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Keilholz lab

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Keilholz lab

We are a translational research group that focuses on the improvement of diagnostics, early detection of treatment resistance and identification of novel molecular therapies to eventually contribute to advance personalized medicine. As part of the DKTK, we access the research power and clinical expertise of 20 institutions and teaching hospitals at 8 different locations in Germany to facilitate the transfer of basic cancer research to clinical applications. Our three main research topics are liquid biopsy, preclinical cancer models and next generation sequencing-based cancer profiling studies.

     

Our main methods

Cell culture methods

  • Establishment and cultivation of 2D cell culture models
  • Establishment and cultivation of 3D patient- and xenograft-derived organoid models (PDO/PDXO)
  • CRISPR/Cas9 genetic engineering
  • Drug testing
  • Cell viability assays

Immunological methods

  • Flow cytometry
  • Fluorescence microscopy
  • Fluorescence microscope-based single cell isolation (micromanipulator, DEPArray NxT)

Molecular biological methods

  • Polymerase chain reactions (PCR, RT-PCR, ddPCR)
  • Sanger Sequencing, Next generation sequencing
  •  Single-cell sequencing (nanowell-based single-cell BD Rhapsody platform)
  • Western Blot

Liquid biopsy

The collection of a tumor tissue sample to identify targetable aberrations and thus the most promising cancer therapy is the current gold standard for tumor diagnosis and characterization. However, not every tumor is accessible for sample collection, nor will every biopsy result in sufficient material for mutational profiling. Two major limitations are that a single tissue biopsy barely reflects inter- and intratumoral heterogeneity and it is rarely repeated during the course of therapy due to the discomfort of the patient and possible adverse effects. In contrast, liquid biopsy (LB) represents a minimal-invasive approach to analyze blood-derived biomarkers originating from primary and metastatic tumor lesions, also allowing serial sampling to monitor subclonal evolution under treatment pressure and the development of therapy resistance. LB includes the analysis of circulating tumor cells (CTCs), which are shed from the tumor into the bloodstream and might give rise to distant metastases. In comparison, cell-free DNA (cfDNA) and extracellular vesicles are both released from healthy and diseased cells, while they can be identified as cancer-derived, when reflecting the molecular profile of the cancer cell of origin. All three LB components exhibit advantages and disadvantages in terms of sample handling and its ability to provide a more comprehensive reflection of the underlying disease. Consequently, the most recent liquid biopsy projects of our group included

  • ddPCR-based detection of BRAF and KRAS mutations in cfDNA in relation to disease stage in colorectal cancer patients (1)
  • ddPCR-based detection of BRAF and KRAS mutations in extracellular vesicles RNA from colorectal cancer and melanoma patients (2)
  • panel sequencing of CTCs and cfDNA to investigate mutation concordance of matched specimens (colorectal cancer, head and neck squamous cell carcinoma and melanoma) (3)
  • ddPCR-based detection of copy number variations in CTCs and cfDNA from patients with neuroendocrine tumors (still ongoing)

 

1.  Liebs S, Keilholz U, Kehler I, Schweiger C, Haybäck J, Nonnenmacher A. Detection of mutations in circulating cell-free DNA in relation to disease stage in colorectal cancer. Cancer Med. 2019;8(8):3761-9.

2.  Yap SA, Münster-Wandowski A, Nonnenmacher A, Keilholz U, Liebs S. Analysis of cancer-related mutations in extracellular vesicles RNA by Droplet Digital™ PCR. Biotechniques. 2020;69(2):99-107.

3.  Liebs S, Eder T, Klauschen F, Schütte M, Yaspo ML, Keilholz U, Tinhofer I, Kidess-Sigal E, Braunholz D. Applicability of liquid biopsies to represent the mutational profile of tumor tissue from different cancer entities. Oncogene. 2021;40(33):5204-12.

Identification of new biomarkers from patients with neuroendocrine cancer using liquid biopsy methods

Neuroendocrine tumors (NETs) arise from neuroendocrine cells, which have traits of both nerve and endocrine cells including the ability to receive signals from the nervous system and to release hormone-like substances as a response. As these cells are resident in all human tissues, this rare cancer type can develop at multiple sites of the body with the vast majority of tumors occurring in the digestive tract, pancreas, rectum, lungs, and appendix. Due to varying tumor localizations and symptoms, NETs are either misdiagnosed at the first diagnosis or even remain undiagnosed for a long period. Despite the fact that they show a promising tumor response to treatment, even resulting in complete remission of some patients, most patients will eventually suffer from metastases. Thus, method establishment to allow for early diagnosis and disease monitoring are of clinical need.

In cooperation with the Department of Hepatology and Gastroenterology, more than 100 patients with NET are enrolled in our liquid biopsy study, aiming to investigate whether cfDNA and CTCs provide clinically relevant information on tumor development and emerging resistance. Our workflow includes CTC enrichment from whole blood samples and immunofluorescence labeling to enable micromanipulator-assisted single cell isolation followed by whole genome amplification. Additionally, DNA is isolated from plasma, archival FFPE tissue and whole blood serving as a germline control. Besides the quantification of CTCs and cfDNA levels, our pipeline offers a technical sensitivity and specificity to detect copy number variations in all specimens in a selected panel of oncogenes using Droplet Digital PCR and eventually correlate results with clinical parameters of patients.

