1. Patients Characteristics
The mean age of the patients with EOC was 57 years old, ranged from 45-71 years. At study closure, 25 (38%) of the patients were alive. Thirty patients with benign ovarian cystadenoma served as controls. The mean age of the controls was 43 years old, ranged from 16-62 years. The detailed information is displayed in Table 1.
Table 1. Patients characteristics
Patients characteristics
|
|
Epithelial ovarian cancer(n=66)
|
n(%)
|
Age
|
|
<50y
|
8
|
50-60y
|
24
|
60-70y
|
32
|
≥70y
|
2
|
Initial CA125
|
|
<35U/ml
|
4
|
≥35U/ml
|
62
|
Pathological type
|
|
Serous adenocarcinoma
|
54
|
Mucinous adenocarcinoma
|
2
|
Endometrioid adenocarcinoma
|
2
|
Clear cell carcinoma
|
8
|
Stage
|
|
I,II
|
8
|
III,IV
|
58
|
Neoadjuvant chemotherapy before surgery
|
32
|
Benign ovarian cystadenoma (n = 30)
|
|
Age
|
|
10-30y
|
5
|
30-50y
|
6
|
>50y
|
19
|
Initial CA125
|
|
<35U/ml
|
26
|
≥35U/ml
|
4
|
Pathological type
|
|
Serous cystadenoma
|
13
|
Mucinous cycstadenoma
|
17
|
2. Immunohistochemistry for HCMV
HCMV-IE protein was detected in tumor specimens from 82% of EOC patients and 36% of those with benign cystadenomas(Table 2). HCMV-IE protein expression was extensive in 61%, focal in 21% and negative in 18% of EOC tissues. Respectively, expression was extensive in 23%, focal in 13% and negative in 64% of benign ovarian cystadenoma(Figure 1). HCMV-pp65 protein was detected in tumor specimens from 97% of EOC patients and 63% of those with benign cystadenomas(Table 2). The expression was extensive in 76%, focal in 21% and negative in 3% in epithelial ovarian cancer tissue. Respectively, expression was extensive in 33%, focal in 30% and negative in 37% in benign ovarian cystadenoma(Figure 2 ).
Table 2. Expression of HCMV-IE and pp65 in Epithelial Ovarian Cancer and Benign Ovarian Cystadenoma
Type of tumor
|
HCMV IE
|
HCMV PP65
|
|
Extensive, n(%)
|
Focal, n(%)
|
Negative, n(%)
|
Extensive, n(%)
|
Focal, n(%)
|
Negative, n(%)
|
EOC
|
40/66(61)
|
14/66(21)
|
12/66(18)
|
50/66(76)
|
14/66(21)
|
2/66(3)
|
Ovarian cystadenoma
|
7/30(23)
|
4/30(13)
|
19/30(64)
|
10/30(33)
|
9/30(30)
|
11/30(37)
|
3.Higher Tumor HCMV Expression is Associated With More Advanced Disease
Next we analyzed the effects of HCMV on the EOC stage. HCMV-IE expression was extensive in 25% of Stage I-II tumors, 66% of Stage III-IV tumors; HCMV-pp65 expression was extensive in 38%, and 64%, respectively. Advanced tumor stage is correlated with extensive expression of HCMV-IE (P =0.0279) (Table 3).
Table 3. Expression of HCMV-IE and pp65 in EOC tissues of different stages
|
Extensive, n(%)
|
Focal/negative, n(%)
|
Chi-square
|
P
|
HCMV-IE
|
|
|
|
|
I,II
|
2(25)
|
6(75)
|
4.834
|
0.0279
|
III,IV
|
38(66)
|
20(34)
|
|
|
HCMV-pp65
|
|
|
|
|
I,II
|
3(38)
|
5(62)
|
2.036
|
0.1536
|
III,IV
|
37(64)
|
21(36)
|
|
|
4.Reactivation of Latent HCMV may be Induced by NACT
HCMV-IE expression was extensive in 75% of cancer tissue with NACT before surgery, 47% of cancer tissue without NACT; HCMV-pp65 expression was extensive in 69%, and 53%, respectively. This observation indicates that reactivation of latent HCMV within the tumor at interval debulking surery (IDS) may be induced with NACT as HCMV-IE viral proteins could be significantly extensive expressed in tumor tissue sections with NACT before surgery(P =0.0279) (Table 4).
Table 4. Expression of HCMV-IE and pp65 in EOC tissues with/witout NACT
|
Extensive, n(%)
|
Focal/negative, n(%)
|
Chi-square
|
P
|
HCMV-IE
|
|
|
|
|
NACT before surgery
|
24(75)
|
8(25)
|
5.390
|
0.0202
|
No NACT before surgery
|
16(47)
|
18(53)
|
|
|
HCMV-pp65
|
|
|
|
|
NACT before surgery
|
22(69)
|
10(31)
|
1.726
|
0.1890
|
No NACT before surgery
|
18(53)
|
16(47)
|
|
|
5.Poor Survival Rate Among EOC Patients With Extensive HCMV-IE Expression
To confirm the effects of HCMV on EOC clinical outcomes, we analyzed the overall survival by the method of Kaplan-Meier survival analysis. At time of study closure, 77% of patients with focal or negative expression of HCMV-IE in their tumors were alive versus 32% of those with extensive expression. The results showed that EOC patients who had focal or negative HCMV-IE expression in their tumors had significantly longer median OS than those with extensive HCMV-IE expression (41 vs.39 months, P = 0.03)(Figure 3A). Similarly, 26% of patients with focal or negative HCMV-pp65 protein expression were alive versus 61% with extensive expression; however, no significant difference in OS was observed(42 vs. 40 months, P = 0.37) (Figure 3B).
To determine whether HCMV-IE expression was an independent risk factor for the OS of EOC patients, we conducted both univariate and multivariate analyses. FIGO stage (P = 0.024) and HCMV-IE expression (P = 0.032) were prognostic factors for OS in patients with EOC, shown from univariate analysis. Furthermore, the multivariate analysis indicated that HCMV-IE expression (P = 0.034) were independent prognostic factors for OS (Table 5).
Table 5 Univariate and multivariate analyses of variables for overall survival
Variable
|
Univariate analysis
|
Multivariate analysis
|
|
HR(95%CI)
|
P
|
HR(95%CI)
|
P
|
Age
|
0.890(0.724-1.134)
|
0.332
|
|
|
Initial CA125
|
1.124(0.767-1.564)
|
0.643
|
|
|
Pathological type
|
0.893(0.546-1.422)
|
0.436
|
|
|
FIGO stage
|
1.097(1.037-1.946)
|
0.024
|
1.154(1.005-1.457)
|
0.167
|
HCMV-IE expression
|
1.008(0.978-1.475)
|
0.032
|
1.012(1.003-1.522)
|
0.034
|