Orsolya Rajky

Programmed death ligand 1 expression and tumor-infiltrating lymphocytes in glioblastoma

BACKGROUND Immune checkpoint inhibitors targeting programmed cell death 1 (PD1) or its ligand (PD-L1) showed activity in several cancer types. METHODS We performed immunohistochemistry for CD3, CD8, CD20, HLA-DR, phosphatase and tensin homolog (PTEN), PD-1, and PD-L1 and pyrosequencing for assessment of the O6-methylguanine-methyltransferase (MGMT) promoter methylation status in 135 glioblastoma specimens (117 initial resection, 18 first local recurrence). PD-L1 gene expression was analyzed in 446 cases from The Cancer Genome Atlas. RESULTS Diffuse/fibrillary PD-L1 expression of variable extent, with or without interspersed epithelioid tumor cells with membranous PD-L1 expression, was observed in 103 of 117 (88.0%) newly diagnosed and 13 of 18 (72.2%) recurrent glioblastoma specimens. Sparse-to-moderate density of tumor-infiltrating lymphocytes (TILs) was found in 85 of 117 (72.6%) specimens (CD3+ 78/117, 66.7%; CD8+ 52/117, 44.4%; CD20+ 27/117, 23.1%; PD1+ 34/117, 29.1%). PD1+ TIL density correlated positively with CD3+ (P < .001), CD8+ (P < .001), CD20+ TIL density (P < .001), and PTEN expression (P = .035). Enrichment of specimens with low PD-L1 gene expression levels was observed in the proneural and G-CIMP glioblastoma subtypes and in specimens with high PD-L1 gene expression in the mesenchymal subtype (P = 5.966e-10). No significant differences in PD-L1 expression or TIL density between initial and recurrent glioblastoma specimens or correlation of PD-L1 expression or TIL density with patient age or outcome were evident. CONCLUSION TILs and PD-L1 expression are detectable in the majority of glioblastoma samples but are not related to outcome. Because the target is present, a clinical study with specific immune checkpoint inhibitors seems to be warranted in glioblastoma.

Invasion patterns in brain metastases of solid cancers

BACKGROUND Brain metastases are generally considered to be well demarcated from the surrounding brain parenchyma, although infiltrative growth patterns have been observed. We systemically investigated infiltration patterns and expression of adhesion molecules in a large and well-defined series of autopsy cases of brain metastases. METHODS Ninety-seven autopsy specimens from 57 brain metastasis patients (primary tumor: 27 lung cancer, 6 breast cancer, 8 melanoma, 2 colorectal cancer, 1 kidney cancer, and 13 other) were evaluated for patterns of invasion into surrounding brain parenchyma. Expression of integrins αv; cytoplasmic β3, αvβ3, αvβ5, αvβ6, and αvβ8; and of E and N cadherin were evaluated using immunohistochemistry. RESULTS Three main invasion patterns were seen: well-demarcated growth (29/57, 51%), vascular co-option (10/57, 18%), and diffuse infiltration (18/57, 32%). There was no statistically significant association of invasion pattern with primary tumor type, although vascular co-option was most common in melanoma brain metastases (4/10). Invasion patterns of different brain metastases of the same patient were highly concordant (P < .001, chi-square test). Distance of infiltration from the main tumor mass ranged from 12.5 µm to 450 µm (median 56.2 µm) and was not significantly different between the vascular co-option and the diffuse infiltration groups. Levels of αvβ6 were significantly higher in the well-demarcated group than in the vascular co-option and the diffuse infiltration groups (P = .033, Kruskal-Wallis test). Expression of αvβ5 in tumor cells was higher in brain metastasis lesions previously treated with stereotactic radiosurgery (P = .034, chi-square test). CONCLUSIONS Distinct invasion patterns of brain metastases into the brain parenchyma are not specific for primary tumor types, seem to be influenced by expression of αv integrin complexes, and may help to guide clinical decision-making.

PD1 (CD279) and PD-L1 (CD274, B7H1) expression in primary central nervous system lymphomas (PCNSL).

