P Kopecková

University of Utah, Salt Lake City, UT, United States

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Publications (151)709.56 Total impact

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    ABSTRACT: Abstract Combination of targeted delivery and controlled release is a powerful technique for cancer treatment. In this paper, we describe the design, synthesis, structure validation and biological properties of targeted and non-targeted N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-docetaxel conjugates. Docetaxel (DTX) was conjugated to HPMA copolymer via a tetrapeptide spacer (-GFLG-). 3-(1,3-dicarboxypropyl)-ureido]pentanedioic acid (DUPA) was used as the targeting moiety to actively deliver DTX for treatment of Prostate-Specific Membrane Antigen (PSMA) expressing prostate cancer. Short and long spacer DUPA monomers were prepared, and four HPMA copolymer - DTX conjugates (non-targeted, two targeted with short spacer of different molecular weight and targeted with long spacer) were prepared via Reversible Addition-Fragmentation Chain Transfer (RAFT) copolymerization. Following confirmation of PSMA expression on C4-2 cell line, the DTX conjugates' in vitro cytotoxicity was tested against C4-2 tumor cells and their anticancer efficacies were assessed in nude mice bearing s.c. human prostate adenocarcinoma C4-2 xenografts. The in vivo results show that the spacer length between targeting moieties and HPMA copolymer backbone can significantly affect the treatment efficacy of DTX conjugates against C4-2 tumor bearing nu/nu mice. Moreover, histological analysis indicated that the DUPA-targeted DTX conjugate with longer spacer had no toxicity in major organs of treated mice.
    Journal of Drug Targeting 12/2013; 21(10):968-80. · 2.77 Impact Factor
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    ABSTRACT: Multiblock, high molecular weight, linear, backbone degradable HPMA copolymer-prostaglandin E1 (PGE1) conjugate has been synthesized by RAFT polymerization mediated by a new bifunctional chain transfer agent (CTA), which contains an enzymatically degradable oligopeptide sequence flanked by two dithiobenzoate groups, followed by postpolymerization aminolysis and thiol-ene chain extension. The multiblock conjugate contains Asp8 as the bone targeting moiety and enzymatically degradable bonds in the polymer backbone; in vivo degradation produces cleavage products that are below the renal threshold. Using an ovariectomized (OVX) rat model, the accumulation in bone and efficacy to promote bone formation was evaluated; low molecular weight conjugates served as control. The results indicated a higher accumulation in bone, greater enhancement of bone density, and higher plasma osteocalcin levels for the backbone degradable conjugate.
    Biomaterials 05/2013; · 8.31 Impact Factor
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    ABSTRACT: Biodistribution, pharmacokinetics, and efficacy of prostate-cancer-targeted HPMA copolymer/DTX conjugates are evaluated in nude mice bearing prostate cancer C4-2 xenografts. PSMA-specific monoclonal antibodies 3F/11 are used as the targeting moiety. Control conjugates tumor accumulation to total background organs (heart, lung, kidney, liver, spleen and blood) accumulation increase substantially with time for the targeted conjugate, and the ratio at 48 h is 7-fold higher than that at 6 h. Preliminary evaluation of the efficacy of the conjugates in vivo show tumor growth inhibition for all HPMA copolymer/DTX conjugates.
    Macromolecular Bioscience 03/2012; 12(3):412-22. · 3.74 Impact Factor
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    ABSTRACT: High-molecular-weight, branched N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers were synthesized and conjugated with Fab' fragments of the anti-CD20 antibody, 1F5. This produced multivalent conjugates with varying valency (amount of Fab' per macromolecule) targeted to the B-cell antigen CD20. The apoptotic activity of the conjugates was screened against several B-cell lymphomas with varied expression levels of CD20 (Raji, Daudi, Ramos, Namalwa, and DG-75). The multivalent conjugates had the strongest activity against cells that had the highest expression of CD20 and failed to demonstrate any measurable activity against lymphomas that did not express the antigen. Furthermore, there was an apparent dose-dependent response to treatment with multivalent conjugates. At optimal valence and concentration, the apoptotic activity of HPMA copolymer-Fab' conjugates superseded that of free anti-CD20 Ab that was hyper-cross-linked with a polyclonal, secondary Ab.
