Copy number variation of ubiquitin- specific proteases genes in blood leukocytes and colorectal cancer

Figures & data

Table 1. Basic characteristics of cases and controls.

Characteristic	No.of case (%)	No.of controls (%)	P value*
Age	60.56 ± 11.06	59.70 ± 10.68	0.23
Gender			0.39
Male	254 (54.3)	254 (54.3)
Female	214 (45.7)	214 (45.7)
BMI (kg/m^2)	23.77 ± 3.84	23.40 ± 4.33	0.53
Education^a			0.63
Primary school and below	259 (59.4)	248 (56.5)
Junior middle school	88 (20.2)	91 (20.7)
Senior middle school and above	89 (20.4)	100 (22.8)
Occupation^a			< 0.001
White collar	88 (19.7)	57 (13.0)
Blue collar	228 (51.1)	264 (60.4)
Both	130 (29.1)	116 (26.5)
Family history of CRC^a			< 0.001
Yes	35 (9.9)	152 (34.0)
No	318 (90.1)	295 (66.0)
Location of primary tumor
Colon	227 (61.9)
Rectum	145 (38.1)
TNM stage ^a
I	36 (11.0)
II	138 (42.1)
III	119 (36.3)
IV	35 (10.6)
I+ II	174 (53.1)
III+ IV Microsatellite instability ^b MSS MSI-L MSI-H	154 (46.9) 249 (81.9) 37 (12.2) 18 (5.9)

^aMissing data of education: 32 cases, 29 controls; occupation: 22 cases, 31 controls; family history of CRC: 115 cases, 21 controls; location of primary tumor: 96 cases; TNM stage: 140 cases.

^bMSI-L, low-level microsatellite instability, MSI-H, high-level microsatellite instability; MSS, microsatellite stability.

*P < 0.05 was considered statistically significant.

Table 2. Associations between CYLD, USP9X and USP11 CNVs and the risk of CRC.

Gene	No.of case (%)	No.of controls (%)	Odds Ratio^a	95%Confidence Interval	P Value*
CYLD^b
Wt	286 (62.7)	312 (68.4)	1.00
Del	54 (11.8)	21 (4.6)	4.18	2.02, 8.62	< 0.001
Amp	116 (25.5)	123 (27.0)	0.94	0.62, 1.43	0.79
Del v.s Wt + Amp			4.26	2.10, 8.67	< 0.001
Del + Wt v.s. Amp			1.30	0.88, 1.94	0.19
Del +Amp v.s. Wt			0.76	0.52, 1.10	0.14
USP9X^b
Wt	151 (33.6)	187 (41.6)	1.00
Del	31 (6.9)	45 (10.0)	0.96	0.48, 1.92	0.92
Amp	268 (59.5)	218 (48.4)	2.30	1.48, 3.57	< 0.001
Wt + Amp v.s Del			1.54	0.81, 2.94	0.19
Amp v.s. Del + Wt			2.32	1.53, 3.53	< 0.001
Del + Amp v.s. Wt			1.93	1.28, 2.90	0.002
USP11^b
Wt	245 (55.8)	258 (58.8)	1.00
Del	68 (15.5)	52 (11.8)	3.49	1.49, 8.64	0.007
Amp	126 (28.7)	129 (29.4)	0.91	0.50, 1.66	0.76
Del v.s Wt + Amp			3.52	1.43, 8.71	0.006
Del + Wt v.s. Amp			1.17	0.64, 2.14	0.59
Del + Amp v.s. Wt			1.42	0.87, 2.31	0.16

^aAdjusted for occupation and family history of CRC. ^bMissing value, CYLD, 24; USP9X, 36; USP11, 58. *P < 0.05 was considered statistically significant.

Table 3. Interactions between three genes CNVs and environmental factors on the risk of CRC.

