Treatment of cancer micrometastasis using a multicomponent chain-like nanoparticle

Abstract

While potent cytotoxic agents are available to oncologists, the clinical utility of these agents is limited due to their non-specific distribution in the body and toxicity to normal tissues leading to use of suboptimal doses for eradication of metastatic disease. Furthermore, treatment of micrometastases is impeded by several biobarriers, including their small size and high dispersion to organs, making them nearly inaccessible to drugs. To circumvent these limitations in treating metastatic disease, we developed a multicomponent, flexible chain-like nanoparticle (termed nanochain) that possesses a unique ability to gain access to and be deposited at micrometastatic sites. Moreover, coupling nanochain particles to radiofrequency (RF)-triggered cargo delivery facilitated widespread delivery of drug into hard-to-reach cancer cells. Collectively, these features synergistically facilitate effective treatment and ultimately eradication of micrometastatic disease using a low dose of a cytotoxic drug.

Keywords: Cancer metastasis; Chain-like nanoparticle; Nanochains; Radiofrequency-triggered drug release; Targeting.

Fig. 1

Illustration of the nChain particle and its therapeutic effect on micrometastasis. (a) Schematic of a linear nChain particle composed of three IO nanospheres and one drug-loaded liposome. (b) TEM image of nChain particles. (c) Illustration of the successful delivery of nanochain-based drug to metastasis via vascular targeting and RF-triggered drug release.

Fig. 2

Reaction scheme of the controlled assembly of multicomponent nChain particles using solid-phase chemistry. (a) In the first step, chemical bifunctionality on the surface of parent IO nanospheres is topologically controlled resulting in nanospheres with two faces, one displaying only amines and the other only thiols. (b) In the second step, the two unique faces on the parent nanosphere serve as fittings to chemically assemble them into nanochains.

Fig. 3

Evaluation of the ability of nChain particles to target metastasis in vivo. (a) The timeline of surgery and BLI imaging are shown with respect to implantation of 4T1 cancer cells into the mammary fat pad of female BALB/c mice. Representative BLI images of an untreated animal indicate the progression of metastatic disease. (b) Representative FMT images of the same mouse show the accumulation of nChain particles in lung metastasis at 2 h post-injection. The nChain particles were injected 11 days after surgical removal of primary tumor (25 days after tumor inoculation), which was the time point of early onset of lung metastasis. (c) Using an NIR fluorophore as a label, the time-course of nanoparticle accumulation in lung metastases was obtained by quantification of the fluorescence signal in the FMT images of mice injected with nChain, integrin-targeting liposome (Lip) and non-targeting liposome (NT-Lip) at a DOX dose of 0.5 mg/kg b.w. (n=4 mice in each group; * P<0.02 by Student’s t-test). The lungs of the animals injected with nChain were excised and digested 5 h after administration and the iron concentration was measured using ICP-OES (data point indicated as ICP in the graph). Control animals were used to correct for background levels of endogenous iron. (d) Using a CRi Maestro fluorescence imaging system, ex vivo imaging of lungs 5 h after injection indicated the colocalization of nChain particles and 4T1 metastatic cells expressing GFP.

Fig. 4

Treatment of breast cancer metastasis using the 4T1 mammary model in mice. (a) The timeline of surgery and schedule of treatments are shown with respect to tumor inoculation. (b) The response of cancer metastasis to treatment was monitored using longitudinal BLI imaging. Representative images are shown for animals treated with DOX/RF, Lip/RF, nChain and nChain/RF. In the case of treatments combined with the RF field, 45 min post-injection, animals were exposed for 60 min to an RF field (amplitude B=6.3 mT, frequency f=10 kHz) using a custom-made solenoid coil. All nanoparticle formulations were administered at 0.5 mg/kg DOX, while free DOX was injected at 5 mg/kg. (c) Quantification of the whole body BLI light emission is shown for the nChain/RF treatment and control treatments including nChain, DOX/RF and Lip/RF. The inset shows the same plot with the y-axis being in logarithmic scale (n=7 mice in each group; * P<0.03 by Student’s t-test). (d) The survival time of the animals treated with nChain/RF, nChain, DOX/RF and Lip/RF is compared to that of the untreated group.

Fig. 5

Histological evaluation of the anticancer effect of the nChain particle on micrometastasis in the liver. (a) Fluorescence imaging of an entire histological section of a lobe of liver 30 min after systemic administration of nChain at 0.5 mg DOX/kg b.w. Nuclei were stained with DAPI (5x magnification). Images of entire histological sections of the organ were obtained using the automated tiling function of the microscope. At 30 min post-injection, the location of metastatic cancer cells is shown with respect to the location of endothelial cells (b), nChain particles and DOX (c) in the same histological section (10x magnification; insets: 20x magnification). (d) At 120 min post-injection, fluorescence imaging of an entire histological section shows the widespread distribution of DOX molecules after a 60-min application of RF employed at 45 min post-injection (5x magnification). At 120 min post-injection, higher magnification imaging shows the distribution of DOX molecules with (e) or without RF (f) with respect to the location of cancer cells and nChain particles (10x magnification; inset: 20x magnification). At 48 h post-injection, fluorescence imaging of entire histological sections shows the distribution of DOX molecules with respect to cancer cells for the nChain/RF (g), nChain (h) and Lip/RF treatment (i). Insets show apoptotic cells in a small portion of the same images. Apoptotic cell nuclei were stained with TUNEL.