerase activity below 5 (P > 0.001). Using immunohistochemistry, the hTERT expression was higher in IBC than non-IBC and no expression at all in normal tissues. Also, a positive correlation was found between the telomerase activity and the number of metastatic lymph nodes in both IBC (r =0.511) and non-IBC (r =0.622). Besides, IBC cell line showed a faster response (53%) to telomerase inhibition by AZT after 24 hrs than non-IBC cell line (13%) at the same conditions. Conclusions: Telomerase could be a promising marker at the diagnostic and therapeutic levels in breast cancer and specifically in IBC.
Mitosis interference of cancer cells during anaphase by electric field from NovoTTF-100A: An update.
Authors: Lee Sze Xian , Tunkyi Adrian , Wong Eric , Swanson Kenneth D.
Published: March 2020

e21078

Background: The NovoTTF-100A is a device that emits an alternating electric field (TTField) that kills mitotic cells and has been FDA-approved for the treatment of recurrent glioblastoma. Exposure to the TTField resulted in impaired cytokinesis and asymmetric chromosome segregation. To further elucidate its mechanism of action, we performed experiments using synchronized tumor cells and TTField-treated xenografts. Methods: Biochemical analysis of cell cycle-specific proteins, timelapse movies, and immunoflourescence microsopy were used to examine the effects of TTField on synchronized tumor cells. Xenografts treated with TTField were analyzed for the presence of phospho-Histone H3 and cell cycle-specific epitopes. Results: Cells exposed to TTField as they transited through M-phase exhibited persistence of phospho-Histone H3 while Securin levels decreased, suggesting that these cells transit through metaphase to anaphase but were delayed in mitotic exit. Immunoflourescence of treated cells showed evidence of impaired cytokinesis together with disordered separation of chromosomes from metaphase plates. Lagging chromosomes, dispersion of chromosomes, chromosomes decondensation in the absence of cytokinesis, and asymmetric chromosomal segregation were also observed in treated samples. The number of 4N cells increased following TTField exposure compared to parallel cultures that had entered the G1-phase of the cell cycle indicating a failure of mitosis. Exposed cells showed no detectable p53 induction indicating that cell death is achieved by a p53-independent mechanism. Lastly, immunostaining of treated xenografts for phospho-Histone H3 revealed the presence of significantly higher numbers of immunopositive mitotic figures compared to untreated samples. Conclusions: These data suggest that TTField exposure resulted in defective entry into anaphase and interfered with orderly exit from mitosis, a mechanism of mitotic perturbation that differs from currently used spindle poisons. NovoTTF-100A may therefore exhibit synergism when combined with conventional cancer treatments. Supported in part by a grant from NovoCure, Inc. and A Reason To Ride research fund.
Association of IL17-expressing γδ t cells with acute radiation-induced pneumonitis.
Authors: Liao Winston W. P. , Chao K. S. Clifford , Hei Tom K , Cheng Simon
Published: March 2020

e21097

Background: Radiation pneumonitis is a substantial cause of morbidity and mortality with thoracic radiation for lung cancer. Little is known about the crucial mechanisms of the inflammatory response. We seek to determine if a key mediator of organ-specific inflammatory disorders and innate immune response, IL-17+ γδ T cells, is associated with radiation pneumonitis. Methods: C3HBe/FeJ mice (7 mice/group) were sham-irradiated as controls or exposed to a single dose of 15 Gy thoracic X-ray to develop pneumonitis. We have previously shown that TGFβ has an immunosuppressive activity in radiation pneumonitis. To potentiate the radiation pneumonitis, one group of mice was administered anti-TGFβ therapy with inhibitory TGFβ mAb (1D11, i.p.10 mg/kg/wk). Bronchoalveolar lavage fluid was assessed for cytology and inflammatory cytokine level. Lung tissues were examined for cell infiltration and histopathological changes. Cell surface marker and intracellular cytokine staining were performed on lymphocytes from the digested lungs by flow cytometry. Results: At 10 weeks post-irradiation, the lungs of the irradiated mice showed substantially more alveolar wall edema and increased infiltration of inflammatory cells compared with sham controls. Pneumonitis-involved lungs contained more IL-17+ γδ T cells (0.85% ± 0.00%) compared with sham controls (0.33% ± 0.02%), p<0.001. Furthermore increased IL-17+ γδ T cells were associated with potentiated radiation pneumonitis with anti-TGFβ therapy. There was a significant increased alveolar inflammation in irradiated mice injected with anti-TGFβ mAb. Anti-TGFβ irradiated lungs also contained significantly more IL-17+ γδ T cells (1.17% ± 0.13%) compared with irradiated controls (0.72% ± 0.13%), p<0.001. There was no increase of other TGFβ-dependent T cell subtypes such as IFNγ+ αβ T cells (Th1), IL-17+ αβ T cells (Th17), CD25+ Foxp3+ Tregs, nor activated macrophages in the potentiated pneumonitis lungs. Conclusions: Our findings implicate a novel role for IL17-expressing γδ T cells in radiation pneumonitis. This study reveals this innate immune response pathway as a potential target for therapeutic intervention in radiation lung injury
Invasive lobular (ILC) versus invasive ductal (IDC) breast cancer (BC): Clinical-pathologic features and clinical outcomes in monoinstitutional series.
Authors: Ferro Antonella , Eccher Claudio , Caldara Alessia , Triolo Renza , Barbareschi Mattia , Cuorvo Lucia Veronica , DiPasquale MariaChiara , Decarli Nicola L , Mussari Salvatore , Galligioni Enzo
Published: March 2020