CNV genotypes		Environmental factors			Interaction
CYLD		Roughage (g/week)
		≤50		>50
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt + Amp	1.00		0.55 (0.37, 0.81)
	Del	3.37 (1.11, 10.25)		3.12 (1.21, 8.08)	1.70 (0.39, 7.39)	0.48
		Fat meat
		NO		YES
		OReg (95% CI)		ORe g(95% CI)		P value*
	Wt+Amp	1.00		1.19 (0.72, 1.83)
	Del	5.16 (2.15, 12.35)		3.38 (1.01, 11.30)	0.57 (0.13, 2.54)	0.46
		Fish (times/week)
		≤1		>1
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt+Amp	1.00		0.63 (0.41, 0.97)
	Del	10.92 (5.81, 20.49)		1.39 (0.69, 2.83)	0.20 (0.04, 0.99)	0.05
		Egg (/week)
		≤3		>3
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt	1.00		1.72 (1.08, 2.73)
	Del+Amp	1.29 (0.70, 2.36)		2.31 (1.38, 3.86)	1.05 (0.47, 2.34)	0.91
	Wt+Amp	1.00		1.90 (1.29, 2.76)
	Del	7.54 (2.37, 24.03)		5.36 (2.17, 13.24)	0.38 (0.09, 1.61)	0.19
		Sausage (times/month)
		≤1		>1
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt	1.00		1.87 (0.87, 3.99)
	Del+Amp	1.29 (0.82, 2.02)		2.79 (1.29, 6.07)	1.16 (0.38, 3.52)	0.79
	Wt+Amp	1.00		1.33 (0.99, 1.82)
	Del	1.68 (1.15, 2.46)		1.98 (1.15, 3.43)	0.89 (0.44, 1.81)	0.75
		Leftovers^b (times/week)
		≤3		>3
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt	1.00		1.96 (1.24, 3.09)
	Del+Amp	1.60 (0.97, 2.62)		2.23 (1.27, 3.92)	0.71 (0.33, 1.53)	0.39
	Wt+Amp	1.00		1.78 (1.22, 2.62)
	Del	5.32 (2.04, 13.86)		5.40 (1.83, 15.92)	0.57 (0.13, 2.45)	0.45
USP9X		Fish (times/week)
		≤1		>1
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt	1.00		1.00 (0.54, 1.85)
	Del+Amp	3.17 (1.93, 5.19)		1.04 (0.56, 1.93)	0.33 (0.14, 0.78)	0.01
		Fat meat
		NO		YES
		OReg (95% CI)		OReg (95% CI)		P value*
	Del + Wt	1.00		1.59 (0.85, 2.96)
	Amp	1.90 (1.10, 3.27)		3.39 (1.88, 6.11)	1.12 (0.52, 3.42)	0.77
	Wt	1.00		1.55 (0.87, 2.75)
	Del + Amp	2.22 (1.30, 3.78)		3.95 (2.20, 7.10)	1.15 (0.53, 2.47)	0.72
		Egg (/week)
		≤3		>3
		OReg (95% CI)		OReg (95% CI)		P value*
	Del + Wt	1.00		2.83 (1.45, 5.51)
	Amp	3.01 (1.55, 5.86)		4.81 (2.45, 9.44)	0.56 (0.26, 1.24)	0.16
	Wt	1.00		2.59 (1.40, 4.79)
	Del +Amp	3.37 (1.76,6.49)		5.11 (2.69,9.72)	0.59 (0.27, 1.27)	0.18
		Sausage (times/month)
		≤1		>1
		OReg (95% CI)		OReg (95% CI)		P value*
	Del + Wt	1.00		3.27 (0.85, 12.61)
	Amp	2.92 (1.31, 4.02)		3.49 (1.67, 7.25)	0.47 (0.10, 2.10)	0.29
	Wt	1.00		2.76 (0.86, 8.85)
	Del +Amp	2.55 (1.53, 4.25)		3.88 (1.86, 8.07)	0.55 (0.13, 2.40)	0.41
		Leftovers^b (times/week)
		≤3	>3
		OReg (95% CI)	OReg (95% CI)			P value*
	Del + Wt	1.00	1.85 (1.03, 3.31)
	Amp	1.98 (1.20, 3.29)	3.77 (2.09, 6.82)		1.03 (0.50, 2.10)	0.94
	Wt	1.00	2.00 (1.14, 3.48)
	Del +Amp	2.51 (1.49, 4.24)	4.64 (2.25, 8.52)		0.93 (0.45, 1.89)	0.83
USP11		Fat meat
		NO	YES
		OReg (95% CI)	OReg (95% CI)			P value*
	Wt	1.00	1.69 (1.05, 2.73)
	Del +Amp	1.48 (0.78, 2.78)	2.46 (1.28, 4.71)		0.98 (0.45, 2.16)	0.97
	Wt + Amp	1.00	1.64 (1.10, 2.45)
	Del	3.28 (1.12, 9.56)	7.44 (2.28, 24.29)		1.38 (0.43, 4.48)	0.59
		Egg (/week)
		≤3	>3
		OReg (95% CI)	OReg (95% CI)			P value*
	Wt	1.00	1.89 (1.16, 3.10)
	Del + Amp	1.52 (0.80, 2.91)	2.97 (1.54, 5.75)		1.03 (0.49, 2.22)	0.93
	Wt + Amp	1.00	1.89 (1.28, 2.80)
	Del	3.56 (1.26, 10.02)	7.83 (2.50, 24.57)		1.16 (0.36, 3.75)	0.80
		Sausage (times/month)
		≤1		>1
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt + Amp	1.00		1.99 (1.03, 3.85)
	Del	3.34 (1.28, 8.72)		8.82 (1.70, 45.74)	1.33 (0.25, 7.05)	0.74
		Leftovers^b(times/week)
		≤3		>3
		OReg (95% CI)		OReg (95% CI)		P value*
	Wt	1.00		1.18 (0.73, 1.89)
	Del + Amp	0.89 (0.48, 1.65)		2.59 (1.39, 4.83)	2.48 (1.13, 5.41)	0.02
	Wt + Amp	1.00		1.47 (0.98, 2.21)
	Del	2.07 (0.68, 6.32)		6.91 (2.34, 20.35)	2.27 (0.67, 7.77)	0.19

