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New class of squalene-based releasable nanoassemblies of paclitaxel, podophyllotoxin, camptothecin and epothilone A
Abstract
The present study reports the preparation of a novel class of squalene conjugates with paclitaxel, podophyllotoxin, camptothecin and epothilone A. The obtained compounds are characterized by a squalene tail that makes them able to self-assemble in water, and by a drug unit connected via a disulfide-containing linker to secure the release inside the cell. All the obtained compounds were effectively able to self-assemble and to release the parent drug in vitro. Disulfide-containing paclitaxel-squalene derivative showed a similar biological activity when compared to the free drug. Immunofluorescence assay shows that this squalene conjugate enters A549 cells and stain microtubule bundles. The results described herein pave the way for different classes of squalene-based releasable nanoassemblies.
... For several years we have been interested in using this kind of NPs to improve the properties of both anticancer and neuroprotective drugs [3][4][5][6][7][8][9][10][11][12][13][14]. We designed conjugates able to form NPs that can release the drug in cellular media, [3,8,11,14] hetero-NPs bearing Based on the above statements, we considered CBD for its potential dual activity (cytotoxic compound and self-assembly inducer) and to conjugate it to well-known tubulin binder drugs, N-desacetyl thiocolchicine (2), podophyllotoxin (3), and paclitaxel (4), through two different linkers, 5a and 5b (Figure 1). ...
... For several years we have been interested in using this kind of NPs to improve the properties of both anticancer and neuroprotective drugs [3][4][5][6][7][8][9][10][11][12][13][14]. We designed conjugates able to form NPs that can release the drug in cellular media, [3,8,11,14] hetero-NPs bearing Based on the above statements, we considered CBD for its potential dual activity (cytotoxic compound and self-assembly inducer) and to conjugate it to well-known tubulin binder drugs, N-desacetyl thiocolchicine (2), podophyllotoxin (3), and paclitaxel (4), through two different linkers, 5a and 5b (Figure 1). The synthesis and characterization of the planned conjugates and their ability to form self-assembled NPs are here reported. ...
... For several years we have been interested in using this kind of NPs to improve the properties of both anticancer and neuroprotective drugs [3][4][5][6][7][8][9][10][11][12][13][14]. We designed conjugates able to form NPs that can release the drug in cellular media, [3,8,11,14] hetero-NPs bearing Based on the above statements, we considered CBD for its potential dual activity (cytotoxic compound and self-assembly inducer) and to conjugate it to well-known tubulin binder drugs, N-desacetyl thiocolchicine (2), podophyllotoxin (3), and paclitaxel (4), through two different linkers, 5a and 5b (Figure 1). The synthesis and characterization of the planned conjugates and their ability to form self-assembled NPs are here reported. ...
Cannabidiol (CBD) is a biologically active compound present in the plants of the Cannabis family, used as anticonvulsant, anti-inflammatory, anti-anxiety, and more recently, anticancer drug. In this work, its use as a new self-assembly inducer in the formation of nanoparticles is validated. The target conjugates are characterized by the presence of different anticancer drugs (namely N-desacetyl thiocolchicine, podophyllotoxin, and paclitaxel) connected to CBD through a linker able to improve drug release. These nanoparticles are formed via solvent displacement method, resulting in monodisperse and stable structures having hydrodynamic diameters ranging from 160 to 400 nm. Their biological activity is evaluated on three human tumor cell lines (MSTO-211H, HT-29, and HepG2), obtaining GI50 values in the low micromolar range. Further biological assays were carried out on MSTO-211H cells for the most effective NP 8B, confirming the involvement of paclitaxel in cytotoxicity and cell death mechanism
... To form self-assembling NPs, the conjugate is usually formed by covalent coupling of the drug with biocompatible lipide moieties [9]. These molecules can be connected through smart linkers, which release the drug when an internal or external stimulus is applied, ensuring targeted delivery and controlled drug release [10]. ...
... Squalene is a lipidic moiety and a natural precursor of many steroids that spontaneously form self-assembling NPs in water, with an unusually high drug content. Since squalene is well tolerated, it makes it an ideal component of the drug delivery system [7,9,13]. Another attractive option commonly used as lipidic moieties is fatty acids due to their biodegradability and biosafety. ...
... In terms of size, it was observed that 12BzRoy-Sq was bigger than the Roy-OA NPs. The size of the 12BzRoy-Sq NPs was different from that previously observed for squalene-based heteronanoparticles [7,9], indicating that the larger nature of the 12BzRoy-Sq NPs could be due to the agglomeration in water; hence, these are not suitable for drug delivery. The zeta potential was negative in both cases, with the Roy-OA being more negative. ...
Different approaches have been reported to enhance penetration of small drugs through physiological barriers; among them is the self-assembly drug conjugates preparation that shows to be a promising approach to improve activity and penetration, as well as to reduce side effects. In recent years, the use of drug-conjugates, usually obtained by covalent coupling of a drug with bio-compatible lipid moieties to form nanoparticles, has gained considerable attention. Natural products isolated from plants have been a successful source of potential drug leads with unique structural diversity. In the present work three molecules derived from natural products were employed as lead molecules for the synthesis of self-assembled nanoparticles. The first molecule is the cyto-toxic royleanone 7α-acetoxy-6β-hydroxyroyleanone (Roy, 1) that has been isolated from hairy co-leus (Plectranthus hadiensis (Forssk.) Schweinf). ex Sprenger leaves in a large amount. This royleanone, its hemisynthetic derivative 7α-acetoxy-6β-hydroxy-12-benzoyloxyroyleanone (12BzRoy, 2) and 6,7-dehydroroyleanone (DHR, 3), isolated from the essential oil of thicket coleus (P. madagascariensis (Pers.) Benth.) were employed in this study. The royleanones were conjugated with squalene (sq), oleic acid (OA), and/or 1-bromododecane (BD) self-assembly inducers. Roy-OA, DHR-sq, and 12BzRoy-sq conjugates were successfully synthesized and characterized. The cyto-toxic effect of DHR-sq was previously assessed on three human cell lines: NCI-H460 (IC50 74.0 ± 2.2 µM), NCI-H460/R (IC50 147.3 ± 3.7 µM), and MRC-5 (IC50 127.3 ± 7.3 µM), and in this work Roy-OA NPs was assayed against Vero-E6 cells at different concentrations (0.05, 0.1, and 0.2 mg/mL). The cytotoxicity of DHR-sq NPs was lower when compared with DHR alone in these cell lines: NCI-H460 (IC50 10.3 ± 0.5 µM), NCI-H460/R (IC50 10.6 ± 0.4 µM), and MRC-5 (IC5016.9 ± 0.5 µM). The same results were observed with Roy-OA NPs against Vero-E6 cells as was found to be less cytotoxic than Roy alone in all the concentrations tested. From the obtained DLS results, 12BzRoy-sq assemblies were not in the nano range, although Roy-OA NP assemblies show a promising size (509.33 nm), Pdl (0.249), zeta potential (-46.2mV), and spherical morphology from SEM. In addition, these NPs had a low release of Roy at physiological pH 7.4 after 24 h. These results suggest the nano assemblies can act as prodrugs for the release of cytotoxic lead molecules. Citation: Ntungwe, E.; Domínguez-Martín, E.M.; Bangay, G.; Garcia, C.; Guerreiro, I.; Colombo, E.; Saraiva, L.; Díaz-Lanza, A.M.; Rosatella, A.; Alves, M.M.; et al. Self-Assembly Nanoparticles of Natural Bioactive Abietane Diterpenes. Int. J. Mol. Sci.
... Organic molecules, such as terpenes, polysaccharides, or polymeric chains with high biocompatibility, are widely used as inducers, which can spontaneously self-assemble into NPs in an aqueous medium without the need for using any additional surfactants to stabilize their colloid suspensions (Bildstein et al., 2011;Delplace et al., 2014). In some instances, connecting the drug and the inducer can be achieved by using a so-called linker moiety, which may further enhance the in vivo release of the therapeutic agent from its conjugate (Borrelli et al., 2014). During the synthetic preparation of bioconjugates, components are coupled together through covalent bonds that are hydrolysable in a biological environment (e.g., esters), so that such self-assembled NPs will typically act as pro-drugs (Cheetham et al., 2017). ...
... Self-assembly inducer squalene was functionalized and conjugated with sebacic acid following previously published procedures, which had allowed us the preparation of compound 5 (Borrelli et al., 2014;Maksimenko et al., 2014). An aliquot of 0.2 mmol of ecdysteroid 3 (112.2 ...
... Therefore, we prepared compound 5 following previously published strategies, by first transforming squalene to 1,1′,2-trisnorsqualene alcohol (Maksimenko et al., 2014). Then we conjugated the terminal hydroxyl moiety of this alcohol with sebacic acid, serving as linker, to yield compound 5 whose carboxylic moiety was required for the preparation of esters (Borrelli et al., 2014). ...
Several ecdysteroid acetonides act as adjuvant chemo-sensitizing agents against various cancer cell lines, and they can be formulated to self-assembling nanoparticle (NP) pro-drugs through a hydrolysable conjugation with squalene. In the bloodstream such squalenoylated nanoparticles dissolve into low-density lipoprotein (LDL) that allows targeting tissues containing high levels of LDL-receptors. In this work, ajugasterone C 2,3;20,22-diacetonide (3) and 11α-hydroxypoststerone 2,3-acetonide (4) were squalenoylated to obtain two new ecdysteroid pro-drugs (6 and 7) and their nano-assemblies (6NP and 7NP ). A complete NMR signal assignment of 6 and 7 was achieved. Interaction of compounds 3 and 4 with chemotherapeutics was studied by the Chou-Talalay method. Compound 3 showed strong synergism with doxorubicin on a multi-drug resistant lymphoma cell line. In contrast, its nanoassembly 6NP significantly decreased the cytotoxicity of doxorubicin on these MDR cells, strongly suggesting that at least the 2,3-acetonide group was cleaved by the acidic pH of lysosomes after endocytosis of the prodrug. Further, compound 4 acted in strong antagonism with paclitaxel on MCF-7 cells and its nanoassemby 7NP also protected MCF-7 cells from the effect of paclitaxel. Our results suggest that acid-resistant A-ring substitution would be crucial to design adjuvant antitumor squalenoylated ecdysteroid prodrugs. Additionally, our results may be considered as a serendipitous discovery of a novel way to deliver cytoprotective, adaptogen ecdysteroids to healthy tissues with upregulated LDL-R.
