Safety,Pharmacokinetics and Biodistribution Studies of a β-galactoside Prodrug of Doxorubicin for Improvement of Tumor Selective Chemotherapy

Anthracycline antibiotics, particularly doxorubicin (DOX) and daunorubicin, have been used extensively in the treatment of human malignancies. However, cardiotoxicity and multidrug resistance are significant problems that limit the clinical efficacy of such agents. Rational design to avoid these side effects includes strategies such as drug targeting and prodrug synthesis. The DOX prodrug N-(β-D-glucopyranosylbenzyloxycarbonyl)-doxorubicin (prodrug 1) was synthesized for specific activation by β-galactosidase, which is expected to release in necrotic areas of tumor lesions. Described here is the safety, pharmacokinetics, and biodistribution studies of a β-galactoside prodrug of DOX. In vivo safety evaluation was done in the Ehrlich Ascites Carcinoma (EAC) tumor model. The dose of DOX was 8 mg/kg and the dose of prodrug was 8 mg/kg and 24 mg/kg of DOX equivalents. Our results on cytotoxicity, which demonstrated compression in the number of EAC cells and their viability, substantiate these data. Prodrug 1 was safe up to a dose of 24 mg/kg of DOX equivalents in EAC mice. The pharmacokinetics and biodistribution of prodrug (300 mg/kg) in normal mice were determined and compared with DOX (20 mg/kg). Administration of DOX in normal mice resulted in a peak plasma concentration of 19.45 μM (t = 30 minutes). Prodrug injection resulted in 3- to 16-fold lower concentrations in the tissues of normal mice. As it is more polar, lower levels were observed in tissues and plasma in contrast to the parent compound DOX. In vivo safety studies have shown that prodrug 1 had a maximum tolerated dose compared with DOX and led to improved pharmacokinetics in normal mice.     

Effect of intratumoral injection on the biodistribution and therapeutic potential of novel chemophor EL-modified single-walled nanotube loading doxorubicin

The purpose of this study is to evaluate in vivo efficacy and loco-regional distribution of a doxorubicin (DOX)-loaded Polyoxyl 35 Castor Oil (Cremophor EL, CrEL) noncovalent modified single-walled carbon nanotubes (SWNTs) formulation in a sarcoma tumor model after intratumoral injection. The drug loaded SWNTs were successfully prepared via physical absorption, which was confirmed by UV-vis-NIR absorbance spectra and dynamic light scattering assay. Solid tumor models were obtained by injecting mouse sarcoma 180 cells into the thighs of ICR mice. CrEL-SWNTs-DOX, CrEL-SWNTs, free DOX and saline (control) were intratumorally injected after 5 days post transplantation. The biodistribution studies demonstrated that intratumoral delivery of CrEL-SWNTs-DOX resulted in longer drug retention time in tumor, higher tumor level (27.6-fold than that of free DOX), as well as less accumulation in other solid tissues, especially in heart. Furthermore, in vivo anti-tumor activity results showed that CrEL-SWNTs-DOX could effectively suppress the tumor growth than free DOX and the control, attributing to its enhanced intratumoral DOX level. The histopathological findings revealed that the new carbon nanomaterials were a safe vehicle for topical drug delivery systems. It is concluded that this noncovalent modification of carbon nanotubes by CrEL for anticancer agents might be a promising alternative for cancer treatment.     

Preparation and properties of a novel drug delivery system with both magnetic and biomolecular targeting

By loading doxorubicin (DOX) on 5-carboxyl-fluorescein (FAM) labeled AGKGTPSLETTP peptide (A54) coupled starch-coated iron oxide nanoparticles (SIONs), we prepared a novel aqueous drug delivery system with both magnetic and biomolecular targeting, which was specific to human hepatocellular carcinoma cell line BEL-7402. The saturated extent of adsorption reached 2.0 mg DOX/mg A54-SIONs at 28°C, which provided a rather high dose of DOX loading for application. Tests in vitro demonstrated the specificity of DOX-loaded A54-SIONs to BEL-7402 cells. The microscopy images proved that DOX-loaded A54-SIONs were successfully targeted to tumor tissue of nude mice with an external magnetic field in vivo. MTT assay showed higher cytostatic effect of DOX-loaded A54-SIONs to hepatocellular carcinoma cells BEL-7402 than that of DOX-loaded SIONs.     

