Biology subjects

Tsao, A.

Publications and source records attributed to Tsao, A..

6 recordsLinked to original sources

bioRxiv2026

An Optimized RNF126-Targeting Covalent Handle for Molecular Glue Degraders

Molecular glue degraders represent a powerful modality for targeting proteins that are refractory to traditional inhibition. However, rational design principles for molecular glue degraders remain poorly defined. Previously, we reported a chemistry-centric strategy to identify covalent degradative handles that, when appended to established ligands, convert non-degradative inhibitors into molecular glue degraders by engaging permissive E3 ligases. This effort identified a fumarate-based electrophilic handle that covalently modified the E3 ligase RNF126, enabling degradation of multiple protein targets when transplanted across diverse ligands. Despite its conceptual impact, the high intrinsic reactivity and cytotoxicity of the fumarate handle limited its translational utility. Here, we report the development of an optimized and metabolically stabilized RNF126-targeting covalent handle incorporating a trans-cyclobutane linker that exhibits reduced glutathione reactivity and diminished cytotoxicity while retaining robust degradative activity. When appended to the BET bromodomain inhibitor JQ1, this optimized handle yielded a potent and selective BRD4 degrader whose activity was dependent on RNF126. Importantly, transplantation of this handle onto a previously non-inhibitory ligand targeting the androgen receptor (AR) and its truncation variant, AR-V7, enabled selective degradation of both AR and AR-V7 in androgen-independent prostate cancer cells, thereby robustly inhibiting AR transcriptional activity beyond the established AR antagonist enzalutamide. Collectively, these findings demonstrate an optimized RNF126-based covalent handle for the rational development of molecular glue degraders against transcriptional regulators, including undruggable variants such as AR-V7.

biochemistry↗

bioRxiv2025

Armoring STEAP1 CAR T cells with IL-18 potentiates antitumor activity in Ewing sarcoma

BackgroundEwing sarcoma (EwS) is a highly aggressive cancer driven by the EWS::FLI1 fusion oncoprotein affecting children, adolescents, and young adults. Six transmembrane epithelial antigen 1 (STEAP1) is a cell surface antigen transcriptionally controlled by EWS::FLI1 that is broadly expressed in EwS, positioning it as a rational immunotherapy target. However, translating CAR T therapy to solid tumors requires overcoming barriers to potency while maintaining safety. MethodsAnalyses of transcriptome and proteome data were performed to evaluate the effects of EWS::FLI1 perturbation on STEAP1 expression at the transcript and protein levels in EwS models. STEAP1 expression was validated in EwS patient tissues by immunohistochemistry. Second-generation STEAP1-BB{zeta} CAR T cells were tested in orthotopic and disseminated EwS xenograft models. To enhance antitumor activity, an IL-18-armored STEAP1 CAR was engineered. Dose-dependent therapeutic efficacy and safety were evaluated through measurement of tumor burden, survival, and observation for gross toxicities. ResultsSTEAP1 was expressed in [~]97% of primary EwS tumors and directly associated with EWS::FLI1 fusion protein expression in EwS cell lines. In orthotopic EwS models, STEAP1 CAR T cells induced complete tumor regression at 5 x 106 cells. In disseminated disease models, responses were dose-dependent with no evidence of antigen loss. Notably, IL-18 armored STEAP1 CAR T cells achieved complete responses in [~]80% of mice at a reduced dose of 106 cells without overt toxicity. ConclusionsThese data establish STEAP1 as a clinically relevant and highly expressed target in EwS and demonstrate that IL-18 armoring significantly improves CAR T cell efficacy by enhancing potency evident through antitumor activity at reduced cell dose. STEAP1 CAR T cells are currently under evaluation in a first-in-human phase 1/2 dose-escalation clinical trial for metastatic castration-resistant prostate cancer (NCT06236139) and these studies support future clinical translation of STEAP1 CAR T cell therapy for relapsed/refractory EwS.

cancer biology↗

bioRxiv2025

Mosaic pattern: lung functional heterogeneity at the alveolus level

Inhaled particles carrying pathogens, pollutants (e.g., microplastics, smoke), therapeutics, and diagnostics are increasingly relevant to public health, yet real-time tracking of aerosol transport in functional alveoli remains challenging. Here, we used the recently developed crystal ribcage to investigate aerosol transport in ex vivo lungs during active ventilation, obtaining the first real-time observations of single aerosol droplet transport and deposition in functional alveoli. We discovered deterministic heterogeneity at both intra- and inter-alveolar levels, with aerosol distribution forming a characteristic "mosaic" pattern in which only specific alveolar clusters received particles. The pattern was consistently formed in vivo during spontaneous breathing and ex vivo using both positive- and negative-pressure ventilation. This pattern was also consistent across a range of aerosols, including small molecules, nanobodies, nanoparticles, microplastics, therapeutics, and pathogens. Additionally, the pattern was observed in murine, porcine, and human lungs, and evolved from birth through aging in mice. The post-deposition stability of the pattern depended on particle type and lung age, lasting from a few minutes for small molecular weight particles to multiple days for cell-binding particles. These alveolar-level heterogeneities may uncover previously unrecognized biological and immunological heterogeneities associated with the mosaic pattern, including its role in postnatal lung development, susceptibility to inhaled airborne hazards such as pollutants and infectious agents, and early pathogenesis and response to inhaled therapeutics in respiratory diseases such as pneumonia, COPD, asthma, and lung cancer.

