A two-component CD20 (non-internalizing) receptor crosslinking system based on the biorecognition of complementary coiled-coil forming peptides was evaluated. Exposure of B cells to Fab'-peptide1 conjugate decorates the cell surface with peptide1; further exposure of the decorated cells to P-(peptide2)x (P is the N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer backbone) results in the formation of coiled-coil heterodimers at the cell surface with concomitant induction of apoptosis. The aim of this study was to determine the potential immunogenicity of this therapeutic system that does not contain low molecular weight drugs. Enantiomeric peptides (L- and D-CCE and L- and D-CCK), HPMA copolymer-peptide conjugates, and Fab' fragment-peptide conjugates were synthesized and the immunological properties of peptide conjugates evaluated in vitro on RAW264.7 macrophages and in vivo on immunocompetent BALB/c mice. HPMA copolymer did not induce immune response in vitro and in vivo. Administration of P-peptide conjugates with strong adjuvant resulted in antibody response directed to the peptide. Fab' was responsible for macrophage activation of Fab'-peptide conjugates and a major factor in the antibody induction following i.v. administration of Fab'-conjugates. There was no substantial difference in the ability of conjugates of D-peptides and conjugates of L-peptides to induce Ab response.

• Klíčová slova
• Coiled-coil peptides, Drug-free macromolecular therapeutics, Enantiomers, Fab' fragment, HPMA copolymer, Immunogenicity,
• MeSH
• akrylamidy aplikace a dávkování   chemie   imunologie   MeSH
• buněčné linie MeSH
• imunoglobuliny - Fab fragmenty aplikace a dávkování   chemie   imunologie   MeSH
• makrofágy účinky léků   imunologie   MeSH
• molekulární sekvence - údaje MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• peptidy aplikace a dávkování   chemie   imunologie   MeSH
• sekvence aminokyselin MeSH
• T-lymfocyty účinky léků   imunologie   MeSH
• tvorba protilátek účinky léků   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• akrylamidy MeSH
• imunoglobuliny - Fab fragmenty MeSH
• N-(2-hydroxypropyl)methacrylamide MeSH Prohlížeč
• peptidy MeSH

To avoid the side effects of the anti-cancer drug doxorubicin (Dox), we conjugated this drug to a N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer backbone. Dox was conjugated via an amide bond (Dox-HPMA(AM), PK1) or a hydrazone pH-sensitive bond (Dox-HPMA(HYD)). In contrast to Dox and Dox-HPMA(HYD), Dox-HPMA(AM) accumulates within the cell's intracellular membranes, including those of the Golgi complex and endoplasmic reticulum, both involved in protein glycosylation. Flow cytometry was used to determine lectin binding and cell death, immunoblot to characterize the presence of CD7, CD43, CD44, and CD45, and high-performance anion exchange chromatography with pulsed amperometric detector analysis for characterization of plasma membrane saccharide composition. Incubation of EL4 cells with Dox-HPMA(AM) conjugate, in contrast to Dox or Dox-HPMA(HYD), increased the amounts of membrane surface-associated glycoproteins, as well as saccharide moieties recognized by peanut agglutinin, Erythrina cristagalli, or galectin-1 lectins. Only Dox-HPMA(AM) increased expression of the highly glycosylated membrane glycoprotein CD43, while expression of others (CD7, CD44, and CD45) was unaffected. The binding sites for galectin-1 are present on CD43 molecule. Furthermore, we present that EL4 treated with Dox-HPMA(AM) possesses increased sensitivity to galectin-1-induced apoptosis. In this study, we demonstrate that Dox-HPMA(AM) treatment changes glycosylation of the EL4 T cell lymphoma surface and sensitizes the cells to galectin-1-induced apoptosis.

• MeSH
• amidy chemie   MeSH
• antibiotika antitumorózní farmakologie   MeSH
• antigeny CD43 metabolismus   MeSH
• apoptóza MeSH
• doxorubicin analogy a deriváty   farmakologie   MeSH
• endoplazmatické retikulum metabolismus   MeSH
• galektin 1 metabolismus   MeSH
• glykosylace MeSH
• Golgiho aparát metabolismus   MeSH
• kyseliny polymethakrylové farmakologie   MeSH
• lymfom T-buněčný farmakoterapie   metabolismus   patologie   MeSH
• myši MeSH
• nádorové buněčné linie účinky léků   MeSH
• nosiče léků MeSH
• proliferace buněk MeSH
• průtoková cytometrie MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• amidy MeSH
• antibiotika antitumorózní MeSH
• antigeny CD43 MeSH
• doxorubicin-N-(2-hydroxypropyl)methacrylamide copolymer conjugate MeSH Prohlížeč
• doxorubicin MeSH
• galektin 1 MeSH
• kyseliny polymethakrylové MeSH
• nosiče léků MeSH