... In recent years we worked on anticancer drug-containing self-assembling drug conjugates that spontaneously form NAs in aqueous media [31]. We reported NAs composed by conjugate releasable compounds [32]; by single and dual drug fluorescent hetero-NAs [33,34], by dual drug hetero-NAs (cyclopamine/taxol [34], cyclopamine/doxorubicin [35], ecdysteroid/doxorubicin [36]), and by self-assembling conjugate dual drug NAs [37]. We prepared compounds containing a squalene [31][32][33][34][35] or a 4-(1,2-diphenylbut-1-en-1-yl)aniline tail [37,38] that leads to NAs ability to self-assemble in water. ...
... We reported NAs composed by conjugate releasable compounds [32]; by single and dual drug fluorescent hetero-NAs [33,34], by dual drug hetero-NAs (cyclopamine/taxol [34], cyclopamine/doxorubicin [35], ecdysteroid/doxorubicin [36]), and by self-assembling conjugate dual drug NAs [37]. We prepared compounds containing a squalene [31][32][33][34][35] or a 4-(1,2-diphenylbut-1-en-1-yl)aniline tail [37,38] that leads to NAs ability to self-assemble in water. We recently reported the assembly and characterization of squalene-thiocolchicine NAs that release cytotoxic, free thiocolchicine in cancer cells through a disulfide bond or a p-hydroxybenzyl moiety [38]. ...
... In recent years we worked on anticancer drug-containing self-assembling drug conjugates that spontaneously form NAs in aqueous media [31]. We reported NAs composed by conjugate releasable compounds [32]; by single and dual drug fluorescent hetero-NAs [33,34], by dual drug hetero-NAs (cyclopamine/taxol [34], cyclopamine/doxorubicin [35], ecdysteroid/doxorubicin [36]), and by selfassembling conjugate dual drug NAs [37]. We prepared compounds containing a squalene [31][32][33][34][35] or a 4-(1,2-diphenylbut-1-en-1-yl)aniline tail [37,38] that leads to NAs ability to self-assemble in water. ...
The disaccharide trehalose is an autophagy inducer, but its pharmacological application is severely limited by its poor pharmacokinetics properties. Thus, trehalose was coupled via suitable spacers with squalene (in 1:2 and 1:1 stoichiometry) and with betulinic acid (1:2 stoichiometry), in order to yield the corresponding nanolipid-trehalose conjugates 1-Sq-mono, 2-Sq-bis and 3-Be-mono. The conjugates were assembled to produce the corresponding nano-assemblies (NAs) Sq-NA1, Sq-NA2 and Be-NA3. The synthetic and assembly protocols are described in detail. The resulting NAs were characterized in terms of loading and structure, and tested in vitro for their capability to induce autophagy. Our results are presented and thoroughly commented upon.
... The introduction of a squalene tail was further exploited for the derivatization of known anticancer compounds such as paclitaxel, podophyllotoxin (16), camptothecin, epothilone A (17) and cyclopamine (18) with the inclusion of an alternative linker [40][41][42]. In this study, the squalene moiety and the active compound were connected by a disulfide-containing linker able to guarantee the drug release inside the cell. ...
... In vivo evaluation demonstrated that NPs show a similar ovarian tumor growth inhibition to paclitaxel and induce no bodyweight loss. The conjugation of paclitaxel with a squalenic tail through a disulfide-containing linker led to the formation of NPs that were biologically evaluated on MCF-7 cells [40]. It was demonstrated that the conjugation to squalene decreased the cytotoxicity of the drug toward MCF-7 cells but it was confirmed that the disulfide bond improved the release of the toxic moiety. ...
... In addition, the functionalization of different known drugs with a cleavable linker and a proper lipophilic chain that induces self-assembly and the generation of hetero-NPs could be exploited for personalized treatment of different types of diseases [42][43][44]. The same strategy can be used for the preparation of fluorescent hetero-NPs that can help follow the dynamics of the delivery and it might be possible to trace these NPs in the cells by imaging [26,40]. ...
... [2,3] The use of nanotechnology can overcome some problems of drug delivery [4][5][6][7][8][9][10] improving the cell internalization of otherwise insoluble, unstable, or unavailable therapeutic compounds and reducing the amount &dosage& of those compounds and the &associated& side effects. [11] In this light, we recently described the preparation of a novel class of squalene-based releasable compounds conjugated with paclitaxel, podophyllotoxin, camptothecin, and epothilone A. Owing to the presence of the squalene tail and the disulfide-containing linker, the derivatives self-assembled in water to form nanoparticles and the drug was released after cell internalization [12] as a possible consequence of the presence of glutathione (GSH) &ok?&. Couvreur et al. reported the possibility to obtain selfassembled fluorescent hetero-nanoparticles. [13] In our case, we recently described the combination of paclitaxel-squalene and fluoresceine-squalene conjugates to trace the delivery of the nanoassemblies and to investigate the internalization mechanism. ...
... In Scheme 1 we report a successful synthesis that relies on the protection of the amino group. [4] Condensation of cyclopamine N-Fmoc (1 b) [24] with dicarboxylic acid monoesters 2 a,b [12] in the presence of EDC-DMAP and subsequent removal of the protecting group led to cyclopamine conjugates 3 a,b.The conjugates obtained were used to form nanoparticles by the solvent-displacement method which were characterized by quasi elastic light scattering (QELS) ( Table 1). ...
... Given the better activity of compound 3 b, we decided to use compound 3 b and compound 4 [12] to form hetero-nanoparticles. Hetero-nanoparticles having different molar ratios of the two compounds were prepared; the results obtained with the more&most?& active nanosuspension (molar ratio 3 b/4 23:1). ...
Cyclopamine- and paclitaxel-containing hetero-nanoparticles generated by self-assembly show combined efficacy in the treatment of three different cancer cell lines. The use of ternary combination with the addition of a dye-squalene conjugate secured the obtainment of fluorescent nanoparticles that permitted the observation of the cellular internalization by confocal microscopy and super-resolution dSTORM (direct stochastic optical reconstruction microscopy).
... Namely, squalene-dithiolinker carboxylate 3 [20] was coupled in a 1:1 ratio with TMSprotected trehalose 2 [21] (step a, Scheme 1). Both TMS-protected mono-squalenylated 4a and bis-squalenylated trehalose 4b were isolated in poor, unoptimized yields after chromatography. ...
... The assembly of both NAs was carried out using a standardized method [20]. Namely, squalene-trehalose conjugates 1a and 1b were dissolved in THF (step a, Scheme 2) and then added dropwise to stirred MilliQ water, reaching a ≈2 mg/mL concentration (step b). ...
The disaccharide trehalose is a well-established autophagy inducer, but its therapeutic application is severely hampered by its low potency and poor pharmacokinetic profile. Thus, we targeted the rational design and synthesis of trehalose-based small molecules and nano objects to overcome such issues. Among several rationally designed trehalose-centered putative autophagy inducers, we coupled trehalose via suitable spacers with known self-assembly inducer squalene to yield two nanolipid-trehalose conjugates. Squalene is known for its propensity, once linked to a bioactive compound, to assemble in aqueous media in controlled conditions, internalizing its payload and forming nanoassemblies with better pharmacokinetics. We assembled squalene conjugates to produce the corresponding nanoassemblies, characterized by a hydrodynamic diameter of 188 and 184 nm and a high stability in aqueous media as demonstrated by the measured Z-potential. Moreover, the nanoassemblies were characterized for their toxicity and capability to induce autophagy in vitro.
... [3][4][5] The characterization of NPs is based on: (1) the sizeincluding mean diameter and size distribution, which influences their physical properties as well as biodistribution and retention in the body, and (2) the surface charge, which is responsible for the interactions with the environment and for the physical and chemical stability. 6 We recently reported the preparation of a novel class of squalene conjugates with paclitaxel, podophyllotoxin, camptothecin and epothilone A. 7 The obtained compounds were composed of a squalene tail that makes them able to selfassemble in water, and a drug unit connected via a disulfidecontaining linker to secure the release inside the cell. 7 We pursued our interest with the preparation of fluorescent hetero-nanoassemblies obtained by mixing a paclitaxelsqualene conjugate with a fluorescein-squalene conjugate. ...
... 6 We recently reported the preparation of a novel class of squalene conjugates with paclitaxel, podophyllotoxin, camptothecin and epothilone A. 7 The obtained compounds were composed of a squalene tail that makes them able to selfassemble in water, and a drug unit connected via a disulfidecontaining linker to secure the release inside the cell. 7 We pursued our interest with the preparation of fluorescent hetero-nanoassemblies obtained by mixing a paclitaxelsqualene conjugate with a fluorescein-squalene conjugate. 8 Next, we moved forward and we demonstrated that cyclopamine and paclitaxel-squalene conjugates are able to selfassemble into hetero-nanoparticles that show combined efficacy in the treatment of three different cancer cell lines. ...
The ability of 4-(1,2-diphenylbut-1-en-1-yl)aniline as a self-assembly inducer is reported. The conjugation of this moiety with aloin or podophyllotoxin resulted in spherical nanoparticles that were characterized by Dynamic Light Scattering (DLS), Transmission Electron Microscopy (TEM) and NanoSight technology. A preliminary biological evaluation on two cancer cell lines is reported.