e21102

Background: ILC is less common than IDC and is usually diagnosed at a later stage Purpose:The aim of our study was to investigate different clinico-biological behavior associated to ILC compared to IDC and to evaluate implications on survival outcomes Methods: We analyzed data from 3749 consecutive cases of IBC treated from 1995 to 2008 and classified as ILC, IDC and mixed/other. Relationships with clinical-pathological variables and the impact of ILC/IDC types on Event Free Survival (EFS), Overall Survival (OS) and Post-Progression Survival (PPS) were analyzed. Results: We have identified 445 ILC BC (12%), 3021 IDC (80.5%), 149 mixed (3.9%) and 134 other histotypes (3.6%). Median age was 61 (25-97) years. ILC presented a significantly (p<0.001) larger tumor size (T≥2: 46% vs 34%) and more frequent axilla involvement (43% vs 37%) compared to IDC. It was usually associated with intermediate grade, low ki67, positive ER and negative HER2 status. Mastectomy was more frequently required for ILC (45%) than IDC (37%). Adjuvant hormonal (± previous chemo) therapy was more frequently given (77 vs 64%) to ILC pts due to higher ER expression than IDC. At a median follow up of 77 (0-272) months (ms), there were not significant differences in EFS (81.4 vs 82.1%, p=0.7) and OS (82.8 vs 84.6%; p=0.19) between ILC vs IDC. Local and Distant Relapses were 15 (3.3%) and 50 (11.2%) in ILC vs 179 (5.9%) and 225 (7.4%) in IDC; preferential sites of first distant relapses were similar in both histotypes. Contralateral and second tumors were 9 (2%) and 22 (5%) in ILC vs 84 (2.8%) and 174 (5.7%) in IDC. Median time to first event was 38.3 ms in ILC vs 35.23 in IDC. PPS was 16.5 ms (0-100) in ILC compared to 22.2 (0-187) in IDC. Prognosis in term of OS was consistently worse for ILC pts, compared to IDC, when considering the luminal A (86.9 vs 93.5%; p=0.003), luminal C (70.4 vs 88.5%; p=0.028) and in triple negative subgroups (50 vs 73.1%; p=0.021). No difference in OS was seen for age, size, hormonal and nodal status, except in N>3 ILC pts, in whom a trend of worse prognosis (55.6 vs 60.2%) was observed. Conclusions: ILC pts did not show in our series a better outcome than IDC pts, despite a quite favorable biological pattern.
EBUS-TBNA for identifying biomarkers.
Authors: Chen Kezhong , Yang Fan , Zhao Hui , Zhou Zu li , Jiang Guanchao , Wang Jun
Published: March 2020

