Introduction
Materials and methods
Patients
Histopathological verification of formalin-fixed and paraffin-embedded cancer specimens, DNA isolation
HPV16 presence and its load
P16 immunostaining
Statistical analysis
Results
Patients
Feature | All N (%)a
| HPV16+ N (%) | HPV16− N (%) |
P value (test χ
2) |
---|---|---|---|---|
All | 109 (100.0) | 36 (33.0) | 73 (67.0) | |
Age | ||||
≤ 58 yearsb
| 51 (46.8) | 12 (33.3) | 39 (53.4) | 0.048 |
> 58 years | 58 (53.2) | 24 (66.7) | 34 (46.6) | |
Gender | ||||
Female | 22 (20.2) | 9 (25.0) | 13 (17.8) | 0.379 |
Male | 87 (79.8) | 27 (75.0) | 60 (82.2) | |
Status in the Karnofsky scale | ||||
≤ 80% | 49 (45.0) | 15 (41.7) | 34 (46.6) | 0.628 |
> 80% | 60 (55.0) | 21 (58.3) | 39 (53.4) | |
Localization | ||||
Oral cavity | 24 (22.0) | 6 (16.6) | 18 (24.7) | 0.501 |
Oropharynx | 68 (62.4) | 26 (72.2) | 42 (57.5) | |
Hypopharynx | 7 (6.4) | 2 (5.6) | 5 (6.8) | |
Larynx | 10 (9.2) | 2 (5.6) | 8 (11.0) | |
T stage | ||||
1 | 2 (1.8) | 0 (0.0) | 2 (2.7) | 0.211 |
2 | 24 (22.0) | 9 (25.0) | 15 (20.6) | |
3 | 60 (55.1) | 23 (63.9) | 37 (50.7) | |
4 | 23 (21.1) | 4 (11.1) | 19 (26.0) | |
N stage | ||||
0 | 19 (17.4) | 5 (13.9) | 14 (19.1) | 0.820 |
1 | 18 (16.5) | 5 (13.9) | 13 (17.8) | |
2 | 61 (56.0) | 22 (61.1) | 39 (53.4) | |
3 | 11 (10.1) | 4 (11.1) | 7 (9.6) | |
Grade | ||||
1 | 40 (36.7) | 12 (33.3) | 28 (38.4) | 0.641 |
2 | 57 (52.3) | 21 (58.3) | 36 (49.3) | |
3 | 12 (11.0) | 3 (8.3) | 9 (12.3) | |
Keratinization | ||||
Yes | 62 (56.9) | 15 (41.7) | 47 (64.0) | 0.024 |
No | 47 (43.1) | 21 (58.3) | 26 (36.0) | |
The level of smoking—Brinkman indexc
| ||||
≤ 520b
| 54 (49.5) | 20 (55.6) | 34 (46.6) | 0.378 |
> 520 | 55 (50.5) | 16 (67.6) | 39 (53.4) | |
The level of drinkingd
| ||||
Low | 50 (45.9) | 20 (55.6) | 30 (41.1) | 0.154 |
High | 59 (54.1) | 16 (44.4) | 43 (58.9) | |
Treatment | ||||
Definitive CRT or surgery + CRT | 46 (42.2) | 19 (52.8) | 27 (37.0) | 0.289 |
Definitive RT or surgery + RT | 38 (34.9) | 10 (27.8) | 28 (38.3) | |
Induction CT + definitive RT | 25 (22.9) | 7 (19.4) | 18 (24.7) | |
Treatment outcome | ||||
Regression of cancer disease | 71 (65.1) | 28 (77.8) | 43 (58.9) | 0.151 |
Treatment failure | 6 (5.5) | 1 (2.8) | 5 (6.9) | |
Local recurrence | 22 (20.2) | 6 (16.7) | 16 (21.9) | |
Distant metastases | 10 (9.2) | 1 (2.8) | 9 (12.3) | |
Survival | ||||
