RESUMO
PURPOSE: To test targeted liposomes in an effort to improve drug transport across cellular barriers into the brain. METHODS: Therefore we prepared Mitoxantrone (MTO) entrapping, rigid and fluid liposomes, equipped with a 19-mer angiopeptide as ligand for LDL lipoprotein receptor related protein (LRP) targeting. RESULTS: Fluid, ligand bearing liposomes showed in vitro the highest cellular uptake and transcytosis and were significantly better than the corresponding ligand-free liposomes and rigid, ligand-bearing vesicles. Treatment of mice, transplanted with human breast cancer cells subcutaneously and into the brain, with fluid membrane liposomes resulted in a significant reduction in the tumor volume by more than 80% and in a clear reduction in drug toxicity. The improvement was mainly depended on liposome fluidity while the targeting contributed only to a minor degree. Pharmacokinetic parameters were also improved for liposomal MTO formulations in comparison to the free drug. So the area under the curve was increased and t(1/2) was extended for liposomes. CONCLUSION: Our data show that it is possible to significantly improve the therapy of brain metastases if MTO-encapsulating, fluid membrane liposomes are used instead of free MTO. This effect could be further enhanced by fluid, ligand bearing liposomes.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/secundário , Encéfalo/efeitos dos fármacos , Proteínas Relacionadas a Receptor de LDL/metabolismo , Lipossomos/metabolismo , Mitoxantrona/uso terapêutico , Sequência de Aminoácidos , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacocinética , Encéfalo/metabolismo , Encéfalo/patologia , Neoplasias Encefálicas/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular , Linhagem Celular Tumoral , Sistemas de Liberação de Medicamentos , Feminino , Humanos , Lipossomos/química , Camundongos , Mitoxantrona/administração & dosagem , Mitoxantrona/farmacocinética , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/metabolismoRESUMO
Alkylphospholipids (APLs) are physiologically active derivatives of lipids with high efficiency in the therapy of experimental human breast cancer. Therefore, they are usually used as liposome formulations, since the application of free, micellar APLs results in serious side effects, such as hemolysis. Since only rough upper estimates of micelle proportions were obtained until now, we developed a new method, based on the spectral decomposition of electron paramagnetic resonance (EPR) spectra. The EPR spectra of different spin-labeled mixed liposome-micelle samples were fitted with a fast, automatic fitting procedure. In this way, we achieved more accurate estimates of the relative proportion of micelles, in comparison to the previously used methods. We confirmed findings of previous studies, which showed that the amount of micelles in OPP liposome formulations increases with a decreasing amount of cholesterol. We analyzed liposome formulations with 29, 39, and 46 mol% of cholesterol and determined the relative proportion of micelles to be 20 ± 9, 11 ± 4, and 5 ± 2, respectively. This study confirms that the efficiency of OPP liposome formulations in experimental breast cancer therapy is not solely a consequence of the amount of micelles present in a liposome formulation, but stems from a propensity of liposomes to interact with cells.
Assuntos
Espectroscopia de Ressonância de Spin Eletrônica/métodos , Lipossomos , Micelas , Fosfolipídeos/química , Neoplasias da Mama/tratamento farmacológico , Simulação por Computador , Fosfolipídeos/efeitos adversos , Fosfolipídeos/uso terapêuticoRESUMO
The process of metastasis formation in cancer is not completely understood and is the main reason cancer therapies fail. Previously, we showed that dual liposomes simultaneously containing the hemostatic inhibitor, dipyridamole and the anticancer drug, perifosine potently inhibited metastasis, causing a 90% reduction in the number of lung metastases in a murine experimental metastasis model. To gain deeper insight into the mechanisms leading to the inhibition of metastasis by these dual liposomes, in the present study, the development of metastases by MT3 breast cancer cells in a mouse xenograft model was analyzed in more detail with regard to tumor cell settlement and metastatic growth. We found that the development of lung metastases by MT3 tumor cells is essentially dependent on the formation of fibrin clots as a precondition for the pulmonary arrest of tumor cells and the subsequent intravascular expansion of micrometastases before their invasion into the surrounding tissue.
