Emerging blood-based biomarkers for detection of gastric cancer

Table 1 Proteomics-based biomarker models for detection of gastric cancer

Biomarkermodel	Approach used	Sample size and type(cancer/controls)	Diagnostic value1	Ref.
Five peaks - 3316, 6629, 3217, 3952, 6431 Da	MB-WCX, MALDI-TOF-MS	T: GC = 32/HC = 32	AUC = 0.86-0.99 for individual features (P < 0.001),	Yang et al[148], 2012
		V: GC = 30	Sn = 79.3%, Sp = 86.5%
		(GC I-II = 8)/HC = 30	Sn = 71.7% for early stage GC
1546 Da (SERPINA1)	MB-WCX, MALDI-TOF-MS; ELISA for validation	T: GC = 70/HC = 72	AUC (1546 Da) = 0.83 (P < 0.001), AUC (5335 Da) = 0.87 (P < 0.001) - calculated for training set; in validation set SERPINA1 concentration was significantly higher for GC patients than for all other controls (P < 0.001) and ENOSF1 concentration was significantly higher for GC patients than HC (P < 0.001)	Yang et al[34], 2015
5335 Da (ENOSF1)		V: GC = 36/HC = 36, BGD = 30, other cancers = 108
Fibrinogen α-chain, apolipo	HPLC, LC-MS/MS	T: GC = 65/HC = 30, BGD = 23	Sn = 90.9%, Sp = 90.6% (P = NA)	Liu W et al[37], 2012
		V: GC = 44/ HC = 30, BGD = 23
Six peaks at 2873, 3163, 4526, 5762, 6121 and 7778 m/z; For stage I three peaks at 2873, 6121 and 7778 m/z	Protein Chip SELDI-TOF-MS	GC = 169 (GCI = 27)/HC = 83	Sn = 93.5%, Sp = 91.6%	Li et al[26], 2012
			Accuracy for stage I - 88.9%, (P = NA)
EGFR, proApoA1, ApoA1, TTR, RANTES, VN, DD, IL-6, A2M, CRP, PAI1	xMAP (Luminex), ELISA	T: GC = 120/BGD = 101, HC = 19	AUC = 0.95, (P < 0.05)	Ahn et al[35], 2012
		V: GC = 95 (GC I-II = 75)/BGD = 43, HC = 8	Sn = 88.8%, Sp = 89.7%
			Sn (I-II ) = 92.3%
			Sn (tumor size ≤ 2 cm) = 81.8%
Four peaks at 1867 (tubulin beta chain), 2701 (thymosin beta4 like protein3), 2094 (cytochrom b-c1 subunit), 1467 Da	MB-WCX, MALDI-TOF-MS	T: GC = 40/HC = 39	AUC (1867 Da) = 1,	Fan et al[30], 2013
		V: GC = 40/GA = 30, HC = 39	AUC (1467 Da) = 0.83
			AUC (2701 Da) = 0.71
			AUC (2094 Da) = 0.70 (P < 0.05)
			Sn = 95.0%, Sp = 97.1%
50 decision trees, 28 masses	Protein Chip SELDI-TOF-MS,	T: GC = 41/HC = 49	Sn = 100%, Sp = 96.7%	Ebert et al[24], 2004
		V: GC = 28; GCI = 9/HC = 30	For stage I Sn = 89.9%
			(P = NA )
Three peaks at 3946, 3503 and 15958 Da			Sn = 92.8%, Sp = 86.7%
			For stage I Sn = 89.9%
			(P = NA )

