In a large number of cancer types, treatment selection depends on the presence of specific tumor biomarkers. Due to the dynamic nature of cancer, very often these predictive biomarkers are not uniformly present in all cancer cells. Tumor heterogeneity represents indeed one of the main causes of therapeutic failure, and its decoding remains a major ongoing challenge in the field. Liquid biopsy is the sampling and analysis of non-solid biological tissue often through rapid and non-invasive methods, which allows the assessment in real-time of the evolving landscape of cancer. Samples can be obtained from blood and most other bodily fluids. A blood-based liquid biopsy can capture circulating tumor cells and leukocytes, as well as circulating tumor-derived nucleic acids. In this review, we discuss the current and possibly future applications of blood-based liquid biopsy in oncology, its advantages and its limitations in clinical practice. We specifically focused on its role as a tool to capture tumor heterogeneity in metastatic cancer patients.

The possibility of detecting and classifying living cells in a label-free and non-invasive manner holds significant theranostic potential. In this work, Hyperspectral Imaging (HSI) has been successfully applied to the analysis of macrophagic polarization, given its central role in several pathological settings, including the regulation of tumour microenvironment. Human monocyte derived macrophages have been investigated using hyperspectral reflectance confocal microscopy, and hyperspectral datasets have been analysed in terms of M1 vs. M2 polarization by Principal Components Analysis (PCA). Following PCA, Linear Discriminant Analysis has been implemented for semi-automatic classification of macrophagic polarization from HSI data. Our results confirm the possibility to perform single-cell-level in vitro classification of M1 vs. M2 macrophages in a non-invasive and label-free manner with a high accuracy (above 98% for cells deriving from the same donor), supporting the idea of applying the technique to the study of complex interacting cellular systems, such in the case of tumour-immunity in vitro models.

Abiraterone acetate (ABI) is associated not only with a significant survival advantage in both chemotherapy-naive and -treated patients with metastatic castration-resistant prostate cancer (mCRPC), but also with a delay in time to development of Skeletal Related Events and in radiological skeletal progression. These bone benefits may be related to a direct effect on prostate cancer cells in bone or to a specific mechanism directed to bone microenvironment. To test this hypothesis we designed an in vitro study aimed to evaluate a potential direct effect of ABI on human primary osteoclasts/osteoblasts (OCLs/OBLs). We also assessed changes in bone turnover markers, serum carboxy-terminal collagen crosslinks (CTX) and alkaline phosphatase (ALP), in 49 mCRPC patients treated with ABI.Our results showed that non-cytotoxic doses of ABI have a statistically significant inhibitory effect on OCL differentiation and activity inducing a down-modulation of OCL marker genes TRAP, cathepsin K and metalloproteinase-9. Furthermore ABI promoted OBL differentiation and bone matrix deposition up-regulating OBL specific genes, ALP and osteocalcin. Finally, we observed a significant decrease of serum CTX values and an increase of ALP in ABI-treated patients.These findings suggest a novel biological mechanism of action of ABI consisting in a direct bone anabolic and anti-resorptive activity.

Bone metastasis remains a major cause of mortality and morbidity in breast cancer. Therefore, there is an urgent need to better select high-risk patients in order to adapt patient’s treatment and prevent bone recurrence. Here, we found that integrin alpha5 (ITGA5) was highly expressed in bone metastases, compared to lung, liver, or brain metastases. High ITGA5 expression in primary tumors correlated with the presence of disseminated tumor cells in bone marrow aspirates from early stage breast cancer patients (n = 268; p = 0.039). ITGA5 was also predictive of poor bone metastasis-free survival in two separate clinical data sets (n = 855, HR = 1.36, p = 0.018 and n = 427, HR = 1.62, p = 0.024). This prognostic value remained significant in multivariate analysis (p = 0.028). Experimentally, ITGA5 silencing impaired tumor cell adhesion to fibronectin, migration, and survival. ITGA5 silencing also reduced tumor cell colonization of the bone marrow and formation of osteolytic lesions in vivo. Conversely, ITGA5 overexpression promoted bone metastasis. Pharmacological inhibition of ITGA5 with humanized monoclonal antibody M200 (volociximab) recapitulated inhibitory effects of ITGA5 silencing on tumor cell functions in vitro and tumor cell colonization of the bone marrow in vivo. M200 also markedly reduced tumor outgrowth in experimental models of bone metastasis or tumorigenesis, and blunted cancer-associated bone destruction. ITGA5 was not only expressed by tumor cells but also osteoclasts. In this respect, M200 decreased human osteoclast-mediated bone resorption in vitro. Overall, this study identifies ITGA5 as a mediator of breast-to-bone metastasis and raises the possibility that volociximab/M200 could be repurposed for the treatment of ITGA5-positive breast cancer patients with bone metastases.

Osteosarcoma (OS) is the most common primary malignant tumor of the bone. Due to its high heterogeneity and to survival signals from bone microenvironment, OS can resist to standard treatments, therefore novel therapies are needed. c-MET oncogene, a tyrosine-kinase receptor, plays a crucial role in OS initiation and progression. The present study aimed to evaluate the effect of c-MET inhibitor cabozantinib (CBZ) on OS both directly and through its action on bone microenvironment. We tested different doses of CBZ in in vitro models of OS alone or in co-culture with bone cells in order to reproduce OS-tumor microenvironment interactions. CBZ is able to decrease proliferation and migration of OS cells, inhibiting ERK and AKT signaling pathways. Furthermore, CBZ leads to the inhibition of the proliferation of OS cells expressing receptor activator of nuclear factor κB (RANK), due to its effect on bone microenvironment, where it causes an overproduction of osteoprotegerin and a decrease of production of RANK ligand by osteoblasts. Overall, our data demonstrate that CBZ might represent a new potential treatment against OS, affecting both OS cells and their microenvironment. In this scenario, RANK expression in OS cells could represent a predictive factor of better response to CBZ treatment.