Engineered urolithin A-laden functional polymer-lipid hybrid nanoparticles prevent cisplatin-induced proximal tubular injury in vitro.

Functional polymer-lipid hybrid nanoparticles (H-NPs) are a promising class of nanocarriers that combine the benefits of polymer and lipid nanoparticles, offering biocompatibility, structural stability, high loading capacity, and, most importantly, superior surface functionalization. Here, we report the synthesis and design of highly functional H-NPs with specificity toward the transferrin receptor (TfR), using a small molecule ligand, gambogic acid (GA). A fluorescence study revealed the molecular orientation of H-NPs, where the lipid-dense core is surrounded by a polymer exterior, functionalized with GA. Urolithin A, an immunomodulator and anti-inflammatory agent, served as a model drug-like compound to prepare H-NPs via traditional emulsion-based techniques, where H-NPs led to smaller particles (132 nm) and superior entrapment efficiencies (70 % at 10 % drug loading) compared to GA-conjugated polymeric nanoparticles (P-NPs) (157 nm and 52 % entrapment efficiency) and solid lipid nanoparticles (L-NPs) (186 nm and 29 % entrapment efficiency). H-NPs showed superior intracellular accumulation compared to individual NPs using human small intestinal epithelial (FHs 74) cells. The in vitro efficacy was demonstrated by flow cytometry analysis, in which UA-laden H-NPs showed excellent anti-inflammatory properties in cisplatin-induced injury in healthy human proximal tubular cell (HK2) model by decreasing the TLR4, NF-κß, and IL-ß expression. This preliminary work highlights the potential of H-NPs as a novel functional polymer-lipid drug delivery system, establishing the foundation for future research on its therapeutic potential in addressing chemotherapy-induced acute kidney injury in cancer patients.

Glucose-Responsive Microgel Comprising Conventional Insulin and Curcumin-Laden Nanoparticles: a Potential Combination for Diabetes Management.

Successful management of type 2 diabetes mellitus (T2DM), a complex and chronic disease, requires a combination of anti-hyperglycemic and anti-inflammatory agents. Here, we have conceptualized and tested an integrated "closed-loop mimic" in the form of a glucose-responsive microgel (GRM) based on chitosan, comprising conventional insulin (INS) and curcumin-laden nanoparticles (nCUR) as a potential strategy for effective management of the disease. In addition to mimicking the normal, on-demand INS secretion, such delivery systems display an uninterrupted release of nCUR to combat the inflammation, oxidative stress, lipid metabolic abnormality, and endothelial dysfunction components of T2DM. Additives such as gum arabic (GA) led to a fivefold increased INS loading capacity compared to GRM without GA. The GRMs showed excellent in vitro on-demand INS release, while a constant nCUR release is observed irrespective of glucose concentrations. Thus, this study demonstrates a promising drug delivery technology that can simultaneously, and at physiological/pathophysiological relevance, deliver two drugs of distinct physicochemical attributes in the same formulation.

Low-Dose Curcumin Nanoparticles Normalise Blood Pressure in Male Wistar Rats with Diet-Induced Metabolic Syndrome.

