Publications by authors named "Felix Oppel"

18 Publications

  • Page 1 of 1

Sensorineural Hearing Loss After Balloon Eustachian Tube Dilatation.

Front Surg 2021 5;8:615360. Epub 2021 Mar 5.

Medical School OWL, Bielefeld University, Klinikum Bielefeld Mitte, Bielefeld, Germany.

Eustachian tube function is of central importance for the ventilation of the middle ear. A dysfunction can be associated with chronic otitis media, and cholesteatoma. Balloon Eustachian tube dilatation (BET) is a treatment option used to solve eustachian tube dysfunction. Although BET is widely performed, little is known about the occurrence rate of the complications associated with BET. The aim of the present study was to observe the rate of sensorineural hearing loss (SNHL) after BET. We retrospectively evaluated in a chart review 1,547 patients and 2,614 procedures of BET performed in a single center between 2015 and 2019 using the Spiggle and Theis, Overath, Germany eustachian tube dilatation system. We observed seven cases of SNHL after BET. In two cases, the SNHL persisted, and in five cases, the SNHL was transient. In two cases of SNHL, a simultaneous tympanoplasty was performed. The overall rate of SNHL per procedure is 0.3%. The rate of permanent SNHL is 0.08%. BET has a low rate of SNHL. Rapid middle ear pressure changes are assumed to cause BET-related hearing loss.
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http://dx.doi.org/10.3389/fsurg.2021.615360DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7973464PMC
March 2021

Disulfiram Acts as a Potent Radio-Chemo Sensitizer in Head and Neck Squamous Cell Carcinoma Cell Lines and Transplanted Xenografts.

Cells 2021 Feb 28;10(3). Epub 2021 Feb 28.

Department of Otolaryngology, Head and Neck Surgery, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Berlin Institute of Health, Charité Campus Benjamin Franklin, Hindenburgdamm 30, 12203 Berlin, Germany.

The poor prognosis of locally advanced and metastatic head and neck squamous cell carcinoma (HNSCC) is primarily mediated by the functional properties of cancer stem cells (CSCs) and resistance to chemoradiotherapy. We investigated whether the aldehyde dehydrogenase (ALDH) inhibitor disulfiram (DSF) can enhance the sensitivity of therapy. Cell viability was assessed by the 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT) and apoptosis assays, and the cell cycle and reactive oxygen species (ROS) levels were evaluated by fluorescence-activated cell sorting (FACS). The radio-sensitizing effect was measured by a colony formation assay. The synergistic effects were calculated by combination index (CI) analyses. The DSF and DSF/Cu inhibited the cell proliferation (inhibitory concentration 50 (IC) of DSF and DSF/Cu were 13.96 μM and 0.24 μM). DSF and cisplatin displayed a synergistic effect (CI values were < 1). DSF or DSF/Cu abolished the cisplatin-induced G2/M arrest (from 52.9% to 40.7% and 41.1%), and combining irradiation (IR) with DSF or DSF/Cu reduced the colony formation and attenuated the G2/M arrest (from 53.6% to 40.2% and 41.9%). The combination of cisplatin, DSF or DSF/Cu, and IR enhanced the radio-chemo sensitivity by inducing apoptosis (42.04% and 32.21%) and ROS activity (46.3% and 37.4%). DSF and DSF/Cu enhanced the sensitivity of HNSCC to cisplatin and IR. Confirming the initial data from patient-derived tumor xenograft (PDX) supported a strong rationale to repurpose DSF as a radio-chemosensitizer and to assess its therapeutic potential in a clinical setting.
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http://dx.doi.org/10.3390/cells10030517DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7999545PMC
February 2021

Chronic inflammation of middle ear cholesteatoma promotes its recurrence via a paracrine mechanism.

Cell Commun Signal 2021 Feb 24;19(1):25. Epub 2021 Feb 24.

Department of Otolaryngology, Head and Neck Surgery, Medical School OWL Campus Klinikum Bielefeld, Bielefeld University, Teutoburger Str. 50, 33604, Bielefeld, Germany.

