Albumin-based nanosystem for dual-modality imaging-guided chem-phototherapy against immune-cold triple-negative breast cancer

Abstract

Triple-negative breast cancer is a highly aggressive and metastatic tumor; diagnosing it in the early stages is still difficult, and the prognosis for conventional radio-chemotherapy and immunotreatment is not promising due to cancer's immunosuppressive microenvironment. The utilization of protein-based nanosystem has proven to be effective in delivering agents with limited adverse effects, yet the combination of diagnosis and treatment remains a difficult challenge. This research took advantage of natural albumin and organic molecules to construct a self-assemble core-shell nanostructure combining with superparamagnetic iron oxide nanocrystals and heptamethine cyanine dye IR780 through non-covalent interactions. This nanocomposite successfully decreased the transverse relaxation time of the magnetic resonance hydrogen nucleus, resulting in outstanding T₂ imaging, as well as emitting near-infrared II fluorescence, thereby the resulting dual-modality imaging tool was applied to improve diagnostic competency. It is noteworthy that the nanocomposites exhibited impressive enzyme-like catalytic and photothermal capabilities, resulting in a successful activation of the immune system to efficiently suppress distant metastatic lesions in vivo. Consequently, this nano-drug-based therapy could be an advantageous asset in reinforcing the immune system and hindering the growth and reappearance of the immune-cold breast cancer.

Keywords: albumin; fluorescence near-infrared II imaging; magnetic resonance imaging; nanozyme; phototherapy.

Graphical abstract

Figure 1.

(A) Synthesis diagram of IR780@BSA@SPIO nanosystem. (B) DLS data and TEM characterization of IR780@BSA@SPIO nanosystem. (C) T₂WI image and the relationship between the relaxation value and concentration of IR780@BSA@SPIO nanosystem. (D) NIR-II fluorescence imaging of IR780@BSA@SPIO and IR780@BSA two systems, and the relationship between the signal intensity and IR780’s concentration. The image on the top shows the fluorescence image of IR780@BSA while the one at the bottom displays the fluorescence image of IR780@BSA@SPIO. (E) Heating absorption and dissipation rocking result of nanosystem under 808 nm laser irradiation at [IR780] 1.5 μg/mL.

Figure 2.

(A) Fluorescent image of 4T1 cells incubated with nanomedicines for different durations. Scale bar =100 μm. (B) Prussian blue staining of the nanosystem treated cells. Scale bar =500 μm. (C) MRI-T₂ imaging of these labeled cells. And the relationship between the T₂-relaxation time and the cell counts. (D) The fluorescence imaging of these labeled cells. And the relation curve between signal intensity and cells’ number. The ‘up’ and ‘down’ image refer to the fluorescence image of IR780@BSA and IR780@BSA@SPIO, respectively. n = 3, ***P < 0.001.

Figure 3.

(A) AM/PI staining of different materials treated 4T1 cells after laser irradiation. (B) The cell viability of 4T1 cells after nanosystem-based phototherapy. The data are expressed as mean ± standard deviation (n = 3). Scale bar =800 μm. **P < 0.01, ***P < 0.001.

Figure 4.

(A) In vivo MR imaging of tumor-bearing mice after injection of IR780@BSA@SPIO nanosystem. Color according to the local gray value. (B) NIR-II fluorescence imaging of tumor-bearing mice administrated with various drugs and isolated organs and tumors after 72 h (H: heart; Li: liver; S: spleen; L: lung; K: kidney; T: tumor). Semi-quantitative analysis of fluorescence intensity from tumor-bearing mice (C) and isolated organs and tumors (D). The data are expressed as mean ± standard deviation (n = 3). Signal intensity differences between BSA-based nanosystems and free IR780 group were analyzed, *P < 0.05.

Figure 5.

(A) Schematic illustration of the in vivo experiments. (B) Primary/distant tumor volume curve of tumor-bearing mice in different treatment groups. The data are expressed as mean ± standard deviation (n = 5). (C) Images of isolated primary and distant tumor from different treatment groups. (D) Foxp3 staining and semi-quantitative analysis of primary and distant tumor sections from different treatment groups. The data are expressed as mean ± standard deviation (n = 3). Scale bar =800 μm. The variation in tumor size between the IR780@BSA@SPIO nanosystem exposed to laser irradiation and the other groups was evaluated (**P < 0.01, ***P < 0.001).

Figure 6.

Immunofluorescence imaging of primary tumor sections in different treatment groups. Scale bar =800 μm.

Figure 7.

(A) Body weight curves of tumor-bearing mice in different treatment groups. The data are expressed as mean ± standard deviation (n = 5). (B) Biochemical analysis of blood components: ALT, AST, BUN, Cre and TBIL. The data are expressed as mean ± standard deviation (n = 3). (C) Hematoxylin and eosin staining of main organs (H: heart; Li: liver; S: spleen; L: lung; K: kidney), primary tumor and distant tumor sections of tumor-bearing mice after treatment. Scale bar =800 μm.