Cyclo(RGD)‐Decorated Reduction‐Responsive Nanogels Mediate Targeted Chemotherapy of Integrin Overexpressing Human Glioblastoma In Vivo

Cyclo(Arg‐Gly‐Asp) peptide (cRGD) decorated disulfide (SS) containing poly(vinyl alcohol) nanogels (cRGD‐SS‐NGs) with an average diameter of 142 nm prepared by inverse nanoprecipitation, “click” reaction, and cRGD conjugation are developed for targeted treatment of integrin overexpressing human glioblastoma in vivo. Doxorubicin (DOX) release from cRGD‐SS‐NGs is highly inhibited under physiological conditions, while accelerated at endosomal pH and in response to cytoplasmic concentration of glutathione. Confocal microscopy shows that cRGD‐SS‐NGs facilitate the cellular uptake and intracellular DOX release in α_vβ₃ integrin overexpressing human glioblastoma U87‐MG cells. DOX‐loaded cRGD‐SS‐NGs present much better killing activity toward U87‐MG cells than that for nontargeted nanogels determined by MTT assay. The in vivo imaging and biodistribution studies reveal that DOX‐loaded cRGD‐SS‐NGs have a much better tumor targetability toward human U87‐MG glioblastoma xenograft in nude mice. Also the tumor growth is effectively inhibited by treatment with DOX‐loaded cRGD‐SS‐NGs, while continuous tumor growth is observed for mice treated with nondecorated nanogels as well as free DOX. Furthermore, the treatment with DOX‐loaded cRGD‐SS‐NGs has much fewer side effects, rendering these nanogels as a new platform for cancer chemotherapy in vivo.     

Programmable Codelivery of Doxorubicin and Apatinib Using an Implantable Hierarchical‐Structured Fiber Device for Overcoming Cancer Multidrug Resistance

Multiple drug resistance (MDR) of cancer cells is a major cause of chemotherapy failure. It is currently a great challenge to develop a direct and effective strategy for continuously inhibiting the P‐glycoprotein (P‐gp) drug pump of MDR tumor cells, thus enhancing the intracellular concentration of the therapeutic agent for effectively killing MDR tumor cells. Here, a new implantable hierarchical‐structured ultrafine fiber device is developed via a microfluidic‐electrospinning technology for localized codelivery of doxorubicin (DOX) and apatinib (AP). An extremely high encapsulation efficiency of ≈99% for the dual drugs is achieved through this strategy. The release of the loaded dual drugs can be controlled in a programmable release model with a rapid release of the micelles, while AP is slowly released. The sustained release of AP can continuously inhibit the P‐gp drug pump of MDR tumor cells, increasing the intracellular DOX accumulation. The in vivo DOX biodistribution displays that the DOX accumulation in the tumor tissues achieves 17.82% after implanting the fiber device for 72 h, which is 6.36‐fold higher than that of the intravenously injected DOX. Importantly, the fiber device shows an excellent antitumor effect on MDR tumor‐bearing mice with low systemic toxicity.     

Physicochemical stability,pharmacokinetic, and biodistribution evaluation of paclitaxel solid dispersion prepared using supercritical antisolvent process

Aim: To investigate the physicochemical stability, pharmacokinetics (PK), and biodistribution of paclitaxel (PTX) from paclitaxel solid dispersion (PSD) prepared by supercritical antisolvent (SAS) process.

Methods: Physicochemical stability was performed in accelerated (40°C 70?±?5% RH) and stress (60°C) storage conditions for a period of 6 months and 4 weeks, respectively. PK and biodistribution studies were performed in rats following i.v. administration of PTX equivalent to 6 and 12?mg/kg formulations.

Results: Physical stability of PSD showed excellent stability with no recrystallization of the amorphous form. Chemical stability of PSD in terms of % PTX remaining was 98.2?±?0.6% at 6 months and 97.9?±?0.3% at 4 weeks of accelerated and stress conditions, respectively. The PK study showed a nonlinear increase in AUC with increasing dose, that is, 100% increase in dose (from 6 to 12?mg/kg) resulted in 405.90% increase in AUC. Unlike PK study, the organ distribution study of PTX from PSD showed linear relationship with dose escalation. The order of organ distribution of PTX from highest to lowest for both PSD and Taxol^® was liver>kidney>lung>brain.

