hydrodynamic radius
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Small molecule Toll-like receptor-7 and -8 agonists (TLR-7/8a) can be used as vaccine adjuvants to induce CD8 T cell immunity but require formulations that prevent systemic toxicity and focus adjuvant activity in lymphoid tissues. Here, we covalently attached TLR-7/8a to polymers of varying composition, chain architecture and hydrodynamic behavior (∼300 nm submicrometer particles, ∼10 nm micelles and ∼4 nm flexible random coils) and evaluated how these parameters of polymer-TLR-7/8a conjugates impact adjuvant activity in vivo. Attachment of TLR-7/8a to any of the polymer compositions resulted in a nearly 10-fold reduction in systemic cytokines (toxicity). Moreover, both lymph node cytokine production and the magnitude of CD8 T cells induced against protein antigen increased with increasing polymer-TLR-7/8a hydrodynamic radius, with the submicrometer particle inducing the highest magnitude responses. Notably, CD8 T cell responses induced by polymer-TLR-7/8a were dependent on CCR2+ monocytes and IL-12, whereas responses by a small molecule TLR-7/8a that unexpectedly persisted in vaccine-site draining lymph nodes (T1/2 = 15 h) had less dependence on monocytes and IL-12 but required Type I IFNs. This study shows how modular properties of synthetic adjuvants can be chemically programmed to alter immunity in vivo through distinct immunological mechanisms.
- MeSH
- adjuvancia imunologická aplikace a dávkování chemie farmakologie MeSH
- aktivace lymfocytů * MeSH
- buněčné linie MeSH
- CD8-pozitivní T-lymfocyty účinky léků imunologie MeSH
- cytokiny metabolismus MeSH
- hydrodynamika MeSH
- kultivované buňky MeSH
- micely * MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- toll-like receptor 7 agonisté MeSH
- toll-like receptor 8 agonisté MeSH
- vazba proteinů MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Intramural MeSH
New amphiphilic diblock polymer nanotherapeutics serving simultaneously as a drug delivery system and an inhibitor of multidrug resistance were designed, synthesized, and evaluated for their physico-chemical and biological characteristics. The amphiphilic character of the diblock polymer, containing a hydrophilic block based on the N-(2-hydroxypropyl)methacrylamide copolymer and a hydrophobic poly(propylene oxide) block (PPO), caused self-assembly into polymer micelles with an increased hydrodynamic radius (Rhof approximately 15nm) in aqueous solutions. Doxorubicin (Dox), as a cytostatic drug, was bound to the diblock polymer through a pH-sensitive hydrazone bond, enabling prolonged circulation in blood, the delivery of Dox into a solid tumor and the subsequent stimuli-sensitive controlled release within the tumor mass and tumor cells at a decreased pH. The applicability of micellar nanotherapeutics as drug carriers was confirmed by an in vivo evaluation using EL4 lymphoma-bearing C57BL/6 mice. We observed significantly higher accumulation of micellar conjugates in a solid tumor because of the EPR effect compared with similar polymer-drug conjugates that do not form micellar structures or with the parent free drug. In addition, highly increased anti-tumor efficacy of the micellar polymer nanotherapeutics, even at a sub-optimal dose, was observed. The presence of PPO in the structure of the diblock polymer ensured, during in vitro tests on human and mouse drug-sensitive and resistant cancer cell lines, the inhibition of P-glycoprotein, one of the most frequently expressed ATP-dependent efflux pump that causes multidrug resistance. In addition, we observed highly increased rate of the uptake of the diblock polymer nanotherapeutics within the cells. We suppose that combination of unique properties based on MDR inhibition, stimuli sensitiveness (pH sensitive activation of drug), improved pharmacokinetics and increased uptake into the cells made the described polymer micelle a good candidate for investigation as potential drug delivery system.
