Desmoplasia in non-small cell lung carcinomas is associated with low programmed death-ligand 1 expression and the absence of tumor-infiltrating lymphocytes.

Programmed death-ligand 1 (PD-L1) is the most widely utilized predictive marker used to identify non-small cell lung carcinoma (NSCLC) patients most suitable for immunotherapy approaches. The relationship between PD-L1 expression, the presence of CD8+ T cells, and other clinicopathological characteristics of NSCLC patients has not been elucidated yet. In this retrospective study, we immunohistochemically determined PD-L1 expression (using clone 22C3) and CD8+ T cell count (using clone c8/144B) in surgical resection specimens from 698 advanced NSCLC patients. Results of PD-L1 expression and CD8+ T cell count were correlated to various clinicopathological characteristics, including the presence of desmoplasia in NSCLC. Regarding the immunological attributes of the tumor microenvironment, we identified major differences between desmoplastic and non-desmoplastic areas in NSCLC. Tumor areas without desmoplasia were significantly more often PD-L1 positive than tumor cell clusters encased in a dense collagenous stroma (p=0.004). Furthermore, the desmoplastic stroma contained significantly less often an immune cell infiltrate rich in CD8+ T cells (p<0.001). Also, the positivity of PD-L1 significantly correlated with advanced N-stage (p<0.001) and poor differentiation in adenocarcinomas (p=0.032) but not with other clinicopathological characteristics. In conclusion, to our knowledge, this is the first study that points to major differences in terms of immunological attributes between desmoplastic and non-desmoplastic areas in NSCLC. The desmoplastic component, therefore, may represent an immunologically distinct tumor area in which PD-L1 immunohistochemistry and CD8+ T cell count should be evaluated separately.

Clinicopathological analysis of programmed death-ligand 1 testing in tumor cells of 325 patients with non-small cell lung cancer: Its predictive and potential prognostic value.

INTRODUCTION: Recent studies on check-point inhibitor therapy, which seems to improve the prognosis of patients with advanced non-small cell lung carcinoma increase the importance of immunohistochemical analyses of the programmed-death receptor and of its ligand, PD-L1 protein. MATERIAL AND METHODS: In our study we present results of PD-L1 immunohistochemical tumor cell expression in a series of 325 lung carcinoma patients biopsies, using the clone 22C3 (and DAKO Link 48 immunostainer). Evaluation of the expression using tissue proportion scoring system allowed to distinguish negative cases (either 0 % or < 1 % of positive tumor cells) versus positive cases in the categories 1-9 %, 10-49 % and ≥ 50 % of positive tumor cells. RESULTS: In association to histopathologic parameters we observed similar rates of positive expression in patients with adenocarcinoma types (47,8 % of all the cases) as well as with squamous cell carcinomas (44,4 %). Within these histological categories, the rates of positivity were similar also in patients with small versus large (resectional) biopsies. In the biopsies of patients with adenocarcinoma we identified differences in the PD-L1 protein expression associated with its histological subtype. In the cases with predominant lepidic pattern the PD-L1 positivity was present in 18,8 %, with predominant acinar or papillary pattern in 40,8 % and in cases with predominant solid or micropapillary component in 74,1 % of the cases resp. Keratinizing squamous cell carcinomas were positive in 38,5 % and non-keratinizing in 53,8 % of all the cases. The hiqhest incidence of an extensive posivity was observed in sarcomatoid carcinoma type. DISCUSSION AND CONCLUSION: Immunohistochemically verified PD-L1 protein expression represents a broadly accepted predictive biomarker for immunotherapy of NSCLC patients. The indicated differences of the expression among various NSCLC types and subtypes require to be verified in larger cohorts of patients in relation with clinical parameters to demonstrate whether it could be plausible to use the PD-L1 expression in a role of a negative prognostic parameter.

Expression of programmed death-ligand 1 protein in pulmonary squamous cell carcinoma correlates with tumour necrosis but not with tumour differentiation.