Conclusions: This study demonstrated excellent physicochemical stability with insight information on the PK and biodistribution of PTX from PSD prepared by SAS process.     

Studies of erythromycin maltobionate, a new derivative of erythromycin

Erythromycin maltobionate, a new water-soluble derivative of erythromycin, was prepared, and its physicochemical and biological properties were evaluated. The derivative has considerable solubility in organic solvents. Its partition coefficient data in different organic solvent-water systems indicate it is possibly well distributed in various tissues in vivo. Antimicrobial potency in vitro of the derivative is 589 μg/mg, and its antimicrobial spectrum is comparable to that of the parent antibiotic. The LD₅₀ value of the new derivative in mice intraperitoneally is 244.7 mg/kg. Results of this and the previous investigation of pharmacokinetics and protein binding indicate that the new derivative erythromycin maltobionate has a potential for possible clinical application.     

Studies of Erythromycin Maltobionate,a New Derivative of Erythromycin

Erythromycin maltobionate, a new water-soluble derivative of erythromycin, was prepared, and its physicochemical and biological properties were evaluated. The derivative has considerable solubility in organic solvents. Its partition coefficient data in different organic solvent-water systems indicate it is possibly well distributed in various tissues in vivo. Antimicrobial potency in vitro of the derivative is 589 μg/mg, and its antimicrobial spectrum is comparable to that of the parent antibiotic. The LD₅₀ value of the new derivative in mice intraperitoneally is 244.7 mg/kg. Results of this and the previous investigation of pharmacokinetics and protein binding indicate that the new derivative erythromycin maltobionate has a potential for possible clinical application.     

pH‐ and NIR Light‐Responsive Polymeric Prodrug Micelles for Hyperthermia‐Assisted Site‐Specific Chemotherapy to Reverse Drug Resistance in Cancer Treatment

Despite the exciting advances in cancer chemotherapy over past decades, drug resistance in cancer treatment remains one of the primary reasons for therapeutic failure. IR‐780 loaded pH‐responsive polymeric prodrug micelles with near infrared (NIR) photothermal effect are developed to circumvent the drug resistance in cancer treatment. The polymeric prodrug micelles are stable in physiological environment, while exhibit fast doxorubicin (DOX) release in acidic condition and significant temperature elevation under NIR laser irradiation. Phosphorylcholine‐based biomimetic micellar shell and acid‐sensitive drug conjugation endow them with prolonged circulation time and reduced premature drug release during circulation to conduct tumor site‐specific chemotherapy. The polymeric prodrug micelles combined with NIR laser irradiation could significantly enhance intracellular DOX accumulation and synergistically induce the cell apoptosis in DOX‐resistant MCF‐7/ADR cells. Meanwhile, the tumor site‐specific chemotherapy combined with hyperthermia effect induces significant inhibition of MCF‐7/ADR tumor growth in tumor‐bearing mice. These results demonstrate that the well‐designed IR‐780 loaded polymeric prodrug micelles for hyperthermia‐assisted site‐specific chemotherapy present an effective approach to reverse drug resistance.     

Pharmacokinetics in rats and efficacy in murine ovarian cancer model for solid lipid nanoparticles loading docetaxel

Docetaxel was used extensively in ovarian cancer treatment in the combination with platinum compound. However, the serious side effect of clinically available formulation limits its further application. The aim of this work was to evaluate the pharmacokinetic behavior, acute toxicity and in vivo antitumor efficacy in murine ovarian cancer model of docetaxel loaded solid lipid nanoparticles (DSN). The particle size of DSN was 97.4 +/- 6.4 nm, and the encapsulation efficiency and loading capacity were 91.1 +/- 1.5% and 3.52 +/- 0.05%, respectively. The release behavior of docetaxel from DSN showed that only 45% of docetaxel was released within 24 h. The pharmacokinetics and biodistribution showed that the half-life (t1/2) and mean residence time (MRT) of docetaxel in DSN treated rats were significantly elongated because of the redistribution of docetaxel from reticulo-endothelial system (RES) to circulation system. Compared with Taxotere, DSN showed more potent in vivo anti-ovarian cancer activity with higher maximum tolerated dose (MTD). Our results suggested for the first time that solid lipid nanoparticles could be a potential candidate to enhance the efficacy of anti-ovarian cancer of docetaxel with low toxicity. The systematical study on pharmacokinetics, biodistribution, in vivo anti-tumor activity and MTD of DSN could improve the understanding of increased antitumor activity of DSN in vivo.     

