Leoma Bere

Myeloid cells are highly prevalent in glioblastoma (GBM), existing in a spectrum of phenotypic and activation states. We now have limited knowledge of the tumor microenvironment (TME) determinants that influence the localization and the functions of the diverse myeloid cell populations in GBM. Here, we have utilized orthogonal imaging mass cytometry with single-cell and spatial transcriptomic approaches to identify and map the various myeloid populations in the human GBM tumor microenvironment (TME). Our results show that different myeloid populations have distinct and reproducible compartmentalization patterns in the GBM TME that is driven by tissue hypoxia, regional chemokine signaling, and varied homotypic and heterotypic cellular interactions. We subsequently identified specific tumor subregions in GBM, based on composition of identified myeloid cell populations, that were linked to patient survival. Our results provide insight into the spatial organization of myeloid cell subpopulations in GBM, and how this is predictive of clinical outcome.

NF2 -related Schwannomatosis ( NF2 SWN) is a rare tumour-predisposition syndrome characterised by the growth of multiple central and peripheral nervous system neoplasms. The drivers of NF2 SWN are pathogenic variants in the tumour suppressor gene NF2 , encoding the protein Merlin, leading to development of bilateral vestibular schwannoma (VS) in >95% of patients. VS tumours are characterised by infiltration of myeloid cells and lymphocytes, highlighting the potential of immunotherapy for VS. However, the immunological landscape in VS and the spatial determinants within the tumour microenvironment that shape the trajectory of disease are presently unknown. In this study, to elucidate the complex immunological networks across VS, we performed imaging mass cytometry (IMC) on clinically annotated VS samples from NF2 SWN patients. We reveal the heterogeneity in neoplastic cell, myeloid cell and T cell populations that co-exist within VS, determining that the cellular composition of VS tumours is independent of NF2 -SWN genetic severity. We show that distinct myeloid cell and Schwann cell populations exist within varied spatial contextures across characteristic Antoni A and B histomorphic niches. Interestingly, we show that T-cell populations associate with tumour-associated macrophages (TAMs) in Antoni A regions, seemingly limiting their ability to interact with tumorigenic Schwann cells. This spatial landscape is altered in Antoni B regions, where T-cell populations appear to interact with PD-L1+ Schwann cells. We also demonstrate that prior bevacizumab treatment (VEGF-A antagonist) preferentially reduces alternatively-activated TAMs, whilst enhancing CD44 expression, in bevacizumab-treated tumours. Together, we describe niche-dependent modes of T-cell regulation in NF2 SWN VS, indicating the potential for microenvironment-altering therapies for VS. Teaser: Imaging mass cytometry and spatial omic analyses illustrate spatially-distinct regions of T-cell regulation in vestibular schwannoma.

Natural killer (NK) cells can kill infected or transformed cells via a lytic immune synapse. Diseased cells may exhibit altered mechanical properties but how this impacts NK cell responsiveness is unknown. We report that human NK cells were stimulated more effectively to secrete granzymes A and B, FasL (also known as FasLG), granulysin and IFNγ, by stiff (142 kPa) compared to soft (1 kPa) planar substrates. To create surrogate spherical targets of defined stiffness, sodium alginate was used to synthesise soft (9 kPa), medium (34 kPa) or stiff (254 kPa) cell-sized beads, coated with antibodies against activating receptor NKp30 (also known as NCR3) and the integrin LFA-1 (also known as ITGAL). Against stiff beads, NK cells showed increased degranulation. Polarisation of the microtubule-organising centre and lytic granules were impaired against soft targets, which instead resulted in the formation of unstable kinapses. Thus, by varying target stiffness to characterise the mechanosensitivity of immune synapses, we identify soft targets as a blind spot in NK cell recognition. This article has an associated First Person interview with the co-first authors of the paper.