Background: Cholesteatoma disease is an expanding lesion in the middle ear. Hearing loss and facial paralysis alongside with other intracranial complications are found. No pharmaceutical treatment is available today and recurrence after surgical extraction occurs. We investigated possible TLR4-based mechanisms promoting recurrence and explore possible treatments strategies.

Methods: We isolated fibroblasts and epidermal stem cells from cholesteatoma tissue and healthy auditory canal skin. Subsequently, their expression under standard culture conditions and after stimulation with LPS was investigated by RT-qPCR. Cell metabolism and proliferation were analysed upon LPS treatment, with and without TLR4 antagonist. An indirect co-culture of fibroblasts and epidermal stem cells isolated from cholesteatoma tissue was utilized to monitor epidermal differentiation upon LPS treatment by RT-qPCR and immunocytochemistry.

Results: Under standard culture conditions, we detected a tissue-independent higher expression of IL-1β and IL-8 in stem cells, an upregulation of KGF and IGF-2 in both cell types derived from cholesteatoma and higher expression of TLR4 in stem cells derived from cholesteatoma tissue. Upon LPS challenge, we could detect a significantly higher expression of IL-1α, IL-1β, IL-6 and IL-8 in stem cells and of TNF-a, GM-CSF and CXCL-5 in stem cells and fibroblasts derived from cholesteatoma. The expression of the growth factors KGF, EGF, EREG, IGF-2 and HGF was significantly higher in fibroblasts, particularly when derived from cholesteatoma. Upon treatment with LPS the metabolism was elevated in stem cells and fibroblasts, proliferation was only enhanced in fibroblasts derived from cholesteatoma. This could be reversed by the treatment with a TLR4 antagonist. The cholesteatoma fibroblasts could be triggered by LPS to promote the epidermal differentiation of the stem cells, while no LPS treatment or LPS treatment without the presence of fibroblasts did not result in such a differentiation.

Conclusion: We propose that cholesteatoma recurrence is based on TLR4 signalling imprinted in the cholesteatoma cells. It induces excessive inflammation of stem cells and fibroblasts, proliferation of perimatrix fibroblasts and the generation of epidermal cells from stem cells thru paracrine signalling by fibroblasts. Treatment of the operation site with a TLR4 antagonist might reduce the chance of cholesteatoma recurrence. Video Abstract.
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http://dx.doi.org/10.1186/s12964-020-00690-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7903614PMC
February 2021

Insights into Nanomedicine for Immunotherapeutics in Squamous Cell Carcinoma of the head and neck.

Int J Biol Sci 2020 19;16(14):2506-2517. Epub 2020 Jul 19.

Department of Clinical Laboratory, Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital); Institute of Cancer and Basic Medicine (IBMC), Chinese Academy of Sciences. Hangzhou, P.R. China.

Immunotherapies such as immune checkpoint blockade benefit only a portion of patients with head and neck squamous cell carcinoma. The multidisciplinary field of nanomedicine is emerging as a promising strategy to achieve maximal anti-tumor effect in cancer immunotherapy and to turn non-responders into responders. Various methods have been developed to deliver therapeutic agents that can overcome bio-barriers, improve therapeutic delivery into the tumor and lymphoid tissues and reduce adverse effects in normal tissues. Additional modification strategies also have been employed to improve targeting and boost cytotoxic T cell-based immune responses. Here, we review the state-of-the-art use of nanotechnologies in the laboratory, in advanced preclinical phases as well as those running through clinical trials assessing their advantages and challenges.
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http://dx.doi.org/10.7150/ijbs.47068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7415431PMC
July 2020

inactivation in - and -deficient zebrafish accelerates the onset of malignant peripheral nerve sheath tumors and expands the spectrum of tumor types.

