scholarly journals BSCI-15. METASTATIC BRAIN TUMOR TARGETING PEPTIDES ISOLATED THROUGH PHAGE DISPLAY BIOPANNING AGAINST BRAIN METASTASIS-INITIATING CELLS

2019 ◽  
Vol 1 (Supplement_1) ◽  
pp. i4-i4
Author(s):  
JongMyung Kim ◽  
James Liu
Keyword(s):  
Lung Cancer ◽  
Brain Metastasis ◽  
Phage Display ◽  
Metastatic Brain Tumors ◽  
Serum Free ◽  
Phage Display Biopanning ◽  
Free Media ◽  
In Vivo Phage Display

Abstract To effectively target metastatic brain tumors (MBTs), the paradigm of treating MBTs after visualization on clinical imaging needs to be shifted to an understanding of the mechanisms that drive the formation and maintenance of brain metastasis-initiating cells (BMICs). Targeting this tumor sub-population, which may form as a result from activation of epithelial-mesenchymal transition, may allow for more effective means of isolating and targeting brain metastasis. In order to isolate BMICs, we have harvested cells from patient derived MBTs originating from lung cancer and cultured the cells using serum-free media conditions. In vivo phage display biopanning was used to isolate 12-amino acid length peptides that specifically target BMICs. Of the peptides recovered, one peptide, LBM4, was tested for specificity of binding to MBTs through in vitro and in vivo binding assays. When comparing patient derived metastatic brain tumors cells against brain metastasis cell lines, we found that both types of cells demonstrated similar morphology when grown in serum media conditions, but when grown in serum-free media, both demonstrated a tumor sphere morphology, similar to a stem cell-like state. LBM4 demonstrated specific binding to MBT cells over primary lung cancer cells in vitro through flow cytometry analysis and immunocytochemistry. Fluorescent tagged LBM4 intravenously injected into mice harboring intracranial BM demonstrated peptide localization to the tumor within the intracranial cavity visualized with live animal imaging. In vivo phage display biopanning is an effective tool that is able to isolate cell specific targeting peptides. MBT targeting peptides can potentially result in a shifting of the clinical treatment paradigm of brain metastases, through the development of more effective targeted therapeutics aimed at BMICs, as well as improving detection of MBT cells which may result in earlier tumor visualization as well as delineation of tumor recurrence versus radiation effects.

scholarly journals STEM-23. METASTATIC BRAIN TUMOR IMAGING THROUGH TARGETED PEPTIDES ISOLATED VIA PHAGE DISPLAY BIOPANNING AGAINST BRAIN METASTASIS-INITIATING CELLS

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi238-vi238
Author(s):  
JongMyung Kim ◽  
James Liu
Keyword(s):  
Lung Cancer ◽  
Brain Metastasis ◽  
Brain Metastases ◽  
Phage Display ◽  
Tumor Imaging ◽  
Mesenchymal Transition ◽  
Phage Display Biopanning ◽  
In Vivo Phage Display

Abstract To effectively target metastatic brain tumors (MBTs), the paradigm of initiating treatment against MBTs following detection on clinical imaging needs to be shifted to an understanding of the mechanisms that drive the formation and maintenance of brain metastasis-initiating cells (BMICs). Targeting this tumor sub-population, which may form as a result of activation of the epithelial-mesenchymal transition, may allow for more effective means of understanding and targeting brain metastases. In order to isolate BMICs, we have harvested cells from patient derived MBTs originating from lung cancer and cultured the cells using stem cell media conditions. We then performed in vitro and in vivo phage display biopanning to isolate 12-amino acid length peptides that specifically target BMICs. Several peptides were isolated from both in vitro and in vivo biopanning strategies. Of the peptides recovered, one peptide, LBM4, demonstrated specific binding to MBT cells over primary lung cancer cells in vitro through flow cytometry analysis and immunocytochemistry. Fluorescent tagged LBM4 intravenously injected into mice harboring intracranial brain metastases demonstrated peptide localization to the tumor within the intracranial cavity visualized with live animal imaging. Peptide imaging of tumor corresponded to MRI imaging confirming that the peptides could serve as an alternative to tumor imaging, with the potential for greater sensitivity resulting from the cellular targeting of MBTs. Our results demonstrate that we can use a combination of in vitro and in vivo phage display biopanning to isolate cell specific targeting peptides. MBT targeting peptides can potentially result in a shifting of the clinical treatment paradigm of brain metastases, through the development of more effective targeted therapeutics aimed at BMICs, as well as improving detection of MBT cells which may result in earlier tumor visualization, as well as delineation of tumor recurrence versus radiation effects.

scholarly journals CBIO-20GLIOBLASTOMA SPECIFIC PROTEINS IDENTIFIED THROUGH COMBINED IN VITRO AND IN VIVO PHAGE DISPLAY BIOPANNING

2015 ◽  
Vol 17 (suppl 5) ◽  
pp. v59.1-v59
Author(s):  
Ping Huang ◽  
Qiulian Wu ◽  
Jeremy Rich ◽  
James Liu
Keyword(s):  
Phage Display ◽  
Specific Proteins ◽  
Phage Display Biopanning ◽  
In Vivo Phage Display

scholarly journals Isolation, Culture and Identification of Choriocarcinoma Stem-Like Cells from the Human Choriocarcinoma Cell-Line JEG-3

