Cyclodextrin-containing Polymers

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Cyclodextrins are cyclic oligosaccharides that are biocompatible, exhibit low toxicity and immunogenicity in humans, and have strong resistance to degradation by human enzymes [115]. They can be incorporated into a cationic linear backbone to form cyclodextrin-containing polymers (CDPs, Figure 2A), which can package siRNA for intracellular delivery.

One prominent example of CDP-based siRNA delivery is the RNAi/Oligonucleotide Nanoparticle Delivery, or RONDEL platform [116]. At the heart of RONDEL are CDPs which consist of 5-6 repeating units of β-cyclodextrin coupled to amidine charge centres and terminal imidazoles (Figure 2A). These CDPs provide several functions to the delivery system: (1) positively charged amidines enable electrostatic interactions with siRNA, (2) CDPs condense siRNAs into nanoparticles and protect them from nuclease degradation, and (3) imidazole groups utilise the proton sponge effect to promote endosomal escape.

CDPs also serve as a structural scaffold onto which additional components can be incorporated, including PEG for steric stabilisation [117] and immunomasking of the delivery system [118], and Tf for preferential tumour-targeting. By coupling PEG and PEG-Tf to adamantane, a cycloalkane which forms high affinity inclusion complexes with β-cyclodextrin [119], the CDP-siRNA core can be non-covalently surface decorated with these ligands (Figure 2B). Assembled nanoparticles are 70 nm in diameter and thus benefit from EPR, and each contains approximately 10,000 CDPs, 2,000 siRNAs, 4,000 PEG-adamantanes and 100 Tf-PEG-adamatanes [120]. Furthermore, the zeta potential of RONDEL particles can be adjusted from +15 mV to -25 mV through incorporation of AD-PEGs modified to contain an anionic charge [121].

RONDEL demonstrated good efficacy when tested in a metastatic murine model of Ewing’s sarcoma, in which RONDEL was used to deliver siRNAs that targeted the EWS/Fli1 fusion oncogene, resulting in effective silencing of the oncogene and significant anti-tumour effects [122]. Further, good tolerability of the delivery system was shown in cynomolgus monkeys at dosages of 3 and 9 mg siRNA/kg [118]. These promising results led to the advancement of RONDEL into clinical trials. Calando Pharmaceuticals’ CALAA-01, a drug candidate currently undergoing phase 1b clinical evaluation [112], utilises RONDEL to deliver systemically siRNA targeted against the M2 subunit of ribonucleotide reductase (RRM2) to solid tumours. Ribonucleotide reductase is required for DNA synthesis and is a key component in the proliferation of cancer cells [123]. CALAA-01 has so far demonstrated promising results in human patients with metastatic melanoma refractory to standard therapies, in which delivered siRNAs localised to tumours, and levels of RRM2 mRNA decreased in a dose-dependent manner [124].