bioengineering↗

bioRxiv2024

Comparison of imaging-based single-cell resolution spatial transcriptomics profiling platforms using formalin-fixed, paraffin-embedded tumor samples

Imaging-based spatial transcriptomics (ST) is evolving rapidly as a pivotal technology in studying the biology of tumors and their associated microenvironments. However, the strengths of the commercially available ST platforms in studying spatial biology have not been systematically evaluated using rigorously controlled experiments. In this study, we used serial 5-m sections of formalin-fixed, paraffin-embedded surgically resected lung adenocarcinoma and pleural mesothelioma tumor samples in tissue microarrays to compare the performance of the single cell ST platforms CosMx, MERFISH, and Xenium (uni/multi-modal) platforms in reference to bulk RNA sequencing, multiplex immunofluorescence, GeoMx Digital Spatial Profiler, and hematoxylin and eosin staining data for the same samples. In addition to objective assessment of automatic cell segmentation and phenotyping, we performed pixel-resolution manual evaluation of phenotyping to carry out pathologically meaningful comparison between ST platforms. Our study detailed the intricate differences between the ST platforms, revealed the importance of parameters such as tissue age and probe design in determining the data quality, and suggested reliable workflows for accurate spatial profiling and molecular discovery.

bioinformatics↗

bioRxiv2023

Hemin-driven chromatin remodelling by atherosclerotic risk gene SMARCA4 switches human blood-derived macrophages from leukocyte disposal to erythrocyte disposal

BackgroundPutative genetic risk loci for atherosclerotic vascular disease include SMARCA4, a chromatin remodeling gene important for gene activation. Its causal role in atherosclerosis has been uncertain. Intraplaque hemorrhage (IPH) is a late event in atherosclerosis that is linked to plaque destabilisation and increased inflammation. IPH is countered by Mhem macrophages, which are directed by hemin-mediated induction of Heme Oxygenase 1 (HMOX1) via Activating Transcription Factor 1 (ATF1). Atf1 deficiency in vivo impairs hematoma clearance, promoting inflammation and oxidative stress. Like its homologue cyclic-adenosine monophosphate response element binding protein 1 (CREB1), ATF1 is normally cyclic-AMP activated. HypothesisHemin-directed chromatin remodelling by SMARCA4 regulates specificity of ATF1 gene-binding, thereby switching between leukocyte disposal and erythrocyte disposal, contributing to its role in atherosclerosis. ResultsWe here show that SMARCA4 is genetically independent of the adjacent LDLR locus (p<0.05). In human blood-derived macrophages, hemin triggered histone acetylation (H3K9Ac) and SMARCA4 recruitment in advance of p-ATF1 recruitment at the HMOX1 enhancer. si-RNA-mediated SMARCA4-knockdown suppressed p-ATF1 binding to HMOX1 but increased its binding to cyclic-AMP responsive genes FOS and NR4A2, with corresponding changes in mRNA levels. This functionally correlated with SMARCA4-knockdown switching hemin to mimic prostacyclin (PGI2), for induced genes and phagocytic disposal of leukocytes rather than erythrocytes. ConclusionsThese data establish SMARCA4 as an independent atherosclerosis risk gene and reveal a novel mechanism in which it switches between disposal of leukocytes or erythrocytes, with important clinical implications for atherosclerotic inflammation and intraplaque hemorrhage including treatment by histone deacetylase inhibitors.

pathology↗

bioRxiv2022

Targeting advanced prostate cancer with STEAP1 chimeric antigen receptor T cell therapy

Six transmembrane epithelial antigen of the prostate 1 (STEAP1) is a compelling tumor-associated cell surface antigen for therapeutic targeting in solid tumors. We identified broad expression of STEAP1 (87% positive) in lethal metastatic prostate cancer, even more so than prostate-specific membrane antigen (PSMA, 60% positive) which is a clinically established diagnostic and therapeutic target. Second-generation chimeric antigen receptor (CAR) T cells were engineered for reactivity against STEAP1 and demonstrated substantial antitumor activity in metastatic human prostate cancer models in immunodeficient mice. Adoptive transfer of STEAP1 CAR T cells was associated with prolonged peripheral persistence and either disease eradication or substantial tumor growth inhibition with progressive disease demonstrating antigen loss. As STEAP1 CAR T cells were also highly active in antigen density conditions as low as [~]1,500 molecules/cell, we generated a human STEAP1 (hSTEAP1) knock-in (KI) mouse to evaluate the potential for on-target off-tumor toxicities. hSTEAP1-KI mice demonstrated a pattern of systemic hSTEAP1 expression akin to that observed in humans with the greatest expression found in the prostate gland. Mouse-in-mouse studies of STEAP1 CAR T cell therapy in immunocompetent hSTEAP1-KI mice engrafted with disseminated mouse prostate cancer showed preliminary safety without evidence of gross toxicity, cytokine storm, or architectural disruption and increased T cell infiltration at sites of systemic hSTEAP1 expression. Tumor responses and extension of survival were appreciated but antigen loss was identified in recurrent and progressive disease. In summary, we report the extent of STEAP1 expression in treatment-refractory metastatic prostate cancer, the generation of a STEAP1 CAR T cell therapy with promising potency and safety in preclinical studies of advanced prostate cancer, and antigen escape as a mechanism of resistance to effective STEAP1 CAR T cell therapy.

cancer biology↗