    Biomacromolecules 02/2012; 13(3):727-35. · 5.37 Impact Factor
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    Kui Luo, Jiyuan Yang, Pavla Kopečková, Jindřich Kopeček
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    ABSTRACT: 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 αω, -diazido oligopeptide (GFLG) sequence via Cu(I) 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.
    Macromolecules 04/2011; 44(8):2481-2488. · 5.93 Impact Factor
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    ABSTRACT: Bone neoplasms, such as osteosarcoma, exhibit a propensity for systemic metastases resulting in adverse clinical outcome. Traditional treatment consisting of aggressive chemotherapy combined with surgical resection, has been the mainstay of these malignances. Therefore, bone-targeted non-toxic therapies are required. We previously conjugated the aminobisphosphonate alendronate (ALN), and the potent anti-angiogenic agent TNP-470 with N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer. HPMA copolymer-ALN-TNP-470 conjugate exhibited improved anti-angiogenic and anti-tumor activity compared with the combination of free ALN and TNP-470 when evaluated in a xenogeneic model of human osteosarcoma. The immune system has major effect on toxicology studies and on tumor progression. Therefore, in this manuscript we examined the safety and efficacy profiles of the conjugate using murine osteosarcoma syngeneic model. Toxicity and efficacy evaluation revealed superior anti-tumor activity and decreased organ-related toxicities of the conjugate compared with the combination of free ALN plus TNP-470. Finally, comparative anti-angiogenic activity and specificity studies, using surrogate biomarkers of circulating endothelial cells (CEC), highlighted the advantage of the conjugate over the free agents. The therapeutic platform described here may have clinical translational relevance for the treatment of bone-related angiogenesis-dependent malignances.
    Biomaterials 03/2011; 32(19):4450-63. · 8.31 Impact Factor
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    ABSTRACT: A new strategy for the synthesis of biodegradable high molecular weight N-(2-hydroxypropyl)methacrylamide (HPMA)-based polymeric carriers has been designed. An enzyme-sensitive, alkyne-functionalized, chain transfer agent (CTA-GFLG-alkyne; Nα-(4-pentynoyl)-Nδ-(4-cyano-4-(phenylcarbonothioylthio)pentanoyl–glycylphenylalanylleucylglycyl)-lysine) was synthesized and used to mediate the reversible addition–fragmentation chain transfer (RAFT) polymerization and copolymerization of HPMA. Post-polymerization modification with 4,4′-azobis(azidopropyl 4-cyanopentanoate) resulted in the formation of heterotelechelic HPMA copolymers containing terminal alkyne and azide groups. Chain extension via click reaction resulted in high molecular weight multiblock copolymers. Upon exposure to papain, these copolymers degraded into the initial blocks. Similar results were obtained for copolymers of HPMA with N-methacryloylglycylphenylalanylleucylglycyl thiazolidine-2-thione and N-methacryloylglycylphenylalanylleucylglycyl-gemcitabine. The new synthetic method presented permits the synthesis of biocompatible, biodegradable high molecular weight HPMA copolymer–anticancer drug conjugates that possess long-circulation times and augmented accumulation in solid tumor tissue due to the enhanced permeability and retention effect.
    Reactive and Functional Polymers 03/2011; 71(3):294-302. · 2.51 Impact Factor
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    ABSTRACT: Telechelic water-soluble HPMA copolymers and HPMA copolymer-doxorubicin (DOX) conjugates have been synthesized by RAFT polymerization mediated by a new bifunctional chain transfer agent (CTA) that contains an enzymatically degradable oligopeptide sequence. Postpolymerization aminolysis followed by chain extension with a bis-maleimide resulted in linear high molecular weight multiblock HPMA copolymer conjugates. These polymers are enzymatically degradable; in addition to releasing the drug (DOX), the degradation of the polymer backbone resulted in products with molecular weights similar to the starting material and below the renal threshold. The new multiblock HPMA copolymers hold potential as new carriers of anticancer drugs.
