Radoslav Mladenov
RWTH Aachen University
Network
Latest external collaboration on country level. Dive into details by clicking on the dots.
Publication
Featured researches published by Radoslav Mladenov.
Green Chemistry | 2010
Philip Engel; Radoslav Mladenov; Helene Wulfhorst; Gernot Jäger; Antje C. Spiess
New strategies are needed to efficiently convert non-food biomass to glucose as a platform chemical. One promising approach is to use ionic liquids to first dissolve lignocellulose. Yet, in the presence of such solvents, the enzymes that catalyze cellulose hydrolysis become compromised in their activity. However, this decreased cellulase activity has not been examined in detail. Thus, the aim of this study was to investigate how the ionic liquid precisely affects cellulase activity and stability with regard to different cellulose substrates. Hereby, four ionic liquids were screened to identify which one best minimized the loss of enzyme activity. Then, this best ionic liquid was tested on one insoluble and two soluble cellulose substrates. Subsequently, the relevant parameters of solution viscosity and ionic strength were evaluated with respect to enzyme activity and stability. Finally the residual ionic liquid concentration from the precipitation of α-cellulose was varied. The best ionic liquid was found to be 1,3-dimethylimidazolium dimethylphosphate with the highest retained activity of 30% on the α-cellulose substrate in the presence of 10% (v/v) ionic liquid. Most importantly, an increase in viscosity and ionic strength contributed to the decrease in enzyme activity which nonetheless retained their stability. The hydrolysis of precipitated α-cellulose from ionic liquid showed significant higher reaction rates but reduced sugar yields when residual ionic liquid was present. None the less, it should be possible to effectively produce glucose from precipitated cellulose without needing to wash off all residual ionic liquid when optimized cellulase mixtures are used.
Toxins | 2012
Dmitrij Hristodorov; Radoslav Mladenov; Michael Huhn; Stefan Barth; Theo Thepen
Diseases caused by chronic inflammation (e.g., arthritis, multiple sclerosis and diabetic ulcers) are multicausal, thus making treatment difficult and inefficient. Due to the age-associated nature of most of these disorders and the demographic transition towards an overall older population, efficient therapeutic intervention strategies will need to be developed in the near future. Over the past decades, elimination of activated macrophages using CD64-targeting immunotoxins has proven to be a promising way of resolving inflammation in animal models. More recent data have shown that the M1-polarized population of activated macrophages in particular is critically involved in the chronic phase. We recapitulate the latest progress in the development of IT. These have advanced from full-length antibodies, chemically coupled to bacterial toxins, into single chain variants of antibodies, genetically fused with fully human enzymes. These improvements have increased the range of possible target diseases, which now include chronic inflammatory diseases. At present there are no therapeutic strategies focusing on macrophages to treat chronic disorders. In this review, we focus on the role of different polarized macrophages and the potential of CD64-based IT to intervene in the process of chronic inflammation.
Cancer Letters | 2013
Sonja Schiffer; Soriba Letzian; Edgar Jost; Radoslav Mladenov; Dmitrij Hristodorov; Michael Huhn; Rainer Fischer; Stefan Barth; Theo Thepen
Immunotoxins are promising targeted therapeutic agents comprising an antibody-based ligand that specifically binds to diseased cells, and a pro-apoptotic protein. Toxic components from bacteria or plants can trigger a neutralizing immune response, so that human effector molecules are more suitable. In this context, the protease granzyme B has been successfully tested in cytotoxicity assays against different cancer cells in vitro and in vivo. Our aim here was to introduce granzyme M as an alternative and novel component of human cytolytic fusion proteins. We fused it to the humanized single-chain antibody fragment (scFv) H22 which specifically binds to CD64, an FcγRI receptor overexpressed on activated myeloid cells and leukemic cells. We show that the humanized cytolytic fusion protein Gm-H22(scFv) specifically targets the acute myeloid leukemia cell line HL60 in vitro and is cytotoxic with an IC50 between 1.2 and 6.4 nM. These findings were confirmed ex vivo using leukemic primary cells from patients, which were killed by granzyme M despite the presence of the granzyme B inhibitor serpin B9. In conclusion, granzyme M is a promising new cell-death inducing component for hCFPs because it specifically and efficiently kills target cells when fused to a targeting component.