2016 ◽  
Vol 39 (4) ◽  
pp. 1421-1432 ◽  
Author(s):  
Jingting Cai ◽  
Tianfang Peng ◽  
Jing Wang ◽  
Jingli Zhang ◽  
Hui Hu ◽  
...  
Keyword(s):  
Cell Line ◽  
Colony Formation ◽  
Soft Agar ◽  
Serum Free ◽  
Significant Difference ◽  
Free Media ◽  
Proliferative Ability ◽  
Choriocarcinoma Cell Line

Background/Aims: Cancer stem cells (CSCs) exhibit enhanced proliferative capacity and resistance to chemotherapy; however, choriocarcinoma CSCs have not yet been reported. In this study the human choriocarcinoma cell line JEG-3 was cultured in serum free media, and the characteristics of suspension and parental adherent JEG-3 cells were compared. Methods: Cell proliferation, colony-formation, soft agar clonogenicity, and transwell invasion assays were performed in vitro, and tumor xenografts in BALB/c nude mice were used to evaluate stem cell properties. Results: In serum-supplemented medium (SSM), JEG-3 cells were 4.51 ± 1.71% CD44+, 7.67 ± 2.67% CD133+, and 13.85 ± 2.95% ABCG2+. In serum-free medium (SFM), the expression of these markers increased to 53.08 ± 3.15%, 47.40 ± 2.67%, and 78.70 ± 7.16%, respectively. Moreover, suspension JEG-3 cells exhibited enhanced colony-formation capability as well as invasive and proliferative ability in vitro, alongside enhanced tumorigenic properties in vivo. Suspension JEG-3 cells also exhibited resistance to the chemotherapeutic drugs methotrexate, fluorouracil and etoposide. When seeded in serum supplemented medium, suspension JEG-3 cells readopted an adherent phenotype and continued to differentiate with no significant difference in the morphology between suspension and parent cells. Conclusion: In this study, choriocarcinoma stem-like cells (CSLCs) were isolated from the human choriocarcinoma JEG-3 cell line by SFM culture and characterized.

An Extract of Morinda citrifolia Interferes with the Serum-Induced Formation of Filamentous Structures in Candida albicans and Inhibits Germination of Aspergillus nidulans

2006 ◽  
Vol 34 (03) ◽  
pp. 503-509 ◽  
Author(s):  
Saswati Banerjee ◽  
Andrew D. Johnson ◽  
Katalin Csiszar ◽  
Daniel L. Wansley ◽  
Paul McGeady
Keyword(s):  
Candida Albicans ◽  
Morphological Change ◽  
Water Soluble ◽  
Morinda Citrifolia ◽  
Filamentous Form ◽  
Serum Free ◽  
Therapeutic Value ◽  
Free Media

An aqueous extract of Morinda citrifolia was shown to interfere with the serum-induced morphological conversion of Candida albicans from a cellular yeast to a filamentous form in vitro. The conversion of C. albicans from a cellular yeast to a filamentous form in vivo is associated with pathogenicity. No significant effect on growth in serum-free media was seen at the concentrations used to interfere with the morphological change. The same extract also inhibited the germination of Apergillus nidulans spores. These results demonstrate that M. citrifolia contains a water-soluble component or components that interfere with the morphological conversion of C. albicans and the germination of A. nidulans and may have potential therapeutic value with regard to candidiasis and aspergillosis.

scholarly journals STEM-02. IN VIVO PHAGE DISPLAY IDENTIFIES PEPTIDE TARGETING N-CADHERIN ON GLIOMA STEM CELLS

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii196-ii196
Author(s):  
Marine Potez ◽  
Jongmyung Kim ◽  
Chunhua She ◽  
Neelkamal Chaudhary ◽  
James Liu
Keyword(s):  
Stem Cells ◽  
Phage Display ◽  
Mass Spectroscopy ◽  
Peptide Binding ◽  
Glioma Stem Cells ◽  
Binding Partner ◽  
Specific Targeting ◽  
In Vivo Phage Display

Abstract Glioblastoma (GBM) is the most aggressive primary brain tumor with high mortality rates and resistance to conventional therapy. Glioma stem cells (GSCs) comprise a sub-population of glioma tumor cells with the ability of self-renewal and tumor recapitulation, and may be responsible for GBM’s treatment resistant properties. Identification of surface receptors that are novel and specific to GSCs may be the key to the development of effective therapeutic strategies. We have selected a GSC specific targeting peptide isolated through in vitro and in vivo phage display biopanning. This screening technique allowed us to determine a peptide (GBM-IC2) which binds specifically to GSCs in vitro, and to GBM tissue in vivo. Although this screening process allows for isolation of cell specific targeting peptides, it does so without identification of the cellular binding partner. Given the specificity of the peptide, identification of the cellular receptor may allow for discovery of novel markers to identify GSCs. To identify the peptide binding partner of GBM-IC2, the biotinylated peptide was incubated with GSC protein lysate. The peptide, along with its binding partner, was isolated using streptavidin agarose resin. The binding partner protein was then identified using mass spectroscopy. This revealed N-cadherin (CDH2) as a potential binding partner for the GBM-IC2 peptide. GBM-IC2 demonstrated specificity for targeting CDH2 compared to control peptide using ELISA. Lentiviral induced overexpression of CDH2 in HEK293 cells allowed for GBM-IC2 peptide binding. Competition assay was performed by applying anti-CDH2 antibody to GBM-IC2 peptide and GSCs in culture. Application of anti-CDH2 antibody decreased peptide binding to GSCs, confirming CDH2 as the binding partner for GBM-IC2. These results demonstrate that cell specific targeting peptides isolated through phage display may lead to the isolation of novel cell specific proteins through immunoprecipitation isolation and mass spectroscopy analysis.

