Tumor-associated autoantibodies are useful biomarkers in immunodiagnosis of α-fetoprotein-negative hepatocellular carcinoma

Table 1 Baseline characteristics of patients in α fetoprotein-negative hepatocellular carcinoma, α fetoprotein-positive hepatocellular carcinoma, chronic liver disease and normal human control groups

Variable	AFP(-)HCC (n = 56)	AFP(+)HCC (n = 86)	CLD (n = 168)	NHC (n = 59)
Age, yr	57 ± 9	56 ± 10	48 ± 14	39 ± 13
Sex, male/female	47/9	66/20	129/39	25/34
AFP, ng/mL	5.38 (3.4-8.2)	795.1(116.1-11244.0)	4.32 (2.5-10.4)	-
HBV/HCV/BC/NBNC	36/4/6/10	75/5/1/5	103/39/3/23
ALT, U/L	37.9 (26.2-53.6)	40.5 (26.8-79.6)	42.9 (24.5-135.1)	-
AST, U/L	41.6 (29.8-66)	63.9 (37.05-146.3)	47.1 (27.8-92.6)	-
TBIL, μmol/L	22.4 (15.9-33.7)	28.4 (17.4-53.7)	23.8 (14.8-49.1)	-
DBIL, μmol/L	5.9 (3.8-10.6)	7.3 (4.5-20.8)	6 (3.6-18.6)
ALB, g/L	34.2 ± 5.9	34.8 ± 5.7	38.1(32.4-43)	-
CR, μmol/L	63 (55.3-73.2)	64 (52.2-74.3)	63.85 (55.1-73.3)	-
INR	1.09 (1-1.21)	1.13 (1.03-1.24)	-	-
PT, s	12.4 (11.4-13.8)	12.7 (11.5-14.1)	-	-
Child-Pugh score	6 (5-8)	7 (6-9)	-	-
Child-Pugh grade, A/B/C	6/1/49	62/19/5	-	-
Meld score	9 (8-11)	10 (8-13)	-	-
BCLC grade, A/B/C/D	24/16/13/3	21/17/42/6	-	-
Tumor size, > 5 cm/< 5 cm	19/37	42/44	-	-
Tumor no., single/double/multiple	4/18/34	31/7/48	-	-
Vascular invasion, yes/no	11/45	44/42	-	-
Metastasis, yes/no	5/51	11/75	-	-
Encephalopathy, non-/1-2/3-4	53/3/0	1/4/81	-	-
Ascites degree, non/low/medium/high	24/25/2/5	31/40/2/13	-	-

Continuous variables are expressed as the mean ± SD or the medians (25^th and 75^th percentile). Count data are described as frequency. AFP: Alpha fetoprotein; CLD: Chronic liver disease; HCC: Hepatocellular carcinoma; NHC: Normal human control; NPM1: Nucleophosmin 1; MDM2: Mouse double minute 2 homolog.

Table 2 Frequency of autoantibodies against NPM1, 14-3-3zeta and MDM2 in human sera by enzyme-linked immunosorbent assay n (%)

Type of sera	No. tested	Frequency of autoantibody
		NPM1	14-3-3zeta	MDM2	Combination of 3 proteins
AFP(-)HCC	56	12 (21.4)b	11 (19.6)b	11 (19.6)ab	17 (30.4)b
AFP(+)HCC	86	9 (10.5)	8 (9.3)	14 (16.3)	18 (20.9)
CLD	168	9 (5.4)	11 (6.5)	15 (8.9)	18 (10.7)
NHC	59	1 (1.7)	0 (0)	1 (1.7)	1 (1.7)

Cut-off value, mean ± 3 SD of NHCs.

^aP < 0.05 and

^bP < 0.01 (value relative to CLD and NHC). AFP: Alpha fetoprotein; CLD: Chronic liver disease; HCC: Hepatocellular carcinoma; NHC: Normal human control; NPM1: Nucleophosmin 1; MDM2: Mouse double minute 2 homolog.

Table 3 Diagnostic value of autoantibodies and different combinations in α-fetoprotein-negative hepatocellular carcinoma

Type of sera	Sensitivity	Specificity	Accuracy
NPM1	21.4%	95.6%	80.9%
14-3-3zeta	19.6%	95.2%	80.2%
MDM2	19.6%	93.0%	78.4%
NPM1 + 14-3-3zeta	25.0%	94.3%	80.7%
NPM1 + MDM2	30.4%	93.0%	80.7%
14-3-3zeta + MDM2	25.0%	92.1%	78.8%
NPM1 + 14-3-3zeta + MDM2	30.4%	91.6%	79.5%

NPM1: Nucleophosmin 1; MDM2: Mouse double minute 2 homolog.

Table 4 Characteristics of patients and three proteins expression in α-fetoprotein-negative hepatocellular carcinoma tissues n (%)

Variable	Age, yr	Sex, male/female	Grade, I-II/III-IV	Frequency
				NPM1	14-3-3zeta	MDM2
Liver cancer	52 ± 9	8/1	6/3	9 (100)	5 (55.6)	3 (33.3)
Normal liver tissue	58 ± 9	9/1	-	2 (20.0)	1 (10.0)	2 (20.0)

Continuous variables are expressed as mean ± SD; Count data are described as frequency.