Assuntos
Antineoplásicos/administração & dosagem , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/patologia , Metástase Neoplásica/prevenção & controle , Inibidores da Agregação Plaquetária/administração & dosagem , Animais , Coagulação Sanguínea/efeitos dos fármacos , Vasos Sanguíneos/efeitos dos fármacos , Linhagem Celular Tumoral , Dipiridamol/administração & dosagem , Feminino , Fibrina , Humanos , Lipossomos , Pulmão/irrigação sanguínea , Metalotioneína 3 , Camundongos , Camundongos Nus , Fosforilcolina/administração & dosagem , Fosforilcolina/análogos & derivados , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
We have investigated interaction of alkyphospholipid (APL) liposomes consisting of 1,1-dimethylpiperidin-1-ium-4-yl) octadecyl phosphate (OPP) and different concentrations of cholesterol (CH) with human MT-3 breast-cancer cells using electron paramagnetic resonance method (EPR) with advanced characterization of EPR spectra of spin labeled liposome membranes. After incubation of OPP liposomes with MT-3 cells, a reduction of liposome entrapped, water soluble spin-probe tempocholine (ASL) was observed, indicating that ASL is released from liposomes and is reduced by oxy-redoxy systems inside the cells. This process is fast if cholesterol content in the bilayer was 29 or 45 mol%, whereas at 56 mol% cholesterol the process is almost stopped. The rate of spin-probe reduction in first 10 min after incubation with cells is even faster as for the free ASL, indicating that liposomes with low amount of cholesterol accelerate penetration of ASL into the cells. A faster release of hydrophilic material from liposomes with low cholesterol content coincides with the presence of domains with highly disordered alkyl chain motion that disappears at 50 mol% of cholesterol. We propose that these highly fluid domains are responsible for interaction of OPP liposomes with cells and fast release of the entrapped material into the cells. These results suggest that micelles are not the only reason for cytotoxic effect of OPP liposome formulations, as it was suggested before. OPP in liposomes, containing 45 mol% cholesterol or less, also contributes to the cytotoxic effect, due to their fast interaction with breast-cancer cells.
Assuntos
Neoplasias da Mama/metabolismo , Colesterol/química , Lipossomos/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Óxidos N-Cíclicos , Espectroscopia de Ressonância de Spin Eletrônica , Feminino , Humanos , Lipossomos/química , Oxirredução , Fosfolipídeos/química , Marcadores de SpinRESUMO
The anti-cancer drug oxaliplatin (OxP) has rarely been used to treat breast carcinoma, as it cannot cross the BBB to treat the frequently subsequent brain metastases. Here, we encapsulated OxP in liposomes prepared to reduce side effects and to simultaneously treat primary tumor and brain metastasis. The angiopep LRP-receptor ligand was bound to the vesicular surface for targeting. Targeted and non-targeted OxP liposomes were tested in vitro (binding, uptake, and transcytosis) and in vivo. Liposomes contained 0.65 mg OxP/mL, their mean diameter was 165 nm, and they released 50% of OxP within 8 days at 4 degrees C and within 22 h at 36 degrees C. MDCK cells were used for uptake and transcytosis quantification. Compared to non-targeted liposomes, targeted liposomes showed 12-fold greater uptake, and 2.25-fold higher transcytosis. In vivo efficacy was tested using human MT-3 breast cancer cells transplanted subcutaneously and intracerebrally into female nude mice, and tumor growth inhibition was measured. OxP was injected (6 mg OxP/kg) four times. The best results were obtained with targeted liposomes (T/C: 21% for subcutaneous and 50% for intracerebral). OxP liposomes with a fluid membrane all inhibited MT-3 tumors significantly better than free OxP, with no significant difference between targeted and non-targeted liposomes. The therapeutic effect was accompanied with strong leukopenia and mild thrombocytopenia with all formulations. The newly developed OxP liposomes significantly improved the treatment of subcutaneously and intracerebrally growing breast cancer, but the targeted angiopep-equipped liposomes showed no superior effect in vivo.
