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Jindřich Kopecek - One of the best experts on this subject based on the ideXlab platform.
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backbone degradable n 2 hydroxypropyl Methacrylamide copolymer conjugates with gemcitabine and paclitaxel impact of molecular weight on activity toward human ovarian carcinoma xenografts
Molecular Pharmaceutics, 2017Co-Authors: Jiyuan Yang, Huaizhong Pan, Rui Zhang, Yixin Fang, Libin Zhang, Jindřich KopecekAbstract:Degradable diblock and multiblock (tetrablock and hexablock) N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer-gemcitabine (GEM) and -paclitaxel (PTX) conjugates were synthesized by reversible addition-fragmentation chain-transter (RAFT) copolymerization followed by click reaction for preclinical investigation. The aim was to validate the hypothesis that long-circulating conjugates are needed to generate a sustained concentration gradient between vasculature and a solid tumor and result in significant anticancer effect. To evaluate the impact of molecular weight of the conjugates on treatment efficacy, diblock, tetrablock, and hexablock GEM and PTX conjugates were administered intravenously to nude mice bearing A2780 human ovarian xenografts. For GEM conjugates, triple doses with dosage 5 mg/kg were given on days 0, 7, and 14 (q7dx3), whereas a single dose regime with 20 mg/kg was applied on day 0 for PTX conjugates treatment. The most effective conjugates for each monotherapy were the diblock ones, 2P-GEM and 2P-PTX (Mw ≈ 100 kDa). Increasing the Mw to 200 or 300 kDa resulted in decrease of activity most probably due to changes in the conformation of the macromolecule because of interaction of hydrophobic residues at side chain termini and formation of "unimer micelles". In addition to monotherapy, a sequential combination treatment of diblock PTX conjugate followed by GEM conjugate (2P-PTX/2P-GEM) was also performed, which showed the best tumor growth inhibition due to synergistic effect: complete remission was achieved after the first treatment cycle. However, because of low dose applied, tumor recurrence was observed 2 weeks after cease of treatment. To assess optimal route of administration, intraperitoneal (i.p.) application of 2P-GEM, 2P-PTX, and their combination was examined. The fact that the highest anticancer efficiency was achieved with diblock conjugates that can be synthesized in one scalable step bodes well for the translation into clinics.
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n 2 hydroxypropyl Methacrylamide copolymer drug conjugates for combination chemotherapy of acute myeloid leukemia
Macromolecular Bioscience, 2016Co-Authors: Rui Zhang, Jiyuan Yang, Yan Zhou, Paul J Shami, Jindřich KopecekAbstract:There is a need for new treatment strategies of acute myeloid leukemia (AML). In this study, four different drugs, including cytarabine, daunorubicin, GDC-0980, and JS-K, were investigated in vitro for the two-drug combinations treatment of AML. The results revealed that cytarabine and GDC-0980 had the strongest synergism. In addition, cell cycle analysis was conducted to investigate the effect of the different combinations on cell division. For future in vivo application, N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer-cytarabine and GDC-0980 conjugates were synthesized, respectively. In vitro studies demonstrated that both conjugates had potent cytotoxicity and their combination also showed strong synergy, suggesting a potential chemotherapeutic strategy for future AML treatment.
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sequential combination therapy of ovarian cancer with degradable n 2 hydroxypropyl Methacrylamide copolymer paclitaxel and gemcitabine conjugates
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Rui Zhang, Jiyuan Yang, Yan Zhou, Monika Sima, Jindřich KopecekAbstract:For rapid and effective clinical translation, polymer-based anticancer therapeutics need long circulating conjugates that produce a sustained concentration gradient between the vasculature and solid tumor. To this end, we designed second-generation backbone-degradable diblock N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer carriers and evaluated sequential combination therapy of HPMA copolymer-paclitaxel and HPMA copolymer-gemcitabine conjugates against A2780 human ovarian carcinoma xenografts. First, extensive in vitro assessment of administration sequence impact on cell cycle, viability, apoptosis, migration, and invasion revealed that treatment with paclitaxel conjugate followed by gemcitabine conjugate was the most effective scheduling strategy. Second, in an in vivo comparison with first-generation (nondegradable, molecular weight below the renal threshold) conjugates and free drugs, the second-generation degradable high-molecular weight conjugates showed distinct advantages, such as favorable pharmacokinetics (three- to five-times half-life compared with the first generation), dramatically enhanced inhibition of tumor growth (complete tumor regression) by paclitaxel and gemcitabine conjugate combination, and absence of adverse effects. In addition, multimodality imaging studies of dual-labeled model conjugates confirmed the efficacy of second-generation conjugates by visualizing more than five-times enhanced tumor accumulation, rapid conjugate internalization, and effective intracellular release of payload. Taken together, the results indicate that the second-generation degradable HPMA copolymer carrier can provide an ideal platform for the delivery of a range of antitumor compounds, which makes it one of the most attractive candidates for potential clinical application.
