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Biomedical subjects

K Ulbrich

Publications and source records attributed to K Ulbrich.

At least 19 recordsLinked to original sources
PubMed2006

Use of synthetic vectors for neutralising antibody resistant delivery of replicating adenovirus DNA.

Use of synthetic vectors to deliver genomes of conditionally replicating lytic viruses combines the strengths of viral and non-viral approaches by enabling neutralising antibody resistant deployment of cancer virotherapy. Adenovirus is particularly suitable for this application since all proteins essential for replication can be expressed from the input DNA, although the presence of terminal protein (TP) covalently linked to the 5' termini of the input virus genomes both improves expression of transgenes encoded in the input DNA and also enhances replication. These roles of TP were distinguished in experiments where E1-deleted Ad(GFP)DNA bearing TP (Ad(GFP)DNA-TP), delivered with DOTAP, gave a two-fold greater frequency of transduction than Ad(GFP)DNA(without TP) in non-complementing A549 cells, while in 293 cells (which support replication of E1-deleted viruses) the presence of TP mediated a much greater differential transgene expression, commensurate with its ability to promote replication. Subsequent studies using AdDNA for virotherapy, therefore, included covalently linked TP. AdDNA-TP delivered to A549 cells using a synthetic polyplex vector was shown to be resistant to levels of neutralising antisera that completely ablated infection by wild-type adenovirus, enabling polyplex/Ad(wild type)DNA-TP to mediate a powerful cytopathic effect. Similarly in vivo, direct injection of a polyplex/Ad(wild type)DNA-TP into A549 tumours was neutralising antibody-resistant and enabled virus replication, whereas intact virus was neutralised by the antibody and failed to infect. The delivery of adenovirus genomes-TP using synthetic vectors should provide a strategy to bypass neutralising antibodies and facilitate clinical application of replicating adenovirus for cancer virotherapy.

Adenoviridae↗
PubMed2006

Chick embryo lethal orphan virus can be polymer-coated and retargeted to infect mammalian cells.

Non-human adenovirus vectors have attractive immunological properties for gene therapy but are frequently restricted by inefficient transduction of human target cells. Using chicken embryo lethal orphan (CELO) virus, we employed a nongenetic mechanism of polymer coating and retargeting with basic fibroblast growth factor (bFGF-pc-CELOluc), a strategy that permits efficient tropism modification of human adenovirus. bFGF-pc-CELOluc showed efficient uptake and transgene expression in chick embryo fibroblasts (CEF), and increased levels of binding and internalization in a variety of human cell lines. Transgene expression was also greater than unmodified CELOluc in PC-3 human prostate cells, although the specific activity (RLU per internalized viral genome) was decreased. In CEF, the specific activity of bFGF-pc-CELOluc was considerably higher than in the human prostate cell line PC-3. Retargeted virus was fully resistant to inhibition by human serum with known adenovirus-neutralizing activity in vitro, while in mice CELOluc was cleared less rapidly from the blood than Adluc following i.v. administration in the presence of adenovirus neutralizing serum. Polymer coating and retargeting with bFGF further reduced rates of clearance for both viruses, suggesting protection against both neutralizing and opsonizing factors. The data indicate that CELO virus may be retargeted to infect human cells via alternative, potentially disease-specific, receptors and resist the effects of pre-existing humoral immunity.

ATPases Associated with Diverse Cellular Activitie↗
PubMed2005

Induction of systemic antitumour resistance with targeted polymers.

Conjugates based on N-(2-hydroxypropyl)methacrylamide (HPMA) represent a new generation of antibody-targeted polymeric anticancer drugs with both cytotoxic and immunoprotecting/immunomobilizing activity. 20-90% of mice that are cured of EL4 mouse T-cell lymphoma, BCL1 mouse B-cell leukaemia and 38C13 mouse B-cell lymphoma by injection of doxorubicin-HPMA conjugate develop a long-lasting memory and systemic antitumour resistance. It is suggested that the main activity of the polymeric drug, directly after application is - due to the high level of the drug - of cytotoxic and cytostatic nature. Thereafter, long-term conjugates persist at low concentration in the circulation, which are capable of mobilizing the defence mechanisms of the host. Until now, seven patients with generalized carcinoma were treated with doxorubicin-HPMA-human-Ig conjugate. Disease stabilization, lasting from 6 to more than 18 months, was recorded.

Animals↗
PubMed2004

Achieving 99.9% proton spin-flip efficiency at higher energy with a small RF dipole.

