The Clinicopathological and Prognostic Significance of Nrf2 and Keap1 Expression in Hepatocellular Carcinoma
Abstract
:1. Introduction
2. Results
2.1. pNrf2 Expression in HCCs and Clinicopathological Correlation
2.2. Cytoplasmic Nrf2 Expression in HCCs and Clinicopathological Correlation
2.3. Keap1 Expression in HCCs and Clinicopathological Correlation
2.4. Multivariate Survival Analysis
3. Discussion
4. Materials and Methods
4.1. Patients
4.2. Tissue Samples and Immunohistochemistry
4.3. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
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Parameters | Frequency (%) or Median (Range) |
---|---|
Sex, n (%) | |
Male | 216 (75.8) |
Female | 69 (24.2) |
Age at operation, median (range, years) | 63 (29–87) |
Etiology, n (%) | |
Hepatitis B virus (HBV) | 202 (70.9) |
Hepatitis C virus (HCV) | 25 (8.8) |
HBV + HCV | 1 (0.3) |
Alcohol | 14 (4.9) |
Non-alcoholic fatty liver disease | 12 (4.2) |
Uncertain etiology | 31 (10.9) |
Tumor size, median (range, cm) | 3.2 (0.9–17) |
Multiplicity present, n (%) | 49 (17.2) |
Edmondson–Steiner grade, n (%) | |
Grade I | 3 (1.0) |
Grade II | 77 (27.0) |
Grade III | 171 (60.0) |
Grade IV | 34 (12.0) |
Microvascular invasion present, n (%) | 113 (39.6) |
Portal vein invasion present, n (%) | 19 (6.7) |
Pathologic T category (AJCC 8th edition), n (%) | |
pT1a | 37 (13.0) |
pT1b | 101 (35.4) |
pT2 | 112 (39.3) |
pT3 | 11 (3.9) |
pT4 | 24 (8.4) |
Extrahepatic metastasis present, n (%) | 55 (19.3) |
Cirrhosis in background liver present, n (%) | 152 (533.3) |
Recurrence on follow-up present, n (%) | 161 (56.5) |
Status at last follow-up, n (%) | |
Alive | 128 (44.9) |
Deceased of disease | 44 (15.4) |
Deceased of other cause | 10 (3.5) |
Follow up loss | 103 (36.2) |
Overall survival, median (range, months) | 66.5 (0–163) |
Disease-free survival, median (range, months) | 32.5 (0–163) |
Parameter | pNrf2 | p-Value | Nrf2 | p-Value | Keap1 | p-Value | |||
---|---|---|---|---|---|---|---|---|---|
Low pNrf2 | High pNrf2 | Low Nrf2 | High Nrf2 | Low Keap1 | High Keap1 | ||||
Frequency | 212 (74.3) | 73 (25.6) | - | 237 (83.2) | 48 (16.8) | - | 124 (43.5) | 161 (56.5) | - |
Sex (male/female) | 159 (75.0)/53 (25.0) | 57 (78.1)/16 (21.9) | 0.638 | 179 (75.5)/58 (24.5) | 37 (77.1)/11 (22.9) | 1.000 | 94 (75.8)/30 (24.2) | 122 (75.8)/39 (24.2) | 1.000 |
Age ≥60 years | 95 (44.8) | 28 (38.4) | 0.411 | 100 (42.2) | 23 (47.9) | 0.542 | 55 (44.7) | 68 (42.2) | 0.809 |
HBV-related etiology | 150 (70.8) | 53 (72.6) | 0.881 | 167 (70.5) | 36 (75.0) | 0.602 | 86 (69.4) | 117 (72.7) | 0.598 |
Size (cm, mean ± SD) | 4.2 ± 3.0 | 4.0 ± 2.4 | 0.274 | 4.4 ± 2.9 | 3.1 ± 1.8 | 0.005 | 4.4 ± 3.1 | 3.9 ± 2.5 | 0.105 |
Multiplicity | 37 (17.5) | 12 (16.4) | 1.000 | 41 (17.3) | 8 (16.7) | 1.000 | 23 (18.5) | 26 (16.1) | 0.636 |
E–S grade III and IV | 155 (73.1) | 50 (68.5) | 0.454 | 173 (73.0) | 32 (66.7) | 0.382 | 92 (74.2) | 113 (70.2) | 0.507 |
Microvascular invasion | 78 (36.8) | 35 (47.9) | 0.098 | 96 (40.5) | 17 (35.4) | 0.628 | 47 (37.9) | 66 (41.0) | 0.627 |
Portal vein invasion | 12 (5.7) | 7 (9.6) | 0.278 | 17 (7.2) | 2 (4.2) | 0.750 | 12 (9.7) | 7 (4.3) | 0.094 |