Nanoparticle formulations improve bioavailability and so may allow low-dose formulations of food-derived compounds such as curcumin to attenuate chronic systemic disease despite intrinsically low oral bioavailability. The current study induced metabolic syndrome in male Wistar rats aged eight-nine weeks using a high-carbohydrate, high-fat diet (H) with corn starch diet (C) as control. Using a reversal protocol, rats were given curcumin as either nanoparticles encapsulated in poly(lactic-co-glycolic acid) (5 mg/kg/day, HCNP) or as an unformulated low dose or high-dose suspension in water (low-dose, 5 mg/kg/day, HC5; high-dose, 100 mg/kg/day, HC100) or blank nanoparticles (HBNP) for the final eight weeks of the 16 week study. We analysed cardiovascular parameters including systolic blood pressure and left ventricular diastolic stiffness along with histopathology, liver parameters including plasma liver enzymes, histopathology and metabolic parameters, including glucose tolerance, blood lipid profile and body composition, and plasma curcumin concentrations. HC100 and HCNP but not HBNP normalised systolic blood pressure (C = 120 ± 4; H = 143 ± 5; HBNP = 141 ± 3; HC5 = 143 ± 4; HC100 = 126 ± 4; HCNP = 128 ± 4 mmHg), left ventricular diastolic stiffness and liver fat deposition. No other improvements were induced in HC100 or HCNP or other intervention groups (HC5 and HBNP). We conclude that 5 mg/kg/day curcumin nanoparticles in H rats showed similar improvements in cardiovascular function as 100 mg/kg/day unformulated curcumin correlating with similar plasma curcumin concentrations.

Oral Drug Delivery Technologies-A Decade of Developments.

Advanced drug delivery technologies, in general, enable drug reformulation and administration routes, together contributing to life-cycle management and allowing the innovator to maintain the product monopoly. Over the years, there has been a steady shift from mere life-cycle management to drug repurposing-applying delivery technologies to tackle solubility and permeability issues in early stages or safety and efficacy issues in the late stages of drug discovery processes. While the drug and the disease in question primarily drive the choice of route of administration, the oral route, for its compliance and safety attributes, is the most preferred route, particularly when it comes to chronic conditions, including pain, which is not considered a disease but a symptom of a primary cause. Therefore, the attempt of this review is to take a stock of the advances in oral delivery technologies that are applicable for injectable to oral transformation, improve risk-benefit profiles of existing orals, and apply them in the early discovery program to minimize the drug attrition rates.

Antacid co-encapsulated polyester nanoparticles for peroral delivery of insulin: development, pharmacokinetics, biodistribution and pharmacodynamics.

The in vitro/in vivo characterization of antacid-insulin co-encapsulated poly(lactide-co-glycolide) (PLGA) nanoparticles is presented here. The optimized nanoparticle composition has 1% surfactant (didodecyl dimethylammonium bromide) and 2% antacid (magnesium hydroxide or zinc carbonate) in the size range ~136-143nm with ~81-85% entrapment of insulin at a 4% (w/w) initial load to that of polymer. Molecular characterization using circular dichroism, fluorescence and Fourier transform infrared spectroscopy showed that the structural integrity of insulin was maintained during formulation. Furthermore, the encapsulated insulin was well protected under in vitro simulated gastric and intestinal fluids. Nanoparticle insulin results in six fold increase in oral bioavailability to that of plain insulin in healthy rats. In diabetic rats, a 120 IU/kg oral dose of insulin nanoparticles achieved an equivalent blood glucose lowering effect to a 20 IU/kg subcutaneous (sc) dose of insulin solution, the nadir in blood glucose concentration occurring 24h and 1h post-administration, respectively. Both sc insulin and oral nanoparticle insulin partially attenuated hyperglycemia-induced inflammation caused by tumor necrosis factor α, but not by interleukin-6 or C-reactive protein; on the other hand, subcutaneous insulin was found to be more effective on lipid profile measured in the form of high density lipoprotein, cholesterol and triglyceride. Successful oral insulin could be beneficial in type II complications.

Towards scale-up and regulatory shelf-stability testing of curcumin encapsulated polyester nanoparticles.

