Autophagic Markers in Chordomas: Immunohistochemical Analysis and Comparison with the Immune Microenvironment of Chordoma Tissues

Abstract

Chordomas are notably resistant to chemotherapy. One of the cytoprotective mechanisms implicated in chemoresistance is autophagy. There are indirect data that autophagy could be implicated in chordomas, but its presence has not been studied in chordoma tissues. Sixty-one (61) chordomas were immunohistochemically studied for autophagic markers and their expression was compared with the expression in notochords, clinicopathological data, as well as the tumor immune microenvironment. All chordomas strongly and diffusely expressed cytoplasmic p62 (sequestosome 1, SQSTM1/p62), whereas 16 (26.2%) tumors also showed nuclear p62 expression. LC3B (Microtubule-associated protein 1A/1B-light chain 3B) tumor cell expression was found in 44 (72.1%) tumors. Autophagy-related 16‑like 1 (ATG16L1) was also expressed by most tumors. All tumors expressed mannose-6-phosphate/insulin-like growth factor 2 receptor (M6PR/IGF2R). LC3B tumor cell expression was negatively associated with tumor size, while no other parameters, such as age, sex, localization, or survival, were associated with the immunohistochemical factors studied. LC3B immune cell expression showed a significant positive association with programmed death-ligand 1 (PD-L1)+ immune cells and with a higher vascular density. ATG16L1 expression was also positively associated with higher vascular density. Notochords (n = 5) showed different immunostaining with a very weak LC3B and M6PR expression, and no p62 expression. In contrast to normal notochords, autophagic factors such as LC3B and ATG16L1 are often present in chordomas, associated with a strong and diffuse expression of p62, suggesting a blocked autophagic flow. Furthermore, PD-L1+ immune cells also express LC3B, suggesting the need for further investigations between autophagy and the immune microenvironment.

Loss of SMARCB1 promotes autophagy and facilitates tumour progression in chordoma by transcriptionally activating ATG5

“… 39 Additionally, a recent IHC analysis showed positive expression of autophagic markers LC3B, P62 and ATG16L1 in chordoma compared with normal notochords, which revealed the potential oncogenic role of autophagy in chordoma. 32 Consistent with prior observations, we found positive expression of ATG5, a vital ATG during autophagy, in chordoma, especially recurrent chordoma, and identified ATG5 as a novel adverse prognostic factor of chordoma patients. Additionally, our in vitro data suggested impairments of proliferation, migration and invasion of chordoma cells after blocking autophagy by ATG5 siRNA or autophagy inhibitor CQ, revealing the tumour promotive effect of autophagy in chordoma.…”

“…The IHC score of ATG5 was evaluated under a Leica Aperio AT2 scanner and defined as the product of the score of positive tumour cells as a percentage (0, no positive tumour cells; 1+, less than 25% cells; 2+, 25%‐50% cells; 3+, >50% cells) × the score of the staining intensity (0, no staining; 1+, weak, light yellow; 2+, moderate, yellow‐brown; 3+, strong, brown). 31 , 32 Low ATG5 expression was defined as an IHC score of <4, and IHC scores of ≥4 were defined as high ATG5 expression.…”

PD-L1 Status in Gastric Cancers, Association with the Transcriptional, Growth Factors, AKT/mTOR Components Change, and Autophagy Initiation

“…The pattern of proteins affecting immunity includes autophagic proteins. It is known that the PD-L1 positive cells express LC3B, suggesting the need for further investigations regarding autophagy and the immune microenvironment [10][11][12].…”

“…The pattern of proteins affecting immunity includes autophagic proteins. It is known the PD-L1 positive cells express LC3B, suggesting the need for further investigation into the modulation of the immune microenvironment [10]. However, misfolded proteins, damaged mitochondria, and other unwanted components in cells can be decomposed and reused via autophagy in some specific cases (including hypoxic stress, low energy states, or nutrient deprivation) [11].…”

Primary central nervous system lymphomas express immunohistochemical factors of autophagy

“…The antibodies were initially tested in a large variety of normal and neoplastic tissues to decide the best immunohistochemical protocol giving no background staining and a range of staining intensities 14 . Thereafter, nerve fibers and normal tonsillar tissue were used as positive controls for LC3B and p62 15 , respectively, as well as tonsillar tissue for M6PR, while omission of the primary antibody was used as a negative control 14 . Normal brain tissue also served as an internal positive control for LC3B and p62 in the current series.…”