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De-structuring of CuS nanomaterials by living cells maintains their therapeutic functions

In theranostics, copper sulfide (CuS) nanomaterials are rising for their high level of therapeutic efficiency. However, they can undergo massive bioprocessing upon internalisation into cells. Using X-ray absorption spectroscopy, this work shows that human cells extensively bioprocess them into CuS materials re-shaped by and within the cell with excellent photothermal potential.

The current development of nanomaterials for the biomedical field aims to provide therapeutic solutions that can overcome the limitations of existing treatments. Because the final target of these materials is generally malignant cells, understanding their mechanism of action and biostability in the demanding cellular environment is vital [1]. Multifunctional copper sulfide (CuS) nano-objects are particularly promising for the biomedical field, with optical properties allowing for a dual photothermal and photodynamic therapeutic function [2], yet few studies have explored the bio-stability of CuS structures. In this work, a spheroid-like model tissue based on human stem cells was used to monitor the intracellular long-term fate of CuS (Figure 83a). Hollow CuS nanoassemblies or rattle-like magnetic@

CuS nanohybrids featuring an iron oxide core were produced and their cellular uptake, short-term toxicity and longer-term impact on stem cell differentiation were investigated.

Electron microscopy observations highlighted a profound de-structuring of the nanomaterials (Figure 83b), which were massively disassembled inside the endosomes of the cells, with no more intact structures observed after three days of cellular processing. Nevertheless, no loss of photothermal conversion was detected over the whole month-period (Figure 83c). Conversely, an almost total loss of magnetism was revealed in the case of the iron oxide @CuS nanohybrids, showing that a nanoparticle can be significantly bioprocessed and transformed to different shapes and sizes while maintaining its therapeutic (here photothermal) function.

X-ray absorption spectroscopy (XAS) measurements at the Fe and Cu K-edge energies (Figure 84) were carried out at the Spanish CRG beamline BM25, confirming strong modification of the iron chemical environment, but no significant change in the copper one. XAS was performed at the spheroid level in the X-ray absorption near-edge structure (XANES) regime. At the Fe K-edge, comparing their spectra with iron oxide references showed that the initial nanomaterials in cells were a maghemite structure. However, after 21 days of tissue maturation, although

Fig. 83: a) Millimetric spheroids made from human stem cells loaded with CuS-based nanomaterials. The spheroids round up along the 21-day culture

period. b) TEM image at day 1 (left) shows still- intact CuS nano-assemblies inside endosomes. In

TEM image at day 21 of spheroid maturation (right), no original nano-assemblies are visible, while the

endosome inner membrane appears decorated with nanomaterial deposits, re-shaped in smaller

structures. c) Laser-induced heating of the CuS-based nanomaterials in spheroids over time: typical infra-

red images of different Cu cellular concentrations (left), at day 1 and 21 of spheroid maturation, after 5 min of 1064 nm laser irradiation at 0.3 W cm-2.

Average time-plots of the temperature increase for all incorporated doses (right).