Activin A circulating levels in patients with bone metastasis from breast or prostate cancer

Recent studies have highlighted that Activin A, a member of the transforming growth factor-β (TGF-β) superfamily, may be involved in the regulation of osteoblastic activity and in osteoclast differentiation. Therefore, we have investigated the clinical significance of its circulating levels in patients with bone metastasis. Activin A serum concentrations were determined, by a commercially available enzyme-linked immunosorbent assay kit, in 72 patients with breast cancer (BC) or prostatic cancer (PC) with (BM+) or without (BM−) bone metastases, in 15 female patients with age-related osteoporosis (OP), in 20 patients with benign prostatic hypertrophy (BPH) and in 48 registered healthy blood donors (HS) of both sex (25 female and 23 male). Activin A serum concentrations were significantly increased in BC or PC patients as compared to OP (P < 0.0001) or BPH (P = 0.045), respectively, or to sex matched HS (P < 0.0001). Additionally, these levels resulted more elevated in PC patients as compared to BC patients (P = 0.032). Interestingly, Activin A was significantly higher in BM+ patients than in BM− patients (BC, P = 0.047; PC, P = 0.016). In BC patients, a significant correlation was observed only between Activin A and number of bone metastases (P = 0.0065) while, in PC patients, Activin A levels were strongly correlated with the Gleason score (P = 0.011) or PSA levels (P = 0.0001) and, to a lessen extent, with the number of bone metastases (P = 0.056). Receiver operating characteristic curve (ROC) analysis showed a fair diagnostic accuracy of Activin A to discriminate between BM+ and BM− patients (BC: AUC = 0.71 ± 0.09, P = 0.03; PC: AUC = 0.73 ± 0.081, P = 0.005). These findings indicate that Activin A may be implicated in the pathogenesis of bone metastasis. Therefore, this cytokine may be considered a novel potential target for a more selective therapeutic approach in the treatment of skeletal metastasis and may be also useful as additional biochemical marker of metastatic bone disease.     

Matrix metalloproteinase activity and osteoclasts in experimental prostate cancer bone metastasis tissue

Previously, we and others showed that broad spectrum pharmaceutical inhibition of matrix metalloproteinase (MMP) activity reduces intraosseous tumor burden and bone degradation in animal models of bone metastasis. Herein, we used specific assays to measure net enzymatic activities of individual MMPs during colonization of bone by prostate cancer cells. PC3 cells were injected into the marrow of human fetal femurs previously implanted in SCID mice. Net MMP-9 activity in bone tissues peaked 2 weeks after injection, coinciding with a wave of osteoclast recruitment. In contrast, MMP-2 and MT1-MMP activity did not change. In vitro, co-culture of PC3 cells with bone tissue led to activation of pro-MMP-9 and increases in secreted net MMP-9 activity. Activation of pro-MMP-9 was prevented by metalloprotease inhibitors but not by inhibitors of other classes of proteases. Ribozyme suppression of MMP-9 expression in PC3 cells did not affect pro-MMP-9 activation or net MMP-9 activity and did not affect the phenotype of bone tumors. siRNA targeting of MMP-9 expression in preosteoclasts in vitro demonstrated that tumor-induced preosteoclast motility was dependent on MMP-9 expression. These data suggest that osteoclast-derived MMP-9 may represent a potential therapeutic target in bone metastasis and provide a rationale for the development of MMP-9-specific inhibitors.     

Species-specific homing mechanisms of human prostate cancer metastasis in tissue engineered bone

The development of effective therapeutic strategies against prostate cancer bone metastases has been impeded by the lack of adequate animal models that are able to recapitulate the biology of the disease in humans. Bioengineered approaches allow researchers to create sophisticated experimentally and physiologically relevant in vivo models to study interactions between cancer cells and their microenvironment under reproducible conditions. The aim of this study was to engineer a morphologically and functionally intact humanized organ bone which can serve as a homing site for human prostate cancer cells. Transplantation of biodegradable tubular composite scaffolds seeded with human mesenchymal progenitor cells and loaded with rhBMP-7 resulted in the development of a chimeric bone construct including a large number of human mesenchymal cells which were shown to be metabolically active and capable of producing extracellular matrix components. Micro-CT analysis demonstrated that the newly formed ossicle recapitulated the morphological features of a physiological organ bone with a trabecular network surrounded by a cortex-like outer structure. This microenvironment was supportive of the lodgement and maintenance of murine haematopoietic cell clusters, thus mimicking a functional organ bone. Bioluminescence imaging demonstrated that luciferase-transduced human PC3 cells reproducibly homed to the humanized tissue engineered bone constructs, proliferated, and developed macro-metastases. This model allows the analysis of interactions between human prostate cancer cells and a functional humanized bone organ within an immuno-incompetent murine host. The system can serve as a reproducible platform to study effects of therapeutics against prostate cancer bone metastases within a humanized microenvironment.     

