The association of genetic factors with serum calretinin levels in asbestos-related diseases

Zupanc, Cita; Franko, Alenka; Strbac, Danijela; Kovac, Viljem; Dolzan, Vita; Goricar, Katja

doi:10.2478/raon-2023-0061

Figures & Tables

Association of selected single nucleotide polymorphisms (SNPs) with serum calretinin concentration: CALB2 rs889704 (A), E2F2 rs2075995 (B), MIR335 rs3807348 (C), NRF1 rs13241028 (D).

Association of investigated single nucleotide polymorphisms (SNPs) with malignant mesothelioma (MM) susceptibility

SNP	Genotype	Subjects without MM (N = 616) N (%)	MM patients (N = 288) N (%)	OR (95% CI)	P	OR (95% CI)_adj	P_adj
CALB2 rs1862818	CC	340 (55.2)	139 (48.3)	Reference		Reference
	CT	226 (36.7)	120 (41.7)	1.30 (0.97–1.75)	0.084	1.35 (0.97–1.87)	0.073
	TT	50 (8.1)	29 (10.1)	1.42 (0.86–2.34)	0.169	1.34 (0.77–2.32)	0.299
	CT+TT	276 (44.8)	149 (51.7)	1.32 (1.00–1.75)	0.052	1.35 (0.99–1.83)	0.059
CALB2 rs889704	CC	485 (78.9) [1]	223 (77.4)	Reference		Reference
	CA	121 (19.7)	61 (21.2)	1.10 (0.78–1.55)	0.602	1.03 (0.70–1.51)	0.899
	AA	9 (1.5)	4 (1.4)	0.97 (0.29–3.17)	0.955	0.55 (0.15–1.94)	0.349
	CA+AA	130 (21.1)	65 (22.6)	1.09 (0.78–1.52)	0.626	0.98 (0.67–1.42)	0.912
CALB2 rs8063760	CC	352 (57.3) [2]	175 (60.8)	Reference		Reference
	CT	222 (36.2)	97 (33.7)	0.88 (0.65–1.19)	0.398	0.91 (0.66–1.26)	0.576
	TT	40 (6.5)	16 (5.6)	0.80 (0.44–1.48)	0.483	0.82 (0.42–1.60)	0.554
	CT+TT	262 (42.7)	113 (39.2)	0.87 (0.65–1.15)	0.329	0.90 (0.65–1.23)	0.493
E2F2 rs2075995	CC	117 (19.0)	70 (24.3)	Reference		Reference
	CA	319 (51.8)	149 (51.7)	0.78 (0.55–1.11)	0.171	0.83 (0.56–1.23)	0.349
	AA	180 (29.2)	69 (24.0)	0.64 (0.43–0.96)	0.032	0.68 (0.44–1.07)	0.093
	CA+AA	499 (81.0)	218 (75.7)	0.73 (0.52–1.02)	0.067	0.78 (0.53–1.13)	0.182
MIR335 rs3807348	GG	158 (25.8) [3]	70 (24.3)	Reference		Reference
	GA	307 (50.1)	139 (48.3)	1.02 (0.72–1.44)	0.902	1.00 (0.68–1.46)	0.98
	AA	148 (24.1)	79 (27.4)	1.20 (0.81–1.78)	0.352	1.22 (0.79–1.87)	0.376
	GA+AA	455 (74.2)	218 (75.7)	1.08 (0.78–1.50)	0.636	1.07 (0.75–1.52)	0.724
NRF1 rs13241028	TT	374 (60.7)	173 (60.1)	Reference		Reference
	TC	210 (34.1)	103 (35.8)	1.06 (0.79–1.43)	0.699	1.08 (0.78–1.50)	0.636
	CC	32 (5.2)	12 (4.2)	0.81 (0.41–1.61)	0.550	0.92 (0.44–1.93)	0.823
	TC+CC	242 (39.3)	115 (39.9)	1.03 (0.77–1.37)	0.853	1.06 (0.78–1.45)	0.711
SEPTIN7 rs3801339	TT	109 (17.7)	55 (19.1)	Reference		Reference
	TC	266 (43.2)	135 (46.9)	1.01 (0.68–1.48)	0.976	1.05 (0.69–1.61)	0.815
	CC	241 (39.1)	98 (34.0)	0.81 (0.54–1.20)	0.291	0.76 (0.49–1.18)	0.218
	TC+CC	507 (82.3)	233 (80.9)	0.91 (0.64–1.30)	0.610	0.91 (0.61–1.35)	0.627

Receiver operating characteristic (ROC) curve analysis according to individual genotypes for selected single nucleotide polymorphisms: comparison of malignant mesothelioma (MM) patients with all other subjects

