Elsevier

Acta Biomaterialia

Volume 86, 1 March 2019, Pages 416-428
Acta Biomaterialia

Full length article
CaP coated mesoporous polydopamine nanoparticles with responsive membrane permeation ability for combined photothermal and siRNA therapy

https://doi.org/10.1016/j.actbio.2019.01.002Get rights and content

Abstract

Combined photothermal and gene therapy provides a promising modality toward cancer treatment, yet facile integration and controlled codelivery of gene payloads and photothermal conversion agents (PTCAs) remains a great challenge. Inspired by the robust wet adhesion of marine mussels, we present a rationally designed nanosystem constructed by using hybrid mesoporous polydopamine nanoparticles (MPDA) with sub-100 nm sizes and a high photothermal conversion efficiency of 37%. The surface of the particles were modified with tertiary amines by the facile Michael addition/Schiff base reactions of PDA to realize high siRNA loading capacity (10 wt%). Moreover, a successful calcium phosphate (CaP) coating via biomineralization was constructed on the cationic nanoparticle to prohibit premature release of siRNA. The CaP coating underwent biodegradation in weakly-acidic subcellular conditions (lysosomes). The synergistic integration of tertiary amines and catechol moieties on the subsequently exposed surfaces was demonstrated to feature the destabilization/disruption ability toward model cellular membranes via the greatly enhanced interfacial adhesion and interactions. Consequently, sufficient permeability of lysosomal membranes, and in turn, a high lysosomal escape efficiency, was realized, which then resulted in high gene silencing efficiencies via sufficient cytosolic delivery of siRNA. When an efficient knocking down (65%) of survivin (an inhibitor of apoptosis proteins) was combined with a subsequent photothermal ablation, remarkably higher therapeutic efficiencies were observed both in vitro and in vivo, as compared with monotherapy. The system may help to pave a new avenue on the utilization of bio-adhesive surfaces for handling the obstacles of combined photothermal and gene therapy.

Statement of Significance

Polydopamine (PDA) based porous photothermal-conversion agent (PTCA) with sufficiently high conversion efficiency was employed to deliver photothermal/gene therapy modalities towards cancer treatment.

CaP coating via PDA-induced biomineralization was constructed to prohibit premature release of siRNA loaded in the pore space of the nanocarriers.

Responsive degradation of CaP also led to the exposure of membrane-lytic surfaces built through the synergistic integration of tertiary amines and catechol moieties, and in turn the significantly enhanced lysosomal escape and cytosol siRNA delivery.

Therapeutic targeting of survivin was successfully applied for activation of apoptosis and programmed cell death. Combined photothermal and gene therapy improved therapeutic effectiveness.

Introduction

Combined cancer therapy, which integrates several therapeutic modalities into a single system with the assistance of nanotechnology, has represented a great potential and experienced solid progress in overcoming the limitation of monotherapy [1]. Among the exploited strategies up to now, the combination of photothermal therapy (PTT) and siRNA based gene therapy (GT) has attracted increasing attention due to the superiority in increasing therapeutic efficacy and decreasing side effects [2], [3], [4]. In this system, facile integration and controlled codelivery of two therapeutic agents, i.e. siRNA payloads and photothermal conversion agents (PTCAs), was pivotal to the improvement of the overall therapeutic outcome. To this end, the idea of using porous PTCAs was considered as a significant success in simplifying the integration, as well as realizing large loading and effective protection of siRNA by the pore space [5]. Notwithstanding the first demonstration of utilizing mesoporous carbon nanoparticles [5], there still exist rational-design challenges in terms of easier surface functionalization, and efficient escape of the nanocarriers from lysosomes after cell uptake, the latter of which has long been recognized as a major bottleneck in cytosolic delivery of siRNA [6], [7], [8].

Recent subversive insights suggested that the classical polyplex-mediated compartmental escape actually relies on the time-dependent membrane permeabilization (via multiple transient pores) by tight apposition of the polyplex and the inner-lysosomal membrane [9], [10]. However, the hydration repulsion between membranes and polar surfaces reduces the mutual perturbation of biomolecular assemblies in the congested cellular environment [11], [12]. Consequently, the abundant body fluids and water in organelles severely hamper the adhesion of siRNA vehicles to inner membrane surfaces via the hydration layers, which has not received enough attention in the past [6], [7], [13]. In this regard, the marine mussel byssal plaque has become a star model for the biomimetic wet adhesion because of the adhesive catecholic amino acid called 3,4-dihydroxyphenylalanine (Dopa) which can form various types of interfacial interactions such as hydrogen bonding, metal chelation, π-π and/or cation-π interaction [14], [15].

