Clinicopathological significance of hyaluronan and hyaluronidase 2 (HYAL2) in breast cancer.

Hyaluronan (HA), as a component of extracellular matrix, has pivotal roles in both physiological and pathological condition. In breast cancer, while high molecular weight HA is produced by hyaluronan synthase, it is degraded by hyaluronidases (hyaluronidase-1 (HYAL1) and hyaluronidase-2 (HYAL2)) into low molecular weight HA (LMW HA), which is considered to have pro-tumorigenic effects in human malignancies. However, HA and HYAL2, the rate-limiting enzyme of HA degradation, have not been comprehensively examined in breast cancer and clinicopathological significance of LMW HA remains to be elucidated in breast cancer. We therefore histochemically localized HA as well as HYAL2 in 116 breast cancer tissues. In addition, we examined size-dependent function of HA on breast cancer cell proliferation and migration using MCF-7 and MDA-MB-231 breast cancer cell lines. HA was localized in both the stroma and breast carcinoma cells, while HYAL2 was predominantly localized in breast carcinoma cells. HA was significantly correlated with cell proliferation and invasion ability as well as increased risk of recurrence especially in HYAL2 positive group. On the other hand, HYAL2 was correlated with breast cancer cell proliferation and increased risk of recurrence. In addition, in vitro analyses revealed that lower molecular weight HA increased sphere forming ability and migration in MCF-7 and MDA-MB-231, whereas higher molecular weight HA inhibited them. It was concluded that HA needs to be degraded by HYAL2 to exert pro-tumorigenic effects and comprehensive HA/HYAL2 status serves as a potent prognostic factor in breast cancer.

Toll-like receptor (TLR) 4 is a potent prognostic factor in prostate cancer associated with proliferation and invasion.

Prostate cancer is one of the most common malignancies in men, and there is a need to explore novel biomarkers or therapeutic targets. Toll-like receptor 4 (TLR4) is expressed not only in antigen-presenting cells but also types of human malignancies, contributing to disease progression, although its clinical significance or functional role in prostate cancer remains unclear. Therefore, we immunolocalized TLR4 in 117 prostate cancer tissues to address its clinicopathological significance. Additionally, we performed in vitro assays to examine the effects of TLR4 on proliferation and migration of prostate cancer cell lines (LNCaP, DU-145 and PC-3). TLR4 immunoreactivity was predominantly detected in the cytoplasm of prostate cancer cells, and it was positively associated with proliferation and invasion abilities, as well as Gleason score. Subsequent in vitro experiments revealed that the inhibition of TLR4 by Sparstolonin B (SsnB) significantly suppressed the proliferation and migration of LNCaP, DU-145 and PC-3 cells. Therefore, we concluded that TLR4 was a potent prognostic factor associated with proliferation and invasion, and it might serve as a therapeutic target in prostate cancer.

Clinicopathological Significance and Prognostic Role of High Mobility Group Box 1 (HMGB1), Toll-Like Receptor (TLR) 2 and TLR4 in Breast Cancer.

High-mobility group box 1 (HMGB1) functions as damage-associated molecular pattern (DAMPs), released into extracellular space during cellular stress. Extracellular HMGB1 act as signal molecules through Toll-like receptor (TLR) 2 or TLR4, exerting diverse functions in both normal cells and malignant cells including breast cancer. However, their comprehensive examination in breast cancer tissues is lacking. Thus, we immunolocalized them in 112 breast cancer tissues, correlating their immunoreactivity with clinicopathological parameters and clinical outcomes to clarify their significance in breast cancer. We demonstrated that nuclear HMGB1 immunoreactivity was correlated with tumor progression and longer disease-free survival. In contrast, TLR2 immunoreactivity was correlated with increased cell proliferation and shorter disease-free survival, dependent on cytoplasmic HMGB1 immunoreactivity. Additionally, TLR4 immunoreactivity correlated with chemoresistance, regardless of cytoplasmic HMGB1 immunoreactivity. It was therefore considered that TLR2 collaboratively contributed to breast cancer progression with HMGB1-DAMPs to become a worse prognostic factor. Meanwhile, TLR4 served as a worse prognostic factor associated with chemoresistance, irrespective of HMGB1.

Kallikrein-Related Peptidase 12 (KLK12) in Breast Cancer as a Favorable Prognostic Marker.

Kallikrein-related peptides (KLKs) form an evolutionally conserved subgroup of secreted serine proteases that consists of 15 members (KLK1-15). Previous studies have shown that KLKs regulate diverse biological processes, but the clinical significance of KLKs remains largely unclear in human breast cancers. We examined the expression profile of 15 KLK genes in breast carcinomas using microarray data. Next, we immunolocalized KLK12 in 140 breast carcinomas and evaluated its clinical significance. Subsequently, we examined the effects of KLK12 on proliferation and migration in breast cancer cell lines. From microarray analyses, it turned out that KLK12 was the most strongly associated with low-grade malignancy in breast carcinomas among the 15 KLK members. Immunohistochemical KLK12 status was positively associated with ER and PR status, while it was inversely associated with stage, pathological T factor, lymph node metastasis, and distant metastasis. Prognostic analyses demonstrated that KLK12 was a favorable prognostic factor for both disease-free and breast cancer-specific survival of the patients. Furthermore, the knockdown of KLK12 significantly increased cell proliferation activity and cell migration of breast cancer cells. These results suggest that KLK12 has antitumorigenic effects associated with proliferation and migration and immunohistochemical KLK12 status as a potent favorable prognostic factor in breast carcinoma patients.

The Pro-Tumorigenic Role of Chemotherapy-Induced Extracellular HSP70 from Breast Cancer Cells via Intratumoral Macrophages.

Tumor-associated macrophages (TAMs) contribute to tumor progression and chemoresistance; it is therefore important to clarify the altered functions of macrophages following chemotherapy. While extracellular heat shock protein (HSP) 70 is associated with therapeutic resistance, the effects of HSP70 on TAMs remain largely unknown. Here, we conducted in vitro experiments and immunohistochemistry in 116 breast carcinoma specimens to determine whether the secretion of HSP70 from breast cancer cells following chemotherapy affects macrophage function. It was revealed that the interaction of epirubicin (EPI)-exposed breast cancer cells with macrophages enhanced tumor progression, and EPI promoted the secretion of extracellular HSP70 from breast cancer cells. The expression of pro-tumorigenic macrophage marker CD163 was decreased in macrophages treated with a conditioned medium (CM) from HSP70-silenced breast cancer cells. Breast cancer cells treated with CM from HSP70-silenced breast cancer cells showed decreased expression of transforming growth factor (TGF)-ß, and the pro-tumorigenic effects of macrophages were impaired when TGF-ß signaling was inhibited. Immunohistochemistry demonstrated that HSP70 served as a poor prognostic factor in conjunction with macrophage infiltration. It was therefore concluded that extracellular HSP70 levels increased following chemotherapy and enhanced the pro-tumorigenic effects of TAMs, either directly or indirectly, by regulating TGF-ß expression in breast cancer cells.