To our knowledge, this is the first study to evaluate the relationship between PD-L1/PD-L2 expression and 18F-FDG uptake on PET in patients with TETs. We found that high expression of PD-L1 and PD-L2 was closely associated with high accumulation of 18F-FDG; in particular, PD-L1/PD-L2 expression levels were significantly correlated with those of glucose metabolism and hypoxia. As angiogenetic markers, VEGFR2 and β2-AR was associated with the expression of PD-L1/PD-L2. Moreover, we confirmed that the expression of PD-L1 and PD-L2 was closely associated with not MTV or TLG but SUVmax on 18F-FDG uptake, confirmed by multivariate analysis. Overall, PD-L1 and PD-L2 expressions indicated a strong correlation with glucose metabolism, as determined by GLUT1. Although SUVmax is closely correlated with MTV and TLG on 18F-FDG uptake, the upregulation of PD-L1 and PD-L2 may play a crucial role in the pathogenesis of tumor glucose metabolism in patients with TETs. Further studies with an experimental approach using thymic tumor cell lines are warranted to elucidate the results of our study.
Several researchers have described that PD-L1 is frequently expressed in TETs, and a WHO classification is closely related to positive PD-L1 expression, but there was some discrepancy regarding the trend for worsened survival 4 16–19. Padda et al. reported that high expression of PD-L1 could predict a significantly worse OS, which was correlated with more aggressive histology 16. However, Yokoyama et al. described that low PD-L1 expression and a high number of PD-1-positive tumor infiltrative lymphocytes (TILs) were significant predictors of worse survival in patients with thymic carcinoma 18. Considering the evidence from previous studies, it is debatable whether PD-L1 could absolutely predict a worse outcome for patients with TETs. As our study also indicated that the expression of PD-L1 was higher in thymic carcinoma than in thymoma, PD-L1 may highly express in human neoplasms with malignant phenotype.
PD-L1 is an important target for PD-1 blockade, whereas PD-L2, as another PD-1 ligand, may also play a crucial role in the inhibition of PD-1 in human neoplasms. A recent study demonstrated that PD-L2 was expressed in all tumor types. The prevalence of PD-L2 was significantly correlated with PD-L1, and PD-L2 status was also a significant predictor of PFS with pembrolizumab, independent of PD-L1 status 20. Although there are some concerns about the clinicopathological significance of PD-L2 expression in human neoplasms, PD-L2 is frequently expressed in tumor cells and seems to play a crucial role in tumor growth and survival, adjusting the immune environment. A previous study reported that GLUT1 expression is associated with better clinical outcomes in advanced-stage classical Hodgkin’s lymphoma and is significantly associated with PD-L1 and PD-L2 expression 21. This study supports the hypothesis that GLUT1-related signaling pathways play an important role in the PD-L1 or PD-L2 pathway. Furthermore, a previous article reported that PD-L2-positive pheochromocytoma and paraganglioma were characterized by higher HIF-1α expression. That study reported the enrichment of transcripts involved in the hypoxic response in relation to PD-L2, but not PD-L1 expression 22. When the researchers considered a broader subset of 200 genes involved in the hypoxic response, PD-L2 upregulation strikingly emerged as a stronger and more substantial determinant of tumor hypoxia than PD-L1, suggesting a potential mechanistic relationship between hypoxia and PD-L2-mediated antitumor immune control. Their data suggest that PD-L2 has a more predominant role than PD-L1 in shaping the immune-tolerogenic environment, given the highly significant association with key pathways involved in innate, adaptive immunity, and inflammation in pheochromocytomas and paragangliomas. However, little is known about the clinicopathological significance of PD-L2 expression in patients with TETs. Recently, Rouquette et al. reported that the PD-L2 antibody stained no tumor epithelial cells in TETs 19. Although we also performed PD-L2 staining using the same antibody, no staining was observed in our study, corresponding to their results 19. Thus, different kinds of PD-L2 clones were explored, and we found the optimal PD-L2 clone for immunohistochemistry. Our investigation is the first study to demonstrate the expression of PD-L2 in TETs. In our study, PD-L2 was closely correlated with the expression of PD-L1 and was associated with histological grade, glucose metabolism, and hypoxia, but not survival.
The aim of our study was to focus on the role of PD-L1/PD-L2 expression as a mechanism underlying 18F-FDG uptake in TETs. Previous investigations have supported the potential of PD-L1 as an alternative target of HIF-1α and suggested that the distribution of glucose metabolism determined by HIF-1α could reflect the immune response reflected by the expression of PD-L1 11 12. In addition, direct blockade of PD-L1 within cancer cells has been reported to diminish glycolysis by inhibiting the mTOR pathway and the expression of glycolysis enzymes 23. Although the close relationship between 18F-FDG uptake and PD-L1 expression was also supported by other evidence, assessments to elucidate its mechanism require further investigation. However, the relationship between 18F-FDG uptake and PD-L2 expression has not yet been supported by experimental evidence. Takada et al. reported the radiological features of PD-L2 expression in 222 patients with lung adenocarcinoma 24. In their study, the SUVmax for 18F-FDG uptake was found to be significantly higher in PD-L2-positive than in PD-L2-negative cases 24. This corresponds to the results of our study. It remains unknown why the expression level of PD-L2 is closely related to 18F-FDG uptake. Our results suggest that the expression of PD-L2 is strongly associated with tumor glucose metabolism, hypoxia and angiogenesis as a mechanism of 18F-FDG uptake. PD-L2 seemed to be more strongly correlated with glucose metabolism, hypoxia and angiogenesis, compared with PD-L1. Further investigation should be conducted to elucidate the relationship between PD-L2 and 18F-FDG uptake from the perspective of basic science.
In the current study, SUVmax as assessment of 18F-FDG uptake on PET was identified as a significant marker for predicting the expression of PD-L1/PD-L2. However, TLG or MTV are also significant indicators of 18F-FDG uptake that can reflect tumor metabolic activity, and are calculated on the basis of SUV and are closely correlated with SUVmax. Our study is a first investigation to evaluate whether MTV or TLG could be correlated with the expression of PD-L1, thus, it remains unclear why SUVmax was chosen as a better marker for the prediction of PD-L1 expression than TLG or MTV. The relationship between metabolic tumor volume and PD-L1 expression may be obscure.
There are several limitations to our study. First, our study had a small sample size, which may have biased the results of our study. Since thymic cancer is a rare neoplasm, only limited numbers of samples were collected. Second, we tried to examine PD-L1 staining using clone 28 − 8; however, there are several kinds of PD-L1 clones. An additional investigation using other clones of PD-L1 may be needed to confirm the results of our study. Finally, the results of our study was not confirmed by experimental investigations. In the level of tumor cell lines, little is known about any data elucidating the association between PD-L1 expression and 18F-FDG uptake. Further examination is needed to approach some basic mechanism.
Further studies are warranted to assess TLG and MTV in the context of the present study’s findings.
In conclusion, the relevance and distribution of 18F-FDG uptake on PET were significantly associated with the expression of PD-L1 and PD-L2 in patients with TETs. In particular, PD-L1 and PD-L2 exhibited a close relationship with upregulation of tumor glucose metabolism (GLUT1) and hypoxia (HIF-1α), which play essential roles in the mechanism of 18F-FDG uptake within tumor cells. Further studies are needed to elucidate why PD-L1 and PD-L2 affect glucose metabolism and hypoxia in thymic tumor cells.