e21103

Background: We assessed the usefulness of endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) to detect EGFR AK-ras AALK mutations from malignant mediastinal nodes of patients with NSCLC. Methods: From February 2010 to December 2011 , 52 specimens diagnosed lung adenocarcinoma by EBUS were used for molecular analysis . Needle puncture site for the EBUS was standard 22G needle, once cytology pathology diagnosed adenocarcinoma, at the same site with the conventional needle puncture method to obtain adequate biopsy specimen for further analysis. The tissue was fixed in 10% formalin and stored in -80°C. When histologic diagnosis confirmed adenocarcinoma, the sample was extracted DNA to detect EGFR mutation by both amplification refractory mutation system( ARMS) and mutation enriched method improved by our department. Partial specimens also extracted DNA or RNA to detect K-ras and ALK mutation. Results: All 52 specimens were successfully extracted DNA, 34 were male and 18 were female, mean age of 62.1 years (34 to 79 years). Somatic mutations of the EGFR gene were detected in 25 of 52 cases (48%), 15 cases of exon 19 mutation, 9 cases of exon 21 mutation, and 1 of T790M mutation in ARMS. The result was of slight difference in mutation enriched method. In the patients who requested other biomarkers analysis, 4 had K-ras mutations and 2 had ALK mutations. 6 patients of 25 cases of patients with EGFR mutations had a radical surgery after neoadjuvant chemotherapy, tumor specimens were tested again with EGFR gene mutation, the same result as the preoperative test results. Conclusions: EBUS-TBNA mediastinal lymph node biopsy in the detection of EGFR AK-ras and ALK mutations in tissue samples was feasible, through the highly sensitive methods, higher detection rate was available, the result can be used as evidence of personalized treatment of lung cancer.
Comprehensive analysis of genetic polymorphisms and irinotecan-induced adverse events in Japanese gastrointestinal cancer patients: A DMET microarray profiling study.
Authors: Kuramochi Hidekazu , Kanno Hitoshi , Uchiyama Tomotaka , Nakajima Go , Saito Kayoko , Hayashi Kazuhiko
Published: March 2020

e21108

Background: Irinotecan is a key drug in the treatment of colorectal and gastric cancer, that may occasionally cause severe adverse events (AEs), especially neutropenia and diarrhea. Although UDP-glucuronosyltransferase (UGT)1A1 polymorphisms are used as biomarkers for predicting AEs, the effect of UGT1A1 polymorphism in clinical use is limited, suggesting that there is a possibility of the existence of other, unknown biomarkers. Methods: Fourteen gastrointestinal cancer (5 gastric, 9 colorectal) patients who had undergone irinotecan-based chemotherapy were enrolled. DNA extracted from peripheral blood cells was genotyped by the DMET Plus microarray system, and 1,931 gene polymorphisms were investigated. The relationship between AEs and polymorphisms was analyzed statistically. Results: Eleven polymorphisms showed the P value < 0.05 with grade 3,4 neutropenia, but no statistically significant polymorphisms were found after the correction of multiple comparisons. With respect to the association with diarrhea, 12 polymorphisms showed P <0.05, and even after the correction of multiple comparisons, CYP2F1_96G>A(P32P) showed a significant relationship (P<0.00001). With respect to the relationship between UGT1A1*6, *28 polymorphisms and AEs, *6 polymorphism showed significant (P=0.044) association with grade 4 neutropenia. Conclusions: CYP2F1_96G>A showed significant association with diarrhea. The association of UGT1A1*6 polymorphism and neutropenia was confirmed, as in previous reports.
The association of plasma TGFβ1 during radiotherapy and genotypes of TGFβ1 pathway in patients with non-small cell lung cancer.
Authors: Yuan Shuanghu , Yu Jinming , Cai Xuwei , Kong Feng-Ming (Spring)
Published: March 2020

e21115

Background: Radiation induced thoracic toxicities (RITT) in lung, esophagus and pericardium are dose-limited toxicities in patients with non-small cell lung cancer (NSCLC). Plasma transforming growth factor-beta1 (TGFβ1) and its change during radiotherapy was reported as a biomarker and single nucleotide polymorphisms (SNP) in TGFβ1 pathway was associated with RITT in our previous studies. In order to explore the mechanism of RITT, we tested whether plasma TGFβ1 was associated with genotypes of TGFβ1, tissue plasminogen activator (tPA) and angiotensin converting enzyme (ACE). Methods: Patients with stage I-III NSCLC enrolled in prospective clinical trials were eligible. All patients received radiotherapy with or without concurrent chemotherapy. Platelet-poor plasma was obtained pre-RT and at 4–5 weeks (40–50 Gy) during RT. Plasma TGF-β1 was measured using an enzyme-linked immunosorbent assay. The DNA samples extracted from blood before treatment were analyzed for the following genetic variations: TGFβ1 509C/T, TPA -7351 C/T, and ACE I/D. Results: 76 NSCLC patients received definitive radiotherapy (median dose 66 Gy) were enrolled. For the entire group of patients, the mean pre-RT TGFß1 level was 10.7±2.3 ng/ml, and the mean during-RT TGFß1 level was 6.0±0.7 ng/ml. No significant TGFß1 level differences were found at pre-RT and during-RT in patients with different genotypes of TGFß1 and tPA. Only ACE DD group had marginally higher pre-RT TGFß1 level than II and ID group (DD 23.6ng/ml vs. II 9.0ng/ml vs. ID 7.5ng/ml, p = 0.05), but no difference at during-RT(p=0.346) or during-RT/pre-RT ratio(p = 0.433). However, patients with TGFß1 509CC had higher elevation of plasma TGF ß1 level at fourth week during-RT than T allele carriers (TGFß1 level ratio of during-RT/pre-RT were CC 1.4±0.2 vs. T allele carriers 0.7±0.1, p=0.047). Conclusions: This exploratory study demonstrated that patients with TGFß1 509CC had higher elevation of plasma TGF ß1 level at fourth week during-RT than T allele carriers. This can help explain the correlation of TGF ß1 level elevation and higher risk of radiation induced lung toxicity in patients with NSCLC.
Development of automated quantitative multiparameter immunocytochemical profiling of circulating tumor cells (CTCs) in breast cancer (BC).
Authors: Miller Brandon L. , Deighan Clayton , Chalmers Jeffrey J. , Lustberg Maryam B.
Published: March 2020

