Biology subjects

Tuano, N.

Publications and source records attributed to Tuano, N..

6 recordsLinked to original sources

bioRxiv2026

Targeted DUX4 base editing improves muscle function in an iPSC-derived model of childhood-onset FSHD

Facioscapulohumeral muscular dystrophy (FSHD) is one of the most common dominant muscular dystrophies and remains without an approved disease modifying therapy. Caused by the aberrant expression of the cytotoxic gene DUX4, FSHD is typically diagnosed in adulthood, however clinical onset in children (<18 years of age) is often associated with a more severe and rapid disease. While clinical trials are underway, a lack of human-specific pre-clinical models limit effective testing of potential therapies, particularly in children. To fill this gap, we describe here the development of induced pluripotent stem cell-derived 2-and 3-dimensional skeletal muscle models of children with clinically defined mild, moderate, and severe FSHD. These iPSC-derived muscle models replicate key features of FSHD, including aberrant DUX4 mRNA expression, muscle atrophy, and weakness, which correlate with the individuals specific disease severity. Next, we assessed the efficacy of adenine base editing (ABE) as a potential gene therapy approach to treat FSHD. DUX4-targeted ABE reduced DUX4 mRNA expression, improved muscle area and force generation in the most severe individual. Together this work supports the use of iPSC-derived skeletal muscle models as a less invasive method to study childhood-onset FSHD and establishes targeted DUX4 gene editing therapies as a potential treatment approach.

cell biology↗

bioRxiv2025

Application of spatial transcriptomics across organoids: a high-resolution spatial whole-transcriptome benchmarking dataset

Stem cell-derived organoids hold promise to model tissue-specific disease. To enable this, it is crucial to assess how transcriptional signatures, cellular organisation and composition of organoids compare to in vivo counterparts. However, technologies which elucidate regional molecular identity, like spatial transcriptomics, have been challenging to apply to organoids. This study presents the first systematic profiling of multiple stem cell derived organoid models (brain, heart muscle, heart valve, kidney, lung, cartilage, and haematopoietic) with Stereo-seq, a full transcriptome, spatial transcriptomics assay using on-chip in situ RNA capture at subcellular resolution. It describes optimisation of this assay to characterise organoids, use of multiple organoid samples on a single chip, assess differences in RNA capture efficiency compared to reference tissues and its limitations. This study introduces a bespoke analysis method that partitions samples into regions and further characterises them. These findings inform future works to characterise organoids using spatial transcriptomics, providing insights in optimising RNA capture of multiple organoids across a chip and novel methods for regional analysis.

bioinformatics↗

bioRxiv2024

Maturation of human cardiac organoids are required for disease modelling and drug discovery

Cardiac maturation is an important developmental phase culminating in profound biological and functional changes to adapt to the high demand environment after birth1,2. Maturation of human pluripotent stem cell-derived human cardiac organoids (hCO) to more closely resemble human heart tissue is critical for understanding disease pathology. Herein, we profile human heart maturation in vivo3 to identify key signalling pathways that drive maturation in hCOs4,5. Transient activation of both the 5 AMP-activated kinase (AMPK) and estrogen-related receptor (ERR) promoted hCO maturation by mimicking the increased functional demands of post-natal development. hCOs cultured under these directed maturation (DM) conditions (DM-hCOs) display robust transcriptional maturation including increased expression of mature sarcomeric and oxidative phosphorylation genes resulting in enhanced metabolic capacity. DM-hCOs have functionally mature properties such as sarcoplasmic reticulum-dependent calcium handling, accurate responses to drug treatments perturbing the excitation-coupling process and ability to detect ectopy CASQ2 and RYR2 mutants. Importantly, DM- hCOs permit modelling of complex human disease processes such as desmoplakin (DSP) cardiomyopathy, which is driven by multiple cell types. Subsequently, we deploy DM-hCOs to demonstrate that bromodomain extra-terminal inhibitor INCB054329 rescues the DSP phenotype. Together, this study demonstrates that recapitulating in vivo development promotes advanced maturation enabling disease modelling and the identification of a therapeutic strategy for DSP- cardiomyopathy.

bioengineering↗

bioRxiv2024

Pooled genetic screens identify breast cancer risk genes involved in evasion from T cell-mediated killing

Genome-wide association studies have identified more than 220 loci associated with breast cancer susceptibility. A major challenge is now to identify the effector genes with plausible functions in the context of breast cancer risk. We have previously performed pooled CRISPR screens to identify target genes at risk loci that drive cancer hallmarks including proliferation or modulating DNA damage response. We now extend these screens to identify genes involved in response to cytotoxic T lymphocyte (CTL) killing. We performed knockout and inhibition screens to identify genes that affect the response of the MCF7 human breast cancer cell line to CTL killing in an in vitro co-culture system. We identified 33 candidate risk genes associated with resistance or sensitisation to T cell-mediated killing. Using single gene perturbation, we showed that deletion of candidate risk genes IRF1, ATF7IP, CCDC170 and CASP8 induced resistance, while ablation of CFLAR, CREBBP, and PRMT7 sensitized cells to CTL killing. We used reporter assays to show that the risk-associated alleles at rs736801 and rs3769821 reduced transactivation of the IRF1 and CASP8 promoters, respectively. We showed that pharmacological inhibition of PRMT7 rendered breast cells sensitive to CTL killing and PRMT7 levels were negatively correlated with CD8+ infiltration and patient survival in luminal A breast cancer patient cohorts. Our results demonstrate that phenotypic pooled CRISPR screens are a useful approach for high throughput functional follow-up of GWAS findings, identifying genes which alter immune responses to breast cancer which offer opportunities to enhance immunotherapy.

genetics↗

bioRxiv2024

SRP19 and the protein secretion machinery is a targetable vulnerability in cancers with APC loss

Adenomatous Polyposis Coli (APC) is a tumour suppressor that is frequently lost in colorectal and other cancers. A common mechanism for APC loss includes heterozygous APC deletion. Here, we show that SRP19, is located near APC and is often co-deleted in these tumours. Heterozygous APC/SRP19 loss leads to lower levels of SRP19 mRNA and protein. Consequently, cells with APC/SRP19 loss are vulnerable to partial suppression of SRP19. We show that SRP19 loss is a unique vulnerability since SRP19 is rate limiting for the formation of the Signal Recognition Particle (SRP), a complex that mediates translocation of proteins to the ER. Consistent with these observations, partial SRP19 knock-down or low dose Arsenic Trioxide treatment induces ER stress and inhibits proliferation in APC/SRP19 loss cancers. Our work identifies a new strategy to treat cancers with APC/SRP19 heterozygous deletions and provides a framework for identifying vulnerabilities associated with loss of a tumour suppressor.

cancer biology↗

bioRxiv2019

Chromatin interactome mapping at 139 independent breast cancer risk signals

Genome-wide association studies have identified 196 high confidence independent signals associated with breast cancer susceptibility. Variants within these signals frequently fall in distal regulatory DNA elements that control gene expression. We designed a Capture Hi-C array to enrich for chromatin interactions between the credible causal variants and target genes in six human mammary epithelial and breast cancer cell lines. We show that interacting regions are enriched for open chromatin, histone marks for active enhancers and transcription factors relevant to breast biology. We exploit this comprehensive resource to identify candidate target genes at 139 independent breast cancer risk signals, and explore the functional mechanism underlying altered risk at the 12q24 risk region. Our results demonstrate the power of combining genetics, computational genomics and molecular studies to rationalize the identification of key variants and candidate target genes at breast cancer GWAS signals.

molecular biology↗