Zhao X, Jing W, Wang G
… +16 more, Liu Z, Xu X, Han M, Fu Z, Zhang Y, Zheng Z, Zhang J, Bo L, Dong X, Li C, Sun Y, Zhang J, Zhao F, Xing N, Zhao K, Jiang X
The antitumor efficacy of immune cell engagers that bind two targets on the same immune cell is limited by structural constraints, leading to incomplete coengagement and uncoordinated signaling. Here, we develop a trispe...The antitumor efficacy of immune cell engagers that bind two targets on the same immune cell is limited by structural constraints, leading to incomplete coengagement and uncoordinated signaling. Here, we develop a trispecific macrophage engager (TrME) that both activates the prophagocytic receptor lipoprotein receptor-related protein 1 (LRP1) and blocks the antiphagocytic receptor signal regulatory protein alpha (SIRPα). This 'activate and block' AND logic gate, when coupled to a tumor-targeting moiety, enables coordinated signaling that enhances macrophage cytotoxicity against solid tumors. The TrME tandemly links monovalent LRP1 activator calreticulin, anti-SIRPα scFv and a tumor-associated antigen (TAA)-targeting arm through flexible linkers. Computational modeling and screening of tandem constructs revealed an optimal conformation for robust cis-targeting, allowing logic-gated control of ratiometric prophagocytic and antiphagocytic signaling. In situ generation of TrME by delivering mRNA encoding TAA-targeting TrME through an optimized lipid nanoparticle system activates macrophages and induces antitumor responses, significantly inhibiting tumor growth and prolonging survival in multiple solid tumor mouse models.
Biomolecular profiling offers a powerful lens into human physiology, yet current diagnostics often rely on invasive sampling and delayed, centralized analysis. Advances in mass spectrometry (MS), particularly untargeted...Biomolecular profiling offers a powerful lens into human physiology, yet current diagnostics often rely on invasive sampling and delayed, centralized analysis. Advances in mass spectrometry (MS), particularly untargeted metabolomics and proteomics, have expanded molecular access to noninvasive biofluids such as sweat, saliva, tears and interstitial fluid, revealing dynamic biomarkers linked to both chronic and acute conditions. In parallel, wearable biosensors enable real-time, on-body chemical sensing, but remain limited to a narrow panel of predefined analytes. This Review highlights how MS-based molecular discovery and wearable sensing serve as complementary approaches-MS enabling high-dimensional untargeted profiling and wearables delivering longitudinal real-time data-and also discusses how their bidirectional integration and co-evolution open new possibilities for personalized noninvasive health monitoring. We discuss advances in sampling strategies, sensing modalities and system integration, and outline criteria for identifying biomarkers amenable to sensor translation. By uniting untargeted discovery with real-world deployment, this convergence shifts personalized noninvasive healthcare from episodic diagnostics to continuous, context-aware monitoring.
TnpB is a diverse family of RNA-guided endonucleases associated with prokaryotic transposons. Because of their small size and putative evolutionary relationship to CRISPR-Cas12, TnpB enzymes hold great potential for geno...TnpB is a diverse family of RNA-guided endonucleases associated with prokaryotic transposons. Because of their small size and putative evolutionary relationship to CRISPR-Cas12, TnpB enzymes hold great potential for genome editing. However, most TnpBs lack robust gene-editing activity. Here, we mapped comprehensive sequence-function landscapes of a TnpB ribonucleoprotein using deep mutational scanning and we discovered activating mutations in both the RNA and the protein. Leveraging the protein's mutational landscape, we constructed a combinatorial library of activating mutations, from which we identified two enhanced TnpB variants. These variants increased editing in human cells, Nicotania benthamiana, pepper and rice. While editing efficiencies varied by target site, engineered variants achieved up to 55% insertion and deletion frequencies (a 50-fold increase over wild type) in N. benthamiana, surpassing ISYmu1 (<7%), AsCas12f-HKRA (<9%) and other compact editors. These findings highlight elements critical for regulating TnpB endonuclease activity and demonstrate latent activity accessible through mutation.
Jozić A, Le Roux C, Kim J
… +12 more, Berchel M, Sahel DK, Bodi EK, Palumbo M, Vasudevan A, Murthy NTV, Eygeris Y, Gautam M, Bloom E, Barnes AP, Jaffrès PA, Sahay G
Endosomal escape is a central barrier to efficient nucleic acid delivery by lipid nanoparticles (LNPs) and remains challenging to quantify in vivo. We report a library of branched ionizable phospholipids that markedly en...Endosomal escape is a central barrier to efficient nucleic acid delivery by lipid nanoparticles (LNPs) and remains challenging to quantify in vivo. We report a library of branched ionizable phospholipids that markedly enhance messenger RNA delivery to the liver. The lead candidate BiP-20 outperformed the clinical benchmark LP01 by eightfold for CRISPR-Cas9 editing of the TTR gene at low dose with rapid pharmacokinetics. To quantify the endosomal escape kinetics of BiP-20, we used LysoTag mice, which allow immunoisolation of liver lysosomes, and our Lysosomal Barcoding method, finding that ~8% of BiP-20 LNPs reach the cytosol within 30 min of administration. Lysosomal proteomics revealed mechanistic regulators of escape and BiP-20-induced alterations in endosomal maturation and recycling pathways. Loss of Rab7, a mediator of late endosomal maturation, increased LNP escape. These findings provide a potent class of ionizable lipids for RNA delivery, a method to quantify endosomal escape in vivo, and mechanistic insight into the endolysosomal determinants of LNP trafficking.