Giuseppe Mangialardi

Stephen Paget and the ‘seed and soil’ theory of metastatic dissemination

The outcome of cancer metastasis depends on multiple interactions between selected metastatic cells and homeostatic mechanisms unique to some organ microenvironments. The English surgeon Stephen Paget (1855–1926) is credited with being the first to postulate the important role played by microenvironment in metastasis formation. The concept of his ‘seed and soil’ theory has been supported and confirmed by numerous publications. This review article summarises the most important literature data about this matter.

Vasculogenic mimicry by bone marrow macrophages in patients with multiple myeloma

Bone marrow macrophages of patients with active and nonactive multiple myeloma (MM), monoclonal gammopathies of undetermined significance (MGUS) and benign anemia (controls) were stimulated for 7 days with vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), and analysed for the expression of endothelial cell (EC) markers by reverse transcription (RT)–PCR, real-time RT–PCR, western blot and immunofluorescence. Their vasculogenic ability was investigated in vitro in a Matrigel assay and in vivo on bone marrow biopsies through dual immunofluorescence and confocal laser microscopy. Active MM macrophages exposed to VEGF and bFGF acquired EC markers and formed capillary-like structures mimicking paired bone marrow ECs (multiple myeloma patient-derived endothelial cells, MMECs), with major responsiveness compared to macrophages from nonactive MM, MGUS or controls. Bone marrow biopsies of active MM harbored ‘mosaic’ vessels, being formed by MMECs, EC-like macrophages and macrophages themselves. These figures were rare in nonactive MM and absent in MGUS or controls. Our data indicate that macrophages contribute to build neovessels in active MM through vasculogenic mimicry, and this ability proceeds parallel to progression of the plasma cell tumors. Macrophages may be a target for the MM antivascular treatment.

Bortezomib and zoledronic acid on angiogenic and vasculogenic activities of bone marrow macrophages in patients with multiple myeloma

Bone marrow neovascularisation supports plasma cell tumour progression in patients with multiple myeloma (MM), and is partially sustained by bone marrow macrophages through their angiogenic and vasculogenic activities. As such, macrophages may be a target for antivascular treatment in MM. Here, we show that bortezomib (BZ) and zoledronic acid (ZOL) display distinct and synergistic inhibitory effects on cell proliferation, adhesion, migration and expression of angiogenic cytokines (i.e.: VEGF, bFGF, HGF and PDGF). Similar effects were found on capillarogenic organisation and expression of vascular markers in cells which became vasculogenic. VEGFR2 and ERK1/2 phosphoactivation as well as NF-kappaB activity were also inhibited. Overall these data provide evidence that the exposure of bone marrow macrophages in MM during the treatment with ZOL and BZ, alone and or in combination, impacts their angiogenic and vasculogenic properties, suggesting that these cells may be considered as a target of both drugs in MM patients.

Zoledronic acid affects over-angiogenic phenotype of endothelial cells in patients with multiple myeloma

Therapeutic doses of zoledronic acid markedly inhibit in vitro proliferation, chemotaxis, and capillarogenesis of bone marrow endothelial cells of patients with multiple myeloma. Zoledronic acid also induces a sizeable reduction of angiogenesis in the in vivo chorioallantoic membrane assay. These effects are partly sustained by gene and protein inhibition of vascular endothelial growth factor and vascular endothelial growth factor receptor 2 in an autocrine loop. Mevastatin, a specific inhibitor of the mevalonate pathway, reverts the zoledronic acid antiangiogenic effect, indicating that the drug halts this pathway. Our results provide evidence of a direct antiangiogenic activity of zoledronic acid on multiple myeloma patient-derived endothelial cells due to at least four different mechanisms identified either in vitro or in vivo. Tentatively, we suggest that the zoledronic acid antitumoral activity in multiple myeloma is also sustained by antiangiogenesis, which would partly account for its therapeutic efficacy in multiple myeloma. [Mol Cancer Ther 2007;6(12):3256–62]

Endothelial Differentiation of Hematopoietic Stem and Progenitor Cells from Patients with Multiple Myeloma