Collaboration partner: Department of Hepatology and Gastroenterology, Charité

Contact person: M. Sc. Gabriela Pachnikova 

Preclinical cancer models

2D cell culture models build the basis in preclinical research for decades. However, validity and significance of drug screening results generated with these models was limited due to the fact that, in vivo, tumor cells grow three-dimensionally in a complex microenvironment, rendering them less accessible to therapeutic agents when compared to cells growing in a monolayer. The generation and analysis of patient-derived organoid models (PDOs), growing in a three-dimensional, scaffold-based setting, will overcome this limitation by reproducing the structure of the original cancer much better than any 2D model. One of our main goals is the establishment of PDOs derived from various tumor entities prior to treatment and after treatment failure for their subsequent use in preclinical drug testing and identification of treatment resistance mechanisms. Further analysis of sensitive and resistant models will identify key players of disease progression, including tumor heterogeneity and subclonal evolution, on the genetic, transcriptomic and proteomic level employing bulk as well as single cell assays. The importance of this work becomes even more evident in regards to Charité’s commitment to the 3R principles which demand a Reduction, Replacement and Refinement of preclinical tests in animals.

Combinatorial drugging of the MAP kinase pathway in colorectal cancer

Colorectal cancer (CRC) still represents one of the most common causes of cancer death. Despite improvements in understanding its biology, treatment options are still limited. Especially CRC with mutations activating the MAPK pathway, represents the most challenging subgroup, with little benefit from current treatments. Our projects aim is to decipher the mechanisms of resistance to conventional drugs, in particular in MAPK-activated CRC, thus identifying potential biomarkers to exploit new combinatorial approaches. In this respect, we use CRC-PDOs and CRC 2D cell lines, on which both drug testings, including combinational treatments, short term and long term proliferation assays, and gene perturbation experiments (shRNA and CRISPR/Cas9 technology) are performed and subsequently analyzed on a molecular and proteomic level (WES, RNAseq, Western Blot, Mass-spectrometry).

Collaboration partners: Prof. Dr. Christine Sers (Department of Pathology, Charité), Dr. Philipp Mertins (Core Unit Proteomics, BIH), Dr. Manuela Benary (CUBI - Core Unit Bioinformatics, BIH), Prof. Stephan Hahn (Department for Molecular Gastroenterological Oncology, Ruhr University Bochum), Dr. Christian Rosé and Dr. Frank Reichenbach (Pierre Fabre Pharma GmbH), Dr. Osumi Hiroki (Department of Gastroenterology, Cancer Institute Hospital of the Japanese Foundation for Cancer Research, Tokyo-Japan), Prof. Ryoji Yao (Department of Cell Biology, Japanese Foundation for Cancer Research, Tokyo-Japan),  Dr. Thomasz Zemojtel (BIH/MDC Genomics Platform, BIH / MDC)

Contact persons: Dr. Loredana Vecchione, M.Sc. Anna Kotarac  

INTEGRATE-TN: Dynamics of therapy response and resistance in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is the most aggressive subtype among breast tumors, accounting for 10-20% of invasive breast cancers. Due to the absence of estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2/neu), which represent therapy targets in hormone receptor-positive breast cancers, treatment options for TNBC patients are very limited. The Deutsche Krebshilfe-funded project INTEGRATE-TN combines the expertise of multiple academic and industrial partner sites to address the unmet medical need for new therapeutic strategies and to identify biomarkers for the adaptive resistance to chemo- and targeted therapy by performing multi-omics approaches (mutational profiling, proteomics) on archival tissue as well as patient-derived organoid models. The focus of our laboratory is the establishment of a biobank of TNBC organoid cultures to perform pre-clinical drug testing for the evaluation of novel combination therapies and the identification of possible drivers of cancer progression.

Collaboration partners: Department for Gynecology (Charité), DKFZ - German Cancer Research Center (Heidelberg), NCT - National Center for Tumor Diseases (Heidelberg), Philipps-University (Marburg), Martin Luther University (Halle-Wittenberg), Technical University Munich (Munich)

Contact person: M. Sc. Sandra Liebs

The functional consequences of BRAF V600E in triple-negative breast cancer

The RAS–RAF–MEK–ERK signaling cascade, a pathway involved in cell proliferation, survival and differentiation, is one of the most frequently altered pathways in human cancer. BRAF mutations are found in 40-50% of melanoma (MEL) and 8–12% of colorectal cancers (CRC), with highest incidence of missense mutation p.V600E. In breast cancer, BRAF V600E is found in 12% of tumors, particularly in estrogen receptor (ER)-/progesterone receptor (PR)‐negative and triple‐negative breast cancers (TNBC). Whereas a combined treatment with BRAF/MEK inhibitors is beneficial for MEL and CRC patients, demonstrating a 60-70% overall response, single BRAF inhibition resulted in therapeutic failure in multiple breast cancer patients. Furthermore, direct and indirect consequences in downstream pathways of BRAF V600E in breast cancer are still unknown.

By analyzing 3D organoid models and genetically engineered 2D cell cultures, we aim to understand the functional consequences of BRAF V600E in TNBC and identify alternative therapy options. To achieve this, we will perform genomic and transcriptomic analyses using Reverse Transcription Quantitative PCR (RT-qPCR) and RNA sequencing as well as proteomic analysis by Western Blot and protein microarrays. Based on our results, we will perform drug treatments and subsequent cell viability and colony formation assays.

Contact persons: M. Sc. Diogo Abreu, M. Sc.Sandra Liebs

Establishing a series of primary and metastatic organoids for personalized therapies of pancreatic cancer

Non-specific symptoms, rapid disease progress, a high rate of metastasis and very little progress in treatment options result in a five-year survival rate of less than 10% of patients with pancreatic cancer. Previous results of the CONKO-001 and -005 study group showed a significantly prolonged overall survival of patients with pulmonary metastasis after initial surgical resection of the locally limited primary tumor compared to patients with hepatic metastasis (30,4 vs. 18,1 months). This indicates a differential physiology of the tumor in association with the metastatic organotropism, suggesting a possible benefit of some subgroups from personalized treatment options.

