Over the last decades, the concept of precision medicine has dramatically renewed the field of medical oncology; the introduction of patient-tailored therapies has significantly improved all measurable outcomes. Liquid biopsy is a revolutionary technique that is opening previously unexpected perspectives. It consists of the detection and isolation of circulating tumor cells, circulating tumor DNA and exosomes, as a source of genomic and proteomic information in patients with cancer. Many technical hurdles have been resolved thanks to newly developed techniques and next-generation sequencing analyses, allowing a broad application of liquid biopsy in a wide range of settings. Initially correlated to prognosis, liquid biopsy data are now being studied for cancer diagnosis, hopefully including screenings, and most importantly for the prediction of response or resistance to given treatments. In particular, the identification of specific mutations in target genes can aid in therapeutic decisions, both in the appropriateness of treatment and in the advanced identification of secondary resistance, aiming to early diagnose disease progression. Still application is far from reality but ongoing research is leading the way to a new era in oncology. This review summarizes the main techniques and applications of liquid biopsy in cancer.

Although statins are lipid-lowering drugs that block cholesterol biosynthesis, they exert immunomodulatory, anti-inflammatory, anti-angiogenic and anti-proliferative functions by reducing the isoprenylation of proteins involved in cell signal transduction such as Ras and RhoA. In this study, we provide evidence that several natural (lovastatin, simvastatin and pravastatin) and synthetic (cerivastatin and atorvastatin) statins exert a cytotoxic effect on human T, B and myeloma tumor cells by promoting their apoptosis. Dissimilar susceptibility to apoptosis has been detected in these lines, presumably in relation to the altered expression of proteins involved in the regulation of cellular signals. Cerivastatin promptly activated the cell death even in doxorubicin resistant cell lines such as MCC-2, whereas pravastatin, a hydrophilic compound, failed to induce any effect on either proliferation or apoptosis. The statin-induced apoptotic pathway in these cell lines was presumably regulated by altered prenylation of either Ras or RhoA, as measured by the defective membrane localization of these small GTPases. In addition the cell proliferation was rescued by both farnesylpyrophosphate (FPP) and geranyl-geranylpyrophosphate (GGPP), whereas no effect was obtained with squalene, a direct precursor of cholesterol. Statins primed apoptosis through its intrinsic pathway involving the mitochondria. In fact, we observed the reduction of mitochondrial membrane potential and the cytosolic release of the second mitochondria-derived activator of caspases (Smac/DIABLO). The apoptotic pathway was caspase-dependent since caspases 9, 3 and 8 were efficiently activated. These results support the potential use of statins in association with conventional treatment as apoptosis-triggering agents in these tumors.

The study was funded by Italian Association for Cancer Research (IG grant 17536), and from the Apulia Region ('Oncogenomic Project' and 'Jonico-Salentino Project'). All Authors declare no competing interests.

Objective. To investigate whether immunologic abnormalities in patients with Behçet's disease (BD) are related to abnormalities of the Th1/Th2 ratio.Methods. Th1/Th2 cytokine production by peripheral blood lymphocytes (PBL) from 31 patients with BD, 11 patients with inflammatory arthritis, and 10 healthy blood donors was evaluated by intracellular immunofluorescence staining. Serum interleukin-12 (IL-12) levels were measured using an enzyme amplified-sensitivity immunoassay. The effect of recombinant IL-12 (rIL-12) on spontaneous and Fas-mediated apoptosis of phytohemagglutinin (PHA)-stimulated PBL was evaluated by flow cytometry using propidium iodide (PI) staining and a bromodeoxyuridine (BrdU)/PI procedure.Results. Intracellular immunofluorescence staining of IL-2, IL-4, and interferon-␥ (IFN␥) in CD3؉ lymphocytes from BD patients demonstrated a strong polarization of the immune response toward the Th1 pathway that correlated with the progression of BD. Peripheral Th1 cells were significantly increased in patients with active disease (n ‫؍‬ 14) as compared with those in patients in complete remission (n ‫؍‬ 17), patients with inflammatory arthritis, and normal donors. In addition, serum IL-12 levels were

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Summary Bone remodelling is severely affected in myeloma bone disease as a consequence of skeletal metastatization of malignant plasma cells. We investigated whether defective bone replacement is dependent on increased osteoblast apoptosis and/or on deregulated events within the bone microenvironment. Circulating tumour necrosis factor (TNF)‐α, interferon‐γ, interleukin (IL)‐1β, and IL‐6 levels were higher in myeloma patients with overt bone disease, whose osteoblasts constitutively overexpressed Fas, DR4/DR5 complex as receptors to TNF‐related apoptosis inducing ligand, intercellular adhesion molecule‐1 (ICAM‐1), and monocyte chemotactic protein‐1 (MCP‐1). They were functionally exhausted and promptly underwent apoptosis in vitro, in contrast to the minor tendency to death detected in control osteoblasts from patients without bone involvement and normal donors. Osteoblasts dramatically enhanced their apoptosis in co‐cultures with MCC‐2 myeloma cells and upregulated both ICAM‐1 and MCP‐1 in a manner similar to control osteoblasts. Pretreating MCC‐2 cells with soluble ICAM‐1 led to a striking inhibition of their adhesion to osteoblasts, suggesting that the ICAM‐1/lymphocyte function‐associated antigen‐1 system plays a role in the reciprocal membrane contact to trigger apoptogenic signals. Our data suggest that, in the myeloma bone microenvironment, both high cytokine levels and physical interaction of malignant plasma cells with osteoblasts drive the accelerated apoptosis in these cells leading to defective new bone formation.

Summary. Typical features of multiple myeloma (MM) are osteolytic lesions and severely affected bone regeneration. This study of 53 MM patients demonstrates an enhancement of osteoblast cytotoxicity by malignant myeloma cells via the upregulation of apoptogenic receptors, including Fas ligand (Fas-L) and tumour-necrosis-factor-related apoptosis inducing ligand (TRAIL). Both were significantly increased in the marrow myeloma cells of patients with extensive osteolytic lesions in a fashion similar to the highly malignant human myeloma cell line MCC-2. Osteoblasts from these subjects over-expressed Fas and death receptor (DR) 4/5 and underwent dramatic apoptosis when co-cultured with either MCC-2 or autologous myeloma cells. In osteoblast and myeloma cell co-cultures, monocyte chemoattractant protein 1 (MCP-1) mRNA was upregulated in osteoblasts from patients with severe bone disease in parallel with increased CC-chemokine receptor R2 (CCR2) expression, the ligand of MCP-1, in the myeloma cells. This chemokine was shown to activate malignant cell migration in vitro. An upregulation of ICAM-1 expression occurred in osteoblasts from patients with active skeleton disease. This upregulation appeared to be an effect of malignant plasma cell contact, as MCC-2 co-culture greatly enhanced ICAM-1 production by resting osteoblasts from patients without skeleton involvement. Our results suggest that osteoblasts in active myeloma are functionally exhausted and promptly undergo apoptosis in the presence of myeloma cells from patients with severe bone disease. It is suggested that this cytotoxic effect plays a pivotal role in the pathogenesis of defective bone repair.