International Journal of Cancer | 2016
Manal Amoury; Radoslav Mladenov; Thomas Nachreiner; Anh-Tuan Pham; Dmitrij Hristodorov; Stefano Di Fiore; Wijnand Helfrich; Alessa Pardo; Georg H. Fey; Michael Schwenkert; Theophilus Thepen; Fabian Kiessling; Ahmad Fawzi Hussain; Rainer Fischer; Katharina Kolberg; Stefan Barth
Chondroitin sulfate proteoglycan 4 (CSPG4) has been identified as a highly promising target antigen for immunotherapy of triple‐negative breast cancer (TNBC). TNBC represents a highly aggressive heterogeneous group of tumors lacking expression of estrogen, progesterone and human epidermal growth factor receptor 2. TNBC is particularly prevalent among young premenopausal women. No suitable targeted therapies are currently available and therefore, novel agents for the targeted elimination of TNBC are urgently needed. Here, we present a novel cytolytic fusion protein (CFP), designated αCSPG4(scFv)‐MAP, that consists of a high affinity CSPG4‐specific single‐chain antibody fragment (scFv) genetically fused to a functionally enhanced form of the human microtubule‐associated protein (MAP) tau. Our data indicate that αCSPG4(scFv)‐MAP efficiently targets CSPG4+ TNBC‐derived cell lines MDA‐MB‐231 and Hs 578T and potently inhibits their growth with IC50 values of ∼200 nM. Treatment with αCSPG(scFv)‐MAP resulted in induction of the mitochondrial stress pathway by activation of caspase‐9 as well as endonuclease G translocation to the nucleus, while induction of the caspase‐3 apoptosis pathway was not detectable. Importantly, in vivo studies in mice bearing human breast cancer xenografts revealed efficient targeting to and accumulation of αCSPG4(scFv)‐MAP at tumor sites resulting in prominent tumor regression. Taken together, this preclinical proof of concept study confirms the potential clinical value of αCSPG4(scFv)‐MAP as a novel targeted approach for the elimination of CSPG4‐positive TNBC.
Molecular Cancer Therapeutics | 2014
Dmitrij Hristodorov; Manal Amoury; Radoslav Mladenov; Judith Niesen; Katharina Arens; Nina Berges; Lea Hein; Stefano Di Fiore; Anh-Tuan Pham; Michael Huhn; Wijnand Helfrich; Rainer Fischer; Theo Thepen; Stefan Barth
In normal epithelia, the epithelial cell adhesion molecule (EpCAM) expression is relatively low and only present at the basolateral cell surface. In contrast, EpCAM is aberrantly overexpressed in various human carcinomas. Therefore, EpCAM is considered to be a highly promising target for antibody-based cancer immunotherapy. Here, we present a new and fully human cytolytic fusion protein (CFP), designated “anti–EpCAM(scFv)-MAP,” that is comprised of an EpCAM-specific antibody fragment (scFv) genetically fused to the microtubule-associated protein tau (MAP). Anti–EpCAM(scFv)-MAP shows potent EpCAM-restricted proapoptotic activity toward rapidly proliferating carcinoma cells. In vitro assays confirmed that treatment with anti–EpCAM(scFv)-MAP resulted in the colocalization and stabilization of microtubules, suggesting that this could be the potential mode of action. Dose-finding experiments indicated that anti–EpCAM(scFv)-MAP is well tolerated in mice. Using noninvasive far-red in vivo imaging in a tumor xenograft mouse model, we further demonstrated that anti–EpCAM(scFv)-MAP inhibited tumor growth in vivo. In conclusion, our data suggest that anti–EpCAM(scFv)-MAP may be of therapeutic value for the targeted elimination of EpCAM+ carcinomas. Mol Cancer Ther; 13(9); 2194–202. ©2014 AACR.
Cancer Letters | 2016
Manal Amoury; Katharina Kolberg; Anh-Tuan Pham; Dmitrij Hristodorov; Radoslav Mladenov; Stefano Di Fiore; Wijnand Helfrich; Fabian Kiessling; Rainer Fischer; Alessa Pardo; Theophilus Thepen; Ahmad Fawzi Hussain; Thomas Nachreiner; Stefan Barth
Triple-negative breast cancer (TNBC) is associated with poor prognosis and high prevalence among young premenopausal women. Unlike in other breast cancer subtypes, no targeted therapy is currently available. Overexpression of epithelial cell adhesion molecule (EpCAM) in 60% of TNBC tumors correlates with poorer prognosis and is associated with cancer stem cell phenotype. Thus, selective elimination of EpCAM(+) TNBC tumor cells is of clinical importance. Therefore, we constructed a fully human targeted cytolytic fusion protein, designated GbR201K-αEpCAM(scFv), in which an EpCAM-selective single-chain antibody fragment (scFv) is genetically fused to a granzyme B (Gb) mutant with reduced sensitivity to its natural inhibitor serpin B9. In vitro studies confirmed its specific binding, internalization and cytotoxicity toward a panel of EpCAM-expressing TNBC cells. Biodistribution kinetics and tumor-targeting efficacy using MDA-MB-468 cells in a human TNBC xenograft model in mice revealed selective accumulation of GbR201K-αEpCAM(scFv) in the tumors after i.v. injection. Moreover, treatment of tumor-bearing mice demonstrated a prominent inhibition of tumor growth of up to 50 % in this proof-of-concept study. Taken together, our results indicate that GbR201K-αEpCAM(scFv) is a promising novel targeted therapeutic for the treatment of TNBC.
