Angiogenesis has a pivotal role in tumor growth and the metastatic process. Molecular imaging was shown to be useful for imaging of tumor-induced angiogenesis. A great variety of radiolabeled peptides have been developed to target αvβ3 integrin, a target structure involved in the tumor-induced angiogenic process. The presented study aimed to synthesize deferoxamine (DFO)-based c(RGD) peptide conjugate for radiolabeling with gallium-68 and perform its basic preclinical characterization including testing of its tumor-imaging potential. DFO-c(RGDyK) was labeled with gallium-68 with high radiochemical purity. In vitro characterization including stability, partition coefficient, protein binding determination, tumor cell uptake assays, and ex vivo biodistribution as well as PET/CT imaging was performed. [68Ga]Ga-DFO-c(RGDyK) showed hydrophilic properties, high stability in PBS and human serum, and specific uptake in U-87 MG and M21 tumor cell lines in vitro and in vivo. We have shown here that [68Ga]Ga-DFO-c(RGDyK) can be used for αvβ3 integrin targeting, allowing imaging of tumor-induced angiogenesis by positron emission tomography.
- MeSH
- deferoxamin analogy a deriváty chemická syntéza chemie MeSH
- glioblastom diagnostické zobrazování MeSH
- integrin alfaVbeta3 metabolismus MeSH
- lidé MeSH
- myši inbrední BALB C MeSH
- myši nahé MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- patologická angiogeneze diagnostické zobrazování MeSH
- počítačová rentgenová tomografie metody MeSH
- pozitronová emisní tomografie metody MeSH
- radioizotopy galia chemie MeSH
- tkáňová distribuce MeSH
- transplantace heterologní MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
In this work, we report two concepts of drug delivery based on small-molecule drug conjugates with the ability of specific targeting and drug release monitoring via ratiometric fluorescence. The functionality of these concepts has been verified by two model systems consisting of three parts: (i) fluorescent aminoBODIPY for real-time detection of conjugate cleavage, (ii) a c(RGDfK) peptide specific for αvβ3 integrin receptors targeting angiogenesis in most solid tumors or redBODIPY for conjugate cleavage monitoring via FRET, and (iii) pegylated-2-phenyl-3-hydroxy-4(1H)-quinolinone (3HQ) as a model drug. The model drug release is based on a self-immolative disulfide linker sensitive to environments containing thiols, especially glutathione, which is overexpressed in cancer cells. The results show effective thiol-mediated cleavage of the fluorescent reporter and the subsequent liberation of the drug in a tube. The conjugate with c(RGDfK) was confirmed to penetrate the cells via interaction with integrin receptors. Drug release from this conjugate is possible to monitor inside the cells. Further, the synthetic approach to the conjugates and the method of fluorescence monitoring of the drug release have also been described.
- MeSH
- fluorescence MeSH
- fluorescenční barviva chemie MeSH
- glutathion metabolismus MeSH
- HeLa buňky MeSH
- hydroxychinoliny aplikace a dávkování farmakokinetika MeSH
- integrin alfaVbeta3 metabolismus MeSH
- lidé MeSH
- nosiče léků chemie farmakologie MeSH
- oligopeptidy chemie farmakologie MeSH
- sloučeniny boru chemie MeSH
- uvolňování léčiv MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Induction of selective thrombosis and infarction in tumor-feeding vessels represents an attractive strategy to combat cancer. Here we took advantage of the unique coagulation properties of staphylocoagulase and genetically engineered it to generate a new fusion protein with novel anti-cancer properties. This novel bi-functional protein consists of truncated coagulase (tCoa) and an NGR (GNGRAHA) motif that recognizes CD13 and αvβ3 integrin receptors, targeting it to tumor endothelial cells. Herein, we report that tCoa coupled by its C-terminus to an NGR sequence retained its normal binding activity with prothrombin and avβ3 integrins, as confirmed in silico and in vitro. Moreover, in vivo biodistribution studies demonstrated selective accumulation of FITC-labeled tCoa-NGR fusion proteins at the site of subcutaneously implanted PC3 tumor xenografts in nude mice. Notably, systemic administration of tCoa-NGR to mice bearing 4T1 mouse mammary xenografts or PC3 human prostate tumors resulted in a significant reduction in tumor growth. These anti-tumor effects were accompanied by massive thrombotic occlusion of small and large tumor vessels, tumor infarction and tumor cell death. From these findings, we propose tCoa-NGR mediated tumor infarction as a novel and promising anti-cancer strategy targeting both CD13 and integrin αvβ3 positive tumor neovasculature.
