High CD49a+ NK cell infiltrate is associated with poor clinical outcomes in Hepatocellular Carcinoma

Abstract

NK cells infiltrating Hepatocellular Carcinoma (HCC) may express residency markers such as Integrin Subunit Alpha 1 (CD49a) that have been associated with nurturing functions in the decidua, and characterized by the production of angiogenic factors as well as loss of cytotoxicity. CIBERSORT, a computational analysis method for quantifying cell fractions from bulk tissue gene expression profiles, was used to estimate the infiltrating immune cell composition of the tumor microenvironment from gene expression profiles of a large cohort of 225 HCCs in the public GEO database. Decidual-like CD49a+ NK cells, in addition to another 22 immune cell populations, were characterized and thoroughly investigated so that HCC cell heterogeneity in a large cohort of 225 HCCs from the public GEO database could be studied. An inverse correlation of the expression of CD49a+ NK-cells and CD8⁺ T-cells suggested a negative association with clinical outcomes. This result was confirmed in a further validation cohort of 100 HCC patients from The Cancer Genome Atlas, Liver Hepatocellular Carcinoma (TCGA-LIHC). Cox regression analysis did not identify CD49a+ cells as a variable independently associated with survival. However, a more abundant infiltrate of this subset was present in patients at a more advanced pathological and clinical HCC stage. In conclusion, we found that NK cells, with a decidual-like gene expression profile, are enriched in HCC, and their abundance increases not only in tumor size but also at advanced stages of the disease suggesting that these cells play a role in tumor growth. For this reason, these NK cells may represent a possible new target for immunotherapeutic approaches in HCC.

Fig. 1

Correlation of the immune cell populations infiltrating HCCs in the GSE14520 cohort. Pearson's correlation matrix: colors represent Pearson's ‘r’ values, ranging from −1.00 to 1.00 from indirect to direct correlation.

Fig. 2

GSE14520 patients survival according to different demographic, clinical, and pathological variables. A. Forest plot showing hazard ratios (HR) and confidence intervals of the survival statistics. *statistically significant hazard ratios. Active viral replication and chronic carrier state are the two categories of HBV viral status. Cut-off values were: cirrhosis (yes or not), CLIP (0 + I and > I), Multinodular (yes or not), tumor size (>< 5 cm) ALT (<> 50UI/ml), AFP (<> 300 ng/ml), BCLC (0 + A and B + C), stage refers to pathologic stage: the comparison is between stage I and combined stage II and III. B. Only survival curves of significant parameters are represented.

Fig. 3

GSE14520 patient survival according to the relative expression of immune cell populations. A. Forest plot showing hazard ratios (HR) and confidence intervals of the survival statistics. *statistically significant hazard ratios. B. Survival curves of significant parameters are represented. Median values of expression of immune cell populations were used as cut-offs.

Fig. 4

TCGA patient survival according to the relative expression of immune cell populations. A. Forest plot showing hazard ratios (HR) and confidence intervals of the survival statistics. *statistically significant hazard ratios. B. Survival curves of significant parameters are represented. Median values of expression of immune cell populations were used as cut-offs.

Fig. 5

Multivariate analysis of immune cell populations from GSE14520, significantly associated with clinical outcome (OS). Forest plot shows hazard ratios (HR) and confidence intervals. *statistically significant hazard ratios.

Fig. 6

CD49a expression in GSE14520 patients HCCs according to tumor size, pathologic and clinical staging. Tumor size below or equal and above 5 cm diameter of the main HCC nodule, tumor stage according to American Joint Committee on Cancer (AJCC), The Cancer of the Liver Italian Program (CLIP) score, Barcelona Clinic Liver Cancer (BCLC).