Pulmonary Codelivery of Doxorubicin and siRNA by pH‐Sensitive Nanoparticles for Therapy of Metastatic Lung Cancer

A pulmonary codelivery system that can simultaneously deliver doxorubicin (DOX) and Bcl2 siRNA to the lungs provides a promising local treatment strategy for lung cancers. In this study, DOX is conjugated onto polyethylenimine (PEI) by using cis‐aconitic anhydride (CA, a pH‐sensitive linker) to obtain PEI‐CA‐DOX conjugates. The PEI‐CA‐DOX/siRNA complex nanoparticles are formed spontaneously via electrostatic interaction between cationic PEI‐CA‐DOX and anionic siRNA. The drug release experiment shows that DOX releases faster at acidic pH than at pH 7.4. Moreover, PEI‐CA‐DOX/Bcl2 siRNA complex nanoparticles show higher cytotoxicity and apoptosis induction in B16F10 cells than those treated with either DOX or Bcl2 siRNA alone. When the codelivery systems are directly sprayed into the lungs of B16F10 melanoma‐bearing mice, the PEI‐CA‐DOX/Bcl2 siRNA complex nanoparticles exhibit enhanced antitumor efficacy compared with the single delivery of DOX or Bcl2 siRNA. Compared with systemic delivery, most drug and siRNA show a long‐term retention in the lungs via pulmonary delivery, and a considerable number of the drug and siRNA accumulate in tumor tissues of lungs, but rarely in normal lung tissues. The PEI‐CA‐DOX/Bcl2 siRNA complex nanoparticles are promising for the treatment of metastatic lung cancer by pulmonary delivery with low side effects on the normal tissues.     

Facile Fabrication of Tumor Redox‐Sensitive Nanoassemblies of Small‐Molecule Oleate Prodrug as Potent Chemotherapeutic Nanomedicine

The conjugate of paclitaxel (PTX) and docosahexaenoic acid has entered into clinical trials. However, the most recent clinical outcomes fell short of expectations, due to the extremely slow drug release from the hydrophobic conjugates. Herein, a novel prodrug‐based nanoplatform self‐assembled by the disulfide bond linked conjugates of PTX and oleic acid for rapid and differential release of PTX in tumor cells is reported. This redox‐responsive prodrug‐nanosystem demonstrates multiple therapeutic advantages, including one‐step facile fabrication, high drug‐loading efficiency (56%, w/w), on‐demand drug release responding to redox stimuli, as well as favorable cellular uptake and biodistribution. These advantages result in significantly enhanced antitumor efficacy in vivo, with the tumor almost completely disappearing in mice. Such a uniquely engineered prodrug‐nanosystem has great potential to be used as potent chemotherapeutic nanomedicine in clinical cancer therapy.     

Effect of intratumoral injection on the biodistribution and therapeutic potential of novel chemophor EL-modified single-walled nanotube loading doxorubicin

The purpose of this study is to evaluate in vivo efficacy and loco-regional distribution of a doxorubicin (DOX)-loaded Polyoxyl 35 Castor Oil (Cremophor EL, CrEL) noncovalent modified single-walled carbon nanotubes (SWNTs) formulation in a sarcoma tumor model after intratumoral injection. The drug loaded SWNTs were successfully prepared via physical absorption, which was confirmed by UV-vis-NIR absorbance spectra and dynamic light scattering assay. Solid tumor models were obtained by injecting mouse sarcoma 180 cells into the thighs of ICR mice. CrEL-SWNTs-DOX, CrEL-SWNTs, free DOX and saline (control) were intratumorally injected after 5 days post transplantation. The biodistribution studies demonstrated that intratumoral delivery of CrEL-SWNTs-DOX resulted in longer drug retention time in tumor, higher tumor level (27.6-fold than that of free DOX), as well as less accumulation in other solid tissues, especially in heart. Furthermore, in vivo anti-tumor activity results showed that CrEL-SWNTs-DOX could effectively suppress the tumor growth than free DOX and the control, attributing to its enhanced intratumoral DOX level. The histopathological findings revealed that the new carbon nanomaterials were a safe vehicle for topical drug delivery systems. It is concluded that this noncovalent modification of carbon nanotubes by CrEL for anticancer agents might be a promising alternative for cancer treatment.     