The aim of this project is to establish patient-derived tumor organoids of the primary and metastatic sides and expose them to various targeted and well-known chemotherapies, followed by proteomic analysis, to classify subgroups and identify potential biomarkers of the tumor. By correlating the in-vitro data to the clinical response rate of these patients, the organoid model will give insight of whether more personalized therapeutic approaches can become future clinical practice.

Collaboration partners: PD Dr. Uwe Pelzer (Department of Hematology, Oncology and Tumor Immunology, Charité), Prof. Dr. Sebastian Stintzing (Department of Hematology, Oncology and Tumor Immunology, Charité)

Contact persons: Dr. Christopher Neumann, Marie Bossen

Implementation of clinical mass spectrometry in precision medicine

The BIH-funded proof-of-principle project implements clinical mass spectrometry into precision medicine and thus, will improve treatment prediction and patient outcome. A large series of patient-derived xenograft (PDX) and PDX-derived organoid (PDXO) models from head and neck cancer patients are subjected to various experimental drugs and comprehensively profiled using mass spectrometric methods ((phospho)-proteomics, imaging mass spectrometry, tissue microarrays, CyTOF, metabolic screening and nucleotide profiling) complemented with single-cell RNA sequencing. This is a precursor project to the MSTARS project (see below).

Cooperation partners:  EPO – Experimental Pharmacology & Oncology GmbH (Berlin), Dr. Philipp Mertins (Core Unit Proteomics, BIH), Prof. Dr. Matthias Selbach (Proteome Dynamics; MDC for Molecular Medicine at Charité, Berlin), Dr. Stefan Kempa (Proteomics and Metabolomics Platform, MDC), Prof. Dr. Nils Blüthgen (Computational Modelling; Department for Pathology, Charité),     BIH - Core Facilities at Charité: Genomics Platform, Technology Platform Metabolomic, MALDI Imaging Core Unit, Flow & Mass Cytometry Core Facility, Dr. Manuela Benary (CUBI - Core Unit Bioinformatics, BIH)

Contact person: M. Sc. Anastasia Dielmann

MSTARS – Multimodal clinical mass spectrometry to target treatment resistance

Precision medicine aims at offering each patient an individualized treatment based on a deep analysis of the biological characteristics of the patient and the disease. The MSTARS research core in Berlin fosters precision medicine by integrating mass spectrometry in the clinical routine aiming for a standardized and high throughput analysis of patient material to support therapy recommendations. The disease foci of MSTARS are different cancer entities (e.g. head and neck squamous cell carcinoma, leukemia, ovarian cancer, neuroblastoma, prostate cancer) and inflammatory diseases (e.g. myocarditis, chronic liver diseases).

The consortium analyzes biosamples by mass spectrometry or mass spectrometry-coupled imaging based technologies to characterize the proteome and metabolome. Bioinformaticians integrate the acquired data and include already existing omics data, such as genomics and transcriptomics, striving for the identification of biomarkers and signatures as indicators for therapy resistances. The acquired knowledge is directly brought back into the clinics supporting future clinical study design and guiding therapeutic decisions.

For further details, please visit the website.

Contact persons: Ulrich Keilholz, Pia-Katharina Larsen (project coordinator), M. Sc. Anastasia Dielmann, M. Sc. Sandra Liebs

Molecular profiling of cancer using next generation sequencing

Over the last two decades, the sequencing of the human genome has driven many advances in cancer research. Especially the introduction of massive parallel sequencing, also known as Next Generation Sequencing (NGS), allowed researchers to gain unprecedented insights into tumor biology by lowering costs and increasing sample throughput by several orders of magnitude. NGS has since become the gold standard for mutational landscape and gene expression analyses. Further refinements have even led to the application of this technology to single cells, which gives scientists a unique window into tumor heterogeneity and the clonal evolution of cells. NGS is routinely employed in our lab to do molecular subtyping of bulk tumors and single tumor cells by mutation and pathway analyses. It is used as a standalone technique but also as a complementary method when analyzing organoids and liquid biopsy samples.

TRUST SIGN-OC

Debulking surgery and chemotherapy constitute the standard treatment of choice for advanced ovarian cancer patients, but there is uncertainty concerning the optimal sequence of therapy regimens. From 2016-2019, the TRUST trial (Trial of Radical Upfront Surgery in Advanced Ovarian Cancer) addressed the role of neoadjuvant chemotherapy in comparison to primary debulking surgery by enrolling patients for both treatments arms. However, for individualized treatment decisions, knowledge of the molecular signature for responsiveness to chemotherapy and long-term outcome would be desirable. To achieve this, we acquired 600 archival FFPE specimens as well as 100 cryopreserved tumor samples of the TRUST trial to analyze the mutational landscape and gene expression profile by a large in-house NGS panel, and in a subset of patients by whole exome and transcriptome sequencing (WES/WTS). Targeted NGS will focus on homology-directed repair, signaling pathways that are associated with therapy resistance and immune response mechanisms. The resulting comprehensive sequencing data will allow defining a specific molecular signature for advanced ovarian cancer to predict treatment responses to debulking surgery and neoadjuvant chemotherapy, thus enabling individualized treatment decisions and novel clinical trials to improve patient outcome.

