  One of the major limitations of nanomedicine is the scarce penetration of
nanoparticles in tumoral tissues. These constrains have been tried to be solved
by different strategies, such as the employ of polyethyleneglycol (PEG) to
avoid the opsonization or reducing the extracellular matrix (ECM) density. Our
research group has developed some strategies to overcome these limitations such
as the employ of pH-sensitive collagenase nanocapsules for the digestion of the
collagen-rich extracellular matrix present in most of tumoral tissues. However,
a deeper understanding of physicochemical kinetics involved in the nanocapsules
degradation process is needed to understand the nanocapsule framework
degradation process produced during the penetration in the tissue. For this, in
this work it has been employed a double-fluorescent labelling strategy of the
polymeric enzyme nanocapsule as a crucial chemical tool which allowed the
analysis of nanocapsules and free collagenase during the diffusion process
throughout a tumour-like collagen matrix. This extrinsic label strategy
provides far greater advantages for observing biological processes. For the
detection of enzyme, collagenase has been labelled with fluorescein
Isothiocyanate (FITC), whereas the nanocapsule surface was labelled with
rhodamine Isothiocyanate (RITC). Thus, it has been possible to monitor the
hydrolysis of nanocapsules and their diffusion throughout a thick 3D Collagen
gel during the time, obtaining a detailed temporal evaluation of the
pH-sensitive collagenase nanocapsule behaviour. These collagenase nanocapsules
displayed a high enzymatic activity in low concentrations at acidic pH, and
their efficiency to penetrate into tissue models pave the way to a wide range
of possible nanomedical applications, especially in cancer therapy.