    Biomacromolecules 01/2011; 12(1):247-52. · 5.37 Impact Factor
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    ABSTRACT: Pulmonary edema and the associated increases in vascular permeability continue to represent a significant clinical problem in the intensive care setting, with no current treatment modality other than supportive care and mechanical ventilation. Therapeutic compound(s) capable of attenuating changes in vascular barrier function would represent a significant advance in critical care medicine. We have previously reported the development of HPMA-based copolymers, targeted to endothelial glycocalyx that are able to enhance barrier function. In this work, we report the refinement of copolymer design and extend our physiological studies to demonstrate that the polymers: 1) reduce both shear stress and pressure-mediated increase in hydraulic conductivity, 2) reduce nitric oxide production in response to elevated hydrostatic pressure and, 3) reduce the capillary filtration coefficient (K(fc)) in an isolated perfused mouse lung model. These copolymers represent an important tool for use in mechanotransduction research and a novel strategy for developing clinically useful copolymers for the treatment of vascular permeability.
    Biomaterials 10/2010; 32(1):288-94. · 8.31 Impact Factor
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    ABSTRACT: Endocytic uptake and subcellular trafficking of a large array of HPMA (N-(2-hydroxypropyl)methacrylamide) based copolymers possessing positively or negatively charged residues, or hydrophobic groups were evaluated by flow cytometry and living cell confocal microscopy in cultured prostate cancer cells. The degrees of cellular uptake of various copolymer fractions with narrow polydispersities were quantified. The copolymer charge was the predominant physicochemical feature in terms of cellular uptake. Fast and efficient uptake occurred in positively charged copolymers due to non-specific adsorptive endocytosis, whereas slow uptake of negatively charged copolymers was observed. The uptake of copolymers was also molecular weight dependent. The copolymers were internalized into the cells through multiple endocytic pathways: positively charged copolymers robustly engaged clathrin-mediated endocytosis, macropinocytosis and dynamin-dependent endocytosis, while weakly negatively charged copolymers weakly employed these pathways; strongly negatively charged copolymers only mobilized macropinocytosis. HPMA copolymer possessing 4 mol% of moderately hydrophobic functional groups did not show preferential uptake. All copolymers ultimately localized in late endosomes/lysosomes via early endosomes; with varying kinetics among the copolymers. This study indicates that cell entry and subsequent intracellular trafficking of polymeric drug carriers are strongly dependent on the physicochemical characteristics of the nanocarrier, such as charge and molecular weight.
    Journal of Controlled Release 04/2010; 143(1):71-9. · 7.63 Impact Factor
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    ABSTRACT: The 17-methoxy group of geldanamycin was substituted with 1,3-diaminopropane and 1,3-diamino-2-hydroxypropane to introduce a primary amino group useful for conjugation with targeting moieties and drug carriers. We have developed a procedure that has provided improved yield and reproducibility of the syntheses. Both geldanamycin derivatives demonstrated antiproliferative activity towards the human ovarian carcinoma cell line, A2780.
    ChemInform 01/2010; 32(45).
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    Jindrich Kopecek, Pavla Kopecková
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    ABSTRACT: The overview covers the discovery of N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers, initial studies on their synthesis, evaluation of biological properties, and explorations of their potential as carriers of biologically active compounds in general and anticancer drugs in particular. The focus is on the research in the authors' laboratory - the development of macromolecular therapeutics for the treatment of cancer and musculoskeletal diseases. In addition, the evaluation of HPMA (co)polymers as building blocks of modified and new biomaterials is presented: the utilization of semitelechelic poly(HPMA) and HPMA copolymers for the modification of biomaterial and protein surfaces and the design of hybrid block and graft HPMA copolymers that self-assemble into smart hydrogels. Finally, suggestions for the design of second-generation macromolecular therapeutics are portrayed.