Cooperation partner: Dr. Manuela Benary (CUBI - Core Unit Bioinformatics, BIH), PD Dr. Florian Heitz (Clinic for Gynecological Oncology, Evang. Clinicals Essen-Mitte), PD Dr. Fabian Trillsch (Department for Obstetrics and Gynecology, Ludwig-Maximilians-University Clinic - LMU München), Prof. Dr. Elena Ioana Braicu (Department of Gynecology, Charité), AG Hummel (Department for Pathology, Charité)

Contact persons: M. Sc. Christoph Hapke, Dr. Soo Ann Yap, Dipl.-Ing. Annika Nonnenmacher

PeDiOn: Personalized and Digital Oncology

The investigation of tumor heterogeneity has become a pivotal field in cancer research by trying to understand the clonal architecture and evolution of distinct cell subpopulations of a single tumor lesion. PeDiOn is a Berlin Institute of Health (BIH)-funded project, which aims to characterize the spatial and temporal diversity and evolution in head and neck squamous cell carcinoma (HNSCC). Whole genome and whole transcriptome sequencing are performed to produce a comprehensive view, displaying the heterogeneity in multi-regional tumor samples from the same patient.

Cooperation partner: Dr. Alexandra Friedrich (Project Management, BIH), Prof. Ingeborg Tinhofer-Keilholz (Department of Radiooncology and Radiotherapy, Charité), Dr. med. dent.  Claudius Steffen/Dr. med. dent. Max Heiland (Department of dental, oral and maxillofacial surgery, Charité), Prof. David Horst/Edda von der Wall, Hedwig Lammert (Department for Pathology, Charité), Dr. Ute Ungethüm (BIH/MDC Genomics Platform at Charité, BIH/MDC ), Dr. Naveed Ishaque and team (Computational Oncology – BIH)

Contact person: M. Sc. Anastasia Dielmann

HNSCC Atlas – The big picture of HNSCC development and evolution through multi-omics analysis

Various DNA mutations lead to selective growth advantages and promote cancer development. Besides such driver mutations, passenger mutations do not directly influence molecular changes in RNA expression or protein levels, but still seem to be of certain relevance for tumor progression. To fully uncover disease development and evolution, multi-omic approaches are becoming more important to unravel overall cancer characteristics concerning the epigenome, transcriptome, proteome, and metabolome. These multi-omics data offer the opportunity to redefine personalized approaches, by moving from simplistic, genomic-focused considerations towards a more comprehensive view of the tumor including its microenvironment.

In this project, multi-regional tumor samples from primary and recurrent head and neck squamous cell carcinoma (HNSCC) patients are analyzed by next-generation sequencing to identify copy number variations (CNVs), mutations, and translocations. Those genetic variations are then completed with transcriptome, proteome and methylome data to understand the cross-linkage between molecular alterations and their effects on gene expression, protein function and eventually on disease development. Gathered findings are compared to immunohistopathological methods of tumor and microenvironment analysis including tissue microarray analysis (TMA). The entire data set will be combined in an HNSCC atlas to improve our understanding of predictive molecular changes in the tumor and its microenvironment and thereby reform personalized treatment decisions. 

Collaboration partners: Dr. med. dent.  Claudius Steffen/Dr. med. dent. Max Heiland (Charité – dental, oral and maxillofacial surgery), Genomic Core Facility (BIH – Berlin Institute of Health), Dr. Naveed Ishaque (Computational Oncology, BIH – Berlin Institute of Health), Prof. Ingeborg Tinhofer-Keilholz (Department of Radiooncology and Radiotherapy – Charité), Dr. Philipp Mertins (Proteomics), David Capper (Methylome analysis), Dr. Simon Schallenberg (Institute of Pathology – Charité), Dr. Manuela Benary (BIH – Core Unit Bioinformatics)

Contact person: M. Sc. Anastasia Dielmann

DKTK Clonal Evolution: Comparison of clonal cancer evolution in patients vs. patient-derived xenograft and organoid models

The OncoTrack program demonstrated a high level of similarity in the clonal selection from the primary tumor to the formed liver metastases and the primary tumor to the derived PDX model. In order to verify the hypothesis that the most aggressive subclone is not only forming the metastasis in the patient, but is also selected in the PDX model, this DKTK-funded project investigates the representation of the metastatic disease in patients with colorectal cancer (CRC), head and neck squamous cell carcinoma (HNSCC) and melanoma (MEL) by PDX and PDO models. Next generation sequencing (NGS) and methylome analysis will be performed on matched clinical samples and preclinical models to investigate patterns of clonal selection. Three NGS panels were designed to cover frequently mutated and therapy resistance-associated cancer genes in CRC, HNSCC and MEL, furthermore including genes for the analysis of tumor mutational burden (TMB) and microsatellite instability (MSI).

Collaboration partners: Prof. Dr. David Capper (Methylome analysis; Department for Neuropathology, Charite), Dr. Manuela Benary (CUBI - Core Unit Bioinformatics, BIH), Dr. Viktor Arnhold (Department of Pathology, Charité), Prof. Franziska Ghoreschi und Prof. Eigentler (Department of Dermatology, Venereology and Allergology, Charité)

Contact persons: M. Sc.Sandra Liebs, M. Sc. Anastasia DielmannDipl.-Ing. Annika Nonnenmacher

BIH Clinical Single Cell Sequencing (CSCS) Pipeline

The analysis of single tumor cells becomes increasingly important in precision oncology. It allows comprehensive analysis of individual cells of a heterogeneous tissue to provide a higher resolution on cell-to-cell variability that might lead to therapy resistance. The BIH Bioportal Single Cells is part of the BIH Clinical Single Cell Sequencing Pipeline, which provides a service workflow including tissue storage, sample processing and next generation sequencing to the entire BIH Faculty. It provides support to scientists and clinicians in planning single cell approaches from tissue collection to sequencing, data analysis, interpretation and storage. The general task of our laboratory includes tissue dissociation and preparation of bead-bound single-cell RNA (scRNA) libraries, including the requisite quality controls.