Alive at the last follow-up | 74 (67.9) | 28 (77.8) | 46 (63.0) | 0.291 |
Death from cancer disease | 19 (17.4) | 4 (11.1) | 15 (20.6) | |
Death from others reasons | 16 (14.7) | 4 (11.1) | 12 (16.4) |
HPV16 infection, viral load, P16 expression—correlation with clinical and histopathological data
Feature | Viral load ≤ 6764.3 copies/cella
N (%)b
| Viral load > 6764.3 copies/cell
N (%) |
P value (test χ
2) | Lack of P16 overexpression N (%) | P16 overexpression N (%) |
P value (test χ
2) |
---|---|---|---|---|---|---|
All | 18 (50.0) | 18 (50.0) | 9 (26.5) | 25 (73.5) | ||
Age | ||||||
≤ 58 yearsa
| 6 (33.3) | 6 (33.3) | 1.000 | 4 (44.4) | 7 (28.0) | 0.366 |
> 58 years | 12 (66.7) | 12 (66.7) | 5 (55.6) | 18 (72.0) | ||
Gender | ||||||
Female | 4 (22.2) | 5 (27.8) | 0.700 | 1 (11.1) | 17 (68.0) | 0.223 |
Male | 14 (77.2) | 13 (72.2) | 8 (88.9) | 8 (32.0) | ||
Status in the Karnofsky scale | ||||||
≤ 80% | 9 (50.0) | 6 (33.3) | 0.310 | 3 (33.3) | 10 (40.0) | 0.724 |
> 80% | 9 (50.0) | 12 (66.7) | 6 (66.7) | 15 (60.0) | ||
Localization | ||||||
Oral cavity | 3 (16.7) | 3 (16.7) | 0.202 | 2 (22.2) | 4 (16.0) | 0.003 |
Oropharynx | 11 (61.1) | 15 (83.3) | 3 (33.4) | 21 (84.0) | ||
Hypopharynx | 2 (11.1) | 0 (0.0) | 2 (22.2) | 0 (0.0) | ||
Larynx | 2 (11.1) | 0 (0.0) | 2 (22.2) | 0 (0.0) | ||
T stage | ||||||
1 | 0 (0.0) | 0 (0.0) | 0.985 | 0 (0.0) | 0 (0.0) | 0.861 |
2 | 4 (22.2) | 5 (27.8) | 3 (33.3) | 6 (24.0) | ||
3 | 12 (66.7) | 11 (61.1) | 5 (55.6) | 16 (64.0) | ||
4 | 2 (11.1) | 2 (11.1) | 1 (11.1) | 3 (12.0) | ||
N stage | ||||||
0 | 4 (22.2) | 1 (5.6) | 0.072 | 4 (44.4) | 1 (4.0) | 0.016 |
1 | 0 (0.0) | 5 (27.8) | 0 (0.0) | 5 (20.0) | ||
2 | 12 (66.7) | 10 (55.6) | 5 (55.6) | 16 (64.0) | ||
3 | 2 (11.1) | 2 (11.1) | 0 (0.0) | 3 (12.0) | ||
Grade | ||||||
1 | 7 (38.9) | 5 (27.8) | 0.700 | 4 (44.4) | 8 (32.0) | 0.597 |
2 | 10 (55.6) | 11 (61.1) | 5 (55.6) | 15 (60.0) | ||
3 | 1 (5.5) | 2 (11.1) | 0 (0.0) | 2 (8.0) | ||
Keratinization | ||||||
Yes | 7 (38.9) | 8 (44.4) | 0.735 | 5 (55.6) | 15 (60.0) | 0.816 |
No | 11 (61.1) | 10 (55.6) | 4 (44.4) | 10 (40.0) | ||
The level of smoking—Brinkman indexc
| ||||||
≤ 520a
| 8 (44.4) | 12 (66.7) | 0.180 | 4 (44.4) | 16 (64.0) | 0.307 |
> 520 | 10 (55.6) | 6 (33.3) | 5 (55.6) | 9 (36.0) | ||
The level of drinkingd
| ||||||
Low | 4 (22.2) | 16 (88.9) | 0.000 | 1 (11.1) | 18 (72.0) | 0.002 |