CI, confidence interval. ^aAdjusted for occupation and family history of CRC. ^b leftovers: leftovers more than 12 hours. *P < 0.05 was considered statistically significant.

Table 4. The AUC of CRC risk-predicting models.

Model	AUC	SE	95% CI	*P value
EF	0.724	0.00730	0.711, 0.737	<0.001
EF + AMP	0.735	0.00717	0.722, 0.747	<0.001
EF + DEL	0.747	0.00703	0.734, 0.759	<0.001
EF + VAR	0.736	0.00719	0.723, 0.748	<0.001
EF + SIG	0.754	0.00695	0.741, 0.766	<0.001

AUC, the area under the receiver operating characteristic curve; SE, standard error; CI, confidence interval. *P < 0.05 was considered statistically significant.

Table 5. The relationships between gene CNVs and prognosis of CRC.

CNV genotypes	Patients (%)	5-year survival (%)	3-year survival (%)	OS (Mean (SD)) (month)	HR (95% CI)^a	P value*
CYLD^b
Wt	192 (62.3)	63	76	54.09 (1.98)	1.00
Del	41 (13.3)	80	80	53.79 (3.14)	0.85 (0.27, 2.67)	0.78
Amp	75 (24.4)	52	79	53.83 (2.60)	0.69 (0.29, 1.63)	0.39
Del v.s Wt +Amp					1.10 (0.35, 3.47)	0.87
Amp v.s. Del + Wt					1.44 (0.61, 3.41)	0.41
Wt v.s Del +Amp					0.73 (0.35, 1.53)	0.41
USP9X^b
Wt	102 (33.2)	64	64	50.37 (2.36)	1.00
Del	22 (7.2)	95	95	63.10 (2.84)	0.10 (0.01, 1.13)	0.11
Amp	183 (9.6)	52	69	53.56 (2.05)	0.56 (0.26, 1.23)	0.10
Del v.s Wt +Amp					6.07 (0.61, 60.34)	0.12
Amp v.s. Del + Wt					1.22 (0.58, 2.57)	0.61
Wt v.s Del +Amp					0.53 (0.24, 1.16)	0.11
USP11^b
Wt	160 (55.7)	88	92	53.28 (1.87)	1.00
Del	51 (17.8)	67	75	52.22 (3.54)	1.35 (0.51, 3.57)	0.54
Amp	76 (26.5)	60	67	53.02 (3.31)	1.14 (0.53, 2.44)	0.75
Del v.s Wt +Amp					1.29 (0.51, 3.28)	0.59
Amp v.s. Del + Wt					0.94 (0.45, 1.95)	0.87
Del + Amp v.s. Wt					0.83 (0.42, 1.64)	0.60

CI, confidence interval; HR, hazard ratio; OS, overall survival. ^aAdjusted for CEA and CA19-9 level before surgery, TNM stage, pathological type, and metastasis. ^bMissing value: CYLD, 2; USP9X, 3; USP11, 23. *P < 0.05 in the survival analysis was considered statistically significant.