pT category | 0.146 | 0.323 | 0.166 | ||||||
pT1a | 31 (14.6) | 6 (8.2) | 33 (13.9) | 4 (8.3) | 18 (14.5) | 19 (11.8) | |||
pT1b | 75 (35.4) | 26 (35.6) | 83 (35.0) | 18 (37.5) | 46 (37.1) | 55 (34.2) | |||
pT2 | 76 (35.8) | 36 (49.3) | 89 (37.6) | 23 (47.9) | 41 (33.1) | 71 (44.1) | |||
pT3 | 10 (4.7) | 1 (1.4) | 11 (4.6) | 0 (0) | 4 (3.2) | 7 (4.3) | |||
pT4 | 20 (9.4) | 4 (5.5) | 21 (8.9) | 3 (6.3) | 15 (12.1) | 9 (5.6) | |||
Recurrence | 118 (55.7) | 43 (58.9) | 0.784 | 133 (56.1) | 28 (58.3) | 0.748 | 64 (51.6) | 97 (60.2) | 0.183 |
Extrahepatic metastasis | 37 (17.6) | 18 (24.7) | 0.229 | 50 (21.2) | 5 (9.1) | 0.109 | 24 (19.5) | 31 (19.4) | 1.000 |
Underlying cirrhosis | 108 (52.9) | 44 (60.3) | 0.338 | 125 (54.1) | 27 (58.7) | 0.628 | 65 (55.1) | 87 (54.7) | 1.000 |
K19 expression | 38 (17.9) | 16 (21.9) | 0.490 | 42 (17.7) | 12 (25.0) | 0.233 | 18 (14.5) | 36 (22.4) | 0.127 |
EpCAM expression | 84 (39.6) | 41 (56.2) | 0.020 | 103 (43.5) | 22 (45.8) | 0.873 | 49 (39.5) | 76 (47.2) | 0.229 |
CAIX expression | 48 (23.4) | 27 (37.5) | 0.030 | 55 (24.0) | 20 (41.7) | 0.019 | 31 (25.4) | 44 (28.4) | 0.589 |
Ezrin expression | 78 (36.8) | 43 (59.7) | 0.001 | 90 (38.1) | 31 (64.6) | 0.001 | 48 (38.7) | 73 (45.6) | 0.277 |
uPAR expression | 58 (27.5) | 26 (36.1) | 0.180 | 70 (29.7) | 14 (29.8) | 1.000 | 29 (23.4) | 55 (34.6) | 0.049 |
E-cadherin loss | 128 (60.4) | 56 (76.7) | 0.015 | 153 (64.6) | 31 (64.6) | 1.000 | 81 (65.3) | 103 (64.0) | 0.901 |
p53 overexpression | 40 (20.5) | 26 (37.1) | 0.009 | 53 (24.1) | 13 (28.9) | 0.570 | 19 (16.1) | 47 (32.0) | 0.004 |
Mitotic index (/10 HPF, mean ± SD) | 10.1 ± 13.4 | 17.4 ± 20.0 | <0.001 | 11.9 ± 15.6 | 12.2 ± 16.1 | 0.623 | 10.5 ± 13.6 | 13.1 ± 16.9 | 0.056 |
Ki-67 labeling index (%, mean ± SD) | 5.1 ± 7.5 | 12.9 ± 12.2 | <0.001 | 7.2 ± 9.6 | 6.7 ± 9.6 | 0.935 | 5.7 ± 8.5 | 8.1 ± 10.2 | 0.035 |
Parameter | Overall Survival | Disease-Free Survival | ||||||
---|---|---|---|---|---|---|---|---|
Univariate Analysis | Multivariate Analysis | Univariate Analysis | Multivariate Analysis | |||||
Parameters | HR (95% CI) | p-Value | HR (95% CI) | p-Value | HR (95% CI) | p-Value | HR (95% CI) | p-Value |
Sex (male/female) | 0.742 (0.345–1.597) | 0.446 | - | - | 0.702 (0.480–1.029) | 0.069 | - | - |
Age (≥60/<60 years) | 0.960 (0.522–1.765) | 0.894 | - | - | 0.848 (0.621–1.157) | 0.299 | - | - |
Underlying HBV | 1.160 (0.586–2.296) | 0.671 | - | - | 1.129 (0.801–1.590) | 0.488 | - | - |
Size (≥5cm/<5cm) | 2.400 (1.315–4.382) | 0.004 | 1.298 (0.650–2.594) | 0.460 | 1.418 (1.012–1.988) | 0.043 | 1.085 (0.765–1.540) | 0.647 |
Multiplicity | 2.812 (1.488–5.312) | 0.001 | 2.156 (1.093–4.253) | 0.027 | 1.991 (1.375–2.884) | <0.001 | 1.487 (1.015–2.180) | 0.042 |
E–S grade III and IV | 0.951 (0.497–1.818) | 0.879 | - | - | 1.095 (0.778–1.542) | 0.601 | - | - |
Microvascular invasion | 1.477 (0.813–2.682) | 0.200 | - | - | 1.308 (0.962–1.779) | 0.087 | - | - |
Portal vein invasion | 3.289 (1.386–7.804) | 0.007 | 2.435 (0.980–6.052) | 0.055 | 1.573 (0.853–2.903) | 0.147 | - | - |
