Route of delivery

1.4.1.1 Systemic administration

This route often fails in the treatment of bladder cancer due to loss of the therapeutic agent from metabolism in the liver and poor tumour penetration. There is a need for a sufficient level and an appropriate amount of time for the therapeutic agent to come into contact with the tumour cells in the bladder mucosa in order to achieve its effect which is not achieved via this route.

1.4.1.2 Intravesical administration

The bladder represents an ideal target for intravesical gene therapy in the management of bladder cancer. The vectors are in direct local tumour contact, eliminating the difficulties associated with systemic administration and ensuring a low risk of gene transfer to other organs. Easy external access is achieved through the urethra which avoids losses from the first pass metabolism in the liver and minimizes the clearance of the vector by the immune system allowing minimal systemic side effects.

The inside of the bladder consists of 6-7 layers of cells which form a permeable transitional epithelium known as the urothelium (Figure1.3). The urothelium acts as a barrier and in healthy circumstances it is almost impermeable. The main part of this barrier is the superficial cell layer of the transitional epithelium made up of umbrella cells. The umbrella cells join to each other with tight junctions and constitute a watertight barrier (Hohlbrugger 1995). The apical surface of the umbrella cells is covered by uroplakins (UPIa, UPIb, UPII, UPIII), which make up the rigid plaques. In addition the umbrella cells are covered by a hydrophilic glycosaminoglycan (GAG) layer. The GAG layer physically blocks the adhesion of foreign substances such as drugs and viruses (Lande et al. 1995; Parsons 1994). Together these layers erect a tough bladder barrier that maintains a high electrochemical gradient between the urine and the blood. The impermeability of the urothelium is frequently exploited for instilling potentially toxic agents into the bladder to achieve localized pharmalogical effect.

Figure 1.3 Structure of bladder transitional epithelium on a H&E staining histology sample. The localisation of tight junction and GAG layer that is not visible under normal microscope is marked in red.

Several strategies have been developed to improve the intravesical delivery of drugs in the bladder. This includes helping them cross the permeability barrier of the urothelium by both physical and chemical enhancement methods and increasing residence time in the bladder. These approaches may also improve the efficacy of some gene therapy vectors.

1.4.1.2.1 Improving permeability using physical methods.

The permeability of bladder cells can be improved with physical approaches like electromotive drug administration (EMDA), thermotherapy or iontophoresis which was further discussed in section 1.2.4.5.

1.4.1.2.2 Improving permeability using chemical approaches.

Several chemical approaches that improve the permeability of the urothelium have been investigated in the literature. Protamine sulfate interacts with the GAG layer to increase the permeability of the urothelium (Cetinel et al. 2003; Tzan et al. 1994) whilst

Bladder lumen

Umbrella cells

Urothelium GAG layer

Tight junction

administration of dimethyl sulfoxide has been described to enhance the absorption of paclitaxel and pirarubicin (Chen et al. 2003; Hashimoto et al. 1992). The administration of 22% ethanol prior to the instillation of an adenoviral vector causes damage of the GAG layer and makes the treatment more effective (Engler et al. 1999). In addition a polyamide containing formulation, syn3, when used as a pre-treatment prior to intravesically administered adenovirus has been reported to enhance viral uptake (Yamashita et al. 2002). Recent studies in preclinical models with the syn3/adenoviral-based strategy have also shown an improved therapeutic effect (Benedict et al. 2004;

Connor et al. 2005). A phase I study of the Big CHAP polyamide and an adenovirus coadministered intravesically led to increased transduction compared to adenovirus alone (Kuball et al. 2002). Although the use of Syn3 appears to be non toxic to the bladder tissue the use of other transducing agents has been shown to have adverse effects on the urothelium (Benedict et al. 2004).

1.4.1.2.3 Increasing residence time in the bladder

Increasing the time of residence in the bladder is an obvious strategy to increase the efficacy of a drug or therapy by increasing the duration of direct contact between drug and abnormal urothelium and ensuring constant drug concentration. This method was further discussed in section 1.2.4.3.

1.4.1.2.4 Bioadhesion

This defines the interaction between a biological surface, such as the urothelium and a polymer such as algin, chitosan, fibrinogen, gelatin, polyethhylene glycol (Ozturk et al.

2004; Tyagi et al. 2006; Ye et al. 2001) . The efficiency of drug absorption is increased after bioadhesion properties are coupled to microspheres, liposomes or nanoparticles.

For example microspheres based on chitosan are strongly mucoadhesive and their instillation was able to increase the residence time and decrease the frequency of administration of acyclovir (Genta et al. 1997).

The residual urine in the bladder before instillation and the accumulated urine during instillation can cause dilution in the concentration of the administered therapeutic agent.

Restricted fluid intake before and during instillation and the complete emptying of bladder is required in any kind of intravesical treatment (Au et al. 2001). Significant

differences in gene expression are achieved by varying physical parameters during intravesical instillation. Increased gene expression associated with larger volume instillation may be responsible for some reported variability of gene transfer to the bladder. Significantly more gene expression was detected in bladders instilled with a higher volume of viral vectors (p <0.05). Likewise, higher instillation pressures resulted in higher transgene expression in distant organs (Siemens et al. 2001).