hENT1 expression is predictive of gemcitabine outcome in pancreatic cancer: A systematic review

Table 1 Characteristics of the identified studies

Ref.	Year of publication	Country	Inclusion period	No. of patients	Study design	Follow-up median (95%CI), mo
Spratlin et al[9]	2004	Canada	1998–2002	21	RS	NR
Giovannetti et al[10]	2006	Italy	2001–2004	1021	RS	11.2 (0.4–32.1)
Farrell et al[12]	2009	United States	1998–2002	91	Post hoc2	NR
Maréchal et al[3]	2009	Belgium	2000–2003	45	RS	21.9 (3.3–107.4)
Fujita et al[8]	2010	Japan	1992–2007	70	RS	15.7 (0.5–114)
Maréchal et al[1]	2012	Belgium	1996–2009	234	RS	55.7 (46.4–61.2)
Kawada et al[2]	2012	Japan	2002–2007	63	RS	31
Morinaga et al[21]	2012	Japan	2006–2008	27	RS	NR
Murata et al[22]	2012	Japan	2005–2010	93	RS	15 (3.5–57.2)
Nakagawa et al[23]	2012	Japan	2002–2011	109	RS	39.7 (2–122)
				Total: 855

¹n = 81 with complete hENT1;

²Post hoc analysis of randomized controlled trial. NR: Not reported; RS: Retrospective.

Table 2 Characteristics of the identified studies

Ref.	Age median, yr (range)	Sex m/tot n (%)	hENT1 method	Chemotherapy	Radiation dose Gy/(Gy/frac)	Outcome measurement	Quality REMARK
Spratlin et al[9]	58 (51-64)1	11 (52)	IHC	Pall Gem	No	OS	10
Giovannetti et al[10]	65 (22-83)	53 (50)	RT-PCR	Pall Gem Adj Gem	45	OS, DFS, TTP, RR	12
Farrell et al[12]	53/63/652	45 (49)	IHC	Adj Gem	50.4	OS, DFS, Tox	17
Maréchal et al[3]	56 (34-83)	23 (51)	IHC	Adj Gem	40-50.4	OS, DFS, Tox	13
Fujita et al[8]	65 (36-86)	42 (60)	RT-PCR	Adj Gem or Resection only	No	OS, DFS	12
Maréchal et al[1]	NR	129 (53)	IHC	Adj Gem	50.4	OS	15
Kawada et al[2]	- (41-81)	33 (52)	IHC	Neo Gem Adj 5-FU	50/2	DSS	9
Morinaga et al[21]	64 (45-74)	17 (63)	IHC	Adj Gem	No	OS, DFS	12
Murata et al[22]	68 (44-87)	38 (69)	IHC	Neo Gem Adj Gem	45/2	OS, DFS, RR	13
Nakagawa et al[23]	67 (41-83)	52 (48)	IHC	Adj Gem + S1	No	OS, DFS	13

¹95%CI;

²Medians in the different hENT1 expression groups. IHC: Immunohistochemistry; RT-PCR: Reverse transcription polymerase chain reaction; Adj: Adjuvant; Neo: Neoadjuvant; Radio: Radiotherapy; Gem: Gemcitabine; Pall: Palliative; OS: Overall survival; DFS: Disease-free survival; Tox: Toxicity; TTP: Time to progression; DSS: Disease-specific survival; RR: Response rate.