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synthesis and characterization of poly e caprolactone block poly n 2 hydroxypropyl Methacrylamide micelles for drug delivery
Macromolecular Bioscience, 2011Co-Authors: S G Krimmer, Huaizhong Pan, Jihua Liu, Jiyuan Yang, Jindřich KopecekAbstract:Amphiphilic block copolymers based on HPMA and e-CL were synthesized by ring-opening polymerization of e-CL followed by RAFT polymerization of HPMA. A copolymer composed of 34 kDa PHPMA and 8.5 kDa PCL associated into micelles with CMC of 5.4 μg · mL−1. A novel retinoid, 3-Cl-AHPC-OMe, was incorporated into micelles with 25 wt.-% loading by dialysis method. The effective diameter of drug loading micelles was 117 nm. Incubation of micelles in PBS at 37 °C indicated 86 wt.-% of the drug was released after 96 h. Cytotoxicity studies performed with C4-2 prostate cancer cells showed the IC50 dose was 1.96 μM after 72 h of incubation, whereas the micelles without drug showed no cytotoxicity. Keywords: block copolymers, drug delivery systems, micelles, reversible addition fragmentation chain transfer (RAFT) Introduction Polymeric micelles are some of the most studied anticancer drug delivery systems.[1] They self-assemble from amphiphilic block copolymers in aqueous solution into nano-aggregates composed of a hydrophobic core and a hydrophilic shell.[2–4] Their advantages include simple preparation, high drug loading capacity, suitability for loading hydrophobic drugs, and accumulation in solid tumors due to enhanced permeability and retention (EPR) effect. Their main challenges are colloidal stability and an appropriate drug release profile.[1] The micelle core is formed by the hydrophobic block. Its chemical structure and molecular weight are important factors that determine the stability, drug loading capacity and drug release profile of micelles.[2] Polyesters, such as poly(e-caprolactone) (PCL)[5,6] and polylactide,[7] polyamino acids like poly(aspartic acid) and poly(β-benzyl-L-aspartate),[8] and poly(propylene oxide)[9] have been widely used as hydrophobic blocks. Poly(ethylene glycol) (PEG) has been the most frequently used hydrophilic shell-forming block,[3] while poly(N-vinyl-2-pyrrolidone) has also been repeatedly used.[10] PEG provides steric stabilization and minimization of non-specific protein interactions.[11] It is used in several products that have been approved by the Federal Drug Administration (FDA).[12] However, the lack of PEG functionality, the accelerated blood clearance (ABC) effect of PEGylated liposomes,[13] and the detection of anti-PEG antibodies[14] encouraged search for PEG alternatives. Poly[N-(2-hydroxypropyl)Methacrylamide] (PHPMA) is a hydrophilic, biocompatible and non-immunogenic water-soluble polymer intensively studied as drug carrier.[15] Like PEG, PHPMA modification has shown to increase blood-circulation times. Examples include semitelechelic PHPMA-modified nanospheres[16] and acetylcholinesterase modified at multiple points with PHPMA.[17] Recently, various micelles were designed with hydrophilic shells formed by PHPMA.[18–24] Interestingly, Hennink’s laboratory used lactate modified PHPMA as the hydrophobic block and PEG as the hydrophilic block to prepare thermosensitive, biodegradable micelles.[25] The multifunctionality of PHPMA demonstrated advantages in the design of different architectures.[2] Lele and Leroux reported the synthesis of amphiphilic triblock[19] and star-shaped[20] copolymers containing PHPMA as a hydrophilic block and PCL, a biodegradable polymer, as the hydrophobic block. Doxorubicin and amphotericin B were loaded into the triblock based micelles (1–4 wt.-%), whereas indomethacin was incorporated into the star-shaped micelles (5–12 wt.-%). Researchers[20,21] used traditional free-radical polymerization to prepare PHPMA blocks. Consequently, the polydispersity of the PHPMA block will be >1.5 and might affect the control of size distribution of the final micelles. In this work, we explore the potential of a novel micellar drug carrier system based on a diblock copolymer of PCL and PHPMA. The block copolymer was synthesized by a combination of ring-opening polymerization (ROP) of e-caprolactone (e-CL) followed by “living” reversible addition-fragmentation chain transfer (RAFT) polymerization of HPMA. Using this approach, the molecular weight and polydispersity can be controlled. Polymeric micelles were formed by self-assembly of copolymers forming a hydrophobic PCL core and a hydrophilic PHPMA shell. To evaluate the feasibility of the micelles for drug delivery, a hydrophobic drug, (E)-4-[3-(1-adamantyl)-4-hydroxyphenyl]-3-chlorocinnamic methyl ester (3-Cl-AHPC-OMe; not used in polymeric micelle-based drug delivery systems before), was chosen and physically incorporated into the hydrophobic micelle core. The micelles were characterized by pyrene assay, dynamic light scattering (DLS), transmission electron microscopy (TEM), and stability measurements. In vitro cytotoxicity of the 3-Cl-AHPC-OMe delivery system was evaluated on the C4-2 prostate cancer cell line.
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biodegradable multiblock poly n 2 hydroxypropyl Methacrylamide via reversible addition fragmentation chain transfer polymerization and click chemistry
Macromolecules, 2011Co-Authors: Kui Luo, Jiyuan Yang, Pavla Kopeckova, Jindřich KopecekAbstract:A new bifunctional chain transfer agent (CTA) containing alkyne end groups was designed, synthesized, and used for direct synthesis of clickable telechelic polymers. Good control of reversible addition−fragmentation chain transfer (RAFT) polymerization of N-(2-hydroxypropyl)Methacrylamide (HPMA) was achieved by using the new CTA, as indicated by a linear increase of number-average molecular weight (Mn) with conversion and low polydispersity (PDI) (<1.1). In particular, enzymatically degradable multiblock HPMA polymers were readily prepared by subsequent reaction with α,ω-diazido-oligopeptide (GFLG) sequence via CuI-catalyzed alkyne−azide cycloaddition. Upon exposure of high molecular weight fractions of multiblock polyHPMA to papain or cathepsin B, the polymer was degraded into segments of molecular weight and narrow polydispersity similar to those of the initial telechelic polyHPMA.
Karel Ulbrich - One of the best experts on this subject based on the ideXlab platform.
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photodynamic therapy and imaging based on tumor targeted nanoprobe polymer conjugated zinc protoporphyrin
Future Science OA, 2015Co-Authors: Jun Fang, Karel Ulbrich, Long Liao, Hongzhuan Yin, Hideaki Nakamura, Vladimir Subr, Hiroshi MaedaAbstract:Aim: To evaluate the potential of tumor-targeted nanoprobe, N-(2-hydroxypropyl)Methacrylamide copolymer-conjugated zinc protoporphyrin (PZP) for photodynamic therapy (PDT) and tumor imaging. Materials & Methods: Different tumor models including carcinogen-induced cancer were used, PZP was intravenously injected followed by irradiation with xenon or blue fluorescent light on tumor. Results: One PZP 20 mg/kg (ZnPP equivalent) dose with two or three treatments of light at an intensity of ≥20 J/cm2 caused necrosis and disappearance of most tumors (>70%) in different tumor models. We also confirmed PZP-based tumor imaging in carcinogen-induced breast tumor and colon cancer models. Conclusion: These findings support the potential application of PZP as a tumor-selective nanoprobe for PDT as well as tumor imaging, by virtue of the enhanced permeability and retention effect.
