Abstract 1314: Evaluation of PD-L1 mRNA and protein expression in non-small cell lung and hepatocellular carcinoma
Authors: Jaeger Savina , Lee Benjamin H. , Mosher Rebecca , Shebnova Olga , Wang Yan , Yu Yenyen , Yang David , Murakami Masato , Greshock Joel , Schlegel Robert , Boral Anthony , Cao Zhu Alexander 
Published: August 2015

Abstract
SJ and BHL contributed equally to this work
BACKGROUND:
Tumor immunotherapy is a unique therapeutic modality against human cancers. The recent success of PD1 antibody treatments highlight the value and potential of this approach. While there is no definitive clinical biomarker that is prognostic of PD1 antibody efficacy, the presence of PD-L1 protein detected by immunohistochemistry has correlated positively to clinical responses to anti-PD1 and PD-L1 therapies. To enable broad examination of cancer indications for PD1/PD-L1 based therapies, we sought to evaluate PD-L1 expression at both the protein and mRNA level in lung and hepatic tumors.
MATERIALS & METHODS:
PD-L1 protein expression was evaluated in a set of formalin-fixed paraffin-embedded non-small cell lung (NSCLC) adenocarcinoma (ACA), NSCLC squamous cell carcinoma (SCC), and hepatocellular carcinoma (HCC) tumors by immunohistochemistry (IHC). PD-L1 expression was scored semi-quantitatively by a manual histo-score (H-score) methodology based on staining intensity and percentage of positive tumor cells. In our IHC analysis, PD-L1 positivity (PD-L1+) was defined as an H-score ≥ 20. In parallel, PD-L1 mRNA expression data was examined from The Cancer Genome Atlas (TCGA) in these same indications (503 NSCLC ACA, 489 NSCLC SCC, and 191 HCC) and analyzed by comparing the expression in matched normal tissues from TCGA.
RESULTS:
With RNAseq analysis, data was calculated as log2 (RPKM+0.1) after RSEM normalization, utilizing OmicSoft RNASeq pipelines across TCGA tumor indications. The expression of PD-L1 is elevated in NSCLC ACA and SCC, relative to that in HCC. By overlaying the distributions and comparing the expression levels across all indications in TCGA, we ranked overexpression profiles for PD-L1 and found the TCGA HCC cohort to have much reduced PD-L1 mRNA levels, with a median level of -0.8 compared to 1.3 for ACA and 1.5 for SCC, which amounts to more than a 2-fold change of median level expression. With RNAseq, our analysis defines 50% of NSCLC adenocarcinoma, 54% of NSCLC squamous cell carcinoma, and 6% of HCC as high expressers for PD-L1.
Tumor cell PD-L1 protein expression was measured in 45 lung adenocarcinoma, 47 lung squamous cell carcinoma, and 69 hepatocellular carcinoma. 16/45 (35.6%) lung ACA, 21/47 (44.7%) lung SCC were PD-L1 positive. In contrast, PD-L1 positivity was seen in only 5/69 (7.2%) HCC samples.
CONCLUSIONS:
In summary, with IHC and RNAseq analysis in large and independent human NSCLC and HCC sample sets, we have found PD-L1 expression to be more enriched in NSCLC than in HCC. Importantly, amongst the large number of samples (161 for IHC and 1183 for RNAseq) in the 3 indications, very good concordance is observed between protein- and mRNA-based analyses. Our finding thus establishes the basis for large scale mRNA-based data mining for indications and patient segments enriched for responses to PD1/PD-L1-based immune therapies.
Citation Format: Savina Jaeger, Benjamin H. Lee, Rebecca Mosher, Olga Shebnova, Yan Wang, Yenyen Yu, David Yang, Masato Murakami, Joel Greshock, Robert Schlegel, Anthony Boral, Zhu Alexander Cao. Evaluation of PD-L1 mRNA and protein expression in non-small cell lung and hepatocellular carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1314. doi:10.1158/1538-7445.AM2015-1314