Cancer stem cells, bone and tumor microenvironment: Key players in bone metastases

Tumor mass is constituted by a heterogeneous group of cells, among which a key role is played by the cancer stem cells (CSCs), possessing high regenerative properties. CSCs directly metastasize to bone, since bone microenvironment represents a fertile environment that protects CSCs against the immune system, and maintains their properties and plasticity. CSCs can migrate from the primary tumor to the bone marrow (BM), due to their capacity to perform the epithelial-to-mesenchymal transition. Once in BM, they can also perform the mesenchymal-to-epithelial transition, allowing them to proliferate and initiate bone lesions. Another factor explaining the osteotropism of CSCs is their ability to recognize chemokine gradients toward BM, through the CXCL12\u2013CXCR4 axis, also known to be involved in tumor metastasis to other organs. Moreover, the expression of CXCR4 is associated with the maintenance of CSCs\u2019 stemness, and CXCL12 expression by osteoblasts attracts CSCs to the BM niches. CSCs localize in the pre-metastatic niches, which are anatomically distinct regions within the tumor microenvironment and govern the metastatic progression. According to the stimuli received in the niches, CSCs can remain dormant for long time or outgrow from dormancy and create bone lesions. This review resumes different aspects of the CSCs\u2019 bone metastastic process and discusses available treatments to target CSCs

Gastric Cancer: The Times they are a-changin’

Gastric cancer is the third leading cause of cancer death worldwide. Even though during these last decades gastric cancer incidence decreased in Western countries, it remains endemic and with a high incidence in Eastern countries. The survival in advanced and metastatic stage of gastric cancer is still very poor. Recently the Cancer Genoma Atlas Research Network identified four subtypes with different molecular profiles to classify gastric cancer in order to offer the optimal targeted therapies for pre-selected patients. Indeed, the key point is still the selection of patients for the right treatment, on basis of molecular tumor characterization. Since chemotherapy reached a plateau of efficacy for gastric cancer, the combination between cytotoxic therapy and biological agents gets a better prognosis and decreases chemotherapeutic toxicity. Currently, Trastuzumab in combination with platinum and fluorouracil is the only approved targeted therapy in the first line for c-erbB2 positive patients, whereas Ramucirumab is the only approved targeted agent for patients with metastatic gastric cancer. New perspectives for an effective treatment derived from the immunotherapeutic strategies. Here, we report an overview on gastric cancer treatments, with particular attention to recent advances in targeted therapies and in immunotherapeutic approach

Osteogenic differentiation modulates the cytokine, chemokine, and growth factor profile of ASCs and SHED

Great efforts have been made to improve bone regeneration techniques owing to a growing variety of sources of stem cells suitable for autologous transplants. Specifically, adipose-derived stem cells (ASCs) and stems cells from human exfoliated deciduous teeth (SHED) hold great potential for bone tissue engineering and cell therapy. After a preliminary characterization of the main biomolecules ASCs and SHED released in their conditioned media, cells were kept both in normal and osteo-inducing conditions. Conventional assays were performed to prove their osteogenic potential such as quantitative real-time polymerase chain reaction (qRT-PCR) (for RUNX-2, collagen type I, osteopontin and osteonectin), alkaline phosphatase activity, osteocalcin production, and von Kossa staining. Conditioned media were tested again after the osteogenic induction and compared to maintaining condition both at base line and after 14 days of culture. The osteogenic condition inhibited the release of all the biomolecules, with the exception, concerning SHED, of growth-regulated alpha protein precursor (GRO\u3b1), and, to a lesser extent, interleukin (IL)-8. In conclusion, our data support that undifferentiated ASCs and SHED may be preferable to committed ones for general cell therapy approaches, due to their higher paracrine activity. Osteoinduction significantly affects the cytokine, chemokine, and growth factor profile in a differential way, as SHED kept a more pronounced pro-angiogenic signature than ASCs

Bone invading NSCLC cells produce IL-7: mice model and human histologic data

<p>Abstract</p> <p>Background</p> <p>Bone metastases are a common and dismal consequence of lung cancer that is a leading cause of death. The role of IL-7 in promoting bone metastases has been previously investigated in NSCLC, but many aspects remain to be disclosed. To further study IL-7 function in bone metastasis, we developed a human-in-mice model of bone aggression by NSCLC and analyzed human bone metastasis biopsies.</p> <p>Methods</p> <p>We used NOD/SCID mice implanted with human bone. After bone engraftment, two groups of mice were injected subcutaneously with A549, a human NSCLC cell line, either close or at the contralateral flank to the human bone implant, while a third control group did not receive cancer cells. Tumor and bone vitality and IL-7 expression were assessed in implanted bone, affected or not by A549. Serum IL-7 levels were evaluated by ELISA. IL-7 immunohistochemistry was performed on 10 human bone NSCLC metastasis biopsies for comparison.</p> <p>Results</p> <p>At 12 weeks after bone implant, we observed osteogenic activity and neovascularization, confirming bone vitality. Tumor aggressive cells implanted close to human bone invaded the bone tissue. The bone-aggressive cancer cells were positive for IL-7 staining both in the mice model and in human biopsies. Higher IL-7 serum levels were found in mice injected with A549 cells close to the bone implant compared to mice injected with A549 cells in the flank opposite to the bone implant.</p> <p>Conclusions</p> <p>We demonstrated that bone-invading cells express and produce IL-7, which is known to promote osteoclast activation and osteolytic lesions. Tumor-bone interaction increases IL-7 production, with an increase in IL-7 serum levels. The presented mice model of bone invasion by contiguous tumor is suitable to study bone-tumor cell interaction. IL-7 plays a role in the first steps of metastatic process.</p

