Biology subjects

Himsworth, C.

Publications and source records attributed to Himsworth, C..

6 recordsLinked to original sources

bioRxiv2026

CD47 blockade augments anti-GD2 driven phagocytosis in vitro but fails to improve in vivo efficacy in immune competent, chemoresistant neuroblastoma preclinical models

CD47 delivers a dominant "Dont Eat Me" signal that inhibits macrophage-mediated clearance of tumour cells. Using immune competent, chemoresistant neuroblastoma (NB) models, we tested a Fc-silent CD47 blocker (ALX301) with anti-GD2 antibody alone and in combination with a clinically aligned temozolomide/irinotecan chemoimmunotherapy backbone. Tumours expressed GD2 and CD47, and bound ALX301. In macrophage coculture assays, anti-GD2 antibody induced dose-dependent phagocytosis, whereas ALX301 or an anti-CD47 antibody alone did not. CD47 blockade in combination with a suboptimal concentration of anti-GD2 antibody showed an additive effect on phagocytosis in vitro. In vivo, however, ALX301 failed to improve tumour control or survival when added to anti-GD2 or to chemoimmunotherapy in two models. Toxicity was acceptable, showing only mild, expected red-cell changes without organ injury. This form of CD47 inhibition is therefore mechanistically active in vitro but insufficient to enhance anti-GD2 antibody-based therapy in immune competent mice bearing a chemoresistant NB, highlighting the potential need for myeloid-reprogramming partners.

immunology↗

bioRxiv2025

An optimised method for generation of murine CAR-T cells by CRISPR/Cas9.

Development of the next generation of chimeric antigen receptor (CAR) T-cells requires assessment in systems that better recapitulate the suppressive tumour microenvironment of solid tumours. CRISPR-Cas9 knock-in of promoter-less homology directed repair templates (HDRT) into the T-cell receptor locus has been shown to result in physiological expression of CARs with improved tumour control. We initially compared the use of dsDNA and adenovirus associated virus (AAV) HDRTs in mouse T cells. We have subsequently developed an optimised method for AAV transduction resulting in high editing efficiencies with minimal toxicity. In contrast with our experience of retroviral transduction of mouse T cells, our CRISPR/Cas9 AAV transduction method results in sustained CAR expression and T cell expansion in vitro as well as in vivo persistence. This approach allows for pre-clinical assessment of individual and libraries of CAR constructs in relevant immune-competent mouse models.

molecular biology↗

bioRxiv2024

Detection of a reassortant swine- and human-origin H3N2 influenza A virus in farmed mink in British Columbia, Canada

In December 2021, influenza A viruses (IAV) were detected in a population of farmed mink in British Columbia, Canada. Based on genomic sequencing and phylogenetic analysis, these IAVs were subtyped as H3N2s that originated from reassortment of swine H3N2 (clade 1990.4h), human seasonal H1N1 (pdm09), and swine H1N2 (clade 1A.1.1.3). This reassortant has been subsequently observed in swine in several Midwest American states, as well as in swine and turkeys in Ontario, suggesting its spillover into farmed mink in British Columbia was incidental to its broader dissemination in North American swine populations. These detections reaffirm the need for extensive genomic surveillance of IAVs in swine populations to monitor reassortments that might become public health concerns. They also highlight the need for closer surveillance of IAVs in mink to preserve animal health, protect agricultural interests, and monitor potential zoonotic threats.