We recently used a new ferrite rf dipole to study spin flipping of a 2.1 GeV/c vertically polarized proton beam stored in the COSY Cooler Synchrotron in Jülich, Germany. We swept the rf dipole's frequency through an rf-induced spin resonance to flip the beam's polarization direction. After determining the resonance's frequency, we varied the frequency range, frequency ramp time, and number of flips. At the rf dipole's maximum strength and optimum frequency range and ramp time, we measured a spin-flip efficiency of 99.92+/-0.04%. This result, along with a similar 0.49 GeV/c IUCF result, indicates that, due to the Lorentz invariance of an rf dipole's transverse integralBdl and the weak energy dependence of its spin-resonance strength, an only 35% stronger rf dipole should allow efficient spin flipping in the 100 GeV BNL RHIC Collider or even the 7 TeV CERN Large Hadron Collider.

Journal Article↗
PubMed2004

HPMA copolymers containing doxorubicin bound by a proteolytically or hydrolytically cleavable bond: comparison of biological properties in vitro.

N-(2-Hydroxypropyl)methacrylamide (HPMA) copolymer carrier containing the anticancer drug doxorubicin bound either by a proteolytically degradable bond (non-targeted PK1 or targeted with alpha-CD71 mAb) or by a hydrolytically degradable bond were synthesised and tested in vivo for various biological properties. Mouse 38C13 B-cell lympoma was used as a well established and defined cell line for this study. 38C13 cells are sensitive to free doxorubicin and IC50 was very low, about 0.014 microM. PK1 showed a strongly decreased cytostatic effect, IC50 being 12.6 microM. alpha-CD71 targeted conjugate, which can be considered as an antibody-targeted form of PK1, had IC50 0.358 microM. HPMA copolymer with doxorubicin bound via a hydrolytically sensitive bond (HYD conjugate) showed a high cytostatic effect with IC50 about 0.052 microM. We demonstrated that HYD conjugate inhibited DNA synthesis and induced p21(Waf1/Cip1) protein expression (p21(Waf1/Cip1) is cyclin-dependent kinase inhibitor which blocks cell cycle progression) as quickly as free doxorubicin, whereas PK1 acted much more slowly. Similarly, apoptosis induction measured by Annexin V binding and Caspase 3 activity was detected later after incubation of cells with PK1 or alpha-CD71 targeted conjugate. Apoptosis was manifested by elevation of bax and bad mRNA levels, which was much more rapid and intense in the case of free doxorubicin and HYD conjugate. Expression of antiapoptotic genes as well as cyclin-dependent kinases was surprisingly not affected.

Acrylamides↗
PubMed2004

Polymeric anticancer drugs with pH-controlled activation.

The paper is dealing with the synthesis and properties of new, nontargeted or antibody-targeted pH-sensitive polymer-doxorubicin (DOX) conjugates designed as anticancer drugs facilitating site-specific therapy. These conjugates are stable and inactive during transport in the body but activate inside target cells as a result of pH changes outside and inside the cells. Cytotoxicity of the conjugates depends on the detailed structure of the polymer and of the spacer between the drug and polymer carrier. In both protective and therapeutic regimes of drug administration, the in vivo antitumor activity of the pH-sensitive conjugates containing DOX was significantly enhanced (T-cell lymphoma EL 4, C57BL/16 mice) in comparison with the free DOX or classic PK1, the PHPMA-DOX conjugate clinically tested at present.

Animals↗
PubMed2004

Polymer-conjugated bovine pancreatic and seminal ribonucleases inhibit growth of human tumors in nude mice.

The hydrophilic poly[N-(2-hydroxypropyl)methacrylamide] (PHPMA) was used for RNase A or BS-RNase modification to prevent their degradation in bloodstream or fast elimination. Two PHPMA chains (classic and star-like) were synthesized and their conjugates with both enzymes were tested on the CD-1 nude mice bearing various human tumors. These RNase conjugates injected intravenously or intraperitoneally into the mice bearing melanoma, neuroblastoma or ovarian tumor caused significant reduction of transplanted tumors following ten daily doses of 2.5 and/or 1 mg/kg, respectively, while free RNase A or BS-RNase injected in doses of 10 mg/kg exerted only negligible antitumor activity. Histological examination confirmed potent cytotoxic effect of RNase A conjugates in ovarian tumor. Despite the antitumor activity observed in vivo, the in vitro cytotoxic activity of RNase A conjugates was not pronounced and did not differ from that caused by the free RNase A. The in vitro experiments with 125I-labeled preparations demonstrated that polymer conjugates were internalized by tumor cells very poorly in contrast to the dose-dependent internalization of the wild enzyme preparation. Surprisingly, mice injected with EL-4 leukemic cells, which were preincubated for 4 h with BS-RNase conjugates, exerted significantly prolonged survival compared with the control non-treated mice. It may be supposed that both BS-RNase and RNase A conjugates with PHPMA act after administration in vivo by a mechanism different from that or those occurring under in vitro conditions because in vivo they exert an antitumor action, whereas in vitro, they are ineffective. The experiments proved that RNase A, when conjugated to PHPMA, produced identical aspermatogenic and antitumor effects as BS-RNase conjugated to this polymer and that this preparation may be regarded as a potential anticancer drug.