Assuntos
Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/secundário , Neoplasias da Mama/tratamento farmacológico , Lipossomos/química , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Compostos Organoplatínicos/administração & dosagem , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Neoplasias Encefálicas/metabolismo , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Difusão , Feminino , Humanos , Metalotioneína 3 , Camundongos , Camundongos Nus , Terapia de Alvo Molecular/métodos , Compostos Organoplatínicos/química , Oxaliplatina , Resultado do TratamentoRESUMO
Alkylphospholipid analogues of perifosine and miltefosine bearing a nitroxide moiety at different positions on an alkyl chain were synthesized as electron paramagnetic resonance (EPR) probes. Their amphiphilic properties were characterized by determining their critical micelle concentration (cmc) and hemolytic activity on erythrocytes both in free and liposomal form. Spin-labeled analogues as membrane components of large unilamellar liposomes containing cholesterol and dicetyl phosphate or in free solution were evaluated using the MTT assay to determine growth inhibition on MT1, MT3, and MCF7 breast cancer cell lines. 4a (IC50 = 56.4 microM) was found to be significantly more active than the perifosine against the MCF-7 cell line. Its high cmc (194.03 microM) and low hemolytic activity shows that its cytotoxic activity might be more specific; therefore, 4a can be an important molecular tool for further EPR investigations.
Assuntos
Antineoplásicos/síntese química , Óxidos N-Cíclicos/síntese química , Organofosfatos/síntese química , Fosfolipídeos/síntese química , Fosforilcolina/análogos & derivados , Marcadores de Spin/síntese química , Tensoativos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Óxidos N-Cíclicos/química , Óxidos N-Cíclicos/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Hemólise , Humanos , Lipossomos , Micelas , Organofosfatos/química , Organofosfatos/farmacologia , Fosfolipídeos/química , Fosfolipídeos/farmacologia , Fosforilcolina/síntese química , Fosforilcolina/química , Fosforilcolina/farmacologia , Soluções , Relação Estrutura-Atividade , Tensoativos/química , Tensoativos/farmacologiaRESUMO
Metastasis is still the most serious reason for the high mortality of cancer patients. It is a complex process in which platelets play a crucial role. Several attempts have been performed to inhibit the metastatic process, some of these using modified liposomes. The aggregation behaviour of human platelets and HT29 colon carcinoma cells in the presence of liposomes with a modified surface has been investigated in the present study. Liposomes (PC/CH/DMPE) were unmodified, sterically stabilized by polyethylene glycol (PEG-DSPE), or equipped with the carbohydrate ligand sialyl Lewis(X) (conjugated to PEG-DMPE or DMPE as anchor) intended to specifically compete with ligands expressed by HT29 cells. We found in vitro that an addition of surface modified liposomes to human platelets in plasma caused an up to 2.9-fold increase in platelet aggregation. In addition, when HT29 tumor cells were mixed with platelets and surface modified liposomes, the number of tumor cells found in aggregates increased significantly from 8.3 % (only tumor cells) to 30.2 %. This result was supported by fluorescence micrographs demonstrating a strong association of platelets and liposomes around the tumor cells. In addition, a clear decrease in number and a change in the distribution of metastases after intravenous injection of HT29 cells in combination with liposomes was observed in vivo. While in control mice metastases in lung, liver and in intestine were prevailing, liposomal treatment resulted in a new localization of metastases in muscles. Taking together, the ability of surface modified liposomes to enhance aggregate formation of platelets and tumor cells has been demonstrated for the first time. The capability of these vesicles to interfere with the metastatic process might have implications for the use of such liposomes for therapeutic applications.
Assuntos
Lipossomos/química , Neoplasias/patologia , Propriedades de Superfície , Plaquetas/citologia , Linhagem Celular Tumoral , Separação Celular , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos , Citometria de Fluxo , Humanos , Ligantes , Lipossomos/metabolismo , Microscopia de Fluorescência , Metástase Neoplásica , Transplante de Neoplasias , Neoplasias/terapia , Oligossacarídeos/química , Fosfatidilcolinas/química , Fosfatidiletanolaminas/química , Agregação Plaquetária , Polietilenoglicóis/química , Ligação Proteica , Antígeno Sialil Lewis X , Fatores de TempoRESUMO
BACKGROUND: The development of efficient drug delivery systems to transport therapeutics across barrier-forming cells remains a challenge. Recently it was shown that liposomes containing perifosine, a synthetic analog of lysophosphatidylcholine, efficiently deliver liposome encapsulated content across barrier-forming cells. METHODS: To elucidate the mechanism of the delivery, fluorescent and spin labeled analog of perifosine were synthesized and their transport from liposomes to the barrier-forming MDKC cells was measured. RESULTS & CONCLUSION: Perifosine analogs are rapidly transported from liposomes into cell membranes. The total amount of perifosine accumulated in plasma membranes seems to be the most important factor in efficient transepithelial transport of liposome-encapsulated substances. Lysolipid-containing liposomal formulations seem to be promising candidates as drug delivery systems in general.
