p16 promoter hypermethylation: A useful serum marker for early detection of gastric cancer

Table 1 Primer sets used in MSP

Primer sets	Sense primer: 5’-3’	Antisense primer: 5’-3’	Size (bp)
p16-W	CAGAGGGTGGGGCGGACCGC	CGGGCCGCGGCCGTGG	140
p16-M	TTATTAGAGGGTGGGGCGGATCGC	GACCCCGAACCGCGACCGTAA	150
p16-U	TTATTAGAGGGTGGGGTGGATTGT	CAACCCCAAACCACAACCATAA	151

W: Unmodified or wild-type primers; M: Methylated-specific primers; U: Unmethylated-specific primers.

Table 2 Results of methylation in tumoral tissue and corresponding serum

	Tissue (+)	Tissue (-)
Serum (+)	14	0
Serum (-)	9	29

(+): Methylated; (–): Unmethylated.

Table 3 Clinicopathological features of p16 promoter hypermethylation

Variable	n (%)	Methylated	Unmethylated	P value
Total		23	29
Gender
Male	38 (73.1)	17	21	0.904
Female	14 (26.9)	6	8
Age (yr)
< 64	26 (50.0)	8	18	0.051
> 64	26 (50.0)	15	11
Pathological grade
1	20 (39.2)	5	15	< 0.05
2	11 (21.6)	7	4
3	20 (39.2)	10	10	(1 vs 2&3)
Pathological type
Intestinal	27 (62.7)	13	14	0.754 (Intestinal vs Diffuse)
Diffuse	14 (27.5)	5	9	0.161 (Intestinal vs Mix)
Mixed	5 (9.8)	4	1	0.140 (Diffuse vs Mix)
Anatomical site
Cardia	22 (44.0)	7	15	0.124
Body	15 (30.0)	9	6
Pylorus	13 (26.0)	6	7	(Cardia vs Noncardia)
Distant metastasis
Absent	39 (75.0)	20	19	0.259
Present	13 (25.0)	9	4
Smoking
Yes	9 (17.6)	4	5	1.000
No	42 (82.4)	19	23
H pylori infection
Positive	30 (60.0)	11	19	0.201
Negative	20 (40.0)	11	9

Table 4 Results of methylation analysis in p16-positive and negative cases

Immunohistochemistry	Methylated	Unmethylated
p16-positive	3	17
p16-negative	20	12