Animals↗
PubMed2004

Extended plasma circulation time and decreased toxicity of polymer-coated adenovirus.

Systemic delivery of adenoviral vectors is a major goal in cancer gene therapy, but is currently prohibited by rapid hepatic uptake of virus following intravenous injection with levels of viable virus in the murine plasma typically falling to less than 0.1% after 30 min. We have used a surface-masking technique based on multivalent copolymers of poly(N-(2-hydroxypropyl)methacrylamide) to ablate all pathways of receptor-mediated infection, combined with dose modulation to achieve partial saturation of nonspecific uptake pathways. Polymer coating gave at least 100-fold decreased hepatic transgene expression at all doses and even high doses of coated virus (pc-virus) showed no weight loss or stimulation of serum transaminases. Low doses of virus and pc-virus (10(9) viral particles (vp)/mouse) were mainly captured by the liver (assessed by quantitative PCR), although higher doses led to greater fractional persistence in the plasma (measured after 30 min). Coated virus at a dose of 6 x 10(11) vp/mouse showed nearly 50% plasma circulation, representing a 3.5-fold greater area under the concentration-time curve (0-30 min) compared to unmodified virus. Such an increase in the bioavailability of adenovirus, coupled with substantial decreases in toxicity and unwanted transgene expression is an important step towards producing systemically available tumour-targeted viruses.

Adenoviridae↗
PubMed2004

Antibody-targeted polymer-doxorubicin conjugates with pH-controlled activation.

The paper is dealing with the synthesis and properties of new non-targeted or antibody-targeted polymer drug conjugates, bearing doxorubicin (DOX) attached via a spacer susceptible to pH-controlled hydrolysis (hydrazone conjugates), designed as anticancer drugs facilitating site-specific therapy. These conjugates are stable in a pH 7.4 buffer, modeling conditions during transport in the body, but release DOX and activate it inside target cells as a result of pH changes when going from outside to inside the cells. Conjugates containing an antibody directed against T lymphocytes bind effectively and specifically T cell lymphoma EL 4 cells. Cytotoxicity of the hydrazone conjugates is higher than that of classic conjugates, depending on the detailed structure of the polymer, the spacer between the drug and polymer carrier and method of antibody conjugation. Cytotoxicity of some of the conjugates is comparable even with that of the free drug. In both protective and therapeutic regimes of drug administration, the in vivo anti-tumor activity of the conjugates containing DOX was enhanced with long-term survivors (T-cell lymphoma EL 4, C57BL/6 mice) in comparison with much less effective free DOX or a classic P(N-(2-hydroxypropyl)methacrylamide)HPMA-DOX conjugate (already clinically tested).

Animals↗
PubMed2003

HPMA copolymer-bound doxorubicin targeted to tumor-specific antigen of BCL1 mouse B cell leukemia.

N-(2-Hydroxypropyl)methacrylamide (HPMA) copolymer carrier containing the anticancer drug doxorubicin and targeted with B1 monoclonal antibody (mAb) to BCL1 leukemia cells was synthesised and tested in vitro and in vivo. BCL1 leukemia growing in syngenic Balb/c mice was selected as a tumor model system. B1 mAb recognising the idiotype of surface IgM on BCL1 cells was used as a targeting moiety. Both B1 mAb and doxorubicin were conjugated to HPMA copolymer carrier by aminolysis through a tetrapeptidic Gly-Phe(D,L)-Leu-Gly spacer to ensure the intracellular delivery and controlled release of the drug. B1 mAb-targeted conjugate was shown to possess strictly tumor-specific binding capacity to target BCL1 cells in vitro. A similar conjugate, but containing human nonspecific Ig (HuIg) instead of B1 mAb, failed to bind to BCL1 cells. In vitro, B1 mAb-targeted conjugate demonstrated 40-fold higher cytotoxic effect than nontargeted or human nonspecific Ig-containing HPMA copolymer-bound doxorubicin. Conjugate targeted with B1 mAb was also shown to bind to target BCL1 cells in vivo. B1 mAb-targeted conjugate was shown to be more efficient in the treatment of established BCL1 leukemia than free doxorubicin, nontargeted and human nonspecific Ig-containing conjugate. Antibody-targeted polymeric drugs are thus promising conjugates for cancer treatment.

