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Biology subjects

Cunnea, P.

Publications and source records attributed to Cunnea, P..

4 recordsLinked to original sources

Multimodal imaging reveals a lysosomal drug reservoir that drives heterogeneous distribution of PARP inhibitors

For all drugs, effective target engagement requires sufficient intracellular concentrations of drug to be reached, but whether tumour heterogeneity impacts drug distribution and efficacy is poorly studied. PARP inhibitors have transformed treatment of high-grade serous ovarian carcinoma (HGSOC), but resistance remains a clinical hurdle in this highly heterogeneous tumour type. We developed a patient-derived explant multi-modal imaging pipeline, which demonstrated that cell-intrinsic PARP inhibitor accumulation is highly variable, both between patients and within tumours. Spatial transcriptomics revealed enrichment of apoptotic and lysosomal signatures in high-drug regions. Rucaparib, an intrinsically fluorescent PARP inhibitor, accumulates heterogeneously at the single-cell level, with rucaparib-high cells demonstrating increased drug response relative to rucaparib low. Mechanistically, lysosomal sequestration creates a rucaparib reservoir that determines drug levels in the nucleus. Perturbation of lysosomal content altered intracellular levels of weak base PARP inhibitors rucaparib and niraparib, but not olaparib. Together these data suggest that lysosomes act as a reservoir for a subset of PARP inhibitor drugs to improve drug response.

cancer biology↗

Cleared tissue dual-view oblique plane microscopy

We present a dual-view oblique plane microscope (dOPM) for imaging thick optically cleared tissue samples using a silicone immersion primary objective. The custom-designed remote refocusing relay utilizes stock optics to achieve remote refocusing in refractive index-matched samples. The spatial resolution of the system was characterized using a series of fluorescent bead phantoms with refractive indices ranging from 1.4-1.5, with the point spread function full width at half maximum measuring [~]0.5 {micro}m laterally and [~]1 {micro}m axially for a refractive index-matched bead sample, with minimal degradation over a >250 {micro}m axial range. We characterize how the remote refocusing performance at sample refractive indices up to n = 1.5 can be partially compensated for using adjustment of the correction collar on the primary objective. We apply the system to imaging a range of biological samples with varied refractive indices. Combined with tiled acquisition, image stitching and multi-view image fusion, the microscope enables multicolour imaging of mm-wide and [~]250 {micro}m thick optically cleared mouse ovarian cancer and colon tissue samples with subcellular resolution. We also image a whole Drosophila melanogaster fruit fly brain. The system offers a platform for fast and high-resolution, multicolour volumetric imaging across spatial scales, integrated on a commercially available microscope frame.

biophysics↗

Myeloid cell networks determine reinstatement of original immune environments in recurrent ovarian cancer

Immunotherapy has produced disappointing results in recurrent ovarian cancer (OC). However, the prognostic value of tumour-infiltrating lymphocytes (TILs) is largely based on the analysis of treatment-naive tumours. To understand the immunobiology of recurrent cancers, and their evolution, we profiled 170 patient-matched primary-recurrent OC samples from 69 patients of two independent cohorts. By capturing heterogeneous TIL distributions, we identified four immune phenotypes associated with differential prognosis, TILs states and TILs:myeloid networks, which dictate malignant evolution after chemotherapy and recurrence. Notably, recurrent tumours recapitulate the immunogenic patterns of original cancers. Mirroring inflamed human OC, preclinical recurrent Brca1mut tumours maintained activated TILs:dendritic cells (DCs) niches and immunostimulatory tumour-associated macrophages (TAMs). Conversely, recurrent Brca1wt tumours displayed loss of TILs:DCs niches and accumulated immunosuppressive myeloid networks featuring Trem2/ApoEhigh TAMs and Nduf4l2high/Galectin3high malignant states. Our study highlights that persistent immunogenicity in recurrent OC is governed by the crosstalk between dissimilar myeloid cells and TILs, which is BRCA-dependent.

immunology↗

Inflammatory state of lymphatic vessels and miRNA profiles associated with relapse in ovarian cancer patients

Lymphogenic spread is associated with poor prognosis in epithelial ovarian cancer (EOC), yet little is known regarding roles of non-peri-tumoural lymphatic vessels (LVs) outside the tumour microenvironment that may impact relapse. The aim of this feasibility study was to assess whether inflammatory status of the LVs and/or changes in the miRNA profile of the LVs have potential prognostic and predictive value for overall outcome and risk of relapse. Samples of macroscopically normal human lymph LVs (n=10) were isolated from the external iliac vessels draining the pelvic region of patients undergoing debulking surgery. This was followed by quantification of the inflammatory state (low, medium and high) and presence of cancer-infiltration of each LV using immunohistochemistry. LV miRNA expression profiling was also performed, and analysed in the context of high versus low inflammation, and cancer-infiltrated versus non-cancer-infiltrated. Results were correlated with clinical outcome data including relapse with an average follow-up time of 13.3 months. The presence of a high degree of inflammation correlated significantly with patient relapse (p=0.033). Cancer-infiltrated LVs showed a moderate but non-significant association with relapse (p=0.07). Differential miRNA profiles were identified in cancer-infiltrated LVs and those with high versus low inflammation. In particular, several members of the let-7 family were consistently down-regulated in highly inflamed LVs (>1.8-fold, p<0.05) compared to the less inflamed ones. Down-regulation of the let-7 family appears to be associated with inflammation, but whether inflammation contributes to or is an effect of cancer-infiltration requires further investigation.

cancer biology↗