Osteoclasts Are Active in Bone Forming Metastases of Prostate Cancer Patients

BACKGROUND: Bone forming metastases are a common and disabling consequence of prostate cancer (CaP). The potential role of osteoclast activity in CaP bone metastases is not completely explained. In this study, we investigated ex vivo whether the osteolytic activity is present and how it is ruled in CaP patients with bone forming metastases. METHODOLOGY: Forty-six patients affected by newly diagnosed CaP and healthy controls were enrolled. At diagnosis, 37 patients had a primary tumour only, while 9 had primary tumour and concomitant bone forming metastases. In all patients there was no evidence of metastasis to other non-bone sites. For all patients and controls we collected blood and urinary samples. We evaluated patients' bone homeostasis; we made peripheral blood mononuclear cell (PBMC) cultures to detect in vitro osteoclastogenesis; we dosed serum expression of molecules involved in cancer induced osteoclatogenesis, such as RANKL, OPG, TNF-alpha, DKK-1 and IL-7. By Real-Time PCR, we quantified DKK-1 and IL-7 gene expression on micro-dissected tumour and healthy tissue sections. PRINCIPAL FINDINGS: CaP bone metastatic patients showed bone metabolism disruption with increased bone resorption and formation compared to non-bone metastatic patients and healthy controls. The CaP PBMC cultures showed an enhanced osteoclastogenesis in bone metastatic patients, due to an increase of RANKL/OPG ratio. We detected increased DKK-1 serum levels and tissue gene expression in patients compared to controls. IL-7 resulted high in patients' sera, but its tissue gene expression was comparable in patients and controls. CONCLUSIONS: We demonstrated ex vivo that osteoclastogenesis is an active mechanism in tumour nesting of bone forming metastatic cancer and that serum DKK-1 levels are increased in CaP patients, suggesting to deeply investigate its role as tumour marker

Isolation and characterization of buccal fat pad and dental pulp mscs from the same donor

Mesenchymal stem cells (MSCs) can be harvested from different sites in the oral cavity, representing a reservoir of cells useful for regenerative purposes. As direct comparisons between at least two types of MSCs deriving from the same patient are surprisingly rare in scientific literature, we isolated and investigated the osteoinductive potential of dental pulp stem cells (DPSCs) and buccal fat pad stem cells (BFPSCs). MSCs were isolated from the third molar dental pulp and buccal fat pads of 12 patients. The number of viable cells was quantified through manual count. Proliferation and osteodifferentiation assays, flow cytometry analysis of cell phenotypes, and osteocalcin release in vitro were performed. The isolation of BFPSCs and DPSCs was successful in 7 out of 12 (58%) and 3 out of 12 (25%) of retrieved samples, respectively. The yield of cells expressing typical stem cell markers and the level of proliferation were higher in BFPSCs than in DPSCs. Both BFP-SCs and DPSCs differentiated into osteoblast-like cells and were able to release a mineralized matrix. The release of osteocalcin, albeit greater for BFPSCs, did not show any significant difference between BFPSCs and DPSCs. The yield of MSCs depends on their site of origin as well as on the protocol adopted for their isolation. Our data show that BFP is a valuable source for the derivation of MSCs that can be used for regenerative treatments

A novel method to optimize autologous adipose tissue recovery with extracellular matrix preservation

This work aims to characterize a new method to recover low-manipulated human adipose tissue, enriched with adipose tissue-derived mesenchymal stem cells (ATD-MSCs) for autologous use in regenerative medicine applications. Lipoaspirated fat collected from patients was processed through Lipocell, a Class II-a medical device for dialysis of adipose tissue, by varying filter sizes and washing solutions. ATD-MSC yield was measured with flow cytometry after stromal vascular fraction (SVF) isolation in fresh and cultured samples. Purification from oil and blood was measured after centrifugationwith spectrophotometer analysis. Extracellularmatrix preservationwas assessed through hematoxylin and eosin (H&amp;E) staining and biochemical assay for total collagen, type-2 collagen, and glycosaminoglycans (GAGs) quantification. Flow cytometry showed a two-fold increase of ATD-MSC yield in treated samples in comparisonwith untreated lipoaspirate; no differenceswhere reportedwhen varying filter size. The association of dialysis and washing thoroughly removed blood and oil from samples. Tissue architecture and extracellular matrix integrity were unaltered after Lipocell processing. Dialysis procedure associated with Ringer's lactate preserves the proliferation ability of ATD-MSCs in cell culture. The characterization of the product showed that Lipocell is an efficient method for purifying the tissue from undesired byproducts and preserving ATD-MSC vitality and extracellular matrix (ECM) integrity, resulting in a promising tool for regenerative medicine applications