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synthesis of poly n 2 hydroxypropyl Methacrylamide conjugates of inhibitors of the abc transporter that overcome multidrug resistance in doxorubicin resistant p388 cells in vitro
Biomacromolecules, 2014Co-Authors: Vladimir Subr, Karel Ulbrich, Jan Strohalm, Blanka řihova, L Sivak, Eva Koziolova, Alena Braunova, Michal Pechar, Martina Kabesova, Marek KovařAbstract:The effects of novel polymeric therapeutics based on water-soluble N-(2-hydroxypropyl)Methacrylamide copolymers (P(HPMA)) bearing the anticancer drug doxorubicin (Dox), an inhibitor of ABC transporters, or both, on the viability and the proliferation of the murine monocytic leukemia cell line P388 (parental cell line) and its doxorubicin-resistant subline P388/MDR were studied in vitro. The inhibitor derivatives 5-methyl-4-oxohexanoyl reversin 121 (MeOHe-R121) and 5-methyl-4-oxohexanoyl ritonavir ester (MeOHe-RIT), showing the highest inhibitory activities, were conjugated to the P(HPMA) via the biodegradable pH-sensitive hydrazone bond, and the ability of these conjugates to block the ATP driven P-glycoprotein (P-gp) efflux pump was tested. The P(HPMA) conjugate P-Ahx-NH-N═MeOHe-R121 showed a dose-dependent increase in the ability to sensitize the P388/MDR cells to Dox from 1.5 to 24 μM, and achieved an approximately 50-fold increase in sensitization at 24 μM. The P(HPMA) conjugate P-Ahx-NH-N═MeOHe-RIT s...
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corrigendum to two step mechanisms of tumor selective delivery of n 2 hydroxypropyl Methacrylamide copolymer conjugated with pirarubicin via an acid cleavable linkage j control release 174 2014 81 87
Journal of Controlled Release, 2014Co-Authors: Hideaki Nakamura, Karel Ulbrich, Jun Fang, Tomas Etrych, Petr Chytil, Manami Ohkubo, Hiroshi MaedaAbstract:Corrigendum to “Two step mechanisms of tumor selective delivery of N-(2-hydroxypropyl) Methacrylamide copolymer conjugated with pirarubicin via an acid-cleavable linkage” [ J. Control. Release. 174 (2014) 81–87] Hideaki Nakamura , Tomas Etrych , Petr Chytil , Manami Ohkubo , Jun Fang , Karel Ulbrich , Hiroshi Maeda b,⁎ a Laboratory of Microbiology and Oncology, Faculty of Pharmaceutical Sciences, Sojo University, Ikeda 4-22-1, Nishi-ku, Kumamoto 860-0082, Japan b Research Institute for Drug Delivery Science, Faculty of Pharmaceutical Sciences, Sojo University, Ikeda 4-22-1, Nishi-ku, Kumamoto 860-0082, Japan c Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic, Heyrovsky Sq. 2, 162 06 Prague 6, Czech Republic
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two step mechanisms of tumor selective delivery of n 2 hydroxypropyl Methacrylamide copolymer conjugated with pirarubicin via an acid cleavable linkage
Journal of Controlled Release, 2014Co-Authors: Hideaki Nakamura, Karel Ulbrich, Jun Fang, Tomas Etrych, Petr Chytil, Manami Ohkubo, Hiroshi MaedaAbstract:N-(2-Hydroxypropyl)Methacrylamide copolymer containing hydrazide groups (PHPMA) conjugated with pirarubicin (THP) via a hydrazone bond (PHPMA-hyd-THP) is a drug conjugate that releases THP in the acidic milieu of a tumor. PHPMA-hyd-THP has an apparent Mw of 40,000 and a hydrodynamic diameter of 8.2±1.7nm but no apparent plasma protein binding. PHPMA-hyd-THP possesses two mechanisms of selectivity toward solid tumors and has potent antitumor action. The first one is drug accumulation in tumors that depends on the enhanced permeability and retention (EPR) effect, which results in a 4-20 times higher concentration of drug in the tumor than in normal tissues such as the heart, lung, and intestine. This accumulation in tumor tissue is in great contrast to that of conventional low-Mw THP. The second one is pH-dependent release of drug from PHPMA-hyd-THP: this conjugate released free THP more efficiently at a lower pH, which exists in tumors, and exerts cytotoxic activity. Free THP is known for its much faster uptake into tumor cells compared with doxorubicin. Thus, in our in vitro study, PHPMA-hyd-THP showed a higher cytotoxicity at the lower pH of tumor tissue than at the neutral pH of normal tissue. Furthermore, much more THP was liberated from the conjugate in acidic tumor tissue than in normal tissue. The EPR effect-dependent accumulation of PHPMA-hyd-THP and tumor-selective THP release in the tumor tissues led to highly tumor-selective drug accumulation, which continued for more than 72h, whereas the lowest free drug concentration was detected in normal tissues at 24h and no longer at a later time. In conclusion, we determined in our study here that the acid-cleavable PHPMA-hyd-THP conjugate had an excellent antitumor effect without appreciable adverse effects.