For further details, please visit the website of the BIH Bioportal Single Cells

Cooperation partner: PD Dr. Markus Morkel (Central Bioportal Single Cell, BIH - Berlin Institute of Health at Charité / Department for Pathology, Charité)

Contact persons: Ulrich Keilholz, M. Sc. Berit Zirkelbach, Lisa Schneider (currently on maternity leave)


Staff of Keilholz lab

Lab Manager

Prof. Dr. med. Ulrich Keilholz

Laborleiter AG Keilholz / Direktor des Charité Comprehensive Cancer Center, Charité

Scientific staff

M. Sc. Sandra Liebs

Doktorandin | Laborleiterin | Labor AG Keilholz

M. Sc. Christoph Hapke

Doktorand | Sicherheitsbeauftragter | Labor AG Keilholz

M. Sc. Diogo de Castro Abreu

Doktorand | Labor AG Keilholz

Dipl.-Ing. Anika Nonnenmacher

Wiss. Mitarbeiterin | Gefahrstoffbeauftragte | Labor AG Keilholz

M. Sc. Anastasia Dielmann

Doktorandin | Labor AG Keilholz

Marie Bossen

Medizinische Doktorandin | Labor AG Keilholz

M. Sc. Gabriela Pachnikova

Doktorandin | Labor AG Keilholz

M. Sc. Anna Kotarac

Wissenschaftliche Mitarbeiterin | Labor AG Keilholz

Sina Löwe

Medizinische Doktorandin | Labor Ag Keilholz

Doctors

Dr. med. Christopher Neumann

Ärztlicher und wissenschaftlicher Mitarbeiter | PostDoc | Labor AG Keilholz

MD, Ph.D, Loredana Vecchione

Ärztliche und wissenschaftliche Mitarbeiterin | PostDoc | Labor AG Keilholz

Dr. med. Soo Ann Yap

Ärztliche und wissenschaftliche Mitarbeiterin | Labor AG Keilholz

Technical staff

Lisa Schneider

Mutterschutz | Technische Angestellte | Labor AG Keilholz

M. Sc. Berit Zirkelbach

Technische Angestellte | Labor AG Keilholz


Selected publications

2022

  • Xu Y., Pachnikova G., Przybilla D., Schäfer R., Cui Y., Zhou D., Chen Z., Zhao A. and Keilholz U. (2022): Evaluation of JQ1 Combined with Docetaxel for the Treatment of Prostate Cancer Cells in 2D- and 3D- Culture Systems. Front. Pharmacol. 13:839620. doi: 10.3389/fphar.2022.839620

2021

  • Regan JL, Schumacher D, Staudte S, Steffen A, Lesche R, Toedling J, Jourdan T, Haybaeck J, Mumberg D, Henderson D, Győrffy B, Regenbrecht CRA, Keilholz U, Schäfer R, Lange M. RNA sequencing of long-term label-retaining colon cancer stem cells identifies novel regulators of quiescence. iScience. 2021 May 24;24(6):102618. doi: 10.1016/j.isci.2021.102618. PMID: 34142064
  • Liebs S, Eder T, Klauschen F, Schütte M, Yaspo ML, Keilholz U, Tinhofer I, Kidess-Sigal E, Braunholz D. Applicability of liquid biopsies to represent the mutational profile of tumor tissue from different cancer entities. Oncogene. 2021 Jul; doi: 10.1038/s41388-021-01928-w, PMID: 34230613
  • Osumi H, Vecchione L, Keilholz U, Vollbrecht C, Alig AHS, von Einem JC, Stahler A, Striefler JK, Kurreck A, Kind A, Modest DP, Stintzing S, Jelas I. NeoRAS wild-type in metastatic colorectal cancer: Myth or truth? - Case series and review of the literature. Eur J Cancer 2021 Aug; 153:86-95, PMID: 34153718
  • Dahlmann M, Gambara G, Brzezicha B, Popp O, Pachmayr E, Wedeken L, Pflaume A, Mokritzkij M, Gül-Klein S, Brandl A, Schweiger-Eisbacher C, Mertins P, Hoffmann J, Keilholz U, Walther W, Regenbrecht C, Rau B, Stein U. Peritoneal metastasis of colorectal cancer (pmCRC): identification of predictive molecular signatures by a novel preclinical platform of matching pmCRC PDX/PD3D models. Mol Cancer. 2021 Oct 21;20(1):129. doi: 10.1186/s12943-021-01430-7. PMID: 34670579
  • Keil M, Conrad T, Becker M, Keilholz U, Yaspo ML, Lehrach H, Schütte M, Haybaeck J, Hoffmann J. Modeling of Personalized Treatments in Colon Cancer Based on Preclinical Genomic and Drug Sensitivity Data. Cancers (Basel). 2021 Nov 30;13(23):6018. doi: 10.3390/cancers13236018. PMID: 34885128

