Diketopyrrolopyrrole-based semiconducting polymer nanoparticles for in vivo second near-infrared window imaging and image-guided tumor surgery

Abstract

A diketopyrrolopyrrole-based semiconducting polymer nanoparticle (PDFT1032) has been developed as a NIR-II (near infrared window II, 1000-1700 nm) fluorescent probe. It shows high photostability, a favorable absorption peak at 809 nm, a large Stokes shift of 223 nm, outstanding biocompatibility and minimal in vivo toxicity. More importantly, the versatile use of PDFT1032 for several important biomedical applications in the NIR-II window has been demonstrated, including the NIR-II optical imaging of tumors on a subcutaneous osteosarcoma model, assessing the vascular embolization therapy of tumors, and NIR-II image-guided orthotopic tumor surgery and sentinel lymph node biopsy (SLNB) with high spatial and temporal resolution. Overall, excellent biocompatibility, favorable hydrophilicity, and desirable chemical and optical properties make the semiconducting polymer nanoparticle PDFT1032 a highly promising NIR-II imaging probe with the potential to be widely applicable in clinical imaging and the surgical treatment of malignancy.

Fig. 1. Synthetic route to PDFT. Detailed synthesis procedures can be found in the ESI.

Fig. 2. Characterization of PDFT1032. (a) Schematic drawing of a PDFT1032 nanoparticle composed of semiconducting polymer DFT and a hydrophilic DSPE-mPEG shell. (b) Absorbance and fluorescence spectrum of PDFT1032 showing an absorption peak at 809 nm and a fluorescence peak at 1032 nm with an 808 nm excitation laser. (c) A DLS spectrum and TEM image (inset) of PDFT1032, showing a hydrodynamic size of 68 nm. (d) NIR-II signals of PDFT1032 with sequential long-pass filters (1000–1200 nm), indicating that the fluorescence signals of PDFT1032 were distinct at the 1000 and 1050 nm filters, and even above 1200 nm. A photostability test of PDFT1032 using NIR-II fluorescence imaging (e) and quantified fluorescence intensity (f) for PDFT1032 in PBS, FBS and DMEM (cell culture media) with 808 nm excitation laser radiation for up to 2 h. The photobleaching of PDFT1032 in PBS, FBS and DMEM culture media was almost negligible.

Fig. 3. NIR-II imaging and image-guided surgery on mice with osteosarcoma using PDFT1032. (a) The images from both the lateral and prone position in mice (n = 3) demonstrated clear visualization of the subcutaneous tumor up to 7 days. (b) High tumor-to-background ratio of the lateral position (TBR, up to 3.5 ± 0.39) and the prone position (up to 2.6 ± 0.16) due to the excellent EPR effect of PDFT1032. (c) Bright field photograph of a nude mouse with an orthotopic osteosarcoma. The whole tumor was visualized using NIR-II imaging with excellent contrast ((d) and (e)). The white dashed circle contours the location of the tumor. (f) The tumor body was resected. The NIR-II signal still remained around the knee joint, indicating that there were adjacent tumor metastases located at the metaphysis of the tibia. The white arrow indicates the residual lesion. (g) The micro-metastasis of the tumor is identified using NIR-II imaging, and (h) the bright field photograph of the surgical scope. (i) The micro-metastasis was then completely resected using NIR-II imaging, which was further confirmed by the histological analysis (j). Finally, the lymph node was also visualized and resected with the help of PDFT1032 (k) and (l). Scale bar: 1.5 cm.

Fig. 4. NIR-II imaging for the assessment of the vascular embolization therapy of osteosarcoma with PDFT1032. (a) The vascular mapping and the hemodynamic status of the tumor and the femoral artery were determined. The white dashed circle contours the location of the tumor. ((b) and (c)) The branch of the femoral artery that supports the tumor and the vascular network of the tumor (exhibited as a claw shape) were clearly identified. (d) A vessel clamp was used to block the blood flow (red arrow) and the signal of the vascular network vanished. (e) After 5 minutes, the clamp was removed and the blood flow of the tumor was still devoid because a temporary thrombus was formed (blue arrowhead). (f) Magnification of (c). The vascular network of the tumor was clearly identified (white arrowheads). (g) Magnification of (d). (h) The major artery was surgically incised (blue arrowhead). (i) NIR-II imaging exhibited the absence of the residual tumor fluorescence and normal circulation (femoral artery) was successfully maintained. Inset is the histological analysis of the osteosarcoma. Scale bar: 8 mm.

Fig. 5. NIR-II image-guided sentinel lymph node biopsy on one of the mice with melanoma. (a) PDFT1032 (green arrowhead) was injected at the margin of the B16F10 melanoma (white dashed circle). (b) Ten minutes later, the two axillary lymph nodes were clearly identified (white arrowhead). (c)–(e) The efferent lymphatic vessel connecting the tumor with the node (blue arrowhead) was clearly identified. The SLN and the secondary lymph node were distinctly identified. (f) The SLN was resected (inserted image). After a positive result for the SLN was confirmed ((g) and (j)), the secondary lymph node was subsequently resected (h). Finally, histological analysis also confirmed the metastasis of melanomas ((i) and (l)). (k) The histological result of a resected melanoma, colored black. For the fluorescence images, scale bar: 8 mm, for the histological analysis, scale bar: 75 μm.