Immune System-Related Changes in Preclinical GL261 Glioblastoma under TMZ Treatment: Explaining MRSI-Based Nosological Imaging Findings with RT-PCR Analyses

Abstract

Glioblastomas (GB) are brain tumours with poor prognosis even after aggressive therapy. Previous work suggests that magnetic resonance spectroscopic imaging (MRSI) could act as a biomarker of efficient immune system attack onto GB, presenting oscillatory changes. Glioma-associated microglia/macrophages (GAMs) constitute the most abundant non-tumour cell type within the GB and can be polarised into anti-tumour (M1) or pro-tumour (M2) phenotypes. One of the mechanisms to mediate immunosuppression in brain tumours is the interaction between programmed cell death-1 ligand 1 (PD-L1) and programmed cell death-1 receptor (PD-1). We evaluated the subpopulations of GAMs in responding and control GB tumours to correlate PD-L1 expression to GAM polarisation in order to explain/validate MRSI-detected findings. Mice were evaluated by MRI/MRSI to assess the extent of response to treatment and with qPCR for GAMs M1 and M2 polarisation analyses. M1/M2 ratios and PD-L1 expression were higher in treated compared to control tumours. Furthermore, PD-L1 expression was positively correlated with the M1/M2 ratio. The oscillatory change in the GAMs prevailing population could be one of the key causes for the differential MRSI-detected pattern, allowing this to act as immune system activity biomarker in future work.

Establishing Imaging Biomarkers of Host Immune System Efficacy during Glioblastoma Therapy Response: Challenges, Obstacles and Future Perspectives

“…Figure 1 summarizes those detected changes. Our results [ 43 , 56 ] show that in a fast-growing GL261 GB tumor before treatment, about 68% of the tissue volume is occupied by fast proliferating tumor cells (average 2900 Ki67+ cells/mm 2 ) and other non-tumor, non Iba1+ cell types, such as myeloid-derived suppressor cells (MDSC) [ 28 ]. The rest of the volume is mostly represented by innate Iba1+ IS cells (microglia/macrophages, 12%), lymphocytes (below 1%), and acellular spaces (also dubbed by other authors microcystic volume [ 58 ]) (19%).…”

“…Microglia/macrophages under different polarization statuses also have different metabolomic patterns [ 54 , 63 ]. In this respect, changes in the microglia/macrophage polarization (M1 vs. M2) have been described in both GB [ 64 , 65 , 66 , 67 ] and, more precisely, in GL261 GB [ 56 , 68 , 69 , 70 ]. Then, the composition of the cell types, their status and tissue architecture, and the changes upon treatment can together contribute to metabolome pattern changes that lead to measurable changes in the TRI ( Figure 1 B).…”

“…Right, a representative TMZ-treated, transiently responding case (C1458, 23 days p.i.) from [ 56 ]. Please note that each studied mouse was identified in the cited studies with a unique code (CXXXX) where XXXX was a correlative number.…”

“…Acellular spaces (dark blue solid line), the Ki67-positive stained area (purple dashed line), and the Iba-1 stained area (brown dashed line) were estimated from fields of histopathological slides from high-level response ( n = 27 fields) and control cases ( n = 40 fields) described in [ 43 ]. M1 and M2 macrophages’ gene level expression changes (light blue and pink dashed lines) were calculated from NOS2 and CD206 obtained from IMS-TMZ treated and control tumor samples ( n = 10 each) described by us in [ 56 ]. ( ii ) Schema of cancer immune cycle (reproduced with permission from [ 57 ]) starting from (A) treatment (e.g., in this example, IMS-TMZ), which triggers immunogenic cell death/damage, (B) release of DAMPs (CRT, calreticulin) [ 72 ] and antigen presentation to host immune system cells, followed by (C) T-cell priming and proliferation, which (D) infiltrates the tumor site and kills tumor cells.…”

Efferocytosis in the Central Nervous System

“…Studies have shown that efferocytosis is closely related to the polarization of the M1/M2 phenotype. Unlike nonneoplastic diseases, the inflammatory defense function of macrophages/microglia can be inhibited in the glioma microenvironment, where they cells are changed from the M1 phenotype (proinflammatory) to the M2 phenotype (anti-inflammatory) ( Calero-Pérez et al, 2021 ). The mainstream view is that anti-inflammatory substances released after efferocytosis induce microglia to polarize to the M2 type.…”