Biology subjects

Minshull, J.

Publications and source records attributed to Minshull, J..

3 recordsLinked to original sources

bioRxiv2025

Cell Line Development for Bispecific Antibodies: Better Predictability Through Transposases

Bispecific antibodies are at the forefront of biopharmaceutical drug development. With over 100 different molecular architectures combined with diverse individual subunit sequences, choosing the most suitable structure and predicting the ideal subunit expression ratios for successful heterodimerization is a significant challenge. In this paper, we demonstrate that the recently described cell line development paradigm shift (Rajendran et al. 2021), enabled by the Leap-In transposon platform, can be extended to the development of bispecific monoclonal antibody-producing cell substrates (stable clones and pools). The key features are 1) Parental pools reliably predict the derivative clonal productivity and clonal heterodimer fractions. 2) Clonal productivity and clonal heterodimer fraction remained stable for at least 60 population doublings. 3) Depending on the products biophysicochemical properties, the stable pools exhibit variable productivity stability. 4) Heterodimer fractions remain stable in the Leap-In mediated stable pools independently of the productivity stability of the pools. 5) Structures and subunit ratios can be triaged at stable pool level, and 6) Due to the homogeneous clonal productivity distribution, only a small number ([~]50) of clones need to be isolated and characterized.

bioengineering↗

bioRxiv2023

Hypoxia coordinates the spatial landscape of myeloid cells within glioblastoma to affect outcome

Myeloid cells are highly prevalent in glioblastoma (GBM), existing in a spectrum of phenotypic and activation states. We currently have limited knowledge of the tumour microenvironment (TME) determinants that influence the localisation and the functions of the diverse myeloid cell populations in GBM. Here we have utilised orthogonal imaging mass cytometry with single cell and spatial transcriptomics approaches to identify and map the various myeloid populations in the human GBM tumour microenvironment (TME). Our results show that different myeloid populations have distinct and reproducible compartmentalisation patterns in the GBM TME that is driven by tissue hypoxia, regional chemokine signalling, and varied homotypic and heterotypic cellular interactions. We subsequently identified specific tumour sub-regions in GBM, based upon composition of identified myeloid cell populations, that were linked to patient survival. Our results provide new insight into the spatial organisation of myeloid cell sub populations in GBM, and how this is predictive of clinical outcome. TeaserMulti-modal mapping reveals that the spatial organisation of myeloid cells in glioblastoma impacts disease outcome.

cancer biology↗

bioRxiv2022

Novel Laser Capture Microdissection-Proteomic Analysis Identifies Spatially Distinct Extracellular Matrix Signatures in the Core and Infiltrating Edge of Human Glioblastoma.

BackgroundGlioblastoma is the most common and aggressive primary brain tumour in adults. Hallmarks of glioblastoma include its intra-tumoural heterogeneity and extensive invasion of the surrounding brain. Glioblastoma is known to remodel the extracellular matrix (ECM) of the brain, resulting in altered mechanical properties and the establishment of a tumour-promoting microenvironment. How changes in the expression and spatial distribution of ECM constituents within glioblastoma contribute to invasion and disease progression is still unclear. MethodsHere we report on a protocol for laser-capture microdissection coupled with mass spectrometry (LCM-proteomics) that allowed a spatially resolved and unbiased analysis of the regional ECM proteome (matrisome) in formalin-fixed and paraffin-embedded (FFPE) samples of human glioblastoma. We investigated five molecularly characterised hemispheric adult glioblastomas where the brain/tumour interface and tumour epicentre were represented in the surgical specimens and snap-frozen tissue was available. LCM-proteomic analysis was validated with immunohistochemistry. ResultsLCM-proteomics identified 53 matrisome proteins in FFPE tissue, demonstrating comparable performance with conventional analysis of snap-frozen tissue. The analysis revealed distinct matrisome components in the brain/tumour interface versus the tumour epicentre. Guided by data from LCM-proteomic analysis, immunostaining for tenascin-R confirmed greater staining in the brain/tumour interface, whilst expression of fibronectin was higher in the tumour epicentre. ConclusionThe protocol described in this work allowed for accurate, spatially resolved analysis of ECM in FFPE tissues, with performance comparable to analysis of snap-frozen tissue. While the focus for this work was on the regional ECM composition of glioblastoma, we found that the LCM-proteomics protocol is also applicable to the study of the wider proteome, and represents a valuable tool for investigating tumour/tissue heterogeneity. This protocol opens the possibility to apply LCM-proteomics to retrospective studies with the advantage of accessing clinical history and follow-up information, providing a valuable resource for translational research in glioblastoma.

cancer biology↗