High Amount of PD-L1+ Tumor Associated Macrophages Is Associated with Better Survival in Patients with Primary Testicular Lymphoma

Introduction:

Primary testicular lymphoma (PTL) is a rare and aggressive extranodal lymphoid malignancy with poor outcome to therapy and largely uncharacterized biology. Majority of PTLs are diffuse large B-cell lymphomas (DLBCL). Tumor associated macrophages (TAMs) are known to have prognostic impact in DLBCLs, but their role in PTLs has not been addressed. The aim of this study was to examine the association of TAMs with disease characteristics and survival in patients with PTL.

Patients and methods:

We identified 74 patients with PTL, all representing DLBCL histologically. Tissue microarray (TMA) was constructed from PTL samples and TMA slides were stained using multiplex immunohistochemistry (mIHC), allowing a detection of four markers (CD68, cMAF, PD-L1 and PD-L2) and nuclei simultaneously. Expression profiles were analyzed quantitatively with the image analysis software Cell Profiler. In addition, CD68, PD-L1, and PD-L2 mRNA levels were measured from 60 PTL samples.

Results:

For the 74 patients in the mIHC cohort, median age at diagnosis was 70 years (36-92 years) and median follow-up 67 months (range 6.7 to 120 months). Majority of the patients represented non-germinal center B-cell (non-GCB) phenotype (76%), low stage (I-II, 64%), and low/intermediate International Prognostic Index (IPI 0-2, 68%). Thirty-five patients (47%) were treated with rituximab containing chemotherapy and 36 patients (49%) received central nervous system (CNS) prophylaxis, most of them (n=34) with CNS penetrating intravenously administered chemotherapy. Altogether 34 deaths, 24 relapses and 24 lymphoma associated deaths occurred during the follow-up. 5-year overall survival (OS), disease specific survival (DSS) and progression free survival (PFS) rates were 56%, 68%, and 53%, respectively.

The median proportions of PD-L1⁺ cells, CD68⁺, PDL1⁺CD68⁺, CD68⁺cMAF⁺ (M2 type), and PD-L1⁺CD68⁺cMAF⁺ macrophages of all cells in the tumor tissue were 15.3%, 23.7%, 9.1%, 1.3%, 0.5%, respectively. In univariate analyses with continuous variables, the proportion of PD-L1⁺ cells (p=0.039) and PDL1⁺CD68⁺ macrophages (p=0.043) correlated with a favorable OS. A trend towards a better OS was also observed for PD-L1⁺CD68⁺cMAF⁺ (p=0.090) macrophages, whereas neither PD-L1^-CD68⁺cMAF⁺ macrophages nor PD-L1⁺CD68^- cells had an impact on survival. Furthermore, no correlation with survival was found for any of the PD-L2⁺ cells. According to Kaplan-Meier estimates, the patients with the high proportion of PD-L1⁺CD68⁺ macrophages had a 5-year OS of 65% in comparison to 41% for those with the lower counts (p=0.026). The difference was more clear in the subgroup of patients with a stage III-IV disease (n=24, 5-year OS, 67% vs 25%, p=ns). When clinical characteristics of the patients were compared according to PD-L1⁺CD68⁺ cell counts, no significant differences in age or molecular subtype were observed between the subgroups. However, in the high PDL1⁺CD68⁺ group, more patients had stage I-II disease (p<0.001) and low/intermediate IPI score (IPI 0-2, p=0.023). PD-L1 and CD68 gene expressions correlated with corresponding cell counts (for both, r_s≥0.673, p<0.001) and favorable OS (PD-L1, p=0.007; CD68, p=0.016).

Conclusion:

Our data demonstrate a significant variation in TAM contents and their phenotypes in PTL, and an association of PD-L1⁺ TAMs with more localized disease and favorable survival in patients with PTL.