Plasma levels of methylated septin 9 are capable of detecting hepatocellular carcinoma and hepatic cirrhosis

Abstract

Hepatocellular carcinoma (HCC) was the third most common cause of cancer‑associated mortality in China in 2015. Early detection of HCC and hepatic cirrhosis (HC) can serve a crucial role in the prevention and therapeutic intervention of these diseases. Current early detection methods rely on less sensitive imaging modalities compared with the pathological examination. In the present study, a total of 64 patients with HCC, 44 patients with HC and 298 individuals with no evidence of disease (NED) were recruited, and the ability of methylated septin 9 (mSEPT9) in diagnosing HCC and HC was investigated. The overall detection sensitivity of mSEPT9 for HCC and HC was 76.7 and 34.1%, respectively, with a 95.9% specificity (HCC vs. NED). The sensitivity of mSEPT9 for HCC was significantly higher than that of α‑fetoprotein (AFP; χ2 test; 56.7%; P<0.05). The areas under the curve from the receiver operating characteristic curves of mSEPT9 for detection of HCC vs. NED, HC vs. NED and HCC vs. HC were 0.85, 0.77 and 0.66, respectively, while those of AFP for the same groups were 0.80, 0.55 and 0.77, respectively. Although both markers exhibited stage‑dependent sensitivity in HCC, mSEPT9 was demonstrated to be more sensitive than AFP. The net reclassification index of mSPET9 for HCC detection was 0.212 compared with AFP, suggesting an improved diagnostic performance of mSEPT9 compared with AFP. In addition, Kaplan‑Meier survival analysis revealed that mSEPT9 is able to predict the long‑term survival of patients with HCC. Further analysis suggested that patients >50 years of age exhibited higher sensitivity compared with those <50 years old in mSEPT9, but not in AFP. No significant difference in sensitivity was observed between compensated and decompensated patients with HC, and in patients with HC with a history of hepatitis B or C virus infection. No difference was observed between male and female subjects in the HC and HCC groups for mSEPT9 and AFP. In conclusion, mSEPT9 may detect HCC with an overall improved sensitivity compared with AFP and may help in predicting the long‑term survival of patients with HCC. The present clinical study was retrospectively registered to the Chinese Clinical Trial Registry on April 4, 2020 (http://www.chictr.org.cn/enIndex.aspx; registration no. ChiCTR2000031547).

Keywords: septin 9; liver; hepatocellular carcinoma; cirrhosis; methylation.

Figure 1.

Scatter plot and ROC curves for HCC and HC detection using the mSEPT9 assay. (A) Scatter plot showing the mSEPT9 ΔΔCq values from the blood of individuals with NED, HC and HCC. ROC curves for mSEPT9 sensitivity in detecting (B) HCC vs. NED, (C) HC vs. NED and (D) HCC vs. HC. The threshold (cutoff value) for ΔΔCq (dotted line) was set at −4.0. Bars represent the mean ± 95% CI. mSEPT9, methylated septin 9; ROC, receiver operating characteristic; HCC, hepatocellular carcinoma; HC, hepatic cirrhosis; NED, no evidence of disease; AUC, area under the curve.

Figure 2.

Scatter plot and ROC curves for HCC and HC detection using AFP. (A) Scatter plot showing the serum AFP concentration of individuals with NED, HC and HCC. ROC curves for AFP sensitivity in detecting (B) HCC vs. NED, (C) HC vs. NED and (D) HCC vs. HC. The threshold (cutoff value) for AFP (dotted line) was set at 20 µg/l (log value at 1.30). Bars represent the mean ± 95% CI. AFP, α-fetoprotein; ROC, receiver operating characteristic; HCC, hepatocellular carcinoma; HC, hepatic cirrhosis; NED, no evidence of disease; AUC, area under the curve.

Figure 3.

Sensitivity of mSEPT9 alone, AFP alone and both combined for the detection of NED, HC and stage A, B and C HCC. Sensitivity of (A) mSEPT9, (B) AFP and (C) both combined. Bonferroni correction was performed for the P-values from the χ² tests when >2 groups were compared. **P<0.01; ***P<0.001 vs. NED; and ^##P<0.01; ^###P<0.001 vs. HC. mSEPT9, methylated septin 9; AFP, α-fetoprotein; HCC, hepatocellular carcinoma; HC, hepatic cirrhosis; NED, no evidence of disease.

Figure 4.

Kaplan-Meier curves for the survival of patients with hepatocellular carcinoma based on mSEPT9 positive or negative detection. Patients were considered as mSEPT9 positive or negative based on a Cq threshold of 41.1. mSEPT9, methylated septin 9.

Figure 5.

Detection sensitivity for HCC and HC using mSEPT9 or AFP levels according to age. mSEPT9, methylated septin 9; AFP, α-fetoprotein; HCC, hepatocellular carcinoma; HC, hepatic cirrhosis.

Figure 6.

Detection sensitivity for HCC and HC using mSEPT9 or AFP levels according to sex. mSEPT9, methylated septin 9; AFP, α-fetoprotein; HCC, hepatocellular carcinoma; HC, hepatic cirrhosis.

Figure 7.

Effect of HBV or HCV infection on HC and compensation status on HC detection sensitivity using mSEPT9 and AFP levels. HBV/HCV, hepatitis B/C virus; com, compensated; decom, decompensated; mSEPT9, methylated septin 9; AFP, α-fetoprotein; HC, hepatic cirrhosis.