Enhanced Release of Drugs from Silicone Elastomers: (IV) Subcutaneous Controlled Release of Indomethacin and in Vivo/In Vitro Correlations

Abstract

The subcutaneous controlled release of indomethacin from implants was studied. The subderraal implants were prepared from silicone elastomers containing various levels of glycerol. The in vitro and in vivo releases of indomethacin were observed to follow a matrix diffusion-control mechanism. The release flux of indomethacin was enhanced when glycerol was incorporated into the silicone elastomers. An in vivo/in vivo correlation coefficient of 0.85 (< 0.05) was obtained for implants containing up to 20% (w/w) of glycerol. The survival rates on the ninth day post-implantation were determined to be 20, 65, and 100%, respectively, for the mice receiving implants containing 20, 10, and 0% glycerol. An LD₅₀ dose of 34 mg/kg was assessed for the subcutaneous controlled administration of indomethacin in CD-1 mice, which is not significantly different from the intraperitoneal LD₅₀ of 28 mg/kg and the intravenous LD₅₀ of 40 mg/kg.     

Studies on Bi-Layer Osmotic Pump Tablets of Water-Insoluble Allopurinol with Large Dose: In Vitro and In Vivo

Controlled release bi-layer osmotic pump tablets (BOPT) of water—insoluble allopurinol with large dose (150 mg/BOPT) were successfully prepared merely with sodium chloride as osmotic promoting agent and polyethylene oxide (PEO) as suspending agent. Formulations of the two kinds of agents were investigated in order to discuss their effects on the release behavior of BOPT, and then the optimal formulation was evaluated. The pharmacokinetics studies of allopurinol and its active metabolite oxypurinol in two-preparation and two-period crossover design relative to the equivalent dose of commercially common allopurinol tablets were evaluated in six Beagle dogs. And the pharmacokinetics results showed that allopurinol BOPT were able to provide a slow release of allopurinol, and oxypurinol were bioequivalent between allopurinol BOPT and common allopurinol tablets. A good in vitro–in vivo correlation of allopurinol was also proved. In conclusion, water-insoluble drugs with large dose can be designed to BOPT for efficacy and safety use.     

Bioinspired Nano‐Prodrug with Enhanced Tumor Targeting and Increased Therapeutic Efficiency

Nanotechnology‐based drug delivery has a great potential to revolutionize cancer treatment by enhancing anticancer drug efficacy and reducing drug toxicity. Here, a bioinspired nano‐prodrug (BiNp) assembled by an antineoplastic peptidic derivative (FA‐KLA‐Hy‐DOX), a folate acid (FA)‐incorporated proapoptotic peptide (KLAKLAK)₂ (KLA) to doxorubicin (DOX) via an acid‐labile hydrozone bond (Hy) is constructed. The hydrophobic antineoplastic agent DOX is efficiently shielded in the core of nano‐prodrug. With FA targeting moieties on the surface, the obtained BiNp shows significant tumor‐targeting ability and enhances the specific uptake of cancer cells. Upon the trigger by the intracellular acidic microenvironment of endosomes, the antineoplastic agent DOX is released on‐demand and promotes the apoptosis of cancer cells. Simultaneously, the liberated FA‐KLA can induce the dysfunction of mitochondria and evoke mitochondria‐dependent apoptosis. In vitro and in vivo results show that the nano‐prodrug BiNp with integrated programmed functions exhibits remarkable inhibition of tumor and achieves a maximized therapeutic efficiency with a minimized side effect.     

