Micellar and Antibody-Targeted Polymer Therapeutics

Summary: Synthesis, physicochemical and some biological properties of new actively targeted antibody-containing and passively targeted micellar polymer - doxorubicin conjugates were investigated. Polymer precursors used for the synthesis of the conjugates were based on semitelechelic N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers with reactive groups situated at the polymer chain end or on multivalent copolymer with groups randomly distributed along the polymer backbone. Micellar HPMA-copolymer-based pharmaceuticals were prepared by self-assembly of copolymer–doxorubicin conjugates containing hydrophobic cholesterol ligands attached to the copolymer via hydrolytically degradable spacer. pH-Controlled release of cholesterol derivative is a key-point for disintegration of the micellar drug carrier after delivering the drug to the tumor tissue. Synthesis of star antibody-targeted polymer conjugates takes advantage of reduction of disulfide bridges in antibody with dithiothreitol followed by conjugation with the semitelechelic copolymer thus avoiding modification of the binding site in the antibody for its antigen. Both conjugates differing in their molecular architecture and mechanism of action are promising candidates for in vivo antitumor therapy.     

Graft and RAFT Reactive Macro Reagents: Bis-[copoly-(divinyl ether-alt-maleic anhydride)]-trithiocarbonate

The alternating cyclocopolymer of maleic anhydride with divinyl ether (MADVE) hydrolyzate, as mimicker of furan related and anionic residues alternation in nucleic acids (NA) backbone, is immune stimulating agonist and competitive antagonist for viral genome NA interventions. The targeted pre-modification of MADVE by antiviral vectors via grafting to the MA anhydride residues before hydrolysis led to more potent and promising antiviral inhibitors. To develop the MADVE capacity for novel modifications we applied the reversible addition – fragmentation chain transfer (RAFT) technique using dibenzyl trithiocarbonate as a RAFT agent. The insertion of trithiocarbonate unit in polymeric chain provided a pseudo living RAFT-polymerization, resulting in: 1) the effective control of polymerization degree (increased with time and conversion), and 2) the narrow dispersive (PDI = 1.1–1.2) products MADVE-S-CS-S-MADVE yield. These products can be used as novel polymeric RAFT-agents for synthesis of new block-copolymers MADVE-(block)-CS₃-(block)-MADVE, for instance with polystyrene blocks. Combined together the RAFT- and graft- reactivity allows both modify the polymer backbone (RAFT-synthesis) and regulate the side groups or branches (grafted to MADVE moieties) with final hydrolysis of unused anhydride units to acidic polyelectrolyte derivatives. This plural reactive capacity of the obtained macro reagents essentially enhances their potential as platform for purposed synthesis of novel (bio-) functional polymeric compounds.     

Syntheses of hydrophobically modified cationic hydrogels by copolymerization of alkyl substituted diallylamine monomers and their use as bile acid sequestrants

A novel class of cationic hydrogels bearing cyclic structures along the polymer backbone has been prepared and these hydrogels have been evaluated as bile acid sequestrants in vivo. The polymeric gels were prepared by the crosslinking cyclocopolymerization of hydrophobically modified dialkyldiallylammonium salts in the presence of different multifunctional crosslinking monomers. The roles of various crosslinking monomers, concentrations of crosslinking monomers, as well as alkyl chain length of dialkyldiallylammonium monomers on the physicochemical properties of the resulting hydrogels have been evaluated. From these preliminary in vivo studies it appears that these cationic hydrogels are excellent bile acid sequestrants and promising cholesterol lowering agents.     

