Detailed Characterization of Immune Cell Infiltrate and Expression of Immune Checkpoint Molecules PD-L1/CTLA-4 and MMR Proteins in Testicular Germ Cell Tumors Disclose Novel Disease Biomarkers

Abstract

Background: The immune infiltrate plays an important part in testicular germ cell tumors, but it remains scarcely studied. We aimed at thoroughly characterizing the immune infiltrate and expression of immune checkpoints PD-L1/CTLA-4 and mismatch repair (MMR) proteins in these neoplasms, seeking for associations with patient outcome. Methods: A total of 162 consecutively diagnosed patients (2005–2018) were included. Immunostaining for PD-L1, CTLA-4 and MMR proteins was independently assessed both in immune cells (ICs) and tumor cells (TCs) of primary tumors and metastases, and characterization of IC populations was pursued. Results: PD-L1 and CTLA-4 positivity in ICs was frequent (85.5% and 96.3%). Patients with absent PD-L1 positive ICs exhibited significantly worse relapse-free survival (hazard ratio = 4.481, 95% CI 1.366–14.697, p = 0.013), both in univariable and multivariable analysis. Lower CD20 and CD3 IC infiltration in seminomas associated with higher disease stage (p = 0.0216, p = 0.0291). CTLA-4 TC intensity was significantly higher in yolk sac tumor, choriocarcinoma and teratoma, while PD-L1 TC positivity was significantly more frequent in choriocarcinoma. Both PD-L1 and CTLA-4 immunoexpression in ICs of metastatic samples was frequent (100% and 88.2%). MMR proteins were differentially expressed among the different tumor subtypes. Conclusions: Immune infiltrate/checkpoints associate with patients’ outcome, constituting novel (potentially targetable) disease biomarkers.

Immunotherapy in Testicular Germ Cell Tumors

“…Characterization of the immune infiltrate in terms of immune checkpoint PD-L1/CTLA-4 expression and its correlation with patient outcome was carried out by Lobo et al A predominant portion of 162 analyzed samples exhibited PD-L1 and CTLA-4 positivity in infiltrating immune cells (ICs) regardless of the histological subtype, while the expression of CTLA-4 in tumor cells (TCs) was significantly higher in yolk sac tumor, choriocarcinoma, and teratoma samples. The most frequent PD-L1 TC positivity was determined in choriocarcinomas (28). These results are in line with data from Sadigh et al, where choriocarcinoma was the only GCT histological subtype expressing PD-L1 in tumor cells, whereas other subtypes primarily expressed varying levels of PD-L1 on tumor-associated macrophages (TAMs) without true PD-L1 expression on tumor cells themselves.…”

“…In addition, a similar pattern was also determined for CTLA-4 immunoexpression in ICs. Namely, higher CTLA-4 expression correlated with good prognostic features, including lymphovascular invasion and lower pT and N stages (28). Moreover, Yamada et al showed the prognostic value of CD66b + tumor-infiltrating neutrophils (TINs) in 102 patients who underwent orchiectomy due to testicular cancer.…”

Efficacy of HDAC Inhibitors Belinostat and Panobinostat against Cisplatin-Sensitive and Cisplatin-Resistant Testicular Germ Cell Tumors

“…As the staining was overall diffuse in all cases, but with great variation in staining intensity, we focused on the latter. The intensity of staining was considered as “weak”, “moderate”, and “strong”, as previously defined [ 65 ].…”

Prediction of relapse in stage I testicular germ cell tumor patients on surveillance: investigation of biomarkers

“…Both the intensity (0, 1, 2 and 3, corresponding to absent, weak, intermediate and strong staining) and percentage of stained tumor cells were assessed independently, as done in previous studies assessing these markers. "weak", "intermediate" and "strong" was defined as indicated in [28]. Overall assessment was reported for mixed tumors, since 17/23 consisted of tumors with > 90% embryonal carcinoma, with only small foci of cells from other distinct subtypes.…”