Dis Model Mech 2020 08 27;13(8). Epub 2020 Aug 27.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115, USA

Polycomb repressive complex 2 (PRC2) is an epigenetic regulator of gene expression that possesses histone methyltransferase activity. PRC2 trimethylates lysine 27 of histone H3 proteins (H3K27me3) as a chromatin modification associated with repressed transcription of genes frequently involved in cell proliferation or self-renewal. Loss-of-function mutations in the PRC2 core subunit SUZ12 have been identified in a variety of tumors, including malignant peripheral nerve sheath tumors (MPNSTs). To determine the consequences of loss in the pathogenesis of MPNST and other cancers, we used CRISPR-Cas9 to disrupt the open reading frame of each of two orthologous genes in zebrafish: and We generated these knockout alleles in the germline of our previously described (also known as )- and deficient zebrafish model of MPNSTs. Loss of significantly accelerated the onset and increased the penetrance of MPNSTs compared to that in control zebrafish. Moreover, in deficient zebrafish, we detected additional types of tumors besides MPNSTs, including leukemia with histological characteristics of lymphoid malignancies, soft tissue sarcoma and pancreatic adenocarcinoma, which were not detected in deficient control fish, and are also contained in the human spectrum of -deficient malignancies identified in the AACR Genie database. The -knockout tumors displayed reduced or abolished H3K27me3 epigenetic marks and upregulation of gene sets reported to be targeted by PRC2. Thus, these zebrafish lines with inactivation of in combination with loss of provide a model of human MPNSTs and multiple other tumor types, which will be useful for mechanistic studies of molecular pathogenesis and targeted therapy with small molecule inhibitors.
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http://dx.doi.org/10.1242/dmm.042341DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7473648PMC
August 2020

The Potential of Tumor Debulking to Support Molecular Targeted Therapies.

Front Oncol 2020 18;10:801. Epub 2020 Jun 18.

Department of Otolaryngology, Head and Neck Surgery, Klinikum Bielefeld, Bielefeld, Germany.

Tumors may consist of billions of cells, which in malignant cases disseminate and form distant metastases. The large number of tumor cells formed by the high number of cell divisions during tumor progression creates a heterogeneous set of genetically diverse tumor cell clones. For cancer therapy this poses unique challenges, as distinct clones have to be targeted in different tissue locations. Recent research has led to the development of specific inhibitors of defined targets in cellular signaling cascades which promise more effective and more tumor-specific therapy approaches. Many of these molecular targeted therapy (MTT) compounds have already been translated into clinics or are currently being tested in clinical studies. However, the outgrowth of tumor cell clones resistant to such inhibitors is a drawback that affects specific inhibitors in a similar way as classical cytotoxic chemotherapeutics, because additionally acquired genetic alterations can enable tumor cells to circumvent the particular regulators of cellular signaling being targeted. Thus, it might be desirable to reduce genetic heterogeneity prior to molecular targeting, which could reduce the statistical chance of tumor relapse initiated by resistant clones. One way to achieve this is employing unspecific methods to remove as much tumor material as possible before MTT, e.g., by tumor debulking (TD). Currently, this is successfully applied in the clinical treatment of ovarian cancer. We believe that TD followed by treatment with a combination of molecular targeted drugs, optimally guided by biomarkers, might advance survival of patients suffering from various cancer types.
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http://dx.doi.org/10.3389/fonc.2020.00801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7314947PMC
June 2020

Stem Cell-Induced Inflammation in Cholesteatoma is Inhibited by the TLR4 Antagonist LPS-RS.

Cells 2020 01 14;9(1). Epub 2020 Jan 14.

Department of Cell Biology, University of Bielefeld, 33619 Bielefeld, Germany.