Innovational combination of hetero-bifunctional N-PEG quinoline scaffolds derivatives with improved anticancer activity against breast and colon cancer cell lines and P-glycoprotein,cytochrome p450 enzyme activity prediction

Polyethylene glycol (PEG) is a polymer that is widely used as a carrier for drug delivery systems (DDS). A library of N-PEGylated quinoline derivatives of PEG molecular weight 200 was prepared rapidly after the activation of PEGs using maleic anhydride. Quinoline with a polymer backbone is essential as new material. PEG is a water-soluble nonionic polymer approved by food and drug organizations for medicine applications. Because of its nontoxic grapheme, it is widely utilized in numerous biochemical, cosmetic, pharmaceutical, and industrialized applications. The modern SwissADME is a web tool that stretches free admittance to a pool of hasty, yet solid, clarifying models for physicochemical properties, pharmacokinetics, and therapeutic science. The present facile synthetic strategy can be a practical approach for incorporating polymeric carriers conjugated with drug moieties, either in the backbone of the polymer or as a terminal and pendant group on the polymer chains.     

Polymeric photosensitizer prodrugs for photodynamic therapy

A targeting strategy based on the selective enzyme-mediated activation of polymeric photosensitizer prodrugs (PPP) within pathological tissue has led to the development of agents with the dual ability to detect and treat cancer. Herein, a detailed study of a simple model system for these prodrugs is described. We prepared "first-generation" PPP by directly tethering the photosensitizer (PS) pheophorbide a to poly-(L)-lysine via epsilon amide links and observed that by increasing the number of PS on a polymer chain, energy transfer between PS units improved leading to better quenching efficiency. Fragmentation of the PPP backbone by trypsin digestion gave rise to a pronounced fluorescence increase and to more efficient generation of reactive oxygen species upon light irradiation. In vitro tests using the T-24 bladder carcinoma cell line and ex vivo experiments using mouse intestines illustrated the remarkable and selective ability of these PPP to fluoresce and induce phototoxicity upon enzymatic activation. This work elucidated the basic physicochemical parameters, such as water solubility and quenching/activation behavior, required for the future elaboration of more adaptable "second-generation" PPP, in which the PS is tethered to a proteolytically stable polymer backbone via enzyme-specific peptide linkers. This polymer architecture offers great flexibility to tailor make the PPP to target any pathological tissue known to over-express a specific enzyme.     

Antitumor Drug Delivery Modulated by A Polymeric Micelle with an Upper Critical Solution Temperature

Thermally sensitive polymeric nanocarriers were developed to optimize the release profile of encapsulated compounds to improve treatment efficiency. However, when referring to thermally sensitive polymeric nanocarriers, this usually means systems fabricated from lower critical solution temperature (LCST) polymers, which have been intensively studied. To extend the field of thermally sensitive polymeric nanocarriers, we for the first time fabricated a polymeric drug delivery system having an upper critical solution temperature (UCST) of 43 °C based on an amphiphilic polymer poly(AAm‐co‐AN)‐g‐PEG. The resulting polymeric micelles could effectively encapsulate doxorubicin and exhibited thermally sensitive drug release both in vitro and in vivo. A drastically improved anticancer efficiency (IC₅₀ decreased from 4.6 to 1.6 μg mL^?1, tumor inhibition rate increased from 55.6 % to 92.8 %) was observed. These results suggest that UCST‐based drug delivery can be an alternative to thermally sensitive LCST‐based drug delivery systems for an enhanced antitumor efficiency.     

Mannan-Based Nanodiagnostic Agents for Targeting Sentinel Lymph Nodes and Tumors

Early detection of metastasis is crucial for successful cancer treatment. Sentinel lymph node (SLN) biopsies are used to detect possible pathways of metastasis spread. We present a unique non-invasive diagnostic alternative to biopsy along with an intraoperative imaging tool for surgery proven on an in vivo animal tumor model. Our approach is based on mannan-based copolymers synergistically targeting: (1) SLNs and macrophage-infiltrated solid tumor areas via the high-affinity DC-SIGN (dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin) receptors and (2) tumors via the enhanced permeability and retention (EPR) effect. The polymer conjugates were modified with the imaging probes for visualization with magnetic resonance (MR) and fluorescence imaging, respectively, and with poly(2-methyl-2-oxazoline) (POX) to lower unwanted accumulation in internal organs and to slow down the biodegradation rate. We demonstrated that these polymer conjugates were successfully accumulated in tumors, SLNs and other lymph nodes. Modification with POX resulted in lower accumulation not only in internal organs, but also in lymph nodes and tumors. Importantly, we have shown that mannan-based polymer carriers are non-toxic and, when applied to an in vivo murine cancer model, and offer promising potential as the versatile imaging agents.     