PET/CT对小动物前列腺癌骨转移的观察研究

目的　探讨小动物PET/CT扫描在小鼠前列腺癌骨转移检测中的可行性。 方法　裸鼠23只,随机分为对照组2只,前列腺原位注射组、左心室注射组、胫骨腔内注射组每组各7只。分别采用前列腺原位注射、左心室内注射、胫骨腔内注射的方法建立前列腺癌骨转移动物模型。建模成功后饲养40天,原位注射及左心室注射组采用PET/CT扫描检测,胫骨腔注射组采用小动物高分辨率CT检测,对可疑的骨转移灶行解剖学观察及HE染色明确。 结果　前列腺原位注射组肿瘤细胞聚集腹腔软组织内生长,均未发现明显骨质破坏（0/7）;左心室注射组均发生皮下等软组织转移（7/7）, PET/CT检测出一例胫骨上端骨质破坏,组织学检测证实为骨转移灶(1/7);胫骨腔注射组所有动物均形成明显骨质破坏（7/7）且高分辨率CT检测见骨破坏分级良好。 结论　小动物PET/CT扫描能够良好的显示转移灶在小鼠体内的定位,且能良好的显示骨破坏情况,因此该技术在检测小动物骨转移上有着良好的可行性。     

Allelic losses on 18q21 are associated with progression and metastasis in human prostate cancer

We analyzed normal/tumor DNA pairs obtained from 46 patients with prostate cancers (stage B, 16 cases; C, 10 cases; D1, 4 cases; and endocrine therapy-resistant cancer-death, 16 cases) for loss of heterozygosity using 32 microsatellite markers on chromosome 18. Seventeen of the 46 cases (37%) showed loss of heterozygosity (LOH) for at least one locus on the long arm. Detailed deletion mapping in these tumors identified a distinct commonly deleted region within a 5-cM interval on 18q21.1. There was a statistical correlation between the frequency of LOH on 18q and clinical stage (χ² = 12.3; P = 0.0064). LOH on 18q was observed more frequently in Stage D1 cases (4/4; 100%) than in Stage B+C cases (5/26; 19%; P = 0.0046, Fisher's exact test). In 8 of 9 (89%) cancer-death patients from whom DNAs were available from both primary and metastatic tumors, the primary tumors had either no detectable abnormality of chromosome 18 or the region involving loss of heterozygosity was limited while the metastatic foci showed more frequent and extended allelic losses on this chromosome. No abnormalities were detected in the DCC and DPC4 genes when their exons were analyzed separately by single strand conformation polymorphism assay. These results suggest that inactivation of one or more putative tumor suppressor genes on 18q21 other than DCC and DPC4 plays an important role in the progression of human prostate cancer. Genes Chromosomes Cancer 20:140–147, 1997. © 1997 Wiley-Liss, Inc.     

Macrophage migration inhibitory factor (MIF) gene polymorphisms are associated with increased prostate cancer incidence

Recurrent or persistent inflammation has emerged as an important factor in cancer development. Overexpression of macrophage migration inhibitory factor (MIF), an upstream regulator of innate immunity with pleiotropic effects on cell proliferation, has been implicated in prostate cancer (CaP). Two polymorphisms in the promoter of the MIF gene (-173G to C transition and seven copies of the -794 CATT repeat) are associated with increased MIF expression in vivo and poor prognosis in autoimmune diseases. We conducted a retrospective analysis of 131 CaP patients and 128 controls from a group of Veterans' Administration patients undergoing routine prostate-specific antigen screening. Patients with CaP were enrolled regardless of treatment. Inclusion criteria for the control group were absence of documented diagnosis of cancer and/or chronic inflammation within patient computerized records. Logistic regression demonstrated a significant association between CaP and the -173G/C, the -173C/C and the -794 7-CATT MIF polymorphisms (P<0.001). Patients with the -794 7-CATT allele had an increased risk of CaP recurrence at 5 years. Individuals with -173G/C, -173C/C and -794 7-CATT MIF genotypes have an increased incidence of CaP and these genotypes may serve as an independent marker for cancer recurrence.     

乳腺微钙化促进乳腺癌骨转移的研究进展

乳腺癌是女性发病率最高的癌症,在过去的十年中,全球发病率持续上升,死亡率高居不下.最新的统计学研究表明,仅2018年,全球范围内检测出约210万乳腺癌病例,其中近63万患者死亡,因而也是女性死亡率最高的癌症[1].然而,许多癌症患者的死亡并非由于肿瘤在原发部位生长,而是在于肿瘤侵袭或转移至其他部位,其中乳腺癌最容易发生...     