Cholesteatoma is a severe non-cancerous lesion of the middle ear characterized by massive inflammation, tissue destruction, and an abnormal growth of keratinized squamous epithelium. We recently demonstrated the presence of pathogenic stem cells within cholesteatoma tissue, unfortunately their potential roles in regulating disease-specific chronic inflammation remain poorly understood. In the presented study, we utilized our established human in vitro cholesteatoma stem cell model for treatments with lipopolysaccharides (LPS), tumor necrosis factor α (TNFα), and the TLR4-antagonist LPS from LPS-RS) followed by qPCR, western blot, and immunocytochemistry. Middle ear cholesteatoma stem cells (ME-CSCs) showed a significantly increased expression of TLR4 accompanied by a significantly enhanced LPS-dependent pro-inflammatory gene expression pattern of TNFα, IL-1α, IL-1ß, IL-6, and IL-8 compared to non-pathogenic control cells. LPS-dependent pro-inflammatory gene expression in ME-CSCs was driven by an enhanced activity of NF-B p65 leading to a TNFα-mediated feed-forward-loop of pro-inflammatory NF-B target gene expression. Functional inactivation of TLR4 via the TLR4-antagonist LPS-RS blocked chronic inflammation in ME-CSCs, resulting in a nearly complete loss of IL-1ß, IL-6, and TNFα expression. In summary, we determined that ME-CSCs mediate the inflammatory environment of cholesteatoma via TLR4-mediated NF-B-signaling, suggesting a distinct role of ME-CSCs as drivers of cholesteatoma progression and TLR4 on ME-CSCs as a therapeutic target.
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http://dx.doi.org/10.3390/cells9010199DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7017370PMC
January 2020

The Therapeutic Effect of 1,8-Cineol on Pathogenic Bacteria Species Present in Chronic Rhinosinusitis.

Front Microbiol 2019 22;10:2325. Epub 2019 Oct 22.

Department of Otolaryngology, Head and Neck Surgery, Klinikum Bielefeld, Bielefeld, Germany.

Chronic rhinosinusitis (CRS) is marked by an inflamed mucosa of sinuses and is accompanied by a significantly reduced quality of live. Since no guidelines for the treatment of CRS are available, long lasting clinical histories with health care costs adding up to dozens of billion $ annually are caused by CRS. The progression of CRS is often induced by bacterial infections and/or a shift in microbiome as well as biofilm formation. The exact microbiome alterations are still unclear and the impenetrable biofilm renders the treatment with common antibiotics ineffective. This study focuses on characterizing the microbiome changes in CRS and investigating the inhibition of biofilm growth by 1,8-Cineol, a small, non-polar and hence biofilm penetrating molecule with known antimicrobial potential. We performed MALDI-TOF MS based characterization of the microbiomes of healthy individuals and CRS patients ( = 50). The microbiome in our test group was shifted to pathogens (, , and ). In contrast to published studies, solely based on cell culture techniques, we could not verify the abundance of in CRS. The inhibition of bacterial proliferation and biofilm growth by 1,8-Cineol was measured for these three pathogens. Interestingly, , the most prominent germ in CRS, showed a biofilm inhibition not simply correlated to its inhibition of proliferation. RT-qPCR confirmed that this was due to the downregulations of major key players in biofilm generation (agrA, SarA and σ) by 1,8-Cineol. Furthermore we verified this high biofilm inhibition potential in a model host system consisting out of biofilm grown on mature respiratory epithelium. A second host model, comprising organotypic slices, was utilized to investigate the reaction of the innate immune system present in the nasal mucosa upon biofilm formation and treatment with 1,8-Cineol. Interestingly , the cause of very common catheter infections, possesses a biofilm generation pathway very similar to and might be treatable in a similar fashion. The two presented model systems might be transferred to combinations of every biofilm forming bacterial with most kind of epithelium and mucosa.
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http://dx.doi.org/10.3389/fmicb.2019.02325DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6821979PMC
October 2019

Mechanisms underlying synergy between DNA topoisomerase I-targeted drugs and mTOR kinase inhibitors in NF1-associated malignant peripheral nerve sheath tumors.

Oncogene 2019 09 23;38(39):6585-6598. Epub 2019 Aug 23.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, 02115, USA.