SNP	Genotype	AUC (95% CI)	P	Calretinin cut-off (ng/ml)¹	Sensitivity	Specificity
Overall analysis in the whole group	/	0.825 (0.781–0.868)	< 0.001	0.32	0.681	0.887
CALB2 rs889704	CC	0.831 (0.782–0.880)	< 0.001	0.32	0.695	0.876
	CA	0.779 (0.667–0.891)	< 0.001	0.31	0.607	0.935
	AA²	0.958 (0.837–1.000)	0.019	0.21	1.000	0.833
	CA+AA	0.801 (0.702–0.901)	< 0.001	0.31	0.625	0.940
E2F2 rs2075995	CC	0.906 (0.845–0.968)	< 0.001	0.26	0.810	0.903
	CA	0.803 (0.736–0.869)	< 0.001	0.32	0.671	0.888
	AA	0.781 (0.686–0.876)	< 0.001	0.33	0.615	0.877
	CA+AA	0.797 (0.742–0.851)	< 0.001	0.32	0.653	0.881
MIR335 rs3807348	GG	0.853 (0.766–0.940)	< 0.001	0.29	0.757	0.872
	GA	0.803 (0.739–0.867)	< 0.001	0.32	0.643	0.892
	AA	0.845 (0.765–0.925)	< 0.001	0.35	0.738	0.881
	GA+AA	0.815 (0.764–0.866)	< 0.001	0.32	0.675	0.881
NRF1 rs13241028	TT	0.812 (0.754–0.871)	< 0.001	0.32	0.693	0.884
	TC	0.868 (0.804–0.931)	< 0.001	0.23	0.818	0.798
	CC³	0.664 (0.406–0.922)	0.203	0.18	0.714	0.700
	TC+CC	0.842 (0.777–0.907)	< 0.001	0.23	0.790	0.785

Clinical characteristics of the subjects included in the study

Characteristic	Category/unit	No disease (N = 83)	Pleural plaques (N = 380)	Asbestosis (N = 153)	MM (N = 288)	P
Sex	Male, N (%)	61 (73.5)	262 (68.9)	119 (77.8)	213 (74.0)	0.180¹
Sex	Female, N (%)	22 (26.5)	118 (31.1)	34 (22.2)	75 (26.0)
Age	Median (25%–75%)	53.4 (48.5–59.2)	54.8 (48.8–62.7)	59.4 (51.3–66.1)	66.0 (59–73)	< 0.001²
Smoking	No, N (%)	46 (55.4)	187 (49.2)	74 (48.4)	158 (56.4) [8]	0.205¹
Smoking	Yes, N (%)	37 (44.6)	193 (50.8)	79 (51.6)	122 (43.6)

Association of selected SNPs with serum calretinin concentration

SNP	Genotype	All subjects			Subjects without MM			MM patients

		Calretinin (ng/ml) Median (25–75%)	P_add	P_dom	Calretinin (ng/ml) Median (25–75%)	P_add	P_dom	Calretinin (ng/ml) Median (25–75%)	P_add	P_dom
CALB2 rs1862818	CC	0.18 (0.11–0.34)	0.622	0.422	0.15 (0.09–0.22)	0.751	0.865	0.64 (0.22–1.45)	0.952	0.802
	CT	0.19 (0.11–0.41)			0.16 (0.09–0.24)			0.51 (0.23–1.41)
	TT	0.18 (0.10–0.37)			0.13 (0.08–0.20)			0.38 (0.21–3.57)
	CT+TT	0.19 (0.11–0.40)			0.15 (0.09–0.24)			0.48 (0.23–1.43)
CALB2 rs889704	CC	0.19 (0.11–0.37)	0.099	0.036	0.15 (0.10–0.23)	0.130	0.069	0.52 (0.25–1.43)	0.508	0.441
	CA	0.17 (0.08–0.27)			0.16 (0.08–0.21)			0.44 (0.14–1.35)
	AA	0.21 (0.05–0.77)			0.10 (0.02–0.21)			1.07 (0.28–1.84)
	CA+AA	0.17 (0.08–0.28)			0.14 (0.07–0.21)			0.50 (0.15–1.51)
CALB2 rs8063760	CC	0.18 (0.11–0.38)	0.955	0.770	0.14 (0.09–0.22)	0.382	0.647	0.53 (0.24–1.44)	0.326	0.768
	CT	0.18 (0.12–0.32)			0.16 (0.1–0.24)			0.44 (0.19–1.30)
	TT	0.21 (0.06–0.51)			0.12 (0.05–0.22)			0.86 (0.50–2.30)
	CT+TT	0.19 (0.11–0.34)			0.16 (0.09–0.24)			0.51 (0.21–1.43)
E2F2 rs2075995	CC	0.19 (0.10–0.46)	0.512	0.481	0.14 (0.08–0.2)	0.161	0.059	0.72 (0.33–1.45)	0.189	0.117
	CA	0.18 (0.12–0.34)			0.16 (0.1–0.23)			0.53 (0.20–1.48)
	AA	0.18 (0.10–0.33)			0.14 (0.09–0.24)			0.40 (0.18–0.90)
	CA+AA	0.18 (0.11–0.34)			0.15 (0.1–0.23)			0.48 (0.20–1.44)
MIR335 rs3807348	GG	0.18 (0.09–0.34)	0.057	0.151	0.14 (0.08–0.2)	0.027	0.081	0.44 (0.26–1.43)	0.400	0.978
	GA	0.18 (0.11–0.34)			0.14 (0.09–0.22)	AA vs. GG P = 0.029		0.50 (0.18–1.16)
	AA	0.21 (0.13–0.39)			0.18 (0.11–0.26)			0.65 (0.27–1.80)
	GA+AA	0.19 (0.11–0.37)			0.15 (0.1–0.23)			0.52 (0.22–1.44)
NRF1 rs13241028	TT	0.19 (0.12–0.36)	0.272	0.144	0.16 (0.1–0.23)	0.096	0.034	0.52 (0.21–1.15)	0.381	0.672
	TC	0.18 (0.10–0.33)			0.14 (0.08–0.21)			0.64 (0.25–1.67)
	CC	0.17 (0.07–0.36)			0.15 (0.07–0.3)			0.24 (0.07–1.18)
	TC+CC	0.18 (0.09–0.34)			0.14 (0.08–0.21)			0.46 (0.24–1.53)
SEPTIN7 rs3801339	TT	0.18 (0.11–0.34)	0.403	0.419	0.14 (0.09–0.2)	0.424	0.288	0.35 (0.17–1.05)	0.079	0.080
	TC	0.18 (0.11–0.33)			0.15 (0.09–0.22)			0.51 (0.21–1.23)
	CC	0.20 (0.11–0.45)			0.16 (0.09–0.25)			0.72 (0.38–1.48)
	TC+CC	0.19 (0.11–0.37)			0.15 (0.09–0.23)			0.64 (0.26–1.45)