e21124

Background: The currently accepted definition of a CTC is a nucleated cell that is positive by qualitative fluorescence for EpCAM, expresses cytokeratin, and is negative for the pan-hematopoietic marker CD45. Such approach has limitations such as operator bias, difficulty distinguishing true signal from autofluoresence or filter bleed-through, and most importantly, it cannot differentiate expression levels that are low, intermediate or high. This is a problem because CTCs are in fact heterogeneous with respect to phenotypic expression of various markers. Methods: Peripheral blood samples were subjected to our previously described, negative depletion technology. Cytospins of enriched samples were stained with 3 to 5 antibodies conjugated to various Alexa Fluor dyes plus a nuclear dye and then analyzed with both confocal microscopy and a traditional epifluorescence microscope equipped with a Nuance (CRi) multispectral imaging camera and associated software. This imaging system allows not only fluorescent signals to be quantified between specific excitation and emission filters, but also the spectral structure between these boundaries through the use of a liquid crystal that takes images in as little as 10nm increments. Such imaging allows spectra from specific dyes to identify from the contributions of other dyes, filter leakage, and auto fluorescence. Results: Using this technology, and appropriate controls, we are developing continuous variable quantification of the expression of: cytokeratins, vimentin, CD45, Her2, CD44, EpCAM and EGFR in negatively enriched, peripheral blood samples from metastatic breast cancer patients. To date, over 10 metastatic patient samples have been quantified and the study is ongoing. Initial quantification indicates that significant variability in expression levels exists in multiple markers including Her2 neu. Conclusions: Using multispectral technology, we have shown that CTCs are heterogeneous, with different CTCs having varying expression levels of EpCAM, HER2 neu, vimentin and EGFR. Further multiparameter quantitative characterization with up to 6 color analysis on the same cell is in progress.
A retrospective study of immunohistochemical detection of Ki67 and correlation with ER, PR, and HER2-neu status in invasive breast cancer.
Authors: Prakash Deepa Meda , Shivakumar Swarna , Bapsy P P
Published: March 2020

e21132

Background: Carcinoma breast is a heterogeneous group of disease. Several prognostic factors are known to predict the biological behavior and clinical outcome of breast cancer, including proliferation index, tumor size, age, histological grade, hormone receptors and Her2Neu status. Of theses proliferative index is the most important parameter in predicting tumor aggressiveness and prognosis. Ki67 is a nuclear protein, tightly linked to the cell cycle and is a marker of cell proliferation. High Ki67 usually correlates with a poor prognosis in breast cancer. Methods: This exploratory study is initiated whether Ki67 will be an additional prognostic factor. Immunohistochemical staining of Ki 67 was performed on formalin fixed paraffin embedded blocks of 50 cases of histologically confirmed breast cancer. Ki 67 expression based on distinct nuclear staining of tumor cells was assessed semiquantitatively as, nil (no immunostaining), low (≤10% immunopositivity), high (>10% immunopositivity) by light microscope. ER, PR, Her 2neu status and clinico-pathological parameters were analyzed and correlated with Ki67 immunostaining using the x2 test. Results: Our study showed high Ki 67 staining in 24 cases(48%), low immunostaining in 18 cases(36%) and was nil in 5 cases (10%) and was inconclusive in 3 cases(6%) (core biopsies). There was an inverse correlation of Ki 67 with ER, PR scores in 32 cases (64%) and positive correlation was seen in 28 cases (56%) with Her2 neu status. A good correlation was also observed with mitotic index in 37cases (74%) and histological grade in 29 cases (58%). Good correlation was seen with large tumor size in 24 cases(48%) and metastatic disease 9 cases (64%). Poor correlation was observed with node positive 22 cases (44%). High Ki 67 expression with good treatment response was seen in 13 cases (26%). Conclusions: Based on our study, Ki 67 score may prove to be an important prognostic factor in breast cancer and thereby, useful in making therapeutic decisions. But this is being explored in a large prospective study.
Analyses of the immunologically relevant gene transcript by qRT-PCR in primary cutaneous melanoma by the young and the elderly.
Authors: Hegde Upendra P. , Dadras Soheil Sam , Kerr Phil , Chakraborty Nitya , grant-Kels Jane , Mukherji Bijay
Published: March 2020

