TLR5 Variants Are Associated with the Risk for COPD and NSCLC Development, Better Overall Survival of the NSCLC Patients and Increased Chemosensitivity in the H1299 Cell Line
Abstract
:1. Introduction
2. Materials and Methods
2.1. Study Participants
2.2. SNP Selection Criterion
2.3. DNA Sample Preparation and Genotyping
2.4. Statistical Analysis for Association Studies
2.5. Cells Cultures
2.6. Generation of Adenoviral Vectors Containing TLR5WT and the TLR5N592S as a Transgenes
2.7. Reporter Gene Assays
2.8. Immunoblot Analysis
2.9. Proliferation Assay
2.10. Serum Concentration of the Cytokines
2.11. RNA-Seq Library Preparation and Sequencing
2.12. Bioinformatics Analysis
2.13. Statistical Analysis
3. Results
3.1. TLR5 Variant N592S Is Associated with an Increased Risk for COPD and NSCLC Development
3.2. TLR5 Variants Are Associated with the Lymph Node Involment (N Status) and Overall Survival in NSCLC Patients
3.3. N592S Variant Affect the Activation of the NF-κB and AP-1 Transcription Factors
3.4. Activation of p38 and ERK Is Affected by the N592S TLR5 Coding Variant in the WI-38 Cell Line
3.5. H1299 Cells Overexpressing N592S Variant Exhibit Increased Chemosensitivity
3.6. ELISA
3.7. Transcriptome Analysis
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Subjects (N, %) | Cases (974) | COPD (500; 51.3%) | COPD + LC (280; 28.7%) | LC (194; 19.9%) | Controls (1283) |
---|---|---|---|---|---|
Age, year (SD 1) | 60 (9.58) | 59 (10.4) | 63 (7.38) | 62 (9.8) | 38 (12) |
Sex (N, %) | |||||
male | 671 (68.9) | 355 (71) | 205 (73) | 111 (57.3) | 1120 (87) |
female | 303 (31.1) | 145 (29) | 75 (26) | 83 (42.7) | 163 (12.7) |
Smoking status (N, %) | |||||
active | 523 (50.8) | 139 (25.8) | 148 (52.5) | 53 (26.3) | 325 (25.3) |
ex-smoker | 347 (33.6) | 317 (57.4) | 124 (43.9) | 90 (43.8) | 4 (0.2) |
non | 104 (9.8) | 44 (8) | 8 (2.5) | 51 (25.3) | 954 (74.4) |
CHARACTERISTICS | COPD (N = 500) | COPD + LC and LConly (N = 474) |
---|---|---|
FEV1 % (N) 1 | ||
80–120% | 55 | 235 |
50–80% | 207 | 185 |
30–50% | 148 | 42 |
≤30% | 90 | 12 |
TYPE OF CANCER | ||
Lung cancer | / | 474 |
NSCLC/AdC | / | 225 |
NSCLC/SQC | / | 168 |
other | / | 81 |
TNM-stage (N) | ||
1 | / | 118 |
2 | / | 100 |
3 | / | 102 |
4 | / | 77 |
nonclasified | / | 77 |
T stage (N) | ||
1 | / | 60 |
2 | / | 149 |
3 | / | 58 |
4 | / | 25 |
nonclasified | / | 182 |
N stage (N) | ||
0 | / | 153 |
1 | / | 55 |
2 | / | 109 |
3 | / | 8 |
nonclasified | / | 149 |
M stage (N) | ||
0 | / | 258 |
1 | / | 60 |
nonclasified | / | 156 |
SNP | Chr. | Position | Allele | Location | MAF 1 | Aa Change | Sift 2 | PolyPhen 3 |
---|---|---|---|---|---|---|---|---|
rs5744174 | 1 | 223284528 | C/T | Exon | 0.45 (T) | F616L | Tolerated | Benign |
rs2072493 | 1 | 223284599 | A/G | Exon | 0.12 (G) | N592S | Tolerated | Benign |
rs725084 | 1 | 223323951 | T/C | promotor | 0.44 (T) | / | / | / |
COPD vs. Healthy | Lung Cancer vs. Healthy | NSCLC vs. Healthy | |||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
SNP | Genotype | Cases (N, %) | Controls | OR (95% CI) | p Value | Cases (N, %) | Controls (N, %) | OR (95% CI) | p Value | Cases (N, %) | Controls (N, %) | OR (95% CI) | p Value |
rs5744174_TC | T/T | 216 (33.4) | 381 (30.8) | 1 | 145 (33.2) | 381 (30.8) | 1 | 132 (32.5) | 381 (30.8) | 1 | |||
(coding F616L) | T/C | 296 (45.7) | 606 (49) | 0.64 (0.32–1.28) | 0.206 | 210 (48) | 606 (49) | 1.09 (0.54–2.18) | 0.8073 | 199 (49) | 606 (49) | 1.33 (0.64–2.74) | 0.4384 |
C/C | 135 (20.9) | 249 (20.2) | 0.97 (0.47–2.01) | 0.9382 | 82 (18.8) | 249 (20.2) | 0.38 (0.13–1.14) | 0.0851 | 75 (18.5) | 249 (20.2) | 0.46 (0.15–1.43) | 0.1803 | |