Association of CALB2 haplotypes with malignant mesothelioma (MM) susceptibility and serum calretinin concentration

Haplotype	Subjects without MM Predicted frequency	MM patients Predicted frequency	OR (95% CI)	P	OR (95% CI)_adj	P_adj	Serum calretinin concentration P
CCC	0.457	0.431	Reference		Reference
TCC	0.245	0.294	1.26 (0.0–991.60)	0.061	1.26 (0.97–1.64)	0.084	0.272
CCT	0.176	0.147	0.88 (0.65–1.20)	0.415	0.94 (0.66–1.34)	0.731	0.125
CAT	0.058	0.066	1.21 (0.77–1.89)	0.408	1.08 (0.64–1.81)	0.782	0.731
CAC	0.045	0.047	1.11 (0.64–1.91)	0.713	0.99 (0.55–1.79)	0.974	0.852

Genotype frequencies of investigated single nucleotide polymorphisms (SNPs) in the whole study group, their variant allele frequency (VAF) and agreement with Hardy-Weinberg equilibrium (HWE) in subjects without any asbestos-related disease (controls)

Gene	SNP	Nucleotide or amino acid change	Predicted function	Genotype	N (%)	VAF (controls)	pHWE (controls)
*CALB2*	rs1862818	c.-828C>T	May influence transcription factor binding, may alter chromatin states and regulatory motifs	CC	479 (53.0)	0.27	0.617
				CT	346 (38.3)
				TT	79 (8.7)
*CALB2*	rs889704	c.-634C>A	May influence transcription factor binding, may alter chromatin states and regulatory motifs	CC	708 (78.4) [1]	0.14	0.814
				CA	182 (20.2)
				AA	13 (1.4)
*CALB2*	rs8063760	c.*138T>C	May influence miRNA binding, may alter regulatory motifs	CC	527 (58.4) [2]	0.23	0.322
				CT	319 (35.4)
				TT	56 (6.2)
*E2F2*	rs2075995	c.678C>A, p.Gln226His	Nonsynonymous, may influence splicing	CC	187 (20.7)	0.61	0.209
				CA	468 (51.8)
				AA	249 (27.5)
*MIR335*	rs3807348	g.130496266G>A	Downstream transcript variant, may influence transcription factor binding	GG	228 (25.3) [3]	0.49	0.376
				GA	446 (49.5)
				AA	227 (25.2)
*NRF1*	rs13241028	c.*1321T>C	May influence miRNA binding	TT	547 (60.5)	0.22	0.061
				TC	313 (34.6)
				CC	44 (4.9)
*SEPTIN7*	rs3801339	c.1168-4451T>C	Genic downstream transcript variant¹	TT	164 (18.1)	0.63	0.187
				TC	401 (44.4)
				CC	339 (37.5)

The association of genetic factors with serum calretinin levels in asbestos-related diseases

Figures & Tables

FIGURE 1.

Association of investigated single nucleotide polymorphisms (SNPs) with malignant mesothelioma (MM) susceptibility

Receiver operating characteristic (ROC) curve analysis according to individual genotypes for selected single nucleotide polymorphisms: comparison of malignant mesothelioma (MM) patients with all other subjects

Clinical characteristics of the subjects included in the study

Association of selected SNPs with serum calretinin concentration

Association of CALB2 haplotypes with malignant mesothelioma (MM) susceptibility and serum calretinin concentration

Genotype frequencies of investigated single nucleotide polymorphisms (SNPs) in the whole study group, their variant allele frequency (VAF) and agreement with Hardy-Weinberg equilibrium (HWE) in subjects without any asbestos-related disease (controls)

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