Maria Fernanda Bahia

Nanotechnology-based systems for the treatment and prevention of HIV/AIDS☆

The HIV/AIDS pandemic is an increasing global burden with devastating health-related and socioeconomic effects. The widespread use of antiretroviral therapy has dramatically improved life quality and expectancy of infected individuals, but limitations of currently available drug regimens and dosage forms, alongside with the extraordinary adapting capacity of the virus, have impaired further success. Alongside, circumventing the escalating number of new infections can only be attained with effective and practical preventative strategies. Recent advances in the field of drug delivery are providing evidence that engineered nanosystems may contribute importantly for the enhancement of current antiretroviral therapy. Additionally, groundwork is also being carried out in the field nanotechnology-based systems for developing preventative solutions for HIV transmission. This manuscript reviews recent advances in the field of nanotechnology-based systems for the treatment and prevention of HIV/AIDS. Particular attention is given to antiretroviral drug targeting to HIV reservoirs and the usefulness of nanosystems for developing topical microbicides and vaccines.

Mucoadhesive nanomedicines: characterization and modulation of mucoadhesion at the nanoscale.

Introduction: The benefits of mucoadhesive systems are related to the increased in situ residence and intimate contact of the delivery vehicle with the mucosa. The recent emergence of nanomedicine and the properties of nanoparticulate systems have created new challenges in understanding the nature and mechanisms of nanoscale mucoadhesion and in the development of methodologies for measuring its mucoadhesive potential. Even when usually regarded as an advantageous property, mucoadhesion can be an inconvenience for nanosystems, and strategies have been developed for minimizing interactions with the mucosal tissues/fluids. Areas covered: This article summarizes the basic concepts of mucoadhesion at the nanoscale, different techniques used for measuring the mucoadhesive potential of nanosystems and strategies for increasing/decreasing mucoadhesive interactions. Expert opinion: The mucoadhesion behavior of materials in bulk and at the nanoscale can significantly differ. Advances in the methodology used for studying the mucoadhesion phenomenon have contributed to its better understanding and, more importantly, the development of strategies to increase/decrease mucoadhesion. However, development of new methodologies for studying mucoadhesion at the nanoscale and the refinement of existing methodologies are still required. Also, a substantial amount of information is still lacking, particularly related to formulation issues, on how to translate lessons learnt at the bench top to the bed side.

Performance of an in vitro mucoadhesion testing method for vaginal semisolids: influence of different testing conditions and instrumental parameters.

The purpose of this work was to develop an in vitro mucoadhesion testing method for vaginal semisolid formulations. The proposed method was based on the measurement of the force (detachment force, Fdt) and the work (work of adhesion, Wad) needed to detach a sample of cow vaginal mucosa from a semisolid formulation, using a commercially available texture analyzer. Several testing conditions and instrumental parameters were tested in order to evaluate the mucoadhesive potential of a model vaginal semisolid formulation (1% Carbopol 974P gel). Also, mucoadhesive potential of several commercially available vaginal semisolid products was evaluated. Obtained results showed that the method is reproducible even when the same cow mucosa sample is used up to six times. The similarity of the fluid used to bathe the vaginal mucosa to the one naturally occurring in the vagina influenced considerably the performance of the test, advising that simulation of vaginal fluid properties is important when measuring mucoadhesive properties. Also, temperature of experiment was an important fact to be considered, as results showed slight but significant differences between body (37 degrees C) and room (20 degrees C) temperature. Fdt and Wad increased with increasing instrumental parameters while a plateau region was observable at higher values of probe speed, probe force, and mucosa/sample contact time. Comparison between results for Fdt and Wad demonstrated that although both parameters are generally in agreement, Wad seems to be more reliable and reproducible when evaluating mucoadhesion. Evaluation of commercially available formulations confirmed that experimental conditions are important features that can influence significantly the determination of mucoadhesive potential, being the proposed method an interesting and useful tool in the in vitro evaluation of vaginal semisolids.

Polymeric Nanoparticles Affect the Intracellular Delivery, Antiretroviral Activity and Cytotoxicity of the Microbicide Drug Candidate Dapivirine

ABSTRACTPurposeTo assess the intracellular delivery, antiretroviral activity and cytotoxicity of poly(ε-caprolactone) (PCL) nanoparticles containing the antiretroviral drug dapivirine.MethodsDapivirine-loaded nanoparticles with different surface properties were produced using three surface modifiers: poloxamer 338 NF (PEO), sodium lauryl sulfate (SLS) and cetyl trimethylammonium bromide (CTAB). The ability of nanoparticles to promote intracellular drug delivery was assessed in different cell types relevant for vaginal HIV transmission/microbicide development. Also, antiretroviral activity of nanoparticles was determined in different cell models, as well as their cytotoxicity.ResultsDapivirine-loaded nanoparticles were readily taken up by different cells, with particular kinetics depending on the cell type and nanoparticles, resulting in enhanced intracellular drug delivery in phagocytic cells. Different nanoparticles showed similar or improved antiviral activity compared to free drug. There was a correlation between increased antiviral activity and increased intracellular drug delivery, particularly when cell models were submitted to a single initial short-course treatment. PEO-PCL and SLS-PCL nanoparticles consistently showed higher selectivity index values than free drug, contrasting with high cytotoxicity of CTAB-PCL.ConclusionsThese results provide evidence on the potential of PCL nanoparticles to affect in vitro toxicity and activity of dapivirine, depending on surface engineering. Thus, this formulation approach may be a promising strategy for the development of next generation microbicides.