- MeSH
- antigeny CD13 metabolismus MeSH
- buněčná smrt fyziologie MeSH
- endoteliální buňky pupečníkové žíly (lidské) MeSH
- integrin alfaVbeta3 metabolismus MeSH
- koagulasa metabolismus MeSH
- lidé MeSH
- myši inbrední C57BL MeSH
- myši nahé MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- nádory mléčné žlázy u zvířat metabolismus patologie MeSH
- nádory prostaty metabolismus patologie MeSH
- oligopeptidy metabolismus MeSH
- patologická angiogeneze metabolismus patologie MeSH
- xenogenní modely - testy protinádorové aktivity 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
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
Positron emission tomography (PET) as well as optical imaging (OI) with peptide receptor targeting probes have proven their value for oncological applications but also show restrictions depending on the clinical field of interest. Therefore, the combination of both methods, particularly in a single molecule, could improve versatility in clinical routine. This proof of principle study aims to show that a chelator, Fusarinine C (FSC), can be utilized as scaffold for novel dimeric dual-modality imaging agents. Two targeting vectors (a minigastrin analogue (MG11) targeting cholecystokinin-2 receptor overexpression (CCK2R) or integrin αVβ3 targeting cyclic pentapeptides (RGD)) and a near-infrared fluorophore (Sulfo-Cyanine7) were conjugated to FSC. The probes were efficiently labeled with gallium-68 and in vitro experiments including determination of logD, stability, protein binding, cell binding, internalization, and biodistribution studies as well as in vivo micro-PET/CT and optical imaging in U-87MG αVβ3- and A431-CCK2R expressing tumor xenografted mice were carried out. Novel bioconjugates showed high receptor affinity and highly specific targeting properties at both receptors. Ex vivo biodistribution and micro-PET/CT imaging studies revealed specific tumor uptake accompanied by slow blood clearance and retention in nontargeted tissues (spleen, liver, and kidneys) leading to visualization of tumors at early (30 to 120 min p.i.). Excellent contrast in corresponding optical imaging studies was achieved especially at delayed time points (24 to 72 h p.i.). Our findings show the proof of principle of chelator scaffolding for hybrid imaging agents and demonstrate FSC being a suitable bifunctional chelator for this approach. Improvements to fine-tune pharmacokinetics are needed to translate this into a clinical setting.
- MeSH
- chelátory chemie farmakokinetika MeSH
- heterografty MeSH
- integrin alfaVbeta3 metabolismus MeSH
- kyseliny hydroxamové farmakokinetika MeSH
- lidé MeSH
- molekulární sondy chemie farmakokinetika MeSH
- multimodální zobrazování metody MeSH
- myši MeSH
- nádorové buňky kultivované MeSH
- nádory diagnostické zobrazování metabolismus MeSH
- PET/CT MeSH
- radioizotopy galia farmakokinetika MeSH
- receptor cholecystokininu B metabolismus MeSH
- železité sloučeniny farmakokinetika MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Cílem naší studie bylo zjistit frekvenci výskytu destickových polymor?smu, které jsou spojovány s aterotrombogenezí u zdravých osob stredního veku v ceské populaci. Metody: K zjištení frekvence polymor?smu GP IIIa (HPA-1; rs5918), P2Y12 (H1/H2 haplotyp; rs2046934), P2Y12 (34C > T; rs6785930), COX-1 (-842A > G; rs10306114), PAR-1 (IVS -14A > T; rs168753), GP VI (13254T > C; rs1613662) a GP Ia (807C > T; rs1126643) bylo provedeno anonymní testování DNA od 1 450 dárcu krve pomocí PCR (polymerázové retezové reakce) a analýzy krivky tání na analyzátoru LightCycler 480 (ROCHE). Výsledky: V testované skupine byly zjišteny tyto frekvence; GP IIIa (HPA-1): 27,38 % heterozygotu, 3,66 % homozygotu a 68,97 % „wild type“ homozygotu; P2Y12 (H1/H2 haplotyp): 25,93 % heterozygotu, 2,62 % homozygotu a 71,45 % „wild type“ homozygotu; P2Y12 (34C > T): 44,03 % heterozygotu, 9,69 % homozygotu a 46,28 % „wild type“ homozygotu; COX-1 (-842A > G): 12,00 % heterozygotu, 0,41 % homozygotu a 87,59 % „wild type“ homozygotu; PAR-1 (IVS -14A > T): 28,55 % heterozygotu, 4,41 % homozygotu a 67,03 % „wild type“ homozygotu; GP VI (13254T > C): 21,45 % heterozygotu, 1,79 % homozygotu a 76,76 % „wild type“ homozygotu; GP Ia (807C > T): 47,27 % heterozygotu, 16,70 % homozygotu a 36,02 % „wild type“ homozygotu. Záver: V Ceské republice byla zjištena 10–40% frekvence testovaných polymor?smu desticek v populaci zdravých osob stredního veku, které jsou spojovány s náchylností k aterotrombogenezi nebo nedostatecnému úcinku protidestickové lécby.