Malignant peripheral nerve sheath tumors (MPNSTs) are soft-tissue sarcomas that frequently arise in patients with neurofibromatosis type 1 (NF1). Most of these tumors are unresectable at diagnosis and minimally responsive to conventional treatment, lending urgency to the identification of new pathway dependencies and drugs with potent antitumor activities. We therefore examined a series of candidate agents for their ability to induce apoptosis in MPNST cells arising in nf1/tp53-deficient zebrafish. In this study, we found that DNA topoisomerase I-targeted drugs and mTOR kinase inhibitors were the most effective single agents in eliminating MPNST cells without prohibitive toxicity. In addition, three members of these classes of drugs, either AZD2014 or INK128 in combination with irinotecan, acted synergistically to induce apoptosis both in vitro and in vivo. In mechanistic studies, irinotecan not only induces apoptosis by eliciting a DNA damage response, but also acts synergistically with AZD2014 to potentiate the hypophosphorylation of 4E-BP1, a downstream target of mTORC1. Profound hypophosphorylation of 4E-BP1 induced by this drug combination causes an arrest of protein synthesis, which potently induces tumor cell apoptosis. Our findings provide a compelling rationale for further in vivo evaluation of the combination of DNA topoisomerase I-targeted drugs and mTOR kinase inhibitors against these aggressive nerve sheath tumors.
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http://dx.doi.org/10.1038/s41388-019-0965-5DOI Listing
September 2019

An Effective Primary Head and Neck Squamous Cell Carcinoma In Vitro Model.

Cells 2019 06 7;8(6). Epub 2019 Jun 7.

Department of Otolaryngology, Head and Neck Surgery, Klinikum Bielefeld, Teutoburger Str. 50, 33604 Bielefeld, Germany.

Head and neck squamous cell carcinoma is a highly malignant disease and research is needed to find new therapeutic approaches. Faithful experimental models are required for this purpose. Here, we describe the specific cell culture conditions enabling the efficient establishment of primary cell culture models. Whereas a classical 10% serum-containing medium resulted in the growth of fibroblast-like cells that outcompeted epithelial cells, we found that the use of specific culture conditions enabled the growth of epithelial tumor cells from HPV+ and HPV- head and neck cancer tissue applicable for research. EpCAM and high Thy-1 positivity on the cell surface were mutually exclusive and distinguished epithelial and fibroblast-like subpopulations in all primary cultures examined and thus can be used to monitor stromal contamination and epithelial cell content. Interestingly, cells of an individual patient developed tumor spheroids in suspension without the use of ultra-low attachment plates, whereas all other samples exclusively formed adherent cell layers. Spheroid cells were highly positive for ALDH1A1 and hence displayed a phenotype reminiscent of tumor stem cells. Altogether, we present a system to establish valuable primary cell culture models from head and neck cancer tissue at high efficiency that might be applicable in other tumor entities as well.
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http://dx.doi.org/10.3390/cells8060555DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6628367PMC
June 2019

Loss of atrx cooperates with p53-deficiency to promote the development of sarcomas and other malignancies.

PLoS Genet 2019 04 10;15(4):e1008039. Epub 2019 Apr 10.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, United States of America.

The SWI/SNF-family chromatin remodeling protein ATRX is a tumor suppressor in sarcomas, gliomas and other malignancies. Its loss of function facilitates the alternative lengthening of telomeres (ALT) pathway in tumor cells, while it also affects Polycomb repressive complex 2 (PRC2) silencing of its target genes. To further define the role of inactivating ATRX mutations in carcinogenesis, we knocked out atrx in our previously reported p53/nf1-deficient zebrafish line that develops malignant peripheral nerve sheath tumors and gliomas. Complete inactivation of atrx using CRISPR/Cas9 was lethal in developing fish and resulted in an alpha-thalassemia-like phenotype including reduced alpha-globin expression. In p53/nf1-deficient zebrafish neither peripheral nerve sheath tumors nor gliomas showed accelerated onset in atrx+/- fish, but these fish developed various tumors that were not observed in their atrx+/+ siblings, including epithelioid sarcoma, angiosarcoma, undifferentiated pleomorphic sarcoma and rare types of carcinoma. These cancer types are included in the AACR Genie database of human tumors associated with mutant ATRX, indicating that our zebrafish model reliably mimics a role for ATRX-loss in the early pathogenesis of these human cancer types. RNA-seq of p53/nf1- and p53/nf1/atrx-deficient tumors revealed that down-regulation of telomerase accompanied ALT-mediated lengthening of the telomeres in atrx-mutant samples. Moreover, inactivating mutations in atrx disturbed PRC2-target gene silencing, indicating a connection between ATRX loss and PRC2 dysfunction in cancer development.
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http://dx.doi.org/10.1371/journal.pgen.1008039DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6476535PMC
April 2019

Systematic Generation of Patient-Derived Tumor Models in Pancreatic Cancer.

