GLORIA

GEOMAR Library Ocean Research Information Access

X

Your email was sent successfully. Check your inbox.

An error occurred while sending the email. Please try again.

Proceed reservation?

Export
  • 1
    Online Resource
    Online Resource
    Abstract 4959: Multi-level analysis of prostate cancer circulating tumor cells allowing IHC-based identification, 6-parameter fluorescence phenotyping, and individual cell molecular analysis
    American Association for Cancer Research (AACR) ; 2016
    In:  Cancer Research Vol. 76, No. 14_Supplement ( 2016-07-15), p. 4959-4959
    Details
    In: Cancer Research, American Association for Cancer Research (AACR), Vol. 76, No. 14_Supplement ( 2016-07-15), p. 4959-4959
    Abstract: Background. Analysis of circulating tumor cells (CTC) allows non-invasive investigation of prostate cancer biology and response to treatment. The primary level of analysis is the CTC count, which has been demonstrated to be prognostic of outcome. Deeper characterization of CTC phenotype and pertinent biomarkers can confirm cancer lineage and identify drug targets or drug-resistance markers. Single cell analysis of individual CTCs can provide genomic insight into cancer heterogeneity. RareCyte has developed AccuCyte® - CyteFinder® (AC-CF), an integrated technology platform for highly sensitive visual identification and retrieval of rare cells in blood by both immunohistochemistry (IHC) and immunofluorescence (IF) staining. Recently we have developed technology allowing 6-marker assays for broader phenotypic analysis. Methods. Normal human whole blood samples were spiked with prostate cancer lines as model CTCs (mCTCs). Blood samples from University of Washington patients with advanced prostate cancer were collected under an IRB-approved protocol. Blood was processed using AccuCyte and the nucleated cell fraction was collected and spread onto microscope slides. Slides were stained on an automated stainer using (1) an IHC assay for cytokeratin, (2) a standard 4-wavelength IF assay (DAPI, CD45, cytokeratin and EpCAM) or (3) a novel 6-parameter IF assay using SYTOX-Orange (nuclear stain), cytokeratin, EpCAM, androgen receptor (AR), prostate-specific membrane antigen (PSMA) and CD45. An assay for AR variant 7 (ARv7) was applied to samples with mCTCs with the ARv7 splice variant. Percent recovery of IHC-stained slides (by blinded pathologist review) was compared to IF-stained slides (by CyteFinder image analysis). Individual IHC-stained CTCs were retrieved after on-slide visual identification and re-visualized after dispensing for confirmation. Whole genome amplification (WGA) of retrieved cells was performed, followed by X- and Y-chromosome gene-specific PCR. Results. There was strong linear correlation between IF and IHC counts of mCTCs over a range of ∼25 - 100 cells/mL (R2 = 0.99). The 6-parameter IF assay was successfully applied to mCTC and clinical samples. AR and PSMA were co-expressed in the majority of epithelial-marker positive clinical CTCs. The ARv7 assay identified mCTCs that express the splice variant. Individual IHC-stained mCTCs spiked into female donor blood were demonstrated to be male after WGA and PCR. Conclusions. Light microscope identification of IHC cytokeratin-positive mCTCs approximated IF identification. 6-parameter phenotyping of prostate cancer CTCs is feasible and allows identification of lineage-specific markers. IHC-stained cells can be individually retrieved from slides for genome amplification and molecular analysis. Citation Format: Daniel Campton, Rachel Needham, Josh Nordberg, Arturo Ramirez, Nick Drovetto, Alisa Clein, Daniel E. Sabath, Jackie Stilwell, Eric Kaldjian. Multi-level analysis of prostate cancer circulating tumor cells allowing IHC-based identification, 6-parameter fluorescence phenotyping, and individual cell molecular analysis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4959.
    Type of Medium: Online Resource
    ISSN: 0008-5472 , 1538-7445
    URL: Article
    DOI: 10.1158/1538-7445.AM2016-4959
    RVK:
    XA 36000
    RVK:
    XA 10000
    Language: English
    Publisher: American Association for Cancer Research (AACR)
    Publication Date: 2016
    detail.hit.zdb_id: 2036785-5
    detail.hit.zdb_id: 1432-1
    detail.hit.zdb_id: 410466-3
    Location Call Number Limitation Availability
    BibTip Others were also interested in ...
    Online Resource
  • 2
    Online Resource
    Online Resource
    Abstract B21: Probing tumor heterogeneity and immune infiltration with Cyclic Immunofluorescence (CycIF), a robust, multiplexed imaging method
    American Association for Cancer Research (AACR) ; 2017
    In:  Cancer Immunology Research Vol. 5, No. 3_Supplement ( 2017-03-01), p. B21-B21
    Details
    In: Cancer Immunology Research, American Association for Cancer Research (AACR), Vol. 5, No. 3_Supplement ( 2017-03-01), p. B21-B21
    Abstract: Tumor microenvironment, defined by surrounding stromal/immune cells as well as blood vessels, plays an important role in disease progression and therapy resistance. Increasing understanding of the heterogeneity in both tumor and its microenvironment will be crucial to development more effective therapies. Recently, several studies employing state-of-the-art single-cell sequencing methods reveal enormous complexity in tumor microenvironment. However, the spatial information and cell-to-cell interaction could not be preserved in these dissociated cells. Immunofluorescence has been widely used in different fields of biological and medical research for decades. The ability to obtain in situ and single-cell information makes this technique particularly important in tumor biology. However, biochemical and optical constraints limit the number of signals that could