e21135

Background: To understand the dynamics of anti tumor immune responses (IR) in the setting of the ageing immune system, we examined immunologically relevant gene transcripts in formalin fixed paraffin embedded (FFPE) primary cutaneous melanoma (PCM) samples derived from five young (age <50 years) and five elderly (age >or=70 years) patients by qRT-PCR. Methods: A total of 10 FFPE tumor blocks were collected between 2004-2007 of patients with PCM at the University of Connecticut Health Center. Total RNA was extracted from FFPE tissue sections and random primed cDNA was generated. Pre amplification using pooled Taqman assays and Universal PCR Master Mix (Applied Biosystems, USA) was performed for 14 cycles to ensure an adequate amount of starting material for qPCR. Using 15 ng total RNA, the quantification of human IR genes (IL-2, INFg, TNFa, TGFb1, IL-10, Foxp3, and IDO-1) and transcripts of succinate dehydrogenase (SDHA) (as endogenous control) was accomplished by qRT-PCR amplification of a cDNA using TaqMan Universal PCR Master Mix and TaqMan gene expressions assay probes. All reaction assays including that of SDHA were performed in triplicate on an ABI 7500 Fast system (Applied Biosystems). Cycle threshold (Ct) values for each mRNA were normalized to SDHA (DCT) and represented as RQ=2-DCT. The average RQ was calculated for all ten samples in both cohorts and Student’s T-test was performed to evaluate the statistical significance of the IR gene expression. Results: While no significant difference of expression of Th1 cytokines (IL-2, IFNg, TNFa) or Th2/3 cytokines (IL-10, TGFb) could be detected within the two groups, FOXp3 expression was significantly decreased in PCM in the elderly. Conclusions: Our results suggest ageing may be associated with less regulatory responses in situ in PCM and that more expanded analyses of the nature of the IR in PCM are likely to provide information with implications on prognosis and immune based treatment options for elderly PCM subjects.
Preoteomic insights in oncology: What have we learned from measuring millions of proteins?
Authors: Brody Ed , Foreman Trudi , Gill Rosalynn D , Mehan Mike , Nikrad Malti , Ostroff Rachel , Stewart Alex , Williams Steve , Zichi Dom
Published: March 2020

e21142

Background: We have assayed over 8400 blood samples from 10 different malignancies, generating over 8 million quantitative protein measurements using SOMAscan, a new proteomic assay based on modified aptamers. It is capable of simultaneously measuring more than a thousand proteins from small volumes of biological samples such as plasma, tissues or cells. Methods: Bioinformatic analysis has revealed biomarker panels for early detection, differential diagnosis or prognosis in 6 of these cancers: lung, mesothelioma, ovarian, pancreatic, bladder, and renal cell carcinoma. Individual classifiers have been built with 7-13 biomarkers and AUC ranges of 0.84-0.99. These analyses have revealed unique biomarkers for each cancer, as well as common markers. Results: Several themes emerge from our extensive survey of oncology proteomic profiles. Many of the proteins correlate with disease burden or histopathological grade, and thus have utility for diagnosis, prognosis and recurrence monitoring. Parallels between serum and lung tumor tissue protein measurements demonstrate that many of the blood biomarkers arise from the tumor environment. Inflammatory response proteins are identified in many cancers and reveal important connections between tumor development and host response. Patterns can be found for biological processes in tumor biology, including angiogenesis, apoptosis, inflammation, metabolism and tumor invasion. Conclusions: With this new proteomics technology – which is fast, economical, highly scalable and flexible – we now have a powerful tool that enables proteome-wide proteomics, biomarker discovery and advancing the next generation of evidence-based, “personalized” diagnostics and therapeutics for oncology.
The detection of circulating tumor cells in patients with renal cell carcinoma using the AccuCyte platform and customized markers.
Authors: Ramirez Arturo , Tykodi Scott S. , Huang Sherry , Drovetto Nicholas Alan , Stilwell Jackie L.
Published: March 2020

