R & D
KINETO Lab develops unique methods to be used along the preclinical drug discovery pipeline to identify and validate new drug targets and to predict in vivo toxicity of anticancer drugs.
The company’s experience in various research grant projects, such as Horizon 2020, the ever biggest research and innovation program of the European Community gives solid scientific basis for high quality research and development.
HORIZON 2020 – EUROPEAN TRAINING NETWORK: PEPTIDE-DRUG CONJUGATES FOR TARGETED DELIVERY IN TUMOR THERAPY (MAGICBULLET)
KINETO Lab is member of HORIZON 2020 Marie Skłodowska-Curie Actions MAGICBULLET” international research consortium built for studying the application of drug conjugates in selective tumor therapy.
Many tumor cells are characterized by the overexpression of certain antigens. Molecules that specifically recognize these structures are suitable as homing devices in tumor therapy. Conjugation of anticancer drugs with such a delivery vector targeting tumors would be a “magic bullet” according to the Nobel laureate Paul Ehrlich. Antibody-drug conjugates (ADC) technology have already been approved for anticancer therapy. However, ADCs have limitations with respect to tumor penetration, high manufacturing costs, and require challenging conjugation chemistry. Peptide-drug conjugates have a high drug loading capacity, easily penetrate tissue, and can be cost efficiently prepared in a homogenous form with straightforward and well-defined conjugation chemistry.
The focus of the European Training Network (ETN) MAGICBULLET is on chemistry-driven approaches toward conjugates between peptides as delivery vectors that recognize tumors and anticancer drugs in order to selectively fight cancer, a topic with a high demand of research activities. The ETN allows to develop and validate an array of new peptide-drug conjugates combining either known tumor-specific peptides or newly discovered tumor-homing peptides with potent cytotoxic drugs. The tumor-selective peptides are designed for cellular uptake mediated either by endocytosis or by cell-penetrating peptides.
The combination of an array of tumor-selective peptides targeting different receptors and different uptake mechanisms with diverse antitumor drugs acting on different cellular targets is a powerful strategy to minimize potential risks on healthy cells and increase the efficacy toward tumors. Because the number of receptors on tumor cells is limited, the combination of different target peptide–drug conjugates may enhance the bioactivity. The influence of the treatment schedule of such combination therapy on the antitumor activity will also be evaluated.
The consortium of the ETN MAGICBULLET brings together interdisciplinary expert knowledge in tumor biology, drug discovery, biochemistry, pharmacology, cell biology, organic chemistry, peptide chemistry, synthetic chemistry, medicinal chemistry, spectroscopy, conformational analysis, and computational chemistry. The training program focuses on multidisciplinary research to explore and validate molecular targets for innovative treatment or investigations on the molecular mechanisms in organ-specific metastatic growth processes. It aims at scientific multilingualism and relies on concerted learning, a combination of introductory training, hands-on learning “on the bench”, teaching by peers, and training in additional skills. This high complementarity is required for the different scientific tasks in the development pipeline.
The ETN MAGICBULLET is a tool that can:
- identify, modify, and validate tumor-selective peptides for known and new cell surface receptor targets (e.g. integrins, Gonadotropin-releasing hormone receptors, CD13, VEGFR, cadherins);
- study different linker systems for release of the anticancer payloads at the appropriate site;
- conjugate known and new anticancer agents or drugs to tumor-selective peptides;
- investigate the biological activity in vitro and in vivo to demonstrate their efficacy.
MAGICBULLET under the Marie Skłodowska-Curie grant agreement No 642004. www.magicbullet.de
HORIZON 2020 – EUROPEAN TRAINING NETWORK: SMALL MOLECULE DRUG CONJUGATES FOR TARGETED DELIVERY IN TUMOR THERAPY (MAGICBULLET::RELOADED)
KINETO Lab continued its membership in the ”MAGICBULLET::RELOADED” international research consortium continued from „MAGICBULLET” built for studying the application of drug conjugates in targeted tumor therapy.
Despite the continuing development of new and more efficient treatments, cancer remains the second cause of premature death worldwide. Multi-faceted interdisciplinary research efforts in industry and academia on different aspects of cancer have provided a knowledge basis for the development of novel therapeutic approaches. An ideal therapeutic agent would be a “magic bullet” that only kills the target cells. This ETN initiative with the title Magicbullet::Reloaded refers to Ehrlich’s bold idea and builds on the previous experience of the ETN MAGICBULLET (2015-2018, grant agreement No. 642004).
The ETN Magicbullet::Reloaded will expand the field of investigation from peptide-drug conjugates (PDCs) to small molecule-drug conjugates (SMDCs) with a special focus on drugs capable to stimulate tumor immune responses and overcome resistance to immune-therapy. The consortium has been substantially expanded to perfectly address the needs of the new research direction. The planned ETN will design and synthesize an array of SMDCs (including PDCs), also targeting less investigated tumor antigens, investigate their pharmacokinetic behavior, their implication on the immune system, as well as their tumor selectivity and antitumor activity.
