Tumor Invasiveness, Not Lymphangiogenesis, Is Correlated with Lymph Node Metastasis and Unfavorable Prognosis in Young Breast Cancer Patients (≤35 Years)

Abstract

The morbidity rate of breast cancer is on the rise, and the age of onset appears to be trending toward a young age. Breast cancer in young women (BCYW) has a number of distinctive features that differ from breast cancer in middle-aged or elderly women (BCMEW). Lymphatic metastasis plays an important role in the spread of BCYW; however, the mechanisms of lymph node metastasis (LNM) in BCYW are not clear. This study aimed to investigate the mechanism of lymphatic metastasis in BCYW and to evaluate the relationships between lymphangiogenesis, the expression of matrix metalloproteinase 9 (MMP-9) and vascular endothelial growth factor C (VEGF-C) expression, clinicopathological characteristics, and prognosis. Using immunohistochemistry, MMP-9, VEGF-C and the level of lymphatic microvessel density (LMVD) were analyzed in 106 cases of breast invasive ductal carcinoma and 20 cases of breast proliferative lesions. Compared with BCMEW, BCYW had higher MMP-9 expression, higher LNM, and more adverse prognoses. In BCYW, high MMP-9 expression was positively correlated with LNM and impaired survival time. However, in BCMEW, MMP-9 expression was not correlated with LNM or survival time. In addition, high VEGF-C expression was positively correlated with a high level of LMVD in both BCYW and BCMEW. Nevertheless, a high level of LMVD was not correlated with LNM or survival time in the two groups. More importantly, univariate and multivariate survival analysis showed that MMP-9 expression and LNM were independent prognostic factors in BCYW. Our present study indicates that lymphangiogenesis induced by VEGF-C is augmented in breast cancer; however, a higher level of lymphangiogenesis has no significant impact on LNM or survival time. We suggest that tumor invasiveness, rather than lymphangiogenesis, plays an important role in LNM among BCYW. Moreover, MMP-9 and LNM were independent prognostic factors for BCYW.

Fig 1. Representative images of LYVE1-positive lymphatic vessels and immunohistochemical staining of VEGF-C and MMP-9.

(A) The hot spots in LYVE1 immuno-stained sections. LYVE1-positive lymphatic vessels were thin-walled structures with irregular lumens, mainly present at the tumor periphery (blue arrows). Blood vessel endothelium near LYVE-1-positive lymph vessels were negatively stained (red arrows). (B) Positive expression of VEGF-C. (C) Positive expression of MMP-9. Diffuse and strong positive VEGF-C and MMP-9 immunostaining was mainly observed in the cytoplasm of breast cancer cells.

Fig 2. Comparisons of LMVD in different subgroups.

(A) LMVD in the BCYW and BCMEW groups and in the control group. LMVD in either BCYW or BCMEW was significantly higher than that in the breast proliferative lesions. However, no significant difference was found between the BCYW and BCMEW groups. (B) LMVD in breast cancer patients with or without LNM was not significantly different. (C) LMVD was not significantly different according to lymph node status in both the BCYW and BCMEW groups (Mann-Whitney U-test).

Fig 3. Kaplan-Meier OS (A and C) and DFS (B and D) curves.

(A) BCYW (age ≤35) had a significantly worse OS compared with BCMEW (age >40). (B) BCYW had a significantly worse DFS compared with BCMEW. (C) Breast cancer patients with LNM had a significantly worse OS compared to those without LNM. (D) Breast cancer patients with LNM had a significantly worse DFS compared to those without LNM.