Cancer Letters | 2015
Hannes Brehm; Dmitrij Hristodorov; Alessa Pardo; Radoslav Mladenov; Judith Niesen; Rainer Fischer; Mehmet Kemal Tur; Stefan Barth
The treatment of rhabdomyosarcoma (RMS) is challenging, and the prognosis remains especially poor for high-grade RMS with metastasis. The conventional treatment of RMS is based on multi-agent chemotherapy combined with resection and radiotherapy, which are often marked by low success rate. Alternative therapeutic options include the combination of standard treatments with immunotherapy. We generated a microtubule-associated protein (MAP)-based fully human cytolytic fusion protein (hCFP) targeting the fetal acetylcholine receptor, which is expressed on RMS cells. We were able to express and purify functional scFv35-MAP from Escherichia coli cells. Moreover, we found that scFv35-MAP is rapidly internalized by target cells after binding its receptor, and exhibits specific cytotoxicity toward FL-OH1 and RD cells in vitro. We also confirmed that scFv35-MAP induces apoptosis in FL-OH1 and RD cells. The in vivo potential of scFv35-MAP will need to be considered in further studies.
mAbs | 2014
Dmitrij Hristodorov; Radoslav Mladenov; Hannes Brehm; Rainer Fischer; Stefan Barth; Theo Thepen
Tumor necrosis factor (TNF) is a pro-inflammatory cytokine that plays a critical role in many inflammatory diseases. Soluble TNF can be neutralized by monoclonal antibodies (mAbs), and this is a widely-used therapeutic approach. However, some patients do not respond to anti-TNF therapy due to the increased expression of CD64 on monocytes and macrophages. A recent study has shown that CD64 captures anti-TNF mAbs via their Fcγ domain, which induces the transcription of pro-inflammatory genes. Specific blocking of CD64 could therefore be a promising strategy to improve the response to anti-TNF therapy. We used the CD64-specific antibody fragment H22(scFv) and tested its activity against the human CD64+ cell line HL-60. When stimulated with interferon gamma (IFN-γ), these cells represent a pro-inflammatory phenotype of the monocyte/macrophage lineage. We found that H22(scFv) binds selectively to and blocks CD64, preventing the capture of anti-TNF mAb. Importantly, H22(scFv) itself does not induce CD64 activation. We also found that transmembrane TNF on HL-60 cells stimulated with IFN-γ also contributes to the capture of anti-TNF mAb, although via their Fab domain. In conclusion, the specific blocking of CD64 by H22(scFv) could be used a possible anti-inflammatory mechanism for potentiating the effect of anti-TNF antibodies.
Biomedicines | 2017
Olusiji Akinrinmade; Sandra Jordaan; Dmitrij Hristodorov; Radoslav Mladenov; Neelakshi Mungra; Shivan Chetty; Stefan Barth
Some of the most promising small molecule toxins used to generate antibody drug conjugates (ADCs) include anti-mitotic agents (e.g., auristatin and its derivatives) which are designed to attack cancerous cells at their most vulnerable state during mitosis. We were interested in identifying a human cystostatic protein eventually showing comparable activities and allowing the generation of corresponding targeted fully human cytolytic fusion proteins. Recently, we identified the human microtubule associated protein tau (MAP tau), which binds specifically to tubulin and modulates the stability of microtubules, thereby blocking mitosis and presumably vesicular transport. By binding and stabilizing polymerized microtubule filaments, MAP tau-based fusion proteins skew microtubule dynamics towards cell cycle arrest and apoptosis. This biological activity makes rapidly proliferating cells (e.g., cancer and inflammatory cells) an excellent target for MAP tau-based targeted treatments. Their superior selectivity for proliferating cells confers additional selectivity towards upregulated tumor-associated antigens at their surface, thereby preventing off-target related toxicity against normal cells bearing tumor-associated antigens at physiologically normal to low levels. In this review, we highlight recent findings on MAP tau-based targeted cytolytic fusion proteins reported in preclinical immunotherapeutic studies.
Oncotarget | 2016
Radoslav Mladenov; Dmitrij Hristodorov; Christian Cremer; Gerrit Gresch; Elena Grieger; Lea Schenke; Diana Klose; Manal Amoury; Mira Woitok; Edgar Jost; Tim H. Brümmendorf; Rolf Fendel; Rainer Fischer; Christoph Stein; Theo Thepen; Stefan Barth
Fc gamma receptor I (FcγRI, CD64) is a well-known target antigen for passive immunotherapy against acute myeloid leukemia and chronic myelomonocytic leukemia. We recently reported the preclinical immunotherapeutic potential of microtubule associated protein tau (MAP) against a variety of cancer types including breast carcinoma and Hodgkins lymphoma. Here we demonstrate that the CD64-directed human cytolytic fusion protein H22(scFv)-MAP kills ex vivo 15–50% of CD64+ leukemic blasts derived from seven myeloid leukemia patients. Furthermore, in contrast to the nonspecific cytostatic agent paclitaxel, H22(scFv)-MAP showed no cytotoxicity towards healthy CD64+ PBMC-derived cells and macrophages. The targeted delivery of this microtubule stabilizing agent therefore offers a promising new strategy for specific treatment of CD64+ leukemia.