Tumor LINE-1 Methylation Level in Association with Survival of Patients with Stage II Colon Cancer
Abstract
:1. Introduction
2. Results
2.1. Patient Characteristics
2.2. Methylation Assay Verification
2.3. LINE-1 Methylation Level and Patient Survival
3. Discussion
4. Materials and Methods
4.1. Patient Selection
4.2. DNA Extraction from Formalin-Fixed Paraffin-Embedded Tumor Tissues and Bisulfite Conversion
4.3. Quantitative Real-Time PCR
4.4. Analysis of LINE-1 Methylation Levels
4.5. MLH1 Methylation Status
4.6. Statistical Analysis
Supplementary Materials
Author Contributions
Conflicts of Interest
Abbreviations
CRC | Colorectal cancer |
DFS | Disease-free survival |
HEX | Hexachloro-Fluorescein |
FAM | 6-Fluorescein amidite |
FFPE | Formalin-fixed paraffin-embedded |
LINE-1 | Long interspersed nucleotide element |
MGB | Minor-groove-binding |
MSI | Microsatellite instable |
OS | Overall survival |
RFS | Recurrence-free survival |
RFU | Relative fluorescence units |
(q)PCR | (Quantitative) polymerase chain reaction |
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Total n = 164 (%) | LINE-1 | p-Value | ||
---|---|---|---|---|
Hypomethylated a n = 54 (%) | Methylated b n = 110 (%) | |||
Gender | ||||
Male | 86 (52.4) | 30 (55.6) | 56 (65.1) | 0.58 |
Female | 78 (47.6) | 24 (44.4) | 54 (49.1) | |
Age median | 68.0 (±13.6) | 69.5 (±12.0) | 68.0 (±14.3) | 0.16 |
Age Groups | ||||
≤65 | 66 (40.2) | 17 (31.5) | 49 (44.5) | 0.11 |
>65 | 98 (59.8) | 37 (68.5) | 61 (55.5) | |
Grade | ||||
Good | 32 (19.5) | 11 (20.4) | 21 (19.1) | 0.24 |
Moderate | 79 (48.2) | 30 (55.6) | 49 (48.2) | |
Poor | 20 (12.2) | 7 (13.0) | 13 (11.8) | |
Unknown | 33 (9.8) | 6 (11.1) | 27 (24.5) | |
Location | ||||
Right | 74 (45.1) | 22 (40.7) | 52 (47.3) | 0.43 |
Left | 90 (54.9) | 32 (59.3) | 58 (52.7) | |
MLH1 Promoter | ||||
Methylated | 30 (18.3) | 6 (11.1) | 24 (21.8) | 0.10 |
Unmethylated | 134 (81.7) | 48 (88.9) | 86 (78.2) | |
Adjuvant Therapy | ||||
No | 156 (95.1) | 50 (92.6) | 106 (96.4) | 0.29 |
Yes | 8 (49) | 4 (7.4) | 4 (3.6) |
Patients n = 164 | Univariate | Multivariate a | |||
---|---|---|---|---|---|
HR (95% CI) | p-Value | HR (95% CI) | p-Value | ||
Overall Survival | |||||
LINE-1 Methylated | 110 | 1.00 (reference) | 0.02 | 1.00 (reference) | 0.04 |
LINE-1 Hypomethylated | 54 | 1.75 (1.09–2.81) | 1.68 (1.03–2.75) | ||
Disease-free Survival b | |||||
LINE-1 Methylated | 110 | 1.00 (reference) | 0.07 | 1.00 (reference) | 0.13 |
LINE-1 Hypomethylated | 54 | 1.52 (0.97–2.40) | 1.43 (0.90–2.29) | ||
Recurrence-free Survival c | |||||
LINE-1 Methylated | 110 | 1.00 (reference) | 0.37 | 1.00 (reference) | 0.33 |
LINE-1 Hypomethylated | 54 | 1.40 (0.66–2.97) | 1.46 (0.68–3.15) | ||