¹Diagnostic values listed for validation set, if not otherwise stated. A2M: Alfa 2 macroglobulin; Apo: Apolipoprotein; AUC: Area under the curve; BGD: Benign gastric diseases; CRP: C reactive protein; DD: D-dimer; ENOSF1: Isoform 2 of mitochondrial enolase superfamily member 1; EGFR: Epidermal growth factor receptor; GA: Gastric adenoma; GC (I-IV): Gastric cancer (TNM stages); HC: Healthy control; HPLC: High performance liquid chromatography; IL-6: Interleukin 6; LC: Liquid chromatography; MALDI-TOF-MS: Matrix-assisted laser desorption/ionization-time of flight-mass spectrometry; MB-WCX: Magnetic bead based weak cation-exchange chromatography; NA: Not available; PAI1: Plasminogen activator inhibitor 1; proApo: Proapolopoprotein; RANTES: Regulated upon activation, normally T-expressed and presumably secreted; SELDI-TOF-MS: Surface-enhanced laser desorption/ionization-time of flight-mass spectrometry; SERPINA1: Serpin peptidase inhibitor clade A (alpha-1 antiproteinase, antitrypsin), member 1; Sn: Sensitivity; Sp: Specificity; T: Training set; TTR: Transthyretrin; V: Validation set; VN: Vitronectin.

Table 2 Autoantibody signatures with diagnostic value for gastric cancer

Biomarker signature description	Technology	Study design	Sample size (GC/controls)	Diagnostic value	Ref.
2 TAAs – p62, Koc	ELISA	GC vs HC	135/82	Sn = 19.3%, Sp = 97.6%, P < 0.01	Zhang et al[62], 2001
3 TSAs - IQGAP3, KRT23 and REG3A	PARSE assay	GC vs HC (age and sex matched)	48/46	Sn = 22.9%, Sp = 100%, P < 0.001	Xu et al[149], 2012
3 TAAs – p16, p53, c-myc	ELISA	GC vs HC	74/82	Sn = 21.6%, Sp = 97.6%; P < 0.001	Looi et al[150], 2006
7 TAAs - p53, C-myc, p16, IMP1, Koc, p62 and Survivin	ELISA	Cardia GC vs HC	88/140	AUC = 0.73, Sn = 64%, Sp = 87%, P < 0.001	Zhou et al[68], 2015
7 TAAs - C-myc, Cyclin B1, IMP1, Koc, P53, p62 and Survivin	ELISA, fixed cut-off	GC vs HC	91/346	Sn = 52.7%, Sp = 89.9%, P < 0.01	Zhang et al[63], 2003
	ELISA, individual cut-off (recursive partitioning)	GC vs HC	91/346	Sn = 98.9%, Sp = 93.1%, P < 0.001	Koziol et al[151], 2003
45 T7 phage-displayed TAA clones (including NY-ESO-1, DDX53, MAGE antigens etc.)	T7 phage displayed TAA microarray	GC vs HC (age and sex matched)	T:100/100	AUC = 0.79, Sn = 59%, Sp = 90%, P < 0.001	Zayakin et al[56], 2013
			V:235/213
		GC vs gastritis	235/100	AUC = 0.64, Sn = 58.7%, Sp = 55%, P < 0.001
		GC vs gastric ulcer	235/54	AUC = 0.76, Sn = 58.7%, Sp = 81.5%, P < 0.001

AUC: Area under the curve; GC: Gastric cancer; HC: Healthy controls; ND: Not determined; Sn: Sensitivity; Sp: Specificity; TAA: Tumor associated antigen; TSA: Tumor specific antigen; T: Training; V: Validation.