Cells 2019 02 10;8(2). Epub 2019 Feb 10.

Department of Translational Medical Oncology, National Center for Tumor Diseases (NCT) Dresden and German Cancer Research Center (DKFZ), 01309 Dresden, Germany.

In highly aggressive malignancies like pancreatic cancer (PC), patient-derived tumor models can serve as disease-relevant models to understand disease-related biology as well as to guide clinical decision-making. In this study, we describe a two-step protocol allowing systematic establishment of patient-derived primary cultures from PC patient tumors. Initial xenotransplantation of surgically resected patient tumors (n = 134) into immunodeficient mice allows for efficient in vivo expansion of vital tumor cells and successful tumor expansion in 38% of patient tumors (51/134). Expansion xenografts closely recapitulate the histoarchitecture of their matching patients' primary tumors. Digestion of xenograft tumors and subsequent in vitro cultivation resulted in the successful generation of semi-adherent PC cultures of pure epithelial cell origin in 43.1% of the cases. The established primary cultures include diverse pathological types of PC: Pancreatic ductal adenocarcinoma (86.3%, 19/22), adenosquamous carcinoma (9.1%, 2/22) and ductal adenocarcinoma with oncocytic IPMN (4.5%, 1/22). We here provide a protocol to establish quality-controlled PC patient-derived primary cell cultures from heterogeneous PC patient tumors. In vitro preclinical models provide the basis for the identification and preclinical assessment of novel therapeutic opportunities targeting pancreatic cancer.
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http://dx.doi.org/10.3390/cells8020142DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6406729PMC
February 2019

Specific Targeting of Oncogenes Using CRISPR Technology.

Cancer Res 2018 10 7;78(19):5506-5512. Epub 2018 Sep 7.

Department of Otolaryngology, Head and Neck Surgery, Klinikum Bielefeld, Bielefeld, Germany.

In recent decades, tools of molecular biology have enabled researchers to genetically modify model organisms, including human cells. RNAi, zinc-finger nucleases, transcription activator-like effector nucleases, CRISPR-Cas9 (clustered regularly-interspaced short palindromic repeats and CRISPR-associated protein 9), retro- or lentiviral gene transfer, and many other methods can be utilized to remove genes, add genes, or change their expression. Within the same timeframe, survival rates for many highly malignant tumor diseases have not improved substantially. If modern medicine could apply even a subset of research methods in clinical management, which are already well established and controllable in basic research laboratories, this could strongly impact patients' prognosis. CRISPR-Cas9 is a method to precisely target and manipulate genomic loci and recent studies have attempted to use this method as a genetic treatment for Duchenne muscular dystrophy, blood disorders, autosomal-dominant hearing loss, and cancer. Some of these approaches target mutant genomic sequences specifically and try to avoid affecting the respective normal loci. Considering obvious genetic risks opposing the objected benefits, data are needed to show whether CRISPR technology is suitable as a future cancer therapy approach or not. Here, we develop strategies for the specific targeting of viral cancer drivers and oncogenes activated by mutation, using the latest CRISPR technology. .
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http://dx.doi.org/10.1158/0008-5472.CAN-18-0571DOI Listing
October 2018

Succession of transiently active tumor-initiating cell clones in human pancreatic cancer xenografts.

