Semiconducting polymers enhance Cerenkov radiation energy transfer for multimodal cancer theranostics

71 Objectives: The therapeutic efficacy of photodynamic therapy is limited by the ability of light to penetrate tissues. Due to this limitation, Cerenkov radiation (CR) from radionuclides has recently been proposed as an alternative light source. The transfer of CR for the activation of nanophotosensitizers, referred to as CRET, is limited by a relatively weak intensity that peaks in the ultraviolet-blue end of the visible light spectrum. Semiconducting polymer nanoparticles (SPNs) have ideal optical properties such as large absorption cross-sections and broad absorbance that can be designed to be in the blue end of the visible light spectrum. SPNs can be doped with photosensitizers and have nearly 100% efficient energy transfer due to multiple energy transfer mechanisms. Here, we aim to prepare SPNs doped with a porphyrin photosensitizer (TPP) as a nanosystem to harness and amplify CR for a cancer theranostics nanosystem. Methods: In this design, CR from the radionuclide (Zr-89; t1/2 = 78.4 h) will be transferred to the SPNs through CRET then to TPP through a combination of Forster resonance energy transfer (FRET) and exciton diffusion. The semiconducting polymers (SPs) poly[(9,9-dihexylfluorenyl-2,7-diyl)-alt-co-(2-methoxy-5-{2-1,4-phenylene)] (MEP) [(9,9-dioctylfluorenyl-2,7-diyl)-alt-(9-hexyl-3,6-carbazole)] (HC), and poly[2,5-dioctyl-1,4-phenylene] (PDP) were used to determine an optimal SPNs nanosystem. SPNs were prepared with the SPs (MEP, HC, PDP), TPP at various doping percentages, an amphiphilic polymer (PEG-b-PPG-b-PEG), and poly(styrene-co-maleic anhydride) using a nanoprecipitation method. The morphological properties of each SPNs nanosystem were measured using TEM and DLS. The optimal TPP doping percentage for the SPNs was determined using fluorescence spectroscopy. The ideal nanosystem was determined by comparing fluorescence intensity following radionuclide activation of the optimized SPNs using IVIS. The optimal SPNs nanosystem (MEP5; MEP SP, 5% TPP doping percentage) was also compared to a nanoparticle prepared using the same method with only TPP (TPP NPs). Deferoxamine was conjugated to MEP5 for longitudinal PET imaging studies in a 4T1 xenograft model. Additionally, the in vivo biodistribution of MEP5 was evaluated using optical imaging based on the fluorescence of TPP (ex: 640 nm, em: 720 nm) and CRET to TPP (ex: closed, ex: 660 nm). Results: SPNs were prepared by doping with TPP at 0-20% relative to the SP concentration during nanoprecipitation and confirmed with UV-VIS spectroscopy. The diameters of the SPNs ranged from 40-60 nm. FL spectroscopy determined the optimized SPNs to be PDP5, MEP5, and HC10, where the number indicates TPP doping percentage. The optimal SPNs formulation was determined to be MEP5 when compared at equivalent TPP concentration and incubated with 100 μCi of Zr-89. Additionally, CRET efficiency for MEP5 was dramatically amplified compared to TPP NPs at equivalent concentrations and remained over a 72 h period. Following intravenous administration, 4T1 tumors could be clearly delineated with PET imaging using 89Zr-Df-MEP5 with ex vivo uptake determined to be 5.13±0.83 %ID/g. The 4T1 tumors were further visualized with optical imaging. Conclusion: SPNs doped with photosensitizers were found to dramatically amplify CRET. Bimodal PET and optical imaging studies clearly visualized tumor uptake of optimized SPNs. This nanosystem has excellent potential as a cancer theranostics nanosystem unabated by tissue penetration limits.