Mapping high-grade glioma immune infiltration to 5-ALA fluorescence levels: TCGA data computation, classical histology, and digital image analysis

Abstract

Purpose Resection of high-grade gliomas has been considerably improved by 5-aminolevulinic acid (5-ALA). However, not all neurobiological properties of 5-ALA are fully understood. Specifically, potential differences in immune infiltration have not been conclusively examined, despite recent reports that immune cells might play a role. Thus, we here provide a systematic mapping of immune infiltration of different 5-ALA fluorescence levels. Methods Tumor-associated macrophages (CD68, CD163), cytotoxic T cells (CD8), and regulatory T cells (FoxP3) were quantified via three methods. First, data from The Cancer Genome Atlas (TCGA) of 172 patients was examined for correlations between 5-ALA fluorescence-related mRNA expression signatures and immune markers. Second, as classical histology, 508 stained slides from 39 high-grade glioma patients were analysed semi-quantitatively by two independent reviewers, generating 1016 data points. Third, digital image analysis was performed with automated scanning and algorithm-based cell quantification. Results TCGA mRNA data from 172 patients showed a direct, significant correlation between 5-ALA signatures and immune markers (p < 0.001). However, we were not able to confirm this finding in the here studied initial set of 39 patient histologies where we found a comparable immune infiltration in different fluorescence levels. Digital image analysis correlated excellently with standard histology. Conclusion With mapping the immune infiltration pattern of different 5-ALA categories, we are adding fundamental basic insights to the field of 5-ALA and glioma biology. The observation that a significant correlation in TCGA data did not fully translate to detectable differences in immune infiltration in first histology data warrants further investigation in larger cohorts.

Fluorescence and immune-cell infiltration of nonneoplastic, postbrachytherapy brain tissue in 5-ALA–guided resection of recurrent anaplastic meningioma: illustrative case

“… 17 A recent study of high-grade glioma gene expression found that tumors with strong 5-ALA fluorescence–related mRNA expression signatures had greater immune cell infiltration, with tumor-associated macrophages as the predominant cell type. 18 This evidence also supports the hypothesis that a portion of tumor fluorescence after 5-ALA administration may be related to myeloid cells such as tumor-associated macrophages that are present in the tumor rather than to tumor-intrinsic fluorescence. Tumor-associated macrophages have previously been shown to make up a large portion of gliomas 19 , 20 and were recently shown to also be present in meningiomas, where they can represent nearly 20% of the tumor.…”

Clinical Application of the Association between Genetic Alteration and Intraoperative Fluorescence Activity of 5-Aminolevulinic Acid during the Resection of Brain Metastasis of Lung Adenocarcinoma

“…To the best of our knowledge, few comprehensive studies have demonstrated the genetic and molecular characteristics of infiltrative BM according to the fluorescence activity of 5-ALA. Several studies have focused on the role of 5-ALA in glioblastoma [ 41 , 42 , 43 , 44 ]. For example, negative 5-ALA fluorescence has been reported to promote temozolomide resistance [ 41 ], and different patterns of immune infiltration have been found according to 5-ALA signatures by analyzing TCGA mRNA data [ 42 , 43 ].…”

“…To the best of our knowledge, few comprehensive studies have demonstrated the genetic and molecular characteristics of infiltrative BM according to the fluorescence activity of 5-ALA. Several studies have focused on the role of 5-ALA in glioblastoma [ 41 , 42 , 43 , 44 ]. For example, negative 5-ALA fluorescence has been reported to promote temozolomide resistance [ 41 ], and different patterns of immune infiltration have been found according to 5-ALA signatures by analyzing TCGA mRNA data [ 42 , 43 ]. Positive 5-ALA gene signatures, which were analyzed by spatially resolved bulk RNA profiling, showed transcriptionally concordant glioblastoma and myeloid cells with mesenchymal subtype to be associated with poor survival and recurrence of glioblastoma [ 44 ].…”

“…However, the present study analyzed a relatively small number of genes included in the NGS panel (323 genes). Moreover, we did not analyze whole genes extensively, as Lang et al [ 42 ] used TCGA mRNA data to determine the association between 5-ALA signatures and immune cell infiltration into glioblastoma. The ONCOaccuPanel ® used in this study did not include many genes associated with immuno-oncology, which is an emerging concern in cancer biology research.…”