Inhibition of the Axl pathway impairs breast and prostate cancer metastasis to the bones and bone remodeling

Clinical & Experimental Metastasis - Approximately 90% of cancer-related deaths result from cancer metastasis. In prostate and breast cancers, bone is the most common site of cancer cell...     

Allelic losses at loci on chromosome 10 are associated with metastasis and progression of human prostate cancer

DNA samples from tumors and paired normal tissues from 48 patients with prostate cancer (stage B, 16 cases; stage C, 14 cases; stage D, 18 cases) were examined with 26 polymorphic markers spanning chromosome 10. Allelic losses were observed in 17 of the 46 cases (37%) that were informative with at least one of the markers. Detailed deletion mapping identified two distict commonly deleted regions on the long arm of chromosome 10 (10q22–q24:7cM and 10q25.1:17cM) and one on 10p, suggesting that at least three tumor suppressor genes associated with prostate cancer are present on this chromosome. We observed loss of heterozygosity more frequently in tumors from fatal cases (stage D, 8/16, 50%) than in localized tumors (stage B, 0/16, 0%; P = 0.001 or stage B + C, 5/30, 17%; P = 0.02 Fisher's exact test). All metastatic tissues showed allelic loss at one or more loci on 10q. In five of the nine patients from whom DNAs were available from both metastatic and primary tumors, the primary cancer foci had no detectable abnormality of chromosome 10, while the metastatic foci showed allelic loss on chromosome 10. These results suggested that inactivation of one or more tumor suppressor genes on chromosome 10 plays an important role in late stages of prostate cancer. Genes Chromosom Cancer 17:245–253 (1996). © 1996 Wiley-Liss, Inc.     

Serum follistatin in patients with prostate cancer metastatic to the bone

The clinical significance of circulating follistatin (FLST), an inhibitor of the multifunctional cytokine activin A (Act A), was investigated in patients with prostate cancer (PCa). The serum concentrations of this molecule were determined by an enzyme-linked immunosorbent assay (ELISA) in PCa patients with (M+) or without (M0) bone metastases, in patients with benign prostate hyperplasia (BPH) and in healthy subjects (HS). The effectiveness of FLST in detecting PCa patients with skeletal metastases was determined by the receiver operating characteristic (ROC) curve analysis. Serum FLST was significantly higher in PCa patients than in BPH patients (P = 0.001) or HS (P = 0.011). Conversely, in BPH patients, FLST levels resulted lower than in HS (P = 0.025). In cancer patients the serum concentrations of FLST significantly correlated with the presence of bone metastases (P = 0.0005) or increased PSA levels (P = 0.04). Interestingly, significant differences in the ratio between FLST and Act A serum concentrations (FLST/Act A) were observed between HS and BPH patients (P = 0.001) or PCa patients (P = 0.0005). Finally, ROC curve analysis, highlighted a sound diagnostic performance of FLST in detecting M+ patients (P = 0.0001). However, the diagnostic effectiveness of FLST did not result significantly superior to that of Act A or PSA. These findings suggest that FLST may be regarded as a potential, molecular target in the treatment of metastatic bone disease while its clinical role as soluble marker in the clinical management of PCa patients with bone metastases needs to be better defined.     

Histopathologic analysis of bone marrow and bone metastasis in murine neuroblastoma

To elucidate the development of bone metastasis in human neuroblastoma, bone marrow and bone metastases were analysed histologically in a hematogeneous metastasis model of murine neuroblastoma. The bone marrow metastasis occurred initially in the bone marrow sinusoid where tumor cells adhered and extravasated to bone marrow parenchyma, resulting in the formation of nodular lesions in the medullary cavity. The nodular lesions eventually progressed to diffuse lesions segmentally occupying the medullary cavity. During the establishment of the diffuse lesions, tumor cells invaded cancellous bone and/or bone cortex, resulting in bone metastasis. Such nodular or diffuse bone marrow metastatic lesions occurred sporadically in a variety of bones. To improve the results of treatment for neuroblastoma, the characteristics of the bone marrow and the bone metastases demonstrated in this study should be considered in the diagnosis and treatment of this disease.     