EMBO Mol Med 2017 07;9(7):918-932

Department of Translational Oncology, National Center for Tumor Diseases (NCT) and German Cancer Research Center (DKFZ), Heidelberg, Germany

Although tumor-initiating cell (TIC) self-renewal has been postulated to be essential in progression and metastasis formation of human pancreatic adenocarcinoma (PDAC), clonal dynamics of TICs within PDAC tumors are yet unknown. Here, we show that long-term progression of PDAC in serial xenotransplantation is driven by a succession of transiently active TICs producing tumor cells in temporally restricted bursts. Clonal tracking of individual, genetically marked TICs revealed that individual tumors are generated by distinct sets of TICs with very little overlap between subsequent xenograft generations. An unexpected functional and phenotypic plasticity of pancreatic TICs underlies the recruitment of inactive TIC clones in serial xenografts. The observed clonal succession of TIC activity in serial xenotransplantation is in stark contrast to the continuous activity of limited numbers of self-renewing TICs within a fixed cellular hierarchy observed in other epithelial cancers and emphasizes the need to target TIC activation, rather than a fixed TIC population, in PDAC.
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http://dx.doi.org/10.15252/emmm.201607354DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5494525PMC
July 2017

Patient-derived xenografts of gastrointestinal cancers are susceptible to rapid and delayed B-lymphoproliferation.

Int J Cancer 2017 03;140(6):1356-1363

Department of Translational Oncology, National Center for Tumor Diseases (NCT) Heidelberg and German Cancer Research Center (DKFZ), Heidelberg, Germany.

Patient-derived cancer xenografts (PDX) are widely used to identify and evaluate novel therapeutic targets, and to test therapeutic approaches in preclinical mouse avatar trials. Despite their widespread use, potential caveats of PDX models remain considerably underappreciated. Here, we demonstrate that EBV-associated B-lymphoproliferations frequently develop following xenotransplantation of human colorectal and pancreatic carcinomas in highly immunodeficient NOD.Cg-Prkdc Il2rg /SzJ (NSG) mice (18/47 and 4/37 mice, respectively), and in derived cell cultures in vitro. Strikingly, even PDX with carcinoma histology can host scarce EBV-infected B-lymphocytes that can fully overgrow carcinoma cells during serial passaging in vitro and in vivo. As serial xenografting is crucial to expand primary tumor tissue for biobanks and cohorts for preclinical mouse avatar trials, the emerging dominance of B-lymphoproliferations in serial PDX represents a serious confounding factor in these models. Consequently, repeated phenotypic assessments of serial PDX are mandatory at each expansion step to verify "bona fide" carcinoma xenografts.
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http://dx.doi.org/10.1002/ijc.30561DOI Listing
March 2017

Phenotypic differentiation does not affect tumorigenicity of primary human colon cancer initiating cells.

Cancer Lett 2016 Feb 8;371(2):326-33. Epub 2015 Dec 8.

Department of Translational Oncology, National Center for Tumor Diseases (NCT) and German Cancer Research Center (DKFZ), Im Neuenheimer Feld 460, 69120 Heidelberg, Germany; German Consortium for Translational Cancer Research (DKTK), Germany. Electronic address:

Within primary colorectal cancer (CRC) a subfraction of all tumor-initiating cells (TIC) drives long-term progression in serial xenotransplantation. It has been postulated that efficient maintenance of TIC activity in vitro requires serum-free spheroid culture conditions that support a stem-like state of CRC cells. To address whether tumorigenicity is indeed tightly linked to such a stem-like state in spheroids, we transferred TIC-enriched spheroid cultures to serum-containing adherent conditions that should favor their differentiation. Under these conditions, primary CRC cells did no longer grow as spheroids but formed an adherent cell layer, up-regulated colon epithelial differentiation markers, and down-regulated TIC-associated markers. Strikingly, upon xenotransplantation cells cultured under either condition equally efficient formed serially transplantable tumors. Clonal analyses of individual lentivirally marked TIC clones cultured under either culture condition revealed no systematic differences in contributing clone numbers, indicating that phenotypic differentiation does not select for few individual clones adapted to unfavorable culture conditions. Our results reveal that CRC TIC can be propagated under conditions previously thought to induce their elimination. This phenotypic plasticity allows addressing primary human CRC TIC properties in experimental settings based on adherent cell growth.
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http://dx.doi.org/10.1016/j.canlet.2015.11.037DOI Listing
February 2016

Stable long-term blood formation by stem cells in murine steady-state hematopoiesis.