Table 3 Cell-free DNA as biomarkers for detection of gastric cancer

Candidate biomarkers	Sample size and type	Method/technology	Diagnostic value/outcome	Ref.
Total cell-free DNA level
β-actin (total cf DNA level)	GC = 53, HC = 21, plasma	qPCR	AUC = 0.75, P < 0.0001	Sai et al[90], 2007
DNA integrity		qPCR (ratio of long vs short b-actin amplicons)	No significant difference between GC and HC
Alu DNA sequences	GC = 54, HC = 59; plasma	Alu81-qPCR	AUC = 0.784, Sn = 75%, Sp = 63%	Park et al[91], 2012
Total cfDNA level	Early GC = 16; advanced GC = 14; HC = 34; plasma	Measurement of cfDNA concentration	AUC = 0.991, Sn = 96.67%, Sp = 94.11% for GC vs HC	Kim et al[92], 2014
Gene amplification
MYC gene copy number (MYC/GAPDH ratio)	GC = 57, HC = 39; tissues and plasma	qPCR	AUC = 0.816; strong positive correlation between MYC levels in GC tissues and plasma (r = 0.342; P = 0.009)	Park et al[99], 2009
HER2 gene copy number (HER2/RPPH1 ratio)	Discovery: GC = 52 (pre and post-operative treatment), HC = 40; plasma and tissues	qPCR	AUC = 0.746, Sn = 53.9%, Sp = 96.7%; Positive correlation between GC tissues and plasma (r = 0.424; P = 0.00721); decrease in post-treatment plasma in HER2 + GC cases	Shoda et al[100], 2014
	Validation: GC = 25 plasma		Sn = 66.7%, Sp = 100%
DNA methylation markers
RPRM (Reprimo)	GC = 43, HC = 31; GC tissues and plasma	MSP	95.3% GC, 9.7% HC, P < 0.00001; Strong correlation between methyl status in tissues and plasma	Bernal et al[107], 2008
RUNX3	GC (preoperative) = 65, GC (postoperative) = 43, HC = 50, tissues and serum	qMSP	AUC = 0.8651, Sn = 95.5%, Sp = 62.5%; decrease after surgical resection	Sakakura et al[152], 2009
KCNA4 + CYP26B1	GC = 46, GPL = 46, HC = 30; serum	Discovery: Methylation microarray in tissues; Testing: MSP	AUC = 0.917, Sn = 91.3%, Sp = 92.1%	Zheng et al[109], 2011
SLC19A3	Discovery: GC = 45, HC = 60; plasma	MSRED-qPCR	Increased in GC, P < 0.0001	Ng et al[153], 2011
	Validation: GC = 20, HC = 20		AUC = 0.82, Sn = 85%, Sp = 85%
FAM5C + MYLK	GC = 58, GPL = 46, HC = 30; serum	Discovery: MeDIP in cell lines; Testing: MSP	AUC = 0.838, Sn = 77.6%, Sp = 90% for GC vs HC; Sn = 30.4% for GPL vs HC; decrease after surgical resection	Chen et al[154], 2012
XAF1	GC = 202, HC = 88, tumor tissues and serum	qMSP	AUC = 0.909, P < 0.0001; 83.9% concordance between GC tissues and serum	Ling et al[108], 2013

AUC: Area under the curve; GC: Gastric cancer; GPL: Gastric precancerous lesions; HC: Healthy controls; MeDIP: Methylated DNA immunoprecipitation; MSP: Methylation-specific PCR; MSRED-qPCR: Methylation-sensitive restriction enzyme digestion and real-time quantitative PCR; Sn: Sensitivity; Sp: Specificity.