This study reports scale-up and shelf-stability of curcumin encapsulated poly(lactic acid-co-glycolic acid) (PLGA) nanoparticles. The curcumin encapsulated PLGA nanoparticles were prepared by emulsification solvent evaporation/diffusion, and large quantities were made by varying the homogenisation time (5, 15 and 30 min). The particle size decreased as the homogenisation duration increased from 5 to 30 min, and the particles were spherical as confirmed by atomic force microscopy. For the large-scale preparations, the mean particles size was found to be 288.7 ± 3.4 (polydispersity index 0.15 ± 0.01) with curcumin entrapment 52.5 ± 4.3 %, which were comparable to the lab-scale preparations. The curcumin encapsulated nanoparticles were freeze-dried using sucrose (5 %, w/v) as a cryoprotectant. The freeze-dried nanoparticles were subjected to 6-month stability study as per the International Conference on Harmonisation guideline at room temperature and refrigerated storage conditions. Intermediate sampling was done (monthly), and the nanoparticles were thoroughly characterised for particle size, entrapment efficiency, surface morphology and crystallinity, which were compared to fresh preparations. The curcumin encapsulated PLGA nanoparticles were found to be stable at refrigerated as well as room temperature storage test conditions indicated by their particle characteristics. X-ray diffraction results confirm amorphous nature of curcumin on nano-encapsulation that stays intact after freeze drying and 6-month stability testing. Together these data offer possibility of producing large quantities of polymer nanoparticles that are suitable for room as well as refrigerated storage conditions opening up possibilities to conduct repeated dosings in a chronic setting or regulatory toxicology studies of such nanomedicines.

The efficacy of aerosol treatment with non-ionic surfactant vesicles containing amphotericin B in rodent models of leishmaniasis and pulmonary aspergillosis infection.

Amphotericin B (AMB) is used to treat both fungal and leishmanial infections, which are of major significance to human health. Clinical use of free AMB is limited by its nephrotoxicity, whereas liposomal AMB is costly and requires parenteral administration, thus development of novel formulations with enhanced efficacy, minimal toxicity and that can be applied via non-invasive routes is required. In this study we analysed the potential of non-ionic surfactant vesicles (NIV) given by nebulisation to deliver AMB to the lungs, liver and skin. Treatment with AMB-NIV resulted in significantly higher drug levels in the lungs and skin (p<0.05) compared to similar treatment with AMB solution but significantly lower plasma levels (p<0.05). Treatment with AMB-NIV resulted in a significant reduction in fungal lung burdens in a rat model of invasive pulmonary aspergillosis (p<0.05) compared to treatment with the carrier alone. Treatment with AMB-NIV but not AMB solution significantly suppressed Leishmania donovani liver parasite burdens (p<0.05) but could not inhibit the growth of cutaneous Leishmania major lesions. The results of this study indicate that aerosolised NIV enhanced pulmonary and hepatic delivery whilst minimising systemic exposure and toxicity.

Tissue localization of nanoparticles is altered due to hypoxia resulting in poor efficacy of curcumin nanoparticles in pulmonary hypertension.

The present study is an attempt to leverage therapeutic benefits of curcumin in pulmonary hypertension by encapsulating it in biodegradable poly(lactide-co-glycolic) acid nanoparticles. Pulmonary hypertension is induced in experimental animals by subjecting them to chronic hypoxic conditions. The ability of curcumin encapsulated nanoparticles to manage pulmonary hypertension is measured by right ventricular hypertrophy, haematocrit, vascular remodelling and target tissue levels of curcumin. Further, single oral dose tissue distribution of the nanoparticulate curcumin was also assessed under normoxic and hypoxic conditions. Orally administered nanoparticulate curcumin failed to offer any protection against hypoxia induced pulmonary hypertension as indicated by insignificant changes in right ventricular hypertrophy and vascular remodelling that are similar to untreated groups. A significant difference in the target tissue levels was observed between normoxic vs. hypoxic rats. The study suggests that hypoxia has a major role in the particle localization in lungs probably due to the altered blood flow, increased barrier properties of the lung vasculature and decreased endocytosis. The target tissue levels of curcumin under hypoxia are much lower to that achieved in normoxic rats probably due to difference in particle dynamics, resulting in the failure of treatment.

Nanoparticulate delivery can improve peroral bioavailability of cyclosporine and match Neoral Cmax sparing the kidney from damage.