... This kind of conjugates is capable to spontaneously assemble in water, forming NPs able to release a payload drug in cellular media. [4][5][6][7][8] Additional modications can be made on this simple design in order to obtain hetero-NPs bearing two different drugs (combining different conjugates that present the same lipidic self-assembling inducer); 9,10 single and dual drug uorescent hetero-NPs (where in one of the conjugates the drug is substituted by a uorescent moiety) 11,12 and NPs formed by selfassembling conjugate dual drugs (in which also the selfassembly inducer is pharmaceutically active). 13,14 Recently, we focused on further improving these NPs, by exploiting targeted drug delivery through folate-containing hetero-NPs. ...
Self-assembled hetero-nanoparticles are obtained by mixing paclitaxel and folic acid conjugates.
... 9−12 These self-assembled nanoparticle systems, which normally contain the drug covalently attached to a lipid moiety, can also profit from the pathways of lipid biochemistry to target specific organs depending on the nanoparticle size. 9 In this work, the structure, thermal stability, tendency for polymorphism, and solubility (phosphate buffer, pH 7.4, 298 and 310 K) of DHR have been investigated. A lipid-DHR conjugate was also synthesized and used to prepare a waterdispersible nanoparticle formulation as a means to enhance the aqueous solubility of DHR. ...
6,7-Dehydroroyleanone (DHR) is a caspase-induced cytotoxic abietane diterpene, frequently found on Plectranthus spp. A pharmaceutical formulation consisting of a DHR-squalene conjugate was synthesized and analyzed by different techniques such as scanning electron microscopy (SEM). The facile production of the dispersion of DHR-squalene conjugate nanoparticles in phosphate buffer (pH 7.4) suggests that this nanodelivery platform may be an effective system to improve the solubility and bioavailability of DHR, so that therapeutical systemic levels may be achieved.
... For instance, the concentrations of PTX and CMPT could be controlled by their loading into the liposome. 1 Borrelli et al. 2 synthesized the squalene conjugates with PTX, CMPT, podophyllotoxin, and epothilone against A549 lung cancer cells. In other study, the combined effect of PTX and CMPT was investigated against melanoma, 3 breast, pancreatic, 4 and medullary thyroid carcinomas. ...
Gold nanoparticles, paclitaxel (PTX), and camptothecin (CMPT) were loaded into the PVA/κ-carrageenan/pegylated-PU composite and core-shell nanofibers prepared by two-nozzle and coaxial electrospinning methods. The capability of composite and core-shell nanofibers was investigated for the targeted delivery of anticancer drugs in lung cancer treatment. In vitro and in vivo release of PTX and CMPT were investigated to find the release mechanism from nanofibers compared to direct administration of pristine PTX and CMPT. The mean fiber diameter for composite and core-shell nanofibers with shell feeding rates of 0.3, 0.5, and 0.7 mL h-1 was about 225, 330, 520, and 640 nm, respectively. In vivo release studies indicated that the blood concentration of CMPT and PTX for rats fed with core-shell nanofibers reached the highest values of 26.8 ± 0.04 μg mL-1, and 26.5 ± 0.05 μg mL-1 in 36 h, and 24 h and reduced slowly within 84 h, and 48 h, respectively. The maximum cytotoxicity was 75% in the presence PVA/κ-carrageenan/CMPT/Au/pegylated-PU/PTX core-shell nanofibers. In vivo antitumor activity results confirmed the synergic effect of Au, CMPT and PTX anticancer drugs on the reduction of tumor volume without change in mouse weight by the PVA/κ-carrageenan/CMPT/Au/pegylated PU/PTX core-shell nanofibers. The obtained results indicated that the simultaneous loading of CMPT and PTX anticancer drugs and Au nanoparticles is more beneficial for lung cancer treatment.
... [51] In similar, when squalene coupled to hydrophobic active compounds, including PTX, podophyllotoxin, camptothecin (CPT) and epothilone A through both disulfide or allhydrocarbon bond to form NPs spontaneously in water, and release the parent drug in vitro, but the size, stability and in vitro activity differed. [52] The monoisoprenoyl was conjugate to PTX to give prodrug, which was further self-assembled into NPs through the surfactant squalene derivative. [53] . ...
This article provides a broad spectrum about the nanoprodrug fabrication advances co‐driven by prodrug and nanotechnology development to potentiate cancer treatment. The nanoprodrug inherits the features of both prodrug concept and nanomedicine know‐how, attempts to solve underexploited challenge in cancer treatment cooperatively. Prodrugs can release bioactive drugs on‐demand at specific sites to reduce systemic toxicity, this is done by using the special properties of the tumor microenvironment, such as pH value, glutathione concentration, and specific overexpressed enzymes; or by using exogenous stimulation, such as light, heat, and ultrasound. The nanotechnology, manipulating the matter within nanoscale, has high relevance to certain biological conditions, and has been widely utilized in cancer therapy. Together, the marriage of prodrug strategy which shield the side effects of parent drug and nanotechnology with pinpoint delivery capability has conceived highly camouflaged Trojan horse to maneuver cancerous threats.
... This kind of self-aggregation might save the potential patient from undesired side effects related to the toxicity of common formulations [412,413] or a burst release of the drug from the formulation [414]. The concept of squalenylation has already been used in the context of drug delivery [415][416][417][418][419] since its first application to improve the delivery of gemcitabine, a cytotoxic nucleoside analogue used in anticancer chemotherapy [420]. ...
Drug candidates derived from oligonucleotides (ON) are receiving increased attention that is supported by the clinical approval of several ON drugs. Such therapeutic ON are designed to alter the expression levels of specific disease-related proteins, e.g., by displaying antigene, antisense, and RNA interference mechanisms. However, the high polarity of the polyanionic ON and their relatively rapid nuclease-mediated cleavage represent two major pharmacokinetic hurdles for their application in vivo. This has led to a range of non-natural modifications of ON structures that are routinely applied in the design of therapeutic ON. The polyanionic architecture of ON often hampers their penetration of target cells or tissues, and ON usually show no inherent specificity for certain cell types. These limitations can be overcome by conjugation of ON with molecular entities mediating cellular ‘targeting’, i.e., enhanced accumulation at and/or penetration of a specific cell type. In this context, the use of small molecules as targeting units appears particularly attractive and promising. This review provides an overview of advances in the emerging field of cellular targeting of ON via their conjugation with small-molecule targeting structures.
... The authors combined the drug with the target molecule in the novel prodrug that showed significant targeting effects on HeLa cells. Borrelli et al. utilized disulfide bonds to link squalene with active drug to construct a conjugation that can self-assemble in water and release the parent drug in vitro ( Fig. 4) [30]. This research also showed that the introduction of disulfide bonds and squalene did not affect the activity of the parent drugs. ...
Combining releasable chemical crosslinkers with photoaffinity probes represents a valuable tool for identifying protein-protein interactions (PPIs). The biomacromolecule photoaffinity probe prepared by using releasable photoaffinity linkers can be used to exploring PPIs by triggering release of the releasable group. More importantly, it can overcome the shortcomings of macromolecular photoaffinity probes without label transfer functionality to accurately confirm defects in specific structural sites. It shows particular promise for research exploring the interaction of unknown proteins and transient-weak PPIs in living organisms to discover new drug targets. In this review, we highlight recent progress in the development and application of chemical releasable linkers in photoaffinity probes. Several comparative studies are described in which the efficiency of various photoaffinity probes are compared.
... 54 Xue et al. demonstrated the nano assembly of the monoterpenoid drug conjugate (citronellol−cabazitaxel) and their enhanced anticancer activity in a redox-stimuli-sensitive manner, 55 and Dosio's research group expanded the squalene technology to design multidrug conjugate NPs that release the drugs under redox stress. 56 ...
Cancer chemotherapy has witnessed translation of good number of lipid-based formulations to clinics based on drug encapsulation strategies. Toxicities associated with release of anticancer drugs in systemic circulation and usage of toxic excipients in these formulations along with poor efficacy, reduced blood circulation time and ineffective tumor-targeting ability are responsible for poor success in patient survival. However, recent advances in bioconjugation strategies for engineering of lipid-drug conjugates uplifted the physicochemical, pharmacokinetic/biodistribution properties and antitumor activities of anticancer drugs with reduced toxicity in preclinical models as compared to traditional lipid formulations. Conjugation of the anticancer drugs to amphiphilic lipid molecules allows the sustained release of the drug under perfect stimuli conditions and their amphiphilic nature helps in formation of self-assembled nanoparticles that can easily be targeted at the tumor site. In this review, we present recent advances in emerging class of lipid-drug conjugates (LDCs) for cancer therapy focusing on their design, tunable drug release at target sites, pharmacokinetics, antitumor activity and toxicology, and provide future directions for their translation into clinics.
... T he use of self-assembled nanoparticles (NPs) is a challenging strategy that could result in a useful and smart approach to effectively treat cancer. 1−3 In particular, we have recently reported the preparation of a novel class of squalene conjugates with paclitaxel, podophyllotoxin, camptothecin, and epothilone A. 4 The compounds obtained were characterized by a squalene tail that makes them able to selfassemble in water, and by a drug unit connected via a disulfidecontaining linker to secure the release inside the cell. The need to trace the delivery of the nanoassemblies and to demonstrate the internalization of the drugs prompted us to prepare fluorescent heteronanoassemblies composed by a paclitaxel− squalene conjugate and fluorescein−squalene conjugate. ...