BACKGROUND Primary central nervous system lymphoma (PCNSL) is a malignant brain tumor with limited treatment options and shows prominent infiltration by tumor infiltrating lymphocytes (TILs) and tumor-associated macrophages (TAMs). Programmed death 1 (PD1; CD279) and its ligand PD-L1 (B7H1, CD274) promote escape of tumor cells from immune surveillance in several tumor types, but no data are available on PCNSL. Agents inhibiting PD1 and PD-L1 are showing compelling antitumor activity in current clinical trials in solid and hematological cancers. METHODS We investigated PD1 (clone NAT ab52587) and PD-L1 (clone 5H1) expression in large neurosurgical resection specimens of 20 immunocompetent historical PCNSL patients using immunohistochemistry. RESULTS We found expression of PD1 and/or PD-L1 on tumor cells, TILs, or TAMs in a total of 18/20 (90%) of PCNSL cases. In 12/20 (60%) cases, intratumoral PD1-positive TILs were present (low density: 9/20, 45%; moderate density: 2/20, 10%; high density: 1/20, 5%) with additional peritumoral accumulation of PD1-positive lymphocytes in all of 12 cases with evaluable adjacent brain tissue on the tissue section. PD-L1 expressing intratumoral TAMs were found in 4/20 (20%) tumors. In 2/20 (10%) and 4/20 (20%) specimens, we observed striking PD-L1 or PD1 expression on PCNSL tumor cells, respectively. Median number of CD8-positive TILs was 517/mm2 (range 75 - 2,470) and did not correlate with PD1 or PD-L1 expression (p > 0.05, Kruskal- Wallis test). CONCLUSIONS PD1 and PDL1 are immunohistochemically detectable in PCNSL and may be involved in creating an immunosuppressive microenvironment. Specific immune checkpoint inhibitors may be considered for experimental therapy approaches in this disease.

Tumour-infiltrating lymphocytes and expression of programmed death ligand 1 (PD-L1) in melanoma brain metastases

In this study we aimed to characterize immune infiltrates and expression of programmed death 1 (PD‐1) and programmed death ligand 1 (PD‐L1) in a series of melanoma BM to provide a basis for experimental therapy using immune checkpoint inhibitors.

Correlation of immune phenotype with IDH mutation in diffuse glioma

Background Tumor infiltrating lymphocytes (TILs) and programmed death ligand 1 (PD-L1) are targets of immune checkpoint inhibitors. Methods Forty-three World Health Organization (WHO) grade II/III gliomas (39 IDH-mutant [mut], 4 IDH-wildtype [wt]) and 14 IDH-mut glioblastomas (GBM) were analyzed for TIL (CD3+; PD1+) infiltration and PD-L1 expression. Results were compared with the data of a previously published series of 117 IDH-wt glioblastomas. PD-L1 gene expression levels were evaluated in 677 diffuse gliomas grades II-IV from The Cancer Genome Atlas (TCGA) database. Results TIL and PD-L1 expression were observed in approximately half of WHO grade II/III gliomas. IDH-wt status was associated with significantly higher TIL infiltration and PD-L1 expression among all (grades II-IV) cases (n = 174, P < 0.001) and within the cohort of glioblastomas (n = 131, P < 0.001). In low-grade glioma (LGG) and glioblastoma cohorts of TCGA, significantly higher PD-L1 gene expression levels were evident in IDH-wt compared with IDH-mut samples (LGG: N = 516; P = 1.933e-11, GBM: N = 161; P < 0.009). Lower PD-L1 gene expression was associated with increased promoter methylation (Spearman correlation coefficient -0.36; P < 0.01) in the LGG cohort of TCGA. IDH-mut gliomas had higher PD-L1 gene promoter methylation levels than IDH-wt gliomas (P < 0.01). Conclusions The immunological tumor microenvironment of diffuse gliomas differs in association with IDH mutation status. IDH-wt gliomas display a more prominent TIL infiltration and higher PD-L1 expression than IDH-mut cases. Mechanistically this may be at least in part due to differential PD-L1 gene promoter methylation levels. Our findings may be relevant for immune modulatory treatment strategies in glioma patients.

PD-1 and PD-L1 expression in HNSCC primary cancer and related lymph node metastasis - impact on clinical outcome

Expression profiles and clinical impact of programmed cell death ligand 1 (PD‐L1) and programmed cell death 1 (PD‐1) expressing tumour infiltrating lymphocytes (TILs) in head and neck squamous cell carcinoma (HNSCC) are not elucidated fully. This study evaluates expression patterns in primary HNSCC and related lymph node metastasis and the impact on patients’ clinical outcome.