Application of biomarkers in sentinel lymph node biopsy for the management of breast cancer

doi:10.3877/cma. j. issn.1674-0807.2012.06.003

Objective

Sentinel lymph node in breast cancer is a predictor of the tumor’s metastatic potential. Cytokines emerging from antitumor immune response might also target sentinel lymph node (SLN). In this study, we evaluated the cytokine profile including interleukin-8 (IL-8), interleukin-12 (IL-12), interferon-gamma (IFN-γ), and tumor necrosis factor alpha (TNF-α) for the T-cell response.

Methods

From January 2008 to August 2010,35 patients with breast mass underwent SLN biopsy in UNIMED Medical Institute. The specimens were formalin-fixed and paraffin-embedded for hematoxylin and eosin staining and immunohistochemistry with following markers: IL-8, IL-12, IFN-γ and TNF-α. The staining intensity was scored according to four grades: negative (-), weak expression (+), moderately positive expression (++), strong positive expression (+++). The correlation between the expression of cytokines and pathological characteristics were also analyzed using the Spearman Rank Correlation test.

Results

Out of 35 patients, SLN metastases were observed in 20 patients and among them, 75% (15/20) were strongly stained (+++) with IL-8 and moderately stained (++) with IL-12, but only 25% (5/20) were strongly stained (+++) with IFN-γ and TNF-α. The expression of IL-8 was significantly associated with all tumor grades (r=0.639, P=0.00), estrogen receptor (r=0.778, P=0.01), progesterone receptor (r=0.759 P=0.00) and HER-2/neu receptor (r=0.863 P=0.00).

Conclusion

The strong association between IL-8 expression and SLN metastases might highlight the prognostic significance of IL-8 in SLN.

关键词: sentinel lymph node, immunochemical staining, cytokines, biomarkers

Table 1 Clinicopathological characteristics of 35 patients

Characteristics	Total number of patients [n(%)]
Menopausal status
Premenopausal	10 (29)
Postmenopausal	25 (71)
SLN status
Negative	15 (43)
Positive	20 (57)
Tumor size
≤2cm (T₁)	18 (51)
2-5cm (T₂)	15 (43)
＞5cm (T₃)	2 (6)
Histology
Ductal	33 (94)
Lobular	2 (6)
Grading
G1	6 (17)
G2	17 (49)
G3	12 (34)
ER status
Negative	17 (49)
Positive	18 (51)
PR status
Negative	17 (49)
Positive	18 (51)
HER-2/neu status
Negative	17 (49)
Positive	18 (51)

Table 1 Clinicopathological characteristics of 35 patients

Characteristics	Total number of patients [n(%)]
Menopausal status
Premenopausal	10 (29)
Postmenopausal	25 (71)
SLN status
Negative	15 (43)
Positive	20 (57)
Tumor size
≤2cm (T₁)	18 (51)
2-5cm (T₂)	15 (43)
＞5cm (T₃)	2 (6)
Histology
Ductal	33 (94)
Lobular	2 (6)
Grading
G1	6 (17)
G2	17 (49)
G3	12 (34)
ER status
Negative	17 (49)
Positive	18 (51)
PR status
Negative	17 (49)
Positive	18 (51)
HER-2/neu status
Negative	17 (49)
Positive	18 (51)

Table 2 Expression of IL-8, IL-12, IFN-γ, and TNF-α in all 35 patients’ SLN

Cytokines	Staining intensity of metastatic lymph node (n=20)				Staining intensity of non-metastatic lymph node (n=15)
Cytokines	-	+	++	+++	-	+	++	+++
IL-8	0	0	5(25%)	15(75%)	15(100%)	0	0	0
IL-12	3(15%)	1(5%)	5(25%)	11(55%)	7(46.7%)	2(13.3%)	4(26.7%)	2(13.3%)
IFN-γ	6(30%)	8(40%)	1(5%)	5(25%)	11(73.3%)	4(26.7%)	0	0
TNF-α	6(30%)	5(25%)	4(20%)	5(25%)	12(80.0%)	3(20.0%)	0	0