The preparation of hetero-nanoparticles (NPs) with doxorubicin (DOXO) and cyclopamine (CYP) conjugates is presented. Biological evaluation on A431 cell lines confirms the maintenance of the activity of the parental drugs. The in vivo study shows that self-assembled NPs reduce tumour growth and toxicity of chemotherapy
... First, the higher anti-proliferative activity of conjugate 18 is mainly due to its strong inhibition of DHFR because it is about 3.4-fold more potent to the target enzyme relative to conjugate 21. Second, cancer cells contain abundant glutathione (GSH) with concentrations ranging from 2 to 20 mM, which are more than 100 times higher than those in the extracellular matrix and blood plasma that range from 2 to 20 lM [25,26]. Thus, the redox-triggered rapid release of MTX from conjugate 18 may have contributed to this phenomenon. ...
... With the increasing public perception of a strong correlation between drugs—safety—and disease prevention, new approaches should be introduced to enrich the biomedical application field with " health promoting substances " . Usually, in the treatment of various diseases it is not sufficient to use highly active drugs, the effectiveness of any active being mainly dependent upon its delivery in an appropriate concentration to target cells and tissue (Borrelli et al., 2014). A great challenge comes from the difficulty to design functional delivery systems which combine low toxicity, lack of immunogenicity and biodegradability, does not accumulate in cells or tissues, and could also bring supplementary biological effects (Jyoti et al., 2015; Lacatusu et al., 2012). ...
... With the increasing public perception of a strong correlation between drugs-safety-and disease prevention, new approaches should be introduced to enrich the biomedical application field with "health promoting substances". Usually, in the treatment of various diseases it is not sufficient to use highly active drugs, the effectiveness of any active being mainly dependent upon its delivery in an appropriate concentration to target cells and tissue (Borrelli et al., 2014). A great challenge comes from the difficulty to design functional delivery systems which combine low toxicity, lack of immunogenicity and biodegradability, does not accumulate in cells or tissues, and could also bring supplementary biological effects (Jyoti et al., 2015;Lacatusu et al., 2012). ...
... With the increasing public perception of a strong correlation between drugs-safety-and disease prevention, new approaches should be introduced to enrich the biomedical application field with "health promoting substances". Usually, in the treatment of various diseases it is not sufficient to use highly active drugs, the effectiveness of any active being mainly dependent upon its delivery in an appropriate concentration to target cells and tissue (Borrelli et al., 2014). A great challenge comes from the difficulty to design functional delivery systems which combine low toxicity, lack of immunogenicity and biodegradability, does not accumulate in cells or tissues, and could also bring supplementary biological effects (Jyoti et al., 2015;Lacatusu et al., 2012). ...
... It was thus expected that this approach could only be applied to hydrophilic compounds. Unexpectedly, two studies showed that the squalenoylation technique may also be appropriate to design nanomedicines loaded with hydrophobic compounds having very low water solubility, like paclitaxel (PTX) [67], podophyllotoxin, camptothecin, and epothilone [68]. It thus appeared that squalenoylation might also be applicable to poorly water soluble compounds, which are difficult to administer intravenously. ...
Some approaches to enhance the entrapment efficiency within nanoparticulate systems, chiefly of hydrophilic molecules, developed mainly by the Turin University Pharmaceutical Technology group, are discussed. Several approaches have been developed: the first entails associating anticancer hydrophilic molecules to liposomes, developing lipophilic prodrugs of these molecules, and encapsulating them in liposomes. The transformation of hydrophilic drugs into lipophilic prodrugs can also overcome problems of poor entrapment efficiency and rapid release from polymer nanocarriers. Examples are nanospheres and nanocapsules produced from various PEGylated poly(alkylcyanoacrylate) copolymers. Strategies have also been developed to enhance hydrophilic drug entrapment in solid lipid nanoparticles (SLN). Hydrophobic ion pairing was designed to enable various antitumor drugs to be entrapped in SLN, produced by the coacervation method. Another technique comprises the covalent linkage of antitumoral or antiviral drugs to a squalenoyl-derived chain, affording bioconjugates that self-assemble as stable nanoparticles. A further development comprises mesoporous silica nanoparticles with immobilized hydrophilic antioxidants, for topical applications: they were complexed with hydrophilic antioxidants (Trolox® or rutin). Polymer-shelled and perfluoropentane-cored nanobubbles have also been designed, as versatile multifunctional carriers for the delivery of gases, drugs, and genes; the size range is below 500 nm, with shell thickness in the 30-50 nm range.
... With the increasing public perception of a strong correlation between drugs-safety-and disease prevention, new approaches should be introduced to enrich the biomedical application field with "health promoting substances". Usually, in the treatment of various diseases it is not sufficient to use highly active drugs, the effectiveness of any active being mainly dependent upon its delivery in an appropriate concentration to target cells and tissue (Borrelli et al., 2014). A great challenge comes from the difficulty to design functional delivery systems which combine low toxicity, lack of immunogenicity and biodegradability, does not accumulate in cells or tissues, and could also bring supplementary biological effects (Jyoti et al., 2015;Lacatusu et al., 2012). ...
... [1] Our continued interest in the field of chemical approaches to target cancer cells [2][3][4][5][6][7][8][9] has led us to study the preparation of a novel class of squalene conjugated with paclitaxel, podophyllotoxin, camptothecin, and epothilone A. These conjugates were all functionalized with a squalene tail that enabled them to self-assemble in water, and to secure the release of the drug inside the cells through a disulfide-containing linker. [10] The need to trace the delivery of the nanoassemblies and to demonstrate the internalization of the drugs prompted us to prepare fluorescent heteronanoassemblies by mixing a paclitaxel-squalene conjugate and a fluorescein-squalene conjugate ( Figure 1). ...
The preparation of fluorescent nanoparticles containing squalenoyl–paclitaxel and a squalene-based fluorescein derivative is presented. The formation of self-assembled heteronanoparticles was confirmed by using the quasielastic light scattering (QELS) technique. The internalization in A549 human lung carcinoma cells was verified by microscopy analysis. Finally, paclitaxel in nanoassemblies maintains its ability to target microtubules.
The conjugation of tetraphenylethylene (TPE) with podophyllotoxin, N-desacetylthiocolchicine, and cabazitaxel through a sebacic acid linker led to the formation of fluorescent nanoparticles. Dynamic light scattering (DLS) and photoluminescence spectroscopy were used for the identification and characterization of the fluorescent nanoparticles. The biological evaluation was determined in three human ovarian (KURAMOCHI, OVCAR3, OVSAHO) and three human breast (MCF7, SKBR 3, and MDA-MB231) cancer cell lines. In the case of cabazitaxel, the nanoparticles maintained the activity of the parent drug, at the low nanomolar range, while exhibiting high blue fluorescence. The internalization of the fluorescent NPs into cells was detected using immunofluorescence assay.
A potent nontoxic antitumor drug, 2-hydroxyoleic acid (6, 2OHOA) used for membrane lipid therapy, was selected as a self-assembly inducer due to its ability to form nanoparticles (NPs) in water. For this purpose, it was conjugated with a series of anticancer drugs through a disulfide-containing linker to enhance cell penetration and to secure drug release inside the cell. The antiproliferative evaluation of the synthesized NP formulations against three human tumor cell lines (biphasic mesothelioma MSTO-211H, colorectal adenocarcinoma HT-29, and glioblastoma LN-229) showed that nanoassemblies 16–22a,bNPs exhibit antiproliferative activity at micromolar and submicromolar concentrations. Furthermore, the ability of the disulfide-containing linker to promote cellular effects was confirmed for most nanoformulations. Finally, 17bNP induced intracellular ROS increase in glioblastoma LN-229 cells similarly to free drug 8, and such elevated production was decreased by pretreatment with the antioxidant N-acetylcysteine. Also, nanoformulations 18bNP and 21bNP confirmed the mechanism of action of the free drugs.
Malignant tumors are potential killers of human life, seriously endangering people’s physical health. As far as the treatment of malignant tumors is concerned, there exist many therapeutic methods such as surgery, radiotherapy, chemotherapy, among which chemotherapy is an important means of antitumor therapy. Chemotherapy antineoplastic agents can be divided into alkylating agents, metalloplatinum, plant drugs, antimetabolite, antineoplastic antibiotics, antineoplastic hormones, molecular targeted agents, and peptides, This article reviews the current drug characteristics, mechanism of action and research directions of various anti-tumor drugs in order to provide references for clinical treatment.
Halobacteria, a type of archaea in high salt environments, have phytanyl ether phospholipid membranes containing up to 50% menaquinone. It is not understood why a high concentration of menaquinone is required and how it influences membrane properties. In this study, menaquinone-8 headgroup and torsion parameters of isoprenoid tail are optimized in the CHARMM36 force field. Molecular dynamics simulations of archaeal bilayers containing 0 to 50% menaquinone characterize the distribution of menaquinone-8 and menaquinol-8, as well as their effects on mechanical properties and permeability. Menaquinone-8 segregates to the membrane midplane above concentrations of 10%, favoring an extended conformation in a fluid state. Menaquinone-8 increases the bilayer thickness but does not significantly alter the area compressibility modulus and lipid chain ordering. Counterintuitively, menaquinone-8 increases water permeability because it lowers the free energy barrier in the midplane. The thickness increase due to menaquinone-8 may help halobacteria ameliorate hyper-osmotic pressure by increasing the membrane bending constant. Simulations of the archaeal membranes with archaerhodopsin-3 show that the local membrane surface adjusts to accommodate the thick membranes. Overall, this study delineates the biophysical landscape of 50% menaquinone in the archaeal bilayer, demonstrates the mixing of menaquinone and menaquinol, and provides atomistic details about menaquinone configurations.
Ionizable cationic lipids are critical components involved in nanoparticle formulations, which are utilized in delivery platforms for RNA therapeutics. While general criteria regarding lipophilicity and measured pKa in formulation are understood to have impacts on utility in vivo, greater granularity with respect to the impacts of the structure on calculated and measured physicochemical parameters and the subsequent performance of those ionizable cationic lipids in in vivo studies would be beneficial. Herein, we describe structural alterations made within a lipid class exemplified by 4, which allow us to tune calculated and measured physicochemical parameters for improved performance, resulting in substantial improvements versus the state of the art at the outset of these studies, resulting in good in vivo activity within a range of measured basicity (pKa = 6.0-6.6) and lipophilicity (cLogD = 10-14).
