In this work, we present the synthesis of NCPs complexes for its potential use as CA of a glioblastoma. The glioblastoma is one of the most common and lethal tumours of the central nervous system.41 It belongs to malignant gliomas which are a group of heterogeneous and invasive brain tumours derived from glial cells. As reported, the 5-year survival rate for glioblastoma is limited to 5%.42 Despite of remarkable advances in basic and clinical research, the clinicians are yet unable to provide realistic therapy for this kind of tumours. For this reason, providing to the community with novel systems which enhances its early diagnosis could help to the increase of the effectiveness of therapies. Nanoparticles have being used in clinical studies and pre-clinical models for different applications in brain tumours for its improvement from a diagnosis and therapy point of view.43-45 The main challenge of a nanosystem to reach the brain is to cross the BBB and the most used strategy is the attachment of peptides on the nanoparticle surface that act as BBB shuttle.46 Nevertheless, some brain diseases in advanced stage have the BBB compromised as in the case of glioblastoma. In this specific pathology, the BBB becomes permeable to different molecules and the microenvironment involved changes, creating a disorganization of vessel structures resulting in an uncontrolled cell migration and in abnormal vascular architecture.47 In this scenario, nanoparticles can play an important role especially in the area of CAs by crossing the BBB and enhancing the tumour imaging.
Currently, most of the examples of nanoplatforms able to cross the BBB for the treatment and diagnosis of tumour brains are related with gold- and iron-based nanoparticles.47,48 Gold nanoparticles has demonstrated their successful accumulation in tumours for hours allowing the imaging of the affected area by MRI. Nevertheless, there is still no evidence of their use in clinical trials mainly due to their unclear interaction with the metabolism and their potential toxic effects on humans. Regarding iron-based nanoparticles, although the SPIONs systems have emerged as a good candidate for brain imaging, their potential us is still limited due to the need of detailed understanding of their in vivo effects.
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As we have seen, toxicity issues related with solid metal nanoparticles force to researchers to develop more efficient, safer alternatives with greater tuneability options. Based on this, we proposed in this work for the first time the use of NCPs as potential candidates for the imaging of glioblastoma tumours. Furthermore, we used MRI as the gold standard equipment for the diagnosis and pre-surgical planning of brain imaging.
4.2.1.5 Present challenges
Despite the progress of DMCAs, some questions remain to be addressed before the clinical application of DMCAs becomes a reality:
i. Facile and cheap synthesis procedures with possibilities for scale up
ii. Chemical stability in biological environment by non- toxicity and non-accumulation in the organism (e. g. avoiding the use of Gd and Mn ions)
iii. High efficiency in different tissues by good biodistribution iv. In vitro and in vivo proof of their performance
These key parameters must be considered during the rational design of novel DMCAs overcoming the main drawbacks of current clinical approved CAs. Therefore, there is still need to overcome the disadvantages of single modality CAs by the preparation of robust DMCAs which are supposed to minimize the risks of ambiguity and improve the diagnostic sensitivity. Furthermore, in case of brain tumours, the information provided by a T2 and especially by a DMCA in its diagnosis or monitoring, is not fully evaluated yet.
4.2.1.6 Our choice
The knowledge of our group in the development of NCPs was the starting point in this work for the successful synthesis of NCPs-based MRI CAs. In this Chapter, the synthetic protocol included the polymerization process of different coordination complexes obtained by the combination of ligands such as 1,4-Bis(imidazol-1-ylmethyl)benzene (L3, Bix), 3,4-dihydroxycinnamic acid (L4, dhc) and diethylenetriaminepentaacetic acid (L5, DTPA) with a variety of metal salts (Fe(II), Mn(II) and Gd(III)) for the synthesis of a set of MRI-active NCPs (Figure 4.8). Through this rationale design, four NCPs complexes were obtained: Fe-NCP, Gd-NCP, Mn-NCP and GdDTPA-NCP. All the systems were fully characterized by different physicochemical techniques and its behaviour in physiological medium in vitro and in vivo was tested.
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Chapter 4.2
Figure 4.8 Scheme for the synthesis of the different NCPs complexes developed in this work. From the L3 together with L4 and combining them with Fe(II), Gd(III) and Mn(II) metal salts, three NCPs complexes were synthesized: Fe-NCP, Gd-NCP and Mn-NCP. Following similar strategy, L3 was combined with L5 and Gd(III) metal salt for the synthesis of GdDTPA-NCP.
Most of the clinically approved CAs are not able to cross the intact BBB but can reach a lesion if at some point the BBB is disrupted as in the case of glioblastoma tumours.49 For this reason, the model selected for the in vivo studies was the GL261 murine model. The immunocompetent GL261 mouse glioma line is one of the most widely used to study the glioblastoma disease and is usually inoculated into C57BL/6 strain, resulting in invasive and infiltrative tumours, characteristics which can be similarly found in human patients with glioblastoma.50
4.2.1.7 Objectives
With the aim to overcome the challenges aforementioned, different objectives were proposed derived from the following two main goals:
Design, synthesis and characterization of novel NCPs based on active MRI ions (Fe(III), Gd(III) and Mn(II)) for their use as DMCAs for glioblastoma imaging
The systems must accomplish i) moderate chemical stability, ii) water-colloidal stability, iii) low toxicity and biocompatibility, iv) intrinsic T1 and T2 imaging activity in vitro and in vivo and v) good biodistribution with low up-take by liver and kidneys.
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Dual T1/T2 Nanoscale Coordination Polymers as Novel Contrast Agents for MRI: A Preclinical Study for Brain Tumour
The steps followed for the achievement of the main goals were the following:
i. Synthesis of complexes Fe-NCP, Gd-NCP, Mn-NCP and GdDTPA-NCP ii. Complete physicochemical and biological characterization in vitro
iii. Pre-clinical in vivo studies with a glioblastoma mice model iv. Comparison with commercial CAs
In this work we show how the coordination chemistry allows the successfully design of novel NCPs that incorporate MRI-active metals polymerised with organic ligands. Furthermore, the dual activity of one of the systems will allow its transfer to pre-clinical studies to evaluate its performance in an in vivo environment.
4.2.2 Results and discussion