MAGICBULLET::RELOADED under the Marie Skłodowska-Curie grant agreement No 861316. www.magicbullet-reloaded.eu
DEVELOPMENT OF DIAGNOSIS AND TARGETED THERAPY OF RASOPATHIES
KRAS is one of the most frequently mutated oncogene in human cancer with an outstanding incidence rate among the most prevalent cancers such as colon and rectum cancer (40%), lung adenocarcinomas (30%) and pancreatic cancer (80%). Moreover, with smaller frequency, other tumor types (myeloma multiplex, head and neck- and breast cancer) display KRAS mutations as well. KRAS mutant human cancers are more aggressive and less sensitive to classical chemotherapeutic agents.
In clinical oncology the new paradigm is the introduction of the molecular targeted therapies exploiting the characteristic genetic alterations of the tumors which is frequently mutation of oncogenes. However, the efficacy of targeted therapies (i.e. anti-EGFR agents) is critically determined by the KRAS mutational status of the tumor, as mutated KRAS leads to resistance to these expensive therapies. Therefore, the mutant oncogenic KRAS in human cancer emerged as arguably the most outstanding issue in clinical oncology that should guide the future of R&D activities.
DEVELOPMENT OF BIOMEDICAL RESEARCH AND SERVICE CAPACITY AND EXPLORE NEW TYPES OF ANTICANCER THERAPEUTIC SOLUTIONS
In this project, KINETO Lab intends to expand its existing tissue culture and animal-based infrastructure so that it can continually be able to isolate larger amount of antibodies several times a year, from the hybridomas created by its own earlier development or from its cells from its academic and industrial partners. The antibodies so obtained are then marketed in various technical solutions (laboratory diagnostic kits, molecular biology reagents, immunohistochemical techniques, etc.).
Testing of new active substances, including therapeutic antibodies, requires the use of better preclinical models. These include molecular biology methods, cell and tissue culture techniques, and animal experiments. This is also the case in oncology. Patient-derived tumor xenografts (PDTX), when implanting a direct surgical sample into specific mice, represent a much better representation of the original tumor because of not only the tumor cells but also their environment are transplanted. The second major objective of the project is to develop such specific models from the least manageable tumor types, subsequent basic and R&D.
In addition, thirdly, with the help of the models, KINETO Lab develops new types of antibodies that are specifically active in tumor vessels, for diagnostic and therapeutic use. Identification of endothelial markers is a novel therapeutic target that can reduce the side effects of anticancer treatments and increase their effectiveness on tumors.
DEVELOPMENT OF TARGETED THERAPIES AGAINST HUMAN TUMORS HARBORING MUTATIONS OF KRAS ONCOGENE
Testing of novel tyrosine kinase inhibitors alone and in combination with farnesyltransferase inhibitors on tumor cells carrying different KRAS mutations (G12C, D, V) on in vitro and in vivo tumor models.
DEVELOPMENT OF DETECTION METHOD FOR CIRCULATING ENDOTHELIAL PROGENITOR CELLS
Growth of new blood vessels, through angiogenesis and vasculogenesis, is required and essential step in the tumor growth as well as in regeneration of hypoxic post-infarction tissues. This process can be achieved through the incorporation of endothelial progenitor cells (EPCs) into the endothelial tube of novel blood capillaries.
Based on several observations, the circulating levels of these cells are associated with the severity of the given disease. In line with that, enumeration of EPCs can help to assess disease progression, prognosis and therapeutic efficacy.
This angiogenesis related project has been successfully completed laying down the principles of developing a detection method for enumeration circulating EPCs.
ID: INNO_08_KM-KLABIN08 (INNOCSEKK_PLUSZ) (NKFIH – ETH Zürich)
DEVICE THAT IS ABLE TO CAPTURE A SPECIFIC CELL TYPE USING COMBINED MARKERS (CELLCOUNTER)
Circulating cells sample or another samples contain many different cell types. All of these cells express several different markers on their surface but these markers are seldom unique to one specific cell type. However, there are combinations of markers that are very specific to one particular cell, e.g. cancer cell, or stem cell.
We propose a microchannel-based device that is able to capture a specific cell type using combined markers for the analysis of cellular content of samples. The device is essentially a set of narrow channels through which the cells can be pumped through. Some areas on channel wall are functionalized with a combination of receptors that are binding to their corresponding markers on the cell’s surface. The flow conditions and the channel dimensions are optimized on the way that if more than one marker is present on the cell surface then the cell gets stuck in the channel. The number of such stuck channels can be determined by optical or electrical means, e.g. using the change in the impedance of the channel.