2020

  • Yap SA, Münster-Wandowski A, Nonnenmacher A, Keilholz U, Liebs S. Analysis of cancer-related mutations in extracellular vesicles RNA by Droplet DigitalTM PCR. BioTechniques 2020; 69:2, 99-107, PMID: 32580578
  • Hess AK, Jöhrens K, Zakarneh A, Balermpas P, Von Der Grün J, Rödel C, Weichert W, Hummel M, Keilholz U, Budach V, Tinhofer I. Characterization of the tumor immune micromilieu and its interference with outcome after concurrent chemoradiation in patients with oropharyngeal carcinomas. Oncoimmunology. 2019 May 25;8(8):1614858. doi: 10.1080/2162402X.2019.1614858. PMID: 31413922
  • Sinn M, Sinn BV, Treue D, Keilholz U, Damm F, Schmuck R, Lohneis P, Klauschen F, Striefler JK, Bahra M, Bläker H, Bischoff S, Pelzer U, Oettle H, Riess H, Budczies J, Denkert C. <i>TP53</i> Mutations Predict Sensitivity to Adjuvant Gemcitabine in Patients with Pancreatic Ductal Adenocarcinoma: Next- Generation Sequencing Results from the CONKO-001 Trial. Clin Cancer Res. 2020 Jul 15;26(14):3732-3739. doi: 10.1158/1078-0432.CCR-19-3034. Epub 2020 Mar 31. PMID: 32234756
  • Joseph JF, Gronbach L, García-Miller J, Cruz LM, Wuest B, Keilholz U, Zoschke C, Parr MK. Automated Real-Time Tumor Pharmacokinetic Profiling in 3D Models: A Novel Approach for Personalized Medicine. Pharmaceutics. 2020 Apr 30;12(5):413. doi: 10.3390/pharmaceutics12050413. PMID: 32366029
  • Schlicker A, Ellappalayam A, Beumer IJ, Snel MHJ, Mittempergher L, Diosdado B, Dreezen C, Tian S, Salazar R, Loupakis F, Pietrantonio F, Santos Vivas C, Martinez-Villacampa MM, Villanueva A, Sanjuán X, Schirripa M, Fassan M, Martinetti A, Fucà G, Lonardi S, Keilholz U, Glas AM, Bernards R, Vecchione L. Investigating the concordance in molecular subtypes of primary colorectal tumors and their matched synchronous liver metastasis. Int J Cancer. 2020 Oct 15;147(8):2303-2315. doi: 10.1002/ijc.33003. Epub 2020 Apr 24. PMID: 32270478
  • Peitz C, Sprüssel A, Linke RB, Astrahantseff K, Grimaldi M, Schmelz K, Toedling J, Schulte JH, Fischer M, Messerschmidt C, Beule D, Keilholz U, Eggert A, Deubzer HE, Lodrini M. Multiplexed Quantification of Four Neuroblastoma DNA Targets in a Single Droplet Digital PCR Reaction. J Mol Diagn. 2020 Aug 26:S1525-1578(20)30446-3. doi: 10.1016/j.jmoldx.2020.07.006. Epub ahead of print. PMID: 32858250
  • Scherr AL, Mock A, Gdynia G, Schmitt N, Heilig CE, Korell F, Rhadakrishnan P, Hoffmeister P, Metzeler KH, Schulze-Osthoff K, Illert AL, Boerries M, Trojan J, Waidmann O, Falkenhorst J, Siveke J, Jost PJ, Bitzer M, Malek NP, Vecchione L, Jelas I, Brors B, Glimm H, Stenzinger A, Grekova SP, Gehrig T, Schulze-Bergkamen H, Jäger D, Schirmacher P, Heikenwalder M, Goeppert B, Schneider M, Fröhling S, Köhler BC. Identification of BCL-XL as highly active survival factor and promising therapeutic target in colorectal cancer. Cell Death Dis. 2020;11(10):875, PMID: 33070156
  • Vecchione L, Stintzing S, Pentheroudakis G, Douillard JY, Lordick F. ESMO management and treatment adapted recommendations in the COVID-19 era: colorectal cancer. ESMO Open. 2020 May;5 (Suppl 3):e000826, PMID: 32457036
  • Klinghammer K, Politz O, Eder T, Otto R, Raguse JD, Albers A, Kaufmann A, Tinhofer I, Hoffmann J, Keller U, Keilholz U. Combination of copanlisib with cetuximab improves tumor response in cetuximab-resistant patient-derived xenografts of head and neck cancer. Oncotarget. 2020 Oct 13;11(41):3688-3697. doi: 10.18632/oncotarget.27763. PMID: 33110476

2019

  • Schumacher D, Andrieux G, Boehnke K, Keil M, Silvestri A, Silvestrov M, Keilholz U, Haybaeck J, Erdmann G, Sachse C, Templin M, Hoffmann J, Boerries M, Schäfer R, Regenbrecht CRA. Heterogeneous pathway activation and drug response modelled in colorectal-tumor-derived 3D cultures. PLoS Genet. 2019 Mar 29;15(3):e1008076. doi: 10.1371/journal.pgen.1008076. Erratum in: PLoS Genet. 2019 May 29;15(5):e1008183. PMID: 30925167
  • Liebs S, Keilholz U, Kehler I, Schweiger C, Haybäck J, Nonnenmacher A. Detection of mutations in circulating cell-free DNA in relation to disease stage in colorectal cancer. Cancer Medicine 2019; 8(8):3761-3769, PMID: 31134762
  • Liebs S, Nonnenmacher A, Klauschen F, Keilholz U, Vecchione L. Liquid biopsy assessment of synchronous malignancies: a case report and review of the literature. ESMO Open 2019; 4:e000528, PMID: 31555482
  • Mulero-Sanchez A, Pogacar Z, Vecchione L.  Importance of genetic screens in precision oncology. ESMO Open 2019, 4, PMID: 31231569