scholarly journals STEM-14. GLIOBLASTOMA STEM CELL TARGETING CHIMERIC ANTIGEN RECEPTOR T CELLS MODIFIED USING PHAGE DISPLAY ISOLATED PEPTIDES

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii199-ii199
Author(s):  
Marine Potez ◽  
Sebastian Snedal ◽  
Jongmyung Kim ◽  
Konrad Thorner ◽  
Maria Cecilia Ramello ◽  
...  
Keyword(s):  
T Cells ◽  
Phage Display ◽  
Conventional Therapy ◽  
Annexin V ◽  
Primary Brain Tumor ◽  
Car T Cells ◽  
Car T ◽  
Phage Display Biopanning

Abstract Glioblastoma (GBM) is the most aggressive primary brain tumor with high mortality rates and resistance to conventional therapy. Their resistance to conventional therapy has been attributed to the presence of cancer stem cells (CSCs), a sub-population of tumor cells capable of self-renewal and tumor initiation. Developing novel strategies to specifically target GSCs may allow more effective therapeutic strategies. Using in vivo phage display biopanning, we have identified several peptides with the potential to selectively target and bind GSCs. We wished to leverage the GSC targeting properties of the peptides to augment therapeutic delivery vehicles for the development of novel targeting strategies. We used a combination of GSC targeting peptides to modify the antigen-binding domain of chimeric antigen receptors, by arranging the peptides in tandem at the N-terminus of the CAR molecule. These tandem peptides were tested for binding to GSCs in vitro and in vivo. The functionality of the CAR-T cells was evaluated by measuring cytokine release in the supernatant after overnight co-culture through ELISA. Apoptosis was evaluated by flow cytometry with Annexin V staining. Two different GSC-targeting peptide CAR-T cells demonstrated specific targeting GSCs. Following co-culture with GSCs, GSC targeting CAR-T cells were activated with release of Interferon gamma and subsequently induced GSCs specific apoptosis. These results demonstrate the use of phage display biopanning to isolate GSC targeting peptides which may be used to develop novel GBM specific cytotoxic therapies.

Overcoming the acquired resistance to gefitinib in lung cancer brain metastasis in vitro and in vivo

2021 ◽  
Author(s):  
Zhongwei Liu ◽  
Neal Shah ◽  
Kent L. Marshall ◽  
Samuel A. Sprowls ◽  
Pushkar Saralkar ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Brain Metastasis ◽  
Acquired Resistance

scholarly journals N6-methyladenosine induced miR-143-3p promotes the brain metastasis of lung cancer via regulation of VASH1

2019 ◽  
Vol 18 (1) ◽  
Author(s):  
Hongsheng Wang ◽  
Qianqian Deng ◽  
Ziyan Lv ◽  
Yuyi Ling ◽  
Xue Hou ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Brain Metastasis ◽  
Expression Profiles ◽  
Primary Lung Cancer ◽  
Lung Cancers ◽  
Lung Cancer Patients ◽  
Cancer Pathogenesis ◽  
Cancer Tissues

Abstract Background Brain metastasis (BM) is one of the principal causes of mortality for lung cancer patients. While the molecular events that govern BM of lung cancer remain frustrating cloudy. Methods The miRNA expression profiles are checked in the paired human BM and primary lung cancer tissues. The effect of miR-143-3p on BM of lung cancer cells and its related mechanisms are investigated. Results miR-143-3p is upregulated in the paired BM tissues as compared with that in primary cancer tissues. It can increase the invasion capability of in vitro blood brain barrier (BBB) model and angiogenesis of lung cancer by targeting the three binding sites of 3’UTR of vasohibin-1 (VASH1) to inhibit its expression. Mechanistically, VASH1 can increase the ubiquitylation of VEGFA to trigger the proteasome mediated degradation, further, it can endow the tubulin depolymerization through detyrosination to increase the cell motility. m6A methyltransferase Mettl3 can increase the splicing of precursor miR-143-3p to facilitate its biogenesis. Moreover, miR-143-3p/VASH1 axis acts as adverse prognosis factors for in vivo progression and overall survival (OS) rate of lung cancer. Conclusions Our work implicates a causal role of the miR-143-3p/VASH1 axis in BM of lung cancers and suggests their critical roles in lung cancer pathogenesis.

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