Association between hypermethylation of DNA repetitive elements in white blood cell DNA and early-onset colorectal cancer

Figures & data

Table 1. Characteristics of CRC-affected cases and healthy controls

Characteristic	Controls		Cases		p-value
	(n = 242)		(n = 539)
Demographic data	Freq	%	Freq	%
Male	128	52.9%	293	54.4%	0.70
Age (years):
At diagnosis/interview*	50	[43,56]	50	[43,55]	0.37
At blood draw*	50	[43,56]	52	[45,57]	0.03
Caucasian	231	95.5%	512	95%	0.78
Smoking status:
Never	110	45.5%	249	46.2%	0.70
Former	87	36%	179	33.2%
Current	45	18.6%	111	20.6%
Drinking status:
Never	32	13.2%	107	19.9%	0.08
Former	21	8.7%	43	8%
Current	189	78.1%	387	72.1%
Body mass index (kg/m^2)*	25.1	[22.4,28.4]	26.5	[23.5,29.7]	< 0.001
Medicines/supplements
2 years ago, taking:
Folic acid	8	3.3%	12	2.3%	0.39
Aspirin	14	5.8%	25	4.6%	0.50
Ibuprofen	11	4.5%	21	3.9%	0.69
Have you ever taken:
Folic acid	39	16.2%	67	12.5%	0.17
Aspirin	30	12.4%	59	11%	0.56
Ibuprofen	50	20.7%	76	14.2%	0.02
Family history:
FDR^∆ with CRC	45	18.6%	130	24.1%	0.09
FDR/SDR^† with CRC	80	33.1%	249	46.2%	0.001
Repetitive DNA elements
Alu PMR*	70	[46,94]	75	[50,110]	0.02
LINE-1 PMR*	72	[48,107]	87	[59,130]	< 0.001
Sat2 PMR*	66	[49,96]	80	[57,111]	< 0.001

* Figures shown are median (Q1, Q3). ^∆ FDR, first degree relative; ^†SDR, second degree relative.

Table 2. PMR values of CRC-affected cases analyzed as quartiles* (Q1 lowest, Q4 highest) and as a continuous variable* (OR per unit increase of logged PMR value) for each repetitive DNA element

	PMR quartile	Q1		Q2		Q3		Q4		Test for trend:		PMR continuous
Sat2	P			0.31		0.01		0.02		0.002		0.001
	95% CI			(0.48, 1.26)		(1.14, 2.85)		(1.10, 2.71)				(1.22, 2.25)
	OR	Ref		0.78		1.81		1.72				1.66
LINE-1	P			0.34		0.02		< 0.001		< 0.001		< 0.001
	95% CI			(0.78, 2.06)		(1.07, 2.71)		(1.48, 3.70)				(1.25, 2.05)
	OR	Ref		1.27		1.71		2.34				1.6
Alu	P			0.59		0.84		0.01		0.005		0.04
	95% CI			(0.72, 1.79)		(0.66, 1.66)		(1.17, 2.86)				(1.01, 1.70)
	OR	Ref		1.13		1.05		1.83				1.31

* Models adjusted for age at blood draw (continuous), sex, body mass index (quartiles), smoking status, drinking status, ethnicity (Caucasian/not), and binary use of folate, aspirin and ibuprofen two years ago, and family history (FDR or SDR) of CRC.

Table 3. Combined effect of hypermethylation over multiple repetitive elements

Number of markers in highest quartile:	OR	95% CI		P
0	Ref
1	1.99	(1.35,	2.94)	< 0.001
2	1.63	(1.04,	2.56)	0.03
3	2.96	(1.56,	5.59)	0.001
LRT test*				0.06

* LRT, likelihood ratio test: compares the categorical PMR variable with the continuous. Model fully adjusted as per Table 2.

Table 4. Comparison of the association between PMR values (on the logarithm scale) and CRC risk between subgroups

Subgroup	Cases	Alu			LINE-1			Sat2
	n	OR	95% CI	P*	OR	95% CI	P*	OR	95% CI	P*
5FU**
No 5FU	173	2.01	(1.41, 2.84)	0.002	1.83	(1.33, 2.53)	0.33	1.91	(1.28, 2.85)	0.68
5FU	196	1.14	(0.81, 1.60)		1.56	(1.15, 2.11)		1.76	(1.20, 2.58)
BRAF
Negative	452	1.38	(1.05, 1.80)	0.41	1.64	(1.27, 2.12)	0.59	1.78	(1.29, 2.44)	0.09
Positive	31	1.08	(0.59, 1.97)		1.4	(0.79, 2.50)		0.98	(0.48, 2.00)
MMR^†
Proficient	451	1.41	(1.08, 1.85)	0.23	1.66	(1.29, 2.15)	0.3	1.73	(1.26, 2.38)	0.78
Deficient	39	1.01	(0.57, 1.78)		1.27	(0.75, 2.15)		1.59	(0.84, 3.00)
Nodes
No positive	224	1.68	(1.25, 2.28)	0.002	1.73	(1.30, 2.30)	0.44	1.87	(1.32, 2.66)	0.24
Positive	204	1.05	(0.76, 1.45)		1.54	(1.14, 2.08)		1.52	(1.05, 2.20)
T-stage
T1/T2	219	1.32	(0.97, 1.79)	0.96	1.47	(1.09, 1.97)	0.29	1.66	(1.16, 2.38)	0.99
T3/T4	320	1.31	(0.98, 1.73)		1.7	(1.29, 2.24)		1.66	(1.19, 2.31)
Age at diagnosis
< 40 y	76	1.37	(0.89, 2.10)	0.35	1.98	(1.30, 3.00)	0.31	1.66	(1.01, 2.73)	0.75
40- < 50 y	170	1.12	(0.80, 1.56)		1.42	(1.03, 1.95)		1.81	(1.23, 2.67)
50–59 y	293	1.41	(1.06, 1.88)		1.62	(1.23, 2.14)		1.58	(1.13, 2.20)
Site
Rectum	189	1.35	(0.97, 1.86)	0.08	1.58	(1.16, 2.15)	0.35	1.88	(1.29, 2.76)	0.81
Left colon	148	1.71	(1.20, 2.42)		1.9	(1.36, 2.66)		1.66	(1.10, 2.49)
Right colon	126	1.09	(0.76, 1.57)		1.46	(1.04, 2.05)		1.7	(1.11, 2.60)
Germline Mutation
None Found	492	1.37	(1.05, 1.78)	0.07	1.63	(1.26, 2.10)	0.69	1.66	(1.22, 2.26)	0.96
MMR^†	37	1.04	(0.58, 1.87)		1.46	(0.84, 2.53)		1.78	(0.94, 3.38)
MUTYH	10	0.46	(0.17, 1.26)		1.09	(0.41, 2.90)		1.51	(0.46, 4.97)
Family History
None	40	1.18	(0.68, 2.04)	0.83	1.53	(0.91, 2.59)	0.97	2.01	(1.07, 3.76)	0.57
FDR^∆ CRC	130	1.26	(0.88, 1.80)		1.64	(1.16, 2.32)		1.74	(1.14, 2.65)
FDR^∆ polypectomy	356	1.35	(1.02, 1.78)		1.6	(1.23, 2.08)		1.52	(1.10, 2.10)

* P, likelihood ratio test p-value, comparing the OR for the PMR marker across subgroups. **Excluding T-stages 1 and 4. ^†MMR, mismatch repair; ^∆ FDR, first degree relative.