AIMS: Pulmonary squamous cell carcinoma (SqCC) represents the second most common non-small cell lung carcinoma type. The mechanisms which regulate programmed death ligand 1 (PD-L1) expression in this form of lung cancer are not fully elucidated yet. METHODS: We immunohistochemically determined the level of PD-L1 expression using the Tumour Proportion Score system in surgical resections of 133 patients with pulmonary SqCC. The results from PD-L1 immunohistochemistry were analysed in relation to tumour differentiation and the presence of necrotic areas comprising at least 20% of the tumour mass. RESULTS: No significant differences in terms of PD-L1 expression were found between SqCC subtypes as defined by the current WHO classification: better differentiated, keratinising tumours (12/24, 50.0 %) compared with less differentiated, non-keratinising and basaloid forms (62/109, 56.9 %) were PD-L1 positive in a comparable proportion of cases (p=0.1903). Contrary to that, SqCCs with the presence of necrosis (51/61, 83.6 %) had significantly more PD-L1-positive cases (p<0.001) compared with SqCCs without necrotic areas (23/72, 32.0 %) CONCLUSIONS: We demonstrated that PD-L1 expression in pulmonary SqCCs does not correlate with the traditionally defined degree of differentiation of these tumours. On the other hand, we found a significant association between the positive result of PD-L1 immunohistochemistry and tumour necrosis. Further investigation regarding the role of hypoxic pathways as presumable inducers of PD-L1 expression in pulmonary SqCCs might contribute to the understanding of this phenomenon.

Non-small cell lung carcinomas with a minor sarcomatoid component and pleomorphic carcinomas are associated with high expression of programmed death ligand 1.

Pleomorphic carcinomas are known to be highly programmed death ligand 1 (PD-L1) positive non-small cell lung cancer (NSCLC) types. However, the level of PD-L1 expression in lung carcinomas with a minor sarcomatoid component, comprising less than 10 % of the tumor mass, has not been determined yet. We hypothesized that NSCLC with a minor sarcomatoid component is more closely related to pleomorphic carcinomas in terms of PD-L1 expression than to NSCLC types without sarcomatoid features. The surgical resections from 690 lung carcinoma patients were retrospectively analyzed for the presence of PD-L1 by means of immunohistochemistry using the 22C3 PharmDx assay. The tumor proportion score system was applied to quantify the level of PD-L1 expression. Membranous staining present in ≥ 1 % of tumor cells was chosen as the cut-off to define a positive result for PD-L1 expression. Tumors were allocated into one of four subgroups: "adenocarcinoma", "squamous cell carcinoma", "pleomorphic carcinoma", or "NSCLC with a minor sarcomatoid component". PD-L1 expression in pleomorphic carcinomas (26/32, 81.3 %) and in the subgroup of NSCLC with a minor sarcomatoid component (35/46, 76.1 %) was identified in a comparable proportion of cases. Pleomorphic carcinomas were significantly more often PD-L1 positive than adenocarcinomas (p < 0.001) or squamous cell carcinomas (p = 0.0015). Accordingly, the proportion of PD-L1 expressing NSCLC with a minor sarcomatoid component was significantly higher than that of the adenocarcinoma (p < 0.001) or squamous cell carcinoma (p = 0.002) subgroup. In summary, we identified a presumable new subgroup of highly PD-L1 positive neoplasms within the NSCLC spectrum that is related to pleomorphic carcinomas in terms of PD-L1 expression. Further investigation regarding genetic relation and mechanism of PD-L1 expression in these two NSCLC categories is recommended.

Programmed death ligand 1 protein expression, histological tumour differentiation and intratumoural heterogeneity in pulmonary adenocarcinoma.

Intratumoural heterogeneity of pulmonary adenocarcinoma challenges the accurate interpretation of programmed death ligand 1 (PD-L1) immunohistochemistry, which is the only validated predictive marker for successful anti-PD-1/PD-L1 immunotherapy. The aim of this study was to determine whether PD-L1 expression is related to adenocarcinoma histological differentiation in a retrospective analysis of tumour biopsies with intratumoural histological heterogeneity. Adenocarcinomas with high intratumoural heterogeneity were categorised as 'mixed adenocarcinomas'. PD-L1 expression was determined immunohistochemically using tumour proportion scores (TPS). In 'mixed adenocarcinomas' PD-L1 scores were assessed across tumour areas with specific histological patterns. Comparisons were performed between histologically distinct differentiated tumours and/or histological areas. Poorly differentiated adenocarcinomas, represented by predominantly solid or micropapillary histological patterns, showed significantly higher expression of PD-L1 than other subtypes (p<0.001). Differentiation of intra-adenocarcinoma components was inversely correlated with PD-L1 expression: there were more PD-L1 positive cells in poorly differentiated areas than less differentiated (p<0.001), or than well differentiated areas (p<0.001), and in less differentiated more than well differentiated areas (p=0.001). In conclusion, PD-L1 expression is associated with poorly differentiated morphology in adenocarcinomas with intratumoural histological heterogeneity. Consequently, a TPS approach may not account for the contribution of more aggressive tumour components with higher levels of PD-L1 expression in within the tumour. Performing spectral analyses of PD-L1 expression across tumours is likely to be more accurate.