Promoter hypermethylation of CDH1, FHIT, MTAP and PLAGL1 in gastric adenocarcinoma in individuals from Northern Brazil

Table 1 Primer sequences (5'-3') for MSP

Gene	Sense	Antisense	Product size	TM
CDH1	M-ATTCGAATTTAGTGGAATTAGAATC	M-CCCAAAACGAAACTAACGAC	125 bp	53°C
	U-GGATTTGAATTTAGTGGAATTAGAATT	U-CTCCCCAAAACAAAACTAACAAC	130 bp
FHIT	M-TTTTCGTTTTTGTTTTTAGATAAGC	M-AAAAATATACCCACTAAATAACCGC	157 bp	52°C
	U-TGGTTTTTGTTTTTGTTTTTAGATAAGT	U-AAAATATACCCACTAAATAACCACC	159 bp
MTAP	M-TGTTTTTTAGGAATTAAGGGAAATAC	M-AACTACAAAATCTAACCCGACGAC	199 bp	52°C
	U-TTTTTAGGAATTAAGGGAAATATGT	U-CAACTACAAAATCTAACCCAACAAC	197 bp
PLAGL1	M-GTTTATTTTGGCGGAGATTTC	M-ACTAAACGACACCCACACGTC	147 bp	51°C
	U-GGTTTATTTTGGTGGAGATTTTG	U-AAAAACTAAACAACACCCACACAT	152 bp

M: methylated sequences; U: unmethylated sequences; TM: melting temperature.

Table 2 Clinico-pathological characteristics of the samples

Variable (n)	Samples			P
	Nonneoplastic n (%)	Intestinal n (%)	Diffuse n (%)
Gender
Male (54)	9 (16.67)	20 (37.03)	25 (46.3)	0.9176
Female (24)	4 (16.66)	10 (41.67)	10 (41.67)
Age (yr)
≤ 50 (23)	7 (30.43)	7 (30.43)	9 (39.14)	0.1056
> 50 (55)	6 (10.91)	23 (41.82)	26 (47.27)
Smoking status
Smoker (27)	1 (3.7)	13 (48.15)	13 (48.15)	0.0717
No smoker (51)	12 (23.53)	17 (33.33)	22 (43.14)
H pylori
Present (36)	6 (16.67)	8 (22.22)	22 (61.11)	0.0142a
Absent (42)	7 (16.67)	22 (52.38)	13 (30.95)
Location
Cardia (25)	-	11 (44)	14 (56)	0.783
Noncardia (40)	-	19 (47.5)	21 (52.5)
T stage
T1, T2 (15)	-	7 (46.67)	8 (53.33)	0.9638
T3, T4 (50)	-	23 (46)	27 (54)
Lymph node metastasis
Present (51)	-	23 (45.1)	28 (54.9)	0.7445
Absent (14)	-	7 (50)	7 (50)
Distant metastasis
Present (6)	-	4 (66.67)	2 (33.33)	0.4649
Absent (55)	-	23 (41.82)	32 (58.18)
Unknown (4)	-	3 (75%)	1 (25%)

^aP < 0.05.

Table 3 Methylation number and frequency in gastric tissue samples, n (%)

Samples	Methylation status
	CDH1		FHIT		MTAP			PLAGL1
	M/U	U	M/U	U	M/U	M	P	M/U	M
Normal tissue (13)	12 (92.31)	1 (7.69)	8 (61.54)	5 (38.46)	11 (84.62)	2 (15.38)		12 (92.31)	1 (7.69)
GC (65)	65 (100)	0 (0)	34 (52.31)	31 (47.69)	49 (75.38)	16 (24.62)		43 (66.15)	22(33.85)
Diffuse-type GC (35)	35 (100)	0 (0)	17 (48.57)	18 (56.25)	22 (62.86)	13 (37.14)	0.0249a	23 (65.71)	12(34.29)
Intestinal-type GC (30)	30 (100)	0 (0)	17 (56.67)	13 (43.33)	27 (90)	3 (10)		20 (66.6)	10(33.33)

U: unmethylated; M/U: methylated and unmethylated; M: methylated.

^aP < 0.05 vs Intestinal-type GC.

Table 4 Number of hypermethylated genes in gastric tissue samples

Samples (n)	Number of hypermethylated genes		P value
	1 or 2	3 or 4
Nonneplastic gastric tissue (13)	12 (92.31%)	1 (7.69%)
Gastric cancer (65)	44 (67.69%)	21 (32.31%)	0.0396b
Diffuse-type GC (35)	21 (60%)	14 (40%)
Intestinal-type GC (30)	23 (76.67%)	7 (23.33%)

N: number of cases;

^bP < 0.05.