Distribution of functional groups grafted onto an ethylene-propylene copolymer

A theoretical model, based on the binomial (Bernoullian) distribution function, was employed for the prediction of functional group distribution in an ethylene-propylene copolymer randomly grafted by maleic anhydride. Using the experimentally determined graft level and molecular weight distribution function, the fraction of polymer molecules with given number of functional groups was calculated. The result was checked experimentally by a fluorescence method based on the excimer formation of pyrene fluorophores attached to the anhydride pendants. The time-resolved fluorescence from the pyrene-labeled copolymer yielded the fraction of polymer molecules with a single functional group. The fraction of singly labeled molecules was compared to the calculated functional group distribution and a reasonable agreement was found between the two. The distribution of grafted maleic anhydride was found to be apparently random among polymer molecules. The distribution of distances was calculated between randomly attached consecutive functional groups along the polymer backbone also. The result indicated that the distance distribution function (similar to a decaying exponential) is dominated by short distances. © 1996 John Wiley & Sons, Inc.     

Cyclodextrin sidechain polyesters — synthesis and inclusion of adamantan derivatives

An efficient synthesis of a cyclodextrin polymer by a polymer-analogous reaction of lithium β-cyclodextrinate with poly[(N-vinyl-2-pyrrolidinone)-co-(maleic anhydride)] is described. Because cyclodextrin polymer is highly water-soluble, its binding of guests, like 1-adamantanamine and l-adamantanecarboxylic acid, could be investigated by titration microcalorimetry. All cyclodextrin moieties are accessible by the guest within the polymer, but binding constants are slightly lower than those for native β-cyclodextrin. Binding constants are influenced by Coulomb interactions between the guest and the anionic polymer backbone.     

Self‐Assembled Polyprodrug Amphiphile for Subcutaneous Xenograft Tumor Inhibition with Prolonged Acting Time In Vivo

Polymeric drug delivery system termed as “polyprodrug amphiphile” poly(2‐methylacryloyloxyethyl phosphorylcholine)‐b‐poly(10‐hydroxy‐camptothecin methacrylate (pMPC‐b‐pHCPT) is developed for the prolonged‐acting cancer therapy. It is obtained by two‐step reversible addition–fragmentation chain transfer polymerization of zwitterionic monomer MPC and an esterase‐responsive polymerizable prodrug methacrylic anhydride–CPT, respectively. This diblock polymer is composed of both antifouling (pMPC) and bioactive (pHCPT) segments and the drug is designed as a building block to construct the polymer skeleton directly. Due to its distinct amphiphilicity, the polymer can self‐assemble into micelles with different dynamic sizes by facilely tuning the ratio of MPC/HCPT under physiological conditions. The outer pMPC shell is superhydrophilic to form dense hydrate layer preventing the nanosystem from unwanted nonspecific protein adsorption, which is the main lead cause of the rapid clearance of nanoparticles in vivo, thus facilitating the accumulation of drugs in tumor sites via enhanced permeability and retention effect. The configuration of the polyprodrug amphiphile is confirmed by several measurements. The resistance to albumin adsorption, prolonged plasma retention time, accumulation in tumor sites, and anticancer activity of the micelles is also investigated in vitro and in vivo. This novel amphiphile can be expected as a promising agent for the passive targeted prolonged‐acting cancer therapy.     