Acrylamides↗
PubMed2003

Measurement of spin-correlation parameters ANN, ASS, and ASL at 2.1 GeV in proton-proton elastic scattering.

At the Cooler Synchrotron COSY/Jülich spin-correlation parameters in elastic proton-proton (pp) scattering have been measured with a 2.11 GeV polarized proton beam and a polarized hydrogen atomic beam target. We report results for A(NN), A(SS), and A(SL) for c.m. scattering angles between 30 degrees and 90 degrees. Our data on A(SS)--the first measurement of this observable above 800 MeV--clearly disagrees with predictions of available pp scattering phase-shift solutions while A(NN) and A(SL) are reproduced reasonably well. We show that in the direct reconstruction of the scattering amplitudes from the body of available pp elastic scattering data at 2.1 GeV the number of possible solutions is considerably reduced.

Journal Article↗
PubMed2003

HPMA copolymers with pH-controlled release of doxorubicin: in vitro cytotoxicity and in vivo antitumor activity.

Data on the synthesis, physicochemical characterisation and in vitro and in vivo biological properties of the new, nontargeted or antibody-targeted polymer-doxorubicin conjugates designed as anticancer drugs are presented. In the conjugates, the anticancer drug doxorubicin (DOX) is attached to the polymer carrier via a simple hydrolytically labile spacer containing either a hydrazone bond or cis-aconitic acid residue. In vitro incubation of the conjugates in buffers led to a fast DOX release from the polymer at pH 5 (modelling intracellular environment) while at pH 7.4 (modelling blood) the conjugates are relatively stable. Cytotoxicity of the conjugates to T cell lymphoma EL4 depended on the detailed structure of the spacer and the method used for antibody attachment and was much higher compared with the effect of similar classic conjugates (DOX attached to the polymer via enzymatically degradable spacer). In both protective and therapeutic regimes of drug administration, the in vivo anti-tumor activity of the hydrazone conjugates containing only DOX was significantly enhanced (T cell lymphoma EL4, C57BL/10 mice) in comparison with free DOX or classic PK1, the PHPMA-DOX conjugate clinically tested at present. Increasing the molecular weight of the polymer carrier resulted in a more pronounced in vivo antitumor effect. Antibody-targeted conjugates with DOX bound via hydrazone bond exhibited even more extensive inhibition of the tumor growth with some long-term survivors. No survivors were observed after treatment of mice with free DOX or the nontargeted PHPMA-DOX conjugate.

Animals↗
PubMed2002

Differences in the intracellular fate of free and polymer-bound doxorubicin.

Internalization and subcellular fate of free doxorubicin or its polymeric conjugates based on poly N-(2-hydroxypropyl)methacrylamide (pHPMA), either non-targeted or targeted with anti-Thy1.2 or anti-CD71 monoclonal antibody was tested on EL-4 mouse T-cell lymphoma, SW620 human colorectal carcinoma and OVCAR-3 human ovarian adenocarcinoma. Doxorubicin fluorescence allowed us to follow the internalization and intracellular distribution of tested conjugates by laser scanning confocal microscopy and/or by fluorescent microscopy. Whereas free doxorubicin was always detectable only in the nuclei of treated cells, detectable fluorescence of doxorubicin bound to a polymeric carrier, targeted or non-targeted, was detectable up to 3 days of incubation only in the cytoplasmatic structures. While free doxorubicin causes apoptosis in the populations of tested cancer cell lines, significant number of apoptotic cells was never found in cell cultures exposed to targeted or non-targeted polymeric conjugates. In contrast to free doxorubicin, which is a strong inducer of p53 expression, increased p53 expression was never observed after the treatment with the polymeric drug. High-performance liquid chromatographic analysis shows that the percentage of cleaved doxorubicin is very low even after 48 h of incubation of tested cells with the polymeric conjugate, and cannot be the only reason for the toxicity of the conjugate. We suggest that: (a) after the treatment with pHPMA-bound drug, the cells die by necrosis and (b) the toxicity of pHPMA-based conjugates is a combination of the toxic effect of released doxorubicin and the toxic effect of doxorubicin in polymer-bound form directed against cell membranes.