Cyclosporine (CsA) as an immunosuppressant has demonstrated immense potential in the field of organ transplantation and autoimmune disorders, despite the nephrotoxicity. The present investigation is an attempt to develop biodegradable nanoparticles entrapping CsA that can match Cmax of Neora, the most potent formulation available to date. The Cmax and AUC0-72 of CsA administered as nanoparticles demonstrated an increase with increase in the dose administered, however the relative bioavailability decreased. The Cmax and AUC0-72 increased from 682 to 1073 ng/ml and 34854 to 55322 ng . h/ml, respectively whereas, the relative bioavailability decreased from 120 to 64%. On the other hand, the increase in initial CsA loading (10-30% w/w of polymer) recorded a proportional increase in Cmax and AUC0-72, from 494 to 1101 ng/ml and 17774 to 51763 ng . h/ml, respectively. The relative bioavailability also increased from 31 to 89%. The CsA nanoparticles at 30% CsA loading w/w of polymer and at 30 mg/Kg single dose demonstrated comparable Cmax of Neoral at 15 mg/Kg, which upon chronic administration over a period of 30 days daily dosing showed low nephrotoxicity to that exhibited by Neoral. The reduced toxicity of nanoparticulate CsA indicated by lower blood urea nitrogen, plasma creatinine and glomerular damage, was due to delay in Tmax as result of slow release of the CsA from the nanoparticles. The investigation unlocks the potential of polymeric nanoparticles in oral delivery of CsA.

Microemulsions for oral delivery of insulin: design, development and evaluation in streptozotocin induced diabetic rats.

Insulin loaded microemulsions were developed adopting a low shear reverse micellar approach using didoceyldimethylammonium bromide (DMAB) as the surfactant, propylene glycol (PG) as the co-surfactant, triacetin (TA) as the oil phase and insulin solution as the aqueous phase. A ternary phase diagram was constructed based on multiple cloud point titration to highlight the reverse micellar region. The droplet sizes of the microemulsions were 161.7±24.7nm with PDI of 0.447±0.076 and insulin entrapment of ∼85%. Transmission electron microscopy (TEM) revealed the spherical nature and size homogeneity of the microemulsion droplets. The conformational stability of the entrapped insulin within microemulsions was confirmed by fluorescence spectroscopy and circular dichroism. The microemulsions displayed a 10-fold enhancement in bioavailability compared with plain insulin solution administered per oral in healthy rats. The short-term in vivo efficacy in STZ induced diabetic rats provided the proof of concept by a modest glucose reduction at a dose of 20IU/kg. Together this preliminary data indicate the promise of microemulsions for oral delivery of insulin.

Evaluating the potential of polymer nanoparticles for oral delivery of paclitaxel in drug-resistant cancer.

The present study was designed to explore the ability of polymeric nanoparticles to restore drug sensitivity to P-glycoprotein over-expressing cancer cells. A multidrug-resistant cell line 2780 AD and its sensitive parent cell line A2780 were studied in cell culture and as a xenografted tumour model. Paclitaxel was incorporated in poly(lactide-co-glycolide) nanoparticles of average diameter 125 nm stabilised by a positively charged surfactant. The nanoparticulate formulation was shown to be about sevenfold more potent than free paclitaxel against cell line A2780 and the poly(lactic-co-glycolic acid) (PLGA) nanoparticles alone were nontoxic to the cells at the concentrations required to deliver the drug. Whilst the oral formulation of paclitaxel was not as potent as the free drug in the A2780 xenografts, it showed significant activity against 2780 AD tumours, which are resistant to the maximum tolerated intravenous dose of paclitaxel. The efficacy of orally delivered paclitaxel in this drug-resistant model supports the concept of exploring nanoparticles for improved drug delivery.

Biodegradable nanoparticles improve oral bioavailability of amphotericin B and show reduced nephrotoxicity compared to intravenous Fungizone.

