Targeting of immunosuppressive myeloid cells from glioblastoma patients by modulation of size and surface charge of lipid nanocapsules.

BACKGROUND: Myeloid derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs) are two of the major players involved in the inhibition of anti-tumor immune response in cancer patients, leading to poor prognosis. Selective targeting of myeloid cells has therefore become an attractive therapeutic strategy to relieve immunosuppression and, in this frame, we previously demonstrated that lipid nanocapsules (LNCs) loaded with lauroyl-modified gemcitabine efficiently target monocytic MDSCs in melanoma patients. In this study, we investigated the impact of the physico-chemical characteristics of LNCs, namely size and surface potential, towards immunosuppressive cell targeting. We exploited myeloid cells isolated from glioblastoma patients, which play a relevant role in the immunosuppression, to demonstrate that tailored nanosystems can target not only tumor cells but also tumor-promoting cells, thus constituting an efficient system that could be used to inhibit their function. RESULTS: The incorporation of different LNC formulations with a size of 100 nm, carrying overall positive, neutral or negative charge, was evaluated on leukocytes and tumor-infiltrating cells freshly isolated from glioblastoma patients. We observed that the maximum LNC uptake was obtained in monocytes with neutral 100 nm LNCs, while positively charged 100 nm LNCs were more effective on macrophages and tumor cells, maintaining at low level the incorporation by T cells. The mechanism of uptake was elucidated, demonstrating that LNCs are incorporated mainly by caveolae-mediated endocytosis. CONCLUSIONS: We demonstrated that LNCs can be directed towards immunosuppressive cells by simply modulating their size and charge thus providing a novel approach to exploit nanosystems for anticancer treatment in the frame of immunotherapy.

The immune suppressive microenvironment of human gliomas depends on the accumulation of bone marrow-derived macrophages in the center of the lesion.

BACKGROUND: Systemic and local immune suppression plays a significant role in glioma progression. Glioma microenvironment contains both brain-resident microglial cells (MG) and bone marrow-derived macrophages (BMDM), but the study of their functional and immune regulatory activity has been hampered until now by the lack of markers allowing a proper identification and isolation to collect pure populations. METHODS: Myeloid and lymphoid infiltrate were characterized in grade II, III and IV gliomas by multicolor flow cytometry, along with the composition of the cell subsets of circulating myeloid cells. Macrophages were sorted and tested for their immunosuppressive ability. Moreover, following preoperative administration of 5-aminolevulinic acid to patients, distinct areas of tumor lesion were surgically removed and analyzed, based on protoporphyrin IX fluorescence emission. RESULTS: The immune microenvironment of grade II to grade IV gliomas contains a large proportion of myeloid cells and a small proportion of lymphocytes expressing markers of dysfunctional activity. BMDM and resident MG cells were characterized through a combination of markers, thus permitting their geographical identification in the lesions, their sorting and subsequent analysis of the functional characteristics. The infiltration by BMDM reached the highest percentages in grade IV gliomas, and it increased from the periphery to the center of the lesion, where it exerted a strong immunosuppression that was, instead, absent in the marginal area. By contrast, MG showed little or no suppression. Functional differences, such as iron metabolism and phagocytosis, characterized resident versus blood-derived macrophages. Significant alterations in circulating monocytes were present in grade IV patients, correlating with accumulation of tumor macrophages. CONCLUSIONS: Grade IV gliomas have an alteration in both circulating and tumor-associated myeloid cells and, differently from grade II and III gliomas, show a significant presence of blood-derived, immune suppressive macrophages. BMDM and MG have different functional properties.

Immunosuppressive activity of tumor-infiltrating myeloid cells in patients with meningioma.

Meningiomas WHO grade I and II are common intracranial tumors in adults that normally display a benign outcome, but are characterized by a great clinical heterogeneity and frequent recurrence of the disease. Although the presence of an immune cell infiltrate has been documented in these tumors, a clear phenotypical and functional characterization of the immune web is missing. Here, we performed an extensive immunophenotyping of peripheral blood and fresh tumor tissue at surgery by multiparametric flow cytometry in 34 meningioma patients, along with immunosuppressive activity of sorted cells of myeloid origin. Four subsets of myeloid cells, phenotypically corresponding to myeloid-derived suppressor cells (MDSCs) are detectable in the blood and in the tumor tissue of patients and three of them are significantly expanded in the blood of patients, but show no evidence of suppressive activity. At the tumor site, a large leukocyte infiltrate is present, predominantly constituted by CD33+ myeloid cells, largely composed of macrophages endowed with suppressive activity and significantly expanded in grade II meningioma patients as compared to grade I.