Cheng Zhu

Metabolic dysfunction–associated steatohepatitis (MASH) is characterized by the excessive accumulation of triglycerides (TG) in the liver, with long‐chain fatty acids (LCFAs) as key TG precursors. CD36, a pivotal transporter, facilitates the uptake of LCFAs into hepatocytes. Currently, there are no effective pharmacological treatments available for early‐stage MASH. To address this, we developed AILITIX, a de novo AI‐designed lipid transport inhibitor. AILITIX exhibits high binding affinity for CD36 and effectively inhibits CD36‐mediated fatty acid uptake. We then synthesized ultrasound‐responsive, liver‐targeted liposomal nanoparticles (NPs) surface‐functionalized with AILITIX and indocyanine green (ICG), and encapsulating the natural flavonoid isoquercitrin (IQ) and the FXR agonist obeticholic acid (OCA). In vitro assays demonstrated that IQ directly binds to and inhibits ACSL4 activity, thereby suppressing SREBF1‐driven de novo lipogenesis. Furthermore, the NPs inhibited the phosphorylation of mTOR, ULK1, and TFEB, activating the autophagy pathway, enhancing lipid degradation, and alleviating oxidative and mitochondrial stress. In murine models, the NPs treatment led to synergistic amelioration of steatosis, oxidative damage, and inflammation. Moreover, single‐cell transcriptomics demonstrated that the intervention suppresses fibrosis‐associated gene expression and reshapes the hepatic immune microenvironment. Collectively, our de novo AI‐designed peptide‐based nanotherapeutics presents a great potential for the precision treatment of MASH.

Due to pronounced tumor heterogeneity and the lack of effective therapeutic options, triple‐negative breast cancer (TNBC) remains a formidable clinical challenge for the development of effective medicines. Here, we report a HER3‐targeted sonogenetic mechano‐immunomodulatory nanoplatform (HSMIN‐LNP), achieving integrated synergistic therapy for TNBC. HSMIN‐LNP employs a de novo AI‐designed HER3‐targeting miniprotein (HTIM) and co‐delivers the TRPV4 (mechanosensory) plasmid together with an NFAT‐promoted IL‐15 gene. The results demonstrated that HTIM directly bound and suppressed HER3 signaling, inhibiting the downstream PI3K/AKT/mTOR pathway. This process markedly impaired tumor cell migratory capacity. The NPs also exhibited a uniform and stable nanostructure with high plasmid encapsulation efficiency and enabled ultrasound (US)‐triggered local IL‐15 induction through TRPV4‐mediated Ca ²⁺ /NFAT signaling. Mechanistically, US‐activated TRPV4 further elicited pronounced Ca ²⁺ overload, reactive oxygen species accumulation, mitochondrial damage, and apoptosis in tumor cells. It simultaneously activated cGAS‐STING‐dependent inflammatory and interferon signaling pathways, thereby contributing to tumor cell death and immune‐associated antitumor responses. In vivo studies further verified that HSMIN‐LNP significantly enhanced dendritic cell maturation, NK cell and CD8 ⁺ T cell responses, as well as memory T cell formation. Collectively, this study provides a promising avenue for integrated precision therapy of TNBC.

Excessive alcohol consumption poses significant health risks and is closely associated with oxidative damage. The KEAP1-NRF2-ARE signaling pathway serves as the primary antioxidant system. However, current small molecule inhibitors are all covalently bound to KEAP1, meaning that once bound, they are not easily dissociated, while continuous inhibition of KEAP1 exhibits severe side effects. In this study, BLI, CETSA, Pull-down, Co-IP, and HDX-MS assay analysis were conducted to detect the KEAP1 binding behavior of natural product, capsaicin (CAP), both in vitro and in cells. The ethanol-induced acute gastric mucosal damage rat model was also established to evaluate the therapeutic effect of CAP. Our findings demonstrated that CAP mitigated mitochondrial damage, facilitated the nuclear translocation of NRF2, leading to the up-regulation of downstream antioxidant response elements, HMOX1, TXN, GSS, and NQO1 in GES-1 cells. Furthermore, CAP directly bind to KEAP1 and inhibit the interaction between KEAP1 and NRF2. In the KEAP1-knockout 293T cells, CAP failed to activate NRF2 expression. We identified that CAP non-covalently bound to the Kelch domain and allosterically regulated three specific regions of KEAP1: L342-L355, D394-G423, and N482-N495. To improve drug solubility and delivery efficiency, we developed IR-Dye800 modified albumin-coated CAP nanoparticles. The nanoparticles significantly reduced the gastric mucosal inflammation and activated NRF2 downstream genes in vivo. Our hypothesis was further verified our hypothesis in Nrf2-knockout mice. This study provides new insights that CAP is a safe and novel NRF2 agonist by allosterically regulating KEAP1, which may contribute to the development of lead drugs for oxidative stress-related illness, e.g., aging, cancer, neurodegenerative, and cardiovascular diseases.