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micelles of zinc protoporphyrin conjugated to n 2 hydroxypropyl Methacrylamide hpma copolymer for imaging and light induced antitumor effects in vivo
Journal of Controlled Release, 2013Co-Authors: Hideaki Nakamura, Karel Ulbrich, Jun Fang, Long Liao, Vladimir Subr, Yuki Hitaka, Kenji Tsukigawa, Hiroshi MaedaAbstract:We synthesized N-(2-hydroxypropyl)Methacrylamide polymer conjugated with zinc protoporphyrin (HPMA-ZnPP) and evaluated its application for tumor detection by imaging and treatment by light exposure using in mouse sarcoma model. To characterize HPMA-ZnPP micelle, we measured its micellar size, surface charge, stability, photochemical, biochemical properties and tissue distribution. In vivo anti-tumor effect and fluorescence imaging were carried out to validate the tumor selective accumulation and therapeutic effect by inducing singlet oxygen by light exposure. HPMA-ZnPP was highly water soluble and formed micelles spontaneously having hydrophobic clustered head group of ZnPP, in aqueous solution, with a hydrodynamic diameter of 82.8±41.8 nm and zeta-potential of +1.12 mV. HPMA-ZnPP had a long plasma half-life and effectively and selectively accumulated in tumors. Although HPMA-ZnPP alone had no toxicity in S-180 tumor-bearing mice, light-irradiation significantly suppressed tumor growth in vivo, similar to the cytotoxicity to HeLa cells in vitro upon endoscopic light-irradiation. HPMA-ZnPP can visualize tumors by fluorescence after i.v. injection, which suggests that this micelle may be useful for both tumor imaging and therapy. Here we describe preparation of a new fluorescence nanoprobe that is useful for simultaneous tumor imaging and treatment, and application to fluorescence endoscopy is now at visible distance.
Ruth Duncan - One of the best experts on this subject based on the ideXlab platform.
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Cell uptake and trafficking behavior of non-covalent, coiled-coil based polymer-drug conjugates.
Macromolecular rapid communications, 2010Co-Authors: Bojana Apostolovic, Ruth Duncan, Samuel P. E. Deacon, Harm-anton KlokAbstract:This paper reports on the cell uptake and trafficking properties of a series of non-covalent polymer-drug conjugates. These nanomedicines are composed of a poly(N-(2-hydroxypropyl)Methacrylamide) backbone functionalized with multiple copies of a drug. The drug moieties are attached to the polymer via a non-covalent, so called coiled coil motif, which is formed by heterodimerization of two complementary peptide strands, one of which is attached to the polymer carrier and the other to the drug. Cytotoxicity and FACS experiments, which were carried out with model anticancer drug or fluorophore conjugates, provided insight into the cell uptake and trafficking behavior of these conjugates.
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n 2 hydroxypropyl Methacrylamide copolymer 6 3 aminopropyl ellipticine conjugates synthesis in vitro and preliminary in vivo evaluation
Bioconjugate Chemistry, 2001Co-Authors: F Searle, S Gacbreton, R Keane, S Dimitrijevic, Steve Brocchini, E A Sausville, Ruth DuncanAbstract:Ellipticine derivatives have potential as anticancer drugs. Their clinical use has been limited, however, by poor solubility and host toxicity. As N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer-anticancer conjugates are showing promise in early clinical trials, a series of novel HPMA copolymer conjugates have been prepared containing the 6-(3-aminopropyl)-ellipticine derivative (APE, NSC176328). Drug was linked to the polymer via GFLG or GG peptide side chains. To optimize biological behavior, HPMA copolymer-GFLG-APE conjugates with different drug loading (total APE: 2.3-7% w/w; free APE: 10-fold). HPMA copolymer-GG-APE did not liberate drug in the presence of isolated lysosomal enzymes (tritosomes), but HPMA copolymer-GFLG-APE released APE to a maximum of 60% after 5 h. The rate of drug release was influenced by drug loading; lower loading led to greater release. Whereas free APE (35 microg/mL) caused significant hemolysis (50% after 1 h), HPMA copolymer-APE conjugates were not hemolytic up to 300 microg/mL (APE-equiv). As would be expected from its cellular pharmacokinetics, HPMA copolymer-GFLG-APE was >75 times less cytotoxic than free drug (IC(50) approximately 0.4 microg/mL) against B16F10 melanoma in vitro. However, in vivo when tested in mice bearing s.c. B16F10 melanoma, HPMA copolymer-GFLG-APE (1-10 mg/kg single dose, APE-equiv) given i.p. was somewhat more active (highest T/C value of 143%) than free APE (1 mg/kg) (T/C =127%). HPMA copolymer-APE conjugates warrant further evaluation as potential anticancer agents.
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phase i clinical and pharmacokinetic study of pk1 n 2 hydroxypropyl Methacrylamide copolymer doxorubicin first member of a new class of chemotherapeutic agents drug polymer conjugates
Clinical Cancer Research, 1999Co-Authors: P Vasey, Ruth Duncan, S B Kaye, R Morrison, Chris Twelves, Peter H Wilson, Alison H Thomson, L S Murray, T E Hilditch, T MurrayAbstract:PK1 comprises doxorubicin covalently bound to N -(2-hydroxypropyl)Methacrylamide copolymer by a peptidyl linker. Following cellular uptake via pinocytosis, the linker is cleaved by lysosomal enzymes, allowing intratumoral drug release. Radically altered plasma and tumor pharmacokinetics, compared to free doxorubicin, and significant activity in animal tumors have been demonstrated preclinically. We aimed to determine the maximum tolerated dose, toxicity profile, and pharmacokinetics of PK1 as an i.v. infusion every 3 weeks to patients with refractory or resistant cancers. Altogether, 100 cycles were administered (range, 20–320 mg/m 2 doxorubicin-equivalent) to 36 patients (20 males and 16 females) with a mean age of 58.3 years (age range, 34–72 years). The maximum tolerated dose was 320 mg/m 2 , and the dose-limiting toxicities were febrile neutropenia and mucositis. No congestive cardiac failure was seen despite individual cumulative doses up to 1680 mg/m 2 . Other anthracycline-like toxicities were attenuated. Pharmacokinetically, PK1 has a distribution t 1/2 of 1.8 h and an elimination t 1/2 averaging 93 h. 131 I-labeled PK1 imaging suggests PK1 is taken up by some tumors. Responses (two partial and two minor responses) were seen in four patients with NSCLC, colorectal cancer, and anthracycline-resistant breast cancer. PK1 demonstrated antitumor activity in refractory cancers, no polymer-related toxicity, and proof of principle that polymer-drug conjugation decreases doxorubicin dose-limiting toxicities. The recommended Phase II dose is 280 mg/m 2 every 3 weeks. Studies are planned in colorectal, NSCLC, and breast cancer patients.
