Biology subjects

Cook, H.

Publications and source records attributed to Cook, H..

3 recordsLinked to original sources

bioRxiv2025

Rictor CRISPR Gene Editing by Lipid Nanoparticle Delivery Stimulates Anti-Tumor Immunity in Breast Cancer Liver Metastasis Model

Triple-negative breast cancer (TNBC) is a highly aggressive subtype, accounting for 10-15% of breast cancer cases in the United States. Liver metastases, common in advanced TNBC, are linked to especially poor outcomes, with a 5-year survival rate of just 11%. Although immune checkpoint inhibitors (ICIs) targeting PD-1 or PD-L1 show promise, durable responses in TNBC remain uncommon. This is largely due to a profoundly immunosuppressive tumor microenvironment (TME), driven by tumor-associated myeloid cells. Tumor-associated macrophages (TAMs) and neutrophils (TANs) polarize into immunosuppressive M2 and N2 phenotypes, respectively, suppressing T cell activity through cytokines, ROS, and checkpoint ligands such as VISTA. Myeloid-derived suppressor cells (MDSCs) further inhibit immunity by depleting nutrients and inducing regulatory T cells. As a result, despite its immunogenic features, TNBC remains resistant to immunotherapy due to persistent myeloid-mediated suppression. Here, we developed ionizable lipid nanoparticles (iLNPs) engineered to deliver the CRISPR-Cas12a ribonuclease complex targeting Rictor, a critical component of mTORC2, for in vivo reprograming of myeloid cells. The intravenous (IV) injection of CRISPR Rictor-targeting iLNP (CR-Ric-LNP) showed efficient uptake by circulating myeloid cells and accumulation into the breast cancer liver metastases. Notably, Rictor gene editing triggered pro-inflammatory activation of myeloid cells in the TME, enhancing antitumor responses. Single-cell RNA sequencing revealed that Rictor silencing treated samples showed induced rapid remodeling of the TME, with a significant reduction in immunosuppressive macrophages within 24 hours of treatment. Concurrently, cytotoxic T-cell populations exhibited increased interferon-gamma (Ifng) production, driving the emergence of specific myeloid clusters that were responsive to Interferon signaling, particularly in macrophages and neutrophils. A shift from an immunosuppressive to an inflammatory TME was further evidenced by an elevated Cxcl10/Spp1 ratio in myeloid cells. CR-Ric-LNP treatment also enhanced T-cell activation, reducing exhausted T cells and regulatory T cells (Tregs) while expanding natural killer (NK) cells, naive CD4+, and CD8+ T cells. These changes correlated with a decreased proportion of tumor cells and proliferating cells, ultimately leading to a significant survival benefit in a 4T1 breast cancer liver metastasis model. Our findings demonstrate that myeloid-targeted Rictor silencing reprograms the TME, promoting antitumor immunity and improving therapeutic outcomes. O_FIG O_LINKSMALLFIG WIDTH=199 HEIGHT=200 SRC="FIGDIR/small/661471v1_ufig1.gif" ALT="Figure 1"> View larger version (109K): org.highwire.dtl.DTLVardef@10a5e7aorg.highwire.dtl.DTLVardef@1c0c005org.highwire.dtl.DTLVardef@17ec6b5org.highwire.dtl.DTLVardef@27c507_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2023

Dominant CIZ1 fragments drive epigenetic instability and are expressed in early stage cancers

CIZ1 is a nuclear matrix protein that is part of the large RNA-dependent supramolecular assembly complexes (SMACs) that form at the inactive X chromosome (Xi) in female cells, and smaller assemblies throughout the nucleus in males and females. It plays a role in maintenance of epigenetic state and gene expression in differentiated cells, via stabilisation of histone post-translational modifications H2AK119ub1 and H3K27me3, added by polycomb repressive complexes (PRC) 1 and 2. Here, we show that expression of the N-terminal replication domain (RD) and C-terminal anchor domain (AD) of human CIZ1 transcript is uncoupled, with consistently elevated AD in early stage breast cancers, and sporadically elevated AD in other common solid tumours. At the protein level CIZ1-Xi SMACs are corrupted in female breast cancers cells, and this is accompanied by elevated AD-encoding transcripts. We modelled the effect of AD fragments in primary murine embryonic fibroblasts and observed dominant-negative interference with CIZ1 SMACs during their assembly in early G1 phase. Mutagenesis identified the matrin 3 homology domain as essential for self-interaction to form stable homodimers in vitro, and as a determinant of its dominant-negative effect in cells, implicating the dimerization interface in CIZ1 SMAC integrity. SMAC disruption was coincident with depletion of PRC1-dependent H2AK119ub1 from Xi chromatin, in a manner abrogated by the PR-deubiquitinase inhibitor PR619, suggesting that CIZ1 SMACs normally stabilise H2AK119ub1 by shielding Xi chromatin from attack by deubiquitinases. Moreover, SMAC disruption was accompanied by changes in gene expression within days. Together, the data suggest that inappropriate expression of CIZ1 AD fragments could drive epigenetic instability in early stage breast cancers by destabilizing the CIZ1 SMACs that normally protect repressed chromatin. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=80 SRC="FIGDIR/small/558821v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@13b1be2org.highwire.dtl.DTLVardef@1a3a16corg.highwire.dtl.DTLVardef@1bac386org.highwire.dtl.DTLVardef@7ac176_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2022

Migratory hoverflies orientate north during spring migration

Migratory hoverflies are long-range migrants that, in the northern hemisphere, move seasonally to higher latitudes in the spring and lower latitudes in the autumn. The preferred migratory direction of hoverflies in the autumn has been the subject of radar and flight simulator studies while spring migration has proved to be more difficult to characterise due to a lack of ground observations. Consequently, the preferred migratory direction during spring has only been inferred from entomological radar studies and patterns of local abundance and currently lacks ground confirmation. Here, during a springtime arrival of migratory insects onto the Isles of Scilly and mainland Cornwall, UK, we provide ground proof that spring hoverfly migrants have an innate northward preference. Captured migratory hoverflies displayed northward vanishing bearings when released under sunny conditions under both favourable wind and zero-wind conditions. In addition, and unlike autumn migrants, spring individuals were also able to orientate when the sun was obscured. Analysis of winds suggests an origin for insects arriving on the Isles of Scilly in from western France. These findings of spring migration routes and preferred migration directions are likely to extend to the diverse set of insects found within the western European migratory assemblage.

zoology↗