Browsing by Author "Targen, Seniye"

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Open Access
Development of a novel zebrafish xenograft model in ache mutants using liver cancer cell lines
(Nature Publishing Group, 2018) Avcı, M. Ender; Keskus, Ayşe Gökçe; Targen, Seniye; Işılak, M. Efe; Öztürk, Mehmet; Çetin-Atalay, Rengül; Adams, Michelle M.; Konu, Özlen
Acetylcholinesterase (AChE), an enzyme responsible for degradation of acetylcholine, has been identified as a prognostic marker in liver cancer. Although in vivo Ache tumorigenicity assays in mouse are present, no established liver cancer xenograft model in zebrafish using an ache mutant background exists. Herein, we developed an embryonic zebrafish xenograft model using epithelial (Hep3B) and mesenchymal (SKHep1) liver cancer cell lines in wild-type and ache sb55 sibling mutant larvae after characterization of cholinesterase expression and activity in cell lines and zebrafish larvae. The comparison of fluorescent signal reflecting tumor size at 3-days post-injection (dpi) revealed an enhanced tumorigenic potential and a reduced migration capacity in cancer cells injected into homozygous ache sb55 mutants when compared with the wild-type. Increased tumor load was confirmed using an ALU based tumor DNA quantification method modified for use in genotyped xenotransplanted zebrafish embryos. Confocal microscopy using the Huh7 cells stably expressing GFP helped identify the distribution of tumor cells in larvae. Our results imply that acetylcholine accumulation in the microenvironment directly or indirectly supports tumor growth in liver cancer. Use of this model system for drug screening studies holds potential in discovering new cholinergic targets for treatment of liver cancers.
Open Access
Development of a specialized zebrafish xenotransplantation database and establishment of ALU-based tumor DNA quantification method in zebrafish: focus on models of overexpression and microenvironment
(2020-09) Targen, Seniye
Successful xenotransplantation of human cancer cells into zebrafish host marked a new era in cancer research enabling high throughput in vivo screens. Zebrafish xenotransplantation literature continues to rapidly accumulate, and this necessitates the development of an interactive database for accommodating the collective data for fined-tune search, visualization and statistical representation purposes. Herein, I have introduced an interactive database, ZenoFishDb v1.1 (https://konulab.shinyapps.io/zenofishdb), housing manually curated details on molecularly-modified cell transplantations, PDXs, stem cell and cancer stem cell transplantation studies as well as transplantation studies bearing modified host details. The database projects collected data in a table format via various attributes and provides graphical representation of the curated details as well as statistical analyses yielding information on incorporated numbers and frequencies of selected attributes; hence can be used for reviews and designing new experiments. Zebrafish PDX studies are separately conceptualized and displayed through ZenoFishDb v1.1. Development of the ZenoFishDb v1.1 leads to a better understanding of tumor analysis methods such as assessment of proliferation and/or tumor growth in xenotransplantation studies and further marks the need for development of novel methods for precise quantification of tumor size. In the light of these findings, I have helped establish a novel qRT-PCRbased proliferation assessment method for xenografts in zebrafish, adapted from previous mouse xenotransplantation studies. Herein, the use and precision of ALU repeat-based quantification of transplanted liver cancer cells in genotyped zebrafish ache mutants and wildtype siblings was shown exemplifying microenvironment as an important factor for tumor growth. I further demonstrated the power of ALU repeatbased quantification in Mineralocorticoid Receptor (MR) overexpressing breast cancer cells (MCF7) injected to the transparent casper zebrafish as a case study. First, I demonstrated that MR expression and signaling was important in breast cancer biology and prognosis based on in silico TCGA and custom RNA sequencing as well as other in vitro and ex vivo assays. I further showed that results obtained from ALU repeatbased quantification of tumor growth in MR-overexpressing MCF7 cells paralleled fluorescent image-based intensity measurements while the former being relatively less time-consuming and more high-throughput. In this study, accurate quantification of MR overexpression in xenografts was also successfully performed by a cDNA-specific primer pair; and the rate of tumor growth based on image analysis, did not correlate with the amount of MR DNA in casper fish xenografts. However, MCF7 cells overexpressing MR exhibited lower cell viability in vitro although some of these effects were due to empty vector (EV) integration. Accordingly, tumor size in xenografts of naïve, EV- and MR-transfected MCF7 cells injected into pigmented AB larvae were quantified for ALU-repeats yet no significant difference was observed due to high within-group variability in vivo. Future studies are needed to assess the role of varying the volume and placement of injected cells along with the amount of MR gene transfected on tumor growth in vivo.
Open Access
Zebrafish xenotransplantation models for studying gene function and drug treatment in hepatocellular carcinoma
(2022-01-15) Konu, Ozlen; Targen, Seniye
Introduction Zebrafish is a promising model organism for human disease including hepatocellular cancer (HCC). Recently, zebrafish has emerged also as a host for xenograft studies of liver cancer cell lines and patient derived tumors of HCC. Zebrafish embryos enable drug screening and gene function studies of xenografted cells via ease of microinjection and visualization of tumor growth and metastasis. Objectives In this review, we aimed to overview zebrafish HCC and liver cancer xenotransplantation studies focusing on ‘gene functional analysis’ and ‘drug/chemical screening’. Methods Herein, a comprehensive literature search was performed for liver and HCC xenografts in zebrafish on PubMed using different key words and filters for molecular modifications or drug exposure. Results Our literature search revealed around 250 studies which were filtered and summarized in a table (Table 1) revealing comprehensive collection of experimental and technical details on microinjection, injected cell lines, molecular modifications of injected cells, types and doses of drug treatments as well as biological assessments. Conclusion This review provides a platform for HCC and liver xenografts and highlights studies performed to understand gene functionality and drug efficacy in vivo in zebrafish.
Open Access
ZenoFishDb v1.1: a database for xenotransplantation studies in zebrafish
(Mary Ann Liebert, 2020) Targen, Seniye; Kaya, Tuğberk; Avcı, M. E.; Güneş, Damla; Keşküş, Ayşe Gökçe; Konu, Özlen
Rapidly accumulating literature has proven feasibility of the zebrafish xenograft models in cancer research. Nevertheless, online databases for searching the current zebrafish xenograft literature are in great demand. Herein, we have developed a manually curated database, called ZenoFishDb v1.1 (https://konulab.shinyapps.io/zenofishdb), based on R Shiny platform aiming to provide searchable information on ever increasing collection of zebrafish studies for cancer cell line transplantation and patient-derived xenografts (PDXs). ZenoFishDb v1.1 user interface contains four modules: DataTable, Visualization, PDX Details, and PDX Charts. The DataTable and Visualization pages represent xenograft study details, including injected cell lines, PDX injections, molecular modifications of cell lines, zebrafish strains, as well as technical aspects of the xenotransplantation procedures in table, bar, and/or pie chart formats. The PDX Details module provides comprehensive information on the patient details in table format and can be searched and visualized. Overall, ZenoFishDb v1.1 enables researchers to effectively search, list, and visualize different technical and biological attributes of zebrafish xenotransplantation studies particularly focusing on the new trends that make use of reporters, RNA interference, overexpression, or mutant gene constructs of transplanted cancer cells, stem cells, and PDXs, as well as distinguished host modifications.