Patent no.: Ep14168842 (Microchannel-based cell capture device for the analysis of cellular content of samples)
DEVELOPMENT OF TECHNOLOGY FOR PRODUCTION OF ANTIBODIES
KINETO Lab intends to expand its existing tissue culture and animal-based infrastructure on the way that it can continually be able to isolate larger amount of antibodies several times a year, from the hybridomas created by its own earlier development or from the cells obtained from the academic and the industrial partners. The antibodies produced are marketed in various technical solutions and can be used as laboratory diagnostic kits, molecular biology reagents, for immunohistochemical techniques, etc.
At the moment we produce CD34 antibody which is validated for……. and available for selling (please see product list page (link) or/and download sheet (link)).
Immunohistochemistry performed on paraffin-embedded human clear cell renal cell carcinoma samples. Magnification: 200x(A-B), 400x (C). Vascular endothelial cells are stained red/brown. Cell nuclei are stained blue.
PDTX Models Development
PATIENT-DERIVED TUMOR XENOGRAFT (PDTX) MODELS
The urge of establishment of effective anticancer compounds lead to the massive number of drug candidates produced. These novel compounds are always going through preclinical testing, involving in vivo antitumor activity testing in rodents. However, traditional xenograft models, while offering good pharmacokinetical data, are limited due to cell line based background, lacking the variety of tumor cells typically present in patients. This often results low success rate in clinical trials after promising preclinical studies. Patient-derived tumor xenografts, originated directly from clinical tumors, offer a suitable solution for the issue: the structure and microenvironment of the tumors are maintained, and tumor heterogeneity is preserved for several generation of host animals.
KINETO Lab is collecting clinical samples of various cancer types, establishing and biobanking tumor samples for basic and applied research use. Our aim is to collect samples from rare malignancies or metastatic sites as well, providing otherwise hardly accessible disease models.
Introduction to PDTX
In vitro cellular assays and in vivo xenograft models are traditionally used in anticancer drug development. However, it turned clear that artificially grown tumor cells or xenografts act dramatically different than clinical tumors. This led to the shift from cell line models to genetically engineered models (GEMs) and patient-derived tumor xenograft models (PDTXs) in the preclinical drug development, not only having better molecular similarity to the clinical disease, but providing similar microenvironment, metabolic and vascularization properties as well.
Our aim is to use this model bank in collaboration with other research groups interested in cancer drug development, tumor phenotype biomarker research, tumor evolution, tumor genetic background, tumor heterogeneity and therapy resistance experiments. As reportedly 95% of drug candidates entering clinical trials fail, mostly because lack of effect and safety, and we believe that using better models drastically reduce the unnecessary clinical testing cost during drug development.
Our PDTX Project
We successfully developed our system to establish, archive and maintain PDTX models in immunodeficient (NOD-SCID or NSG) mice in order to provide hundreds of options to our partners to use the best preclinical models for in vivo drug testing. Also we establish PDTX derived cell line cultures (PDTX-C) from the models for the investigation of similarities and differences between cell cultures and serial xenografting. We are in connection with multiple clinical centers, allowing us to collect samples from broad variety of tumor types. Additionally, we are focusing on the collection of metastatic tumors in order to facilitate metastasis-targeting research.
The figures above show that PDTX tumours are preserving the morphological and immunohistochemical properties of the clinical tumor, while assessment of a derived cell line – similarly to other traditional cell line xenografts – comes along with a drastic change both tissue structure and molecular marker pattern.
Our workflow – PDTX establishment
We work in cooperation with 10 clinical oncology centers in Hungary in order to access various types of clinical tumor samples. Serial xenografting in immunodeficient mice, and biobanking of different outcome samples allows us to provide complete documentation about our PDTX models.
Our workflow – PDTX utilization
Based on our PDTX database, we are developing our online interface to see all relevant information about our models, including basic clinical data, results of past assays performed on the model, characterization of the model, and its availability. In vivo tumor experiments, molecular biology samples, or cell lines are equally reachable in order to optimize the tools for every research needs.
We are building our biobank to develop many different PDTX models by the end of 2024. This ongoing project is open to any collaborations and drug development experiments.
In case you are interested in PDTX models, do not hesitate to contact us:
József Tóvári, PhD, tovari.jozsef@KINETOlab.hu
Mihály Cserepes, PhD, cserepes.mihaly@KINETOlab.hu
DEVELOPMENT OF TECHNOLOGY FOR PRODUCTION OF ANTIBODIES
KINETO Lab develops new types of antibodies that are specifically active in tumor vessels, for diagnostic and therapeutic use. Identification of endothelial markers as a novel therapeutic target that can reduce the side effects of anticancer treatments and increase their effectiveness on tumors.
IDENTIFICATION OF ENDOTHELIAL MARKERS