Assuntos
Membrana Celular/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Lipossomos/química , Fosforilcolina/análogos & derivados , Transporte Biológico/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Células Cultivadas , Química Farmacêutica , Liberação Controlada de Fármacos , Fluoresceínas/química , Humanos , Fosforilcolina/química , Fosforilcolina/farmacologiaRESUMO
Enhanced levels of the inflammatory chemokine CCL2 are known to correlate with increased tumorigenesis and metastases, and thereby poor prognosis for cancer patients. The CCL2-CCR2 chemokine axis was shown to facilitate the metastatic initiation through the recruitment of inflammatory monocytes and the activation of endothelial cells at metastatic sites. Both steps are required for efficient cancer cell trans-endothelial migration and seeding in the targeted tissue. The translation of preclinical evidence proved to be challenging due to systemic effects of chemokine inhibition and limited target specificity. Here we tested an approach of a targeted delivery of the CCR2 antagonist Teijin Compound 1 to metastatic sites. VCAM-1 binding peptide tagged liposomes carrying the CCR2 antagonist enabled a specific delivery to cancer cell-activated endothelium. The subsequent binding of target-sensitive liposomes triggered the release of the Teijin Compound 1 and thereby local inhibition of CCR2 in the lungs. Blocking of CCR2 resulted in reduced induction of the lungs vascular permeability, and thereby reduced tumor cell extravasation. However, the recruitment of inflammatory monocytes to the pre-metastatic lungs remained unaltered. Endothelial VCAM-1 targeted delivery of the CCR2 antagonist resulted in inhibition of pulmonary metastases both in a murine (MC-38GFP cells) and a human xenograft (patient-derived cells) model. Thus, timely- and spatially-defined inhibition of CCR2 signaling represents a potential therapeutic approach for treatment of metastasis without affecting homeostatic functions.
Assuntos
Antineoplásicos/administração & dosagem , Benzamidas/administração & dosagem , Células Endoteliais/efeitos dos fármacos , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Peptídeos/metabolismo , Artéria Pulmonar/efeitos dos fármacos , Pirrolidinas/administração & dosagem , Receptores CCR2/antagonistas & inibidores , Animais , Antineoplásicos/química , Antineoplásicos/metabolismo , Benzamidas/química , Benzamidas/metabolismo , Permeabilidade Capilar/efeitos dos fármacos , Técnicas de Cocultura , Composição de Medicamentos , Células Endoteliais/metabolismo , Feminino , Humanos , Lipossomos , Neoplasias Pulmonares/metabolismo , Camundongos , Peptídeos/química , Artéria Pulmonar/metabolismo , Pirrolidinas/química , Pirrolidinas/metabolismo , Receptores CCR2/metabolismo , Fatores de Tempo , Migração Transendotelial e Transepitelial/efeitos dos fármacos , Células U937 , Molécula 1 de Adesão de Célula Vascular/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
In this work, we evaluated combinations of doxorubicin with INNO-206, a (6-maleimidocaproyl)hydrazone derivative of doxorubicin (DOXO-EMCH) that is currently undergoing two phase II clinical trials, in a primarily chemoresistant tumor indication, i.e. pancreatic cancer. Thus, we compared the antitumor efficacy and tolerability of the following weekly intravenous treatments in the MIA PaCa-2 xenograft model: 3×6 mg doxorubicin (MTD), 3×24 mg/kg DOXO-EMCH (doxorubicin equivalents, MTD), 3×3 mg/kg doxorubicin followed 6h later by 3×12 mg/kg DOXO-EMCH, and 3×12 mg/kg DOXO-EMCH followed 6 h later by 3×3 mg/kg doxorubicin. Whereas therapy with doxorubicin only produced a moderate tumor inhibition, all other therapy arms induced complete and partial remissions up to the end of the experiment on day 43. Although the total amount of doxorubicin equivalents is 72 mg/kg when DOXO-EMCH is administered alone, but only 45 mg/kg doxorubicin equivalents