生物医用高分子在癌症药物治疗中的应用

利用生物医用高分子作载体,化学结合或物理包裹抗肿瘤化学药物、生物工程药物和放射药物,制剂通过植入或靶向运输至肿瘤区域。可增强药物在运输及吸收过程中的稳定性,提高药物的生物利用度,药物以一定速率从制剂中缓慢释放,可简化服用程序,在肿瘤区域维持较高的药物浓度,同时降低药物对全身的毒副作用。本文综述了生物医用高分子在高分子导向药物、抗肿瘤药物聚合物微球制剂,植入制剂以及肿瘤栓塞治疗中的应用。     

Polymer materials for constructing therapeutical nanoparticles in photothermal therapy

Photothermal therapy (PTT) ablates tumors by thermal effects of photothermal agents (PTAs), and attracts wide attention due to the non-invasive characteristic. The ideal PTAs are expected to have high photothermal conversion effect under NIR irradiation, as well as targeting abilities and good biocompatibility satisfying the need of application in vivo. Nanoparticles (NPs) are commonly used as anti-tumor materials, and plenty of researches on therapeutical NPs for PTT treatment have been developed. Among various building blocks for photothermal NPs, polymer materials for biomedical applications have great advantages due to their negligible toxicity, flexibility for functional modification, and ability to integrate multiple therapeutic strategies. This review focuses on the polymer materials utilized in photothermal NP designing, including their application as excellent carriers and powerful PTAs with great PTT effects. Furthermore, the synergy therapy based on polymeric nanoplatform for enhancing PTT therapeutic efficiency will be introduced.     

Properties of a nanocomposite polymer electrolyte from an amorphous comb-branch polymer and nanoparticles

Three fully amorphous comb-branch polymers based on poly(styrene-co-maleic anhydride) as a backbone and poly(ethylene glycol) methyl ether of different molecular weights as side chains were synthesized. SiO₂ nanoparticles of various contents and the salt LiCF₃SO₃ were added to these comb-branch polymers to obtain nanocomposite polymer electrolytes. The thermal and transport properties of the samples have been characterized. The maximum conductivity of 2.8×10^–4 S cm^–1 is obtained at 28 °C. In the system the longer side chain of the comb-branch polymer electrolyte increases in ionic conductivity after the addition of nanoparticles. To account for the role of the ceramic fillers in the nanocomposite polymer electrolyte, a model based on a fully amorphous comb-branch polymer matrix in enhancing transport properties of Li⁺ ions is proposed.     

Poly-2-vinylfuran,synthesis, characterization,and diels-alder reaction with maleic anhydride

Poly-2-vinylfuran, synthesized by free-radical polymerization of 2-vinylfuran, was characterized by nuclear magnetic resonance (NMR) and infrared (IR) spectroscopy. The pendant furan rings on the polymer backbone were then used as the diene component of a Diels-Alder reaction with maleic anhydride. The juxtaposition of the furan rings at first suggested an “avalanche” Diels-Alder reaction, in which the product of one cyclization would be the reactant of the next. A lack of polymer stereoregularity and the reversibility of the Diels-Alder reaction, however, prevented its formation. On the other hand, when the dienophile was used in a 1:1 molar ratio with respect to furan the smooth reaction produced a new polymer, the maleic anhydride adduct of poly-2-vinylfuran, which characterized by NMR and IR spectroscopy, was air stable and soluble in a number of solvents up to 70% transformation. When heated to 160°C the polymer reverted to maleic anhydride and somewhat decomposed poly-2-vinylfuran.     

Reversibly self-assembled pH-responsive PEG-p(CL-g-TMC) polymersomes

Polymersomes have gained much interest within the biomedical field as drug delivery systems due to their ability to transport and protect cargo from the harsh environment inside the body. For an improved drug efficacy, control over cargo release is however also an important factor to take into account. An often employed method is to incorporate pH sensitive groups in the vesicle membrane, which induce disassembly and content release when the particles have reached a target site in the body with the appropriate pH, such as the acidic microenvironment of tumor tissue or the endosome. In this paper, biodegradable poly(ethylene glycol)-poly(caprolactone-gradient-trimethylene carbonate)-based polymeric vesicles have been developed with disassembly features at mild acidic conditions. Modifying the polymer backbone with imidazole moieties results in vesicle disassembly upon protonation due to the lowered pH. Furthermore, upon increasing the pH efficient re-assembly into vesicles is observed due to the switchable amphiphilic nature of the polymer. When this re-assembly process is conducted in presence of cargo, enhanced encapsulation is achieved. Furthermore, the potency of the polymeric system for future biomedical applications such as adjuvant delivery is demonstrated.     

