Complete destruction of deep-tissue buried tumors via combination of gene silencing and gold nanoechinus-mediated photodynamic therapy
Introduction
Treating deep tissue-buried tumors with the existing medical technologies is one of the long-lasting challenges in the cancer therapy [1], [2]. Clinically, it is sometimes impossible to treat the deep tissue-buried tumors (for example, lung and liver cancers), especially when a patient's physical condition cannot co-operate and tolerate with the invasive surgical treatments [3]. Therefore, there is a great demand for the development of less or non-invasive modalities that can treat the complicated deeply seated tumors. In the recent years, phototherapies were widely explored for treating various types of cancers [4], [5]. The major advantages of phototherapies are (1) non-invasiveness, (2) superior tissue penetration, (3) reduced side effects, and (4) more cost-effective than conventional treatment modalities. Photodynamic therapy (PDT) was emerged where it involves the use of an organic photosensitizer (PS) molecule to absorb and transfer the photon energy to the normal tissue oxygen (3O2) to generate cytotoxic singlet oxygen (1O2), which is able to kill cancer cells [6], [7]. Most of the clinical PDT treatments are restricted to surface tumors, because organic PS can be photochemically excited by either uv or visible light, which have very short tissue penetration depths. To treat deep tissue-buried tumors using PDT, one has to use NIR light as the light source since NIR light has large tissue penetration depths. However, organic PS able to be excited by NIR light is very rare. Therefore, it leaves a grand challenge to treat deep seated tumors with the existing photodynamic therapeutic drug molecules.
To overcome the absorption of NIR light and excitation of organic photosensitizers, upconversion nanoparticles (UCNPs) co-doped with organic PS were developed to absorb and convert the deeply penetrating NIR light to visible wavelengths, which was then used to photochemically excite the co-loaded organic PS to facilitate organic PS-mediated PDT in treating deep tissue buried tumors [8], [9], [10]. In such kind of nanomaterial design, partial tumor suppression or delay tumor growths were usually achieved. Many limitations are associated with the design of UCNPs-doped organic PS-mediated PDT, for example, (1) only a single choice of wavelength of 980 nm can be used to excite UCNPs; (2) high incident laser power (0.33–2.5 W/cm2) [8], [9], [10] has to be used due to the very low extinction coefficient of UCNPs at 980 nm; (3) water has very strong absorption at 980 nm wavelength and thus creates the overheating problem; (4) very low singlet O2 generation efficiency was obtained due to the multi-step processes (via UCNPs light absorption efficiency x upconversion emission quantum yield × efficiency of light absorption of organic photosensitizers x singlet O2 sensitization efficiency of organic photosensitizers); and (5) only delay growth or partial suppressions, rather than complete destruction, of tumors was achieved [8], [9], [10]. Previously, gold nanorods-in-shell particles were reported to be able to absorb and convert 1064 nm light (NIR II) to heat and mediate photothermal therapy (PTT) effect for the treatment of surface tumors. However, only partial tumor suppression was achieved, albeit a very strong incident laser power of 3 W/cm2 was used. The use of high laser power, beyond the skin tolerance threshold value (0.42 W/cm2) prescribed by the American National Standards Institute (ANSI), severely limits its practical clinical applications [11]. Previously, we have reported a morphology of a gold nanostructure, called gold nanoechinus (Au NEs), which possesses ultra-high molar extinction coefficients (∼1012 M−1cm−1) and can also sensitize formation of singlet oxygen in the NIR windows I and II to exert in vivo nanomaterial-mediated photodynamic and photothermal therapeutic (NmPDT & NmPTT) for the destruction of surface tumors in mice [12]. Due to its exceptionally high extinction coefficients in the NIR windows I and II, this unique nanomaterial might have great potential for treating deep tissue-buried tumors, which is not yet proven or demonstrated.
It is often highly difficult to achieve successful therapeutic destruction of deep tissue-buried tumors using a single treatment modality. Gene therapy involves knocking down or silencing of specific oncogenes, and plays a vital role in treating surface tumors [13], [14]. Superoxide dismutase 1 (SOD1) is one of the very effective anti-apoptotic and self-defending genes, and can destroy the free radicals or reactive oxygen species in the human body. Knocking down of anti-apoptotic genes can trigger apoptosis and induce cellular deaths very efficiently [15]. Using gene silencing alone, partial suppression, rather than complete destruction, of tumor growth was usually observed [13], [14], [15]. Combining gene therapy and NmPDT for the treatment of deep tissue buried tumors might be a promising modality for treatment of solid tumors, and was not yet reported or demonstrated. To the best of our knowledge, the current work is the first demonstration that complete destruction of deep tissue buried tumors can be achieved with the ultra-low doses of NIR light by combining SOD1 gene silencing and Au NEs-mediated PDT in the second biological window (NIR II; 1064 nm).
Section snippets
Preparation of lipid-coated Au NEs
Gold nanoechinus were synthesized according to the previously reported literature procedure [12]. For the preparation of lipid-coated Au NEs, the as-synthesized gold nanoechinus were washed using DI water for two times. Then, Au NEs were re-dispersed in a Lipofectamine 2000 (LP-2000) reagent (100 μL, Invitrogen, USA)-containing aqueous solution to promote the formation of lipid-bilayer coated Au NEs.
Detection of singlet oxygen phosphorescence emission
The lipid-coated Au NEs (1 mg/mL) dispersed in D2O was used for the singlet oxygen
In vitro combination of Au NEs-mediated photodynamic therapy and SOD1 gene silencing
The synthesis of Au NEs was carried out according to the previously reported modified seed-growth procedure [12]. The detailed structure characterizations of Au NEs were shown in the Fig. S1. The UV–Vis–NIR spectrum of Au NEs exhibits a broad NIR absorption extending to 1700 nm. The molar extinction coefficients for Au NEs were determined to be 1.51 × 1012 M−1cm−1 at 915 nm (NIR I) and 1.54 × 1012 M−1cm−1 at 1064 nm (NIR II), respectively. The excitation spectrum for the singlet O2
Conclusions
In summary, we have achieved complete destruction of deep tissue-buried tumors (0.5 cm depth) using ultra-low doses of NIR light via combination of highly efficient gene silencing effects of SOD1 gene and Au NEs-mediated PDT in the first and the second biological windows (NIR I & II). Upon transfecting siRNA-lipid-coated Au NEs in HeLa cells, ultra-high gene silencing efficiencies (∼86%) of SOD1 were achieved. Under photoirradiation, significantly higher amounts of cellular deaths were observed
Acknowledgments
The authors are grateful to the financial support from the Frontier Research Center on Fundamental and Applied Sciences of Matters, National Tsing Hua University and the Ministry of Science and Technology, Taiwan (103-2113-M-007-011-MY3).
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2019, Coordination Chemistry ReviewsCitation Excerpt :Advances in research regarding cellular mechanisms of PDT have shed light on numerous possible combination treatments. To date, PDT has been combined with ionizing radiation [162–164], chemotherapy [133,165–167], photothermal therapy (PTT) [168–172], and gene therapy [173–175] in cells and animals and even show additive benefits. Considering the NIR-light-conversion function of UCNPs, the use of NIR-excitable UCNPs as PS carriers provides a robust strategy to overcome the main drawback in conventional PDT, and meanwhile, UCNP-PS with ingenious design can be used as the agents for combined treatment with synergistic anticancer effects.