Table 6. The relationships between gene CNVs and prognosis of colon cancer and rectal cancer.

	Colon cancer						Rectal cancer
	Patients (%)	5-year survival (%)	3-year survival (%)	OS (Mean (SD)) (month)	HR (95% CI)	P value*	Patients (%)	5-year survival (%)	3-year survival (%)	OS (Mean (SD)) (month)	HR (95% CI)	P value*
CYLD^a
Wt	61 (59.2)	72	80	53.67 (2.97)	1.00		125 (64.1)	60	75	52.29 (2.42)	1.00
Del	12 (11.7)	100	100		0.02 (0.00, 1116.04)	0.49	28 (14.4)	71	71	49.78 (4.26)	1.70 (0.46, 6.34)	0.42
Amp	30 (29.1)	69	78	51.53 (4.61)	1.16 (0.22, 6.11)	0.86	42 (21.5)	58	84	54.79 (3.22)	0.49 (0.13, 1.86)	0.29
Del v.s Wt+Amp					641.18 (0.00, 93629.36)	0.47					0.54 (0.14, 1.99)	0.26
Amp v.s. Del +Wt					0.68 (0.13, 3.48)	0.64					2.14 (0.57, 8.02)	0.69
Wt v.s Del+Amp					0.77 (0.17, 3.52)	0.77					0.83 (0.32, 2.13)	0.54
USP9X^a
Wt	34 (33.3)	73	73	51.35 (4.47)	1.00		66 (33.8)	60	60	47.63 (2.63)	1.00
Del	10 (8.8)	100	100		0.01(0.00, 4719.17)	0.51	13 (6.7)	92	92	60.92 (4.87)	0.39 (0.13, 8.18)	0.97
Amp	59 (57.8)	40	76	52.91 (3.12)	0.32(0.05, 2.27)	0.25	116 (59.5)	58	67	53.77 (2.46)	0.60 (0.25, 1.42)	0.25
Del v.s Wt+Amp					159.20 (0.00, 1.26E+15)	0.74					0.71 (0.09, 5.48)	0.74
Amp v.s. Del+Wt					1.80 (0.34, 9.44)	0.49					1.67 (0.72, 3.88)	0.23
Wt v.s Del+Amp					0.31 (0.04, 2.26)	0.25					0.62 (0.26, 1.45)	0.27
USP11^a
Wt	62 (62.6)	43	71	53.67 (2.71)	1.00		95 (52.8)	65	65	52.71 (2.42)	1.00
Del	16 (16.2)	92	92	61.33 (4.47)	0.00 (0.00, 3873.97)	0.30	33 (18.3)	55	69	47.14 (4.07)	3.37 (1.15, 9.87)	0.03
Amp	21 (21.2)	40	40	41.85 (7.30)	3.50 (0.26, 47.58)	0.35	52 (28.9)	64	73	55.51 (3.52)	1.63 (0.63, 4.18)	0.31
Del v.s Wt+Amp					41740.8 (0.00, 1.15E+19)	0.53					0.36 (0.14, 0.96)	0.04
Amp v.s. Del+Wt					0.61 (0.11, 3.43)	0.56					2.06 (0.90, 4.71)	0.69
Wt v.s Del+Amp					0.28 (0.04, 1.82)	0.18					0.84 (0.36, 1.98)	0.09

Note: In the analysis stratified by tumor location, there was a total of 300 cases included due to the 10 cases who missed detail information about tumor location. ^aMissing value, CYLD, 2 in colon cancer; USP9X, 2 in colon cancer; USP11, 6 in colon cancer and 15 in rectal cancer. *P < 0.025 in the stratified survival analysis by location was considered statistically significant.