are administered in the combination regimens, the antitumor efficacy in all treated groups was essentially identical, a surprising finding of this study. However, there were significant differences in the tolerability as assessed by the body weight changes: whereas therapy at the MTD of DOXO-EMCH (3×24 mg/kg) produced a body weight loss of -16% including one death, therapy with 3×12 mg/kg DOXO-EMCH followed 6h later by 3×3 mg/kg doxorubicin produced -7% body weight loss, and 3×3 mg/kg doxorubicin followed 6h later by 3×12 mg/kg DOXO-EMCH produced a body weight gain of +2% as a clear indication of minimal systemic toxicity. In addition, cell culture experiments revealed additive to synergistic effects when MIA PaCa-2 cells were exposed to doxorubicin followed 6h later to exposure of the albumin-bound form of DOXO-EMCH spanning a ratio of 1:5 to 5:1 (analyzed for synergistic, additive or antagonistic effects using the software program CalcuSyn(®)). This animal study demonstrates that the time-dependent schedule of an albumin-binding prodrug and a free drug has a critical influence on the overall tolerability. A combination of doxorubicin and DOXO-EMCH is currently being investigated in a phase Ib study.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Doxorrubicina/análogos & derivados , Hidrazonas/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Animais , Antibióticos Antineoplásicos/administração & dosagem , Antibióticos Antineoplásicos/toxicidade , Peso Corporal/efeitos dos fármacos , Relação Dose-Resposta a Droga , Doxorrubicina/administração & dosagem , Doxorrubicina/farmacologia , Doxorrubicina/toxicidade , Sinergismo Farmacológico , Feminino , Humanos , Hidrazonas/administração & dosagem , Hidrazonas/toxicidade , Injeções Intravenosas , Camundongos , Camundongos Nus , Neoplasias Pancreáticas/patologia , Pró-Fármacos , Indução de Remissão , Albumina Sérica/metabolismo , Fatores de Tempo , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
INTRODUCTION: The integrin VLA-4-mediated binding is important for the metastatic dissemination of melanoma cells. Recently we found that heparin possesses a binding capacity to VLA-4. This could contribute to the heparin function to attenuate metastasis in a selectin-dependent manner. Aiming to a purposive, anti-adhesive heparin application, structural requirements of heparin for VLA-4 recognition have to be elucidated. MATERIALS AND METHODS: A series of non-anticoagulant heparin derivatives were investigated concerning their inhibitory capacities for VLA-4 mediated binding of human melanoma MV3 cells to VCAM-1 under physiological flow conditions in vitro. A surface acoustic wave biosensor was applied to detect kinetic constants of selected derivatives binding to both, VLA-4 or P- and L-selectin. RESULTS: Experimental metastasis of MV3 cells in mice confirmed the relevance of VLA-4 for metastatic dissemination. LMWHs (enoxaparin, tinzaparin) efficiently blocked VLA-4 cell binding, dominantly via the integrin`s α-chain. Desulfation at 2-O-position, N-acetylation or a size smaller than tetradecasaccharide disfavoured VLA-4 inhibition. Glycol-splitting of heparin and thus higher chain flexibility is a tolerable parameter. A derivative with 50% 6-O-desulfation appeared promising and exceeded tinzaparin in VLA-4 inhibition, both compounds displayed binding affinities to VLA-4 in the low micromolar range. CONCLUSIONS: These findings provide structure-activity relationships for heparin VLA-4 binding, which partly differ from P- and L-selectin requirements. The data confirm that anti-coagulative and anti-adhesive function of heparin can be distinguished favouring applications of non-anticoagulant heparins in antimetastatic approaches without the risk of bleeding complications. The 50% 6-O-desulfated heparin-derivative appears promising to further evaluate the interference with selectin and VLA-4 binding functions in vivo.