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niosomes containing n 2 hydroxypropyl Methacrylamide copolymer doxorubicin pk1 effect of method of preparation and choice of surfactant on niosome characteristics and a preliminary study of body distribution
International Journal of Pharmaceutics, 1997Co-Authors: Ijeoma F Uchegbu, Ruth DuncanAbstract:Abstract PK1 is an N -(2-hydroxypropyl)Methacrylamide (HPMA) copolymer-doxorubicin conjugate currently in early clinical development. Niosome encapsulation is a means to increase PK1 blood residence time, potentially promote tumour uptake and produce a slow, sustained release of active drug. Factors effecting encapsulation efficiency and size of PK1-niosome formulations were studied. Five surfactants were used to prepare PK1-niosomes; hexadecyl poly-5-oxyethylene ether (C 16 EO 5 ); octadecyl poly-5-oxyethylene ether (C 18 EO 5 ); hexadecyl diglycerol ether (C 16 G 2 ); sorbitan monopalmitate (Span 40) and sorbitan monostearate (Span 60). All were mixed in equimolar ratio with cholesterol and varying amounts of Solulan C24 (a cholesteryl poly-24-oxyethylene ether) (9–39 mol%). Dicetylphosphate (DCP) was also added (2 mol%). Passive association of PK1 with preformed C 16 G 2 and Span 60 vesicles was low (3–4%) while subsequent dehydration (freeze drying) followed by rehydration of the formulation increased the entrapment to 61% in the C 16 G 2 formulation. Transmission electron microscopy revealed that these niosomes had an electron dense core, evidence of intravesicular concentration of PK1. Increasing Solulan C24 content resulted in decreased PK1 entrapment after freeze drying, and the vesicle size was also decreased. Solulan C24 (39 mol%) caused pronounced vesicle aggregation on freeze drying, whereas at lower levels (9 mol%), PK1 appeared to act as a cryoptrotectant and the mean size of C 16 G 2 niosomes was 235 nm. A PK1:surfactant/lipid ratio of 0.3 (11.2 mg ml −1 doxorubicin) was achieved with Span 60 niosomes. This formulation, and the C 16 G 2 niosomes, did not induce red blood cell lysis at the proposed dose for in vivo use. Preliminary in vivo biodistribution studies showed PK1-C 16 G 2 niosomes to be mainly taken up by the liver and spleen. After 24 h, 25 and 3% of dose administered was present as free doxorubicin in these organs respectively.
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gamma scintigraphy of the biodistribution of 123i labelled n 2 hydroxypropyl Methacrylamide copolymer doxorubicin conjugates in mice with transplanted melanoma and mammary carcinoma
Journal of Drug Targeting, 1996Co-Authors: M V Pimm, Karel Ulbrich, A C Perkins, Jiri Strohalm, Ruth DuncanAbstract:AbstractAn N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer-doxorubicin conjugate is currently under clinical evaluation as a new antitumour agent. It has been shown previously that such conjugates exhibit selective tumour accumulation. In this study HPMA copolymer doxorubicin conjugates of low (LMW) or high (HMW) molecular weight were synthesised (which had a weight average molecular weight (Mw) of 25,000 and 94,000 respectively) and additionally contained a small amount (1 mol%) of the comonomer methacryloyltyrosinamide to permit labelling with [123I or 125I]iodide. Gamma camera imaging using the I-labelled probes was used to follow time-dependent biodistribution after intraperitoneal (i.p.) or intravenous (i.v.) administration to mice bearing subcutaneously either B16F10 melanoma or a mammary carcinoma. Imaging showed more rapid clearance of LMW conjugate from the peritoneal cavity than HMW conjugate. The images of mice given the LMW conjugate revealed rapid urinary excretion of radioactivity after b...
Jiyuan Yang - One of the best experts on this subject based on the ideXlab platform.
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backbone degradable n 2 hydroxypropyl Methacrylamide copolymer conjugates with gemcitabine and paclitaxel impact of molecular weight on activity toward human ovarian carcinoma xenografts
Molecular Pharmaceutics, 2017Co-Authors: Jiyuan Yang, Huaizhong Pan, Rui Zhang, Yixin Fang, Libin Zhang, Jindřich KopecekAbstract:Degradable diblock and multiblock (tetrablock and hexablock) N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer-gemcitabine (GEM) and -paclitaxel (PTX) conjugates were synthesized by reversible addition-fragmentation chain-transter (RAFT) copolymerization followed by click reaction for preclinical investigation. The aim was to validate the hypothesis that long-circulating conjugates are needed to generate a sustained concentration gradient between vasculature and a solid tumor and result in significant anticancer effect. To evaluate the impact of molecular weight of the conjugates on treatment efficacy, diblock, tetrablock, and hexablock GEM and PTX conjugates were administered intravenously to nude mice bearing A2780 human ovarian xenografts. For GEM conjugates, triple doses with dosage 5 mg/kg were given on days 0, 7, and 14 (q7dx3), whereas a single dose regime with 20 mg/kg was applied on day 0 for PTX conjugates treatment. The most effective conjugates for each monotherapy were the diblock ones, 2P-GEM and 2P-PTX (Mw ≈ 100 kDa). Increasing the Mw to 200 or 300 kDa resulted in decrease of activity most probably due to changes in the conformation of the macromolecule because of interaction of hydrophobic residues at side chain termini and formation of "unimer micelles". In addition to monotherapy, a sequential combination treatment of diblock PTX conjugate followed by GEM conjugate (2P-PTX/2P-GEM) was also performed, which showed the best tumor growth inhibition due to synergistic effect: complete remission was achieved after the first treatment cycle. However, because of low dose applied, tumor recurrence was observed 2 weeks after cease of treatment. To assess optimal route of administration, intraperitoneal (i.p.) application of 2P-GEM, 2P-PTX, and their combination was examined. The fact that the highest anticancer efficiency was achieved with diblock conjugates that can be synthesized in one scalable step bodes well for the translation into clinics.
