Abstract 4330: Immune checkpoint expression score is an independent prognostic biomarker in resectable non-small cell lung cancer
Authors: Usó Marta , Jantus-Lewintre Eloisa , Sirera Rafael , Calabuig Silvia , Pastor Enrique , Forteza Jerónimo , Camps Carlos 
Published: August 2015

Abstract
BACKGROUND
Immune checkpoints blockade, which activate antitumor immunity, has demonstrated promising clinical results in NSCLC. In this study we have investigated the prognostic role of immune checkpoint expression markers and its correlation with immune-cells infiltration and clinico-pathological characteristics in a cohort of resectable NSCLC patients.
MATERIAL AND METHODS
RNA was isolated from fresh-frozen lung specimens (tumor and normal lung) (n = 178). RTqPCR was performed to analyze the expression of CTLA-4, PD-1 and PD-L1 by the use of hydrolysis probes. Relative gene expression was assessed by Pfaffl formula and normalized by the use of CDKN1B, GUS and ACTB as endogenous genes (selected by GeNorm algorithm). These data was used to develop a gene expression score. Furthermore, the presence of CD4+, CD8+ and FOXP3+ lymphocytes was also assessed in FFPE samples from 63 of these patients by immunohistochemistry (IHC). All statistical analysis were considered significant at p&lt; 0.05.
RESULTS
Patient's median age was 65 years [26-85], 86.5% were male and 43.8% were adenocarcinomas (ADC). Since CTLA-4 and PD-1 were moderately associated with prognosis based on COX regression analysis (|Z-score|&lt;1.5), a multivariate model including these two genes was created. Absolute regression coefficients from this analysis were used in order to calculate the immune checkpoint score: (PD1 × 0.116) + (CTLA4 × 0.058) for each case. We found a significant association between the high immune checkpoint score and the presence of cytotoxic (CD8+) infiltrating lymphocytes in the tumor microenvironment (p = 0.030). Kaplan-Meier survival analysis showed that patients with high immune checkpoint score have longer overall survival (OS) [NR vs 40.4 months, p = 0.008] and longer progression free survival (PFS) [82.6 vs 23 months, p = 0.009]. A stratified analysis by histology was performed, showing a strong association between the high immune checkpoint score and better outcomes [OS p = 0.002; PFS p&lt;0.001] in the group of ADCs (n = 78). Multivariate analysis in the entire cohort indicated that the immune checkpoint score was an independent biomarker of prognosis for OS [HR: 0.308; 95%CI, 0.156-0.609; p = 0.001] and PFS [HR: 0.527; 95%CI, 0.298-0.933; p = 0.028] in early-stage NSCLC patients.
CONCLUSIONS
The immune checkpoint score based on the expression levels of CTLA-4 and PD-1 correlates with the presence of CD8+ infiltrating lymphocytes in the tumor microenvironment. This score provides relevant prognostic information for a better characterization of early-stage NSCLC patients with strikingly different outcomes who may be candidates for immune-based therapies.
Supported by grants PS09-01149 and RD12/0036/0025 from ISCIII.
Citation Format: Marta Usó, Eloisa Jantus-Lewintre, Rafael Sirera, Silvia Calabuig, Enrique Pastor, Jerónimo Forteza, Carlos Camps. Immune checkpoint expression score is an independent prognostic biomarker in resectable non-small cell lung cancer. [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 4330. doi:10.1158/1538-7445.AM2015-4330