Synthesis and Preliminary Biological Evaluation of Novel Functionalised Poly(ethylene glycol)‐block‐poly(ester‐carbonate) Copolymers as Biodegradable Polymeric Carriers

Polymer conjugates of anticancer drugs containing bound active moieties are used to improve pharmacokinetics by prolonging antitumour activity and decreasing protein immunogenicity. The first generation of polymer conjugates, N‐(2‐hydroxylpropyl)methacrylamide copolymer or poly(ethylene glycol) conjugates, cannot be considered optimised since their polymer chain lacks biodegradability. The polymeric system proposed here is based on high molecular weight water‐soluble functionalised polyesters containing fumarate or maleate units linked by poly(ethylene glycol) segments by means of carbonate bonds. It shows the following features: a) high molecular weight; b) biodegradable backbone (rate of degradation was determined viscometrically in vitro, under conditions mimicking physiological conditions and c) water solubility. Also the biocompatibility of the polymer and its precursors were assessed using a B16F10 murine melanoma cell line and a haemolysis assay.     

A study of the ring size in the copolymer of divinyl ether and maleic anhydride by H-NMR spectroscopy

The well-known alternating 1:2 cyclocopolymer of divinyl ether (DVE) and maleic anhydride (MA) possesses a wide spectrum of biological activities, including antitumor. Recent research on the structure of a variety of cyclopolymers has raised a question about the ring size of this cyclocopolymer. In this article we report on an extensive spectroscopic study of its structure. By use of deuterated monomers the H-NMR peaks at δ 2.31, 3.47, 4.06, and 4.49 ppm with an area ratio of 2:1:1:1 were assigned to the hydrogens of methylenes, methines on the backbone anhydride unit, methines on the ring anhydride unit, and methines adjacent to oxygen on the cyclic ether ring, respectively. By examination of the possible isomeric structures of the bicyclic ring, the splitting of each peak group was further assigned for cis and trans disubstitutions on the anhydride unit. The splitting pattern from the 300-MHz NMR spectrum of the DVE-2,3-dideuteriomaleic anhydride (DMA) copolymer confirmed the unsymmetrical ring structure. ¹³C-NMR spectra were consistent with the conclusion from the H-NMR spectra. A chair-form, six-membered ring with predominantly trans geometry in the anhydride ring was assigned to the structure of DVE–MA copolymer. On the basis of little or no change in the ¹³C-NMR spectra of the copolymers prepared at different temperatures it was concluded that there was no significant change in structure with temperature. This led to the assignment of the energetically favored, six-membered ring structure to the copolymer prepared under these conditions. A mechanism for cyclocopolymerization, based on the HOMO–LUMO interaction of the comonomers and the intramolecular radical addition on the preoriented double bond, was proposed. This mechanism leads to the formation of the six-membered ring structure of the copolymer as the only product. A ¹³C-NMR study of the structure of the copolymer prepared in chloroform by Kunitake and Tsukino is being published as a companion article.     

SYNTHESIS AND CHARACTERIZATION OF POLYMER DERIVATIVES OF cis-PLATINUM COMPLEXES

Cis-platinum complexes used as antitumor drugs were attached onto preformed polymers toreduce the toxicity. Either alternating copolymer of styrene and maleic anhydride (CPSMA) orpolyacrylic acid (PAA) is readily dissolved in alkaline solution. The cis-platinum complexes areeasily converted into water soluble species when treated by silver nitrate in water. The expectedproducts could be obtained when the two aqueous solutions were brought together. The productswere characterized by elementary analysis, IR and XPS. The polymer derivatives could exchangeligands with nucleophilic groups in biological environment and also exhibit antineoplastic activity.     