High | 14 (77.8) | 2 (11.1) | 8 (88.9) | 7 (28.0) | ||
Viral load | ||||||
≤ 6764.3 copies/cella
| – | – | 9 (100.0) | 7 (28.0) | 0.000 | |
> 6764.3 copies/cell | – | – | 0 (0.0) | 18 (72.0) | ||
Treatment | ||||||
Definitive CRT or surgery + CRT | 5 (27.8) | 14 (77.8) | 0.009 | 3 (33.3) | 16 (64.0) | 0.283 |
Definitive RT or surgery + RT | 7 (38.9) | 3 (16.7) | 4 (44.4) | 6 (24.0) | ||
Induction CT + definitive RT | 6 (33.3) | 1 (5.5) | 2 (22.2) | 3 (12.0) | ||
Treatment outcome | ||||||
Regression of cancer disease | 12 (66.8) | 16 (88.9) | 0.504 | 6 (66.7) | 21 (84.0) | 0.140 |
Treatment failure | 1 (5.5) | 2 (11.1) | 0 (0.0) | 1 (4.0) | ||
Local recurrence | 4 (22.2) | 0 (0.0) | 3 (33.3) | 2 (8.0) | ||
Distant metastases | 1 (5.5) | 0 (0.0) | 0 (0.0) | 1 (4.0) | ||
Survival | ||||||
Alive at the last follow-up | 12 (66.7) | 16 (88.9) | 0.102 | 3 (33.3) | 19 (76.0) | 0.072 |
Death from cancer disease | 4 (22.2) | 0 (0.0) | 3 (33.3) | 3 (12.0) | ||
Death from others reasons | 2 (11.1) | 2 (11.1) | 3 (33.3) | 3 (12.0) |
HPV16 infection, viral load, P16 expression—survival analysis
Overall survival | Disease-free survival | |||||||
---|---|---|---|---|---|---|---|---|
Response N (%)a
| HR | 95% CI | Log-rank P
| Response N (%)a
| HR | 95% CI | Log-rank P
| |
Age | ||||||||
≤ 58 yearsb
| 22/51 (43.1) | 1.354 | 33/51 (64.7) | 1.119 | ||||
> 58 years | 34/58 (58.6) | 1.000 | 0.430–1.269 | 0.265 | 40/58 (69.0) | 1.000 | 0.465–1.718 | 0.734 |
Gender | ||||||||
Female | 17/22 (77.3) | 1.000 | 18/22 (81.8) | 1.000 | ||||
Male | 39/87 (44.8) | 3.230 | 0.122–0.780 | 0.004 | 55/87 (63.2) | 2.664 | 0.132–1.064 | 0.038 |
Status in the Karnofsky scale | ||||||||
≤ 80% | 21/49 (42.9) | 1.712 | 29/49 (59.2) | 1.912 | ||||
> 80% | 35/60 (58.3) | 1.000 | 0.340–1.004 | 0.051 | 44/60 (73.3) | 1.000 | 0.271–1.011 | 0.052 |
Localization | ||||||||
Oral cavity | 10/24 (41.7) | 5.962 | 0.781–45.497 | 15/24 (62.5) | 2.034 | 0.439–9.418 | ||
Oropharynx | 34/68 (50.0) | 4.458 | 0.614–32.829 | 48/68 (70.6) | 1.329 | 0.310–5.693 | ||
Hypopharynx | 3/7 (42.9) | 5.118 | 0.568–46.127 | 2/7 (28.6) | 4.469 | 0.832–24.006 | ||
Larynx | 9/10 (90.0) | 1.000 | 0.253 | 8/10 (80.0) | 1.000 | 0.082 | ||
T stage | ||||||||
1 + 2 | 18/26 (69.2) | 1.000 | 22/26 (84.6) | 1.000 | ||||
3 + 4 | 38/83 (45.8) | 2.242 | 1.055–4.763 | 0.021 | 51/83 (61.4) | 3.148 | 1.111–8.914 | 0.015 |
