Introduction

Silicon particles, on the nanometer to micron size scale, are potentially well-suited to function as targeted molecular imaging agents, due to their biocompatibility, biodegradability, and simple surface chemistry that is amenable to the addition of targeting moieties and therapeutic drugs [1,2]. Hyperpolarization (HP) of these particles through solid-state Dynamic Nuclear Polarization (DNP) [3] increases magnetic resonance imaging (MRI) signals by 4-5 orders of magnitude through temporary alterations to the nuclear spin distribution. This process is aided by endogenous electronic defects that are present on the particle surface, thus precluding the need for additional exogenous radical species [4]. The resulting enhanced ²⁹Si MR signal lasts for significantly longer than other hyperpolarized agents in vivo (tens of minutes, compared to <1 minute for many other HP species) because the spin polarization is relatively protected within the core of the particle [5]. Currently, we report our efforts to develop hyperpolarized silicon microparticles for targeted molecular imaging of early stage colorectal cancer (CRC). In the United States, colorectal cancer is the second leading cause of cancer-related deaths amongst men and women (combined) [6], despite the availability of active preventative screening measures, such as the traditional colonoscopy. As such, there is a clinical need to develop advanced imaging methods for the early detection, observation of treatment efficacy, and monitoring of disease recurrence concerning colorectal cancer. In this work, we show that the hyperpolarization process is not affected by the addition of an antibody to the silicon particle surface, and the enhanced ²⁹Si MRI signal lasts for tens of minutes after injection into a CRC mouse model.

Methods

Commercially available silicon particles (average mean diameter ~2 μm) were investigated. The particles were surface functionalized with 3-aminopropyltriethoxysilane (APTES), polyethylene glycol (PEG8), and 214D4 antibody that targets the glycosylated ectodomain of human MUC1—a transmembrane Mucin protein that is overexpressed in some forms of CRC. The particles were hyperpolarized using a laboratory-constructed ²⁹Si solid-state DNP device [7], which is located adjacent to a 7 T small animal MRI system. Initial ²⁹Si MR signals and room temperature T₁ decays were recorded spectroscopically (α = 10°) at 7 T for particles both with and without surface functionalization. Separately, functionalized particles were injected (intratumoral) into nude mice harboring a subcutaneous, human MUC1-expressing colorectal tumor (HT29-MTX-E12). Co-registered [¹H:²⁹Si] MR imaging was performed using a dual-tuned ²⁹Si/¹H Litz coil: in vivo ²⁹Si imaging was performed with a coronal RARE sequence (α = 90°; TR/TE: 60 ms/1.8 ms; 6.4 cm FOV; 2 mm resolution), while ¹H imaging is performed with a coronal RARE scan (α = 90°), TR/TE: 1927 ms/9.5 ms with a RARE factor of 8; 6.4 cm FOV (0.25 mm resolution) and 4 averages.

Results

PEGylated silicon microparticles (no antibody) highlighted the colon morphology from the rectum to the cecum of a normal mouse 5 minutes after rectal administration, demonstrating the proof-of-concept for this technique (Fig. 1). Comparing bare (no surface treatments) vs. antibody-functionalized particles resulted in similar initial ²⁹Si MRS signal and hyperpolarized decay constant (T₁ ~19 minutes) for both samples (Fig. 2). Particles functionalized with the 214D4 antibody were hyperpolarized and then injected directly into a subcutaneous CRC tumor, and provided ²⁹Si signal within the tumor 20 minutes after injection (Fig. 3).

Discussion

Demonstration that the conjugation of targeting antibodies to the silicon particle surface does not affect the hyperpolarization dynamics is a critical step in developing silicon particles as a diagnostic platform for cancer medicine, and the ability to image the particles for tens of minutes following injection demonstrates an enhanced in vivo imaging window compared to other hyperpolarized contrast agents. MUC1 is an attractive target, as it is a transmembrane glycoprotein that is expressed on the cancer cell epithelium—thus the particles do not need to be internalized by the cells for effective targeting. While these are positive strides forward, the large microparticles also proved to have limited mobility, thus necessitating the administrative method of intratumoral injection. Future studies will look to translate these advances to nano-scale particles, which are expected to exhibit improved solubility and mobility—allowing for true molecular targeting. Preliminary in vitro cell-binding studies indicate that exposure to the harsh conditions of DNP do not negatively affect the targeting ability of the antibody following hyperpolarization.

Conclusion

We present our most recent work developing hyperpolarized silicon microparticles for targeted molecular imaging of colorectal cancer. When fully developed, these particles are engineered to be a platform system, where different targeting agents and therapeutic drugs can be attached for advanced molecular imaging and therapeutic interventions in the clinic.

Acknowledgements

This work was funded by the MDACC Odyssey Program, NCI R25T CA057730, DoD PC131680, NCI R25E CA056452, CPRIT RP150701, MDACC Colorectal Cancer Moonshot, MDACC Institutional Research Grants, MDACC Institutional Startup, U54 CA151668, Leukemia and Brain SPORE Developmental Research Awards, NCI R21 CA185536, Gulf Coast Consortium, and NCI Cancer Center Support Grant CA016672.

References

[1] Park, et al. Nat. Mat. (2009); [2] Tasciotti, et al. Nat. Nano. (2008); [3] Cassidy, et al. Nat. Nano. (2013); [4] Dementyev, et al. Phys. Rev. Lett. (2008); [5] Aptekar, et al. ACS Nano (2009); [6] SEER Cancer Statistics Review (2016); [7] Whiting, et al. J. Med. Imag. (2016).