    Advanced drug delivery reviews 11/2009; 62(2):122-49. · 11.96 Impact Factor
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    ABSTRACT: The antitumor activity of a colon-specific N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer - 9-aminocamptothecin (9-AC) conjugate (P-9-AC) was assessed in orthotopic and subcutaneous animal (HT29 xenograft) tumor models. P-9-AC treatment of mice bearing orthotopic colon tumors, with a dose of 3 mg/kg of 9-AC equivalent every other day for 6 weeks, resulted in regression of tumors in 9 of 10 mice. A lower dose of P-9-AC (1.25 mg/kg of 9-AC equivalent) every other day for 8 weeks inhibited subcutaneous tumor growth in all mice. No liver metastases were observed. Colon-specific release of 9-AC from polymer conjugates enhanced antitumor activity and minimized the systemic toxicity.
    Macromolecular Bioscience 09/2009; 9(11):1135-42. · 3.74 Impact Factor
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    ABSTRACT: Acute changes in lung capillary permeability continue to complicate procedures such as cardiopulmonary bypass, solid organ transplant, and major vascular surgery and precipitate the more severe disease state Adult Respiratory Distress Syndrome (ARDS). To date there is no treatment targeted directly to the lung microvasculature. We hypothesized that biomimetic polymers could be used to enhance passive barrier function by reducing the porosity of the endothelial glycocalyx and attenuate mechanotransduction by restricting the motion of the glycoproteins implicated in signal transduction. To this end, cationic copolymers containing methacrylamidopropyl trimethylammonium chloride (P-TMA Cl) have been developed as an infusible therapy to target the lung capillary glycocalyx in order to mechanically enhance the capillary barrier and turn off pressure-induced mechanotransduction. Copolymers were tested for functional efficacy by measuring both albumin permeability (P(DA)) and hydraulic conductivity (L(p)) across cultured endothelial monolayers. P-TMA Cl significantly decreased P(DA) in normal and inflamed cells and attenuated pressure-induced increases in L(p). Decreases in L(p) across endothelial monolayers in the presence of P-TMA Cl is evidence of a dampening of mechanotransduction-induced barrier dysfunction. We show the potential for biomimetic polymers targeted to lung endothelium as a viable therapy to enhance endothelial barrier function thereby attenuating a major component of vascular inflammation.
    Biomaterials 08/2009; 30(29):5885-91. · 8.31 Impact Factor
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    ABSTRACT: A new generation of antibodies against the prostate specific membrane antigen (PSMA) has been proven to bind specifically to PSMA molecules on the surface of living prostate cancer cells. To explore the potential of anti-PSMA antibodies as targeting moieties for macromolecular therapeutics for prostate cancer, fluorescently labeled HPMA (N-(2-hydroxypropyl)methacrylamide) copolymer-anti-PSMA antibody conjugates (P-anti-PSMA) were synthesized and the mechanisms of their endocytosis and subcellular trafficking in C4-2 prostate cancer cells were studied. Radioimmunoassays showed the dissociation constants of P-anti-PSMA for C4-2 prostate cancer cells in the low nanomolar range, close to values for free anti-PSMA. It indicated that conjugation of anti-PSMA to HPMA copolymers did not compromise their binding affinity. The rate of endocytosis of P-anti-PSMA was much faster than that of control HPMA copolymer conjugates containing nonspecific IgG. Selective pathway inhibitors of clathrin-mediated endocytosis and of macropinocytosis inhibited the internalization of P-anti-PSMA. Inhibition of clathrin-mediated endocytosis was further evidenced by down-regulation of clathrin heavy chain expression by siRNA. Using a dominant-negative mutant of dynamin (Dyn K44A) to abolish the clathrin-, caveolae-independent endocytic pathway, we found that some of P-anti-PSMA adopted this pathway to be endocytosed into C4-2 cells. Thus multiple receptor-mediated endocytic pathways, including clathrin-mediated endocytosis, macropinocytosis, and clathrin-, caveolae-independent endocytosis, were involved in the internalization of P-anti-PSMA. The extent of the participation of each pathway in P-anti-PSMA endocytosis was estimated. Membrane vesicles containing P-anti-PSMA rapidly colocalized with membrane vesicles overexpressing Rab7, a late endosome localized protein, demonstrating that a part of P-anti-PSMA was transported to late endosomes.