Table 3 hENT1 expression levels, cut-offs and grouping

Ref.	Method	Grading	Reference cells	Groups (n)
Spratlin et al[9]	IHC	0-2 based on relative intensities of staining.	Langerhans cells, lymphocytes.	Dichotomized:
		0 = absence of staining		Low = 0 (12)
		1 = intermediate staining		High = 1 and 2 (9)
		2 = most intense staining
Giovannetti et al[10]	RT-PCR	Gene-expression ratio with GAPDH, expressed as tertiles		Gene expression tertiles:
		GAPDH/target gene ratio		Low < 1.06 (27)
				Intermediate 1.06-1.38 (28)
				High ≥ 1.38 (26)
				Dichotomized:
				By medians
				Low < 1.23 (44)
				High ≥ 1.23 (37)
Farrell et al[12]	IHC	Based on relative intensities.	Lymphocytes	Dichotomized:
		High = strong reactivity in > 50% of neoplastic cells.		No (18)1
		No = no staining in > 50%		vs
		Low = all cases between High and No.		Low/high (73)1
Maréchal et al[3]	IHC	0-3 based on staining intensities	Langerhans cells	Dichotomized:
		0 = no staining	Lymphocytes	Low < 80 (26)
		1 = weakly positive		(final score)
		2 = moderately positive
		3 = strongly positive		High = ≥ 80 (19)
		Final score calculated: multiplying intensity score and the percentage of the specimen. Weighted score 0-300
Fujita et al[8]	RT-PCR	Level of mRNA calculated from standard curve constructed with total RNA from Capan-1, a human pancreatic cancer cell line		mRNA split into high/low groups using recursive descent partitioning. Cut-off 0.5
				Low (26)1
				High (14)
Maréchal et al[1]	IHC	0-2 based on staining intensities	Lymphocytes	Dichotomized:
		Quantified as Farrell		Low/moderate (136)1
				High (86)1
Kawada et al[2]	IHC	0-2 based on staining intensities.	Langerhans cells	Negative = 0-1 (41)
		1 = same intensity as control.		Positive = 2 (22)
Morinaga et al[21]	IHC	Staining intensity and percentage of positive tumor cells scored and given a hENT1-score by calculating the two		Low = hENT1 score 0-3 (11)
		Staining 0-3 where		High = hENT1 score 4-6 (16)
		0 = no
		1= weakly pos
		2 = moderately pos
		3 = strongly pos
		Percentage:
		0 = no positive
		1 ≤ 50% positive cells
		2 = 50%-80% positive cells
		3 = ≥ 80%
Murata et al[22]	IHC	Staining intensity + extent of positive staining	Langerhans cells	Dichotomized:
		Intensity:		Negative = low and intermediate (16)
		0 = no staining
		1 = weakly positive		Positive = high (39)
		2 = moderately positive
		3 = strongly positive
		Extent staining:
		High = score 3 > 50% cells
		Low = score 0 or 1 > 50%
		Intermediate = all others
Nakagawa et al[23]	IHC	Staining intensities:	Langerhans cells	Low = grade 0 or 1 in > 50% (31)
		0 = not stained		High = grade 2 or 3 in > 50% of cells (78)
		1 = faintly stained
		2 = weakly stained
		3 = as strongly as islet cells

¹In gem arm. GAPDH: Glyceraldehyde 3-phosphate dehydrogenase.