The aim of our study was to assess the frequencies of platelet gene polymorphisms associated with atherothrombogenesis in the healthy middle-aged Czech population. Methods: Anonymous testing of 1,450 blood donor DNA by PCR followed by melting curve analysis using a LightCycler 480 analyzer (ROCHE) were used to determine the frequencies of GP IIIa (HPA-1; rs5918), P2Y12 (H1/H2 haplotyp; rs2046934), P2Y12 (34C > T; rs6785930), COX-1 (-842A > G; rs10306114), PAR-1 (IVS -14A > T; rs168753), GP VI (13254T > C; rs1613662) and GP Ia (807C > T; rs1126643) polymorphisms. Results: The frequencies of GP IIIa (HPA-1) were: 27.38% of heterozygotes, 3.66% of homozygotes, and 68.97% of “wild type“ homozygotes; frequencies of P2Y12 (H1/H2 haplotype) were: 25.93% of heterozygotes, 2.62% of homozygotes, and 71.45% of “wild type“ homozygotes; frequencies of P2Y12 (34C > T) were: 44.03% of heterozygotes, 9.69% of homozygotes and 46.28% of “wild type“ homozygotes; frequencies of COX-1 (-842A > G) were: 12.00% of heterozygotes, 0.41% of homozygotes, and 87.59% of “wild type“ homozygotes; frequencies of PAR-1 (IVS -14A > T) were: 28.55% of heterozygotes, 4.41% of homozygotes, and 67.03% of “wild type“ homozygotes, frequencies of GP VI (13254T > C) were: 21.45% of heterozygotes, 1.79% of homozygotes, and 76.76% of “wild type“ homozygotes; frequencies of GP Ia (807C > T) were: 47.27% of heterozygotes, 16.70% of homozygotes, and 36.02% of “wild type“ homozygotes in the tested group of healthy persons. Conclusion: The frequencies of tested platelets gene polymorphisms implicated in in=uencing susceptibility to atherothrombogenesis, or to failure of antiplatelet therapy, were 10–40% in the healthy middle-aged Czech population.
- MeSH
- agregace trombocytů fyziologie MeSH
- ateroskleróza etiologie genetika MeSH
- dospělí MeSH
- epidemiologické studie MeSH
- inhibitory agregace trombocytů metabolismus MeSH
- integrin alfaVbeta3 genetika MeSH
- integrin beta3 genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- polymorfismus genetický MeSH
- trombocytový glykoproteinový komplex IIb-IIIa genetika MeSH
- trombóza etiologie genetika MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- Geografické názvy
- Česká republika MeSH
- MeSH
- bisfosfonáty farmakologie terapeutické užití MeSH
- hodnocení léčiv MeSH
- integrin alfaVbeta3 antagonisté a inhibitory MeSH
- kalcitonin terapeutické užití MeSH
- kathepsiny antagonisté a inhibitory MeSH
- kombinovaná farmakoterapie MeSH
- lidé MeSH
- ligand RANK antagonisté a inhibitory MeSH
- monoklonální protilátky terapeutické užití MeSH
- organokovové sloučeniny terapeutické užití MeSH
- parathormon terapeutické užití MeSH
- postmenopauzální osteoporóza farmakoterapie MeSH
- remodelace kosti účinky léků MeSH
- selektivní modulátory estrogenních receptorů farmakologie terapeutické užití MeSH
- Check Tag
- lidé MeSH