Amplification and overexpression of vinculin are associated with increased tumour cell proliferation and progression in advanced prostate cancer

Androgen withdrawal is the standard treatment for advanced prostate cancer. Although this therapy is initially effective, nearly all prostate cancers become refractory to it. Approximately 15% of these castration-resistant prostate cancers harbour a genomic amplification at 10q22. The aim of this study was to explore the structure of the 10q22 amplicon and to determine the major driving genes. Application of high-resolution array-CGH using the 244k Agilent microarrays to cell lines with 10q22 amplification allowed us to narrow down the common amplified region to a region of 5.8 megabases. We silenced each of the genes of this region by an RNAi screen in the prostate cancer cell lines PC-3 and 22Rv1. We selected genes with a significant growth reduction in the 10q22 amplified cell line PC-3, but not in the non-amplified 22Rv1 cells, as putative target genes of this amplicon. Immunohistochemical analysis of the protein expression of these candidate genes on a tissue microarray enriched for 10q22 amplified prostate cancers revealed vinculin as the most promising target of this amplicon. We found a strong association between vinculin gene amplification and overexpression (p < 0.001). Further analysis of 443 specimens from across all stages of prostate cancer progression showed that vinculin expression was highest in castration-resistant prostate cancers, but negative or very low in benign prostatic hyperplasia (p < 0.0001). Additionally, high tumour cell proliferation measured by Ki67 expression was significantly associated with high vinculin expression in prostate cancer (p < 0.0001). Our data suggest that vinculin is a major driving gene of the 10q22 amplification in prostate cancer and that vinculin overexpression might contribute to prostate cancer progression by enhancing tumour cell proliferation.     

A novel orthotopic model of breast cancer metastasis to bone

Breast cancer affects approximately one woman in twelve and kills more women than any other cancer. If detected early, patients have a five year survival rate of 66%, but once metastatic disease has developed, there is no effective treatment. About 70% of patients with metastatic disease have bone involvement, while lungs and liver are the other common targets. Bone metastases cause severe pain, pathological fractures and hypercalcaemia and thus are a significant clinical problem. The development of new therapies for metastatic breast carcinoma depends on a better understanding of the mechanism of homing of the tumour cells to bone, liver and lungs and the factors required for their growth in these organs. Research on mechanisms of breast cancer metastasis, particularly to bone, has relied on in vitro studies or on tumour models in which the inoculation route is designed to promote delivery of tumour cells to a specific organ. Metastases in bone are achieved by inoculation into the right ventricle of the heart. To our knowledge there has been no report of a model of metastatic spread from the mammary gland to distant sites which reliably includes bone. In this paper, we describe our recent development of a novel murine model of metastatic breast carcinoma. The new model is unique in that the pattern of metastatic spread closely resembles that observed in human breast cancer. In particular, these murine breast tumours metastasise to bone from the primary breast site and cause hypercalcaemia, characteristics not normally found in murine tumours, but common in human disease. Furthermore, in a preliminary characterisation of this model, we show that secretion of parathyroid hormone-related protein, a role for which has been implicated in breast cancer spread to bone, correlates with metastasis to bone. This model therefore provides an excellent experimental system in which to investigate the factors that control metastatic spread of breast cancer to specific sites, particularly bone. The special advantage of this system is that it involves the whole metastasis process, beginning from the primary site. Existing models consider mechanisms that pertain to growth of tumour once the site has been reached. An understanding of the regulation of these factors by potential therapeutic agents could lead to improvement in therapies designed to combat metastatic disease. For the first time, this development will allow exploration of the molecular basis of site-specific metastasis of breast cancer to bone in a clinically relevant model.     

Formation of vasculogenic mimicry in bone metastasis of prostate cancer: Correlation with cell apoptosis and senescence regulation pathways

Vasculogenic mimicry (VM) has been found in prostate cancer (PCa) as an independent marker of poor prognosis. To investigate the correlation between VM and bone metastasis in PCa, a total of 80 cases were analyzed by CD31 and PAS dual-staining as well as the follow-up data. All cases were divided into two groups: VM-positive and VM-negative (VM-pos/VM-neg). Immunohistochemical staining for investigating the expression of Casepase-3, Bcl-2/Bax, and SA-β-gal was performed. 28 of the 80 PCa cases exhibited VM structure (35.0%). The incidence of bone metastasis in the VM-pos and VM-neg was 67.9% (19/28) and 38.5% (20/52), respectively. The positive rate of Casepase-3 and Bcl-2 expression was significantly different of the two groups (Caspases-3: VM-pos 71.4%, 20/28 vs VM-neg 42.3%, 22/52; Bcl-2: VM-pos 35.7%, 10/28 vs VM-neg 65.4%, 34/52). Bcl-2/Bax ratio of the VM-pos (0.71 ± 0.22) was lower than that of the VM-pos (0.89 ± 0.13). In addition, a higher frequency of SA-β-gal was detected in VM-pos (64.29 ± 86.42) than in VM-neg (25.37 ± 72.21). Taken together, our findings demonstrate that PCa with VM has the tendency to develop bone metastasis. Activations of cell apoptosis and senescence regulation pathways may play important roles in the formation process of VM structure.