Table 4 Results

Ref.	Median survival all patients (95%CI)	OS		DFS		Main conclusions
		Univariate analysis median (95%CI) or HR (95%CI) P-value	Multivariate analysis HR (95%CI)	Univariate analysis median (95%CI) or HR (95%CI) P-value	Multivariate analysis HR (95%CI)
Spratlin et al[9]	11.01 (6.8-17.5)	(mo):		NR		Pat with detectable hENT1 had sig longer OS compared with pat with low hENT1
		High = 13 (4.2-20.4)
	5.01 (2.8-12.2)	Low = 4 (1.5-6.9)
		P = 0.01
Giovannetti et al[10]	13.3 (10.9-15.7)	(mo):	Low = 5.34 (2.28-12.50)	Palliative (mo):		hENT1 expression was significantly correlated with outcome - pat with high hENT1 had longer OS
		Low = 8.48 (7.01-9.95)		Low = 5.85 (2.75-8.95)
		Inter = 15.74 (13.84-17.63)	Inter = 1.07 (0.46-2.49)	Inter = 10.09 (9.63-10.54)
		High = 25.69 (17.64-33.74)	High = 1	High = 12.68 (2.89-22.47)
		P ≤ 0.001	P < 0.0001	P = 0.02
			2 groups:
		2 groups:	HR = 4.21	Adjuvant (mo):
		Low = 12.42 (8.18-16.66)	P ≤ 0.001	Low = 9.26 (3.86-14.67)
		High = 22.34 (16.34-28.34)		Inter = 12.91 (9.31-16.51)
		P ≤ 0.001		High = 20.43 (13.27-27.60)
				P ≤ 0.01
Farrell et al[12]	NR	(HR):	Low/high = 0.40 (0.22-0.75)	(HR):	Low/high = 0.39 (0.21-0.73)	hENT1 expression was ass with longer OS, DFS in pat receiving gem. hENT1 is a relevant predictive marker for gem outcome
		Low/High = 0.51 (0.29-0.91)		Low/High = 0.57 (0.32-1.001)
		No = 1	No = 1	No = 1	No = 1
		P = 0.02	P = 0.03	P = 0.05	P = 0.003
Maréchal et al[3]	21.9 (3.3-107.4)	(HR):	High = 1	(HR):	High = 1	Pat with high hENT1 had sig longer OS and DFS compared to low hENT1
		High = 1	Low = 3.42 (1.44-8.81)	High = 1	Low = 3.17 (1.43-6.73)
		Low = 3.88 (1.78-8.92)	P = 0.0005	Low = 3.55 (1.65-7.63)	P = 0.0004
		P = 0.0007		P = 0.02
Fujita et al[8]	NR	(mo):	(RR):	(mo):	NR	Low hENT1 ass with shorter OS in gem-group
		High = 45	Low = 2.980 (0.964-10.86)	High = 25
		Low = 16.5		Low = 8
		P = 0.011	P = 0.2 (not sig)	P = 0.11 (not sig)
Maréchal et al[1]	32.0 (26.4-34.3)	(HR):	n = 2222	NR	NR	High hENT1 predicts longer OS in pat treated with adj gem. Absence of gem - hENT1 lacks prognostic value
		High = 0.43 (0.29-0.63)	High = 0.34 (0.22-0.53)
	(GEM-group)	Low/Mod = 1	Low/Mod = 1
		P < 0.0001	P < 0.0001
Kawada et al[2]	NR	Positive vs negative	Positive/negative	NR	NR	DSS tended to be better in the hENT1-neg group but not statistically sig
		P = 0.352	P = 0.503
Morinaga et al[21]	NR	(mo):	Low = 1	(mo):	Low = 1	High hENT1 sig ass with longer OS in pat receiving adj gem after resection
		Low = 11.8 (6.9-16.6)	High = 0.327 (0.128-0.835)	Low = 7.3 (3.6-11.1)	High = 0.558 (0.214-1.452)
		High = 22.2 (11.5-32.9)		High = 9.3 (4.2-14.5)
		P = 0.024	P = 0.019	P = 0.022	P = 0.232
		(HR):		(HR):
		Low = 1		Low = 1
		High = 0.366 (0.148-0.906)		High = 0.362 (0.146-0.898)
		P = 0.030		P = 0.028
Murata et al[22]	24.3	(HR):	Positive = 1	(HR):	Positive = 1	Sig longer OS, RFS in pat with pos hENT1
		Positive = 1	Negative = 3.15 (1.35-7.37)	Positive = 1	Negative = 1.76 (0.85-3.66)
		Negative = 3.04 (1.45-6.37)		Negative = 2.34 (1.22-4-47)
		P = 0.0037	P = 0.008	P = 0.011	P = 0.129
Nakagawa et al[23]	OS: 34.9	(5y-SR %):	High = 1	(5y-SR %):	High = 1	hEN1 expression is predictive of the efficacy of adj gem-based chemotherapy after resection
	DFS: 17.8	High = 38	Low = 3.16 (1.65-6.06)	High = 30	Low = 2.70 (1.52-4.83)
		Low = 13		Low = 17
		P = 0.001	P = 0.001	P = 0.004	P = 0.001

¹From diagnosis/from treatment;

²n = 222 in multivariate analysis. Ass: Associated; pat: Patient; sig: Significant; adj: pos: Positive; op: Operation; HR: Hazard ratio; SR: Survival rate; RFS: Recurrence-free survival.