Richard A. Graves

Poly(D,L-lactide-co-glycolide) encapsulated poly(vinyl alcohol) hydrogel as a drug delivery system.

AbstractPurpose. The efficiency of encapsulation of water-soluble drugs in biodegradable polymer is often low and occasionally these microcapsules are associated with high burst effect. The primary objective of this study is to develop a novel microencapsulation technique with high efficiency of encapsulation and low burst effect. Method. Pentamidine was used as a model drug in this study. Pentamidine/polyvinyl alcohol (PVA) hydrogel was prepared by freeze-thaw technique. Pentamidine loaded hydrogel was later microencapsulated in poly(lactide-co-glycolide) (PLGA) using solvent evaporation technique. The microcapsules were evaluated for the efficiency of encapsulation, particle size, surface morphology, thermal characteristic, and drug release. Results. Scanning Electron Microscope (SEM) studies revealed that the microcapsules were porous. The microcapsules were uniform in size and shape with the median size of the microcapsules ranging between 27 and 94 μm. The samples containing 10% PLGA showed nearly three times increase in drug loading (18-53%) by increasing the hydrogel content from 0-6%. The overall drug release from the microencapsulated hydrogel, containing 3% and 6% PVA, respectively, was significantly lower than the control batches. Conclusions. The use of a crosslinked hydrogel such as PVA can significantly increase the drug loading of highly water-soluble drugs. In addition, incorporation of the PVA hydrogel significantly reduced the burst effect and overall dissolution of pentamidine.

Porous biodegradable microparticles for delivery of pentamidine.

The primary objective of this study was to develop a method for the preparation of porous biodegradable controlled release formulation of poly(lactide/glycolide) (PLGA). The model drug used for this study was pentamidine. Scanning electron microscopy pictures showed that these microparticles are highly porous and spherical in shape. A comparison of particle size reveals a similar median particle size (54-68 microm) in all six batches. The particles are all smaller than 90 microm. Differential scanning calorimetry thermograms revealed that pentamidine was mostly present in the crystalline form in the microparticles and did not dissolve in PLGA. The efficiency of encapsulation of pentamidine was higher than 58% in all six batches. The amount of drug released from these microparticles was at least 12% within the first 60 min. At least 50% of the total drug was released within the first 4 h. Drug release from these microparticles continued for up to 12 h. This faster drug dissolution was due to the highly porous surface. This highly porous surface will allow large molecules to release at a much faster rate than the regular microcapsules/microspheres.

Oral delivery of spray dried PLGA/amifostine nanoparticles

Amifostine (Ethyol, WR‐2721) is a cytoprotective drug approved by the US Food & Drug Administration for intravenous administration in cancer patients receiving radiation therapy and certain forms of chemotherapy. The primary objective of this project was to develop orally active amifostine nanoparticles using spray drying technique. Two different nanoparticle formulations (Amifostine‐PLGA (0.4:1.0 and 1.0:1.0)) were prepared using a Buchi B191 Mini Spray Dryer. A water‐in‐oil emulsion of amifostine and PLGA (RG 502) was spray dried using an airflow of 600 Lh−1 and input temperature of 55°C. A tissue distribution study in mice was conducted following oral administration of the formulation containing drug‐polymer (0.4:1.0). The efficiency of encapsulation was 90% and 100%, respectively, for the two formulations while the median particle sizes were 257 and 240 nm, with 90% confidence between 182 and 417 nm. Since amifostine is metabolized to its active form, WR‐1065, by intracellular alkaline phosphatase, the tissue levels of WR‐1065 were measured, instead of WR‐2721. WR‐1065 was detected in significant amounts in all tissues, including bone marrow, jejunum and the kidneys, and there was some degree of selectivity in its distribution in various tissues. This work demonstrates the feasibility of developing an orally effective formulation of amifostine that can be used clinically.

Cellular delivery of PEGylated PLGA nanoparticles

Objectives  The objective of this study was to investigate the efficiency of uptake of PEGylated polylactide‐co‐gycolide (PLGA) nanoparticles by breast cancer cells.