Hepatocellular carcinoma (HCC) ranks as the fifth most common and the second deadliest cancer worldwide. HCC is extremely resistant to the conventional chemotherapeutics. Hence, it is vital to develop new treatment options. Chalcones were previously shown to have anticancer activities in other cancer types. In this study, 11 chalcones along with quercetin, papaverin, catechin, Sorafenib and 5FU were analyzed for their bioactivities on 6 HCC cell lines and on dental pulp stem cells (DPSC) which differentiates into hepatocytes, and is used as a model for untransformed control cells. 3 of the chalcones (1, 9 and 11) were selected for further investigation due to their high cytotoxicity against liver cancer cells and compared to the other clinically established compounds. Chalcones did not show significant bioactivity (IC50>20μM) on dental pulp stem cells. Cell cycle analysis revealed that these 3 chalcone-molecules induced SubG1/G1 arrest. Akt protein phosphorylation was inhibited by these molecules in PTEN deficient, drug resistant, mesenchymal like Mahlavu cells leading to the activation of p21 and the inhibition of NFκB-p65 transcription factor. Hence the chalcones induced apoptotic cell death pathway through NFκB-p65 inhibition. On the other hand, these molecules triggered p21 dependent activation of Rb protein and thereby inhibition of cell cycle and cell growth in liver cancer cells. Involvement of PI3K/Akt pathway hyperactivation was previously described in survival of liver cancer cells as carcinogenic event. Therefore, our results indicated that these chalcones can be considered as candidates for liver cancer therapeutics particularly when PI3K/Akt pathway involved in tumor development.
Lipid drug conjugates (LDCs) which may also be addressed as lipoidal prodrug, are drug molecules that have been covalently linked with lipids like fatty acids, glycerides or phospholipids. Lipid drug conjugates are fabricated with the aim of increasing drug payload and thus prevent leakage of a highly polar drug from a lipophilic matrix. Conjugating lipids to drug molecules improves lipophilicity and also changes other characteristics of drugs.The conjugates possess numerous advantages including enhanced tumor targeting, improved lymphatic system targeting, improved systemic bioavailability and decreased toxicity. Different conjugation approaches, chemical linkers and spacers can be used to synthesize LDCs based on the chemical nature of lipids and bioactive(s). Linkers and/or conjugation techniques determine how bioactive(s) are released from LDCs and considered as a critical parameter for the optimal performance of the LDCs. The purpose of this review is to discuss widely the use of LDCs as an approach for improving therapeutic drug delivery. In this review different conjugation methods, various lipids used for preparing LDCs and advantages of using LDCs are summarized. Even though LDCs might be administered without a delivery system, most of them are incorporated in appropriate delivery system. The lipid moiety in the conjugates may significantly improve loading of bioactive(s) into hydrophobic compartments. This results in formulations with high drug loading and greater stability. Different delivery carriers such as polymeric nanoparticles, micelles, liposomes, emulsions and lipid nanoparticles which have been explored are discussed in this review.
Betulinic acid is validated as a new self-assembly inducer for the formation of nanoparticles (NPs) in combination to different drugs. The target compounds are characterized by the presence of anticancer drugs acting on tubulin dynamics and as a linker a carbon chain or a triazole-based one. Nanoparticles formed are characterized and their biological activity is evaluated.
Enkephalin is an endogenous pentapeptide producing potent analgesia by activating opioid receptors located on central and peripheral neuronal cell membranes. However, its clinical use has historically been limited due to pharmacokinetic issues, including restricted plasma stability and blood brain barrier impermeability. The aim of this project is to create a new enkephalin-based nanomedicine targeting pain, using biocompatible and biodegradable materials for drug delivery and targeting purposes, such as squalene (squalenoylation nanotechnology). This nanotechnology presents a new concept with numerous advantages in comparison with the conventional nanocarriers, such as high drug loading and absence of “burst release”. Here, we show for the first time, that the rapidly metabolized Leu-enkephalin (LENK) neuropeptide may become pharmacologically efficient owing to its simple conjugation with the squalene (SQ) using three different chemical linkers, i.e., dioxycarbonyl (Diox), diglycolate (Dig), or amide bond (Am). The resulting prodrugs were able to self-assemble in nanoparticles in aqueous media. This new squalene-based nanoformulation prevented rapid plasma degradation of LENK and conferred to the released neuropeptide a significant anti-hyperalgesic effect in a carrageenan-induced paw edema model in rats (Hargreaves test). It should be stressed that this effect lasted 3 times longer than morphine. Pretreatment with brain impermeant opioid receptor antagonist naloxone methiodide (Nal-M) reversed the nanoparticles induced anti-hyperalgesia, indicating that LENK-SQ NPs acted through peripherally located opioid receptors. Moreover, the biodistribution of DiD-fluorescently labeled LENK-SQ NPs showed a strong accumulation of the fluorescence within the inflamed paw as well as in the liver, spleen, and lung, while no signal could be detected in the brain, confirming the peripheral effect of LENK-SQ NPs. Toxicological studies showed that despite nanoparticles accumulation in the liver, the levels of aspartate transaminase (AST) and alanine transaminase (ALT) were not increased after i.v. injection of LENK-SQ NPs, highlighting thus their safety. This study represents a novel drug targeting approach, allowing the specific delivery of LENK neuropeptide into inflamed tissues for pain alleviation.
The design and the synthesis of new self-assembling conjugates is reported. The target compounds are characterized by the presence of a self-immolative linker that secures a controlled release induced by lipase cleavage. 4-(1,2-Diphenylbut-1-en-1-yl)aniline is used as a self-assembling inducer and amino-thiocolchicine as prototype of drug. The release of thiocolchicine derivative has been demonstrated in vitro in the presence of porcine pancreatic lipase (PPL) and celite-supported lipase (PS). The formation of nanoparticles is confirmed by DLS, atomic force microscopy (AFM) and fluorescence microscopy. The antiproliferative activity has been proved on two human cancer cell lines.
Lipid-drug conjugates (LDCs) are drug molecules that have been covalently modified with lipids. The conjugation of lipids to drug molecules increases lipophilicity and also changes other properties of drugs. The conjugates demonstrate several advantages including improved oral bioavailability, improved targeting to the lymphatic system, enhanced tumor targeting, and reduced toxicity. Based on the chemical nature of drugs and lipids, various conjugation strategies and chemical linkers can be utilized to synthesize LDCs. Linkers and/or conjugation methods determine how drugs are released from LDCs and are critical for the optimal performance of LDCs. In this review, different lipids used for preparing LDCs and various conjugation strategies are summarized. Although LDCs can be administered without a delivery carrier, most of them are loaded into appropriate delivery systems. The lipid moiety in the conjugates can significantly enhance drug loading into hydrophobic components of delivery carriers and thus generating formulations with high drug loading and superior stability. Different delivery carriers such as emulsions, liposomes, micelles, lipid nanoparticles, and polymer nanoparticles are also discussed in this review.
The synthesis of three macrotriazoles that are analogues of migrastatin is reported. The synthesis is based on copper(I)- and ruthenium catalyzed azide alkyne cycloaddition. The preparation of the enantiopure terminal alkyne derivative is based onTrost desymmetrisation, Brown alkoxyallylation and the efficient Colvin reaction. Biological evaluation of the obtained compounds evidenced a promising efficacy in reducing the ability of MDA-MB-361 cell line to migrate.
Various lipid particulate drug delivery systems with liposomes as forefront have attracted increasing attention in cancer treatment, especially breast cancer therapy. Multidrug resistance and off-target cytotoxicity are among major obstacles faced by researchers in this area. To address these problems, various strategies, such as composition modifications, pegylation and attaching the targeting ligands, have been investigated. In this regard, a number of liposomal therapeutics could successfully reach the market and also several preclinical and clinical approaches are ongoing to upgrade lipid particles to multipotential carriers and potentiate their therapeutic efficacies. The current review is mainly concentrated on recent approaches in liposomal drug delivery systems intended for breast cancer therapy, and will have a glance at novel attributed studies based on other particulate lipid carriers.
There has been significant interest in the methodologies of controlled release for a diverse range of applications spanning drug delivery, biological and chemical sensors, and diagnostics. The advancement in novel substrate-polymer coupling moieties has led to the discovery of self-immolative linkers. This new class of linker has gained popularity in recent years in polymeric release technology as a result of stable bond formation between protecting and leaving groups, which becomes labile upon activation, leading to the rapid disassembly of the parent polymer. This ability has prompted numerous studies into the design and development of self-immolative linkers and the kinetics surrounding their disassembly. This review details the main concepts that underpin self-immolative linker technologies that feature in polymeric or dendritic conjugate systems and outlines the chemistries of amplified self-immolative elimination.
Modern combinatorial chemistry is used to discover compounds with desired function by an alternative strategy, in which the biological target is directly involved in the choice of ligands assembled from a pool of smaller fragments. Herein, we present the first experimental result where the use of in situ click chemistry has been successfully applied to probe the ligand-binding site of Abl and the ability of this enzyme to form its inhibitor. Docking studies show that Abl is able to allow the in situ click chemistry between specific azide and alkyne fragments by binding to Abl-active sites. This report allows medicinal chemists to use protein-directed in situ click chemistry for exploring the conformational space of a ligand-binding pocket and the ability of the protein to guide its inhibitor. This approach can be a novel, valuable tool to guide drug design synthesis in the field of tyrosine kinases.