2008 to 2018

2018

  • Gambara G, Gaebler M, Keilholz U, Regenbrecht CRA, Silvestri A. From Chemotherapy to Combined Targeted Therapeutics: In Vitro and in Vivo Models to Decipher Intra-tumor Heterogeneity. Front Pharmacol. 2018 Feb 14;9:77. doi: 10.3389/fphar.2018.00077. PMID: 29491834
  • Meucci S, Keilholz U, Heim D, Klauschen F, Cacciatore S. Somatic genome alterations in relation to age in lung squamous cell carcinoma. Oncotarget. 2018 Aug 14;9(63):32161-32172. doi: 10.18632/oncotarget.25848. PMID: 30181806


2017

  • Schütte M, Risch T, Abdavi-Azar N, Boehnke K, Schumacher D, Keil M, Yildiriman R, Jandrasits C, Borodina T, Amstislavskiy V, Worth CL, Schweiger C, Liebs S, Lange M, Warnatz HJ, Butcher LM, Barrett JE, Sultan M, Wierling C, Golob-Schwarzl N, Lax S, Uranitsch S, Becker M, Welte Y, Regan JL, Silvestrov M, Kehler I, Fusi A, Kessler T, Herwig R, Landegren U, Wienke D, Nilsson M, Velasco JA, Garin-Chesa P, Reinhard C, Beck S, Schäfer R, Regenbrecht CRA, Henderson D, Lange B, Haybaeck J, Keilholz U, Hoffmann J, Lehrach H, Yaspo ML. Molecular dissection of colorectal cancer in pre-clinical models identifies biomarkers predicting sensitivity to EGFR inhibitors. Nature Communications 2017. 8:14262, PMID: 28186126
  • Procaccio L, Schirripa M, Fassan M, Vecchione L, Bergamo F, Prete AA, Intini R, Manai C, Dadduzio V, Boscolo A, Zagonel V, Lonardi S. Immunotherapy in Gastrointestinal Cancers  BioMed Research International, vol. 2017, Article ID 4346576, PMID: 28758114
  • Liu Y, Cheng G, Qian J, Ju H, Zhu Y, Stefano M, Keilholz U, Li D. Expression of guanylyl cyclase C in tissue samples and the circulation of rectal cancer patients. Oncotarget. 2017 Jun 13;8(24):38841-38849. doi: 10.18632/oncotarget.16406. PMID: 28418917
  • Klinghammer K, Otto R, Raguse JD, Albers AE, Tinhofer I, Fichtner I, Leser U, Keilholz U, Hoffmann J. Basal subtype is predictive for response to cetuximab treatment in patient-derived xenografts of squamous cell head and neck cancer. Int J Cancer. 2017 Sep 15;141(6):1215-1221. doi: 10.1002/ijc.30808. Epub 2017 Jun 21. PMID: 28560858
  • Liu Y, Meucci S, Sheng L, Keilholz U. Meta-analysis of the mutational status of circulation tumor cells and paired primary tumor tissues from colorectal cancer patients. Oncotarget. 2017 May 26;8(44):77928-77941. doi: 10.18632/oncotarget.18272. PMID: 29100436
  • Lodrini M, Sprüssel A, Astrahantseff K, Tiburtius D, Konschak R, Lode HN, Fischer M, Keilholz U, Eggert A, Deubzer HE. Using droplet digital PCR to analyze MYCN and ALKcopy number in plasma from patients with neuroblastoma. Oncotarget. 2017 Jul 7;8(49):85234-85251. doi: 10.18632/oncotarget.19076. PMID: 29156716
  • Regan JL, Schumacher D, Staudte S, Steffen A, Haybaeck J, Keilholz U, Schweiger C, Golob-Schwarzl N, Mumberg D, Henderson D, Lehrach H, Regenbrecht CRA, Schäfer R, Lange M. Non-Canonical Hedgehog Signaling Is a Positive Regulator of the WNT Pathway and Is Required for the Survival of Colon Cancer Stem Cells. Cell Rep. 2017 Dec 5;21(10):2813-2828. doi: 10.1016/j.celrep.2017.11.025. PMID: 29212028
  • Thuss-Patience P, Vecchione L, Keilholz U. Should cT2 esophageal cancer get neoadjuvant treatment before surgery? J Thorac Dis. 2017 Sep;9(9):2819-2823. doi: 10.21037/jtd.2017.08.143. PMID: 29221247


2016

  • Boehnke K, Iversen PW, Schumacher D, Lallena MJ, Haro R, Amat J, Haybaeck J, Liebs S, Lange M, Schäfer R, Regenbrecht CRA, Reinhard C, Velasco JA. Assay establishment and validation of a high-throughput screening platform for three-dimensional patient-derived colon cancer organoid cultures. Journal of Biomolecular Screening 2016; 21(9):931-41, PMID: 27233291
  • Dielmann A, Letsch A, Nonnenmacher A, Miller K, Keilholz U, Busse A. Favorable prognostic influence of T-box transcription factor Eomesodermin in metastatic renal cell cancer patients. Cancer Immunol Immunother. 2016 Feb;65(2):181-92, PMID: 26753694