e21148

Background: Enumeration of circulating tumor cells (CTCs) in peripheral blood represents a prognostic marker for select solid tumors. Commercially available technologies for CTC detection rely on enrichment of EpCAM-expressing cells. However, not all tumor histologies demonstrate high constitutive EpCAM expression, including renal cell carcinoma (RCC). New techniques for more complex immunophenotypic characterization of CTCs are therefore of great interest. Methods: The AccuCyte system for characterization of rare blood cell populations retains all nucleated cells for analysis. Fluorochrome-conjugated monoclonal antibodies (mAbs) are added to 3 ml peripheral blood samples processed by density separation with a specialized tube and float. Centrifugation deposits all nucleated cells (Buffy coat) onto a surface designed for imaging with an automated fluorescence microscope. Sensitivity for CTC identification was measured by spiking blood from healthy donors with defined numbers of MDA-MD-453 breast tumor cells analyzed with mAbs to EpCAM and CD45. Peripheral blood samples from 4 patients (pts) with metastatic RCC and whose primary tumor had been resected were analyzed with a mAb specific for the RCC-associated antigen carbonic anhydrase IX (CAIX), as well as mAbs for CD10, EGFR and EpCAM. Results: In 13 blood samples from healthy donors, an average of 89% of spiked-in MDA-MD-453 tumor cells defined as EpCAM+/CD45- were detected by AccuCyte analysis with linearity from 0 to 139 cells (slope=0.8899; r2=0.9351) and a sensitivity of approximately 1 CTC per 106 nucleated blood cells. 3 of 4 peripheral blood samples from pts with clear cell RCC contained putative CTCs (range 1-6 cells/ml) defined as CD45-/CAIX+ nucleated cells. 3 CTCs from one patient were also CD10+ or EGFR+. All putative CTCs were EpCAM-. Conclusions: The detection of EpCAM-/CAIX+ cells in RCC pts demonstrates the adaptability of the AccuCyte platform for CTC (or other rare blood cell) identification defined by customized markers. Further development of marker panels for CTC analyses in pts with RCC and other tumor histologies as well as automation of image analysis is ongoing.
Primary results from EMILIA, a phase III study of trastuzumab emtansine (T-DM1) versus capecitabine (X) and lapatinib (L) in HER2-positive locally advanced or metastatic breast cancer (MBC) previously treated with trastuzumab (T) and a taxane.
Authors: Blackwell Kimberly L. , Miles David , Gianni Luca , Krop Ian E , Welslau Manfred , Baselga José , Pegram Mark D. , Oh Do-Youn , Dieras Veronique , Olsen Steven R. , Fang Liang , Lu Michael W , Guardino Ellie , Verma Sunil
Published: March 2020

LBA1

The full, final text of this abstract will be available at abstract.asco.org at 12:01 AM (EDT) on Sunday, June 3, 2012, and in the Annual Meeting Proceedings online supplement to the June 20, 2012, issue of Journal of Clinical Oncology. Onsite at the Meeting, this abstract will be printed in the Sunday edition of ASCO Daily News.
A randomized, multicenter trial of epirubicin, oxaliplatin, and capecitabine (EOC) plus panitumumab in advanced esophagogastric cancer (REAL3).
Authors: Waddell Tom Samuel , Chau Ian , Barbachano Yolanda , Gonzalez de Castro David , Wotherspoon Andrew , Saffery Claire , Middleton Gary William , Wadsley Jonathan , Ferry David Raymond , Mansoor Wasat , Crosby Tom David Lewis , Coxon Fareeda Y , Smith David , Waters Justin S. , Iveson Timothy , Falk Stephen , Slater Sarah , Okines Alicia Frances Clare , Cunningham David
Published: March 2020

LBA4000

The full, final text of this abstract will be available at abstract.asco.org at 12:01 AM (EDT) on Saturday, June 2, 2012, and in the Annual Meeting Proceedings online supplement to the June 20, 2012, issue of Journal of Clinical Oncology. Onsite at the Meeting, this abstract will be printed in the Saturday edition of ASCO Daily News.
Updated analysis of the phase III, double-blind, randomized, multinational study of radium-223 chloride in castration-resistant prostate cancer (CRPC) patients with bone metastases (ALSYMPCA).
Authors: Parker Chris , Nilsson Sten , Heinrich Daniel , O'Sullivan Joe M. , Fossa Sophie D. , Chodacki Ales , Wiechno Pawel J. , Logue John P. , Seke Mihalj , Widmark Anders , Johannessen Dag Clement , Hoskin Peter , Bottomley David , Coleman Robert Edward , Vogelzang Nicholas J. , O'Bryan-Tear C. Gillies , Garcia-Vargas Jose E. , Shan Minghua , Sartor A. Oliver
Published: March 2020

