Evaluation of the Use of Methylation as a New Tool for the Diagnostics and Progression of Squamous Intraepithelial Lesions
Abstract
:1. Introduction
2. Results
2.1. Comparison Between Patients with a Positive (+) and Negative Outcome of Methylation (−) in the Research Group
2.2. Effectiveness of Methylation in Predicting Final Outcome in the Research Group
2.3. Effectiveness of Methylation in Predicting Final Outcome in Both Study Groups (Research Group + Control Group)
3. Discussion
4. Materials and Methods
4.1. Study Design
4.2. Specimen Collection and Handling
4.2.1. HPV Genotyping Test and LBC
4.2.2. Sample Preparation for Methylation Test
- Isolation from cytological cervical material
- B.
- DNA bisulfite conversion and methylation
- Denaturation and bisulfite conversion;
- Binding of the DNA to the spin column and desulfonation;
- Washing the spin column bound DNA and elimination of ethanol;
- Elution of converted DNA.
4.3. Colposcopy, Punch Biopsy and LEEP-Conisation
4.4. Statistical Analysis
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Characteristics | Research Group | Control Group | MD (95% CI) | p |
---|---|---|---|---|
N | 85 (100.0) | 23 (100.0) | - | - |
Age, years, Me (Q1; Q3) | 37.97 (33.79;41.92) | 49.47 (41.87;58.52) | −11.50 (−16.39;−5.82) | <0.001 |
Methylation, n (%) | ||||
Positive | 37 (43.5) | 2 (8.7) | - | 0.004 |
Negative | 48 (56.5) | 21 (91.3) | ||
Pap smear, n (%) | ||||
NILM | 9 (10.6) | 13 (56.5) | - | <0.001 |
ASC-US | 15 (17.6) | 4 (17.4) | ||
LSIL | 21 (24.7) | 3 (13.0) | ||
ASC-H | 22 (25.9) | 1 (4.3) | ||
HSIL | 14 (16.5) | 0 (0.0) | ||
AGC | 2 (2.4) | 1 (4.3) | ||
Clinically suspicious image of the cervix | 2 (2.4) | 1 (4.3) | ||
HPV, n (%) | ||||
Positive | 73 (85.9) | 8 (34.8) | - | <0.001 |
Negative | 12 (14.1) | 15 (65.2) | ||
HPV genotype, n (%) * | ||||
HR | 34 (40.0) | 6 (26.1) | - | 0.326 |
16 | 38 (44.7) | 3 (13.0) | - | 0.011 |
18 | 4 (4.7) | 0 (0.0) | - | 0.576 |
31 | 14 (16.5) | 0 (0.0) | - | 0.038 |
45 | 5 (5.9) | 0 (0.0) | - | 0.582 |
Biopsy, n (%) | ||||
LSIL | 16 (18.8) | 0 (0.0) | - | <0.001 |
HSIL | 58 (68.2) | 0 (0.0) | ||
Adenocarcinoma | 2 (2.4) | 0 (0.0) | ||
Squamous cell carcinoma | 2 (2.4) | 0 (0.0) | ||
No signs of atypia | 3 (3.5) | 13 (56.5) | ||
Not performed | 4 (4.7) | 10 (43.5) | ||
LEEP/hysterectomy, n (%) | ||||
LSIL | 15 (17.6) | 0 (0.0) | - | <0.001 |
HSIL | 56 (65.9) | 0 (0.0) | ||
Adenocarcinoma | 2 (2.4) | 0 (0.0) | ||
Squamous cell carcinoma | 2 (2.4) | 0 (0.0) | ||
No signs of atypia | 7 (8.2) | 5 (21.7) | ||
Not performed | 3 (3.5) | 18 (78.3) | ||
Final, n (%) | ||||
LSIL | 15 (17.6) | 0 (0.0) | - | <0.001 |