Radioprotection in mice following oral delivery of amifostine nanoparticles

Purpose: Amifostine (Ethyol®) is an approved cytoprotective agent prescribed to reduce certain side-effects in the chemotherapy of ovarian or non-small cell lung cancer, or in radiation treatment of head-and-neck cancer. The usefulness of this drug is further hampered, because it is not effective when given orally. The objective of this part of the project was to evaluate the radioprotective efficacy of orally active amifostine nanoparticles. Materials and methods: Radioprotective efficacy was evaluated by measuring the ability of the amifostine nanoparticles (equivalent to 500 mg/Kg) to inhibit whole-body gamma irradiation -induced injury in mice. All mice received acute whole-body gamma irradiation from a Cesium-137 source and the radioprotective efficacy of the formulation was determined by measuring 30-day survival at 9 Gy, bone marrow hemopoeitic progenitor cell survival at 9 Gy and 8 Gy, and intestinal crypt cell survival at 11 Gy. Results: Thirty-day survival, hemopoietic progenitor cell survival, as well as the jejunal crypt cell survival were all significantly enhanced when the mice were treated orally with the amifostine nanoparticles 1 h prior to irradiation. Conclusions: These results clearly and unequivocally demonstrate that the amifostine nanoparticles developed in our laboratory provides significant protection from acute whole-body gamma irradiation injury in mice.

Spray-Dried Chitosan as a Direct Compression Tableting Excipient

The objective of this study was to prepare and evaluate a novel spray-dried tableting excipient using a mixture of chitosan and lactose. Three different grades of chitosan (low-, medium-, and high-molecular-weight) were used for this study. Propranolol hydrochloride was used as a model drug. A specific amount of chitosan (1, 1.9, and 2.5 g, respectively) was dissolved in 50 mL of an aqueous solution of citric acid (1%) and later mixed with 50 mL of an aqueous solution containing lactose (20, 19.1, and 18.5 g, respectively) and propanolol (2.2 g). The resultant solution was sprayed through a laboratory spray drier at 1.4 mL/min. The granules were evaluated for bulk density, tap density, Carr index, particle size distribution, surface morphology, thermal properties, and tableting properties. Bulk density of the granules decreased from 0.16 to 0.13 g/mL when the granules were prepared using medium- or high-molecular-weight chitosan compared with the low-molecular-weight chitosan. The relative proportion of chitosan also showed a significant effect on the bulk density. The granules prepared with 1 g of low-molecular-weight chitosan showed the minimum Carr index (11.1%) indicating the best flow properties among all five formulations. All three granules prepared with 1 g chitosan, irrespective of their molecular weight, showed excellent flow properties. Floating tablets prepared by direct compression of these granules with sodium bicarbonate showed 50% drug release between 30 and 35 min. In conclusion, the spray-dried granules prepared with chitosan and lactose showed excellent flow properties and were suitable for tableting.

Preparation and in vitro characterization of amifostine biodegradable microcapsules

The purpose of this project was to develop sustained release microcapsules of amifostine. The microcapsules were prepared using solvent evaporation technique. The effect of several formulation variables on the characteristics of the microcapsules was studied. The formulation variables studied were drug loading, polymer (polylactide-co-glycolide) (PLGA) concentration, and the amount of gelatin in the initial aqueous phase. The drug loading was studied at three different levels (5, 10, and 25 mg); the PLGA concentration was studied at two levels (500 and 1000 mg); and the amount of gelatin used ranged from 2 to 14 mg. In general, the microcapsules were less than 155 microm in diameter with median size between 50 and 80 microm. While the use of higher amounts of PLGA significantly increased the median size of the microcapsules, using higher amounts of amifostine had no significant effect, irrespective of the amount of PLGA. The use of gelatin, within the range 2-14 mg, did not show any significant effect on the particle size distribution. Scanning electron microscopy (SEM) of the microcapsules revealed that all nine formulations yielded spherical particles. The use of 500 mg PLGA with 10 or 25 mg amifostine yielded microcapsules with porous surfaces. The surface pores, however, were not present in microcapsules prepared using 1000 mg PLGA. The efficiency of encapsulation decreased significantly from 63 to 24% when the amount of amifostine increased from 5 to 25 mg in the formulations using 500 mg PLGA. Similarly, the efficiency of encapsulation decreased from 87 to 23% when the amount of PLGA was doubled to 1000 mg. An increase in the amount of amifostine in the formulation using 500 mg PLGA also resulted in a significant increase in initial drug release (from 20 to 62%) within the first hour. These results were consistent with the porous morphology of these microcapsules. In general, all batches of microcapsules showed 24-96 h sustained drug release.