LBA4512

The full, final text of this abstract will be available at abstract.asco.org at 12:01 AM (EDT) on Monday, June 4, 2012, and in the Annual Meeting Proceedings online supplement to the June 20, 2012, issue of Journal of Clinical Oncology. Onsite at the Meeting, this abstract will be printed in the Monday edition of ASCO Daily News.
Open-label phase III randomized controlled trial comparing taxane-based chemotherapy (Tax) with lapatinib (L) or trastuzumab (T) as first-line therapy for women with HER2+ metastatic breast cancer: Interim analysis (IA) of NCIC CTG MA.31/GSK EGF 108919.
Authors: Gelmon Karen A. , Boyle Frances , Kaufman Bella , Huntsman David , Manikhas Alexey , Di Leo Angelo , Martin Miguel , Schwartzberg Lee Steven , Dent Susan Faye , Ellard Susan , Tonkin Katia Sonia , Nagarwala Yasir M. , Pritchard Kathleen I. , Whelan Timothy Joseph , Nomikos Dora , Chapman Judy-Anne W. , Parulekar Wendy
Published: March 2020

LBA671

The full, final text of this abstract will be available at abstract.asco.org at 12:01 AM (EDT) on Sunday, June 3, 2012, and in the Annual Meeting Proceedings online supplement to the June 20, 2012, issue of Journal of Clinical Oncology. Onsite at the Meeting, this abstract will be printed in the Sunday edition of ASCO Daily News.
TAILOR: Phase III trial comparing erlotinib with docetaxel in the second-line treatment of NSCLC patients with wild-type (wt) EGFR.
Authors: Garassino Marina Chiara , Martelli Olga , Bettini Anna , Floriani Irene , Copreni Elena , Lauricella Calogero , Ganzinelli Monica , Marabese Mirko , Broggini Massimo , Veronese Silvio , Gherardi Giorgio , Longo Flavia , Fabbri Maria Agnese , Tomirotti Maurizio , Alabiso Oscar , Sarobba Maria Giuseppa , Labianca Roberto , Marsoni Silvia , Farina Gabriella , Scanni Alberto
Published: March 2020

LBA7501

The full, final text of this abstract will be available at abstract.asco.org at 12:01 AM (EDT) on Monday, June 4, 2012, and in the Annual Meeting Proceedings online supplement to the June 20, 2012, issue of Journal of Clinical Oncology. Onsite at the Meeting, this abstract will be printed in the Monday edition of ASCO Daily News.
Phase III, randomized, open-label, multicenter trial (BREAK-3) comparing the BRAF kinase inhibitor dabrafenib (GSK2118436) with dacarbazine (DTIC) in patients with BRAFV600E-mutated melanoma.
Authors: Hauschild Axel , Grob Jean Jacques , Demidov Lev V. , Jouary Thomas , Gutzmer Ralf , Millward Michael , Rutkowski Piotr , Blank Christian U. , Mirakhur Beloo , Guckert Mary E. , Swann R. Suzanne , Haney Patricia , Martin Anne-Marie , Ouellet Danielle , Grotzinger Kelly , Goodman Vicki L. , Chapman Paul B.
Published: March 2020

LBA8500^

The full, final text of this abstract will be available at abstract.asco.org at 12:01 AM (EDT) on Monday, June 4, 2012, and in the Annual Meeting Proceedings online supplement to the June 20, 2012, issue of Journal of Clinical Oncology. Onsite at the Meeting, this abstract will be printed in the Monday edition of ASCO Daily News.
METRIC phase III study: Efficacy of trametinib (T), a potent and selective MEK inhibitor (MEKi), in progression-free survival (PFS) and overall survival (OS) compared with chemotherapy (C) in patients (pts) with BRAFV600/k mutant advanced or metastatic melanoma (MM).
Authors: Robert Caroline , Flaherty Keith T. , Hersey Peter , Nathan Paul D. , Garbe Claus , Milhem Mohammed M. , Demidov Lev V. , Hassel Jessica C. , Rutkowski Piotr , Mohr Peter , Dummer Reinhard , Trefzer Uwe , Larkin James M. G. , Utikal Jochen , Casey Michelle , Sherman Laurie Jill , Crist Wendy A. , Wu Frank S. , Patel Kiran , Schadendorf Dirk
Published: March 2020