Extrahepatic metastasis | 4.458 (2.445–8.130) | <0.001 | 3.256 (1.742–6.083) | <0.001 | 4.799 (3.411–6.753) | <0.001 | 4.475 (3.152–6.354) | <0.001 |
Underlying cirrhosis | 1.282 (0.702–2.340) | 0.419 | - | - | 1.031 (0.758–1.404) | 0.844 | - | - |
K19 | 1.860 (0.957–3.614) | 0.067 | - | - | 1.209 (0.826–1.771) | 0.329 | - | - |
EpCAM | 1.727 (0.951-3.137) | 0.073 | - | - | 0.889 (0.613–1.292) | 0.538 | - | - |
CAIX | 1.854 (1.015–3.386) | 0.044 | 1.402 (0.738–2.664) | 0.302 | 1.043 (0.743–1.464) | 0.809 | - | - |
Ezrin | 3.160 (1.674–5.966) | <0.001 | 2.893 (1.527–5.483) | 0.001 | 1.090 (0.766–1.552) | 0.632 | - | - |
uPAR | 1.492 (0.812–2.743) | 0.198 | 1.058 (0.756–1.480) | 0.742 | ||||
E-cadherin loss | 1.037 (0.556–1.936) | 0.909 | 0.853 (0.622–1.169) | 0.323 | ||||
p53 overexpression | 1.777 (0.961–3.288) | 0.067 | - | - | 1.219 (0.854–1.740) | 0.276 | - | - |
Nrf2 | 3.329 (1.738–6.374) | <0.001 | 4.151 (2.025–8.508) | <0.001 | 1.315 (0.879–1.968) | 0.183 | - | - |
pNrf2 | 1.894 (1.037–3.458) | 0.038 | 1.181 (0.950–3.450) | 0.071 | 0.933 (0.661–1.317) | 0.693 | - | - |
Keap1 | 1.061 (0.582–1.937) | 0.846 | - | - | 1.290 (0.944–1.762) | 0.109 | - | - |
Antibody | Description | Source | Dilution | Method |
---|---|---|---|---|
Nrf2 | Rabbit monoclonal (EP1808Y) | Abcam | 1:50 | Autostainer |
(Cambridge, UK) | ||||
pNrf2 | Rabbit monoclonal (EP1809Y) | Abcam | 1:100 | Autostainer |
(Cambridge, UK) | ||||
Keap1 | Rabbit polyclonal | Proteintech | 1:300 | Autostainer |
(Manchester, UK) | ||||
Keratin 19 (K19) | Mouse monoclonal (RCK108) | Dako | 1:200 | Autostainer |
(Glostrup, Denmark) | ||||
EpCAM | Mouse monoclonal (VU-1D9) | Millipore | 1:1500 | Autostainer |
(Temecula, CA, USA) | ||||
Ezrin | Mouse monoclonal (3C12) | Abcam | 1:100 | Manual (pH 6.0) |
(Cambridge, UK) | ||||
uPAR | Mouse monoclonal (R-4) | Abcam | 1:40 | Manual (pH 6.0) |
(Cambridge, UK) | ||||
E-cadherin | Mouse monoclonal (36B5) | Novocastra | 1:100 | Autostainer |
(Newcastle, UK) | ||||
CAIX | Rabbit polyclonal | Abcam | 1:500 | Autostainer |
(Cambridge, UK) | ||||
p53 | Mouse monoclonal (DO-7) | Dako | 1:1000 | Autostainer |
(Glostrup, Denmark) | ||||
Ki-67 | Mouse monoclonal (MIB-1) | Dako | 1:100 | Autostainer |
(Glostrup, Denmark) |
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Share and Cite
Lee, K.; Kim, S.; Lee, Y.; Lee, H.; Lee, Y.; Park, H.; Nahm, J.H.; Ahn, S.; Yu, S.J.; Lee, K.; et al. The Clinicopathological and Prognostic Significance of Nrf2 and Keap1 Expression in Hepatocellular Carcinoma. Cancers 2020, 12, 2128. https://doi.org/10.3390/cancers12082128
Lee K, Kim S, Lee Y, Lee H, Lee Y, Park H, Nahm JH, Ahn S, Yu SJ, Lee K, et al. The Clinicopathological and Prognostic Significance of Nrf2 and Keap1 Expression in Hepatocellular Carcinoma. Cancers. 2020; 12(8):2128. https://doi.org/10.3390/cancers12082128
Chicago/Turabian StyleLee, Kiryang, Seunghye Kim, Yangkyu Lee, Hyejung Lee, Youngeun Lee, Hyunjin Park, Ji Hae Nahm, Soomin Ahn, Su Jong Yu, Kyoungbun Lee, and et al. 2020. "The Clinicopathological and Prognostic Significance of Nrf2 and Keap1 Expression in Hepatocellular Carcinoma" Cancers 12, no. 8: 2128. https://doi.org/10.3390/cancers12082128