N stage | ||||||||
0 + 1 | 23/37 (62.2) | 1.000 | 27/37 (73.0) | 1.000 | ||||
2 + 3 | 33/72 (45.8) | 1.902 | 1.029–3.515 | 0.030 | 46/72 (63.9) | 1.652 | 0.795–3.432 | 0.158 |
Grade | ||||||||
1 + 2 | 53/96 (55.2) | 1.000 | 68/96 (70.8) | 1.000 | ||||
3 | 3/13 (23.1) | 1.625 | 0.816–3.234 | 0.149 | 5/13 (38.5) | 2.197 | 0.999–4.832 | 0.046 |
Keratinization | ||||||||
Yes | 27/62 (43.6) | 1.734 | 38/62 (61.3) | 1.863 | ||||
No | 29/47 (61.7) | 1.000 | 0.981–3.065 | 0.051 | 35/47 (74.5) | 1.000 | 0.931–3.730 | 0.069 |
The level of smoking—Brinkman indexc
| ||||||||
≤ 520b
| 32/54 (59.3) | 1.000 | 42/54 (77.8) | 1.000 | ||||
> 520 | 24/55 (43.6) | 1.646 | 0.951–2.848 | 0.070 | 31/55 (56.4) | 2.420 | 1.208–4.850 | 0.010 |
The level of drinkingd
| ||||||||
Low | 29/50 (58.0) | 1.000 | 38/50 (76.0) | 1.000 | ||||
High | 27/59 (45.8) | 1.588 | 0.914–2.760 | 0.094 | 35/59 (59.3) | 2.106 | 1.051–4.221 | 0.030 |
HPV16 infection | ||||||||
Present | 23/36 (63.9) | 1.000 | 30/36 (83.3) | 1.000 | ||||
Absent | 33/73 (45.2) | 1.844 | 0.290–1.015 | 0.041 | 43/73 (58.9) | 3.128 | 0.133–0.769 | 0.005 |
Treatment | ||||||||
Definitive CRT or surgery + CRT | 31/46 (67.4) | 1.000 | 35/46 (76.1) | 1.000 | ||||
Definitive RT or surgery + RT | 16/38 (42.1) | 1.778 | 0.919–3.439 | 25/38 (65.8) | 1.531 | 0.686–3.418 | ||
Induction CT + definitive RT | 9/25 (36.0) | 2.460 | 1.212–4.994 | 0.050 | 13/25 (52.0) | 2.405 | 1.058–5.470 | 0.103 |
HR | 95% CI |
P valuea
| |
---|---|---|---|
Overall survival—109 patients | |||
Gender | |||
Female | 1.000 | ||
Male | 3.461 | 0.114–0.732 | 0.009 |
N stage | |||
0 + 1 | 1.000 | ||
2 + 3 | 2.221 | 1.187–4.157 | 0.013 |
HPV16 infection | |||
Present | 1.000 | ||
Absent | 2.134 | 0.247–0.887 | 0.020 |
Disease-free survival—109 patients | |||
T stage | |||
1 + 2 | 1.000 | ||
3 + 4 | 3.229 | 1.137–9.170 | 0.028 |
The level of smoking—Brinkman indexb
| |||
≤ 520c
| 1.000 | ||
> 520 | 2.149 | 1.069–4.319 | 0.032 |
HPV16 infection | |||
Present | 1.000 | ||
Absent | 3.083 | 0.134–0.783 | 0.012 |
Disease-free survival—36 patients with HPV16 positivity (assessed by amplification of viral gene E6 fragment) | |||
The level of smoking—Brinkman indexb
| |||
≤ 520c
| 1.000 | ||
> 520 | 2.253 | 1.124–4.514 | 0.023 |
Viral load | |||
> 6764.3 copies/cellc
| 1.000 | ||
≤ 6764.3 copies/cell | 2.795 | 1.060–1.121 | 0.045 |