This short review aims at presenting some recent illustrative examples of spontaneous nucleolipids self-assembly. High-resolution
structural investigations reveal the diversity and complexity of assemblies formed by these bioinspired amphiphiles, resulting
from the interplay between aggregation of the lipid chains and base–base interactions. Nucleolipids supramolecular assemblies
are promising soft drug delivery systems, particularly for nucleic acids. Regarding prodrugs, squalenoylation is an innovative
concept for improving efficacy and delivery of nucleosidic drugs.
Using the anticancer compound paclitaxel as a model drug, this study investigates the potential of the squalenoylation technology (i.e., bioconjugation with the natural lipid squalene) in addressing the drug ability and delivery issues of poorly soluble therapeutic agents. In this view, a variety of novel squalene-based prodrugs of the anticancer compound paclitaxel were synthesized, which produced nanoparticles in water. These prodrugs were obtained by covalent coupling of the paclitaxel 2'-hydroxyl group as direct ester, as well as with a succinate or a diglycolate ester as cleavable linker to the 1,1',2-tris-norsqualenoic acid. The hydrophilicity of these paclitaxel bioconjugates was increased by placing poly(ethylene glycol) chains of different lengths between paclitaxel and the squalenoyl moiety. All these prodrugs self-assembled into nanosized aggregates in aqueous solution as characterized by dynamic light scattering, atomic force microscopy, and transmission electron microscopy. The critical aggregation concentration was very low, ranging from 0.09 to 0.4 mg/L. Zeta potential measurements revealed that all squalenoyl-paclitaxel nanoassemblies (NA) held a global negative charge and appeared stable in water for several weeks as determined by particle size measurement. The release of paclitaxel from NA was evaluated in different conditions and in the presence of serum and depended on the nature of the linker used. Preliminary biological assessment showed that these squalenoyl-paclitaxel NA induced the formation of microtubule bundles in HT-29 and KB-31 cells, and additionally displayed notable cytotoxicity on a lung tumor cell line. Furthermore, the cytotoxic activity of these different prodrugs correlated closely with the observed linker stability. Overall, the squalenoylation nanotechnology opens up interesting perspectives for the development of injectable prodrugs of poorly soluble therapeutic compounds by addressing the associated physicochemical and biopharmaceutical challenges.
The synthesis and biological evaluation of 9 dimeric compounds obtained by condensation of thiocolchicine and/or podophyllotoxin with 6 different dicarboxylic acids is described. In particular, tubulin assembly assay and immunofluorescence analysis results are reported. The biological data highlighted three compounds as being more active than the others, having a marked ability to inhibit the polymerization of tubulin in vitro and causing significant disruption to the microtubule network in vivo. The spacer unit was found to have a significant effect on biological activity, reinforcing the importance of the design of conjugate compounds to create new biologically active molecules in which the spacer could be useful to improve the solubility and to modulate the efficacy of well known anticancer drugs.
Nanocarriers offer unique possibilities to overcome cellular barriers in order to improve the delivery of various drugs and drug candidates, including the promising therapeutic biomacromolecules (i.e., nucleic acids, proteins). There are various mechanisms of nanocarrier cell internalization that are dramatically influenced by nanoparticles' physicochemical properties. Depending on the cellular uptake and intracellular trafficking, different pharmacological applications may be considered. This review will discuss these opportunities, starting with the phagocytosis pathway, which, being increasingly well characterized and understood, has allowed several successes in the treatment of certain cancers and infectious diseases. On the other hand, the non-phagocytic pathways encompass various complicated mechanisms, such as clathrin-mediated endocytosis, caveolae-mediated endocytosis and macropinocytosis, which are more challenging to control for pharmaceutical drug delivery applications. Nevertheless, various strategies are being actively investigated in order to tailor nanocarriers able to deliver anticancer agents, nucleic acids, proteins and peptides for therapeutic applications by these non-phagocytic routes.
Gemcitabine (2',2'-difluorodeoxyribofuranosylcytosine; dFdC) is an anticancer nucleoside analog active against wide variety of solid tumors. However, this compound is rapidly inactivated by enzymatic deamination and can also induce drug resistance. To overcome the above drawbacks, we recently designed a new squalenoyl nanomedicine of dFdC [4-N-trisnorsqualenoyl-gemcitabine (SQdFdC)] by covalently coupling gemcitabine with the 1,1',2-trisnorsqualenic acid; the resultant nanomedicine displayed impressively greater anticancer activity compared with the parent drug in an experimental murine model. In the present study, we report that SQdFdC nanoassemblies triggered controlled and prolonged release of dFdC and displayed considerably greater t(1/2) (approximately 3.9-fold), mean residence time (approximately 7.5-fold) compared with the dFdC administered as a free drug in mice. It was also observed that the linkage of gemcitabine to the 1,1',2-trisnorsqualenic acid noticeably delayed the metabolism of dFdC into its inactive difluorodeoxyuridine (dFdU) metabolite, compared with dFdC. Additionally, the elimination of SQdFdC nanoassemblies was considerably lower compared with free dFdC, as indicated by lower radioactivity found in urine and kidneys, in accordance with the plasmatic concentrations of dFdU. SQdFdC nanoassemblies also underwent considerably higher distribution to the organs of the reticuloendothelial system, such as spleen and liver (p < 0.05), both after single- or multiple-dose administration schedule. Herein, this paper brings comprehensive pharmacokinetic and biodistribution insights that may explain the previously observed greater efficacy of SQdFdC nanoassemblies against experimental leukemia.
Three H2O-soluble prodrugs of taxol have been prepared using a sulfonate group as the H2O-solubilizing group. The first two contain taurine or 3-amino-1-sulfopropionic acid linked to the 2′-position of taxol through a succinate group, and a novel method of preparing these derivatives is described. The third derivative is prepared by the Michael addition of bisulfite to 2′-acryloyltaxol. A 2′-(γ-aminobutyryl)taxol salt was too unstable to be isolable.
The introduction of a hydrophobic group at position 7 of 9-fluorenone-2-carboxylic acid generates new tubulin binders. The design is suggested by modelling studies. The synthesis is based on the use of 2,7-dibromo- -fluorenone as starting material. The antiproliferative activity on two different cell lines, fluorescent microscopy, flow cytometry and sedimentation assay tests confirmed the supposed mechanism.
Azasqualenes, bis-azasqualenes and derivatives, designed as inhibitors of squalene 2,3-epoxide cyclase, a key enzyme in sterol biosynthesis, were synthesized and their in vitro activities against a variety of yeasts, fungi, gram-positive and gram-negative bacteria were determined.The synthesis involves a new method of squalene degradation, together with an unusual procedure for the aminative reduction of lipophilic aldehydes.A study of the structure—activity relationship was attempted for different biological parameters: anti-bacterial and anti-fungal activities (MIC), inhibition of mycelial growth (GTT), surfactant activity (CMC) and membrane perturbation activity (induction of leakage in liposomes).RésuméLes azasqualènes, bis-azasqualènes et dérivés ont été conçus pour inhiber l'époxydo-2,3 squalène cyclase, une enzyme clé dans la biosynthèse des stérols. Ces produits ont été synthétisés et leur activité testée in vitro vis-4-vis d'une grande variété de champignons, de levures et de bactéries gram-positives et -négatives.La synthèse chimique de ces composés fait appel à une nouvelle méthode de dégradation du squalène ainsi qu'à une technique particulière pour l'amination réductrice d'aldéhydes hydrophobes.Nous avons tenté d'effectuer une étude de relations structure—activité avec les différents paramètres mesurés: activité bactéricide et fongicide (MIC), inhibition de la croissance du mycélium (GTT), activité surfactante (CMC) et perturbation membranaire (perméabilisation de liposomes).
Thiocolchicine, taxol and cephalomannine have been used as building blocks for the preparation of four different fluorescent compounds designed to image the tubulin/microtubule network in cells. Thiocolchicine gave the best results and, in particular, the compound derived from conjugation with fluorescein minimally inhibits tubulin polymerization, is cell permeable and binds microtubules. Thus, it meets some of the demanding requirements for a new fluorescent dye that can secure a direct assay to evidence the tubulin/microtubules network in cells.
The use of tubulin as a target to influence the composition of the mixture from a dynamic combinatorial library, based on the disulfide bond exchange reaction, is described. ESI-FT-ICR-MS was used to determine the composition of the library. The heterodimeric compound amplified by this approach was used to design the homologous derivative with a two-carbon spacer in place of the disulfide function. The ability of the compounds to inhibit tubulin polymerization is reported and compared to thiocolchicine.
A series of new lipid prodrugs of paclitaxel, which can be formulated as nanoassemblies, are described. These prodrugs which are designed to overcome the limitations due to the systemic toxicity and low water solubility of paclitaxel consist of a squalene chain bound to the 2'-OH of paclitaxel through a 1,4-cis,cis-dienic linker. This design allows the squalene-conjugates to self-assemble as nanoparticular systems while preserving an efficient release of the free drug, thanks to the dienic spacer. The size, steric hindrance, and functional groups of the spacer have been modulated. All these prodrugs self-assemble into nanosized aggregates in aqueous solution as characterized by dynamic light scattering and transmission electron microscopy and appear stable in water for several days as determined by particle size measurement. In vitro biological assessment shows that these squalenoyl-paclitaxel nanoparticles display notable cytotoxicity on several tumor cell lines including A549 lung cell line, colon cell line HT-29, or KB 3.1 nasopharyngeal epidermoid cell line. The cis,cis-squalenyl-deca-5,8-dienoate prodrug show improved activity over simple 2'-squalenoyl-paclitaxel prodrug highlighting the favourable effect of the dienic linker. The antitumor efficacy of the nanoassemblies constructed with the more active prodrugs has been investigated on human lung (A549) carcinoma xenograft model in mice. The prodrug bearing the cis,cis-deca-5,8-dienoyl linker shows comparable antitumor efficacy to the parent drug, but reveals a much lower subacute toxicity as seen in body weight loss. Thus, nanoparticles with the incorporated squalenoyl paclitaxel prodrug may prove useful for replacement of the toxic Cremophor EL.
