Cyanine derivative as a suitable marker for thermosensitive in situ gelling delivery systems: In vitro and in vivo validation of a sustained buccal drug delivery

Abstract

Buccal administration route is a promising way for a large number of drugs exhibiting a low oral bioavailability. The present work describes the formulation and evaluation of a mucoadhesive and thermosensitive in situ gelling delivery system based on poloxamer 407, poloxamer 188 and xanthan gum for buccal drug delivery. First, the mucoadhesion properties were evaluated using a tensile test. The effect of xanthan gum on the mucoadhesion force was demonstrated. Then, to assess the buccal residence time which reflects the mucoadhesion properties, the validation of a fluorescence probe for in vivo optical imaging experiment was conducted. Methyl-Cyanine 5 derivative (Me-Cy5) was used to label the hydrogels, dissolution tests and permeation studies through buccal epithelium cells showed that Me-Cy5 release from hydrogels was mainly due to an erosion mechanism and presented a limited penetration across epithelium cells. These results suggest that, Me-Cy5 is a suitable marker for thermosensitive in situ gelling delivery systems as the probe mostly stays entrapped in the hydrogel and do not cross the epithelial barrier. Buccal residence performance of the hydrogel was evaluated for the first time by non-invasive optical imaging after administration to mice. This technique is an interesting alternative compared to visual observations and sacrifice involved experiments, which could also be exploited to various administration routes.

Introduction

For a large number of drugs, from small chemical compounds to large and complex structured proteins, the relatively low oral bioavailability has conducted researchers to find new administration routes (Sudhakar et al., 2006). Instinctively, buccal administration of drugs appeared as an alternative in terms of patient compliance and ease of administration. Furthermore, buccal drug delivery is considered as an interesting route of administration for the populations with swallowing disorders, such as pediatric patients and elderly people (Lam et al., 2014). Buccal mucosa provides many advantages for local or systemic therapy (Hearnden et al., 2012). Indeed, buccal administration of drugs allows avoiding the first pass hepatic metabolism as well as the proteolytic activity of the gastrointestinal tract (Bhattarai et al., 2010) increasing drastically the bioavailability (Junginger et al., 1999, Patel et al., 2011). Therefore, buccal administration route has raised a large interest during the last two decades. One of the major issues in buccal drug delivery is to obtain an optimal localized concentration of the active pharmaceutical ingredient in the buccal cavity over a sufficiently long period of time. Unfortunately, this is difficult because of the salivation and tongue movements resulting in a rapid evacuation (Rathbone et al., 1994) and therefore, in the absorption of a small fraction of drugs by the buccal mucosa (Madhav et al., 2009, Rossi et al., 2005). Actually, buccal administration of drugs by conventional forms, often leads to low bioavailability.

To overcome the drawbacks of existing buccal forms, a wide variety of solid and semi-solid delivery systems that adhere to the buccal mucosa and remain in place for a considerable time, such as bioadhesive patches and films (Diaz del Consuelo et al., 2007, Dixit and Puthli, 2009, Morales and McConville, 2011, Nair et al., 2013), adhesive tablets (Choi et al., 2000, Choi and Kim, 2000, Ìkinci et al., 2004), bioadhesive wafers and matrices (Ayensu et al., 2012, Cafaggi et al., 2005, Giannola et al., 2007, Kianfar et al., 2013), or hydrogels (Nagai and Machida, 1993) have been developed. These pharmaceutical forms enable the sustained release of the drug and improve its bioavailability.

As another promising way to achieve efficient buccal drug delivery, a few investigators have considered the use of in situ gelling delivery systems as potential buccal dosage forms (Ajazuddin et al., 2012, Das et al., 2010). As a result of their physicochemical properties, in situ gelling systems allow a liquid administration into the buccal cavity and exhibit a sol to gel transition once in the buccal environment. Most of the time, this phase transition is observed for stimuli responsive polymeric formulations under variations of pH or temperature. On the one hand, the liquid phase allows the formulation spreading over a larger area than solid forms, resulting in a larger surface of absorption. On the other hand, the gel phase allows prolonging the time residence of the formulation in the buccal cavity. Thus, in situ gelling systems exhibit the main requirements of a buccal dosage form. The developed hydrogels should exhibit suitable mechanical and mucoadhesive properties including appropriate viscosity, ease of dispersion at the mucosa surface, and extended residence time in the buccal cavity.

In the literature, several studies show the impact of adding mucoadhesive polymers into a buccal form to enhance its time residence in the buccal cavity (Gandhi and Robinson, 1994, Nafee et al., 2004, Salamat-Miller et al., 2005). The adhesive polymers can be classified as synthetic and natural molecules, charged and uncharged types. The natural bioadhesive polymers, like chitosan (Ilium, 1998), hyaluronic acid (Liao et al., 2005) and various gums (guar, alginate, xanthan, carrageenan) (Bhardwaj et al., 2000, Mughal et al., 2011), are considerably developed because of their biocompatibility, biodegradability and low toxicity. Xanthan gum, obtained by the fermentation of the bacterium Xanthomonas campestris, is an anionic polysaccharide. It represents an attractive biopolymer for a variety of biomedical and pharmaceutical applications (Rowe et al., 2005). It has also been demonstrated to possess mucoadhesive properties in the development of ophthalmic devices (Bhowmik et al., 2013, Ceulemans et al., 2002).

The first part of this work focuses on the measurement of the mucoadhesive properties of a previously described thermosensitive in situ gelling system for buccal administration (Zeng et al., 2014). A series of hydrogels based on the mixture of poloxamer analogs and xanthan gum have been studied for buccal delivery of Salbutamol. Their thermal, mechanical, rheological and drug release properties had been investigated. The combination of P407, P188 and Satiaxane^® (xanthan gum) provided not only an optimal sol-gel transition temperature (28–34 °C), but also an enhanced gel strength, which sustained the drug release especially in the presence of Satiaxane^®.

The determination of the adhesive strength between the formulation and the biological substrate is typically used to assess mucoadhesion capacity. Different apparatus have been described to determine forces of detachment using shearing, tensile or peeling methods (Andrews et al., 2009). Many works reported the use of animal mucosa tissue (Al-Dhubiab et al., 2014, Kanjanabat and Pongjanyakul, 2011, Karavana et al., 2009). However, using animal mucosa tissue raises conservation problems and requires an intimate fixation of the tissue on the dynamometer plate which is very difficult to achieve in a reproducible way. In this context, a mucin film technique was used in this study to avoid the concerns of tissue conservation and proved effective to demonstrate Satiaxane^® mucoadhesive properties (Zeng et al., 2014).

The second part of this work focuses on the validation of a Cyanine derivative as a suitable marker for the in vivo visualization of the previously developed hydrogel (Zeng et al., 2014). In order to visualize and track the hydrogel in vivo, a derivative of Cyanine 5 was synthesized in order to interact with the hydrophobic part of the pluronic triblock copolymers and incorporated into the hydrogel to label the form. To validate the methyl cyanine 5 derivative (Me-Cy5) as a suitable marker of our hydrogels, and detect the hydrogels in vivo, dissolution test and epithelium permeation test on Transwell^® device were carried out. The human buccal epithelium was formed by culturing TR146 cells, which originate from a neck node metastasis of a human buccal epithelium carcinoma (Jacobsen et al., 1999). Finally, in vivo study in mice was performed by optical imaging, the time-response profile of fluorescence intensity in the buccal zone reflects the duration of the formulation residence.