HSIL | 66 (77.6) | 0 (0.0) | ||
Adenocarcinoma | 2 (2.4) | 0 (0.0) | ||
Squamous cell carcinoma | 2 (2.4) | 0 (0.0) | ||
No signs of atypia | 0 (0.0) | 23 (100.0) |
Characteristics | Research Group | Control Group | ||||
---|---|---|---|---|---|---|
Methylation | p | Methylation | p | |||
+ | − | + | − | |||
Pap smear, n (%) | ||||||
NILM | 3 (8.1) | 6 (12.5) | 0.016 | 1 (50.0) | 12 (57.1) | 0.332 |
ASC-US | 3 (8.1) | 12 (25.0) | 0 (0.0) | 4 (19.0) | ||
LSIL | 6 (16.2) | 15 (31.2) | 0 (0.0) | 3 (14.3) | ||
ASC-H | 12 (32.4) | 10 (20.8) | 1 (50.0) | 0 (0.0) | ||
HSIL | 11 (29.7) | 3 (6.2) | 0 (0.0) | 0 (0.0) | ||
AGC | 1 (2.7) | 1 (2.1) | 0 (0.0) | 1 (4.8) | ||
Clinically suspicious image of the cervix | 1 (2.7) | 1 (2.1) | 0 (0.0) | 1 (4.8) | ||
HPV negative, n (%) | 3 (8.1) | 9 (18.8) | 0.279 | 1 (50.0) | 14 (66.7) | >0.999 |
HPV genotype, n (%) * | ||||||
16 | 22 (59.5) | 16 (33.3) | 0.029 | 0 (0.0) | 3 (14.3) | >0.999 |
18 | 2 (5.4) | 2 (4.2) | >0.999 | 0 (0.0) | 0 (0.0) | - |
31 | 5 (13.5) | 9 (18.8) | 0.726 | 0 (0.0) | 0 (0.0) | - |
Biopsy, n (%) | ||||||
LSIL | 5 (13.5) | 11 (22.9) | 0.208 | 0 (0.0) | 0 (0.0) | >0.999 |
HSIL | 25 (67.6) | 33 (68.8) | 0 (0.0) | 0 (0.0) | ||
Adenocarcinoma | 2 (5.4) | 0 (0.0) | 0 (0.0) | 0 (0.0) | ||
Squamous cell carcinoma | 2 (5.4) | 0 (0.0) | 0 (0.0) | 0 (0.0) | ||
No signs of atypia | 2 (5.4) | 1 (2.1) | 1 (50.0) | 12 (57.1) | ||
Not performed | 1 (2.7) | 3 (6.2) | 1 (50.0) | 9 (42.9) | ||
LEEP/hysterectomy, n (%) | ||||||
LSIL | 3 (8.1) | 12 (25.0) | 0.019 | 0 (0.0) | 0 (0.0) | 0.395 |
HSIL | 28 (75.7) | 28 (58.3) | 0 (0.0) | 0 (0.0) | ||
Adenocarcinoma | 2 (5.4) | 0 (0.0) | 0 (0.0) | 0 (0.0) | ||
Squamous cell carcinoma | 2 (5.4) | 0 (0.0) | 0 (0.0) | 0 (0.0) | ||
No signs of atypia | 1 (2.7) | 6 (12.5) | 1 (50.0) | 4 (19.0) | ||
Not performed | 1 (2.7) | 2 (4.2) | 1 (50.0) | 17 (81.0) | ||
Final, n (%) | ||||||
LSIL | 4 (10.8) | 11 (22.9) | 0.048 | 0 (0.0) | 0 (0.0) | - |
HSIL | 29 (78.4) | 37 (77.1) | 0 (0.0) | 0 (0.0) | ||
Adenocarcinoma | 2 (5.4) | 0 (0.0) | 0 (0.0) | 0 (0.0) | ||
Squamous cell carcinoma | 2 (5.4) | 0 (0.0) | 0 (0.0) | 0 (0.0) | ||
No signs of atypia | 0 (0.0) | 0 (0.0) | 2 (100.0) | 21 (100.0) |
Final Outcome | Sensitivity, % | Specificity, % | PPV, % | NPV, % | Accuracy, % | |||
---|---|---|---|---|---|---|---|---|
HSIL/Adenocarcinoma/Squamous Cell Carcinoma | LSIL | |||||||
Methylation | + | 33 | 4 | 47.14 (35.09–59.45) | 73.33 (44.90–92.21) | 89.19 (77.47–95.19) | 22.92 (16.94–30.24) | 51.76 (40.66–62.74) |
− | 37 | 11 |
Final Outcome | Sensitivity, % | Specificity, % | PPV, % | NPV, % | Accuracy, % | |||
---|---|---|---|---|---|---|---|---|
