Mn²⁺-doped silica nanoparticles for hepatocyte-targeted detection of liver cancer in T₁-weighted MRI

Abstract

With an aim to examine the possibility of developing a liver-specific MRI contrast agent that takes advantages of brightly enhanced MR images by Mn²⁺ whilst making up the limitations of the pre-developed contrast agent, the Mn²⁺-doped SiO₂ nanoparticles (Mn–SiO₂) were synthesized and their characteristics as MR contrast agents were investigated. The in vitro and in vivo investigations showed that Mn–SiO₂ has unique MR contrast-enhancing characteristics that activate positive contrast enhancement in T₁-weighted MR images only under low pH conditions by liberating Mn²⁺ ions from MR inactive nanoparticles. The administration of Mn–SiO₂ to an orthotopic xenograft model of human hepatocellular carcinoma (HCC) resulted in a differentiation of enhancement periods between HCC and normal parenchyma tissues on T₁-weighted MR images and consequently presented the duplicates of the highly contrast-enhanced liver image with an equal liver-to-HCC contrast ratio but opposite contrast. The Mn–SiO₂-enhanced MR imaging therefore allowed for the repetitive detection of the HCC within a single MR imaging session, which can help us to achieve more reliable diagnosis and characterization of liver lesions than is possible with any currently used Mn²⁺-based contrast agent. In addition, the in vivo biodistribution study also supported the effectiveness of Mn–SiO₂ nanoparticles as a liver-specific MRI contrast agent, which efficiently delivers and releases the T₁-contrasting Mn²⁺ ions to targeted hepatocytes.

Introduction

The use of contrast agents in magnetic resonance (MR) imaging improves the detection of liver lesions by increasing the lesion-to-liver contrast and, in some situations, facilitating the lesion characterization [1], [2], [3], [4], [5]. Non-specific extracellular fluid (ECF) agents, that rely on differential blood flow of paramagnetic Gd³⁺ chelate complexes (Gd) between liver and lesion, have been the most widely employed contrast agents for liver MR imaging [6], [7], [8]. However, even well-performed dynamic MR imaging using ECF agents often requires difficult operations, such as rapid imaging acquisition and highly accurate initiation timing. Of even greater concern is the possibility of nephrogenic systemic fibrosis (NSF) resulting from the presence of dechelated Gd³⁺ ions [9], [10]. Recently, liver-specific contrast agents such as superparamagnetic iron oxide nanoparticles (SPIO) and mangafodipir trisodium (MnDPDP), which target the main cell populations of the liver, were developed to overcome the limitations of ECF agents. SPIO-enhanced MR imaging, which exploits the preferential uptake of SPIOs into a normal liver via Kupffer cells, was once employed in liver diagnosis using T₂-weighted MR imaging [11], [12], [13], [14], [15], [16], [17]. However, these agents are no longer clinically used, likely owing to a number of drawbacks, including ambiguity in distinguishing a hypointense contrast effect by the SPIOs from that of bleeding or calcifications [18]. MnDPDP was developed as a hepatobiliary contrast agent that is taken up by hepatocytes and excreted in bile. [19], [20], [21], [22], [23], [24] The intravenously administrated MnDPDPs are transmetallated by Zn²⁺ ions in the bloodstream and release the free Mn²⁺ ions [25]. The Mn²⁺ ions are then taken up by hepatocytes, resulting in a hyperintense signal-intensity change on T₁-weighted MR images. The MnDPDP-enhanced MR imaging is known to be advantageous in characterizing lesions as hepatocellular or non-hepatocellular. Whereas, because any hepatocellular tissues, including lesions, are enhanced in a similar time course, by the non-specifically incorporated Mn²⁺ ions, the MnDPDP-enhanced MRI usually does not give a high lesion-to-liver contrast ratio. In terms of toxicity, the increased blood concentration of free Mn²⁺ ions, which interfere with myocardial processing of Ca²⁺, may result in a toxic effect on the cardiovascular and brain systems [26], [27].

The aim of the present study was to examine the possibility of developing a liver-specific MRI contrast agent that takes advantages of brightly enhanced MR images by Mn²⁺, without the limitations of the pre-developed MnDPDP (Teslascan). For this purpose, we incorporated Mn²⁺ ions in an amorphous SiO₂ nanoparticle, which is generally considered safe by the U. S. Federal Drug Administration (FDA) [28], [29], by using a pH-responsive material that liberates Mn²⁺ ions only under acidic conditions [30], [31], [32], [33], [34]. It was hypothesized that the engulfment of the Mn²⁺-doped SiO₂ (Mn–SiO₂) nanoparticles in acidic endosomal vesicles by Kupffer cells would trigger the release of Mn²⁺ ions [35], [36], enhancing normal liver tissues with a high density of Kupffer cells over lesions, which are devoid of Kupffer cells, on T₁-weighted MR imaging. Therefore, the administration of Mn–SiO₂ nanoparticles may lead to the time-sequential signal enhancement of normal and lesion tissues, enhancing their contrast ratio and improving the lesion visibility in liver MR imaging. In addition, the use of Mn–SiO₂ nanoparticles may allow for efficient, targeted delivery of a contrast agent to the liver, without releasing the potentially toxic Mn²⁺ ions in the nearby neutral bloodstream.