Biology subjects

Nishikii, H.

Publications and source records attributed to Nishikii, H..

3 recordsLinked to original sources

bioRxiv2026

Spatial profiling of CAR protein organization reveals in vivo remodeling during CAR-T therapy

Chimeric antigen receptor (CAR) T cell therapy utilizes genetically engineered patient-derived T cells to target cancer cells. Despite its clinical successes in multiple cancer types, the underlying molecular mechanisms by which molecules on CAR-T cells and surrounding cells interact with other proteins and collectively determine treatment efficacy remain elusive. Most previous studies have relied on transcriptome profiling, which does not fully reflect protein-level organization and interactions. In this study, we developed an antibody-oligonucleotide conjugate targeting the FMC63 region of CAR and integrated it into molecular pixelation (MPX). This approach enabled profiling of the dynamics of CAR molecules on cell surfaces as well as their colocalization with other proteins at the single-cell level. By applying MPX to longitudinal samples from three patients undergoing CAR-T cell therapy, we characterized the dynamic changes in CAR-associated protein organization in both pre-infusion CAR products and post-infusion peripheral blood. While CAR protein abundance and polarization showed limited variation across clinical courses, remodeling of a CAR-centered co-localization network was observed over time, including different retentions of specific molecular associations between patients with different clinical outcomes. Although derived from a limited cohort, our study identifies insights from this methodological framework beyond those gained by conventional omics analyses and offers results of a systematic investigation to predict and enhance CAR therapeutic outcomes. Key pointsO_LIMolecular pixelation was applied for chimeric antigen receptor (CAR) profiling at single-molecule and single-cell resolutions. C_LIO_LIProtein and transcriptome analyses of the CAR molecule showed dynamic remodeling during CAR-T therapy in patients with non-Hodgkin lymphoma. C_LI

genomics↗

bioRxiv2025

A bright synthetic near-infrared luciferin enhances the capabilities of deep-tissue bioluminescence imaging using firefly luciferases

Synthetic bioluminescence reactions exhibiting near-infrared (NIR)-shifted spectra have been explored to improve deep-tissue imaging through the design of firefly luciferin analogues. Although the NIR bioluminescence reactions improve the tissue penetration of bioluminescence signals from deep tissues, their photon output is markedly lower compared to the natural reaction with D-luciferin and firefly luciferase (Fluc), often by an order of magnitude or more. Consequently, in most instances, the sensitivity of NIR bioluminescence imaging (NIR-BLI) has not yet substantially surpassed that of BLI with the natural firefly reaction. Here, we present a synthetic firefly luciferin, named AkaSuke, that generates intense NIR bioluminescence ({lambda}max = 680 nm) in reaction with Fluc, greatly improving the detection sensitivity beyond that of the D-luciferin/Fluc reaction for targeting deep tissue. AkaSuke enables sensitive visualizations of ectopic hematogenesis through entire tissues of mice over time following transplantation of bone marrow stem cells labeled with Fluc. We additionally identify a Japanese firefly luciferase, DkumLuc1, that displays higher catalytic activities for bioluminescence emission of AkaSuke compared to typical Fluc, resulting in detection sensitivity comparable to that of AkaLumine/Akaluc reaction, one of the most sensitive bioluminescence systems for deep tissue imaging. We further propose the potential of the AkaSuke/DkumLuc1 reaction as an orthogonal pair with the AkaLumine/Akaluc for sensitive dual-target tracking in mice. Overall results suggest that AkaSuke enhances the capabilities of deep-tissue bioluminescence imaging using Fluc and its variant, and could serve as an emerging benchmark for the molecular design of NIR luciferin analogues.

cancer biology↗

bioRxiv2017

Correlation of gene expressions between nucleus and cytoplasm reflects single-cell physiology

BackgroundEukaryotes transcribe RNAs in nuclei and transport them to the cytoplasm through multiple steps of post-transcriptional regulation. Existing single-cell sequencing technologies, however, are unable to analyse nuclear (nuc) and cytoplasmic (cyt) RNAs separately and simultaneously. Hence, there remain challenges to discern correlation, localisation, and translocation between them.\n\nResultsHere we report a microfluidic system that physically separates nucRNA and cytRNA from a single cell and enables single-cell integrated nucRNA and cytRNA-sequencing (SINC-seq). SINC-seq constructs two individual RNA-seq libraries, nucRNA and cytRNA per cell, quantifies gene expression in the subcellular compartments and combines them to create a novel single-cell RNA-seq data enabled by our system, which we here term in-silico single cell.\n\nConclusionsLeveraging SINC-seq, we discovered three distinct natures of correlation among cytRNA and nucRNA that reflected the physiological state of single cells: The cell-cycle-related genes displayed highly correlated expression pattern with minor differences; RNA splicing genes showed lower nucRNA-to-cytRNA correlation, suggesting a retained intron may be implicated in inhibited mRNA transport; A chemical perturbation, sodium butyrate treatment, transiently distorted the correlation along differentiating human leukemic cells to erythroid cells. These data uniquely provide insights into the regulatory network of mRNA from nucleus toward cytoplasm at the single cell level.

genomics↗