Assuntos
Anticoagulantes/metabolismo , Heparina/análogos & derivados , Integrina alfa4beta1/antagonistas & inibidores , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Acetilação , Animais , Anticorpos Monoclonais Humanizados/farmacologia , Anticoagulantes/farmacologia , Adesão Celular/fisiologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Enoxaparina/química , Enoxaparina/farmacocinética , Enoxaparina/farmacologia , Feminino , Técnicas de Silenciamento de Genes , Heparina/química , Heparina/farmacocinética , Heparina/farmacologia , Heparina de Baixo Peso Molecular/química , Heparina de Baixo Peso Molecular/farmacocinética , Heparina de Baixo Peso Molecular/farmacologia , Humanos , Integrina alfa4beta1/genética , Integrina alfa4beta1/metabolismo , Melanoma/patologia , Camundongos , Natalizumab , Metástase Neoplásica , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/genética , Relação Estrutura-Atividade , Tinzaparina , Molécula 1 de Adesão de Célula Vascular/química , Molécula 1 de Adesão de Célula Vascular/metabolismoRESUMO
Two new water-soluble folate receptor-targeted drug conjugates that contain the highly active doxorubicin derivative N-(5,5-diacetoxybut-1-yl)doxorubicin were designed and evaluated for their biological activity against folate receptor positive tumours. The prodrugs were designed to contain an acid-sensitive hydrazone bond KO019 or in addition a disulphide bond KO013 in order to elucidate the importance of the pre-determined breaking point for their in vitro and in vivo properties. Fluorescence microscopy studies confirmed higher uptake of the prodrugs in folate receptor positive KB cells than in the folate receptor negative A549 lung cancer cells. In subsequent in vivo studies in the folate receptor positive KB xenograft model, KO019 was as active as the free drug but significantly less toxic when dosed at twice the dose of the free drug whereas KO013 showed no anticancer efficacy. As an explanation, we could show by HPLC that the prodrug KO013 that additionally contains a disulphide bond undergoes rapid disulphide exchange in murine plasma in the order of 40% after 5h at 37°C in contrast to KO019 which was essentially stable after a 5h incubation.
Assuntos
Doxorrubicina/análogos & derivados , Receptores de Folato com Âncoras de GPI/metabolismo , Ácido Fólico/análogos & derivados , Pró-Fármacos/farmacologia , Pterinas/farmacologia , Alcaloides de Vinca/farmacologia , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Receptores de Folato com Âncoras de GPI/antagonistas & inibidores , Ácido Fólico/farmacologia , Camundongos , Camundongos Nus , Pró-Fármacos/química , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The successful treatment of brain tumors or metastases in the brain is still hampered by the very efficient blood-brain barrier, which prevents the cerebral accumulation of a pharmacologically sufficient amount of a drug. Beside the possibility of disintegrating the functionality of this effective working barrier, a nanocarrier-mediated transport is presently an interesting and promising method to increase the drug concentration in the brain. Nanocarriers are small vesicles (<200 nm) and can be prepared by polymerization, resulting in nanoparticles, or by producing superficial lipid structures to incorporate the drug. In this context, liposomes are of importance owing to their ability to adapt their properties to the pharmacological requirements. In this article, we will give an overview of current possibilities of enhancing anticancer drug transport across the blood-brain barrier, based on its structure and functionality. Special consideration will be given to recent liposomal approaches that use active targeting for receptor-mediated transport across this physiological barrier.
Assuntos
Antineoplásicos/metabolismo , Barreira Hematoencefálica/metabolismo , Portadores de Fármacos/metabolismo , Animais , Antineoplásicos/administração & dosagem , Transporte Biológico/fisiologia , Barreira Hematoencefálica/citologia , Barreira Hematoencefálica/efeitos dos fármacos , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/metabolismo , Portadores de Fármacos/administração & dosagem , Humanos , LipossomosRESUMO
The interaction between circulating tumor cells and blood components, mainly platelets, plays an important role during metastasis. In this study, we prepared liposomes containing the platelet aggregation inhibitor Cilostazol (Cil-L). The objective of this study was to investigate the effect of this Cil-L on platelet aggregation and complex formation with murine 4T1 breast cancer cells in vitro and to determine their anti-metastatic potency in a spontaneous metastasis model of 4T1 breast cancer. Cil-L significantly inhibited the aggregation of platelets by up to 78% and completely abolished the complex formation of 4T1 tumor cells in the presence of activated platelets in vitro. Intravenous (i.v.) injection of Cil-L into mice significantly reduced the aggregability of mouse platelets by 60% measured ex vivo. To gain deeper insight into the mode of metastasis formation in a spontaneous metastasis model, 4T1 breast cancer cells were transplanted into the mammary fad pad of mice and metastasis to the mouse lungs was investigated with regard to tumor cell settlement and metastatic growth. We could demonstrate that the formation of pulmonary metastases was significantly reduced by 55% when mice were treated intravenously with 100 nmol Cil-L 6 h before tumor cell inoculation and then daily for 2 weeks. We conclude that Cil-L reduced metastasis by restricting the aggregability of mouse platelets, which probably prevents the interaction between circulating 4T1 tumor cells and platelets, making the Cil-L a useful tool for the inhibition of breast cancer metastasis in mice.
