A photoactivatable upconverting nanodevice boosts the lysosomal escape of PROTAC degraders for enhanced combination therapy.

PROteolysis TArgeting Chimeras have received increasing attention due to their capability to induce potent degradation of various disease-related proteins. However, the effective and controlled cytosolic delivery of current small-molecule PROTACs remains a challenge, primarily due to their intrinsic shortcomings, including unfavorable solubility, poor cell permeability, and limited spatiotemporal precision. Here, we develop a near-infrared light-controlled PROTAC delivery device (abbreviated as USDPR) that allows the efficient photoactivation of PROTAC function to achieve enhanced protein degradation. The nanodevice is constructed by encapsulating the commercial BRD4-targeting PROTACs (dBET6) in the hollow cavity of mesoporous silica-coated upconversion nanoparticles, followed by coating a Rose Bengal (RB) photosensitizer conjugated poly-L-lysine (PLL-RB). This composition enables NIR light-activatable generation of cytotoxic reactive oxygen species due to the energy transfer from the UCNPs to PLL-RB, which boosts the endo/lysosomal escape and subsequent cytosolic release of dBET6. We demonstrate that USDPR is capable of effectively degrading BRD4 in a NIR light-controlled manner. This in combination with NIR light-triggered photodynamic therapy enables an enhanced antitumor effect both in vitro and in vivo. This work thus presents a versatile strategy for controlled release of PROTACs and codelivery with photosensitizers using an NIR-responsive nanodevice, providing important insight into the design of effective PROTAC-based combination therapy.

Photoactivatable Aptamer-CRISPR Nanodevice Enables Precise Profiling of Interferon-Gamma Release in Humanized Mice.

Real-time dynamic imaging of immunoactivation-related cytokines is crucial for evaluating the efficacy of immune checkpoint blockade therapy and optimizing the treatment regimen. We introduce herein a spatiotemporally controlled nanodevice that allows in situ photoactivated imaging of interferon-gamma (IFN-γ) secretion from T cells in vitro and in vivo. The nanodevice is constructed by rational engineering of an aptamer-embedded, UV-cleavable PC-DNA probe and further integration with upconversion nanoparticles- and CRISPR-Cas12a-enhanced fluorescence systems. Using human peripheral blood mononuclear cells (PBMC)-engrafted mouse models, this nanodevice allows for the quantitative imaging of endogenous IFN-γ and its intratumoral dynamics responding to antiprogrammed cell death receptor 1 (anti-PD-1) therapy. This study thus provides a toolbox for boosting the sensitivity and precision of cytokine imaging during immune checkpoint blockade therapy, enlightening research toward imaging-guided tumor therapy.

An orthogonally activatable CRISPR-Cas13d nanoprodrug to reverse chemoresistance for enhanced chemo-photodynamic therapy.

Orthogonal therapy that combines CRISPR-based gene editing and prodrug-based chemotherapy is a promising approach to combat multidrug-resistant cancer. However, its potency to precisely regulate different therapeutic modalities in vivo is limited due to the lack of an integrated platform with high spatiotemporal resolution. Taking advantage of CRISPR technology, a Pt(iv)-based prodrug and orthogonal emissive upconversion nanoparticles (UCNPs), we herein rationally designed the first logic-gated CRISPR-Cas13d-based nanoprodrug for orthogonal photomodulation of gene editing and prodrug release for enhanced cancer therapy. The nanoprodrug (URL) was constructed by encapsulating a green light-activatable Pt(iv) prodrug and UV light-activatable Cas13d gene editing tool into UCNPs. We demonstrated that URL maintained excellent orthogonal emission behaviors under 808 and 980 nm excitations, allowing wavelength-selective photoactivation of Cas13d and the prodrug for downregulation of the resistance-related gene and induction of chemo-photodynamic therapy, respectively. Moreover, the photomodulation superiority of URL for overcoming drug resistance was highlighted by integrating it with a Boolean logic gate for programmable modulation of multiple cell behaviors. Importantly, in vivo studies demonstrated that URL can promote Pt(iv) prodrug activation and ROS generation and massively induce on-target drug accumulation by Cas13d-mediated drug resistance attenuation, delivering an ultimate chemo-photodynamic therapeutic performance in efficiently eradicating primary tumors and preventing further liver metastasis. Collectively, our results suggest that URL expands the Cas13d-based genome editing toolbox into prodrug nanomedicine and accelerates the discovery of new orthogonal therapeutic approaches.

Near-Infrared-Light-Mediated DNA-Logic Nanomachine for Bioorthogonal Cascade Imaging of Endogenous Interconnected MicroRNAs and Metal Ions.

The interconnection of microRNAs (miRNAs) and metal ions governs multiple biological processes in disease development and progression. However, developing multiplexed tools for dynamic imaging of these regulators remains a significant challenge. Herein, we report a conceptual approach for the design of an optically controlled DNA nanomachine by introducing a ternary DNAzyme-based, UV light-cleavable DNA scaffold and upconversion nanoparticle to the activatable hybrid chain reaction. We demonstrate that this nanomachine is capable of being effectively operated either in the presence of an endogenous miRNA target or the coexistence of intracellular Zn2+ and external near-infrared light, resulting in enhanced fluorescence resonance energy transfer signals. With this design, the logic-gated imaging of endogenous miR-21 and Zn2+ is demonstrated in living cells. More importantly, taking advantages of photoacoustic imaging modality, a combinational logic circuit (AND/OR) is constructed for the bioorthogonal cascade imaging of miR-21 and Zn2+ in vivo, realizing dynamic monitoring of the correlation of miRNA and metal ions levels. Collectively, our results suggest that this conceptual design possesses the ability to expand the DNA nanomachine toolbox for visualizing a broad spectrum of interconnected molecules and thus provides new perspectives to improve the diagnostic and therapeutic outcomes.

Spatiotemporal-Resolved Hyperspectral Raman Imaging of Plasmon-Assisted Reactions at Single Hotspots.

Raman spectroscopy facilitates the study of reacting molecules on single nanomaterials. In recent years, the temporal resolution of Raman spectral measurement has been remarkably reduced to the millisecond level. However, the classic scan-based imaging mode limits the application in the dynamical study of reactions at multiple nanostructures. In this paper, we propose a spatiotemporal-resolved Raman spectroscopy (STRS) technology to achieve fast (∼40 ms) and high spatial resolution (∼300 nm) hyperspectral Raman imaging of single nanostructures. With benefits of the outstanding electromagnetic field enhancement factor by surface plasmon resonance (∼1012) and the snapshot hyperspectral imaging strategy, we demonstrate the observation of stepwise Raman signals from single-particle plasmon-assisted reactions. Results reveal that the reaction kinetics is strongly affected by not only the surface plasmon-polariton generation but also the density of Raman molecules. In consideration of the spatiotemporal resolving capability of STRS, we anticipate that it provides a potential platform for further extending the application of Raman spectroscopy methods in the dynamic study of 1D or 2D nanostructures.