2015

  • Tinhofer I, Niehr F, Konschak R, Liebs S, Munz M, Stenzinger A, Weichert W, Keilholz U, Budach V. Next-generation sequencing: hype and hope for development or personalized radiation therapy? Radiation Oncology 2015; 10:183, PMID: 26316159
  • Riechardt AI, Maier AK, Nonnenmacher A, Reichhart N, Keilholz U, Kociok N, Strauss O, Joussen AM, Gundlach E. B-Raf inhibition in conjunctival melanoma cell lines with PLX 4720. Br J Ophthalmol. 2015 Dec;99(12):1739-45, PMID: 26347528
  • Pfisterer K, Fusi A, Klinghammer K, Knödler M, Nonnenmacher A, Keilholz U. PI3K/PTEN/AKT/mTOR polymorphisms: association with clinical outcome in patients with head and neck squamous cell carcinoma receiving cetuximab-docetaxel. Head Neck. 2015 Apr;37(4):471-8, PMID: 24421178


2014

  • Gross A, Niemetz-Rahn A, Nonnenmacher A, Tucholski J, Keilholz U, Fusi A. Expression and activity of EGFR in human cutaneous melanoma cell lines and influence of vemurafenib on the EGFR pathway. Target Oncol. 2015 Mar;10(1):77-84, PMID: 24824730
     

2013

  • Busse A, Letsch A, Fusi A, Nonnenmacher A, Stather D, Ochsenreither S, Regenbrecht CR, Keilholz U. Characterization of small spheres derived from various solid tumor cell lines: are they suitable targets for T cells? Clin Exp Metastasis. 2013 Aug;30(6):781-91, PMID: 23519726
  • Busse A, Rapion J, Fusi A, Suciu S, Nonnenmacher A, Santinami M, Kruit WH, Testori A, Punt CJ, Dalgleish AG, Spatz A, Eggermont AM, Keilholz U. Analysis of surrogate gene expression markers in peripheral blood of melanoma patients to predict treatment outcome of adjuvant pegylated interferon alpha 2b (EORTC 18991 side study). Cancer Immunol Immunother. 2013 Jul;62(7):1223-33, PMID: 23624802
     

2012

  • --

2011

  • Fusi A, Berdel R, Havemann S, Nonnenmacher A, Keilholz U. Enhanced detection of BRAF-mutants by pre-PCR cleavage of wild-type sequences revealed circulating melanoma cells heterogeneity. Eur J Cancer. 2011 Sep;47(13):1971-6, PMID: 21570823
  • Liu Z, Fusi A, Klopocki E, Schmittel A, Tinhofer I, Nonnenmacher A, Keilholz U. Negative enrichment by immunomagnetic nanobeads for unbiased characterization of circulating tumor cells from peripheral blood of cancer patients. J Transl Med. 2011 May 19;9:70, PMID: 21595914
  • Busse A, Asemissen AM, Nonnenmacher A, Braun F, Ochsenreither S, Stather D, Fusi A, Schmittel A, Miller K, Thiel E, Keilholz U. Immunomodulatory effects of sorafenib on peripheral immune effector cells in metastatic renal cell carcinoma. Eur J Cancer. 2011 Mar;47(5):690-6, PMID: 21215610
  • Busse A, Asemissen A, Nonnenmacher A, Ochsenreither S, Fusi A, Braun F, Stather D, Schmittel A, Miller K, Thiel E, Keilholz U. Systemic immune tuning in renal cell carcinoma: favorable prognostic impact of TGF-β1 mRNA expression in peripheral blood mononuclear cells. J Immunother. 2011 Jan;34(1):113-9, PMID: 21150720
  • Fusi A, Reichelt U, Busse A, Ochsenreither S, Rietz A, Maisel M, Keilholz U. Expression of the stem cell markers nestin and CD133 on circulating melanoma cells. J Invest Dermatol. 2011 Feb;131(2):487-94, PMID: 20882037
     

2010

  • Busse A, Letsch A, Scheibenbogen C, Nonnenmacher A, Ochsenreither S, Thiel E, Keilholz U. Mutation or loss of Wilms' tumor gene 1 (WT1) are not major reasons for immune escape in patients with AML receiving WT1 peptide vaccination. J Transl Med. 2010 Jan 21;8:5, PMID: 20092642
  • Fusi A, Ochsenreither S, Busse A, Rietz A, Keilholz U. Expression of the stem cell marker nestin in peripheral blood of patients with melanoma. Br J Dermatol. 2010 Jul;163(1):107-14, PMID: 20346020
     

2009

  • Fusi A, Collette S, Busse A, Suciu S, Rietz A, Santinami M, Kruit WH, Testori A, Punt CJ, Dalgleish AG, Spatz A, Eggermont AM, Keilholz U. Circulating melanoma cells and distant metastasis-free survival in stage III melanoma patients with or without adjuvant interferon treatment (EORTC 18991 side study). Eur J Cancer. 2009 Dec;45(18):3189-97, PMID: 19793643
  • Busse A, Rietz A, Schwartz S, Thiel E, Keilholz U. An intron 9 containing splice variant of PAX2. J Transl Med. 2009 May 25;7:36, PMID: 19467152
  • Busse A, Gökbuget N, Siehl JM, Hoelzer D, Schwartz S, Rietz A, Thiel E, Keilholz U. Wilms' tumor gene 1 (WT1) expression in subtypes of acute lymphoblastic leukemia (ALL) of adults and impact on clinical outcome. Ann Hematol. 2009 Dec;88(12):1199-205, PMID: 19404640
     

2008

  • Busse A, Kraus M, Na IK, Rietz A, Scheibenbogen C, Driessen C, Blau IW, Thiel E, Keilholz U. Sensitivity of tumor cells to proteasome inhibitors is associated with expression levels and composition of proteasome subunits. Cancer. 2008 Feb 1;112(3):659-70, PMID: 18181098