PURPOSE: In vivo efficacy and safety of HPMA-based copolymers armed with doxorubicin via a spacer containing pH-sensitive linkage that can be prepared within a broad range of attached drug contents (1) was tested in murine tumor models. METHODS: Mice bearing T cell lymphoma EL4 or B cell lymphoma 38C13 were treated with a single dose of the conjugate (15, 25, and 75 mg Dox eq./kg i.v.) in a therapeutic regime. Anti-tumor resistance of the cured animals was proved by a second challenge with a lethal dose of tumor cells without additional treatment. RESULTS: The content of drug bound to the polymer is an important parameter in relation to the conjugate therapeutic efficacy. The best anti-tumor effects were produced by conjugates with 10 - 13 wt% of bound doxorubicin. Free doxorubicin up to 4.6% relative to total drug content had no impact on the treatment efficacy and acute toxicity. The conjugates induced a complete cure of mice and regular treatment-dependent development of specific anti-tumor resistance. No myelosuppression or organ damage was observed. CONCLUSIONS: A well-defined HPMA copolymer-doxorubicin conjugate with pH-sensitive drug release is a good candidate for clinical trials as it has remarkable anti-tumor efficacy and a favorable safety profile.

• MeSH
• antibiotika antitumorózní farmakokinetika   farmakologie   MeSH
• doxorubicin analogy a deriváty   chemická syntéza   farmakokinetika   farmakologie   MeSH
• imunomodulace účinky léků   MeSH
• koncentrace vodíkových iontů MeSH
• kyseliny polymethakrylové chemická syntéza   farmakokinetika   farmakologie   MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nosiče léků chemická syntéza   farmakokinetika   farmakologie   MeSH
• polymery * chemická syntéza   farmakokinetika   farmakologie   MeSH
• proliferace buněk účinky léků   MeSH
• xenogenní modely - testy antitumorózní aktivity MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antibiotika antitumorózní MeSH
• doxorubicin-N-(2-hydroxypropyl)methacrylamide copolymer conjugate MeSH Prohlížeč
• doxorubicin MeSH
• kyseliny polymethakrylové MeSH
• nosiče léků MeSH
• polymery * MeSH

Linkage of doxorubicin (Dox) to a water-soluble synthetic N-(2-hydroxypropyl)methacrylamide copolymer (PHPMA) eliminates most of the systemic toxicity of the free drug. In EL-4 lymphoma-bearing C57BL/6 mice, a complete regression of pre-established tumours has been achieved upon treatment with Dox-PHPMA-HuIg conjugate. The treatment was effective using a range of regimens and dosages, ranging from 62.5 to 100% cured mice treated with a single dose of 10-20 mg of Dox eq./kg, respectively. Fractionated dosages producing lower levels of the conjugate for a prolonged time period had substantial curative capacity as well. The cured mice developed anti-tumour protection as they rejected subsequently re-transplanted original tumour. The proportion of tumour-protected mice inversely reflected the effectiveness of the primary treatment. The treatment protocol leading to 50% of cured mice produced only protected mice, while no mice treated with early treatment regimen (i.e. starting on day 1 after tumour transplantation) rejected the re-transplanted tumour. Exposure of the host to the cancer cells was a prerequisite for developing protection. The anti-tumour memory was long lasting and specific against the original tumour, as the cured mice did not reject another syngeneic tumour, melanoma B16-F10. The immunity was transferable to naïve recipients in in vivo neutralization assay by spleen cells or CD8(+) lymphocytes derived from cured animals. We propose an effective treatment strategy which eradicates tumours without harming the protective immune anti-cancer responses.

• MeSH
• antibiotika antitumorózní terapeutické užití   MeSH
• doxorubicin analogy a deriváty   terapeutické užití   MeSH
• imunoglobuliny terapeutické užití   MeSH
• imunologická tolerance * MeSH
• kyseliny polymethakrylové terapeutické užití   MeSH
• lidé MeSH
• lymfom T-buněčný farmakoterapie   imunologie   prevence a kontrola   MeSH
• melanom experimentální farmakoterapie   imunologie   metabolismus   MeSH
• míra přežití MeSH
• myši inbrední C57BL MeSH
• myši nahé MeSH
• myši MeSH
• nádorové buňky kultivované transplantace   MeSH
• nádory kůže farmakoterapie   imunologie   metabolismus   MeSH
• nosiče léků MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antibiotika antitumorózní MeSH
• doxorubicin-N-(2-hydroxypropyl)methacrylamide copolymer conjugate MeSH Prohlížeč
• doxorubicin MeSH
• imunoglobuliny MeSH
• kyseliny polymethakrylové MeSH
• nosiče léků MeSH

