(F-220) Stimuli-responsive Magnetic-plasmonic core-shell nanosensors for trimodal US/PA/MR spatial and functional tracking of cell therapy

Introduction:: Stem cells (SCs) have shown significant potential in the restoration of degenerative, damaged, or aging tissues through remodeling and repair. Promising preclinical outcomes of SC therapy (SCT) have been reported for many degenerative diseases, yet many limitations hinder the clinical translation of SCT. A major safety roadblock for clinical SCT is the lack of techniques to monitor transplanted cell status in vivo – such as location, viability, differentiation, etc. Current methods to determine the fate of transplanted cells are invasive end-point analyses, such as biopsies and histological staining, which cause damage to the regenerating tissue and do not reflect the real-time dynamics of the microenvironment. To improve the safety and efficacy of SCT, there is a critical need to track transplanted SCs non-invasively and monitor their fate in vivo.

While magnetic resonance (MR) cell tracking has been widely employed in clinical SC trials for spatial monitoring, it does not provide functional information about cell status or allow intraprocedural guidance. Alternatively, ultrasound-guided photoacoustic (US/PA) imaging is capable of the molecular imaging of cell status non-invasively but suffers from deep tissue imaging limitations. However, when combined, US/PA/MR imaging can perform molecular imaging of cell status along with deep tissue imaging. In this project, iron-gold core-shell nanoparticles (IGNPs) acting as trimodal US/PA/MR contrast agents were synthesized. IGNPs modified with stimuli-responsive triblock peptides (termed SRIGs) will hydrophobically aggregate when exposed to a target biomarker, resulting in PA signal changes due to interparticle plasmon coupling and MR contrast changes due to magnetic dipole-dipole interactions.


Materials and Methods:: To create uniform (200 nm) IGNPs, silica-coated iron oxide nanospheres were synthesized and decorated with gold seeds on the surface followed by subsequent gold shell growth in the presence of poly(vinylpyrrolidone) (A). The surface of IGNPs were modified with a triblock peptide consisting of a cell penetrating sequence, an enzyme-cleavable sequence, and a hydrophobic aggregation-inducing sequence designed to cause hydrophobic aggregation in the presence of the target intracellular biomarker of interest. Successful synthesis, morphology, and aggregation of stimuli-responsive IGNPs (SRIGs) was confirmed by transmission electron microscopy (TEM) (B). For MRI, SRIGs were imaged in a tissue-mimicking gelatin phantom and images were acquired using a multi-slice multi-echo pulse sequence (7T Bruker PharmaScan; 38 mm coil). The relaxation time (TR) was held constant at 2000 ms. The echo time (TE) started at 7 ms and was ramped up in 7 ms increments for 32 time steps. The average T2 relaxation time was calculated using the built-in software and parametric fit algorithm. Next, aggregation-responsive PA signal change was tested in a tube phantom using naïve and aggregated SRIG solutions (20 MHz, 800 nm, Vevo/LAZR, VisualSonics Inc.). Samples were 3D imaged over 10.21 mm at 0.152 mm intervals and reconstructed in the coronal view using maximum intensity projections (VevoLAB 5.7.0).


Results, Conclusions, and Discussions:: SRIGs generated high negative contrast under T2-weighted MRI. The T2-weighted relaxation time and the MR signal intensity both showed differences in unaggregated vs. aggregated SRIGs (C). The T2 relaxation upon aggregation showed similar trends to unaggregated SRIGs, but the average relaxation time decreased, which reflects higher negative contrast (D-E). US/PA imaging of the SRIG solutions showed strong PA contrast, and a clear signal difference upon aggregation, which was quantified to show a ~45% change in PA signal at 800 nm (F-G). These results demonstrate the potential of SRIG as a trimodal contrast agent and sensor. Future work will involve modification of IGNPs with different peptide sequences to target particular biomarkers of interest, use the designed SRIGs to label cells, study cell-nanosensor interactions, and perform in vivo cell monitoring through combined US/PA/MR imaging.

In summary, our pilot studies showcased a US/PA/MRI contrast agent functionalized with a triblock peptide as a nanosensor blueprint that can detect a variety of target biomarkers by exploiting hydrophobic aggregation-driven plasmonic coupling and magnetic dipole-dipole signal changes. Using this nanosensor we can perform spatial and functional cell tracking through combined US/PA/MR imaging and exploit multimodal advantages, such as intraprocedural and molecular imaging through US/PA coupled with deep tissue pre- and post-procedural monitoring through MR imaging. Following synthesis, the nanosensor will be modified with (1) triblock peptides which are responsive to SC apoptosis using capsase-3 as a biomarker and (2) triblock peptides responsive to SC differentiation. The ability of our sensor to detect SC location and fate will be validated in relevant small animal models via combined US/PA/MR imaging. Our approach to perform in vivo US/PA/MR guided spatial and functional tracking of transplanted cell status through cell nanoengineering and non-invasive bioimaging has potential to optimize SC transplantation towards safer and improved SCT for degenerative diseases through real-time therapeutic planning.


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