Rheological properties of vaginal hydrophilic polymer gels.

The objective of this work was to investigate the main theological features of vaginal hydrophilic polymer gels and to elucidate about the relationship between these characteristics and gels composition, and their general influence in therapeutic/usage purpose. Flow and dynamic oscillatory properties of four commercially available (Conceptrol, Gynol II, RepHresh, and Replens) and two investigational vaginal gels were determined by cone-and-plate rheometry, at body temperature. Several parameters (apparent viscosity, complex viscosity, storage modulus, loss modulus, critical oscillatory stress, tan delta, thixotropy and yield stress) were measured and/or calculated. Gels presented non-Newtonian, pseudoplastic, thixotropic behavior, with yield stress. Overall viscosities varied between 13500 Pa.s and approximately 80 Pa.s within a biologically relevant shear rate interval (0.01-100 s(-1)). Yield stress values were variable between different determination methods but coherent in terms of ranking. Also, tested gels showed viscoelastic properties, being characterized by predominant elastic solid-like behavior. Rheological behavior of vaginal gels strongly depended on the type of gelling agent used, which potentially influences their spreading and retention properties when administered in the vaginal canal. Small variations in gels composition can result in substantial changes in their features, namely viscosity, yield stress and thixotropy. Rheological properties of tested gels appeared to be correlated with their therapeutic/usage purpose.

Local treatment of vulvovaginal candidosis : general and practical considerations.

Vulvovaginal candidosis is a common worldwide female medical problem, occurring mostly in women of childbearing age. Currently available options for the treatment of this condition include local and oral (systemic) therapy. Both alternatives have been considered equally effective in the treatment of uncomplicated vulvovaginal candidosis, although oral regimens are often preferred by physicians and women. However, local treatment presents several advantageous and unique features that may favour this therapeutic approach. The availability of numerous antifungal drugs and products for topical administration makes the selection quite challenging as this task is mostly based on personal experience or anecdotal data. Also, recent advances have been made in topical antifungal formulations and there is an increasing availability of over-the-counter products. Therefore, a review of both general and practical considerations related to the local treatment of vulvovaginal candidosis is timely.In summary, azoles and short-term regimens are usually recommended for the local treatment of vulvovaginal candidosis, with nystatin and boric acid considered as second-line alternatives. Unconventional approaches may also be regarded as suitable in patients refractory to usual treatments. In addition to the susceptibility of implicated Candida spp. to the antifungal agents, this choice should take into consideration other important issues such as particular situations (e.g. pregnancy, menopause, drug hypersensitivity), women’s preferences, and the availability, particularities and cost of antifungal formulations.

In Vitro and Ex Vivo Evaluation of Polymeric Nanoparticles for Vaginal and Rectal Delivery of the Anti-HIV Drug Dapivirine

Prevention strategies such as the development of microbicides are thought to be valuable in the fight against HIV/AIDS. Despite recent achievements, there is still a long road ahead in the field, particularly at the level of drug formulation. Drug nanocarriers based on polymers may be useful in enhancing local drug delivery while limiting systemic exposure. We prepared differently surface-engineered poly(ε-caprolactone) (PCL) nanoparticles (NPs) and tested their ability to modulate the permeability and retention of dapivirine in cell monolayers and pig vaginal and rectal mucosa. NPs coated with poly(ethylene oxide) (PEO) were shown able to reduce permeability across monolayers/tissues, while modification of nanosystems with cetyl trimethylammonium bromide (CTAB) enhanced transport. In the case of coating NPs with sodium lauryl sulfate (SLS), dapivirine permeability was unchanged. All NPs increased monolayer/tissue drug retention as compared to unformulated dapivirine. This fact was associated, at least partially, to the ability of NPs to be taken up by cells or penetrate mucosal tissue. Cell and tissue toxicity was also affected differently by NPs: PEO modification decreased the in vitro (but not ex vivo) toxicity of dapivirine, while higher toxicity was generally observed for NPs coated with SLS or CTAB. Overall, presented results support that PCL nanoparticles are capable of modulating drug permeability and retention in cell monolayers and mucosal tissues relevant for vaginal and rectal delivery of microbicides. In particular, PEO-modified dapivirine-loaded PCL NPs may be advantageous in increasing drug residence at epithelial cell lines/mucosal tissues, which may potentially increase the efficacy of microbicide drugs.

