Tissue-based biomarkers and prognosticators in nasopharyngeal carcinoma

Abstract

The histological classification of nasopharyngeal carcinoma (NPC) (keratinizing and Epstein-Barr virus (EBV)-related non-keratinizing subtypes) does not take into account the underlying heterogeneity in molecular processes and tumor micro-environment. Also, treatment decisions for NPC are made on stage and not subtype. This impedes better understanding of NPC subtypes, but also development of more precise and tailored treatment and follow-up strategies. We found that all patients regardless of NPC subtype have a higher risk of developing second primary malignancies (SPM), however, SPM were especially prevalent in keratinizing NPC. The differences in risks suggest differences in tumor biology. What we also see is that positive EBV-status is not related to development of other EBV-related cancers. Thus, other underlying (genetic) defects/risk factors are probably also important, as there still is a higher risk of SPM in non-keratinizing/EBV positive NPC patients. We also concluded that close clinical follow-up of patients with all NPC subtypes, with specific attention on the detection of SPM, is warranted. We also studied the prognostic effect of molecules in the EGFR-pathway in NPC, because EGFR is strongly induced by LMP1-CTAR1, a viral oncogene. We found overexpression of EGFR to predict worse survival. We additionally assessed gene copy number alterations between EBV positive and negative NPCs and see that they are distinct entities with CCNL1 gain/amplification conferring worse prognosis in EBV positive NPC. We also show EBV positive and negative NPCs to have important differences in hypermethylation patterns, with hypermethylation of RASSF1A being more prevalent and of prognostic importance in EBV positive NPC. EBV negative NPC correlated strongly with APC hypermethylation which has been described in other (smoking/alcohol related) HNSCC suggesting a common pathogenesis. We also assessed the tumor-microenvironment. There are differences in tumor-infiltrating lymphocytes (TIL) between NPC subtypes. We found that not only the amount, but also the composition of the infiltrate is important for a positive effect on survival. In the EBV positive NPC CD4 and CD8 seem to predict better OS despite PDL1 expression, and in EBV negative NPC absence of CD8 seems to predicts worse OS despite also less PD1 expression. Thus, immunotherapy can possibly also be of use in treating patients with NPC. In addition, there were differences in tumor-associated macrophages between EBV positive and negative NPC with also functional differences in the macrophages. The macrophage infiltrate in EBV negative NPC was unable to recruit Tregs. Also, a higher FOXP3 count seems to be an independent prognostic factor for better OS in the whole NPC group. Finally, we studied specific characteristics in EBV infected NPC and found a correlation between positive EBV status and immunohistochemical SSTR2A positivity. Thus, 68Ga-DOTA-TOC PET/CT can be used in the follow-up of patients with EBV positive NPC. Also, treatment with (177Lu) DOTA-TATE and/ or (90)Y-DOTATOC, and somatostatin analogues needs study. As we have shown differences in risks of developing SPMs, genetics, epigenetics, but also the tumor micro-environment and novel markers, we strongly suggest that the various NPC subtypes are different entities, and require different approaches to treatment and follow-up.