    Molecular Pharmaceutics 05/2009; 6(3):959-70. · 4.57 Impact Factor
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    ABSTRACT: BACKGROUND: There is an immense clinical need for novel therapeutics for the treatment of angiogenesis-dependent calcified neoplasms such as osteosarcomas and bone metastases. We developed a new therapeutic strategy to target bone metastases and calcified neoplasms using combined polymer-bound angiogenesis inhibitors. Using an advanced "living polymerization" technique, the reversible addition-fragmentation chain transfer (RAFT), we conjugated the aminobisphosphonate alendronate (ALN), and the potent anti-angiogenic agent TNP-470 with N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer through a Glycine-Glycine-Proline-Norleucine linker, cleaved by cathepsin K, a cysteine protease overexpressed at resorption sites in bone tissues. In this approach, dual targeting is achieved. Passive accumulation is possible due to the increase in molecular weight following polymer conjugation of the drugs, thus extravasating from the tumor leaky vessels and not from normal healthy vessels. Active targeting to the calcified tissues is achieved by ALN's affinity to bone mineral. METHODS AND FINDING: The anti-angiogenic and antitumor potency of HPMA copolymer-ALN-TNP-470 conjugate was evaluated both in vitro and in vivo. We show that free and conjugated ALN-TNP-470 have synergistic anti-angiogenic and antitumor activity by inhibiting proliferation, migration and capillary-like tube formation of endothelial and human osteosarcoma cells in vitro. Evaluation of anti-angiogenic, antitumor activity and body distribution of HPMA copolymer-ALN-TNP-470 conjugate was performed on severe combined immunodeficiency (SCID) male mice inoculated with mCherry-labeled MG-63-Ras human osteosarcoma and by modified Miles permeability assay. Our targeted bi-specific conjugate reduced VEGF-induced vascular hyperpermeability by 92% and remarkably inhibited osteosarcoma growth in mice by 96%. CONCLUSIONS: This is the first report to describe a new concept of a narrowly-dispersed combined polymer therapeutic designed to target both tumor and endothelial compartments of bone metastases and calcified neoplasms at a single administration. This new approach of co-delivery of two synergistic drugs may have clinical utility as a potential therapy for angiogenesis-dependent cancers such as osteosarcoma and bone metastases.
    PLoS ONE 02/2009; 4(4):e5233. · 3.73 Impact Factor
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    Russell N Johnson, Pavla Kopecková, Jindrich Kopecek
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    ABSTRACT: Several drug delivery designs combine synthetic drug carriers with covalently conjugated targeting moieties. Such modifications of monoclonal antibodies (mAb), or their Fab' fragments, inevitably result in diminished affinity for their targeted tissue. In an attempt to overcome this limitation, high molecular weight, branched N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers were synthesized and conjugated with Fab' fragments of the anti-CD20 antibody, 1F5. This produced multivalent conjugates with varying valency (amount of Fab' per macromolecule) targeted to the B-cell antigen CD20. Evaluation of a multivalent effect was done by determining the apparent K(D) at low concentrations of conjugates, the Sips heterogeneity factor, a, and the binding enhancement factors of each construct. The results clearly indicated that multivalency could improve the affinity of the HPMA copolymer-Fab' conjugates to that of unconjugated mAb.
