To obtain optical molecular images that reflect changes in biomarker expression requires the ability to deliver contrast agents to cells and tissues of interest, as well as the ability to wash away unbound contrast agent. Two primary routes of administration exist for delivery of imaging agents: topical application and iv. injection. Topical application is only appropriate for certain epithelial tissues, but provides some distinct advantages in these cases. Typically, smaller amounts of contrast agent are required for topical application; this reduces potential toxicity and clearance concerns associated with iv. injection. However, achieving sufficient epithelial permeation of topically applied contrast agents is still a major barrier, especially for larger nanoparticle-based agents. Using iv. delivery can help enhance contrast agent accumulation in tumors via the enhanced permeability and retention effect. However, iv. delivery typically requires higher dosages of contrast agents and can lead to a higher nonspecific background signal.
The major barriers to delivery are shown in . Topically applied contrast agents must permeate tight junctions in the epithelium and in some cases a tough, protective keratinized epithelium further hampers delivery. By contrast, agents administered through iv. injection must evade degradation and the immune system to reach the target organ. These iv. agents must then localize to the area of interest and leave the circulatory system by passing through endothelial tight junctions. If the iv. contrast agent targets epithelial cells, it must also be transported through any stromal barriers. In addition, cytoplasmic- and nuclear-targeted agents delivered via either mechanism must also traverse the cell and nuclear membranes, respectively, to reach the intended target.
Barriers to contrast agent delivery
The tight junctions of the epithelium serve as a barrier to exclude foreign agents, so crossing this delivery barrier is not trivial. Chemical permeation enhancers such as Triton™-X and modified chitosan have been used to improve permeation in excised tissues [110
]. Alternatively, pulsed ultrasound has been used to enhance permeation of 20 nm particles into the core of MCF-7 breast cancer spheroids compared with those not exposed to ultrasound [113
]. These results show the potential for increasing delivery of imaging and therapeutic agents to tumor tissue and the effect of particle properties on penetration. Thick keratinized surfaces are not affected by many permeation enhancers and alternatives are needed for topical delivery of contrast agents to keratinizing epithelial tissues.
Contrast agent design and modification is especially important for iv. agents, as they must evade the immune system and avoid degradation. Size, shape and surface chemistry are critical for effective delivery. Polyethlyene glycol is commonly used to create ‘stealth’ particles that evade immune clearance. For delivery to tumors, iv. delivery can leverage contrast agent accumulation by utilizing the enhanced permeability and retention effect due to poorly formed capillary networks near the tumor. However, in early neoplasia, agents must traverse the robust vascular endothelium to reach the targeted neoplastic cells. Early epithelial neoplasias near the tissue surface may not be accessible through the vascular network at all.
Delivery to cytoplasmic and nuclear targets adds an additional level of complexity. Peptides are a popular tool to overcome these barriers as many of them are inspired by viral systems that have successfully evolved to overcome barriers to intracellular delivery. Cytosolic delivery of gold nanoparticles (20 nm) was accomplished by conjugating a targeting antibody against actin and a TAT-HA2 peptide sequence to the surface of the particle [70
]. The actin antibody targets the nanoparticle to a specific structure in the cell, while the TAT portion of the peptide enables endocytosis of the particles, and HA2 disrupts the endosomal lipid membrane enabling endosomal escape to the cytosol. Kang et al.
used the nuclear localization signal peptide to target gold nanoparticles to the nucleus, causing cytokinesis arrest and resulting in the apoptosis of cancer cells in vitro
]. A nuclear localization signal was also used by Oyelere et al.
to improve delivery to the nucleus in vitro
]. However, these particular particles will probably face other barriers following iv. delivery. The size of these particles would hinder iv. delivery due to retention in the blood pool followed by uptake in the liver [116