Purpose: Vasculogenesis is a physiologic process typical of fetal development in which new blood vessels develop from undifferentiated precursors (or angioblasts). In tumors, near angiogenesis, vasculogenesis contributes to the formation of the microvascular plexus that is important for diffusion. Here, we show that hematopoietic stem and progenitor cells (HSPC) of multiple myeloma (MM) patients are able to differentiate into cells with endothelial phenotype on exposure to angiogenic cytokines. Experimental Design: Circulating HSPCs were purified with an anti-CD133 antibody from patients with newly diagnosed MM before autologous transplantation and exposed to vascular endothelial growth factor (VEGF), fibroblast growth factor-2 and insulin-like growth factor in a 3-week culture. Results: HSPCs gradually lost CD133 expression and acquired VEGF receptor-2, factor VIII–related antigen, and vascular endothelial-cadherin expression. The expression pattern overlapped with paired MM endothelial cells (MMEC). During culture, cells adhered to fibronectin, spread, and acquired an endothelial cell shape. Differentiated HSPCs also became capillarogenic in the Matrigel assay with maximal activity at the third week of culture. Bone marrow biopsies revealed HSPCs inside the neovessel wall in patients with MM but not in those with monoclonal gammopathy of undetermined significance. Conclusions: In patients with MM, but not in those with monoclonal gammopathy of undetermined significance, HSPCs contribute to the neovessel wall building together with MMECs. Therefore, besides angiogenesis, HSPC-linked vasculogenesis contributes to neovascularization in MM patients. Tentatively, we hypothesize that in HSPC cultures a multipotent cell population expressing low VEGF receptor-2 levels corresponds to the endothelial progenitor cell precursor and seems to be the MMEC precursor.

Angiogenesis and progression in human melanoma.

In tumor growth, angiogenesis, the process of new-formation of blood vessels from pre-existing ones, is uncontrolled and unlimited in time. The vascular phase is characterized by the new-formation of vascular channels that enhances tumor cell proliferation, local invasion and hematogenous metastasis. Human malignant melanoma is a highly metastatic tumor with poor prognosis, and high resistance to treatment. Parallel with progression, melanoma acquires a rich vascular network, whereas an increasing number of tumor cells express the laminin receptor, which enables their adhesion to the vascular wall, favouring tumor cell extravasation and metastases. Melanoma neovascularization has been correlated with poor prognosis, overall survival, ulceration and increased rate of relapse. Secretion of various angiogenic cytokines, i.e. VEGF-A, FGF-2, PGF-1 and -2, IL-8, and TGF-1 by melanoma cells promote the angiogenic switch and has been correlated to transition from the radial to the vertical growth phase, and to the metastatic phase. Moreover, melanoma cells overexpress αvβ3, αvβ5, α2β1 and α5β1 integrins and release, together with stromal cells, higher amount of metalloproteases that increasing their invasive potential and angiogenesis. Basing on these observations, different molecular targets of antiangiogenic molecules has be recognized and various antiangiogenic agents are currently in preclinical and clinical trials for melanoma.

Antiangiogenic therapeutic approaches in multiple myeloma.

Angiogenesis is a constant hallmark of multiple myeloma (MM) progression and has prognostic potential. The pathophysiology of MM-induced angiogenesis involves both direct production of angiogenic cytokines by plasma cells and their induction within the bone marrow microenvironment. An improved understanding of the importance of angiogenesis-related signaling in MM has allowed for the rational use of antiangiogenic therapies in this tumor. This review article summarizes the literature data concerning the employment of the most important antiangiogenic therapeutic agents actually used in preclinical models and clinical settings for the treatment of MM.

Induction Therapy and Stem Cell Mobilization in Patients with Newly Diagnosed Multiple Myeloma

Autologous stem cell transplantation (ASCT) is considered the standard therapy for younger patients with newly diagnosed symptomatic multiple myeloma (MM). The introduction into clinical practice of novel agents, such as the proteasome inhibitor bortezomib and the immunomodulatory derivatives (IMiDs) thalidomide and lenalidomide, has significantly contributed to major advances in MM therapy and prognosis. These novel agents are incorporated into induction regimens to enhance the depth of response before ASCT and further improve post-ASCT outcomes. Between January 2000 and November 2011, 65 patients with MM were transplanted in the Department of Biomedical Science and Clinical Oncology at the University of Bari. According to Durie-Salmon, 60 patients had stage III of disease and 5 stage II. Only 7 patients were in stage B (renal failure). Induction regimens that were administered in two or more cycles were VAD (vincristine, adriamycin, and dexamethasone), Thal-Dex (thalidomide, dexamethasone), Len-Dex (lenalidomide, dexamethasone), Vel-Dex (bortezomib, dexamethasone), VTD (bortezomib, thalidomide, and dexamethasone), and PAD (bortezomib, pegylated liposomal doxorubicin, and dexamethasone). In mobilization procedure, the patients received cyclophosphamide and granulocyte colony-stimulating factor (G-CSF). The number of cells collected through two or more leukapheresess, response after induction, and toxicity were evaluated to define the more adequate up-front induction regimen in transplantation-eligible MM patients.