Overall Survival by MLH1 Status | |||||
MLH1 Unmethylated | 134 | ||||
LINE-1 Methylated | 86 | 1.00 (reference) | 0.02 | 1.00 (reference) | 0.09 |
LINE-1 Hypomethylated | 48 | 1.85 (1.10–3.11) | 1.56 (0.93–2.63) | ||
MLH1 Methylated | 30 | ||||
LINE-1 Methylated | 24 | 1.00 (reference) | 0.89 | - | - |
LINE-1 Hypomethylated | 6 | 1.09 (0.29–4.07) | - |
Patients n = 164 | Univariate | Multivariate a | p-Value for Interaction b | |||
---|---|---|---|---|---|---|
HR (95%CI) | p-Value | HR (95% CI) | p-Value | |||
Overall Survival | ||||||
≤65 years | 66 | 0.24 | ||||
LINE-1 Methylated | 49 | 1.00 (reference) | 0.82 | 1.00 (reference) | 0.80 | |
LINE-1 Hypomethylated | 17 | 0.81 (0.22–3.04) | 0.84 (0.23–3.16) | |||
>65 years | 98 | |||||
LINE-1 Methylated | 61 | 1.00 (reference) | 0.01 | 1.00 (reference) | 0.02 | |
LINE-1 Hypomethylated | 37 | 1.93 (1.15–3.26) | 2.00 (1.13–3.52) | |||
Disease-Free Survival c | ||||||
≤65 years | 66 | 0.31 | ||||
LINE-1 Methylated | 49 | 1.00 (reference) | 0.76 | 1.00 (reference) | 0.71 | |
LINE-1 Hypomethylated | 17 | 0.84 (0.27–2.56) | 0.71 (0.26–2.53) | |||
>65 years | 98 | |||||
LINE-1 Methylated | 61 | 1.00 (reference) | 0.05 | 1.00 (reference) | 0.04 | |
LINE-1 Hypomethylated | 37 | 1.67 (1.00–2.77) | 1.76 (1.02–3.05) | |||
Recurrence-Free Survival | ||||||
≤65 years | 66 | 0.35 | ||||
LINE-1 Methylated | 49 | 1.00 (reference) | 0.93 | 1.00 (reference) | 0.99 | |
LINE-1 Hypomethylated | 17 | 0.94 (0.25–3.55) | 1.00 (0.26–3.08) | |||
>65 years | 98 | |||||
LINE-1 Methylated | 61 | 1.00 (reference) | 0.28 | 1.00 (reference) | 0.11 | |
LINE-1 Hypomethylated | 37 | 1.67 (0.66–4.23) | 2.32 (0.83–6.52) |
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Swets, M.; Zaalberg, A.; Boot, A.; Van Wezel, T.; Frouws, M.A.; Bastiaannet, E.; Gelderblom, H.; Van de Velde, C.J.H.; Kuppen, P.J.K. Tumor LINE-1 Methylation Level in Association with Survival of Patients with Stage II Colon Cancer. Int. J. Mol. Sci. 2017, 18, 36. https://doi.org/10.3390/ijms18010036
Swets M, Zaalberg A, Boot A, Van Wezel T, Frouws MA, Bastiaannet E, Gelderblom H, Van de Velde CJH, Kuppen PJK. Tumor LINE-1 Methylation Level in Association with Survival of Patients with Stage II Colon Cancer. International Journal of Molecular Sciences. 2017; 18(1):36. https://doi.org/10.3390/ijms18010036
Chicago/Turabian StyleSwets, Marloes, Anniek Zaalberg, Arnoud Boot, Tom Van Wezel, Martine A. Frouws, Esther Bastiaannet, Hans Gelderblom, Cornelis J. H. Van de Velde, and Peter J. K. Kuppen. 2017. "Tumor LINE-1 Methylation Level in Association with Survival of Patients with Stage II Colon Cancer" International Journal of Molecular Sciences 18, no. 1: 36. https://doi.org/10.3390/ijms18010036