肝组织特异性非离子型高分子磁共振造影剂的制备及性能评价

L-酪氨酸甲酯与DTPA双N-羟基琥珀酰亚胺活性酯(SuO-DTPA-OSu)反应,合成了含L-酪氨酸甲酯残基的DTPA单N-羟基琥珀酰亚胺活性酯(SuO-DTPA-Tyr).以EDC/NHS为媒介,通过一步反应将不同量的乳糖酸偶联到α,β-聚[(2-氨乙基)-L-天冬酰胺]上,然后将含有苯环的结构的SuO-DTPA-Tyr与联有D-半乳糖的α,β-聚[(2-氨乙基)-L-天冬酰胺]反应,合成了3种含糖量不同的大分子配体,并制备了其Gd(III)螯合物.结果表明,EDC/NHS方法操作简便,产率高且易于提纯;而SuO-DTPA-Tyr上L-酪氨酸甲酯残基的引入,不仅实现了高分子造影剂的非离子化,同时L-酪氨酸甲酯残基上的苯环结构也可以方便核磁氢谱的指认.大分子配体的细胞毒性随含糖量降低而增加,但均小于同浓度聚(L-赖氨酸)的毒性;大分子螯合物的细胞毒性与商用小分子造影剂的细胞毒性相当,但其弛豫率明显高于小分子造影剂的弛豫率;大分子螯合物在小白鼠肝部有比商用小分子造影剂(钆喷酸葡胺)更好的成像增强效果及更长的停留时间,且在注射后前6h,含D-半乳糖酸残基的大分子螯合物比不含D-半乳糖酸残基的大分子螯合物在小白鼠肝部的代谢速率慢,并呈现出更清晰的造影效果.     

Studies on the Cure Reaction and Relationship between Network Structure/Thermal Properties of Styrene Copolymers Based on Adypic/Sebacic Acid Modified Unsaturated (Epoxy) Polyesters

The studies on the relationship between network structure/thermal properties of styrene copolymers based on adypic/sebacic acid modified unsaturated (epoxy) polyesters cured using different hardeners as well as the course of the cure reaction of polyesters with styrene have been presented. The adypic/sebacic acid modified unsaturated polyesters (UP) prepared from 4-cyclohexene-1,2-dicarboxylic anhydride (THPA), maleic anhydride (MA), adypic acid (AA) or sebacic acid (SA) and ethylene glycol (EG) and their epoxy derivatives: adypic/sebacic acid modified unsaturated epoxy polyesters (UEP) were subjected to the cure process with styrene using diacyl peroxide: benzoyl peroxide (BPO) or the mixture of BPO/suitable acid anhydride: 4-cyclohexene-1,2-dicarboxylic anhydride (THPA) or glutaric anhydride (GA). Thermal properties were evaluated by means of DSC, TG and DMA analyses. It was proved that studied properties were significantly depended on polyester's structure and the type of applied curing system. Generally, higher values of E'_20°C, tgδ_max, E”, ν_e, IDT, T_k for styrene copolymers based on UEP were obtained. It was connected with more cross-linked polymer network structure due to the possible copolymerization reaction of carbon-carbon double bonds of polyester with styrene and additional polyaddition of epoxy to anhydride groups or thermal curing of epoxy groups. The additional connections between polyester's chains led to obtain more stiff and thermal stable polymeric materials. Moreover, the increase of saturated aliphatic acid's chain length in polyester backbone caused the decrease of E'_20°C, tgδ_max, E”, ν_e, IDT, T_k values of styrene copolymers. It suggested that copolymers based on polyesters prepared from acid containing more methylene groups in their structure were characterized by more flexible polymer network due to the “laxity” effect of aliphatic chains.