Mussel-inspired polydopamine (PDA), has attracted immense attention in the community of drug delivery nanocarriers, owing to its advantages in wet adhesion, photothermal conversion, and easy functionalization [16]. Among representative breakthroughs, mesoporous PDA nanoparticles are being developed by our group and others [17], [18], while findings in PDA-assisted biomineralization [19], [20], [21] holds a great promise in simple surface encapsulation. Besides, continuous research efforts are also being devoted to exploring new application properties originated from PDA’s bioadhesive feature. Noteworthily, the binding strength of Dopa containing molecules to surfaces was demonstrated to be significantly enhanced by neighboring positively charged lysine or arginine residues [22], [23]. Molecular-level insights into the adhesive mechanisms attributed such a synergy to the role of catechols and amines in cooperative displacing surface-bound salt ions, and in turn the hydration, which allows for effective adhesion and interfacial interactions [24], [25]. Therefore, we anticipated that surface modification of amines on porous PDA-based nanocarriers would address the challenge in lysosomal escape.

Herein, we report an efficient siRNA-delivery and PTT system of hybrid mesoporous nanoparticles integrating functions of biomimetic wet-adhesion and biomineralization encapsulation via the combination of the instinct properties from bio-inspired PDA. As shown in Scheme 1, mesoporous PDA nanoparticles (MPDA) with sub-100 nm sizes were prepared as a PTCA with the surface modification of tertiary amines, which facilitated the siRNA loading inside mesopores. Premature release of siRNA was prohibited by calcium phosphate (CaP) coating, which was realized by PDA-induced biomineralization. Facilitated by efficient cellular uptake, pH-responsive CaP degradation, and destabilization of lysosome membrane, an efficient lysosome escape was subsequently demonstrated by in vitro studies. As a consequence, high gene silencing efficiencies can be obtained. The efficiency in siRNA induced therapy was revealed by downregulation of survivin (an inhibitor of apoptosis proteins). The subsequent photothermal ablation resulted in substantially improved therapeutic effectiveness. The novel delivery system is expected to provide new pathways and considerations on the utilization of bio-adhesive surfaces for nanocarriers mediated GT and PTT combination.

Section snippets

Materials

Unless otherwise noted, all reagent-grade chemicals were used as received, and distilled water was used for the preparation of all aqueous solutions. 2-Morpholinoethanesulfonic acid (MES), Ethanol (AR) and acetone (AR) were purchased from Fluka. Pluronic® F-127 and FITC-dextran (M.W. 10,000) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Dopamine hydrochloride (98%, AR), fluorescein isothiocyanate (FITC, 90%), tris (hydroxymethyl) aminomethane (Tris, 99.9%), Acridine Orange, N,N

Preparation, characterization, and siRNA loading of MPDA nanocarriers

MPDA was first synthesized based on the assembly of primary PDA particles and Pluronic F127 stabilized emulsion droplets on water/1,3,5-trimethylbenzene (TMB) interfaces, according to the procedure in a previous study by us [17]. To impart the particle surface with positive charges and high affinity of siRNA within mesopores, MPDA was modified by N,N-dimethylethylenediamine (DMEA) bearing two different amine groups. Herein, the primary amine of DMEA facilitates the conjugation towards PDA by

Conclusion

In summary, we have demonstrated that the synergistic integration of tertiary amines and catechol groups on surfaces of porous nanocarriers can offer a simple and highly efficient solution to handle the obstacle of combined photothermal and gene therapy on the basis of porous photothermal conversion agents. Application of this nanosystem gave an example on cooperative integration of siRNA delivery and heat generation by a porous photothermal conversion agent (PTCA). Most importantly, the

Acknowledgements

This work was supported in part by the National Natural Science Foundation of China (NSFC, Grant Nos. 51773022, 21734002, 51825302), Graduate research and innovation foundation of Chongqing, China (Grant No. CYB18028), Basic Advanced Research Project of Chongqing (Grant No. cstc2015jcyjA10051), the 100 Talents Program of Chongqing University (J.Z.), and Innovation Team in University of Chongqing Municipal Government (CXTDX201601002).

Declarations of interest

None.

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