    Bioconjugate Chemistry 02/2009; 20(1):129-37. · 4.58 Impact Factor
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    ABSTRACT: Enzyme-based hybrid hydrogels were prepared by covalently incorporating an adenylate kinase mutant, possessing two thiol groups, into HPMA copolymer- or PEG-based hydrogel structures. The nanoscale conformational change of enzyme, triggered by substrate recognition, translated into macroscopic motion of hydrogels.
    Journal of the American Chemical Society 12/2008; 130(47):15760-1. · 10.68 Impact Factor
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    ABSTRACT: A wide variety of therapeutic agents may benefit by specifically directing them to the mitochondria in tumor cells. The current work aimed to design delivery systems that would enable a combination of tumor and mitochondrial targeting for such therapeutic entities. To this end, novel HPMA copolymer-based delivery systems that employ triphenylphosphonium (TPP) ions as mitochondriotropic agents were developed. Constructs were initially synthesized with fluorescent labels substituting for drug and were used for validation experiments. Microinjection and incubation experiments performed using these fluorescently labeled constructs confirmed the mitochondrial targeting ability. Subsequently, HPMA copolymer-drug conjugates were synthesized using a photosensitizer mesochlorin e 6 (Mce 6). Mitochondrial targeting of HPMA copolymer-bound Mce 6 enhanced cytotoxicity as compared to nontargeted HPMA copolymer-Mce 6 conjugates. Minor modifications may be required to adapt the current design and allow for tumor site-specific mitochondrial targeting of other therapeutic agents.
    Molecular Pharmaceutics 10/2008; 5(5):776-86. · 4.57 Impact Factor
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    ABSTRACT: Macromolecular delivery systems have therapeutic uses because of their ability to deliver and release drugs to specific tissues. The uptake and localization of HPMA copolymers using Asp(8) as the bone-targeting moiety was determined in aged, ovariectomized (ovx) rats. PGE(1) was attached via a cathepsin K-sensitive linkage to HPMA copolymer-Asp(8) conjugate and was tested to determine if it could promote bone formation. The uptake of FITC-labeled HPMA copolymer-Asp(8) conjugate (P-Asp(8)-FITC) on bone surfaces was compared with the mineralization marker, tetracycline. Then a targeted PGE(1)-HPMA copolymer conjugate (P-Asp(8)-FITC-PGE(1)) was given as a single injection and its effects on bone formation were measured 4 weeks later. P-Asp(8)-FITC preferentially deposited on resorption surfaces, unlike tetracycline. A single injection of P-Asp(8)-FITC-PGE(1) resulted in greater indices of bone formation in aged, ovx rats. HPMA copolymers can be targeted to bone surfaces using Asp(8), with preferential uptake on resorption surfaces. Additionally, PGE(1) attached to the Asp(8)-targeted HPMA copolymers and given by a single injection resulted in greater bone formation measured 4 weeks later. This initial in vivo study suggests that macromolecular delivery systems targeted to bone may offer some therapeutic opportunities and advantages for the treatment of skeletal diseases.
    Pharmaceutical Research 09/2008; 25(12):2889-95. · 4.74 Impact Factor

Publication Stats

3k Citations
709.56 Total Impact Points

Institutions

  • 1988–2013
    • University of Utah
      • • Department of Pharmaceutics and Pharmaceutical Chemistry
      • • Department of Radiobiology
      • • Department of BioEngineering
      Salt Lake City, UT, United States
  • 2007
    • University of Nebraska at Omaha
      • Department of Pharmaceutical Sciences
      Omaha, NE, United States
  • 2001–2004
    • Hebrew University of Jerusalem
      • School of Pharmacy
      Jerusalem, Jerusalem District, Israel
  • 1995–1998
    • Academy of Sciences of the Czech Republic
      • Hydrobiologický ústav
      Praha, Hlavni mesto Praha, Czech Republic
  • 1988–1993
    • Keele University
      Newcastle-under-Lyme, England, United Kingdom