HO-1 Interactors Involved in the Colonization of the Bone Niche: Role of ANXA2 in Prostate Cancer Progression

Abstract

Background: Prostate cancer (PCa) dissemination shows a tendency to develop in the bone, where heme oxygenase 1 (HO-1) plays a critical role in bone remodeling. Previously by LC/ESI-MSMS, we screened for HO-1 interacting proteins and identified annexin 2 (ANXA2). The aim of this study was to analyze the relevance of ANXA2/HO-1 in PCa and bone metastasis. Methods: We assessed ANXA2 levels using a co-culture transwell system of PC3 cells (pre-treated or not with hemin, an HO-1 specific inducer) and the pre-osteoclastic Raw264.7 cell line. Results: Under co-culture conditions, ANXA2 mRNA levels were significantly modulated in both cell lines. Immunofluorescence analysis unveiled a clear ANXA2 reduction in cell membrane immunostaining for Raw264.7 under the same conditions. This effect was supported by the detection of a decrease in Ca 2+ concentration in the conditioned medium. HO-1 induction in tumor cells prevented both, the ANXA2 intracellular relocation and the decrease in Ca 2+ concentration. Further, secretome analysis revealed urokinase (uPA) as a key player in the communication between osteoclast progenitors and PC3 cells. To assess the clinical significance of ANXA2/HO-1, we performed a bioinformatics analysis and identified that low expression of each gene strongly associated with poor prognosis in PCa regardless of the clinico-pathological parameters assessed. Further, these genes appear to behave in a dependent manner. Conclusions: ANXA2/HO-1 rises as a critical axis in PCa.Biomolecules 2020, 10, 467 2 of 24 microenvironment communicate and interact with each other to develop a fertile niche for the promotion of the metastatic process [3][4][5].Evidence has recognized inflammation as a risk factor for this neoplastic disease [6]. Heme oxygenase 1 (HO-1), encoded by the HMOX1 gene, is a stress response protein and a critical mediator of cellular homeostasis [7]. Although the role of HO-1 in cancer is controversial [8,9], we have shown that its pharmacologic or genetic upregulation is associated with a less aggressive phenotype in PCa [10]. HO-1 impairs tumor growth and angiogenesis in vivo and downregulates the expression of target genes associated with inflammation in PCa [11,12]. In the metastatic bone site, we demonstrated that HO-1 is capable of modulating signaling pathways relevant to skeletal PCa metastasis, such as FoxO/β-catenin and promotes bone remodeling when human tumor cells are transplanted into the femur of SCID mice [13]. Moreover, we have shown the direct effect of HO-1 on bone turnover and remodeling. When assessing the physiological impact of Hmox1 gene knockout on bone metabolism in vivo, histomorphometric analysis of Hmox1−/− mice bones exhibited significantly decreased bone density. A positive correlation between Hmox1 expression and key bone markers was observed in primary mouse osteoblasts (PMOs) [14]. These observations highlight the importance of HO-1 expression in bone, not only for the physiology of bone cells but also in the modulation of the communication between PMOs and PC...

EphA2–YES1–ANXA2 pathway promotes gastric cancer progression and metastasis

“…Ser phosphorylation of ANXA2 seems to be an essential event of the secretory process [ 23 ], while Tyr24 phosphorylation of ANXA2 (p-ANXA2-Y24) has been related to the localization of ANXA2 to the endocytic membrane system and the nucleus, and is associated with malignant transformation, EMT, tumor invasion, and metastasis [ 23 – 26 ]. ANXA2 has recently been linked to the metastasis of several types of cancer, such as prostate [ 27 ], colorectal [ 28 ], and breast cancer [ 29 ]. High expression of ANXA2 promotes the development of GC and is negatively correlated with the clinical prognosis [ 30 , 31 ].…”

HO-1 Modulates Aerobic Glycolysis through LDH in Prostate Cancer Cells

“…Regarding HMOX1 , we previously performed a similar analysis using these three databases, which revealed no significant differences in mRNA expression levels among tumoral and non-tumoral prostate tissue, but demonstrated that PCa patients with higher expression of HMOX1 showed longer RFS (HR: 0.5, p = 0.021) [ 36 ]. Herein, we found that the association between higher HMOX1 levels and longer RFS was not modified by LDHA expression ( Figure 6 D).…”

“…Furthermore, we have reported that HO-1 modulates bone remodeling and alters the communication between PCa and bone cells [ 35 ]. We have also demonstrated that HO-1 induction affects annexin 2 localization and calcium metabolism in osteoclasts co-cultured with PCa cells, thus suggesting the capability of HO-1 to modify the bone tumor niche [ 36 ].…”

Myxovirus Resistance Protein 1 (MX1), a Novel HO-1 Interactor, Tilts the Balance of Endoplasmic Reticulum Stress towards Pro-Death Events in Prostate Cancer

“…Total cell lysates and immunoblot analysis were carried out as previously described [ 18 ]. Briefly, cells were lysed with RIPA buffer (Tris HCl 50 mM pH 7.4; NaCl 150 mM; ethylenediaminetetraacetic acid (EDTA) 20 mM pH 8; sodium deoxycholate 1%; SDS 0.1%; Triton X-100 1%, 1 mM Na 3 VO 4 , 20 mM NaF, and 1 mM Na 4 P 2 O 7 , pH 7.9) and homogenized.…”