Table 4 Cell-free RNAs as biomarkers for detection of gastric cancer

Candidate biomarkers	Sample size and type	Method/technology	Diagnostic value/outcome	Ref.
Circulating cell-free miRNAs
miR-106a/let-7a ratio	GC = 69, HC = 30; plasma	qRT-PCR	AUC = 0.879, Sn = 85.5%, Sp = 80%	Tsujiura et al[117], 2010
5-miRNA signature: miR-1, miR-20a, miR-27a, miR-34, miR-423-5p	Discovery: GC = 20, HC = 20; Validation: GC = 142, HC = 105; Serum	Discovery: Solexa sequencing; Testing: qRT-PCR	AUC = 0.831 (validation set)	Liu et al[155], 2011
miR-451	Discovery: pre- and post-operative plasma, GC = 3; Validation: GC = 56, HC = 30	Discovery: microarray	AUC = 0.96, Sn = 96%, Sp = 100%; decreased in 90% of post-operation plasma samples	Konishi et al[119], 2012
miR-486		Testing: qRT-PCR	AUC = 0.92, Sn = 86%, Sp = 97%; decreased in 93% of post-operation plasma samples
miR-378	Discovery: GC = 7, CRC = 7, HC = 10;	Discovery: microarray	AUC = 0.861, Sn = 87.5%, Sp = 70.73%;	Liu et al[122], 2012
	Validation: GC = 40, HC = 41; serum	Testing: qRT-PCR	No significant differences across stages I-IV
miR-223	Test set: GC = 10, HC = 10; Validation: GC = 60, HC = 60; plasma	qRT-PCR	AUC = 0.9089	Li et al[121], 2012
miR-21			AUC = 0.7944
miR-218			AUC = 0.7432
3 miRNA combined			AUC = 0.9531, Sn = 84.29%, Sp = 92.86%
			No significant differences across stages I-IV
3-miRNA signature: miR-221, miR-744, and miR-376c	Discovery: GC = 14, HC = 14;	Discovery: TaqMan array, validation: qRT-PCR	Sn = 82.4%, Sp = 58.8% (for GC vs HC)	Song et al[124], 2012
	Validation I: GC = 68, HC = 68		Sn = 73.3% (for early GC)
	Validation II: DYS = 46, HC = 46		miR-221 elevated in DYS, no difference from HC for miR-376c and miR-744;
	Pre-diagnosis serum samples, GC = 58		Increase during GC development;
			Sn = 79.3% (for GC 2-5 years before diagnosis)
miR-106b	Discovery: GC = 30, HC = 30	qRT-PCR	AUC = 0.773 (all in validation set)	Cai et al[156], 2013
miR-20a	Validation: GC = 60, HC = 60;		AUC = 0.859
miR-221	plasma		AUC = 0.796
miR-223	GC = 50, HC = 47; serum	qRT-PCR	AUC = 0.85, Sn = 81%, Sp = 78%;	Wang et al[157], 2014
			Increased in advanced stages
miR-16			AUC = 0.90, Sn = 79%, Sp = 78%
miR-100			AUC = 0.71, Sn = 71%, Sp = 58%
			Increased in advanced stages
miR-16	Discovery: stage I non-cardia GC = 40, HC = HC; Validation: stage I non-cardia GC = 48, HC = 102	Discovery: TaqMan array, validation: qRT-PCR	AUC = 0.768 (all in validation set)	Zhu et al[120], 2014
miR-25			AUC = 0.694
miR-92a			AUC = 0.732
miR-451			AUC = 0.790
miR-486-5p			AUC = 0.779
5 miRNA combined			AUC = 0.812, Sn = 72.9%, Sp = 89.2%; In vitro evidence that miR-16, miR-25 and miR92a but not miR-451 and miR486-5p are secreted from cancer cells
miR-222	GC = 114, HC = 56; plasma	qRT-PCR	AUC = 0.850, Sn = 66.1%, Sp = 88.3%	Fu et al[158], 2014
miR-18a	GC = 82, HC = 65, plasma	qRT-PCR	AUC = 0.907, Sn = 80.5%, Sp = 84.6%; no association with stage	Su et al[123], 2014
miR-18a	GC = 104, HC = 65, plasma and GC tissues	qRT-PCR	AUC = 0.8059, Sn = 84.6%, Sp = 69.2%	Tsujiura et al[159], 2015
			Overexpressed in GC; in vitro evidence that miR-18a is released by cancer cells; decreased in postoperative plasma
Circulating cell-free mRNAs and long non-coding RNAs
hTERT mRNA	GC = 118, CAG = 40, HC = 58; plasma	qRT-PCR	AUC = 0.891, Sn = 66%, Sp = 87%; strong positive correlation with advanced stage of GC	Kang et al[125], 2013
MACC1 mRNA	GC = 76, HC = 54, plasma	qRT-PCR	Sn = 68%, Sp = 89%	Burock et al[160], 2015
LINC00152	Pre- and post-operative plasma GC = 79, GED = 31, HC = 81	qRT-PCR	AUC = 0.657, Sn = 48.1%, Sp = 85.2%	Li et al[131], 2015

AUC: Area under the curve; DYS: Intestinal dysplasia; GC: Gastric cancer; HC: Healthy controls; Sn: Sensitivity; Sp: Specificity.