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n 2 hydroxypropyl Methacrylamide copolymer drug conjugates for combination chemotherapy of acute myeloid leukemia
Macromolecular Bioscience, 2016Co-Authors: Rui Zhang, Jiyuan Yang, Yan Zhou, Paul J Shami, Jindřich KopecekAbstract:There is a need for new treatment strategies of acute myeloid leukemia (AML). In this study, four different drugs, including cytarabine, daunorubicin, GDC-0980, and JS-K, were investigated in vitro for the two-drug combinations treatment of AML. The results revealed that cytarabine and GDC-0980 had the strongest synergism. In addition, cell cycle analysis was conducted to investigate the effect of the different combinations on cell division. For future in vivo application, N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer-cytarabine and GDC-0980 conjugates were synthesized, respectively. In vitro studies demonstrated that both conjugates had potent cytotoxicity and their combination also showed strong synergy, suggesting a potential chemotherapeutic strategy for future AML treatment.
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sequential combination therapy of ovarian cancer with degradable n 2 hydroxypropyl Methacrylamide copolymer paclitaxel and gemcitabine conjugates
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Rui Zhang, Jiyuan Yang, Yan Zhou, Monika Sima, Jindřich KopecekAbstract:For rapid and effective clinical translation, polymer-based anticancer therapeutics need long circulating conjugates that produce a sustained concentration gradient between the vasculature and solid tumor. To this end, we designed second-generation backbone-degradable diblock N-(2-hydroxypropyl)Methacrylamide (HPMA) copolymer carriers and evaluated sequential combination therapy of HPMA copolymer-paclitaxel and HPMA copolymer-gemcitabine conjugates against A2780 human ovarian carcinoma xenografts. First, extensive in vitro assessment of administration sequence impact on cell cycle, viability, apoptosis, migration, and invasion revealed that treatment with paclitaxel conjugate followed by gemcitabine conjugate was the most effective scheduling strategy. Second, in an in vivo comparison with first-generation (nondegradable, molecular weight below the renal threshold) conjugates and free drugs, the second-generation degradable high-molecular weight conjugates showed distinct advantages, such as favorable pharmacokinetics (three- to five-times half-life compared with the first generation), dramatically enhanced inhibition of tumor growth (complete tumor regression) by paclitaxel and gemcitabine conjugate combination, and absence of adverse effects. In addition, multimodality imaging studies of dual-labeled model conjugates confirmed the efficacy of second-generation conjugates by visualizing more than five-times enhanced tumor accumulation, rapid conjugate internalization, and effective intracellular release of payload. Taken together, the results indicate that the second-generation degradable HPMA copolymer carrier can provide an ideal platform for the delivery of a range of antitumor compounds, which makes it one of the most attractive candidates for potential clinical application.
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biodegradable multiblock poly n 2 hydroxypropyl Methacrylamide gemcitabine and paclitaxel conjugates for ovarian cancer cell combination treatment
International Journal of Pharmaceutics, 2013Co-Authors: Nate Larson, Hamidreza Ghandehari, Jiyuan Yang, Abhijit Ray, Darwin L Cheney, Jindrich KopecekAbstract:The synthesis, characterization, and in vitro evaluation of a combination delivery of multiblock poly(N-2-Hydroxypropyl)Methacrylamide (HPMA), gemcitabine (GEM) and paclitaxel (PTX) conjugates is described in this study. Multiblock copolymer conjugates of a large molecular weight (Mw > 200 kDa) were studied and compared to traditional, small molecular weight (Mw < 45 kDa) conjugates. Stability of the conjugates in different pH was assessed, and their cytotoxicity in combination toward A2780 human ovarian cancer cells was evaluated by combination index analysis. Treatment duration (4 and 72 h) and sequence of addition were explored. In addition, an HPMA copolymer conjugate with both GEM and PTX in the side chains was evaluated in a similar manner and compared to a physical mixture of individual conjugates. Conjugates with narrow molecular weight distribution (Mw/Mn < 1.1) were obtained via RAFT polymerization, and drug loadings of between 5.5 and 9.2 wt% were achieved. Conjugates demonstrated moderate stability with less than 65% release over 24 h at pH 7.4, and near complete drug release in the presence of the lysosomal enzyme cathepsin B in 3 h. In combination, the cytotoxic effects of a mixture of the conjugates were primarily additive. Synergistic effects were observed when A2780 human ovarian cancer cells were treated simultaneously for 4 h with multiblock conjugates (CI < 0.7). When both GEM and PTX were conjugated to the same copolymer backbone, moderate antagonism (CI 1.3–1.6) was observed. These results demonstrate that multiblock HPMA copolymer–GEM and –PTX conjugates, when delivered as a mixture of individual agents, are promising for the treatment of ovarian cancer.