LBA8509

The full, final text of this abstract will be available at abstract.asco.org at 12:01 AM (EDT) on Monday, June 4, 2012, and in the Annual Meeting Proceedings online supplement to the June 20, 2012, issue of Journal of Clinical Oncology. Onsite at the Meeting, this abstract will be printed in the Monday edition of ASCO Daily News.
Emerging findings in the Cancer Research UK stratified medicine program.
Authors: Shaw Emily , Tuff Alice , Sharpe Rowena , Jones Louise K. , Turtiaien Tarita , Griffiths Michael , Butler Rachel , Gonzalez de Castro David , Mason Malcolm David , Collins V. Peter , Rae Frances , Evans T.R. Jeffry , Johnston Stephen R. D. , Rogan Jane , Hanby Andrew , Peach James , Johnson Peter W. M.
Published: March 2020

TPS10633^

Background: Molecular analysis of tumours may be used to identify those predicted to benefit from novel targeted therapies. The Cancer Research UK programme is piloting plans to apply such testing broadly across the UK healthcare system, linking molecular phenotype to clinical outcomes. Methods: The Stratified Medicine Programme (SMP) aims to develop a model for high quality, large-scale molecular characterization of cancer specimens through an initiative developed in partnership with AstraZeneca, Pfizer, the UK Department of Health and academic researchers. Phase One of the SMP is a two year feasibility study. It aims to demonstrate the submission of consented blood samples and sections of surplus diagnostic formalin-fixed paraffin-embedded tumour tissue from 9,000 patients at centres across the UK to one of three ‘technology hubs’ for mutation testing of genes of potential clinical interest (KRAS, BRAF, NRAS, PIK3CA, TP53, PTEN, TMPRSS2-ERG, EGFR, EML4-ALK and KIT) in six selected tumour types. The tests are technically validated and will be completed in clinically relevant timescales. Data including pathological and treatment information and clinical outcome is also collected for the recruited patients, linked to the genetic data and stored in a central data repository hosted within the National Cancer Registration Service. The study opened in September 2011 at 7 sites across the UK and by the end of 2011, 760 patientswith breast, lung, prostate, colorectal, ovarian cancer or metastatic malignant melanoma had consented to participate. 142 sets of molecular results had been returned to clinical teams. Updated figures will be presented at the meeting, by which time the programme is projected to have accrued 4000 subjects. By 2013, we hope to have developed a scalable model for routine, high quality, prospective molecular characterisation of tumours for NHS cancer patients, with consent for the collection, storage and research use of population-scale genetic and clinical outcome data. We will report the emerging results from the Stratified Medicines Programme and early insights into implications for wider implementation across the UK healthcare system.
Clinical pharmacokinetics (PK) of BMS-936558, a fully human anti-PD-1 monoclonal antibody.
Authors: Agrawal Shruti , Feng Yan , Kollia Georgia , Saeger Sally , Ullmann Martin , McDonald Dan , Gupta Ashok Kumar , Roy Amit , Masson Eric
Published: March 2020

TPS2622

Background: BMS-936558 is a fully human IgG4 monoclonal antibody targeted against human Programed Death-1 (PD1) receptor on activated T and B lymphocytes. Blocking of PD-1 prevents these activated cells from becoming anergic, thereby maintaining anti-tumor immunologic activity. Methods: The PK of BMS-936558 was characterized with data from a single ascending dose (SAD) and a multiple ascending dose (MAD) study in subjects with advanced solid malignancies. Subjects received a single IV infusion of 0.3 to 10 mg/kg BMS-936558 in the SAD study (MDX-1106-01 Study) and 0.1 to 10 mg/kg BMS-936558 every two weeks in the MAD study (MDX-1106-03/CA209003 Study). The PK of BMS-936558 was characterized by non-compartmental analysis of intensively sampled PK data from a SAD study, as well as by population PK analysis of available intensive and sparse PK data from the SAD and MAD studies, respectively. Results: The PK of BMS-936558 is linear in the studied dose range with dose-proportional increase in Cmax and AUC with low to moderate (20-44%) inter-subject variability. Geometric mean clearance ranged from 0.13 - 0.19 mL/h/kg, whereas mean volume of distribution ranged from 83 -113 mL/kg. The range of mean terminal elimination half-life of BMS-936558 is 17 to 25 days which is consistent with half-life of endogenous IgG4. BMS-936558 PK was adequately described by a linear two-compartment model, and there was no evidence of a contribution of target mediated drug disposition to drug elimination within tested dose range. Body weight and gender are potentially clinically significant covariate for both clearance and volume of distribution (> 20% effect); and baseline CRP, LDH, and albumin are potentially clinically significant covariates for CL. The body weight normalized dosing employed produces trough concentrations that are approximately constant over a wide range of body weights. Conclusions: The pharmacokinetics of BMS-936558 is dose-proportional and body weight normalized dosing is appropriate to ensure consistent exposure over a wide range of body weights.