be captured simultaneously within the same sample. We have developed the CycIF (Cyclic Immunofluorescence), an easy and low-cost method to increase the multiplexity of conventional immunofluorescence. The CycIF method has been first applied in pre-clinical drug discovery, cancer and stem cell biology with adherent cell cultures. In here, we modified the original CycIF method for IHC/IF on FFPE samples, and used that to probe tumor heterogeneity, microenvironment and immune infiltration in various types of tumors. Up to 30 different antigens/markers could be simultaneously detected in different tumor samples, and these makers represent a wide range of biological processes, including the key molecules for lymphocyte surface makers (CD45, CD4, CD8, CD20, CD11c), immune checkpoints (PD-1, PD-L1), stromal/EMT proteins (E-Cadherin, Vimentin), cell cycle regulators (CycD1, PCNA, Ki67, pRB, p21/CIP), signaling proteins (EGFR, pERK, pS6) and apoptosis mediators (p53, Bax, Bcl-2, Survivin). Our study not only provides the first detailed map of tumor and its immune microenvironment, but also illustrates a robust multiplexed imaging platform for probing tumor heterogeneity. Citation Format: Jia-Ren Lin, Benjamin Izar, Sabrina Hawthorne, Josh Nordberg, Eric Kaldjian, Peter Sorger. Probing tumor heterogeneity and immune infiltration with Cyclic Immunofluorescence (CycIF), a robust, multiplexed imaging method. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr B21.
    Type of Medium: Online Resource
    ISSN: 2326-6066 , 2326-6074
    URL: Article
    DOI: 10.1158/2326-6074.TUMIMM16-B21
    Language: English
    Publisher: American Association for Cancer Research (AACR)
    Publication Date: 2017
    detail.hit.zdb_id: 2732517-9
    Location Call Number Limitation Availability
    BibTip Others were also interested in ...
    Online Resource
  • 3
    Online Resource
    Online Resource
    Abstract 3375: Molecular subtyping of circulating tumor cells in patients with small cell lung cancer
    American Association for Cancer Research (AACR) ; 2022
    In:  Cancer Research Vol. 82, No. 12_Supplement ( 2022-06-15), p. 3375-3375
    Details
    In: Cancer Research, American Association for Cancer Research (AACR), Vol. 82, No. 12_Supplement ( 2022-06-15), p. 3375-3375
    Abstract: Background: Recently, a new molecular classification of small cell lung cancer (SCLC), defined by expression of four key transcription regulators - ASCL1, NEUROD1, YAP1 and POU2F3 - has been proposed. These molecular subtypes may confer differential biology and therapeutic vulnerabilities, however, studies to date have been constrained by the paucity of available longitudinal tumor biopsy samples obtained from patients with SCLC. Herein, we utilized circulating tumor cells (CTCs) to characterize subtype marker heterogeneity at the time of diagnosis and evaluate the dynamic nature of marker expression during treatment in patients with SCLC. Methods: Informed consent was obtained from patients with SCLC using an IRB approved protocol (IRB#030763). We have collected blood samples from 32 patients, including treatment naïve samples from the entire cohort as well as on-treatment samples (median of 4 collections per patient). We utilized the RareCyte rare-cell analysis platform to isolate and quantify CTCs, which were defined as CK/EpCAM positive and CD45 negative. Each sample was also evaluated for expression of neuroendocrine markers (ASCL1, NEUROD1) and non-neuroendocrine markers (YAP1, POU2F3). Each set of neuroendocrine and non-neuroendocrine markers were co-stained along with the CTC markers. Mean fluorescence intensity of each marker was extracted using a python program. Results: To date, we have analyzed 22/32 (69%) treatment naïve samples, including n=6 patients with early stage disease and n=16 patients with advanced/metastatic disease. We detected CTCs from 16/22 (73%) patients, with quantities ranging from 1 - 3406 (median = 66) per 7.5ml of blood. CTCs were positive for the neuroendocrine markers ASCL1 in 16/16 (100%) and NEUROD1 in 10/16 (62%); all NEUROD1 positive CTCs were also positive for ASCL1. The non-neuroendocrine markers YAP1 and POU2F3 were detected in 6/16 (37%) and 11/16 (69%) samples, respectively, including YAP1/POU2F3 double positives in 6/16 (37%). Analysis of on-treatment samples is ongoing. Conclusion: We have developed a protocol for monitoring expression of subtype markers on CTCs isolated from patients with SCLC. We found CTCs expressing both neuroendocrine and non-neuroendocrine markers at the time of diagnosis. Ongoing work will attempt to delineate the dynamic nature of subtype marker expression during therapy, to correlate with clinical outcomes. These studies have the potential to offer critical insights regarding the heterogeneity and evolution of SCLC, a tumor type in which novel disease insights and innovative treatment strategies are urgently needed to improve patient survival. Citation Format: Prasad R. Kopparapu, Melinda Duplessis, Edward Lo, Yingjun Yan, Wade T. Iams, Josh Nordberg, Arturo Ramirez, Tad George, Christine Lovly. Molecular subtyping of circulating tumor cells in patients with small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3375.
    Type of Medium: Online Resource
    ISSN: 1538-7445
    URL: Article
    DOI: 10.1158/1538-7445.AM2022-3375
    Language: English
    Publisher: American Association for Cancer Research (AACR)
    Publication Date: 2022
    detail.hit.zdb_id: 2036785-5
    detail.hit.zdb_id: 1432-1
    detail.hit.zdb_id: 410466-3
    Location Call Number Limitation Availability
    BibTip Others were also interested in ...
    Online Resource
Close ⊗
This website uses cookies and the analysis tool Matomo. More information can be found here...