Nanomaterial‐Based Organelles Protect Normal Cells against Chemotherapy‐Induced Cytotoxicity

Chemotherapy‐induced cytotoxicity in normal cells and organs triggers undesired lesions. Although targeted delivery is used extensively, more than half of the chemotherapy dose still concentrates in normal tissues, especially in the liver. Enabling normal cells or organs to defend against cytotoxicity represents an alternative method for improving chemotherapy. Herein, rationally designed nanomaterials are used as artificial organelles to remove unexpected cytotoxicity in normal cells. Nanocomposites of gold‐oligonucleotides (Au‐ODN) can capture intracytoplasmic doxorubicin (DOX), a standard chemotherapy drug, blocking the drug's access into the cell nucleus. Cells with implanted Au‐ODN are more robust since their viability is maintained during DOX treatment. In vivo experiments confirm that the Au‐ODN nanomaterials selectively concentrate in hepatocytes and eliminate DOX‐induced hepatotoxicity, increasing the cell's capacity to resist the threatening chemotherapeutic milieu. The finding suggests that introducing functional materials as biological devices into living systems may be a new strategy for improving the regulation of cell fate in more complex conditions and for manufacturing super cells.     

Studies on bi-layer osmotic pump tablets of water-insoluble allopurinol with large dose: in vitro and in vivo

Controlled release bi-layer osmotic pump tablets (BOPT) of water—insoluble allopurinol with large dose (150 mg/BOPT) were successfully prepared merely with sodium chloride as osmotic promoting agent and polyethylene oxide (PEO) as suspending agent. Formulations of the two kinds of agents were investigated in order to discuss their effects on the release behavior of BOPT, and then the optimal formulation was evaluated. The pharmacokinetics studies of allopurinol and its active metabolite oxypurinol in two-preparation and two-period crossover design relative to the equivalent dose of commercially common allopurinol tablets were evaluated in six Beagle dogs. And the pharmacokinetics results showed that allopurinol BOPT were able to provide a slow release of allopurinol, and oxypurinol were bioequivalent between allopurinol BOPT and common allopurinol tablets. A good in vitro-in vivo correlation of allopurinol was also proved. In conclusion, water-insoluble drugs with large dose can be designed to BOPT for efficacy and safety use.     

Multihydroxylated [Gd@C82(OH)22]n nanoparticles: antineoplastic activity of high efficiency and low toxicity

[Gd@C82(OH)22]n particles (22 nm in a saline solution) of a dose level as low as 10(-7) mol/kg exhibit a very high antineoplastic efficiency ( approximately 60%) in mice. A dose increment of 1 x 10(-7) mol/kg increases the tumor inhibition rate 26%. [Gd@C82(OH)22]n particles have a strong capacity to improve immunity and interfere with tumor invasion in normal muscle cells, nearly without toxicity in vivo and in vitro. Unlike conventional antineoplastic chemicals, the high antitumor efficiency of nanoparticles is not due to toxic effects to cells because they do not kill the tumor cells directly and only about 0.05% of the used dose is found in the tumor tissues. Results suggest that fullerene derivatives with proper surface modifications and sizes may help realize the dream of tumor chemotherapeutics of high-efficacy and low-toxicity.     

Octreotide-modification enhances the delivery and targeting of doxorubicin-loaded liposomes to somatostatin receptors expressing tumor in vitro and in vivo

Octreotide is believed to be the ligand of somatostatin receptors (SSTRs) which are widely used in tumor diagnosis and clinical therapy. In the present work, a new targeting conjugate, octreotide-polyethylene glycol-phosphatidylethanolamine (Oct-PEG-PE), was developed for the assembling of liposome, and the effect of octreotide-modification on the enhancement of the delivery and targeting of doxorubicin-loaded liposomes was investigated in vitro and in vivo. Oct-PEG-PE was synthesized by a three-step reaction involving two derivative intermediate formations of bis (p-nitrophenyl carbonate)-PEG ((pNP)(2)-PEG) and pNP-PEG-PE. The Oct-modified and unmodified liposomes (DOX-OL and DOX-CL) were prepared by the ammonium sulfate gradient method. Both drug uptake assay and cell apoptosis assay suggested that DOX-OL noticeably increased the uptake of DOX in SMMC-7721 cells and showed a more significant cytotoxicity, compared with DOX-CL. The effect of DOX-OL was remarkably inhibited by free octreotide. In contrast, no significant difference in drug cytotoxicity was found between DOX-OL and DOX-CL in CHO cells without obvious expression of SSTRs. The study of ex vivo fluorescence tissues imaging of BALB/c mice and in vivo tissue distribution of B16 tumor-bearing mice indicated that DOX-OL caused remarkable accumulation of DOX in melanoma tumors and the pancreas, in which the SSTRs are highly expressed.