Increased foxp3+ regulatory T cells predicts for superior relapse-free and overall survival in early stage hepatocelullar carcinoma

Abstract

4642 Background: Hepatocellular carcinoma (HCC) endemic in Asia is aggressive and mainly hepatitis B-related. In ovarian and breast cancer, higher levels of foxp3+ regulatory T cells predicts for a poorer clinical outcome. The reverse is true with follicular lymphoma and Hodgkin’s disease. We hypothesize that foxp3+ CD4+ regulatory T lymphocyte (Treg) frequency predicts for clinical outcome in HCC. We also examined the recently identified IL17-producing, RORγt-expressing, CD4+ pathogenic Th17 cells in HCC. Methods: We analyzed Treg cells (n=116) and CD8+ T cells (n=24) expression by CD3/Foxp3 double immunoenzymatic using a anti-Foxp3 murine monoclonal antibody (Nuffield Department of Clinical Laboratory Sciences, Oxford University, UK) and CD3/CD8 double immunofluorescence, respectively. Treg cells and CD8 cells expression were reported as a percentage of CD3+ T cells. RORγt (for Th17) and Foxp3 mRNA were measured using real-time PCR. Results: Median age (n=116) was 62 years, 65.5% were hepatitis B+ (n=76), 4.3% were hepatitis C+ (n=5), 1.7% were both hepatitis B+ and C+ (n=2) and 28.4% were not virus-related (n=33). Treg numbers were significantly higher in all 116 HCC (median 9.3% of CD3+ T cells) compared to paired adjacent liver (median 1.9%) (p<0.001). Conversely, CD8+ T cell numbers were significantly lower in 24 HCC (median 21.1%) vs paired liver (median 49.5%) (p<0.001). Stage 1 and II HCC patients (n=24) with superior overall survival (= 6 years) compared with stage I and II patients (n=29) with inferior survival (≤ 1 year) had a significantly higher Treg frequency (median 10.9% versus 8.4%) (p=0.035). Stage I and II HCC patients (n=66) who were relapse-free at ≥ 1 year vs Stage I and II HCC patients (n=21) who were relapse-free at ≤ 0.5 years had higher Treg frequency (median 10.2% vs 6.8%) (p = 0.005). Hepatitis B+ HCC patients (n=76) had significantly higher Treg (median 10.2%) compared to non-viral associated HCC patients (n=33) (p=0.013). No correlation was seen between ROR and foxp3 mRNA. ROR mRNA was over-expressed (> 1 fold) in 20 (54%) HCC (analysis ongoing). Conclusions: Treg expression was significantly higher in HCC compared to paired liver. Increased Treg frequency predicts for a superior relapse-free and overall survival in HCC. No significant financial relationships to disclose.