Assuntos
Plaquetas/fisiologia , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/patologia , Inibidores da Agregação Plaquetária/farmacologia , Agregação Plaquetária/efeitos dos fármacos , Tetrazóis/farmacologia , Animais , Linhagem Celular Tumoral , Cilostazol , Lipossomos , Camundongos , Camundongos Nus , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The aim of this study was to investigate the effect of liposomal membrane properties on cellular uptake and transcytosis across a tight Madin-Darby canine kidney (MDCK) cell barrier in vitro. More than 25 small vesicles were prepared by lipid film hydration/extrusion to generate small unilamellar vesicles. The fluorescence marker calcein was encapsulated to mimic hydrophilic drug transport. Marker uptake by MDCK cells seems to be mediated by different mechanisms for the liposomes used. It was mainly depending on membrane fluidity and vesicle charge. Liposomes L2 with a positive charge (325 +/- 3 pmol/well) and vesicles L3 containing the helper lipid dioleylphosphatidylethanolamine (DOPE) in their membrane (216 +/- 42 pmol/well) were taken up to the most. Selected liposomes were tested for their transcytotic transport across a MDCK monolayer. Liposomes L4 containing equimolar DOPE and octadecyl-1,1-dimethylpiperidin-1-ium-4-yl phosphate (OPP) were the most efficient vesicles for transcellular transport resulting in 808 +/- 30 pmol calcein/cm(2) in the basal medium (28.1% of total liposomal marker added). Transcytosis was positively correlated with membrane fluidity in the outer part of the bilayer, as electron paramagnetic resonance measurements revealed. We expect that an increase in membrane fluidity of vesicles should also improve the restricted transport of hydrophilic drugs across the blood-brain barrier.
Assuntos
Portadores de Fármacos/química , Células Epiteliais/metabolismo , Fluidez de Membrana , Nanopartículas/química , Lipossomas Unilamelares/química , Animais , Transporte Biológico , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Cães , Portadores de Fármacos/farmacocinética , Composição de Medicamentos , Espectroscopia de Ressonância de Spin Eletrônica , Células Epiteliais/efeitos dos fármacos , Fluoresceínas/administração & dosagem , Corantes Fluorescentes/administração & dosagem , Microscopia Confocal , Microscopia de Fluorescência , Lipossomas Unilamelares/farmacocinéticaRESUMO
The interaction between tumour cells and blood components, mainly platelets, plays an important role in metastasis. In this study, the anti-metastatic effect of vesicles containing the cytotoxic drug perifosine (OPP) and the haemostatic inhibitor dipyridamole (DIP) was tested. These dual liposomes (DIP/OPP-L) encapsulating up to 400microg DIP/ml and 6mM OPP were prepared by extrusion technique. In vitro, DIP/OPP-L significantly inhibited the aggregation of platelets and reduced their adhesion to immobilized MT3 cells by up to 60% and 24.7%, respectively. Complex formation between platelets and tumour cells in vitro was completely prevented by DIP/OPP-vesicles. These combinatory liposomes also inhibited the metastatic capacity of circulating tumour cells by reducing the complex formation with platelets. Formation of lung and extrapulmonary metastases after intravenous administration of MT3 breast cancer cells was significantly reduced when mice were treated with a single intravenous dose of DIP/OPP-L containing 100nmol lipid 6h before tumour cell inoculation.