microbiology↗

bioRxiv2024

Blocking MIF secretion enhances CAR T-cell efficacy against neuroblastoma

While chimeric antigen receptor (CAR) T-cell therapies are showing highly promising first results in neuroblastoma, immunosuppressive tumor microenvironments (TME) limit T cell persistence and durable clinical efficacy. To improve CAR T-cell efficacy further, we applied a multi-omics approach including single-cell RNA sequencing and proteomics, which identified 13 targetable immunosuppressive factors in neuroblastoma. Of these, macrophage migration inhibitory factor (MIF) and midkine (MDK) were validated across multiple published RNA datasets. Moreover, they were secreted in high abundance by neuroblastoma tumoroids. Functional validation experiments revealed MIF as a potent inhibitor of CAR T-cells, in vitro and in vivo. Degradation of MIF by PROTAC technology significantly enhanced CAR T-cell activation targeting GPC2 and B7-H3, providing a potential intervention against MIF. By defining the immunosuppressive effects of neuroblastomas TME on CAR T-cell efficacy, particularly the pivotal role of MIF, we provide a therapeutic strategy for improving adoptive cell therapies for this pediatric malignancy.

cancer biology↗

bioRxiv2024

Functional avidity of anti-B7H3 CAR-T constructs predicts antigen density thresholds for triggering effector function

Chimeric Antigen receptor T cell (CAR-T) treatments for solid cancers have been compromised by limited expansion and survival in the tumor microenvironment following interaction with antigen-expressing target cells. Using B7H3 as a model antigen with broad clinical applicability, we evaluated the relationship between the antibody/antigen affinity of three clinical candidate binders and the three following functional characteristics: functional avidity, prolonged cytotoxicity in tumoroid re-stimulation assays, and in vivo anti-tumoral responses. BEHAV3D video-microscopy assessed distinct CAR-T cell behaviors at single cell resolution. T cell exhaustion did not dictate effector function. Rather, we demonstrated a threshold avidity of CAR-T / tumor cell interaction, characterized by longer cumulative CD8+ CAR-T / tumor target interaction times, and required for adequate CAR-T cell expansion to result in sustained tumor control upon re-challenge. These results provide new insights into design of CAR-T cells for antigen-dim cell targeting, and avoidance of antigen-dim tumor relapse.

bioengineering↗

bioRxiv2022

ProbeTools: Designing hybridization probes for targeted genomic sequencing of diverse and hypervariable viral taxa

BackgroundSequencing viruses in many specimens is hindered by excessive background material from hosts, microbiota, and environmental organisms. Consequently, enrichment of target genomic material is necessary for practical high-throughput viral genome sequencing. Hybridization probes are widely used for enrichment in many fields, but their application to viral sequencing faces a major obstacle: it is difficult to design panels of probe oligo sequences that broadly target many viral taxa due to their rapid evolution, extensive diversity, and genetic hypervariability. To address this challenge, we created ProbeTools, a package of bioinformatic tools for generating effective viral capture panels, and for assessing coverage of target sequences by probe panel designs in silico. In this study, we validated ProbeTools by designing a panel of 3,600 probes for subtyping the hypervariable haemagglutinin (HA) and neuraminidase (NA) genome segments of avian-origin influenza A viruses (AIVs). Using in silico assessment of AIV reference sequences and in vitro capture on egg-cultured viral isolates, we demonstrated effective performance by our custom AIV panel and ProbeTools suitability for challenging viral probe design applications. ResultsBased on ProbeTools in silico analysis, our panel provided broadly inclusive coverage of 14,772 HA and 11,967 NA reference sequences. 90% of these HA and NA references sequences had 90.8% and 95.1% of their nucleotide positions covered in silico by the panel respectively. We also observed effective in vitro capture on a representative collection of 23 egg-cultured AIVs that included isolates from wild birds, poultry, and humans and representatives from all HA and NA subtypes. 42 of 46 HA and NA segments had over 98.3% of their nucleotide positions significantly enriched by our custom panel. These in vitro results were further used to validate ProbeTools in silico coverage assessment algorithm; 89.2% of in silico predictions were concordant with in vitro results. ConclusionsProbeTools generated an effective panel for subtyping AIVs that can be deployed for genomic surveillance, outbreak prevention, and pandemic preparedness. Effective probe design against hypervariable AIV targets also validated ProbeTools design and coverage assessment algorithms, demonstrating their suitability for other challenging viral capture applications.

genomics↗