The introduction of a methylenthiol group at position 7 of camptothecin was carried out in four steps. This preparation also yielded the corresponding disulfide, which behaves as a prodrug due to its reactivity with glutathione. Assessment of their antiproliferative activities, investigations of their mechanism of action, and molecular modeling analysis indicated that the 7-modified camptothecin derivatives described herein maintain the biological activity and drug-target interactions of the parent compound.
A series of new polyisoprenoyl prodrugs of gemcitabine, which can be formulated as nanoassemblies are described. These prodrugs were designed to improve gemcitabine efficacy and to overcome the limitations due to the systemic toxicity of this anticancer compound. In vitro biological assessment showed that these polyisoprenoyl gemcitabine nanoassemblies displayed notable cytotoxicity on several cancer cell lines, including murine melanoma cell line B16F10, human pancreatic carcinoma cell line MiaPaCa-2, human lung carcinoma cell line A549 and human breast adenocarcinoma cell line MCF7. Interestingly, it was observed that the anticancer efficacy of these nanoassemblies was dependant on the size of polyisoprenoyl moiety. The polyisoprenoyl prodrug of gemcitabine containing three isoprene units (2d) was the more active on all the cancer cell lines tested. The antitumor efficacy of the nanoassemblies (NAs) constructed with the most active prodrug 2d was further evaluated on a human pancreatic (MiaPaCa-2) carcinoma xenograft model in mice. The prodrug 2d NAs showed an increased antitumor efficacy as compared to free gemcitabine or to squalene-gemcitabine (SQ-gem, 2a) nanoassemblies. Interestingly, MiaPaCa-2 tumors that did not respond to gemcitabine were inhibited by 76% after treatment with prodrug 2d NAs, whereas SQ-gem-treated MiaPaCa-2 tumor xenografts decreased only by 41% compared to saline or to gemcitabine-treated mice. Together, these findings demonstrated that the modulation of the length of nanoassemblies polyisoprenoyl moiety made tumor cells more sensitive to gemcitabine treatment without flagrant toxicity, thus providing a significant improvement in the drug therapeutic index.
Transition metal complexes provide a promising avenue for designing new therapeutic and diagnostic agents. In particular, ruthenium(II) polypyridyl complexes are useful for studying cellular uptake, due to their easy synthesis and unique photophysical properties. Dyes are frequently combined with material substrates to modulate their properties, enhance stability, reduce toxicity, and improve delivery. A novel Ru polypyridyl complex linked to a derivative of the natural lipid squalene (Ru-BIPPBI-hx-SQ) is described. Using the solvent displacement method, Ru-BIPPBI-hx-SQ easily self-assembles into nanosized aggregates in aqueous solution, as characterized by dynamic light scattering. The nanoassemblies exhibit long-lived and intense luminescence. Preliminary biological assessment showed them to be non-toxic; they are efficiently and rapidly transported across the cell membrane without requiring its permeabilization. Ru-labeled nanoassemblies are likely to be significant cellular-imaging tools, probing cellular events at very low concentrations. Moreover co-nanoassembly, with drug-derivatives based on squalenoylation technology, including gemcitabine and paclitaxel, has given interesting preliminary results.
We describe here new nanoparticles based on the bioconjugation of penicillin G to squalene in order to overcome severe intracellular infections by pathogen bacteria whose mechanism of resistance arises from the poor intracellular diffusion of several antibiotics. Two different squalene-penicillin G conjugates were synthesized (pH-sensitive and pH-insensitive), and their self-assembly as nanoparticles was investigated through morphology and stability studies. These nanoparticles had a size of 140 ± 10 nm (polydispersity index of 0.1) and a negative charge, and they did not display any supramolecular organization. Furthermore, they were found stable in water and in different culture medium. The cellular uptake and localization of these fluorescently labeled nanoparticles were explored on the macrophage cell line J774 by flow cytometry and confocal microscopy analysis. The squalenoylated nanoparticles were found to be cell internalized through clathrin-dependent and -independent endocytic pathways. Moreover, they induced an improved intracellular antibacterial activity on the facultative intracellular pathogen S. aureus, compared with free penicillin G, despite the absence of co-localization between the bacteria and the nanoparticles in the cells. This study suggests that the bioconjugation of an antibiotic to a squalene template could be a valuable approach for overcoming the antibiotic resistance due to intracellular bacterial infections.
The Met receptor tyrosine kinase is a promising target in anticancer therapies for its role during tumor evolution and resistance to treatment. It is characterized by an unusual structural plasticity as its active site accepts different inhibitor binding modes. Such feature can be exploited to identify distinct agents targeting tumor dependence and/or resistance by oncogenic Met. Here we report the identification of bioactive agents, featuring a new 4-(imidazo[2,1-b]benzothiazol-2-yl)phenyl moiety, targeting cancer cells dependent on oncogenic Met. One of these compounds (7c; Triflorcas) impairs survival, anchorage-independent growth, and in vivo tumorigenesis, without showing side effects. Our medicinal chemistry strategy was based on an in-house Met-focused library of aminoacid-amide derivatives enriched through structure-based computer modeling, taking into account the Met multiple-binding-mode feature. Altogether, our findings show how a rational structure-based drug design approach coupled to cell-based drug evaluation strategies can be applied in medicinal chemistry to identify new agents targeting a given oncogenic-dependency setting.
N-[2-Methyl-5-(triazol-1-yl)phenyl]pyrimidin-2-amine derivatives were synthesized and evaluated in vitro for their potential use as inhibitors of Bcr-Abl. The design is based on the bioisosterism between the 1,2,3-triazole ring and the amide group. The synthesis involves a copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) as the key step, with the exclusive production of anti-(1,4)-triazole derivatives. One of the compounds obtained shows general activity similar to that of imatinib; in particular, it was observed to be more effective in decreasing the fundamental function of cdc25A phosphatases in the K-562 cell line.
Anticancer agents continue to be a preferred therapeutic option for several malignancies. Despite their effectiveness, oncologists are continually looking for tumor-specific anticancer agents to prevent adverse effects in patients. Targeting of imaging agents to cancerous tissue is another area that is enthusiastically explored to circumvent some of the drawbacks that current imaging agents possess, including the inability to target small tumor cells, inadequate imaging period, and the risk of renal damage. Formulation scientists have explored nanotechnology-based delivery systems for targeting anticancer agents and tumor-imaging agents to cancer tissue. Targeting with nanotechnology-based delivery systems has been investigated by both passive and active mechanisms with significant clinical success. This review presents a discussion on targeting strategies used for the delivery of nanoparticles by passive and active mechanisms, focusing more specifically on active targeting of nanoparticles using albumin, folic acid, transferrin, and aptamers as targeting ligands.
Squalene is a triterpene widely distributed in nature that is an intermediate in the cholesterol biosynthesis pathway. The remarkable dynamic folded conformation of squalene has been used to chemically conjugate this lipid with various therapeutic molecules to construct nanoassemblies of 100-300 nm. In this review, we discuss the new concept of "squalenoylation" through application to anticancer (i.e. gemcitabine, paclitaxel, cisplatin etc.…) or antiviral (ddI, ddC) compounds. In a lego-type approach, it is also possible to construct multifunctional nanoparticles endowed with additional imaging functionalities (i.e. "Nanotheragnostics"). This new nanotechnology platform is expected to have important applications in pharmacology.
Folate (FOL) mediated poly-lactide-co-glycolide-polyethylene glycol nanoparticles (FOL-PEG-PLGA NPs) bearing paclitaxel (PTX) were prepared for the effective delivery of drug to endometrial carcinoma. The average size, zeta potential and encapsulation efficiency of FOL-targeted NPs were found to be around 220 nm, -30.43 mV and 95.6%. Cellular uptake was observed. The accumulation of FOL-targeted NPs depends on dual effects of passive and active targeting. The FOL-targeted PTX NPs showed a greater cytotoxicity against HEC-1A cancer cells in vitro and in vivo, which might be induced by apoptosis. H&E staining did not showed apparent tissue damage to liver and kidney of the mice after injecting NPs intravenously. These results suggest that the novel FOL-PEG-PLGA NPs could be a potential delivery system with excellent therapeutic efficacy for targeting the drugs to cancer cells.
We report the conjugation of the natural lipid squalene (SQ) with a small interfering RNA (siRNA), against the junction oncogene RET/PTC1, usually found in papillary thyroid carcinoma (PTC). The acyclic isoprenoid chain of squalene has been covalently coupled with siRNA RET/PTC1 at the 3'-terminus of the sense strand via maleimide-sulfhydryl chemistry. Remarkably, the linkage of siRNA RET/PTC1 to squalene led to an amphiphilic molecule that self-organized in H(2)O as siRNA-SQ RET/PTC1 nanoparticles (NPs). The siRNA-SQ RET/PTC1 NPs, stable in H(2)O, were used for biological studies. In vitro, they did not show any cytotoxicity. Interestingly, in vivo, on a mice xenografted RET/PTC1 experimental model, RET/PTC1-SQ NPs were found to inhibit tumor growth and RET/PTC1 oncogene and oncoprotein expression after 2.5 mg/kg cumulative dose intravenous injections. In conclusion, these results showed that the "squalenoylation" offers a new noncationic plate-form for the siRNA delivery.
Since activation of p53 in response to cytotoxic stress may have proapoptotic or protective effects depending on the nature of the injury, inhibitors of p53 may have therapeutic interest as modulators of chemotherapy toxicity or efficacy. In an attempt to identify novel p53 inhibitors, a quality collection of compounds structurally related to pifithrin-β were designed and synthesized as potential inhibitors of p53. The biochemical and biological evaluations supported that compounds of the tetrahydrobenzothiazole series were inhibitors of the p53 transcriptional activity and were effective in enhancing paclitaxel-induced apoptosis. In contrast, in spite of the increased cytotoxic potency, selected compounds of the benzothiazole series were not able to modulate the transcriptional activity of p53, as indicated by lack of change of p21 expression. The therapeutic interest of the compounds of the former series in combination with taxanes was confirmed in a human tumor xenograft model.
