On-command drug release from nanochains inhibits growth of breast tumors

Abstract

Purpose: To evaluate the ability of radiofrequency (RF)-triggered drug release from a multicomponent chain-shaped nanoparticle to inhibit the growth of an aggressive breast tumor.

Methods: A two-step solid phase chemistry was employed to synthesize doxorubicin-loaded nanochains, which were composed of three iron oxide nanospheres and one doxorubicin-loaded liposome assembled in a 100-nm-long linear nanochain. The nanochains were tested in the 4T1-LUC-GFP orthotopic mouse model, which is a highly aggressive breast cancer model. The 4T1-LUC-GFP cell line stably expresses firefly luciferase, which allowed the non-invasive in vivo imaging of tumor response to the treatment using bioluminescence imaging (BLI).

Results: Longitudinal BLI imaging showed that a single nanochain treatment followed by application of RF resulted in an at least 100-fold lower BLI signal compared to the groups treated with nanochains (without RF) or free doxorubicin followed by RF. A statistically significant increase in survival time of the nanochain-treated animals followed by RF (64.3 days) was observed when compared to the nanochain-treated group without RF (35.7 days), free doxorubicin-treated group followed by RF (38.5 days), and the untreated group (30.5 days; n=5 animals per group).

Conclusions: These studies showed that the combination of RF and nanochains has the potential to effectively treat highly aggressive cancers and prolong survival.

Figure 1

(a) Illustration of the required steps for the successful delivery of nanoparticle-based drug to tumors. (b) A cartoon shows a linear nanochain composed of three IO nanospheres and one liposome. The constituent nanospheres display two distinctive faces in terms of chemical functionality, which allows them to be assembled into a linear nanochain. (c) The illustration of the triggered release mechanism shows that defects on the liposome are caused by the ‘vibration’ of the IO tail of the nanochain particle under an RF field.

Figure 2

Synthetic scheme of the partial modification of the functional groups on a nanospheres’ surface (step 1) and the controlled assembly of linear nanochains (step 2).

Figure 3

Monitoring the progression of tumor growth and metastasis using BLI imaging. Luc-GFP-4T1 cells (0.5 ×10⁻⁶) were implanted into the mammary fat pad of female BALB/c mice. Animals were imaged every 2–3 days over the period of 6 weeks. Besides an untreated group, animals were treated with DOX-loaded nanochains or free DOX (followed by RF) at a dose of 0.5 mg/kg DOX at day 14 after tumor inoculation. Representative images of longitudinal imaging are shown for (a) an untreated animal, (c) DOX-treated animal followed by application of an RF field 18 hours after injection (e) nanochain-treated animal, (g) and nanochain-treated animal followed by application of an RF field 18 hours after injection. Quantification of BLI light emission from primary and metastatic tumors over the six-week period is shown for (b) the untreated group (inset: the same plot is shown with the y-axis being in a standard linear scale), (d) the DOX-treated group followed by RF, (f) the nanochain-treated group, (h) and the nanochain-treated group followed by RF. Y-axis is in logarithmic scale. The blue tick mark indicates that all the data points between the two curves are statistically different (P<0.005; n=5 animals per group).

Figure 4

Overall comparison of the BLI signal indicating the progression of (a) the primary and (b) metastatic spread of the untreated group (control), DOX-treated group (DOX with RF), the nanochain-treated group (NC-no RF), and the nanochain-treated group followed by RF (NC with RF). Y-axis is in logarithmic scale. Data points marked with asterisks are statistically different between the “NC with RF” group and the other groups. Data points marked with crosses are statistically different between the “NC(no RF)” group and the control group (* and † P<0.05; n=5–6 animals per group).

Figure 5

(a) Evaluation of distribution of metastases using ex vivo imaging of organs. Organs from one animal per condition were excised 30 days after tumor inoculation and imaged using a CRi Maestro fluorescence imaging system. Green indicates the presence of Luc-GFP-4T1 cells. (b) Fluorescence images of entire histological sections of liver and lung lobes from the untreated animal and the nanochain-treated animal followed by RF (5x magnification; blue: nuclear stain (DAPI); green: Luc-GFP-4T1 cells). Images of entire histological sections of the organs were obtained using the automated tiling function of the microscope.

Figure 6

The survival of the group treated with DOX-loaded nanochain followed by RF (n=5) was significantly prolonged when compared to the DOX-treated group followed by RF (n=6), the nanochain-treated group (n=5) and the untreated group (n=5).