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synthesis and characterization of poly e caprolactone block poly n 2 hydroxypropyl Methacrylamide micelles for drug delivery
Macromolecular Bioscience, 2011Co-Authors: S G Krimmer, Huaizhong Pan, Jihua Liu, Jiyuan Yang, Jindřich KopecekAbstract:Amphiphilic block copolymers based on HPMA and e-CL were synthesized by ring-opening polymerization of e-CL followed by RAFT polymerization of HPMA. A copolymer composed of 34 kDa PHPMA and 8.5 kDa PCL associated into micelles with CMC of 5.4 μg · mL−1. A novel retinoid, 3-Cl-AHPC-OMe, was incorporated into micelles with 25 wt.-% loading by dialysis method. The effective diameter of drug loading micelles was 117 nm. Incubation of micelles in PBS at 37 °C indicated 86 wt.-% of the drug was released after 96 h. Cytotoxicity studies performed with C4-2 prostate cancer cells showed the IC50 dose was 1.96 μM after 72 h of incubation, whereas the micelles without drug showed no cytotoxicity. Keywords: block copolymers, drug delivery systems, micelles, reversible addition fragmentation chain transfer (RAFT) Introduction Polymeric micelles are some of the most studied anticancer drug delivery systems.[1] They self-assemble from amphiphilic block copolymers in aqueous solution into nano-aggregates composed of a hydrophobic core and a hydrophilic shell.[2–4] Their advantages include simple preparation, high drug loading capacity, suitability for loading hydrophobic drugs, and accumulation in solid tumors due to enhanced permeability and retention (EPR) effect. Their main challenges are colloidal stability and an appropriate drug release profile.[1] The micelle core is formed by the hydrophobic block. Its chemical structure and molecular weight are important factors that determine the stability, drug loading capacity and drug release profile of micelles.[2] Polyesters, such as poly(e-caprolactone) (PCL)[5,6] and polylactide,[7] polyamino acids like poly(aspartic acid) and poly(β-benzyl-L-aspartate),[8] and poly(propylene oxide)[9] have been widely used as hydrophobic blocks. Poly(ethylene glycol) (PEG) has been the most frequently used hydrophilic shell-forming block,[3] while poly(N-vinyl-2-pyrrolidone) has also been repeatedly used.[10] PEG provides steric stabilization and minimization of non-specific protein interactions.[11] It is used in several products that have been approved by the Federal Drug Administration (FDA).[12] However, the lack of PEG functionality, the accelerated blood clearance (ABC) effect of PEGylated liposomes,[13] and the detection of anti-PEG antibodies[14] encouraged search for PEG alternatives. Poly[N-(2-hydroxypropyl)Methacrylamide] (PHPMA) is a hydrophilic, biocompatible and non-immunogenic water-soluble polymer intensively studied as drug carrier.[15] Like PEG, PHPMA modification has shown to increase blood-circulation times. Examples include semitelechelic PHPMA-modified nanospheres[16] and acetylcholinesterase modified at multiple points with PHPMA.[17] Recently, various micelles were designed with hydrophilic shells formed by PHPMA.[18–24] Interestingly, Hennink’s laboratory used lactate modified PHPMA as the hydrophobic block and PEG as the hydrophilic block to prepare thermosensitive, biodegradable micelles.[25] The multifunctionality of PHPMA demonstrated advantages in the design of different architectures.[2] Lele and Leroux reported the synthesis of amphiphilic triblock[19] and star-shaped[20] copolymers containing PHPMA as a hydrophilic block and PCL, a biodegradable polymer, as the hydrophobic block. Doxorubicin and amphotericin B were loaded into the triblock based micelles (1–4 wt.-%), whereas indomethacin was incorporated into the star-shaped micelles (5–12 wt.-%). Researchers[20,21] used traditional free-radical polymerization to prepare PHPMA blocks. Consequently, the polydispersity of the PHPMA block will be >1.5 and might affect the control of size distribution of the final micelles. In this work, we explore the potential of a novel micellar drug carrier system based on a diblock copolymer of PCL and PHPMA. The block copolymer was synthesized by a combination of ring-opening polymerization (ROP) of e-caprolactone (e-CL) followed by “living” reversible addition-fragmentation chain transfer (RAFT) polymerization of HPMA. Using this approach, the molecular weight and polydispersity can be controlled. Polymeric micelles were formed by self-assembly of copolymers forming a hydrophobic PCL core and a hydrophilic PHPMA shell. To evaluate the feasibility of the micelles for drug delivery, a hydrophobic drug, (E)-4-[3-(1-adamantyl)-4-hydroxyphenyl]-3-chlorocinnamic methyl ester (3-Cl-AHPC-OMe; not used in polymeric micelle-based drug delivery systems before), was chosen and physically incorporated into the hydrophobic micelle core. The micelles were characterized by pyrene assay, dynamic light scattering (DLS), transmission electron microscopy (TEM), and stability measurements. In vitro cytotoxicity of the 3-Cl-AHPC-OMe delivery system was evaluated on the C4-2 prostate cancer cell line.
Wim E Hennink - One of the best experts on this subject based on the ideXlab platform.