This study reports the design of a novel theragnostic nanomedicine which combines (i) the ability to target a prodrug of gemcitabine to an experimental solid tumor under the influence of a magnetic field with (ii) the imaging of the targeted tumoral nodule. This concept is based on the inclusion of magnetite nanocrystals into nanoparticles (NPs) constructed by self-assembling molecules of the squalenoyl gemcitabine (SQgem) bioconjugate. The nanocomposites are characterized by an unusually high drug loading, a significant magnetic susceptibility, and a low burst release. When injected to the L1210 subcutaneous mice tumor model, these magnetite/SQgem NPs were magnetically guided, and they displayed considerably greater anticancer activity than the other anticancer treatments (magnetite/SQgem NPs nonmagnetically guided, SQgem NPs, or gemcitabine free in solution). The histology and immunohistochemistry investigation of the tumor biopsies clearly evidenced the therapeutic superiority of the magnetically guided nanocomposites, while Prussian blue staining confirmed their accumulation at the tumor periphery. The superior therapeutic activity and enhanced tumor accumulation has been successfully visualized using T(2)-weighted imaging in magnetic resonance imaging (MRI). This concept was further enlarged by (i) the design of squalene-based NPs containing the T(1) Gd(3+) contrast agent instead of magnetite and (ii) the application to other anticancer squalenoyls, such as, cisplatin, doxorubicin, and paclitaxel. Thus, by combining different anticancer medicines as well as contrast imaging agents in NPs, we open the door toward generic conceptual framework for cancer treatment and diagnosis. This new theragnostic nanotechnology platform is expected to have important applications in cancer therapy.
The preparation and biological evaluation of a novel series of dimeric camptothecin derivatives are described. All the new compounds showed a significant ability to inhibit human tumor cell growth with IC(50) values ranging from 0.03 to 12.2 μM. The interference with the activity of the nuclear enzymes topoisomerases has been demonstrated, highlighting the poison effect of one of the obtained byproducts toward topoisomerase I. A moderate antiangiogenic activity has been demonstrated for one of the obtained compounds. Moreover, the effects of four new compounds on caspases activity and ROS generation have been studied on transgenic mouse cell.
Design and synthesis of an HDAC inhibitor and its merger with three tubulin binders to create releasable conjugate compounds is described. The biological evaluation includes: (a) in vitro reactivity with glutathione, (b) antiproliferative activity, (c) cell cycle analysis and (d) quantification of protein acetylation. The cellular pharmacology study indicated that the HDAC-inhibitor-drug conjugates retained antimitotic and proapoptotic activity with a reduced potency.
The preparation and biological evaluation of a novel series of dimeric epothilone A derivatives (1-6) are described. Two types of diacyl spacers were introduced to establish the various dimeric epothilone A constructs. The effect of these compounds on tubulin polymerization and their cytotoxicity against four different cancer cell lines are reported. Several of the newly synthesized compounds inhibit endothelial cell differentiation and endothelial cell migration that are key steps of the angiogenic process.
Differential scanning calorimetry was used to study the interaction of acyclovir and its prodrug squalenoyl-acyclovir (obtained by conjugation of 1,1',2-tris-nor-squalene acid (squaleneCOOH) with acyclovir) with biomembrane models made of DMPC multilamellar vesicles with the aim to verify whether a stronger interaction of the prodrug with respect to the free drug can be obtained. Multilamellar vesicles were prepared in the presence of increasing molar fractions of acyclovir, squaleneCOOH or prodrug and the effect of the compounds on the thermotropic behavior of vesicles was researched, revealing no effect of acyclovir but a strong effect of squaleneCOOH and prodrug. To evaluate if acyclovir, squaleneCOOH and prodrug can be absorbed by the biomembrane model, an experiment was carried out in which the considered compounds were left in contact with the biomembrane model and their eventual uptake was evaluated analyzing the effect on the thermotropic behavior of the biomembrane model. A very small uptake was revealed for all the compounds. To check the potential use of liposomes as a delivery system for the prodrug, the biomembrane models were incubated with liposomes loaded with the compounds and the compounds transferring from the loaded liposomes to the unloaded biomembrane model was followed. The results suggest that liposomes could be used to deliver the squalenoyl-acyclovir to the biomembrane model.
This review highlights the properties of nanoparticles used in targeted drug delivery, including delivery to cells as well as organelle targets, some of the known pharmacokinetic properties of nanoparticles, and their typical modifications to allow for therapeutic delivery. Nanoparticles exploit biological pathways to achieve payload delivery to cellular and intracellular targets, including transport past the blood-brain barrier. As illustrative examples of their utility, the evaluation of targeted nanoparticles in the treatment of cancers and diseases of the central nervous system, such as glioblastoma multiforme, neurovascular disorders, and neurodegenerative diseases, is discussed.
The Leu-enkephalin dalargin and the Met-enkephalin kyotorphin normally do not cross the blood-brain barrier (BBB) when given systemically. To transport these neuropeptides across the BBB they were adsorbed onto the surface of poly(butylcyanoacrylate) nanoparticles (NPs) and the NPs were coated with polysorbate 80. Central analgesia was measured by the hot plate test in mice. The antidepressant amitriptyline, which normally penetrates the BBB, was used to examine the versatility of the NP method. The concentration of amitriptyline in serum and brain of mice was determined by a gas chromatographic method. Furthermore, NPs were fabricated with different stabilizers. After the adsorption of the peptides on polysorbate 85-stabilized NPs, analgesia was noted after intravenous application when NPs were not coated. The amitriptyline level was significantly enhanced in brain when the substance was adsorbed onto the NP and coated or when the particles were stabilized with polysorbate 85.
Tyrosine nitration of proteins is emerging as a post-translational modification playing a role in physiological conditions. Looking for the molecular events triggered by nitric oxide in nerve growth factor-induced neuronal differentiation, we now find that nitration occurs on the microtubule-associated protein tau. In differentiated PC12 cells, we have identified as tau a nitrated protein that co-immunoprecipitates with alpha-tubulin and indicated that the modified protein is associated with the cytoskeleton but it is confined to a restricted cell region. This paper supplies the first evidence that nitration of tau occurs in a physiological process and suggests that it could play a role in neuronal differentiation.
The sulforhodamine B (SRB) assay was developed by Skehan and colleagues to measure drug-induced cytotoxicity and cell proliferation for large-scale drug-screening applications. Its principle is based on the ability of the protein dye sulforhodamine B to bind electrostatically and pH dependent on protein basic amino acid residues of trichloroacetic acid-fixed cells. Under mild acidic conditions it binds to and under mild basic conditions it can be extracted from cells and solubilized for measurement. Results of the SRB assay were linear with cell number and cellular protein measured at cellular densities ranging from 1 to 200% of confluence. Its sensitivity is comparable with that of several fluorescence assays and superior to that of Lowry or Bradford. The signal-to-noise ratio is favorable and the resolution is 1000-2000 cells/well. It performed similarly compared to other cytotoxicity assays such as MTT or clonogenic assay. The SRB assay possesses a colorimetric end point and is nondestructive and indefinitely stable. These practical advances make the SRB assay an appropriate and sensitive assay to measure drug-induced cytotoxicity even at large-scale application.
Nucleoside analogues display significant anticancer or antiviral activity by interfering with DNA synthesis. However, there are some serious restrictions to their use, including their rapid metabolism and the induction of resistance. We have discovered that the linkage of nucleoside analogues to squalene leads to amphiphilic molecules that self-organize in water as nanoassemblies of 100-300 nm, irrespective of the nucleoside analogue used. The squalenoyl gemcitabine exhibited superior anticancer activity in vitro in human cancer cells and gemcitabine-resistant murine leukemia cells, and in vivo in experimental leukemia both after intravenous and oral administration. The squalenoylation of other antiretroviral nucleosides also led to more potent drugs when tested in primary cultures of HIV-infected lymphocytes. Thus, the squalenoylation is an original technology platform for generating more potent anticancer and antiviral nanomedicines.
A series of novel hybrid compounds obtained by the attachment of anhydrovinblastine, vinorelbine, and vindoline to thiocolchicine, podophyllotoxin, and baccatin III are described. Two types of diacyl spacers are introduced. The influence of the hybrid compounds on tubulin polymerization is reported. The results highlight the importance of the length of the spacer. Immunofluorescence microscopy and flow cytometry measurements that compound with the best in vitro activity could disrupt microtubule networks in cell and prevent the formation of the proper spindle apparatus, thereby causing cell cycle arrest in the G2/M phase. The newly synthesized compounds were tested in the human lung cancer cell line A549.
Cellular uptake of vitamin folic acid occurs via folate-receptor mediated endocytosis. Many types of cancer cells express high levels of folate receptors as they need continuous supply of this vitamin for their proliferation. With an objective to use folic acid as a 'Trojan Horse' to transport anticancer drugs into cancer cells, a novel heterobifunctional disulfide-containing linker was synthesized and utilized to covalently link an amino- and hydroxyl-containing anticancer drug, and an appropriately functionalized folic acid to create novel targetable folate-drug conjugates that are shown to release free drugs under biologically relevant pH via sulfhydryl-assisted cleavage of the self-immolative disulfide-containing linker.
Nanocarriers' entry into the cell: relevance to drug delivery
- H Hillaireau
- P Couvreur
H. Hillaireau, P. Couvreur, Nanocarriers' entry into the cell: relevance to drug
delivery, Cell. Mol. Life Sci. 66 (2009) 2873e2896.