T/C + C/C | 431 (66.6) | 855 (69.2) | 0.87 (0.51–1.51) | 0.6314 | 292 (66.8) | 855 (69.2) | 0.88 (0.45–1.71) | 0.6987 | 274 (67.5) | 855 (69.2) | 0.96 (0.56–1.64) | 0.8854 | |
rs725084_TC | T/T | 208 (32.4) | 325 (26.6) | 1 | 124 (28.8) | 325 (26.6) | 1 | 111 (28.1) | 325 (26.6) | 1 | |||
(promoter) | T/C | 301 (46.9) | 628 (51.4) | 0.85 (0.62–1.16) | 0.3126 | 217 (50) | 628 (51.4) | 1.12 (0.78–1.59) | 0.5431 | 201 (50.7) | 628 (51.4) | 1.07 (0.72–1.57) | 0.744 |
C/C | 133 (20.7) | 269 (22) | 0.83 (0.57–1.22) | 0.3502 | 91 (21.1) | 269 (22) | 0.81 (0.52–1.26) | 0.3496 | 84 (21.2) | 269 (22) | 0.82 (0.51–1.33) | 0.4311 | |
T/C + C/C | 434 (67.6) | 897 (73.4) | 0.85 (0.63–1.13) | 0.2635 | 308 (71.1) | 897 (73.4) | 1.02 (0.72–1.43) | 0.9213 | 285 (71.9) | 897 (73.4) | 0.99 (0.68–1.43) | 0.95 | |
rs2072493_AG | A/A | 413 (64.5) | 967 (80) | 1 | 248 (57.7) | 967 (80) | 1 | 226 (57.1) | 967 (80) | 1 | |||
(coding N592S) | A/G | 196 (30.6) | 226 (18.7) | 4.48 (2.34–8.57) | <0.0001 | 150 (34.9) | 226 (18.7) | 4.63 (2.06–10.39) | 0.0002 | 142 (35.8) | 226 (18.7) | 4.93 (2.12–11.49) | 0.0002 |
G/G | 31 (4.9) | 15 (1.3) | 4.08 (0.72–23.13) | 0.1122 | 32 (7.4) | 15 (1.3) | 3.94 (0.77–20.14) | 0.0995 | 28 (7.1) | 15 (1.3) | 8.44 (1.25–56.85) | 0.0284 | |
A/G + G/G | 227 (35.5) | 241 (20) | 4.41 (2.36–8.24) | <0.0001 | 182 (42.9) | 241 (20) | 4.61 (2.15–9.87) | 0.0001 | 170 (42.9) | 241 (20) | 5.17 (2.37–11.31) | <0.0001 |
NSCLC + COPD (Cases) vs. COPD (Controls) | |||||
---|---|---|---|---|---|
SNP | Genotype | Cases (N, %) | Controls (N, %) | OR (95% CI) | p Value |
rs5744174_TC | T/T | 43 (17.7) | 88 (22.6) | 1 | |
(coding F616L) | C/T | 118 (48.6) | 173 (44.3) | 1.41 (0.87–2.28) | 0.1558 |
C/C | 82 (33.7) | 129 (33.1) | 1.43 (0.86–2.36) | 0.1594 | |
C/T + C/C | 200 (82.3) | 302 (77.4) | 1.41 (0.90–2.19) | 0.1312 | |
rs725084_TC | T/T | 45 (18.7) | 84 (21.6) | 1 | |
(promoter) | C/T | 121 (50.4) | 174 (44.8) | 1.18 (0.73–1.91) | 0.4912 |
C/C | 74 (30.9) | 130 (33.6) | 1.06 (0.64–1.73) | 0.8156 | |
C/T + T/T | 195 (81.3) | 304 (78.4) | 1.13 (0.73–1.76) | 0.5683 | |
rs2072493_AG | A/A | 144 (59.5) | 264 (68.6) | 1 | |
(coding N592S) | A/G | 80 (33.1) | 110 (28.6) | 1.52 (1.02–2.25) | 0.0365 |
G/G | 18 (7.4) | 11 (2.8) | 4.49 (1.9–10.61) | 0.0006 | |
A/G + G/G | 98 (40.5) | 121 (31.4) | 1.75 (1.21–2.54) | 0.0031 |
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Baranašić, J.; Šutić, M.; Catalano, C.; Drpa, G.; Huhn, S.; Majhen, D.; Nestić, D.; Kurtović, M.; Rumora, L.; Bosnar, M.; et al. TLR5 Variants Are Associated with the Risk for COPD and NSCLC Development, Better Overall Survival of the NSCLC Patients and Increased Chemosensitivity in the H1299 Cell Line. Biomedicines 2022, 10, 2240. https://doi.org/10.3390/biomedicines10092240
Baranašić J, Šutić M, Catalano C, Drpa G, Huhn S, Majhen D, Nestić D, Kurtović M, Rumora L, Bosnar M, et al. TLR5 Variants Are Associated with the Risk for COPD and NSCLC Development, Better Overall Survival of the NSCLC Patients and Increased Chemosensitivity in the H1299 Cell Line. Biomedicines. 2022; 10(9):2240. https://doi.org/10.3390/biomedicines10092240
Chicago/Turabian StyleBaranašić, Jurica, Maja Šutić, Calogerina Catalano, Gordana Drpa, Stefanie Huhn, Dragomira Majhen, Davor Nestić, Matea Kurtović, Lada Rumora, Martina Bosnar, and et al. 2022. "TLR5 Variants Are Associated with the Risk for COPD and NSCLC Development, Better Overall Survival of the NSCLC Patients and Increased Chemosensitivity in the H1299 Cell Line" Biomedicines 10, no. 9: 2240. https://doi.org/10.3390/biomedicines10092240