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preclinical evaluation of thermosensitive poly n 2 hydroxypropyl Methacrylamide mono dilactate grafted liposomes for cancer thermochemotherapy
International Journal of Pharmaceutics, 2018Co-Authors: Merel Van Elk, Tina Vermonden, Wim E Hennink, Gert Storm, Joep Van Den Dikkenberg, Michal HegerAbstract:Thermosensitive liposomes grafted with cholesterol-conjugated poly(N-(2-hydroxypropyl) Methacrylamide mono/dilactate) (chol-pHPMAlac) have been developed for heat-induced release of doxorubicin (DOX). These liposomes release DOX completely during mild hyperthermia, but their interaction with blood cells and cancer cells has not been studied. Following intravenous administration, liposomes may interact with plasma proteins and various types of cells (e.g., endothelial cells, platelets, and macrophages), which would reduce their disposition in the tumor stroma. Interaction between liposomes and platelets may further cause platelet activation and thrombosis, which could lead to vascular occlusion and thromboembolic complications. The aim was to investigate DOX release kinetics in the presence of serum, stability, in vitro uptake by and toxicity to cancer cells and somatic cells, and platelet activating potential of the chol-pHPMAlac liposomes. DOX release was determined spectrofluorometrically. Liposome stability was determined in buffer and serum by dynamic light scattering and nanoparticle tracking analysis. Association with/uptake by and toxicity of empty liposomes to AML-12, HepG2 (both hepatocyte-derived cancer cells), RAW 264.7 (macrophages), and HUVEC (endothelial) cells was assayed in vitro. Platelet activation was determined by analysis of P-selectin expression and fibrinogen binding. DOPE:EPC liposomes (diameter = 135 nm) grafted with 5% chol-pHPMAlac (cloud point (CP) = 16 °C; Mn = 8.5 kDa) released less than 10% DOX at 37 °C in 30 min, whereas complete release took place at 47 °C or higher within 10 min. The size of these liposomes remained stable in buffer and serum during 24 h at 37 °C. Fluorescently labeled but DOX-lacking chol-pHPMAlac-liposomes exhibited poor association with/uptake by all cells under investigation, were not cytotoxic, and did not activate platelets in both buffered solution and whole blood. In conclusion, thermosensitive chol-pHPMAlac-grafted liposomes rapidly release DOX during mild hyperthermia. The liposomes are stable in a physiological milieu, are not taken up by cells that are encountered in an in vivo setting, and are non-antagonistic towards platelets. Chol-pHPMAlac-grafted liposomes are therefore good candidates for DOX delivery to tumors and temperature-triggered release in tumor stroma.
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triggered release of doxorubicin from temperature sensitive poly n 2 hydroxypropyl Methacrylamide mono dilactate grafted liposomes
Biomacromolecules, 2014Co-Authors: Merel Van Elk, Tina Vermonden, Roel Deckers, Chris Oerlemans, Yang Shi, Gerrit Storm, Wim E HenninkAbstract:The objective of this study was to design temperature-sensitive liposomes with tunable release characteristics that release their content at an elevated temperature generated by high intensity focused ultrasound (HIFU) exposure. To this end, thermosensitive polymers of N-(2-hydroxypropyl)Methacrylamide mono/dilactate of different molecular weights and composition with a cholesterol anchor (chol-pHPMAlac) were synthesized and grafted onto liposomes loaded with doxorubicin (DOX). The liposomes were incubated at different temperatures and their release kinetics were studied. A good correlation between the release-onset temperature of the liposomes and the cloud point (CP) of chol-pHPMAlac was found. However, release took place at significantly higher temperatures than the CP of chol-pHPMAlac, likely at the CP, the dehydration and thus hydrophobicity is insufficient to penetrate and permeabilize the liposomal membrane. Liposomes grafted with chol-pHPMAlac with a CP of 11.5 °C released 89% DOX within 5 min at 42 °C while for the liposomes grafted with a polymer with CP of 25.0 °C, a temperature of 52 °C was needed to obtain the same extent of DOX release. At a fixed copolymer composition, an increase in molecular weight from 6.5 to 14.5 kDa decreased the temperature at which DOX was released with a release-onset temperature from 52 to 42 °C. Liposomes grafted with 5% chol-pHPMAlac exhibited a rapid release to a temperature increase, while at a grafting density of 2 and 10%, the liposomes were less sensitive to an increase in temperature. Sequential release of DOX was obtained by mixing liposomes grafted with chol-pHPMAlac having different CPs. Chol-pHPMAlac grafted liposomes released DOX nearly quantitatively after pulsed wave HIFU. In conclusion, the release of DOX from liposomes grafted with thermosensitive polymers of N-(2-hydroxypropyl)Methacrylamide mono/dilactate can be tuned to the characteristics and the grafting density of chol-pHPMAlac, making these liposomes attractive for local drug delivery using hyperthermia.
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temperature sensitive poly n 2 hydroxypropyl Methacrylamide mono dilactate coated liposomes for triggered contents release
Bioconjugate Chemistry, 2007Co-Authors: Lauri Paasonen, Birgit Romberg, Gert Storm, Marjo Yliperttula, Arto Urtti, Wim E HenninkAbstract:We prepared thermosensitive poly(N-(2-hydroxypropyl)Methacrylamide mono/dilactate) (pHPMA mono/dilactate) polymer and studied temperature-triggered contents release from polymer-coated liposomes. HPMA mono/dilactate polymer was synthesized with a cholesterol anchor suitable for incorporation in the liposomal bilayers and with a cloud point (CP) temperature of the polymer slightly above normal body temperature (42 °C). Dynamic light scattering (DLS) measurements showed that whereas the size of noncoated liposomes remained stable upon raising the temperature from 25 to 46 °C, polymer-coated liposomes aggregated around 43 °C. Also, noncoated liposomes loaded with calcein showed hardly any leakage of the fluorescent marker when heated to 46 °C. However, polymer-coated liposomes showed a high degree of temperature-triggered calcein release above the CP of the polymer. Likely, liposome aggregation and bilayer destabilization are triggered because of the precipitation of the thermosensitive polymer above its CP ...
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poly n 2 hydroxypropyl Methacrylamide mono di lactate a new class of biodegradable polymers with tuneable thermosensitivity
Biomacromolecules, 2004Co-Authors: Osamu Soga, Cornelus F Van Nostrum, Wim E HenninkAbstract:A novel class of thermosensitive and biodegradable polymers, poly(N-(2-hydroxypropyl) Methacrylamide mono/di lactate) (poly(HPMAm-mono/di lactate)), was synthesized. The cloud points (CP) of poly(HPMAm-monolactate) and poly(HPMAm-dilactate) in water were 65 and 13 °C, respectively. The lower CP for poly(HPMAm-dilactate) is likely due the greater hydrophobicity of the dilactate side group over the monolactate side group. The CP of poly(HPMAm-monolactate-co-HPMAm-dilactate) increased linearly with mol % of HPMA-monolactate, which demonstrates that the CP is tuneable by the copolymer composition.
