PREOPERATIVE PLASMA CONCENTRATION OF MMP-9/TIMP-1 COMPLEXES IS NOT ASSOCIATED WITH DISEASE OUTCOME IN PRIMARY BREAST CANCER (N=483)

Anne-Sofie Schrohl1, Sidse Ørnbjerg Würtz1, Susanne Møller2, Birgitte Sander Nielsen1, Henning T. Mouridsen2, and Nils Brünner1

1 University of Copenhagen, Faculty of Life Sciences & Sino-Danish Breast Cancer Research Centre, Copenhagen, Denmark2 Danish Breast Cancer Cooperative Group, Rigshospitalet, Copenhagen, Denmark

Contact: sofie@life.ku.dk

RESULTS

ASSAY AND SAMPLES

AIM & HYPOTHESIS

Plasma MMP-9/TIMP-1 complex levels were not associated with age, lymph node status, steroid hormone receptor status, or tumor size. Significant associations with tumor grade (p=0.04) and menopausal status (p=0.001) were found.

Table 1. Patient and tumor characteristics and association with MMP-9/TIMP-1 complexes

CharacteristicAll

patients(N=483)

MMP-9/TIMP-1

P(Χ2-test)Q1-25

(%)Q25-50

(%)Q50-75

(%)Q75-100

(%)

Age≤ 50 years50-69 years≥ 70 years

107 271105

362523

152625

272331

222721

0.06

Lymph node statusPositiveNegative

228255

2528

2224

2625

2723

0.70

Hormone rec. statusPositiveNegativeUnknown

39583

5

2631

2517

2524

2428 0.21

Tumor size0-20 mm> 20 mm

316167

2726

2422

2526

2426

0.92

Tumor gradeGrade 1Grade 2 and 3Unknown

206244

33

2628

2918

2329

2226 0.04

Menopausal statusPremenopausalPostmenopausal

145338

3523

1527

2625

2325

0.001

CONCLUSIONS

ACKNOWLEDGEMENTS

Variable Hazard ratio 95% CI P

MMP-9/TIMP-1 complexQ25-50 vs. Q1-25Q50-75 vs. Q1-25Q75-100 vs. Q1-25

0.951.010.92

0.55-1.620.61-1.680.55-1.57

0.840.960.77

Age50-69 years vs. < 50 years≥ 70 years vs. < 50 years

0.992.48

0.45-2.211.61-3.83

0.99< 0.0001

Lymph node statusPositive vs. negative 1.18 0.80-1.75 0.40

Hormone rec. statusPositive vs. negative 0.46 0.29-0.73 0.001

Tumor size> 20 mm vs. 0-20 mm 1.31 0.88-1.95 0.18

Tumor gradeGrade 2 and 3 vs. grade 1 1.34 0.86-2.08 0.20

Menopausal statusPost- vs. premenopausal 1.34 0.64-2.84 0.44

Only age ≥ 70 years and steroid hormone receptor status contributed significantly to the multivariable model. MMP-9/TIMP-1 complex levels were not associated with recurrence-free survival.

Table 2. Multivariable Cox proportional hazards model of recurrence-free survival

Figure 1. Recurrence-free survival of patients grouped in quartiles by plasma MMP-9/TIMP-1 level

Plasma levels of MMP-9/TIMP-1 complexes were not associated with recurrence-free survival (logrank test p=0.9565).

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No association was found between plasma concentration of MMP-9/TIMP-1 complexes and disease outcome

These results suggest that other fractions of MMP-9 and TIMP-1 carry prognostic information, e.g. free MMP-9 and TIMP-1 or MMP-9 and TIMP-1 bound to other plasma proteins

This work was supported by The Danish Cancer Society

Anne-Sofie Schrohl is supported by the Danish Council for Independent Research | Medical Sciences

Both Matrix Metalloproteinase-9 (MMP-9) and Tissue Inhibitor of Metalloproteinases-1 (TIMP-1) are associated with a poor prognosis in breast cancer.

Here, our aim was to measure the concentration of MMP-9/TIMP-1 complexes in plasma from primary breast cancer patients and to evaluate whether these levels are associated with disease outcome.

The hypothesis was that high plasma levels of MMP-9/TIMP-1 complexes are associated with increased recurrence risk in primary breast cancer patients.

MMP-9/TIMP-1 COMPLEX ELISA

The human MMP-9/TIMP-1 Complex DuoSet® ELISA Development System (R&D Systems, Inc.) was validated for measurement of MMP-9/TIMP-1 complex levels in human plasma. The validation process included investigation of reproducibility (intra-assay, inter-assay), recovery, and linearity upon dilution of samples.

PLASMA SAMPLES

Preoperative EDTA plasma samples from 483 consecutively enrolled primary breast cancer patients were analyzed. Prior to analysis, samples were diluted in reagent diluent as recommended by the manufacturer (1:3).

The median MMP-9/TIMP-1 complex concentration was 2.06 ng/mL (range, 0.11 - >14.8 ng/mL).

DATA ANALYSIS

The possible associations between MMP-9/TIMP-1 plasma level and patient- and tumor characteristics were analyzed using a Χ2 test. The possible association with recurrence-free survival was investigated using a logrank test and a multivariable Cox proportional hazards model.

In the analyses, patients were grouped in quartiles based on their plasma MMP-9/TIMP-1 complex level.

Recurrence-free survival was defined as survival without breast cancer relapse, contralateral breast cancer, other malignant disease and death without a previous relapse.

REFERENCESLi H. et al (2004). “Prognostic value of matrix metalloproteinases (MMP-2 and MMP-9) in patients with lymph node-negative breast carcinoma.” Breast Cancer Res Treat 88: 75-85

Somiari S.B. et al (2006). “Circulating MMP2 and MMP9 in breast cancer – Potential role in classification of patients into low risk, high risk, benign disease and breast cancer categories.” Int J Cancer 119: 1403-1411

Würtz S.Ø. et al (2008). “Plasma and serum levels of tissue inhibitor of metalloproteinases-1 are associated with prognosis in node-negative breast cancer: a prospective study.” Mol Cell Proteomics 7(2): 424-430