Table 2 Expression of IL-8, IL-12, IFN-γ, and TNF-α in all 35 patients’ SLN

Cytokines	Staining intensity of metastatic lymph node (n=20)				Staining intensity of non-metastatic lymph node (n=15)
Cytokines	-	+	++	+++	-	+	++	+++
IL-8	0	0	5(25%)	15(75%)	15(100%)	0	0	0
IL-12	3(15%)	1(5%)	5(25%)	11(55%)	7(46.7%)	2(13.3%)	4(26.7%)	2(13.3%)
IFN-γ	6(30%)	8(40%)	1(5%)	5(25%)	11(73.3%)	4(26.7%)	0	0
TNF-α	6(30%)	5(25%)	4(20%)	5(25%)	12(80.0%)	3(20.0%)	0	0

Table 3 Correlation between the expressions of IL-8 in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	IL-8(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.639	0.00
G1	6	0	0	0
G2	7	1	1	8
G3	0	1	4	7
ER					0.778	0.00
Positive	1	1	5	13
Negative	12	1	0	2
PR					0.759	0.00
Positive	1	1	5	13
Negative	12	1	0	2
HER-2/neu					0.863	0.00
Positive	0	1	2	15
Negative	13	1	3	0

Table 3 Correlation between the expressions of IL-8 in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	IL-8(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.639	0.00
G1	6	0	0	0
G2	7	1	1	8
G3	0	1	4	7
ER					0.778	0.00
Positive	1	1	5	13
Negative	12	1	0	2
PR					0.759	0.00
Positive	1	1	5	13
Negative	12	1	0	2
HER-2/neu					0.863	0.00
Positive	0	1	2	15
Negative	13	1	3	0

Table 4 Correlation between the expressions of IL-12 in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	IL-12(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.187	0.28
G1	1	3	0	2
G2	6	1	4	6
G3	3	1	1	7
ER					0.441	0.01
Positive	4	0	5	11
Negative	6	5	0	4
PR					0.399	0.02
Positive	4	0	5	11
Negative	6	5	0	4
HER-2/neu					0.415	0.01
Positive	3	1	5	9
Negative	7	4	0	6

Table 4 Correlation between the expressions of IL-12 in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	IL-12(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.187	0.28
G1	1	3	0	2
G2	6	1	4	6
G3	3	1	1	7
ER					0.441	0.01
Positive	4	0	5	11
Negative	6	5	0	4
PR					0.399	0.02
Positive	4	0	5	11
Negative	6	5	0	4
HER-2/neu					0.415	0.01
Positive	3	1	5	9
Negative	7	4	0	6

Table 5 Correlation between the expressions of IFN-γ in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	IFN-γ(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.155	0.37
G1	2	4	0	0
G2	10	4	0	3
G3	4	5	1	2
ER					0.367	0.03
Positive	8	6	1	5
Negative	8	7	0	0
PR					0.283	0.10
Positive	8	6	1	5
Negative	8	7	0	0
HER-2/neu					0.326	0.06
Positive	6	7	1	1
Negative	10	6	0	4

Table 5 Correlation between the expressions of IFN-γ in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	IFN-γ(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.155	0.37
G1	2	4	0	0
G2	10	4	0	3
G3	4	5	1	2
ER					0.367	0.03
Positive	8	6	1	5
Negative	8	7	0	0
PR					0.283	0.10
Positive	8	6	1	5
Negative	8	7	0	0
HER-2/neu					0.326	0.06
Positive	6	7	1	1
Negative	10	6	0	4

Table 6 Correlation between the expressions of TNF-α in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	TNF-α(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.286	0.10
G1	4	2	0	0
G2	8	5	2	2
G3	5	2	2	3
ER					0.393	0.02
Positive	7	4	4	5
Negative	10	5	0	0
PR					0.419	0.01
Positive	7	4	4	5
Negative	10	5	0	0
HER-2/neu					0.451	0.01
Positive	5	5	4	4
Negative	12	4	0	1

Table 6 Correlation between the expressions of TNF-α in sentinel lymph nodes and pathological characteristics of breast tumor

Characteristics	TNF-α(n)				r value	P value
Characteristics	-	+	++	+++	r value	P value
Grade					0.286	0.10
G1	4	2	0	0
G2	8	5	2	2
G3	5	2	2	3
ER					0.393	0.02
Positive	7	4	4	5
Negative	10	5	0	0
PR					0.419	0.01
Positive	7	4	4	5
Negative	10	5	0	0
HER-2/neu					0.451	0.01
Positive	5	5	4	4
Negative	12	4	0	1

Figure 1 Example of sentinel lymph node biopsy with HE staining (a:×20; b:×10)

Figure 2 Example of IL-8(a) and TNF-α(b) of strong expression with immunohistochemistry staining

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