- MeSH
- akrylamidy aplikace a dávkování chemie farmakokinetika terapeutické užití MeSH
- chemorezistence účinky léků MeSH
- doxorubicin aplikace a dávkování chemie farmakokinetika terapeutické užití MeSH
- hydrofobní a hydrofilní interakce MeSH
- lidé MeSH
- micely * MeSH
- mnohočetná léková rezistence účinky léků MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- nádory farmakoterapie metabolismus patologie MeSH
- nosiče léků aplikace a dávkování chemie farmakokinetika terapeutické užití MeSH
- polymery aplikace a dávkování chemie farmakokinetika terapeutické užití MeSH
- propylenglykoly aplikace a dávkování chemie farmakokinetika terapeutické užití MeSH
- protinádorová antibiotika aplikace a dávkování chemie farmakokinetika terapeutické užití MeSH
- tumor burden účinky léků MeSH
- uvolňování léčiv MeSH
- viabilita buněk účinky léků MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Mutual diffusion coefficients, D, were determined for aqueous solutions of sodium hyaluronate (NaHy) at 25 °C and concentrations ranging from 0.00 to 1.00 g·dm(-3) using the Taylor dispersion technique. From these experimental data, it was possible to estimate some parameters, such as the hydrodynamic radius Rh, and the diffusion coefficient at infinitesimal concentration, D0, of hyaluronate ion, permitting us to have a better understanding of the structure of these systems of sodium hyaluronate in aqueous solutions. The additional viscosity measurements were done and Huggins constant, kH, and limiting viscosity number, [η], were computed for interaction NaHy/water and NaHy/NaHy determination.
The molecular weight and molecular architecture of soluble polymer drug carriers significantly influence the biodistribution and anti-tumour activities of their doxorubicin (DOX) conjugates in tumour-bearing mice. Biodistribution of N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-DOX conjugates of linear and star architectures were compared in EL4 T-cell lymphoma-bearing mice. Biodistribution, including tumour accumulation, and anti-tumour activity of the conjugates strongly depended on conjugate molecular weight (MW), polydispersity, hydrodynamic radius (R(h)) and molecular architecture. With increasing MW, renal clearance decreased, and the conjugates displayed extended blood circulation and enhanced tumour accumulation. The linear conjugates with flexible polymer chains were eliminated by kidney clearance more quickly than the highly branched star conjugates with comparable MWs. Interestingly, the data suggested different mechanisms of renal filtration for star and linear conjugates. Only star conjugates with MWs below 50,000g.mo(-1) were removed by kidney filtration, while linear polymer conjugates with MWs near 70,000g.mol(-1), exceeding the generally accepted limit for renal elimination, were detected in the urine 36-96h after injection. Additionally, survival of tumour-bearing mice was strongly dependent on molecular weight and polymer conjugate architecture. Treatment of mice with the lower MW conjugate at a dose of 10mg DOX eq./kg resulted in 12% long-term surviving animals, while treatment with the corresponding star conjugate enabled 75% survival of animals.
- MeSH
- akrylamidy chemie farmakokinetika terapeutické užití MeSH
- doxorubicin chemie farmakokinetika terapeutické užití MeSH
- Kaplanův-Meierův odhad MeSH
- lymfom farmakoterapie MeSH
- metabolická clearance MeSH
- molekulární struktura MeSH
- molekulová hmotnost MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- protinádorová antibiotika chemie farmakokinetika MeSH
- stabilita léku MeSH
- tkáňová distribuce MeSH
- transplantace nádorů MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
To assist in overcoming the inherent instability of nucleic acid-containing polyplexes in physiological solutions, we have here set out to develop removable nanocoatings for modifying the surface of siRNA-based nanoparticles. Here, N-(2-hydroxypropyl)methacrylamide (HPMA) based copolymers containing carbonylthiazolidine-2-thione (TT) reactive groups in their side chains bound via disulfide spacers to the polymeric backbone were synthesized, and these copolymers were used to coat the surface of polyplexes formed by the self-assembly of anti-Luciferase siRNA with the polycations polyethylene imine (PEI) and poly(HPMA)-grafted poly(l-lysine) (GPL). The coating process was monitored by analyzing changes in the weight-average molecular weight (M(w)), the hydrodynamic radius (R(h)), and the zeta-potential (ζ) of the polyplexes, using both static (SLS) and dynamic (DLS) light scattering methods. The outlined methods resulted in the attachment of, on average, 28 polymer molecules to the surface of the polyplexes, forming a ∼5-nm-thick hydrophilic stealth coating. Initial efforts to develop RGD-targeted coated polyplexes are also described. Atomic force microscopy (AFM) showed an angular polyplex structure and confirmed the narrow size distribution of the coated nanoparticles. The stability of the polymer-coated and uncoated polyplexes was evaluated by gel electrophoresis and by turbidity measurements, and it was found that modifying the surface of the siRNA-containing polyplexes substantially improved their stability in physiological solutions. Using polymer-coated GPL-based polyplexes containing anti-Luciferase siRNA, we finally also obtained some initial proof-of-principle for time- and concentration-dependent target-specific gene silencing, suggesting that these systems hold significant potential for further in vitro and in vivo evaluation.
