1. CIC biomaGUNE and CIBER-BBN, Paseo de Miramón 182, 20014 Donostia-San Sebastián, Spain
2. Nanomedicine Lab, Faculty of Biology, Medicine & Health and National Graphene Institute, The University of Manchester, AV Hill Building, Manchester M13 9PT, UK
3. Ikerbasque, Basque Foundation for Science, 48013 Bilbao, Spain
*Current address: M.Q.: Materials Physics Department, Universidad Autónoma de Madrid. Avda. Francisco Tomás y Valiente, 7. 28049, Madrid, Spain. I.d L.: 1. Wyss Institute for Biologically Inspired Engineering at Harvard University, Cambridge, MA 02138, USA, 2. John A. Paulson School of Engineering and Applied Sciences, Harvard University, Cambridge, MA, USA
The control of temperature during photothermal therapy is key to preventing unwanted damage in surrounding tissue or post-treatment inflammatory responses. Lack of accurate thermal control is indeed one of the main limitations that hyperthermia techniques present to allow their translation into therapeutic applications. We developed a nanoprobe that allows controlled local heating, combined with in situ nanothermometry. The design of the probe follows a practical rationale that aims at simplifying experimental requirements and exploits exclusively optical wavelengths matching the first and second biological windows in the near-infrared.
Methods: Hybrid nanostructures were chemically synthesized, and combine gold nanostars (photothermal agents) with CaF2:Nd3+,Y3+ nanoparticles (luminescent nanothermometers). Both components were simultaneously excited in the near-infrared range, at 808 nm. Following the goal of simplifying the thermal monitoring technique, the luminescent signal was recorded with a portable near-infrared detector. The performance of the probes was tested in 3D tumor spheroids from a human glioblastoma (U87MG) cell line. The location of the beads within the spheroids was determined measuring Nd3+ emission in a commercial Lightsheet microscope, modified in-house to be able to select the required near-infrared wavelengths. The temperature achieved inside the tumor spheroids was deduced from the luminescence of Nd3+, following a protocol that we developed to provide reliable thermal readings.
Results: The choice of materials was shown to work as an optically excited hybrid probe. Depending on the illumination parameters, temperature can be controlled in a range between 37 ºC and 100 ºC. The near-infrared emission of nanothermometers also allows microscopic tracking of the hybrid nanostructures, confirming that the probes can penetrate deeper into the spheroid mass. We observed that, application of optical thermometry in biological environments requires often neglected considerations, since the optical signal changes along the optical path. Accordingly, we developed data analysis protocols that guarantee reliable thermal readings.
Conclusions: The prepared hybrid probes are internalized in 3D tumor spheroids and can be used to induce cell death through photothermal effects, while simultaneously measuring the local temperature in situ. We show that luminescent thermometry in biomedical applications requires the development of protocols that guarantee accurate readings. Regarding photothermal treatments, we observe a sharp thermal threshold at around 55 ºC (for 10 min treatments) that separates high survival ratio from complete cell death.
Keywords: photothermal therapy, nanothermometry, plasmonic heating, brain cancer, luminescence sensing, near-infrared