This study compares the toxic effects of native cyclosporia A (CyA) with those of targeted CyA that is conjugated with the anti-rat-thymocyte antibody of rabbit origin via the N-(2-hydroxypropyl)methacrylamide (HPMA) carrier bearing digestible, reactive oligopeptide side chains. Ten toxic doses of native CyA (50 mg/kg i.p.) given to young adult rats in the course of 14 d produced a severe renal lesion-diffuse microvacuolization of the proximal tubules in the deep cortex, and hypergranulation of juxtaglomerular regions. Severe atrophy of the thymic medulla was documented by morphometry. In the cortex the epithelial reticular (but not deep interdigitating) cells showed ultrastructural signs of severe degeneration and lysis. The immature CD4+8+ double-positive cortical lymphocytes were preserved whereas the single-positive medullary thymocytes were greatly depleted; there was also a restriction of MHC class II antigen expression in the medulla. The number of medullary B cells was increased. The cytokeratin net was focally shrunken in the cortex and almost negative in the medulla, with loss of Hassall's corpuscles. After ten corresponding doses of antibody-targeted conjugated CyA no damage to the renal tubules and arterioles appeared and the antiGBM or immune-complex deposition was absent. The thymus had a normal medulla with numerous mature thymocytes and the cortical epithelial reticulum remained well preserved. Thus, the main toxic effects of CyA could be eliminated by targeting. The T-cell-targeted drug was tested for preserved immunosuppressive properties and non-toxic character of HPMA copolymer carrier.

• MeSH
• cyklosporin toxicita   MeSH
• cyklosporiny toxicita   MeSH
• imunokonjugáty toxicita   MeSH
• imunosupresiva toxicita   MeSH
• krysa rodu Rattus MeSH
• ledviny účinky léků   MeSH
• nosiče léků toxicita   MeSH
• potkani Wistar MeSH
• T-lymfocyty účinky léků   imunologie   MeSH
• thymus účinky léků   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• cyklosporin MeSH
• cyklosporiny MeSH
• imunokonjugáty MeSH
• imunosupresiva MeSH
• nosiče léků MeSH

Drug targeting is an attractive new approach to killing cancer cells while leaving normal tissue unharmed. Recently we have developed a new generation of antibody-targeted immunosuppressive (cyclosporin A) and cytostatic (daunomycin, doxorubicin) drugs and photosensitizers (chlorin e6) effective in vitro and in vivo. The drugs and the targeting antibody (polyclonal and monoclonal) are conjugated to the oligopeptidic side chains of a water-soluble synthetic carrier, copolymer of N-(2-hydroxypropyl)methacrylamide. The composition of the side chains ensures the stability of the linkage between the drug and the polymeric carrier in the bloodstream and its intralysosomal degradability which is a prerequisite for the pharmacological activity of the preparation. Antibody-targeted polymer bound drugs show considerably decreased hepatotoxicity, cardiotoxicity, myelotoxicity and nephrotoxicity. Two adriamycin-HPMA copolymers are in Phase I/II clinical trials in United Kingdom.

• MeSH
• akrylamidy aplikace a dávkování   škodlivé účinky   chemie   farmakokinetika   MeSH
• antibiotika antitumorózní aplikace a dávkování   škodlivé účinky   farmakokinetika   MeSH
• antigeny Thy-1 imunologie   MeSH
• antitumorózní látky aplikace a dávkování   škodlivé účinky   farmakokinetika   MeSH
• chlorofylidy MeSH
• cyklosporin aplikace a dávkování   škodlivé účinky   farmakokinetika   MeSH
• daunomycin terapeutické užití   MeSH
• doxorubicin analogy a deriváty   terapeutické užití   MeSH
• histokompatibilita - antigeny třídy II imunologie   MeSH
• imunokonjugáty * aplikace a dávkování   škodlivé účinky   chemie   farmakokinetika   MeSH
• imunosupresiva aplikace a dávkování   škodlivé účinky   chemie   farmakokinetika   MeSH
• klinické zkoušky jako téma MeSH
• kyseliny polymethakrylové terapeutické užití   MeSH
• lidé MeSH
• lyzozomy metabolismus   MeSH
• monoklonální protilátky aplikace a dávkování   chemie   imunologie   farmakokinetika   MeSH
• myši inbrední BALB C MeSH
• myši nahé MeSH
• myši MeSH
• nádory farmakoterapie   MeSH
• nemoci imunitního systému farmakoterapie   MeSH
• porfyriny aplikace a dávkování   škodlivé účinky   farmakokinetika   MeSH
• radiosenzibilizující látky aplikace a dávkování   škodlivé účinky   chemie   farmakokinetika   MeSH
• T-lymfocyty - podskupiny imunologie   MeSH
• tkáňová distribuce MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• akrylamidy MeSH
• antibiotika antitumorózní MeSH
• antigeny Thy-1 MeSH
• antitumorózní látky MeSH
• chlorofylidy MeSH
• cyklosporin MeSH
• daunomycin-N-(2-hydroxypropyl)methacrylamide copolymer conjugate MeSH Prohlížeč
• daunomycin MeSH
• doxorubicin-N-(2-hydroxypropyl)methacrylamide copolymer conjugate MeSH Prohlížeč
• doxorubicin MeSH
• histokompatibilita - antigeny třídy II MeSH
• imunokonjugáty * MeSH
• imunosupresiva MeSH
• kyseliny polymethakrylové MeSH
• monoklonální protilátky MeSH
• N-(2-hydroxypropyl)methacrylamide MeSH Prohlížeč
• phytochlorin MeSH Prohlížeč
• porfyriny MeSH
• radiosenzibilizující látky MeSH