PURPOSE: Amphotericin B (AMB), an effective antifungal and antileishmanial agent associated with low oral bioavailability (0.3%) and severe nephrotoxicity, was entrapped into poly(lactide-co-glycolide) (PLGA) nanoparticles to improve the oral bioavailability and to minimize the adverse effects associated with it. MATERIALS AND METHODS: The AMB-nanoparticles (AMB-NP) were prepared by nanoprecipitation method employing Vitamin E-TPGS as a stabilizer. In vitro release was carried out using membrane dialysis method. The in vitro hemolytic activity of AMB-NP was evaluated by incubation with red blood cells (RBCs). The acute nephrotoxicity profile and oral bioavailability of AMB-NP were evaluated in rats. RESULTS: The prepared AMB-NP formulation contained monodispersed particles in the size range of 165.6 +/- 2.9 nm with 34.5 +/- 2.1% entrapment at 10% w/w initial drug loading. AMB-NP formulation showed biphasic drug release, an initial rapid release followed by a sustained release. The AMB-NP formulation exerted lower hemolysis and nephrotoxicity as compared to Fungizone. The relative oral bioavailability of the AMB-NP was found to be approximately 800% as compared to Fungizone. CONCLUSION: Together, these results offer a possibility of treating systemic fungal infection and leishmaniasis with oral AMB-NP, which could revolutionize the infectious disease treatment modalities.

High-performance liquid chromatographic analysis of amphotericin B in rat plasma using alpha-naphthol as an internal standard.

A simple, sensitive and accurate reverse phase high-performance liquid chromatographic (RP-HPLC) method with photo-diode array detector (PDA) was developed and validated for the determination of amphotericin B (AMB) in the rat plasma using a new internal standard (IS) alpha-naphthol. The plasma samples were subjected to protein precipitation with methanol prior to a HPLC analysis. Chromatographic separations were achieved on a Nucleosil 100-5C18 (150 mm x 4.6 mm) column. The mobile phase consisted of acetonitrile and sodium acetate buffer (pH 4; 10mM) in a gradient mode. Detection was carried out at a wavelength of 407 and 294 nm for AMB and IS, respectively. The retention times of AMB and IS were about 6.8 and 7.8 min, respectively. The calibration curve was linear in the range of 10-2000 ngmL(-1) for AMB (r(2)>0.998). No significant matrix effect was observed on quantification of AMB or IS. At three quality control concentrations of 20, 500, and 2000 ngmL(-1), the intra-day and inter-day relative standard deviation ranged from 1.13% to 4.91%. The limit of detection (LOD) was 5 ngmL(-1) and the limit of quantification (LOQ) was 10 ngmL(-1) for AMB in rat plasma. This method is simple, sensitive, rapid and does not require any extensive sample purification before injecting into HPLC.

Design of biodegradable nanoparticles for oral delivery of doxorubicin: in vivo pharmacokinetics and toxicity studies in rats.

PURPOSE: Doxorubicin, a potent anticancer drug associated with cardiotoxicity and low oral bioavailability, was loaded into nanoparticles with a view to improve its performance. METHODS: Doxorubicin loaded PLGA nanoparticles were prepared by a double emulsion method. The pH dependent stability of nanoparticles in simulated fluids was evaluated. DSC and XRD studies were carried out in order to ascertain the nature of doxorubicin in formulations in conjunction with accelerated stability studies. The in vitro release was investigated in phosphate buffer. The pharmacokinetic and toxicity studies were conducted in rats. RESULTS: Nanoparticles had an average size of 185 nm, with 49% entrapment at 10% w/w of polymer. The particles displayed good pH dependent stability in the pH range 1.1-7.4. DSC and XRD studies revealed the amorphous nature of doxorubicin in nanoparticles and the accelerated stability studies revealed the integrity of formulations. Initial biphasic release (20%) followed by a sustained release (80%) for 24 days was observed under in vitro conditions. The doxorubicin loaded nanoparticles demonstrated superior performance in vivo as evident by enhanced bioavailability and lower toxicity. CONCLUSIONS: Together, the data indicates the potential of doxorubicin loaded nanoparticles for oral chemotherapy. Further, these formulations could be explored for new indications like leishmaniasis.