A Novel Peptide Derived from Human Apolipoprotein E Is an Inhibitor of Tumor Growth and Ocular Angiogenesis

Angiogenesis is a hallmark of tumor development and metastasis and now a validated target for cancer treatment. We previously reported that a novel dimer peptide (apoEdp) derived from the receptor binding region of human apolipoprotein E (apoE) inhibits virus-induced angiogenesis. However, its role in tumor anti-angiogenesis is unknown. This study demonstrates that apoEdp has anti-angiogenic property in vivo through reduction of tumor growth in a mouse model and ocular angiogenesis in a rabbit eye model. Our in vitro studies show that apoEdp inhibits human umbilical vein endothelial cell proliferation, migration, invasion and capillary tube formation. We document that apoEdp inhibits vascular endothelial growth factor-induced Flk-1 activation as well as downstream signaling pathways that involve c-Src, Akt, eNOS, FAK, and ERK1/2. These in vitro data suggest potential sites of the apoE dipeptide inhibition that could occur in vivo. This is the first evidence that a synthetic dimer peptide mimicking human apoE has anti-angiogenesis functions and could be an anti-tumor drug candidate.

In Vitro Dissolution Method for Evaluation of Buprenorphine In Situ Gel Formulation: A Technical Note

ConclusionThe in situ gel formulation of buprenorphine showed sustained drug release for a prolonged period of time. The drug release from RG 502 followed a linear pattern throughout the dissolution without any significant burst release. The amount of buprenorphine released during the first 30 minutes, irrespective of the type of Resomer or dissolution medium, was less than 3%. Drug release continued over 55 days in phosphate buffer and 35 days in Tween 80. The in vitro dissolution method developed during this study was capable of identifying formulation differences and thus will be useful for routine drug delivery research, particularly in situ gel formulation development research. In situ gel formulations are routinely compared using animal models,21, so development of such an in vitro method will expedite formulation evaluation.

Development and in vitro evaluation of a nanoemulsion for transcutaneous delivery

Abstract Objective: The purpose of this study is to develop a nanoemulsion formulation for its use as a transcutaneous vaccine delivery system. Materials and methods: With bovine albumin-fluorescein isothiocyanate conjugate (FITC-BSA) as a vaccine model, formulations were selected with the construction of pseudo-ternary phase diagrams and a short-term stability study. The size of the emulsion droplets was furthered optimized with high-pressure homogenization. The optimized formulation was evaluated for its skin permeation efficiency. In vitro skin permeation studies were conducted with shaved BALB/c mice skin samples with a Franz diffusion cell system. Different drug concentrations were compared, and the effect of the nanoemulsion excipients on the permeation of the FITC-BSA was also studied. Results: The optimum homogenization regime was determined to be five passes at 20 000 psi, with no evidence of protein degradation during processing. With these conditions, the particle diameter was 85.2 nm ± 15.5 nm with a polydispersity index of 0.186 ± 0.026 and viscosity of 14.6 cP ± 1.2 cP. The optimized formulation proved stable for 1 year at 4 °C. In vitro skin diffusion studies show that the optimized formulation improves the permeation of FITC-BSA through skin with an enhancement ratio of 4.2 compared to a neat control solution. Finally, a comparison of the skin permeation of the nanoemulsion versus only the surfactant excipients resulted in a steady state flux of 23.44 μg/cm2/h for the nanoemulsion as opposed to 6.10 μg/cm2/h for the emulsifiers. Conclusion: A novel nanoemulsion with optimized physical characteristics and superior skin permeation compared to control solution was manufactured. The formulation proposed in this study has the flexibility for the incorporation of a variety of active ingredients and warrants further development as a transcutaneous vaccine delivery vehicle.