Gadolinium-Loaded Solid Lipid Nanoparticles As A Tumor- Absorbable Contrast Agent For Early Diagnosis Of Colorectal Tumors Using Magnetic Resonance Colonography

ObjectiveMagnetic resonance (MR) contrast agents with special physiological functions are becoming realiaty in improvingaccurate cancer diagnosis at the early stage. Here, we designed multifunctional solid lipid nanoparticles (SLNs) with simultaneous loading of gadolinium (Gd) diethylenetriaminepentaacetic acid (Gd-DTPA) and octadecylamine fluorescein isothiocyanate (FITC) to achieve Gd-FITC-SLNs as a tumor-absorbablenanoparticle contrast agent for specific MR imaging (MRI) of colorectal walls.T1-weightedSE sequence MRI was performed on mice after transrectal infusion of Gd-FITC-SLNs, with the final goal ofopening new avenues for the early diagnosis of colorectaltumors.Materials and MethodsGd-FITC-SLNs were synthesized by monostearin loading of Gd-DTPA and FITC in the solvent diffusion method.We first evaluated the particle size, surface charge,Gd-DTPA entrapment efficiency and the stability of these SLNs.Then,colorectal tumors were evaluated in vitro via direct uptake of this contrast agent by human colon carcinoma, cells (HT29) and mouse models with colon carcinomas (CT26),aiming toexplore the absorption,cytotoxicity and MR imaging mechanism ofGd-FITC-SLNs. Continuely, we estimated the in vivo toxicity of Gd-FITC-SLNs using the MTD assay in C57/BL mice. Forty C57/BL mice were randomized into two groups, one receiving a single enema of Gd-FITC-SLNs (10 g/kg) and one receiving the same volume of water (negative control) after overnight fasting,then the mice were observed for 24 h after and assessed for toxic reactions or death responses.During in vivo MRI, eight female C57/BL mice with azoxymethane(AOM)/dextran sulfate sodium(DSS)-induced colorectal high-grade intraepithelial neoplasia were randomized into two groups, one receiving a transrectal enema of Gd-FITC-SLNs (40 mg/mL, n=4) and one receiving a transrectal enema of Gd-DTPA (13.92 mg/ml, n=4).3.0T MRI of colorectal tomors were achieved by using T1-weighted SE sequenc, TR/TE,550/15 ms and different TI (500,650,800,950 ms).MR colonography was performed before and 20,40,60, and 80 min after the liquid intracolonic enema. MRI findings were confirmed by subsequent histology correlation.ResultsThe average particle size of Gd-FITC-SLNs was 220 nm. The mean Gd-DTPA loading rate of Gd-FITC-SLNs was 29.76 ± 4.48%(w/w), and the encapsulation efficiencies of Gd-DTPA-SLNs was 55.00%. For in vitro confirmation experiments in evaluating-cell uptake of the Gd-FITC-SLNs, we explore contrast agent absorption by the two types of cancer cells (HT29&CT26) in the safe concentration range of 0-200 μg/mL and 0-600 ug/mL(P> 0.05), and demonstrated that contrast agent absorption by the two types of cells was concentration-dependent in the safe concentration range(P< 0.001). Meanwhile, in vivo toxicity of Gd-FITC-SLNs showed that there was no obvious death response or toxic reaction after the administration of a single enema. In vivo MRI with transrectal infusion of Gd-FITC-SLNs showed more significant enhancement at the tumor site compared with the infusion of Gd-DTPA in female C57/BL mice of colorectal high-grade intraepithelial neoplasia(P< 0.001). The MRI manifestations were correlated well by histology, and subsequent confocal fluorescence microscopy demonstrated Gd-FITC-SLNs as highly concentrated green fluorescent spots, which were distributed from the tumor capsule into the tumor.Conclusions1. Characteristics of intestine-absorbable Gd-FITC-SLNs include high loading rate of Gd-DTPA and detection by MR and histology.2. The novel MR Colonography with SLN contrast agent is feasible and it is helpful in the tumor display.3. The nanoparticle contrast has no significant cytotoxicity both in vivo and in vitro experiments at the safe concentration range which is expected to achieve clinical transformation.4. This study establishes the "proofs-of-principle" of a new MRI technique which enhances colorectal tumors based on absorption of nanoparticle contrast agents by colorectal wall. Solid lipid nanoparticle-based MRI may open new avenues for the early diagnosis of colorectal tumors.