Pharmacodynamic evaluation of oral estradiol nanoparticles in estrogen deficient (ovariectomized) high-fat diet induced hyperlipidemic rat model.

PURPOSE: It is believed that estrogen deficiency contributes importantly to the pathogenesis of menopausal metabolic syndrome and symptoms can be ameliorated with estradiol therapy. The present study reports efficacy of 17-beta estradiol encapsulated nanoparticles in treating the postmenopausal dyslipidemic condition. MATERIALS AND METHODS: Estradiol encapsulated poly(lactide-co-glycolide) (PLGA) nanoparticles were prepared by emulsion-diffusion-evaporation method and evaluated in estrogen deficient (ovariectomized) high fat diet induced hyperlipidemic rat model. RESULTS: The results obtained showed that estradiol nanoparticles were equally/more effective in treatment of estrogen deficient hyperlipidemic conditions at three times reduced dose and frequency in comparison to that of drug suspension administered orally. CONCLUSION: Together, these results demonstrate the ability of nanoparticles in improving oral bioavailability/efficacy of estradiol.

Estradiol loaded PLGA nanoparticles for oral administration: effect of polymer molecular weight and copolymer composition on release behavior in vitro and in vivo.

The present investigation was aimed at optimization of estradiol loaded PLGA nanoparticulate formulations resulting in improved oral bioavailability and sustained release of estradiol by varying the molecular weight and copolymer composition of PLGA. Nanoparticles were prepared following emulsion-diffusion-evaporation method employing didodecyldimethyl ammonium bromide (DMAB) as stabilizer. The effect of polymer molecular weight and copolymer composition on particle properties and release behavior (in vitro and in vivo) has been reported. Drug release in vitro decreased with increase in molecular weight and lactide content of PLGA. Zero order release was obtained with low molecular weight (14,500 and 45,000 Da) PLGA, while high molecular weight (85,000 and 213,000 Da) and different copolymer compositions followed square root of time (Higuchi's pattern) dependent release. The bioavailability of estradiol from nanoparticles was assessed in male Sprague Dawley (SD) rats at a dose of 1 mg estradiol/rat. The in vivo performance of the nanoparticles was found to be dependent on the particle size, polymer molecular weight and copolymer composition. The C(max) of drug in the plasma was dependent on the polymer molecular weight and composition while particle size was found to influence the duration of release, suggesting smaller is better. The histopathological examination revealed absence of any inflammatory response with the formulations prepared of low/high molecular weight or high lactide content polymers for the studied period. Together, these results indicate that nanoparticulate formulations are ideal carriers for oral administration of estradiol having great potential to address the dose related issues of estradiol.

Design and evaluation of biodegradable, biosensitive in situ gelling system for pulsatile delivery of insulin.

Biodegradable glucose-sensitive in situ gelling system based on chitosan for pulsatile delivery of insulin was developed. The sols/gels were thoroughly characterized for swelling properties, rheology, texture analysis and water content. The developed glucose-sensitive gels responded to varied glucose concentrations in vitro indicating their ability to function as environment-sensitive systems. Insulin load onto the gels was optimized and was found to affect the rheological behavior of these gels, the final preparation used for in vitro contained 1IU/200mul of the sol. These gels released the entrapped insulin in a pulsatile manner in response to the glucose concentration in vitro. Furthermore, the formulations when evaluated for their in vivo efficacy in streptozotocin-induced diabetic rats at a dose of 3IU/kg, demonstrated their ability to release insulin in response to glucose concentration and were preferred much better against subcutaneously given plain insulin formulation used as the control. Together, these preliminary results indicate that biosensitive chitosan in situ gelling systems have substantial potential as pulsatile delivery systems for insulin.