Stem Cells 2012 Sep;30(9):1961-70

Department of Translational Oncology, National Center for Tumor Diseases and German Cancer Research Center, Heidelberg, Germany.

Hematopoietic stem cells (HSCs) generate all mature blood cells during the whole lifespan of an individual. However, the clonal contribution of individual HSC and progenitor cells in steady-state hematopoiesis is poorly understood. To investigate the activity of HSCs under steady-state conditions, murine HSC and progenitor cells were genetically marked in vivo by integrating lentiviral vectors (LVs) encoding green fluorescent protein (GFP). Hematopoietic contribution of individual marked clones was monitored by determination of lentiviral integration sites using highly sensitive linear amplification-mediated-polymerase chain reaction. A remarkably stable small proportion of hematopoietic cells expressed GFP in LV-injected animals for up to 24 months, indicating stable marking of murine steady-state hematopoiesis. Analysis of the lentiviral integration sites revealed that multiple hematopoietic clones with both myeloid and lymphoid differentiation potential contributed to long-term hematopoiesis. In contrast to intrafemoral vector injection, intravenous administration of LV preferentially targeted short-lived progenitor cells. Myelosuppressive treatment of mice prior to LV-injection did not affect the marking efficiency. Our study represents the first continuous analysis of clonal behavior of genetically marked hematopoietic cells in an unmanipulated system, providing evidence that multiple clones are simultaneously active in murine steady-state hematopoiesis.
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http://dx.doi.org/10.1002/stem.1151DOI Listing
September 2012

SOX2-RNAi attenuates S-phase entry and induces RhoA-dependent switch to protease-independent amoeboid migration in human glioma cells.

Mol Cancer 2011 Nov 9;10:137. Epub 2011 Nov 9.

Department of Neurosurgery, Section Experimental Neurosurgery and Tumor Immunology, University Hospital Carl Gustav Carus, TU Dresden, Dresden, Fetscherstr 74, 01307 Dresden, Germany.

Background: SOX2, a high mobility group (HMG)-box containing transcription factor, is a key regulator during development of the nervous system and a persistent marker of neural stem cells. Recent studies suggested a role of SOX2 in tumor progression. In our previous work we detected SOX2 in glioma cells and glioblastoma specimens. Herein, we aim to explore the role of SOX2 for glioma malignancy in particular its role in cell proliferation and migration.

Methods: Retroviral shRNA-vectors were utilized to stably knockdown SOX2 in U343-MG and U373-MG cells. The resulting phenotype was investigated by Western blot, migration/invasion assays, RhoA G-LISA, time lapse video imaging, and orthotopic xenograft experiments.

Results: SOX2 depletion results in pleiotropic effects including attenuated cell proliferation caused by decreased levels of cyclinD1. Also an increased TCF/LEF-signaling and concomitant decrease in Oct4 and Nestin expression was noted. Furthermore, down-regulation of focal adhesion kinase (FAK) signaling and of downstream proteins such as HEF1/NEDD9, matrix metalloproteinases pro-MMP-1 and -2 impaired invasive proteolysis-dependent migration. Yet, cells with knockdown of SOX2 switched to a RhoA-dependent amoeboid-like migration mode which could be blocked by the ROCK inhibitor Y27632 downstream of RhoA-signaling. Orthotopic xenograft experiments revealed a higher tumorigenicity of U343-MG glioma cells transduced with shRNA targeting SOX2 which was characterized by increased dissemination of glioma cells.

Conclusion: Our findings suggest that SOX2 plays a role in the maintenance of a less differentiated glioma cell phenotype. In addition, the results indicate a critical role of SOX2 in adhesion and migration of malignant gliomas.
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http://dx.doi.org/10.1186/1476-4598-10-137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3228695PMC
November 2011