Circularly permuted green fluorescent protein (cpGFP)-based high-performance cAMP sensors have enabled real-time monitoring of cAMP dynamics with high spatiotemporal resolution in living animals. However, their utility is hampered by significant spectral overlap with other green/yellow fluorescent indicators and blue/cyan light-activated optogenetic actuators, limiting their compatibility in multiplexed imaging applications. While existing red cAMP sensors offer great spectral separation, they often suffer from a limited dynamic range ( < 1.5-fold in HEK293T cells), low cellular brightness, aggregation, or significant blue-light-induced photoactivation. Here, we report R-Flamp1, a red cAMP sensor with a large dynamic range ( > 10-fold in HEK293T cells), enhanced cellular brightness, appropriate cAMP affinity (K_d ~1.9 μM), subsecond response kinetics, and minimal photoactivation under blue or cyan light exposure. Using R-Flamp1, we visualized region-specific cAMP dynamics, and when paired with green indicators, revealed differential activation patterns between cAMP and neuromodulators or calcium using two-photon imaging and fiber photometry during various behaviors. These findings provide valuable insights into the role of cAMP signaling in complex behaviors.

The severity of the recent Coronavirus Disease 2019 pandemic stresses the importance of analytical and biosensor research aimed at determining and curbing disease severity. It demanded an easy-to-adapt method to reflect the infectivity of viruses. The critical role of 3C-Like protease (3CL^Pro) in the replication cycle of coronaviruses, such as SARS-CoV-2, highlights its potential as an effective target correlated with viral activities. In this study, we aimed to develop a modular and orthogonal analytical tool for the detection and quantification of the essential protease component of coronaviruses, focusing on the enzymatic activity of the viral 3CL^Pro, a key component for coronavirus replication. Our approach leveraged the high sequence conservation of 3CL^Pro across global coronavirus strains, particularly in its substrate recognition pocket, to computationally design optimal substrate peptides. The designed sequences were orthogonal to any natural viral protein sequences. When incorporated into Gluc or FlipGFP proteins, our designs achieved modular gain-of-signal or loss-of-signal detections of 3CL^Pro expression levels. These antigen-focused molecular tools could facilitate the screening of effective treatments and quantitative visualization of virus-infected cells.

Antimicrobial peptides (AMPs) are attractive candidates to combat antibiotic resistance for their capability to target biomembranes and restrict a wide range of pathogens. It is a daunting challenge to discover novel AMPs due to their sparse distributions in a vast peptide universe, especially for peptides that demonstrate potencies for both bacterial membranes and viral envelopes. Here, we establish a de novo AMP design framework by bridging a deep generative module and a graph-encoding activity regressor. The generative module learns hidden ‘grammars’ of AMP features and produces candidates sequentially pass antimicrobial predictor and antiviral classifiers. We discovered 16 bifunctional AMPs and experimentally validated their abilities to inhibit a spectrum of pathogens in vitro and in animal models. Notably, P076 is a highly potent bactericide with the minimal inhibitory concentration of 0.21 μM against multidrug-resistant Acinetobacter baumannii, while P002 broadly inhibits five enveloped viruses. Our study provides feasible means to uncover the sequences that simultaneously encode antimicrobial and antiviral activities, thus bolstering the function spectra of AMPs to combat a wide range of drug-resistant infections.