Assuntos
Antineoplásicos/administração & dosagem , Neoplasias da Mama/patologia , Dipiridamol/administração & dosagem , Fosforilcolina/análogos & derivados , Inibidores da Agregação Plaquetária/administração & dosagem , Animais , Antineoplásicos/uso terapêutico , Plaquetas/efeitos dos fármacos , Plaquetas/fisiologia , Linhagem Celular Tumoral , Dipiridamol/uso terapêutico , Combinação de Medicamentos , Feminino , Humanos , Lipossomos , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Metalotioneína 3 , Camundongos , Camundongos Nus , Metástase Neoplásica , Transplante de Neoplasias , Células Neoplásicas Circulantes/efeitos dos fármacos , Fosforilcolina/administração & dosagem , Fosforilcolina/uso terapêutico , Inibidores da Agregação Plaquetária/uso terapêuticoRESUMO
To study whether membrane fluidity of tumor cells have an influence on metastasis, MT3 breast cancer cells harvested during exponential growth and under confluent conditions were compared. Electron paramagnetic resonance (EPR) data revealed that, in comparison to growing cells, confluent cells have a significant higher fluidity in their membrane related to a higher relative portion of disordered domains and a reduced portion of the most ordered domains. Further, sialyl Lewis X and/or A ligand-mediated adhesion of these cells was 2-fold enhanced. Confocal laser scanning microscopy further demonstrated a higher motility of ligands in the membrane of confluent cells, together with an accumulation of these ligands in distinct areas. Both facts are suggested to be responsible for an enhanced cell adhesion observed. Finally, an increased number of large distinct metastatic foci was registered in lungs of mice after i.v. inoculation of confluent cells. The results indicate that domain organization and fluidity of the cell membrane affect tumor cell adhesion and can have in this way also an impact on the malignancy of breast cancer cells.
Assuntos
Neoplasias da Mama/fisiopatologia , Neoplasias da Mama/secundário , Adesão Celular , Fluidez de Membrana , Animais , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Humanos , Metalotioneína 3 , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Invasividade Neoplásica , Estatística como AssuntoRESUMO
The domain structure of human cancer cells membranes was investigated by electron paramagnetic resonance (EPR) in different phases of cell growth, and the results were compared to those obtained for nonmalignant cells. On the basis of computer simulation of the EPR spectra using a newly developed GHOST condensation routine it was suggested that plasma membranes of cancer cells have less lateral lipid domain types at confluent conditions than in the exponential growing phase, while in nonmalignant cells the domain structure does not change significantly during cell growth. In accordance to our experimental data we propose a membrane switch hypothesis: disappearance of certain membrane domain types might act as a switch promoting the clustering of membrane constituents into the active units in a common lipid membrane domain and thus influencing the physiology of cells.
Assuntos
Membrana Celular/química , Membrana Celular/fisiologia , Neoplasias/química , Neoplasias/ultraestrutura , Contagem de Células , Linhagem Celular , Linhagem Celular Tumoral , Simulação por Computador , Espectroscopia de Ressonância de Spin Eletrônica , Humanos , Fluidez de Membrana , Proteínas de Membrana/metabolismoRESUMO
We investigated whether it is possible to reduce anti-estrogen resistance using liposomally encapsulated tamoxifen in vivo. Small liposomal vesicles containing up to 5.1 mg tamoxifen/ml liposomal suspension, together with an alkylphospholipid to enhance the cellular uptake, were prepared and characterized. Mice transplanted with different tumor models were treated with tamoxifen liposomes administered i.p. or orally as a bolus dose of 50 mg/kg once a week or as a daily dose of 10 mg/kg/day, both during a 4-week period. After orally administered tamoxifen liposomes, tumor growth was significantly reduced for the 3366/tamoxifen (acquired resistance) and for the MCF-7 (inherent resistance) models to 47 and 16%, respectively (treated to control value of relative tumor volume). Intraperitoneal treatment with tamoxifen liposomes revealed similar results. Investigation of biodistribution revealed especially an accumulation of liposomal tamoxifen in MCF-7 tumors and livers of the treated mice. These liposomes had uterotrophic properties comparable to the dissolved compound. This study demonstrates for the first time that a liposomal formulation of tamoxifen was able to induce pharmacological effects and to improve the therapeutic efficacy in several anti-estrogen-resistant xenografts.