Animals↗
PubMed2002

Acquired and specific immunological mechanisms co-responsible for efficacy of polymer-bound drugs.

We present data providing new evidence that poly[N-(2-hydroxypropyl)methacrylamide] (PHPMA)-bound drugs, unlike free drugs, have both cytostatic and immunomobilizing activity (CIA). Immediately after injection, due to the high level of the drug, the main activity of the polymeric conjugate is cytotoxic and cytostatic. Later on, long-term circulating PHPMA-bound drug, at concentrations lower than its minimal inhibitory levels, mobilizes the defense mechanisms of the host. Cytotoxic and cytostatic effects of drug-PHPMA were repeatedly confirmed. The following data support the concept of the immunomobilizing activity of the N-(2-hydroxypropyl)methacrylamide (HPMA) conjugates: (a) pre-treatment with free drugs (doxorubicin, cyclosporin A) accelerates the appearance of EL4 mouse T-cell lymphoma while a similar pre-treatment with doxorubicin-PHPMA induces limited but definitive mobilization of the host's defense mechanisms; (b) mice cured of EL4 mouse T-cell lymphoma, BCL1 mouse B-cell leukemia and 38C13 mouse B-cell lymphoma by injection of doxorubicin-PHPMA conjugate targeted with monoclonal antibodies (anti-Thy 1.2 for EL4, anti-B1 for BCL1 and anti-CD71 for 38C13) and re-transplanted with a lethal dose of the same cancer cells survive without any treatment considerably longer than control mice; (c) increased NK activity and anti-cancer antibody was detected only in animals treated with doxorubicin-PHPMA conjugate; and (d) considerably increased NK and LAK activity was seen in a human patient treated for generalized breast carcinoma with doxorubicin-PHPMA-IgG.

Animals↗
PubMed2002

HPMA-hydrogels result in prolonged delivery of anticancer drugs and are a promising tool for the treatment of sensitive and multidrug resistant leukaemia.

Treatment of an established BCL1 leukaemia in mice showed that the use of hydrogels is advantageous in comparison with free doxorubicin (DOX), partially due to the different pharmacokinetic profile of the drug release. Pharmacologically active concentrations ranging from 100 to 800 ng/ml were detectable in the bloodstream for more than 4 days when DOX-loaded hydrogels were implanted into mice. Animals treated with free DOX survived for 35 days, survival of hydrogel-DOX treated animals increased up to 60 days and long-term survivors were achieved, when the second hydrogel was implanted 2 weeks after the first one. Hydrogels containing vinblastine (VLB) were ineffective. N-(2-hydroxypropyl)methacrylamide (HPMA) hydrogels were also used in combined therapy against multidrug resistant leukaemia P388-MDR to achieve a synergistic effect of both the cytostatic drug and chemosensitising agent. It was shown that when 4 times the maximal tolerated dose (MTD) of free DOX was incorporated into HPMA-hydrogels, tumour volume was reduced by approximately 50% after implantation of the hydrogel containing DOX and cyclosporine A (CsA) and survival was slightly prolonged.

Animals↗
PubMed2002

In vitro and in vivo effect of HPMA copolymer-bound doxorubicin targeted to transferrin receptor of B-cell lymphoma 38C13.

N-(2-Hydroxypropyl)methacrylamide (HPMA) copolymers containing the anticancer agent doxorubicin and targeted to the transferrin receptor either with anti-mouse CD71 monoclonal antibody (mAb) or with transferrin were synthesized to evaluate their binding and anti-proliferative activity in vitro and anti-tumor potential against 38C13 B-cell lymphoma in vivo. Both the doxorubicin and the targeting moieties were bound to HPMA copolymer chain by aminolysis via a Gly-Phe(D,L)-Leu-Gly spacer to ensure controlled intracellular release of the conjugated drug. We demonstrated that HPMA copolymer-bound doxorubicin targeted to the transferrin receptor with anti-mouse CD71 mAb strongly retards tumor growth, prolongs the survival and completely cures three out of nine experimental mice with established 38C13 tumors. The conjugate targeted with transferrin was less effective in vitro as well as in vivo. It completely cured only one out of seven experimental mice. Free or non-targeted HPMA copolymer-bound doxorubicin showed only a mild anti-tumor effect within the therapeutic schedule used. In vitro, HPMA copolymer-bound doxorubicin targeted with anti-mouse CD71 mAb shows approximately 4-fold higher cytotoxic effect than HPMA copolymer-bound doxorubicin targeted with transferin and 9-fold higher cytotoxic effect than non-targeted HPMA copolymer-bound doxorubicin.

Animals↗