HSIL/Adenocarcinoma/Squamous Cell Carcinoma | LSIL/No Signs of Atypia | |||||||
Methylation | + | 33 | 6 | 47.14 (35.09–59.45) | 84.62 (69.47–94.14) | 84.62 (71.67–92.28) | 47.14 (40.78–53.60) | 60.55 (50.73–69.78) |
− | 37 | 33 |
Characteristics | Biopsy = HSIL and LEEP = LSIL/No Signs of Atypia n = 12 | Other Patients n = 67 | p |
---|---|---|---|
Methylation, n (%) | |||
Negative | 11 (91.7) | 32 (47.8) | 0.013 |
Positive | 1 (8.3) | 35 (52.2) |
Biopsy = HSIL and LEEP = LSIL/No Signs of Atypia | Other Patients | Sensitivity, % | Specificity, % | PPV, % | NPV, % | Accuracy, % | ||
---|---|---|---|---|---|---|---|---|
Methylation | − | 11 | 32 | 91.67 (61.52–99.79) | 52.24 (39.67–64.60) | 25.58 (20.25–31.76) | 97.22 (84.09–99.57) | 58.23 (46.59–69.23) |
− | 1 | 35 |
Characteristics | Biopsy = LSIL and LEEP = LSIL/No Signs of Atypia n = 7 | Other Patients n = 72 | p |
---|---|---|---|
Methylation, n (%) | |||
Negative | 6 (85.7) | 37 (51.4) | 0.119 |
Positive | 1 (14.3) | 35 (48.6) |
Biopsy = LSIL and LEEP = LSIL/No Signs of Atypia | Other Patients | Sensitivity, % | Specificity, % | PPV, % | NPV, % | Accuracy, % | ||
---|---|---|---|---|---|---|---|---|
Methylation | − | 6 | 37 | 85.71 (42.13–99.64) | 48.61 (36.65–60.69) | 13.95 (10.01–19.12) | 97.22 (84.88–99.54) | 51.90 (40.36–63.29) |
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Pruski, D.; Millert-Kalińska, S.; Lis, A.; Pelc, E.; Konopelski, P.; Jach, R.; Przybylski, M. Evaluation of the Use of Methylation as a New Tool for the Diagnostics and Progression of Squamous Intraepithelial Lesions. Int. J. Mol. Sci. 2024, 25, 11863. https://doi.org/10.3390/ijms252211863
Pruski D, Millert-Kalińska S, Lis A, Pelc E, Konopelski P, Jach R, Przybylski M. Evaluation of the Use of Methylation as a New Tool for the Diagnostics and Progression of Squamous Intraepithelial Lesions. International Journal of Molecular Sciences. 2024; 25(22):11863. https://doi.org/10.3390/ijms252211863
Chicago/Turabian StylePruski, Dominik, Sonja Millert-Kalińska, Agata Lis, Ewa Pelc, Przemysław Konopelski, Robert Jach, and Marcin Przybylski. 2024. "Evaluation of the Use of Methylation as a New Tool for the Diagnostics and Progression of Squamous Intraepithelial Lesions" International Journal of Molecular Sciences 25, no. 22: 11863. https://doi.org/10.3390/ijms252211863
APA StylePruski, D., Millert-Kalińska, S., Lis, A., Pelc, E., Konopelski, P., Jach, R., & Przybylski, M. (2024). Evaluation of the Use of Methylation as a New Tool for the Diagnostics and Progression of Squamous Intraepithelial Lesions. International Journal of Molecular Sciences, 25(22), 11863. https://doi.org/10.3390/ijms252211863