Interactions of Microbicide Nanoparticles with a Simulated Vaginal Fluid

The interaction with cervicovaginal mucus presents the potential to impact the performance of drug nanocarriers. These systems must migrate through this biological fluid in order to deliver their drug payload to the underlying mucosal surface. We studied the ability of dapivirine-loaded polycaprolactone (PCL)-based nanoparticles (NPs) to interact with a simulated vaginal fluid (SVF) incorporating mucin. Different surface modifiers were used to produce NPs with either negative (poloxamer 338 NF and sodium lauryl sulfate) or positive (cetyltrimethylammonium bromide) surface charge. Studies were performed using the mucin particle method, rheological measurements, and real-time multiple particle tracking. Results showed that SVF presented rheological properties similar to those of human cervicovaginal mucus. Analysis of NP transport indicated mild interactions with mucin and low adhesive potential. In general, negatively charged NPs underwent subdiffusive transport in SVF, i.e., hindered as compared to their diffusion in water, but faster than for positively charged NPs. These differences were increased when the pH of SVF was changed from 4.2 to 7.0. Diffusivity was 50- and 172-fold lower in SVF at pH 4.2 than in water for negatively charged and positively charged NPs, respectively. At pH 7.0, this decrease was around 20- and 385-fold, respectively. The estimated times required to cross a layer of SVF were equal to or lower than 1.7 h for negatively charged NPs, while for positively charged NPs these values were equal to or higher than 7 h. Overall, our results suggest that negatively charged PCL NPs may be suitable to be used as carriers in order to deliver dapivirine and potentially other antiretroviral drugs to the cervicovaginal mucosal lining. Also, they further reinforce the importance in characterizing the interactions of nanosystems with mucus fluids or surrogates when considering mucosal drug delivery.

Protective effect of Castanea sativa and Quercus robur leaf extracts against oxygen and nitrogen reactive species

Topical natural antioxidants are a useful strategy for the prevention of photoaging and oxidative stress mediated skin diseases. In view of this underlying principle, the screening of natural plant extracts with scavenging activity for pro-oxidant reactive species is a primary requirement for the development of new topical antioxidant formulations. In the present study, an ethanol:water (7:3) extract from Castanea sativa leaves and a ethanol:water (2:3) extract from Quercus robur leaves were evaluated for their putative in vitro scavenging effects on reactive oxygen species (ROS) namely superoxide radical (O(2)(-)), hydroxyl radical (HO()), peroxyl radical (ROO()), hydrogen peroxide (H(2)O(2)) and singlet oxygen ((1)O(2)) as well as on reactive nitrogen species (RNS) namely nitric oxide (()NO) and peroxynitrite (ONOO(-)). The extracts presented a high potency to scavenge the tested reactive species, all the IC(50)s being found at the microg/mL level. IC(50)s (mean+/-SE) for the ROS O(2)(-),HO(),H(2)O(2) and (1)O(2) were 13.6+/-1.8; 216+/-4; 410+/-8; 12.3+/-0.7 microug/mL, respectively, for C. sativa, and 11.0+/-0.5; 285+/-22; 251+/-32; 7.90+/-0.56 microg/mL, respectively, for Q. robur. The ORAC values obtained for ROO() were 1.24+/-0.13 for C. sativa and 1.09+/-0.06 for Q. robur. The IC(50)s (mean+/-SE) for ()NO and ONOO(-) were 3.10+/-0.14 and 1.49+/-0.10 microg/mL, respectively, for C. sativa and 3.13+/-0.11 and 0.95+/-0.02 microg/mL, respectively, for Q. robur. The content of total phenolics for C. sativa and Q. robur were 284+/-9 and 346+/-4 mg of gallic acid equivalents (GAE)/g of lyophilized extract respectively. The observed effects might be of relevance considering the putative interest of these extracts as topical antioxidants.

Injectability of a Bone Filler System Based on Hydroxyapatite Microspheres and a Vehicle With in situ Gel-Forming Ability

The aim of this study was to test the injectability of a bone filler system based on the combination of ceramic microspheres with a gel-like vehicle, for noninvasive surgery. Porous hydroxyapatite microspheres with a uniform size and an average diameter of 535 +/- 38 mum were prepared, and their compression strength and friability were tested. The sodium-alginate solution with a concentration of 7.25% (w/v) was used as the vehicle. To promote its in situ gelation, calcium carbonate and D-gluconic-delta-lactone were added to the solution. Microspheres were mixed with the vehicle at different percentages (20-40 wt %). Gelation times in the range of 8-20 min, were obtained, depending on the formulation. Mixtures of HAp microspheres with alginate solution at 7.25% originating a gel in 11 min present an adequate handling time to perform an injection. Their injectability was evaluated using an injection device commonly employed in vertebroplasty surgical procedures, coupled to a texturometer in compression mode. Using an extrusion rate of 0.1 mm/s, the force required to extrude any of the mixtures tested was lower than 100 N. For an extrusion rate of 1 mm/s mixtures with 40 wt % of microspheres were very difficult to inject. Mixtures with 35 wt % of microspheres presented the best compromise between injectability and compression strength of the gelled system. MicroCT analysis revealed a homogeneous distribution of the microspheres inside the vehicle, as well as full interconnection of the intra-microspheres spaces. The compression strength for the gelled systems ranged from 80 kPa (gel itself) to 600 kPa (composite with 40 wt % of microspheres).